Acute alcohol intoxication-induced microvascular leakage.

PubMed

Doggett, Travis M; Breslin, Jerome W

2014-09-01

Alcohol intoxication can increase inflammation and worsen injury, yet the mechanisms involved are not clear. We investigated whether acute alcohol intoxication increases microvascular permeability and investigated potential signaling mechanisms in endothelial cells that may be involved. Conscious rats received a 2.5 g/kg alcohol bolus via gastric catheters to produce acute intoxication. Microvascular leakage of intravenously administered fluorescein isothiocyanate (FITC)-conjugated albumin (FITC-albumin) from the mesenteric microcirculation was assessed by intravital microscopy. Endothelial-specific mechanisms were studied using cultured endothelial cell monolayers. Transendothelial electrical resistance (TER) served as an index of barrier function, before and after treatment with alcohol or its metabolite acetaldehyde. Pharmacologic agents were used to test the roles of alcohol metabolism, oxidative stress, p38 mitogen-activated protein kinase (MAPK), myosin light-chain kinase (MLCK), rho kinase (ROCK), and exchange protein activated by cAMP (Epac). VE-cadherin localization was investigated to assess junctional integrity. Rac1 and RhoA activation was assessed by ELISA assays. Alcohol significantly increased FITC-albumin extravasation from the mesenteric microcirculation. Alcohol also significantly decreased TER and disrupted VE-cadherin organization at junctions. Acetaldehyde significantly decreased TER, but inhibition of alcohol dehydrogenase or application of a superoxide dismutase mimetic failed to prevent alcohol-induced decreases in TER. Inhibition of p38 MAPK, but not MLCK or ROCK, significantly attenuated the alcohol-induced barrier dysfunction. Alcohol rapidly decreased GTP-bound Rac1 but not RhoA during the drop in TER. Activation of Epac increased TER, but did not prevent alcohol from decreasing TER. However, activation of Epac after initiation of alcohol-induced barrier dysfunction quickly resolved TER to baseline levels. Our results suggest that alcohol intoxication increases microvascular permeability to plasma proteins. The data also suggest the endothelial-specific mechanism involves the p38 MAPK, Rac1, and reorganization of VE-cadherin at junctions. Last, activation of Epac can quickly resolve alcohol-induced endothelial barrier dysfunction. Copyright © 2014 by the Research Society on Alcoholism.

High glucose, glucose fluctuation and carbonyl stress enhance brain microvascular endothelial barrier dysfunction: Implications for diabetic cerebral microvasculature.

PubMed

Li, Wei; Maloney, Ronald E; Aw, Tak Yee

2015-08-01

We previously demonstrated that in normal glucose (5mM), methylglyoxal (MG, a model of carbonyl stress) induced brain microvascular endothelial cell (IHEC) dysfunction that was associated with occludin glycation and prevented by N-acetylcysteine (NAC). Herein, we investigated the impact of high glucose and low GSH, conditions that mimicked the diabetic state, on MG-induced IHEC dysfunction. MG-induced loss of transendothelial electrical resistance (TEER) was potentiated in IHECs cultured for 7 or 12 days in 25 mM glucose (hyperglycemia); moreover, barrier function remained disrupted 6h after cell transfer to normal glucose media (acute glycemic fluctuation). Notably, basal occludin glycation was elevated under these glycemic states. TEER loss was exaggerated by inhibition of glutathione (GSH) synthesis and abrogated by NAC, which corresponded to GSH decreases and increases, respectively. Significantly, glyoxalase II activity was attenuated in hyperglycemic cells. Moreover, hyperglycemia and GSH inhibition increased MG accumulation, consistent with a compromised capacity for MG elimination. α-Oxoaldehydes (MG plus glyoxal) levels were elevated in streptozotocin-induced diabetic rat plasma. Immunohistochemistry revealed a prevalence of MG-positive, but fewer occludin-positive microvessels in the diabetic brain in vivo, and Western analysis confirmed an increase in MG-occludin adducts. These results provide the first evidence that hyperglycemia and acute glucose fluctuation promote MG-occludin formation and exacerbate brain microvascular endothelial dysfunction. Low occludin expression and high glycated-occludin contents in diabetic brain in vivo are factors that would contribute to the dysfunction of the cerebral microvasculature during diabetes. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

High glucose, glucose fluctuation and carbonyl stress enhance brain microvascular endothelial barrier dysfunction: Implications for diabetic cerebral microvasculature

PubMed Central

Li, Wei; Maloney, Ronald E.; Aw, Tak Yee

2015-01-01

We previously demonstrated that in normal glucose (5 mM), methylglyoxal (MG, a model of carbonyl stress) induced brain microvascular endothelial cell (IHEC) dysfunction that was associated with occludin glycation and prevented by N-acetylcysteine (NAC). Herein, we investigated the impact of high glucose and low GSH, conditions that mimicked the diabetic state, on MG-induced IHEC dysfunction. MG-induced loss of transendothelial electrical resistance (TEER) was potentiated in IHECs cultured for 7 or 12 days in 25 mM glucose (hyperglycemia); moreover, barrier function remained disrupted 6 h after cell transfer to normal glucose media (acute glycemic fluctuation). Notably, basal occludin glycation was elevated under these glycemic states. TEER loss was exaggerated by inhibition of glutathione (GSH) synthesis and abrogated by NAC, which corresponded to GSH decreases and increases, respectively. Significantly, glyoxalase II activity was attenuated in hyperglycemic cells. Moreover, hyperglycemia and GSH inhibition increased MG accumulation, consistent with a compromised capacity for MG elimination. α-Oxoaldehydes (MG plus glyoxal) levels were elevated in streptozotocin-induced diabetic rat plasma. Immunohistochemistry revealed a prevalence of MG-positive, but fewer occludin-positive microvessels in the diabetic brain in vivo, and Western analysis confirmed an increase in MG–occludin adducts. These results provide the first evidence that hyperglycemia and acute glucose fluctuation promote MG–occludin formation and exacerbate brain microvascular endothelial dysfunction. Low occludin expression and high glycated-occludin contents in diabetic brain in vivo are factors that would contribute to the dysfunction of the cerebral microvasculature during diabetes. PMID:25867911

Hypothyroidism Is Associated With Coronary Endothelial Dysfunction in Women.

PubMed

Sara, Jaskanwal D; Zhang, Ming; Gharib, Hossein; Lerman, Lilach O; Lerman, Amir

2015-07-29

Hypothyroidism is associated with an increased risk of coronary artery disease, beyond that which can be explained by its association with conventional cardiovascular risk factors. Coronary endothelial dysfunction precedes atherosclerosis, has been linked to adverse cardiovascular events, and may account for some of the increased risk in patients with hypothyroidism. The aim of this study was to determine whether there is an association between epicardial and microvascular coronary endothelial dysfunction and hypothyroidism. In 1388 patients (mean age 50.5 [12.3] years, 34% male) presenting with stable chest pain to Mayo Clinic, Rochester, MN for diagnostic coronary angiography, and who were found to have nonobstructive coronary artery disease (<40% stenosis), we invasively assessed coronary artery endothelial-dependent microvascular and epicardial function by evaluating changes in coronary blood flow (% Δ CBF Ach) and diameter (% Δ CAD Ach), respectively, in response to intracoronary infusions of acetylcholine. Patients were divided into 2 groups: hypothyroidism, defined as a documented history of hypothyroidism or a thyroid-stimulating hormone (TSH) >10.0 mU/mL, n=188, and euthyroidism, defined as an absence of a history of hypothyroidism in the clinical record and/or 0.3

Resveratrol recruits rat muscle microvasculature via a nitric oxide-dependent mechanism that is blocked by TNFα

PubMed Central

Wang, Nasui; Ko, Seung-Hyun; Chai, Weidong; Li, Guolian; Barrett, Eugene J.; Tao, Lijian; Cao, Wenhong

2011-01-01

Resveratrol, a polyphenol found in many plants, has antioxidant and anti-inflammatory actions. It also improves endothelial function and may be cardioprotective. Tumor necrosis factor-α (TNFα) causes oxidative stress and microvascular endothelial dysfunction. Whether resveratrol affects microvascular function in vivo and, if so, whether inflammatory cytokines antagonize its microvascular action are not clear. In cultured bovine aortic endothelial cells (BAECs), reserveratrol (100 nM) increased the phosphorylation of protein kinase B (Akt), endothelial nitric oxide (NO) synthase (eNOS), and ERK1/2 within 15 min by more than twofold, and this effect lasted for at least 2 h. Treatment of BAECs with TNFα (10 ng/ml) significantly increased the NADPH oxidase activity and the production of hydrogen peroxide and superoxide. Pretreatment of cells with resveratrol (100 nM) prevented each of these. Injection (ip) of resveratrol in rats potently increased muscle microvascular blood volume (MBV; P = 0.007) and flow (MBF; P < 0.02) within 30 min, and this was sustained for at least 2 h. The phosphorylation of Akt in liver or muscle was unchanged. Superimposed systemic infusion of l-NAME (NOS inhibitor) completely abolished resveratrol-induced increases in MBV and MBF. Similarly, systemic infusion of TNFα prevented resveratrol-induced muscle microvascular recruitment. In conclusion, resveratrol activates eNOS and increases muscle microvascular recruitment via an NO-dependent mechanism. Despite the potent antioxidant effect of resveratrol, TNFα at concentrations that block insulin-mediated muscle microvascular recruitment completely neutralized resveratrol's microvascular action. Thus, chronic inflammation, as seen in type 2 diabetes, may limit resveratrol's vasodilatory actions on muscle microvasculature. PMID:20978231

The presence of African American race predicts improvement in coronary endothelial function after supplementary L-arginine.

PubMed

Houghton, Jan L; Philbin, Edward F; Strogatz, David S; Torosoff, Mikhail T; Fein, Steven A; Kuhner, Patricia A; Smith, Vivienne E; Carr, Albert A

2002-04-17

The purpose of our study was to determine if the presence of African American ethnicity modulates improvement in coronary vascular endothelial function after supplementary L-arginine. Endothelial dysfunction is an early stage in the development of coronary atherosclerosis and has been implicated in the pathogenesis of hypertension and cardiomyopathy. Amelioration of endothelial dysfunction has been demonstrated in patients with established coronary atherosclerosis or with risk factors in response to infusion of L-arginine, the precursor of nitric oxide. Racial and gender patterns in L-arginine responsiveness have not, heretofore, been studied. Invasive testing of coronary artery and microvascular reactivity in response to graded intracoronary infusions of acetylcholine (ACh) +/- L-arginine was carried out in 33 matched pairs of African American and white subjects with no angiographic coronary artery disease. Pairs were matched for age, gender, indexed left ventricular mass, body mass index and low-density lipoprotein cholesterol. In addition to the matching parameters, there were no significant differences in peak coronary blood flow (CBF) response to intracoronary adenosine or in the peak CBF response to ACh before L-arginine infusion. However, absolute percentile improvement in CBF response to ACh infusion after L-arginine, as compared with before, was significantly greater among African Americans as a group (45 +/- 10% vs. 4 +/- 6%, p = 0.0016) and after partitioning by gender. The mechanism of this increase was mediated through further reduction in coronary microvascular resistance. L-arginine infusion also resulted in greater epicardial dilator response after ACh among African Americans. We conclude that intracoronary infusion of L-arginine provides significantly greater augmentation of endothelium-dependent vascular relaxation in those of African American ethnicity when compared with matched white subjects drawn from a cohort electively referred for coronary angiography. Our findings suggest that there are target populations in which supplementary L-arginine may be of therapeutic benefit in the amelioration of microvascular endothelial dysfunction. In view of the excess prevalence of cardiomyopathy among African Americans, pharmacologic correction of microcirculatory endothelial dysfunction in this group is an important area of further investigation and may ultimately prove to be clinically indicated.

A novel effective method for the assessment of microvascular function in male patients with coronary artery disease: a pilot study using laser speckle contrast imaging.

PubMed

Borges, J P; Lopes, G O; Verri, V; Coelho, M P; Nascimento, P M C; Kopiler, D A; Tibirica, E

2016-09-01

Evaluation of microvascular endothelial function is essential for investigating the pathophysiology and treatment of cardiovascular and metabolic diseases. Although laser speckle contrast imaging technology is well accepted as a noninvasive methodology for assessing microvascular endothelial function, it has never been used to compare male patients with coronary artery disease with male age-matched healthy controls. Thus, the aim of this study was to determine whether laser speckle contrast imaging could be used to detect differences in the systemic microvascular functions of patients with established cardiovascular disease (n=61) and healthy age-matched subjects (n=24). Cutaneous blood flow was assessed in the skin of the forearm using laser speckle contrast imaging coupled with the transdermal iontophoretic delivery of acetylcholine and post-occlusive reactive hyperemia. The maximum increase in skin blood flow induced by acetylcholine was significantly reduced in the cardiovascular disease patients compared with the control subjects (74 vs 116%; P<0.01). With regard to post-occlusive reactive hyperemia-induced vasodilation, the patients also presented reduced responses compared to the controls (0.42±0.15 vs 0.50±0.13 APU/mmHg; P=0.04). In conclusion, laser speckle contrast imaging can identify endothelial and microvascular dysfunctions in male individuals with cardiovascular disease. Thus, this technology appears to be an efficient non-invasive technique for evaluating systemic microvascular and endothelial functions, which could be valuable as a peripheral marker of atherothrombotic diseases in men.

Endothelial dysfunction as a predictor of cardiovascular disease in type 1 diabetes

PubMed Central

Bertoluci, Marcello C; Cé, Gislaine V; da Silva, Antônio MV; Wainstein, Marco V; Boff, Winston; Puñales, Marcia

2015-01-01

Macro and microvascular disease are the main cause of morbi-mortality in type 1 diabetes (T1DM). Although there is a clear association between endothelial dysfunction and atherosclerosis in type 2 diabetes, a cause-effect relationship is less clear in T1DM. Although endothelial dysfunction (ED) precedes atherosclerosis, it is not clear weather, in recent onset T1DM, it may progress to clinical macrovascular disease. Moreover, endothelial dysfunction may either be reversed spontaneously or in response to intensive glycemic control, long-term exercise training and use of statins. Acute, long-term and post-prandial hyperglycemia as well as duration of diabetes and microalbuminuria are all conditions associated with ED in T1DM. The pathogenesis of endothelial dysfunction is closely related to oxidative-stress. NAD(P)H oxidase over activity induces excessive superoxide production inside the mitochondrial oxidative chain of endothelial cells, thus reducing nitric oxide bioavailability and resulting in peroxynitrite formation, a potent oxidant agent. Moreover, oxidative stress also uncouples endothelial nitric oxide synthase, which becomes dysfunctional, inducing formation of superoxide. Other important mechanisms are the activation of both the polyol and protein kinase C pathways as well as the presence of advanced glycation end-products. Future studies are needed to evaluate the potential clinical applicability of endothelial dysfunction as a marker for early vascular complications in T1DM. PMID:26069717

Brain endothelial dysfunction in cerebral adrenoleukodystrophy.

PubMed

Musolino, Patricia L; Gong, Yi; Snyder, Juliet M T; Jimenez, Sandra; Lok, Josephine; Lo, Eng H; Moser, Ann B; Grabowski, Eric F; Frosch, Matthew P; Eichler, Florian S

2015-11-01

See Aubourg (doi:10.1093/awv271) for a scientific commentary on this article.X-linked adrenoleukodystrophy is caused by mutations in the ABCD1 gene leading to accumulation of very long chain fatty acids. Its most severe neurological manifestation is cerebral adrenoleukodystrophy. Here we demonstrate that progressive inflammatory demyelination in cerebral adrenoleukodystrophy coincides with blood-brain barrier dysfunction, increased MMP9 expression, and changes in endothelial tight junction proteins as well as adhesion molecules. ABCD1, but not its closest homologue ABCD2, is highly expressed in human brain microvascular endothelial cells, far exceeding its expression in the systemic vasculature. Silencing of ABCD1 in human brain microvascular endothelial cells causes accumulation of very long chain fatty acids, but much later than the immediate upregulation of adhesion molecules and decrease in tight junction proteins. This results in greater adhesion and transmigration of monocytes across the endothelium. PCR-array screening of human brain microvascular endothelial cells after ABCD1 silencing revealed downregulation of both mRNA and protein levels of the transcription factor c-MYC (encoded by MYC). Interestingly, MYC silencing mimicked the effects of ABCD1 silencing on CLDN5 and ICAM1 without decreasing the levels of ABCD1 protein itself. Together, these data demonstrate that ABCD1 deficiency induces significant alterations in brain endothelium via c-MYC and may thereby contribute to the increased trafficking of leucocytes across the blood-brain barrier as seen in cerebral adrenouleukodystrophy. © The Author (2015). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Loss of the Endothelial Glycocalyx Links Albuminuria and Vascular Dysfunction

PubMed Central

Ferguson, Joanne K.; Burford, James L.; Gevorgyan, Haykanush; Nakano, Daisuke; Harper, Steven J.; Bates, David O.; Peti-Peterdi, Janos

2012-01-01

Patients with albuminuria and CKD frequently have vascular dysfunction but the underlying mechanisms remain unclear. Because the endothelial surface layer, a meshwork of surface-bound and loosely adherent glycosaminoglycans and proteoglycans, modulates vascular function, its loss could contribute to both renal and systemic vascular dysfunction in proteinuric CKD. Using Munich-Wistar-Fromter (MWF) rats as a model of spontaneous albuminuric CKD, multiphoton fluorescence imaging and single-vessel physiology measurements revealed that old MWF rats exhibited widespread loss of the endothelial surface layer in parallel with defects in microvascular permeability to both water and albumin, in both continuous mesenteric microvessels and fenestrated glomerular microvessels. In contrast to young MWF rats, enzymatic disruption of the endothelial surface layer in old MWF rats resulted in neither additional loss of the layer nor additional changes in permeability. Intravenous injection of wheat germ agglutinin lectin and its adsorption onto the endothelial surface layer significantly improved glomerular albumin permeability. Taken together, these results suggest that widespread loss of the endothelial surface layer links albuminuric kidney disease with systemic vascular dysfunction, providing a potential therapeutic target for proteinuric kidney disease. PMID:22797190

Adult males with haemophilia have a different macrovascular and microvascular endothelial function profile compared with healthy controls.

PubMed

Sun, H; Yang, M; Fung, M; Chan, S; Jawi, M; Anderson, T; Poon, M-C; Jackson, S

2017-09-01

Endothelial function has been identified as an independent predictor of cardiovascular risk in the general population. It is unclear if the haemophilia population has a different endothelial function profile compared to the healthy population. This prospective study aims to assess if there is a difference in endothelial function between haemophilia patients and healthy controls, and the impact of endothelial function on vascular outcomes in the haemophilia population. Baseline cardiovascular risk factors and endothelial function were presented. Adult males with haemophilia A or B recruited from the British Columbia and Southern Alberta haemophilia treatment centres were matched to healthy male controls by age and cardiovascular risk factors. Macrovascular endothelial function was assessed by brachial artery flow-mediated dilation (FMD) and nitroglycerin-mediated dilation (NMD), and microvascular endothelial function was assessed by hyperaemic velocity time integral (VTI). Multivariable linear regression was used to assess the association between haemophilia and endothelial function. A total of 81 patients with haemophilia and 243 controls were included. Patients with haemophilia had a similar FMD and NMD compared to controls, although haemophilia was associated with higher FMD on multivariable analysis. Haemophilia was associated with significantly lower VTI on univariate and multivariable analyses, regardless of haemophilia type and severity. Adult males with haemophilia appear to have lower microvascular endothelial function compared to healthy controls. Future studies to assess the impact of endothelial dysfunction on cardiovascular events in the haemophilia population are needed. © 2017 John Wiley & Sons Ltd.

A novel effective method for the assessment of microvascular function in male patients with coronary artery disease: a pilot study using laser speckle contrast imaging

PubMed Central

Borges, J.P.; Lopes, G.O.; Verri, V.; Coelho, M.P.; Nascimento, P.M.C.; Kopiler, D.A.; Tibirica, E.

2016-01-01

Evaluation of microvascular endothelial function is essential for investigating the pathophysiology and treatment of cardiovascular and metabolic diseases. Although laser speckle contrast imaging technology is well accepted as a noninvasive methodology for assessing microvascular endothelial function, it has never been used to compare male patients with coronary artery disease with male age-matched healthy controls. Thus, the aim of this study was to determine whether laser speckle contrast imaging could be used to detect differences in the systemic microvascular functions of patients with established cardiovascular disease (n=61) and healthy age-matched subjects (n=24). Cutaneous blood flow was assessed in the skin of the forearm using laser speckle contrast imaging coupled with the transdermal iontophoretic delivery of acetylcholine and post-occlusive reactive hyperemia. The maximum increase in skin blood flow induced by acetylcholine was significantly reduced in the cardiovascular disease patients compared with the control subjects (74 vs 116%; P<0.01). With regard to post-occlusive reactive hyperemia-induced vasodilation, the patients also presented reduced responses compared to the controls (0.42±0.15 vs 0.50±0.13 APU/mmHg; P=0.04). In conclusion, laser speckle contrast imaging can identify endothelial and microvascular dysfunctions in male individuals with cardiovascular disease. Thus, this technology appears to be an efficient non-invasive technique for evaluating systemic microvascular and endothelial functions, which could be valuable as a peripheral marker of atherothrombotic diseases in men. PMID:27599202

2-Chlorohexadecanoic acid induces ER stress and mitochondrial dysfunction in brain microvascular endothelial cells.

PubMed

Bernhart, Eva; Kogelnik, Nora; Prasch, Jürgen; Gottschalk, Benjamin; Goeritzer, Madeleine; Depaoli, Maria Rosa; Reicher, Helga; Nusshold, Christoph; Plastira, Ioanna; Hammer, Astrid; Fauler, Günter; Malli, Roland; Graier, Wolfgang F; Malle, Ernst; Sattler, Wolfgang

2018-05-01

Peripheral leukocytes induce blood-brain barrier (BBB) dysfunction through the release of cytotoxic mediators. These include hypochlorous acid (HOCl) that is formed via the myeloperoxidase-H 2 O 2 -chloride system of activated phagocytes. HOCl targets the endogenous pool of ether phospholipids (plasmalogens) generating chlorinated inflammatory mediators like e.g. 2-chlorohexadecanal and its conversion product 2-chlorohexadecanoic acid (2-ClHA). In the cerebrovasculature these compounds inflict damage to brain microvascular endothelial cells (BMVEC) that form the morphological basis of the BBB. To follow subcellular trafficking of 2-ClHA we synthesized a 'clickable' alkyne derivative (2-ClHyA) that phenocopied the biological activity of the parent compound. Confocal and superresolution structured illumination microscopy revealed accumulation of 2-ClHyA in the endoplasmic reticulum (ER) and mitochondria of human BMVEC (hCMEC/D3 cell line). 2-ClHA and its alkyne analogue interfered with protein palmitoylation, induced ER-stress markers, reduced the ER ATP content, and activated transcription and secretion of interleukin (IL)-6 as well as IL-8. 2-ClHA disrupted the mitochondrial membrane potential and induced procaspase-3 and PARP cleavage. The protein kinase R-like ER kinase (PERK) inhibitor GSK2606414 suppressed 2-ClHA-mediated activating transcription factor 4 synthesis and IL-6/8 secretion, but showed no effect on endothelial barrier dysfunction and cleavage of procaspase-3. Our data indicate that 2-ClHA induces potent lipotoxic responses in brain endothelial cells and could have implications in inflammation-induced BBB dysfunction. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Uric acid is associated with inflammation, coronary microvascular dysfunction, and adverse outcomes in postmenopausal women

PubMed Central

Prasad, Megha; Matteson, Eric L.; Herrmann, Joerg; Gulati, Rajiv; Rihal, Charanjit S.; Lerman, Lilach O.; Lerman, Amir

2016-01-01

Uric acid is a risk factor for coronary artery disease (CAD) in postmenopausal women but the association with inflammation and coronary microvascular endothelial dysfunction (CED) is not well-defined. The aim of this study was to determine the relationship of serum uric acid (SUA), inflammatory markers and CED. In this prospective cohort study, serum uric acid, hsCRP levels, and neutrophil count were measured in 229 postmenopausal women who underwent diagnostic catheterization, were found to have no obstructive CAD and underwent coronary microvascular function testing, to measure coronary blood flow (CBF) response to intracoronary acetylcholine. The average age was 58 years (IQR 52, 66) years. Hypertension was present in 48%, type 2 diabetes mellitus in 5.6%, and hyperlipidemia in 61.8%. CED was diagnosed in 59% of postmenopausal women. Mean uric acid level was 4.7 ± 1.3 mg/dL. Postmenopausal women with CED had significantly higher SUA compared to patients without CED (4.9 ± 1.3 vs. 4.4 ± 1.3 mg/dL; p=0.02). There was a significant correlation between SUA and % change in CBF to acetylcholine (p=0.009), and this correlation persisted in multivariable analysis. SUA levels were significantly associated with increased neutrophil count (p=0.02) and hsCRP levels (p=0.006) among patients with CED, but not those without CED. Serum uric acid is associated with coronary microvascular endothelial dysfunction in postmenopausal women and may be related to inflammation. These findings link serum uric acid levels to early coronary atherosclerosis in postmenopausal women. PMID:27993955

Inhibitory effect of melatonin on necroptosis via repressing the Ripk3-PGAM5-CypD-mPTP pathway attenuates cardiac microvascular ischemia reperfusion injury.

PubMed

Zhou, Hao; Li, Dandan; Zhu, Pingjun; Ma, Qiang; Sam, Toan; Wang, Jin; Hu, Shunying; Chen, Yundai; Zhang, Yingmei

2018-05-16

The molecular features of necroptosis in cardiac ischemia reperfusion (IR) injury have been extensively explored. However, there have been no studies investigating the physiological regulatory mechanisms of melatonin acting on necroptosis in cardiac IR injury. This study was designed to determine the role of necroptosis in microvascular IR injury, and investigate the contribution of melatonin in repressing necroptosis and preventing IR-mediated endothelial system collapse. Our results demonstrated that Ripk3 was primarily activated by IR injury and consequently aggravated endothelial necroptosis, microvessel barrier dysfunction, capillary hyperpermeability, the inflammation response, microcirculatory vasospasms and microvascular perfusion defects. However, administration of melatonin prevented Ripk3 activation and provided a pro-survival advantage for the endothelial system in the context of cardiac IR injury, similar to the results obtained via genetic ablation of Ripk3. Functional investigations clearly illustrated that activated Ripk3 upregulated PGAM5 expression, and the latter repressed CypD phosphorylation, which obligated endothelial cells to undergo necroptosis via augmenting mPTP (mitochondrial permeability transition pore) opening. Interestingly, melatonin supplementation suppressed mPTP opening and interrupted endothelial necroptosis via blocking the Ripk3-PGAM5-CypD signal pathways. Taken together, our studies identified the Ripk3-PGAM5-CypD-mPTP axis as a new pathway responsible for reperfusion-mediated microvascular damage via initiating endothelial necroptosis. In contrast, melatonin treatment inhibited the Ripk3-PGAM5-CypD-mPTP cascade and thus reduced cellular necroptosis, conferring a protective advantage to the endothelial system in IR stress. These findings establish a new paradigm in microvascular IR injury and update the concept for cell death management handled by melatonin under the burden of reperfusion attack. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Oxytocin inhibits ox-LDL-induced adhesion of monocytic THP-1 cells to human brain microvascular endothelial cells.

PubMed

Liu, Shuyan; Pan, Shengying; Tan, Jing; Zhao, Weina; Liu, Fengguo

2017-12-15

The attachment of monocytes to human brain microvascular endothelial cells (HBMVEs) caused by oxidized low-density lipoprotein (ox-LDL) is associated with an early event and the pathological progression of cerebrovascular diseases. Oxytocin (OT) is a human peptide hormone that is traditionally used as a medication to facilitate childbirth. However, little information is available regarding the physiological function of OT in brain endothelial dysfunction. In the present study, our results indicate that the oxytocin receptor (OTR) was expressed in human brain microvascular endothelial cells (HBMVEs) and was upregulated in response to ox-LDL in a concentration-dependent manner. Notably, OT significantly suppressed ox-LDL-induced attachment of THP-1 monocytes to HBMVEs. Furthermore, we found that OT reduced the expression of adhesion molecules, such as VCAM-1 and E-selectin. Interestingly, it was shown that OT could restore ox-LDL-induced reduction of KLF4 in HBMVEs. Importantly, knockdown of KLF4 abolished the inhibitory effects of OT on ox-LDL-induced expressions of VCAM-1 and E-selectin as well as the adhesion of human monocytic THP-1 cells to endothelial HBMVEs. Mechanistically, we found that the stimulatory effects of OT on KLF4 expression are mediated by the MEK5/MEF2A pathway. Copyright © 2017. Published by Elsevier Inc.

Regulation of Human Brain Microvascular Endothelial Cell Adhesion and Barrier Functions by Memantine.

PubMed

Wang, Fei; Zou, Zhirong; Gong, Yi; Yuan, Dong; Chen, Xun; Sun, Tao

2017-05-01

Vascular risk factors have been linked to cognitive decline and dementia in the elderly. Microvascular inflammation, especially of the endothelium, may contribute to the progression of neurodegenerative events in Alzheimer's disease (AD). Memantine, an uncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist, is a licensed drug used for the treatment of moderate to severe AD. However, little information is available regarding its anti-inflammatory effects on the endothelium. In this study, we investigated the effects of memantine on human brain microvascular endothelial dysfunction induced by the pro-inflammatory cytokine tumor necrosis factor-α (TNF-α). Our results show that memantine prevents the attachment of monocyte THP-1 cells to human brain microvascular endothelial cells (HBMVEs). An in vitro BBB model experiment displayed that memantine could rescue TNF-α-induced disruption of the in vitro BBB model. In addition, memantine also interferes with monocyte transmigration across the BBB model. Our results indicate that TNF-α significantly increased the expression of cell adhesion molecules, such as ICAM-1, VCAM-1, and E-selectin, which was prevented by pretreatment with memantine. Mechanistically, memantine reversed activation of the transcription factor NF-κB by preventing the phosphorylation and degradation of its inhibitor IκBα. Our data is the first to describe a novel anti-inflammatory mechanism driven by the endothelial cell-mediated neuroprotective effects of memantine.

Poly(I:C) Induces Human Lung Endothelial Barrier Dysfunction by Disrupting Tight Junction Expression of Claudin-5

DOE PAGES

Huang, Li -Yun; Stuart, Christine; Takeda, Kazuyo; ...

2016-08-09

Viral infections are often accompanied by pulmonary microvascular leakage and vascular endothelial dysfunction via mechanisms that are not completely defined. Here, we investigated the effect of the Toll-like receptor 3 (TLR3) ligand polyinosinic-polycytidylic acid [Poly(I:C)], a synthetic analog of viral double-stranded RNA (dsRNA) commonly used to simulate viral infections, on the barrier function and tight junction integrity of primary human lung microvascular endothelial cells. Poly(I:C) stimulated IL-6, IL-8, TNFα, and IFNβ production in conjunction with the activation of NF-κB and IRF3 confirming the Poly(I:C)-responsiveness of these cells. Poly(I:C) increased endothelialmonolayer permeability with a corresponding dose- and time-dependent decrease in themore » expression of claudin-5, a transmembrane tight junction protein and reduction of CLDN5 mRNA levels. Immunofluorescence experiments revealed disappearance of membrane-associated claudin-5 and co-localization of cytoplasmic claudin-5 with lysosomal-associated membrane protein 1. Chloroquine and Bay11-7082, inhibitors of TLR3 and NF-κB signaling, respectively, protected against the loss of claudin-5. Altogether, these findings provide new insight on how dsRNA-activated signaling pathways may disrupt vascular endothelial function and contribute to vascular leakage pathologies.« less

Effects of low- and high-advanced glycation endproduct meals on macro- and microvascular endothelial function and oxidative stress in patients with type 2 diabetes mellitus.

PubMed

Negrean, Monica; Stirban, Alin; Stratmann, Bernd; Gawlowski, Thomas; Horstmann, Tina; Götting, Christian; Kleesiek, Knut; Mueller-Roesel, Michaela; Koschinsky, Theodor; Uribarri, Jaime; Vlassara, Helen; Tschoepe, Diethelm

2007-05-01

An advanced glycation endproducts (AGEs)-rich diet induces significant increases in inflammatory and endothelial dysfunction markers in type 2 diabetes mellitus (T2DM). The aim was to investigate the acute effects of dietary AGEs on vascular function in T2DM patients. Twenty inpatients with T2DM [x (+/-SEM) age: 55.4 +/- 2.2 y; glycated hemoglobin: 8.8 +/- 0.5%] were investigated. In a randomized crossover design, the effects of a low-AGE (LAGE) and high-AGE (HAGE) meal on macrovascular [by flow-mediated dilatation (FMD)] and microvascular (by Laser-Doppler flowmetry) function, serum markers of endothelial dysfunction (E-selectin, intracellular adhesion molecule 1, and vascular cell adhesion molecule 1), oxidative stress, and serum AGE were assessed. The meals had identical ingredients but different AGE amounts (15.100 compared with 2.750 kU AGE for the HAGE and LAGE meals, respectively), which were obtained by varying the cooking temperature and time. The measurements were performed at baseline and 2, 4, and 6 h after each meal. After the HAGE meal, FMD decreased by 36.2%, from 5.77 +/- 0.65% (baseline) to 3.93 +/- 0.48 (2 h), 3.70 +/- 0.42 (4 h), and 4.42 +/- 0.54% (6 h) (P<0.01 for all compared with baseline). After the LAGE meal, FMD decreased by 20.9%, from 6.04 +/- 0.68% (baseline) to 4.75 +/- 0.48% (2 h), 4.69 +/- 0.51% (4 h), and 5.62 +/- 0.63% (6 h), respectively (P<0.01 for all compared with baseline; P<0.001 for all compared with the HAGE meal). This impairment of macrovascular function after the HAGE meal was paralleled by an impairment of microvascular function (-67.2%) and increased concentrations of serum AGE and markers of endothelial dysfunction and oxidative stress. In patients with T2DM, a HAGE meal induces a more pronounced acute impairment of vascular function than does an otherwise identical LAGE meal. Therefore, chemical modifications of food by means of cooking play a major role in influencing the extent of postprandial vascular dysfunction.

Structural and functional changes in the microcirculation of lepromatous leprosy patients - Observation using orthogonal polarization spectral imaging and laser Doppler flowmetry iontophoresis

PubMed Central

Treu, Curt; de Souza, Maria das Graças Coelho; Lupi, Omar; Sicuro, Fernando Lencastre; Maranhão, Priscila Alves; Kraemer-Aguiar, Luiz Guilherme; Bouskela, Eliete

2017-01-01

Leprosy is a chronic granulomatous infection of skin and peripheral nerves caused by Mycobacterium leprae and is considered the main infectious cause of disability worldwide. Despite the several studies regarding leprosy, little is known about its effects on microvascular structure and function in vivo. Thus, we have aimed to compare skin capillary structure and functional density, cutaneous vasomotion (spontaneous oscillations of arteriolar diameter), which ensures optimal blood flow distribution to skin capillaries) and cutaneous microvascular blood flow and reactivity between ten men with lepromatous leprosy (without any other comorbidity) and ten age- and gender-matched healthy controls. Orthogonal polarization spectral imaging was used to evaluate skin capillary morphology and functional density and laser Doppler flowmetry to evaluate blood flow, vasomotion and spectral analysis of flowmotion (oscillations of blood flow generated by vasomotion) and microvascular reactivity, in response to iontophoresis of acetylcholine and sodium nitroprusside. The contribution of different frequency components of flowmotion (endothelial, neurogenic, myogenic, respiratory and cardiac) was not statistically different between groups. However, endothelial-dependent and -independent vasodilatations elicited by acetylcholine and sodium nitroprusside iontophoresis, respectively, were significantly reduced in lepromatous leprosy patients compared to controls, characterizing the existence of microvascular dysfunction. These patients also presented a significant increase in the number of capillaries with morphological abnormalities and in the diameters of the dermal papilla and capillary bulk when compared to controls. Our results suggest that lepromatous leprosy causes severe microvascular dysfunction and significant alterations in capillary structure. These structural and functional changes are probably induced by exposure of the microvascular bed to chronic inflammation evoked by the Mycobacterium leprae. PMID:28419120

Microvascular dysfunction in the immediate aftermath of chronic total coronary occlusion recanalization.

PubMed

Ladwiniec, Andrew; Cunnington, Michael S; Rossington, Jennifer; Thackray, Simon; Alamgir, Farquad; Hoye, Angela

2016-05-01

The aim of this study was to compare microvascular resistance under both baseline and hyperemic conditions immediately after percutaneous coronary intervention (PCI) of a chronic total occlusion (CTO) with an unobstructed reference vessel in the same patient Microvascular dysfunction has been reported to be prevalent immediately after CTO PCI. However, previous studies have not made comparison with a reference vessel. Patients with a CTO may have global microvascular and/or endothelial dysfunction, making comparison with established normal values misleading. After successful CTO PCI in 21 consecutive patients, coronary pressure and flow velocity were measured at baseline and hyperemia in distal segments of the CTO/target vessel and an unobstructed reference vessel. Hemodynamics including hyperemic microvascular resistance (HMR), basal microvascular resistance (BMR), and instantaneous minimal microvascular resistance at baseline and hyperemia were calculated and compared between reference and target/CTO vessels. After CTO PCI, BMR was reduced in the target/CTO vessel compared with the reference vessel: 3.58 mm Hg/cm/s vs 4.94 mm Hg/cm/s, difference -1.36 mm Hg/cm/s (-2.33 to -0.39, p = 0.008). We did not detect a difference in HMR: 1.82 mm Hg/cm/s vs 2.01 mm Hg/cm/s, difference -0.20 (-0.78 to 0.39, p = 0.49). Instantaneous minimal microvascular resistance correlated strongly with the length of stented segment at baseline (r = 0.63, p = 0.005) and hyperemia (r = 0.68, p = 0.002). BMR is reduced in a recanalized CTO in the immediate aftermath of PCI compared to an unobstructed reference vessel; however, HMR appears to be preserved. A longer stented segment is associated with increased microvascular resistance. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Treatment of Angina and Microvascular Coronary Dysfunction

PubMed Central

Samim, Arang; Nugent, Lynn; Mehta, Puja K.; Shufelt, Chrisandra; Merz, C. Noel Bairey

2014-01-01

Opinion statement Microvascular coronary dysfunction (MCD) is an increasingly recognized cause of cardiac ischemia and angina, more commonly diagnosed in women. Patients with MCD present with the triad of persistent chest pain, ischemic changes on stress testing, and no obstructive coronary artery disease (CAD) on cardiac catheterization. Data from National Heart, Lung and Blood Institute (NHLBI)-sponsored Women’s Ischemia Syndrome Evaluation (WISE) study has shown that the diagnosis of MCD is not benign, with a 2.5% annual risk of adverse cardiac events including myocardial infarction, stroke, congestive heart failure, or death. The gold standard diagnostic test for MCD is an invasive coronary reactivity test (CRT), which uses acetylcholine, adenosine, and nitroglycerin to test the endothelial dependent and independent, microvascular and macrovascular coronary function. The CRT allows for diagnostic and treatment options as well as further risk stratifying patients for future cardiovascular events. Treatment of angina and MCD should be aimed at ischemia disease management to reduce risk of adverse cardiac events, ameliorating symptoms to improve quality of life, and to decrease the morbidity from unnecessary and repeated cardiac catheterization in patients with open coronary arteries. A comprehensive treatment approach aimed at risk factor managment, including lifestyle counseling regarding smoking cessation, nutrition and physical activity should be initiated. Current pharmacotherapy for MCD can include the treatment of microvascular endothelial dysfunction (statins, angiotensin-converting enzyme inhibitor, low dose aspirin), as well as treatment for angina and myocardial ischemia (beta blockers, calcium channel blockers, nitrates, ranolazine). Additional symptom management techniques can include tri-cyclic medication, enhanced external counterpulsation, autogenic training, and spinal cord stimulation. While our current therapies are effective in the treatment of angina and MCD, large randomized outcome trials are needed to optimize strategies to improve morbidity and mortality. PMID:20842559

Carbachol inhibits TNF-α-induced endothelial barrier dysfunction through alpha 7 nicotinic receptors.

PubMed

Li, Yu-zhen; Liu, Xiu-hua; Rong, Fei; Hu, Sen; Sheng, Zhi-yong

2010-10-01

To test whether carbachol can influence endothelial barrier dysfunction induced by tumor necrosis factor (TNF)-α and whether the alpha 7 nicotinic receptor can mediate this process. Rat cardiac microvascular endothelial cells were exposed to carbachol followed by TNF-α treatment in the presence or the absence of α-bungarotoxin (an antagonist of the alpha 7 nicotinic receptor). Permeability of endothelial cells cultured on Transwell filters was assayed using FITC-albumin. F-actin was stained with FITC- phalloidin. Expression of vascular endothelial cadherin, intercellular adhesion molecule 1 (ICAM-1), phosphor-ERK1/2 and phosphor-JNK was detected using Western blot. Carbachol (2 μmol/L-2 mmol/L) prevented increase in endothelial cell permeability induced by TNF-α (500 ng/mL) in a dose-dependent manner. Further, it attenuated the down-regulation of vascular endothelial cadherin and the up-regulation of ICAM-1 induced by TNF-α. In addition, treatment of endothelial cells with carbachol decreased phosphor-ERK1/2 and phosphor-JNK. These effects of carbachol were blocked by α-bungarotoxin 3 μg/mL. These data suggest that the inhibitory effect of carbachol on TNF-α-induced endothelial barrier dysfunction mediated by the alpha 7 nicotinic receptor.

Carbachol inhibits TNF-α-induced endothelial barrier dysfunction through alpha 7 nicotinic receptors

PubMed Central

Li, Yu-zhen; Liu, Xiu-hua; Rong, Fei; Hu, Sen; Sheng, Zhi-yong

2010-01-01

Aim: To test whether carbachol can influence endothelial barrier dysfunction induced by tumor necrosis factor (TNF)-α and whether the alpha 7 nicotinic receptor can mediate this process. Methods: Rat cardiac microvascular endothelial cells were exposed to carbachol followed by TNF-α treatment in the presence or the absence of α-bungarotoxin (an antagonist of the alpha 7 nicotinic receptor). Permeability of endothelial cells cultured on Transwell filters was assayed using FITC-albumin. F-actin was stained with FITC- phalloidin. Expression of vascular endothelial cadherin, intercellular adhesion molecule 1 (ICAM-1), phosphor-ERK1/2 and phosphor-JNK was detected using Western blot. Results: Carbachol (2 μmol/L-2 mmol/L) prevented increase in endothelial cell permeability induced by TNF-α (500 ng/mL) in a dose-dependent manner. Further, it attenuated the down-regulation of vascular endothelial cadherin and the up-regulation of ICAM-1 induced by TNF-α. In addition, treatment of endothelial cells with carbachol decreased phosphor-ERK1/2 and phosphor-JNK. These effects of carbachol were blocked by α-bungarotoxin 3 μg/mL. Conclusion: These data suggest that the inhibitory effect of carbachol on TNF-α-induced endothelial barrier dysfunction mediated by the alpha 7 nicotinic receptor. PMID:20871620

A role for long chain myosin light chain kinase (MLCK-210) in microvascular hyperpermeability during severe burns.

PubMed

Reynoso, Rashell; Perrin, Rachel M; Breslin, Jerome W; Daines, Dayle A; Watson, Katherine D; Watterson, D Martin; Wu, Mack H; Yuan, Sarah

2007-11-01

Microvascular leakage has been implicated in the pathogenesis of multiple organ dysfunction during trauma. Previous studies suggest the involvement of myosin light chain (MLC) phosphorylation-triggered endothelial contraction in the development of microvascular hyperpermeability. Myosin light chain kinase (MLCK) plays a key role in the control of MLC-phosphorylation status; thus, it is thought to modulate barrier function through its regulation of intracellular contractile machinery. The aim of this study was to further investigate the endothelial mechanism of MLC-dependent barrier injury in burns, focusing on the long isoform of MLCK (MLCK-210) that has recently been identified as the predominant isoform expressed in vascular endothelial cells. An MLCK-210 knockout mouse model was subjected to third-degree scald burn covering 25% total body surface area. The mesenteric microcirculation was observed using intravital microscopy, and the microvascular permeability was assessed by measuring the transvenular flux of fluorescein isothiocyanate-albumin. In a separate experiment, in vivo mesenteric hydraulic conductivity (Lp) was measured using the modified Landis technique. The injury caused a profound microvascular leakage, as indicated by a 2-fold increase in albumin flux and 4-fold increase in Lp at the early stages, which was associated with a high mortality within the 24-h period. Compared with wild-type control, the MLCK-210-deficient mice displayed a significantly improved survival with a greatly attenuated microvascular hyperpermeability response to albumin and fluid. These results provide direct evidence for a role of MLCK-210 in mediating burn-induced microvascular barrier injury and validate MLCK-210 as a potential therapeutic target in the treatment of burn edema.

Force control of endothelium permeability in mechanically stressed pulmonary micro-vascular endothelial cells.

PubMed

Wang, Bin; Caluch, Adam; Fodil, Redouane; Féréol, Sophie; Zadigue, Patricia; Pelle, Gabriel; Louis, Bruno; Isabey, Daniel

2012-01-01

Mechanical factors play a key role in the pathogenesis of Acute Respiratory Distress Syndrome (ARDS) and Ventilator-Induced Lung Injury (VILI) as contributing to alveolo-capillary barrier dysfunction. This study aims at elucidating the role of the cytoskeleton (CSK) and cell-matrix adhesion system in the stressed endothelium and more precisely in the loss of integrity of the endothelial barrier. We purposely develop a cellular model made of a monolayer of confluent Human Pulmonary Microvascular Endothelial Cells (HPMVECs) whose cytoskeleton (CSK) is directly exposed to sustained cyclic mechanical stress for 1 and 2 h. We used RGD-coated ferromagnetic beads and measured permeability before and after stress application. We find that endothelial permeability increases in the stressed endothelium, hence reflecting a loss of integrity. Structural and mechanical results suggest that this endothelial barrier alteration would be due to physically-founded discrepancies in latero-basal reinforcement of adhesion sites in response to the global increase in CSK stiffness or centripetal intracellular forces. Basal reinforcement of adhesion is presently evidenced by the marked redistribution of αvβ3 integrin with cluster formation in the stressed endothelium.

Pericyte-derived sphingosine 1-phosphate induces the expression of adhesion proteins and modulates the retinal endothelial cell barrier.

PubMed

McGuire, Paul G; Rangasamy, Sampathkumar; Maestas, Joann; Das, Arup

2011-12-01

The mechanisms that regulate the physical interaction of pericytes and endothelial cells and the effects of these interactions on interendothelial cell junctions are not well understood. We determined the extent to which vascular pericytes could regulate pericyte-endothelial adhesion and the consequences that this disruption might have on the function of the endothelial barrier. Human retinal microvascular endothelial cells were cocultured with pericytes, and the effect on the monolayer resistance of endothelial cells and expression of the cell junction molecules N-cadherin and VE-cadherin were measured. The molecules responsible for the effect of pericytes or pericyte-conditioned media on the endothelial resistance and cell junction molecules were further analyzed. Our results indicate that pericytes increase the barrier properties of endothelial cell monolayers. This barrier function is maintained through the secretion of pericyte-derived sphingosine 1-phosphate. Sphingosine 1-phosphate aids in maintenance of microvascular stability by upregulating the expression of N-cadherin and VE-cadherin, and downregulating the expression of angiopoietin 2. Under normal circumstances, the retinal vascular pericytes maintain pericyte-endothelial contacts and vascular barrier function through the secretion of sphingosine 1-phosphate. Alteration of pericyte-derived sphingosine 1-phosphate production may be an important mechanism in the development of diseases characterized by vascular dysfunction and increased permeability.

RECOMBINANT FACTOR XIII DIMINISHES MULTIPLE ORGAN DYSFUNCTION IN RATS CAUSED BY GUT ISCHEMIA-REPERFUSION INJURY

PubMed Central

Zaets, Sergey B.; Xu, Da-Zhong; Lu, Qi; Feketova, Eleonora; Berezina, Tamara L.; Gruda, Maryann; Malinina, Inga V.; Deitch, Edwin A.; Olsen, Eva H. N.

2010-01-01

Plasma factor XIII (FXIII) is responsible for stabilization of fibrin clot at the final stage of blood coagulation. Because FXIII has also been shown to modulate inflammation and endothelial permeability, we hypothesized that FXIII diminishes multiple organ dysfunction caused by gut I/R injury. A model of superior mesenteric artery occlusion (SMAO) was used to induce gut I/R injury. Rats were subjected to 45-min SMAO or sham SMAO and treated with recombinant human FXIII A2 subunit (rFXIII) or placebo at the beginning of the reperfusion period. Lung permeability, lung and gut myeloperoxidase activity, gut histology, neutrophil respiratory burst, and microvascular blood flow in the liver and muscles were measured after a 3-h reperfusion period. The effect of activated rFXIII on transendothelial resistance of human umbilical vein endothelial cells was tested in vitro. Superior mesenteric artery occlusion–induced lung permeability as well as lung and gut myeloperoxidase activity was significantly lower in rFXIII-treated versus untreated animals. Similarly, rFXIII-treated rats had lower neutrophil respiratory burst activity and ileal mucosal injury. Rats treated with rFXIII also had higher liver microvascular blood flow compared with the placebo group. Superior mesenteric artery occlusion did not cause FXIII consumption during the study period. In vitro, activated rFXIII caused a dose-dependent increase in human umbilical vein endothelial cell monolayer resistance to thrombin-induced injury. Thus, administration of rFXIII diminishes SMAO-induced multiple organ dysfunction in rats, presumably by preservation of endothelial barrier function and the limitation of polymorphonuclear leukocyte activation. PMID:18948851

Smooth muscle‐generated methylglyoxal impairs endothelial cell‐mediated vasodilatation of cerebral microvessels in type 1 diabetic rats

PubMed Central

Alomar, Fadhel; Singh, Jaipaul; Jang, Hee‐Seong; Rozanzki, George J; Shao, Chun Hong; Padanilam, Babu J; Mayhan, William G

2016-01-01

Background and Purpose Endothelial cell‐mediated vasodilatation of cerebral arterioles is impaired in individuals with Type 1 diabetes (T1D). This defect compromises haemodynamics and can lead to hypoxia, microbleeds, inflammation and exaggerated ischaemia‐reperfusion injuries. The molecular causes for dysregulation of cerebral microvascular endothelial cells (cECs) in T1D remains poorly defined. This study tests the hypothesis that cECs dysregulation in T1D is triggered by increased generation of the mitochondrial toxin, methylglyoxal, by smooth muscle cells in cerebral arterioles (cSMCs). Experimental Approach Endothelial cell‐mediated vasodilatation, vascular transcytosis inflammation, hypoxia and ischaemia‐reperfusion injury were assessed in brains of male Sprague‐Dawley rats with streptozotocin‐induced diabetes and compared with those in diabetic rats with increased expression of methylglyoxal‐degrading enzyme glyoxalase‐I (Glo‐I) in cSMCs. Key Results After 7–8 weeks of T1D, endothelial cell‐mediated vasodilatation of cerebral arterioles was impaired. Microvascular leakage, gliosis, macrophage/neutrophil infiltration, NF‐κB activity and TNF‐α levels were increased, and density of perfused microvessels was reduced. Transient occlusion of a mid‐cerebral artery exacerbated ischaemia‐reperfusion injury. In cSMCs, Glo‐I protein was decreased, and the methylglyoxal‐synthesizing enzyme, vascular adhesion protein 1 (VAP‐1) and methylglyoxal were increased. Restoring Glo‐I protein in cSMCs of diabetic rats to control levels via gene transfer, blunted VAP‐1 and methylglyoxal increases, cECs dysfunction, microvascular leakage, inflammation, ischaemia‐reperfusion injury and increased microvessel perfusion. Conclusions and Implications Methylglyoxal generated by cSMCs induced cECs dysfunction, inflammation, hypoxia and exaggerated ischaemia‐reperfusion injury in diabetic rats. Lowering methylglyoxal produced by cSMCs may be a viable therapeutic strategy to preserve cECs function and blunt deleterious downstream consequences in T1D. PMID:27611446

Pharmacokinetic-Pharmacodynamic Model for the Effect of l-Arginine on Endothelial Function in Patients with Moderately Severe Falciparum Malaria

PubMed Central

Brussee, Janneke M.; Yeo, Tsin W.; Lampah, Daniel A.; Anstey, Nicholas M.

2015-01-01

Impaired organ perfusion in severe falciparum malaria arises from microvascular sequestration of parasitized cells and endothelial dysfunction. Endothelial dysfunction in malaria is secondary to impaired nitric oxide (NO) bioavailability, in part due to decreased plasma concentrations of l-arginine, the substrate for endothelial cell NO synthase. We quantified the time course of the effects of adjunctive l-arginine treatment on endothelial function in 73 patients with moderately severe falciparum malaria derived from previous studies. Three groups of 10 different patients received 3 g, 6 g, or 12 g of l-arginine as a half-hour infusion. The remaining 43 received saline placebo. A pharmacokinetic-pharmacodynamic (PKPD) model was developed to describe the time course of changes in exhaled NO concentrations and reactive hyperemia-peripheral arterial tonometry (RH-PAT) index values describing endothelial function and then used to explore optimal dosing regimens for l-arginine. A PK model describing arginine concentrations in patients with moderately severe malaria was extended with two pharmacodynamic biomeasures, the intermediary biochemical step (NO production) and endothelial function (RH-PAT index). A linear model described the relationship between arginine concentrations and exhaled NO. NO concentrations were linearly related to RH-PAT index. Simulations of dosing schedules using this PKPD model predicted that the time within therapeutic range would increase with increasing arginine dose. However, simulations demonstrated that regimens of continuous infusion over longer periods would prolong the time within the therapeutic range even more. The optimal dosing regimen for l-arginine is likely to be administration schedule dependent. Further studies are necessary to characterize the effects of such continuous infusions of l-arginine on NO and microvascular reactivity in severe malaria. PMID:26482311

Pericyte Derived Sphinogosine 1-Phosphate Induces the Expression of Adhesion Proteins and Modulates the Retinal Endothelial Cell Barrier

PubMed Central

McGuire, P.G.; Rangasamy, S.; Maestas, J.; Das, A.

2011-01-01

Objective The mechanisms that regulate the physical interaction of pericytes and endothelial cells and the effects of these interactions on interendothelial cell junctions are not well understood. We determined the extent to which vascular pericytes could regulate pericyte-endothelial adhesion and the consequences that this disruption might have on the function of the endothelial barrier. Methods and Results Human retinal microvascular endothelial cells were co-cultured with pericytes, and the effect on the monolayer resistance of endothelial cells and expression of the cell junction molecules N-cadherin and VE-cadherin were measured. The molecules responsible for the effect of pericytes or pericyte conditioned media on the endothelial resistance and cell junction molecules were further analyzed. Our results indicate that pericytes increase the barrier properties of endothelial cell monolayers. This barrier function is maintained through the secretion of pericyte-derived sphingosine 1-phosphate (S1P). S1P aids in maintenance of microvascular stability by up-regulating the expression of N-cadherin and VE-cadherin, and down-regulating the expression of angiopoietin 2. Conclusion Under normal circumstances, the retinal vascular pericytes maintain pericyte-endothelial contacts and vascular barrier function through the secretion of S1P. Alteration of pericyte-derived S1P production may be an important mechanism in the development of diseases characterized by vascular dysfunction and increased permeability. PMID:21940944

Evaluation of Vitamin C for Adjuvant Sepsis Therapy

PubMed Central

2013-01-01

Abstract Significance: Evidence is emerging that parenteral administration of high-dose vitamin C may warrant development as an adjuvant therapy for patients with sepsis. Recent Advances: Sepsis increases risk of death and disability, but its treatment consists only of supportive therapies because no specific therapy is available. The characteristics of severe sepsis include ascorbate (reduced vitamin C) depletion, excessive protein nitration in microvascular endothelial cells, and microvascular dysfunction composed of refractive vasodilation, endothelial barrier dysfunction, and disseminated intravascular coagulation. Parenteral administration of ascorbate prevents or even reverses these pathological changes and thereby decreases hypotension, edema, multiorgan failure, and death in animal models of sepsis. Critical Issues: Dehydroascorbic acid appears to be as effective as ascorbate for protection against microvascular dysfunction, organ failure, and death when injected in sepsis models, but information about pharmacodynamics and safety in human subjects is only available for ascorbate. Although the plasma ascorbate concentration in critically ill and septic patients is normalized by repletion protocols that use high doses of parenteral ascorbate, and such doses are tolerated well by most healthy subjects, whether such large amounts of the vitamin trigger adverse effects in patients is uncertain. Future Directions: Further study of sepsis models may determine if high concentrations of ascorbate in interstitial fluid have pro-oxidant and bacteriostatic actions that also modify disease progression. However, the ascorbate depletion observed in septic patients receiving standard care and the therapeutic mechanisms established in models are sufficient evidence to support clinical trials of parenteral ascorbate as an adjuvant therapy for sepsis. Antioxid. Redox Signal. 19, 2129–2140. PMID:23682970

Hyperglycemia and Diabetes Downregulate the Functional Expression of TRPV4 Channels in Retinal Microvascular Endothelium

PubMed Central

Monaghan, Kevin; McNaughten, Jennifer; McGahon, Mary K.; Kelly, Catriona; Kyle, Daniel; Yong, Phaik Har

2015-01-01

Retinal endothelial cell dysfunction is believed to play a key role in the etiology and pathogenesis of diabetic retinopathy. Numerous studies have shown that TRPV4 channels are critically involved in maintaining normal endothelial cell function. In the current paper, we demonstrate that TRPV4 is functionally expressed in the endothelium of the retinal microcirculation and that both channel expression and activity is downregulated by hyperglycaemia. Quantitative PCR and immunostaining demonstrated molecular expression of TRPV4 in cultured bovine retinal microvascular endothelial cells (RMECs). Functional TRPV4 activity was assessed in cultured RMECs from endothelial Ca2+-responses recorded using fura-2 microfluorimetry and electrophysiological recordings of membrane currents. The TRPV4 agonist 4α-phorbol 12,13-didecanoate (4-αPDD) increased [Ca2+]i in RMECs and this response was largely abolished using siRNA targeted against TRPV4. These Ca2+-signals were completely inhibited by removal of extracellular Ca2+, confirming their dependence on influx of extracellular Ca2+. The 4-αPDD Ca2+-response recorded in the presence of cyclopiazonic acid (CPA), which depletes the intracellular stores preventing any signal amplification through store release, was used as a measure of Ca2+-influx across the cell membrane. This response was blocked by HC067047, a TRPV4 antagonist. Under voltage clamp conditions, the TRPV4 agonist GSK1016790A stimulated a membrane current, which was again inhibited by HC067047. Following incubation with 25mM D-glucose TRPV4 expression was reduced in comparison with RMECs cultured under control conditions, as were 4αPDD-induced Ca2+-responses in the presence of CPA and ion currents evoked by GSK1016790A. Molecular expression of TRPV4 in the retinal vascular endothelium of 3 months’ streptozotocin-induced diabetic rats was also reduced in comparison with that in age-matched controls. We conclude that hyperglycaemia and diabetes reduce the molecular and functional expression of TRPV4 channels in retinal microvascular endothelial cells. These changes may contribute to diabetes induced endothelial dysfunction and retinopathy. PMID:26047504

Omega-3 fatty acids attenuate constitutive and insulin-induced CD36 expression through a suppression of PPAR α/γ activity in microvascular endothelial cells.

PubMed

Madonna, Rosalinda; Salerni, Sara; Schiavone, Deborah; Glatz, Jan F; Geng, Yong-Jian; De Caterina, Raffaele

2011-09-01

Microvascular dysfunction occurs in insulin resistance and/or hyperinsulinaemia. Enhanced uptake of free fatty acids (FFA) and oxidised low-density lipoproteins (oxLDL) may lead to oxidative stress and microvascular dysfunction interacting with CD36, a PPARα/γ-regulated scavenger receptor and long-chain FFA transporter. We investigated CD36 expression and CD36-mediated oxLDL uptake before and after insulin treatment in human dermal microvascular endothelial cells (HMVECs), ± different types of fatty acids (FA), including palmitic, oleic, linoleic, arachidonic, eicosapentaenoic (EPA), and docosahexaenoic (DHA) acids. Insulin (10(-8) and 10(-7) M) time-dependently increased DiI-oxLDL uptake and CD36 surface expression (by 30 ± 13%, p<0.05 vs. untreated control after 24 hours incubation), as assessed by ELISA and flow cytometry, an effect that was potentiated by the PI3-kinase inhibitor wortmannin and reverted by the ERK1/2 inhibitor PD98059 and the PPARα/γ antagonist GW9662. A ≥ 24 hour exposure to 50 μM DHA or EPA, but not other FA, blunted both the constitutive (by 23 ± 3% and 29 ± 2%, respectively, p<0.05 for both) and insulin-induced CD36 expressions (by 45 ± 27 % and 12 ± 3 %, respectively, p<0.05 for both), along with insulin-induced uptake of DiI-oxLDL and the downregulation of phosphorylated endothelial nitric oxide synthase (P-eNOS). At gel shift assays, DHA reverted insulin-induced basal and oxLDL-stimulated transactivation of PPRE and DNA binding of PPARα/γ and NF-κB. In conclusion, omega-3 fatty acids blunt the increased CD36 expression and activity promoted by high concentrations of insulin. Such mechanisms may be the basis for the use of omega-3 fatty acids in diabetic microvasculopathy.

Venous gas emboli are involved in post-dive macro, but not microvascular dysfunction.

PubMed

Lambrechts, Kate; Balestra, Costantino; Theron, Michaël; Henckes, Anne; Galinat, Hubert; Mignant, Fanny; Belhomme, Marc; Pontier, Jean-Michel; Guerrero, François

2017-02-01

Previous studies have shown vascular dysfunction of main conductance arteries and microvessels after diving. We aim to evaluate the impact of bubble formation on vascular function and haemostasis. To achieve this, we used a vibration preconditioning to influence bubble levels without changing any other parameters linked to the dive. Twentty-six divers were randomly assigned to one of three groups: (1) the "vibrations-dive" group (VD; n = 9) was exposed to a whole-body vibration session 30 min prior the dive; (2) the "diving" group (D; n = 9) served as a control for the effect of the diving protocol; (3) The "vibration" protocol (V; n = 8) allowed us to assess the effect of vibrations without diving. Macro- and microvascular function was assessed for each subject before and after the dive, subsequently. Bubble grades were monitored with Doppler according to the Spencer grading system. Blood was taken before and after the protocol to assess any change of platelets or endothelial function. Bubble formation was lower in the VD than the diving group. The other measured parameters remained unchanged after the "vibration" protocol alone. Diving alone induced macrovascular dysfunction, and increased PMP and thrombin generation. Those parameters were no longer changed in the VD group. Conversely, a microvascular dysfunction persists despite a significant decrease of circulating bubbles. Finally, the results of this study suggest that macro- but not microvascular impairment results at least partly from bubbles, possibly related to platelet activation and generation of pro-coagulant microparticles.

Functional vascular contributions to cognitive impairment and dementia: mechanisms and consequences of cerebral autoregulatory dysfunction, endothelial impairment, and neurovascular uncoupling in aging

PubMed Central

Toth, Peter; Tarantini, Stefano; Csiszar, Anna

2017-01-01

Increasing evidence from epidemiological, clinical and experimental studies indicate that age-related cerebromicrovascular dysfunction and microcirculatory damage play critical roles in the pathogenesis of many types of dementia in the elderly, including Alzheimer’s disease. Understanding and targeting the age-related pathophysiological mechanisms that underlie vascular contributions to cognitive impairment and dementia (VCID) are expected to have a major role in preserving brain health in older individuals. Maintenance of cerebral perfusion, protecting the microcirculation from high pressure-induced damage and moment-to-moment adjustment of regional oxygen and nutrient supply to changes in demand are prerequisites for the prevention of cerebral ischemia and neuronal dysfunction. This overview discusses age-related alterations in three main regulatory paradigms involved in the regulation of cerebral blood flow (CBF): cerebral autoregulation/myogenic constriction, endothelium-dependent vasomotor function, and neurovascular coupling responses responsible for functional hyperemia. The pathophysiological consequences of cerebral microvascular dysregulation in aging are explored, including blood-brain barrier disruption, neuroinflammation, exacerbation of neurodegeneration, development of cerebral microhemorrhages, microvascular rarefaction, and ischemic neuronal dysfunction and damage. Due to the widespread attention that VCID has captured in recent years, the evidence for the causal role of cerebral microvascular dysregulation in cognitive decline is critically examined. PMID:27793855

Human microvascular dysfunction and apoptotic injury induced by AL amyloidosis light chain proteins.

PubMed

Migrino, Raymond Q; Truran, Seth; Gutterman, David D; Franco, Daniel A; Bright, Megan; Schlundt, Brittany; Timmons, Mitchell; Motta, Angelica; Phillips, Shane A; Hari, Parameswaran

2011-12-01

Light chain amyloidosis (AL) involves overproduction of amyloidogenic light chain proteins (LC) leading to heart failure, yet the mechanisms underlying tissue toxicity remain unknown. We hypothesized that LC induces endothelial dysfunction in non-AL human microvasculature and apoptotic injury in human coronary artery endothelial cells (HCAECs). Adipose arterioles (n = 34, 50 ± 3 yr) and atrial coronary arterioles (n = 19, 68 ± 2 yr) from non-AL subjects were cannulated. Adipose arteriole dilator responses to acetylcholine/papaverine were measured at baseline and 1 h exposure to LC (20 μg/ml) from biopsy-proven AL subjects (57 ± 11 yr) without and with antioxidant cotreatment. Coronary arteriole dilation to bradykinin/papaverine was measured post-LC exposure. HCAECs were exposed to 1 or 24 h of LC. LC reduced dilation to acetylcholine (10(-4) M: 41.6 ± 7 vs. 85.8 ± 2.2% control, P < 0.001) and papaverine (81.4 ± 4.6 vs. 94.8 ± 1.3% control, P < 0.01) in adipose arterioles and to bradykinin (10(-6) M: 68.6 ± 6.2 vs. 90.9 ± 1.6% control, P < 0.001) but not papaverine in coronary arterioles. There was an increase in superoxide and peroxynitrite in arterioles treated with LC. Adipose arteriole dilation was restored by cotreatment with polyethylene glycol-superoxide dismutase and tetrahydrobiopterin but only partially restored by mitoquinone (mitochondria-targeted antioxidant) and gp91ds-tat (NADPH oxidase inhibitor). HCAECs exposed to LC showed reduced NO and increased superoxide, peroxynitrite, annexin-V, and propidium iodide compared with control. Brief exposure to physiological amounts of LC induced endothelial dysfunction in human adipose and coronary arterioles and increased apoptotic injury in coronary artery endothelial cells likely as a result of oxidative stress, reduced NO bioavailability, and peroxynitrite production. Microvascular dysfunction and injury is a novel mechanism underlying AL pathobiology and is a potential target for therapy.

Publisher Correction: Eternal blood vessels

NASA Astrophysics Data System (ADS)

Hindson, Jordan

2018-05-01

This article was originally published with an incorrect reference for the original article. The reference has been amended. Please see the correct reference below. Qiu, Y. et al. Microvasculature-on-a-chip for the long-term study of endothelial barrier dysfunction and microvascular obstruction in disease. Nat. Biomed. Eng. https://doi.org/10.1038/s41551-018-0224-z (2018)

Assessment of macrovascular endothelial function using pulse wave analysis and its association with microvascular reactivity in healthy subjects.

PubMed

Ibrahim, N N I N; Rasool, A H G

2017-08-01

Pulse wave analysis (PWA) and laser Doppler fluximetry (LDF) are non-invasive methods of assessing macrovascular endothelial function and microvascular reactivity respectively. The aim of this study was to assess the correlation between macrovascular endothelial function assessed by PWA and microvascular reactivity assessed by LDF. 297 healthy and non-smoking subjects (159 females, mean age (±SD) 23.56 ± 4.54 years) underwent microvascular reactivity assessment using LDF followed by macrovascular endothelial function assessments using PWA. Pearson's correlation showed no correlation between macrovascular endothelial function and microvascular reactivity (r = -0.10, P = 0.12). There was no significant correlation between macrovascular endothelial function assessed by PWA and microvascular reactivity assessed by LDF in healthy subjects. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Dobesilate enhances endothelial nitric oxide synthase-activity in macro- and microvascular endothelial cells

PubMed Central

Suschek, Christoph; Kolb, Hubert; Kolb-Bachofen, Victoria

1997-01-01

Dobesilate is used for normalizing vascular dysfunction in a number of diseases. In search for an effect on endothelial NO production, macrovascular endothelial cells from rat aorta, microvascular endothelial cells from rat exocrine pancreatic tissue, and capillary endothelial cells from rat islets, were cultured in the presence or absence of Mg-Dobesilate. The activity of constitutive nitric oxide synthase (ecNOS) in resident cells as well as of inducible nitric oxide synthase (iNOS) in cytokine-activated cells was measured indirectly by recording the citrulline concentrations in culture supernatants.In each of the different endothelial cells Mg-Dobesilate incubation (0.25–1 mM) for 24 h led to a significant and concentration-dependent increase in ecNOS-activities. With cytokine-activated endothelial cell cultures only moderate effects were seen with little or no concentration-dependency. Addition of the NOS-inhibitor NG-monomethyl-L-arginine led to a significant suppression of citrulline formation in all cultures as an evidence for the enzyme specificity of these effects.iNOS- and ecNOS-specific reverse transcription and semi-quantitative polymerase chain reaction (RT–PCR) with RNA from resident or cytokine-activated endothelial cells gave no evidence for an increase in NOS-specific mRNA after Mg-Dobesilate-treatment. Furthermore, Dobesilate-mediated enhancement of NO synthesis in resting endothelial cells was not due to iNOS induction in these cells, as no iNOS-specific signal was found by RT–PCR. PMID:9421302

Globular adiponectin ameliorates metabolic insulin resistance via AMPK-mediated restoration of microvascular insulin responses

PubMed Central

Zhao, Lina; Fu, Zhuo; Wu, Jing; Aylor, Kevin W; Barrett, Eugene J; Cao, Wenhong; Liu, Zhenqi

2015-01-01

Abstract Hypoadiponectinaemia is closely associated with endothelial dysfunction and insulin resistance, and microvasculature plays a critical role in the regulation of insulin action in muscle. Here we tested whether adiponectin replenishment could improve metabolic insulin sensitivity in male rats fed a high-fat diet (HFD) via the modulation of microvascular insulin responses. Male Sprague–Dawley rats were fed either a HFD or low-fat diet (LFD) for 4 weeks. Small resistance artery myograph changes in tension, muscle microvascular recruitment and metabolic response to insulin were determined. Compared with rats fed a LFD, HFD feeding abolished the vasodilatory actions of globular adiponectin (gAd) and insulin on pre-constricted distal saphenous arteries. Pretreatment with gAd improved insulin responses in arterioles isolated from HFD rats, which was blocked by AMP-activated protein kinase (AMPK) inhibition. Similarly, HFD abolished microvascular responses to either gAd or insulin and decreased insulin-stimulated glucose disposal by ∼60%. However, supplementing gAd fully rescued insulin’s microvascular action and significantly improved the metabolic responses to insulin in HFD male rats and these actions were abolished by inhibition of either AMPK or nitric oxide production. We conclude that HFD induces vascular adiponectin and insulin resistance but gAd administration can restore vascular insulin responses and improve insulin’s metabolic action via an AMPK- and nitric oxide-dependent mechanism in male rats. Key points Adiponectin is an adipokine with anti-inflammatory and anti-diabetic properties. Hypoadiponectinaemia is closely associated with endothelial dysfunction and insulin resistance in obesity and diabetes. Insulin resistance is present in muscle microvasculature and this may contribute to decreased insulin delivery to, and action in, muscle. In this study we examined whether adiponectin ameliorates metabolic insulin resistance by affecting muscle microvascular recruitment. We demonstrated that a high-fat diet induces vascular adiponectin and insulin resistance but globular adiponectin administration can restore vascular insulin responses and improve insulin’s metabolic action via an AMPK- and nitric oxide-dependent mechanism. This suggests that globular adiponectin might have a therapeutic potential for improving insulin resistance and preventing cardiovascular complications in patients with diabetes via modulation of microvascular insulin responses. PMID:26108677

Angiotensin II receptor blockade or deletion of vascular endothelial ACE does not prevent vascular dysfunction and remodeling in 20-HETE-dependent hypertension.

PubMed

Garcia, Victor; Joseph, Gregory; Shkolnik, Brian; Ding, Yan; Zhang, Frank Fan; Gotlinger, Katherine; Falck, John R; Dakarapu, Rambabu; Capdevila, Jorge H; Bernstein, Kenneth E; Schwartzman, Michal Laniado

2015-07-01

Increased vascular 20-HETE is associated with hypertension and activation of the renin-angiotensin system (RAS) through induction of vascular angiotensin-converting enzyme (ACE) expression. Cyp4a12tg mice, whose Cyp4a12-20-HETE synthase expression is under the control of a tetracycline (doxycycline, DOX) promoter, were used to assess the contribution of ACE/RAS to microvascular remodeling in 20-HETE-dependent hypertension. Treatment of Cyp4a12tg mice with DOX increased systolic blood pressure (SBP; 136 ± 2 vs. 102 ± 1 mmHg; P < 0.05), and this increase was prevented by administration of 20-HEDGE, lisinopril, or losartan. DOX-induced hypertension was associated with microvascular dysfunction and remodeling of preglomerular microvessels, which was prevented by 20-HEDGE, a 20-HETE antagonist, yet only lessened, but not prevented, by lisinopril or losartan. In ACE 3/3 mice, which lack vascular endothelial ACE, administration of 5α-dihydrotestosterone (DHT), a known inducer of 20-HETE production, increased SBP; however, the increase was about 50% of that in wild-type (WT) mice (151 ± 1 vs. 126 ± 1 mmHg). Losartan and 20-HEDGE prevented the DHT-induced increase in SBP in WT and ACE 3/3 mice. DHT treatment increased 20-HETE production and microvascular remodeling in WT and ACE 3/3 mice; however, remodeling was attenuated in the ACE 3/3 mice as opposed to WT mice (15.83 ± 1.11 vs. 22.17 ± 0.92 μm; P < 0.05). 20-HEDGE prevented microvascular remodeling in WT and ACE 3/3 mice, while losartan had no effect on microvascular remodeling in ACE 3/3. Taken together, these results suggest that RAS contributes to 20-HETE-mediated microvascular remodeling in hypertension and that 20-HETE-driven microvascular remodeling independent of blood pressure elevation does not fully rely on ACE activity in the vascular endothelium. Copyright © 2015 the American Physiological Society.

Diabetes mellitus: The linkage between oxidative stress, inflammation, hypercoagulability and vascular complications.

PubMed

Domingueti, Caroline Pereira; Dusse, Luci Maria Sant'Ana; Carvalho, Maria das Graças; de Sousa, Lirlândia Pires; Gomes, Karina Braga; Fernandes, Ana Paula

2016-01-01

Vascular complications are the leading cause of morbidity and mortality among patients with type 1 and type 2 diabetes mellitus. These vascular abnormalities result of a chronic hyperglycemic state, which leads to an increase in oxidative stress and inflammatory responses. This review addresses the relationships among endothelial dysfunction, hypercoagulability and inflammation and their biomarkers in the development of vascular complications in type 1 and type 2 diabetes. Inflammation, endothelial dysfunction, and hypercoagulability are correlated to each other, playing an important role in the development of vascular complications in diabetic patients. Moreover, it has been observed that several endothelial, inflammatory and pro-coagulant biomarkers, such as VWF, IL-6, TNF-α, D-dimer and PAI-1, are increased in diabetic patients who have microvascular and macrovascular complications, including nephropathy or cardiovascular disease. It is promising the clinical and laboratory use of endothelial, inflammatory and pro-coagulant biomarkers for predicting the risk of cardiovascular and renal complications in diabetic patients and for monitoring these patients. Copyright © 2016 Elsevier Inc. All rights reserved.

Inflammation-induced microvascular insulin resistance is an early event in diet-induced obesity.

PubMed

Zhao, Lina; Fu, Zhuo; Wu, Jing; Aylor, Kevin W; Barrett, Eugene J; Cao, Wenhong; Liu, Zhenqi

2015-12-01

Endothelial dysfunction and vascular insulin resistance usually coexist and chronic inflammation engenders both. In the present study, we investigate the temporal relationship between vascular insulin resistance and metabolic insulin resistance. We assessed insulin responses in all arterial segments, including aorta, distal saphenous artery and the microvasculature, as well as the metabolic insulin responses in muscle in rats fed on a high-fat diet (HFD) for various durations ranging from 3 days to 4 weeks with or without sodium salicylate treatment. Compared with controls, HFD feeding significantly blunted insulin-mediated Akt (protein kinase B) and eNOS [endothelial nitric oxide (NO) synthase] phosphorylation in aorta in 1 week, blunted vasodilatory response in small resistance vessel in 4 weeks and microvascular recruitment in as early as 3 days. Insulin-stimulated whole body glucose disposal did not begin to progressively decrease until after 1 week. Salicylate treatment fully inhibited vascular inflammation, prevented microvascular insulin resistance and significantly improved muscle metabolic responses to insulin. We conclude that microvascular insulin resistance is an early event in diet-induced obesity and insulin resistance and inflammation plays an essential role in this process. Our data suggest microvascular insulin resistance contributes to the development of metabolic insulin resistance in muscle and muscle microvasculature is a potential therapeutic target in the prevention and treatment of diabetes and its related complications. © 2015 Authors; published by Portland Press Limited.

Inflammation-induced microvascular insulin resistance is an early event in diet-induced obesity

PubMed Central

Zhao, Lina; Fu, Zhuo; Wu, Jing; Aylor, Kevin W.; Barrett, Eugene J.; Cao, Wenhong

2015-01-01

Endothelial dysfunction and vascular insulin resistance usually coexist and chronic inflammation engenders both. In the present study, we investigate the temporal relationship between vascular insulin resistance and metabolic insulin resistance. We assessed insulin responses in all arterial segments, including aorta, distal saphenous artery and the microvasculature, as well as the metabolic insulin responses in muscle in rats fed on a high-fat diet (HFD) for various durations ranging from 3 days to 4 weeks with or without sodium salicylate treatment. Compared with controls, HFD feeding significantly blunted insulin-mediated Akt (protein kinase B) and eNOS [endothelial nitric oxide (NO) synthase] phosphorylation in aorta in 1 week, blunted vasodilatory response in small resistance vessel in 4 weeks and microvascular recruitment in as early as 3 days. Insulin-stimulated whole body glucose disposal did not begin to progressively decrease until after 1 week. Salicylate treatment fully inhibited vascular inflammation, prevented microvascular insulin resistance and significantly improved muscle metabolic responses to insulin. We conclude that microvascular insulin resistance is an early event in diet-induced obesity and insulin resistance and inflammation plays an essential role in this process. Our data suggest microvascular insulin resistance contributes to the development of metabolic insulin resistance in muscle and muscle microvasculature is a potential therapeutic target in the prevention and treatment of diabetes and its related complications. PMID:26265791

What can we learn about treating heart failure from the heart's response to acute exercise? Focus on the coronary microcirculation.

PubMed

Heinonen, Ilkka; Sorop, Oana; de Beer, Vincent J; Duncker, Dirk J; Merkus, Daphne

2015-10-15

Coronary microvascular function and cardiac function are closely related in that proper cardiac function requires adequate oxygen delivery through the coronary microvasculature. Because of the close proximity of cardiomyocytes and coronary microvascular endothelium, cardiomyocytes not only communicate their metabolic needs to the coronary microvasculature, but endothelium-derived factors also directly modulate cardiac function. This review summarizes evidence that the myocardial oxygen balance is disturbed in the failing heart because of increased extravascular compressive forces and coronary microvascular dysfunction. The perturbations in myocardial oxygen balance are exaggerated during exercise and are due to alterations in neurohumoral influences, endothelial function, and oxidative stress. Although there is some evidence from animal studies that the myocardial oxygen balance can partly be restored by exercise training, it is largely unknown to what extent the beneficial effects of exercise training include improvements in endothelial function and/or oxidative stress in the coronary microvasculature and how these improvements are impacted by risk factors such as diabetes, obesity, and hypercholesterolemia. Copyright © 2015 the American Physiological Society.

Phloretin ameliorates 2-chlorohexadecanal-mediated brain microvascular endothelial cell dysfunction in vitro

PubMed Central

Üllen, Andreas; Fauler, Günter; Bernhart, Eva; Nusshold, Christoph; Reicher, Helga; Leis, Hans-Jörg; Malle, Ernst; Sattler, Wolfgang

2012-01-01

2-Chlorohexadecanal (2-ClHDA), a chlorinated fatty aldehyde, is formed via attack on ether-phospholipids by hypochlorous acid (HOCl) that is generated by the myeloperoxidase–hydrogen peroxide–chloride system of activated leukocytes. 2-ClHDA levels are elevated in atherosclerotic lesions, myocardial infarction, and neuroinflammation. Neuroinflammatory conditions are accompanied by accumulation of neutrophils (an ample source of myeloperoxidase) in the brain. Microvessel damage by inflammatory mediators and/or reactive oxidants can induce blood–brain barrier (BBB) dysfunction, a pathological condition leading to cerebral edema, brain hemorrhage, and neuronal death. In this in vitro study we investigated the impact of 2-ClHDA on brain microvascular endothelial cells (BMVEC), which constitute the morphological basis of the BBB. We show that exogenously added 2-ClHDA is subject to rapid uptake and metabolism by BMVEC. Using C16 structural analogues of 2-ClHDA we found that the cytotoxic potential decreases in the following order: 2-ClHDA>hexadecanal>palmitic acid>2-ClHDA-dimethylacetal. 2-ClHDA induces loss of barrier function, mitochondrial dysfunction, apoptosis via activation of caspase 3, and altered intracellular redox balance. Finally we investigated potential protective effects of several natural polyphenols on in vitro BBB function. Of the compounds tested, phloretin almost completely abrogated 2-ClHDA-induced BMVEC barrier dysfunction and cell death. These data suggest that 2-ClHDA has the potential to induce BBB breakdown under inflammatory conditions and that phloretin confers protection in this experimental setting. PMID:22982051

Microvascular function in pre-eclampsia is influenced by insulin resistance and an imbalance of angiogenic mediators.

PubMed

Ghosh, Anshuman; Freestone, Nicholas S; Anim-Nyame, Nicholas; Arrigoni, Francesca I F

2017-04-01

In preeclampsia, maternal microvascular function is disrupted and angiogenesis is dysfunctional. Insulin resistance that occurs in some pregnancies also pathologically affects microvascular function. We wished to examine the relationship of angiogenic mediators and insulin resistance on microvascular health in pregnancy. We performed a nested, case-control study of 16 women who developed preeclampsia with 17 normal pregnant controls. We hypothesized that the impaired microvascular blood flow in preeclamptic women associated with an increased ratio of the antiangiogenic factors; (s-endoglin [sEng] and soluble fms-like tyrosine kinase-1 [sFlt-1]) and proangiogenic molecule (placental growth factor [PlGF]) could be influenced by insulin resistance. Serum samples taken after 28 weeks of gestation were measured for the angiogenic factors, insulin, and glucose alongside the inflammatory marker; tumor necrosis factor-α and endothelial activation, namely; soluble vascular cell adhesion molecule 1, intercellular adhesion molecule-1, and e-selectin. Maternal microvascular blood flow, measured by strain gauge plethysmography, correlated with ratios of pro- and antiangiogenic mediators independently of preeclampsia. Decreased microvascular function measured in preeclampsia strongly correlated with both the antiangiogenic factor (sFlt-1 + sEng): PlGF ratio and high levels of insulin resistance, and combining insulin resistance with antiangiogenic factor ratios further strengthened this relationship. In pregnancy, microvascular blood flow is strongly associated with perturbations in pro- and antiangiogenic mediators. In preeclampsia, the relationship of maternal microvascular dysfunction with antiangiogenic mediators is strengthened when combined with insulin resistance. © 2017 Kingston University. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.

Cyclooxygenase inhibition improves endothelial vasomotor dysfunction of visceral adipose arterioles in human obesity

PubMed Central

Farb, Melissa G.; Tiwari, Stephanie; Karki, Shakun; Ngo, Doan TM; Carmine, Brian; Hess, Donald T.; Zuriaga, Maria A.; Walsh, Kenneth; Fetterman, Jessica L.; Hamburg, Naomi M.; Vita, Joseph A.; Apovian, Caroline M.; Gokce, Noyan

2013-01-01

Objective The purpose of this study was to determine whether cyclooxygenase inhibition improves vascular dysfunction of adipose microvessels from obese humans. Design and Methods In 20 obese subjects (age 37±12 yrs, BMI 47±8 kg/m2) we collected subcutaneous and visceral fat during bariatric surgery and characterized adipose depot-specific gene expression, endothelial cell phenotype, and microvascular function. Vasomotor function was assessed in response to endothelium-dependent agonists using videomicroscopy of small arterioles from fat. Results Arterioles from visceral fat exhibited impaired endothelium-dependent, acetylcholine-mediated vasodilation, compared to the subcutaneous depot (p<0.001). Expression of mRNA transcripts relevant to the cyclooxygenase pathway were upregulated in visceral compared to subcutaneous fat. Pharmacological inhibition of cyclooxygenase with indomethacin improved endothelium-dependent vasodilator function of arterioles from visceral fat by 2-fold (p=0.01), whereas indomethacin had no effect in the subcutaneous depot. Indomethacin increased activation via serine-1177 phosphorylation of endothelial nitric oxide synthase in response to acetylcholine in endothelial cells from visceral fat. Inhibition of endothelial nitric oxide synthase with Nω-nitro-L-arginine methyl ester abrogated the effects of cyclooxygenase-inhibition suggesting that vascular actions of indomethacin were related to increased nitric oxide bioavailability. Conclusions Our findings suggest that cyclooxygenase-mediated vasoconstrictor prostanoids partly contribute to endothelial dysfunction of visceral adipose arterioles in human obesity. PMID:23640904

Habitual aerobic exercise does not protect against micro- or macrovascular endothelial dysfunction in healthy estrogen-deficient postmenopausal women.

PubMed

Santos-Parker, Jessica R; Strahler, Talia R; Vorwald, Victoria M; Pierce, Gary L; Seals, Douglas R

2017-01-01

Aging causes micro- and macrovascular endothelial dysfunction, as assessed by endothelium-dependent dilation (EDD), which can be prevented and reversed by habitual aerobic exercise (AE) in men. However, in estrogen-deficient postmenopausal women, whole forearm microvascular EDD has not been studied, and a beneficial effect of AE on macrovascular EDD has not been consistently shown. We assessed forearm blood flow in response to brachial artery infusions of acetylcholine (FBF ACh ), a measure of whole forearm microvascular EDD, and brachial artery flow-mediated dilation (FMD), a measure of macrovascular EDD, in 12 premenopausal sedentary women (Pre-S; 24 ± 1 yr; V̇o 2max = 37.5 ± 1.6 ml·kg -1 ·min -1 ), 25 estrogen-deficient postmenopausal sedentary women (Post-S; 62 ± 1 yr; V̇o 2max = 24.7 ± 0.9 ml·kg -1 ·min -1 ), and 16 estrogen-deficient postmenopausal AE-trained women (Post-AE; 59 ± 1 yr; V̇o 2max = 40.4 ± 1.4 ml·kg -1 ·min -1 ). FBF ACh was lower in Post-S and Post-AE compared with Pre-S women (135 ± 9 and 116 ± 17 vs. 193 ± 21 AUC, respectively, both P < 0.008), whereas Post-S and Post-AE women were not different (P = 0.3). Brachial artery FMD was 34% (5.73 ± 0.67%) and 45% (4.79 ± 0.57%) lower in Post-S and Post-AE, respectively, vs. Pre-S women (8.69 ± 0.95%, both P ≤ 0.01), but not different between Post-S and Post-AE women (P = 0.3). Post-AE women had lower circulating C-reactive protein and oxidized low-density lipoprotein compared with Post-S women (0.5 ± 0.1 vs. 1.1 ± 0.2 mg/l and 40 ± 4 vs. 55 ± 3 U/l, respectively, both P = 0.01), but these markers were not correlated to FBF ACh (P = 0.3) or brachial artery FMD (P = 0.8). These findings are consistent with the idea that habitual AE does not protect against age/menopause-related whole forearm micro- and macrovascular endothelial dysfunction in healthy nonobese estrogen-deficient postmenopausal women, despite being associated with lower systemic markers of inflammation and oxidative stress. This is the first study to demonstrate that habitual aerobic exercise may not protect against age/menopause-related whole forearm microvascular endothelial dysfunction in healthy nonobese estrogen-deficient postmenopausal women, consistent with recent findings regarding macrovascular endothelial function. This is in contrast to what is observed in healthy middle-aged and older aerobic exercise-trained men. Copyright © 2017 the American Physiological Society.

Specific role of impaired glucose metabolism and diabetes mellitus in endothelial progenitor cell characteristics and function.

PubMed

Yiu, Kai-Hang; Tse, Hung-Fat

2014-06-01

The disease burden of diabetes mellitus (DM) and its associated cardiovascular complications represent a growing and major global health problem. Recent studies suggest that circulating exogenous endothelial progenitor cells (EPCs) play an important role in endothelial repair and neovascularization at sites of injury or ischemia. Both experimental and clinical studies have demonstrated that hyperglycemia related to DM can induce alterations to EPCs. The reduction and dysfunction of EPCs related to DM correlate with the occurrence and severity of microvascular and macrovascular complications, suggesting a close mechanistic link between EPC dysfunction and impaired vascular function/repair in DM. These alterations to EPCs, likely mediated by multiple pathophysiological mechanisms, including inflammation, oxidative stress, and alterations in Akt and the nitric oxide pathway, affect EPCs at multiple stages: differentiation and mobilization in the bone marrow, trafficking and survival in the circulation, and homing and neovascularization. Several different therapeutic approaches have consequently been proposed to reverse the reduction and dysfunction of EPCs in DM and may represent a novel therapeutic approach to prevent and treat DM-related cardiovascular complications. © 2014 American Heart Association, Inc.

Ascorbate protects endothelial barrier function during septic insult: Role of protein phosphatase type 2A.

PubMed

Han, Min; Pendem, Suresh; Teh, Suet Ling; Sukumaran, Dinesh K; Wu, Feng; Wilson, John X

2010-01-01

Endothelial barrier dysfunction contributes to morbidity in sepsis. We tested the hypothesis that raising the intracellular ascorbate concentration protects the endothelial barrier from septic insult by inhibiting protein phosphatase type 2A. Monolayer cultures of microvascular endothelial cells were incubated with ascorbate, dehydroascorbic acid (DHAA), the NADPH oxidase inhibitors apocynin and diphenyliodonium, or the PP2A inhibitor okadaic acid and then were exposed to septic insult (lipopolysaccharide and interferon-gamma). Under standard culture conditions that depleted intracellular ascorbate, septic insult stimulated oxidant production and PP2A activity, dephosphorylated phosphoserine and phosphothreonine residues in the tight junction-associated protein occludin, decreased the abundance of occludin at cell borders, and increased monolayer permeability to albumin. NADPH oxidase inhibitors prevented PP2A activation and monolayer leak, showing that these changes required reactive oxygen species. Okadaic acid, at a concentration that inhibited PP2A activity and monolayer leak, prevented occludin dephosphorylation and redistribution, implicating PP2A in the response of occludin to septic insult. Incubation with ascorbate or DHAA raised intracellular ascorbate concentrations and mitigated the effects of septic insult. In conclusion, ascorbate acts within microvascular endothelial cells to inhibit septic stimulation of oxidant production by NADPH oxidase and thereby prevents PP2A activation, PP2A-dependent dephosphorylation and redistribution of occludin, and disruption of the endothelial barrier. Copyright 2009 Elsevier Inc. All rights reserved.

FTY720 Protects Cardiac Microvessels of Diabetes: A Critical Role of S1P1/3 in Diabetic Heart Disease

PubMed Central

Wei, Liping; Gao, Haokao; Zhang, Rongqing; Tao, Ling; Cao, Feng; Wang, Haichang

2012-01-01

Background: Diabetes is associated with an increased risk of cardiac microvascular disease. The mechanisms by which this damage occurs are unknown. However, research suggests that signaling through the sphingosine-1-phosphates receptor 1 and 3 (S1P1/3) by FTY720, a sphiongolipid drug that is structually similar to SIP, may play a role in the treatment on cardiac microvascular dysfunction in diabetes. We hypothesized that FTY720 might exert the cardioprotective effects of S1P1 and S1P3 viaprotein kinase C-beta (PKCβ II) signaling pathway. Methodology/Principal Findings: Transthoracic echocardiography was performed to detect the change of cardiac function. Scanning and transmission electron microscope with lanthanum tracer were used to determine microvascular ultrastructure and permeability in vivo. Apoptosis was detected by TUNEL and CD31 dual labeling in paraffin-embedded sections. Laser capture miscrodissection was used to assess cardiac micovascular endothelial cells (CMECs) in vivo. RT-PCR and Western blot analysis were used to determine the mRNA levels and protein expression of S1P1, S1P3, and PKCβ II. In the diabetic rats vs. controls, cardiac capillaries showed significantly higher density; CD31 positive endothelial cells were significantly reduced; the apoptosis index of cardiac endothlial cells was significantly higher. And FTY720 could increase the expressional level of S1P1 and boost S1P3 trasnslocation from membrane to nuclear, then ameliorate cardiac microvascular barrier impairment and pathologic angiogenesis induced by diabetes. In addition, overexpression of PKCβ II significantly decreased the protective effect of FTY720. Conclusions: Our study represents that the deregulation of S1P1 and S1P3 is an important signalresponsible for cardiac microvascular dysfunction in diabetes. FTY720 might be competent to serve as a potential therapeutic approach for diabetic heart disease through ameliorating cardiac microvascular barrier impairment and pathologic angiogenesis, which might be partly dependent on PKCβII-mediated signaling pathway. PMID:22916176

Dietary sodium restriction reverses vascular endothelial dysfunction in middle-aged/older adults with moderately elevated systolic blood pressure

PubMed Central

Jablonski, Kristen L.; Racine, Matthew L.; Geolfos, Candace J.; Gates, Phillip E.; Chonchol, Michel; McQueen, Matthew B.; Seals, Douglas R.

2013-01-01

Objectives We determined the efficacy of dietary sodium restriction (DSR) for improving vascular endothelial dysfunction in middle-aged/older adults with moderately elevated systolic blood pressure (SBP; 130–159 mmHg) and the associated physiological mechanisms. Background Vascular endothelial dysfunction develops with advancing age and elevated SBP, contributing to increased cardiovascular risk. DSR lowers BP, but its effect on vascular endothelial function and mechanisms involved are unknown. Methods Seventeen subjects (11M/6F; 62±7 yrs, mean±S.D.) completed a randomized, crossover study of 4 weeks of both low and normal sodium intake. Vascular endothelial function (endothelium-dependent dilation; EDD), nitric oxide (NO)/tetrahydrobiopterin (BH4) bioavailability and oxidative stress-associated mechanisms were assessed following each condition. Results Urinary sodium excretion was reduced by ~50% (to 70±30 mmol/day), and conduit (brachial artery flow-mediated dilation [FMDBA]) and resistance (forearm blood flow responses to acetylcholine [FBFACh]) artery EDD were 68% and 42% (peak FBFACh) higher following the low sodium diet (p<0.005). Low sodium markedly enhanced NO- mediated EDD (greater ΔFBFACh with endothelial NO synthase [eNOS] inhibition) without changing eNOS expression/activation (Ser1177 phosphorylation), restored BH4 bioactivity (less ΔFMDBA with acute BH4), abolished tonic superoxide suppression of EDD (less ΔFMDBA and ΔFBFACh with ascorbic acid infusion), and increased circulating superoxide dismutase activity (p<0.05). These effects were independent of ΔSBP. Other subject characteristics/dietary factors and endothelium-independent dilation were unchanged. Conclusions DSR largely reverses both macro- and microvascular endothelial dysfunction by enhancing NO and BH4 bioavailability and reducing oxidative stress. Our findings support the emerging concept that DSR induces “vascular protection” beyond that attributable to its BP-lowering effects. PMID:23141486

Phloretin ameliorates 2-chlorohexadecanal-mediated brain microvascular endothelial cell dysfunction in vitro.

PubMed

Ullen, Andreas; Fauler, Günter; Bernhart, Eva; Nusshold, Christoph; Reicher, Helga; Leis, Hans-Jörg; Malle, Ernst; Sattler, Wolfgang

2012-11-01

2-Chlorohexadecanal (2-ClHDA), a chlorinated fatty aldehyde, is formed via attack on ether-phospholipids by hypochlorous acid (HOCl) that is generated by the myeloperoxidase-hydrogen peroxide-chloride system of activated leukocytes. 2-ClHDA levels are elevated in atherosclerotic lesions, myocardial infarction, and neuroinflammation. Neuroinflammatory conditions are accompanied by accumulation of neutrophils (an ample source of myeloperoxidase) in the brain. Microvessel damage by inflammatory mediators and/or reactive oxidants can induce blood-brain barrier (BBB) dysfunction, a pathological condition leading to cerebral edema, brain hemorrhage, and neuronal death. In this in vitro study we investigated the impact of 2-ClHDA on brain microvascular endothelial cells (BMVEC), which constitute the morphological basis of the BBB. We show that exogenously added 2-ClHDA is subject to rapid uptake and metabolism by BMVEC. Using C16 structural analogues of 2-ClHDA we found that the cytotoxic potential decreases in the following order: 2-ClHDA>hexadecanal>palmitic acid>2-ClHDA-dimethylacetal. 2-ClHDA induces loss of barrier function, mitochondrial dysfunction, apoptosis via activation of caspase 3, and altered intracellular redox balance. Finally we investigated potential protective effects of several natural polyphenols on in vitro BBB function. Of the compounds tested, phloretin almost completely abrogated 2-ClHDA-induced BMVEC barrier dysfunction and cell death. These data suggest that 2-ClHDA has the potential to induce BBB breakdown under inflammatory conditions and that phloretin confers protection in this experimental setting. Copyright © 2012 Elsevier Inc. All rights reserved.

Anesthetic propofol overdose causes endothelial cytotoxicity in vitro and endothelial barrier dysfunction in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Ming-Chung; Department of Anesthesiology, Chi Mei Medical Center, Liouying, Tainan, Taiwan; Chen, Chia-Ling

An overdose and a prolonged treatment of propofol may cause cellular cytotoxicity in multiple organs and tissues such as brain, heart, kidney, skeletal muscle, and immune cells; however, the underlying mechanism remains undocumented, particularly in vascular endothelial cells. Our previous studies showed that the activation of glycogen synthase kinase (GSK)-3 is pro-apoptotic in phagocytes during overdose of propofol treatment. Regarding the intravascular administration of propofol, we therefore hypothesized that propofol overdose also induces endothelial cytotoxicity via GSK-3. Propofol overdose (100 μg/ml) inhibited growth in human arterial and microvascular endothelial cells. After treatment, most of the endothelial cells experienced caspase-independent necrosis-likemore » cell death. The activation of cathepsin D following lysosomal membrane permeabilization (LMP) determined necrosis-like cell death. Furthermore, propofol overdose also induced caspase-dependent apoptosis, at least in part. Caspase-3 was activated and acted downstream of mitochondrial transmembrane potential (MTP) loss; however, lysosomal cathepsins were not required for endothelial cell apoptosis. Notably, activation of GSK-3 was essential for propofol overdose-induced mitochondrial damage and apoptosis, but not necrosis-like cell death. Intraperitoneal administration of a propofol overdose in BALB/c mice caused an increase in peritoneal vascular permeability. These results demonstrate the cytotoxic effects of propofol overdose, including cathepsin D-regulated necrosis-like cell death and GSK-3-regulated mitochondrial apoptosis, on endothelial cells in vitro and the endothelial barrier dysfunction by propofol in vivo. Highlights: ► Propofol overdose causes apoptosis and necrosis in endothelial cells. ► Propofol overdose triggers lysosomal dysfunction independent of autophagy. ► Glycogen synthase kinase-3 facilitates propofol overdose-induced apoptosis. ► Propofol overdose causes an increase in peritoneal vascular permeability.« less

Protein kinase C-α and arginase I mediate pneumolysin-induced pulmonary endothelial hyperpermeability.

PubMed

Lucas, Rudolf; Yang, Guang; Gorshkov, Boris A; Zemskov, Evgeny A; Sridhar, Supriya; Umapathy, Nagavedi S; Jezierska-Drutel, Agnieszka; Alieva, Irina B; Leustik, Martin; Hossain, Hamid; Fischer, Bernhard; Catravas, John D; Verin, Alexander D; Pittet, Jean-François; Caldwell, Ruth B; Mitchell, Timothy J; Cederbaum, Stephen D; Fulton, David J; Matthay, Michael A; Caldwell, Robert W; Romero, Maritza J; Chakraborty, Trinad

2012-10-01

Antibiotics-induced release of the pore-forming virulence factor pneumolysin (PLY) in patients with pneumococcal pneumonia results in its presence days after lungs are sterile and is a major factor responsible for the induction of permeability edema. Here we sought to identify major mechanisms mediating PLY-induced endothelial dysfunction. We evaluated PLY-induced endothelial hyperpermeability in human lung microvascular endothelial cells (HL-MVECs) and human lung pulmonary artery endothelial cells in vitro and in mice instilled intratracheally with PLY. PLY increases permeability in endothelial monolayers by reducing stable and dynamic microtubule content and modulating VE-cadherin expression. These events, dependent upon an increased calcium influx, are preceded by protein kinase C (PKC)-α activation, perturbation of the RhoA/Rac1 balance, and an increase in myosin light chain phosphorylation. At later time points, PLY treatment increases the expression and activity of arginase in HL-MVECs. Arginase inhibition abrogates and suppresses PLY-induced endothelial barrier dysfunction by restoring NO generation. Consequently, a specific PKC-α inhibitor and the TNF-derived tonoplast intrinsic protein peptide, which blunts PLY-induced PKC-α activation, are able to prevent activation of arginase in HL-MVECs and to reduce PLY-induced endothelial hyperpermeability in mice. Arginase I (AI)(+/-)/arginase II (AII)(-/-) C57BL/6 mice, displaying a significantly reduced arginase I expression in the lungs, are significantly less sensitive to PLY-induced capillary leak than their wild-type or AI(+/+)/AII(-/-) counterparts, indicating an important role for arginase I in PLY-induced endothelial hyperpermeability. These results identify PKC-α and arginase I as potential upstream and downstream therapeutic targets in PLY-induced pulmonary endothelial dysfunction.

Protein Kinase C-α and Arginase I Mediate Pneumolysin-Induced Pulmonary Endothelial Hyperpermeability

PubMed Central

Yang, Guang; Gorshkov, Boris A.; Zemskov, Evgeny A.; Sridhar, Supriya; Umapathy, Nagavedi S.; Jezierska-Drutel, Agnieszka; Alieva, Irina B.; Leustik, Martin; Hossain, Hamid; Fischer, Bernhard; Catravas, John D.; Verin, Alexander D.; Pittet, Jean-François; Caldwell, Ruth B.; Mitchell, Timothy J.; Cederbaum, Stephen D.; Fulton, David J.; Matthay, Michael A.; Caldwell, Robert W.; Romero, Maritza J.; Chakraborty, Trinad

2012-01-01

Antibiotics-induced release of the pore-forming virulence factor pneumolysin (PLY) in patients with pneumococcal pneumonia results in its presence days after lungs are sterile and is a major factor responsible for the induction of permeability edema. Here we sought to identify major mechanisms mediating PLY-induced endothelial dysfunction. We evaluated PLY-induced endothelial hyperpermeability in human lung microvascular endothelial cells (HL-MVECs) and human lung pulmonary artery endothelial cells in vitro and in mice instilled intratracheally with PLY. PLY increases permeability in endothelial monolayers by reducing stable and dynamic microtubule content and modulating VE-cadherin expression. These events, dependent upon an increased calcium influx, are preceded by protein kinase C (PKC)-α activation, perturbation of the RhoA/Rac1 balance, and an increase in myosin light chain phosphorylation. At later time points, PLY treatment increases the expression and activity of arginase in HL-MVECs. Arginase inhibition abrogates and suppresses PLY-induced endothelial barrier dysfunction by restoring NO generation. Consequently, a specific PKC-α inhibitor and the TNF-derived tonoplast intrinsic protein peptide, which blunts PLY-induced PKC-α activation, are able to prevent activation of arginase in HL-MVECs and to reduce PLY-induced endothelial hyperpermeability in mice. Arginase I (AI)+/−/arginase II (AII)−/− C57BL/6 mice, displaying a significantly reduced arginase I expression in the lungs, are significantly less sensitive to PLY-induced capillary leak than their wild-type or AI+/+/AII−/− counterparts, indicating an important role for arginase I in PLY-induced endothelial hyperpermeability. These results identify PKC-α and arginase I as potential upstream and downstream therapeutic targets in PLY-induced pulmonary endothelial dysfunction. PMID:22582175

Verocytotoxin-induced apoptosis of human microvascular endothelial cells.

PubMed

Pijpers, A H; van Setten, P A; van den Heuvel, L P; Assmann, K J; Dijkman, H B; Pennings, A H; Monnens, L A; van Hinsbergh, V W

2001-04-01

The pathogenesis of the epidemic form of hemolytic uremic syndrome is characterized by endothelial cell damage. In this study, the role of apoptosis in verocytotoxin (VT)-mediated endothelial cell death in human glomerular microvascular endothelial cells (GMVEC), human umbilical vein endothelial cells, and foreskin microvascular endothelial cells (FMVEC) was investigated. VT induced apoptosis in GMVEC and human umbilical vein endothelial cells when the cells were prestimulated with the inflammatory mediator tumor necrosis factor-alpha (TNF-alpha). FMVEC displayed strong binding of VT and high susceptibility to VT under basal conditions, which made them suitable for the study of VT-induced apoptosis without TNF-alpha interference. On the basis of functional (flow cytometry and immunofluorescence microscopy using FITC-conjugated annexin V and propidium iodide), morphologic (transmission electron microscopy), and molecular (agarose gel electrophoresis of cellular DNA fragments) criteria, it was documented that VT induced programmed cell death in microvascular endothelial cells in a dose- and time-dependent manner. Furthermore, whereas partial inhibition of protein synthesis by VT was associated with a considerable number of apoptotic cells, comparable inhibition of protein synthesis by cycloheximide was not. This suggests that additional pathways, independent of protein synthesis inhibition, may be involved in VT-mediated apoptosis in microvascular endothelial cells. Specific inhibition of caspases by Ac-Asp-Glu-Val-Asp-CHO, but not by Ac-Tyr-Val-Ala-Asp-CHO, was accompanied by inhibition of VT-induced apoptosis in FMVEC and TNF-alpha-treated GMVEC. These data indicate that VT can induce apoptosis in human microvascular endothelial cells.

The flow dependency of Tie2 expression in endotoxemia.

PubMed

Kurniati, Neng F; Jongman, Rianne M; vom Hagen, Franziska; Spokes, Katherine C; Moser, Jill; Regan, Erzsébet Ravasz; Krenning, Guido; Moonen, Jan-Renier A J; Harmsen, Martin C; Struys, Michel M R F; Hammes, Hans-Peter; Zijlstra, Jan G; Aird, William C; Heeringa, Peter; Molema, Grietje; van Meurs, Matijs

2013-07-01

Tie2 is predominantly expressed by endothelial cells and is involved in vascular integrity control during sepsis. Changes in Tie2 expression during sepsis development may contribute to microvascular dysfunction. Understanding the kinetics and molecular basis of these changes may assist in the development of therapeutic intervention to counteract microvascular dysfunction. To investigate the molecular mechanisms underlying the changes in Tie2 expression upon lipopolysaccharide (LPS) challenge. Studies were performed in LPS and pro-inflammatory cytokine challenged mice as well as in mice subjected to hemorrhagic shock, primary endothelial cells were used for in vitro experiments in static and flow conditions. Eight hours after LPS challenge, Tie2 mRNA loss was observed in all major organs, while loss of Tie2 protein was predominantly observed in lungs and kidneys, in the capillaries. A similar loss could be induced by secondary cytokines TNF-α and IL-1β. Ang2 protein administration did not affect Tie2 protein expression nor was Tie2 protein rescued in LPS-challenged Ang2-deficient mice, excluding a major role for Ang2 in Tie2 down regulation. In vitro, endothelial loss of Tie2 was observed upon lowering of shear stress, not upon LPS and TNF-α stimulation, suggesting that inflammation related haemodynamic changes play a major role in loss of Tie2 in vivo, as also hemorrhagic shock induced Tie2 mRNA loss. In vitro, this loss was partially counteracted by pre-incubation with a pharmacologically NF-кB inhibitor (BAY11-7082), an effect further substantiated in vivo by pre-treatment of mice with the NF-кB inhibitor prior to the inflammatory challenge. Microvascular bed specific loss of Tie2 mRNA and protein in vivo upon LPS, TNFα, IL-1β challenge, as well as in response to hemorrhagic shock, is likely an indirect effect caused by a change in endothelial shear stress. This loss of Tie2 mRNA, but not Tie2 protein, induced by TNFα exposure was shown to be controlled by NF-кB signaling. Drugs aiming at restoring vascular integrity in sepsis could focus on preventing the Tie2 loss.

PEGylated-nanoliposomal clusterin for amyloidogenic light chain-induced endothelial dysfunction.

PubMed

Guzman-Villanueva, Diana; Migrino, Raymond Q; Truran, Seth; Karamanova, Nina; Franco, Daniel A; Burciu, Camelia; Senapati, Subhadip; Nedelkov, Dobrin; Hari, Parameswaran; Weissig, Volkmar

2018-06-01

Light chain (AL) amyloidosis is a disease associated with significant morbidity and mortality arising from multi-organ injury induced by amyloidogenic light chain proteins (LC). There is no available treatment to reverse the toxicity of LC. We previously showed that chaperone glycoprotein clusterin (CLU) and nanoliposomes (NL), separately, restore human microvascular endothelial function impaired by LC. In this work, we aim to prepare PEGylated-nanoliposomal clusterin (NL-CLU) formulations that could allow combined benefit against LC while potentially enabling efficient delivery to microvascular tissue, and test efficacy on human arteriole endothelial function. NL-CLU was prepared by a conjugation reaction between the carboxylated surface of NL and the primary amines of the CLU protein. NL were made of phosphatidylcholine (PC), cholesterol (Chol) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[carboxy(polyethylene glycol)-2000] (DSPE-PEG 2000 carboxylic acid) at 70:25:5 mol%. The protective effect of NL-CLU was tested by measuring the dilation response to acetylcholine and papaverine in human adipose arterioles exposed to LC. LC treatment significantly reduced the dilation response to acetylcholine and papaverine; co-treatment of LC with PEGylated-nanoliposomal CLU or free CLU restored the dilator response. NL-CLU is a feasible and promising approach to reverse LC-induced endothelial damage.

Priming Mesenchymal Stem Cells with Endothelial Growth Medium Boosts Stem Cell Therapy for Systemic Arterial Hypertension

PubMed Central

de Oliveira, Lucas Felipe; Almeida, Thalles Ramos; Ribeiro Machado, Marcus Paulo; Cuba, Marilia Beatriz; Alves, Angélica Cristina; da Silva, Marcos Vinícius; Rodrigues Júnior, Virmondes; Dias da Silva, Valdo José

2015-01-01

Systemic arterial hypertension (SAH), a clinical syndrome characterized by persistent elevation of arterial pressure, is often associated with abnormalities such as microvascular rarefaction, defective angiogenesis, and endothelial dysfunction. Mesenchymal stem cells (MSCs), which normally induce angiogenesis and improve endothelial function, are defective in SAH. The central aim of this study was to evaluate whether priming of MSCs with endothelial growth medium (EGM-2) increases their therapeutic effects in spontaneously hypertensive rats (SHRs). Adult female SHRs were administered an intraperitoneal injection of vehicle solution (n = 10), MSCs cultured in conventional medium (DMEM plus 10% FBS, n = 11), or MSCs cultured in conventional medium followed by 72 hours in EGM-2 (pMSC, n = 10). Priming of the MSCs reduced the basal cell death rate in vitro. The administration of pMSCs significantly induced a prolonged reduction (10 days) in arterial pressure, a decrease in cardiac hypertrophy, an improvement in endothelium-dependent vasodilation response to acetylcholine, and an increase in skeletal muscle microvascular density compared to the vehicle and MSC groups. The transplanted cells were rarely found in the hearts and kidneys. Taken together, our findings indicate that priming of MSCs boosts stem cell therapy for the treatment of SAH. PMID:26300922

Angiogenic dysfunction in bone marrow-derived early outgrowth cells from diabetic animals is attenuated by SIRT1 activation.

PubMed

Yuen, Darren A; Zhang, Yanling; Thai, Kerri; Spring, Christopher; Chan, Lauren; Guo, Xiaoxin; Advani, Andrew; Sivak, Jeremy M; Gilbert, Richard E

2012-12-01

Impaired endothelial repair is a key contributor to microvascular rarefaction and consequent end-organ dysfunction in diabetes. Recent studies suggest an important role for bone marrow-derived early outgrowth cells (EOCs) in mediating endothelial repair, but the function of these cells is impaired in diabetes, as in advanced age. We sought to determine whether diabetes-associated EOC dysfunction might be attenuated by pharmacological activation of silent information regulator protein 1 (SIRT1), a lysine deacetylase implicated in nutrient-dependent life span extension in mammals. Despite being cultured in normal (5.5 mM) glucose for 7 days, EOCs from diabetic rats expressed less SIRT1 mRNA, induced less endothelial tube formation in vitro and neovascularization in vivo, and secreted less of the proangiogenic ELR(+) CXC chemokines CXCL1, CXCL3, and CXCL5. Ex vivo SIRT1 activation restored EOC chemokine secretion and increased the in vitro and in vivo angiogenic activity of EOC conditioned medium derived from diabetic animals to levels similar to that derived from control animals. These findings suggest a pivotal role for SIRT1 in diabetes-induced EOC dysfunction and that its pharmacologic activation may provide a new strategy for the restoration of EOC-mediated repair mechanisms.

The Krebs cycle and mitochondrial mass are early victims of endothelial dysfunction: proteomic approach.

PubMed

Addabbo, Francesco; Ratliff, Brian; Park, Hyeong-Cheon; Kuo, Mei-Chuan; Ungvari, Zoltan; Csiszar, Anna; Ciszar, Anna; Krasnikov, Boris; Krasnikof, Boris; Sodhi, Komal; Zhang, Fung; Nasjletti, Alberto; Goligorsky, Michael S

2009-01-01

Endothelial cell dysfunction is associated with bioavailable nitric oxide deficiency and an excessive generation of reactive oxygen species. We modeled this condition by chronically inhibiting nitric oxide generation with subpressor doses of N(G)-monomethyl-L-arginine (L-NMMA) in C57B6 and Tie-2/green fluorescent protein mouse strains. L-NMMA-treated mice exhibited a slight reduction in vasorelaxation ability, as well as detectable abnormalities in soluble adhesion molecules (soluble intercellular adhesion molecule-1 and vascular cellular adhesion molecule-1, and matrix metalloproteinase 9), which represent surrogate indicators of endothelial dysfunction. Proteomic analysis of the isolated microvasculature using 2-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy revealed abnormal expression of a cluster of mitochondrial enzymes, which was confirmed using immunodetection. Aconitase-2 and enoyl-CoA-hydratase-1 expression levels were decreased in L-NMMA-treated animals; this phenotype was absent in nitric oxide synthase-1 and -3 knockout mice. Depletion of aconitase-2 and enoyl-CoA-hydratase-1 resulted in the inhibition of the Krebs cycle and enhanced pyruvate shunting toward the glycolytic pathway. To assess mitochondrial mass in vivo, co-localization of green fluorescent protein and MitoTracker fluorescence was detected by intravital microscopy. Quantitative analysis of fluorescence intensity showed that L-NMMA-treated animals exhibited lower fluorescence of MitoTracker in microvascular endothelia as a result of reduced mitochondrial mass. These findings provide conclusive and unbiased evidence that mitochondriopathy represents an early manifestation of endothelial dysfunction, shifting cell metabolism toward "metabolic hypoxia" through the selective depletion of both aconitase-2 and enoyl-CoA-hydratase-1. These findings may contribute to an early preclinical diagnosis of endothelial dysfunction.

In vitro model of cerebral ischemia by using brain microvascular endothelial cells derived from human induced pluripotent stem cells.

PubMed

Kokubu, Yasuhiro; Yamaguchi, Tomoko; Kawabata, Kenji

2017-04-29

Brain-derived microvascular endothelial cells (BMECs), which play a central role in blood brain barrier (BBB), can be used for the evaluation of drug transport into the brain. Although human BMEC cell lines have already been reported, they lack original properties such as barrier integrity. Pluripotent stem cells (PSCs) can be used for various applications such as regenerative therapy, drug screening, and pathological study. In the recent study, an induction method of BMECs from PSCs has been established, making it possible to more precisely study the in vitro human BBB function. Here, using induced pluripotent stem (iPS) cell-derived BMECs, we examined the effects of oxygen-glucose deprivation (OGD) and OGD/reoxygenation (OGD/R) on BBB permeability. OGD disrupted the barrier function, and the dysfunction was rapidly restored by re-supply of the oxygen and glucose. Interestingly, TNF-α, which is known to be secreted from astrocytes and microglia in the cerebral ischemia, prevented the restoration of OGD-induced barrier dysfunction in an apoptosis-independent manner. Thus, we could establish the in vitro BBB disease model that mimics the cerebral ischemia by using iPS cell-derived BMECs. Copyright © 2017 Elsevier Inc. All rights reserved.

The relationship between insulin resistance and endothelial dysfunction in obese adolescents.

PubMed

Brar, Preneet Cheema; Patel, Payal; Katz, Stuart

2017-05-24

Insulin resistance and endothelial dysfunction share a reciprocal relationship that links the metabolic and cardiovascular sequelae of obesity. We characterized the brachial artery reactivity testing (BART) and carotid artery-intima media thickness (CIMT) in adolescents categorized as obese insulin resistant (OIR) and obese not insulin resistant (ONIR). Lipoprotein particle (p) analysis and inflammatory cytokines in OIR and ONIR groups were also analyzed. Obese adolescents (n=40; mean body mass index [BMI] 35.6) were categorized as ONIR and OIR based on their homeostatic model assessment of insulin resistance (HOMA-IR) calculation (≤or> than 3.4). Ultrasound measured conduit arterial function BART, microvascular function (post-ischemic hyperemia) and conduit artery structure CIMT. BART did not differ according to IR status (mean±SD: 7.0±4.3% vs. 5.9±3.4% in ONIR and OIR, respectively, p=0.3, but post-ischemic hyperemia was significantly greater in the ONIR group (4.5±2.2 vs. 3.5±3, p=0.04). Atherogenic lipoprotein particles; large VLDL particles and small LDL particles were higher in the OIR compared to ONIR group. OIR adolescents demonstrate an inflamed atherogenic milieu compared to the ONIR adolescents. Microvascular function, but not conduit vessel structure or function, was impaired in association with IR.

Classical cardiovascular disease risk factors associate with vascular function and morphology in rheumatoid arthritis: a six-year prospective study

PubMed Central

2013-01-01

Introduction Patients with rheumatoid arthritis (RA) are at an increased risk for cardiovascular disease (CVD). An early manifestation of CVD is endothelial dysfunction which can lead to functional and morphological vascular abnormalities. Classical CVD risk factors and inflammation are both implicated in causing endothelial dysfunction in RA. The objective of the present study was to examine the effect of baseline inflammation, cumulative inflammation, and classical CVD risk factors on the vasculature following a six-year follow-up period. Methods A total of 201 RA patients (155 females, median age (25th to 75th percentile): 61 years (53 to 67)) were examined at baseline (2006) for presence of classical CVD risk factors and determination of inflammation using C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR). At follow-up (2012) patients underwent assessments of microvascular and macrovascular endothelium-dependent and endothelium-independent function, along with assessment of carotid atherosclerosis. The CRP and ESR were recorded from the baseline study visit to the follow-up visit for each patient to calculate cumulative inflammatory burden. Results Classical CVD risk factors, but not RA disease-related inflammation, predicted microvascular endothelium-dependent and endothelium-independent function, macrovascular endothelium-independent function and carotid atherosclerosis. These findings were similar in a sub-group of patients free from CVD, and not receiving non-steroidal anti-inflammatory drugs, cyclooxygenase 2 inhibitors or biologics. Cumulative inflammation was not associated with microvascular and macrovascular endothelial function, but a weak association was apparent between area under the curve for CRP and carotid atherosclerosis. Conclusions Classical CVD risk factors may be better long-term predictors of vascular function and morphology than systemic disease-related inflammation in patients with RA. Further studies are needed to confirm if assessments of vascular function and morphology are predictive of long-term CV outcomes in RA. PMID:24289091

Microvascular disorders in obese Zucker rats are restored by a rice bran diet.

PubMed

Justo, M L; Claro, C; Vila, E; Herrera, M D; Rodriguez-Rodriguez, R

2014-05-01

Nutritional-based approaches aimed to prevent microvascular dysfunction associated to obesity present potential advantages over pharmacological strategies. Our aim was to test whether a rice bran enzymatic extract (RBEE)-supplemented diet could attenuate microvascular alterations in obese rats. Lean and obese Zucker rats were fed standard diet supplemented or not with 1% and 5% RBEE for 20 weeks. Functional studies were performed in small mesenteric arteries in isometric myograph. Immunoblotting and fluorescence studies were made in arterial homogenates and arterial sections, respectively. RBEE-supplementation restored microvascular function in obese rats through a marked increase in NO and endothelial-derived hyperpolarizing factor contribution by up-regulation of eNOS and calcium-activated potassium channels expression, respectively, in association to a substantial reduction of microvascular inflammation and superoxide anion formation. These data agrees with the beneficial actions of RBEE on dyslipidemia, hyperinsulinemia and hypertension in obesity. The multi-factorial properties of RBEE-diet, especially for restoring the function of small resistance arteries shows this dietary-based approach to be a promising candidate for prevention of microvascular alterations in obesity, which are crucial in cardiovascular events in obese subjects. Copyright © 2013 Elsevier B.V. All rights reserved.

Expression of PKA inhibitor (PKI) gene abolishes cAMP-mediated protection to endothelial barrier dysfunction.

PubMed

Lum, H; Jaffe, H A; Schulz, I T; Masood, A; RayChaudhury, A; Green, R D

1999-09-01

We investigated the hypothesis that cAMP-dependent protein kinase (PKA) protects against endothelial barrier dysfunction in response to proinflammatory mediators. An E1-, E3-, replication-deficient adenovirus (Ad) vector was constructed containing the complete sequence of PKA inhibitor (PKI) gene (AdPKI). Infection of human microvascular endothelial cells (HMEC) with AdPKI resulted in overexpression of PKI. Treatment with 0.5 microM thrombin increased transendothelial albumin clearance rate (0.012 +/- 0.003 and 0.035 +/- 0.005 microl/min for control and thrombin, respectively); the increase was prevented with forskolin + 3-isobutyl-1-methylxanthine (F + I) treatment. Overexpression of PKI resulted in abrogation of the F + I-induced inhibition of the permeability increase. However, with HMEC infected with ultraviolet-inactivated AdPKI, the F + I-induced inhibition was present. Also, F + I treatment of HMEC transfected with reporter plasmid containing the cAMP response element-directed transcription of the luciferase gene resulted in an almost threefold increase in luciferase activity. Overexpression of PKI inhibited this induction of luciferase activity. The results show that Ad-mediated overexpression of PKI in endothelial cells abrogated the cAMP-mediated protection against increased endothelial permeability, providing direct evidence that cAMP-dependent protein kinase promotes endothelial barrier function.

Cardiac Remodeling: Endothelial Cells Have More to Say Than Just NO

PubMed Central

Segers, Vincent F. M.; Brutsaert, Dirk L.; De Keulenaer, Gilles W.

2018-01-01

The heart is a highly structured organ consisting of different cell types, including myocytes, endothelial cells, fibroblasts, stem cells, and inflammatory cells. This pluricellularity provides the opportunity of intercellular communication within the organ, with subsequent optimization of its function. Intercellular cross-talk is indispensable during cardiac development, but also plays a substantial modulatory role in the normal and failing heart of adults. More specifically, factors secreted by cardiac microvascular endothelial cells modulate cardiac performance and either positively or negatively affect cardiac remodeling. The role of endothelium-derived small molecules and peptides—for instance NO or endothelin-1—has been extensively studied and is relatively well defined. However, endothelial cells also secrete numerous larger proteins. Information on the role of these proteins in the heart is scattered throughout the literature. In this review, we will link specific proteins that modulate cardiac contractility or cardiac remodeling to their expression by cardiac microvascular endothelial cells. The following proteins will be discussed: IL-6, periostin, tenascin-C, thrombospondin, follistatin-like 1, frizzled-related protein 3, IGF-1, CTGF, dickkopf-3, BMP-2 and−4, apelin, IL-1β, placental growth factor, LIF, WISP-1, midkine, and adrenomedullin. In the future, it is likely that some of these proteins can serve as markers of cardiac remodeling and that the concept of endothelial function and dysfunction might have to be redefined as we learn more about other factors secreted by ECs besides NO. PMID:29695980

Blood-brain barrier dysfunction and amyloid precursor protein accumulation in microvascular compartment following ischemia-reperfusion brain injury with 1-year survival.

PubMed

Pluta, R

2003-01-01

This study examined the late microvascular consequences of brain ischemia due to cardiac arrest in rats. In reacted vibratome sections scattered foci of extravasated horseradish peroxidase were noted throughout the brain and did not appear to be restricted to any specific area of brain. Ultrastructural investigation of leaky sites frequently presented platelets adhering to the endothelium of venules and capillaries. Endothelial cells demonstrated pathological changes with evidence of perivascular astrocytic swelling. At the same time, we noted C-terminal of amyloid precursor protein/beta-amyloid peptide (CAPP/betaA) deposits in cerebral blood vessels, with a halo of CAPP/betaA immunoreactivity in the surrounding parenchyma suggested diffusion of CAPP/betaA out of the vascular compartment. Changes predominated in the hippocampus, cerebral and entorhinal cortex, corpus callosum, thalamus, basal ganglia and around the lateral ventricles. These data implicate delayed abnormal endothelial function of vessels following ischemia-reperfusion brain injury as a primary event in the pathogenesis of the recurrent cerebral infarction.

Hydrogen sulfide replacement therapy protects the vascular endothelium in hyperglycemia by preserving mitochondrial function.

PubMed

Suzuki, Kunihiro; Olah, Gabor; Modis, Katalin; Coletta, Ciro; Kulp, Gabriella; Gerö, Domokos; Szoleczky, Petra; Chang, Tuanjie; Zhou, Zongmin; Wu, Lingyun; Wang, Rui; Papapetropoulos, Andreas; Szabo, Csaba

2011-08-16

The goal of the present studies was to investigate the role of changes in hydrogen sulfide (H(2)S) homeostasis in the pathogenesis of hyperglycemic endothelial dysfunction. Exposure of bEnd3 microvascular endothelial cells to elevated extracellular glucose (in vitro "hyperglycemia") induced the mitochondrial formation of reactive oxygen species (ROS), which resulted in an increased consumption of endogenous and exogenous H(2)S. Replacement of H(2)S or overexpression of the H(2)S-producing enzyme cystathionine-γ-lyase (CSE) attenuated the hyperglycemia-induced enhancement of ROS formation, attenuated nuclear DNA injury, reduced the activation of the nuclear enzyme poly(ADP-ribose) polymerase, and improved cellular viability. In vitro hyperglycemia resulted in a switch from oxidative phosphorylation to glycolysis, an effect that was partially corrected by H(2)S supplementation. Exposure of isolated vascular rings to high glucose in vitro induced an impairment of endothelium-dependent relaxations, which was prevented by CSE overexpression or H(2)S supplementation. siRNA silencing of CSE exacerbated ROS production in hyperglycemic endothelial cells. Vascular rings from CSE(-/-) mice exhibited an accelerated impairment of endothelium-dependent relaxations in response to in vitro hyperglycemia, compared with wild-type controls. Streptozotocin-induced diabetes in rats resulted in a decrease in the circulating level of H(2)S; replacement of H(2)S protected from the development of endothelial dysfunction ex vivo. In conclusion, endogenously produced H(2)S protects against the development of hyperglycemia-induced endothelial dysfunction. We hypothesize that, in hyperglycemic endothelial cells, mitochondrial ROS production and increased H(2)S catabolism form a positive feed-forward cycle. H(2)S replacement protects against these alterations, resulting in reduced ROS formation, improved endothelial metabolic state, and maintenance of normal endothelial function.

CORM-A1 prevents blood-brain barrier dysfunction caused by ionotropic glutamate receptor-mediated endothelial oxidative stress and apoptosis.

PubMed

Basuroy, Shyamali; Leffler, Charles W; Parfenova, Helena

2013-06-01

In cerebral microvascular endothelial cells (CMVEC) of newborn pigs, glutamate at excitotoxic concentrations (mM) causes apoptosis mediated by reactive oxygen species (ROS). Carbon monoxide (CO) produced by CMVEC or delivered by a CO-releasing molecule, CORM-A1, has antioxidant properties. We tested the hypothesis that CORM-A1 prevents cerebrovascular endothelial barrier dysfunction caused by glutamate excitotoxicity. First, we identified the glutamate receptors (GluRs) and enzymatic sources of ROS involved in the mechanism of endothelial apoptosis. In glutamate-exposed CMVEC, ROS formation and apoptosis were blocked by rotenone, 2-thenoyltrifluoroacetone (TTFA), and antimycin, indicating that mitochondrial complexes I, II, and III are the major sources of oxidative stress. Agonists of ionotropic GluRs (iGluRs) N-methyl-D-aspartate (NMDA), cis-ACPD, AMPA, and kainate increased ROS production and apoptosis, whereas iGluR antagonists exhibited antiapoptotic properties, suggesting that iGluRs mediate glutamate-induced endothelial apoptosis. The functional consequences of endothelial injury were tested in the model of blood-brain barrier (BBB) composed of CMVEC monolayer on semipermeable membranes. Glutamate and iGluR agonists reduced transendothelial electrical resistance and increased endothelial paracellular permeability to 3-kDa dextran. CORM-A1 exhibited potent antioxidant and antiapoptotic properties in CMVEC and completely prevented BBB dysfunction caused by glutamate and iGluR agonists. Overall, the endothelial component of the BBB is a cellular target for excitotoxic glutamate that, via a mechanism involving a iGluR-mediated activation of mitochondrial ROS production and apoptosis, leads to BBB opening that may be prevented by the antioxidant and antiapoptotic actions of CORMs. Antioxidant CORMs therapy may help preserve BBB functional integrity in neonatal cerebrovascular disease.

Longitudinal assessment of maternal endothelial function and markers of inflammation and placental function throughout pregnancy in lean and obese mothers.

PubMed

Stewart, Frances M; Freeman, Dilys J; Ramsay, Jane E; Greer, Ian A; Caslake, Muriel; Ferrell, William R

2007-03-01

Obesity in pregnancy is increasing and is a risk factor for metabolic pathology such as preeclampsia. In the nonpregnant, obesity is associated with dyslipidemia, vascular dysfunction, and low-grade chronic inflammation. Our aim was to measure microvascular endothelial function in lean and obese pregnant women at intervals throughout their pregnancies and at 4 months after delivery. Plasma markers of endothelial function, inflammation, and placental function and their association with microvascular function were also assessed. Women in the 1st trimester of pregnancy were recruited, 30 with a body mass index (BMI) less than 30 kg/m(2) and 30 with a BMI more than or equal to 30 kg/m(2) matched for age, parity, and smoking status. In vivo endothelial-dependent and -independent microvascular function was measured using laser Doppler imaging in the 1st, 2nd, and 3rd trimesters of pregnancy and at 4 months postnatal. Plasma markers of endothelial activation [soluble intercellular cell adhesion molecule-1 (sVCAM-1), soluble vascular cell adhesion molecule-1 (sVCAM-1), von Willebrand factor (vWF), and plasminogen activator inhibitor (PAI)-1], inflammation (IL-6, TNFalpha, C-reactive protein, and IL-10), and placental function (PAI-1/PAI-2 ratio) were also assessed at each time point. The pattern of improving endothelial function during pregnancy was the same for lean and obese, but endothelial-dependent vasodilation was significantly lower (P < 0.05) in the obese women at each trimester (51, 41, and 39%, respectively). In the postpartum period, the improvement in endothelial-dependent vasodilation persisted in the lean women but declined to near 1st trimester levels in the obese (lean/obese difference, 115%; P < 0.01). There was a small but significant difference in endothelial-independent vasodilation between the two groups, lean response being greater than obese (P = 0.021), and response declined in both groups in the postpartum period. In multivariate analysis, time of sampling had the most impact on endothelial-independent function [18.5% (adjusted sum of squares expressed as a percentage of total means squared), P < 0.001 for sodium nitroprusside response; 9.8%, P < 0.001 for acetylcholine response], and obesity had the most impact on endothelial-dependent microvascular function (1.7%, P = 0.046 for sodium nitroprusside response; 19.3%, P < 0.001 for acetylcholine response). Time of sampling (11.2%, P < 0.001), IL-6 (4.0%, P = 0.002), and IL-10 (2.4%, P = 0.018) were significant independent contributors to variation in endothelial-dependent microvascular function. When obesity was entered into the model, the association with IL-6 and IL-10 was no longer significant, and obesity explained 6.8% (P < 0.001) of the variability in endothelial-dependent microvascular function. In the 1st trimester, obese women had a significantly higher PAI-1/PAI-2 ratio [obese median (interquartile range), 0.87 (0.54-1.21) vs. lean 0.30 (0.21-0.47), P < 0.001), reflecting the lower PAI-2 levels in obese pregnant women. In a multivariate analysis, 1st trimester BMI (7.6%, P = 0.012), IL-10 (8.2%, P < 0.001), and sVCAM-1 (0.73%, P = 0.007) contributed to the 1st trimester PAI-1/PAI-2 ratio. Obese mothers have a lower endothelium-dependent and -independent vasodilation when compared with lean counterparts. There was a higher PAI-1/ PAI-2 ratio in the 1st trimester in obese women, which improved later in pregnancy. Obese pregnancy is associated with chronic preexisting endothelial activation and impairment of endothelial function secondary to increased production of inflammatory T-helper cells-2 cytokines.

Monosialoganglioside-Containing Nanoliposomes Restore Endothelial Function Impaired by AL Amyloidosis Light Chain Proteins.

PubMed

Franco, Daniel A; Truran, Seth; Weissig, Volkmar; Guzman-Villanueva, Diana; Karamanova, Nina; Senapati, Subhadip; Burciu, Camelia; Ramirez-Alvarado, Marina; Blancas-Mejia, Luis M; Lindsay, Stuart; Hari, Parameswaran; Migrino, Raymond Q

2016-06-13

Light chain amyloidosis (AL) is associated with high mortality, especially in patients with advanced cardiovascular involvement. It is caused by toxicity of misfolded light chain proteins (LC) in vascular, cardiac, and other tissues. There is no treatment to reverse LC tissue toxicity. We tested the hypothesis that nanoliposomes composed of monosialoganglioside, phosphatidylcholine, and cholesterol (GM1 ganglioside-containing nanoliposomes [NLGM1]) can protect against LC-induced human microvascular dysfunction and assess mechanisms behind the protective effect. The dilator responses of ex vivo abdominal adipose arterioles from human participants without AL to acetylcholine and papaverine were measured before and after exposure to LC (20 μg/mL) with or without NLGM1 (1:10 ratio for LC:NLGM1 mass). Human umbilical vein endothelial cells were exposed for 18 to 20 hours to vehicle, LC with or without NLGM1, or NLGM1 and compared for oxidative and nitrative stress response and cellular viability. LC impaired arteriole dilator response to acetylcholine, which was restored by co-treatment with NLGM1. LC decreased endothelial cell nitric oxide production and cell viability while increasing superoxide and peroxynitrite; these adverse effects were reversed by NLGM1. NLGM1 increased endothelial cell protein expression of antioxidant enzymes heme oxygenase 1 and NAD(P)H quinone dehydrogenase 1 and increased nuclear factor, erythroid 2 like 2 (Nrf-2) protein. Nrf-2 gene knockdown reduced antioxidant stress response and reversed the protective effects of NLGM1. NLGM1 protects against LC-induced human microvascular endothelial dysfunction through increased nitric oxide bioavailability and reduced oxidative and nitrative stress mediated by Nrf-2-dependent antioxidant stress response. These findings point to a potential novel therapeutic approach for light chain amyloidosis. © 2016 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

Microvascular endothelial function and cognitive performance: The ELSA-Brasil cohort study.

PubMed

Brant, Luisa; Bos, Daniel; Araujo, Larissa Fortunato; Ikram, M Arfan; Ribeiro, Antonio Lp; Barreto, Sandhi M

2018-06-01

Impaired microvascular endothelial function may be implicated in the etiology of cognitive decline. Yet, current data on this association are inconsistent. Our objective is to investigate the relation of microvascular endothelial function to cognitive performance in the ELSA-Brasil cohort study. A total of 1521 participants from ELSA-Brasil free of dementia underwent peripheral arterial tonometry (PAT) to quantify microvascular endothelial function (PAT-ratio and mean baseline pulse amplitude (BPA)) and cognitive tests that covered the domains of memory, verbal fluency, and executive function at baseline. Cognitive tests in participants aged 55 years old and above were repeated during the second examination (mean follow-up: 3.5 (0.3) years). Linear regression and generalized linear models were used to evaluate the association between endothelial function, global cognitive performance, and performance on specific cognitive domains. In unadjusted cross-sectional analyses, we found that BPA and PAT-ratio were associated with worse global cognitive performance (mean difference for BPA: -0.07, 95% CI: -0.11; -0.03, p<0.01; mean difference for PAT-ratio: 0.11, 95% CI: 0.01; 0.20, p=0.02), worse performance on learning, recall, and word recognition tests (BPA: -0.87, 95% CI: -1.21; -0.52, p<0.01; PAT-ratio: 1.58, 95% CI: 0.80; 2.36, p<0.01), and only BPA was associated with worse performance in verbal fluency tests (-0.70, 95% CI: -1.19; -0.21, p<0.01). Adjustments for age, sex, and level of education rendered the associations statistically non-significant. Longitudinally, there was no association between microvascular endothelial and cognitive functions. The associations between microvascular endothelial function and cognition are explained by age, sex, and educational level. Measures of microvascular endothelial function may be of limited value with regard to preclinical cognitive deficits.

The effects of anti-obesity intervention with orlistat and sibutramine on microvascular endothelial function.

PubMed

Al-Tahami, Belqes Abdullah Mohammad; Ismail, Ab Aziz Al-Safi; Bee, Yvonne Tee Get; Awang, Siti Azima; Salha Wan Abdul Rani, Wan Rimei; Sanip, Zulkefli; Rasool, Aida Hanum Ghulam

2015-01-01

Obesity is associated with impaired microvascular endothelial function. We aimed to determine the effects of orlistat and sibutramine treatment on microvascular endothelial function, anthropometric and lipid profile, blood pressure (BP), and heart rate (HR). 76 subjects were recruited and randomized to receive orlistat 120 mg three times daily or sibutramine 10 mg daily for 9 months. Baseline weight, BMI, BP, HR and lipid profile were taken. Microvascular endothelial function was assessed using laser Doppler fluximetry and iontophoresis process. Maximum change (max), percent change (% change) and peak flux (peak) in perfusion to acetylcholine (ACh) and sodium nitroprusside (SNP) iontophoresis were used to quantify endothelium dependent and independent vasodilatations. 24 subjects in both groups completed the trial. After treatment, weight and BMI were decreased for both groups. AChmax, ACh % change and ACh peak were increased in orlistat-treated group but no difference was observed for sibutramine-treated group. BP and total cholesterol (TC) were reduced for orlistat-treated group. HR was reduced for orlistat-treated group but was increased in sibutramine-treated group. 9 months treatment with orlistat significantly improved microvascular endothelial function. This was associated with reductions in weight, BMI, BP, HR, TC and low density lipoprotein cholesterol. No effect was seen in microvascular endothelial function with sibutramine.

Listeriolysin O mediates cytotoxicity against human brain microvascular

USDA-ARS?s Scientific Manuscript database

Penetration of the brain microvascular endothelial layer is one of the routes L. monocytogenes use to breach the blood-brain barrier. Because host factors in the blood severely limit direct invasion of human brain microvascular endothelial cells (HBMECs) by L. monocytogenes, alternative mechanisms m...

Alda-1 Protects Against Acrolein-Induced Acute Lung Injury and Endothelial Barrier Dysfunction.

PubMed

Lu, Qing; Mundy, Miles; Chambers, Eboni; Lange, Thilo; Newton, Julie; Borgas, Diana; Yao, Hongwei; Choudhary, Gaurav; Basak, Rajshekhar; Oldham, Mahogany; Rounds, Sharon

2017-12-01

Inhalation of acrolein, a highly reactive aldehyde, causes lung edema. The underlying mechanism is poorly understood and there is no effective treatment. In this study, we demonstrated that acrolein not only dose-dependently induced lung edema but also promoted LPS-induced acute lung injury. Importantly, acrolein-induced lung injury was prevented and rescued by Alda-1, an activator of mitochondrial aldehyde dehydrogenase 2. Acrolein also dose-dependently increased monolayer permeability, disrupted adherens junctions and focal adhesion complexes, and caused intercellular gap formation in primary cultured lung microvascular endothelial cells (LMVECs). These effects were attenuated by Alda-1 and the antioxidant N-acetylcysteine, but not by the NADPH inhibitor apocynin. Furthermore, acrolein inhibited AMP-activated protein kinase (AMPK) and increased mitochondrial reactive oxygen species levels in LMVECs-effects that were associated with impaired mitochondrial respiration. AMPK total protein levels were also reduced in lung tissue of mice and LMVECs exposed to acrolein. Activation of AMPK with 5-aminoimidazole-4-carboxamide-1-β-4-ribofuranoside blunted an acrolein-induced increase in endothelial monolayer permeability, but not mitochondrial oxidative stress or inhibition of mitochondrial respiration. Our results suggest that acrolein-induced mitochondrial dysfunction may not contribute to endothelial barrier dysfunction. We speculate that detoxification of acrolein by Alda-1 and activation of AMPK may be novel approaches to prevent and treat acrolein-associated acute lung injury, which may occur after smoke inhalation.

Coronary microvascular function in patients with isolated systolic and combined systolic/diastolic hypertension.

PubMed

Bozbas, Huseyin; Pirat, Bahar; Yildirir, Aylin; Eroglu, Serpil; Simsek, Vahide; Sade, Elif; Atar, Ilyas; Aydinalp, Alp; Ozin, Bulent; Muderrisoglu, Haldun

2012-12-01

Isolated systolic hypertension (ISH) is a common condition in the elderly that is associated with endothelial dysfunction. Concerning the effect of type of hypertension on coronary microvascular function, coronary flow reserve (CFR) in patients with ISH was evaluated and the results were compared with patients with combined systolic/diastolic hypertension (SDH). Seventy-six elderly patients (older than 60 years) who were free of coronary artery disease and diabetes mellitus were enrolled in the study (38 with ISH and 38 with combined SDH). Using transthoracic Doppler echocardiography, CFR was calculated as the ratio of hyperemic to baseline diastolic peak flow velocities. A CFR value of >2 was accepted as normal. The mean age was 68.6±6.3 years and the groups had similar features with regard to demographic and clinical characteristics. Patients with ISH had significantly lower CFR values compared with those with combined SDH (2.22±0.51 vs 2.49±0.56, respectively; P=.03). On multivariate regression analysis, ISH (β=-0.40, P=.004) and dyslipidemia (β=-0.29, P=.04) were the independent predictors of CFR. These findings indicate that CFR, an indicator of coronary microvascular/endothelial function, is impaired more profoundly in patients with ISH than in patients with combined SDH. © 2012 Wiley Periodicals, Inc.

Increase in acid sphingomyelinase level in human retinal endothelial cells and CD34+ circulating angiogenic cells isolated from diabetic individuals is associated with dysfunctional retinal vasculature and vascular repair process in diabetes

PubMed Central

Kady, Nermin; Yan, Yuanqing; Salazar, Tatiana; Wang, Qi; Chakravarthy, Harshini; Huang, Chao; Beli, Eleni; Navitskaya, Svetlana; Grant, Maria; Busik, Julia

2017-01-01

Background Diabetic retinopathy (DR) is a microvascular disease that results from retinal vascular degeneration and defective repair due to diabetes induced endothelial progenitor dysfunction. Objective Understanding key molecular factors involved in vascular degeneration and repair is paramount for developing effective DR treatment strategies. We propose that diabetes-induced activation of acid sphingomyelinase (ASM) plays essential role in retinal endothelial and CD34+ circulating angiogenic cell (CAC) dysfunction in diabetes. Methods Human retinal endothelial cells (HRECs) isolated from control and diabetic donor tissue and human CD34+ CACs from control and diabetic patients were used in this study. ASM mRNA and protein expression was assessed by quantitative PCR and ELISA, respectively. To evaluate the effect of diabetes-induced ASM on HRECs and CD34+ CACs function, tube formation, CAC incorporation into endothelial tubes, and diurnal release of CD34+ CACs in diabetic individuals was determined. Results ASM expression level was significantly increased in HRECs isolated from diabetic compared to control donor tissue, as well as CD34+CACs and plasma of diabetic patients. A significant decrease in tube area was observed in HRECs from diabetic donors as compared to control HRECs. The tube formation deficiency was associated with increased expression of ASM in diabetic HRECs. Moreover, diabetic CD34+ CACs with high ASM showed defective incorporation into endothelial tubes. Diurnal release of CD34+ CACs was disrupted with the rhythmicity lost in diabetic patients. Conclusion Collectively, these findings support that diabetes-induced ASM upregulation has a marked detrimental effect on both retinal endothelial cells and CACs. PMID:28457994

Regulation of human cerebro-microvascular endothelial baso-lateral adhesion and barrier function by S1P through dual involvement of S1P1 and S1P2 receptors.

PubMed

Wiltshire, Rachael; Nelson, Vicky; Kho, Dan Ting; Angel, Catherine E; O'Carroll, Simon J; Graham, E Scott

2016-01-27

Herein we show that S1P rapidly and acutely reduces the focal adhesion strength and barrier tightness of brain endothelial cells. xCELLigence biosensor technology was used to measure focal adhesion, which was reduced by S1P acutely and this response was mediated through both S1P1 and S1P2 receptors. S1P increased secretion of several pro-inflammatory mediators from brain endothelial cells. However, the magnitude of this response was small in comparison to that mediated by TNFα or IL-1β. Furthermore, S1P did not significantly increase cell-surface expression of any key cell adhesion molecules involved in leukocyte recruitment, included ICAM-1 and VCAM-1. Finally, we reveal that S1P acutely and dynamically regulates microvascular endothelial barrier tightness in a manner consistent with regulated rapid opening followed by closing and strengthening of the barrier. We hypothesise that the role of the S1P receptors in this process is not to cause barrier dysfunction, but is related to controlled opening of the endothelial junctions. This was revealed using real-time measurement of barrier integrity using ECIS ZΘ TEER technology and endothelial viability using xCELLigence technology. Finally, we show that these responses do not occur simply though the pharmacology of a single S1P receptor but involves coordinated action of S1P1 and S1P2 receptors.

Long Noncoding RNA-GAS5: A Novel Regulator of Hypertension-Induced Vascular Remodeling.

PubMed

Wang, Yang-Ning-Zhi; Shan, Kun; Yao, Mu-Di; Yao, Jin; Wang, Jia-Jian; Li, Xiang; Liu, Ban; Zhang, Yang-Yang; Ji, Yong; Jiang, Qin; Yan, Biao

2016-09-01

Vascular remodeling is an important pathological feature of hypertension, leading to increased vascular resistance and reduced compliance. Endothelial cell (EC) and vascular smooth muscle cell (VSMC) dysfunction is involved in vascular remodeling. Long noncoding RNAs are potential regulators of EC and VSMC function. Herein, we determined whether long noncoding RNA-growth arrest-specific 5 (GAS5) is involved in hypertension-related vascular remodeling. We revealed that GAS5 knockdown aggravated hypertension-induced microvascular dysfunction as shown by increased retinal neovascularization and capillary leakage. GAS5 regulated the remodeling of arteries, including caudal arteries, carotid arteries, renal arteries, and thoracic arteries. GAS5 was mainly expressed in ECs and VSMCs, and its expression was significantly downregulated in hypertension. GAS5 knockdown affected endothelial activation, endothelial proliferation, VSMC phenotypic conversion, and EC-VSMC communication in vivo and in vitro. Mechanistically, GAS5 regulated EC and VSMC function through β-catenin signaling. This study identified GAS5 as a critical regulator in hypertension and demonstrated the potential of gene therapy and drug development for treating hypertension. © 2016 American Heart Association, Inc.

Fetoplacental Vascular Endothelial Dysfunction as an Early Phenomenon in the Programming of Human Adult Diseases in Subjects Born from Gestational Diabetes Mellitus or Obesity in Pregnancy

PubMed Central

Leiva, Andrea; Pardo, Fabián; Ramírez, Marco A.; Farías, Marcelo; Casanello, Paola; Sobrevia, Luis

2011-01-01

Gestational diabetes mellitus (GDM) and obesity in pregnancy (OP) are pathological conditions associated with placenta vascular dysfunction coursing with metabolic changes at the fetoplacental microvascular and macrovascular endothelium. These alterations are seen as abnormal expression and activity of the cationic amino acid transporters and endothelial nitric oxide synthase isoform, that is, the “endothelial L-arginine/nitric oxide signalling pathway.” Several studies suggest that the endogenous nucleoside adenosine along with insulin, and potentially arginases, are factors involved in GDM-, but much less information regards their role in OP-associated placental vascular alterations. There is convincing evidence that GDM and OP prone placental endothelium to an “altered metabolic state” leading to fetal programming evidenced at birth, a phenomenon associated with future development of chronic diseases. In this paper it is suggested that this pathological state could be considered as a metabolic marker that could predict occurrence of diseases in adulthood, such as cardiovascular disease, obesity, diabetes mellitus (including gestational diabetes), and metabolic syndrome. PMID:22144986

Decreased endothelial nitric oxide bioavailability, impaired microvascular function, and increased tissue oxygen consumption in children with falciparum malaria.

PubMed

Yeo, Tsin W; Lampah, Daniel A; Kenangalem, Enny; Tjitra, Emiliana; Weinberg, J Brice; Granger, Donald L; Price, Ric N; Anstey, Nicholas M

2014-11-15

Endothelial nitric oxide (NO) bioavailability, microvascular function, and host oxygen consumption have not been assessed in pediatric malaria. We measured NO-dependent endothelial function by using peripheral artery tonometry to determine the reactive hyperemia index (RHI), and microvascular function and oxygen consumption (VO2) using near infrared resonance spectroscopy in 13 Indonesian children with severe falciparum malaria and 15 with moderately severe falciparum malaria. Compared with 19 controls, children with severe malaria and those with moderately severe malaria had lower RHIs (P = .03); 12% and 8% lower microvascular function, respectively (P = .03); and 29% and 25% higher VO2, respectively. RHIs correlated with microvascular function in all children with malaria (P < .001) and all with severe malaria (P < .001). Children with malaria have decreased endothelial and microvascular function and increased oxygen consumption, likely contributing to the pathogenesis of the disease. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Overexpression of hexokinase 2 reduces mitochondrial calcium overload in coronary endothelial cells of type 2 diabetic mice.

PubMed

Pan, Minglin; Han, Ying; Basu, Aninda; Dai, Anzhi; Si, Rui; Willson, Conor; Balistrieri, Angela; Scott, Brian T; Makino, Ayako

2018-03-07

Coronary microvascular rarefaction due to endothelial cell (EC) dysfunction is one of the causes of increased morbidity and mortality in diabetes. Coronary ECs in diabetes are more apoptotic due partly to mitochondrial calcium overload. This study was designed to investigate the role of hexokinase 2 (HK2, an endogenous inhibitor of voltage-dependent anion channel) in coronary endothelial dysfunction in type 2 diabetes. We used mouse coronary ECs (MCECs) isolated from type 2 diabetic mice and human coronary ECs (HCECs) from type 2 diabetic patients to examine protein levels and mitochondrial functions. ECs were more apoptotic and capillary density was lower in the left ventricle of diabetic mice than the control. MCECs from diabetic mice exhibited significant increase in mitochondrial Ca 2+ concentration ([Ca 2+ ] mito ) compared to the control. Among several regulatory proteins for [Ca 2+ ] mito , HK1 and HK2 were significantly lower in MCECs from diabetic mice than control MCECs. We also found that the level of HK2 ubiquitination was higher in MCECs from diabetic mice than in control MCECs. In line with the data from MCECs, HCECs from diabetic patients showed lower HK2 protein levels than HCECs from non-diabetic patients. High-glucose treatment, but not high-fat treatment, significantly decreased HK2 protein levels in the MCEC. HK2 overexpression in MCECs of diabetic mice not only lowered the level of [Ca 2+ ] mito , but also reduced mitochondrial ROS production toward the level seen in control MCECs. These data suggest that HK2 is a potential therapeutic target for coronary microvascular disease in diabetes by restoring mitochondrial function in coronary ECs.

Intravenous Lipid Infusion Induces Endoplasmic Reticulum Stress in Endothelial Cells and Blood Mononuclear Cells of Healthy Adults.

PubMed

Tampakakis, Emmanouil; Tabit, Corey E; Holbrook, Monika; Linder, Erika A; Berk, Brittany D; Frame, Alissa A; Bretón-Romero, Rosa; Fetterman, Jessica L; Gokce, Noyan; Vita, Joseph A; Hamburg, Naomi M

2016-01-11

Endoplasmic reticulum (ER) stress and the subsequent unfolded protein response may initially be protective, but when prolonged, have been implicated in atherogenesis in diabetic conditions. Triglycerides and free fatty acids (FFAs) are elevated in patients with diabetes and may contribute to ER stress. We sought to evaluate the effect of acute FFA elevation on ER stress in endothelial and circulating white cells. Twenty-one healthy subjects were treated with intralipid (20%; 45 mL/h) plus heparin (12 U/kg/h) infusion for 5 hours. Along with increased triglyceride and FFA levels, intralipid/heparin infusion reduced the calf reactive hyperemic response without a change in conduit artery flow-mediated dilation consistent with microvascular dysfunction. To investigate the short-term effects of elevated triglycerides and FFA, we measured markers of ER stress in peripheral blood mononuclear cells (PBMCs) and vascular endothelial cells (VECs). In VECs, activating transcription factor 6 (ATF6) and phospho-inositol requiring kinase 1 (pIRE1) proteins were elevated after infusion (both P<0.05). In PBMCs, ATF6 and spliced X-box-binding protein 1 (XBP-1) gene expression increased by 2.0- and 2.5-fold, respectively (both P<0.05), whereas CHOP and GADD34 decreased by ≈67% and 74%, respectively (both P<0.01). ATF6 and pIRE1 protein levels also increased (both P<0.05), and confocal microscopy revealed the nuclear localization of ATF6 after infusion, suggesting activation. Along with microvascular dysfunction, intralipid infusion induced an early protective ER stress response evidenced by activation of ATF6 and IRE1 in both leukocytes and endothelial cells. Our results suggest a potential link between metabolic disturbances and ER stress that may be relevant to vascular disease. © 2016 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

Biology of portal hypertension.

PubMed

McConnell, Matthew; Iwakiri, Yasuko

2018-02-01

Portal hypertension develops as a result of increased intrahepatic vascular resistance often caused by chronic liver disease that leads to structural distortion by fibrosis, microvascular thrombosis, dysfunction of liver sinusoidal endothelial cells (LSECs), and hepatic stellate cell (HSC) activation. While the basic mechanisms of LSEC and HSC dysregulation have been extensively studied, the role of microvascular thrombosis and platelet function in the pathogenesis of portal hypertension remains to be clearly characterized. As a secondary event, portal hypertension results in splanchnic and systemic arterial vasodilation, leading to the development of a hyperdynamic circulatory syndrome and subsequently to clinically devastating complications including gastroesophageal varices and variceal hemorrhage, hepatic encephalopathy from the formation of portosystemic shunts, ascites, and renal failure due to the hepatorenal syndrome. This review article discusses: (1) mechanisms of sinusoidal portal hypertension, focusing on HSC and LSEC biology, pathological angiogenesis, and the role of microvascular thrombosis and platelets, (2) the mesenteric vasculature in portal hypertension, and (3) future directions for vascular biology research in portal hypertension.

Is endothelial microvascular function equally impaired among patients with chronic Chagas and ischemic cardiomyopathy?

PubMed

Borges, Juliana Pereira; Mendes, Fernanda de Souza Nogueira Sardinha; Lopes, Gabriella de Oliveira; Sousa, Andréa Silvestre de; Mediano, Mauro Felippe Felix; Tibiriçá, Eduardo

2018-08-15

Chronic Chagas cardiomyopathy (CCC) and cardiomyopathies due to other etiologies involve differences in pathophysiological pathways that are still unclear. Systemic microvascular abnormalities are associated with the pathogenesis of ischemic heart disease. However, systemic microvascular endothelial function in CCC remains to be elucidated. Thus, we compared the microvascular endothelial function of patients presenting with CCC to those with ischemic cardiomyopathy disease. Microvascular reactivity was assessed in 21 patients with cardiomyopathy secondary to Chagas disease, 21 patients with cardiomyopathy secondary to ischemic disease and 21 healthy controls. Microvascular blood flow was assessed in the skin of the forearm using laser speckle contrast imaging coupled with iontophoresis of acetylcholine (ACh). Peak increase in forearm blood flow with ACh iontophoresis in relation to baseline was greater in healthy controls than in patients with heart disease (controls: 162.7 ± 58.4% vs. ischemic heart disease: 74.1 ± 48.3% and Chagas: 85.1 ± 68.1%; p < 0.0001). Patients with Chagas and ischemic cardiomyopathy presented similar ACh-induced changes from baseline in skin blood flow (p = 0.55). Endothelial microvascular function was equally impaired among patients with CCC and ischemic cardiomyopathy. Copyright © 2018 Elsevier B.V. All rights reserved.

Flow in the left anterior descending coronary artery in patients with migraine headache.

PubMed

Aslan, Gamze; Sade, Leyla Elif; Yetis, Begum; Bozbas, Huseyin; Eroglu, Serpil; Pirat, Bahar; Can, Ufuk; Muderrisoglu, Haldun

2013-11-15

Migraine is a common neurovascular disorder characterized by attacks of severe headache, autonomic and neurologic symptoms. Migraine can affect many systems in the body, yet its effects on cardiovascular system are unclear. We hypothesized that migraine and coronary microvascular angina may be manifestations of a common systemic microvascular dysfunction and clinically associated. Forty patients with migraine and 35 healthy volunteers were included into the study. Using transthoracic Doppler echocardiography, coronary flow was visualized in the middle or distal part of the left anterior descending artery. Coronary diastolic peak flow velocities were measured with pulse wave Doppler at baseline and after dipyridamole infusion (0.56 mg/kg/4 min). Coronary flow reserve of <2 was considered normal. In addition, thorough 2-dimensional and Doppler echocardiographic examinations were also performed. Fifty-two women and 23 men were included. Coronary flow reserve was significantly lesser in the migraine group than in the control group (1.99 ± 0.3 vs 2.90 ± 0.5, p <0.05). In addition, mitral annular velocities were lower and the ratio of early mitral inflow velocity to early mitral annular velocity (E/E' lateral and E/E' septal) was higher in migraineurs than in the control group (p <0.05 for all), indicating diastolic function abnormalities in the migraine group. In conclusion, these findings suggest that there is an association between coronary microvascular dysfunction and migraine independently of the metabolic state of the patients. A common pathophysiologic pathway of impaired endothelial vasodilatation, vasomotor dysfunction, and increased systemic inflammatory factors may play a role in these 2 clinical conditions and could be the underlying cause of subclinical systolic and diastolic left ventricular dysfunction in migraineurs. Copyright © 2013 Elsevier Inc. All rights reserved.

Dysfunction of annexin A2 contributes to hyperglycaemia-induced loss of human endothelial cell surface fibrinolytic activity.

PubMed

Dai, Haibin; Yu, Zhanyang; Fan, Xiang; Liu, Ning; Yan, Min; Chen, Zhong; Lo, Eng H; Hajjar, Katherine A; Wang, Xiaoying

2013-06-01

Hyperglycaemia impairs fibrinolytic activity on the surface of endothelial cells, but the underlying mechanisms are not fully understood. In this study, we tested the hypothesis that hyperglycaemia causes dysfunction of the endothelial membrane protein annexin A2, thereby leading to an overall reduction of fibrinolytic activity. Hyperglycaemia for 7 days significantly reduced cell surface fibrinolytic activity in human brain microvascular endothelial cells (HBMEC). Hyperglycaemia also decreased tissue type plasminogen activator (t-PA), plasminogen, and annexin A2 mRNA and protein expression, while increasing plasminogen activator inhibitor-1 (PAI-1). No changes in p11 mRNA or protein expression were detected. Hyperglycaemia significantly increased AGE-modified forms of total cellular and membrane annexin A2. The hyperglycemia-associated reduction in fibrinolytic activity was fully restored upon incubation with recombinant annexin A2 (rA2), but not AGE-modified annexin A2 or exogenous t-PA. Hyperglycaemia decreased t-PA, upregulated PAI-1 and induced AGE-related disruption of annexin A2 function, all of which contributed to the overall reduction in endothelial cell surface fibrinolytic activity. Further investigations to elucidate the underlying molecular mechanisms and pathophysiological implications of A2 derivatisation might ultimately lead to a better understanding of mechanisms of impaired vascular fibrinolysis, and to development of new interventional strategies for the thrombotic vascular complications in diabetes.

Dysfunction of annexin A2 contributes to hyperglycaemia-induced loss of human endothelial cell surface fibrinolytic activity

PubMed Central

Dai, Haibin; Yu, Zhanyang; Fan, Xiang; Liu, Ning; Yan, Min; Chen, Zhong; Lo, Eng H.; Hajjar, Katherine A.; Wang, Xiaoying

2014-01-01

Summary Hyperglycaemia impairs fibrinolytic activity on the surface of endothelial cells, but the underlying mechanisms are not fully understood. In this study, we tested the hypothesis that hyperglycaemia causes dysfunction of the endothelial membrane protein annexin A2, thereby leading to an overall reduction of fibrinolytic activity. Hyperglycaemia for 7 days significantly reduced cell surface fibrinolytic activity in human brain microvascular endothelial cells (HBMEC). Hyperglycaemia also decreased tissue type plasminogen activator (t-PA), plasminogen, and annexin A2 mRNA and protein expression, while increasing plasminogen activator inhibitor-1 (PAI-1). No changes in p11 mRNA or protein expression were detected. Hyperglycaemia significantly increased AGE-modified forms of total cellular and membrane annexin A2. The hyperglycemia-associated reduction in fibrinolytic activity was fully restored upon incubation with recombinant annexin A2 (rA2), but not AGE-modified annexin A2 or exogenous t-PA. Hyperglycaemia decreased t-PA, upregulated PAI-1 and induced AGE-related disruption of annexin A2 function, all of which contributed to the overall reduction in endothelial cell surface fibrinolytic activity. Further investigations to elucidate the underlying molecular mechanisms and pathophysiological implications of A2 derivatisation might ultimately lead to a better understanding of mechanisms of impaired vascular fibrinolysis, and to development of new interventional strategies for the thrombotic vascular complications in diabetes. PMID:23572070

Hypoxia/Aglycemia-Induced Endothelial Barrier Dysfunction and Tight Junction Protein Downregulation Can Be Ameliorated by Citicoline

PubMed Central

Pan, Qunwen; Zhao, Yuhui; Chen, Ji; Zhao, Bin; Chen, Yanfang

2013-01-01

This study explores the effect of citicoline on the permeability and expression of tight junction proteins (TJPs) in endothelial cells under hypoxia/aglycemia conditions. Hypoxia or oxygen and glucose deprivation (OGD) was utilized to induce endothelial barrier breakdown model on human umbilical vein endothelial cells (HUVECs) and mouse brain microvascular endothelial cells (bEnd.3s). The effect of citicoline on endothelial barrier breakdown models was determined at either low or high concentrations. FITC-Dextran flux was used to examine the endothelial permeability. The expression of TJPs was measured by immunofluorescence, Real-time PCR and Western Blot methods. Results showed that hypoxia or OGD increased the permeability of HUVECs accompanied with down-regulation of occludens-1 (ZO-1) and occludin at both mRNA and protein levels. Similarly in bEnd.3s, hypoxia increased the permeability and decreased the expression of ZO-1 and claudin-5. Citicoline treatment dose-dependently decreased the permeability in these two models, which paralleled with elevated expression of TJPs. The data demonstrate that citicoline restores the barrier function of endothelial cells compromised by hypoxia/aglycemia probably via up-regulating the expression of TJPs. PMID:24358213

Responses of brain and non-brain endothelial cells to meningitis-causing Escherichia coli K1.

PubMed

Paul-Satyaseela, Maneesh; Xie, Yi; Di Cello, Francescopaolo; Kim, Kwang Sik

2006-03-31

Bacterial interaction with specific host tissue may contribute to its propensity to cause an infection in a particular site. In this study, we examined whether meningitis-causing Escherichia coli K1 interaction with human brain microvascular endothelial cells, which constitute the blood-brain barrier, differed from its interaction with non-brain endothelial cells derived from skin and umbilical cord. We showed that E. coli K1 association was significantly greater with human brain microvascular endothelial cells than with non-brain endothelial cells. In addition, human brain microvascular endothelial cells maintained their morphology and intercellular junctional resistance in response to E. coli K1. In contrast, non-brain endothelial cells exhibited decreased transendothelial electrical resistance and detachment from the matrix upon exposure to E. coli K1. These different responses of brain and non-brain endothelial cells to E. coli K1 may form the basis of E. coli K1's propensity to cause meningitis.

TNF-alpha inhibition could reduce biomarkers of endothelial dysfunction in patients with moderate to severe psoriasis: A 52-week echo-Doppler based quasi-experimental study.

PubMed

Molina-Leyva, Alejandro; Garrido-Pareja, Fermín; Ruiz-Carrascosa, José Carlos; Ruiz-Villaverde, Ricardo

2018-06-22

Psoriasis is associated to endothelial dysfunction, which causes impaired vascular functioning. TNF-α blockers have shown the ability to improve vascular functioning in psoriasis. The nailfold vessel resistance index (NVRI) assesses microvascular functioning at nailfold. The objectives of the study is to assess the effect of the TNF-α inhibition with adalimumab on NVRI. Quasi-experimental study. Fifteen patients with moderate-severe psoriasis received adalimumab 40mg sc according to label information. Participants were assessed at baseline and at 12, 24 and 52 weeks after study intervention. A reduction of -0.09±0.02 (P<.01) in NVRI and a -11.2±2,41ng/ml (P<.001) in E-selectin was observed at week 52. Adalimumab could produce a progressive and sustained reduction of vessel resistance at nailfold and E-selectin in patients with psoriasis. Copyright © 2017 Elsevier España, S.L.U. All rights reserved.

Endurance, interval sprint, and resistance exercise training: impact on microvascular dysfunction in type 2 diabetes

PubMed Central

Laughlin, M. Harold

2015-01-01

Type 2 diabetes (T2D) alters capillary hemodynamics, causes capillary rarefaction in skeletal muscle, and alters endothelial and vascular smooth muscle cell phenotype, resulting in impaired vasodilatory responses. These changes contribute to altered blood flow responses to physiological stimuli, such as exercise and insulin secretion. T2D-induced microvascular dysfunction impairs glucose and insulin delivery to skeletal muscle (and other tissues such as skin and nervous), thereby reducing glucose uptake and perpetuating hyperglycemia and hyperinsulinemia. In patients with T2D, exercise training (EX) improves microvascular vasodilator and insulin signaling and attenuates capillary rarefaction in skeletal muscle. EX-induced changes subsequently augment glucose and insulin delivery as well as glucose uptake. If these adaptions occur in a sufficient amount of tissue, and skeletal muscle in particular, chronic exposure to hyperglycemia and hyperinsulinemia and the risk of microvascular complications in all vascular beds will decrease. We postulate that EX programs that engage as much skeletal muscle mass as possible and recruit as many muscle fibers within each muscle as possible will generate the greatest improvements in microvascular function, providing that the duration of the stimulus is sufficient. Primary improvements in microvascular function occur in tissues (skeletal muscle primarily) engaged during exercise, and secondary improvements in microvascular function throughout the body may result from improved blood glucose control. We propose that the added benefit of combined resistance and aerobic EX programs and of vigorous intensity EX programs is not simply “more is better.” Rather, we believe the additional benefit is the result of EX-induced adaptations in and around more muscle fibers, resulting in more muscle mass and the associated microvasculature being changed. Thus, to acquire primary and secondary improvements in microvascular function and improved blood glucose control, EX programs should involve upper and lower body exercise and modulate intensity to augment skeletal muscle fiber recruitment. Under conditions of limited mobility, it may be necessary to train skeletal muscle groups separately to maximize whole body skeletal muscle fiber recruitment. PMID:26408541

Vascular Function, Insulin Action and Exercise: An Intricate Interplay

PubMed Central

Zheng, Chao; Liu, Zhenqi

2015-01-01

Insulin enhances the compliance of conduit arteries, relaxes resistance arterioles to increase tissue blood flow and dilates precapillary arterioles to expand muscle microvascular blood volume. These actions are impaired in the insulin resistant states. Exercise ameliorates endothelial dysfunction and improves insulin responses in insulin resistant patients, but the precise underlying mechanisms remain unclear. The microvasculature critically regulates insulin action in muscle by modulating insulin delivery to the capillaries nurturing the myocytes and trans-endothelial insulin transport. Recent data suggest that exercise may exert its insulin-sensitizing effect via recruiting muscle microvasculature to increase insulin delivery to and action in muscle. The current review focuses on how the interplay among exercise, insulin action and the vasculature contributes to exercise-mediated insulin sensitization in muscle. PMID:25735473

Late gestational hypoxia and a postnatal high salt diet programs endothelial dysfunction and arterial stiffness in adult mouse offspring.

PubMed

Walton, Sarah L; Singh, Reetu R; Tan, Tiffany; Paravicini, Tamara M; Moritz, Karen M

2016-03-01

Gestational hypoxia and high dietary salt intake have both been associated with impaired vascular function in adulthood. Using a mouse model of prenatal hypoxia, we examined whether a chronic high salt diet had an additive effect in promoting vascular dysfunction in offspring. Pregnant CD1 dams were placed in a hypoxic chamber (12% O2) or housed under normal conditions (21% O2) from embryonic day 14.5 until birth. Gestational hypoxia resulted in a reduced body weight for both male and female offspring at birth. This restriction in body weight persisted until weaning, after which the animals underwent catch-up growth. At 10 weeks of age, a subset of offspring was placed on a high salt diet (5% NaCl). Pressurized myography of mesenteric resistance arteries at 12 months of age showed that both male and female offspring exposed to maternal hypoxia had significantly impaired endothelial function, as demonstrated by impaired vasodilatation to ACh but not sodium nitroprusside. Endothelial dysfunction caused by prenatal hypoxia was not exacerbated by postnatal consumption of a high salt diet. Prenatal hypoxia increased microvascular stiffness in male offspring. The combination of prenatal hypoxia and a postnatal high salt diet caused a leftward shift in the stress-strain relationship in both sexes. Histopathological analysis of aortic sections revealed a loss of elastin integrity and increased collagen, consistent with increased vascular stiffness. These results demonstrate that prenatal hypoxia programs endothelial dysfunction in both sexes. A chronic high salt diet in postnatal life had an additive deleterious effect on vascular mechanics and structural characteristics in both sexes. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

Effects of Riot Control Training on Systemic Microvascular Reactivity and Capillary Density.

PubMed

Pereira, Flavio; de Moraes, Roger; Van Bavel, Diogo; De Lorenzo, Andrea; Tibirica, Eduardo

2018-03-14

The main aim of the present study is to evaluate the effects of strenuous exercise, related to special military training for riot control, on systemic microvascular endothelial function and skin capillary density. Endothelium-dependent microvascular reactivity was evaluated in the forearm skin of healthy military trainees (age 23.4 ± 2.3 yr; n = 15) using laser speckle contrast imaging coupled with cutaneous acetylcholine (ACh) iontophoresis and post-occlusive reactive hyperemia (PORH). Functional capillary density was assessed using high-resolution, intra-vital color microscopy in the dorsum of the middle phalanx. Capillary recruitment (capillary reserve) was evaluated using PORH. Microcirculatory tests were performed before and after a 5-wk special military training for riot control. Microvascular endothelium-dependent vasodilatory responses were markedly and significantly reduced after training, compared with values obtained before training. The peak values of microvascular conductance obtained during iontophoresis of ACh or PORH before training (0.84 ± 0.22 and 0.94 ± 0.72 APU/mmHg, respectively) were markedly reduced after training (0.47 ± 0.11 and 0.71 ± 0.14 APU/mmHg; p < 0.0001 and p = 0.0037, respectively). Endothelium-dependent capillary recruitment was significantly reduced after training (before 101 ± 9 and after 95 ± 8 capillaries/mm2; p = 0.0007). The present study showed that a 5-wk strenuous military training, performed in unfavorable climatic conditions, induces marked systemic microvascular dysfunction, mainly characterized by reduced endothelium-dependent microvascular vasodilation and blunted capillary recruitment.

LPS causes pericyte loss and microvascular dysfunction via disruption of Sirt3/angiopoietins/Tie-2 and HIF-2α/Notch3 pathways.

PubMed

Zeng, Heng; He, Xiaochen; Tuo, Qin-Hui; Liao, Duan-Fang; Zhang, Guo-Qiang; Chen, Jian-Xiong

2016-02-12

Recent studies reveal a crucial role of pericyte loss in sepsis-associated microvascular dysfunction. Sirtuin 3 (SIRT3) mediates histone protein post-translational modification related to aging and ischemic disease. This study investigated the involvement of SIRT3 in LPS-induced pericyte loss and microvascular dysfunction. Mice were exposed to LPS, expression of Sirt3, HIF-2α, Notch3 and angiopoietins/Tie-2, pericyte/endothelial (EC) coverage and vascular permeability were assessed. Mice treated with LPS significantly reduced the expression of SIRT3, HIF-2α and Notch3 in the lung. Furthermore, exposure to LPS increased Ang-2 while inhibited Ang-1/Tie-2 expression with a reduced pericyte/EC coverage. Intriguingly, knockout of Sirt3 upregulated Ang-2, but downregulated Tie-2 and HIF-2α/Notch3 expression which resulted in a dramatic reduction of pericyte/EC coverage and exacerbation of LPS-induced vascular leakage. Conversely, overexpression of Sirt3 reduced Ang-2 expression and increased Ang-1/Tie-2 and HIF-2α/Notch3 expression in the LPS treated mice. Overexpression of Sirt3 further prevented LPS-induced pericyte loss and vascular leakage. This was accompanied by a significant reduction of the mortality rate. Specific knockout of prolyl hydroxylase-2 (PHD2) increased HIF-2α/Notch3 expression, improved pericyte/EC coverage and reduced the mortality rate in the LPS-treated mice. Our study demonstrates the importance of SIRT3 in preserving vascular integrity by targeting pericytes in the setting of LPS-induced sepsis.

The Decay of Stem Cell Nourishment at the Niche

PubMed Central

de Mora, Jaime Font

2013-01-01

Abstract One of the main features of human aging is the loss of adult stem cell homeostasis. Organs that are very dependent on adult stem cells show increased susceptibility to aging, particularly organs that present a vascular stem cell niche. Reduced regenerative capacity in tissues correlates with reduced stem cell function, which parallels a loss of microvascular density (rarefraction) and plasticity. Moreover, the age-related loss of microvascular plasticity and rarefaction has significance beyond metabolic support for tissues because stem cell niches are regulated co-ordinately with the vascular cells. In addition, microvascular rarefaction is related to increased inflammatory signals that may negatively regulate the stem cell population. Thus, the processes of microvascular rarefaction, adult stem cell dysfunction, and inflammation underlie the cycle of physiological decline that we call aging. Observations from new mouse models and humans are discussed here to support the vascular aging theory. We develop a novel theory to explain the complexity of aging in mammals and perhaps in other organisms. The connection between vascular endothelial tissue and organismal aging provides a potential evolutionary conserved mechanism that is an ideal target for the development of therapies to prevent or delay age-related processes in humans. PMID:23937078

Cardioprotective effects of red wine and vodka in a model of endothelial dysfunction

PubMed Central

Lassaletta, Antonio D; Chu, Louis M; Elmadhun, Nassrene Y; Burgess, Thomas A; Feng, Jun; Robich, Michael P; Sellke, Frank W

2012-01-01

Background Moderate alcohol consumption is largely believed to be cardioprotective, while red wine is hypothesized to offer benefit in part due to the pro-angiogenic and antioxidant properties of polyphenols. We investigated the cardiovascular effects of both red wine and vodka in a swine model of endothelial dysfunction. Methods Twenty-seven male Yorkshire swine fed a high-fat/cholesterol diet were divided into three groups and received either no alcohol (Control), red wine, or vodka. After seven weeks, myocardial perfusion was measured, and ventricular tissue was analyzed for microvascular reactivity, and immunohistochemical studies. Results There were no differences in myocardial perfusion, in arteriolar or capillary density, or in VEGF expression among groups. Total protein oxidation as well as expression of superoxide dismutase-1 and -2 (SOD1, SOD2) and NADPH-oxidase (NOX2) was decreased in both treatment groups compared to controls. Endothelium-dependent microvessel relaxation, however, was significantly improved only in the red wine-supplemented group. Conclusions Supplementation with both red wine and vodka decreased oxidative stress by several measures, implicating the effects of ethanol in reducing oxidative stress in the myocardium. However, it was only in the red wine-supplemented group that an improvement in microvessel function was observed. This suggests that a component of red wine, independent of ethanol, possibly a polyphenol such as resveratrol, may confer cardioprotection by normalizing endothelial dysfunction induced by an atherogenic diet. PMID:22748601

Digital ulcers in systemic sclerosis: role of flow-mediated dilatation and capillaroscopy as risk assessment tools.

PubMed

Silva, Ivone; Loureiro, Tiago; Teixeira, Andreia; Almeida, Isabel; Mansilha, Armando; Vasconcelos, Carlos; Almeida, Rui

2015-01-01

The aim of this study was to evaluate macrovascular endothelial dysfunction and microvascular damage as clinical markers of peripheral microangiopathy in patients with Raynaud's phenomenon (RP). Seventy-seven secondary RP with systemic sclerosis, 32 primary RP and 34 healthy controls were included in our study. Secondary RP patients were divided into two subgroups: 39 with digital ulcers (DU) and 38 without digital ulcers (non-DU). Patients with DU had significantly lower flow-mediated dilatation values (5.34 ± 7.49%) compared to non-DU patients (16.21 ± 11.31%), primary RP (17.96 ± 12.78%) and controls (20.17 ± 8.86%), p<0.001, favouring macrovascular endothelium dysfunction. Regarding microvascular damage, the DU group had a predominately capillaroscopic late pattern (71.1%) whereas non-DU patients had an active pattern (56.4%). The microangiopathy evolution score was significantly higher in the DU group compared to the non-DU group (4.79 ± 1.82 vs. 1.79 ± 1.56, p<0.001). Flow-mediated dilation was significantly lower in late pattern (6.13 ± 7.09%) compared to active (12.58 ± 10.66%) and early patterns (17.72 ± 14.90%), p = 0.016 and p = 0.044 respectively. Low flow-mediated dilatation and microvascular damage in capillaroscopy are early clinical markers of DU risk in RP patients.

Telomerase reverse transcriptase protects against angiotensin II-induced microvascular endothelial dysfunction.

PubMed

Ait-Aissa, Karima; Kadlec, Andrew O; Hockenberry, Joseph; Gutterman, David D; Beyer, Andreas M

2018-05-01

A rise in reactive oxygen species (ROS) may contribute to cardiovascular disease by reducing nitric oxide (NO) levels, leading to loss of NO's vasodilator and anti-inflammatory effects. Although primarily studied in larger conduit arteries, excess ROS release and a corresponding loss of NO also occur in smaller resistance arteries of the microcirculation, but the underlying mechanisms and therapeutic targets have not been fully characterized. We examined whether either of the two subunits of telomerase, telomerase reverse transcriptase (TERT) or telomerase RNA component (TERC), affect microvascular ROS production and peak vasodilation at baseline and in response to in vivo administration to angiotensin II (ANG II). We report that genetic loss of TERT [maximal dilation: 52.0 ± 6.1% with vehicle, 60.4 ± 12.9% with N ω -nitro-l-arginine methyl ester (l-NAME), and 32.2 ± 12.2% with polyethylene glycol-catalase (PEG-Cat) ( P < 0.05), means ± SD, n = 9-19] but not TERC [maximal dilation: 79 ± 5% with vehicle, 10.7 ± 9.8% with l-NAME ( P < 0.05), and 86.4 ± 8.4% with PEG-Cat, n = 4-7] promotes flow-induced ROS formation. Moreover, TERT knockout exacerbates the microvascular dysfunction resulting from in vivo ANG II treatment, whereas TERT overexpression is protective [maximal dilation: 88.22 ± 4.6% with vehicle vs. 74.0 ± 7.3% with ANG II (1,000 ng·kg -1 ·min -1 ) ( P = not significant), n = 4]. Therefore, loss of TERT but not TERC may be a key contributor to the elevated microvascular ROS levels and reduced peak dilation observed in several cardiovascular disease pathologies. NEW & NOTEWORTHY This study identifies telomerase reverse transcriptase (TERT) but not telomerase RNA component as a key factor regulating endothelium-dependent dilation in the microcirculation. Loss of TERT activity leads to microvascular dysfunction but not conduit vessel dysfunction in first-generation mice. In contrast, TERT is protective in the microcirculation in the presence of prolonged vascular stress. Understanding the mechanism of how TERT protects against vascular stress represents a novel target for the treatment of vascular disorders.

Protective Effect of Bendavia (SS-31) Against Oxygen/Glucose-Deprivation Stress-Induced Mitochondrial Damage in Human Brain Microvascular Endothelial Cells.

PubMed

Imai, Takahiko; Mishiro, Keisuke; Takagi, Toshinori; Isono, Aoi; Nagasawa, Hideko; Tsuruma, Kazuhiro; Shimazawa, Masamitsu; Hara, Hideaki

2017-01-01

Mitochondria play a key role in cell survival by perfoming functions such as adenosine tri-phosphate (ATP) synthesis, regulation of apoptotic cell death, calcium storage. Hypoxic conditions induce mitochondrial dysfunction, which leads to endothelial injury in cerebral ischemia. Functional disorders include the following: collapse of mitochondrial membrane potential, reduction of ATP synthesis, and generation of reactive oxygen species (ROS). Bendavia, a novel tetra-peptide, has been reported to restrict the uncoupling of the mitochondrial membrane chain, protect the synthesis of ATP, and inhibit ROS generation. In the present study, we investigated whether bendavia protects mitochondria under hypoxic and starved conditions by using human brain microvascular endothelial cells (HBMVECs). After pre-treatment with bendavia, we exposed HBMVECs to oxygen glucose deprivation (OGD) for 6 h. We then assessed cell viability, the level of caspase-3/7 activity, ROS generation, mitochondrial membrane potential, ATP contents, and the number of mitochondria. Bendavia recovered cell viability and reduced the caspase-3/7 activity induced by OGDinduced damage. Bendavia also recovered mitochondrial functions. These results suggest that bendavia protects mitochondrial function against OGD-induced injury and inhibits apoptosis in HBMVECs. Consequently, our findings indicate that bendavia might become the new therapeutic drug of choice to target mitochondria in case of cerebral ischemia. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Influenza Infects Lung Microvascular Endothelium Leading to Microvascular Leak: Role of Apoptosis and Claudin-5

PubMed Central

Armstrong, Susan M.; Wang, Changsen; Tigdi, Jayesh; Si, Xiaoe; Dumpit, Carlo; Charles, Steffany; Gamage, Asela; Moraes, Theo J.; Lee, Warren L.

2012-01-01

Severe influenza infections are complicated by acute lung injury, a syndrome of pulmonary microvascular leak. The pathogenesis of this complication is unclear. We hypothesized that human influenza could directly infect the lung microvascular endothelium, leading to loss of endothelial barrier function. We infected human lung microvascular endothelium with both clinical and laboratory strains of human influenza. Permeability of endothelial monolayers was assessed by spectrofluorimetry and by measurement of the transendothelial electrical resistance. We determined the molecular mechanisms of flu-induced endothelial permeability and developed a mouse model of severe influenza. We found that both clinical and laboratory strains of human influenza can infect and replicate in human pulmonary microvascular endothelium, leading to a marked increase in permeability. This was caused by apoptosis of the lung endothelium, since inhibition of caspases greatly attenuated influenza-induced endothelial leak. Remarkably, replication-deficient virus also caused a significant degree of endothelial permeability, despite displaying no cytotoxic effects to the endothelium. Instead, replication-deficient virus induced degradation of the tight junction protein claudin-5; the adherens junction protein VE-cadherin and the actin cytoskeleton were unaffected. Over-expression of claudin-5 was sufficient to prevent replication-deficient virus-induced permeability. The barrier-protective agent formoterol was able to markedly attenuate flu-induced leak in association with dose-dependent induction of claudin-5. Finally, mice infected with human influenza developed pulmonary edema that was abrogated by parenteral treatment with formoterol. Thus, we describe two distinct mechanisms by which human influenza can induce pulmonary microvascular leak. Our findings have implications for the pathogenesis and treatment of acute lung injury from severe influenza. PMID:23115643

Blood transfusion improves renal oxygenation and renal function in sepsis-induced acute kidney injury in rats.

PubMed

Zafrani, Lara; Ergin, Bulent; Kapucu, Aysegul; Ince, Can

2016-12-20

The effects of blood transfusion on renal microcirculation during sepsis are unknown. This study aimed to investigate the effect of blood transfusion on renal microvascular oxygenation and renal function during sepsis-induced acute kidney injury. Twenty-seven Wistar albino rats were randomized into four groups: a sham group (n = 6), a lipopolysaccharide (LPS) group (n = 7), a LPS group that received fluid resuscitation (n = 7), and a LPS group that received blood transfusion (n = 7). The mean arterial blood pressure, renal blood flow, and renal microvascular oxygenation within the kidney cortex were recorded. Acute kidney injury was assessed using the serum creatinine levels, metabolic cost, and histopathological lesions. Nitrosative stress (expression of endothelial (eNOS) and inducible nitric oxide synthase (iNOS)) within the kidney was assessed by immunohistochemistry. Hemoglobin levels, pH, serum lactate levels, and liver enzymes were measured. Fluid resuscitation and blood transfusion both significantly improved the mean arterial pressure and renal blood flow after LPS infusion. Renal microvascular oxygenation, serum creatinine levels, and tubular damage significantly improved in the LPS group that received blood transfusion compared to the group that received fluids. Moreover, the renal expression of eNOS was markedly suppressed under endotoxin challenge. Blood transfusion, but not fluid resuscitation, was able to restore the renal expression of eNOS. However, there were no significant differences in lactic acidosis or liver function between the two groups. Blood transfusion significantly improved renal function in endotoxemic rats. The specific beneficial effect of blood transfusion on the kidney could have been mediated in part by the improvements in renal microvascular oxygenation and sepsis-induced endothelial dysfunction via the restoration of eNOS expression within the kidney.

Advanced Glycation End-Products Induce Apoptosis in Pancreatic Islet Endothelial Cells via NF-κB-Activated Cyclooxygenase-2/Prostaglandin E2 Up-Regulation

PubMed Central

Lan, Kuo-Cheng; Chiu, Chen-Yuan; Kao, Chia-Wei; Huang, Kuo-How; Wang, Ching-Chia; Huang, Kuo-Tong; Tsai, Keh-Sung

2015-01-01

Microvascular complications eventually affect nearly all patients with diabetes. Advanced glycation end-products (AGEs) resulting from hyperglycemia are a complex and heterogeneous group of compounds that accumulate in the plasma and tissues in diabetic patients. They are responsible for both endothelial dysfunction and diabetic vasculopathy. The aim of this study was to investigate the cytotoxicity of AGEs on pancreatic islet microvascular endothelial cells. The mechanism underlying the apoptotic effect of AGEs in pancreatic islet endothelial cell line MS1 was explored. The results showed that AGEs significantly decreased MS1 cell viability and induced MS1 cell apoptosis in a dose-dependent manner. AGEs dose-dependently increased the expressions of cleaved caspase-3, and cleaved poly (ADP-ribose) polymerase in MS1 cells. Treatment of MS1 cells with AGEs also resulted in increased nuclear factor (NF)-κB-p65 phosphorylation and cyclooxygenase (COX)-2 expression. However, AGEs did not affect the expressions of endoplasmic reticulum (ER) stress-related molecules in MS1 cells. Pretreatment with NS398 (a COX-2 inhibitor) to inhibit prostaglandin E2 (PGE2) production reversed the induction of cleaved caspase-3, cleaved PARP, and MS1 cell viability. Moreover, AGEs significantly increased the receptor for AGEs (RAGE) protein expression in MS1 cells, which could be reversed by RAGE neutralizing antibody. RAGE Neutralizing antibody could also reverse the induction of cleaved caspase-3 and cleaved PARP and decreased cell viability induced by AGEs. These results implicate the involvement of NF-κB-activated COX-2/PGE2 up-regulation in AGEs/RAGE-induced islet endothelial cell apoptosis and cytotoxicity. These findings may provide insight into the pathological processes within the pancreatic islet microvasculature induced by AGEs accumulation. PMID:25898207

Endothelial cell culture in microfluidic devices for investigating microvascular processes.

PubMed

Mannino, Robert G; Qiu, Yongzhi; Lam, Wilbur A

2018-07-01

Numerous conditions and disease states such as sickle cell disease, malaria, thrombotic microangiopathy, and stroke significantly impact the microvasculature function and its role in disease progression. Understanding the role of cellular interactions and microvascular hemodynamic forces in the context of disease is crucial to understanding disease pathophysiology. In vivo models of microvascular disease using animal models often coupled with intravital microscopy have long been utilized to investigate microvascular phenomena. However, these methods suffer from some major drawbacks, including the inability to tightly and quantitatively control experimental conditions, the difficulty of imaging multiple microvascular beds within a living organism, and the inability to isolate specific microvascular geometries such as bifurcations. Thus, there exists a need for in vitro microvascular models that can mitigate the drawbacks associated with in vivo systems. To that end, microfluidics has been widely used to develop such models, as it allows for tight control of system inputs, facile imaging, and the ability to develop robust and repeatable systems with well-defined geometries. Incorporating endothelial cells to branching microfluidic models allows for the development of "endothelialized" systems that accurately recapitulate physiological microvessels. In this review, we summarize the field of endothelialized microfluidics, specifically focusing on fabrication methods, limitations, and applications of these systems. We then speculate on future directions and applications of these cutting edge technologies. We believe that this review of the field is of importance to vascular biologists and bioengineers who aim to utilize microfluidic technologies to solve vascular problems.

Capsule independent uptake of the fungal pathogen Cryptococcus neoformans into brain microvascular endothelial cells.

PubMed

Sabiiti, Wilber; May, Robin C

2012-01-01

Cryptococcosis is a life-threatening fungal disease with a high rate of mortality among HIV/AIDS patients across the world. The ability to penetrate the blood-brain barrier (BBB) is central to the pathogenesis of cryptococcosis, but the way in which this occurs remains unclear. Here we use both mouse and human brain derived endothelial cells (bEnd3 and hCMEC/D3) to accurately quantify fungal uptake and survival within brain endothelial cells. Our data indicate that the adherence and internalisation of cryptococci by brain microvascular endothelial cells is an infrequent event involving small numbers of cryptococcal yeast cells. Interestingly, this process requires neither active signalling from the fungus nor the presence of the fungal capsule. Thus entry into brain microvascular endothelial cells is most likely a passive event that occurs following 'trapping' within capillary beds of the BBB.

Capsule Independent Uptake of the Fungal Pathogen Cryptococcus neoformans into Brain Microvascular Endothelial Cells

PubMed Central

Sabiiti, Wilber; May, Robin C.

2012-01-01

Cryptococcosis is a life-threatening fungal disease with a high rate of mortality among HIV/AIDS patients across the world. The ability to penetrate the blood-brain barrier (BBB) is central to the pathogenesis of cryptococcosis, but the way in which this occurs remains unclear. Here we use both mouse and human brain derived endothelial cells (bEnd3 and hCMEC/D3) to accurately quantify fungal uptake and survival within brain endothelial cells. Our data indicate that the adherence and internalisation of cryptococci by brain microvascular endothelial cells is an infrequent event involving small numbers of cryptococcal yeast cells. Interestingly, this process requires neither active signalling from the fungus nor the presence of the fungal capsule. Thus entry into brain microvascular endothelial cells is most likely a passive event that occurs following ‘trapping’ within capillary beds of the BBB. PMID:22530025

Inhibition of caspase activity prevents CD95-mediated hepatic microvascular perfusion failure and restores Kupffer cell clearance capacity.

PubMed

Wanner, G A; Mica, L; Wanner-Schmid, E; Kolb, S A; Hentze, H; Trentz, O; Ertel, W

1999-07-01

Using a murine model, we studied the effect of agonistic anti-CD95 antibodies (aCD95) on sinusoidal lining cells and a potential protection by caspase inhibition. C3H/HeN mice were intravenously administered aCD95 (10 microgram/mouse) or unspecific IgG (control) in the presence or absence of the caspase inhibitor z-VAD-fmk. Analysis of hepatic microcirculation using intravital fluorescence microscopy revealed severe (P<0.01) sinusoidal perfusion failure and reduced (P<0.05) phagocytic activity of Kupffer cells (KC) within 2 h. Transmission electron micrographs demonstrated loss of integrity of sinusoidal endothelial cells as early as 1 h after aCD95 application, whereas histological manifestation of hepatocellular apoptosis and hemorrhagic necrosis was most pronounced at 6 h. Blocking of caspase activity attenuated (P<0.01) both hepatic microvascular perfusion failure and KC dysfunction. Accordingly, full protection of the liver from apoptotic damage and intact microarchitecture was observed in histological sections after z-VAD-fmk treatment. Mortality rate was 40% 6 h after aCD95 administration, whereas all animals survived in the z-VAD-fmk group (P<0.05). The activation of caspases through CD95 may primarily lead to damage of sinusoidal endothelial cells and hepatic microvascular perfusion failure. Moreover, reduced phagocytic capacity of KC may contribute to accumulation of toxic metabolites released by dying cells at the local site of inflammation, further aggravating liver injury.

Diabetes and sexual dysfunction: current perspectives

PubMed Central

Maiorino, Maria Ida; Bellastella, Giuseppe; Esposito, Katherine

2014-01-01

Diabetes mellitus is one of the most common chronic diseases in nearly all countries. It has been associated with sexual dysfunction, both in males and in females. Diabetes is an established risk factor for sexual dysfunction in men, as a threefold increased risk of erectile dysfunction was documented in diabetic men, as compared with nondiabetic men. Among women, evidence regarding the association between diabetes and sexual dysfunction are less conclusive, although most studies have reported a higher prevalence of female sexual dysfunction in diabetic women as compared with nondiabetic women. Female sexual function appears to be more related to social and psychological components than to the physiological consequence of diabetes. Hyperglycemia, which is a main determinant of vascular and microvascular diabetic complications, may participate in the pathogenetic mechanisms of sexual dysfunction in diabetes. Moreover, diabetic people may present several clinical conditions, including hypertension, overweight and obesity, metabolic syndrome, cigarette smoking, and atherogenic dyslipidemia, which are themselves risk factors for sexual dysfunction, both in men and in women. The adoption of healthy lifestyles may reduce insulin resistance, endothelial dysfunction, and oxidative stress – all of which are desirable achievements in diabetic patients. Improved well-being may further contribute to reduce and prevent sexual dysfunction in both sexes. PMID:24623985

Sex Differences Influencing Micro- and Macrovascular Endothelial Phenotype In Vitro.

PubMed

Huxley, Virginia H; Kemp, Scott S; Schramm, Christine; Sieveking, Steve; Bingaman, Susan; Yu, Yang; Zaniletti, Isabella; Stockard, Kevin; Wang, Jianjie

2018-06-09

Endothelial dysfunction is an early hallmark of multiple disease states that also display sex differences with respect to age of onset, frequency, and severity. Results of in vivo studies of basal and stimulated microvascular barrier function revealed sex differences difficult to ascribe to specific cells or environmental factors. The present study evaluated endothelial cells (EC) isolated from macro- and/or microvessels of reproductively mature rats under the controlled conditions of low-passage culture to test the assumption that EC phenotype would be sex-independent. The primary finding was that EC, regardless of where they are derived, retain a sex-bias in low-passage culture, independent of varying levels of reproductive hormones. Implications of the work include the fallacy of expecting a universal set of mechanisms derived from study of EC from one sex and/or one vascular origin to apply uniformly to all EC under unstimulated conditions no less in the disease state. Vascular endothelial cells (EC) are heterogeneous with respect to phenotype reflecting at least organ of origin, location within the vascular network, and physical forces. Sex, as an independent influence on EC functions in health or etiology, susceptibility, and progression of dysfunction in numerous disease states, has been largely ignored. The current study focussed on EC isolated from aorta (macrovascular) and skeletal muscle vessels (microvascular) of age-matched male and female rats under identical conditions of short term (passage 4) culture. We tested the hypothesis that genomic sex would not influence endothelial growth, wound healing, morphology, lactate production, or messenger RNA and protein expression of key proteins (sex hormone receptors for androgen (AR) and oestrogen (ERα and ERβ); PECAM-1 and VE-CAD mediating barrier function; α v β 3 and N-Cadherin influencing matrix interactions; ICAM-1 and VCAM-1 mediating EC/white cell adhesion). The hypothesis was rejected as EC origin (macro- versus microvessel) and sex influenced multiple phenotypic characteristics. Statistical model analysis of EC growth demonstrated an hierarchy of variable importance, recapitulated for other phenotypic characteristics, wherein predictions assuming EC homogeneity < Sex < Vessel Origin < Sex and Vessel Origin. Further, patterns of EC mRNA expression by vessel origin and by sex did not predict protein expression. Overall the study demonstrated that accurate assessment of sex-linked EC dysfunction first requires understanding of EC function by position in the vascular tree and by sex. Results from a single EC tissue source/species/sex cannot provide universal insight into the mechanisms regulating in vivo endothelial function in health, no less disease. (250) This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Time of correlation of low-frequency fluctuations in the regional laser Doppler flow signal from human skin

NASA Astrophysics Data System (ADS)

Folgosi-Correa, M. S.; Nogueira, G. E. C.

2012-06-01

The laser Doppler flowmetry allows the non-invasive assessment of the skin perfusion in real-time, being an attractive technique to study the human microcirculation in clinical settings. Low-frequency oscillations in the laser Doppler blood flow signal from the skin have been related to the endothelial, endothelial-metabolic, neurogenic and myogenic mechanisms of microvascular flow control, in the range 0.005-0.0095 Hz, 0.0095-0.021 Hz, 0.021-0.052 Hz and 0.052- 0.145 Hz respectively. The mean Amplitude (A) of the periodic fluctuations in the laser Doppler blood flow signal, in each frequency range, derived from the respective wavelet-transformed coefficients, has been used to assess the function and dysfunctions of each mechanism of flow control. Known sources of flow signal variances include spatial and temporal variability, diminishing the discriminatory capability of the technique. Here a new time domain method of analysis is proposed, based on the Time of Correlation (TC) of flow fluctuations between two adjacent sites. Registers of blood flow from two adjacent regions, for skin temperature at 32 0C (basal) and thermally stimulated (42 0C) of volar forearms from 20 healthy volunteers were collected and analyzed. The results obtained revealed high time of correlation between two adjacent regions when thermally stimulated, for signals in the endothelial, endothelial-metabolic, neurogenic and myogenic frequency ranges. Experimental data also indicate lower variability for TC when compared to A, when thermally stimulated, suggesting a new promising parameter for assessment of the microvascular flow control.

Tumor Response to Radiotherapy Regulated by Endothelial Cell Apoptosis

NASA Astrophysics Data System (ADS)

Garcia-Barros, Monica; Paris, Francois; Cordon-Cardo, Carlos; Lyden, David; Rafii, Shahin; Haimovitz-Friedman, Adriana; Fuks, Zvi; Kolesnick, Richard

2003-05-01

About 50% of cancer patients receive radiation therapy. Here we investigated the hypothesis that tumor response to radiation is determined not only by tumor cell phenotype but also by microvascular sensitivity. MCA/129 fibrosarcomas and B16F1 melanomas grown in apoptosis-resistant acid sphingomyelinase (asmase)-deficient or Bax-deficient mice displayed markedly reduced baseline microvascular endothelial apoptosis and grew 200 to 400% faster than tumors on wild-type microvasculature. Thus, endothelial apoptosis is a homeostatic factor regulating angiogenesis-dependent tumor growth. Moreover, these tumors exhibited reduced endothelial apoptosis upon irradiation and, unlike tumors in wild-type mice, they were resistant to single-dose radiation up to 20 grays (Gy). These studies indicate that microvascular damage regulates tumor cell response to radiation at the clinically relevant dose range.

Anti-TNFα therapy transiently improves high density lipoprotein cholesterol levels and microvascular endothelial function in patients with rheumatoid arthritis: a Pilot Study

PubMed Central

2012-01-01

Background Rheumatoid arthritis (RA) is associated with increased morbidity and mortality from cardiovascular disease (CVD). This can be only partially attributed to traditional CVD risk factors such as dyslipidaemia and their downstream effects on endothelial function. The most common lipid abnormality in RA is reduced levels of high-density lipoprotein (HDL) cholesterol, probably due to active inflammation. In this longitudinal study we hypothesised that anti-tumor necrosis factor-α (anti-TNFα) therapy in patients with active RA improves HDL cholesterol, microvascular and macrovascular endothelial function. Methods Twenty-three RA patients starting on anti-TNFα treatment were assessed for HDL cholesterol level, and endothelial-dependent and -independent function of microvessels and macrovessels at baseline, 2-weeks and 3 months of treatment. Results Disease activity (CRP, fibrinogen, DAS28) significantly decreased during the follow-up period. There was an increase in HDL cholesterol levels at 2 weeks (p < 0.05) which was paralleled by a significant increase in microvascular endothelial-dependent function (p < 0.05). However, both parameters returned towards baseline at 12 weeks. Conclusion Anti-TNFα therapy in RA patients appears to be accompanied by transient but significant improvements in HDL cholesterol levels, which coexists with an improvement in microvascular endothelial-dependent function. PMID:22824166

Disparate effects of single endothelin A and B receptor blocker therapy on the progression of renal injury in advanced renovascular disease

PubMed Central

Chade, Alejandro R.; Stewart, Nicholas J.; Peavy, Patrick R.

2013-01-01

We hypothesized that chronic specific endothelin (ET)-A receptor blockade therapy would reverse renal dysfunction and injury in advanced experimental renovascular disease. To test this, unilateral renovascular disease was induced in 19 pigs and after 6 weeks, single-kidney hemodynamics and function was quantified in vivo using computed-tomography. All pigs with renovascular disease were divided such that 7 were untreated, 7 were treated with ET-A blockers, and 5 were treated with ET-B blockers. Four weeks later, all pigs were re-studied in vivo, then euthanized and ex vivo studies performed on the stenotic kidney to quantify microvascular density, remodeling, renal oxidative stress, inflammation, and fibrosis. RBF, GFR, and redox status were significantly improved in the stenotic kidney after ET-A but not ET-B blockade. Furthermore, only ET-A blockade therapy reversed renal microvascular rarefaction and diminished remodeling, which was accompanied by a marked decreased in renal inflammatory and fibrogenic activity. Thus, ET-A but not ET-B blockade ameliorated renal injury in pigs with advanced renovascular disease by stimulating microvascular proliferation and decreasing the progression of microvascular remodeling, renal inflammation and fibrosis in the stenotic kidney. These effects were functionally consequential since ET-A blockade improved single kidney microvascular endothelial function, RBF, and GFR, and decreased albuminuria. PMID:24352153

Low intensity shear stress increases endothelial ELR+ CXC chemokine production via a focal adhesion kinase-p38{beta} MAPK-NF-{kappa}B pathway.

PubMed

Shaik, Sadiq S; Soltau, Thomas D; Chaturvedi, Gaurav; Totapally, Balagangadhar; Hagood, James S; Andrews, William W; Athar, Mohammad; Voitenok, Nikolai N; Killingsworth, Cheryl R; Patel, Rakesh P; Fallon, Michael B; Maheshwari, Akhil

2009-02-27

CXC chemokines with a glutamate-leucine-arginine (ELR) tripeptide motif (ELR(+) CXC chemokines) play an important role in leukocyte trafficking into the tissues. For reasons that are not well elucidated, circulating leukocytes are recruited into the tissues mainly in small vessels such as capillaries and venules. Because ELR(+) CXC chemokines are important mediators of endothelial-leukocyte interaction, we compared chemokine expression by microvascular and aortic endothelium to investigate whether differences in chemokine expression by various endothelial types could, at least partially, explain the microvascular localization of endothelial-leukocyte interaction. Both in vitro and in vivo models indicate that ELR(+) CXC chemokine expression is higher in microvascular endothelium than in aortic endothelial cells. These differences can be explained on the basis of the preferential activation of endothelial chemokine production by low intensity shear stress. Low shear activated endothelial ELR(+) CXC chemokine production via cell surface heparan sulfates, beta(3)-integrins, focal adhesion kinase, the mitogen-activated protein kinase p38beta, mitogen- and stress-associated protein kinase-1, and the transcription factor.

Inhibition of c-Src protects paraquat induced microvascular endothelial injury by modulating caveolin-1 phosphorylation and caveolae mediated transcellular permeability.

PubMed

Huang, Yu; He, Qing

2017-06-01

The mechanisms underlying paraquat induced acute lung injury (ALI) is still not clear. C-Src plays an important role in the regulation of microvascular endothelial barrier function and the pathogenesis of ALI. In the present study, we found that paraquat induced cell toxicity and an increase of reactive oxygen species (ROS) in endothelium. Paraquat exposure also induced significant increase of caveolin-1 phosphorylation, caveolae trafficking and albumin permeability in endothelial monolayers. C-Src depletion by siRNA significantly attenuate paraquat induced cell toxicity, caveolin-1 phosphorylation, caveolae formation and endothelial hyperpermeability. N-acetylcysteine (NAC) failed to protect endothelial monolayers against paraquat induced toxicity. Thus, our findings suggest that paraquat exposure increases paracellular endothelial permeability by increasing caveolin-1 phosphorylation in a c-Src dependant manner. The depletion of c-Src might protect microvascular endothelial function by regulating caveolin-1 phosphorylation and caveolae trafficking during paraquat exposure, and might have potential therapeutic effects on paraquat induced ALI. Copyright © 2017 Elsevier B.V. All rights reserved.

Vascular complications in diabetes: Microparticles and microparticle associated microRNAs as active players.

PubMed

Alexandru, Nicoleta; Badila, Elisabeta; Weiss, Emma; Cochior, Daniel; Stępień, Ewa; Georgescu, Adriana

2016-03-25

The recognition of the importance of diabetes in vascular disease has greatly increased lately. Common risk factors for diabetes-related vascular disease include hyperglycemia, insulin resistance, dyslipidemia, inflammation, hypercoagulability, hypertension, and atherosclerosis. All of these factors contribute to the endothelial dysfunction which generates the diabetic complications, both macro and microvascular. Knowledge of diabetes-related vascular complications and of associated mechanisms it is becoming increasingly important for therapists. The discovery of microparticles (MPs) and their associated microRNAs (miRNAs) have opened new perspectives capturing the attention of basic and clinical scientists for their potential to become new therapeutic targets and clinical biomarkers. MPs known as submicron vesicles generated from membranes of apoptotic or activated cells into circulation have the ability to act as autocrine and paracrine effectors in cell-to-cell communication. They operate as biological vectors modulating the endothelial dysfunction, inflammation, coagulation, angiogenesis, thrombosis, subsequently contributing to the progression of macro and microvascular complications in diabetes. More recently, miRNAs have started to be actively investigated, leading to first exciting reports, which suggest their significant role in vascular physiology and disease. The contribution of MPs and also of their associated miRNAs to the development of vascular complications in diabetes was largely unexplored and undiscussed. In essence, with this review we bring light upon the understanding of impact diabetes has on vascular biology, and the significant role of MPs and MPs associated miRNAs as novel mediators, potential biomarkers and therapeutic targets in vascular complications in diabetes. Copyright © 2016 Elsevier Inc. All rights reserved.

Coronary flow reserve is impaired in patients with aortic valve calcification.

PubMed

Bozbas, Huseyin; Pirat, Bahar; Yildirir, Aylin; Simşek, Vahide; Sade, Elif; Eroglu, Serpil; Atar, Ilyas; Altin, Cihan; Demirtas, Saadet; Ozin, Bulent; Muderrisoglu, Haldun

2008-04-01

Calcific aortic valve disease is an active and progressive condition. Data indicate that aortic valve calcification (AVC) is associated with endothelial dysfunction and accepted as a manifestation of atherosclerosis. Coronary flow reserve (CFR) determined by transthoracic echocardiography has been introduced as a reliable indicator for coronary microvascular function. In this study we aimed to evaluate CFR in patients with AVC. Eighty patients, aged more than 60 years, without coronary heart disease or diabetes mellitus were included: 40 had AVC without significant stenosis (peak gradient across the valve <25 mm Hg) and 40 had normal aortic valves (controls). Using transthoracic Doppler echocardiography, we measured coronary diastolic peak flow velocities (PFV) at baseline and after dipyridamole infusion. CFR was calculated as the ratio of hyperemic to baseline diastolic PFV and was compared between groups. Mean ages for patients with AVC and controls were 68.9+/-6.2 and 67.6+/-5.9 years (P=.3). There were no significant differences regarding clinical characteristics, laboratory findings, ejection fraction, or peak aortic valve gradients. Mean diastolic PFV at baseline and during hyperemia were 28.4+/-4.2 and 59.2+/-7.8 cm/s for AVC and 27.7+/-3.9 and 68.5+/-10.5 cm/s for controls. Compared with controls, patients with AVC had significantly lower CFR values (2.12+/-0.41 versus 2.51+/-0.51; P<.0001). CFR is impaired in patients with AVC before valve stenosis develops, suggesting that microvascular-endothelial dysfunction is present during the early stages of the calcific aortic valve disease.

Laminar shear stress modulates endothelial luminal surface stiffness in a tissue-specific manner.

PubMed

Merna, Nick; Wong, Andrew K; Barahona, Victor; Llanos, Pierre; Kunar, Balvir; Palikuqi, Brisa; Ginsberg, Michael; Rafii, Shahin; Rabbany, Sina Y

2018-04-17

Endothelial cells form vascular beds in all organs and are exposed to a range of mechanical forces that regulate cellular phenotype. We sought to determine the role of endothelial luminal surface stiffness in tissue-specific mechanotransduction of laminar shear stress in microvascular mouse cells and the role of arachidonic acid in mediating this response. Microvascular mouse endothelial cells were subjected to laminar shear stress at 4 dynes/cm 2 for 12 hours in parallel plate flow chambers that enabled real-time optical microscopy and atomic force microscopy measurements of cell stiffness. Lung endothelial cells aligned parallel to flow, while cardiac endothelial cells did not. This rapid alignment was accompanied by increased cell stiffness. The addition of arachidonic acid to cardiac endothelial cells increased alignment and stiffness in response to shear stress. Inhibition of arachidonic acid in lung endothelial cells and embryonic stem cell-derived endothelial cells prevented cellular alignment and decreased cell stiffness. Our findings suggest that increased endothelial luminal surface stiffness in microvascular cells may facilitate mechanotransduction and alignment in response to laminar shear stress. Furthermore, the arachidonic acid pathway may mediate this tissue-specific process. An improved understanding of this response will aid in the treatment of organ-specific vascular disease. © 2018 John Wiley & Sons Ltd.

Microvascular dysfunction following multi-walled carbon nanotube exposure is mediated by thrombospondin-1 receptor CD47.

PubMed

Mandler, W Kyle; Nurkiewicz, Timothy R; Porter, Dale W; Kelley, Eric E; Olfert, I Mark

2018-05-21

Pulmonary exposure to multi-walled carbon nanotubes (MWCNT) disrupts peripheral microvascular function. Thrombospondin-1 (TSP-1) is highly expressed during lung injury and has been shown to alter microvascular reactivity. It is unclear exactly how TSP-1 exerts effects on vascular function, but we hypothesized that the TSP-1 receptor CD47 may mediate changes in vasodilation.Wildtype (WT) or CD47 knockout (CD47 KO) C57B6/J-background animals were exposed to 50 µg of MWCNT or saline control via pharyngeal aspiration. Twenty-four hours post-exposure, intravital microscopy was performed to assess arteriolar dilation and venular leukocyte adhesion and rolling. To assess tissue redox status, electron paramagnetic resonance and NOx measurements were performed, while inflammatory biomarkers were measured via multiplex assay.Vasodilation was impaired in the WT+MWCNT group compared to control (57±9% vs 90±2% relaxation), while CD47 KO animals showed no impairment (108±8% relaxation). Venular leukocyte adhesion and rolling increased by > 2-fold, while the CD47 KO group showed no change. Application of the antioxidant apocynin rescued normal leukocyte activity in the WT+MWCNT group. Lung and plasma NOx were reduced in the WT+MWCNT group by 47% and 32%, respectively, while the CD47 KO groups were unchanged from control. Some inflammatory cytokines were increased in the CD47+MWCNT group only.In conclusion, TSP-1 is an important ligand mediating MWCNT-induced microvascular dysfunction, and CD47 is a component of this dysregulation. CD47 activation likely disrupts nitric oxide (•NO) signaling and promotes leukocyte-endothelial interactions. Impaired •NO production, signaling, and bioavailability is linked to a variety of cardiovascular diseases in which TSP-1/CD47 may play an important role.

Associations between microvascular function and short-term exposure to traffic-related air pollution and particulate matter oxidative potential.

PubMed

Zhang, Xian; Staimer, Norbert; Tjoa, Tomas; Gillen, Daniel L; Schauer, James J; Shafer, Martin M; Hasheminassab, Sina; Pakbin, Payam; Longhurst, John; Sioutas, Constantinos; Delfino, Ralph J

2016-07-26

Short-term exposure to ambient air pollution has been associated with acute increases in cardiovascular hospitalization and mortality. However, causative chemical components and underlying pathophysiological mechanisms remain to be clarified. We hypothesized that endothelial dysfunction would be associated with mobile-source (traffic) air pollution and that pollutant components with higher oxidative potential to generate reactive oxygen species (ROS) would have stronger associations. We carried out a cohort panel study in 93 elderly non-smoking adults living in the Los Angeles metropolitan area, during July 2012-February 2014. Microvascular function, represented by reactive hyperemia index (RHI), was measured weekly for up to 12 weeks (N = 845). Air pollutant data included daily data from regional air-monitoring stations, five-day average PM chemical components and oxidative potential in three PM size-fractions, and weekly personal nitrogen oxides (NOx). Linear mixed-effect models estimated adjusted changes in microvascular function with exposure. RHI was inversely associated with traffic-related pollutants such as ambient PM2.5 black carbon (BC), NOx, and carbon monoxide (CO). An interquartile range change increase (1.06 μg/m(3)) in 5-day average BC was associated with decreased RHI, -0.093 (95 % CI: -0.151, -0.035). RHI was inversely associated with other mobile-source components/tracers (polycyclic aromatic hydrocarbons, elemental carbon, and hopanes), and PM oxidative potential as quantified in two independent assays (dithiothreitol and in vitro macrophage ROS) in accumulation and ultrafine PM, and transition metals. Our findings suggest that short-term exposures to traffic-related air pollutants with high oxidative potential are major components contributing to microvascular dysfunction.

Similar NF-κB Gene Signatures in TNF-α Treated Human Endothelial Cells and Breast Tumor Biopsies

PubMed Central

Perrot-Applanat, Martine; Vacher, Sophie; Toullec, Aurore; Pelaez, Irma; Velasco, Guillaume; Cormier, Françoise; Saad, Hanan El Sheikh; Lidereau, Rosette; Baud, Véronique; Bièche, Ivan

2011-01-01

Background Endothelial dysfunction has been implicated in the pathogenesis of diverse pathologies ranging from vascular and immune diseases to cancer. TNF-α is one of the mediators of endothelial dysfunction through the activation of transcription factors, including NF-κB. While HUVEC (macrovascular cells) have been largely used in the past, here, we documented an NF-κB gene signature in TNFα-stimulated microvascular endothelial cells HMEC often used in tumor angiogenesis studies. Methodology/Principal Findings We measured mRNA expression of 55 NF-κB related genes using quantitative RT-PCR in HUVEC and HMEC. Our study identified twenty genes markedly up-regulated in response to TNFα, including adhesion molecules, cytokines, chemokines, and apoptosis regulators, some of them being identified as TNF-α-inducible genes for the first time in endothelial cells (two apoptosis regulators, TNFAIP3 and TNFRSF10B/Trail R2 (DR5), the chemokines GM-CSF/CSF2 and MCF/CSF1, and CD40 and TNF-α itself, as well as NF-κB components (RELB, NFKB1or 50/p105 and NFKB2 or p52/p100). For eight genes, the fold induction was much higher in HMEC, as compared to HUVEC. Most importantly, our study described for the first time a connection between NF-κB activation and the induction of most, if not all, of these genes in HMEC as evaluated by pharmacological inhibition and RelA expression knock-down by RNA interference. Moreover, since TNF-α is highly expressed in tumors, we further applied the NF-κB gene signature documented in TNFα-stimulated endothelial cells to human breast tumors. We found a significant positive correlation between TNF and the majority (85 %) of the identified endothelial TNF-induced genes in a well-defined series of 96 (48 ERα positive and 48 ERα negative) breast tumors. Conclusion/Significance Taken together these data suggest the potential use of this NF-κB gene signature in analyzing the role of TNF-α in the endothelial dysfunction, as well as in breast tumors independently of the presence of ERα. PMID:21754991

Endothelin-1 Mediates Brain Microvascular Dysfunction Leading to Long-Term Cognitive Impairment in a Model of Experimental Cerebral Malaria

PubMed Central

Freeman, Brandi D.; Martins, Yuri C.; Akide-Ndunge, Oscar B.; Bruno, Fernando P.; Wang, Hua; Tanowitz, Herbert B.; Spray, David C.; Desruisseaux, Mahalia S.

2016-01-01

Plasmodium falciparum infection causes a wide spectrum of diseases, including cerebral malaria, a potentially life-threatening encephalopathy. Vasculopathy is thought to contribute to cerebral malaria pathogenesis. The vasoactive compound endothelin-1, a key participant in many inflammatory processes, likely mediates vascular and cognitive dysfunctions in cerebral malaria. We previously demonstrated that C57BL6 mice infected with P. berghei ANKA, our fatal experimental cerebral malaria model, sustained memory loss. Herein, we demonstrate that an endothelin type A receptor (ETA) antagonist prevented experimental cerebral malaria-induced neurocognitive impairments and improved survival. ETA antagonism prevented blood-brain barrier disruption and cerebral vasoconstriction during experimental cerebral malaria, and reduced brain endothelial activation, diminishing brain microvascular congestion. Furthermore, exogenous endothelin-1 administration to P. berghei NK65-infected mice, a model generally regarded as a non-cerebral malaria negative control for P. berghei ANKA infection, led to experimental cerebral malaria-like memory deficits. Our data indicate that endothelin-1 is critical in the development of cerebrovascular and cognitive impairments with experimental cerebral malaria. This vasoactive peptide may thus serve as a potential target for adjunctive therapy in the management of cerebral malaria. PMID:27031954

Increased pulmonary arteriolar tone associated with lung oxidative stress and nitric oxide in a mouse model of Alzheimer's disease.

PubMed

Roberts, Andrew M; Jagadapillai, Rekha; Vaishnav, Radhika A; Friedland, Robert P; Drinovac, Robert; Lin, Xingyu; Gozal, Evelyne

2016-09-01

Vascular dysfunction and decreased cerebral blood flow are linked to Alzheimer's disease (AD). Loss of endothelial nitric oxide (NO) and oxidative stress in human cerebrovascular endothelium increase expression of amyloid precursor protein (APP) and enhance production of the Aβ peptide, suggesting that loss of endothelial NO contributes to AD pathology. We hypothesize that decreased systemic NO bioavailability in AD may also impact lung microcirculation and induce pulmonary endothelial dysfunction. The acute effect of NO synthase (NOS) inhibition on pulmonary arteriolar tone was assessed in a transgenic mouse model (TgAD) of AD (C57BL/6-Tg(Thy1-APPSwDutIowa)BWevn/Mmjax) and age-matched wild-type controls (C57BL/6J). Arteriolar diameters were measured before and after the administration of the NOS inhibitor, L-NAME Lung superoxide formation (DHE) and formation of nitrotyrosine (3-NT) were assessed as indicators of oxidative stress, inducible NOS (iNOS) and tumor necrosis factor alpha (TNF-α) expression as indicators of inflammation. Administration of L-NAME caused either significant pulmonary arteriolar constriction or no change from baseline tone in wild-type (WT) mice, and significant arteriolar dilation in TgAD mice. DHE, 3-NT, TNF-α, and iNOS expression were higher in TgAD lung tissue, compared to WT mice. These data suggest L-NAME could induce increased pulmonary arteriolar tone in WT mice from loss of bioavailable NO In contrast, NOS inhibition with L-NAME had a vasodilator effect in TgAD mice, potentially caused by decreased reactive nitrogen species formation, while significant oxidative stress and inflammation were present. We conclude that AD may increase pulmonary microvascular tone as a result of loss of bioavailable NO and increased oxidative stress. Our findings suggest that AD may have systemic microvascular implications beyond central neural control mechanisms. © 2016 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

Short-term regular aerobic exercise reduces oxidative stress produced by acute in the adipose microvasculature.

PubMed

Robinson, Austin T; Fancher, Ibra S; Sudhahar, Varadarajan; Bian, Jing Tan; Cook, Marc D; Mahmoud, Abeer M; Ali, Mohamed M; Ushio-Fukai, Masuko; Brown, Michael D; Fukai, Tohru; Phillips, Shane A

2017-05-01

High blood pressure has been shown to elicit impaired dilation in the vasculature. The purpose of this investigation was to elucidate the mechanisms through which high pressure may elicit vascular dysfunction and determine the mechanisms through which regular aerobic exercise protects arteries against high pressure. Male C57BL/6J mice were subjected to 2 wk of voluntary running (~6 km/day) for comparison with sedentary controls. Hindlimb adipose resistance arteries were dissected from mice for measurements of flow-induced dilation (FID; with or without high intraluminal pressure exposure) or protein expression of NADPH oxidase II (NOX II) and superoxide dismutase (SOD). Microvascular endothelial cells were subjected to high physiological laminar shear stress (20 dyn/cm 2 ) or static condition and treated with ANG II + pharmacological inhibitors. Cells were analyzed for the detection of ROS or collected for Western blot determination of NOX II and SOD. Resistance arteries from exercised mice demonstrated preserved FID after high pressure exposure, whereas FID was impaired in control mouse arteries. Inhibition of ANG II or NOX II restored impaired FID in control mouse arteries. High pressure increased superoxide levels in control mouse arteries but not in exercise mouse arteries, which exhibited greater ability to convert superoxide to H 2 O 2 Arteries from exercised mice exhibited less NOX II protein expression, more SOD isoform expression, and less sensitivity to ANG II. Endothelial cells subjected to laminar shear stress exhibited less NOX II subunit expression. In conclusion, aerobic exercise prevents high pressure-induced vascular dysfunction through an improved redox environment in the adipose microvasculature. NEW & NOTEWORTHY We describe potential mechanisms contributing to aerobic exercise-conferred protection against high intravascular pressure. Subcutaneous adipose microvessels from exercise mice express less NADPH oxidase (NOX) II and more superoxide dismutase (SOD) and demonstrate less sensitivity to ANG II. In microvascular endothelial cells, shear stress reduced NOX II but did not influence SOD expression.

GM-CSF ameliorates microvascular barrier integrity via pericyte-derived Ang-1 in wound healing.

PubMed

Yan, Min; Hu, Yange; Yao, Min; Bao, Shisan; Fang, Yong

2017-11-01

Skin wound healing involves complex coordinated interactions of cells, tissues, and mediators. Maintaining microvascular barrier integrity is one of the key events for endothelial homeostasis during wound healing. Vasodilation is observed after vasoconstriction, which causes blood vessels to become porous, facilitates leukocyte infiltration and aids angiogenesis at the wound-area, postinjury. Eventually, vessel integrity has to be reestablished for vascular maturation. Numerous studies have found that granulocyte macrophage colony-stimulating factor (GM-CSF) accelerates wound healing by inducing recruitment of repair cells into the injury area and releases of cytokines. However, whether GM-CSF is involving in the maintaining of microvascular barrier integrity and the underlying mechanism remain still unclear. Aim of this study was to investigate the effects of GM-CSF on modulation of microvascular permeability in wound healing and underlying mechanisms. Wound closure and microvascular leakage was investigated using a full-thickness skin wound mouse model after GM-CSF intervention. The endothelial permeability was measured by Evans blue assay in vivo and in vitro endothelium/pericyte co-culture system using a FITC-Dextran permeability assay. To identify the source of angiopoietin-1 (Ang-1), double staining is used in vivo and ELISA and qPCR are used in vitro. To determine the specific effect of Ang-1 on GM-CSF maintaining microvascular stabilization, Ang-1 siRNA was applied to inhibit Ang-1 production in vivo and in vitro. Wound closure was significantly accelerated and microvascular leakage was ameliorated after GM-CSF treatment in mouse wound sites. GM-CSF decreased endothelial permeability through tightening endothelial junctions and increased Ang-1 protein level that was derived by perictye. Furthermore, applications of siRNAAng-1 inhibited GM-CSF mediated protection of microvascular barrier integrity both in vivo and in vitro. Our data indicate that GM-CSF ameliorates microvascular barrier integrity via pericyte-derived Ang-1 during wound healing. © 2018 by the Wound Healing Society.

The neutrophil-to-lymphocyte ratio as a marker of systemic endothelial dysfunction in asymptomatic subjects.

PubMed

Martínez-Urbistondo, Diego; Beltrán, Almudena; Beloqui, Oscar; Huerta, Ana

2016-01-01

The neutrophil-to-lymphocyte ratio has demonstrated to be a prognostic inflammatory marker in cardiovascular disease. The objective of this study is to evaluate the association between neutrophil-to-lymphocyte ratio and pathologic urinary albumin/creatinine ratio as an early marker of cardiovascular risk and systemic endothelial dysfunction, associated with microvascular disease, in asymptomatic subjects. A unicenter cross-sectional study was conducted, including 1816 asymptomatic subjects. Patients with previous cardiovascular disease, those who were treated with ACE inhibitors and/or angiotensin II receptor blockers and patients with albumin/creatinine ratio over 300mg/g were excluded. The outcome of the study was the presence of a pathologic urinary albumin/creatinine ratio. The neutrophil-to-lymphocyte ratio was significantly associated with altered urinary albumin/creatinine ratio in the univariate analysis and after adjustment for other known endothelial and cardiovascular risk factors (age, hypertension, dyslipidaemia, diabetes or altered glomerular filtration rate). Based on the sensitivity and specificity of different neutrophil-to-lymphocyte ratio thresholds, 3 risk groups were created for altered urinary albumin/creatinine ratio: low risk in those with neutrophil-to-lymphocyte ratio < 1.5, intermediate risk in patients between 1.5 and 3, and high risk in those with neutrophil-to-lymphocyte ratio > 3. These groups were found to have a statistically significant and independent prognostic power for altered urinary albumin/creatinine ratio in asymptomatic patients. The neutrophil-to-lymphocyte ratio appears to be a cost-efficient, non-invasive and independent potential marker of systemic endothelial dysfunction in asymptomatic subjects. Copyright © 2015 Sociedad Española de Nefrología. Published by Elsevier España, S.L.U. All rights reserved.

Reversal of anemia with allogenic RBC transfusion prevents post-cardiopulmonary bypass acute kidney injury in swine

PubMed Central

Patel, Nishith N.; Lin, Hua; Toth, Tibor; Welsh, Gavin I.; Jones, Ceri; Ray, Paramita; Satchell, Simon C.; Sleeman, Philippa; Angelini, Gianni D.

2011-01-01

Anemia during cardiopulmonary bypass (CPB) is strongly associated with acute kidney injury in clinical studies; however, reversal of anemia with red blood cell (RBC) transfusions is associated with further renal injury. To understand this paradox, we evaluated the effects of reversal of anemia during CPB with allogenic RBC transfusion in a novel large-animal model of post-cardiac surgery acute kidney injury with significant homology to that observed in cardiac surgery patients. Adult pigs undergoing general anesthesia were allocated to a Sham procedure, CPB alone, Sham+RBC transfusion, or CPB+RBC transfusion, with recovery and reassessment at 24 h. CPB was associated with dilutional anemia and caused acute kidney injury in swine characterized by renal endothelial dysfunction, loss of nitric oxide (NO) bioavailability, vasoconstriction, medullary hypoxia, cortical ATP depletion, glomerular sequestration of activated platelets and inflammatory cells, and proximal tubule epithelial cell stress. RBC transfusion in the absence of CPB also resulted in renal injury. This was characterized by endothelial injury, microvascular endothelial dysfunction, platelet activation, and equivalent cortical tubular epithelial phenotypic changes to those observed in CPB pigs, but occurred in the absence of severe intrarenal vasoconstriction, ATP depletion, or reductions in creatinine clearance. In contrast, reversal of anemia during CPB with RBC transfusion prevented the reductions in creatinine clearance, loss of NO bioavailability, platelet activation, inflammation, and epithelial cell injury attributable to CPB although it did not prevent the development of significant intrarenal vasoconstriction and endothelial dysfunction. In conclusion, contrary to the findings of observational studies in cardiac surgery, RBC transfusion during CPB protects pigs against acute kidney injury. Our study underlines the need for translational research into indications for transfusion and prevention strategies for acute kidney injury. PMID:21653630

Ghrelin stimulates angiogenesis in human microvascular endothelial cells: Implications beyond GH release

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li Aihua; Cheng Guangli; Zhu Genghui

Ghrelin, a peptide hormone isolated from the stomach, releases growth hormone and stimulates appetite. Ghrelin is also expressed in pancreas, kidneys, cardiovascular system and in endothelial cells. The precise role of ghrelin in endothelial cell functions remains unknown. We examined the expression of ghrelin and its receptor (GHSR1) mRNAs and proteins in human microvascular endothelial cells (HMVEC) and determined whether ghrelin affects in these cells proliferation, migration and in vitro angiogenesis; and whether MAPK/ERK2 signaling is important for the latter action. We found that ghrelin and GHSR1 are constitutively expressed in HMVEC. Treatment of HMVEC with exogenous ghrelin significantly increasedmore » in these cells proliferation, migration, in vitro angiogenesis and ERK2 phosphorylation. MEK/ERK2 inhibitor, PD 98059 abolished ghrelin-induced in vitro angiogenesis. This is First demonstration that ghrelin and its receptor are expressed in human microvascular endothelial cells and that ghrelin stimulates HMVEC proliferation, migration, and angiogenesis through activation of ERK2 signaling.« less

The regulation of vascular endothelial growth factor-induced microvascular permeability requires Rac and reactive oxygen species.

PubMed

Monaghan-Benson, Elizabeth; Burridge, Keith

2009-09-18

Vascular permeability is a complex process involving the coordinated regulation of multiple signaling pathways in the endothelial cell. It has long been documented that vascular endothelial growth factor (VEGF) greatly enhances microvascular permeability; however, the molecular mechanisms controlling VEGF-induced permeability remain unknown. Treatment of microvascular endothelial cells with VEGF led to an increase in reactive oxygen species (ROS) production. ROS are required for VEGF-induced permeability as treatment with the free radical scavenger, N-acetylcysteine, inhibited this effect. Additionally, treatment with VEGF caused ROS-dependent tyrosine phosphorylation of both vascular-endothelial (VE)-cadherin and beta-catenin. Rac1 was required for the VEGF-induced increase in permeability and adherens junction protein phosphorylation. Knockdown of Rac1 inhibited VEGF-induced ROS production consistent with Rac lying upstream of ROS in this pathway. Collectively, these data suggest that VEGF leads to a Rac-mediated generation of ROS, which, in turn, elevates the tyrosine phosphorylation of VE-cadherin and beta-catenin, ultimately regulating adherens junction integrity.

Endothelial safety of radiological contrast media: why being concerned.

PubMed

Scoditti, Egeria; Massaro, Marika; Montinari, Maria Rosa

2013-01-01

Iodinated radiocontrast media have been the most widely used pharmaceuticals for intravascular administration in diagnostic and interventional angiographic procedures. Although they are regarded as relatively safe drugs and vascular biocompatibility of contrast media has been progressively improved, severe adverse reactions may occur, among which acute nephropathy is one of the most clinically significant complications after intravascular administration of contrast media and a powerful predictor of poor early and long-term outcomes. Since radiocontrast media are given through the arterial or the venous circulation in vascular procedures, morphological and functional changes of the microvascular and macrovascular endothelial cells substantially contribute to the pathogenesis of organ-specific and systemic adverse reactions of contrast media. Endothelial toxicity of contrast media seems to be the result of both direct proapoptotic effects and morphological derangements, as well as endothelial dysfunction and induction of inflammation, oxidative stress, thrombosis, and altered vasomotor balance, with predominant vasoconstrictive response in atherosclerotic coronary arteries and kidney microcirculation. Further understanding of pathogenetic mechanisms underlying contrast media-induced adverse reactions in cellular targets, including endothelial cells, will hopefully lead to the development of novel preventive strategies appropriately curbing the pathogenesis of contrast media vasotoxicity. Copyright © 2012 Elsevier Inc. All rights reserved.

Mechanisms of lung endothelial barrier disruption induced by cigarette smoke: role of oxidative stress and ceramides.

PubMed

Schweitzer, Kelly S; Hatoum, Hadi; Brown, Mary Beth; Gupta, Mehak; Justice, Matthew J; Beteck, Besem; Van Demark, Mary; Gu, Yuan; Presson, Robert G; Hubbard, Walter C; Petrache, Irina

2011-12-01

The epithelial and endothelial cells lining the alveolus form a barrier essential for the preservation of the lung respiratory function, which is, however, vulnerable to excessive oxidative, inflammatory, and apoptotic insults. Whereas profound breaches in this barrier function cause pulmonary edema, more subtle changes may contribute to inflammation. The mechanisms by which cigarette smoke (CS) exposure induce lung inflammation are not fully understood, but an early alteration in the epithelial barrier function has been documented. We sought to investigate the occurrence and mechanisms by which soluble components of mainstream CS disrupt the lung endothelial cell barrier function. Using cultured primary rat microvascular cell monolayers, we report that CS induces endothelial cell barrier disruption in a dose- and time-dependent manner of similar magnitude to that of the epithelial cell barrier. CS exposure triggered a mechanism of neutral sphingomyelinase-mediated ceramide upregulation and p38 MAPK and JNK activation that were oxidative stress dependent and that, along with Rho kinase activation, mediated the endothelial barrier dysfunction. The morphological changes in endothelial cell monolayers induced by CS included actin cytoskeletal rearrangement, junctional protein zonula occludens-1 loss, and intercellular gap formation, which were abolished by the glutathione modulator N-acetylcysteine and ameliorated by neutral sphingomyelinase inhibition. The direct application of ceramide recapitulated the effects of CS, by disrupting both endothelial and epithelial cells barrier, by a mechanism that was redox and apoptosis independent and required Rho kinase activation. Furthermore, ceramide induced dose-dependent alterations of alveolar microcirculatory barrier in vivo, measured by two-photon excitation microscopy in the intact rat. In conclusion, soluble components of CS have direct endothelial barrier-disruptive effects that could be ameliorated by glutathione modulators or by inhibitors of neutral sphingomyelinase, p38 MAPK, JNK, and Rho kinase. Amelioration of endothelial permeability may alleviate lung and systemic vascular dysfunction associated with smoking-related chronic obstructive lung diseases.

The protective role of isorhamnetin on human brain microvascular endothelial cells from cytotoxicity induced by methylglyoxal and oxygen-glucose deprivation.

PubMed

Li, Wenlu; Chen, Zhigang; Yan, Min; He, Ping; Chen, Zhong; Dai, Haibin

2016-02-01

As the first target of stroke, cerebral endothelial cells play a key role in brain vascular repair and maintenance, and their function is impeded in diabetes. Methylglyoxal (MGO), a reactive dicarbonyl produced during glucose metabolism, accumulates in diabetic patients. MGO and MGO-induced advanced glycation end-products (AGEs) could ameliorate stroke-induced brain vascular damage, closely related with ECs dysfunction. Using MGO plus oxygen-glucose deprivation (OGD) to mimic diabetic stroke, we reported the protective effect of isorhamnetin on OGD-induced cytotoxicity after MGO treatment on primary human brain microvascular endothelial cells (HBMEC) and explored the underlying mechanisms. Treatment of MGO for 24 h significantly enhanced 3-h OGD-induced HBMEC toxic effect, which was inhibited by pretreatment of isorhamnetin (100 μmol/L). Moreover, the protective effect of isorhamnetin is multiple function dependent, which includes anti-inflammation, anti-oxidative stress and anti-apoptosis effects. Besides its well-known inhibition on the mitochondria-dependent or intrinsic apoptotic pathway, isorhamnetin also reduced activation of the extrinsic apoptotic pathway, as characterized by the decreased expression and activity of caspase 3 and caspase 8. Furthermore, pretreatment with isorhamnetin specifically inhibited FAS/FASL expression and suppressed nuclear factor-kappa B nuclear translocation. Taken together, our results indicated that isorhamnetin protected against OGD-induced cytotoxicity after MGO treatment in cultured HBMEC due to its multiple protective effects and could inhibit Fas-mediated extrinsic apoptosis. Therefore, isorhamnetin is a promising reagent for the treatment of hyperglycemia and ischemia-induced cerebral vascular degeneration. A proposed model of the potential protective mechanism of isorhamnetin, a metabolite of quercetin, on methylglyoxal (MGO) treatment plus oxygen-glucose deprivation (OGD) exposure-induced cytotoxicity in cultured human brain microvascular endothelial cells. Isorhamnetin inhibits FasL-mediated extrinsic apoptosis and neurotrophic factor κB (NF-κB) nuclear translocation, which can induce the cell DNA damage. Therefore, the protective effect of isorhamnetin occurs through multiple functions, including anti-inflammation, anti-oxidative stress and anti-apoptosis. Therefore, isorhamnetin is a promising reagent for the treatment of hyperglycemia and ischemia-induced cerebral vascular degeneration. © 2015 International Society for Neurochemistry.

Cytotoxicity, permeability, and inflammation of metal oxide nanoparticles in human cardiac microvascular endothelial cells: cytotoxicity, permeability, and inflammation of metal oxide nanoparticles.

PubMed

Sun, Jing; Wang, Shaochuang; Zhao, Dong; Hun, Fei Han; Weng, Lei; Liu, Hui

2011-10-01

Wide applications and extreme potential of metal oxide nanoparticles (NPs) increase occupational and public exposure and may yield extraordinary hazards for human health. Exposure to NPs has a risk for dysfunction of the vascular endothelial cells. The objective of this study was to assess the cytotoxicity of six metal oxide NPs to human cardiac microvascular endothelial cells (HCMECs) in vitro. Metal oxide NPs used in this study included zinc oxide (ZnO), iron(III) oxide (Fe(2)O(3)), iron(II,III) oxide (Fe(3)O(4)), magnesium oxide (MgO), aluminum oxide (Al(2)O(3)), and copper(II) oxide (CuO). The cell viability, membrane leakage of lactate dehydrogenase, intracellular reactive oxygen species, permeability of plasma membrane, and expression of inflammatory markers vascular cell adhesion molecule-1, intercellular adhesion molecule-1, macrophage cationic peptide-1, and interleukin-8 in HCMECs were assessed under controlled and exposed conditions (12-24 h and 0.001-100 μg/ml of exposure). The results indicated that Fe(2)O(3), Fe(3)O(4), and Al(2)O(3) NPs did not have significant effects on cytotoxicity, permeability, and inflammation response in HCMECs at any of the concentrations tested. ZnO, CuO, and MgO NPs produced the cytotoxicity at the concentration-dependent and time-dependent manner, and elicited the permeability and inflammation response in HCMECs. These results demonstrated that cytotoxicity, permeability, and inflammation in vascular endothelial cells following exposure to metal oxide nanoparticles depended on particle composition, concentration, and exposure time. © Springer Science+Business Media B.V. 2011

Endothelial Dysfunction Exacerbates Renal Interstitial Fibrosis through Enhancing Fibroblast Smad3 Linker Phosphorylation in the Mouse Obstructed Kidney

PubMed Central

Sun, Yu Bo Yang; Qu, Xinli; Li, Xueling; Nikolic-Paterson, David J.; Li, Jinhua

2013-01-01

Endothelial dysfunction and enhanced transforming growth factor-β (TGF-β)/Smad3 signalling are common features of progressive renal fibrosis. This study investigated a potential link between these mechanisms. In unilateral ureteric obstruction (UUO) we observed an acute (6 hr) down-regulation of nitric oxide synthase 3 (NOS3/eNOS) levels and increased phosphorylation of the linker region of Smad3 at T179 and S208 in Smad3/JNK complexes. These events preceded Smad3 C-terminal domain phosphorylation and the induction of myofibroblast proliferation at 48 hrs. Mice deficient in NOS3 showed enhanced myofibroblast proliferation and collagen accumulation compared to wild type mice in a 7 day UUO model. This was associated with enhanced phosphorylation of Smad3 T179 and S208 by 92% and 88%, respectively, whereas Smad3-C-terminal phosphorylation was not affected. Resolvin D1 (RvD1) can suppress renal fibrosis in the UUO model, and further analysis herein showed that RvD1 protected against endothelial dysfunction and suppressed Smad3/JNK complex formation with a consequent reduction in phosphorylation of Smad3 T179 and S208 by 78% and 65%, respectively, while Smad3 C-terminal phosphorylation was unaltered. In vitro, conditioned media from mouse microvascular endothelial cells (MMEC) treated with a general inhibitor of nitric oxide synthase (L-NAME) augmented the proliferation and collagen production of renal fibroblasts (NRK49F cells) compared to control MMEC media and this was associated with increased phosphorylation of JNK and Smad3 T179 and S208, whereas Smad3-C-terminal domain phosphorylation was unaffected. The addition of RvD1 to L-NAME treated MMEC abrogated these effects of the conditioned media on renal fibroblasts. Finally, Smad3 T179/V and S208/A mutations significantly inhibit TGF-β1 induced up-regulation collagen I promoter. In conclusion, these data suggest that endothelial dysfunction can exacerbate renal interstitial fibrosis through increased fibroblast proliferation and collagen production via enhanced Smad3 linker phosphorylation. PMID:24391884

Endothelial dysfunction exacerbates renal interstitial fibrosis through enhancing fibroblast Smad3 linker phosphorylation in the mouse obstructed kidney.

PubMed

Sun, Yu Bo Yang; Qu, Xinli; Li, Xueling; Nikolic-Paterson, David J; Li, Jinhua

2013-01-01

Endothelial dysfunction and enhanced transforming growth factor-β (TGF-β)/Smad3 signalling are common features of progressive renal fibrosis. This study investigated a potential link between these mechanisms. In unilateral ureteric obstruction (UUO) we observed an acute (6 hr) down-regulation of nitric oxide synthase 3 (NOS3/eNOS) levels and increased phosphorylation of the linker region of Smad3 at T179 and S208 in Smad3/JNK complexes. These events preceded Smad3 C-terminal domain phosphorylation and the induction of myofibroblast proliferation at 48 hrs. Mice deficient in NOS3 showed enhanced myofibroblast proliferation and collagen accumulation compared to wild type mice in a 7 day UUO model. This was associated with enhanced phosphorylation of Smad3 T179 and S208 by 92% and 88%, respectively, whereas Smad3-C-terminal phosphorylation was not affected. Resolvin D1 (RvD1) can suppress renal fibrosis in the UUO model, and further analysis herein showed that RvD1 protected against endothelial dysfunction and suppressed Smad3/JNK complex formation with a consequent reduction in phosphorylation of Smad3 T179 and S208 by 78% and 65%, respectively, while Smad3 C-terminal phosphorylation was unaltered. In vitro, conditioned media from mouse microvascular endothelial cells (MMEC) treated with a general inhibitor of nitric oxide synthase (L-NAME) augmented the proliferation and collagen production of renal fibroblasts (NRK49F cells) compared to control MMEC media and this was associated with increased phosphorylation of JNK and Smad3 T179 and S208, whereas Smad3-C-terminal domain phosphorylation was unaffected. The addition of RvD1 to L-NAME treated MMEC abrogated these effects of the conditioned media on renal fibroblasts. Finally, Smad3 T179/V and S208/A mutations significantly inhibit TGF-β1 induced up-regulation collagen I promoter. In conclusion, these data suggest that endothelial dysfunction can exacerbate renal interstitial fibrosis through increased fibroblast proliferation and collagen production via enhanced Smad3 linker phosphorylation.

Endothelial Progenitor Cells in Diabetic Microvascular Complications: Friends or Foes?

PubMed

Yu, Cai-Guo; Zhang, Ning; Yuan, Sha-Sha; Ma, Yan; Yang, Long-Yan; Feng, Ying-Mei; Zhao, Dong

2016-01-01

Despite being featured as metabolic disorder, diabetic patients are largely affected by hyperglycemia-induced vascular abnormality. Accumulated evidence has confirmed the beneficial effect of endothelial progenitor cells (EPCs) in coronary heart disease. However, antivascular endothelial growth factor (anti-VEGF) treatment is the main therapy for diabetic retinopathy and nephropathy, indicating the uncertain role of EPCs in the pathogenesis of diabetic microvascular disease. In this review, we first illustrate how hyperglycemia induces metabolic and epigenetic changes in EPCs, which exerts deleterious impact on their number and function. We then discuss how abnormal angiogenesis develops in eyes and kidneys under diabetes condition, focusing on "VEGF uncoupling with nitric oxide" and "competitive angiopoietin 1/angiopoietin 2" mechanisms that are shared in both organs. Next, we dissect the nature of EPCs in diabetic microvascular complications. After we overview the current EPCs-related strategies, we point out new EPCs-associated options for future exploration. Ultimately, we hope that this review would uncover the mysterious nature of EPCs in diabetic microvascular disease for therapeutics.

The association of endothelial function and tone by digital arterial tonometry with MRI left ventricular mass in African Americans: the Jackson Heart Study.

PubMed

Tripathi, Avnish; Benjamin, Emelia J; Musani, Solomon K; Hamburg, Naomi M; Tsao, Connie W; Saraswat, Arti; Vasan, Ramachandran S; Mitchell, Gary F; Fox, Ervin R

2017-05-01

Peripheral vascular endothelial dysfunction assessed by digital peripheral arterial tonometry (PAT) has been associated with risk for adverse cardiovascular events. We examined the relations of peripheral microvascular dysfunction and left ventricular mass in a community-based cohort of African Americans. We examined participants of the Jackson Heart Study who had PAT and cardiac magnetic resonance imaging evaluations between 2007 and 2013. Consistent with pertinent literature, left ventricular mass index (LVMI) was adjusted for body size by indexing to height 2.7 . Pearson's correlation and general linear regression analyses were used to relate reactive hyperemia index, baseline pulse amplitude (BPA), and augmentation index (markers of microvascular vasodilator function, baseline vascular pulsatility, and relative wave reflection, respectively) to LVMI after adjusting for traditional cardiovascular risk factors. A total of 440 participants (mean age 59 ± 10 years, 60% women) were included. Age- and sex-adjusted Pearson's correlation analysis suggested that natural log transformed LVMI was negatively correlated with reactive hyperemia index (coefficient: -0.114; P = .02) and positively correlated with BPA (coefficient: 0.272; P < .001). In multivariable analyses, higher log e LVMI was associated with higher BPA (β: 0.210; P = .03) after accounting for age, sex, body mass index, diabetes, hypertension, ratio of total cholesterol and high-density lipoprotein cholesterol, smoking, and history of cardiovascular disease. In a community-based sample of African Americans, higher baseline pulsatility measured by PAT was associated with higher LVMI by cardiac magnetic resonance imaging after adjusting for traditional risk factors. Copyright © 2017 American Society of Hypertension. Published by Elsevier Inc. All rights reserved.

Reduction of vascular leakage by imatinib is associated with preserved microcirculatory perfusion and reduced renal injury markers in a rat model of cardiopulmonary bypass.

PubMed

Koning, N J; de Lange, F; van Meurs, M; Jongman, R M; Ahmed, Y; Schwarte, L A; van Nieuw Amerongen, G P; Vonk, A B A; Niessen, H W; Baufreton, C; Boer, C

2018-06-01

Cardiopulmonary bypass during cardiac surgery leads to impaired microcirculatory perfusion. We hypothesized that vascular leakage is an important contributor to microcirculatory dysfunction. Imatinib, a tyrosine kinase inhibitor, has been shown to reduce vascular leakage in septic mice. We investigated whether prevention of vascular leakage using imatinib preserves microcirculatory perfusion and reduces organ injury markers in a rat model of cardiopulmonary bypass. Male Wistar rats underwent cardiopulmonary bypass after treatment with imatinib or vehicle (n=8 per group). Cremaster muscle microcirculatory perfusion and quadriceps microvascular oxygen saturation were measured using intravital microscopy and reflectance spectroscopy. Evans Blue extravasation was determined in separate experiments. Organ injury markers were determined in plasma, intestine, kidney, and lungs. The onset of cardiopulmonary bypass decreased the number of perfused microvessels by 40% in the control group [9.4 (8.6-10.6) to 5.7 (4.8-6.2) per microscope field; P<0.001 vs baseline], whereas this reduction was not seen in the imatinib group. In the control group, the number of perfused capillaries remained low throughout the experiment, whilst perfusion remained normal after imatinib administration. Microvascular oxygen saturation was less impaired after imatinib treatment compared with controls. Imatinib reduced vascular leakage and decreased fluid resuscitation compared with control [3 (3-6) vs 12 ml (7-16); P=0.024]. Plasma neutrophil-gelatinase-associated-lipocalin concentrations were reduced by imatinib. Prevention of endothelial barrier dysfunction using imatinib preserved microcirculatory perfusion and oxygenation during and after cardiopulmonary bypass. Moreover, imatinib-induced protection of endothelial barrier integrity reduced fluid-resuscitation requirements and attenuated renal and pulmonary injury markers. Copyright © 2017 British Journal of Anaesthesia. Published by Elsevier Ltd. All rights reserved.

Binding of human endothelium to Ulex europaeus I-coated Dynabeads: application to the isolation of microvascular endothelium.

PubMed

Jackson, C J; Garbett, P K; Nissen, B; Schrieber, L

1990-06-01

A major problem encountered when isolating human microvascular endothelium is the presence of contaminating cells such as fibroblasts that rapidly over-grow the endothelial cells. We describe here a simple, rapid technique for purifying endothelial cells derived from the microvasculature of neonatal foreskin and osteoarthritic and rheumatoid arthritic synovium. This technique is based on the selective binding of the lectin Ulex europaeus I (UEA I) to the endothelial cell surface via fucose residues. Initially UEA I was covalently bound to tosyl-activated super-paramagnetic polystyrene beads (Dynabeads) by incubation for 24 h at room temperature. Cells were isolated by extracting microvascular segments from enzyme-treated (trypsin and Pronase) cubes of tissue. The mixed population of cells obtained were purified by incubating them at 4 degrees C for 10 min with the UEA I-coated Dynabeads. Endothelium bound to the beads whilst contaminating cells were removed by five washes with HBSS using a magnetic particle concentrator. The endothelial cells thus obtained grew to confluence as a cobblestone-like monolayer and expressed von Willebrand factor antigen. The cells were released from the Dynabeads by the competitive binding of fucose (10 min at 4 degrees C). This new method is simple and reproducible and allows pure human microvascular endothelial cells to be cultured within 2 h of obtaining a specimen.

Multiple protocadherins are expressed in brain microvascular endothelial cells and might play a role in tight junction protein regulation.

PubMed

Dilling, Christina; Roewer, Norbert; Förster, Carola Y; Burek, Malgorzata

2017-10-01

Protocadherins (Pcdhs) are a large family of cadherin-related molecules. They play a role in cell adhesion, cellular interactions, and development of the central nervous system. However, their expression and role in endothelial cells has not yet been characterized. Here, we examined the expression of selected clustered Pcdhs in endothelial cells from several vascular beds. We analyzed human and mouse brain microvascular endothelial cell (BMEC) lines and primary cells, mouse myocardial microvascular endothelial cell line, and human umbilical vein endothelial cells. We examined the mRNA and protein expression of selected Pcdhs using RT-PCR, Western blot, and immunostaining. A strong mRNA expression of Pcdhs was observed in all endothelial cells tested. At the protein level, Pcdhs-gamma were detected using an antibody against the conserved C-terminal domain of Pcdhs-gamma or an antibody against PcdhgC3. Deletion of highly expressed PcdhgC3 led to differences in the tight junction protein expression and mRNA expression of Wnt/mTOR (mechanistic target of rapamycin) pathway genes as well as lower transendothelial electrical resistance. Staining of PcdhgC3 showed diffused cytoplasmic localization in mouse BMEC. Our results suggest that Pcdhs may play a critical role in the barrier-stabilizing pathways at the blood-brain barrier.

Mechanism of action of vitamin C in sepsis: Ascorbate modulates redox signaling in endothelium

PubMed Central

Wilson, John X.

2009-01-01

Circulating levels of vitamin C (ascorbate) are low in patients with sepsis. Parenteral administration of ascorbate raises plasma and tissue concentrations of the vitamin and may decrease morbidity. In animal models of sepsis, intravenous ascorbate injection increases survival and protects several microvascular functions, namely, capillary blood flow, microvascular permeability barrier, and arteriolar responsiveness to vasoconstrictors and vasodilators. The effects of parenteral ascorbate on microvascular function are both rapid and persistent. Ascorbate quickly accumulates in microvascular endothelial cells, scavenges reactive oxygen species, and acts through tetrahydrobiopterin to stimulate nitric oxide production by endothelial nitric oxide synthase. A major reason for the long duration of the improvement in microvascular function is that cells retain high levels of ascorbate, which alter redox-sensitive signaling pathways to diminish septic induction of NADPH oxidase and inducible nitric oxide synthase. These observations are consistent with the hypothesis that microvascular function in sepsis may be improved by parenteral administration of ascorbate as an adjuvant therapy. PMID:19319840

The role of nailfold capillaroscopy in the assessment of internal organ involvement in systemic sclerosis: A critical review.

PubMed

Soulaidopoulos, Stergios; Triantafyllidou, Eva; Garyfallos, Alexandros; Kitas, George D; Dimitroulas, Theodoros

2017-08-01

Endothelial dysfunction and microvascular damage constitute the hallmarks of systemic sclerosis (SSc), explaining much of the pathophysiology and clinical manifestations of the disease. Nailfold videocapillaroscopy (NVC) is an established method for the assessment of the microvasculature, aiding in distinguishing different types of structural vascular abnormalities. Until recently, NVC was used in the diagnosis of SSc as well as in the assessment and follow-up of peripheral digital vasculopathy. On the top of digital ulcers, internal organ involvement such as myocardial dysfunction, pulmonary vascular and/or parenchymal lung disease characterizes severe SSc imparting a high risk of mortality. There is growing evidence suggesting that the extent of peripheral microvascular changes reflects the severity of the disease, especially in terms of life-threatening cardiopulmonary complications. The possible use of nailfold videocapillaroscopy as a useful, non-invasive modality to improve the ability to identify patients at higher risk for these devastating complications of the disease remains to be established. The aim of this review is to critically summarize and discuss current literature regarding the relationship between morphological alterations of nailfold dermal papillary vessels and several manifestations of SSc, focusing on visceral organ involvement, as well as their association with surrogate markers of macrovascular disease. Copyright © 2017 Elsevier B.V. All rights reserved.

Hyperglycaemia promotes human brain microvascular endothelial cell apoptosis via induction of protein kinase C-ßI and prooxidant enzyme NADPH oxidase.

PubMed

Shao, Beili; Bayraktutan, Ulvi

2014-01-01

Blood-brain barrier disruption represents a key feature in hyperglycaemia-aggravated cerebral damage after an ischaemic stroke. Although the underlying mechanisms remain largely unknown, activation of protein kinase C (PKC) is thought to play a critical role. This study examined whether apoptosis of human brain microvascular endothelial cells (HBMEC) might contribute to hyperglycaemia-evoked barrier damage and assessed the specific role of PKC in this phenomenon. Treatments with hyperglycaemia (25 mM) or phorbol myristate acetate (PMA, a protein kinase C activator, 100 nM) significantly increased NADPH oxidase activity, O2 (•-) generation, proapoptotic protein Bax expression, TUNEL-positive staining and caspase-3/7 activities. Pharmacological inhibition of NADPH oxidase, PKC-a, PKC-ß or PKC-ßI via their specific inhibitors and neutralisation of O2 (•-) by a cell-permeable superoxide dismutase mimetic, MnTBAP normalised all the aforementioned increases induced by hyperglycaemia. Suppression of these PKC isoforms also negated the stimulatory effects of hyperglycaemia on the protein expression of NADPH oxidase membrane-bound components, Nox2 and p22-phox which determine the overall enzymatic activity. Silencing of PKC-ßI gene through use of specific siRNAs abolished the effects of both hyperglycaemia and PMA on endothelial cell NADPH oxidase activity, O2 (•-) production and apoptosis and consequently improved the integrity and function of an in vitro model of human cerebral barrier comprising HBMEC, astrocytes and pericytes. Hyperglycaemia-mediated apoptosis of HBMEC contributes to cerebral barrier dysfunction and is modulated by sequential activations of PKC-ßI and NADPH oxidase.

Impaired coronary flow reserve in metastatic cancer patients treated with sunitinib.

PubMed

Sen, F; Yildiz, I; Basaran, M; Ekenel, M; Oz, F; Kilic, L; Toz, B; Gurdal, A; Camlica, H; Bavbek, S; Oflaz, H

2013-01-01

Hypertension is one of the major side effects of sunitinib, an angiogenesis inhibitor used in the treatment of metastatic renal cell carcinomas (mRCC) and gastrointestinal stromal tumors (GIST). Endothelial dysfunction, an early and reversible event in the pathogenesis of atherosclerosis, is suggested to be one of the possible underlying mechanisms of hypertension caused by angiogenesis inhibitors. Coronary flow reserve (CFR) measurement by trans-thoracic Doppler echocardiography (TTDE) reflects coronary microvascular and endothelial functions, as a cheaper and an easy screening test. We have used TTDE to evaluate endothelial function and coronary microvascular function in mRCC and GIST patients under sunitinib treatment. Eighteen metastatic cancer patients (16 mRCC and 2 GIST) on sunitinib treatment and 27 healthy subjects were enrolled in this cross-sectional study. Thyroid stimulating hormone (TSH), lipid profile, creatinine, hemoglobin, glucose, C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), anthropometric and physical parameters of patients were recorded. CFR recordings were performed by the Vivid 7 echocardiography device. CFR was significantly lower in patients when compared with controls (1.82±0.4 vs 2.71±0.8, respectively; p < 0.001). Impaired CFR was found in 13 (72%) patients whereas all controls had normal CFR values. CFR was inversely correlated with the duration of sunitinib treatment (r=-0.36, p =0.01), high sensitivite (hs) CRP (r = -0.574, p =0.01) and ESR (r = - 0.5, p = 0.02). Our findings indicate that CFR is significantly impaired in cancer patients on sunitinib treatment. There is an inverse correlation between CFR and duration of sunitinib treatment and inflammation markers.

Reduced Microvascular Density in Omental Biopsies of Children with Chronic Kidney Disease

PubMed Central

Grabe, Niels; Lahrmann, Bernd; Nasser, Hamoud; Freise, Christian; Schneider, Axel; Lingnau, Anja; Degenhardt, Petra; Ranchin, Bruno; Sallay, Peter; Cerkauskiene, Rimante; Malina, Michal; Ariceta, Gema; Schmitt, Claus Peter; Querfeld, Uwe

2016-01-01

Background Endothelial dysfunction is an early manifestation of cardiovascular disease (CVD) and consistently observed in patients with chronic kidney disease (CKD). We hypothesized that CKD is associated with systemic damage to the microcirculation, preceding macrovascular pathology. To assess the degree of “uremic microangiopathy”, we have measured microvascular density in biopsies of the omentum of children with CKD. Patients and Methods Omental tissue was collected from 32 healthy children (0–18 years) undergoing elective abdominal surgery and from 23 age-matched cases with stage 5 CKD at the time of catheter insertion for initiation of peritoneal dialysis. Biopsies were analyzed by independent observers using either a manual or an automated imaging system for the assessment of microvascular density. Quantitative immunohistochemistry was performed for markers of autophagy and apoptosis, and for the abundance of the angiogenesis-regulating proteins VEGF-A, VEGF-R2, Angpt1 and Angpt2. Results Microvascular density was significantly reduced in uremic children compared to healthy controls, both by manual imaging with a digital microscope (median surface area 0.61% vs. 0.95%, p<0.0021 and by automated quantification (total microvascular surface area 0.89% vs. 1.17% p = 0.01). Density measured by manual imaging was significantly associated with age, height, weight and body surface area in CKD patients and healthy controls. In multivariate analysis, age and serum creatinine level were the only independent, significant predictors of microvascular density (r2 = 0.73). There was no immunohistochemical evidence for apoptosis or autophagy. Quantitative staining showed similar expression levels of the angiogenesis regulators VEGF-A, VEGF-receptor 2 and Angpt1 (p = 0.11), but Angpt2 was significantly lower in CKD children (p = 0.01). Conclusions Microvascular density is profoundly reduced in omental biopsies of children with stage 5 CKD and associated with diminished Angpt2 signaling. Microvascular rarefaction could be an early systemic manifestation of CKD-induced cardiovascular disease. PMID:27846250

Comparison of apoptosis in human primary pulmonary endothelial cells and a brain microvascular endothelial cell line co-cultured with Plasmodium falciparum field isolates.

PubMed

Essone, Jean Claude Biteghe Bi; N'Dilimabaka, Nadine; Ondzaga, Julien; Lekana-Douki, Jean Bernard; Mba, Dieudonné Nkoghe; Deloron, Philippe; Mazier, Dominique; Gay, Frédrérick; Touré Ndouo, Fousseyni S

2017-06-27

Plasmodium falciparum infection can progress unpredictably to severe forms including respiratory distress and cerebral malaria. The mechanisms underlying the variable natural course of malaria remain elusive. The cerebral microvascular endothelial cells-D3 and lung endothelial cells both from human were cultured separately and challenged with P. falciparum field isolates taken directly from malaria patients or 3D7 strain (in vitro maintained culture). The capacity of these P. falciparum isolates to induce endothelial cell apoptosis via cytoadherence or not was then assessed. Overall, 27 P. falciparum isolates were collected from patients with uncomplicated malaria (n = 25) or severe malaria (n = 2). About half the isolates (n = 17) were able to bind brain endothelial cells (12 isolates, 44%) or lung endothelial cells (17 isolates, 63%) or both (12 isolates, 44%). Sixteen (59%) of the 27 isolates were apoptogenic for brain and/or lung endothelial cells. The apoptosis stimulus could be cytoadherence, direct cell-cell contact without cytoadherence, or diffusible soluble factors. While some of the apoptogenic isolates used two stimuli (direct contact with or without cytoadherence, plus soluble factors) to induce apoptosis, others used only one. Among the 16 apoptogenic isolates, eight specifically targeted brain endothelial cells, one lung endothelial cells, and seven both. These results indicate that the brain microvascular cell line was more susceptible to apoptosis triggered by P. falciparum than the primary pulmonary endothelial cells and may have relevance to host-parasite interaction.

Atrial fibrillation: effects beyond the atrium?

PubMed

Wijesurendra, Rohan S; Casadei, Barbara

2015-03-01

Atrial fibrillation (AF) is the most common sustained clinical arrhythmia and is associated with significant morbidity, mostly secondary to heart failure and stroke, and an estimated two-fold increase in premature death. Efforts to increase our understanding of AF and its complications have focused on unravelling the mechanisms of electrical and structural remodelling of the atrial myocardium. Yet, it is increasingly recognized that AF is more than an atrial disease, being associated with systemic inflammation, endothelial dysfunction, and adverse effects on the structure and function of the left ventricular myocardium that may be prognostically important. Here, we review the molecular and in vivo evidence that underpins current knowledge regarding the effects of human or experimental AF on the ventricular myocardium. Potential mechanisms are explored including diffuse ventricular fibrosis, focal myocardial scarring, and impaired myocardial perfusion and perfusion reserve. The complex relationship between AF, systemic inflammation, as well as endothelial/microvascular dysfunction and the effects of AF on ventricular calcium handling and oxidative stress are also addressed. Finally, consideration is given to the clinical implications of these observations and concepts, with particular reference to rate vs. rhythm control. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Cardiology.

Pulmonary microvascular dysfunction and pathological changes induced by blast injury in a rabbit model.

PubMed

Wu, Si Yu; Han, Geng Fen; Kang, Jian Yi; Zhang, Liang Chao; Wang, Ai Min; Wang, Jian Min

2016-09-01

Vascular leakage has been proven to play a critical role in the incidence and development of explosive pulmonary barotrauma. Quantitatively investigated in the present study was the severity of vascular leakage in a gradient blast injury series, as well as ultrastructural evidence relating to pulmonary vascular leakage. One hundred adult male New Zealand white rabbits were randomly divided into 5 groups according to distance from the detonator (10 cm, 15 cm, 20 cm, 30 cm, and sham control). Value of pulmonary vascular leakage was monitored by a radioactive 125I-albumin labeling method. Pathological changes caused by the blast wave were examined under light and electron microscopes. Transcapillary escape rate of 125I-albumin and residual radioactivity in both lungs increased significantly at the distances of 10 cm, 15 cm, and 20 cm, suggesting increased severity of vascular leakage in these groups. Ultrastructural observation showed swelling of pulmonary capillary endothelial cells and widened gap between endothelial cells in the 10-cm and 15-cm groups. Primary blast wave can result in pulmonary capillary blood leakage. Blast wave can cause swelling of pulmonary capillary endothelial cells and widened gap between endothelial cells, which may be responsible for pulmonary vascular leakage.

Angiogenic factors and their soluble receptors predict organ dysfunction and mortality in post-cardiac arrest syndrome.

PubMed

Wada, Takeshi; Jesmin, Subrina; Gando, Satoshi; Yanagida, Yuichiro; Mizugaki, Asumi; Sultana, Sayeeda N; Zaedi, Sohel; Yokota, Hiroyuki

2012-09-29

Post-cardiac arrest syndrome (PCAS) often leads to multiple organ dysfunction syndrome (MODS) with a poor prognosis. Endothelial and leukocyte activation after whole-body ischemia/reperfusion following resuscitation from cardiac arrest is a critical step in endothelial injury and related organ damage. Angiogenic factors, including vascular endothelial growth factor (VEGF) and angiopoietin (Ang), and their receptors play crucial roles in endothelial growth, survival signals, pathological angiogenesis and microvascular permeability. The aim of this study was to confirm the efficacy of angiogenic factors and their soluble receptors in predicting organ dysfunction and mortality in patients with PCAS. A total of 52 resuscitated patients were divided into two subgroups: 23 survivors and 29 non-survivors. The serum levels of VEGF, soluble VEGF receptor (sVEGFR)1, sVEGFR2, Ang1, Ang2 and soluble Tie2 (sTie2) were measured at the time of admission (Day 1) and on Day 3 and Day 5. The ratio of Ang2 to Ang1 (Ang2/Ang1) was also calculated. This study compared the levels of angiogenic factors and their soluble receptors between survivors and non-survivors, and evaluated the predictive value of these factors for organ dysfunction and 28-day mortality. The non-survivors demonstrated more severe degrees of organ dysfunction and a higher prevalence of MODS. Non-survivors showed significant increases in the Ang2 levels and the Ang2/Ang1 ratios compared to survivors. A stepwise logistic regression analysis demonstrated that the Ang2 levels or the Ang2/Ang1 ratios on Day 1 independently predicted the 28-day mortality. The receiver operating characteristic curves of the Ang2 levels, and the Ang2/Ang1 ratios on Day 1 were good predictors of 28-day mortality. The Ang2 levels also independently predicted increases in the Sequential Organ Failure Assessment (SOFA) scores. We observed a marked imbalance between Ang1 and Ang2 in favor of Ang2 in PCAS patients, and the effect was more prominent in non-survivors. Angiogenic factors and their soluble receptors, particularly Ang2 and Ang2/Ang1, are considered to be valuable predictive biomarkers in the development of organ dysfunction and poor outcomes in PCAS patients.

Effect of Antimicrobial Compounds on Balamuthia mandrillaris Encystment and Human Brain Microvascular Endothelial Cell Cytopathogenicity▿

PubMed Central

Siddiqui, Ruqaiyyah; Matin, Abdul; Warhurst, David; Stins, Monique; Khan, Naveed Ahmed

2007-01-01

Cycloheximide, ketoconazole, or preexposure of organisms to cytochalasin D prevented Balamuthia mandrillaris-associated cytopathogenicity in human brain microvascular endothelial cells, which constitute the blood-brain barrier. In an assay for inhibition of cyst production, these three agents prevented the production of cysts, suggesting that the biosynthesis of proteins and ergosterol and the polymerization of actin are important in cytopathogenicity and encystment. PMID:17875991

Vascular and inflammatory high fat meal responses in young healthy men; a discriminative role of IL-8 observed in a randomized trial.

PubMed

Esser, Diederik; Oosterink, Els; op 't Roodt, Jos; Henry, Ronald M A; Stehouwer, Coen D A; Müller, Michael; Afman, Lydia A

2013-01-01

High fat meal challenges are known to induce postprandial low-grade inflammation and endothelial dysfunction. This assumption is largely based on studies performed in older populations or in populations with a progressed disease state and an appropriate control meal is often lacking. Young healthy individuals might be more resilient to such challenges. We therefore aimed to characterize the vascular and inflammatory response after a high fat meal in young healthy individuals. In a double-blind randomized cross-over intervention study, we used a comprehensive phenotyping approach to determine the vascular and inflammatory response after consumption of a high fat shake and after an average breakfast shake in 20 young healthy subjects. Both interventions were performed three times. Many features of the vascular postprandial response, such as FMD, arterial stiffness and micro-vascular skin blood flow were not different between shakes. High fat/high energy shake consumption was associated with a more pronounced increase in blood pressure, heart rate, plasma concentrations of IL-8 and PBMCs gene expression of IL-8 and CD54 (ICAM-1), whereas plasma concentrations of sVCAM1 were decreased compared to an average breakfast. Whereas no difference in postprandial response were observed on classical markers of endothelial function, we did observe differences between consumption of a HF/HE and an average breakfast meal on blood pressure and IL-8 in young healthy volunteers. IL-8 might play an important role in dealing with high fat challenges and might be an early marker for endothelial stress, a stage preceding endothelial dysfunction.

Direct Isolation, Culture and Transplant of Mouse Skeletal Muscle Derived Endothelial Cells with Angiogenic Potential

PubMed Central

Ieronimakis, Nicholas; Balasundaram, Gayathri; Reyes, Morayma

2008-01-01

Background Although diseases associated with microvascular endothelial dysfunction are among the most prevalent illnesses to date, currently no method exists to isolate pure endothelial cells (EC) from skeletal muscle for in vivo or in vitro study. Methodology By utilizing multicolor fluorescent-activated cell sorting (FACS), we have isolated a distinct population of Sca-1+, CD31+, CD34dim and CD45− cells from skeletal muscles of C57BL6 mice. Characterization of this population revealed these cells are functional EC that can be expanded several times in culture without losing their phenotype or capabilities to uptake acetylated low-density lipoprotein (ac-LDL), produce nitric oxide (NO) and form vascular tubes. When transplanted subcutaneously or intramuscularly into the tibialis anterior muscle, EC formed microvessels and integrated with existing vasculature. Conclusion This method, which is highly reproducible, can be used to study the biology and role of EC in diseases such as peripheral vascular disease. In addition this method allows us to isolate large quantities of skeletal muscle derived EC with potential for therapeutic angiogenic applications. PMID:18335025

Interleukin 2 Activates Brain Microvascular Endothelial Cells Resulting in Destabilization of Adherens Junctions.

PubMed

Wylezinski, Lukasz S; Hawiger, Jacek

2016-10-28

The pleiotropic cytokine interleukin 2 (IL2) disrupts the blood-brain barrier and alters brain microcirculation, underlying vascular leak syndrome that complicates cancer immunotherapy with IL2. The microvascular effects of IL2 also play a role in the development of multiple sclerosis and other chronic neurological disorders. The mechanism of IL2-induced disruption of brain microcirculation has not been determined previously. We found that both human and murine brain microvascular endothelial cells express constituents of the IL2 receptor complex. Then we established that signaling through this receptor complex leads to activation of the transcription factor, nuclear factor κB, resulting in expression of proinflammatory interleukin 6 and monocyte chemoattractant protein 1. We also discovered that IL2 induces disruption of adherens junctions, concomitant with cytoskeletal reorganization, ultimately leading to increased endothelial cell permeability. IL2-induced phosphorylation of vascular endothelial cadherin (VE-cadherin), a constituent of adherens junctions, leads to dissociation of its stabilizing adaptor partners, p120-catenin and β-catenin. Increased phosphorylation of VE-cadherin was also accompanied by a reduction of Src homology 2 domain-containing protein-tyrosine phosphatase 2, known to maintain vascular barrier function. These results unravel the mechanism of deleterious effects induced by IL2 on brain microvascular endothelial cells and may inform the development of new measures to improve IL2 cancer immunotherapy, as well as treatments for autoimmune diseases affecting the central nervous system. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Effects of Dietary Supplementation with Brazil Nuts on Microvascular Endothelial Function in Hypertensive and Dyslipidemic Patients: A Randomized Crossover Placebo-Controlled Trial.

PubMed

Huguenin, Grazielle V B; Moreira, Annie S B; Siant'Pierre, Tatiana D; Gonçalves, Rodrigo A; Rosa, Glorimar; Oliveira, Glaucia M M; Luiz, Ronir R; Tibirica, Eduardo

2015-11-01

To investigate the effects of dietary supplementation with GBNs on microvascular endothelial function in hypertensive and dyslipidemic patients. Ninety-one patients of both sexes aged 62.1 ± 9.3 years received 13 g/day of GBNs or a placebo for three months with a washout period of one month between treatments. Microvascular endothelial function was assessed using LSCI coupled with iontophoresis of ACh and PORH. We also used skin video capillaroscopy to measure capillary density and recruitment at rest and during PORH. Plasma concentrations of NOx were also measured as a marker of nitric oxide bioavailability. Supplementation with GBNs significantly increased the plasma levels of Se (p < 0.05) and NOx (p < 0.05). However, we did not observe any effects of GBN consumption on microvascular vasodilator responses to ACh or PORH (p > 0.05), and GBNs did not improve capillary density at baseline or recruitment during PORH (p > 0.05). Supplementation with GBNs induced significant increases in the plasma Se concentration and systemic bioavailability of nitric oxide. Nevertheless, GBN supplementation did not lead to any improvement in systemic microvascular reactivity or density in patients with arterial hypertension and dyslipidemia who were undergoing multiple drug therapies. © 2015 John Wiley & Sons Ltd.

Vascular endothelial growth factor is upregulated by l-dopa in the parkinsonian brain: implications for the development of dyskinesia

PubMed Central

Francardo, Veronica; Lindgren, Hanna S.; Sillivan, Stephanie E.; O’Sullivan, Sean S.; Luksik, Andrew S.; Vassoler, Fair M.; Lees, Andrew J.; Konradi, Christine

2011-01-01

Angiogenesis and increased permeability of the blood–brain barrier have been reported to occur in animal models of Parkinson’s disease and l-dopa-induced dyskinesia, but the significance of these phenomena has remained unclear. Using a validated rat model of l-dopa-induced dyskinesia, this study demonstrates that chronic treatment with l-dopa dose dependently induces the expression of vascular endothelial growth factor in the basal ganglia nuclei. Vascular endothelial growth factor was abundantly expressed in astrocytes and astrocytic processes in the proximity of blood vessels. When co-administered with l-dopa, a small molecule inhibitor of vascular endothelial growth factor signalling significantly attenuated the development of dyskinesia and completely blocked the angiogenic response and associated increase in blood–brain barrier permeability induced by the treatment. The occurrence of angiogenesis and vascular endothelial growth factor upregulation was verified in post-mortem basal ganglia tissue from patients with Parkinson’s disease with a history of dyskinesia, who exhibited increased microvascular density, microvascular nestin expression and an upregulation of vascular endothelial growth factor messenger ribonucleic acid. These congruent findings in the rat model and human patients indicate that vascular endothelial growth factor is implicated in the pathophysiology of l-dopa-induced dyskinesia and emphasize an involvement of the microvascular compartment in the adverse effects of l-dopa pharmacotherapy in Parkinson’s disease. PMID:21771855

Endothelial sirtuin 1 inactivation enhances capillary rarefaction and fibrosis following kidney injury through Notch activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kida, Yujiro; Zullo, Joseph A.; Renal Research Institute, Department of Physiology, New York Medical College, Valhalla, NY

Peritubular capillary (PTC) rarefaction along with tissue fibrosis is a hallmark of chronic kidney disease (CKD). However, molecular mechanisms of PTC loss have been poorly understood. Previous studies have demonstrated that functional loss of endothelial sirtuin 1 (SIRT1) impairs angiogenesis during development and tissue damage. Here, we found that endothelial SIRT1 dysfunction causes activation of endothelial Notch1 signaling, which leads to PTC rarefaction and fibrosis following kidney injury. In mice lacking functional SIRT1 in the endothelium (Sirt1 mutant), kidney injury enhanced apoptosis and senescence of PTC endothelial cells with impaired endothelial proliferation and expanded myofibroblast population and collagen deposition. Comparedmore » to wild-type kidneys, Sirt1 mutant kidneys up-regulated expression of Delta-like 4 (DLL4, a potent Notch1 ligand), Hey1 and Hes1 (Notch target genes), and Notch intracellular domain-1 (NICD1, active form of Notch1) in microvascular endothelial cells (MVECs) post-injury. Sirt1 mutant primary kidney MVECs reduced motility and vascular assembly and enhanced senescence compared to wild-type kidney MVECs. This difference in the phenotype was negated with Notch inhibition. Concurrent stimulation of DLL4 and transforming growth factor (TGF)-β1 increased trans-differentiation of primary kidney pericytes into myofibroblast more than TGF-β1 treatment alone. Collectively, these results indicate that endothelial SIRT1 counteracts PTC rarefaction by repression of Notch1 signaling and antagonizes fibrosis via suppression of endothelial DLL4 expression. - Highlights: • SIRT1 represses Notch1 signaling in capillary endothelial cells in the kidney. • Endothelial SIRT1 is depleted in the kidney following injury. • Activation of endothelial Notch impairs angiogenesis in the kidney. • Increased expression of endothelial DLL4 enhances renal fibrosis.« less

Expression of Biglycan in First Trimester Chorionic Villous Sampling Placental Samples and Altered Function in Telomerase-Immortalized Microvascular Endothelial Cells.

PubMed

Chui, Amy; Gunatillake, Tilini; Brennecke, Shaun P; Ignjatovic, Vera; Monagle, Paul T; Whitelock, John M; van Zanten, Dagmar E; Eijsink, Jasper; Wang, Yao; Deane, James; Borg, Anthony J; Stevenson, Janet; Erwich, Jan Jaap; Said, Joanne M; Murthi, Padma

2017-06-01

Biglycan (BGN) has reduced expression in placentae from pregnancies complicated by fetal growth restriction (FGR). We used first trimester placental samples from pregnancies with later small for gestational age (SGA) infants as a surrogate for FGR. The functional consequences of reduced BGN and the downstream targets of BGN were determined. Furthermore, the expression of targets was validated in primary placental endothelial cells isolated from FGR or control pregnancies. APPROACH AND RESULTS: BGN expression was determined using real-time polymerase chain reaction in placental tissues collected during chorionic villous sampling performed at 10 to 12 weeks' gestation from pregnancies that had known clinical outcomes, including SGA. Short-interference RNA reduced BGN expression in telomerase-immortalized microvascular endothelial cells, and the effect on proliferation, angiogenesis, and thrombin generation was determined. An angiogenesis array identified downstream targets of BGN, and their expression in control and FGR primary placental endothelial cells was validated using real-time polymerase chain reaction. Reduced BGN expression was observed in SGA placental tissues. BGN reduction decreased network formation of telomerase-immortalized microvascular endothelial cells but did not affect thrombin generation or cellular proliferation. The array identified target genes, which were further validated: angiopoetin 4 ( ANGPT4 ), platelet-derived growth factor receptor α ( PDGFRA ), tumor necrosis factor superfamily member 15 ( TNFSF15 ), angiogenin ( ANG ), serpin family C member 1 ( SERPIN1 ), angiopoietin 2 ( ANGPT2 ), and CXC motif chemokine 12 ( CXCL12 ) in telomerase-immortalized microvascular endothelial cells and primary placental endothelial cells obtained from control and FGR pregnancies. This study reports a temporal relationship between altered placental BGN expression and subsequent development of SGA. Reduction of BGN in vascular endothelial cells leads to disrupted network formation and alterations in the expression of genes involved in angiogenesis. Therefore, differential expression of these may contribute to aberrant angiogenesis in SGA pregnancies. © 2017 American Heart Association, Inc.

Effects of cryopreservation on excretory function, cellular adhesion molecules and vessel lumen formation in human umbilical vein endothelial cells.

PubMed

Cai, Guoping; Lai, Binbin; Hong, Huaxing; Lin, Peng; Chen, Weifu; Zhu, Zhong; Chen, Haixiao

2017-07-01

Cryopreservation is widely used in regenerative medicine for tissue preservation. In the present study, the effects of cryopreservation on excretory function, cellular adhesion molecules and vessel lumen formation in human umbilical vein endothelial cells (HUVECs) were investigated. After 0, 4, 8, 12 or 24 weeks of cryopreservation in liquid nitrogen, the HUVECs were thawed. The excretory functions markers (endothelin‑1, prostaglandin E1, von Willebrand factor and nitric oxide) of HUVECs were measured by ELISA assay. The expression of intercellular adhesion molecule‑1 (ICAM‑1) in HUVECs was analyzed using flow cytometry. An angiogenesis assay was used to determine the angiogeneic capabilities of the thawed HUVECs. The results demonstrated that cryopreserved/thawed and recultivated HUVECs were unsuitable for tissue‑engineered microvascular construction. Specifically, the excretory function of the cells was significantly decreased in the post‑cryopreserved HUVECs at 24 weeks. In addition, the level of ICAM‑1 in HUVECs was significantly upregulated from the fourth week of cryopreservation. Furthermore, the tube‑like structure‑forming potential was weakened with increasing cryopreservation duration, and the numbers of lumen and the length of the pipeline were decreased in the thawed HUVECs, in a time‑dependent manner. In conclusion, the results of the present study revealed that prolonged cryopreservation may lead to HUVEC dysfunction and did not create stable cell lines for tissue‑engineered microvascular construction.

Targeting Microvascular Pericytes in Angiogenic Vessels of Prostate Cancer

DTIC Science & Technology

2005-04-01

endothelial cells (BEC) were identified using a cocktail of rat antibodies against mouse endoglin ( CD 105), PECAM-1 (CD3 1), and VEGF receptor-2 (flk...immunohistochemistry, and imaging angiogenic tufts composed of endothelial cells positive for endoglin ( CD 105), PECAM- I (CD3 1), and VEGF receptor...TYPE AND DATES COVERED (Leave blank) - April 2005 Annual (1 Apr 2004 - 31 Mar 2005) 4. TITLE AND SUBTITLE 5. FUNDING NUMBERS Targeting Microvascular

Microvascular pericytes in healthy and diseased kidneys

PubMed Central

Pan, Szu-Yu; Chang, Yu-Ting; Lin, Shuei-Liong

2014-01-01

Pericytes are interstitial mesenchymal cells found in many major organs. In the kidney, microvascular pericytes are defined anatomically as extensively branched, collagen-producing cells in close contact with endothelial cells. Although many molecular markers have been proposed, none of them can identify the pericytes with satisfactory specificity or sensitivity. The roles of microvascular pericytes in kidneys were poorly understood in the past. Recently, by using genetic lineage tracing to label collagen-producing cells or mesenchymal cells, the elusive characteristics of the pericytes have been illuminated. The purpose of this article is to review recent advances in the understanding of microvascular pericytes in the kidneys. In healthy kidney, the pericytes are found to take part in the maintenance of microvascular stability. Detachment of the pericytes from the microvasculature and loss of the close contact with endothelial cells have been observed during renal insult. Renal microvascular pericytes have been shown to be the major source of scar-forming myofibroblasts in fibrogenic kidney disease. Targeting the crosstalk between pericytes and neighboring endothelial cells or tubular epithelial cells may inhibit the pericyte–myofibroblast transition, prevent peritubular capillary rarefaction, and attenuate renal fibrosis. In addition, renal pericytes deserve attention for their potential to produce erythropoietin in healthy kidneys as pericytes stand in the front line, sensing the change of oxygenation and hemoglobin concentration. Further delineation of the mechanisms underlying the reduced erythropoietin production occurring during pericyte–myofibroblast transition may be promising for the development of new treatment strategies for anemia in chronic kidney disease. PMID:24465134

The endogenous zinc finger transcription factor, ZNF24, modulates the angiogenic potential of human microvascular endothelial cells

PubMed Central

Jia, Di; Huang, Lan; Bischoff, Joyce; Moses, Marsha A.

2015-01-01

We have previously identified a zinc finger transcription factor, ZNF24 (zinc finger protein 24), as a novel inhibitor of tumor angiogenesis and have demonstrated that ZNF24 exerts this effect by repressing the transcription of VEGF in breast cancer cells. Here we focused on the role of ZNF24 in modulating the angiogenic potential of the endothelial compartment. Knockdown of ZNF24 by siRNA in human primary microvascular endothelial cells (ECs) led to significantly decreased cell migration and invasion compared with control siRNA. ZNF24 knockdown consistently led to significantly impaired VEGF receptor 2 (VEGFR2) signaling and decreased levels of matrix metalloproteinase-2 (MMP-2), with no effect on levels of major regulators of MMP-2 activity such as the tissue inhibitors of metalloproteinases and MMP-14. Moreover, silencing ZNF24 in these cells led to significantly decreased EC proliferation. Quantitative PCR array analyses identified multiple cell cycle regulators as potential ZNF24 downstream targets which may be responsible for the decreased proliferation in ECs. In vivo, knockdown of ZNF24 specifically in microvascular ECs led to significantly decreased formation of functional vascular networks. Taken together, these results demonstrate that ZNF24 plays an essential role in modulating the angiogenic potential of microvascular ECs by regulating the proliferation, migration, and invasion of these cells.— Jia, D., Huang, L., Bischoff, J., Moses, M. A. The endogenous zinc finger transcription factor, ZNF24, modulates the angiogenic potential of human microvascular endothelial cells. PMID:25550468

Coronary microvascular dysfunction equivalent to left main coronary artery disease.

PubMed

Panç, Cafer; Kocaağa, Mehmet; Erdoğan, Onur; Sarıkaya, Remzi; Umman, Sabahattin

2017-04-01

Coronary microvascular dysfunction, also known as cardiac syndrome X, is a clinical syndrome presenting with typical angina and evidence of myocardial ischemia in the absence of flow-limiting stenosis on coronary angiography. Of patients undergoing coronary angiography due to suspected myocardial ischemia, 50% are found to have normal or near-normal coronary arteries. Described in this case report is a patient who developed hypotension and ST segment depressions during treadmill exercise test. Left main coronary artery or multivessel disease was suspected. Coronary angiography was normal, but coronary flow reserve measurement revealed severe microvascular dysfunction.

Ethnic differences in microvascular function in apparently healthy South African men and women.

PubMed

Pienaar, P R; Micklesfield, L K; Gill, J M R; Shore, A C; Gooding, K M; Levitt, N S; Lambert, E V

2014-07-01

Microvascular dysfunction precedes the clinical manifestations of cardiovascular disease. Given the ethnic disparities in cardiovascular disease, we aimed to investigate ethnic differences in microvascular endothelial function in a group of young (18-33 years old), apparently healthy individuals (n = 33, nine Black African, 12 mixed ancestry and 12 Caucasian). Microvascular endothelium-dependent and -independent function was assessed by laser Doppler imagery and iontophoresis of ACh and sodium nitroprusside (SNP), respectively, adjusting for skin resistance. Microvascular reactivity was expressed as maximum absolute perfusion, percentage change from baseline and area under the curve (AUC). Skin resistance was significantly lower in the Caucasian group in response to ACh (Caucasian, mean 0.16 ± 0.03 Ω versus Black, 0.21 ± 0.04 Ω and mixed ancestry, 0.20 ± 0.02 Ω, P < 0.01) and SNP (Caucasian, 0.08 ± 0.01 Ω versus Black, 0.11 ± 0.02 Ω and mixed ancestry, 0.12 ± 0.01 Ω, P < 0.01). Microvascular function in response to ACh was significantly higher in the Caucasian group compared with the other two groups; however, after adjusting for skin resistance these differences were no longer significant. Conversely, the microvascular SNP response remained significantly higher in the Caucasian group, even after adjusting for skin resistance (P < 0.01). Diastolic blood pressure was inversely associated with the AUC of ACh (r = -0.4) and all SNP responses (r = -0.3 to -0.6). Skin resistance was inversely associated with AUC and maximum absolute ACh response (r = -0.59 and -0.64, respectively) and all SNP responses (r = -0.37 to -0.79). Ethnic differences in endothelium-independent microvascular function may contribute to ethnic disparities in cardiovascular disease. Moreover, skin resistance plays a significant role in the interpretation of the microvascular response to outcomes of iontophoresis in a multiethnic group. © 2014 The Authors. Experimental Physiology © 2014 The Physiological Society.

Beneficial effects of intracoronary nicorandil on microvascular dysfunction after primary percutaneous coronary intervention: demonstration of its superiority to nitroglycerin in a cross-over study.

PubMed

Ito, Noritoshi; Nanto, Shinsuke; Doi, Yasuji; Kurozumi, Yuma; Natsukawa, Tomoaki; Shibata, Hiroyuki; Morita, Masaya; Kawata, Atsushi; Tsuruoka, Ayumu; Sawano, Hirotaka; Okada, Ken-ichiro; Sakata, Yasuhiko; Kai, Tatsuro; Hayashi, Toru

2013-08-01

In patients undergoing primary percutaneous coronary intervention (PCI) for the treatment of ST-segment elevation myocardial infarction (STEMI), coronary microvascular dysfunction is associated with poor prognosis. Coronary microvascular resistance is predominantly regulated by ATP-sensitive potassium (KATP) channels. The aim of this study was to clarify whether nicorandil, a hybrid KATP channel opener and nitric oxide donor, may be a good candidate for improving microvascular dysfunction even when administered after primary PCI. We compared the beneficial effects of nicorandil and nitroglycerin on microvascular function in 60 consecutive patients with STEMI. After primary PCI, all patients received single intracoronary administrations of nitroglycerin (250 μg) and nicorandil (2 mg) in a randomized order; 30 received nicorandil first, while the other 30 received nitroglycerin first. Microvascular dysfunction was evaluated with the index of microcirculatory resistance (IMR), defined as the distal coronary pressure multiplied by the hyperemic mean transit time. As a first administration, nicorandil decreased IMR significantly more than did nitroglycerin (median [interquartile ranges]: 10.8[5.2-20.7] U vs. 2.1[1.0-6.0] U, p=0.0002).As a second administration, nicorandil further decreased IMR, while nitroglycerin did not (median [interquartile ranges]: 6.0[1.3-12.7] U vs. -1.4[-2.6 to 1.3] U, p<0.0001). The IMR after the second administration was significantly associated with myocardial blush grade, angiographic TIMI frame count after the procedure, and peak creatine kinase level. Intracoronary nicorandil reduced microvascular dysfunction after primary PCI more effectively than did nitroglycerin in patients with STEMI, probably via its KATP channel-opening effect.

Prolonged superficial local cryotherapy attenuates microcirculatory impairment, regional inflammation, and muscle necrosis after closed soft tissue injury in rats.

PubMed

Schaser, Klaus-Dieter; Disch, Alexander C; Stover, John F; Lauffer, Annette; Bail, Herman J; Mittlmeier, Thomas

2007-01-01

Closed soft tissue injury induces progressive microvascular dysfunction and regional inflammation. The authors tested the hypothesis that adverse trauma-induced effects can be reduced by local cooling. While superficial cooling reduces swelling, pain, and cellular oxygen demand, the effects of cryotherapy on posttraumatic microcirculation are incompletely understood. Controlled laboratory study. After a standardized closed soft tissue injury to the left tibial compartment, male rats were randomly subjected to percutaneous perfusion for 6 hours with 0.9% NaCL (controls; room temperature) or cold NaCL (cryotherapy; 8 degrees C) (n = 7 per group). Uninjured rats served as shams (n = 7). Microcirculatory changes and leukocyte adherence were determined by intravital microscopy. Intramuscular pressure was measured, and invasion of granulocytes and macrophages was assessed by immunohistochemistry. Edema and tissue damage was quantified by gravimetry and decreased desmin staining. Closed soft tissue injury significantly decreased functional capillary density (240 +/- 12 cm(-1)); increased microvascular permeability (0.75 +/- 0.03), endothelial leukocyte adherence (995 +/- 77/cm(2)), granulocyte (182.0 +/- 25.5/mm(2)) and macrophage infiltration, edema formation, and myonecrosis (ratio: 2.95 +/- 0.45) within the left extensor digitorum longus muscle. Cryotherapy for 6 hours significantly restored diminished functional capillary density (393 +/- 35), markedly decreased elevated intramuscular pressure, reduced the number of adhering (462 +/- 188/cm(2)) and invading granulocytes (119 +/- 28), and attenuated tissue damage (ratio: 1.7 +/- 0.17). The hypothesis that prolonged cooling reduces posttraumatic microvascular dysfunction, inflammation, and structural impairment was confirmed. These results may have therapeutic implications as cryotherapy after closed soft tissue injury is a valuable therapeutic approach to improve nutritive perfusion and attenuate leukocyte-mediated tissue destruction. The risk for evolving compartment syndrome may be reduced, thereby preventing further irreversible aggravation.

Uterine microvascular sensitivity to nanomaterial inhalation: An in vivo assessment☆

PubMed Central

Stapleton, P.A.; McBride, C.R.; Yi, J.; Nurkiewicz, T.R.

2015-01-01

With the tremendous number and diverse applications of engineered nanomaterials incorporated in daily human activity, exposure can no longer be solely confined to occupational exposures of healthy male models. Cardiovascular and endothelial cell dysfunction have been established using in vitro and in situ preparations, but the translation to intact in vivo models is limited. Intravital microscopy has been used extensively to understand microvascular physiology while maintaining in vivo neurogenic, humoral, and myogenic control. However, a tissue specific model to assess the influences of nanomaterial exposure on female reproductive health has not been fully elucidated. Female Sprague Dawley (SD) rats were exposed to nano-TiO2 aerosols (171 ± 6 nm, 10.1 ± 0.39 mg/m3, 5 h) 24-hours prior to experimentation, leading to a calculated deposition of 42.0 ± 1.65 μg. After verifying estrus status, vital signs were monitored and the right horn of the uterus was exteriorized, gently secured over an optical pedestal, and enclosed in a warmed tissue bath using intravital microscopy techniques. After equilibration, significantly higher leukocyte-endothelium interactions were recorded in the exposed group. Arteriolar responsiveness was assessed using ionophoretically applied agents: muscarinic agonist acetylcholine (0.025 M; ACh; 20, 40, 100, and 200 nA), and nitric oxide donor sodium nitroprusside (0.05 M; SNP; 20, 40, and 100 nA), or adrenergic agonist phenylephrine (0.05 M; PE; 20, 40, and 100 nA) using glass micropipettes. Passive diameter was established by tissue superfusion with 10−4 M adenosine. Similar to male counterparts, female SD rats present systemic microvascular dysfunction; however the ramifications associated with female health and reproduction have yet to be elucidated. PMID:26375943

Predictive value of vascular disease biomarkers for digital ulcers in systemic sclerosis patients.

PubMed

Silva, Ivone; Teixeira, Andreia; Oliveira, José; Almeida, Isabel; Almeida, Rui; Vasconcelos, Carlos

2015-01-01

To investigate the role of endothelial dysfunction and angiogenesis vascular biomarkers as risk factors and their predictive value for digital ulcers in systemic sclerosis patients. Endothelin-1 (ET-1), asymmetric dimethylarginine (ADMA), vascular endothelial growth factor (VEGF), endostatin and endoglin were measured in an observational prospective cohort of 77 SSc patients. The primary outcome was the occurrence of one or more new ischaemic digital ulcers during a planned 3-year follow-up. After the 3-year follow-up, 40 patients developed new digital ulcers. Logistic regression confirmed VEGF (HR 1.128, 95% CI 1.010-1.260, p=0.033) and ADMA (HR 0.995, 95% CI 0.991-0.998, p=0.006) as independent predictors of new digital ulcers. Patients with serum levels of ET-1>11.9 pmol/ml (p<0.001) and VEGF<422.47 pg/ml (p=0.028) had significantly more DU in the 3-year follow-up. Although not significant, a trend towards increased serum levels of endoglin>4.215 ng/ml (p=0.053) was associated to a new DU episode. No predictive serum value was found for ADMA (p=0.075) and endostatin (p=0.130). Endothelial dysfunction and angiogenic vascular biomarkers have an important role in the underlying and in the progression of microvascular disease in systemic sclerosis. Increased serum levels of ET-1, ADMA and VEGF are strong predictors of severe microangiopathy complications, namely ischaemic digital ulcers.

Hypokalemia correlated with arterial stiffness but not microvascular endothelial function in patients with primary aldosteronism.

PubMed

Chang, Yi-Yao; Chen, Aaron; Chen, Ying-Hsien; Hung, Chi-Sheng; Wu, Vin-Cent; Wu, Xue-Ming; Lin, Yen-Hung; Ho, Yi-Lwun; Wu, Kwan-Dun

2015-06-01

Hypokalemia in primary aldosteronism (PA) patients correlates with higher levels of cardiovascular events and altered left ventricular geometry. However, the influence of aldosterone on microvascular endothelial function and the effect of hypokalemia on the vascular structure still remain unclear. We investigated the peripheral arterial functions, including the endothelial function of microvasculature and arterial stiffness in PA and essential hypertension (EH) patients, and the correlation between hypokalemia and peripheral arterial function among PA patients. Twenty patients diagnosed as EH and 37 patients with PA were enrolled in this study. Reactive hyperemia index (RHI) and the augmentation index (AI) were obtained by non-invasive peripheral arterial tonometry. Twenty EH patients and a total of 37 PA patients, including 21 patients with normokalemia and 16 patients with hypokalemia, were enrolled and divided into groups 1, 2 and 3 respectively. PA patients had significantly higher AI (p=0.024) but not RHI than EH patients. RHI showed no difference between groups 1, 2 and 3. Group 3 had higher AI than either group 1 or group 2. In the whole study population, serum potassium level, after multivariate regression analysis testing, was the only factor associated with AI (ß= -0.102; p=0.002). In PA patients, serum potassium level was the only significant factor correlated with AI. (r= -0.458; p=0.004) CONCLUSIONS: PA patients had higher arterial stiffness but comparable microvascular endothelial function to EH patients. Hypokalemia correlated with arterial stiffness but not microvascular endothelial function in PA patients. © The Author(s) 2014.

TPL2 (Therapeutic Targeting Tumor Progression Locus-2)/ATF4 (Activating Transcription Factor-4)/SDF1α (Chemokine Stromal Cell-Derived Factor-α) Axis Suppresses Diabetic Retinopathy.

PubMed

Lai, De-Wei; Lin, Keng-Hung; Sheu, Wayne Huey-Herng; Lee, Maw-Rong; Chen, Chung-Yu; Lee, Wen-Jane; Hung, Yi-Wen; Shen, Chin-Chang; Chung, Tsung-Ju; Liu, Shing-Hwa; Sheu, Meei-Ling

2017-09-01

Diabetic retinopathy is characterized by vasopermeability, vascular leakage, inflammation, blood-retinal barrier breakdown, capillary degeneration, and neovascularization. However, the mechanisms underlying the association between diabetes mellitus and progression retinopathy remain unclear. TPL2 (tumor progression locus 2), a serine-threonine protein kinase, exerts a pathological effect on vascular angiogenesis. This study investigated the role of N ε -(carboxymethyl)lysine, a major advanced glycation end products, and the involved TPL2-related molecular signals in diabetic retinopathy using models of in vitro and in vivo and human samples. Serum N ε -(carboxymethyl)lysine levels and TPL2 kinase activity were significantly increased in clinical patients and experimental animals with diabetic retinopathy. Intravitreal administration of pharmacological blocker or neutralizing antibody inhibited TPL2 and effectively suppressed the pathological characteristics of retinopathy in streptozotocin-induced diabetic animal models. Intravitreal VEGF (vascular endothelial growth factor) neutralization also suppressed the diabetic retinopathy in diabetic animal models. Mechanistic studies in primary human umbilical vein endothelial cells and primary retinal microvascular endothelial cells from streptozotocin-diabetic rats, db/db mice, and samples from patients with diabetic retinopathy revealed a positive parallel correlation between N ε -(carboxymethyl)lysine and the TPL2/chemokine SDF1α (stromal cell-derived factor-α) axis that is dependent on endoplasmic reticulum stress-related molecules, especially ATF4 (activating transcription factor-4). This study demonstrates that inhibiting the N ε -(carboxymethyl)lysine-induced TPL2/ATF4/SDF1α axis can effectively prevent diabetes mellitus-mediated retinal microvascular dysfunction. This signaling axis may include the therapeutic potential for other diseases involving pathological neovascularization or macular edema. © 2017 American Heart Association, Inc.

Hyperglycaemia promotes human brain microvascular endothelial cell apoptosis via induction of protein kinase C-ßI and prooxidant enzyme NADPH oxidase

PubMed Central

Shao, Beili; Bayraktutan, Ulvi

2014-01-01

Blood–brain barrier disruption represents a key feature in hyperglycaemia-aggravated cerebral damage after an ischaemic stroke. Although the underlying mechanisms remain largely unknown, activation of protein kinase C (PKC) is thought to play a critical role. This study examined whether apoptosis of human brain microvascular endothelial cells (HBMEC) might contribute to hyperglycaemia-evoked barrier damage and assessed the specific role of PKC in this phenomenon. Treatments with hyperglycaemia (25 mM) or phorbol myristate acetate (PMA, a protein kinase C activator, 100 nM) significantly increased NADPH oxidase activity, O2•- generation, proapoptotic protein Bax expression, TUNEL-positive staining and caspase-3/7 activities. Pharmacological inhibition of NADPH oxidase, PKC-a, PKC-ß or PKC-ßI via their specific inhibitors and neutralisation of O2•- by a cell-permeable superoxide dismutase mimetic, MnTBAP normalised all the aforementioned increases induced by hyperglycaemia. Suppression of these PKC isoforms also negated the stimulatory effects of hyperglycaemia on the protein expression of NADPH oxidase membrane-bound components, Nox2 and p22-phox which determine the overall enzymatic activity. Silencing of PKC-ßI gene through use of specific siRNAs abolished the effects of both hyperglycaemia and PMA on endothelial cell NADPH oxidase activity, O2•- production and apoptosis and consequently improved the integrity and function of an in vitro model of human cerebral barrier comprising HBMEC, astrocytes and pericytes. Hyperglycaemia-mediated apoptosis of HBMEC contributes to cerebral barrier dysfunction and is modulated by sequential activations of PKC-ßI and NADPH oxidase. PMID:24936444

Biomarkers of Endothelial Activation Are Associated with Poor Outcome in Critical Illness.

PubMed

Mikacenic, Carmen; Hahn, William O; Price, Brenda L; Harju-Baker, Susanna; Katz, Ronit; Kain, Kevin C; Himmelfarb, Jonathan; Liles, W Conrad; Wurfel, Mark M

2015-01-01

Endothelial activation plays a role in organ dysfunction in the systemic inflammatory response syndrome (SIRS). Angiopoietin-1 (Ang-1) promotes vascular quiescence while angiopoietin-2 (Ang-2) mediates microvascular leak. Circulating levels of Ang-1 and Ang-2 in patients with SIRS could provide insight on risks for organ dysfunction and death distinct from inflammatory proteins. In this study, we determined if biomarkers of endothelial activation and inflammation exhibit independent associations with poor outcomes in SIRS. We studied 943 critically ill patients with SIRS admitted to an Intensive Care Unit (ICU) of an academic medical center. We measured plasma levels of endothelial markers (Ang-1, Ang-2, soluble vascular cell adhesion molecule-1 (sVCAM-1)) and inflammatory markers (interleukin-6 (IL-6), interleukin-8 (IL-8), granulocyte-colony stimulating factor (G-CSF), soluble tumor necrosis factor receptor-1 (sTNFR-1)) within 24 hours of enrollment. We tested for associations between each marker and 28 day mortality, shock, and day 3 sequential organ failure assessment (SOFA) score. For 28 day mortality, we performed sensitivity analysis for those subjects with sepsis and those with sterile inflammation. We used multivariate models to adjust for clinical covariates and determine if associations identified with endothelial activation markers were independent of those observed with inflammatory markers. Higher levels of all biomarkers were associated with increased 28 day mortality except levels of Ang-1 which were associated with lower mortality. After adjustment for comorbidities and sTNFR-1 concentration, a doubling of Ang-1 concentration was associated with lower 28 day mortality (Odds ratio (OR) = 0.81; p<0.01), shock (OR = 0.82; p<0.001), and SOFA score (β = -0.50; p<0.001), while Ang-2 concentration was associated with increased mortality (OR = 1.55; p<0.001), shock (OR = 1.51; p<0.001), and SOFA score (β = +0.63; p<0.001). sVCAM-1 was not independently associated with SIRS outcomes. In critically ill patients with SIRS, early measurements of Ang-1 and Ang-2 are associated with death and organ dysfunction independently of simultaneously-measured markers of inflammation.

A Novel ex vivo Mouse Mesometrium Culture Model for Investigating Angiogenesis in Microvascular Networks.

PubMed

Suarez-Martinez, Ariana D; Bierschenk, Susanne; Huang, Katie; Kaplan, Dana; Bayer, Carolyn L; Meadows, Stryder M; Sperandio, Markus; Murfee, Walter L

2018-05-18

The development of models that incorporate intact microvascular networks enables the investigation of multicellular dynamics during angiogenesis. Our laboratory introduced the rat mesentery culture model as such a tool, which would be enhanced with mouse tissue. Since mouse mesentery is avascular, an alternative is mouse mesometrium, the connective tissue of uterine horns. The study's objective was to demonstrate that mouse mesometrium contains microvascular networks that can be cultured to investigate multicellular dynamics during angiogenesis. Harvested mesometrium tissues from C57Bl/6 female mice were cultured in media with serum for up to 7 days. PECAM, NG2, αSMA, and LYVE-1 labeling identified endothelial cells, pericytes, smooth muscle cells, and lymphatic endothelial cells, respectively. These cells comprised microvascular networks with arterioles, venules, and capillaries. Compared to day 0, capillary sprouts per vascular length were increased by 3 and 5 days in culture (day 0, 0.08 ± 0.01; day 3, 3.19 ± 0.78; day 5, 2.49 ± 0.05 sprouts/mm; p < 0.05). Time-lapse imaging of cultured tissues from FlkEGFP mice showcases the use of the model for lineage studies. The impact is supported by the identification of endothelial cell jumping from one sprout to another. These results introduce a novel culture model for investigating multicellular dynamics during angiogenesis in real-time ex vivo microvascular networks. © 2018 S. Karger AG, Basel.

Antiproliferative effect of elevated glucose in human microvascular endothelial cells

NASA Technical Reports Server (NTRS)

Kamal, K.; Du, W.; Mills, I.; Sumpio, B. E.

1998-01-01

Diabetic microangiopathy has been implicated as a fundamental feature of the pathological complications of diabetes including retinopathy, neuropathy, and diabetic foot ulceration. However, previous studies devoted to examining the deleterious effects of elevated glucose on the endothelium have been performed largely in primary cultured cells of macrovessel origin. Difficulty in the harvesting and maintenance of microvascular endothelial cells in culture have hindered the study of this relevant population. Therefore, the objective of this study was to characterize the effect of elevated glucose on the proliferation and involved signaling pathways of an immortalized human dermal microvascular endothelial cell line (HMEC-1) that possess similar characteristics to their in vivo counterparts. Human dermal microvascular endothelial cells (HMEC-1) were grown in the presence of normal (5 mM) or high D-glucose (20 mM) for 14 days. The proliferative response of HMEC-1 was compared under these conditions as well as the cAMP and PKC pathways by in vitro assays. Elevated glucose significantly inhibited (P < 0.05) HMEC-1 proliferation after 7, 10, and 14 days. This effect was not mimicked by 20 mM mannitol. The antiproliferative effect was more pronounced with longer exposure (1-14 days) to elevated glucose and was irreversible 4 days after a 10-day exposure. The antiproliferative effect was partially reversed in the presence of a PKA inhibitor, Rp-cAMP (10-50 microM), and/or a PKC inhibitor, Calphostin C (10 nM). HMEC-1 exposed to elevated glucose (20 mM) for 14 days caused an increase in cyclic AMP accumulation, PKA, and PKC activity but was not associated with the activation of downstream events such as CRE and AP-1 binding activity. These data support the hypothesis that HMEC-1 is a suitable model to study the deleterious effects of elevated glucose on microvascular endothelial cells. Continued studies with HMEC-1 may prove advantageous in delineation of the molecular pathophysiology associated with diabetic microangiopathy.

Clathrin-dependent entry and vesicle-mediated exocytosis define insulin transcytosis across microvascular endothelial cells

PubMed Central

Azizi, Paymon M.; Zyla, Roman E.; Guan, Sha; Wang, Changsen; Liu, Jun; Bolz, Steffen-Sebastian; Heit, Bryan; Klip, Amira; Lee, Warren L.

2015-01-01

Transport of insulin across the microvasculature is necessary to reach its target organs (e.g., adipose and muscle tissues) and is rate limiting in insulin action. Morphological evidence suggests that insulin enters endothelial cells of the microvasculature, and studies with large vessel–derived endothelial cells show insulin uptake; however, little is known about the actual transcytosis of insulin and how this occurs in the relevant microvascular endothelial cells. We report an approach to study insulin transcytosis across individual, primary human adipose microvascular endothelial cells (HAMECs), involving insulin uptake followed by vesicle-mediated exocytosis visualized by total internal reflection fluorescence microscopy. In this setting, fluorophore-conjugated insulin exocytosis depended on its initial binding and uptake, which was saturable and much greater than in muscle cells. Unlike its degradation within muscle cells, insulin was stable within HAMECs and escaped lysosomal colocalization. Insulin transcytosis required dynamin but was unaffected by caveolin-1 knockdown or cholesterol depletion. Instead, insulin transcytosis was significantly inhibited by the clathrin-mediated endocytosis inhibitor Pitstop 2 or siRNA-mediated clathrin depletion. Accordingly, insulin internalized for 1 min in HAMECs colocalized with clathrin far more than with caveolin-1. This study constitutes the first evidence of vesicle-mediated insulin transcytosis and highlights that its initial uptake is clathrin dependent and caveolae independent. PMID:25540431

Lipopolysaccharide-induced pulmonary endothelial barrier disruption and lung edema: critical role for bicarbonate stimulation of AC10.

PubMed

Nickols, Jordan; Obiako, Boniface; Ramila, K C; Putinta, Kevin; Schilling, Sarah; Sayner, Sarah L

2015-12-15

Bacteria-induced sepsis is a common cause of pulmonary endothelial barrier dysfunction and can progress toward acute respiratory distress syndrome. Elevations in intracellular cAMP tightly regulate pulmonary endothelial barrier integrity; however, cAMP signals are highly compartmentalized: whether cAMP is barrier-protective or -disruptive depends on the compartment (plasma membrane or cytosol, respectively) in which the signal is generated. The mammalian soluble adenylyl cyclase isoform 10 (AC10) is uniquely stimulated by bicarbonate and is expressed in pulmonary microvascular endothelial cells (PMVECs). Elevated extracellular bicarbonate increases cAMP in PMVECs to disrupt the endothelial barrier and increase the filtration coefficient (Kf) in the isolated lung. We tested the hypothesis that sepsis-induced endothelial barrier disruption and increased permeability are dependent on extracellular bicarbonate and activation of AC10. Our findings reveal that LPS-induced endothelial barrier disruption is dependent on extracellular bicarbonate: LPS-induced barrier failure and increased permeability are exacerbated in elevated bicarbonate compared with low extracellular bicarbonate. The AC10 inhibitor KH7 attenuated the bicarbonate-dependent LPS-induced barrier disruption. In the isolated lung, LPS failed to increase Kf in the presence of minimal perfusate bicarbonate. An increase in perfusate bicarbonate to the physiological range (24 mM) revealed the LPS-induced increase in Kf, which was attenuated by KH7. Furthermore, in PMVECs treated with LPS for 6 h, there was a dose-dependent increase in AC10 expression. Thus these findings reveal that LPS-induced pulmonary endothelial barrier failure requires bicarbonate activation of AC10. Copyright © 2015 the American Physiological Society.

Lipopolysaccharide-induced pulmonary endothelial barrier disruption and lung edema: critical role for bicarbonate stimulation of AC10

PubMed Central

Nickols, Jordan; Obiako, Boniface; Ramila, K. C.; Putinta, Kevin; Schilling, Sarah

2015-01-01

Bacteria-induced sepsis is a common cause of pulmonary endothelial barrier dysfunction and can progress toward acute respiratory distress syndrome. Elevations in intracellular cAMP tightly regulate pulmonary endothelial barrier integrity; however, cAMP signals are highly compartmentalized: whether cAMP is barrier-protective or -disruptive depends on the compartment (plasma membrane or cytosol, respectively) in which the signal is generated. The mammalian soluble adenylyl cyclase isoform 10 (AC10) is uniquely stimulated by bicarbonate and is expressed in pulmonary microvascular endothelial cells (PMVECs). Elevated extracellular bicarbonate increases cAMP in PMVECs to disrupt the endothelial barrier and increase the filtration coefficient (Kf) in the isolated lung. We tested the hypothesis that sepsis-induced endothelial barrier disruption and increased permeability are dependent on extracellular bicarbonate and activation of AC10. Our findings reveal that LPS-induced endothelial barrier disruption is dependent on extracellular bicarbonate: LPS-induced barrier failure and increased permeability are exacerbated in elevated bicarbonate compared with low extracellular bicarbonate. The AC10 inhibitor KH7 attenuated the bicarbonate-dependent LPS-induced barrier disruption. In the isolated lung, LPS failed to increase Kf in the presence of minimal perfusate bicarbonate. An increase in perfusate bicarbonate to the physiological range (24 mM) revealed the LPS-induced increase in Kf, which was attenuated by KH7. Furthermore, in PMVECs treated with LPS for 6 h, there was a dose-dependent increase in AC10 expression. Thus these findings reveal that LPS-induced pulmonary endothelial barrier failure requires bicarbonate activation of AC10. PMID:26475732

Time-dependent effect of clonidine on microvascular permeability during endotoxemia.

PubMed

Schmidt, Karsten; Hernekamp, Jochen Frederick; Philipsenburg, Christoph; Zivkovic, Aleksandar R; Brenner, Thorsten; Hofer, Stefan

2015-09-01

Endothelial leakage with accompanying tissue edema and increased leukocyte adhesion are characteristics of the vascular inflammatory response. Tissue edema formation is a key mechanism in sepsis pathophysiology contributing to impaired tissue oxygenation and the development of shock. Sepsis mortality is directly associated with the severity of these microcirculatory alterations. Dysfunction of the sympathetic nervous system can have deleterious effects in generalized inflammation. This study evaluated the effect of the adrenergic alpha 2 agonist clonidine on microvascular permeability and leukocyte adhesion during endotoxemia. Macromolecular leakage, leukocyte adhesion, and venular wall shear rate were examined in mesenteric postcapillary venules of rats by using intravital microscopy (IVM). Lipopolysaccharide (LPS) (4mg/kg/h) or equivalent volumes of saline were continuously infused following baseline IVM at 0min. IVM was repeated after 60 and 120min in endotoxemic and nonendotoxemic animals. Clonidine (10μg/kg) was applied as an i.v. bolus. Animals received either (i) saline alone, (ii) clonidine alone, (iii) clonidine 45min prior to LPS, (iv) clonidine 10min prior to LPS, (v) clonidine 30min after LPS, or (vi) LPS alone. Due to nonparametric data distribution, Wilcoxon test and Dunn's multiple comparisons test were used for data analysis. Data were considered statistically significant at p<0.05. LPS significantly increased microvascular permeability and leukocyte adhesion and decreased venular wall shear rate. Clonidine significantly reduced microvascular permeability when applied 45min before or 30min after LPS administration. Leukocyte adhesion and venular wall shear rate were not affected by clonidine during endotoxemia. Clonidine reduces microvascular permeability in endotoxemic animals in a time-dependent manner. Adrenergic alpha 2 agonists might prove beneficial in stabilizing capillary leakage during inflammation. Copyright © 2015 Elsevier Inc. All rights reserved.

Static mechanical strain induces capillary endothelial cell cycle re-entry and sprouting.

PubMed

Zeiger, A S; Liu, F D; Durham, J T; Jagielska, A; Mahmoodian, R; Van Vliet, K J; Herman, I M

2016-08-16

Vascular endothelial cells are known to respond to a range of biochemical and time-varying mechanical cues that can promote blood vessel sprouting termed angiogenesis. It is less understood how these cells respond to sustained (i.e., static) mechanical cues such as the deformation generated by other contractile vascular cells, cues which can change with age and disease state. Here we demonstrate that static tensile strain of 10%, consistent with that exerted by contractile microvascular pericytes, can directly and rapidly induce cell cycle re-entry in growth-arrested microvascular endothelial cell monolayers. S-phase entry in response to this strain correlates with absence of nuclear p27, a cyclin-dependent kinase inhibitor. Furthermore, this modest strain promotes sprouting of endothelial cells, suggesting a novel mechanical 'angiogenic switch'. These findings suggest that static tensile strain can directly stimulate pathological angiogenesis, implying that pericyte absence or death is not necessarily required of endothelial cell re-activation.

Choroid Sprouting Assay: An Ex Vivo Model of Microvascular Angiogenesis

PubMed Central

Shao, Zhuo; Friedlander, Mollie; Hurst, Christian G.; Cui, Zhenghao; Pei, Dorothy T.; Evans, Lucy P.; Juan, Aimee M.; Tahir, Houda; Duhamel, François; Chen, Jing; Sapieha, Przemyslaw; Chemtob, Sylvain; Joyal, Jean-Sébastien; Smith, Lois E. H.

2013-01-01

Angiogenesis of the microvasculature is central to the etiology of many diseases including proliferative retinopathy, age-related macular degeneration and cancer. A mouse model of microvascular angiogenesis would be very valuable and enable access to a wide range of genetically manipulated tissues that closely approximate small blood vessel growth in vivo. Vascular endothelial cells cultured in vitro are widely used, however, isolating pure vascular murine endothelial cells is technically challenging. A microvascular mouse explant model that is robust, quantitative and can be reproduced without difficulty would overcome these limitations. Here we characterized and optimized for reproducibility an organotypic microvascular angiogenesis mouse and rat model from the choroid, a microvascular bed in the posterior of eye. The choroidal tissues from C57BL/6J and 129S6/SvEvTac mice and Sprague Dawley rats were isolated and incubated in Matrigel. Vascular sprouting was comparable between choroid samples obtained from different animals of the same genetic background. The sprouting area, normalized to controls, was highly reproducible between independent experiments. We developed a semi-automated macro in ImageJ software to allow for more efficient quantification of sprouting area. Isolated choroid explants responded to manipulation of the external environment while maintaining the local interactions of endothelial cells with neighboring cells, including pericytes and macrophages as evidenced by immunohistochemistry and fluorescence-activated cell sorting (FACS) analysis. This reproducible ex vivo angiogenesis assay can be used to evaluate angiogenic potential of pharmacologic compounds on microvessels and can take advantage of genetically manipulated mouse tissue for microvascular disease research. PMID:23922736

Endothelial E-type prostanoid 4 receptors promote barrier function and inhibit neutrophil trafficking.

PubMed

Konya, Viktoria; Üllen, Andreas; Kampitsch, Nora; Theiler, Anna; Philipose, Sonia; Parzmair, Gerald P; Marsche, Gunther; Peskar, Bernhard A; Schuligoi, Rufina; Sattler, Wolfgang; Heinemann, Akos

2013-02-01

Increased vascular permeability is a fundamental characteristic of inflammation. Substances that are released during inflammation, such as prostaglandin (PG) E(2), can counteract vascular leakage, thereby hampering tissue damage. In this study we investigated the role of PGE(2) and its receptors in the barrier function of human pulmonary microvascular endothelial cells and in neutrophil trafficking. Endothelial barrier function was determined based on electrical impedance measurements. Neutrophil recruitment was assessed based on adhesion and transendothelial migration. Morphologic alterations are shown by using immunofluorescence microscopy. We observed that activation of E-type prostanoid (EP) 4 receptor by PGE(2) or an EP4-selective agonist (ONO AE1-329) enhanced the barrier function of human microvascular lung endothelial cells. EP4 receptor activation prompted similar responses in pulmonary artery and coronary artery endothelial cells. These effects were reversed by an EP4 antagonist (ONO AE3-208), as well as by blocking actin polymerization with cytochalasin B. The EP4 receptor-induced increase in barrier function was independent of the classical cyclic AMP/protein kinase A signaling machinery, endothelial nitric oxide synthase, and Rac1. Most importantly, EP4 receptor stimulation showed potent anti-inflammatory activities by (1) facilitating wound healing of pulmonary microvascular endothelial monolayers, (2) preventing junctional and cytoskeletal reorganization of activated endothelial cells, and (3) impairing neutrophil adhesion to endothelial cells and transendothelial migration. The latter effects could be partially attributed to reduced E-selectin expression after EP4 receptor stimulation. These data indicate that EP4 agonists as anti-inflammatory agents represent a potential therapy for diseases with increased vascular permeability and neutrophil extravasation. Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

Lycium barbarum polysaccharide protects against LPS-induced ARDS by inhibiting apoptosis, oxidative stress, and inflammation in pulmonary endothelial cells.

PubMed

Chen, Lan; Li, Wen; Qi, Di; Wang, Daoxin

2018-04-01

Acute respiratory distress syndrome (ARDS) is a heterogenous syndrome characterised by diffuse alveolar damage, with an increase in lung endothelial and epithelial permeability. Lycium barbarum polysaccharide (LBP), the most biologically active fraction of wolfberry, possesses antiapoptotic and antioxidative effects in distinct situations. In the present study, the protective effects and potential molecular mechanisms of LBP against lipopolysaccharide (LPS)-induced ARDS were investigated in the mice and in the human pulmonary microvascular endothelial cells (HPMECs). The data indicated that pretreatment with LBP significantly attenuated LPS-induced lung inflammation and pulmonary oedema in vivo. LBP significantly reversed LPS-induced decrease in cell viability, increase in apoptosis and oxidative stress via inhibiting caspase-3 activation and intracellular reactive oxygen species (ROS) production in vitro. Moreover, the scratch assay verified that LBP restored the dysfunction of endothelial cells (ECs) migration induced by LPS stimulation. Furthermore, LBP also significantly suppressed LPS-induced NF-κB activation, and subsequently reversed the release of cytochrome c. These results showed the antiapoptosis and antioxidant LBP could partially protect against LPS-induced ARDS through promoting the ECs survival and scavenging ROS via inhibition of NF-κB signalling pathway. Thus, LBP could be potentially used for ARDS against pulmonary inflammation and pulmonary oedema.

Drugs of abuse and blood-brain barrier endothelial dysfunction: A focus on the role of oxidative stress

PubMed Central

Sajja, Ravi K; Rahman, Shafiqur

2015-01-01

Psychostimulants and nicotine are the most widely abused drugs with a detrimental impact on public health globally. While the long-term neurobehavioral deficits and synaptic perturbations are well documented with chronic use of methamphetamine, cocaine, and nicotine, emerging human and experimental studies also suggest an increasing incidence of neurovascular complications associated with drug abuse. Short- or long-term administration of psychostimulants or nicotine is known to disrupt blood-brain barrier (BBB) integrity/function, thus leading to an increased risk of brain edema and neuroinflammation. Various pathophysiological mechanisms have been proposed to underlie drug abuse-induced BBB dysfunction suggesting a central and unifying role for oxidative stress in BBB endothelium and perivascular cells. This review discusses drug-specific effects of methamphetamine, cocaine, and tobacco smoking on brain microvascular crisis and provides critical assessment of oxidative stress-dependent molecular pathways focal to the global compromise of BBB. Additionally, given the increased risk of human immunodeficiency virus (HIV) encephalitis in drug abusers, we have summarized the synergistic pathological impact of psychostimulants and HIV infection on BBB integrity with an emphasis on unifying role of endothelial oxidative stress. This mechanistic framework would guide further investigations on specific molecular pathways to accelerate therapeutic approaches for the prevention of neurovascular deficits by drugs of abuse. PMID:26661236

Sialic acids regulate microvessel permeability, revealed by novel in vivo studies of endothelial glycocalyx structure and function

PubMed Central

Betteridge, Kai B.; Arkill, Kenton P.; Neal, Christopher R.; Harper, Steven J.; Foster, Rebecca R.; Satchell, Simon C.; Bates, David O.

2017-01-01

Key points We have developed novel techniques for paired, direct, real‐time in vivo quantification of endothelial glycocalyx structure and associated microvessel permeability.Commonly used imaging and analysis techniques yield measurements of endothelial glycocalyx depth that vary by over an order of magnitude within the same vessel.The anatomical distance between maximal glycocalyx label and maximal endothelial cell plasma membrane label provides the most sensitive and reliable measure of endothelial glycocalyx depth.Sialic acid residues of the endothelial glycocalyx regulate glycocalyx structure and microvessel permeability to both water and albumin. Abstract The endothelial glycocalyx forms a continuous coat over the luminal surface of all vessels, and regulates multiple vascular functions. The contribution of individual components of the endothelial glycocalyx to one critical vascular function, microvascular permeability, remains unclear. We developed novel, real‐time, paired methodologies to study the contribution of sialic acids within the endothelial glycocalyx to the structural and functional permeability properties of the same microvessel in vivo. Single perfused rat mesenteric microvessels were perfused with fluorescent endothelial cell membrane and glycocalyx labels, and imaged with confocal microscopy. A broad range of glycocalyx depth measurements (0.17–3.02 μm) were obtained with different labels, imaging techniques and analysis methods. The distance between peak cell membrane and peak glycocalyx label provided the most reliable measure of endothelial glycocalyx anatomy, correlating with paired, numerically smaller values of endothelial glycocalyx depth (0.078 ± 0.016 μm) from electron micrographs of the same portion of the same vessel. Disruption of sialic acid residues within the endothelial glycocalyx using neuraminidase perfusion decreased endothelial glycocalyx depth and increased apparent solute permeability to albumin in the same vessels in a time‐dependent manner, with changes in all three true vessel wall permeability coefficients (hydraulic conductivity, reflection coefficient and diffusive solute permeability). These novel technologies expand the range of techniques that permit direct studies of the structure of the endothelial glycocalyx and dependent microvascular functions in vivo, and demonstrate that sialic acid residues within the endothelial glycocalyx are critical regulators of microvascular permeability to both water and albumin. PMID:28524373

Peroxynitrite Disrupts Endothelial Caveolae Leading to eNOS Uncoupling and Diminished Flow-Mediated Dilation in Coronary Arterioles of Diabetic Patients

PubMed Central

Cassuto, James; Dou, Huijuan; Czikora, Istvan; Szabo, Andras; Patel, Vijay S.; Kamath, Vinayak; Belin de Chantemele, Eric; Feher, Attila; Romero, Maritza J.; Bagi, Zsolt

2014-01-01

Peroxynitrite (ONOO−) contributes to coronary microvascular dysfunction in diabetes mellitus (DM). We hypothesized that in DM, ONOO− interferes with the function of coronary endothelial caveolae, which plays an important role in nitric oxide (NO)-dependent vasomotor regulation. Flow-mediated dilation (FMD) of coronary arterioles was investigated in DM (n = 41) and non-DM (n = 37) patients undergoing heart surgery. NO-mediated coronary FMD was significantly reduced in DM patients, which was restored by ONOO− scavenger, iron-(III)-tetrakis(N-methyl-4'pyridyl)porphyrin-pentachloride, or uric acid, whereas exogenous ONOO− reduced FMD in non-DM subjects. Immunoelectron microscopy demonstrated an increased 3-nitrotyrosine formation (ONOO−-specific protein nitration) in endothelial plasma membrane in DM, which colocalized with caveolin-1 (Cav-1), the key structural protein of caveolae. The membrane-localized Cav-1 was significantly reduced in DM and also in high glucose–exposed coronary endothelial cells. We also found that DM patients exhibited a decreased number of endothelial caveolae, whereas exogenous ONOO− reduced caveolae number. Correspondingly, pharmacological (methyl-β-cyclodextrin) or genetic disruption of caveolae (Cav-1 knockout mice) abolished coronary FMD, which was rescued by sepiapterin, the stable precursor of NO synthase (NOS) cofactor, tetrahydrobiopterin. Sepiapterin also restored coronary FMD in DM patients. Thus, we propose that ONOO− selectively targets and disrupts endothelial caveolae, which contributes to NOS uncoupling, and, hence, reduced NO-mediated coronary vasodilation in DM patients. PMID:24353182

The anti-apoptotic effect of fluid mechanics preconditioning by cells membrane and mitochondria in rats brain microvascular endothelial cells.

PubMed

Tian, Shan; Zhu, Fengping; Hu, Ruiping; Tian, Song; Chen, Xingxing; Lou, Dan; Cao, Bing; Chen, Qiulei; Li, Bai; Li, Fang; Bai, Yulong; Wu, Yi; Zhu, Yulian

2018-01-01

Exercise preconditioning is a simple and effective way to prevent ischemia. This paper further provided the mechanism in hemodynamic aspects at the cellular level. To study the anti-apoptotic effects of fluid mechanics preconditioning, Cultured rats brain microvascular endothelial cells were given fluid intervention in a parallel plate flow chamber before oxygen glucose deprivation. It showed that fluid mechanics preconditioning could inhibit the apoptosis of endothelial cells, and this process might be mediated by the shear stress activation of Tie-2 on cells membrane surface and Bcl-2 on the mitochondria surface. Copyright © 2017 Elsevier B.V. All rights reserved.

Escherichia coli K1 invasion of human brain microvascular endothelial cells.

PubMed

Loh, Lip Nam; Ward, Theresa H

2012-01-01

The pathogenic Escherichia coli strain E. coli K1 is a primary causative agent of neonatal meningitis. Understanding how these bacteria cross the blood-brain barrier is vital to develop therapeutics. Here, we describe the use of live-cell imaging techniques to study E. coli K1 interactions with cellular markers following infection of human brain microvascular endothelial cells, a model system of the blood-brain barrier. We also discuss optimization of endothelial cell transfection conditions using nonviral transfection technique, bacterial labeling techniques, and in vitro assays to screen for fluorescent bacteria that retain their ability to invade host cells. Copyright © 2012 Elsevier Inc. All rights reserved.

Monitoring skin microvascular dysfunction of type 1 diabetic mice using in vivo skin optical clearing

NASA Astrophysics Data System (ADS)

Feng, Wei; Shi, Rui; Zhu, Dan

2018-02-01

To monitor skin microvascular dysfunction of alloxan-induced type 1 diabetic mice model. In this work, we used laser speckle contrast imaging and hyperspectral imaging through in vivo skin optical clearing method to simultaneously monitor the noradrenaline-induced response of microvascular blood flow and blood oxygen with the development of diabetes. The main results showed that venous and arterious blood flow steadily decreased without recovery after injecting noradrenaline (NE), furthermore the influence of NE-induced arterious blood oxygen response greatly decreased, especially for 2-weeks and 4-weeks diabetic mice. This study demonstrated that skin microvascular function was a potential research biomarker for early warning in the occurrence and development of diabetes. And it provides a feasible solution to realize visualization of cutaneous microvessels for monitoring microvascular reactivity.

Treatment of Angina Pectoris Associated with Coronary Microvascular Dysfunction.

PubMed

Ong, Peter; Athanasiadis, Anastasios; Sechtem, Udo

2016-08-01

Treatment of angina pectoris associated with coronary microvascular dysfunction is challenging as the underlying mechanisms are often diverse and overlapping. Patients with type 1 coronary microvascular dysfunction (i.e. absence of epicardial coronary artery disease and myocardial disease) should receive strict control of their cardiovascular risk factors and thus receive statins and ACE-inhibitors in most cases. Antianginal medication consists of ß-blockers and/or calcium channel blockers. Second line drugs are ranolazine and nicorandil with limited evidence. Despite individually titrated combinations of these drugs up to 30 % of patients have refractory angina. Rho-kinase inhibitors and endothelin-receptor antagonists represent potential drugs that may prove useful in these patients in the future.

Endothelial network formed with human dermal microvascular endothelial cells in autologous multicellular skin substitutes.

PubMed

Ponec, Maria; El Ghalbzouri, Abdoelwaheb; Dijkman, Remco; Kempenaar, Johanna; van der Pluijm, Gabri; Koolwijk, Pieter

2004-01-01

A human skin equivalent from a single skin biopsy harboring keratinocytes and melanocytes in the epidermal compartment, and fibroblasts and microvascular dermal endothelial cells in the dermal compartment was developed. The results of the study revealed that the nature of the extracellular matrix of the dermal compartments plays an important role in establishment of endothelial network in vitro. With rat-tail type I collagen matrices only lateral but not vertical expansion of endothelial networks was observed. In contrast, the presence of extracellular matrix of entirely human origin facilitated proper spatial organization of the endothelial network. Namely, when human dermal fibroblasts and microvascular endothelial cells were seeded on the bottom of an inert filter and subsequently epidermal cells were seeded on top of it, fibroblasts produced extracellular matrix throughout which numerous branched tubes were spreading three-dimensionally. Fibroblasts also facilitated the formation of basement membrane at the epidermal/matrix interface. Under all culture conditions, fully differentiated epidermis was formed with numerous melanocytes present in the basal epidermal cell layer. The results of the competitive RT-PCR revealed that both keratinocytes and fibroblasts expressed VEGF-A, -B, -C, aFGF and bFGF mRNA, whereas fibroblasts also expressed VEGF-D mRNA. At protein level, keratinocytes produced 10 times higher amounts of VEGF-A than fibroblasts did. The generation of multicellular skin equivalent from a single human skin biopsy will stimulate further developments for its application in the treatment of full-thickness skin defects. The potential development of biodegradable, biocompatible material suitable for these purposes is a great challenge for future research.

Transcription factor fos-related antigen-2 induces progressive peripheral vasculopathy in mice closely resembling human systemic sclerosis.

PubMed

Maurer, Britta; Busch, Nicole; Jüngel, Astrid; Pileckyte, Margarita; Gay, Renate E; Michel, Beat A; Schett, Georg; Gay, Steffen; Distler, Jörg; Distler, Oliver

2009-12-08

Microvascular damage is one of the first pathological changes in systemic sclerosis. In this study, we investigated the role of Fos-related antigen-2 (Fra-2), a transcription factor of the activator protein-1 family, in the peripheral vasculopathy of systemic sclerosis and examined the underlying mechanisms. Expression of Fra-2 protein was significantly increased in skin biopsies of systemic sclerosis patients compared with healthy controls, especially in endothelial and vascular smooth muscle cells. Fra-2 transgenic mice developed a severe loss of small blood vessels in the skin that was paralleled by progressive skin fibrosis at 12 weeks of age. The reduction in capillary density was preceded by a significant increase in apoptosis in endothelial cells at week 9 as detected by immunohistochemistry. Similarly, suppression of Fra-2 by small interfering RNA prevented human microvascular endothelial cells from staurosporine-induced apoptosis and improved both the number of tubes and the cumulative tube lengths in the tube formation assay. In addition, cell migration in the scratch assay and vascular endothelial growth factor-dependent chemotaxis in a modified Boyden chamber assay were increased after transfection of human microvascular endothelial cells with Fra-2 small interfering RNA, whereas proliferation was not affected. Fra-2 is present in human systemic sclerosis and may contribute to the development of microvasculopathy by inducing endothelial cell apoptosis and by reducing endothelial cell migration and chemotaxis. Fra-2 transgenic mice are a promising preclinical model to study the mechanisms and therapeutic approaches of the peripheral vasculopathy in systemic sclerosis.

Quantification of Malignant Breast Cancer Cell MDA-MB-231 Transmigration across Brain and Lung Microvascular Endothelium

PubMed Central

Fan, Jie; Fu, Bingmei M.

2015-01-01

Tumor cell extravasation through the endothelial barrier forming the microvessel wall is a crucial step during tumor metastasis. However, where, how and how fast tumor cells transmigrate through endothelial barriers remain unclear. Using an in vitro transwell model, we performed a transmigration assay of malignant breast tumor cells (MDA-MB-231) through brain and lung microvascular endothelial monolayers under control and pathological conditions. The locations and rates of tumor cell transmigration as well as the changes in the structural components (integrity) of endothelial monolayers were quantified by confocal microscopy. Endothelial monolayer permeability to albumin Palbumin was also quantified under the same conditions. We found that about 98% of transmigration occurred at the joints of endothelial cells instead of cell bodies; tumor cell adhesion and transmigration degraded endothelial surface glycocalyx and disrupted endothelial junction proteins, consequently increased Palbumin; more tumor cells adhered to and transmigrated through the endothelial monolayer with higher Palbumin; Palbumin and tumor transmigration were increased by vascular endothelial growth factor (VEGF), a representative of cytokines, and lipopolysaccharides (LPS), a typical systemic inflammatory factor, but reduced by adenosine 3′, 5′-cyclic monophosphate (cAMP). These results suggest that reinforcing endothelial structural integrity is an effective approach for inhibiting tumor extravasation. PMID:26603751

HIF-2α Expression Regulates Sprout Formation into 3D Fibrin Matrices in Prolonged Hypoxia in Human Microvascular Endothelial Cells.

PubMed

Nauta, Tessa D; Duyndam, Monique C A; Weijers, Ester M; van Hinsbergh, Victor M W; Koolwijk, Pieter

2016-01-01

During short-term hypoxia, Hypoxia Inducible Factors (particular their subunits HIF-1α and HIF-2α) regulate the expression of many genes including the potent angiogenesis stimulator VEGF. However, in some pathological conditions chronic hypoxia occurs and is accompanied by reduced angiogenesis. We investigated the effect of prolonged hypoxia on the proliferation and sprouting ability of human microvascular endothelial cells and the involvement of the HIFs and Dll4/Notch signaling. Human microvascular endothelial cells (hMVECs), cultured at 20% oxygen for 14 days and seeded on top of 3D fibrin matrices, formed sprouts when stimulated with VEGF-A/TNFα. In contrast, hMVECs precultured at 1% oxygen for 14 days were viable and proliferative, but did not form sprouts into fibrin upon VEGF-A/TNFα stimulation at 1% oxygen. Silencing of HIF-2α with si-RNA partially restored the inhibition of endothelial sprouting, whereas HIF-1α or HIF-3α by si-RNA had no effect. No involvement of Dll4/Notch pathway in the inhibitory effect on endothelial sprouting by prolonged hypoxia was found. In addition, hypoxia decreased the production of urokinase-type plasminogen activator (uPA), needed for migration and invasion, without a significant effect on its inhibitor PAI-1. This was independent of HIF-2α, as si-HIF-2α did not counteract uPA reduction. Prolonged culturing of hMVECs at 1% oxygen inhibited endothelial sprouting into fibrin. Two independent mechanisms contribute. Silencing of HIF-2α with si-RNA partially restored the inhibition of endothelial sprouting pointing to a HIF-2α-dependent mechanism. In addition, reduction of uPA contributed to reduced endothelial tube formation in a fibrin matrix during prolonged hypoxia.

Nitric Oxide-Dependent Endothelial Dysfunction and Reduced Arginine Bioavailability in Plasmodium vivax Malaria but No Greater Increase in Intravascular Hemolysis in Severe Disease.

PubMed

Barber, Bridget E; William, Timothy; Grigg, Matthew J; Piera, Kim A; Chen, Youwei; Wang, Hao; Weinberg, J Brice; Yeo, Tsin W; Anstey, Nicholas M

2016-11-15

 Pathogenesis of severe Plasmodium vivax malaria is poorly understood. Endothelial dysfunction and reduced nitric oxide (NO) bioavailability characterize severe falciparum malaria, but have not been assessed in severe vivax malaria.  In patients with severe vivax malaria (n = 9), patients with nonsevere vivax malaria (n = 58), and healthy controls (n = 79), we measured NO-dependent endothelial function by using reactive hyperemia-peripheral arterial tonometry (RH-PAT) and assessed associations with arginine, asymmetric dimethylarginine (ADMA), and hemolysis.  The L-arginine level and the L-arginine to ADMA ratio (a measure of L-arginine bioavailability) were reduced in patients with severe vivax malaria and those with nonsevere vivax malaria, compared with healthy controls (median L-arginine level, 65, 66, and 98 µmol/mL, respectively [P = .0001]; median L-arginine to ADMA ratio, 115, 125, and 187, respectively [P = .0001]). Endothelial function was impaired in proportion to disease severity (median RH-PAT index, 1.49, 1.73, and 1.97 in patients with severe vivax malaria, those with nonsevere vivax malaria, and healthy controls, respectively; P = .018) and was associated with the L-arginine to ADMA ratio. While the posttreatment fall in hemoglobin level was greater in severe vivax malaria as compared to nonsevere vivax malaria (2.5 vs 1 g/dL; P = .0001), markers of intravascular hemolysis were not higher in severe disease.  Endothelial function is impaired in nonsevere and severe vivax malaria, is associated with reduced L-arginine bioavailability, and may contribute to microvascular pathogenesis. Severe disease appears to be more associated with extravascular hemolysis than with intravascular hemolysis. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Engineering a Microvascular Capillary Bed in a Tissue-Like Collagen Construct

PubMed Central

Unger, Ronald E.; Brochhausen, Christoph; Brown, Robert A.; Kirkpatrick, James C.

2014-01-01

Previous studies have shown that plastic compression (PC) of collagen gels allows a rapid and controlled fabrication of matrix- and cell-rich constructs in vitro that closely mimic the structure and characteristics of tissues in vivo. Microvascular endothelial cells, the major cell type making up the blood vessels in the body, were added to the PC collagen to determine whether cells attach, survive, grow, and express endothelial cell characteristics when seeded alone or in coculture with other cells. Endothelial cells seeded on the PC collagen containing human foreskin fibroblasts (HFF) or human osteoblasts (HOS) formed vessel-like structures over 3 weeks in culture without the addition of exogenous growth factors in the medium. In contrast, on the PC scaffolds without HFF or HOS, human dermal microvascular endothelial cells (HDMEC) exhibited a typical cobblestone morphology for 21 days under the same conditions. We propose that the coculture of primary endothelial cells with PC collagen constructs, containing a stromal cell population, is a valuable technique for in vitro modeling of proangiogenic responses toward such biomimetic constructs in vivo. A major observation in the cocultures was the absence of gel contraction, even after 3 weeks of fibroblast culture. This collagen form could, for example, be of great value in tissue engineering of the skin, as contractures are both aesthetically and functionally disabling. PMID:24684395

Mitral annular calcification associated with impaired coronary microvascular function.

PubMed

Bozbas, Huseyin; Pirat, Bahar; Yildirir, Aylin; Simşek, Vahide; Sade, Elif; Altin, Cihan; Muderrisoglu, Haldun

2008-05-01

Mitral annular calcification (MAC) has been shown to be associated with atherosclerosis, and is a predictor of cardiovascular events. Coronary flow reserve (CFR) determined by transthoracic echocardiography has been introduced as a reliable indicator for coronary microvascular function. In this study we sought to investigate CFR in patients with and without MAC. Seventy patients (mean age, 68.2+/-6.6 years) who were free of coronary artery disease or diabetes mellitus were involved; 35 patients with MAC constituted the experimental group while 35 patients without MAC served as controls. Using transthoracic Doppler echocardiography coronary peak flow velocities were measured at baseline and after dipyridamole infusion. CFR was calculated as the ratio of hyperemic to baseline diastolic peak flow velocities. The clinical and demographic characteristics including age, sex, and traditional coronary risk factors did not differ between the groups (P>.05). The mean value of CFR was significantly lower in participants with mitral annular calcification than it was in controls (2.25+/-0.41 vs. 2.64+/-0.57; P<.0001). Multivariable regression analysis identified MAC (beta=-0.40, P=.004), smoking (beta=-0.36, P=.007), and C-reactive protein levels (beta=-0.28, P=.04) as the independent variables significantly associated with CFR. Our results demonstrate that CFR is impaired in patients with mitral annular calcification suggesting that coronary microvascular-endothelial dysfunction, an early finding of atherosclerosis, is present in these patients.

Uterine microvascular sensitivity to nanomaterial inhalation: An in vivo assessment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stapleton, P.A.; McBride, C.R.; Yi, J.

With the tremendous number and diverse applications of engineered nanomaterials incorporated in daily human activity, exposure can no longer be solely confined to occupational exposures of healthy male models. Cardiovascular and endothelial cell dysfunction have been established using in vitro and in situ preparations, but the translation to intact in vivo models is limited. Intravital microscopy has been used extensively to understand microvascular physiology while maintaining in vivo neurogenic, humoral, and myogenic control. However, a tissue specific model to assess the influences of nanomaterial exposure on female reproductive health has not been fully elucidated. Female Sprague Dawley (SD) rats weremore » exposed to nano-TiO{sub 2} aerosols (171 ± 6 nm, 10.1 ± 0.39 mg/m{sup 3}, 5 h) 24-hours prior to experimentation, leading to a calculated deposition of 42.0 ± 1.65 μg. After verifying estrus status, vital signs were monitored and the right horn of the uterus was exteriorized, gently secured over an optical pedestal, and enclosed in a warmed tissue bath using intravital microscopy techniques. After equilibration, significantly higher leukocyte-endothelium interactions were recorded in the exposed group. Arteriolar responsiveness was assessed using ionophoretically applied agents: muscarinic agonist acetylcholine (0.025 M; ACh; 20, 40, 100, and 200 nA), and nitric oxide donor sodium nitroprusside (0.05 M; SNP; 20, 40, and 100 nA), or adrenergic agonist phenylephrine (0.05 M; PE; 20, 40, and 100 nA) using glass micropipettes. Passive diameter was established by tissue superfusion with 10{sup −4} M adenosine. Similar to male counterparts, female SD rats present systemic microvascular dysfunction; however the ramifications associated with female health and reproduction have yet to be elucidated. - Highlights: • Female reproductive health associated with nanomaterial exposure is understudied. • We examined uterine microvascular alterations 24-hours after nano-TiO{sub 2} inhalation. • There is an increase in uterine venular leukocyte activity post nano-TiO{sub 2} exposure. • Nano-TiO{sub 2} exposure perturbs uterine arteriolar endothelium-dependent dilation. • Intravital microscopy is a powerful tool for future toxicological assessments.« less

Effect of diabetes on the cutaneous microcirculation of the feet in patients with intermittent claudication.

PubMed

Klonizakis, M; Manning, G; Lingam, K; Donnelly, R; Yeung, J M C

2015-01-01

To evaluate endothelial-dependent and - independent cutaneous vasodilator responses in the feet of patients with peripheral arterial disease (PAD) with or without Type 2 diabetes. Cutaneous microvascular responses in the dorsum of both lower limbs were measured in the supine position using Laser Doppler Fluximetry combined with iontophoretic administration of endothelial-dependent (acetylcholine, Ach) and -independent (sodium nitroprusside, SNP) vasodilators in diabetic (n = 19) and non diabetic (n = 17) patients with PAD (presenting as unilateral calf intermittent claudication (IC). In patients with diabetes and IC, endothelial-dependent vasodilation was significantly impaired in the symptomatic limb [74 (57,105) vs 68 (24,81) PU, Z =-2.79, p = 0.005] compared to the asymptomatic limb. Patients without diabetes showed no impairment of vasodilation. Resting ankle-brachial pressure index did not identify the presence of abnormalities in microvascular function. The combination of diabetes and PAD is associated with a reduction in endothelial-dependent cutaneous vasodilation in the feet without an associated reduction in endothelial independent vasodilation.

Cocaine Inhibits Store-Operated Ca2+ Entry in Brain Microvascular Endothelial Cells: Critical Role for Sigma-1 Receptors

PubMed Central

Brailoiu, G. Cristina; Deliu, Elena; Console-Bram, Linda M; Soboloff, Jonathan; Abood, Mary E; Unterwald, Ellen M; Brailoiu, Eugen

2015-01-01

Sigma-1 receptor (Sig-1R) is an intracellular chaperone protein with many ligands, located at the endoplasmic reticulum. Binding of cocaine to Sig-1R has previously been found to modulate endothelial functions. In the present study, we show that cocaine dramatically inhibits store-operated Ca2+ entry (SOCE), a Ca2+ influx mechanism promoted by depletion of intracellular Ca2+ stores, in rat brain microvascular endothelial cells. Using either Sig-1R shRNA or pharmacological inhibition with the unrelated Sig-1R antagonists BD-1063 and NE-100, we show that cocaine-induced SOCE inhibition is dependent on Sig-1R. In addition to revealing new insight into fundamental mechanisms of cocaine-induced changes in endothelial function, these studies provide an unprecedented role for Sig-1R as a SOCE inhibitor. PMID:26467159

Involvement of PI3K and ROCK signaling pathways in migration of bone marrow-derived mesenchymal stem cells through human brain microvascular endothelial cell monolayers.

PubMed

Lin, Mei-Na; Shang, De-Shu; Sun, Wei; Li, Bo; Xu, Xin; Fang, Wen-Gang; Zhao, Wei-Dong; Cao, Liu; Chen, Yu-Hua

2013-06-04

Bone marrow-derived mesenchymal stem cells (MSC) represent an important and easily available source of stem cells for potential therapeutic use in neurological diseases. The entry of circulating cells into the central nervous system by intravenous administration requires, firstly, the passage of the cells across the blood-brain barrier (BBB). However, little is known of the details of MSC transmigration across the BBB. In the present study, we employed an in vitro BBB model constructed using a human brain microvascular endothelial cell monolayer to study the mechanism underlying MSC transendothelial migration. Transmigration assays, transendothelial electrical resistance (TEER) and horseradish peroxidase (HRP) flux assays showed that MSC could transmigrate through human brain microvascular endothelial cell monolayers by a paracellular pathway. Cell fractionation and immunofluorescence assays confirmed the disruption of tight junctions. Inhibition assays showed that a Rho-kinase (ROCK) inhibitor (Y27632) effectively promoted MSC transendothelial migration; conversely, a PI3K inhibitor (LY294002) blocked MSC transendothelial migration. Interestingly, adenovirus-mediated interference with ROCK in MSC significantly increased MSC transendothelial migration, and overexpression of a PI3K dominant negative mutant in MSC cells could block transendothelial migration. Our findings provide clear evidence that the PI3K and ROCK pathways are involved in MSC migration through human brain microvascular endothelial cell monolayers. The information yielded by this study may be helpful in constructing gene-modified mesenchymal stem cells that are able to penetrate the BBB effectively for cell therapy. Copyright © 2013 Elsevier B.V. All rights reserved.

Perfluorooctane sulfonate (PFOS) induces reactive oxygen species (ROS) production in human microvascular endothelial cells: role in endothelial permeability.

PubMed

Qian, Yong; Ducatman, Alan; Ward, Rebecca; Leonard, Steve; Bukowski, Valerie; Lan Guo, Nancy; Shi, Xianglin; Vallyathan, Val; Castranova, Vincent

2010-01-01

Perfluorooctane sulfonate (PFOS) is a member of the perfluoroalkyl acids (PFAA) containing an eight-carbon backbone. PFOS is a man-made chemical with carbon-fluorine bonds that are among the strongest in organic chemistry, and PFOS is widely used in industry. Human occupational and environmental exposure to PFOS occurs globally. PFOS is non-biodegradable and is persistent in the human body and environment. In this study, data demonstrated that exposure of human microvascular endothelial cells (HMVEC) to PFOS induced the production of reactive oxygen species (ROS) at both high and low concentrations. Morphologically, it was found that exposure to PFOS induced actin filament remodeling and endothelial permeability changes in HMVEC. Furthermore, data demonstrated that the production of ROS plays a regulatory role in PFOS-induced actin filament remodeling and the increase in endothelial permeability. Our results indicate that the generation of ROS may play a role in PFOS-induced aberrations of the endothelial permeability barrier. The results generated from this study may provide a new insight into the potential adverse effects of PFOS exposure on humans at the cellular level.

Effects of the PPAR{gamma} agonist troglitazone on endothelial cells in vivo and in vitro: Differences between human and mouse

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kakiuchi-Kiyota, Satoko; Vetro, Joseph A.; Suzuki, Shugo

2009-05-15

Peroxisome proliferator-activated receptor gamma (PPAR{gamma}) agonists and PPAR{gamma}/{alpha} dual agonists have been or are being developed for clinical use in the treatment of type 2 diabetes mellitus and hyperlipidemias. A common tumor finding in rodent carcinogenicity studies for these agonists is hemangioma/hemangiosarcoma in mice but not in rats. We hypothesized that increased endothelial cell proliferation may be involved in the mechanism of PPAR agonist-induced vascular tumors in mice, and we investigated the effects on endothelial cells utilizing troglitazone, the first clinically used PPAR{gamma} agonist, in vivo and in vitro. Troglitazone (400 and 800 mg/kg/day) induced hemangiosarcomas in mice in amore » 2-year bioassay. We showed that troglitazone increased endothelial cell proliferation in brown and white adipose tissue and liver in mice at sarcomagenic doses after 4 weeks of treatment. Troglitazone was cytotoxic both to human dermal microvascular endothelial cells (HMEC1) and mouse mammary fat pad microvascular endothelial cells (MFP MVEC) at high concentrations. However, MFP MVEC were more resistant to the cytotoxic effects of troglitazone based on the much lower LC{sub 50} in HMEC1 (17.4 {mu}M) compared to MFP MVEC (92.2 {mu}M). Troglitazone increased the proliferation and survival of MFP MVEC but not HMEC1 in growth factor reduced conditions. Our data demonstrate that troglitazone may induce hemangiosarcomas in mice, at least in part, through enhancement of survival and proliferation of microvascular endothelial cells. Such an effect does not occur with human cells, suggesting that human may react differently to exposure to PPAR agonists compared with mice.« less

Astrocytes increase barrier properties and ZO-1 expression in retinal vascular endothelial cells.

PubMed

Gardner, T W; Lieth, E; Khin, S A; Barber, A J; Bonsall, D J; Lesher, T; Rice, K; Brennan, W A

1997-10-01

Diabetic retinopathy and other diseases associated with retinal edema are characterized by increased microvascular leakage. Astrocytes have been proposed to maintain endothelial function in the brain, suggesting that glial impairment may underlie the development of retinal edema. The purpose of this study was to test the effects of astrocytes on barrier properties in retinal microvascular endothelial cells. Bovine retinal microvascular endothelial cells were exposed to conditioned media from rat brain astrocytes. Transendothelial electrical resistance (TER) was determined on 24-mm Transwell (Cambridge, MA) polycarbonate filters with the End-Ohm device (World Precision Instruments, Sarasota, FL). ZO-1 protein content was quantified by microtiter enzyme-linked immunosorbent assay. Astrocyte-conditioned medium (ACM) significantly increased TER (P < 0.0001) and ZO-1 content (P < 0.01). Both serum-containing and serum-free N1B defined ACM increased ZO-1 expression, but heating abolished the effect. Serum-free ACM decreased cell proliferation by 16%. Astrocytes release soluble, heat-labile factors that increase barrier properties and tight junction protein content. These results suggest that astrocytes enhance blood-retinal barrier properties, at least in part by increasing tight junction protein expression. Our findings suggest that glial malfunction plays an important role in the pathogenesis of vasogenic retinal edema.

Association of Microvascular Function and Endothelial Biomarkers With Clinical Outcome in Dengue: An Observational Study

PubMed Central

Yacoub, Sophie; Lam, Phung Khanh; Vu, Le Hoang Mai; Le, Thi Lien; Ha, Ngo Thanh; Toan, Tran Thi; Van, Nguyen Thu; Quyen, Nguyen Than Ha; Le Duyen, Huynh Thi; Van Kinh, Nguyen; Fox, Annette; Mongkolspaya, Juthathip; Wolbers, Marcel; Simmons, Cameron Paul; Screaton, Gavin Robert; Wertheim, Heiman; Wills, Bridget

2016-01-01

Background. The hallmark of severe dengue is increased microvascular permeability, but alterations in the microcirculation and their evolution over the course of dengue are unknown. Methods. We conducted a prospective observational study to evaluate the sublingual microcirculation using side-stream dark-field imaging in patients presenting early (<72 hours after fever onset) and patients hospitalized with warning signs or severe dengue in Vietnam. Clinical findings, microvascular function, global hemodynamics assessed with echocardiography, and serological markers of endothelial activation were determined at 4 time points. Results. A total of 165 patients were enrolled. No difference was found between the microcirculatory parameters comparing dengue with other febrile illnesses. The proportion of perfused vessels (PPV) and the mean flow index (MFI) were lower in patients with dengue with plasma than those without leakage (PPV, 88.1% vs 90.6% [P = .01]; MFI, 2.1 vs 2.4 [P = .007]), most markedly during the critical phase. PPV and MFI were correlated with the endothelial activation markers vascular cell adhesion molecule 1 (P < .001 for both) and angiopoietin 2 (P < .001 for both), negatively correlated. Conclusions. Modest microcirculatory alterations occur in dengue, are associated with plasma leakage, and are correlate with molecules of endothelial activation, angiopoietin 2 and vascular cell adhesion molecule 1. PMID:27230099

Sphingosine kinase inhibition alleviates endothelial permeability induced by thrombin and activated neutrophils.

PubMed

Itagaki, Kiyoshi; Zhang, Qin; Hauser, Carl J

2010-04-01

Inflammation and microvascular thrombosis are interrelated causes of acute lung injury in the systemic inflammatory response syndrome. Neutrophils (polymorphonuclear neutrophil [PMN]) and endothelial cells (EC) activated by systemic inflammatory response syndrome interact to increase pulmonary vascular permeability, but the interactions between PMN and EC are difficult to study. Recently, we reported that sphingosine 1-phosphate is a second messenger eliciting store-operated calcium entry (SOCE) in response to inflammatory agonists in both PMN and EC. Store-operated calcium entry is therefore a target mechanism for the therapeutic modulation of inflammatory PMN-EC interactions. Here, we isolated, modeled, and studied the effects of pharmacologic SOCE inhibition using real-time systems to monitor EC permeability after exposure to activated PMN. We created systems to continuously assess permeability of human pulmonary artery endothelial cells and human microvascular endothelial cells from lung. Endothelial cells show increased permeability after challenge by activated PMN. Such permeability increases can be attenuated by exposure of the cocultures to sphingosine kinase (SK) inhibitors (SKI-2, N,N-dimethylsphingosine [DMS]) or Ca2+ entry inhibitors (Gd3+, MRS-1845). Human microvascular endothelial cells from lung pretreated with SKI-2 or DMS showed decreased permeability when later exposed to activated PMN. Likewise, when PMNs were activated with thapsigargin (TG) in the presence of SKI-2, DMS, Gd, or MRS-1845, their ability to cause EC permeability subsequently was reduced. SKI-2 also inhibited the activation of human pulmonary artery ECs by thrombin. These studies will provide a firm mechanistic foundation for understanding how systemic SOCE inhibition may be used to prevent acute lung injury in vivo.

Altered decorin leads to disrupted endothelial cell function: a possible mechanism in the pathogenesis of fetal growth restriction?

PubMed

Chui, A; Murthi, P; Gunatillake, T; Brennecke, S P; Ignjatovic, V; Monagle, P T; Whitelock, J M; Said, J M

2014-08-01

Fetal growth restriction (FGR) is a key cause of adverse pregnancy outcome where maternal and fetal factors are identified as contributing to this condition. Idiopathic FGR is associated with altered vascular endothelial cell functions. Decorin (DCN) has important roles in the regulation of endothelial cell functions in vascular environments. DCN expression is reduced in FGR. The objectives were to determine the functional consequences of reduced DCN in a human microvascular endothelial cell line model (HMVEC), and to determine downstream targets of DCN and their expression in primary placental microvascular endothelial cells (PLECs) from control and FGR-affected placentae. Short-interference RNA was used to reduce DCN expression in HMVECs and the effect on proliferation, angiogenesis and thrombin generation was determined. A Growth Factor PCR Array was used to identify downstream targets of DCN. The expression of target genes in control and FGR PLECs was performed. DCN reduction decreased proliferation and angiogenesis but increased thrombin generation with no effect on apoptosis. The array identified three targets of DCN: FGF17, IL18 and MSTN. Validation of target genes confirmed decreased expression of VEGFA, MMP9, EGFR1, IGFR1 and PLGF in HMVECs and PLECs from control and FGR pregnancies. Reduction of DCN in vascular endothelial cells leads to disrupted cell functions. The targets of DCN include genes that play important roles in angiogenesis and cellular growth. Therefore, differential expression of these may contribute to the pathogenesis of FGR and disease states in other microvascular circulations. Copyright © 2014 Elsevier Ltd. All rights reserved.

Clinical Features, Diagnosis, and Management of Patients With Anderson-Fabry Cardiomyopathy.

PubMed

Yogasundaram, Haran; Kim, Daniel; Oudit, Omar; Thompson, Richard B; Weidemann, Frank; Oudit, Gavin Y

2017-07-01

Anderson-Fabry disease (AFD) is an X-linked recessive, multisystem disease of lysosomal storage. A mutation in the gene encoding the hydrolase enzyme α-galactosidase A results in its deficiency, or complete absence of activity. Subsequent progressive intracellular accumulation of glycosphingolipids, predominantly globotriaosylceramide, in various tissues, results in progressive organ dysfunction and failure, most commonly affecting the kidneys, nervous system, skin, eyes, vascular endothelium, and the heart. Cardiac involvement in AFD represents a leading cause of morbidity and mortality. Globotriaosylceramide accumulation affects cardiomyocytes, smooth muscle cells, vascular endothelial cells, and fibroblasts leading to various pathologies including valvular regurgitation, conduction disease and arrhythmias, coronary microvascular dysfunction, and right and left ventricular hypertrophy (LVH) leading to early diastolic dysfunction and late-stage systolic impairment. Diagnosis is on the basis of decreased plasma α-galactosidase activity in men and positive genetic testing in women. Contemporary large-scale screening studies have revealed a prevalence of 1%-5% in patients with unexplained LVH in multiple cohorts. Cardiac magnetic resonance imaging, with its unique tissue characterization capabilities, is the most important imaging modality to assess for cardiomyopathy in patients with AFD. Enzyme replacement therapy is indicated in AFD patients with significant organ involvement, and has been shown to clear sphingolipids from endothelial cells in other organs, as well as to reduce left ventricular mass as early as 6 months after starting treatment. There is increasing evidence that enzyme replacement therapy might be more effective if given at earlier stages of disease, before the development of LVH and myocardial fibrosis. Copyright © 2017 Canadian Cardiovascular Society. Published by Elsevier Inc. All rights reserved.

Endothelial dysfunction in metabolic and vascular disorders.

PubMed

Polovina, Marija M; Potpara, Tatjana S

2014-03-01

Vascular endothelium has important regulatory functions in the cardiovascular system and a pivotal role in the maintenance of vascular health and metabolic homeostasis. It has long been recognized that endothelial dysfunction participates in the pathogenesis of atherosclerosis from early, preclinical lesions to advanced, thrombotic complications. In addition, endothelial dysfunction has been recently implicated in the development of insulin resistance and type 2 diabetes mellitus (T2DM). Considering that states of insulin resistance (eg, metabolic syndrome, impaired fasting glucose, impaired glucose tolerance, and T2DM) represent the most prevalent metabolic disorders and risk factors for atherosclerosis, it is of considerable scientific and clinical interest that both metabolic and vascular disorders have endothelial dysfunction as a common background. Importantly, endothelial dysfunction has been associated with adverse outcomes in patients with established cardiovascular disease, and a growing body of evidence indicates that endothelial dysfunction also imparts adverse prognosis in states of insulin resistance. In this review, we discuss the association of insulin resistance and T2DM with endothelial dysfunction and vascular disease, with a focus on the underlying mechanisms and prognostic implications of the endothelial dysfunction in metabolic and vascular disorders. We also address current therapeutic strategies for the improvement of endothelial dysfunction.

Endothelial actions of atrial and B-type natriuretic peptides.

PubMed

Kuhn, Michaela

2012-05-01

The cardiac hormone atrial natriuretic peptide (ANP) is critically involved in the maintenance of arterial blood pressure and intravascular volume homeostasis. Its cGMP-producing GC-A receptor is densely expressed in the microvascular endothelium of the lung and systemic circulation, but the functional relevance is controversial. Some studies reported that ANP stimulates endothelial cell permeability, whereas others described that the peptide attenuates endothelial barrier dysfunction provoked by inflammatory agents such as thrombin or histamine. Many studies in vitro addressed the effects of ANP on endothelial proliferation and migration. Again, both pro- and anti-angiogenic properties were described. To unravel the role of the endothelial actions of ANP in vivo, we inactivated the murine GC-A gene selectively in endothelial cells by homologous loxP/Cre-mediated recombination. Our studies in these mice indicate that ANP, via endothelial GC-A, increases endothelial albumin permeability in the microcirculation of the skin and skeletal muscle. This effect is critically involved in the endocrine hypovolaemic, hypotensive actions of the cardiac hormone. On the other hand the homologous GC-A-activating B-type NP (BNP), which is produced by cardiac myocytes and many other cell types in response to stressors such as hypoxia, possibly exerts more paracrine than endocrine actions. For instance, within the ischaemic skeletal muscle BNP released from activated satellite cells can improve the regeneration of neighbouring endothelia. This review will focus on recent advancements in our understanding of endothelial NP/GC-A signalling in the pulmonary versus systemic circulation. It will discuss possible mechanisms accounting for the discrepant observations made for the endothelial actions of this hormone-receptor system and distinguish between (patho)physiological and pharmacological actions. Lastly it will emphasize the potential therapeutical implications derived from the actions of NPs on endothelial permeability and regeneration. © 2012 The Author. British Journal of Pharmacology © 2012 The British Pharmacological Society.

Subendocardial viability ratio in patients with rheumatoid arthritis: comparison with healthy controls and identification of prognostic factors.

PubMed

Anyfanti, Panagiota; Triantafyllou, Areti; Gkaliagkousi, Eugenia; Triantafyllou, Georgios; Koletsos, Nikolaos; Chatzimichailidou, Sophia; Panagopoulos, Panagiotis; Botis, Ioannis; Aslanidis, Spyros; Douma, Stella

2017-06-01

Cardiac involvement is common in rheumatoid arthritis. Subendocardial viability ratio (SEVR) is a non-invasive measure of microvascular coronary perfusion, yet it remains unclear whether it is affected in rheumatoid arthritis patients. We additionally sought predictors of SEVR in rheumatoid arthritis among a wide range of disease-related parameters, cardiac and hemodynamic factors, and markers of atherosclerosis, arteriosclerosis, and endothelial dysfunction. SEVR was estimated in rheumatoid arthritis patients and healthy controls by applanation tonometry, which was also used to evaluate arterial stiffness (pulse wave velocity and augmentation index). In the rheumatoid arthritis group, carotid intima-media thickness (cIMT) was additionally estimated by ultrasound, cardiac and hemodynamic parameters by impedance cardiography, and endothelial dysfunction by measurement of asymmetric dimethylarginine (ADMA). In a total of 122 participants, SEVR was lower among 91 patients with rheumatoid arthritis compared to 31 controls (141.4 ± 21.9 vs 153.1 ± 18.7%, p = 0.009) and remained so among 29 rheumatoid arthritis patients without hypertension, diabetes, or cardiovascular diseases, compared to the control group (139.7 ± 21.7 vs 153.1 ± 18.7%, p = 0.013). SEVR did not significantly correlate with arterial stiffness, cIMT, ADMA, or disease-related parameters. Multivariate analysis revealed gender (p = 0.007), blood pressure (p = 0.028), heart rate (p = 0.025), cholesterol levels (p = 0.008), cardiac index (p < 0.001) and left ventricular ejection time (p = 0.004) as independent predictors of SEVR among patients with rheumatoid arthritis. Patients with rheumatoid arthritis exhibit lower values of SEVR compared to healthy individuals. Cardiac and hemodynamic parameters, rather than functional indices of endothelial and macrovascular dysfunction, may be useful as predictors of myocardial perfusion in rheumatoid arthritis.

miR-98 and let-7g* protect the blood–brain barrier under neuroinflammatory conditions

PubMed Central

Rom, Slava; Dykstra, Holly; Zuluaga-Ramirez, Viviana; Reichenbach, Nancy L; Persidsky, Yuri

2015-01-01

Pathologic conditions in the central nervous system, regardless of the underlying injury mechanism, show a certain level of blood–brain barrier (BBB) impairment. Endothelial dysfunction is the earliest event in the initiation of vascular damage caused by inflammation due to stroke, atherosclerosis, trauma, or brain infections. Recently, microRNAs (miRNAs) have emerged as a class of gene expression regulators. The relationship between neuroinflammation and miRNA expression in brain endothelium remains unexplored. Previously, we showed the BBB-protective and anti-inflammatory effects of glycogen synthase kinase (GSK) 3β inhibition in brain endothelium in in vitro and in vivo models of neuroinflammation. Using microarray screening, we identified miRNAs induced in primary human brain microvascular endothelial cells after exposure to the pro-inflammatory cytokine, tumor necrosis factor-α, with/out GSK3β inhibition. Among the highly modified miRNAs, let-7 and miR-98 were predicted to target the inflammatory molecules, CCL2 and CCL5. Overexpression of let-7 and miR-98 in vitro and in vivo resulted in reduced leukocyte adhesion to and migration across endothelium, diminished expression of pro-inflammatory cytokines, and increased BBB tightness, attenuating barrier ‘leakiness' in neuroinflammation conditions. For the first time, we showed that miRNAs could be used as a therapeutic tool to prevent the BBB dysfunction in neuroinflammation. PMID:26126865

miR-98 and let-7g* protect the blood-brain barrier under neuroinflammatory conditions.

PubMed

Rom, Slava; Dykstra, Holly; Zuluaga-Ramirez, Viviana; Reichenbach, Nancy L; Persidsky, Yuri

2015-12-01

Pathologic conditions in the central nervous system, regardless of the underlying injury mechanism, show a certain level of blood-brain barrier (BBB) impairment. Endothelial dysfunction is the earliest event in the initiation of vascular damage caused by inflammation due to stroke, atherosclerosis, trauma, or brain infections. Recently, microRNAs (miRNAs) have emerged as a class of gene expression regulators. The relationship between neuroinflammation and miRNA expression in brain endothelium remains unexplored. Previously, we showed the BBB-protective and anti-inflammatory effects of glycogen synthase kinase (GSK) 3β inhibition in brain endothelium in in vitro and in vivo models of neuroinflammation. Using microarray screening, we identified miRNAs induced in primary human brain microvascular endothelial cells after exposure to the pro-inflammatory cytokine, tumor necrosis factor-α, with/out GSK3β inhibition. Among the highly modified miRNAs, let-7 and miR-98 were predicted to target the inflammatory molecules, CCL2 and CCL5. Overexpression of let-7 and miR-98 in vitro and in vivo resulted in reduced leukocyte adhesion to and migration across endothelium, diminished expression of pro-inflammatory cytokines, and increased BBB tightness, attenuating barrier 'leakiness' in neuroinflammation conditions. For the first time, we showed that miRNAs could be used as a therapeutic tool to prevent the BBB dysfunction in neuroinflammation.

Analysis of vascular endothelial dysfunction genes and related pathways in obesity through systematic bioinformatics.

PubMed

Zhang, Hui; Wang, Jing; Sun, Ling; Xu, Qiuqin; Hou, Miao; Ding, Yueyue; Huang, Jie; Chen, Ye; Cao, Lei; Zhang, Jianmin; Qian, Weiguo; Lv, Haitao

2015-01-01

Obesity has become an increasingly serious health problem and popular research topic. It is associated with many diseases, especially cardiovascular disease (CVD)-related endothelial dysfunction. This study analyzed genes related to endothelial dysfunction and obesity and then summarized their most significant signaling pathways. Genes related to vascular endothelial dysfunction and obesity were extracted from a PubMed database, and analyzed by STRING, DAVID, and Gene-Go Meta-Core software. 142 genes associated with obesity were found to play a role in endothelial dysfunction in PubMed. A significant pathway (Angiotensin system maturation in protein folding and maturation) associated with obesity and endothelial dysfunction was explored. The genes and the pathway explored may play an important role in obesity. Further studies about preventing vascular endothelial dysfunction obesity should be conducted through targeting these loci and pathways.

Gene variations of nitric oxide synthase regulate the effects of a saturated fat rich meal on endothelial function

USDA-ARS?s Scientific Manuscript database

Objective: Endothelial nitric oxide synthase gene variations have been linked to a higher risk for cardiovascular diseases by unknown mechanisms. Our aim was to determine if two SNPs located in NOS3 (E298D and i19342) interfere with microvascular endothelial function (MEF) and/or oxidative stress du...

Diabetic ketoacidosis elicits systemic inflammation associated with cerebrovascular endothelial cell dysfunction.

PubMed

Close, Taylor E; Cepinskas, Gediminas; Omatsu, Tatsushi; Rose, Keeley L; Summers, Kelly; Patterson, Eric K; Fraser, Douglas D

2013-08-01

To determine if the DKA-induced inflammation in juvenile mice provokes activation and dysfunction of CVECs. DKA in juvenile mice was induced with administration of STZ and ALX. Blood from DKA mice was assessed for cytokines and soluble cell adhesion proteins, and either DKA plasma or exogenous compounds were applied to immortalized bEND3. DKA increased circulating levels of IL-6, IL-8(KC), MCP-1, IL-10, sE-selectin, sICAM-1, and sVCAM-1. Stimulation of bEND3 with DKA plasma caused cellular activation (increased ROS and activation of NF-κΒ), upregulation of a proadhesive phenotype (E-selectin, ICAM-1, and VCAM-1), and increased leukocyte-bEND3 interaction (leukocyte rolling/adhesion). TEER, a measure of bEND3 monolayer integrity, was decreased by DKA plasma. Activation and dysfunction of bEND3 with DKA plasma were suppressed by plasma heat treatment (56°C, 1 hour) and replicated with the application of DKA recombinant cytomix (IL-6, IL-8[KC], MCP-1, and IL-10), implicating circulating inflammatory protein(s) as mediators. Treatment of bEND3 with β-OH-butyrate, the main ketone elevated in DKA, failed to mimic the DKA plasma-induced activation and dysfunction of bEND3. DKA elicits systemic inflammation associated with CVEC activation and dysfunction, possibly contributing to DKA-associated intracranial microvascular complications. © 2013 John Wiley & Sons Ltd.

N-n-butyl Haloperidol Iodide Protects against Hypoxia/Reoxygenation Injury in Cardiac Microvascular Endothelial Cells by Regulating the ROS/MAPK/Egr-1 Pathway

PubMed Central

Lu, Shishi; Zhang, Yanmei; Zhong, Shuping; Gao, Fenfei; Chen, Yicun; Li, Weiqiu; Zheng, Fuchun; Shi, Ganggang

2017-01-01

Endothelium dysfunction induced by reactive oxygen species (ROS) is an important initial event at the onset of myocardial ischemia/reperfusion in which the Egr-1 transcription factor often serves as a master switch for various damage pathways following reperfusion injury. We hypothesized that an intracellular ROS/MAPK/Egr-1 signaling pathway is activated in cardiac microvascular endothelial cells (CMECs) following hypoxia/reoxygenation (H/R). ROS generation, by either H/R or the ROS donor xanthine oxidase-hypoxanthine (XO/HX) activated all three MAPKs (ERK1/2, JNK, p38), and induced Egr-1 expression and Egr-1 DNA-binding activity in CMECs, whereas ROS scavengers (EDA and NAC) had the opposite effect following H/R. Inhibitors of all three MAPKs individually inhibited induction of Egr-1 expression by H/R in CMECs. Moreover, N-n-butyl haloperidol (F2), previously shown to protect cardiomyocytes subjected to I/R, dose-dependently downregulated H/R-induced ROS generation, MAPK activation, and Egr-1 expression and activity in CMECs, whereas XO/HX and MAPK activators (EGF, anisomycin) antagonized the effects of F2. Inhibition of the ROS/MAPK/Egr-1 signaling pathway, by either F2, NAC, or inhibition of MAPK, increased CMEC viability and the GSH/GSSG ratio, and decreased Egr-1 nuclear translocation. These results show that the ROS/MAPK/Egr-1 signaling pathway mediates H/R injury in CMECs, and F2 blocks this pathway to protect against H/R injury and further alleviate myocardial I/R injury. PMID:28111550

Microvascular and mitochondrial dysfunction in the female F1 generation after gestational TiO2 nanoparticle exposure

PubMed Central

Stapleton, Phoebe A.; Nichols, Cody E.; Yi, Jinghai; McBride, Carroll R.; Minarchick, Valerie C.; Shepherd, Danielle L.; Hollander, John M.; Nurkiewicz, Timothy R.

2016-01-01

Due to the ongoing evolution of nanotechnology, there is a growing need to assess the toxicological outcomes in under-studied populations in order to properly consider the potential of engineered nanomaterials (ENM) and fully enhance their safety. Recently, we and others have explored the vascular consequences associated with gestational nanomaterial exposure, reporting microvascular dysfunction within the uterine circulation of pregnant dams and the tail artery of fetal pups. It has been proposed (via work derived by the Barker Hypothesis) that mitochondrial dysfunction and subsequent oxidative stress mechanisms as a possible link between a hostile gestational environment and adult disease. Therefore, in this study, we exposed pregnant Sprague-Dawley rats to nanosized titanium dioxide aerosols after implantation (gestational day 6). Pups were delivered, and the progeny grew into adulthood. Microvascular reactivity, mitochondrial respiration and hydrogen peroxide production of the coronary and uterine circulations of the female offspring were evaluated. While there were no significant differences within the maternal or litter characteristics, endothelium-dependent dilation and active mechanotransduction in both coronary and uterine arterioles were significantly impaired. In addition, there was a significant reduction in maximal mitochondrial respiration (state 3) in the left ventricle and uterus. These studies demonstrate microvascular dysfunction and coincide with mitochondrial inefficiencies in both the cardiac and uterine tissues, which may represent initial evidence that prenatal ENM exposure produces microvascular impairments that persist throughout multiple developmental stages. PMID:25475392

The lung in liver disease: old problem, new concepts.

PubMed

Fallon, Michael B; Zhang, Junlan

2013-01-01

Liver dysfunction has been recognized to influence the lung in many different clinical situations, although the mechanisms for these effects are not well understood. One increasingly recognized interaction, the hepatopulmonary syndrome (HPS) occurs in the context of cirrhosis and results when alveolar microvascular dilation causes arterial gas exchange abnormalities and hypoxemia. HPS occurs in up to 30% of patients with cirrhosis and significantly increases mortality in affected patients. Currently, liver transplantation is the only curative therapy. Experimental biliary cirrhosis induced by common bile duct ligation (CBDL) in the rat reproduces the pulmonary vascular and gas exchange abnormalities of human HPS and has been contrasted with other experimental models of cirrhosis in which HPS does not develop. Microvascular dilation, intravascular monocyte infiltration, and angiogenesis in the lung have been identified as pathologic features that drive gas exchange abnormalities in experimental HPS. Our recent studies have identified biliary epithelium and activation and interaction between the endothelin-1 (ET-1)/endothelial endothelin B (ETB) receptor and CX3CL1/CX3CR1 pathways as important mechanisms for the observed pathologic events. These studies define novel interactions between the lung and liver in cirrhosis and may lead to effective medical therapies.

Brain vascular heterogeneity: implications for disease pathogenesis and design of in vitro blood-brain barrier models.

PubMed

Noumbissi, Midrelle E; Galasso, Bianca; Stins, Monique F

2018-04-23

The vertebrate blood-brain barrier (BBB) is composed of cerebral microvascular endothelial cells (CEC). The BBB acts as a semi-permeable cellular interface that tightly regulates bidirectional molecular transport between blood and the brain parenchyma in order to maintain cerebral homeostasis. The CEC phenotype is regulated by a variety of factors, including cells in its immediate environment and within functional neurovascular units. The cellular composition of the brain parenchyma surrounding the CEC varies between different brain regions; this difference is clearly visible in grey versus white matter. In this review, we discuss evidence for the existence of brain vascular heterogeneity, focusing on differences between the vessels of the grey and white matter. The region-specific differences in the vasculature of the brain are reflective of specific functions of those particular brain areas. This BBB-endothelial heterogeneity may have implications for the course of pathogenesis of cerebrovascular diseases and neurological disorders involving vascular activation and dysfunction. This heterogeneity should be taken into account when developing BBB-neuro-disease models representative of specific brain areas.

Platelet lysate gel and endothelial progenitors stimulate microvascular network formation in vitro: tissue engineering implications.

PubMed

Fortunato, Tiago M; Beltrami, Cristina; Emanueli, Costanza; De Bank, Paul A; Pula, Giordano

2016-05-04

Revascularisation is a key step for tissue regeneration and complete organ engineering. We describe the generation of human platelet lysate gel (hPLG), an extracellular matrix preparation from human platelets able to support the proliferation of endothelial colony forming cells (ECFCs) in 2D cultures and the formation of a complete microvascular network in vitro in 3D cultures. Existing extracellular matrix preparations require addition of high concentrations of recombinant growth factors and allow only limited formation of capillary-like structures. Additional advantages of our approach over existing extracellular matrices are the absence of any animal product in the composition hPLG and the possibility of obtaining hPLG from patients to generate homologous scaffolds for re-implantation. This discovery has the potential to accelerate the development of regenerative medicine applications based on implantation of microvascular networks expanded ex vivo or the generation of fully vascularised organs.

Platelet lysate gel and endothelial progenitors stimulate microvascular network formation in vitro: tissue engineering implications

PubMed Central

Fortunato, Tiago M.; Beltrami, Cristina; Emanueli, Costanza; De Bank, Paul A.; Pula, Giordano

2016-01-01

Revascularisation is a key step for tissue regeneration and complete organ engineering. We describe the generation of human platelet lysate gel (hPLG), an extracellular matrix preparation from human platelets able to support the proliferation of endothelial colony forming cells (ECFCs) in 2D cultures and the formation of a complete microvascular network in vitro in 3D cultures. Existing extracellular matrix preparations require addition of high concentrations of recombinant growth factors and allow only limited formation of capillary-like structures. Additional advantages of our approach over existing extracellular matrices are the absence of any animal product in the composition hPLG and the possibility of obtaining hPLG from patients to generate homologous scaffolds for re-implantation. This discovery has the potential to accelerate the development of regenerative medicine applications based on implantation of microvascular networks expanded ex vivo or the generation of fully vascularised organs. PMID:27141997

Loss of endothelial barrier antigen immunoreactivity as a marker of Clostridium perfringens type D epsilon toxin-induced microvascular damage in rat brain.

PubMed

Finnie, J W; Manavis, J; Chidlow, G

2014-01-01

The epsilon toxin elaborated by Clostridium perfringens type D in the intestine of domestic livestock is principally responsible for the neurological disease produced after its absorption in excessive quantities into the systemic circulation. The fundamental basis of the cerebral damage induced by epsilon toxin appears to be microvascular injury with ensuing severe, diffuse vasogenic oedema. Endothelial barrier antigen (EBA), which is normally expressed by virtually all capillaries and venules in the rat brain, was used in this study as a marker of blood-brain barrier (BBB) integrity. After exposure to high levels of circulating epsilon toxin, there was substantial loss of EBA in many brain microvessels, attended by widespread plasma albumin extravasation. These results support microvascular injury and subsequent BBB breakdown as a key factor in the pathogenesis of epsilon toxin-induced neurological disease. Copyright © 2014 Elsevier Ltd. All rights reserved.

Role of Vascular and Lymphatic Endothelial Cells in Hantavirus Pulmonary Syndrome Suggests Targeted Therapeutic Approaches

PubMed Central

Gorbunova, Elena E.; Dalrymple, Nadine A.; Gavrilovskaya, Irina N.

2013-01-01

Abstract Background Hantaviruses in the Americas cause a highly lethal acute pulmonary edema termed hantavirus pulmonary syndrome (HPS). Hantaviruses nonlytically infect microvascular and lymphatic endothelial cells and cause dramatic changes in barrier functions without disrupting the endothelium. Hantaviruses cause changes in the function of infected endothelial cells that normally regulate fluid barrier functions. The endothelium of arteries, veins, and lymphatic vessels are unique and central to the function of vast pulmonary capillary beds that regulate pulmonary fluid accumulation. Results We have found that HPS-causing hantaviruses alter vascular barrier functions of microvascular and lymphatic endothelial cells by altering receptor and signaling pathway responses that serve to permit fluid tissue influx and clear tissue edema. Infection of the endothelium provides several mechanisms for hantaviruses to cause acute pulmonary edema, as well as potential therapeutic targets for reducing the severity of HPS disease. Conclusions Here we discuss interactions of HPS-causing hantaviruses with the endothelium, roles for unique lymphatic endothelial responses in HPS, and therapeutic targeting of the endothelium as a means of reducing the severity of HPS disease. PMID:24024573

Role of vascular and lymphatic endothelial cells in hantavirus pulmonary syndrome suggests targeted therapeutic approaches.

PubMed

Mackow, Erich R; Gorbunova, Elena E; Dalrymple, Nadine A; Gavrilovskaya, Irina N

2013-09-01

Hantaviruses in the Americas cause a highly lethal acute pulmonary edema termed hantavirus pulmonary syndrome (HPS). Hantaviruses nonlytically infect microvascular and lymphatic endothelial cells and cause dramatic changes in barrier functions without disrupting the endothelium. Hantaviruses cause changes in the function of infected endothelial cells that normally regulate fluid barrier functions. The endothelium of arteries, veins, and lymphatic vessels are unique and central to the function of vast pulmonary capillary beds that regulate pulmonary fluid accumulation. We have found that HPS-causing hantaviruses alter vascular barrier functions of microvascular and lymphatic endothelial cells by altering receptor and signaling pathway responses that serve to permit fluid tissue influx and clear tissue edema. Infection of the endothelium provides several mechanisms for hantaviruses to cause acute pulmonary edema, as well as potential therapeutic targets for reducing the severity of HPS disease. Here we discuss interactions of HPS-causing hantaviruses with the endothelium, roles for unique lymphatic endothelial responses in HPS, and therapeutic targeting of the endothelium as a means of reducing the severity of HPS disease.

Human brain microvascular endothelial cells resist elongation due to shear stress.

PubMed

Reinitz, Adam; DeStefano, Jackson; Ye, Mao; Wong, Andrew D; Searson, Peter C

2015-05-01

Endothelial cells in straight sections of vessels are known to elongate and align in the direction of flow. This phenotype has been replicated in confluent monolayers of bovine aortic endothelial cells and human umbilical vein endothelial cells (HUVECs) in cell culture under physiological shear stress. Here we report on the morphological response of human brain microvascular endothelial cells (HBMECs) in confluent monolayers in response to shear stress. Using a microfluidic platform we image confluent monolayers of HBMECs and HUVECs under shear stresses up to 16 dyne cm(-2). From live-cell imaging we quantitatively analyze the cell morphology and cell speed as a function of time. We show that HBMECs do not undergo a classical transition from cobblestone to spindle-like morphology in response to shear stress. We further show that under shear stress, actin fibers are randomly oriented in the cells indicating that there is no cytoskeletal remodeling. These results suggest that HBMECs are programmed to resist elongation and alignment under shear stress, a phenotype that may be associated with the unique properties of the blood-brain barrier. Copyright © 2015 Elsevier Inc. All rights reserved.

Better microvascular function on long-term treatment with lisinopril than with nifedipine in renal transplant recipients.

PubMed

Asberg, A; Midtvedt, K; Vassbotn, T; Hartmann, A

2001-07-01

The prevalence of hypertension in renal transplant recipients is high but the pathophysiology is poorly defined. Impaired endothelial function may be a factor of major importance. The present study addresses the effects of long-term treatment with either lisinopril or slow-release nifedipine on microvascular function and plasma endothelin in renal transplant recipients on cyclosporin A (CsA). Seventy-five hypertensive renal transplant recipients were double-blind randomized to receive slow-release nifedipine (NIF, n=40) or lisinopril (LIS, n=35). Ten normotensive, age-matched recipients served as controls. All patients received CsA-based immunosuppressive therapy including prednisolone and azathioprine. Microvascular function was assessed in the forearm skin vasculature, using laser Doppler flowmetry in combination with post-occlusive reactive hyperaemia and endothelial-dependent function during local acetylcholine (ACh) stimulation. The analysis of microvascular function (AUC(rh)) showed that nifedipine-treated patients had significantly lower responses compared with lisinopril-treated patients (20+/-17 and 43+/-20 AU x min respectively, P=0.0016). Endothelial function was borderline significantly lower in the NIF group compared with the LIS group (640+/-345 and 817+/-404 AU x min respectively, P=0.056). The responses in the LIS group were comparable with those in non-hypertensive controls (AUC(rh) was 37+/-16 and AUC(ACh) was 994+/-566 AU x min). Plasma endothelin-1 concentrations were significantly higher in the NIF group compared with the LIS group (0.44+/-0.19 vs. 0.34+/-0.10 fmol/ml respectively, P=0.048), and were 0.29+/-0.09 fmol/ml in the control patients. AUC(ACh) was associated with plasma endothelin-1 (P=0.0053), while AUC(rh) was not (P=0.080). The study indicates that long-term treatment with lisinopril, when compared with nifedipine, yields a more beneficial effect on microvascular function in hypertensive renal transplant recipients on CsA. The beneficial microvascular effect may be mediated in part by an endothelin-1-associated effect on the endothelium.

Defibrotide prevents the activation of macrovascular and microvascular endothelia caused by soluble factors released to blood by autologous hematopoietic stem cell transplantation.

PubMed

Palomo, Marta; Diaz-Ricart, Maribel; Rovira, Montserrat; Escolar, Ginés; Carreras, Enric

2011-04-01

Endothelial activation and damage occur in association with autologous hematopoietic stem cell transplantation (HSCT). Several of the early complications associated with HSCT seem to have a microvascular location. Through the present study, we have characterized the activation and damage of endothelial cells of both macro (HUVEC) and microvascular (HMEC) origin, occurring early after autologous HSCT, and the potential protective effect of defibrotide (DF). Sera samples from patients were collected before conditioning (Pre), at the time of transplantation (day 0), and at days 7, 14, and 21 after autologous HSCT. Changes in the expression of endothelial cell receptors at the surface, presence and reactivity of extracellular adhesive proteins, and the signaling pathways involved were analyzed. The expression of ICAM-1 at the cell surface increased progressively in both HUVEC and HMEC. However, a more prothrombotic profile was denoted for HMEC, in particular at the time of transplantation (day 0), reflecting the deleterious effect of the conditioning treatment on the endothelium, especially at a microvascular location. Interestingly, this observation correlated with a higher increase in the expression of both tissue factor and von Willebrand factor on the extracellular matrix, together with activation of intracellular p38 MAPK and Akt. Previous exposure and continuous incubation of cells with DF prevented the signs of activation and damage induced by the autologous sera. These observations corroborate that conditioning treatment in autologous HSCT induces a proinflammatory and a prothrombotic phenotype, especially at a microvascular location, and indicate that DF has protective antiinflammatory and antithrombotic effects in this setting. Copyright © 2011 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.

The angiopoietin1-Akt pathway regulates barrier function of the cultured spinal cord microvascular endothelial cells through Eps8.

PubMed

Liu, Xinchun; Zhou, Xiaoshu; Yuan, Wei

2014-10-15

In mammalian central nervous system (CNS), the integrity of the blood-spinal cord barrier (BSCB), formed by tight junctions (TJs) between adjacent microvascular endothelial cells near the basement membrane of capillaries and the accessory structures, is important for relatively independent activities of the cellular constituents inside the spinal cord. The barrier function of the BSCB are tightly regulated and coordinated by a variety of physiological or pathological factors, similar with but not quite the same as its counterpart, the blood-brain barrier (BBB). Herein, angiopoietin 1 (Ang1), an identified ligand of the endothelium-specific tyrosine kinase receptor Tie-2, was verified to regulate barrier functions, including permeability, junction protein interactions and F-actin organization, in cultured spinal cord microvascular endothelial cells (SCMEC) of rat through the activity of Akt. Besides, these roles of Ang1 in the BSCB in vitro were found to be accompanied with an increasing expression of epidermal growth factor receptor pathway substrate 8 (Eps8), an F-actin bundling protein. Furthermore, the silencing of Eps8 by lentiviral shRNA resulted in an antagonistic effect vs. Ang1 on the endothelial barrier function of SCMEC. In summary, the Ang1-Akt pathway serves as a regulator in the barrier function modulation of SCMEC via the actin-binding protein Eps8. Copyright © 2014 Elsevier Inc. All rights reserved.

Selective endothelin ETA and dual ET(A)/ET(B) receptor blockade improve endothelium-dependent vasodilatation in patients with type 2 diabetes and coronary artery disease.

PubMed

Rafnsson, Arnar; Shemyakin, Alexey; Pernow, John

2014-11-24

Endothelin-1 contributes to endothelial dysfunction in patients with atherosclerosis and type 2 diabetes. In healthy arteries the ETA receptor mediates the main part of the vasoconstriction induced by endothelin-1 whilst the ETB receptor mediates vasodilatation. The ETB receptor expression is upregulated on vascular smooth muscle cells in atherosclerosis and may contribute to the increased vasoconstrictor tone and endothelial dysfunction observed in this condition. Due to these opposing effects of the ETB receptor it remains unclear whether ETB blockade together with ETA blockade may be detrimental or beneficial. The aim was therefore to compare the effects of selective ETA and dual ETA/ETB blockade on endothelial function in patients with type 2 diabetes and coronary artery disease. Forearm endothelium-dependent and endothelium-independent vasodilatation was assessed by venous occlusion plethysmography in 12 patients before and after selective ETA or dual ETA/ETB receptor blockade. Dual ETA/ETB receptor blockade increased baseline forearm blood flow by 30±14% (P<0.01) whereas selective ETA blockade did not (14±8%). Both selective ETA blockade and dual ETA/ETB blockade significantly improved endothelium-dependent vasodilatation. The improvement did not differ between the two treatments. There was also an increase in endothelium-independent vasodilatation with both treatments. Dual ETA/ETB blockade did not significantly increase microvascular flow but improved transcutaneous pO2. Both selective ETA and dual ETA/ETB improve endothelium-dependent vasodilatation in patients with type 2 diabetes and coronary artery disease. ETB blockade increases basal blood flow but does not additionally improve endothelium-dependent vasodilatation. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

BAMBI Elimination Enhances Alternative TGF-β Signaling and Glomerular Dysfunction in Diabetic Mice

PubMed Central

Fan, Ying; Li, Xuezhu; Xiao, Wenzhen; Fu, Jia; Harris, Ray C.; Lindenmeyer, Maja; Cohen, Clemens D.; Guillot, Nicolas; Baron, Margaret H.; Wang, Niansong; Lee, Kyung; He, John C.; Chuang, Peter Y.

2015-01-01

BMP, activin, membrane-bound inhibitor (BAMBI) acts as a pseudo-receptor for the transforming growth factor (TGF)-β type I receptor family and a negative modulator of TGF-β kinase signaling, and BAMBI−/− mice show mild endothelial dysfunction. Because diabetic glomerular disease is associated with TGF-β overexpression and microvascular alterations, we examined the effect of diabetes on glomerular BAMBI mRNA levels. In isolated glomeruli from biopsies of patients with diabetic nephropathy and in glomeruli from mice with type 2 diabetes, BAMBI was downregulated. We then examined the effects of BAMBI deletion on streptozotocin-induced diabetic glomerulopathy in mice. BAMBI−/− mice developed more albuminuria, with a widening of foot processes, than BAMBI+/+ mice, along with increased activation of alternative TGF-β pathways such as extracellular signal–related kinase (ERK)1/2 and Smad1/5 in glomeruli and cortices of BAMBI−/− mice. Vegfr2 and Angpt1, genes controlling glomerular endothelial stability, were downmodulated in glomeruli from BAMBI−/− mice with diabetes. Incubation of glomeruli from nondiabetic BAMBI+/+ or BAMBI−/− mice with TGF-β resulted in the downregulation of Vegfr2 and Angpt1, effects that were more pronounced in BAMBI−/− mice and were prevented by a MEK inhibitor. The downregulation of Vegfr2 in diabetes was localized to glomerular endothelial cells using a histone yellow reporter under the Vegfr2 promoter. Thus, BAMBI modulates the effects of diabetes on glomerular permselectivity in association with altered ERK1/2 and Smad1/5 signaling. Future therapeutic interventions with inhibitors of alternative TGF-β signaling may therefore be of interest in diabetic nephropathy. PMID:25576053

Linagliptin reduces effects of ET-1 and TLR2-mediated cerebrovascular hyperreactivity in diabetes.

PubMed

Hardigan, Trevor; Abdul, Yasir; Ergul, Adviye

2016-08-15

The anti-hyperglycemic agent linagliptin, a dipeptidyl peptidase-4 inhibitor, has been shown to reduce inflammation and improve endothelial cell function. In this study, we hypothesized that DPP-IV inhibition with linagliptin would improve impaired cerebral blood flow in diabetic rats through improved insulin-induced cerebrovascular relaxation and reversal of pathological cerebrovascular remodeling that subsequently leads to improvement of cognitive function. Male type-2 diabetic Goto-Kakizaki (GK) and nondiabetic Wistar rats were treated with linagliptin, and ET-1 plasma levels and dose response curves to ET-1 (0.1-100nM) in basilar arteries were assessed. The impact of TLR2 antagonism on ET-1 mediated basilar contraction and endothelium-dependent relaxation to acetylcholine (ACh, 1nM-1M) in diabetic GK rats was examined with antibody directed against the TLR2 receptor (Santa Cruz, 5μg/mL). The expression of TLR2 in middle cerebral arteries (MCAs) from treated rats and in brain microvascular endothelial cells (BMVEC) treated with 100nM linagliptin was assessed. Linagliptin lowered plasma ET-1 levels in diabetes, and reduced ET-1-induced vascular contraction. TLR2 antagonism in diabetic basilar arteries reduced ET-1-mediated cerebrovascular dysfunction and improved endothelium-dependent vasorelaxation. Linagliptin treatment in the BMVEC was able to reduce TLR2 expression in cells from both diabetic and nondiabetic rats. These results suggest that inhibition of DPPIV using linagliptin improves the ET-1-mediated cerebrovascular dysfunction observed in diabetes through a reduction in ET-1 plasma levels and reduced cerebrovascular hyperreactivity. This effect is potentially a result of linagliptin causing a decrease in endothelial TLR2 expression and a subsequent increase in NO bioavailability. Copyright © 2016 Elsevier Inc. All rights reserved.

Human severe sepsis cytokine mixture increases β2-integrin-dependent polymorphonuclear leukocyte adhesion to cerebral microvascular endothelial cells in vitro.

PubMed

Blom, Chris; Deller, Brittany L; Fraser, Douglas D; Patterson, Eric K; Martin, Claudio M; Young, Bryan; Liaw, Patricia C; Yazdan-Ashoori, Payam; Ortiz, Angelica; Webb, Brian; Kilmer, Greg; Carter, David E; Cepinskas, Gediminas

2015-04-07

Sepsis-associated encephalopathy (SAE) is a state of acute brain dysfunction in response to a systemic infection. We propose that systemic inflammation during sepsis causes increased adhesion of leukocytes to the brain microvasculature, resulting in blood-brain barrier dysfunction. Thus, our objectives were to measure inflammatory analytes in plasma of severe sepsis patients to create an experimental cytokine mixture (CM), and to use this CM to investigate the activation and interactions of polymorphonuclear leukocytes (PMN) and human cerebrovascular endothelial cells (hCMEC/D3) in vitro. The concentrations of 41 inflammatory analytes were quantified in plasma obtained from 20 severe sepsis patients and 20 age- and sex-matched healthy controls employing an antibody microarray. Two CMs were prepared to mimic severe sepsis (SSCM) and control (CCM), and these CMs were then used for PMN and hCMEC/D3 stimulation in vitro. PMN adhesion to hCMEC/D3 was assessed under conditions of flow (shear stress 0.7 dyn/cm(2)). Eight inflammatory analytes elevated in plasma obtained from severe sepsis patients were used to prepare SSCM and CCM. Stimulation of PMN with SSCM led to a marked increase in PMN adhesion to hCMEC/D3, as compared to CCM. PMN adhesion was abolished with neutralizing antibodies to either β2 (CD18), αL/β2 (CD11α/CD18; LFA-1) or αM/β2 (CD11β/CD18; Mac-1) integrins. In addition, immune-neutralization of the endothelial (hCMEC/D3) cell adhesion molecule, ICAM-1 (CD54) also suppressed PMN adhesion. Human SSCM up-regulates PMN pro-adhesive phenotype and promotes PMN adhesion to cerebrovascular endothelial cells through a β2-integrin-ICAM-1-dependent mechanism. PMN adhesion to the brain microvasculature may contribute to SAE.

Horizontal alignment of 5' -> 3' intergene distance segment tropy with respect to the gene as the conserved basis for DNA transcription.

PubMed

Sarin, Hemant

2017-03-01

To study the conserved basis for gene expression in comparative cell types at opposite ends of the cell pressuromodulation spectrum, the lymphatic endothelial cell and the blood microvascular capillary endothelial cell. The mechanism for gene expression is studied in terms of the 5' -> 3' direction paired point tropy quotients ( prpT Q s) and the final 5' -> 3' direction episodic sub-episode block sums split-integrated weighted average-averaged gene overexpression tropy quotient ( esebssiwaagoT Q ). The final 5' -> 3' esebssiwaagoT Q classifies an lymphatic endothelial cell overexpressed gene as a supra-pressuromodulated gene ( esebssiwaagoT Q ≥ 0.25 < 0.75) every time and classifies a blood microvascular capillary endothelial cell overexpressed gene every time as an infra-pressuromodulated gene ( esebssiwaagoT Q < 0.25) (100% sensitivity; 100% specificity). Horizontal alignment of 5' -> 3' intergene distance segment tropy wrt the gene is the basis for DNA transcription in the pressuromodulated state.

Beta-adrenergic stimulation contributes to maintenance of endothelial barrier functions under baseline conditions.

PubMed

Spindler, Volker; Waschke, Jens

2011-02-01

cAMP signaling within the endothelium is known to reduce paracellular permeability and to protect against loss of barrier functions under various pathological conditions. Because activation of β-adrenergic receptors elevates cellular cAMP, we tested whether β-adrenergic receptor signaling contributes to the maintenance of baseline endothelial barrier properties. We compared hydraulic conductivity of rat postcapillary venules in vivo with resistance measurements and with reorganization of endothelial adherens junctions in cultured microvascular endothelial cells downstream of β-adrenergic receptor-mediated changes of cAMP levels. Inhibition of β-adrenergic receptors by propranolol increased hydraulic conductivity, reduced both cAMP levels and TER of microvascular endothelial cell monolayers and induced fragmentation of VE-cadherin staining. In contrast, activation by epinephrine both increased cAMP levels and TER and resulted in linearized VE-cadherin distribution, however this was not sufficient to block barrier-destabilization by propranolol. Similarly, PDE inhibition did not prevent propranolol-induced TER reduction and VE-cadherin reorganization whereas increased cAMP formation by AC activation enhanced endothelial barrier functions under baseline conditions and under conditions of propranolol treatment. Our results indicate that generation of cAMP mediated by activation of β-adrenergic receptor signaling contributes to the maintenance of endothelial barrier properties under baseline conditions. © 2011 John Wiley & Sons Ltd.

Tumour necrosis factor-α-mediated disruption of cerebrovascular endothelial barrier integrity in vitro involves the production of proinflammatory interleukin-6.

PubMed

Rochfort, Keith D; Collins, Laura E; McLoughlin, Alisha; Cummins, Philip M

2016-02-01

The co-involvement of tumour necrosis factor-α (TNF-α) and interleukin-6 (IL-6) during blood-brain barrier (BBB) injury has been reported in various models of neuroinflammation, although the precise functional interplay between these archetypal proinflammatory cytokines remains largely undefined within this context. In the current paper, we tested the hypothesis that TNF-α-mediated BBB disruption is measurably attributable in-part to induction of microvascular endothelial IL-6 production. In initial experiments, we observed that treatment of human brain microvascular endothelial cells (HBMvECs) with TNF-α (0-100 ng/mL, 0-24 h) robustly elicited both time- and dose-dependent induction of IL-6 expression and release, as well as expression of the IL-6 family receptor, GP130. Further experiments demonstrated that the TNF-α-dependent generation of reactive oxygen species, down-regulation of adherens/tight junction proteins, and concomitant elevation of HBMvEC permeability, were all significantly attenuated by blockade of IL-6 signalling using either an anti-IL-6 neutralizing antibody or an IL-6 siRNA. Based on these observations, we conclude that TNF-α treatment of HBMvECs in vitro activates IL-6 production and signalling, events that were shown to synergize with TNF-α actions to elicit HBMvEC permeabilization. These novel findings offer a constructive insight into the specific contribution of downstream cytokine induction to the injurious actions of TNF-α at the BBB microvascular endothelium interface. The co-involvement of tumour necrosis factor-α (TNF-α) and interleukin-6 (IL-6) during blood-brain barrier (BBB) injury has been widely reported. Using human brain microvascular endothelial cells (HBMvEC), we show that TNF-α-mediated BBB disruption is measurably attributable in-part to induction of endothelial IL-6 production and signalling. We demonstrate that the TNF-α-dependent generation of reactive oxygen species (ROS), down-regulation of interendothelial junctions, and concomitant elevation of HBMvEC permeability, could be significantly attenuated by using either an IL-6 neutralizing antibody or an IL-6-specific siRNA. These findings provide insight into the complex nature of proinflammatory cytokine injury at the BBB microvascular endothelium interface. © 2015 International Society for Neurochemistry.

Lipopolysaccharide-induced endothelial barrier breakdown is cyclic adenosine monophosphate dependent in vivo and in vitro.

PubMed

Schlegel, Nicolas; Baumer, Yvonne; Drenckhahn, Detlev; Waschke, Jens

2009-05-01

To determine whether cyclic adenosine monophosphate (cAMP) is critically involved in lipopolysaccharide (LPS)-induced breakdown of endothelial barrier functions in vivo and in vitro. Experimental laboratory research. Research laboratory. Wistar rats and cultured human microvascular endothelial cells. Permeability measurements in single postcapillary venules in vivo and permeability measurements and cell biology techniques in vitro. We demonstrate that within 120 minutes LPS increased endothelial permeability in rat mesenteric postcapillary venules in vivo and caused a barrier breakdown in human dermal microvascular endothelial cells in vitro. This was associated with the formation of large intercellular gaps and fragmentation of vascular endothelial cadherin immunostaining. Furthermore, claudin 5 immunostaining at cell borders was drastically reduced after LPS treatment. Interestingly, activity of the small GTPase Rho A, which has previously been suggested to mediate the LPS-induced endothelial barrier breakdown, was not increased after 2 hours. However, activity of Rac 1, which is known to be important for maintenance of endothelial barrier functions, was significantly reduced to 64 +/- 8% after 2 hours. All LPS-induced changes of endothelial cells were blocked by a forskolin-mediated or rolipram-mediated increase of cAMP. Consistently, enzyme-linked immunosorbent assay-based measurements demonstrated that LPS significantly decreased intracellular cAMP. In summary, our data demonstrate that LPS disrupts endothelial barrier properties by decreasing intracellular cAMP. This mechanism may involve inactivation of Rac 1 rather than activation of Rho A.

Ripk3 regulates cardiac microvascular reperfusion injury: The role of IP3R-dependent calcium overload, XO-mediated oxidative stress and F-action/filopodia-based cellular migration.

PubMed

Zhou, Hao; Wang, Jin; Zhu, Pingjun; Hu, Shunying; Ren, Jun

2018-05-01

Ripk3-mediated cellular apoptosis is a major contributor to the pathogenesis of myocardial ischemia reperfusion (IR) injury. However, the mechanisms by which Ripk3 influences microvascular homeostasis and endothelial apoptosis are not completely understood. In this study, loss of Ripk3 inhibited endothelial apoptosis, alleviated luminal swelling, maintained microvasculature patency, reduced the expression of adhesion molecules and limited the myocardial inflammatory response. In vitro, Ripk3 deficiency protected endothelial cells from apoptosis and migratory arrest induced by HR injury. Mechanistically, Ripk3 had the ability to migrate onto the endoplasmic reticulum (ER), leading to ER damage, as evidenced by increased IP3R and XO expression. The higher IP3R content was associated with cellular calcium overload, and increased XO expression was involved in cellular oxidative injury. Furthermore, IP3R-mediated calcium overload and XO-dependent oxidative damage were able to initiate cellular apoptosis. More importantly, IP3R and XO also caused F-actin degradation into G-actin via post-transcriptional modification of cofilin, impairing the formation of the filopodia and limiting the migratory response of endothelial cells. Altogether, our data confirmed that Ripk3 was involved in microvascular IR injury via regulation of IP3R-mediated calcium overload, XO-dependent oxidative damage and filopodia-related cellular migration, ultimately leading to endothelial apoptosis and migratory inhibition. These findings provide a potential target for treating cardiac microcirculatory IR injury. Copyright © 2018 Elsevier Inc. All rights reserved.

Vascular endothelial growth factor receptor 1 contributes to Escherichia coli K1 invasion of human brain microvascular endothelial cells through the phosphatidylinositol 3-kinase/Akt signaling pathway.

PubMed

Zhao, Wei-Dong; Liu, Wei; Fang, Wen-Gang; Kim, Kwang Sik; Chen, Yu-Hua

2010-11-01

Escherichia coli is the most common Gram-negative organism causing neonatal meningitis. Previous studies demonstrated that E. coli K1 invasion of brain microvascular endothelial cells (BMEC) is required for penetration into the central nervous system, but the microbe-host interactions that are involved in this process remain incompletely understood. Here we report the involvement of vascular endothelial growth factor receptor 1 (VEGFR1) expressed on human brain microvascular endothelial cells (HBMEC) in E. coli K1 invasion of HBMEC. Our results showed that treatment of confluent HBMEC with pan-VEGFR inhibitors significantly inhibited E. coli K1 invasion of HBMEC. Immunofluorescence results indicated the colocalization of VEGFR1 with E. coli K1 during bacterial invasion of HBMEC. The E. coli-induced actin cytoskeleton rearrangements in HBMEC were blocked by VEGFR inhibitors but not by VEGFR2-specific inhibitors. The small interfering RNA (siRNA) knockdown of VEGFR1 in HBMEC significantly attenuated E. coli invasion and the concomitant actin filament rearrangement. Furthermore, we found an increased association of VEGFR1 with the p85 subunit of phosphatidylinositol 3-kinase (PI3K) in HBMEC infected with E. coli K1 and that E. coli K1-triggered Akt activation in HBMEC was blocked by VEGFR1 siRNA and VEGFR inhibitors. Taken together, our results demonstrate that VEGFR1 contributes to E. coli K1 invasion of HBMEC via recruitment of the PI3K/Akt signaling pathway.

Targeting Microvascular Pericytes in Angiogenic Vessels of Prostate Cancer

DTIC Science & Technology

2006-04-01

Schlingemann RO. 2004. In vivo angiogenic phenotype of endothelial cells and pericytes induced by vascular endothelial growth factor -a. J Histochem Cytochem...R, McDonald DM. Age-related changes in vascular endothelial growth factor dependency and angiopoietin-1-induced plasti- city of adult blood vessels...hematopoietic progenitor cells and their progeny in vivo . We used the basic fibroblast growth factor (bFGF)- induced mouse corneal neovascularization

Recombinant Factor XIII Mitigates Hemorrhagic Shock-Induced Organ Dysfunction

PubMed Central

Zaets, Sergey B.; Xu, Da-Zhong; Lu, Qi; Feketova, Eleonora; Berezina, Tamara L.; Malinina, Inga V.; Deitch, Edwin A.; Olsen, Eva H.

2012-01-01

Background Plasma factor XIII (FXIII) is responsible for stabilization of fibrin clot at the final stage of blood coagulation. Since FXIII has also been shown to modulate inflammation, endothelial permeability, as well as diminish multiple organ dysfunction (MOD) after gut ischemia-reperfusion injury, we hypothesized that FXIII would reduce MOD caused by trauma-hemorrhagic shock (THS). Materials and methods Rats were subjected to a 90 min THS or trauma sham shock (TSS) and treated with either recombinant human FXIII A2 subunit (rFXIII) or placebo immediately after resuscitation with shed blood or at the end of the TSS period. Lung permeability, lung and gut myeloperoxidase (MPO) activity, gut histology, neutrophil respiratory burst, microvascular blood flow in the liver and muscles, and cytokine levels were measured 3 h after the THS or TSS. FXIII levels were measured before THS or TSS and after the 3-h post-shock period. Results THS-induced lung permeability as well as lung and gut MPO activity was significantly lower in rFXIII-treated than in placebo-treated animals. Similarly, rFXIII-treated rats had lower neutrophil respiratory burst activity and less ileal mucosal injury. rFXIII-treated rats also had a higher liver microvascular blood flow compared with the placebo group. Cytokine response was more favorable in rFXIII-treated animals. Trauma-hemorrhagic shock did not cause a drop in FXIII activity during the study period. Conclusions Administration of rFXIII diminishes THS-induced MOD in rats, presumably by preservation of the gut barrier function, limitation of polymorphonuclear leukocyte (PMN) activation, and modulation of the cytokine response. PMID:21276979

Biophysical markers of the peripheral vasoconstriction response to pain in sickle cell disease

PubMed Central

Khaleel, Maha; Sunwoo, John; Shah, Payal; Detterich, Jon A.; Kato, Roberta M.; Thuptimdang, Wanwara; Meiselman, Herbert J.; Sposto, Richard; Tsao, Jennie; Wood, John C.; Zeltzer, Lonnie; Coates, Thomas D.; Khoo, Michael C. K.

2017-01-01

Painful vaso-occlusive crisis (VOC), a complication of sickle cell disease (SCD), occurs when sickled red blood cells obstruct flow in the microvasculature. We postulated that exaggerated sympathetically mediated vasoconstriction, endothelial dysfunction and the synergistic interaction between these two factors act together to reduce microvascular flow, promoting regional vaso-occlusions, setting the stage for VOC. We previously found that SCD subjects had stronger vasoconstriction response to pulses of heat-induced pain compared to controls but the relative degrees to which autonomic dysregulation, peripheral vascular dysfunction and their interaction are present in SCD remain unknown. In the present study, we employed a mathematical model to decompose the total vasoconstriction response to pain into: 1) the neurogenic component, 2) the vascular response to blood pressure, 3) respiratory coupling and 4) neurogenic-vascular interaction. The model allowed us to quantify the contribution of each component to the total vasoconstriction response. The most salient features of the components were extracted to represent biophysical markers of autonomic and vascular impairment in SCD and controls. These markers provide a means of phenotyping severity of disease in sickle-cell anemia that is based more on underlying physiology than on genotype. The marker of the vascular component (BMv) showed stronger contribution to vasoconstriction in SCD than controls (p = 0.0409), suggesting a dominant myogenic response in the SCD subjects as a consequence of endothelial dysfunction. The marker of neurogenic-vascular interaction (BMn-v) revealed that the interaction reinforced vasoconstriction in SCD but produced vasodilatory response in controls (p = 0.0167). This marked difference in BMn-v suggests that it is the most sensitive marker for quantifying combined alterations in autonomic and vascular function in SCD in response to heat-induced pain. PMID:28542469

New therapeutic modality for corneal endothelial disease using Rho-associated kinase inhibitor eye drops.

PubMed

Koizumi, Noriko; Okumura, Naoki; Ueno, Morio; Kinoshita, Shigeru

2014-11-01

Corneal endothelial dysfunction accompanied by visual disturbance is a primary indication for corneal endothelial transplantation. However, despite the value and potential of endothelial graft surgery, a strictly pharmacological approach for treating corneal endothelial dysfunction remains an attractive proposition. Previously, we reported that the selective Rho-associated kinase (ROCK) inhibitor Y-27632 promotes cell adhesion and proliferation, and inhibits the apoptosis of primate corneal endothelial cells in culture. These findings have led us to develop a novel medical treatment for the early phase of corneal endothelial disease using ROCK inhibitor eye drops. In rabbit and monkey models of partial endothelial dysfunction, we showed that corneal endothelial wound healing was accelerated via the topical application of ROCK inhibitor to the ocular surface, resulting in the regeneration of a corneal endothelial monolayer with a high endothelial cell density. Based on these animal studies, we are now attempting to advance the clinical application of ROCK inhibitor eye drops for patients with corneal endothelial dysfunction. A pilot clinical study was performed at the Kyoto Prefectural University of Medicine, and the effects of Y-27632 eye drops after transcorneal freezing were evaluated in 8 patients with corneal endothelial dysfunction. We observed a positive effect of ROCK inhibitor eye drops in treating patients with central edema caused by Fuchs corneal endothelial dystrophy. We believe that our new findings will contribute to the establishment of a new approach for the treatment of corneal endothelial dysfunction.

Mitochondrial Fission Triggered by Hyperglycemia Is Mediated by ROCK1 Activation in Podocytes and Endothelial Cells

PubMed Central

Wang, Wenjian; Wang, Yin; Long, Jianyin; Wang, Jinrong; Haudek, Sandra B.; Overbeek, Paul; Chang, Benny H.J.; Schumacker, Paul T.; Danesh, Farhad R.

2012-01-01

SUMMARY Several lines of evidence suggest that mitochondrial dysfunction plays a critical role in the pathogenesis of microvascular complications of diabetes, including diabetic nephropathy. However, the signaling pathways by which hyperglycemia leads to mitochondrial dysfunction are not fully understood. Here we examined the role of Rho-associated coiled-coil containing protein kinase 1 (ROCK1) on mitochondrial dynamics by generating two diabetic mouse models with targeted deletions of ROCK1, and an inducible podocyte-specific knock-in mouse expressing a constitutively active (cA) mutant of ROCK1. Our findings suggest that ROCK1 mediates hyperglycemia-induced mitochondrial fission by promoting dynamin-related protein-1 (Drp1) recruitment to the mitochondria. Deletion of ROCK1 in diabetic mice prevented mitochondrial fission, whereas podocyte-specific cA-ROCK1 mice exhibited increased mitochondrial fission. Importantly, we found that ROCK1 triggers mitochondrial fission by phosphorylating Drp1 at Serine 600 residue. These findings provide insights into the unexpected role of ROCK1 in a signaling cascade that regulates mitochondrial dynamics. PMID:22326220

Diabetic retinopathy: loss of neuroretinal adaptation to the diabetic metabolic environment.

PubMed

Abcouwer, Steven F; Gardner, Thomas W

2014-04-01

Diabetic retinopathy (DR) impairs vision of patients with type 1 and type 2 diabetes, associated with vascular dysfunction and occlusion, retinal edema, hemorrhage, and inappropriate growth of new blood vessels. The recent success of biologic treatments targeting vascular endothelial growth factor (VEGF) demonstrates that treating the vascular aspects in the later stages of the disease can preserve vision in many patients. It would also be highly desirable to prevent the onset of the disease or arrest its progression at a stage preceding the appearance of overt microvascular pathologies. The progression of DR is not necessarily linear but may follow a series of steps that evolve over the course of multiple years. Abundant data suggest that diabetes affects the entire neurovascular unit of the retina, with an early loss of neurovascular coupling, gradual neurodegeneration, gliosis, and neuroinflammation occurring before observable vascular pathologies. In this article, we consider the pathology of DR from the point of view that diabetes causes measurable dysfunctions in the complex integral network of cell types that produce and maintain human vision. © 2014 New York Academy of Sciences.

Diabetic retinopathy: loss of neuroretinal adaptation to the diabetic metabolic environment

PubMed Central

Abcouwer, Steven F.; Gardner, Thomas W.

2014-01-01

Diabetic retinopathy (DR) impairs vision of patients with type 1 and type 2 diabetes, associated with vascular dysfunction and occlusion, retinal edema, hemorrhage, and inappropriate growth of new blood vessels. The recent success of biologic treatments targeting vascular endothelial growth factor (VEGF) demonstrates that treating the vascular aspects in the later stages of the disease can preserve vision in many patients. It would also be highly desirable to prevent the onset of the disease or arrest its progression at a stage preceding the appearance of overt microvascular pathologies. The progression of DR is not necessarily linear but may follow a series of steps that evolve over the course of multiple years. Abundant data suggest that diabetes affects the entire neurovascular unit of the retina, with an early loss of neurovascular coupling, gradual neurodegeneration, gliosis, and neuroinflammation before observable vascular pathologies. In this article, we consider the pathology of diabetic retinopathy from the point of view that diabetes causes measurable dysfunctions in the complex integral network of cell types that produce and maintain human vision. PMID:24673341

Penile involvement in Systemic Sclerosis: New Diagnostic and Therapeutic Aspects

PubMed Central

Aversa, Antonio; Bruzziches, Roberto; Francomano, Davide; Rosato, Edoardo; Salsano, Felice; Spera, Giovanni

2010-01-01

Systemic Sclerosis (SSc) is a connective tissue disorder featuring vascular alterations and an immunological activation leading to a progressive and widespread fibrosis of several organs such as the skin, lung, gastrointestinal tract, heart, and kidney. Men with SSc are at increased risk of developing erectile dysfunction (ED) because of the evolution of early microvascular tissutal damage into corporeal fibrosis. The entity of penile vascular damage in SSc patients has been demonstrated by using Duplex ultrasonography and functional infra-red imaging and it is now clear that this is a true clinical entity invariably occurring irrespective of age and disease duration and constituting the ‘‘sclerodermic penis”. Once-daily phosphodiesterase type-5 (PDE5) inhibitors improve both sexual function and vascular measures of cavernous arteries by improving surrogate markers of endothelial dysfunction, that is, plasma endothelin-1 and adrenomedullin levels, which may play a potential role in preventing progression of penile fibrosis and ED. Also, the beneficial effect of long-term PDE5i add-on therapy to SSc therapy in the treatment of Raynaud's phenomenon is described. PMID:20981315

Endothelial Barrier Protection by Local Anesthetics: Ropivacaine and Lidocaine Block Tumor Necrosis Factor-α–induced Endothelial Cell Src Activation

PubMed Central

Piegeler, Tobias; Votta-Velis, E. Gina; Bakhshi, Farnaz R.; Mao, Mao; Carnegie, Graeme; Bonini, Marcelo G.; Schwartz, David E.; Borgeat, Alain; Beck-Schimmer, Beatrice; Minshall, Richard D.

2014-01-01

Background Pulmonary endothelial barrier dysfunction mediated in part by Src-kinase activation plays a crucial role in acute inflammatory disease. Proinflammatory cytokines, such as tumor necrosis factor-α (TNFα), activate Src via phosphatidylinositide 3-kinase/Akt-dependent nitric oxide generation, a process initiated by recruitment of phosphatidylinositide 3-kinase regulatory subunit p85 to TNF-receptor-1. Because amide-linked local anesthetics have well-established anti-inflammatory effects, the authors hypothesized that ropivacaine and lidocaine attenuate inflammatory Src signaling by disrupting the phosphatidylinositide 3-kinase–Akt–nitric oxide pathway, thus blocking Src-dependent neutrophil adhesion and endothelial hyperpermeability. Methods Human lung microvascular endothelial cells, incubated with TNFα in the absence or presence of clinically relevant concentrations of ropivacaine and lidocaine, were analyzed by Western blot, probing for phosphorylated/activated Src, endothelial nitric oxide synthase, Akt, intercellular adhesion molecule-1, and caveolin-1. The effect of ropivacaine on TNFα-induced nitric oxide generation, co-immunoprecipitation of TNF-receptor-1 with p85, neutrophil adhesion, and endothelial barrier disruption were assessed. Results Ropivacaine and lidocaine attenuated TNFα-induced Src activation (half-maximal inhibitory concentration [IC50] = 8.611 × 10−10 M for ropivacaine; IC50 = 5.864 × 10−10 M for lidocaine) and endothelial nitric oxide synthase phosphorylation (IC50 = 7.572 × 10−10 M for ropivacaine; IC50 = 6.377 × 10−10 M for lidocaine). Akt activation (n = 7; P = 0.006) and stimulus-dependent binding of TNF-receptor-1 and p85 (n = 6; P = 0.043) were blocked by 1 nM of ropivacaine. TNFα-induced neutrophil adhesion and disruption of endothelial monolayers via Src-dependent intercellular adhesion molecule-1- and caveolin-1-phosphorylation, respectively, were also attenuated. Conclusions Ropivacaine and lidocaine effectively blocked inflammatory TNFα signaling in endothelial cells by attenuating p85 recruitment to TNF-receptor-1. The resultant decrease in Akt, endothelial nitric oxide synthase, and Src phosphorylation reduced neutrophil adhesion and endothelial hyperpermeability. This novel anti-inflammatory “side-effect” of ropivacaine and lidocaine may provide therapeutic benefit in acute inflammatory disease. PMID:24525631

Endothelial barrier protection by local anesthetics: ropivacaine and lidocaine block tumor necrosis factor-α-induced endothelial cell Src activation.

PubMed

Piegeler, Tobias; Votta-Velis, E Gina; Bakhshi, Farnaz R; Mao, Mao; Carnegie, Graeme; Bonini, Marcelo G; Schwartz, David E; Borgeat, Alain; Beck-Schimmer, Beatrice; Minshall, Richard D

2014-06-01

Pulmonary endothelial barrier dysfunction mediated in part by Src-kinase activation plays a crucial role in acute inflammatory disease. Proinflammatory cytokines, such as tumor necrosis factor-α (TNFα), activate Src via phosphatidylinositide 3-kinase/Akt-dependent nitric oxide generation, a process initiated by recruitment of phosphatidylinositide 3-kinase regulatory subunit p85 to TNF-receptor-1. Because amide-linked local anesthetics have well-established anti-inflammatory effects, the authors hypothesized that ropivacaine and lidocaine attenuate inflammatory Src signaling by disrupting the phosphatidylinositide 3-kinase-Akt-nitric oxide pathway, thus blocking Src-dependent neutrophil adhesion and endothelial hyperpermeability. Human lung microvascular endothelial cells, incubated with TNFα in the absence or presence of clinically relevant concentrations of ropivacaine and lidocaine, were analyzed by Western blot, probing for phosphorylated/activated Src, endothelial nitric oxide synthase, Akt, intercellular adhesion molecule-1, and caveolin-1. The effect of ropivacaine on TNFα-induced nitric oxide generation, co-immunoprecipitation of TNF-receptor-1 with p85, neutrophil adhesion, and endothelial barrier disruption were assessed. Ropivacaine and lidocaine attenuated TNFα-induced Src activation (half-maximal inhibitory concentration [IC50] = 8.611 × 10 M for ropivacaine; IC50 = 5.864 × 10 M for lidocaine) and endothelial nitric oxide synthase phosphorylation (IC50 = 7.572 × 10 M for ropivacaine; IC50 = 6.377 × 10 M for lidocaine). Akt activation (n = 7; P = 0.006) and stimulus-dependent binding of TNF-receptor-1 and p85 (n = 6; P = 0.043) were blocked by 1 nM of ropivacaine. TNFα-induced neutrophil adhesion and disruption of endothelial monolayers via Src-dependent intercellular adhesion molecule-1- and caveolin-1-phosphorylation, respectively, were also attenuated. Ropivacaine and lidocaine effectively blocked inflammatory TNFα signaling in endothelial cells by attenuating p85 recruitment to TNF-receptor-1. The resultant decrease in Akt, endothelial nitric oxide synthase, and Src phosphorylation reduced neutrophil adhesion and endothelial hyperpermeability. This novel anti-inflammatory "side-effect" of ropivacaine and lidocaine may provide therapeutic benefit in acute inflammatory disease.

Regulation of Coronary Blood Flow in Health and Ischemic Heart Disease

PubMed Central

Duncker, Dirk J.; Koller, Akos; Merkus, Daphne; Canty, John M.

2018-01-01

The major factors determining myocardial perfusion and oxygen delivery have been elucidated over the past several decades, and this knowledge has been incorporated into the management of patients with ischemic heart disease (IHD). The basic understanding of the fluid mechanical behavior of coronary stenoses has also been translated to the cardiac catheterization laboratory where measurements of coronary pressure distal to a stenosis and coronary flow are routinely obtained. However, the role of perturbations in coronary microvascular structure and function, due to myocardial hypertrophy or coronary microvascular dysfunction, in IHD is becoming increasingly recognized. Future studies should therefore be aimed at further improving our understanding of the integrated coronary microvascular mechanisms that control coronary blood flow, and of the underlying causes and mechanisms of coronary microvascular dysfunction. This knowledge will be essential to further improve the treatment of patients with IHD. PMID:25475073

Microvascular Remodeling and Wound Healing: A Role for Pericytes

PubMed Central

Dulmovits, Brian M.; Herman, Ira M.

2012-01-01

Physiologic wound healing is highly dependent on the coordinated functions of vascular and non-vascular cells. Resolution of tissue injury involves coagulation, inflammation, formation of granulation tissue, remodeling and scarring. Angiogenesis, the growth of microvessels the size of capillaries, is crucial for these processes, delivering blood-borne cells, nutrients and oxygen to actively remodeling areas. Central to angiogenic induction and regulation is microvascular remodeling, which is dependent upon capillary endothelial cell and pericyte interactions. Despite our growing knowledge of pericyte-endothelial cell crosstalk, it is unclear how the interplay among pericytes, inflammatory cells, glia and connective tissue elements shape microvascular injury response. Here, we consider the relationships that pericytes form with the cellular effectors of healing in normal and diabetic environments, including repair following injury and vascular complications of diabetes, such as diabetic macular edema and proliferative diabetic retinopathy. In addition, pericytes and stem cells possessing “pericyte-like” characteristics are gaining considerable attention in experimental and clinical efforts aimed at promoting healing or eradicating ocular vascular proliferative disorders. As the origin, identification and characterization of microvascular pericyte progenitor populations remains somewhat ambiguous, the molecular markers, structural and functional characteristics of pericytes will be briefly reviewed. PMID:22750474

Inhaled nitric oxide pretreatment but not posttreatment attenuates ischemia-reperfusion-induced pulmonary microvascular leak.

PubMed

Chetham, P M; Sefton, W D; Bridges, J P; Stevens, T; McMurtry, I F

1997-04-01

Ischemia-reperfusion (I/R) pulmonary edema probably reflects a leukocyte-dependent, oxidant-mediated mechanism. Nitric oxide (NO) attenuates leukocyte-endothelial cell interactions and I/R-induced microvascular leak. Cyclic adenosine monophosphate (cAMP) agonists reverse and prevent I/R-induced microvascular leak, but reversal by inhaled NO (INO) has not been tested. In addition, the role of soluble guanylyl cyclase (sGC) activation in the NO protection effect is unknown. Rat lungs perfused with salt solution were grouped as either I/R, I/R with INO (10 or 50 ppm) on reperfusion, or time control. Capillary filtration coefficients (Kfc) were estimated 25 min before ischemia (baseline) and after 30 and 75 min of reperfusion. Perfusate cell counts and lung homogenate myeloperoxidase activity were determined in selected groups. Additional groups were treated with either INO (50 ppm) or isoproterenol (ISO-10 microM) after 30 min of reperfusion. Guanylyl cyclase was inhibited with 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one (ODQ-15 microM), and Kfc was estimated at baseline and after 30 min of reperfusion. (1) Inhaled NO attenuated I/R-induced increases in Kfc. (2) Cell counts were similar at baseline. After 75 min of reperfusion, lung neutrophil retention (myeloperoxidase activity) and decreased perfusate neutrophil counts were similar in all groups. (3) In contrast to ISO, INO did not reverse microvascular leak. (4) 8-bromoguanosine 3',5'-cyclic monophosphate (8-br-cGMP) prevented I/R-induced microvascular leak in ODQ-treated lungs, but INO was no longer effective. Inhaled NO attenuates I/R-induced pulmonary microvascular leak, which requires sGC activation and may involve a mechanism independent of inhibition of leukocyte-endothelial cell interactions. In addition, INO is ineffective in reversing I/R-induced microvascular leak.

Protective effect of Growth Hormone-Releasing Hormone agonist in bacterial toxin-induced pulmonary barrier dysfunction.

PubMed

Czikora, Istvan; Sridhar, Supriya; Gorshkov, Boris; Alieva, Irina B; Kasa, Anita; Gonzales, Joyce; Potapenko, Olena; Umapathy, Nagavedi S; Pillich, Helena; Rick, Ferenc G; Block, Norman L; Verin, Alexander D; Chakraborty, Trinad; Matthay, Michael A; Schally, Andrew V; Lucas, Rudolf

2014-01-01

Antibiotic treatment of patients infected with G(-) or G(+) bacteria promotes release of the toxins lipopolysaccharide (LPS) and pneumolysin (PLY) in their lungs. Growth Hormone-releasing Hormone (GHRH) agonist JI-34 protects human lung microvascular endothelial cells (HL-MVEC), expressing splice variant 1 (SV-1) of the receptor, from PLY-induced barrier dysfunction. We investigated whether JI-34 also blunts LPS-induced hyperpermeability. Since GHRH receptor (GHRH-R) signaling can potentially stimulate both cAMP-dependent barrier-protective pathways as well as barrier-disruptive protein kinase C pathways, we studied their interaction in GHRH agonist-treated HL-MVEC, in the presence of PLY, by means of siRNA-mediated protein kinase A (PKA) depletion. Barrier function measurements were done in HL-MVEC monolayers using Electrical Cell substrate Impedance Sensing (ECIS) and VE-cadherin expression by Western blotting. Capillary leak was assessed by Evans Blue dye (EBD) incorporation. Cytokine generation in broncho-alveolar lavage fluid (BALF) was measured by multiplex analysis. PKA and PKC-α activity were assessed by Western blotting. GHRH agonist JI-34 significantly blunts LPS-induced barrier dysfunction, at least in part by preserving VE-cadherin expression, while not affecting inflammation. In addition to activating PKA, GHRH agonist also increases PKC-α activity in PLY-treated HL-MVEC. Treatment with PLY significantly decreases resistance in control siRNA-treated HL-MVEC, but does so even more in PKA-depleted monolayers. Pretreatment with GHRH agonist blunts PLY-induced permeability in control siRNA-treated HL-MVEC, but fails to improve barrier function in PKA-depleted PLY-treated monolayers. GHRH signaling in HL-MVEC protects from both LPS and PLY-mediated endothelial barrier dysfunction and concurrently induces a barrier-protective PKA-mediated and a barrier-disruptive PKC-α-induced pathway in the presence of PLY, the former of which dominates the latter.

Integrin β4 Signaling Promotes Mammary Tumor Cell Adhesion to Brain Microvascular Endothelium by Inducing ErbB2-mediated Secretion of VEGF

PubMed Central

Fan, Jie; Cai, Bin; Zeng, Min; Hao, Yanyan

2015-01-01

Prior studies have indicated that the β4 integrin promotes mammary tumor invasion and metastasis by combining with ErbB2 and amplifying its signaling capacity. However, the effector pathways and cellular functions by which the β4 integrin exerts these effects are incompletely understood. To examine if β4 signaling plays a role during mammary tumor cell adhesion to microvascular endothelium, we have examined ErbB2-transformed mammary tumor cells expressing either a wild-type (WT) or a signaling-defective form of β4 (1355T). We report that WT cells adhere to brain microvascular endothelium in vitro to a significantly larger extent as compared to 1355T cells. Interestingly, integrin β4 signaling does not exert a direct effect on adhesion to the endothelium or the underlying basement membrane. Rather, it enhances ErbB2-dependent expression of VEGF by tumor cells. VEGF in turn disrupts the tight and adherens junctions of endothelial monolayers, enabling the exposure of underlying basement membrane and increasing the adhesion of tumor cells to the intercellular junctions of endothelium. Inhibition of ErbB2 on tumor cells or the VEGFR-2 on endothelial cells suppresses mammary tumor cell adhesion to microvascular endothelium. Our results indicate that β4 signaling regulates VEGF expression by the mammary tumor cells thereby enhancing their adhesion to microvascular endothelium. PMID:21556948

Doxycycline Attenuates Lipopolysaccharide-Induced Microvascular Endothelial Cell Derangements.

PubMed

Wiggins-Dohlvik, Katie; Stagg, Hayden W; Han, Min Suk; Alluri, Himakarnika; Oakley, Ryan P; Anasooya Shaji, Chinchusha; Davis, Matthew L; Tharakan, Binu

2016-06-01

Lipopolysaccharide (LPS) is known to induce vascular derangements. The pathophysiology involved therein is unknown, but matrix metalloproteinases (MMPs) may be an important mediator. We hypothesized that in vitro LPS provokes vascular permeability, damages endothelial structural proteins, and increases MMP activity; that in vivo LPS increases permeability and fluid requirements; and that the MMP inhibitor doxycycline mitigates such changes. Rat lung microvascular endothelial cells were divided into four groups: control, LPS, LPS plus doxycycline, and doxycycline. Permeability, structural proteins β-catenin and Filamentous-actin, and MMP-9 activity were examined. Sprauge Dawley rats were divided into sham, IV LPS, and IV LPS plus IV doxycycline groups. Mesenteric postcapillary venules were observed. Blood pressure was measured as animals were resuscitated and fluid requirements were compared. Statistical analysis was conducted using Student's t-test and ANOVA. In vitro LPS increased permeability, damaged adherens junctions, induced actin stress fiber formation, and increased MMP-9 enzyme activity. In vivo, IV LPS administration induced vascular permeability. During resuscitation, significantly more fluid was necessary to maintain normotension in the IV LPS group. Doxycycline mitigated all derangements observed. We conclude that LPS increases permeability, damages structural proteins, and increases MMP-9 activity in endothelial cells. Additionally, endotoxemia induces hyperpermeability and increases the amount of IV fluid required to maintain normotension in vivo. Doxycycline mitigates such changes both in vitro and in vivo. Our findings illuminate the possible role of matrix metalloproteinases in the pathophysiology of lipopolysaccharide-induced microvascular hyperpermeability and pave the way for better understanding and treatment of this process.

Cyclic adenosine monophosphate levels and the function of skin microvascular endothelial cells.

PubMed

Tuder, R M; Karasek, M A; Bensch, K G

1990-02-01

The maintenance of the normal epithelioid morphology of human dermal microvascular endothelial cells (MEC) grown in vitro depends strongly on the presence of factors that increase intracellular levels of cyclic AMP. Complete removal of dibutyryl cAMP and isobutylmethylxanthine (IMX) from the growth medium results in a progressive transition from an epithelioid to a spindle-shaped cell line. This transition cannot be reversed by the readdition of dibutyryl cAMP and IMX to the growth medium or by addition of agonists that increase cAMP levels. Spindle-shaped MEC lose the ability to express Factor VIII rAG and DR antigens and to bind peripheral blood mononuclear leukocyte (PBML). Ultrastructural analyses of transitional cells and spindle-shaped cells show decreased numbers of Weibel-Palade bodies in transitional cells and their complete absence in spindle-shaped cells. Interferon-gamma alters several functional properties of both epithelioid and spindle-shaped cells. In the absence of dibutyryl cAMP it accelerates the transition from epithelial to spindle-shaped cells, whereas in the presence of cyclic AMP interferon-gamma increases the binding of PBMLs to both epithelioid and spindle-shaped MEC and the endocytic activity of the endothelial cells. These results suggest that cyclic AMP is an important second messenger in the maintenance of several key functions of microvascular endothelial cells. Factors that influence the levels of this messenger in vivo can be expected to influence the angiogenic and immunologic functions of the microvasculature.

Brain Invasion by Mouse Hepatitis Virus Depends on Impairment of Tight Junctions and Beta Interferon Production in Brain Microvascular Endothelial Cells

PubMed Central

Bleau, Christian; Filliol, Aveline; Samson, Michel

2015-01-01

ABSTRACT Coronaviruses (CoVs) have shown neuroinvasive properties in humans and animals secondary to replication in peripheral organs, but the mechanism of neuroinvasion is unknown. The major aim of our work was to evaluate the ability of CoVs to enter the central nervous system (CNS) through the blood-brain barrier (BBB). Using the highly hepatotropic mouse hepatitis virus type 3 (MHV3), its attenuated variant, 51.6-MHV3, which shows low tropism for endothelial cells, and the weakly hepatotropic MHV-A59 strain from the murine coronavirus group, we investigated the virus-induced dysfunctions of BBB in vivo and in brain microvascular endothelial cells (BMECs) in vitro. We report here a MHV strain-specific ability to cross the BBB during acute infection according to their virulence for liver. Brain invasion was observed only in MHV3-infected mice and correlated with enhanced BBB permeability associated with decreased expression of zona occludens protein 1 (ZO-1), VE-cadherin, and occludin, but not claudin-5, in the brain or in cultured BMECs. BBB breakdown in MHV3 infection was not related to production of barrier-dysregulating inflammatory cytokines or chemokines by infected BMECs but rather to a downregulation of barrier protective beta interferon (IFN-β) production. Our findings highlight the importance of IFN-β production by infected BMECs in preserving BBB function and preventing access of blood-borne infectious viruses to the brain. IMPORTANCE Coronaviruses (CoVs) infect several mammals, including humans, and are associated with respiratory, gastrointestinal, and/or neurological diseases. There is some evidence that suggest that human respiratory CoVs may show neuroinvasive properties. Indeed, the severe acute respiratory syndrome coronavirus (SARS-CoV), causing severe acute respiratory syndrome, and the CoVs OC43 and 229E were found in the brains of SARS patients and multiple sclerosis patients, respectively. These findings suggest that hematogenously spread CoVs may gain access to the CNS at the BBB level. Herein we report for the first time that CoVs exhibit the ability to cross the BBB according to strain virulence. BBB invasion by CoVs correlates with virus-induced disruption of tight junctions on BMECs, leading to BBB dysfunction and enhanced permeability. We provide evidence that production of IFN-β by BMECs during CoV infection may prevent BBB breakdown and brain viral invasion. PMID:26202229

Brain Invasion by Mouse Hepatitis Virus Depends on Impairment of Tight Junctions and Beta Interferon Production in Brain Microvascular Endothelial Cells.

PubMed

Bleau, Christian; Filliol, Aveline; Samson, Michel; Lamontagne, Lucie

2015-10-01

Coronaviruses (CoVs) have shown neuroinvasive properties in humans and animals secondary to replication in peripheral organs, but the mechanism of neuroinvasion is unknown. The major aim of our work was to evaluate the ability of CoVs to enter the central nervous system (CNS) through the blood-brain barrier (BBB). Using the highly hepatotropic mouse hepatitis virus type 3 (MHV3), its attenuated variant, 51.6-MHV3, which shows low tropism for endothelial cells, and the weakly hepatotropic MHV-A59 strain from the murine coronavirus group, we investigated the virus-induced dysfunctions of BBB in vivo and in brain microvascular endothelial cells (BMECs) in vitro. We report here a MHV strain-specific ability to cross the BBB during acute infection according to their virulence for liver. Brain invasion was observed only in MHV3-infected mice and correlated with enhanced BBB permeability associated with decreased expression of zona occludens protein 1 (ZO-1), VE-cadherin, and occludin, but not claudin-5, in the brain or in cultured BMECs. BBB breakdown in MHV3 infection was not related to production of barrier-dysregulating inflammatory cytokines or chemokines by infected BMECs but rather to a downregulation of barrier protective beta interferon (IFN-β) production. Our findings highlight the importance of IFN-β production by infected BMECs in preserving BBB function and preventing access of blood-borne infectious viruses to the brain. Coronaviruses (CoVs) infect several mammals, including humans, and are associated with respiratory, gastrointestinal, and/or neurological diseases. There is some evidence that suggest that human respiratory CoVs may show neuroinvasive properties. Indeed, the severe acute respiratory syndrome coronavirus (SARS-CoV), causing severe acute respiratory syndrome, and the CoVs OC43 and 229E were found in the brains of SARS patients and multiple sclerosis patients, respectively. These findings suggest that hematogenously spread CoVs may gain access to the CNS at the BBB level. Herein we report for the first time that CoVs exhibit the ability to cross the BBB according to strain virulence. BBB invasion by CoVs correlates with virus-induced disruption of tight junctions on BMECs, leading to BBB dysfunction and enhanced permeability. We provide evidence that production of IFN-β by BMECs during CoV infection may prevent BBB breakdown and brain viral invasion. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Approaches to improve angiogenesis in tissue-engineered skin.

PubMed

Sahota, Parbinder S; Burn, J Lance; Brown, Nicola J; MacNeil, Sheila

2004-01-01

A problem with tissue-engineered skin is clinical failure due to delays in vascularization. The aim of this study was to explore a number of simple strategies to improve angiogenesis/vascularization using a tissue-engineered model of skin to which small vessel human dermal microvascular endothelial cells were added. For the majority of these studies, a modified Guirguis chamber was used, which allowed the investigation of several variables within the same experiment using the same human dermis; cell type, angiogenic growth factors, the influence of keratinocytes and fibroblasts, mechanical penetration of the human dermis, the site of endothelial cell addition, and the influence of hypoxia were all examined. A qualitative scoring system was used to assess the impact of these factors on the penetration of endothelial cells throughout the dermis. Similar results were achieved using freshly isolated small vessel human dermal microvascular endothelial cells or an endothelial cell line and a minimum cell seeding density was identified. Cell penetration was not influenced by the addition of angiogenic growth factors (vascular endothelial growth factor and basic fibroblast growth factor); similarly, including epidermal keratinocytes or dermal fibroblasts did not encourage endothelial cell entry, and neither did mechanical introduction of holes throughout the dermis. Two factors were identified that significantly enhanced endothelial cell penetration into the dermis: hypoxia and the site of endothelial cell addition. Endothelial cells added from the papillary surface entered into the dermis much more effectively than when cells were added to the reticular surface of the dermis. We conclude that this model is valuable in improving our understanding of how to enhance vascularization of tissue-engineered grafts.

The association between diabetes and dermal microvascular dysfunction non-invasively assessed by laser Doppler with local thermal hyperemia: a systematic review with meta-analysis.

PubMed

Fuchs, Dagmar; Dupon, Pepijn P; Schaap, Laura A; Draijer, Richard

2017-01-19

Diabetes and cardiovascular disease develop in concert with metabolic abnormalities mirroring and causing changes in the vasculature, particularly the microcirculation. The microcirculation can be affected in different parts of the body of which the skin is the most easily accessible tissue. The association between diabetes and dermal microvascular dysfunction has been investigated in observational studies. However, the strength of the association is unknown. Therefore we conducted a systematic review with meta-analysis on the association between diabetes and dermal microvascular dysfunction as assessed by laser Doppler/laser speckle contrast imaging with local thermal hyperaemia as non-invasive indicator of microvascular functionality. PubMed and Ovid were  systematically searched for eligible studies through March 2015. During the first selection, studies were included if they were performed in humans and were related to diabetes or glucose metabolism disorders and to dermal microcirculation. During the second step we selected studies based on the measurement technique, measurement location (arm or leg) and the inclusion of a healthy control group. A random effects model was used with the standardised mean difference as outcome measure. Calculations and imputation of data were done according to the Cochrane Handbook. Of the 1445 studies found in the first search, thirteen cross-sectional studies were included in the meta-analysis, comprising a total of 857 subjects. Resting blood flow was similar between healthy control subjects and diabetes patients. In contrast, the microvascular response to local skin heating was reduced in diabetic patients compared to healthy control subjects [pooled effect of -0.78 standardised mean difference (95% CI -1.06, -0.51)]. This effect is considered large according to Cohen's effect size definition. The variability in effect size was high (heterogeneity 69%, p < 0.0001). However, subgroup analysis revealed no difference between the type and duration of diabetes and other health related factors, indicating that diabetes per se causes the microvascular dysfunction. Our meta-analysis shows that diabetes is associated with a large reduction of dermal microvascular function in diabetic patients. The local thermal hyperaemia methodology may become a valuable non-invasive tool for diagnosis and assessing progress of diabetes-related microvascular complications, but standardisation of the technique and quality of study conduct is urgently required.

Endothelial dysfunction is associated with activation of the type I interferon system and platelets in patients with systemic lupus erythematosus

PubMed Central

Tydén, Helena; Lood, Christian; Gullstrand, Birgitta; Nielsen, Christoffer Tandrup; Heegaard, Niels H H; Kahn, Robin; Jönsen, Andreas; Bengtsson, Anders A

2017-01-01

Objectives Endothelial dysfunction may be connected to cardiovascular disease (CVD) in systemic lupus erythematosus (SLE). Type I interferons (IFNs) are central in SLE pathogenesis and are suggested to induce both endothelial dysfunction and platelet activation. In this study, we investigated the interplay between endothelial dysfunction, platelets and type I IFN in SLE. Methods We enrolled 148 patients with SLE and 79 sex-matched and age-matched healthy controls (HCs). Type I IFN activity was assessed with a reporter cell assay and platelet activation by flow cytometry. Endothelial dysfunction was assessed using surrogate markers of endothelial activation, soluble vascular cell adhesion molecule-1 (sVCAM-1) and endothelial microparticles (EMPs), and finger plethysmograph to determine Reactive Hyperaemia Index (RHI). Results In patients with SLE, type I IFN activity was associated with endothelial activation, measured by high sVCAM-1 (OR 1.68, p<0.01) and elevated EMPs (OR 1.40, p=0.03). Patients with SLE with high type I IFN activity had lower RHI than HCs (OR 2.61, p=0.04), indicating endothelial dysfunction. Deposition of complement factors on platelets, a measure of platelet activation, was seen in patients with endothelial dysfunction. High levels of sVCAM-1 were associated with increased deposition of C4d (OR 4.57, p<0.01) and C1q (OR 4.10, p=0.04) on platelets. High levels of EMPs were associated with C4d deposition on platelets (OR 3.64, p=0.03). Conclusions Endothelial dysfunction was associated with activation of platelets and the type I IFN system. We suggest that an interplay between the type I IFN system, injured endothelium and activated platelets may contribute to development of CVD in SLE. PMID:29119007

Effect of shear stress on iPSC-derived human brain microvascular endothelial cells (dhBMECs).

PubMed

DeStefano, Jackson G; Xu, Zinnia S; Williams, Ashley J; Yimam, Nahom; Searson, Peter C

2017-08-04

The endothelial cells that form the lumen of capillaries and microvessels are an important component of the blood-brain barrier. Cell phenotype is regulated by transducing a range of biomechanical and biochemical signals in the local microenvironment. Here we report on the role of shear stress in modulating the morphology, motility, proliferation, apoptosis, and protein and gene expression, of confluent monolayers of human brain microvascular endothelial cells derived from induced pluripotent stem cells. To assess the response of derived human brain microvascular endothelial cells (dhBMECs) to shear stress, confluent monolayers were formed in a microfluidic device. Monolayers were subjected to a shear stress of 4 or 12 dyne cm -2 for 40 h. Static conditions were used as the control. Live cell imaging was used to assess cell morphology, cell speed, persistence, and the rates of proliferation and apoptosis as a function of time. In addition, immunofluorescence imaging and protein and gene expression analysis of key markers of the blood-brain barrier were performed. Human brain microvascular endothelial cells exhibit a unique phenotype in response to shear stress compared to static conditions: (1) they do not elongate and align, (2) the rates of proliferation and apoptosis decrease significantly, (3) the mean displacement of individual cells within the monolayer over time is significantly decreased, (4) there is no cytoskeletal reorganization or formation of stress fibers within the cell, and (5) there is no change in expression levels of key blood-brain barrier markers. The characteristic response of dhBMECs to shear stress is significantly different from human and animal-derived endothelial cells from other tissues, suggesting that this unique phenotype that may be important in maintenance of the blood-brain barrier. The implications of this work are that: (1) in confluent monolayers of dhBMECs, tight junctions are formed under static conditions, (2) the formation of tight junctions decreases cell motility and prevents any morphological transitions, (3) flow serves to increase the contact area between cells, resulting in very low cell displacement in the monolayer, (4) since tight junctions are already formed under static conditions, increasing the contact area between cells does not cause upregulation in protein and gene expression of BBB markers, and (5) the increase in contact area induced by flow makes barrier function more robust.

Blood-brain barrier KCa3.1 channels: evidence for a role in brain Na uptake and edema in ischemic stroke.

PubMed

Chen, Yi-Je; Wallace, Breanna K; Yuen, Natalie; Jenkins, David P; Wulff, Heike; O'Donnell, Martha E

2015-01-01

KCa3.1, a calcium-activated potassium channel, regulates ion and fluid secretion in the lung and gastrointestinal tract. It is also expressed on vascular endothelium where it participates in blood pressure regulation. However, the expression and physiological role of KCa3.1 in blood-brain barrier (BBB) endothelium has not been investigated. BBB endothelial cells transport Na(+) and Cl(-) from the blood into the brain transcellularly through the co-operation of multiple cotransporters, exchangers, pumps, and channels. In the early stages of cerebral ischemia, when the BBB is intact, edema formation occurs by processes involving increased BBB transcellular Na(+) transport. This study evaluated whether KCa3.1 is expressed on and participates in BBB ion transport. The expression of KCa3.1 on cultured cerebral microvascular endothelial cells, isolated microvessels, and brain sections was evaluated by Western blot and immunohistochemistry. Activity of KCa3.1 on cerebral microvascular endothelial cells was examined by K(+) flux assays and patch-clamp. Magnetic resonance spectroscopy and MRI were used to measure brain Na(+) uptake and edema formation in rats with focal ischemic stroke after TRAM-34 treatment. KCa3.1 current and channel protein were identified on bovine cerebral microvascular endothelial cells and freshly isolated rat microvessels. In situ KCa3.1 expression on BBB endothelium was confirmed in rat and human brain sections. TRAM-34 treatment significantly reduced Na(+) uptake, and cytotoxic edema in the ischemic brain. BBB endothelial cells exhibit KCa3.1 protein and activity and pharmacological blockade of KCa3.1 seems to provide an effective therapeutic approach for reducing cerebral edema formation in the first 3 hours of ischemic stroke. © 2014 American Heart Association, Inc.

Apigenin and naringenin ameliorate PKCβII-associated endothelial dysfunction via regulating ROS/caspase-3 and NO pathway in endothelial cells exposed to high glucose.

PubMed

Qin, Weiwei; Ren, Bei; Wang, Shanshan; Liang, Shujun; He, Baiqiu; Shi, Xiaoji; Wang, Liying; Liang, Jingyu; Wu, Feihua

2016-10-01

Endothelial dysfunction is a key event in the progression of atherosclerosis with diabetes. Increasing cell apoptosis may lead to endothelial dysfunction. Apigenin and naringenin are two kinds of widely used flavones. In the present study, we investigated whether and how apigenin and naringenin reduced endothelial dysfunction induced by high glucose in endothelial cells. We showed that apigenin and naringenin protected against endothelial dysfunction via inhibiting phosphorylation of protein kinase C βII (PKCβII) expression and downstream reactive oxygen species (ROS) production in endothelial cells exposed to high glucose. Furthermore, we demonstrated that apigenin and naringenin reduced high glucose-increased apoptosis, Bax expression, caspase-3 activity and phosphorylation of NF-κB in endothelial cells. Moreover, apigenin and naringenin effectively restored high glucose-reduced Bcl-2 expression and Akt phosphorylation. Importantly, apigenin and naringenin significantly increased NO production in endothelial cells subjected to high glucose challenge. Consistently, high glucose stimulation impaired acetylcholine (ACh)-mediated vasodilation in the rat aorta, apigenin and naringenin treatment restored the impaired endothelium-dependent vasodilation via dramatically increasing eNOS activity and nitric oxide (NO) level. Taken together, our results manifest that apigenin and naringenin can ameliorate endothelial dysfunction via regulating ROS/caspase-3 and NO pathway. Copyright © 2016 Elsevier Inc. All rights reserved.

Strategies to reverse endothelial progenitor cell dysfunction in diabetes.

PubMed

Petrelli, Alessandra; Di Fenza, Raffaele; Carvello, Michele; Gatti, Francesca; Secchi, Antonio; Fiorina, Paolo

2012-01-01

Bone-marrow-derived cells-mediated postnatal vasculogenesis has been reported as the main responsible for the regulation of vascular homeostasis in adults. Since their discovery, endothelial progenitor cells have been depicted as mediators of postnatal vasculogenesis for their peculiar phenotype (partially staminal and partially endothelial), their ability to differentiate in endothelial cell line and to be incorporated into the vessels wall during ischemia/damage. Diabetes mellitus, a condition characterized by cardiovascular disease, nephropathy, and micro- and macroangiopathy, showed a dysfunction of endothelial progenitor cells. Herein, we review the mechanisms involved in diabetes-related dysfunction of endothelial progenitor cells, highlighting how hyperglycemia affects the different steps of endothelial progenitor cells lifetime (i.e., bone marrow mobilization, trafficking into the bloodstream, differentiation in endothelial cells, and homing in damaged tissues/organs). Finally, we review preclinical and clinical strategies that aim to revert diabetes-induced dysfunction of endothelial progenitor cells as a means of finding new strategies to prevent diabetic complications.

Fabrication of a reticular poly(lactide-co-glycolide) cylindrical scaffold for the in vitro development of microvascular networks

NASA Astrophysics Data System (ADS)

Tung, Yen-Ting; Chang, Cheng-Chung; Ju, Jyh-Cherng; Wang, Gou-Jen

2017-12-01

The microvascular network is a simple but critical system that is responsible for a range of important biological mechanisms in the bodies of all animals. The ability to generate a functional microvessel not only makes it possible to engineer vital tissue of considerable size but also serves as a platform for biomedical studies. However, most of the current methods for generating microvessel networks in vitro use rectangular channels which cannot represent real vessels in vivo and have dead zones at their corners, hence hindering the circulation of culture medium. We propose a scaffold-wrapping method which enables fabrication of a customized microvascular network in vitro in a more biomimetic way. By integrating microelectromechanical techniques with thermal reflow, we designed and fabricated a microscale hemi-cylindrical photoresist template. A replica mold of polydimethylsiloxane, produced by casting, was then used to generate cylindrical scaffolds with biodegradable poly(lactide-co-glycolide) (PLGA). Human umbilical vein endothelial cells were seeded on both sides of the PLGA scaffold and cultured using a traditional approach. The expression of endothelial cell marker CD31 and intercellular junction vascular endothelial cadherin on the cultured cell demonstrated the potential of generating a microvascular network with a degradable cylindrical scaffold. Our method allows cells to be cultured on a scaffold using a conventional culture approach and monitors cell conditions continuously. We hope our cell-covered scaffold can serve as a framework for building large tissues or can be used as the core of a vascular chip for in vitro circulation studies.

Toll-like receptor 4-induced endoplasmic reticulum stress contributes to endothelial dysfunction

USDA-ARS?s Scientific Manuscript database

Impairment of vasodilator action of insulin is associated with endothelial dysfunction and insulin resistance. Endoplasmic reticulum (ER) stress is implicated as one of the mechanisms for pathophysiology of various cardiometabolic syndromes, including insulin resistance and endothelial dysfunction. ...

S-nitrosylation of VASP at cysteine 64 mediates the inflammation-stimulated increase in microvascular permeability.

PubMed

Zamorano, Patricia; Marín, Natalie; Córdova, Francisco; Aguilar, Alejandra; Meininger, Cynthia; Boric, Mauricio P; Golenhofen, Nikola; Contreras, Jorge E; Sarmiento, José; Durán, Walter N; Sánchez, Fabiola A

2017-07-01

We tested the hypothesis that platelet-activating factor (PAF) induces S -nitrosylation of vasodilator-stimulated phosphoprotein (VASP) as a mechanism to reduce microvascular endothelial barrier integrity and stimulate hyperpermeability. PAF elevated S -nitrosylation of VASP above baseline levels in different endothelial cells and caused hyperpermeability. To ascertain the importance of endothelial nitric oxide synthase (eNOS) subcellular location in this process, we used ECV-304 cells transfected with cytosolic eNOS (GFPeNOSG2A) and plasma membrane eNOS (GFPeNOSCAAX). PAF induced S -nitrosylation of VASP in cells with cytosolic eNOS but not in cells wherein eNOS is anchored to the cell membrane. Reconstitution of VASP knockout myocardial endothelial cells with cysteine mutants of VASP demonstrated that S -nitrosylation of cysteine 64 is associated with PAF-induced hyperpermeability. We propose that regulation of VASP contributes to endothelial cell barrier integrity and to the onset of hyperpermeability. S -nitrosylation of VASP inhibits its function in barrier integrity and leads to endothelial monolayer hyperpermeability in response to PAF, a representative proinflammatory agonist. NEW & NOTEWORTHY Here, we demonstrate that S -nitrosylation of vasodilator-stimulated phosphoprotein (VASP) on C64 is a mechanism for the onset of platelet-activating factor-induced hyperpermeability. Our results reveal a dual role of VASP in endothelial permeability. In addition to its well-documented function in barrier integrity, we show that S -nitrosylation of VASP contributes to the onset of endothelial permeability. Copyright © 2017 the American Physiological Society.

Escherichia coli K1 invasion increases human brain microvascular endothelial cell monolayer permeability by disassembling vascular-endothelial cadherins at tight junctions.

PubMed

Sukumaran, Sunil K; Prasadarao, Nemani V

2003-11-01

We investigated the permeability changes that occur in the human brain microvascular endothelial cell (HBMEC) monolayer, an in vitro model of the blood-brain barrier, during Escherichia coli K1 infection. An increase in permeability of HBMECs and a decrease in transendothelial electrical resistance were observed. These permeability changes occurred only when HBMECs were infected with E. coli expressing outer membrane protein A (OmpA) and preceded the traversal of bacteria across the monolayer. Activated protein kinase C (PKC)-alpha interacts with vascular-endothelial cadherins (VECs) at the tight junctions of HBMECs, resulting in the dissociation of beta-catenins from VECs and leading to the increased permeability of the HBMEC monolayer. Overexpression of a dominant negative form of PKC-alpha in HBMECs blocked the E. coli-induced increase in permeability of HBMECs. Anti-OmpA and anti-OmpA receptor antibodies exerted inhibition of E. coli-induced permeability of HBMEC monolayers. This inhibition was the result of the absence of PKC-alpha activation in HBMECs treated with the antibodies.

Comparative study between microvascular tone regulation and rheological properties of blood in patients with type 2 diabetes mellitus.

PubMed

Antonova, N; Tsiberkin, K; Podtaev, S; Paskova, V; Velcheva, I; Chaushev, N

2016-01-01

The aim of the study is to investigate the changes of the skin blood flow responses to cold stress in patients with diabetes mellitus type 2 through wavelet analysis of the peripheral skin temperature oscillations and to estimate their relationship with the blood viscosity values. The amplitudes of the skin temperature pulsations (ASTP) were monitored by "Microtest" device ("FM-Diagnostics", Russia); the whole blood viscosity and the shear stresses were measured by Contraves LS30 viscometer (Switzerland) at a steady flow in 9 healthy subjects and in 30 patients with type 2 diabetes mellitus. Power law and Herschel-Bulkley (HB) equations were applied to describe the blood rheology. Both models include consistency (k) and flow index (m), and the HB also gives the yield stress (τ0). The Spearman rank correlations between these parameters and the ASTP in the frequency ranges, corresponding to the myogenic, neurogenic and endothelial mechanisms of the microcirculation tone regulation were calculated. The ASTP values decreased when the blood viscosity increased. The correlation analysis revealed good ASTP-m (r > 0.5) and ASTP-k (r < -0.5) relationships in the endothelial range, while the ASTP-τ0 correlation was weaker (r≈-0.4). These correlations became lower for the ASTP during the cold stress. The results prompt manifestation of endothelial dysfunction in patients with type 2 diabetes.

Vaspin protects against LPS-induced ARDS by inhibiting inflammation, apoptosis and reactive oxygen species generation in pulmonary endothelial cells via the Akt/GSK-3β pathway

PubMed Central

Qi, Di; Wang, Daoxin; Zhang, Chunrong; Tang, Xumao; He, Jing; Zhao, Yan; Deng, Wang; Deng, Xinyu

2017-01-01

Acute respiratory distress syndrome (ARDS) is characterized by uncontrolled extravasation of protein-rich fluids, which is caused by disruption and dysfunction of the barrier of pulmonary endothelial cells (ECs). Visceral adipose tissue-derived serine protease inhibitor (vaspin) is a novel adipokine with pleiotropic properties, which has been reported to exert beneficial effects against obesity-associated systemic vascular diseases; however, its effects on ARDS remain unknown. In the present study, mice were subjected to systemic administration of adenoviral vector expressing vaspin (Ad-vaspin) to examine its effects on lipopolysaccharide (LPS)-induced ARDS in vivo. Histological analysis was then conducted, and cytokine [tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-10] levels, and intercellular cell adhesion molecule-1 (ICAM-1) and adherens junctions (AJs) expression were detected. In addition, human pulmonary microvascular ECs (HPMECs) were treated with recombinant human (rh)-vaspin to further investigate its molecular basis and underlying mechanism. The mRNA expression levels of inflammatory cytokines (TNF-α and IL-6) and endothelial-specific adhesion markers [vascular cell adhesion molecule-1 and E-selectin], activation of nuclear factor-κB, and cell viability and apoptosis were then examined. Furthermore, the expression of AJs and organization of the cytoskeleton, as well as expression and activity of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and generation of reactive oxygen species (ROS) were determined. The results indicated that Ad-vaspin protected against LPS-induced ARDS by alleviating the pulmonary inflammatory response and pulmonary EC barrier dysfunction in mice, which was accompanied by activation of the protein kinase B (Akt)/glycogen synthase kinase (GSK)-3β pathway. In addition, pretreatment of HPMECs with rh-vaspin attenuated inflammation, apoptosis and ROS generation without alterations in AJs and cytoskeletal organization following LPS insult, which was accompanied by activation of the Akt/GSK3β pathway. In conclusion, the present study demonstrated that vaspin protects against LPS-induced ARDS by reversing EC barrier dysfunction via the suppression of inflammation, apoptosis and ROS production in pulmonary ECs, at least partially via activation of the Akt/GSK3β pathway. These findings provide evidence of a causal link between vaspin and EC dysfunction in ARDS, and suggest a potential therapeutic intervention for patients with ARDS. PMID:29039444

CXCL4L1 and CXCL4 signaling in human lymphatic and microvascular endothelial cells and activated lymphocytes: involvement of mitogen-activated protein (MAP) kinases, Src and p70S6 kinase.

PubMed

Van Raemdonck, Katrien; Gouwy, Mieke; Lepers, Stefanie Antoinette; Van Damme, Jo; Struyf, Sofie

2014-07-01

CXC chemokines influence a variety of biological processes, such as angiogenesis, both in a physiological and pathological context. Platelet factor-4 (PF-4)/CXCL4 and its variant PF-4var/CXCL4L1 are known to favor angiostasis by inhibiting endothelial cell proliferation and chemotaxis. CXCL4L1 in particular is a potent inhibitor of angiogenesis with anti-tumoral characteristics, both through regulation of neovascularization and through attraction of activated lymphocytes. However, its underlying signaling pathways remain to be elucidated. Here, we have identified various intracellular pathways activated by CXCL4L1 in comparison with other CXCR3 ligands, including CXCL4 and interferon-γ-induced protein 10/CXCL10. Signaling experiments show involvement of the mitogen-activated protein kinase (MAPK) family in CXCR3A-transfected cells, activated lymphocytes and human microvascular endothelial cells (HMVEC). In CXCR3A transfectants, CXCL4 and CXCL4L1 activated p38 MAPK, as well as Src kinase within 30 and 5 min, respectively. Extracellular signal-regulated kinase (ERK) phosphorylation occurred in activated lymphocytes, yet was inhibited in microvascular and lymphatic endothelial cells. CXCL4L1 and CXCL4 counterbalanced the angiogenic chemokine stromal cell-derived factor-1/CXCL12 in both endothelial cell types. Notably, inhibition of ERK signaling by CXCL4L1 and CXCL4 in lymphatic endothelial cells implies that these chemokines might also regulate lymphangiogenesis. Furthermore, CXCL4, CXCL4L1 and CXCL10 slightly enhanced forskolin-stimulated cAMP production in HMVEC. Finally, CXCL4, but not CXCL4L1, induced activation of p70S6 kinase within 5 min in HMVEC. Our findings confirm that the angiostatic chemokines CXCL4L1 and CXCL4 activate both CXCR3A and CXCR3B and bring new insights into the complexity of their signaling cascades.

Associations of low grade inflammation and endothelial dysfunction with depression - The Maastricht Study.

PubMed

van Dooren, Fleur E P; Schram, Miranda T; Schalkwijk, Casper G; Stehouwer, Coen D A; Henry, Ronald M A; Dagnelie, Pieter C; Schaper, Nicolaas C; van der Kallen, Carla J H; Koster, Annemarie; Sep, Simone J S; Denollet, Johan; Verhey, Frans R J; Pouwer, Frans

2016-08-01

The pathogenesis of depression may involve low-grade inflammation and endothelial dysfunction. We aimed to evaluate the independent associations of inflammation and endothelial dysfunction with depressive symptoms and depressive disorder, and the role of lifestyle factors in this association. In The Maastricht Study, a population-based cohort study (n=852, 55% men, m=59.8±8.5years), depressive symptoms were assessed with the Patient Health Questionnaire-9 and (major and minor) depressive disorder with the Mini-International Neuropsychiatric Interview. Plasma biomarkers of inflammation (hsCRP, SAA, sICAM-1, IL-6, IL-8, TNF-α) and endothelial dysfunction (sVCAM-1, sICAM-1, sE-selectin, vWF) were measured with sandwich immunoassays and combined into two standardized sum scores. Biomarkers of inflammation (hsCRP, TNF-α, SAA, sICAM-1) and endothelial dysfunction (sICAM-1, sE-Selectin) were univariately associated with depressive symptoms and depressive disorder. The sum scores of inflammation and endothelial dysfunction were associated with depressive disorder after adjustment for age, sex, type 2 diabetes, kidney function and prior cardiovascular disease (OR 1.54, p=0.001 and 1.40, p=0.006). Both sum scores remained significantly associated with depressive disorder after additional adjustment for lifestyle factors smoking, alcohol consumption and body mass index. The sum score of inflammation was also independently associated with depressive symptoms, while the sum score of endothelial dysfunction was not. Inflammation and endothelial dysfunction are both associated with depressive disorder, independent of lifestyle factors. Our results might suggest that inflammation and endothelial dysfunction are involved in depression. Copyright © 2016. Published by Elsevier Inc.

Nrf2 Deficiency Exacerbates Obesity-Induced Oxidative Stress, Neurovascular Dysfunction, Blood-Brain Barrier Disruption, Neuroinflammation, Amyloidogenic Gene Expression, and Cognitive Decline in Mice, Mimicking the Aging Phenotype.

PubMed

Tarantini, Stefano; Valcarcel-Ares, M Noa; Yabluchanskiy, Andriy; Tucsek, Zsuzsanna; Hertelendy, Peter; Kiss, Tamas; Gautam, Tripti; Zhang, Xin A; Sonntag, William E; de Cabo, Rafael; Farkas, Eszter; Elliott, Michael H; Kinter, Michael T; Deak, Ferenc; Ungvari, Zoltan; Csiszar, Anna

2018-06-14

Obesity has deleterious effects on cognitive function in the elderly adults. In mice, aging exacerbates obesity-induced oxidative stress, microvascular dysfunction, blood-brain barrier (BBB) disruption, and neuroinflammation, which compromise cognitive health. However, the specific mechanisms through which aging and obesity interact to remain elusive. Previously, we have shown that Nrf2 signaling plays a critical role in microvascular resilience to obesity and that aging is associated with progressive Nrf2 dysfunction, promoting microvascular impairment. To test the hypothesis that Nrf2 deficiency exacerbates cerebromicrovascular dysfunction induced by obesity Nrf2+/+ and Nrf2-/-, mice were fed an adipogenic high-fat diet (HFD). Nrf2 deficiency significantly exacerbated HFD-induced oxidative stress and cellular senescence, impairment of neurovascular coupling responses, BBB disruption, and microglia activation, mimicking the aging phenotype. Obesity in Nrf2-/- mice elicited complex alterations in the amyloidogenic gene expression profile, including upregulation of amyloid precursor protein. Nrf2 deficiency and obesity additively reduced long-term potentiation in the CA1 area of the hippocampus. Collectively, Nrf2 dysfunction exacerbates the deleterious effects of obesity, compromising cerebromicrovascular and brain health by impairing neurovascular coupling mechanisms, BBB integrity and synaptic function and promoting neuroinflammation. These results support a possible role for age-related Nrf2 dysfunction in the pathogenesis of vascular cognitive impairment and Alzheimer's disease.

Effect of Melilotus suaveolens extract on pulmonary microvascular permeability by downregulating vascular endothelial growth factor expression in rats with sepsis

PubMed Central

LIU, MING-WEI; SU, MEI-XIAN; ZHANG, WEI; WANG, YUN HUI; QIN, LAN-FANG; LIU, XU; TIAN, MAO-LI; QIAN, CHUAN-YUN

2015-01-01

A typical indicator of sepsis is the development of progressive subcutaneous and body-cavity edema, which is caused by the breakdown of endothelial barrier function, leading to a marked increase in vascular permeability. Microvascular leakage predisposes to microvascular thrombosis, breakdown of microcirculatory flow and organ failure, which are common events preceding mortality in patients with severe sepsis. Melilotus suaveolens (M. suaveolens) is a Traditional Tibetan Medicine. Previous pharmacological studies have demonstrated that an ethanolic extract of M. suaveolens has powerful anti-inflammatory activity and leads to an improvement in capillary permeability. However, the mechanisms underlying its pharmacological activity remain elusive. The present study aimed to assess the impact of M. suaveolens extract tablets on pulmonary vascular permeability, and their effect on regulating lung inflammation and the expression of vascular endothelial growth factor (VEGF) in the lung tissue of rats with sepsis. A cecal ligation and puncture (CLP) sepsis model was established for both the control and treatment groups. ~2 h prior to surgery, 25 mg/kg of M. suaveolens extract tablet was administered to the treatment group. Polymerase chain reaction and western blot analyses were used to assess the expression of nuclear factor (NF)-κB and VEGF in the lung tissue, and ELISA was applied to detect changes in serum tumor necrosis factor-α as well as interleukins (IL) -1, -4, -6, and -10. The lung permeability, wet/dry weight ratio and lung pathology were determined. The results demonstrated that in the lung tissue of CLP-rats with sepsis, M. suaveolens extract inhibited the expression of NF-κB, reduced the inflammatory response and blocked the expression of VEGF, and thus significantly decreased lung microvascular permeability. The effects of M. Suaveolens extract may be of potential use in the treatment of CLP-mediated lung microvascular permeability. PMID:25571852

GSK-3Beta-Dependent Activation of GEF-H1/ROCK Signaling Promotes LPS-Induced Lung Vascular Endothelial Barrier Dysfunction and Acute Lung Injury.

PubMed

Yi, Lei; Huang, Xiaoqin; Guo, Feng; Zhou, Zengding; Chang, Mengling; Huan, Jingning

2017-01-01

The bacterial endotoxin or lipopolysaccharide (LPS) leads to the extensive vascular endothelial cells (EC) injury under septic conditions. Guanine nucleotide exchange factor-H1 (GEF-H1)/ROCK signaling not only involved in LPS-induced overexpression of pro-inflammatory mediator in ECs but also implicated in LPS-induced endothelial hyper-permeability. However, the mechanisms behind LPS-induced GEF-H1/ROCK signaling activation in the progress of EC injury remain incompletely understood. GEF-H1 localized on microtubules (MT) and is suppressed in its MT-bound state. MT disassembly promotes GEF-H1 release from MT and stimulates downstream ROCK-specific GEF activity. Since glycogen synthase kinase (GSK-3beta) participates in regulating MT dynamics under pathologic conditions, we examined the pivotal roles for GSK-3beta in modulating LPS-induced activation of GEF-H1/ROCK, increase of vascular endothelial permeability and severity of acute lung injury (ALI). In this study, we found that LPS induced human pulmonary endothelial cell (HPMEC) monolayers disruption accompanied by increase in GSK-3beta activity, activation of GEF-H1/ROCK signaling and decrease in beta-catenin and ZO-1 expression. Inhibition of GSK-3beta reduced HPMEC monolayers hyper-permeability and GEF-H1/ROCK activity in response to LPS. GSK-3beta/GEF-H1/ROCK signaling is implicated in regulating the expression of beta-catenin and ZO-1. In vivo , GSK-3beta inhibition attenuated LPS-induced activation of GEF-H1/ROCK pathway, lung edema and subsequent ALI. These findings present a new mechanism of GSK-3beta-dependent exacerbation of lung micro-vascular hyper-permeability and escalation of ALI via activation of GEF-H1/ROCK signaling and disruption of intracellular junctional proteins under septic condition.

Endothelial microparticles interact with and support the proliferation of T cells.

PubMed

Wheway, Julie; Latham, Sharissa L; Combes, Valery; Grau, Georges E R

2014-10-01

Endothelial cells closely interact with circulating lymphocytes. Aggression or activation of the endothelium leads to an increased shedding of endothelial cell microparticles (MP). Endothelial MP (EMP) are found in high plasma levels in numerous immunoinflammatory diseases, such as atherosclerosis, sepsis, multiple sclerosis, and cerebral malaria, supporting their role as effectors and markers of vascular dysfunction. Given our recently described role for human brain microvascular endothelial cells (HBEC) in modulating immune responses, we investigated how HBEC-derived MP could interact with and support the proliferation of T cells. Like their mother cells, EMP expressed molecules important for Ag presentation and T cell costimulation, that is, β2-microglobulin, MHC II, CD40, and ICOSL. HBEC were able to take up fluorescently labeled Ags with EMP also containing fluorescent Ags, suggestive of Ag carryover from HBEC to EMP. In cocultures, fluorescently labeled EMP from resting or cytokine-stimulated HBEC formed conjugates with both CD4(+) and CD8(+) subsets, with higher proportions of T cells binding EMP from cytokine-stimulated cells. The increased binding of EMP from cytokinestimulated HBEC to T cells was VCAM-1 and ICAM-1 dependent. Finally, in CFSE T cell proliferation assays using anti-CD3 mAb or T cell mitogens, EMP promoted the proliferation of CD4(+) T cells and that of CD8(+) T cells in the absence of exogenous stimuli and in the T cell mitogenic stimulation. Our findings provide novel evidence that EMP can enhance T cell activation and potentially ensuing Ag presentation, thereby pointing toward a novel role for MP in neuroimmunological complications of infectious diseases. Copyright © 2014 by The American Association of Immunologists, Inc.

Cancer Cells Regulate Biomechanical Properties of Human Microvascular Endothelial Cells*

PubMed Central

Mierke, Claudia Tanja

2011-01-01

Metastasis is a key event of malignant tumor progression. The capability to metastasize depends on the ability of the cancer cell to migrate into connective tissue, adhere, and possibly transmigrate through the endothelium. Previously we reported that the endothelium does not generally act as barrier for cancer cells to migrate in three-dimensional extracellular matrices (3D-ECMs). Instead, the endothelium acts as an enhancer or a promoter for the invasiveness of certain cancer cells. How invasive cancer cells diminish the endothelial barrier function still remains elusive. Therefore, this study investigates whether invasive cancer cells can decrease the endothelial barrier function through alterations of endothelial biomechanical properties. To address this, MDA-MB-231 breast cancer cells were used that invade deeper and more numerous into 3D-ECMs when co-cultured with microvascular endothelial cells. Using magnetic tweezer measurements, MDA-MB-231 cells were found to alter the mechanical properties of endothelial cells by reducing endothelial cell stiffness. Using spontaneous bead diffusion, actin cytoskeletal remodeling dynamics were shown to be increased in endothelial cells co-cultured with MDA-MB-231 cells compared with mono-cultured endothelial cells. In addition, knockdown of the α5 integrin subunit in highly transmigrating α5β1high cells derived from breast, bladder, and kidney cancer cells abolished the endothelial invasion-enhancing effect comparable with the inhibition of myosin light chain kinase. These results indicate that the endothelial invasion-enhancing effect is α5β1 integrin-dependent. Moreover, inhibition of Rac-1, Rho kinase, MEK kinase, and PI3K reduced the endothelial invasion-enhancing effect, indicating that signaling via small GTPases may play a role in the endothelial facilitated increased invasiveness of cancer cells. In conclusion, decreased stiffness and increased cytoskeletal remodeling dynamics of endothelial cells may account for the breakdown of endothelial barrier function, suggesting that biomechanical alterations are sufficient to facilitate the transmigration and invasion of invasive cancer cells into 3D-ECMs. PMID:21940631

Leptin promotes endothelial dysfunction in chronic kidney disease through AKT/GSK3β and β-catenin signals

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ding, Nannan; Liu, Bing; Song, Jiaguang

Endothelial dysfunction (ED) is a well-recognized instigator of cardiovascular diseases and develops in chronic kidney disease (CKD) with high rate. Recent studies have implicated that leptin is associated with endothelial dysfunction. We investigated the relationship between leptin and markers of ED in CKD patients and how leptin contributed to endothelial damage. 140 CKD patients and 140 healthy subjects were studied. Serum leptin levels were significantly higher in CKD than in controls and displayed significantly positive association with the increase levels of sICAM-1 and sVCAM-1 but negative correlation with flow-mediated dilatation (FMD) reduction in patients. Our in vitro study demonstrated that leptinmore » induced overexpression of ICAM-1 and VCAM-1, led to f-actin reorganization and vinculin assembly, increased endothelial monolayer permeability for FITC-dextran, and accelerated endothelial cell migration; these changes were markedly reversed when the cells were transfected with AKT or β-catenin shRNA vectors. Notably, high leptin resulted in hyper-phosphorylation of AKT and GSK3β, along with nuclear accumulation of β-catenin. In conclusion, serum leptin was elevated in CKD patients and it might contribute to endothelial dysfunction by disarrangement of f-actin cytoskeleton via a mechanism involving the AKT/GSK3β and β-catenin pathway. - Highlights: • Serum leptin was elevated in CKD patients and it was associated with endothelial dysfunction. • Leptin induced endothelial dysfunction by remodeling cytoskeleton in HUVECs. • Leptin promoted endothelial dysfunction via a mechanism involving the AKT/GSK3β and β-catenin signals.« less

Effect of Vandetanib on Andes virus survival in the hamster model of Hantavirus pulmonary syndrome.

PubMed

Bird, Brian H; Shrivastava-Ranjan, Punya; Dodd, Kimberly A; Erickson, Bobbie R; Spiropoulou, Christina F

2016-08-01

Hantavirus pulmonary syndrome (HPS) is a severe disease caused by hantavirus infection of pulmonary microvascular endothelial cells leading to microvascular leakage, pulmonary edema, pleural effusion and high case fatality. Previously, we demonstrated that Andes virus (ANDV) infection caused up-regulation of vascular endothelial growth factor (VEGF) and concomitant downregulation of the cellular adhesion molecule VE-cadherin leading to increased permeability. Analyses of human HPS-patient sera have further demonstrated increased circulating levels of VEGF. Here we investigate the impact of a small molecule antagonist of the VEGF receptor 2 (VEGFR-2) activation in vitro, and overall impact on survival in the Syrian hamster model of HPS. Copyright © 2016. Published by Elsevier B.V.

TMEM16A Contributes to Endothelial Dysfunction by Facilitating Nox2 NADPH Oxidase-Derived Reactive Oxygen Species Generation in Hypertension.

PubMed

Ma, Ming-Ming; Gao, Min; Guo, Kai-Min; Wang, Mi; Li, Xiang-Yu; Zeng, Xue-Lin; Sun, Lu; Lv, Xiao-Fei; Du, Yan-Hua; Wang, Guan-Lei; Zhou, Jia-Guo; Guan, Yong-Yuan

2017-05-01

Ca 2+ -activated Cl - channels play a crucial role in various physiological processes. However, the role of TMEM16A in vascular endothelial dysfunction during hypertension is unclear. In this study, we investigated the specific involvement of TMEM16A in regulating endothelial function and blood pressure and the underlying mechanism. Reverse transcription-polymerase chain reaction, Western blotting, coimmunoprecipitation, confocal imaging, patch-clamp recordings, and TMEM16A endothelial-specific transgenic and knockout mice were used. We found that TMEM16A was expressed abundantly and functioned as a Ca 2+ -activated Cl - channel in endothelial cells. Angiotensin II induced endothelial dysfunction with an increase in TMEM16A expression. The knockout of endothelial-specific TMEM16A significantly lowered the blood pressure and ameliorated endothelial dysfunction in angiotensin II-induced hypertension, whereas the overexpression of endothelial-specific TMEM16A resulted in the opposite effects. These results were related to the increased reactive oxygen species production, Nox2-containing NADPH oxidase activation, and Nox2 and p22phox protein expression that were facilitated by TMEM16A on angiotensin II-induced hypertensive challenge. Moreover, TMEM16A directly bound with Nox2 and reduced the degradation of Nox2 through the proteasome-dependent degradation pathway. Therefore, TMEM16A is a positive regulator of endothelial reactive oxygen species generation via Nox2-containing NADPH oxidase, which induces endothelial dysfunction and hypertension. Modification of TMEM16A may be a novel therapeutic strategy for endothelial dysfunction-associated diseases. © 2017 American Heart Association, Inc.

Non-Acute Coronary Syndrome Anginal Chest Pain

PubMed Central

Agarwal, Megha; Mehta, Puja K.; Merz, C. Noel Bairey

2010-01-01

Anginal chest pain is one of the most common complaints in the outpatient setting. While much of the focus has been on identifying obstructive atherosclerotic coronary artery disease (CAD) as the cause of anginal chest pain, it is clear that microvascular coronary dysfunction (MCD) can also cause anginal chest pain as a manifestation of ischemic heart disease (IHD), and carries an increased cardiovascular risk. Epicardial coronary vasospasm, aortic stenosis, left ventricular hypertrophy, congenital coronary anomalies, mitral valve prolapse and abnormal cardiac nociception can also present as angina of cardiac origin. For non-acute coronary syndrome (ACS) stable chest pain, exercise treadmill testing (ETT) remains the primary tool for diagnosis of ischemia and cardiac risk stratification; however, in certain subsets of patients, such as women, ETT has a lower sensitivity and specificity for identifying obstructive CAD. When combined with an imaging modality, such as nuclear perfusion or echocardiography testing, the sensitivity and specificity of stress testing for detection of obstructive CAD improves significantly. Advancements in stress cardiac magnetic resonance imaging (MRI) enables detection of perfusion abnormalities in a specific coronary artery territory, as well as subendocardial ischemia associated with MCD. Coronary computed tomography angiography (CCTA) enables visual assessment of obstructive CAD, albeit with a higher radiation dose. Invasive coronary angiography (CA) remains the gold standard for diagnosis and treatment of obstructive lesions that cause medically refractory stable angina. Furthermore, in patients with normal coronary angiograms, the addition of coronary reactivity testing (CRT) can help diagnose endothelial dependent and independent microvascular dysfunction. Life-style modification and pharmacologic intervention remains the cornerstone of therapy to reduce morbidity and mortality in patients with stable angina. This review focuses on the pathophysiology, diagnosis, and treatment of stable, non-ACS anginal chest pain. PMID:20380951

Microcirculatory effects of zinc on fructose-fed hamsters.

PubMed

Castiglione, R C; Barros, C M M R; Boa, B C S; Bouskela, E

2016-04-01

Fructose is a major dietary component directly related to vascular dysfunction and diseases such as obesity, diabetes, and hypertension. Zinc is considered a non-pharmacological alternative for treating diabetes due to its antioxidant and hyperglycemia-lowering effects in diabetic animals. Therefore, the aim of this study was to evaluate the effects of dietary zinc supplementation on the microcirculatory parameters of fructose-fed hamsters. Male hamsters (Mesocricetus auratus) were fed drinking water substituted by 10% fructose solution for 60 days, whereas control animals were fed drinking water alone. Their microcirculatory function was evaluated using cheek pouch preparation, as well as their blood glucose and serum insulin levels. Their microcirculatory responses to acetylcholine (ACh, an endothelium-dependent vasodilator) and to sodium nitroprusside (SNP, an endothelium-independent vasodilator) as well as the increase in macromolecular permeability induced by 30 min of ischemia/reperfusion (I/R) were noted. Endothelium-dependent vasodilation was significantly increased in control animals with high zinc supplementation compared to the groups without zinc supplementation. Zinc was able to protect against plasma leakage induced by I/R in all control and fructose-fed groups, although the microvascular permeability was higher in animals fed drinking water substituted by 10% fructose solution compared to those fed filtered drinking water alone. Our results indicate that dietary zinc supplementation can improve microvascular dysfunction by increasing endothelial-dependent dilatation and reducing the increase in macromolecular permeability induced by I/R in fructose-fed animals. Copyright © 2015 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.

Loss of the endothelial glycocalyx is associated with increased E-selectin mediated adhesion of lung tumour cells to the brain microvascular endothelium.

PubMed

Rai, Srijana; Nejadhamzeeigilani, Zaynab; Gutowski, Nicholas J; Whatmore, Jacqueline L

2015-09-25

Arrest of metastasising lung cancer cells to the brain microvasculature maybe mediated by interactions between ligands on circulating tumour cells and endothelial E-selectin adhesion molecules; a process likely to be regulated by the endothelial glycocalyx. Using human cerebral microvascular endothelial cells and non-small cell lung cancer (NSCLC) cell lines, we describe how factors secreted by NSCLC cells i.e. cystatin C, cathepsin L, insulin-like growth factor-binding protein 7 (IGFBP7), vascular endothelial growth factor (VEGF) and tumour necrosis factor-alpha (TNF-α), damage the glycocalyx and enhance initial contacts between lung tumour and cerebral endothelial cells. Endothelial cells were treated with tumour secreted-proteins or lung tumour conditioned medium (CM). Surface levels of E-selectin were quantified by ELISA. Adhesion of A549 and SK-MES-1 cells was examined under flow conditions (1 dyne/cm(2)). Alterations in the endothelial glycocalyx were quantified by binding of fluorescein isothiocyanate-linked wheat germ agglutinin (WGA-FITC). A549 and SK-MES-1 CM and secreted-proteins significantly enhanced endothelial surface E-selectin levels after 30 min and 4 h and tumour cell adhesion after 30 min, 4 and 24 h. Both coincided with significant glycocalyx degradation; A549 and SK-MES-1 CM removing 55 ± 12 % and 58 ± 18.7 % of WGA-FITC binding, respectively. Inhibition of E-selectin binding by monoclonal anti-E-selectin antibody completely attenuated tumour cell adhesion. These data suggest that metastasising lung cancer cells facilitate their own adhesion to the brain endothelium by secreting factors that damage the endothelial glycocalyx, resulting in exposure of the previously shielded adhesion molecules and engagement of the E-selectin-mediated adhesion axis.

Novel critical role of Toll-like receptor 4 in lung ischemia-reperfusion injury and edema

PubMed Central

Zanotti, Giorgio; Casiraghi, Monica; Abano, John B.; Tatreau, Jason R.; Sevala, Mayura; Berlin, Hilary; Smyth, Susan; Funkhouser, William K.; Burridge, Keith; Randell, Scott H.; Egan, Thomas M.

2009-01-01

Toll-like receptors (TLRs) of the innate immune system contribute to noninfectious inflammatory processes. We employed a murine model of hilar clamping (1 h) with reperfusion times between 15 min and 3 h in TLR4-sufficient (C3H/OuJ) and TLR4-deficient (C3H/HeJ) anesthetized mice with additional studies in chimeric and myeloid differentiation factor 88 (MyD88)- and TLR4-deficient mice to determine the role of TLR4 in lung ischemia-reperfusion injury. Human pulmonary microvascular endothelial monolayers were subjected to simulated warm ischemia and reperfusion with and without CRX-526, a competitive TLR4 inhibitor. Functional TLR4 solely on pulmonary parenchymal cells, not bone marrow-derived cells, mediates early lung edema following ischemia-reperfusion independent of MyD88. Activation of MAPKs and NF-κB was significantly blunted and/or delayed in lungs of TLR4-deficient mice as a consequence of ischemia-reperfusion injury, but edema development appeared to be independent of activation of these signaling pathways. Pretreatment with a competitive TLR4 inhibitor prevented edema in vivo and reduced actin cytoskeletal rearrangement and gap formation in pulmonary microvascular endothelial monolayers subjected to simulated warm ischemia and reperfusion. In addition to its well-accepted role to alter gene transcription, functioning TLR4 on pulmonary parenchymal cells plays a key role in very early and profound pulmonary edema in murine lung ischemia-reperfusion injury. This may be due to a novel mechanism: regulation of endothelial cell cytoskeleton affecting microvascular endothelial cell permeability. PMID:19376887

MicroRNA Signature of Human Microvascular Endothelium Infected with Rickettsia rickettsii

PubMed Central

Sahni, Abha; Narra, Hema P.; Patel, Jignesh; Sahni, Sanjeev K.

2017-01-01

MicroRNAs (miRNAs) mediate gene silencing by destabilization and/or translational repression of target mRNA. Infection of human microvascular endothelial cells as primary targets of Rickettsia rickettsii, the etiologic agent of Rocky Mountain spotted fever, triggers host responses appertaining to alterations in cellular gene expression. Microarray-based profiling of endothelial cells infected with R. rickettsii for 3 or 24 h revealed differential expression of 33 miRNAs, of which miRNAs129-5p, 200a-3p, 297, 200b-3p, and 595 were identified as the top five up-regulated miRNAs (5 to 20-fold, p ≤ 0.01) and miRNAs 301b-3p, 548a-3p, and 377-3p were down-regulated (2 to 3-fold, p ≤ 0.01). Changes in the expression of selected miRNAs were confirmed by q-RT-PCR in both in vitro and in vivo models of infection. As potential targets, expression of genes encoding NOTCH1, SMAD2, SMAD3, RIN2, SOD1, and SOD2 was either positively or negatively regulated. Using a miRNA-specific mimic or inhibitor, NOTCH1 was determined to be a target of miRNA 200a-3p in R. rickettsii-infected human dermal microvascular endothelial cells (HMECs). Predictive interactome mapping suggested the potential for miRNA-mediated modulation of regulatory gene networks underlying important host cell signaling pathways. This first demonstration of altered endothelial miRNA expression provides new insights into regulatory elements governing mechanisms of host responses and pathogenesis during human rickettsial infections. PMID:28698491

Hydrophilic bile acids protect human blood-brain barrier endothelial cells from disruption by unconjugated bilirubin: an in vitro study

PubMed Central

Palmela, Inês; Correia, Leonor; Silva, Rui F. M.; Sasaki, Hiroyuki; Kim, Kwang S.; Brites, Dora; Brito, Maria A.

2015-01-01

Ursodeoxycholic acid and its main conjugate glycoursodeoxycholic acid are bile acids with neuroprotective properties. Our previous studies demonstrated their anti-apoptotic, anti-inflammatory, and antioxidant properties in neural cells exposed to elevated levels of unconjugated bilirubin (UCB) as in severe jaundice. In a simplified model of the blood-brain barrier, formed by confluent monolayers of a cell line of human brain microvascular endothelial cells, UCB has shown to induce caspase-3 activation and cell death, as well as interleukin-6 release and a loss of blood-brain barrier integrity. Here, we tested the preventive and restorative effects of these bile acids regarding the disruption of blood-brain barrier properties by UCB in in vitro conditions mimicking severe neonatal hyperbilirubinemia and using the same experimental blood-brain barrier model. Both bile acids reduced the apoptotic cell death induced by UCB, but only glycoursodeoxycholic acid significantly counteracted caspase-3 activation. Bile acids also prevented the upregulation of interleukin-6 mRNA, whereas only ursodeoxycholic acid abrogated cytokine release. Regarding barrier integrity, only ursodeoxycholic acid abrogated UCB-induced barrier permeability. Better protective effects were obtained by bile acid pre-treatment, but a strong efficacy was still observed by their addition after UCB treatment. Finally, both bile acids showed ability to cross confluent monolayers of human brain microvascular endothelial cells in a time-dependent manner. Collectively, data disclose a therapeutic time-window for preventive and restorative effects of ursodeoxycholic acid and glycoursodeoxycholic acid against UCB-induced blood-brain barrier disruption and damage to human brain microvascular endothelial cells. PMID:25821432

Cardiovascular risk reduction by reversing endothelial dysfunction:ARBs, ACE inhibitors, or both? Expectations from The ONTARGET Trial Programme

PubMed Central

Ruilope, Luis Miguel; Redón, Josep; Schmieder, Roland

2007-01-01

Endothelial dysfunction is the initial pathophysiological step in a progression of vascular damage that leads to overt cardiovascular and chronic kidney disease. Angiotensin II, the primary agent of the renin–angiotensin system (RAS), has a central role in endothelial dysfunction. Therefore, RAS blockade with an angiotensin receptor blocker (ARB) and/or angiotensin-converting enzyme (ACE) inhibitor provides a rational approach to reverse endothelial dysfunction, reduce microalbuminuria, and, thus, improves cardiovascular and renal prognosis. ARBs and ACE inhibitors act at different points in the RAS pathway and recent evidence suggests that there are differences regarding their effects on endothelial dysfunction. In addition to blood pressure lowering, studies have shown that ARBs reduce target-organ damage, including improvements in endothelial dysfunction, arterial stiffness, the progression of renal dysfunction in patients with type 2 diabetes, proteinuria, and left ventricular hypertrophy. The ONgoing Telmisartan Alone in combination with Ramipril Global Endpoint Trial (ONTARGET) Programme is expected to provide the ultimate evidence of whether improved endothelial function translates into reduced cardiovascular and renal events in high-risk patients, and to assess possible differential outcomes with telmisartan, the ACE inhibitor ramipril, or a combination of both (dual RAS blockade). Completion of ONTARGET is expected in 2008. PMID:17583170

Daily muscle stretching enhances blood flow, endothelial function, capillarity, vascular volume and connectivity in aged skeletal muscle.

PubMed

Hotta, Kazuki; Behnke, Bradley J; Arjmandi, Bahram; Ghosh, Payal; Chen, Bei; Brooks, Rachael; Maraj, Joshua J; Elam, Marcus L; Maher, Patrick; Kurien, Daniel; Churchill, Alexandra; Sepulveda, Jaime L; Kabolowsky, Max B; Christou, Demetra D; Muller-Delp, Judy M

2018-05-15

In aged rats, daily muscle stretching increases blood flow to skeletal muscle during exercise. Daily muscle stretching enhanced endothelium-dependent vasodilatation of skeletal muscle resistance arterioles of aged rats. Angiogenic markers and capillarity increased in response to daily stretching in muscles of aged rats. Muscle stretching performed with a splint could provide a feasible means of improving muscle blood flow and function in elderly patients who cannot perform regular aerobic exercise. Mechanical stretch stimuli alter the morphology and function of cultured endothelial cells; however, little is known about the effects of daily muscle stretching on adaptations of endothelial function and muscle blood flow. The present study aimed to determine the effects of daily muscle stretching on endothelium-dependent vasodilatation and muscle blood flow in aged rats. The lower hindlimb muscles of aged Fischer rats were passively stretched by placing an ankle dorsiflexion splint for 30 min day -1 , 5 days week -1 , for 4 weeks. Blood flow to the stretched limb and the non-stretched contralateral limb was determined at rest and during treadmill exercise. Endothelium-dependent/independent vasodilatation was evaluated in soleus muscle arterioles. Levels of hypoxia-induced factor-1α, vascular endothelial growth factor A and neuronal nitric oxide synthase were determined in soleus muscle fibres. Levels of endothelial nitric oxide synthase and superoxide dismutase were determined in soleus muscle arterioles, and microvascular volume and capillarity were evaluated by microcomputed tomography and lectin staining, respectively. During exercise, blood flow to plantar flexor muscles was significantly higher in the stretched limb. Endothelium-dependent vasodilatation was enhanced in arterioles from the soleus muscle from the stretched limb. Microvascular volume, number of capillaries per muscle fibre, and levels of hypoxia-induced factor-1α, vascular endothelial growth factor and endothelial nitric oxide synthase were significantly higher in the stretched limb. These results indicate that daily passive stretching of muscle enhances endothelium-dependent vasodilatation and induces angiogenesis. These microvascular adaptations may contribute to increased muscle blood flow during exercise in muscles that have undergone daily passive stretch. © 2018 The Authors. The Journal of Physiology © 2018 The Physiological Society.

Regulation of human feto-placental endothelial barrier integrity by vascular endothelial growth factors: competitive interplay between VEGF-A165a, VEGF-A165b, PIGF and VE-cadherin.

PubMed

Pang, Vincent; Bates, David O; Leach, Lopa

2017-12-01

The human placenta nourishes and protects the developing foetus whilst influencing maternal physiology for fetal advantage. It expresses several members of the vascular endothelial growth factor (VEGF) family including the pro-angiogenic/pro-permeability VEGF-A 165 a isoform, the anti-angiogenic VEGF-A 165 b, placental growth factor (PIGF) and their receptors, VEGFR1 and VEGFR2. Alterations in the ratio of these factors during gestation and in complicated pregnancies have been reported; however, the impact of this on feto-placental endothelial barrier integrity is unknown. The present study investigated the interplay of these factors on junctional occupancy of VE-cadherin and macromolecular leakage in human endothelial monolayers and the perfused placental microvascular bed. Whilst VEGF-A 165 a (50 ng/ml) increased endothelial monolayer albumin permeability ( P <0.0001), equimolar concentrations of VEGF-A 165 b ( P >0.05) or PlGF ( P >0.05) did not. Moreover, VEGF-A 165 b (100 ng/ml; P <0.001) but not PlGF (100 ng/ml; P >0.05) inhibited VEGF-A 165 a-induced permeability when added singly. PlGF abolished the VEGF-A 165 b-induced reduction in VEGF-A 165 a-mediated permeability ( P >0.05); PlGF was found to compete with VEGF-A 165 b for binding to Flt-1 at equimolar affinity. Junctional occupancy of VE-cadherin matched alterations in permeability. In the perfused microvascular bed, VEGF-A 165 b did not induce microvascular leakage but inhibited and reversed VEGF-A 165 a-induced loss of junctional VE-cadherin and tracer leakage. These results indicate that the anti-angiogenic VEGF-A 165 b isoform does not increase permeability in human placental microvessels or HUVEC primary cells and can interrupt VEGF-A 165 a-induced permeability. Moreover, the interplay of these isoforms with PIGF (and s-flt1) suggests that the ratio of these three factors may be important in determining the placental and endothelial barrier in normal and complicated pregnancies. © 2017 The Author(s).

C-reactive protein induces release of both endothelial microparticles and circulating endothelial cells in vitro and in vivo: further evidence of endothelial dysfunction.

PubMed

Devaraj, Sridevi; Kumaresan, Pappanaicken R; Jialal, Ishwarlal

2011-12-01

Inflammation is pivotal in atherosclerosis. A key early event in atherosclerosis is endothelial dysfunction. C-reactive protein (CRP), the prototypic marker of inflammation in humans, is a risk marker for cardiovascular disease, and there is mounting evidence to support its role in atherothrombosis. CRP has been shown to promote endothelial dysfunction both in vitro and in vivo. Emerging biomarkers of endothelial dysfunction include circulating endothelial cells (CECs) and endothelial microparticles (EMPs). However, there is a paucity of data examining the effect of CRP on CEC and EMP production in vitro and in vivo. In this report, we treated human aortic endothelial cells (HAECs) with increasing concentrations of CRP (0-50 μg/mL) or boiled CRP. We counted CECs and EMPs by flow cytometry. Although CRP treatment resulted in a significant increase in release of both CECs and EMPs, boiled CRP failed to have an effect. Pretreatment of HAECs with sepiapterin or diethylenetriamine NONOate, both of which preserve nitric oxide (NO), resulted in attenuation of CRP's effects on CECs and EMPs. CD32 and CD64 blocking antibodies but not CD16 antibody or lectin-like oxidized LDL receptor 1 small interfering RNA (LOX-1 siRNA) prevented CRP-induced production of CECs and EMPs. Furthermore, delivery of human CRP to Wistar rats compared with human serum albumin resulted in significantly increased CECs and EMPs, corroborating the in vitro findings. We provide novel data that CRP, via NO deficiency, promotes endothelial dysfunction by inducing release of CECs and EMPs, which are biomarkers of endothelial dysfunction.

Losartan increases muscle insulin delivery and rescues insulin's metabolic action during lipid infusion via microvascular recruitment

PubMed Central

Wang, Nasui; Chai, Weidong; Zhao, Lina; Tao, Lijian; Cao, Wenhong

2013-01-01

Insulin delivery and transendothelial insulin transport are two discrete steps that limit muscle insulin action. Angiotensin II type 1 receptor (AT1R) blockade recruits microvasculature and increases glucose use in muscle. Increased muscle microvascular perfusion is associated with increased muscle delivery and action of insulin. To examine the effect of acute AT1R blockade on muscle insulin uptake and action, rats were studied after an overnight fast to examine the effects of losartan on muscle insulin uptake (protocol 1), microvascular perfusion (protocol 2), and insulin's microvascular and metabolic actions in the state of insulin resistance (protocol 3). Endothelial cell insulin uptake was assessed, using 125I-insulin as tracer. Systemic lipid infusion was used to induce insulin resistance. Losartan significantly increased muscle insulin uptake (∼60%, P < 0.03), which was associated with a two- to threefold increase in muscle microvascular blood volume (MBV; P = 0.002) and flow (MBF; P = 0.002). Losartan ± angiotensin II had no effect on insulin internalization in cultured endothelial cells. Lipid infusion abolished insulin-mediated increases in muscle MBV and MBF and lowered insulin-stimulated whole body glucose disposal (P = 0.0001), which were reversed by losartan administration. Inhibition of nitric oxide synthase abolished losartan-induced muscle insulin uptake and reversal of lipid-induced metabolic insulin resistance. We conclude that AT1R blockade increases muscle insulin uptake mainly via microvascular recruitment and rescues insulin's metabolic action in the insulin-resistant state. This may contribute to the clinical findings of decreased cardiovascular events and new onset of diabetes in patients receiving AT1R blockers. PMID:23299501

Long-term effects of bariatric surgery on peripheral endothelial function and coronary microvascular function.

PubMed

Tarzia, Pierpaolo; Lanza, Gaetano A; Sestito, Alfonso; Villano, Angelo; Russo, Giulio; Figliozzi, Stefano; Lamendola, Priscilla; De Vita, Antonio; Crea, Filippo

We previously demonstrated that bariatric surgery (BS) leads to a short-term significant improvement of endothelial function and coronary microvascular function. In this study we assessed whether BS maintains its beneficial effect at long-term follow up. We studied 19 morbidly obese patients (age 43±9years, 12 women) without any evidence of cardiovascular disease who underwent BS. Patients were studied before BS, at 3 months and at 4.0±1.5years follow up. Peripheral vascular function was assessed by flow-mediated dilation (FMD) and nitrate-mediated dilation (NMD), i.e., brachial artery diameter changes in response to post-ischemic forearm hyperhaemia and to nitroglycerin administration, respectively. Coronary microvascular function was assessed by measuring coronary blood flow (CBF) response to intravenous adenosine and to cold pressor test (CPT) in the left anterior descending coronary artery. Together with improvement of anthropometric and metabolic profile, at long-term follow-up patients showed a significant improvement of FMD (6.43±2.88 vs. 8.21±1.73%, p=0.018), and CBF response to both adenosine (1.73±0.48 vs. 2.58±0.54; p<0.01) and CPT (1.43±0.30 vs. 2.23±0.48; p<0.01), compared to basal values. No differences in vascular end-points were shown at 3-month and 4-year follow-up after BS. Our data show that, in morbidly obese patients, BS exerts beneficial and long lasting effects on peripheral endothelial function and on coronary microvascular dilator function. Copyright © 2016 Asia Oceania Association for the Study of Obesity. Published by Elsevier Ltd. All rights reserved.

Sprint interval and endurance training are equally effective in increasing muscle microvascular density and eNOS content in sedentary males

PubMed Central

Cocks, Matthew; Shaw, Christopher S; Shepherd, Sam O; Fisher, James P; Ranasinghe, Aaron M; Barker, Thomas A; Tipton, Kevin D; Wagenmakers, Anton J M

2013-01-01

Sprint interval training (SIT) has been proposed as a time efficient alternative to endurance training (ET) for increasing skeletal muscle oxidative capacity and improving certain cardiovascular functions. In this study we sought to make the first comparisons of the structural and endothelial enzymatic changes in skeletal muscle microvessels in response to ET and SIT. Sixteen young sedentary males (age 21 ± SEM 0.7 years, BMI 23.8 ± SEM 0.7 kg m−2) were randomly assigned to 6 weeks of ET (40–60 min cycling at ∼65%, 5 times per week) or SIT (4–6 Wingate tests, 3 times per week). Muscle biopsies were taken from the m. vastus lateralis before and following 60 min cycling at 65% to measure muscle microvascular endothelial eNOS content, eNOS serine1177 phosphorylation, NOX2 content and capillarisation using quantitative immunofluorescence microscopy. Whole body insulin sensitivity, arterial stiffness and blood pressure were also assessed. ET and SIT increased skeletal muscle microvascular eNOS content (ET 14%; P < 0.05, SIT 36%; P < 0.05), with a significantly greater increase observed following SIT (P < 0.05). Sixty minutes of moderate intensity exercise increased eNOS ser1177 phosphorylation in all instances (P < 0.05), but basal and post-exercise eNOS ser1177 phosphorylation was lower following both training modes. All microscopy measures of skeletal muscle capillarisation (P < 0.05) were increased with SIT or ET, while neither endothelial nor sarcolemmal NOX2 was changed. Both training modes reduced aortic stiffness and increased whole body insulin sensitivity (P < 0.05). In conclusion, in sedentary males SIT and ET are effective in improving muscle microvascular density and eNOS protein content. PMID:22946099

In vivo microvascular and macrovascular endothelial function is not associated with circulating dimethylarginines in patients with rheumatoid arthritis: a prospective analysis of the DRACCO cohort.

PubMed

Dimitroulas, Theodoros; Sandoo, Aamer; Hodson, James; Smith, Jacqueline P; Kitas, George D

2016-07-01

To examine associations between asymmetric (ADMA), symmetric dimethylarginine (SDMA) and ADMA:SDMA ratio with assessments of endothelial function and coronary artery perfusion in RA patients. ADMA and SDMA levels were measured in 197 RA individuals [144 (77.4%) females, median age: 66 years (quartiles: 59-73)]. Patients underwent assessments of microvascular endothelium-dependent and endothelium-independent function, macrovascular endothelium-dependent and endothelium-independent function and vascular morphology (pulse wave analysis, carotid intima-media thickness (cIMT), and carotid plaque). Coronary perfusion was assessed by subendocardial viability ratio (SEVR). SEVR correlated with SDMA (r = 0.172, p = 0.026) and ADMA:SDMA (r = -0.160, p = 0.041) in univariable analysis, but not in multivariable analysis accounting for confounding factors. Neither ADMA:SDMA ratio nor SDMA were significantly correlated with microvascular or macrovascular endothelial function, or with arterial stiffness and cIMT. Within subgroup of patients (n = 26) with high inflammatory markers, a post-hoc analysis showed that SDMA and the ADMA:SDMA ratio were significantly associated with endothelium-dependent microvascular function in univariable analysis, with Pearson's r correlation coefficients of -0.440 (p = 0.031) and 0.511 (p = 0.011), respectively. Similar finding were established between ADMA:SDMA ratio and arterial stiffness in univariable analysis, with Pearson's r of 0.493, (p = 0.024). Dimethylarginines were not found to be significantly associated with several assessments of vascular function and morphology in patients with RA, however, post-hoc analysis indicates that there may be associations in patients with raised inflammatory markers. Our results suggest that dysregulated NO metabolism may not be the sole mechanism for the development of preclinical atherosclerosis in RA.

Low intrinsic exercise capacity in rats predisposes to age-dependent cardiac remodeling independent of macrovascular function

PubMed Central

Ritchie, Rebecca H.; Leo, Chen Huei; Qin, Chengxue; Stephenson, Erin J.; Bowden, Marissa A.; Buxton, Keith D.; Lessard, Sarah J.; Rivas, Donato A.; Koch, Lauren G.; Britton, Steven L.; Woodman, Owen L.

2013-01-01

Rats selectively bred for low (LCR) or high (HCR) intrinsic running capacity simultaneously present with contrasting risk factors for cardiovascular and metabolic disease. However, the impact of these phenotypes on left ventricular (LV) morphology and microvascular function, and their progression with aging, remains unresolved. We tested the hypothesis that the LCR phenotype induces progressive age-dependent LV remodeling and impairments in microvascular function, glucose utilization, and β-adrenergic responsiveness, compared with HCR. Hearts and vessels isolated from female LCR (n = 22) or HCR (n = 26) were studied at 12 and 35 wk. Nonselected N:NIH founder rats (11 wk) were also investigated (n = 12). LCR had impaired glucose tolerance and elevated plasma insulin (but not glucose) and body-mass at 12 wk compared with HCR, with early LV remodeling. By 35 wk, LV prohypertrophic and glucose transporter GLUT4 gene expression were up- and downregulated, respectively. No differences in LV β-adrenoceptor expression or cAMP content between phenotypes were observed. Macrovascular endothelial function was predominantly nitric oxide (NO)-mediated in both phenotypes and remained intact in LCR for both age-groups. In contrast, mesenteric arteries microvascular endothelial function, which was impaired in LCR rats regardless of age. At 35 wk, endothelial-derived hyperpolarizing factor-mediated relaxation was impaired whereas the NO contribution to relaxation is intact. Furthermore, there was reduced β2-adrenoceptor responsiveness in both aorta and mesenteric LCR arteries. In conclusion, diminished intrinsic exercise capacity impairs systemic glucose tolerance and is accompanied by progressive development of LV remodeling. Impaired microvascular perfusion is a likely contributing factor to the cardiac phenotype. PMID:23262135

Proton Pump Inhibitors Decrease Soluble fms-Like Tyrosine Kinase-1 and Soluble Endoglin Secretion, Decrease Hypertension, and Rescue Endothelial Dysfunction.

PubMed

Onda, Kenji; Tong, Stephen; Beard, Sally; Binder, Natalie; Muto, Masanaga; Senadheera, Sevvandi N; Parry, Laura; Dilworth, Mark; Renshall, Lewis; Brownfoot, Fiona; Hastie, Roxanne; Tuohey, Laura; Palmer, Kirsten; Hirano, Toshihiko; Ikawa, Masahito; Kaitu'u-Lino, Tu'uhevaha; Hannan, Natalie J

2017-03-01

Preeclampsia is a severe complication of pregnancy. Antiangiogenic factors soluble fms-like tyrosine kinase-1 (sFlt-1) and soluble endoglin are secreted in excess from the placenta, causing hypertension, endothelial dysfunction, and multiorgan injury. Oxidative stress and vascular inflammation exacerbate the endothelial injury. A drug that can block these pathophysiological steps would be an attractive treatment option. Proton pump inhibitors (PPIs) are safe in pregnancy where they are prescribed for gastric reflux. We performed functional studies on primary human tissues and animal models to examine the effects of PPIs on sFlt-1 and soluble endoglin secretion, vessel dilatation, blood pressure, and endothelial dysfunction. PPIs decreased sFlt-1 and soluble endoglin secretion from trophoblast, placental explants from preeclamptic pregnancies, and endothelial cells. They also mitigated tumor necrosis factor-α-induced endothelial dysfunction: PPIs blocked endothelial vascular cell adhesion molecule-1 expression, leukocyte adhesion to endothelium, and disruption of endothelial tube formation. PPIs decreased endothelin-1 secretion and enhanced endothelial cell migration. Interestingly, the PPI esomeprazole vasodilated maternal blood vessels from normal pregnancies and cases of preterm preeclampsia, but its vasodilatory effects were lost when the vessels were denuded of their endothelium. Esomeprazole decreased blood pressure in a transgenic mouse model where human sFlt-1 was overexpressed in placenta. PPIs upregulated endogenous antioxidant defenses and decreased cytokine secretion from placental tissue and endothelial cells. We have found that PPIs decrease sFlt-1 and soluble endoglin secretion and endothelial dysfunction, dilate blood vessels, decrease blood pressure, and have antioxidant and anti-inflammatory properties. They have therapeutic potential for preeclampsia and other diseases where endothelial dysfunction is involved. © 2017 American Heart Association, Inc.

Decellularized Human Kidney Cortex Hydrogels Enhance Kidney Microvascular Endothelial Cell Maturation and Quiescence.

PubMed

Nagao, Ryan J; Xu, Jin; Luo, Ping; Xue, Jun; Wang, Yi; Kotha, Surya; Zeng, Wen; Fu, Xiaoyun; Himmelfarb, Jonathan; Zheng, Ying

2016-10-01

The kidney peritubular microvasculature is highly susceptible to injury from drugs and toxins, often resulting in acute kidney injury and progressive chronic kidney disease. Little is known about the process of injury and regeneration of human kidney microvasculature, resulting from the lack of appropriate kidney microvascular models that can incorporate the proper cells, extracellular matrices (ECMs), and architectures needed to understand the response and contribution of individual vascular components in these processes. In this study, we present methods to recreate the human kidney ECM (kECM) microenvironment by fabricating kECM hydrogels derived from decellularized human kidney cortex. The majority of native matrix proteins, such as collagen-IV, laminin, and heparan sulfate proteoglycan, and their isoforms were preserved in similar proportions as found in normal kidneys. Human kidney peritubular microvascular endothelial cells (HKMECs) became more quiescent when cultured on this kECM gel compared with culture on collagen-I-assessed using phenotypic, genotypic, and functional assays; whereas human umbilical vein endothelial cells became stimulated on kECM gels. We demonstrate for the first time that human kidney cortex can form a hydrogel suitable for use in flow-directed microphysiological systems. Our findings strongly suggest that selecting the proper ECM is a critical consideration in the development of vascularized organs on a chip and carries important implications for tissue engineering of all vascularized organs.

Ionophore and Biometal Modulation of P-glycoprotein Expression and Function in Human Brain Microvascular Endothelial Cells.

PubMed

McInerney, Mitchell P; Volitakis, Irene; Bush, Ashley I; Banks, William A; Short, Jennifer L; Nicolazzo, Joseph A

2018-03-05

Biometals such as zinc and copper have been shown to affect tight junction expression and subsequently blood-brain barrier (BBB) integrity. Whether these biometals also influence the expression and function of BBB transporters such as P-glycoprotein (P-gp) however is currently unknown. Using the immortalised human cerebral microvascular endothelial (hCMEC/D3) cell line, an in-cell western assay (alongside western blotting) assessed relative P-gp expression after treatment with the metal ionophore clioquinol and biometals zinc and copper. The fluorescent P-gp substrate rhodamine-123 was employed to observe functional modulation, and inductively coupled plasma mass spectrometry (ICP-MS) provided information on biometal trafficking. A 24-h treatment with clioquinol, zinc and copper (0.5, 0.5 and 0.1 μM) induced a significant upregulation of P-gp (1.7-fold) assessed by in-cell western and this was confirmed with western blotting (1.8-fold increase). This same treatment resulted in a 23% decrease in rhodamine-123 accumulation over a 1 h incubation. ICP-MS demonstrated that while t8his combination treatment had no effect on intracellular zinc concentrations, the treatment significantly enhanced bioavailable copper (4.6-fold). Enhanced delivery of copper to human brain microvascular endothelial cells is associated with enhanced expression and function of the important efflux pump P-gp, which may provide therapeutic opportunities for P-gp modulation.

Are there Race-Dependent Endothelial Cell Responses to Exercise?

PubMed Central

Brown, Michael D.; Feairheller, Deborah L.

2013-01-01

African Americans have endothelial dysfunction which likely contributes to their high prevalence of hypertension. Endothelial cell responses to stimuli could play a role in the development of endothelial dysfunction and hypertension. High physiological levels of vascular laminar shear stress can profoundly alter endothelial cell phenotype. It is not known whether there are race-dependent endothelial cell responses to laminar shear stress. PMID:23262464

Exercise performance and peripheral vascular insufficiency improve with AMPK activation in high-fat diet-fed mice.

PubMed

Baltgalvis, Kristen A; White, Kathy; Li, Wei; Claypool, Mark D; Lang, Wayne; Alcantara, Raniel; Singh, Baljit K; Friera, Annabelle M; McLaughlin, John; Hansen, Derek; McCaughey, Kelly; Nguyen, Henry; Smith, Ira J; Godinez, Guillermo; Shaw, Simon J; Goff, Dane; Singh, Rajinder; Markovtsov, Vadim; Sun, Tian-Qiang; Jenkins, Yonchu; Uy, Gerald; Li, Yingwu; Pan, Alison; Gururaja, Tarikere; Lau, David; Park, Gary; Hitoshi, Yasumichi; Payan, Donald G; Kinsella, Todd M

2014-04-15

Intermittent claudication is a form of exercise intolerance characterized by muscle pain during walking in patients with peripheral artery disease (PAD). Endothelial cell and muscle dysfunction are thought to be important contributors to the etiology of this disease, but a lack of preclinical models that incorporate these elements and measure exercise performance as a primary end point has slowed progress in finding new treatment options for these patients. We sought to develop an animal model of peripheral vascular insufficiency in which microvascular dysfunction and exercise intolerance were defining features. We further set out to determine if pharmacological activation of 5'-AMP-activated protein kinase (AMPK) might counteract any of these functional deficits. Mice aged on a high-fat diet demonstrate many functional and molecular characteristics of PAD, including the sequential development of peripheral vascular insufficiency, increased muscle fatigability, and progressive exercise intolerance. These changes occur gradually and are associated with alterations in nitric oxide bioavailability. Treatment of animals with an AMPK activator, R118, increased voluntary wheel running activity, decreased muscle fatigability, and prevented the progressive decrease in treadmill exercise capacity. These functional performance benefits were accompanied by improved mitochondrial function, the normalization of perfusion in exercising muscle, increased nitric oxide bioavailability, and decreased circulating levels of the endogenous endothelial nitric oxide synthase inhibitor asymmetric dimethylarginine. These data suggest that aged, obese mice represent a novel model for studying exercise intolerance associated with peripheral vascular insufficiency, and pharmacological activation of AMPK may be a suitable treatment for intermittent claudication associated with PAD.

Bicarbonate disruption of the pulmonary endothelial barrier via activation of endogenous soluble adenylyl cyclase, isoform 10

PubMed Central

Obiako, Boniface; Calchary, Wendy; Xu, Ningyong; Kunstadt, Ryan; Richardson, Bianca; Nix, Jessica

2013-01-01

It is becoming increasingly apparent that cAMP signals within the pulmonary endothelium are highly compartmentalized, and this compartmentalization is critical to maintaining endothelial barrier integrity. Studies demonstrate that the exogenous soluble bacterial toxin, ExoY, and heterologous expression of the forskolin-stimulated soluble mammalian adenylyl cyclase (AC) chimera, sACI/II, elevate cytosolic cAMP and disrupt the pulmonary microvascular endothelial barrier. The barrier-disruptive effects of cytosolic cAMP generated by exogenous soluble ACs are in contrast to the barrier-protective effects of subplasma membrane cAMP generated by transmembrane AC, which strengthens endothelial barrier integrity. Endogenous soluble AC isoform 10 (AC10 or commonly known as sAC) lacks transmembrane domains and localizes within the cytosolic compartment. AC10 is uniquely activated by bicarbonate to generate cytosolic cAMP, yet its role in regulation of endothelial barrier integrity has not been addressed. Here we demonstrate that, within the pulmonary circulation, AC10 is expressed in pulmonary microvascular endothelial cells (PMVECs) and pulmonary artery endothelial cells (PAECs), yet expression in PAECs is lower. Furthermore, pulmonary endothelial cells selectively express bicarbonate cotransporters. While extracellular bicarbonate generates a phosphodiesterase 4-sensitive cAMP pool in PMVECs, no such cAMP response is detected in PAECs. Finally, addition of extracellular bicarbonate decreases resistance across the PMVEC monolayer and increases the filtration coefficient in the isolated perfused lung above osmolality controls. Collectively, these findings suggest that PMVECs have a bicarbonate-sensitive cytosolic cAMP pool that disrupts endothelial barrier integrity. These studies could provide an alternative mechanism for the controversial effects of bicarbonate correction of acidosis of acute respiratory distress syndrome patients. PMID:23686854

Label-free quantitative cell division monitoring of endothelial cells by digital holographic microscopy

NASA Astrophysics Data System (ADS)

Kemper, Björn; Bauwens, Andreas; Vollmer, Angelika; Ketelhut, Steffi; Langehanenberg, Patrik; Müthing, Johannes; Karch, Helge; von Bally, Gert

2010-05-01

Digital holographic microscopy (DHM) enables quantitative multifocus phase contrast imaging for nondestructive technical inspection and live cell analysis. Time-lapse investigations on human brain microvascular endothelial cells demonstrate the use of DHM for label-free dynamic quantitative monitoring of cell division of mother cells into daughter cells. Cytokinetic DHM analysis provides future applications in toxicology and cancer research.

Targeting Endothelial Function to Treat Heart Failure with Preserved Ejection Fraction: The Promise of Exercise Training

PubMed Central

Lemmens, Katrien; Vrints, Christiaan J.

2017-01-01

Although the burden of heart failure with preserved ejection fraction (HFpEF) is increasing, there is no therapy available that improves prognosis. Clinical trials using beta blockers and angiotensin converting enzyme inhibitors, cardiac-targeting drugs that reduce mortality in heart failure with reduced ejection fraction (HFrEF), have had disappointing results in HFpEF patients. A new “whole-systems” approach has been proposed for designing future HFpEF therapies, moving focus from the cardiomyocyte to the endothelium. Indeed, dysfunction of endothelial cells throughout the entire cardiovascular system is suggested as a central mechanism in HFpEF pathophysiology. The objective of this review is to provide an overview of current knowledge regarding endothelial dysfunction in HFpEF. We discuss the molecular and cellular mechanisms leading to endothelial dysfunction and the extent, presence, and prognostic importance of clinical endothelial dysfunction in different vascular beds. We also consider implications towards exercise training, a promising therapy targeting system-wide endothelial dysfunction in HFpEF. PMID:28706575

Sirt6 mRNA-incorporated endothelial microparticles (EMPs) attenuates DM patient-derived EMP-induced endothelial dysfunction

PubMed Central

Jing, Tong; Ya-Shu, Kuang; Xue-Jun, Wang; Han-Jing, Hou; Yan, Lai; Yi-An, Yao; Fei, Chen; Xue-Bo, Liu

2017-01-01

Background Endothelial microparticles (EMPs) are small vesicles released by endothelial cells (ECs); they are considered biomarkers for endothelial dysfunction and therapeutic targets in diabetes-related vascular disease. Sirtuins have also been shown to play important roles in diabetes by regulating endothelial dysfunction. However, the effect of sirtuin-incorporated EMPs on their parental ECs remains unknown. Aim The present study aims to investigate the diagnostic value of EMPs in diabetes and detect the protective effects of sirtuin 6 (Sirt6) mRNA -incorporated EMPs on endothelial dysfunction. Methods EMPs were prepared from cultured HUVECs and venous blood from patients with diabetes (n=10) and from healthy volunteers (n=6) after sequential centrifugation. Adv-Sirt6 or Sirt6 siRNA was used to alter Sirt6 expression. EC angiogenesis, inflammatory phenotypes, nitric oxide (NO) formation and eNOS phosphorylation were used to evaluate endothelial dysfunction. Results The levels of EMPs in diabetic patients and high glucose-cultured HUVECs are high, whereas Sirt6 expression in plasma and EMPs is low. EMPs generated from diabetic patients or high glucose-cultured HUVECs increase inflammatory chemokine release and blunt EC angiogenesis. Furthermore, EMPs enriched with Sirt6 mRNA induces EC angiogenesis, increases eNOS phosphorylation and impedes inflammatory chemokine release. Inhibition of Sirt6 mRNA expression in EMPs by siRNA hinders angiogenesis and eNOS phosphorylation but increases cellular inflammation. Conclusion The Sirt6 mRNA-carrying EMPs may ameliorate endothelial dysfunction in diabetic patients. PMID:29371988

Sirt6 mRNA-incorporated endothelial microparticles (EMPs) attenuates DM patient-derived EMP-induced endothelial dysfunction.

PubMed

Jing, Tong; Ya-Shu, Kuang; Xue-Jun, Wang; Han-Jing, Hou; Yan, Lai; Yi-An, Yao; Fei, Chen; Xue-Bo, Liu

2017-12-26

Endothelial microparticles (EMPs) are small vesicles released by endothelial cells (ECs); they are considered biomarkers for endothelial dysfunction and therapeutic targets in diabetes-related vascular disease. Sirtuins have also been shown to play important roles in diabetes by regulating endothelial dysfunction. However, the effect of sirtuin-incorporated EMPs on their parental ECs remains unknown. The present study aims to investigate the diagnostic value of EMPs in diabetes and detect the protective effects of sirtuin 6 ( Sirt6 ) mRNA -incorporated EMPs on endothelial dysfunction. EMPs were prepared from cultured HUVECs and venous blood from patients with diabetes (n=10) and from healthy volunteers (n=6) after sequential centrifugation. Adv- Sirt6 or Sirt6 siRNA was used to alter Sirt6 expression. EC angiogenesis, inflammatory phenotypes, nitric oxide (NO) formation and eNOS phosphorylation were used to evaluate endothelial dysfunction. The levels of EMPs in diabetic patients and high glucose-cultured HUVECs are high, whereas Sirt6 expression in plasma and EMPs is low. EMPs generated from diabetic patients or high glucose-cultured HUVECs increase inflammatory chemokine release and blunt EC angiogenesis. Furthermore, EMPs enriched with Sirt6 mRNA induces EC angiogenesis, increases eNOS phosphorylation and impedes inflammatory chemokine release. Inhibition of Sirt6 mRNA expression in EMPs by siRNA hinders angiogenesis and eNOS phosphorylation but increases cellular inflammation. The Sirt6 mRNA -carrying EMPs may ameliorate endothelial dysfunction in diabetic patients.

High-spatial-resolution mapping of the oxygen concentration in cortical tissue (Conference Presentation)

NASA Astrophysics Data System (ADS)

Jaswal, Rajeshwer S.; Yaseen, Mohammad A.; Fu, Buyin; Boas, David A.; Sakadžic, Sava

2016-03-01

Due to a lack of imaging tools for high-resolution imaging of cortical tissue oxygenation, the detailed maps of the oxygen partial pressure (PO2) around arterioles, venules, and capillaries remain largely unknown. Therefore, we have limited knowledge about the mechanisms that secure sufficient oxygen delivery in microvascular domains during brain activation, and provide some metabolic reserve capacity in diseases that affect either microvascular networks or the regulation of cerebral blood flow (CBF). To address this challenge, we applied a Two-Photon PO2 Microscopy to map PO2 at different depths in mice cortices. Measurements were performed through the cranial window in the anesthetized healthy mice as well as in the mouse models of microvascular dysfunctions. In addition, microvascular morphology was recorded by the two-photon microscopy at the end of each experiment and subsequently segmented. Co-registration of the PO2 measurements and exact microvascular morphology enabled quantification of the tissue PO2 dependence on distance from the arterioles, capillaries, and venules at various depths. Our measurements reveal significant spatial heterogeneity of the cortical tissue PO2 distribution that is dominated by the high oxygenation in periarteriolar spaces. In cases of impaired oxygen delivery due to microvascular dysfunction, significant reduction in tissue oxygenation away from the arterioles was observed. These tissue domains may be the initial sites of cortical injury that can further exacerbate the progression of the disease.

microRNAs as Pharmacological Targets in Endothelial Cell Function and Dysfunction

PubMed Central

Chamorro-Jorganes, Aránzazu; Araldi, Elisa; Suárez, Yajaira

2013-01-01

Endothelial cell dysfunction is a term which implies the dysregulation of normal endothelial cell functions, including impairment of the barrier functions, control of vascular tone, disturbance of proliferative, migratory and morphogenic capacities of endothelial cells, as well as control of leukocyte trafficking. MicroRNAs (miRNAs) are short non-coding RNAs that have emerged as critical regulators of gene expression acting predominantly at the post-transcriptional level. This review summarizes the latest insights in the identification of endothelial-specific miRNAs and their targets, as well as their roles in controlling endothelial cell functions in both autocrine and paracrine manner. In addition, we discuss the therapeutic potential for the treatment of endothelial cell dysfunction and associated vascular pathophysiological conditions. PMID:23603154

Oxidative Stress and Modification of Renal Vascular Permeability Are Associated with Acute Kidney Injury during P. berghei ANKA Infection

PubMed Central

Elias, Rosa Maria; Correa-Costa, Matheus; Barreto, Claudiene Rodrigues; Silva, Reinaldo Correia; Hayashida, Caroline Y.; Castoldi, Ângela; Gonçalves, Giselle Martins; Braga, Tarcio Teodoro; Barboza, Renato; Rios, Francisco José; Keller, Alexandre Castro; Cenedeze, Marcos Antonio; Hyane, Meire Ioshie; D'Império-Lima, Maria Regina; Figueiredo-Neto, Antônio Martins; Reis, Marlene Antônia; Marinho, Cláudio Romero Farias; Pacheco-Silva, Alvaro; Câmara, Niels Olsen Saraiva

2012-01-01

Malaria associated-acute kidney injury (AKI) is associated with 45% of mortality in adult patients hospitalized with severe form of the disease. However, the causes that lead to a framework of malaria-associated AKI are still poorly characterized. Some clinical studies speculate that oxidative stress products, a characteristic of Plasmodium infection, as well as proinflammatory response induced by the parasite are involved in its pathophysiology. Therefore, we aimed to investigate the development of malaria-associated AKI during infection by P. berghei ANKA, with special attention to the role played by the inflammatory response and the involvement of oxidative stress. For that, we took advantage of an experimental model of severe malaria that showed significant changes in the renal pathophysiology to investigate the role of malaria infection in the renal microvascular permeability and tissue injury. Therefore, BALB/c mice were infected with P. berghei ANKA. To assess renal function, creatinine, blood urea nitrogen, and ratio of proteinuria and creatininuria were evaluated. The products of oxidative stress, as well as cytokine profile were quantified in plasma and renal tissue. The change of renal microvascular permeability, tissue hypoxia and cellular apoptosis were also evaluated. Parasite infection resulted in renal dysfunction. Furthermore, we observed increased expression of adhesion molecule, proinflammatory cytokines and products of oxidative stress, associated with a decrease mRNA expression of HO-1 in kidney tissue of infected mice. The measurement of lipoprotein oxidizability also showed a significant increase in plasma of infected animals. Together, our findings support the idea that products of oxidative stress, as well as the immune response against the parasite are crucial to changes in kidney architecture and microvascular endothelial permeability of BALB/c mice infected with P. berghei ANKA. PMID:22952850

Compromised renal microvascular reactivity of angiotensin type 1 double null mice.

PubMed

Park, Sungmi; Bivona, Benjamin J; Harrison-Bernard, Lisa M

2007-07-01

Angiotensin type 1A (AT(1A)) and 1B (AT(1B)) receptor deletion (AT1DKO) results in renal microvascular disease, tubulointerstitial injury, and reduced blood pressure. To test the hypothesis that renal preglomerular responses to angiotensin (ANG) II are mediated by AT(1A) and AT(1B) receptors, experiments were performed in AT1DKO mice using the in vitro blood perfused juxtamedullary nephron technique. Kidneys were harvested from AT1DKO and wild-type (WT) mice and bathed with ANG II (1-100 nM), norepinephrine (NE; 100-1,000 nM), or acetylcholine (ACh; 10 microM). Baseline diameters of afferent (19.5 +/- 0.7 and 13.9 +/- 0.7 microm, n = 17 and 16) and efferent (15.5 +/- 2.1 and 10.8 +/- 1.0 microm, n = 4 and 7) arterioles of AT1DKO were significantly larger than WT. Afferent and efferent arteriolar responses to ANG II, 100, and 300 nM NE were absent in AT1DKO; although significant constriction to 1 microM NE was observed (-17 +/- 5 and -23 +/- 6%, respectively). Afferent arterioles of WT mice dilated significantly in response to ACh (15.1 +/- 0.6 to 17.0 +/- 1.2 microm, n = 6); however, arterioles from AT1DKO tended to contract (19.9 +/- 1.2 to 17.8 +/- 1.6 microm; n = 6, P = 0.06). In summary, loss of ANG II-induced contraction, reduced vasoconstriction to NE, and endothelial cell dysfunction contribute to the renal vascular phenotype of AT1DKO mice. We conclude that ANG II signaling via the AT(1) receptor plays a pivotal role in basal renal microvascular tone and effectiveness to respond to vasoconstrictor and vasodilator agonists.

Methylmercury promotes prostacyclin release from cultured human brain microvascular endothelial cells via induction of cyclooxygenase-2 through activation of the EGFR-p38 MAPK pathway by inhibiting protein tyrosine phosphatase 1B activity.

PubMed

Yoshida, Eiko; Kurita, Masaru; Eto, Komyo; Kumagai, Yoshito; Kaji, Toshiyuki

2017-12-01

Methylmercury is an environmental pollutant that exhibits neurotoxicity when ingested, primarily in the form of neuropathological lesions that localize along deep sulci and fissures, in addition to edematous and inflammatory changes in patient cerebrums. These conditions been known to give rise to a variety of ailments that have come to be collectively termed Minamata disease. Since prostaglandins I 2 and E 2 (PGI 2 and PGE 2 ) increase vascular permeability and contribute to the progression of inflammatory changes, we hypothesize that methylmercury induces the synthesis of these prostaglandins in brain microvascular endothelial cells and pericytes. To test this theory, human brain microvascular endothelial cells and pericytes were cultured and treated with methylmercury, after which the PGI 2 and PGE 2 released from endothelial cells and/or pericytes were quantified by enzyme-linked immunosorbent assay while protein and mRNA expressions in endothelial cells were analyzed by western blot analysis and real-time reverse transcription polymerase chain reaction, respectively. Experimental results indicate that methylmercury inhibits the activity of protein tyrosine phosphatase 1B, which in turn activates the epidermal growth factor receptor-p38 mitogen-activated protein kinase pathway that induces cyclooxygenase-2 expression. It was also found that the cyclic adenosine 3',5'-monophosphate pathway, which can be activated by PGI 2 and PGE 2 , is involved in methylmercury-induced cyclooxygenase-2 expression. Since it appears that protein tyrosine phosphatase 1 B serves as a sensor protein for methylmercury in these mechanisms, it is our belief that the results of the present study may provide additional insights into the molecular mechanisms responsible for edematous and inflammatory changes in the cerebrum of patients with Minamata disease. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

VEGF isoforms have differential effects on permeability of human pulmonary microvascular endothelial cells.

PubMed

Ourradi, Khadija; Blythe, Thomas; Jarrett, Caroline; Barratt, Shaney L; Welsh, Gavin I; Millar, Ann B

2017-06-02

Alternative splicing of Vascular endothelial growth factor-A mRNA transcripts (commonly referred as VEGF) leads to the generation of functionally differing isoforms, the relative amounts of which have potentially significant physiological outcomes in conditions such as acute respiratory distress syndrome (ARDS). The effect of such isoforms on pulmonary vascular permeability is unknown. We hypothesised that VEGF 165 a and VEGF 165 b isoforms would have differing effects on pulmonary vascular permeability caused by differential activation of intercellular signal transduction pathways. To test this hypothesis we investigated the physiological effect of VEGF 165 a and VEGF 165 b on Human Pulmonary Microvascular Endothelial Cell (HPMEC) permeability using three different methods: trans-endothelial electrical resistance (TEER), Electric cell-substrate impedance sensing (ECIS) and FITC-BSA passage. In addition, potential downstream signalling pathways of the VEGF isoforms were investigated by Western blotting and the use of specific signalling inhibitors. VEGF 165 a increased HPMEC permeability using all three methods (paracellular and transcellular) and led to associated VE-cadherin and actin stress fibre changes. In contrast, VEGF 165 b decreased paracellular permeability and did not induce changes in VE-cadherin cell distribution. Furthermore, VEGF 165 a and VEGF 165 b had differing effects on both the phosphorylation of VEGF receptors and downstream signalling proteins pMEK, p42/44MAPK, p38 MAPK, pAKT and peNOS. Interestingly specific inhibition of the pMEK, p38 MAPK, PI3 kinase and eNOS pathways blocked the effects of both VEGF 165 a and VEGF 165 b on paracellular permeability and the effect of VEGF 165 a on proliferation/migration, suggesting that this difference in cellular response is mediated by an as yet unidentified signalling pathway(s). This study demonstrates that the novel isoform VEGF 165 a and VEGF 165 b induce differing effects on permeability in pulmonary microvascular endothelial cells.

Curcumin inhibits VEGF-mediated angiogenesis in human intestinal microvascular endothelial cells through COX-2 and MAPK inhibition.

PubMed

Binion, D G; Otterson, M F; Rafiee, P

2008-11-01

Angiogenesis, the growth of new blood vessels, is a critical homeostatic mechanism which regulates vascular populations in response to physiological requirements and pathophysiological demand, including chronic inflammation and cancer. The importance of angiogenesis in gastrointestinal chronic inflammation and cancer has been defined, as antiangiogenic therapy has demonstrated benefit in models of inflammatory bowel disease and colon cancer treatment. Curcumin is a natural product undergoing evaluation for the treatment of chronic inflammation, including inflammatory bowel disease (IBD). The effect of curcumin on human intestinal angiogenesis is not defined. The antiangiogenic effect of curcumin on in vitro angiogenesis was examined using primary cultures of human intestinal microvascular endothelial cells (HIMECs), stimulated with vascular endothelial growth factor (VEGF). Curcumin inhibited proliferation, cell migration and tube formation in HIMECs induced by VEGF. Activation of HIMECs by VEGF resulted in enhanced expression of cyclo-oxygenase-2 (COX-2) mRNA, protein and prostaglandin E(2) (PGE(2)) production. Pretreatment of HIMECs with 10 microM curcumin as well as 1 microM NS398, a selective inhibitor of COX-2, resulted in inhibition of COX-2 at the mRNA and protein level and PGE(2) production. Similarly COX-2 expression in HIMECs was significantly inhibited by Jun N-terminal kinase (JNK; SP600125) and p38 mitogen-activated protein kinase (MAPK; SB203580) inhibitors and was reduced by p44/42 MAPK inhibitor (PD098059). Taken together, these data demonstrate an important role for COX-2 in the regulation of angiogenesis in HIMECs via MAPKs. Moreover, curcumin inhibits microvascular endothelial cell angiogenesis through inhibition of COX-2 expression and PGE(2) production, suggesting that this natural product possesses antiangiogenic properties, which warrants further investigation as adjuvant treatment of IBD and cancer.

Rosetting Plasmodium falciparum-infected erythrocytes bind to human brain microvascular endothelial cells in vitro, demonstrating a dual adhesion phenotype mediated by distinct P. falciparum erythrocyte membrane protein 1 domains.

PubMed

Adams, Yvonne; Kuhnrae, Pongsak; Higgins, Matthew K; Ghumra, Ashfaq; Rowe, J Alexandra

2014-03-01

Adhesion interactions between Plasmodium falciparum-infected erythrocytes (IE) and human cells underlie the pathology of severe malaria. IE cytoadhere to microvascular endothelium or form rosettes with uninfected erythrocytes to survive in vivo by sequestering IE in the microvasculature and avoiding splenic clearance mechanisms. Both rosetting and cytoadherence are mediated by the parasite-derived IE surface protein family Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1). Rosetting and cytoadherence have been widely studied as separate entities; however, the ability of rosetting P. falciparum strains to cytoadhere has received little attention. Here, we show that IE of the IT/R29 strain expressing a rosette-mediating PfEMP1 variant (IT4var09) cytoadhere in vitro to a human brain microvascular endothelial cell line (HBEC-5i). Cytoadherence was inhibited by heparin and by treatment of HBEC-5i with heparinase III, suggesting that the endothelial receptors for IE binding are heparan sulfate proteoglycans. Antibodies to the N-terminal regions of the IT4var09 PfEMP1 variant (NTS-DBL1α and DBL2γ domains) specifically inhibited and reversed cytoadherence down to low concentrations (<10 μg/ml of total IgG). Surface plasmon resonance experiments showed that the NTS-DBLα and DBL2γ domains bind strongly to heparin, with half-maximal binding at a concentration of ∼0.5 μM in both cases. Therefore, cytoadherence of IT/R29 IE is distinct from rosetting, which is primarily mediated by NTS-DBL1α interactions with complement receptor 1. These data show that IT4var09-expressing parasites are capable of dual interactions with both endothelial cells and uninfected erythrocytes via distinct receptor-ligand interactions.

Arginase Inhibitor in the Pharmacological Correction of Endothelial Dysfunction

PubMed Central

Pokrovskiy, Mihail V.; Korokin, Mihail V.; Tsepeleva, Svetlana A.; Pokrovskaya, Tatyana G.; Gureev, Vladimir V.; Konovalova, Elena A.; Gudyrev, Oleg S.; Kochkarov, Vladimir I.; Korokina, Liliya V.; Dudina, Eleonora N.; Babko, Anna V.; Terehova, Elena G.

2011-01-01

This paper is about a way of correction of endothelial dysfunction with the inhibitor of arginase: L-norvaline. There is an imbalance between vasoconstriction and vasodilatation factors of endothelium on the basis of endothelial dysfunction. Among vasodilatation agents, nitrogen oxide plays the basic role. Amino acid L-arginine serves as a source of molecules of nitrogen oxide in an organism. Because of the high activity of arginase enzyme which catalyzes the hydrolysis of L-arginine into ornithine and urea, the bioavailability of nitrogen oxide decreases. The inhibitors of arginase suppress the activity of the given enzyme, raising and production of nitrogen oxide, preventing the development of endothelial dysfunction. PMID:21747978

Silver nanoparticles induce tight junction disruption and astrocyte neurotoxicity in a rat blood-brain barrier primary triple coculture model.

PubMed

Xu, Liming; Dan, Mo; Shao, Anliang; Cheng, Xiang; Zhang, Cuiping; Yokel, Robert A; Takemura, Taro; Hanagata, Nobutaka; Niwa, Masami; Watanabe, Daisuke

2015-01-01

Silver nanoparticles (Ag-NPs) can enter the brain and induce neurotoxicity. However, the toxicity of Ag-NPs on the blood-brain barrier (BBB) and the underlying mechanism(s) of action on the BBB and the brain are not well understood. To investigate Ag-NP suspension (Ag-NPS)-induced toxicity, a triple coculture BBB model of rat brain microvascular endothelial cells, pericytes, and astrocytes was established. The BBB permeability and tight junction protein expression in response to Ag-NPS, NP-released Ag ions, and polystyrene-NP exposure were investigated. Ultrastructural changes of the microvascular endothelial cells, pericytes, and astrocytes were observed using transmission electron microscopy (TEM). Global gene expression of astrocytes was measured using a DNA microarray. A triple coculture BBB model of primary rat brain microvascular endothelial cells, pericytes, and astrocytes was established, with the transendothelial electrical resistance values >200 Ω·cm(2). After Ag-NPS exposure for 24 hours, the BBB permeability was significantly increased and expression of the tight junction (TJ) protein ZO-1 was decreased. Discontinuous TJs were also observed between microvascular endothelial cells. After Ag-NPS exposure, severe mitochondrial shrinkage, vacuolations, endoplasmic reticulum expansion, and Ag-NPs were observed in astrocytes by TEM. Global gene expression analysis showed that three genes were upregulated and 20 genes were downregulated in astrocytes treated with Ag-NPS. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that the 23 genes were associated with metabolic processes, biosynthetic processes, response to stimuli, cell death, the MAPK pathway, and so on. No GO term and KEGG pathways were changed in the released-ion or polystyrene-NP groups. Ag-NPS inhibited the antioxidant defense of the astrocytes by increasing thioredoxin interacting protein, which inhibits the Trx system, and decreasing Nr4a1 and Dusp1. Meanwhile, Ag-NPS induced inflammation and apoptosis through modulation of the MAPK pathway or B-cell lymphoma-2 expression or mTOR activity in astrocytes. These results draw our attention to the importance of Ag-NP-induced toxicity on the neurovascular unit and provide a better understanding of its toxicological mechanisms on astrocytes.

Functional Differences Between Placental Micro- and Macrovascular Endothelial Colony-Forming Cells

PubMed Central

Solomon, Ioana; O’Reilly, Megan; Ionescu, Lavinia; Alphonse, Rajesh S.; Rajabali, Saima; Zhong, Shumei; Vadivel, Arul; Shelley, W. Chris; Yoder, Mervin C.

2016-01-01

Alterations in the development of the placental vasculature can lead to pregnancy complications, such as preeclampsia. Currently, the cause of preeclampsia is unknown, and there are no specific prevention or treatment strategies. Further insight into the placental vasculature may aid in identifying causal factors. Endothelial colony-forming cells (ECFCs) are a subset of endothelial progenitor cells capable of self-renewal and de novo vessel formation in vitro. We hypothesized that ECFCs exist in the micro- and macrovasculature of the normal, term human placenta. Human placentas were collected from term pregnancies delivered by cesarean section (n = 16). Placental micro- and macrovasculature was collected from the maternal and fetal side of the placenta, respectively, and ECFCs were isolated and characterized. ECFCs were CD31+, CD105+, CD144+, CD146+, CD14−, and CD45−, took up 1,1′-dioctadecyl-3,3,3′,3′-tetramethyl-indocarbocyanine perchlorate-labeled acetylated low-density lipoprotein, and bound Ulex europaeus agglutinin 1. In vitro, macrovascular ECFCs had a greater potential to generate high-proliferative colonies and formed more complex capillary-like networks on Matrigel compared with microvascular ECFCs. In contrast, in vivo assessment demonstrated that microvascular ECFCs had a greater potential to form vessels. Macrovascular ECFCs were of fetal origin, whereas microvascular ECFCs were of maternal origin. ECFCs exist in the micro- and macrovasculature of the normal, term human placenta. Although macrovascular ECFCs demonstrated greater vessel and colony-forming potency in vitro, this did not translate in vivo, where microvascular ECFCs exhibited a greater vessel-forming ability. These important findings contribute to the current understanding of normal placental vascular development and may aid in identifying factors involved in preeclampsia and other pregnancy complications. Significance This research confirms that resident endothelial colony-forming cells (ECFCs) exist in the micro- and macrovasculature of the normal, term human placenta. Their isolation from two different anatomical locations yields two functionally different ECFC populations. Investigation of these ECFC populations during placental pathologies, such as preeclampsia, may lead to a better understanding of the disease process and aid in developing new therapies. PMID:26819255

C5a induces caspase-dependent apoptosis in brain vascular endothelial cells in experimental lupus.

PubMed

Mahajan, Supriya D; Tutino, Vincent M; Redae, Yonas; Meng, Hui; Siddiqui, Adnan; Woodruff, Trent M; Jarvis, James N; Hennon, Teresa; Schwartz, Stanley; Quigg, Richard J; Alexander, Jessy J

2016-08-01

Blood-brain barrier (BBB) dysfunction complicates central nervous system lupus, an important aspect of systemic lupus erythematosus. To gain insight into the underlying mechanism, vascular corrosion casts of brain were generated from the lupus mouse model, MRL/lpr mice and the MRL/MpJ congenic controls. Scanning electron microscopy of the casts showed loss of vascular endothelial cells in lupus mice compared with controls. Immunostaining revealed a significant increase in caspase 3 expression in the brain vascular endothelial cells, which suggests that apoptosis could be an important mechanism causing cell loss, and thereby loss of BBB integrity. Complement activation occurs in lupus resulting in increased generation of circulating C5a, which caused the endothelial layer to become 'leaky'. In this study, we show that C5a and lupus serum induced apoptosis in cultured human brain microvascular endothelial cells (HBMVECs), whereas selective C5a receptor 1 (C5aR1) antagonist reduced apoptosis in these cells, demonstrating C5a/C5aR1-dependence. Gene expression of initiator caspases, caspase 1 and caspase 8, and pro-apoptotic proteins death-associated protein kinase 1, Fas-associated protein (FADD), cell death-inducing DNA fragmentation factor 45 000 MW subunit A-like effector B (CIDEB) and BCL2-associated X protein were increased in HBMVECs treated with lupus serum or C5a, indicating that both the intrinsic and extrinsic apoptotic pathways could be critical mediators of brain endothelial cell apoptosis in this setting. Overall, our findings suggest that C5a/C5aR1 signalling induces apoptosis through activation of FADD, caspase 8/3 and CIDEB in brain endothelial cells in lupus. Further elucidation of the underlying apoptotic mechanisms mediating the reduced endothelial cell number is important in establishing the potential therapeutic effectiveness of C5aR1 inhibition that could prevent and/or reduce BBB alterations and preserve the physiological function of BBB in central nervous system lupus. © 2016 John Wiley & Sons Ltd.

Physical activity on endothelial and erectile dysfunction: a literature review.

PubMed

Leoni, Luís Antônio B; Fukushima, André R; Rocha, Leandro Y; Maifrino, Laura B M M; Rodrigues, Bruno

2014-09-01

Physical inactivity, diabetes, hypertension, dyslipidemia, smoking and obesity were associated with imbalance in oxidative stress, leading to endothelial dysfunction. Such dysfunction is present in both cardiovascular disease (CVD) and erectile dysfunction (ED). ED is the persistent inability to achieve or sustain an erection sufficient for satisfactory sexual performance and is one of the first manifestations of endothelial damage in men with CVD risk factors. The purpose of this article is to review the results of studies involving physical activity, CVD, endothelial dysfunction and ED in order to verify its applicability for improving the health and quality of life of men with such disorders. There is consistent evidence that endothelial damage is intimately linked to ED, and this manifestation seems to be associated with the appearance CVDs. On the other hand, physical activity has been pointed out as an important clinical strategy in the prevention and treatment of CVDs and ED mainly associated with improvement of endothelial function. However, further experimental and clinical prospective investigations are needed to test the role of physical exercises in the modulation of endothelial function and their implications on erectile function and the appearance of CVDs.

Age-related changes in endothelial function and blood flow regulation.

PubMed

Toda, Noboru

2012-02-01

Vascular endothelial dysfunction is regarded as a primary phenotypic expression of normal human aging. This senescence-induced disorder is the likely culprit underlying the increased cardiovascular and metabolic disease risks associated with aging. The rate of this age-dependent deterioration is largely influenced by the poor-quality lifestyle choice, such as smoking, sedentary daily life, chronic alcohol ingestion, high salt intake, unbalanced diet, and mental stress; and it is accelerated by cardiovascular and metabolic diseases. Although minimizing these detrimental factors is the best course of action, nonetheless chronological age steadily impairs endothelial function through reduced endothelial nitric oxide synthase (eNOS) expression/action, accelerated nitric oxide (NO) degradation, increased phosphodiesterase activity, inhibition of NOS activity by endogenous NOS inhibitors, increased production of reactive oxygen species, inflammatory reactions, decreased endothelial progenitor cell number and function, and impaired telomerase activity or telomere shortening. Endothelial dysfunction in regional vasculatures results in cerebral hypoperfusion triggering cognitive dysfunction and Alzheimer's disease, coronary artery insufficiency, penile erectile dysfunction, and circulatory failures in other organs and tissues. Possible prophylactic measures to minimize age-related endothelial dysfunction are also summarized in this review. Copyright © 2011 Elsevier Inc. All rights reserved.

Vascular Endothelial Growth Factor (VEGF) and Platelet (PF-4) Factor 4 Inputs Modulate Human Microvascular Endothelial Signaling in a Three-Dimensional Matrix Migration Context*

PubMed Central

Hang, Ta-Chun; Tedford, Nathan C.; Reddy, Raven J.; Rimchala, Tharathorn; Wells, Alan; White, Forest M.; Kamm, Roger D.; Lauffenburger, Douglas A.

2013-01-01

The process of angiogenesis is under complex regulation in adult organisms, particularly as it often occurs in an inflammatory post-wound environment. As such, there are many impacting factors that will regulate the generation of new blood vessels which include not only pro-angiogenic growth factors such as vascular endothelial growth factor, but also angiostatic factors. During initial postwound hemostasis, a large initial bolus of platelet factor 4 is released into localized areas of damage before progression of wound healing toward tissue homeostasis. Because of its early presence and high concentration, the angiostatic chemokine platelet factor 4, which can induce endothelial anoikis, can strongly affect angiogenesis. In our work, we explored signaling crosstalk interactions between vascular endothelial growth factor and platelet factor 4 using phosphotyrosine-enriched mass spectrometry methods on human dermal microvascular endothelial cells cultured under conditions facilitating migratory sprouting into collagen gel matrices. We developed new methods to enable mass spectrometry-based phosphorylation analysis of primary cells cultured on collagen gels, and quantified signaling pathways over the first 48 h of treatment with vascular endothelial growth factor in the presence or absence of platelet factor 4. By observing early and late signaling dynamics in tandem with correlation network modeling, we found that platelet factor 4 has significant crosstalk with vascular endothelial growth factor by modulating cell migration and polarization pathways, centered around P38α MAPK, Src family kinases Fyn and Lyn, along with FAK. Interestingly, we found EphA2 correlational topology to strongly involve key migration-related signaling nodes after introduction of platelet factor 4, indicating an influence of the angiostatic factor on this ambiguous but generally angiogenic signal in this complex environment. PMID:24023389

Glutathione adducts on sarcoplasmic/endoplasmic reticulum Ca2+ ATPase Cys-674 regulate endothelial cell calcium stores and angiogenic function as well as promote ischemic blood flow recovery.

PubMed

Thompson, Melissa D; Mei, Yu; Weisbrod, Robert M; Silver, Marcy; Shukla, Praphulla C; Bolotina, Victoria M; Cohen, Richard A; Tong, Xiaoyong

2014-07-18

The sarco/endoplasmic reticulum Ca(2+) ATPase (SERCA) is key to Ca(2+) homeostasis and is redox-regulated by reversible glutathione (GSH) adducts on the cysteine (C) 674 thiol that stimulate Ca(2+) uptake activity and endothelial cell angiogenic responses in vitro. We found that mouse hind limb muscle ischemia induced S-glutathione adducts on SERCA in both whole muscle tissue and endothelial cells. To determine the role of S-glutathiolation, we used a SERCA 2 C674S heterozygote knock-in (SKI) mouse lacking half the key thiol. Following hind limb ischemia, SKI animals had decreased SERCA S-glutathione adducts and impaired blood flow recovery. We studied SKI microvascular endothelial cells in which total SERCA 2 expression was unchanged. Cultured SKI microvascular endothelial cells showed impaired migration and network formation compared with wild type (WT). Ca(2+) studies showed decreased nitric oxide (·NO)-induced (45)Ca(2+) uptake into the endoplasmic reticulum (ER) of SKI cells, while Fura-2 studies revealed lower Ca(2+) stores and decreased vascular endothelial growth factor (VEGF)- and ·NO-induced Ca(2+) influx. Adenoviral overexpression of calreticulin, an ER Ca(2+) binding protein, increased ionomycin-releasable stores, VEGF-induced Ca(2+) influx and endothelial cell migration. Taken together, these data indicate that the redox-sensitive Cys-674 thiol on SERCA 2 is required for normal endothelial cell Ca(2+) homeostasis and ischemia-induced angiogenic responses, revealing a novel redox control of angiogenesis via Ca(2+) stores. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Hydroquinone stimulates inflammatory functions in microvascular endothelial cells via NF-κB nuclear activation.

PubMed

Hebeda, Cristina Bichels; Pinedo, Fernanda Júdice; Vinolo, Marco Aurélio Ramirez; Curi, Rui; Farsky, Sandra Helena Poliselli

2011-11-01

Hydroquinone impairs several leucocyte cell functions, which alter the immune response. Although endothelial cell functions are important for the development of immune responses, hydroquinone actions on endothelial cell have not been shown. Therefore, the effect of hydroquinone exposure (10 or 100 μM for 2 hr) on primary culture of microvascular endothelial cells (PMECs) obtained from the cremaster muscle of Wistar rats incubated in the presence or absence of lipopolysaccharide (LPS, 2 μg/mL) was investigated. Hydroquinone treatment induced the membrane expression of cell adhesion molecules (CAMs) from the immunoglobulin superfamilies ICAM-1 (intercellular), VCAM-1(vascular) and PECAM-1 (platelet endothelial) and induced the secretion of cytokines interleukin-1β (IL-1β) and tumour necrosis factor-α (TNF-α). The effects were dependent on transcriptional modifications because enhanced CAM mRNA expression as well as both cytokines and nuclear factor κB (NF-κB) nuclear activation was found. These effects may be due to the direct action of hydroquinone rather than its quinone metabolites, because endothelial cells do not present myeloperoxidase enzyme and hydroquinone incubation did not induce the expression of cytochrome P450 2E1 (CYP2E1) or prostaglandin H synthase 1. In addition, the incubation of endothelial cells with benzoquinone (10 μM, 2 hr) impaired PECAM-1 expression and did not modify NF-κB nuclear activation. Taken together, the data herein presented reveal that hydroquinone evokes pro-inflammatory properties in endothelial cells that are triggered by the enhancement of NF-κB nuclear translocation-dependent gene transcription. © 2011 The Authors. Basic & Clinical Pharmacology & Toxicology © 2011 Nordic Pharmacological Society.

Flavanol-rich cocoa ameliorates lipemia-induced endothelial dysfunction.

PubMed

Westphal, Sabine; Luley, Claus

2011-09-01

Consumption of flavanols improves chronic endothelial dysfunction. We investigated whether it can also improve acute lipemia-induced endothelial dysfunction. In this randomized, placebo-controlled, double-blind, crossover trial, 18 healthy subjects received a fatty meal with cocoa either rich in flavanols (918 mg) or flavanol-poor. Flow-mediated dilation (FMD), triglycerides, and free fatty acids were then determined over 6 h. After the flavanol-poor fat loading, the FMD deteriorated over 4 h. The consumption of flavanol-rich cocoa, in contrast, improved this deterioration in hours 2, 3, and 4 without abolishing it completely. Flavanols did not have any influence on triglycerides or on free fatty acids. Flavanol-rich cocoa can alleviate the lipemia-induced endothelial dysfunction, probably through an improvement in endothelial NO synthase.

[The severity of gestational diabetes mellitus affects microvascular dysfunction measured three years after pregnancy that may be related to increased oxidative stress].

PubMed

Horváth, Eszter Mária; Mágenheim, Rita; Domján, Beatrix Annamária; Ferencz, Viktória; Tänczer, Tímea; Szabó, Eszter; Benkő, Rita; Szabó, Csaba; Tabák, Ádám; Somogyi, Anikó

2015-11-22

Oxidative-nitrative stress and poly(ADP-ribose) polymerase activation observed in gestational diabetes may play role in the increased cardiovascular risk in later life. The present study aimed to examine the influence of the severity of previous gestational diabetes (insulin need) on vascular function three years after delivery. Furthermore, the authors investigated the relation of vascular function with oxidative-nitrative stress and poly(ADP-ribose) polymerase activation. Macrovascular function was measured by applanation tonometry; microvascular reactivity was assessed by provocation tests during Laser-Doppler flowmetry in 40 women who had gestational diabetes 3 years before the study. Oxidative-nitrative stress and poly(ADP-ribose) polymerase activity in blood components were determined by colorimetry and immunohistochemistry. Three years after insulin treated gestational diabetes impaired microvascular function and increased oxidative stress was observed compared to mild cases. The severity of previous gestational diabetes affects microvascular dysfunction that is accompanied by elevated oxidative stress. Nitrative stress and poly(ADP-ribose) polymerase activity correlates with certain vascular factors not related to the severity of the disease.

A positive circuit of VEGF increases Glut-1 expression by increasing HIF-1α gene expression in human retinal endothelial cells.

PubMed

Choi, Yoon Kyung

2017-12-01

Treatment of human retinal microvascular endothelial cells (HRMECs) with vascular endothelial growth factor 165 (VEGF 165 ) increased hypoxia-inducible factor 1α (HIF-1α), VEGF, and glucose transporter 1 (Glut-1) mRNA expression and Glut-1 protein localization to the membrane. In contrast, treatment of human retinal pigment epithelium cells with VEGF 165 did not induce HIF-1α, VEGF, and Glut-1 gene expression. Microvascular endothelial cells are surrounded by astrocytic end feet in the retina. Astrocyte-derived A-kinase anchor protein 12 overexpression during hypoxia downregulated VEGF secretion, and this conditioned medium reduced VEGF and Glut-1 expression in HRMECs, suggesting that communications between astrocytes and endothelial cells may be the determinants of the blood vessel network. In HRMECs, HIF-1α small interfering RNA transfection blocked the VEGF 165 -mediated increase in VEGF and Glut-1 gene expression. Inhibition of protein kinase C (PKC) with inhibitor GF109203X or with a small interfering RNA targeting PKCζ attenuated the VEGF 165 -induced Glut-1 protein expression and VEGF and Glut-1 mRNA expression. In addition, results of an immunoprecipitation assay imply an interaction between VEGF receptor 2 (VEGFR2) and PKCζ in HRMECs. Therefore, VEGF secretion by hypoxic astrocytes may upregulate HIF-1α gene expression, inducing VEGF and Glut-1 expression via the VEGFR2-PKCζ axis in HRMECs.

A physiological model for interpretation of arterial spin labeling reactive hyperemia of calf muscles.

PubMed

Chen, Hou-Jen; Wright, Graham A

2017-01-01

To characterize and interpret arterial spin labeling (ASL) reactive hyperemia of calf muscles for a better understanding of the microcirculation in peripheral arterial disease (PAD), we present a physiological model incorporating oxygen transport, tissue metabolism, and vascular regulation mechanisms. The model demonstrated distinct effects between arterial stenoses and microvascular dysfunction on reactive hyperemia, and indicated a higher sensitivity of 2-minute thigh cuffing to microvascular dysfunction than 5-minute cuffing. The recorded perfusion responses in PAD patients (n = 9) were better differentiated from the normal subjects (n = 7) using the model-based analysis rather than characterization using the apparent peak and time-to-peak of the responses. The analysis results suggested different amounts of microvascular disease within the patient group. Overall, this work demonstrates a novel analysis method and facilitates understanding of the physiology involved in ASL reactive hyperemia. ASL reactive hyperemia with model-based analysis may be used as a noninvasive microvascular assessment in the presence of arterial stenoses, allowing us to look beyond the macrovascular disease in PAD. A subgroup who will have a poor prognosis after revascularization in the patients with critical limb ischemia may be associated with more severe microvascular diseases, which may potentially be identified using ASL reactive hyperemia.

Nitric Oxide-Mediated Coronary Flow Regulation in Patients with Coronary Artery Disease: Recent Advances

PubMed Central

Toda, Noboru; Tanabe, Shinichi; Nakanishi, Sadanobu

2011-01-01

Nitric oxide (NO) formed via endothelial NO synthase (eNOS) plays crucial roles in the regulation of coronary blood flow through vasodilatation and decreased vascular resistance, and in inhibition of platelet aggregation and adhesion, leading to the prevention of coronary circulatory failure, thrombosis, and atherosclerosis. Endothelial function is impaired by several pathogenic factors including smoking, chronic alcohol intake, hypercholesterolemia, obesity, hyperglycemia, and hypertension. The mechanisms underlying endothelial dysfunction include reduced NO synthase (NOS) expression and activity, decreased NO bioavailability, and increased production of oxygen radicals and endogenous NOS inhibitors. Atrial fibrillation appears to be a risk factor for endothelial dysfunction. Endothelial dysfunction is an important predictor of coronary artery disease (CAD) in humans. Penile erectile dysfunction, associated with impaired bioavailability of NO produced by eNOS and neuronal NOS, is also considered to be highly predictive of ischemic heart disease. There is evidence suggesting an important role of nitrergic innervation in coronary blood flow regulation. Prophylactic and therapeutic measures to eliminate pathogenic factors inducing endothelial and nitrergic nerve dysfunction would be quite important in preventing the genesis and development of CAD. PMID:22942627

A nanobiosensor for dynamic single cell analysis during microvascular self-organization.

PubMed

Wang, S; Sun, J; Zhang, D D; Wong, P K

2016-10-14

The formation of microvascular networks plays essential roles in regenerative medicine and tissue engineering. Nevertheless, the self-organization mechanisms underlying the dynamic morphogenic process are poorly understood due to a paucity of effective tools for mapping the spatiotemporal dynamics of single cell behaviors. By establishing a single cell nanobiosensor along with live cell imaging, we perform dynamic single cell analysis of the morphology, displacement, and gene expression during microvascular self-organization. Dynamic single cell analysis reveals that endothelial cells self-organize into subpopulations with specialized phenotypes to form microvascular networks and identifies the involvement of Notch1-Dll4 signaling in regulating the cell subpopulations. The cell phenotype correlates with the initial Dll4 mRNA expression level and each subpopulation displays a unique dynamic Dll4 mRNA expression profile. Pharmacological perturbations and RNA interference of Notch1-Dll4 signaling modulate the cell subpopulations and modify the morphology of the microvascular network. Taken together, a nanobiosensor enables a dynamic single cell analysis approach underscoring the importance of Notch1-Dll4 signaling in microvascular self-organization.

Perturbations in Endothelial Dysfunction-Associated Pathways in the Nitrofen-Induced Congenital Diaphragmatic Hernia Model.

PubMed

Zhaorigetu, Siqin; Bair, Henry; Lu, Jonathan; Jin, Di; Olson, Scott D; Harting, Matthew T

2018-01-01

Although it is well known that nitrofen induces congenital diaphragmatic hernia (CDH), including CDH-associated lung hypoplasia and pulmonary hypertension (PH) in rodents, the mechanism of pathogenesis remains largely unclear. It has been reported that pulmonary artery (PA) endothelial cell (EC) dysfunction contributes to the development of PH in CDH. Thus, we hypothesized that there is significant alteration of endothelial dysfunction-associated proteins in nitrofen-induced CDH PAs. Pregnant SD rats received either nitrofen or olive oil on gestational day 9.5. The newborn rats were sacrificed and divided into a CDH (n = 81) and a control (n = 23) group. After PA isolation, the expression of PA endothelial dysfunction-associated proteins was assessed on Western blot and immunostaining. We demonstrate that the expression of C-reactive protein and endothelin-1 and its receptors, ETA and ETB, were significantly increased in the CDH PAs. Levels of phosphorylated myosin light chain were significantly elevated, but those of phosphorylated endothelial nitric oxide synthase, caveolin-1, and mechanistic target of rapamycin were significantly decreased in the CDH PAs. In this work, we elucidate alterations in the expression of endothelial dysfunction-associated proteins specific to nitrofen-induced CDH rodent PAs, thereby advancing our understanding of the critical role of endothelial dysfunction-associated pathways in the pathogenesis of nitrofen-induced CDH. © 2017 S. Karger AG, Basel.

Endothelial mineralocorticoid receptor activation mediates endothelial dysfunction in diet-induced obesity.

PubMed

Schäfer, Nicola; Lohmann, Christine; Winnik, Stephan; van Tits, Lambertus J; Miranda, Melroy X; Vergopoulos, Athanasios; Ruschitzka, Frank; Nussberger, Jürg; Berger, Stefan; Lüscher, Thomas F; Verrey, François; Matter, Christian M

2013-12-01

Aldosterone plays a crucial role in cardiovascular disease. 'Systemic' inhibition of its mineralocorticoid receptor (MR) decreases atherosclerosis by reducing inflammation and oxidative stress. Obesity, an important cardiovascular risk factor, is an inflammatory disease associated with increased plasma aldosterone levels. We have investigated the role of the 'endothelial' MR in obesity-induced endothelial dysfunction, the earliest stage in atherogenesis. C57BL/6 mice were exposed to a normal chow diet (ND) or a high-fat diet (HFD) alone or in combination with the MR antagonist eplerenone (200 mg/kg/day) for 14 weeks. Diet-induced obesity impaired endothelium-dependent relaxation in response to acetylcholine, whereas eplerenone treatment of obese mice prevented this. Expression analyses in aortic endothelial cells isolated from these mice revealed that eplerenone attenuated expression of pro-oxidative NADPH oxidase (subunits p22phox, p40phox) and increased expression of antioxidative genes (glutathione peroxidase-1, superoxide dismutase-1 and -3) in obesity. Eplerenone did not affect obesity-induced upregulation of cyclooxygenase (COX)-1 or prostacyclin synthase. Endothelial-specific MR deletion prevented endothelial dysfunction in obese (exhibiting high 'endogenous' aldosterone) and in 'exogenous' aldosterone-infused lean mice. Pre-incubation of aortic rings from aldosterone-treated animals with the COX-inhibitor indomethacin restored endothelial function. Exogenous aldosterone administration induced endothelial expression of p22phox in the presence, but not in the absence of the endothelial MR. Obesity-induced endothelial dysfunction depends on the 'endothelial' MR and is mediated by an imbalance of oxidative stress-modulating mechanisms. Therefore, MR antagonists may represent an attractive therapeutic strategy in the increasing population of obese patients to decrease vascular dysfunction and subsequent atherosclerotic complications.

Obesity-Induced Endoplasmic Reticulum Stress Causes Lung Endothelial Dysfunction and Promotes Acute Lung Injury.

PubMed

Shah, Dilip; Romero, Freddy; Guo, Zhi; Sun, Jianxin; Li, Jonathan; Kallen, Caleb B; Naik, Ulhas P; Summer, Ross

2017-08-01

Obesity is a significant risk factor for acute respiratory distress syndrome. The mechanisms underlying this association are unknown. We recently showed that diet-induced obese mice exhibit pulmonary vascular endothelial dysfunction, which is associated with enhanced susceptibility to LPS-induced acute lung injury. Here, we demonstrate that lung endothelial dysfunction in diet-induced obese mice coincides with increased endoplasmic reticulum (ER) stress. Specifically, we observed enhanced expression of the major sensors of misfolded proteins, including protein kinase R-like ER kinase, inositol-requiring enzyme α, and activating transcription factor 6, in whole lung and in primary lung endothelial cells isolated from diet-induced obese mice. Furthermore, we found that primary lung endothelial cells exposed to serum from obese mice, or to saturated fatty acids that mimic obese serum, resulted in enhanced expression of markers of ER stress and the induction of other biological responses that typify the lung endothelium of diet-induced obese mice, including an increase in expression of endothelial adhesion molecules and a decrease in expression of endothelial cell-cell junctional proteins. Similar changes were observed in lung endothelial cells and in whole-lung tissue after exposure to tunicamycin, a compound that causes ER stress by blocking N-linked glycosylation, indicating that ER stress causes endothelial dysfunction in the lung. Treatment with 4-phenylbutyric acid, a chemical protein chaperone that reduces ER stress, restored vascular endothelial cell expression of adhesion molecules and protected against LPS-induced acute lung injury in diet-induced obese mice. Our work indicates that fatty acids in obese serum induce ER stress in the pulmonary endothelium, leading to pulmonary endothelial cell dysfunction. Our work suggests that reducing protein load in the ER of pulmonary endothelial cells might protect against acute respiratory distress syndrome in obese individuals.

Intracellular pH (pHin) and cytosolic calcium ([Ca2+]cyt) regulation via ATPases: studies in cell populations, single cells, and subcellular compartments

NASA Astrophysics Data System (ADS)

Rojas, Jose D.; Sanka, Shankar C.; Gyorke, Sandor; Wesson, Donald E.; Minta, Akwasi; Martinez-Zaguilan, Raul

1999-07-01

Changes in pHin and (Ca2+)cyt are important in the signal transduction mechanisms leading to many physiological responses including cell growth, motility, secretion/exocytosis, etc. The concentrations of these ions are regulated via primary and secondary ion transporting mechanisms. In diabetes, specific pH and Ca2+ regulatory mechanism might be altered. To study these ions, we employ fluorescence spectroscopy, and cell imagin spectroscopy/confocal microscopy. pH and Ca2+ indicators are loaded in the cytosol with acetoxymethyl ester forms of dyes, and in endosomal/lysosomal (E/L) compartments by overnight incubation of cells with dextran- conjugated ion fluorescent probes. We focus on specific pH and Ca2+ regulatory systems: plasmalemmal vacuolar- type H+-ATPases (pm V-ATPases) and sarcoplasmic/endoplasmic reticulum Ca2+-ATPases (SERCA). As experimental models, we employ vascular smooth muscle (VSM) and microvascular endothelial cells. We have chosen these cells because they are important in blood flow regulation and in angiogenesis. These processes are altered in diabetes. In many cell types, ion transport processes are dependent on metabolism of glucose for maximal activity. Our main findings are: (a) glycolysis coupling the activity of SERCA is required for cytosolic Ca2+ homeostasis in both VSM and microvascular endothelial cells; (b) E/L compartments are important for pH and Ca2+ regulation via H+-ATPases and SERCA, respectively; and (c) pm-V- ATPases are important for pHin regulation in microvascular endothelial cells.

PGC-1α dictates endothelial function through regulation of eNOS expression

PubMed Central

Craige, Siobhan M.; Kröller-Schön, Swenja; Li, Chunying; Kant, Shashi; Cai, Shenghe; Chen, Kai; Contractor, Mayur M.; Pei, Yongmei; Schulz, Eberhard; Keaney, John F.

2016-01-01

Endothelial dysfunction is a characteristic of many vascular related diseases such as hypertension. Peroxisome proliferator activated receptor gamma, coactivator 1α (PGC-1α) is a unique stress sensor that largely acts to promote adaptive responses. Therefore, we sought to define the role of endothelial PGC-1α in vascular function using mice with endothelial specific loss of function (PGC-1α EC KO) and endothelial specific gain of function (PGC-1α EC TG). Here we report that endothelial PGC-1α is suppressed in angiotensin-II (ATII)-induced hypertension. Deletion of endothelial PGC-1α sensitized mice to endothelial dysfunction and hypertension in response to ATII, whereas PGC-1α EC TG mice were protected. Mechanistically, PGC-1α promotes eNOS expression and activity, which is necessary for protection from ATII-induced dysfunction as mice either treated with an eNOS inhibitor (LNAME) or lacking eNOS were no longer responsive to transgenic endothelial PGC-1α expression. Finally, we determined that the orphan nuclear receptor, estrogen related receptor α (ERRα) is required to coordinate the PGC-1α -induced eNOS expression. In conclusion, endothelial PGC-1α expression protects from vascular dysfunction by promoting NO• bioactivity through ERRα induced expression of eNOS. PMID:27910955

Traumatic Brain Injury Causes Endothelial Dysfunction in the Systemic Microcirculation through Arginase-1-Dependent Uncoupling of Endothelial Nitric Oxide Synthase.

PubMed

Villalba, Nuria; Sackheim, Adrian M; Nunez, Ivette A; Hill-Eubanks, David C; Nelson, Mark T; Wellman, George C; Freeman, Kalev

2017-01-01

Endothelial dysfunction is a hallmark of many chronic diseases, including diabetes and long-term hypertension. We show that acute traumatic brain injury (TBI) leads to endothelial dysfunction in rat mesenteric arteries. Endothelial-dependent dilation was greatly diminished 24 h after TBI because of impaired nitric oxide (NO) production. The activity of arginase, which competes with endothelial NO synthase (eNOS) for the common substrate l-arginine, were also significantly increased in arteries, suggesting that arginase-mediated depletion of l-arginine underlies diminished NO production. Consistent with this, substrate restoration by exogenous application of l-arginine or inhibition of arginase recovered endothelial function. Moreover, evidence for increased reactive oxygen species production, a consequence of l-arginine starvation-dependent eNOS uncoupling, was detected in endothelium and plasma. Collectively, our findings demonstrate endothelial dysfunction in a remote vascular bed after TBI, manifesting as impaired endothelial-dependent vasodilation, with increased arginase activity, decreased generation of NO, and increased O 2 - production. We conclude that blood vessels have a "molecular memory" of neurotrauma, 24 h after injury, because of functional changes in vascular endothelial cells; these effects are pertinent to understanding the systemic inflammatory response that occurs after TBI even in the absence of polytrauma.

Endothelial microparticles interact with and support the proliferation of T cells

PubMed Central

Wheway, Julie; Latham, Sharissa L; Combes, Valery; Grau, Georges ER

2014-01-01

Endothelial cells (EC) closely interact with circulating lymphocytes. Aggression or activation of the endothelium leads to an increased shedding of EC microparticles (MP). Endothelial MP (EMP) are found in high plasma levels in numerous immunoinflammatory diseases, e.g. atherosclerosis, sepsis, multiple sclerosis and cerebral malaria, supporting their role as effectors and markers of vascular dysfunction. Given our recently described role for human brain microvascular endothelial cells (HBEC) in modulating immune responses we investigated how HBEC-derived MP could interact with and support the proliferation of T cells. Like their mother cells, EMP expressed molecules important for antigen presentation and T cell co-stimulation, i.e., β2-microglobulin, MHC II, CD40 and ICOSL. HBEC were able to take up fluorescently labeled antigens with EMP also containing fluorescent antigens suggestive of antigen carryover from HBEC to EMP. In co-cultures, fluorescently labeled EMP from resting or cytokine-stimulated HBEC formed conjugates with both CD4+ and CD8+ subsets, with higher proportions of T cells binding EMP from cytokine stimulated cells. The increased binding of EMP from cytokine stimulated HBEC to T cells was VCAM-1 and ICAM-1-dependent. Finally, in CFSE T cell proliferation assays using anti-CD3 mAb or T cell mitogens, EMP promoted the proliferation of CD4+ T cells and that of CD8+ T cells in the absence of exogenous stimuli and in the T cell mitogenic stimulation. Our findings provide novel evidence that EMP can enhance T cell activation and potentially ensuing antigen presentation, thereby pointing towards a novel role for MP in neuro-immunological complications of infectious diseases. PMID:25187656

Endothelial C-Type Natriuretic Peptide Acts on Pericytes to Regulate Microcirculatory Flow and Blood Pressure.

PubMed

Špiranec, Katarina; Chen, Wen; Werner, Franziska; Nikolaev, Viacheslav O; Naruke, Takashi; Koch, Franziska; Werner, Andrea; Eder-Negrin, Petra; Diéguez-Hurtado, Rodrigo; Adams, Ralf H; Baba, Hideo A; Schmidt, Hannes; Schuh, Kai; Skryabin, Boris V; Movahedi, Kiavash; Schweda, Frank; Kuhn, Michaela

2018-04-06

Background -Peripheral vascular resistance has a major impact on arterial blood pressure levels. Endothelial C-type natriuretic peptide (CNP) participates in the local regulation of vascular tone but the target cells remain controversial. The cGMP-producing guanylyl cyclase-B (GC-B) receptor for CNP is expressed in vascular smooth muscle cells (VSMC). However, whereas endothelial cell-specific CNP knockout mice are hypertensive, mice with deletion of GC-B in VSMC have unaltered blood pressure. Methods -We analyzed whether the vasodilating response to CNP changes along the vascular tree, i.e. whether the GC-B receptor is expressed in microvascular types of cells. Mice with a floxed GC-B ( Npr2 ) gene were interbred with Tie2-Cre or PDGF-Rβ-Cre ERT2 lines to develop mice lacking GC-B in endothelial cells or in precapillary arteriolar SMC and capillary pericytes. Intravital microscopy, (non)invasive hemodynamics, fluorescence energy transfer studies of pericyte's cAMP levels in situ and renal physiology were combined to dissect whether and how CNP/GC-B/cGMP signaling modulates microcirculatory tone and blood pressure. Results -Intravital microscopy studies revealed that the vasodilatatory effect of CNP increases towards small-diameter arterioles and capillaries. Consistently, CNP did not prevent endothelin-1-induced acute constrictions of proximal arterioles but fully reversed endothelin effects in precapillary arterioles and capillaries. Here, the GC-B receptor is expressed both in endothelial and mural cells, i.e. in pericytes. Notably, the vasodilatatory effects of CNP were preserved in mice with endothelial GC-B deletion but abolished in mice lacking GC-B in microcirculatory SMC and pericytes. CNP, via GC-B/cGMP signaling modulates two signaling cascades in pericytes: it activates cGMP-dependent protein kinase I to phosphorylate downstream targets such as the cytoskeleton-associated vasodilator activated phosphoprotein; and it inhibits phosphodiesterase 3A, thereby enhancing pericyte's cAMP levels. Ultimately these pathways prevent endothelin-induced increases of pericyte calcium levels and pericyte contraction. Mice with deletion of GC-B in microcirculatory SMC and pericytes have elevated peripheral resistance and chronic arterial hypertension without a change in renal function. Conclusions -Our studies indicate that endothelial CNP regulates distal arteriolar and capillary blood flow. CNP-induced GC-B/cGMP signaling in microvascular SMC and pericytes is essential for the maintenance of normal microvascular resistance and blood pressure.

Interactions of neuropathogenic Escherichia coli K1 (RS218) and its derivatives lacking genomic islands with phagocytic Acanthamoeba castellanii and nonphagocytic brain endothelial cells.

PubMed

Yousuf, Farzana Abubakar; Yousuf, Zuhair; Iqbal, Junaid; Siddiqui, Ruqaiyyah; Khan, Hafsa; Khan, Naveed Ahmed

2014-01-01

Here we determined the role of various genomic islands in E. coli K1 interactions with phagocytic A. castellanii and nonphagocytic brain microvascular endothelial cells. The findings revealed that the genomic islands deletion mutants of RS218 related to toxins (peptide toxin, α -hemolysin), adhesins (P fimbriae, F17-like fimbriae, nonfimbrial adhesins, Hek, and hemagglutinin), protein secretion system (T1SS for hemolysin), invasins (IbeA, CNF1), metabolism (D-serine catabolism, dihydroxyacetone, glycerol, and glyoxylate metabolism) showed reduced interactions with both A. castellanii and brain microvascular endothelial cells. Interestingly, the deletion of RS218-derived genomic island 21 containing adhesins (P fimbriae, F17-like fimbriae, nonfimbrial adhesins, Hek, and hemagglutinin), protein secretion system (T1SS for hemolysin), invasins (CNF1), metabolism (D-serine catabolism) abolished E. coli K1-mediated HBMEC cytotoxicity in a CNF1-independent manner. Therefore, the characterization of these genomic islands should reveal mechanisms of evolutionary gain for E. coli K1 pathogenicity.

Interactions of Neuropathogenic Escherichia coli K1 (RS218) and Its Derivatives Lacking Genomic Islands with Phagocytic Acanthamoeba castellanii and Nonphagocytic Brain Endothelial Cells

PubMed Central

Yousuf, Farzana Abubakar; Yousuf, Zuhair; Iqbal, Junaid; Siddiqui, Ruqaiyyah; Khan, Hafsa; Khan, Naveed Ahmed

2014-01-01

Here we determined the role of various genomic islands in E. coli K1 interactions with phagocytic A. castellanii and nonphagocytic brain microvascular endothelial cells. The findings revealed that the genomic islands deletion mutants of RS218 related to toxins (peptide toxin, α-hemolysin), adhesins (P fimbriae, F17-like fimbriae, nonfimbrial adhesins, Hek, and hemagglutinin), protein secretion system (T1SS for hemolysin), invasins (IbeA, CNF1), metabolism (D-serine catabolism, dihydroxyacetone, glycerol, and glyoxylate metabolism) showed reduced interactions with both A. castellanii and brain microvascular endothelial cells. Interestingly, the deletion of RS218-derived genomic island 21 containing adhesins (P fimbriae, F17-like fimbriae, nonfimbrial adhesins, Hek, and hemagglutinin), protein secretion system (T1SS for hemolysin), invasins (CNF1), metabolism (D-serine catabolism) abolished E. coli K1-mediated HBMEC cytotoxicity in a CNF1-independent manner. Therefore, the characterization of these genomic islands should reveal mechanisms of evolutionary gain for E. coli K1 pathogenicity. PMID:24818136

Molecular Mechanisms Underlying the Cardiovascular Benefits of SGLT2i and GLP-1RA.

PubMed

Khat, Dorrin Zarrin; Husain, Mansoor

2018-06-09

In addition to their effects on glycemic control, two specific classes of relatively new anti-diabetic drugs, namely the sodium glucose co-transporter-2 inhibitors (SGLT2i) and glucagon-like peptide-1 receptor agonists (GLP-1RA) have demonstrated reduced rates of major adverse cardiovascular events (MACE) in subjects with type 2 diabetes (T2D) at high risk for cardiovascular disease (CVD). This review summarizes recent experimental results that inform putative molecular mechanisms underlying these benefits. SGLT2i and GLP-1RA exert cardiovascular effects by targeting in both common and distinctive ways (A) several mediators of macro- and microvascular pathophysiology: namely (A1) inflammation and atherogenesis, (A2) oxidative stress-induced endothelial dysfunction, (A3) vascular smooth muscle cell reactive oxygen species (ROS) production and proliferation, and (A4) thrombosis. These agents also exhibit (B) hemodynamic effects through modulation of (B1) natriuresis/diuresis and (B2) the renin-angiotensin-aldosterone system. This review highlights that while GLP-1RA exert direct effects on vascular (endothelial and smooth muscle) cells, the effects of SGLT2i appear to include the activation of signaling pathways that prevent adverse vascular remodeling. Both SGLT2i and GLP-1RA confer hemodynamic effects that counter adverse cardiac remodeling.

Complement inhibition prevents gut ischemia and endothelial cell dysfunction after hemorrhage/resuscitation.

PubMed

Fruchterman, T M; Spain, D A; Wilson, M A; Harris, P D; Garrison, R N

1998-10-01

Complement, a nonspecific immune response, is activated during hemorrhage/resuscitation (HEM/RES) and is involved in cellular damage. We hypothesized that activated complement injures endothelial cells (ETCs) and is responsible for intestinal microvascular hypoperfusion after HEM/RES. Four groups of rats were studied by in vivo videomicroscopy of the intestine: SHAM, HEM/RES, HEM/RES + sCR1 (complement inhibitor, 15 mg/kg intravenously given before resuscitation), and SHAM + sCR1. Hemorrhage was to 50% of mean arterial pressure for 60 minutes followed by resuscitation with shed blood plus an equal volume of saline. ETC function was assessed by response to acetylcholine. Resuscitation restored central hemodynamics to baseline after hemorrhage. After resuscitation, inflow A1 and premucosal A3 arterioles progressively constricted (-24% and -29% change from baseline, respectively), mucosal blood flow was reduced, and ETC function was impaired. Complement inhibition prevented postresuscitation vasoconstriction and gut ischemia. This protective effect appeared to involve preservation of ETC function in the A3 vessels (SHAM 76% of maximal dilation, HEM/RES 61%, HEM/RES + sCR1 74%, P < .05). Complement inhibition preserved ETC function after HEM/RES and maintained gut perfusion. Inhibition of complement activation before resuscitation may be a useful adjunct in patients experiencing major hemorrhage and might prevent the sequelae of gut ischemia.

Compartmentalized, functional role of angiogenin during spotted fever group rickettsia-induced endothelial barrier dysfunction: evidence of possible mediation by host tRNA-derived small noncoding RNAs

PubMed Central

2013-01-01

Background Microvascular endothelial barrier dysfunction is the central enigma in spotted fever group (SFG) rickettsioses. Angiogenin (ANG) is one of the earliest identified angiogenic factors, of which some are relevant to the phosphorylation of VE-cadherins that serve as endothelial adherens proteins. Although exogenous ANG is known to translocate into the nucleus of growing endothelial cells (ECs) where it plays a functional role, nuclear ANG is not detected in quiescent ECs. Besides its nuclear role, ANG is thought to play a cytoplasmic role, owing to its RNase activity that cleaves tRNA to produce small RNAs. Recently, such tRNA-derived RNA fragments (tRFs) have been shown to be induced under stress conditions. All these observations raise an intriguing hypothesis about a novel cytoplasmic role of ANG, which is induced upon infection with Rickettsia and generates tRFs that may play roles in SFG rickettsioses. Methods C3H/HeN mice were infected intravenously with a sublethal dose of R. conorii. At days 1, 3, and 5 post infection (p.i.), liver, lung and brain were collected for immunofluorescence (IF) studies of R. conorii and angiogenin (ANG). Human umbilical vein endothelial cells (HUVECs) were infected with R. conorii for 24, 48, and 72 hrs before incubation with 1μg/ml recombinant human ANG (rANG) in normal medium for 2 hrs. HUVEC samples were subjected to IF, exogenous ANG translocation, endothelial permeability, and immunoprecipitation phosphorylation assays. To identify small non-coding RNAs (sncRNAs) upon rickettsial infection, RNAs from pulverized mouse lung tissues and HUVECs were subjected to library preparation and deep sequencing analysis using an Illumina 2000 instrument. Identified sncRNAs were confirmed by Northern hybridization, and their target mRNAs were predicted in silico using BLAST and RNA hybrid programs. Results In the present study, we have demonstrated endothelial up-regulation of ANG, co-localized with SFG rickettsial infection in vivo. We also have provided direct evidence that rickettsial infection sensitizes human ECs to the translocation of exogenous ANG in a compartmentalized pattern at different times post-infection. Typically, exogenous ANG translocates into the nucleus at 24 hrs and to the cytoplasm at 72 hrs post-infection. The ANG cytoplasmic translocation enhances phosphorylation and destabilization of VE-cadherin and attenuates endothelial barrier function. Of note, deep sequencing analysis detected tRFs, mostly derived from the 5'-halves of host tRNAs, that are induced by ANG. Northern hybridization validates the two most abundantly cloned tRFs derived from tRNA-ValGTG and tRNA-GlyGCC, in both mouse tissues and human cells. Bioinformatics analysis predicted that these tRFs may interact with transcripts associated with the endothelial barrier, the host cell inflammatory response, and autophagy. Conclusions Our data provide new insight into the role of compartmentalized ANG during SFG rickettsioses, and highlight its possible mediation through tRFs. PMID:23800282

Alterations in triglyceride rich lipoproteins are related to endothelial dysfunction in metabolic syndrome.

PubMed

Lucero, Diego; López, Graciela I; Gorzalczany, Susana; Duarte, Mariano; González Ballerga, Esteban; Sordá, Juan; Schreier, Laura; Zago, Valeria

2016-08-01

Our aim was to analyze the effect of circulating triglyceride rich lipoprotein (TRL) on endothelial function in metabolic syndrome (MetS). We studied 40 patients with MetS (ATPIII), divided into those presenting normal endothelial function (n=19) and those with endothelial dysfunction (n=21) by means of the evaluation of pulse wave velocity, before and after brachial artery ischemia. In fasting serum we measured lipid and lipoprotein profile, insulin and glucose (HOMA-IR). Moreover, isolated TRL (d<1006g/l) were chemically characterized. In parallel, using randomly selected TRL from MetS patients with endothelial dysfunction (n=6) and MetS patients with normal endothelial function (n=6), the ability of TRL to inhibit ACh-induced vasorelaxation (10(-9)-10(-5)mM) on aortic rings previously pre-contracted by noradrenaline (10(-8)mM) was evaluated. Interestingly, TRL isolated from MetS patients presenting endothelial dysfunction showed triglyceride over-enrichment (59.1±4.8 vs. 54.1±4.7%; p=0.04), even after adjusting by potential confounders (p=0.05). In addition, while TRL resulting from both MetS groups significantly inhibited endothelium dependent vasorelaxation (p<0.001), TRL from MetS patients with endothelial dysfunction showed a strong tendency to a greater inhibition of vasorelaxation (p=0.06). Moreover, TRL-triglyceride (%) showed a strong tendency to correlate with the grade of vasorelaxation inhibition exerted by TRL (r=0.60; p=0.05). These results, taken together, would allow inferring for the first time that the predominance of triglyceride over-enriched TRL in circulation in MetS would induce endothelial dysfunction, contributing to the inherent cardiovascular risk of MetS. Copyright © 2016 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Integrated analysis of long noncoding RNA and mRNA profiling ox-LDL-induced endothelial dysfunction after atorvastatin administration.

PubMed

Jiang, Ling-Yu; Jiang, Yue-Hua; Qi, Ying-Zi; Shao, Lin-Lin; Yang, Chuan-Hua

2018-06-01

Long noncoding RNAs (lncRNAs) play a key role in the development of endothelial dysfunction. However, few lncRNAs associated with endothelial dysfunction after atorvastatin administration have been reported. In the present study, differentially expressed (DE) genes in ox-LDL versus control and ox-LDL + atorvastatin versus control were detected. Bioinformatics analysis and integrated analysis of mRNAs and lncRNAs were conducted to study the mechanisms of endothelial dysfunction after atorvastatin administration and to explore the regulation functions of lncRNAs. Here, 532 DE mRNAs and 532 DE lncRNAs were identified (among them, 195 mRNAs and 298 lncRNAs were upregulated, 337 mRNAs and 234 lncRNAs were downregulated) after ox-LDL treatment for 24 hours (fold change ≥2.0, P < .05). After ox-LDL treatment following atorvastatin administration, 750 DE mRNAs and 502 DE lncRNAs were identified (among them, 149 mRNAs and 218 lncRNAs were upregulated and 601 mRNAs and 284 lncRNAs were downregulated). After atorvastatin administration, 167 lncRNAs and 262 mRNAs were still DE. Q-PCR validated the results of microarrays. Chronic inflammatory response, nitric oxide biosynthetic process, microtubule cytoskeleton, cell proliferation and cell migration are regulated by lncRNAs, which also participated in the mainly molecular function and biological processes underlying endothelial dysfunction. Atorvastatin partly improved endothelial dysfunction, but the aspects beyond recovery were mainly concentrated in cell cycle, mitosis, and metabolism. Further exploration is required to explicit the mechanism by which lncRNAs participate in endothelial dysfunction.

Hyperoxic sheep pulmonary microvascular endothelial cells generate free radicals via mitochondrial electron transport.

PubMed Central

Sanders, S P; Zweier, J L; Kuppusamy, P; Harrison, S J; Bassett, D J; Gabrielson, E W; Sylvester, J T

1993-01-01

Free radical generation by hyperoxic endothelial cells was studied using electron paramagnetic resonance (EPR) spectroscopy and the spin trap 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). Studies were performed to determine the radical species produced, whether mitochondrial electron transport was involved, and the effect of the radical generation on cell mortality. Sheep pulmonary microvascular endothelial cell suspensions exposed to 100% O2 for 30 min exhibited prominent DMPO-OH and, occasionally, additional smaller DMPO-R signals thought to arise from the trapping of superoxide anion (O2-.), hydroxyl (.OH), and alkyl (.R) radicals. Superoxide dismutase (SOD) quenched both signals suggesting that the observed radicals were derived from O2-.. Studies with deferoxamine suggested that the generation of .R occurred secondary to the formation of .OH from O2-. via an iron-mediated Fenton reaction. Blocking mitochondrial electron transport with rotenone (20 microM) markedly decreased radical generation. Cell mortality increased slightly in oxygen-exposed cells. This increase was not significantly altered by SOD or deferoxamine, nor was it different from the mortality observed in air-exposed cells. These results suggest that endothelial cells exposed to hyperoxia for 30 min produce free radicals via mitochondrial electron transport, but under the conditions of these experiments, this radical generation did not appear cause cell death. PMID:8380815

Pharmacological modulation of endothelial cell-associated adhesion molecule expression: implications for future treatment of dermatological diseases.

PubMed

Foster, C A; Dreyfuss, M; Mandak, B; Meingassner, J G; Naegeli, H U; Nussbaumer, A; Oberer, L; Scheel, G; Swoboda, E M

1994-11-01

Skin diseases with an inflammatory component, regardless of their etiology, are characterized at some point by the extravasation and subsequent infiltration of leukocytes into the dermal and/or epidermal compartments. This trafficking pattern is determined by a complex series of events whereby the leukocytes interact with cell adhesion molecules (CAM), particularly those induced on endothelial cells following activation with various inflammatory mediators. Vascular CAMs belonging to the selectin family (i.e., P-selectin and E-selectin) are thought to mediate early and reversible events involving leukocyte rolling and margination along the lumenal surface of microvascular cells (post-capillary venules). Certain members of the immunoglobulin supergene family (i.e., VCAM-1 and ICAM-1) regulate later and irreversible steps which lead to firm attachment and subsequent diapedesis of leukocytes. Accumulating evidence suggests that if one blocks the ligand-binding sites between leukocytes and endothelial cells, or inhibits vascular CAM expression, hematopoietic cell extravasation and progressive inflammatory events can be greatly diminished. To identify such inhibitors we developed a cell-based Elisa using the human microvascular cell line HMEC-1. As reported in the present paper, this approach yielded a naturally-occurring, low molecular weight compound which potently inhibits cytokine-induced adhesion molecule expression on cultured endothelial cells, without modulating "house-keeping" proteins.

Ibrolipim attenuates high glucose-induced endothelial dysfunction in cultured human umbilical vein endothelial cells via PI3K/Akt pathway.

PubMed

Xiao, Guohua; Wang, Zongbao; Zeng, Huaicai; Yu, Jian; Yin, Weidong; Zhang, Sujun; Wang, Yueting; Zhang, Yali

2011-10-01

Endothelial dysfunction is a key event in the onset and progression of atherosclerosis associated with diabetes. Increasing cell apoptosis may lead to endothelial dysfunction and contribute to vascular complications. Therefore, we aimed to elucidate the possible role and mechanism of ibrolipim in preventing endothelial dysfunction induced by high glucose. Human umbilical vein endothelial cells (HUVECs) were cultured respectively under normal glucose level (5.5mM), high glucose level (33mM), and high glucose level with ibrolipim treatment. Endothelial dysfunction was identified by the expression of ET-1 and vWF through reverse transcription PCR (RT-PCR). HUVECs apoptosis was assessed by fluorescent staining with Hoechst 33258. Akt activity was analyzed by western blot. High glucose condition significantly increased the rate of apoptotic cells, weakened cell viability, and decreased the expression of ET-1 and vWF. Ibrolipim treatment significantly attenuated these alterations of endothelial dysfunction. The lower concentrations (2, 4, 8 microM) of ibrolipim inhibited apoptosis of cultured HUVECs, improved cell viability, down-regulated the mRNA levels of ET-1, vWF, and attenuated the cytotoxicity; however, higher concentration (16, 32 microM) of ibrolipim aggravated the damage of HUVECs cultured under high glucose level. Meanwhile, high glucose induced a decrease of Akt activity which led to apoptosis, and ibrolipim prevented the decrease and attenuated apoptotic effect induced by high glucose. Furthermore, the PI3K inhibitor LY294002 significantly abolished the anti-apoptotic effect of ibrolipim, and decreased Akt phosphorylation. Although, the expression of Akt mRNA and total protein were not altered in cultured HUVECs. Ibrolipim at lower concentrations can inhibit high glucose-induced apoptosis in cultured HUVECs, which might be related to the alternation of Akt activity. Ibrolipim has the potential to attenuate endothelial dysfunction and lower the risk of diabetes-associated vascular diseases. And it might be a therapeutic agent for diabetic vascular complications.

A microengineered model of RBC transfusion-induced pulmonary vascular injury.

PubMed

Seo, Jeongyun; Conegliano, David; Farrell, Megan; Cho, Minseon; Ding, Xueting; Seykora, Thomas; Qing, Danielle; Mangalmurti, Nilam S; Huh, Dongeun

2017-06-13

Red blood cell (RBC) transfusion poses significant risks to critically ill patients by increasing their susceptibility to acute respiratory distress syndrome. While the underlying mechanisms of this life-threatening syndrome remain elusive, studies suggest that RBC-induced microvascular injury in the distal lung plays a central role in the development of lung injury following blood transfusion. Here we present a novel microengineering strategy to model and investigate this key disease process. Specifically, we created a microdevice for culturing primary human lung endothelial cells under physiological flow conditions to recapitulate the morphology and hemodynamic environment of the pulmonary microvascular endothelium in vivo. Perfusion of the microengineered vessel with human RBCs resulted in abnormal cytoskeletal rearrangement and release of intracellular molecules associated with regulated necrotic cell death, replicating the characteristics of acute endothelial injury in transfused lungs in vivo. Our data also revealed the significant effect of hemodynamic shear stress on RBC-induced microvascular injury. Furthermore, we integrated the microfluidic endothelium with a computer-controlled mechanical stretching system to show that breathing-induced physiological deformation of the pulmonary microvasculature may exacerbate vascular injury during RBC transfusion. Our biomimetic microsystem provides an enabling platform to mechanistically study transfusion-associated pulmonary vascular complications in susceptible patient populations.

Endothelial microparticles: Pathogenic or passive players in endothelial dysfunction in autoimmune rheumatic diseases?

PubMed

McCarthy, E M; Wilkinson, F L; Parker, B; Alexander, M Y

2016-11-01

Autoimmune rheumatic diseases are characterised by systemic inflammation and complex immunopathology, with an increased risk of cardiovascular disease, initiated by endothelial dysfunction in a chronic inflammatory environment. Endothelial microparticles (EMPs) are released into the circulation from activated endothelial cells and may therefore, reflect disease severity, vascular and endothelial dysfunction, that could influence disease pathogenesis via autocrine/paracrine signalling. The exact function of EMPs in rheumatic disease remains unknown, and this has initiated research to elucidate EMP composition and function, which may be determined by the mode of endothelial activation and the micro environment. To date, EMPs are thought to play a role in angiogenesis, thrombosis and inflammation by transferring specific proteins and microRNAs (miRs) to target cells. Here, we review the mechanisms underlying the generation and composition of EMPs and the clinical and experimental studies describing the involvement of EMPs in rheumatic diseases, since we have previously shown endothelial dysfunction and an elevated risk of cardiovascular disease are characteristics in systemic lupus erythematosus. We will also discuss the potential of EMPs as future biomarkers of cardiovascular risk in these diseases. Copyright © 2016 Elsevier Inc. All rights reserved.

Effects of dietary creatine supplementation on systemic microvascular density and reactivity in healthy young adults.

PubMed

Moraes, Roger de; Van Bavel, Diogo; Moraes, Beatriz Serpa de; Tibiriçá, Eduardo

2014-12-15

Dietary creatine supplementation (CrS) is a practice commonly adopted by physically active individuals. However, the effects of CrS on systemic microvascular reactivity and density have never been reported. Additionally, CrS is able to influence blood levels of homocysteine, resulting in presumed effects on vascular endothelial function. Thus, we investigated the effects of CrS on the systemic microcirculation and on homocysteine levels in healthy young individuals. This open-label study was performed on a group of 40 healthy male, moderately physically active subjects aged 27.7 ± 13.4 years who received one week of CrS at a dose of 20 g/day of commercially available micronized creatine monohydrate. Laser speckle contrast imaging was used in the evaluation of cutaneous microvascular reactivity, and intra-vital video microscopy was used to evaluate skin capillary density and reactivity, before and after CrS. CrS did not alter plasma levels of homocysteine, although CrS increased creatinine (p = 0.0001) and decreased uric acid (p = 0.0004) plasma levels. Significant changes in total cholesterol (p = 0.0486) and LDL-cholesterol (p = 0.0027) were also observed along with a reduction in plasma levels of T3 (p = 0.0074) and an increase in T4 levels (p = 0.0003). Skin functional capillary density (p = 0.0496) and capillary recruitment during post-occlusive reactive hyperemia (p = 0.0043) increased after CrS. Increases in cutaneous microvascular vasodilation induced by post-occlusive reactive hyperemia (p = 0.0078) were also observed. Oral supplementation with creatine in healthy, moderately physically active young adults improves systemic endothelial-dependent microvascular reactivity and increases skin capillary density and recruitment. These effects are not concurrent with changes in plasma homocysteine levels.

Renal Hypoxia and Dysoxia After Reperfusion of the Ischemic Kidney

PubMed Central

Legrand, Matthieu; Mik, Egbert G; Johannes, Tanja; Payen, Didier; Ince, Can

2008-01-01

Ischemia is the most common cause of acute renal failure. Ischemic-induced renal tissue hypoxia is thought to be a major component in the development of acute renal failure in promoting the initial tubular damage. Renal oxygenation originates from a balance between oxygen supply and consumption. Recent investigations have provided new insights into alterations in oxygenation pathways in the ischemic kidney. These findings have identified a central role of microvascular dysfunction related to an imbalance between vasoconstrictors and vasodilators, endothelial damage and endothelium–leukocyte interactions, leading to decreased renal oxygen supply. Reduced microcirculatory oxygen supply may be associated with altered cellular oxygen consumption (dysoxia), because of mitochondrial dysfunction and activity of alternative oxygen-consuming pathways. Alterations in oxygen utilization and/or supply might therefore contribute to the occurrence of organ dysfunction. This view places oxygen pathways’ alterations as a potential central player in the pathogenesis of acute kidney injury. Both in regulation of oxygen supply and consumption, nitric oxide seems to play a pivotal role. Furthermore, recent studies suggest that, following acute ischemic renal injury, persistent tissue hypoxia contributes to the development of chronic renal dysfunction. Adaptative mechanisms to renal hypoxia may be ineffective in more severe cases and lead to the development of chronic renal failure following ischemia-reperfusion. This paper is aimed at reviewing the current insights into oxygen transport pathways, from oxygen supply to oxygen consumption in the kidney and from the adaptation mechanisms to renal hypoxia. Their role in the development of ischemia-induced renal damage and ischemic acute renal failure are discussed. PMID:18488066

Association of parental blood pressure with retinal microcirculatory abnormalities indicative of endothelial dysfunction in children.

PubMed

Islam, Muhammad; Jafar, Tazeen H; Bux, Rasool; Hashmi, Shiraz; Chaturvedi, Nish; Hughes, Alun D

2014-03-01

Microcirculatory abnormalities precede the onset of hypertension and may explain its familial nature. We examined the relationship between parental blood pressure (BP) and offspring retinal microvasculature in Pakistani trios [father, mother, and child (aged 9-14 years)]. This is a substudy of a population-based trial of BP reduction. Data were available on 358 normotensive, and 410 offspring of at least one hypertensive parent. Retinal vessel characteristics were measured from digital images. Multivariable linear regression models were built to assess the associations between maternal and paternal BP and offspring retinal microvasculature. Optimality deviation was greatest in offspring of two hypertensive parents, compared with those with one or no hypertensive parent (P=0.030 for trend). Paternal SBP and DBP were each significantly associated with optimality deviation in offspring (P=0.023 and P=0.006, respectively). This relationship persisted after accounting for offspring cardiovascular risk factors [increase in optimality deviation (95% confidence interval, CI) 0.0053 (0.0001-0.0106, P=0.047) and 0.0109 (0.0025-0.0193, P=0.011), for each 10 mmHg increase in paternal SBP and DBP, respectively]. Maternal DBP was inversely associated with offspring arteriovenous ratio -0.0102 (-0.0198 to -0.0007, P=0.035). Microvascular endothelial dysfunction in children is associated with increasing levels of parental hypertension. The association with paternal BP is independent of other cardiovascular risk factors, including the child's BP. Higher maternal DBP is associated with evidence of arteriolar narrowing in offspring. These early microcirculatory changes may help explain familial predisposition to hypertension in people of Pakistani origin at an early age. :

Retinopathy in type 1 diabetes mellitus: Major differences between rural and urban dwellers in northwest Ethiopia.

PubMed

Alemu, Shitaye; Dessie, Abere; Tsegaw, Asamere; Patterson, Christopher C; Parry, Eldryd H O; Phillips, David I W; Trimble, Elisabeth R

2015-07-01

To audit levels of diabetes-related eye disease in Type 1 diabetes mellitus (T1DM) patients in northwest Ethiopia. In particular to establish whether, despite identical clinical goals, major differences between the physically demanding life-style of rural subsistence farmers and the sedentary life-style of urban dwellers would influence the prevalence of diabetes-related eye complications. A robust infrastructure for chronic disease management that comprehensively includes all rural dwellers was a pre-requisite for the investigation. A total of 544 T1DM were examined, representing 80% of all T1DM patients under regular review at both the urban and rural clinics and representative of patient age and gender (62.1% male, 37.9% female) of T1DM patients from this region; all were supervised by the same clinical team. Eye examinations were performed for visual acuity, cataract and retinal changes (retinal photography). HbA1c levels and the presence or absence of hypertension were recorded. Urban and rural groups had similar prevalences of severe visual impairment/blindness (7.0% urban, 5.2% rural) and cataract (7.3% urban, 7.1% rural). By contrast, urban dwellers had a significantly higher prevalence of retinopathy compared to rural patients, 16.1% and 5.0%, respectively (OR 2.9, p<0.02, after adjustment for duration, age, gender and hypertension). There was a 3-fold greater prevalence of hypertension in urban patients, whereas HbA1c levels were similar in the two groups. Since diabetic retinopathy is closely associated with microvascular disease and endothelial dysfunction, the possible influences of hypertension to increase and of sustained physical activity to reduce endothelial dysfunction are discussed. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Nitric oxide-induced changes in endothelial expression of phosphodiesterases 2, 3, and 5.

PubMed

Schankin, Christoph J; Kruse, Lars S; Reinisch, Veronika M; Jungmann, Steffen; Kristensen, Julie C; Grau, Stefan; Ferrari, Uta; Sinicina, Inga; Goldbrunner, Roland; Straube, Andreas; Kruuse, Christina

2010-03-01

To investigate nitric oxide (NO)-mediated changes in expression of cyclic nucleotide degrading phosphodiesterases 2A (PDE2A), PDE3B, and PDE5A in human endothelial cells. Nitric oxide induces production of cyclic guanosine monophosphate (cGMP), which along with cyclic adenosine monophosphate (cAMP) is degraded by PDEs. NO donors and selective inhibitors of PDE3 and PDE5 induce migraine-like headache and play a role in endothelial dysfunction during stroke. The current study investigates possible NO modulation of cGMP-related PDEs relevant to headache induction in a cell line containing such PDEs. Real time polymerase chain reaction and Western blots were used to show expression of PDE2A, PDE3B, and PDE5A in a stable cell line of human brain microvascular endothelial cells. Effects of NO on PDE expression were analyzed at specific time intervals after continued DETA NONOate administration. This study shows the expression of PDE2A, PDE3B, and PDE5A mRNA and PDE3B and PDE5A protein in human cerebral endothelial cells. Long-term DETA NONOate administration induced an immediate mRNA up-regulation of PDE5A (1.9-fold, 0.5 hour), an early peak of PDE2A (1.4-fold, 1 and 2 hours) and later up-regulation of both PDE3B (1.6-fold, 4 hours) and PDE2A (1.7-fold, 8 hours and 1.2-fold after 24 hours). Such changes were, however, not translated into significant changes in protein expression indicating few, if any, functional effects. Long-term NO stimulation modulated PDE3 and PDE5 mRNA expression in endothelial cells. However, PDE3 and PDE5 protein levels were unaffected by NO. The presence of PDE3 or PDE5 in endothelial cells indicates that selective inhibitors may have functional effects in such cells. A complex interaction of cGMP and cAMP in response to NO administration may take place if the mRNA translates into active protein. Whether or not this plays a role in the headache mechanisms remains to be investigated.

Olive Oil Supplements Ameliorate Endothelial Dysfunction Caused by Concentrated Ambient Particulate Matter Exposure in Healthy Human Volunteers

EPA Science Inventory

Context: Exposure to ambient particulate matter (PM) induces endothelial dysfunction, a risk factor for clinical cardiovascular events and progression of atherosclerosis. Dietary supplements such as olive oil and fish oil have beneficial effects on endothelial function, and ther...

Constitutive production and thrombin-induced release of vascular endothelial growth factor by human megakaryocytes and platelets

PubMed Central

Möhle, Robert; Green, David; Moore, Malcolm A. S.; Nachman, Ralph L.; Rafii, Shahin

1997-01-01

We have shown that coculture of bone marrow microvascular endothelial cells with hematopoietic progenitor cells results in proliferation and differentiation of megakaryocytes. In these long-term cultures, bone marrow microvascular endothelial cell monolayers maintain their cellular integrity in the absence of exogenous endothelial growth factors. Because this interaction may involve paracrine secretion of cytokines, we evaluated megakaryocytic cells for secretion of vascular endothelial growth factor (VEGF). Megakaryocytes (CD41a+) were generated by ex vivo expansion of hematopoietic progenitor cells with kit-ligand and thrombopoietin for 10 days and further purified with immunomagnetic microbeads. Using reverse transcription–PCR, we showed that megakaryocytic cell lines (Dami, HEL) and purified megakaryocytes expressed mRNA of the three VEGF isoforms (121, 165, and 189 amino acids). Large quantities of VEGF (>1 ng/106 cells/3 days) were detected in the supernatant of Dami cells, ex vivo-generated megakaryocytes, and CD41a+ cells isolated from bone marrow. The constitutive secretion of VEGF by CD41a+ cells was stimulated by growth factors of the megakaryocytic lineage (interleukin 3, thrombopoietin). Western blotting of heparin–Sepharose-enriched supernatant mainly detected the isoform VEGF165. In addition, immunohistochemistry showed intracytoplasmic VEGF in polyploid megakaryocytes. Thrombin stimulation of megakaryocytes and platelets resulted in rapid release of VEGF within 30 min. We conclude that human megakaryocytes produce and secrete VEGF in an inducible manner. Within the bone marrow microenvironment, VEGF secreted by megakaryocytes may contribute to the proliferation of endothelial cells. VEGF delivered to sites of vascular injury by activated platelets may initiate angiogenesis. PMID:9012841

Systemic influences contribute to prolonged microvascular rarefaction after brain irradiation: a role for endothelial progenitor cells

PubMed Central

Ashpole, Nicole M.; Warrington, Junie P.; Mitschelen, Matthew C.; Yan, Han; Sosnowska, Danuta; Gautam, Tripti; Farley, Julie A.; Csiszar, Anna; Ungvari, Zoltan

2014-01-01

Whole brain radiation therapy (WBRT) induces profound cerebral microvascular rarefaction throughout the hippocampus. Despite the vascular loss and localized cerebral hypoxia, angiogenesis fails to occur, which subsequently induces long-term deficits in learning and memory. The mechanisms underlying the absence of vessel recovery after WBRT are unknown. We tested the hypotheses that vascular recovery fails to occur under control conditions as a result of loss of angiogenic drive in the circulation, chronic tissue inflammation, and/or impaired endothelial cell production/recruitment. We also tested whether systemic hypoxia, which is known to promote vascular recovery, reverses these chronic changes in inflammation and endothelial cell production/recruitment. Ten-week-old C57BL/6 mice were subjected to a clinical series of fractionated WBRT: 4.5-Gy fractions 2 times/wk for 4 wk. Plasma from radiated mice increased in vitro endothelial cell proliferation and adhesion compared with plasma from control mice, indicating that WBRT did not suppress the proangiogenic drive. Analysis of cytokine levels within the hippocampus revealed that IL-10 and IL-12(p40) were significantly increased 1 mo after WBRT; however, systemic hypoxia did not reduce these inflammatory markers. Enumeration of endothelial progenitor cells (EPCs) in the bone marrow and circulation indicated that WBRT reduced EPC production, which was restored with systemic hypoxia. Furthermore, using a bone marrow transplantation model, we determined that bone marrow-derived endothelial-like cells home to the hippocampus after systemic hypoxia. Thus, the loss of production and homing of EPCs have an important role in the prolonged vascular rarefaction after WBRT. PMID:25038144

Gestational diabetes, preeclampsia and cytokine release: similarities and differences in endothelial cell function.

PubMed

Rao, Rashmi; Sen, Suvajit; Han, Bing; Ramadoss, Sivakumar; Chaudhuri, Gautam

2014-01-01

Gestational diabetes, pre-eclampsia as well as intra-uterine infection during pregnancy affects the function of the endothelium both in the mother and the fetus leading to endothelial dysfunction. Gestational diabetes is also associated with an increased incidence of pre-eclampsia and it is likely that both the hyperglycemia as well as the release of cytokines especially TNFα during hyperglycemia may play an important role in the pathogenesis of endothelial dysfunction leading to preeclampsia. Similarly, some but not all studies have suggested that infection of the mother under certain circumstances can also lead to preeclampsia as women with either a bacterial or viral infection were at a higher risk of developing preeclampsia, compared to women without infection and infection also leads to a release in TNFα. Endothelial cells exposed to either high glucose or TNFα leads to an increase in the production of H2O2 and to a decrease in endothelial cell proliferation. The cellular and molecular mechanisms involved in this phenomenon are discussed.Gestational diabetes, pre-eclampsia as well as intra-uterine infection during pregnancy has profound effects on the fetus and long term effects on the neonate. All three conditions affect the function of the endothelium both in the mother and the fetus leading to endothelial dysfunction. Gestational diabetes is also associated with an increased incidence of pre-eclampsia and it is likely that both the hyperglycemia as well as the release of cytokines especially TNFα during hyperglycemia may play an important role in the pathogenesis of endothelial dysfunction leading to preeclampsia. It has also been suggested although not universally accepted that under certain circumstances maternal infection may also predispose to pre-eclampsia. Pre-eclampsia is also associated with the release of TNFα and endothelial dysfunction. However, the cellular and molecular mechanism(s) leading to the endothelial dysfunction by either hyperglycemia or by the cytokine TNFα appear to be different. In this chapter, we explore some of the similarities and differences leading to endothelial dysfunction by both hyperglycemia and by the inflammatory cytokine TNFα and the cellular and molecular mechanism(s) involved.

Rhubarb Antagonizes Matrix Metalloproteinase-9-induced Vascular Endothelial Permeability

PubMed Central

Cui, Yun-Liang; Zhang, Sheng; Tian, Zhao-Tao; Lin, Zhao-Fen; Chen, De-Chang

2016-01-01

Background: Intact endothelial structure and function are critical for maintaining microcirculatory homeostasis. Dysfunction of the latter is an underlying cause of various organ pathologies. In a previous study, we showed that rhubarb, a traditional Chinese medicine, protected intestinal mucosal microvascular endothelial cells in rats with metastasizing septicemia. In this study, we investigated the effects and mechanisms of rhubarb on matrix metalloproteinase-9 (MMP9)-induced vascular endothelial (VE) permeability. Methods: Rhubarb monomers were extracted and purified by a series of chromatography approaches. The identity of these monomers was analyzed by hydrogen-1 nuclear magnetic resonance (NMR), carbon-13 NMR, and distortionless enhancement by polarization transfer magnetic resonance spectroscopy. We established a human umbilical vein endothelial cell (HUVEC) monolayer on a Transwell insert. We measured the HUVEC permeability, proliferation, and the secretion of VE-cadherin into culture medium using fluorescein isothiocyanate-dextran assay, 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay, and enzyme-linked immunosorbent assay, respectively, in response to treatment with MMP9 and/or rhubarb monomers. Results: A total of 21 rhubarb monomers were extracted and identified. MMP9 significantly increased the permeability of the HUVEC monolayer, which was significantly reduced by five individual rhubarb monomer (emodin, 3,8-dihydroxy-1-methyl-anthraquinone-2-carboxylic acid, 1-O-caffeoyl-2-(4-hydroxyl-O-cinnamoyl)-β-D-glucose, daucosterol linoleate, and rhein) or a combination of all five monomers (1 μmol/L for each monomer). Mechanistically, the five-monomer mixture at 1 μmol/L promoted HUVEC proliferation. In addition, MMP9 stimulated the secretion of VE-cadherin into the culture medium, which was significantly inhibited by the five-monomer mixture. Conclusions: The rhubarb mixture of emodin, 3,8-dihydroxy-1-methyl-anthraquinone-2-carboxylic acid, 1-O-caffeoyl-2-(4-hydroxyl-O-cinnamoyl)-β-D-glucose, daucosterol linoleate, and rhein, at a low concentration, antagonized the MMP9-induced HUVEC monolayer permeability by promoting HUVEC proliferation and reducing extracellular VE-cadherin concentrations. PMID:27411464

Loss of Endothelial Barrier Antigen Immunoreactivity in Rat Retinal Microvessels is Correlated with Clostridium perfringens Type D Epsilon Toxin-induced Damage to the Blood-Retinal Barrier.

PubMed

Mander, K A; Finnie, J W

2018-01-01

Clostridium perfringens type D epsilon toxin (ETX) is a potent neurotoxin producing a severe, and often fatal, neurological disorder in ruminant livestock. Microvascular damage appears to be the fundamental action of ETX in the brain and, recently, similar vascular injury, with subsequent severe vasogenic oedema, has been reported in the retina of rats given ETX. Endothelial barrier antigen (EBA) is a useful marker of an intact blood-brain barrier in rats and it has been shown that loss of EBA immunoreactivity is correlated with ETX-induced cerebral microvascular damage in this species. This paper reports, for the first time, that loss of EBA immunoexpression also occurs in rat retinal microvessels exposed to ETX, the marked reduction in EBA immunopositivity acting as a useful marker for blood-retinal barrier breakdown produced by this neurotoxin. Copyright © 2017 Elsevier Ltd. All rights reserved.

How Cryptococcus interacts with the blood-brain barrier.

PubMed

Tseng, Hsiang-Kuang; Huang, Tseng-Yu; Wu, Alice Ying-Jung; Chen, Hsin-Hong; Liu, Chang-Pan; Jong, Ambrose

2015-01-01

Cryptococcus demonstrates predilection for invasion of the brain, but the mechanism by which Cryptococcus crosses the blood-brain barrier (BBB) to cause brain invasion is largely unknown. In order for Cryptococcus to cross the BBB, there must be a way to either cross human brain microvascular endothelial cells, which are the main constitute of the BBB, or go in between tight junctions. Recent evidence of human brain microvascular endothelial cell responses to transcellular brain invasions includes membrane rearrangements, intracellular signaling pathways and cytoskeletal activations. Several Cryptococcal genes related to the traversal of BBB have been identified, including CPS1, ITR1a, ITR3c, PLB1, MPR1, FNX1 and RUB1. In addition, Cryptococcus neoformans-derived microvesicles may contribute to cryptococcal brain invasion. Paracellularly, Cryptococcus may traverse across BBB using either routes utilizing plasmin, ammonia or macrophages in a Trojan horse mechanism.

Which side of the balance determines the frequency of vaso-occlusive crises in children with sickle cell anemia: Blood viscosity or microvascular dysfunction?

PubMed

Charlot, Keyne; Romana, Marc; Moeckesch, Berenike; Jumet, Stéphane; Waltz, Xavier; Divialle-Doumdo, Lydia; Hardy-Dessources, Marie-Dominique; Petras, Marie; Tressières, Benoît; Tarer, Vanessa; Hue, Olivier; Etienne-Julan, Maryse; Antoine-Jonville, Sophie; Connes, Philippe

2016-01-01

Vascular resistance and tissue perfusion may be both affected by impaired vascular function and increased blood viscosity. Little is known about the effects of vascular function on the occurrence of painful vaso-occlusive crises (VOC) in children with sickle cell anemia (SCA). The aim of the present study was to determine which side of the balance (blood viscosity or vascular function) is the most deleterious in SCA and increases the risk for frequent hospitalized VOC. Microvascular function, microcirculatory oxygenation and blood viscosity were determined in a group of 22 SCA children/adolescents at steady state and a group of 13 healthy children/adolescents. Univariate analyses demonstrated blunted microvascular reactivity during local thermal heating test and decreased microcirculatory oxygenation in SCA children compared to controls. Multivariate analysis revealed that increased blood viscosity and decreased microcirculatory oxygenation were independent risk factors of frequent VOC in SCA. In contrast, the level of microvascular dysfunction does not predict VOC rate. In conclusion, increased blood viscosity is usually well supported in healthy individuals where vascular function is not impaired. However, in the context of SCA, microvascular function is impaired and any increase of blood viscosity or decrease in microcirculatory oxygenation would increase the risks for frequent VOC. Copyright © 2015 Elsevier Inc. All rights reserved.

Inhibition of dipeptidyl peptidase 4 regulates microvascular endothelial growth induced by inflammatory cytokines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takasawa, Wataru; Ohnuma, Kei; Hatano, Ryo

2010-10-08

Research highlights: {yields} TNF-{alpha} or IL-1{beta} induces EC proliferation with reduction of CD26 expression. {yields} CD26 siRNA or DPP-4 inhibition enhances TNF-{alpha} or IL-1{beta}-induced EC proliferation. {yields} Loss of CD26/DPP-4 enhances aortic sprouting induced by TNF-{alpha} or IL-1{beta}. {yields} Capillary formation induced by TNF-{alpha} or IL-1{beta} is enahced in the CD26{sup -/-} mice. -- Abstract: CD26/DPP-4 is abundantly expressed on capillary of inflamed lesion as well as effector T cells. Recently, CD26/dipeptidyl peptidase 4 (DPP-4) inhibition has been used as a novel oral therapeutic approach for patients with type 2 diabetes. While accumulating data indicate that vascular inflammation is amore » key feature of both micro- and macro-vascular complications in diabetes, the direct role of CD26/DPP-4 in endothelial biology is to be elucidated. We herein showed that proinflammatory cytokines such as tumor necrosis factor or interleukin-1 reduce expression of CD26 on microvascular endothelial cells, and that genetical or pharmacological inhibition of CD26/DPP-4 enhances endothelial growth both in vitro and in vivo. With DPP-4 inhibitors being used widely in the treatment of type 2 diabetes, our data strongly suggest that DPP-4 inhibition plays a pivotal role in endothelial growth and may have a potential role in the recovery of local circulation following diabetic vascular complications.« less

Endothelial mineralocorticoid receptor activation mediates endothelial dysfunction in diet-induced obesity

PubMed Central

Schäfer, Nicola; Lohmann, Christine; Winnik, Stephan; van Tits, Lambertus J.; Miranda, Melroy X.; Vergopoulos, Athanasios; Ruschitzka, Frank; Nussberger, Jürg; Berger, Stefan; Lüscher, Thomas F.; Verrey, François; Matter, Christian M.

2013-01-01

Received 22 July 2012; revised 29 January 2013; accepted 4 March 2013 Aims Aldosterone plays a crucial role in cardiovascular disease. ‘Systemic’ inhibition of its mineralocorticoid receptor (MR) decreases atherosclerosis by reducing inflammation and oxidative stress. Obesity, an important cardiovascular risk factor, is an inflammatory disease associated with increased plasma aldosterone levels. We have investigated the role of the ‘endothelial’ MR in obesity-induced endothelial dysfunction, the earliest stage in atherogenesis. Methods and results C57BL/6 mice were exposed to a normal chow diet (ND) or a high-fat diet (HFD) alone or in combination with the MR antagonist eplerenone (200 mg/kg/day) for 14 weeks. Diet-induced obesity impaired endothelium-dependent relaxation in response to acetylcholine, whereas eplerenone treatment of obese mice prevented this. Expression analyses in aortic endothelial cells isolated from these mice revealed that eplerenone attenuated expression of pro-oxidative NADPH oxidase (subunits p22phox, p40phox) and increased expression of antioxidative genes (glutathione peroxidase-1, superoxide dismutase-1 and -3) in obesity. Eplerenone did not affect obesity-induced upregulation of cyclooxygenase (COX)-1 or prostacyclin synthase. Endothelial-specific MR deletion prevented endothelial dysfunction in obese (exhibiting high ‘endogenous’ aldosterone) and in ‘exogenous’ aldosterone-infused lean mice. Pre-incubation of aortic rings from aldosterone-treated animals with the COX-inhibitor indomethacin restored endothelial function. Exogenous aldosterone administration induced endothelial expression of p22phox in the presence, but not in the absence of the endothelial MR. Conclusion Obesity-induced endothelial dysfunction depends on the ‘endothelial’ MR and is mediated by an imbalance of oxidative stress-modulating mechanisms. Therefore, MR antagonists may represent an attractive therapeutic strategy in the increasing population of obese patients to decrease vascular dysfunction and subsequent atherosclerotic complications. PMID:23594590

Effects of a Physical Activity Program on Markers of Endothelial Dysfunction, Oxidative Stress, and Metabolic Status in Adolescents with Metabolic Syndrome

PubMed Central

Camarillo-Romero, Eneida; Dominguez-Garcia, Ma Victoria; Amaya-Chavez, Araceli; Camarillo-Romero, Maria del Socorro; Talavera-Piña, Juan; Huitron-Bravo, Gerardo; Majluf-Cruz, Abraham

2012-01-01

The metabolic syndrome (MetS) is a precursor of diabetes. Physical activity (PA) improves endothelial dysfunction and may benefit patients with MetS. Aims. To evaluate the effect of a physical activity (PA) program on markers of endothelial dysfunction and oxidative stress in adolescents with (MetS). Methods. We carried out a cohort study of 38 adolescents with and without MetS (18 females and 20 males). All participants completed a 3-month PA program. All variables of the MetS as well as markers of endothelial dysfunction and oxidative stress tests were evaluated. Results. Females with and without MetS showed significant differences for almost all components of the MetS, whereas males were significantly different in half of the components. After the PA program, components of the MetS were not different from baseline values except for HDL-C levels. Some baseline endothelial dysfunction markers were significantly different among adolescents with and without MetS; however, after the PA program, most of these markers significantly improved in subjects with and without MetS. Conclusion. PA improves the markers of endothelial dysfunction in adolescents with MetS although other changes in the components of the MetS were not observed. Perhaps the benefits of PA on all components of MetS would appear after a PA program with a longer duration. PMID:22888450

Febuxostat attenuates paroxysmal atrial fibrillation-induced regional endothelial dysfunction.

PubMed

Li, YanGuang; Chen, FuKun; Deng, Long; Lin, Kun; Shi, Xiangmin; Zhaoliang, Shan; Wang, YuTang

2017-01-01

Paroxysmal atrial fibrillation (PAF) can increase thrombogenesis risk, especially in the left atrium (LA). The exact mechanism is still unclear. We assessed the effects of PAF on endothelial function, and investigated if febuxostat (FX) can attenuate endothelial dysfunction by inhibition of xanthine oxidase (XO). Eighteen male New Zealand white rabbits were divided randomly into sham-operated (S), PAF (P) or FX+pacing (FP) groups. Group P and group FP received rapid atrial pacing (RAP). Group FP was administered febuxostat (FX) for 7days before RAP. Post-procedure, blood samples were collected from the LA, right atrium (RA) and peripheral circulation. Tissues from the LA and RA were obtained. Endothelial dysfunction (thrombomodulin [TM], von Willebrand factor [VWF], asymmetric dimethylarginine [ADMA]), and indirect thrombin generation (thrombin-antithrombin complex [TAT], prothrombin fragment 1+2 [F1.2]) and oxidative stress in atrial tissue (xanthine oxidase [XO], superoxide dismutase [SOD], malondialdehyde [MDA]) were measured using an Enzyme-linked immunosorbent assay. Atrial endothelial expression of TM and VWF was measured by histology/western blotting. Endothelial dysfunction (TM, VWF, ADMA), TAT generation and oxidative stress (XO, SOD, MDA) in group P were more significant compared with that in group S (p<0.05, respectively). In group P, all of these changes occurred to a greater extent in the LA compared with those in the RA or peripheral circulation. In group FP, FX attenuated endothelial dysfunction and reduced TAT levels by inhibition of XO-mediated oxidative stress. PAF can lead to endothelial dysfunction and TAT generation by XO-mediated oxidative stress. The LA is more susceptible to these effects. FX can attenuate these changes by inhibition XO and XO-mediated oxidative stress. Copyright © 2016. Published by Elsevier Ltd.

You're Only as Old as Your Arteries: Translational Strategies for Preserving Vascular Endothelial Function with Aging

PubMed Central

Kaplon, Rachelle E.; Gioscia-Ryan, Rachel A.; LaRocca, Thomas J.

2014-01-01

Endothelial dysfunction develops with age and increases the risk of age-associated vascular disorders. Nitric oxide insufficiency, oxidative stress, and chronic low-grade inflammation, induced by upregulation of adverse cellular signaling processes and imbalances in stress resistance pathways, mediate endothelial dysfunction with aging. Healthy lifestyle behaviors preserve endothelial function with aging by inhibiting these mechanisms, and novel nutraceutical compounds that favorably modulate these pathways hold promise as a complementary approach for preserving endothelial health. PMID:24985329

Endothelial dysfunction in the regulation of portal hypertension

PubMed Central

Iwakiri, Yasuko

2013-01-01

Portal hypertension is caused by an increased intrahepatic resistance, a major consequence of cirrhosis. Endothelial dysfunction in liver sinusoidal endothelial cells (LSECs) decreases the production of vasodilators, such as nitric oxide (NO) and favors vasoconstriction. This contributes to an increased vascular resistance in the intrahepatic/sinusoidal microcirculation. Portal hypertension, once developed, causes endothelial cell (EC) dysfunction in the extrahepatic, i.e. splanchnic and systemic, circulation. Unlike LSEC dysfunction, EC dysfunction in the splanchnic and systemic circulation overproduces vasodilator molecules, leading to arterial vasodilatation. In addition, portal hypertension leads to the formation of portosystemic collateral vessels. Both arterial vasodilatation and portosystemic collateral vessel formation exacerbate portal hypertension by increasing the blood flow through the portal vein. Pathologic consequences, such as esophageal varices and ascites, result. While the sequence of pathological vascular events in cirrhosis and portal hypertension have been elucidated, the underlying cellular and molecular mechanisms causing EC dysfunctions are not yet fully understood. This review article summarizes the current cellular and molecular studies on EC dysfunctions found during the development of cirrhosis and portal hypertension with a focus on intra- and extrahepatic circulation. The article ends by discussing future directions of study for EC dysfunctions. PMID:21745318

Early microvascular changes in the preterm neonate: a comparative study of the human and guinea pig.

PubMed

Dyson, Rebecca M; Palliser, Hannah K; Lakkundi, Anil; de Waal, Koert; Latter, Joanna L; Clifton, Vicki L; Wright, Ian M R

2014-09-17

Dysfunction of the transition from fetal to neonatal circulatory systems may be a major contributor to poor outcome following preterm birth. Evidence exists in the human for both a period of low flow between 5 and 11 h and a later period of increased flow, suggesting a hypoperfusion-reperfusion cycle over the first 24 h following birth. Little is known about the regulation of peripheral blood flow during this time. The aim of this study was to conduct a comparative study between the human and guinea pig to characterize peripheral microvascular behavior during circulatory transition. Very preterm (≤28 weeks GA), preterm (29-36 weeks GA), and term (≥37 weeks GA) human neonates underwent laser Doppler analysis of skin microvascular blood flow at 6 and 24 h from birth. Guinea pig neonates were delivered prematurely (62 day GA) or at term (68-71 day GA) and laser Doppler analysis of skin microvascular blood flow was assessed every 2 h from birth. In human preterm neonates, there is a period of high microvascular flow at 24 h after birth. No period of low flow was observed at 6 h. In preterm animals, microvascular flow increased after birth, reaching a peak at 10 h postnatal age. Blood flow then steadily decreased, returning to delivery levels by 24 h. Preterm birth was associated with higher baseline microvascular flow throughout the study period in both human and guinea pig neonates. The findings do not support a hypoperfusion-reperfusion cycle in the microcirculation during circulatory transition. The guinea pig model of preterm birth will allow further investigation of the mechanisms underlying microvascular function and dysfunction during the initial extrauterine period. © 2014 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

In Vitro Endothelialization Test of Biomaterials Using Immortalized Endothelial Cells.

PubMed

Kono, Ken; Hiruma, Hitomi; Kobayashi, Shingo; Sato, Yoji; Tanaka, Masaru; Sawada, Rumi; Niimi, Shingo

2016-01-01

Functionalizing biomaterials with peptides or polymers that enhance recruitment of endothelial cells (ECs) can reduce blood coagulation and thrombosis. To assess endothelialization of materials in vitro, primary ECs are generally used, although the characteristics of these cells vary among the donors and change with time in culture. Recently, primary cell lines immortalized by transduction of simian vacuolating virus 40 large T antigen or human telomerase reverse transcriptase have been developed. To determine whether immortalized ECs can substitute for primary ECs in material testing, we investigated endothelialization on biocompatible polymers using three lots of primary human umbilical vein endothelial cells (HUVEC) and immortalized microvascular ECs, TIME-GFP. Attachment to and growth on polymer surfaces were comparable between cell types, but results were more consistent with TIME-GFP. Our findings indicate that TIME-GFP is more suitable for in vitro endothelialization testing of biomaterials.

Withaferin A protects against palmitic acid-induced endothelial insulin resistance and dysfunction through suppression of oxidative stress and inflammation

PubMed Central

Batumalaie, Kalaivani; Amin, Muhammad Arif; Murugan, Dharmani Devi; Sattar, Munavvar Zubaid Abdul; Abdullah, Nor Azizan

2016-01-01

Activation of inflammatory pathways via reactive oxygen species (ROS) by free fatty acids (FFA) in obesity gives rise to insulin resistance and endothelial dysfunction. Withaferin A (WA), possesses both antioxidant and anti-inflammatory properties and therefore would be a good strategy to suppress palmitic acid (PA)-induced oxidative stress and inflammation and hence, insulin resistance and dysfunction in the endothelium. Effect of WA on PA-induced insulin resistance in human umbilical vein endothelial cells (HUVECs) was determined by evaluating insulin signaling mechanisms whilst effect of this drug on PA-induced endothelial dysfunction was determined in acetylcholine-mediated relaxation in isolated rat aortic preparations. WA significantly inhibited ROS production and inflammation induced by PA. Furthermore, WA significantly decreased TNF-α and IL-6 production in endothelial cells by specifically suppressing IKKβ/NF-κβ phosphorylation. WA inhibited inflammation-stimulated IRS-1 serine phosphorylation and improved the impaired insulin PI3-K signaling, and restored the decreased nitric oxide (NO) production triggered by PA. WA also decreased endothelin-1 and plasminogen activator inhibitor type-1 levels, and restored the impaired endothelium-mediated vasodilation in isolated aortic preparations. These findings suggest that WA inhibited both ROS production and inflammation to restore impaired insulin resistance in cultured endothelial cells and improve endothelial dysfunction in rat aortic rings. PMID:27250532

Endothelial Dysfunction in Human Diabetes Is Mediated by Wnt5a-JNK Signaling.

PubMed

Bretón-Romero, Rosa; Feng, Bihua; Holbrook, Monika; Farb, Melissa G; Fetterman, Jessica L; Linder, Erika A; Berk, Brittany D; Masaki, Nobuyuki; Weisbrod, Robert M; Inagaki, Elica; Gokce, Noyan; Fuster, Jose J; Walsh, Kenneth; Hamburg, Naomi M

2016-03-01

Endothelial dysfunction is linked to insulin resistance, inflammatory activation, and increased cardiovascular risk in diabetes mellitus; however, the mechanisms remain incompletely understood. Recent studies have identified proinflammatory signaling of wingless-type family member (Wnt) 5a through c-jun N-terminal kinase (JNK) as a regulator of metabolic dysfunction with potential relevance to vascular function. We sought to gain evidence that increased activation of Wnt5a-JNK signaling contributes to impaired endothelial function in patients with diabetes mellitus. We measured flow-mediated dilation of the brachial artery and characterized freshly isolated endothelial cells by protein expression, eNOS activation, and nitric oxide production in 85 subjects with type 2 diabetes mellitus (n=42) and age- and sex-matched nondiabetic controls (n=43) and in human aortic endothelial cells treated with Wnt5a. Endothelial cells from patients with diabetes mellitus displayed 1.3-fold higher Wnt5a levels (P=0.01) along with 1.4-fold higher JNK activation (P<0.01) without a difference in total JNK levels. Higher JNK activation was associated with lower flow-mediated dilation, consistent with endothelial dysfunction (r=0.53, P=0.02). Inhibition of Wnt5a and JNK signaling restored insulin and A23187-mediated eNOS activation and improved nitric oxide production in endothelial cells from patients with diabetes mellitus. In endothelial cells from nondiabetic controls, rWnt5a treatment inhibited eNOS activation replicating the diabetic endothelial phenotype. In human aortic endothelial cells, Wnt5a-induced impairment of eNOS activation and nitric oxide production was reversed by Wnt5a and JNK inhibition. Our findings demonstrate that noncanonical Wnt5a signaling and JNK activity contribute to vascular insulin resistance and endothelial dysfunction and may represent a novel therapeutic opportunity to protect the vasculature in patients with diabetes mellitus. © 2016 American Heart Association, Inc.

Utilization of Microgravity Bioreactor for Differentiation and Growth of Human Vascular Endothelial Cells

NASA Technical Reports Server (NTRS)

Chen, Chu-Huang; Pellis, Neal R.

1997-01-01

The goal was to delineate mechanisms of genetic responses to angiogenic stimulation of human coronary arterial and dermal microvascular endothelial cells during exposure to microgravity. The NASA-designed rotating-wall vessel was used to create a three-dimensional culture environment with low shear-stress and microgravity simulating that in space. The primary specific aim was to determine whether simulated microgravity enhances endothelial cell growth and whether the growth enhancement is associated by augmented expression of Basic Fibroblast Growth Factor (BFGF) and c-fos, an immediate early gene and component of the transcription factor AP-1.

Differential Effects of Leptin and Adiponectin in Endothelial Angiogenesis

PubMed Central

Adya, Raghu; Tan, Bee K.; Randeva, Harpal S.

2015-01-01

Obesity is a major health burden with an increased risk of cardiovascular morbidity and mortality. Endothelial dysfunction is pivotal to the development of cardiovascular disease (CVD). In relation to this, adipose tissue secreted factors termed “adipokines” have been reported to modulate endothelial dysfunction. In this review, we focus on two of the most abundant circulating adipokines, that is, leptin and adiponectin, in the development of endothelial dysfunction. Leptin has been documented to influence a multitude of organ systems, that is, central nervous system (appetite regulation, satiety factor) and cardiovascular system (endothelial dysfunction leading to atherosclerosis). Adiponectin, circulating at a much higher concentration, exists in different molecular weight forms, essentially made up of the collagenous fraction and a globular domain, the latter being investigated minimally for its involvement in proinflammatory processes including activation of NF-κβ and endothelial adhesion molecules. The opposing actions of the two forms of adiponectin in endothelial cells have been recently demonstrated. Additionally, a local and systemic change to multimeric forms of adiponectin has gained importance. Thus detailed investigations on the potential interplay between these adipokines would likely result in better understanding of the missing links connecting CVD, adipokines, and obesity. PMID:25650072

Septic encephalopathy and septic encephalitis‬‬.

PubMed

Tauber, Simone C; Eiffert, Helmut; Brück, Wolfgang; Nau, Roland

2017-02-01

During the last two decades, septic encephalopathy (SE) was recognized as a clinically relevant problem with a high prevalence in patients at admission and during their hospital stay. SE is a condition associated with increased mortality and morbidity such as long-term cognitive impairment. Areas covered: This review illustrates the pathophysiology of sepsis-associated encephalopathy and encephalitis involving blood-brain-barrier dysfunction and neuroinflammation caused by endothelial and microglial activation by endogenous or pathogen-derived compounds, hypoxia by impaired microvascular regulation and septic shock as well as imbalance of neurotransmitters. The continuum between septic-embolic and septic-metastatic encephalitis and SE is underlined by histological findings. The options of technical examinations and biomarkers to diagnose SE are discussed together with established therapeutic options as well as current experimental approaches. Expert commentary: An outlook for clinicians is provided including promising diagnostic approaches by means of new imaging techniques. Clinical trials with drugs already established for other indications such as statins, erythropoietin and minocycline are warranted in the future.

Pathogenesis of Systemic Sclerosis

PubMed Central

Pattanaik, Debendra; Brown, Monica; Postlethwaite, Bradley C.; Postlethwaite, Arnold E.

2015-01-01

Systemic scleroderma (SSc) is one of the most complex systemic autoimmune diseases. It targets the vasculature, connective tissue-producing cells (namely fibroblasts/myofibroblasts), and components of the innate and adaptive immune systems. Clinical and pathologic manifestations of SSc are the result of: (1) innate/adaptive immune system abnormalities leading to production of autoantibodies and cell-mediated autoimmunity, (2) microvascular endothelial cell/small vessel fibroproliferative vasculopathy, and (3) fibroblast dysfunction generating excessive accumulation of collagen and other matrix components in skin and internal organs. All three of these processes interact and affect each other. The disease is heterogeneous in its clinical presentation that likely reflects different genetic or triggering factor (i.e., infection or environmental toxin) influences on the immune system, vasculature, and connective tissue cells. The roles played by other ubiquitous molecular entities (such as lysophospholipids, endocannabinoids, and their diverse receptors and vitamin D) in influencing the immune system, vasculature, and connective tissue cells are just beginning to be realized and studied and may provide insights into new therapeutic approaches to treat SSc. PMID:26106387

Association Between the Female Athlete Triad and Endothelial Dysfunction in Dancers

PubMed Central

Hoch, Anne Z.; Papanek, Paula; Szabo, Aniko; Widlansky, Michael E.; Schimke, Jane E.; Gutterman, David D.

2013-01-01

Objective To determine the prevalence of the 3 components of the female athlete triad [disordered eating, menstrual dysfunction, low bone mineral density (BMD)] and their relationships with brachial artery flow-mediated dilation in professional dancers. Design Prospective study. Setting Academic institution in the Midwest. Participants Twenty-two professional ballet dancers volunteered for this study. Interventions The prevalence of the female athlete triad and its relationship to endothelial dysfunction. Main Outcome Measures Subjects completed questionnaires to assess disordered eating and menstrual status/history. They also completed a 3-day food record and wore an accelerometer for 3 days to determine energy availability. Serum baseline thyrotropin, prolactin, and hormonal concentrations were obtained. Bone mineral density and body composition were measured with a GE Lunar Prodigy dual-energy X-ray absorptiometry. Endothelial function was determined as flow-mediated vasodilation measured by high-frequency ultrasound in the brachial artery. An increase in brachial diameter <5% to hyperemic flow stimulus was defined a priori as endothelial dysfunction. Results Seventeen dancers (77%) had evidence of low/negative energy availability. Thirty-two percent had disordered eating (EDE-Q score). Thirty-six percent had menstrual dysfunction and 14% were currently using hormone contraception. Twenty-three percent had evidence of low bone density (Z-score < −1.0). Sixty-four percent had abnormal brachial artery flow-mediated dilation (<5%). Flow-mediated dilation values were significantly correlated with serum estrogen and whole-body and lumbar BMD. All the 3 components of the triad plus endothelial dysfunction were present in 14% of the subjects. Conclusions Endothelial dysfunction was correlated with reduced BMD, menstrual dysfunction, and low serum estrogen. These findings may have profound implications for cardiovascular and bone health in professional women dancers. PMID:21358502

Endothelial dysfunction and amyloid-β-induced neurovascular alterations

PubMed Central

Koizumi, Kenzo; Wang, Gang; Park, Laibaik

2015-01-01

Alzheimer's disease (AD) and cerebrovascular diseases share common vascular risk factors that have disastrous effects on cerebrovascular regulation. Endothelial cells, lining inner walls of cerebral blood vessels, form a dynamic interface between the blood and the brain and are critical for the maintenance of neurovascular homeostasis. Accordingly, injury in endothelial cells is regarded as one of the earliest symptoms of impaired vasoregulatory mechanisms. Extracellular buildup of amyloid-β (Aβ) is a central pathogenic factor in AD. Aβ exerts potent detrimental effects on cerebral blood vessels and impairs endothelial structure and function. Recent evidence implicates vascular oxidative stress and activation of the nonselective cationic channel transient receptor potential melastatin (TRPM)-2 on endothelial cells in the mechanisms of Aβ-induced neurovascular dysfunction. Thus, Aβ triggers opening of TRPM2 channels in endothelial cells leading to intracellular Ca2+ overload and vasomotor dysfunction. The cerebrovascular dysfunction may contribute to AD pathogenesis by reducing the cerebral blood supply, leading to increased susceptibility to vascular insufficiency, and by promoting Aβ accumulation. The recent realization that vascular factors contribute to AD pathobiology suggests new targets for the prevention and treatment of this devastating disease. PMID:26328781

Proteomic study of endothelial dysfunction induced by AGEs and its possible role in diabetic cardiovascular complications.

PubMed

Banarjee, Reema; Sharma, Akshay; Bai, Shakuntala; Deshmukh, Arati; Kulkarni, Mahesh

2018-06-20

Endothelial dysfunction is one of the primary steps in the development of diabetes associated cardiovascular diseases. Hyperglycemic condition in diabetes promotes accumulation of advanced glycation end products (AGEs) in the plasma, that interact with the receptor for AGEs (RAGE) present on the endothelial cells and negatively affect their function. Using Human umbilical vascular endothelial cells (HUVECs) in culture, the effect of glycated human serum albumin on global proteomic changes was studied by SWATH-MS, a label free quantitative proteomic approach. Out of the 1860 proteins identified, 161 showed higher abundance while 123 showed lesser abundance in cells treated with glycated HSA. Bioinformatic analysis revealed that the differentially regulated proteins were involved in various processes such as apoptosis, oxidative stress etc. that are associated with endothelial dysfunction. Furthermore, the iRegulon analysis and immunofuorescence studies indicated that several of the differentially regulated proteins were transcriptionally regulated by NF-κB, that is downstream to AGE-RAGE axis. Some of the important differentially regulated proteins include ICAM1, vWF, PAI-1that affect important endothelial functions like cell adhesion and blood coagulation. qPCR analysis showed an increase in expression of the AGE receptor RAGE along with other genes involved in endothelial function. AGE treatment to HUVEC cells led to increased oxidative stress and apoptosis. This is the first proteomics study that provides insight into proteomic changes downstream to AGE-RAGE axis leading to endothelial dysfunction and predisposing to cardiovascular complications. Cardiovascular disease (CVD) is a major pathological outcome in diabetic patients and it is important to address ways that target its development before the onset. Elevated plasma AGEs in diabetes can affect endothelial function and can continue to show their effects even after blood glucose levels are back to normal. Since endothelial dysfunction acts as one of the initiating factors for the development of CVD, understanding how AGEs affect the endothelial cell proteome to cause dysfunction will provide insight into the mechanisms involved and aid designing new therapeutic approaches. Copyright © 2018. Published by Elsevier B.V.

Implications Enzymatic Degradation of the Endothelial Glycocalyx on the Microvascular Hemodynamics and the Arteriolar Red Cell Free Layer of the Rat Cremaster Muscle.

PubMed

Yalcin, Ozlem; Jani, Vivek P; Johnson, Paul C; Cabrales, Pedro

2018-01-01

The endothelial glycocalyx is a complex network of glycoproteins, proteoglycans, and glycosaminoglycans; it lines the vascular endothelial cells facing the lumen of blood vessels forming the endothelial glycocalyx layer (EGL). This study aims to investigate the microvascular hemodynamics implications of the EGL by quantifying changes in blood flow hydrodynamics post-enzymatic degradation of the glycocalyx layer. High-speed intravital microscopy videos of small arteries (around 35 μm) of the rat cremaster muscle were recorded at various time points after enzymatic degradation of the EGL. The thickness of the cell free layer (CFL), blood flow velocity profiles, and volumetric flow rates were quantified. Hydrodynamic effects of the presence of the EGL were observed in the differences between the thickness of CFL in microvessels with an intact EGL and glass tubes of similar diameters. Maximal changes in the thickness of CFL were observed 40 min post-enzymatic degradation of the EGL. Analysis of the frequency distribution of the thickness of CFL allows for estimation of the thickness of the endothelial surface layer (ESL), the plasma layer, and the glycocalyx. Peak flow, maximum velocity, and mean velocity were found to statistically increase by 24, 27, and 25%, respectively, after enzymatic degradation of the glycocalyx. The change in peak-to-peak maximum velocity and mean velocity were found to statistically increase by 39 and 32%, respectively, after 40 min post-enzymatic degradation of the EGL. The bluntness of blood flow velocity profiles was found to be reduced post-degradation of the EGL, as the exclusion volume occupied by the EGL increased the effective volume impermeable to RBCs in microvessels. This study presents the effects of the EGL on microvascular hemodynamics. Enzymatic degradation of the EGL resulted in a decrease in the thickness of CFL, an increase in blood velocity, blood flow, and decrease of the bluntness of the blood flow velocity profile in small arterioles. In summary, the EGL functions as a molecular sieve to solute transport and as a lubrication layer to protect the endothelium from red blood cell (RBC) motion near the vessel wall, determining wall shear stress.

In vitro analysis of human periodontal microvascular endothelial cells.

PubMed

Tsubokawa, Mizuki; Sato, Soh

2014-08-01

Endothelial cells (ECs) participate in key aspects of vascular biology, such as maintenance of capillary permeability, initiation of coagulation, and regulation of inflammation. According to previous reports, ECs have revealed highly specific characteristics depending on the organs and tissues. However, some reports have described the characteristics of the capillaries formed by human periodontal ECs. Therefore, the aim of the present study is to examine the functional characteristics of the periodontal microvascular ECs in vitro. Human periodontal ligament-endothelial cells (HPDL-ECs) and human gingiva-endothelial cells (HG-ECs) were isolated by immunoprecipitation with magnetic beads conjugated to a monoclonal anti-CD31 antibody. The isolated HPDL-ECs and HG-ECs were characterized to definitively demonstrate that these cell cultures represented pure ECs. Human umbilical-vein ECs and human dermal microvascular ECs were used for comparison. These cells were compared according to the proliferation potential, the formation of capillary-like tubes, the transendothelial electric resistance (TEER), and the expression of tight junction proteins. HPDL-ECs and HG-ECs with characteristic cobblestone monolayer morphology were obtained, as determined by light microscopy at confluence. Furthermore, the HPDL-ECs and HG-ECs expressed the EC markers platelet endothelial cell adhesion molecule-1 (also known as CD31), von Willebrand factor, and Ulex europaeus agglutinin 1, and the cells stained strongly positive for CD31 and CD309. In addition, the HPDL-ECs and HG-ECs were observed to form capillary-like tubes, and they demonstrated uptake of acetylated low-density lipoprotein. Functional analyses of the HPDL-ECs and HG-ECs showed that, compared to the control cells, tube formation persisted for only a brief period of time, and TEER was substantially reduced at confluence. Furthermore, the cells exhibited delocalization of zonula occludens-1 and occludin at cell-cell contact sites. The present results provide new evidence that HPDL-ECs and HG-ECs have characteristics of fenestrated capillaries. Therefore, capillaries in human periodontal tissues have functional characteristics of fenestrated capillaries, which might be related to the onset and the progression of systemic diseases and inflammation.

The Metalloprotease, Mpr1, Engages AnnexinA2 to Promote the Transcytosis of Fungal Cells across the Blood-Brain Barrier

PubMed Central

Na Pombejra, Sarisa; Salemi, Michelle; Phinney, Brett S.; Gelli, Angie

2017-01-01

Eukaryotic pathogens display multiple mechanisms for breaching the blood-brain barrier (BBB) and invading the central nervous system (CNS). Of the fungal spp., that cause disease in mammals, only some cross brain microvascular endothelial cells which constitute the BBB, and invade the brain. Cryptococcus neoformans, the leading cause of fungal meningoencephalitis, crosses the BBB directly by transcytosis or by co-opting monocytes. We previously determined that Mpr1, a secreted fungal metalloprotease, facilitates association of fungal cells to brain microvascular endothelial cells and we confirmed that the sole expression of CnMPR1 endowed S. cerevisiae with an ability to cross the BBB. Here, the gain of function conferred onto S. cerevisiae by CnMPR1 (i.e., Sc strain) was used to identify targets of Mpr1 that might reside on the surface of the BBB. Following biotin-labeling of BBB surface proteins, Sc-associated proteins were identified by LC-MS/MS. Of the 62 proteins identified several were cytoskeleton-endocytosis-associated including AnnexinA2 (AnxA2). Using an in vitro model of the human BBB where AnxA2 activity was blocked, we found that the lack of AnxA2 activity prevented the movement of S. cerevisiae across the BBB (i.e., transcytosis of Sc strain) but unexpectedly, TEM analysis revealed that AnxA2 was not required for the association or the internalization of Sc. Additionally, the co-localization of AnxA2 and Sc suggest that successful crossing of the BBB is dependent on an AxnA2-Mpr1-mediated interaction. Collectively the data suggest that AnxA2 plays a central role in fungal transcytosis in human brain microvascular endothelial cells. The movement and exocytosis of Sc is dependent on membrane trafficking events that involve AnxA2 but these events appear to be independent from the actions of AnxA2 at the host cell surface. We propose that Mpr1 activity promotes cytoskeleton remodeling in brain microvascular endothelial cells and thereby engages AnxA2 in order to facilitate fungal transcytosis of the BBB. PMID:28713781

The Metalloprotease, Mpr1, Engages AnnexinA2 to Promote the Transcytosis of Fungal Cells across the Blood-Brain Barrier.

PubMed

Na Pombejra, Sarisa; Salemi, Michelle; Phinney, Brett S; Gelli, Angie

2017-01-01

Eukaryotic pathogens display multiple mechanisms for breaching the blood-brain barrier (BBB) and invading the central nervous system (CNS). Of the fungal spp., that cause disease in mammals, only some cross brain microvascular endothelial cells which constitute the BBB, and invade the brain. Cryptococcus neoformans , the leading cause of fungal meningoencephalitis, crosses the BBB directly by transcytosis or by co-opting monocytes. We previously determined that Mpr1, a secreted fungal metalloprotease, facilitates association of fungal cells to brain microvascular endothelial cells and we confirmed that the sole expression of Cn MPR1 endowed S. cerevisiae with an ability to cross the BBB. Here, the gain of function conferred onto S. cerevisiae by Cn MPR1 (i.e., Sc strain) was used to identify targets of Mpr1 that might reside on the surface of the BBB. Following biotin-labeling of BBB surface proteins, Sc-associated proteins were identified by LC-MS/MS. Of the 62 proteins identified several were cytoskeleton-endocytosis-associated including AnnexinA2 (AnxA2). Using an in vitro model of the human BBB where AnxA2 activity was blocked, we found that the lack of AnxA2 activity prevented the movement of S. cerevisiae across the BBB (i.e., transcytosis of Sc strain) but unexpectedly, TEM analysis revealed that AnxA2 was not required for the association or the internalization of Sc. Additionally, the co-localization of AnxA2 and Sc suggest that successful crossing of the BBB is dependent on an AxnA2-Mpr1-mediated interaction. Collectively the data suggest that AnxA2 plays a central role in fungal transcytosis in human brain microvascular endothelial cells. The movement and exocytosis of Sc is dependent on membrane trafficking events that involve AnxA2 but these events appear to be independent from the actions of AnxA2 at the host cell surface. We propose that Mpr1 activity promotes cytoskeleton remodeling in brain microvascular endothelial cells and thereby engages AnxA2 in order to facilitate fungal transcytosis of the BBB.

Silver nanoparticles induce tight junction disruption and astrocyte neurotoxicity in a rat blood–brain barrier primary triple coculture model

PubMed Central

Xu, Liming; Dan, Mo; Shao, Anliang; Cheng, Xiang; Zhang, Cuiping; Yokel, Robert A; Takemura, Taro; Hanagata, Nobutaka; Niwa, Masami; Watanabe, Daisuke

2015-01-01

Background Silver nanoparticles (Ag-NPs) can enter the brain and induce neurotoxicity. However, the toxicity of Ag-NPs on the blood–brain barrier (BBB) and the underlying mechanism(s) of action on the BBB and the brain are not well understood. Method To investigate Ag-NP suspension (Ag-NPS)-induced toxicity, a triple coculture BBB model of rat brain microvascular endothelial cells, pericytes, and astrocytes was established. The BBB permeability and tight junction protein expression in response to Ag-NPS, NP-released Ag ions, and polystyrene-NP exposure were investigated. Ultrastructural changes of the microvascular endothelial cells, pericytes, and astrocytes were observed using transmission electron microscopy (TEM). Global gene expression of astrocytes was measured using a DNA microarray. Results A triple coculture BBB model of primary rat brain microvascular endothelial cells, pericytes, and astrocytes was established, with the transendothelial electrical resistance values >200 Ω·cm2. After Ag-NPS exposure for 24 hours, the BBB permeability was significantly increased and expression of the tight junction (TJ) protein ZO-1 was decreased. Discontinuous TJs were also observed between microvascular endothelial cells. After Ag-NPS exposure, severe mitochondrial shrinkage, vacuolations, endoplasmic reticulum expansion, and Ag-NPs were observed in astrocytes by TEM. Global gene expression analysis showed that three genes were upregulated and 20 genes were downregulated in astrocytes treated with Ag-NPS. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that the 23 genes were associated with metabolic processes, biosynthetic processes, response to stimuli, cell death, the MAPK pathway, and so on. No GO term and KEGG pathways were changed in the released-ion or polystyrene-NP groups. Ag-NPS inhibited the antioxidant defense of the astrocytes by increasing thioredoxin interacting protein, which inhibits the Trx system, and decreasing Nr4a1 and Dusp1. Meanwhile, Ag-NPS induced inflammation and apoptosis through modulation of the MAPK pathway or B-cell lymphoma-2 expression or mTOR activity in astrocytes. Conclusion These results draw our attention to the importance of Ag-NP-induced toxicity on the neurovascular unit and provide a better understanding of its toxicological mechanisms on astrocytes. PMID:26491287

[Epithelial dysfunction associated with pyo-inflammatory diseases of the ENT organs].

PubMed

Petukhova, N A

The modern concept of epithelial-endothelial dysfunction and epithelial-endothelial distress-syndrome associated with pyo-inflammatory ENT diseases is presented. It has provided a basis for the analysis of the initial stages of etiopathogenesis of acute and chronic inflammation in the ENT system including the mucous and associated lymphoid tissues as well as the Pirogov-Waldeyer limphopharyngeal ring making up the first protective barrier. The leading role of dysbiosis of synanthropic microflora and endotoxins of the Gram-negative bacteria in the mechanisms of regional responsiveness of the organism to the infection and chronic endotoxic aggression is demonstrated. The regional and synthetic mechanisms underlying the interaction between the external and internal media of the organism are subjected to the analysis with special reference to those operating in epithelium. The possible variants of the outcome of these processes are considered including both the recovery and the development of chronic inflammation. It has been proved that the exhaustion of the internal reserves for the stabilization of the epithelium-associated lymphoid tissue system including the Pirogov-Waldeyer limphopharyngeal ring leads to the formation of epithelial dysfunction as the initial stage of epithelial-endothelial dysfunction and epithelial-endothelial distress-syndrome. It is concluded that the modern concept of epithelial-endothelial dysfunction and epithelial-endothelial distress-syndrome is a fundamental interdisciplinary phenomenon.

Balance of autonomic nervous system in children having signs of endothelial dysfunction, that were born and are domiciled in contaminated territories.

PubMed

Kondrashova, V G; Kolpakov, I E; Vdovenko, V Yu; Leonovych, O S; Lytvynets, O M; Stepanova, E I

2014-09-01

Objective. The study examined the features of functional state of the autonomic nervous system in children having endothelial dysfunction and permanently residing in contaminated areas. Materials and methods. Clinical and instrumental examination of 101 children aged 7-18 years that were born and are domiciled in contaminated territories, including 37 persons with signs of endothelial dysfunction (subgroup IA) and 64 ones with no signs of endothelial dysfunction (IB subgroup) was conducted. The control group being comparable to the subgroups IA and IB by age, gender and clinical examination results included 37 children neither been domiciled in contaminated areas nor were belonging to the contingent of Chornobyl accident survivors. There were 20 apparently healthy children also examined. Results. Due to peculiarities of physiological pathways providing adaptive responses the children having signs of endothelial dysfunction are characterized by a more pronounced dysregulation of autonomous nervous system both in a resting state and under a functional load simulation, and also by a high strain of adaptation pathways. The lack of autonomous support of cardiovascular system is caused by inadequate adaptive responses of both central regulatory bodies (hypothalamus, vasomotor center) and peripheral receptors. Mainly the failure of segmental autonomous (parasympathetic) structures was revealed. The mode of their response to stress in this case corresponds to that in healthy individuals but at a lower functional level. There is a reduced aerobic capacity of the organism by the Robinson index, contributing to low adaptive range to non-specific stress in children being domiciled on contaminated territories including children having the endothelial dysfunction. Conclusions. Endothelial dysfunction was associated with more pronounced manifestations of autonomic dysregulation and reduced aerobic capacity of the organism being the risk factors of development of a range of somatic diseases requiring the development of prevention measures in children permanently residing in contaminated areas. autonomous nervous system balance, endothelial dysfunction, children, Chornobyl accident. V. G. Kondrashova, I. E. Kolpakov, V. Yu. Vdovenko, O. S. Leonovych, O. M. Lytvynets, E. I. Stepanova.

Monoamine oxidases are mediators of endothelial dysfunction in the mouse aorta.

PubMed

Sturza, Adrian; Leisegang, Matthias S; Babelova, Andrea; Schröder, Katrin; Benkhoff, Sebastian; Loot, Annemarieke E; Fleming, Ingrid; Schulz, Rainer; Muntean, Danina M; Brandes, Ralf P

2013-07-01

Monoamine oxidases (MAOs) generate H(2)O(2) as a by-product of their catalytic cycle. Whether MAOs are mediators of endothelial dysfunction is unknown and was determined here in the angiotensin II and lipopolysaccharide-models of vascular dysfunction in mice. Quantitative real-time polymerase chain reaction revealed that mouse aortas contain enzymes involved in catecholamine generation and MAO-A and MAO-B mRNA. MAO-A and -B proteins could be detected by Western blot not only in mouse aortas but also in human umbilical vein endothelial cells. Ex vivo incubation of mouse aorta with recombinant MAO-A increased H(2)O(2) formation and induced endothelial dysfunction that was attenuated by polyethylene glycol-catalase and MAO inhibitors. In vivo lipopolysaccharide (8 mg/kg IP overnight) or angiotensin II (1 mg/kg per day, 2 weeks, minipump) treatment induced vascular MAO-A and -B expressions and resulted in attenuated endothelium-dependent relaxation of the aorta in response to acetylcholine. MAO inhibitors reduced the lipopolysaccharide- and angiotensin II-induced aortic reactive oxygen species formation by 50% (ferrous oxidation xylenol orange assay) and partially normalized endothelium-dependent relaxation. MAO-A and MAO-B inhibitors had an additive effect; combined application completely restored endothelium-dependent relaxation. To determine how MAO-dependent H(2)O(2) formation induces endothelial dysfunction, cyclic GMP was measured. Histamine stimulation of human umbilical vein endothelial cells to activate endothelial NO synthase resulted in an increase in cyclic GMP, which was almost abrogated by MAO-A exposure. MAO inhibition prevented this effect, suggesting that MAO-induced H(2)O(2) formation is sufficient to attenuate endothelial NO release. Thus, MAO-A and MAO-B are both expressed in the mouse aorta, induced by in vivo lipopolysaccharide and angiotensin II treatment and contribute via the generation of H(2)O(2) to endothelial dysfunction in vascular disease models.

Inhibition of HMGCoA reductase by simvastatin protects mice from injurious mechanical ventilation.

PubMed

Manitsopoulos, Nikolaos; Orfanos, Stylianos E; Kotanidou, Anastasia; Nikitopoulou, Ioanna; Siempos, Ilias; Magkou, Christina; Dimopoulou, Ioanna; Zakynthinos, Spyros G; Armaganidis, Apostolos; Maniatis, Nikolaos A

2015-02-14

Mortality from severe acute respiratory distress syndrome exceeds 40% and there is no available pharmacologic treatment. Mechanical ventilation contributes to lung dysfunction and mortality by causing ventilator-induced lung injury. We explored the utility of simvastatin in a mouse model of severe ventilator-induced lung injury. Male C57BL6 mice (n = 7/group) were pretreated with simvastatin or saline and received protective (8 mL/kg) or injurious (25 mL/kg) ventilation for four hours. Three doses of simvastatin (20 mg/kg) or saline were injected intraperitoneally on days -2, -1 and 0 of the experiment. Lung mechanics, (respiratory system elastance, tissue damping and airway resistance), were evaluated by forced oscillation technique, while respiratory system compliance was measured with quasi-static pressure-volume curves. A pathologist blinded to treatment allocation scored hematoxylin-eosin-stained lung sections for the presence of lung injury. Pulmonary endothelial dysfunction was ascertained by bronchoalveolar lavage protein content and lung tissue expression of endothelial junctional protein Vascular Endothelial cadherin by immunoblotting. To assess the inflammatory response in the lung, we determined bronchoalveolar lavage fluid total cell content and neutrophil fraction by microscopy and staining in addition to Matrix-Metalloprotease-9 by ELISA. For the systemic response, we obtained plasma levels of Tumor Necrosis Factor-α, Interleukin-6 and Matrix-Metalloprotease-9 by ELISA. Statistical hypothesis testing was undertaken using one-way analysis of variance and Tukey's post hoc tests. Ventilation with high tidal volume (HVt) resulted in significantly increased lung elastance by 3-fold and decreased lung compliance by 45% compared to low tidal volume (LVt) but simvastatin abrogated lung mechanical alterations of HVt. Histologic lung injury score increased four-fold by HVt but not in simvastatin-pretreated mice. Lavage pleocytosis and neutrophilia were induced by HVt but were significantly attenuated by simvastatin. Microvascular protein permeability increase 20-fold by injurious ventilation but only 4-fold with simvastatin. There was a 3-fold increase in plasma Tumor Necrosis Factor-α, a 7-fold increase in plasma Interleukin-6 and a 20-fold increase in lavage fluid Matrix-Metalloprotease-9 by HVt but simvastatin reduced these levels to control. Lung tissue vascular endothelial cadherin expression was significantly reduced by injurious ventilation but remained preserved by simvastatin. High-dose simvastatin prevents experimental hyperinflation lung injury by angioprotective and anti-inflammatory effects.

Shiga Toxin 2 and Lipopolysaccharide Induce Human Microvascular Endothelial Cells To Release Chemokines and Factors That Stimulate Platelet Function

PubMed Central

Guessous, Fadila; Marcinkiewicz, Marek; Polanowska-Grabowska, Renata; Kongkhum, Sudawadee; Heatherly, Daniel; Obrig, Tom; Gear, Adrian R. L.

2005-01-01

Shiga toxins (Stxs) produced by Shigella dysenteriae type 1 and enterohemorrhagic Escherichia coli are the most common cause of hemolytic-uremic syndrome (HUS). It is well established that vascular endothelial cells, mainly those located in the renal microvasculature, are targets for Stxs. The aim of the present research was to evaluate whether E. coli-derived Shiga toxin 2 (Stx2) incubated with human microvascular endothelial cells (HMEC-1) induces release of chemokines and other factors that might stimulate platelet function. HMEC-1 were exposed for 24 h in vitro to Stx2, lipopolysaccharide (LPS), or the Stx2-LPS combination, and chemokine production was assessed by immunoassay. More interleukin-8 was released than stromal cell-derived factor 1α (SDF-1α) or SDF-1β and RANTES. The Stx2-LPS combination potentiated chemokine release, but Stx2 alone caused more release of SDF-1α at 24 h than LPS or Stx2-LPS did. In the presence of low ADP levels, HMEC-1 supernatants activated platelet function assessed by classical aggregometry, single-particle counting, granule secretion, P-selectin exposure, and the formation of platelet-monocyte aggregates. Supernatants from HMEC-1 exposed only to Stx2 exhibited enhanced exposure of platelet P-selectin and platelet-THP-1 cell interactions. Blockade of platelet cyclooxygenase by indomethacin prevented functional activation. The chemokine RANTES enhanced platelet aggregation induced by SDF-1α, macrophage-derived chemokine, or thymus and activation-regulated chemokine in the presence of very low ADP levels. These data support the hypothesis that microvascular endothelial cells exposed to E. coli O157:H7-derived Stx2 and LPS release chemokines and other factors, which when combined with low levels of primary agonists, such as ADP, cause platelet activation and promote the renal thrombosis associated with HUS. PMID:16299328

Phthalimide neovascular factor 1 (PNF1) modulates MT1-MMP activity in human microvascular endothelial cells

PubMed Central

Wieghaus, Kristen A.; Gianchandani, Erwin P.; Neal, Rebekah A.; Paige, Mikell A.; Brown, Milton L.; Papin, Jason A.; Botchwey, Edward A.

2009-01-01

We are creating synthetic pharmaceuticals with angiogenic activity and potential to promote vascular invasion. We previously demonstrated that one of these molecules, phthalimide neovascular factor 1 (PNF1), significantly expands microvascular networks in vivo following sustained release from poly(lactic-co-glycolic acid) (PLAGA) films. In addition, to probe PNF1 mode-of-action, we recently applied a novel pathway-based compendium analysis to a multi-timepoint, controlled microarray dataset of PNF1-treated (versus control) human microvascular endothelial cells (HMVECs), and we identified induction of tumor necrosis factor-alpha (TNF-α) and, subsequently, transforming growth factor-beta (TGF-β) signaling networks by PNF1. Here we validate this microarray data-set with quantitative real-time polymerase chain reaction (RT-PCR) analysis. Subsequently, we probe this dataset and identify three specific TGF-β-induced genes with regulation by PNF1 conserved over multiple timepoints—amyloid beta (A4) precursor protein (APP), early growth response 1 (EGR-1), and matrix metalloproteinase 14 (MMP14 or MT1-MMP)—that are also implicated in angiogenesis. We further focus on MMP14 given its unique role in angiogenesis, and we validate MT1-MMP modulation by PNF1 with an in vitro fluorescence assay that demonstrates the direct effects that PNF1 exerts on functional metalloproteinase activity. We also utilize endothelial cord formation in collagen gels to show that PNF1-induced stimulation of endothelial cord network formation in vitro is in some way MT1-MMP-dependent. Ultimately, this new network analysis of our transcriptional footprint characterizing PNF1 activity 1–48 h post-supplementation in HMVECs coupled with corresponding validating experiments suggests a key set of a few specific targets that are involved in PNF1 mode-of-action and important for successful promotion of the neovascularization that we have observed by the drug in vivo. PMID:19326468

Phthalimide neovascular factor 1 (PNF1) modulates MT1-MMP activity in human microvascular endothelial cells.

PubMed

Wieghaus, Kristen A; Gianchandani, Erwin P; Neal, Rebekah A; Paige, Mikell A; Brown, Milton L; Papin, Jason A; Botchwey, Edward A

2009-07-01

We are creating synthetic pharmaceuticals with angiogenic activity and potential to promote vascular invasion. We previously demonstrated that one of these molecules, phthalimide neovascular factor 1 (PNF1), significantly expands microvascular networks in vivo following sustained release from poly(lactic-co-glycolic acid) (PLAGA) films. In addition, to probe PNF1 mode of action, we recently applied a novel pathway-based compendium analysis to a multi-timepoint, controlled microarray data set of PNF1-treated (vs. control) human microvascular endothelial cells (HMVECs), and we identified induction of tumor necrosis factor-alpha (TNF-alpha) and, subsequently, transforming growth factor-beta (TGF-beta) signaling networks by PNF1. Here we validate this microarray data set with quantitative real-time polymerase chain reaction (RT-PCR) analysis. Subsequently, we probe this data set and identify three specific TGF-beta-induced genes with regulation by PNF1 conserved over multiple timepoints-amyloid beta (A4) precursor protein (APP), early growth response 1 (EGR-1), and matrix metalloproteinase 14 (MMP14 or MT1-MMP)-that are also implicated in angiogenesis. We further focus on MMP14 given its unique role in angiogenesis, and we validate MT1-MMP modulation by PNF1 with an in vitro fluorescence assay that demonstrates the direct effects that PNF1 exerts on functional metalloproteinase activity. We also utilize endothelial cord formation in collagen gels to show that PNF1-induced stimulation of endothelial cord network formation in vitro is in some way MT1-MMP-dependent. Ultimately, this new network analysis of our transcriptional footprint characterizing PNF1 activity 1-48 h post-supplementation in HMVECs coupled with corresponding validating experiments suggests a key set of a few specific targets that are involved in PNF1 mode of action and important for successful promotion of the neovascularization that we have observed by the drug in vivo.

Analysis of correlations between selected endothelial cell activation markers, disease activity, and nailfold capillaroscopy microvascular changes in systemic lupus erythematosus patients.

PubMed

Ciołkiewicz, Mariusz; Kuryliszyn-Moskal, Anna; Klimiuk, Piotr Adrian

2010-02-01

The aim of the study was to evaluate the correlation between selected serum endothelial cell activation markers such as vascular endothelial growth factor (VEGF), endothelin-1 (ET-1), soluble thrombomodulin (sTM), soluble E-selectin (sE-selectin), disease activity, and microvascular changes determined by nailfold capillaroscopy in patients with systemic lupus erythematosus (SLE). Serum levels of VEGF, ET-1, sTM, and sE-selectin were determined by an enzyme-linked immunosorbent assay in 80 SLE patients. The disease activity was measured with Systemic Lupus Erythematosus Disease Activity Index score. Nailfold capillaroscopy was performed in all patients. Positive correlation was found between VEGF and both ET-1 (r = 0.294, p < 0.01) and sE-selectin (r = 0.274, p < 0.05) serum levels as well as between sTM and ET-1 (r = 0.273, p < 0.05) serum concentrations. We noticed also positive correlation between VEGF (r = 0.224, p < 0.05) and ET-1 (r = 0.471, p < 0.001) serum levels and disease activity, and also between VEGF serum concentration and grade of morphological changes observed by nailfold capillaroscopy (r = 0.458, p < 0.001). There was also positive correlation between capillaroscopic score and disease activity (r = 0.339, p < 0.01). Our data suggest that correlation between VEGF and both ET-1 and E-selectin serum levels as well as between sTM and ET-1 serum concentrations may reflect their participation in the pathogenesis of SLE. VEGF seems to reflect changes in microcirculation in the course of SLE, visualised by nailfold capillaroscopy. The relationship between changes in nailfold capillaroscopy, endothelial cell activation markers, and the clinical activity of SLE points to an important role of microvascular abnormalities in the clinical manifestation of the disease.

Restoration of Autophagy in Endothelial Cells from Patients with Diabetes Mellitus Improves Nitric Oxide Signaling

PubMed Central

Fetterman, Jessica L.; Holbrook, Monica; Flint, Nir; Feng, Bihua; Bretón-Romero, Rosa; Linder, Erika A.; Berk, Brittany D.; Duess, Mai-Ann; Farb, Melissa G.; Gokce, Noyan; Shirihai, Orian S.; Hamburg, Naomi M.; Vita, Joseph A.

2016-01-01

Background Endothelial dysfunction contributes to cardiovascular disease in diabetes mellitus. Autophagy is a multistep mechanism for removal of damaged proteins and organelles from the cell. Under diabetic conditions, inadequate autophagy promotes cellular dysfunction and insulin resistance in non-vascular tissue. We hypothesized that impaired autophagy contributes to endothelial dysfunction in diabetes mellitus. Methods and Results We measured autophagy markers and endothelial nitric oxide synthase (eNOS) activation in freshly isolated endothelial cells from diabetic subjects (n=45) and non-diabetic controls (n=41). p62 levels were higher in cells from diabetics (34.2±3.6 vs. 20.0±1.6, P=0.001), indicating reduced autophagic flux. Bafilomycin inhibited insulin-induced activation of eNOS (−21±5% vs. 64±22%, P=0.003) in cells from controls, confirming that intact autophagy is necessary for eNOS signaling. In endothelial cells from diabetics, activation of autophagy with spermidine restored eNOS activation, suggesting that impaired autophagy contributes to endothelial dysfunction (P=0.01). Indicators of autophagy initiation including the number of LC3-bound puncta and beclin 1 expression were similar in diabetics and controls, whereas an autophagy terminal phase indicator, the lysosomal protein Lamp2a, was higher in diabetics. In endothelial cells under diabetic conditions, the beneficial effect of spermidine on eNOS activation was blocked by autophagy inhibitors bafilomycin or 3-methyladenine. Blocking the terminal stage of autophagy with bafilomycin increased p62 (P=0.01) in cells from diabetics to a lesser extent than in cells from controls (P=0.04), suggesting ongoing, but inadequate autophagic clearance. Conclusion Inadequate autophagy contributes to endothelial dysfunction in patients with diabetes and may be a target for therapy of diabetic vascular disease. PMID:26926601

Restoration of autophagy in endothelial cells from patients with diabetes mellitus improves nitric oxide signaling.

PubMed

Fetterman, Jessica L; Holbrook, Monica; Flint, Nir; Feng, Bihua; Bretón-Romero, Rosa; Linder, Erika A; Berk, Brittany D; Duess, Mai-Ann; Farb, Melissa G; Gokce, Noyan; Shirihai, Orian S; Hamburg, Naomi M; Vita, Joseph A

2016-04-01

Endothelial dysfunction contributes to cardiovascular disease in diabetes mellitus. Autophagy is a multistep mechanism for the removal of damaged proteins and organelles from the cell. Under diabetic conditions, inadequate autophagy promotes cellular dysfunction and insulin resistance in non-vascular tissue. We hypothesized that impaired autophagy contributes to endothelial dysfunction in diabetes mellitus. We measured autophagy markers and endothelial nitric oxide synthase (eNOS) activation in freshly isolated endothelial cells from diabetic subjects (n = 45) and non-diabetic controls (n = 41). p62 levels were higher in cells from diabetics (34.2 ± 3.6 vs. 20.0 ± 1.6, P = 0.001), indicating reduced autophagic flux. Bafilomycin inhibited insulin-induced activation of eNOS (64.7 ± 22% to -47.8 ± 8%, P = 0.04) in cells from controls, confirming that intact autophagy is necessary for eNOS signaling. In endothelial cells from diabetics, activation of autophagy with spermidine restored eNOS activation, suggesting that impaired autophagy contributes to endothelial dysfunction (P = 0.01). Indicators of autophagy initiation including the number of LC3-bound puncta and beclin 1 expression were similar in diabetics and controls, whereas an autophagy terminal phase indicator, the lysosomal protein Lamp2a, was higher in diabetics. In endothelial cells under diabetic conditions, the beneficial effect of spermidine on eNOS activation was blocked by autophagy inhibitors bafilomycin or 3-methyladenine. Blocking the terminal stage of autophagy with bafilomycin increased p62 (P = 0.01) in cells from diabetics to a lesser extent than in cells from controls (P = 0.04), suggesting ongoing, but inadequate autophagic clearance. Inadequate autophagy contributes to endothelial dysfunction in patients with diabetes and may be a target for therapy of diabetic vascular disease. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Aging impairs transcriptional regulation of vascular endothelial growth factor in human microvascular endothelial cells: implications for angiogenesis and cell survival.

PubMed

Ahluwalia, A; Jones, M K; Szabo, S; Tarnawski, A S

2014-04-01

In some tissues, aging impairs angiogenesis and reduces expression of vascular endothelial growth factor A (VEGF), a fundamental regulator of angiogenesis. We previously examined angiogenesis in aging and young gastric mucosa in vivo and in vitro and showed that an imbalance between expressions of VEGF (pro-angiogenic factor) and endostatin (anti-angiogenic protein) results in an aging-related impairment of angiogenesis in rats. However, the human relevance of these findings, and whether these mechanisms apply to endothelial cells derived from other tissues, is not clear. Since P-STAT3 and P-CREB are transcription factors that, in association with HIF-1α, can activate VEGF gene expression in some cells (e.g., liver cancer cells, vascular smooth muscle cells), we examined the expression of these two proteins in human dermal microvascular endothelial cells (HMVECs) derived from aging and neonatal individuals. We examined and quantified in vitro angiogenesis, expression of VEGF, P-STAT3, P-CREB and importin-α in HMVECs isolated from neonates (neonatal) and a 66 year old subject (aging). We also examined the effects of treatment with exogenous VEGF and endostatin on in vitro angiogenesis in these cells. Endothelial cells isolated from aging individuals had impaired angiogenesis (vs. neonatal endothelial cells) and reduced expression of VEGF mRNA and protein. Aged HMVECs also had reduced importin-α expression, and reduced expression and nuclear translocation of P-STAT3 and P-CREB. Reduced VEGF gene expression in aged HMVECs strongly correlated with the decreased levels of P-STAT3, P-CREB and importin-α in these cells. Our study clearly demonstrates that endothelial cells from aging individuals have impaired angiogenesis and reduced expression of VEGF likely due to impaired nuclear transport of P-STAT3 and P-CREB transcription factors in these cells.

Metabolic Profiling in Association with Vascular Endothelial Cell Dysfunction Following Non-Toxic Cadmium Exposure

PubMed Central

Li, Xiaofei; Nong, Qingjiao; Mao, Baoyu; Pan, Xue

2017-01-01

This study aimed to determine the metabolic profile of non-toxic cadmium (Cd)-induced dysfunctional endothelial cells using human umbilical vein endothelial cells (HUVECs). HUVECs (n = 6 per group) were treated with 0, 1, 5, or 10 μM cadmium chloride (CdCl2) for 48 h. Cell phenotypes, including nitric oxide (NO) production, the inflammatory response, and oxidative stress, were evaluated in Cd-exposed and control HUVECs. Cd-exposed and control HUVECs were analysed using gas chromatography time-of-flight/mass spectrometry. Compared to control HUVECs, Cd-exposed HUVECs were dysfunctional, exhibiting decreased NO production, a proinflammatory state, and non-significant oxidative stress. Further metabolic profiling revealed 24 significantly-altered metabolites in the dysfunctional endothelial cells. The significantly-altered metabolites were involved in the impaired tricarboxylic acid (TCA) cycle, activated pyruvate metabolism, up-regulated glucogenic amino acid metabolism, and increased pyrimidine metabolism. The current metabolic findings further suggest that the metabolic changes linked to TCA cycle dysfunction, glycosylation of the hexosamine biosynthesis pathway (HBP), and compensatory responses to genomic instability and energy deficiency may be generally associated with dysfunctional phenotypes, characterized by decreased NO production, a proinflammatory state, and non-significant oxidative stress, in endothelial cells following non-toxic Cd exposure. PMID:28872622

Endothelial Dysfunction in Human Diabetes is mediated by Wnt5a-JNK Signaling

PubMed Central

Bretón-Romero, Rosa; Feng, Bihua; Holbrook, Monika; Farb, Melissa G.; Fetterman, Jessica L.; Linder, Erika A.; Berk, Brittany D.; Masaki, Nobuyuki; Weisbrod, Robert M.; Inagaki, Elica; Gokce, Noyan; Fuster, Jose J.; Walsh, Kenneth; Hamburg, Naomi M.

2016-01-01

Objectives Endothelial dysfunction is linked to insulin resistance, inflammatory activation and increased cardiovascular risk in diabetes mellitus; however the mechanisms remain incompletely understood. Recent studies have identified pro-inflammatory signaling of Wnt5a through JNK as a regulator of metabolic dysfunction with potential relevance to vascular function. We sought to gain evidence that increased activation of Wnt5a-JNK signaling contributes to impaired endothelial function in patients with diabetes mellitus. Approach We measured flow-mediated dilation of the brachial artery and characterized freshly isolated endothelial cells by protein expression, eNOS activation, and nitric oxide production in from 85 subjects with Type 2 diabetes mellitus (n=42) and age- and sex-matched non-diabetic controls (n=43) and in human aortic endothelial cells treated with Wnt5a. Results Endothelial cells from patients with diabetes displayed 1.3-fold higher Wnt5a levels (P=0.01) along with 1.4-fold higher JNK activation (P<0.01) without a difference in total JNK levels. Higher JNK activation was associated with lower flow-mediated dilation, consistent with endothelial dysfunction (r=0.53, P=0.02). Inhibition of Wnt5a and JNK signaling restored insulin and A23187-mediated eNOS activation and improved nitric oxide production in endothelial cells from patients with diabetes. In endothelial cells from non-diabetic controls, rWnt5a treatment inhibited eNOS activation replicating the diabetic endothelial phenotype. In HAECs, Wnt5a-induced impairment of eNOS activation and nitric oxide production was reversed by Wnt5a and JNK inhibition. Conclusions Our findings demonstrate that non-canonical Wnt5a signaling and JNK activity contributes to vascular insulin resistance and endothelial dysfunction and may represent a novel therapeutic opportunity to protect the vasculature in patients with diabetes. PMID:26800561

Let-7i attenuates human brain microvascular endothelial cell damage in oxygen glucose deprivation model by decreasing toll-like receptor 4 expression.

PubMed

Xiang, Wei; Tian, Canhui; Peng, Shunli; Zhou, Liang; Pan, Suyue; Deng, Zhen

2017-11-04

The let-7 family of microRNAs (miRNAs) plays an important role on endothelial cell function. However, there have been few studies on their role under ischemic conditions. In this study, we demonstrate that let-7i, belonging to the let-7 family, rescues human brain microvascular endothelial cells (HBMECs) in an oxygen-glucose deprivation (OGD) model. Our data show that the expression of let-7 family miRNAs was downregulated after OGD. Overexpression of let-7i significantly alleviated cell death and improved survival of OGD-treated HBMECs. Let-7i also protected permeability in an in vitro blood brain barrier (BBB) model. Further, let-7i downregulated the expression of toll-like receptor 4 (TLR4), an inflammation trigger. Moreover, overexpression of let-7i decreased matrix metallopeptidase 9 (MMP9) and inducible nitric oxide synthase (iNOS) expression under OGD. Upon silencing TLR4 expression in HBMECs, the anti-inflammatory effect of let-7i was abolished. Our research suggests that let-7i promotes OGD-induced inflammation via downregulating TLR4 expression. Copyright © 2017 Elsevier Inc. All rights reserved.

Endoglin (CD105) expression in the development of haemorrhoids.

PubMed

Chung, Y-C; Hou, Y-C; Pan, A C-H

2004-02-01

Conventional pathogenesis of haemorrhoid emphasized the anchoring connective tissue system, whereas the vascular changes were ignored. The aim of this study was to clarify vascular changes of haemorrhoid disease. Forty-six samples of grade III and grade IV haemorrhoid tissue were selected for an in vitro study. We assessed the expressions in endoglin (CD105), an accessory protein in transforming growth factor-beta receptor complex, in CD34 and in vascular endothelial growth factor by using an immunohistochemical method. Microvascular density was calculated to correlate the expression of endoglin. Microvascular density was higher in haemorrhoid tissue than in normal anal and lower rectal tissues. CD34 was demonstrated in whole vessels in the haemorrhoids. However, endoglin, a proliferative marker of neovascularization, was present in only 25 of 46 (54%) haemorrhoidal vessels, and its immunoactivity was prominent in venules larger than 100 micro m. Thrombosis formation and stromal vascular endothelial growth factor was significantly associated with the presence of endoglin immunoactivity. The results of this study suggest that neovascularization is one important phenomenon of haemorrhoid disease, along with conventional venous dilatation and arteriovenous communication. In addition, thrombosis and stromal vascular endothelial growth factor might be important factors in promoting vascular proliferation.

Cerebral Microvascular Endothelial Cell Apoptosis after Ischemia: Role of Enolase-Phosphatase 1 Activation and Aci-Reductone Dioxygenase 1 Translocation

PubMed Central

Zhang, Yuan; Wang, Ting; Yang, Ke; Xu, Ji; Ren, Lijie; Li, Weiping; Liu, Wenlan

2016-01-01

Enolase-phosphatase 1 (ENOPH1), a newly discovered enzyme of the methionine salvage pathway, is emerging as an important molecule regulating stress responses. In this study, we investigated the role of ENOPH1 in blood brain barrier (BBB) injury under ischemic conditions. Focal cerebral ischemia induced ENOPH1 mRNA and protein expression in ischemic hemispheric microvessels in rats. Exposure of cultured brain microvascular endothelial cells (bEND3 cells) to oxygen-glucose deprivation (OGD) also induced ENOPH1 upregulation, which was accompanied by increased cell death and apoptosis reflected by increased 3-(4, 5-Dimethylthiazol-2-yl)-2, 5- diphenyltetrazolium bromide formation, lactate dehydrogenase release and TUNEL staining. Knockdown of ENOPH1 expression with siRNA or overexpressing ENOPH1 with CRISPR-activated plasmids attenuated or potentiated OGD-induced endothelial cell death, respectively. Moreover, ENOPH1 knockdown or overexpression resulted in a significant reduction or augmentation of reactive oxygen species (ROS) generation, apoptosis-associated proteins (caspase-3, PARP, Bcl-2 and Bax) and Endoplasmic reticulum (ER) stress proteins (Ire-1, Calnexin, GRP78 and PERK) in OGD-treated endothelial cells. OGD upregulated the expression of ENOPH1’s downstream protein aci-reductone dioxygenase 1 (ADI1) and enhanced its interaction with ENOPH1. Interestingly, knockdown of ENOPH1 had no effect on OGD-induced ADI1 upregulation, while it potentiated OGD-induced ADI1 translocation from the nucleus to the cytoplasm. Lastly, knockdown of ENOPH1 significantly reduced OGD-induced endothelial monolayer permeability increase. In conclusion, our data demonstrate that ENOPH1 activation may contribute to OGD-induced endothelial cell death and BBB disruption through promoting ROS generation and the activation of apoptosis associated proteins, thus representing a new therapeutic target for ischemic stroke. PMID:27630541

Cerebral Microvascular Endothelial Cell Apoptosis after Ischemia: Role of Enolase-Phosphatase 1 Activation and Aci-Reductone Dioxygenase 1 Translocation.

PubMed

Zhang, Yuan; Wang, Ting; Yang, Ke; Xu, Ji; Ren, Lijie; Li, Weiping; Liu, Wenlan

2016-01-01

Enolase-phosphatase 1 (ENOPH1), a newly discovered enzyme of the methionine salvage pathway, is emerging as an important molecule regulating stress responses. In this study, we investigated the role of ENOPH1 in blood brain barrier (BBB) injury under ischemic conditions. Focal cerebral ischemia induced ENOPH1 mRNA and protein expression in ischemic hemispheric microvessels in rats. Exposure of cultured brain microvascular endothelial cells (bEND3 cells) to oxygen-glucose deprivation (OGD) also induced ENOPH1 upregulation, which was accompanied by increased cell death and apoptosis reflected by increased 3-(4, 5-Dimethylthiazol-2-yl)-2, 5- diphenyltetrazolium bromide formation, lactate dehydrogenase release and TUNEL staining. Knockdown of ENOPH1 expression with siRNA or overexpressing ENOPH1 with CRISPR-activated plasmids attenuated or potentiated OGD-induced endothelial cell death, respectively. Moreover, ENOPH1 knockdown or overexpression resulted in a significant reduction or augmentation of reactive oxygen species (ROS) generation, apoptosis-associated proteins (caspase-3, PARP, Bcl-2 and Bax) and Endoplasmic reticulum (ER) stress proteins (Ire-1, Calnexin, GRP78 and PERK) in OGD-treated endothelial cells. OGD upregulated the expression of ENOPH1's downstream protein aci-reductone dioxygenase 1 (ADI1) and enhanced its interaction with ENOPH1. Interestingly, knockdown of ENOPH1 had no effect on OGD-induced ADI1 upregulation, while it potentiated OGD-induced ADI1 translocation from the nucleus to the cytoplasm. Lastly, knockdown of ENOPH1 significantly reduced OGD-induced endothelial monolayer permeability increase. In conclusion, our data demonstrate that ENOPH1 activation may contribute to OGD-induced endothelial cell death and BBB disruption through promoting ROS generation and the activation of apoptosis associated proteins, thus representing a new therapeutic target for ischemic stroke.

[The physiopathology of critical ischemia of the lower limbs].

PubMed

Novo, S; Abrignani, M G; Liquori, M; Sangiorgi, G B; Strano, A

1993-10-01

Peripheral obstructive arterial disease (POAD) of the lower limbs is the third main complication of atherosclerosis, after coronary artery disease and cerebrovascular disease. In 15-20% of cases POAD have an unfavourable evolution toward critical leg ischemia (CLI). This clinical condition is characterized by the onset of rest pain and/or trophic cutaneous lesions until gangrene appears. In some cases amputation is needed. The pathophysiological, clinical and therapeutic aspects of CLI were recently discussed in two Consensus Conferences held in Berlin in 1989 and in Rudesheim in 1991, with the elaboration of a final draft published on circulation. CLI appears when peripheral perfusion critically decreases due to macro and microcirculatory alterations. Atherosclerotic plaque is the primum movens, but often there are more plaques in sequence along the ilio-femoro-popliteal axis. The pathophysiological and clinical consequences are more severe if the stenosis is haemodynamically important, after a rapid progression of plaque growth or when thrombotic complications develop. The reduction in distal perfusion induces troubles in the microcirculation and an embalancement between the microvascular defense system (MDS) and the microvascular flow regulating system (MFRS) with endothelial dysfunction, platelet and leucocytes activation, worsening of blood viscosity due to the increase in fibrinogen levels and to the red cells deformability changes, activation of coagulation and impairment of fibrinolysis. So, a vicious circle appears with further worsening of distal perfusion and onset of trophic lesions. A further worsening of CLI can derive from local recurrent infections particularly frequent in diabetic patients.

New TiAg composite coating for bone prosthesis engineering shows promising microvascular compatibility in the murine dorsal skinfold chamber model.

PubMed

Behrendt, Ann-Kathrin; Beythien, Maximilian; Huber, Jakob; Zufraß, Thorsten; Butschkau, Antje; Mittlmeier, Thomas; Vollmar, Brigitte

2015-01-01

The incorporation of antimicrobial substances like silver into implant surface coatings is one promising concept against primary infections of endoprosthesis, especially for immunocompromised patients as well as against reinfection after revision operations. However, besides good antimicrobial and mechanical properties it is equally important that the implant material does not disturb the local microvascular perfusion of muscle tissue to enable microbial host defense and tissue repair processes. In this study the biocompatibility of a newly developed TiAg-composite coating applied on conventional titanium via physical vapor deposition was analysed. To evaluate the local microvascular and inflammatory response of striated muscle tissue upon implantation of TiAg-coated plates the murine dorsal skinfold chamber model was used. We repetitively examined local capillary and venular perfusion, endothelial integrity as well as leucocyte activation by intravital fluorescence microscopy at 1 h, 24 h as well as 3 and 7 days after implantation. TiAg-implants were well tolerated by the vascular system as indicated by intact functional capillary density and endothelial integrity compared to pure titanium plates and controls without a metal implant. Furthermore, quantification of rolling and adherent leucocytes did not reveal signs of inflammation upon TiAg-implantation.

Mechanisms of Endothelial Dysfunction in Hypertensive Pregnancy and Preeclampsia

PubMed Central

Possomato-Vieira, José S.; Khalil, Raouf A.

2016-01-01

Preeclampsia is a pregnancy-related disorder characterized by hypertension, and could lead to maternal and fetal morbidity and mortality. Although the causative factors and pathophysiological mechanisms are unclear, endothelial dysfunction is a major hallmark of preeclampsia. Clinical tests and experimental research have suggested that generalized endotheliosis in the systemic, renal, cerebral and hepatic circulation could decrease endothelium-derived vasodilators such as nitric oxide, prostacyclin and hyperpolarization factor and increase vasoconstrictors such as endothelin-1 and thromboxane A2, leading to increased vasoconstriction, hypertension and other manifestation of preeclampsia. In search for the upstream mechanisms that could cause endothelial dysfunction, certain genetic, demographic and environmental risk factors have been suggested to cause abnormal expression of uteroplacental integrins, cytokines and matrix metalloproteinases, leading to decreased maternal tolerance, apoptosis of invasive trophoblast cells, inadequate spiral arteries remodeling, reduced uterine perfusion pressure (RUPP), and placental ischemia/hypoxia. RUPP may cause imbalance between the anti-angiogenic factors soluble fms-like tyrosine kinase-1 and soluble endoglin and the pro-angiogenic factors vascular endothelial growth factor and placental growth factor, or stimulate the release of other circulating bioactive factors such as inflammatory cytokines, hypoxia-inducible factor-1, reactive oxygen species, and angiotensin AT1 receptor agonistic autoantibodies. These circulating factors could then target endothelial cells and cause generalized endothelial dysfunction. Therapeutic options are currently limited, but understanding the factors involved in endothelial dysfunction could help design new approaches for prediction and management of preeclampsia. PMID:27451103

Endothelial Cells Derived from the Blood-Brain Barrier and Islets of Langerhans Differ in their Response to the Effects of Bilirubin on Oxidative Stress Under Hyperglycemic Conditions.

PubMed

Kapitulnik, Jaime; Benaim, Clara; Sasson, Shlomo

2012-01-01

Unconjugated bilirubin (UCB) is a neurotoxic degradation product of heme. Its toxic effects include induction of apoptosis, and ultimately neuronal cell death. However, at low concentrations, UCB is a potent antioxidant that may protect cells and tissues against oxidative stress by neutralizing toxic metabolites such as reactive oxygen species (ROS). High glucose levels (hyperglycemia) generate reactive metabolites. Endothelial cell dysfunction, an early vascular complication in diabetes, has been associated with hyperglycemia-induced oxidative stress. Both glucose and UCB are substrates for transport proteins in microvascular endothelial cells of the blood-brain barrier (BBB). In the current study we show that UCB (1-40 μM) induces apoptosis and reduces survival of bEnd3 cells, a mouse brain endothelial cell line which serves as an in vitro model of the BBB. These deleterious effects of UCB were enhanced in the presence of high glucose (25 mM) levels. Interestingly, the bEnd3 cells exhibited an increased sensitivity to the apoptotic effects of UCB when compared to the MS1 microcapillary endothelial cell line. MS1 cells originate from murine pancreatic islets of Langerhans, and are devoid of the barrier characteristics of BBB-derived endothelial cells. ROS production was increased in both bEnd3 and MS1 cells exposed to high glucose, as compared with cells exposed to normal (5.5 mM) glucose levels. While UCB (0.1-40 μM) did not alter ROS production in cells exposed to normal glucose, relatively low ("physiological") UCB concentrations (0.1-5 μM) attenuated ROS generation in both cell lines exposed to high glucose levels. Most strikingly, higher UCB concentrations (20-40 μM) increased ROS generation in bEnd3 cells exposed to high glucose, but not in similarly treated MS1 cells. These results may be of critical importance for understanding the vulnerability of the BBB endothelium upon exposure to increasing UCB levels under hyperglycemic conditions.

Endothelial Cells Derived from the Blood-Brain Barrier and Islets of Langerhans Differ in their Response to the Effects of Bilirubin on Oxidative Stress Under Hyperglycemic Conditions

PubMed Central

Kapitulnik, Jaime; Benaim, Clara; Sasson, Shlomo

2012-01-01

Unconjugated bilirubin (UCB) is a neurotoxic degradation product of heme. Its toxic effects include induction of apoptosis, and ultimately neuronal cell death. However, at low concentrations, UCB is a potent antioxidant that may protect cells and tissues against oxidative stress by neutralizing toxic metabolites such as reactive oxygen species (ROS). High glucose levels (hyperglycemia) generate reactive metabolites. Endothelial cell dysfunction, an early vascular complication in diabetes, has been associated with hyperglycemia-induced oxidative stress. Both glucose and UCB are substrates for transport proteins in microvascular endothelial cells of the blood-brain barrier (BBB). In the current study we show that UCB (1–40 μM) induces apoptosis and reduces survival of bEnd3 cells, a mouse brain endothelial cell line which serves as an in vitro model of the BBB. These deleterious effects of UCB were enhanced in the presence of high glucose (25 mM) levels. Interestingly, the bEnd3 cells exhibited an increased sensitivity to the apoptotic effects of UCB when compared to the MS1 microcapillary endothelial cell line. MS1 cells originate from murine pancreatic islets of Langerhans, and are devoid of the barrier characteristics of BBB-derived endothelial cells. ROS production was increased in both bEnd3 and MS1 cells exposed to high glucose, as compared with cells exposed to normal (5.5 mM) glucose levels. While UCB (0.1–40 μM) did not alter ROS production in cells exposed to normal glucose, relatively low (“physiological”) UCB concentrations (0.1–5 μM) attenuated ROS generation in both cell lines exposed to high glucose levels. Most strikingly, higher UCB concentrations (20–40 μM) increased ROS generation in bEnd3 cells exposed to high glucose, but not in similarly treated MS1 cells. These results may be of critical importance for understanding the vulnerability of the BBB endothelium upon exposure to increasing UCB levels under hyperglycemic conditions. PMID:22811666

Cell-SELEX-Based Identification of a Human and Mouse Cross-Reactive Endothelial Cell-Internalizing Aptamer.

PubMed

Dua, Pooja; Kang, Sinae; Shin, Hye-Soo; Kim, Soyoun; Lee, Dong-Ki

2018-04-02

Increased interest and insights gained by researchers on the roles of endothelial cells in the pathophysiology of cancer, inflammatory, and cardiovascular diseases have led to the design of pharmacological interventions aimed at the endothelium lining in the diseased sites. Toward this end, we used established brain microvascular endothelial cell lines mouse (bEND3), human (hCMEC/D3), and Toggle Cell-SELEX to identify a species cross-reactive, endothelial cell-internalizing aptamer R11-3. This 2'F-modified RNA aptamer is specific for endothelial cells as no internalization was seen with cells of nonendothelial origin. R11-3 was truncated in size, and its potential in endothelial targeted therapeutics was established using VEGFR2 targeting long interfering RNA (liRNA) aptamer chimera. Due to its specificity for both mouse and human endothelial cells, we believe that this aptamer not only fits for development of endothelial targeted drug development for human diseases but is also suitable for preclinical evaluation in mice.

Prediabetes and Type 2 Diabetes Are Associated With Generalized Microvascular Dysfunction: The Maastricht Study.

PubMed

Sörensen, Ben M; Houben, Alfons J H M; Berendschot, Tos T J M; Schouten, Jan S A G; Kroon, Abraham A; van der Kallen, Carla J H; Henry, Ronald M A; Koster, Annemarie; Sep, Simone J S; Dagnelie, Pieter C; Schaper, Nicolaas C; Schram, Miranda T; Stehouwer, Coen D A

2016-11-01

Type 2 diabetes (T2DM) is associated with an increased risk of cardiovascular disease. This can be partly explained by large-artery dysfunction, which already occurs in prediabetes ("ticking clock hypothesis"). Whether a similar phenomenon also applies to microvascular dysfunction is not known. We therefore tested the hypothesis that microvascular dysfunction is already present in prediabetes and is more severe in T2DM. To do so, we investigated the associations of prediabetes, T2DM, and measures of hyperglycemia with microvascular function measured as flicker light-induced retinal arteriolar dilation and heat-induced skin hyperemia. In the Maastricht Study, a T2DM-enriched population-based cohort study (n=2213, 51% men, aged [mean±standard deviation] 59.7±8.2 years), we determined flicker light-induced retinal arteriolar %-dilation (Dynamic Vessel Analyzer), heat-induced skin %-hyperemia (laser-Doppler flowmetry), and glucose metabolism status (oral glucose tolerance test; normal glucose metabolism [n=1269], prediabetes [n=335], or T2DM [n=609]). Differences were assessed with multivariable regression analyses adjusted for age, sex, body mass index, smoking, physical activity, systolic blood pressure, lipid profile, retinopathy, estimated glomerular filtration rate, (micro)albuminuria, the use of lipid-modifying and blood pressure-lowering medication, and prior cardiovascular disease. Retinal arteriolar %-dilation was (mean±standard deviation) 3.4±2.8 in normal glucose metabolism, 3.0±2.7 in prediabetes, and 2.3±2.6 in T2DM. Adjusted analyses showed a lower arteriolar %-dilation in prediabetes (B=-0.20, 95% confidence interval -0.56 to 0.15) with further deterioration in T2DM (B=-0.61 [-0.97 to -0.25]) versus normal glucose metabolism (P for trend=0.001). Skin %-hyperemia was (mean±standard deviation) 1235±810 in normal glucose metabolism, 1109±748 in prediabetes, and 937±683 in T2DM. Adjusted analyses showed a lower %-hyperemia in prediabetes (B=-46 [-163 to 72]) with further deterioration in T2DM (B=-184 [-297 to -71]) versus normal glucose metabolism (P for trend=0.001). In addition, higher glycohemoglobin A1c and fasting plasma glucose were associated with lower retinal arteriolar %-dilation and skin %-hyperemia in fully adjusted models (for glycohemoglobin A1c, standardized B=-0.10 [-0.15 to -0.05], P<0.001 and standardized B=-0.13 [-0.19 to -0.07], P<0.001, respectively; for fasting plasma glucose, standardized B=-0.09 [-0.15 to -0.04], P<0.001 and standardized B=-0.10 [-0.15 to -0.04], P=0.002, respectively). Prediabetes, T2DM, and measures of hyperglycemia are independently associated with impaired microvascular function in the retina and skin. These findings support the concept that microvascular dysfunction precedes and thus may contribute to T2DM-associated cardiovascular disease and other complications, which may in part have a microvascular origin such as impaired cognition and heart failure. © 2016 American Heart Association, Inc.

Increased Angiotensin II Sensitivity Contributes to Microvascular Dysfunction in Women Who Have Had Preeclampsia.

PubMed

Stanhewicz, Anna E; Jandu, Sandeep; Santhanam, Lakshmi; Alexander, Lacy M

2017-08-01

Women who have had preeclampsia have increased cardiovascular disease risk; however, the mechanism(s) responsible for this association remain unclear. Microvascular damage sustained during a preeclamptic pregnancy may persist postpartum. The putative mechanisms mediating this dysfunction include a reduction in NO-dependent dilation and an increased sensitivity to angiotensin II. In this study, we evaluated endothelium-dependent dilation, angiotensin II sensitivity, and the therapeutic effect of angiotensin II receptor blockade (losartan) on endothelium-dependent dilation in vivo in the microvasculature of women with a history of preeclampsia (n=12) and control women who had a healthy pregnancy (n=12). We hypothesized that preeclampsia would have (1) reduced endothelium-dependent dilation, (2) reduced NO-mediated dilation, and (3) increased sensitivity to angiotensin II. We further hypothesized that localized losartan would increase endothelium-dependent vasodilation in preeclampsia. We assessed microvascular endothelium-dependent vasodilator function by measurement of cutaneous vascular conductance responses to graded infusion of acetylcholine (acetylcholine; 10 -7 -102 mmol/L) and a standardized local heating protocol in control sites and sites treated with 15 mmol/L L-NAME ( N G -nitro-l-arginine methyl ester; NO-synthase inhibitor) or 43 µmol/L losartan. Further, we assessed microvascular vasoconstrictor sensitivity to angiotensin II (10 -20 -10 -4 mol/L). Preeclampsia had significantly reduced endothelium-dependent dilation (-0.3±0.5 versus -1.0±0.4 log EC50 ; P <0.001) and NO-dependent dilation (16±3% versus 39±6%; P =0.006). Preeclampsia also had augmented vasoconstrictor sensitivity to angiotensin II (-10.2±1.3 versus -8.3±0.5; P =0.006). Angiotensin II type I receptor inhibition augmented endothelium-dependent vasodilation and NO-dependent dilation in preeclampsia but had no effect in healthy pregnancy. These data suggest that women who have had preeclampsia have persistent microvascular dysfunction postpartum, mediated, in part, by increased sensitivity to angiotensin II. © 2017 American Heart Association, Inc.

Role of endoplasmic reticulum stress in 12/15-lipoxygenase-induced retinal microvascular dysfunction in a mouse model of diabetic retinopathy.

PubMed

Elmasry, Khaled; Ibrahim, Ahmed S; Saleh, Heba; Elsherbiny, Nehal; Elshafey, Sally; Hussein, Khaled A; Al-Shabrawey, Mohamed

2018-05-01

Our earlier studies have established the role of 12/15-lipoxygenase (LO) in mediating the inflammatory reaction in diabetic retinopathy. However, the exact mechanism is still unclear. The goal of the current study was to identify the potential role of endoplasmic reticulum (ER) stress as a major cellular stress response in the 12/15-LO-induced retinal changes in diabetic retinopathy. We used in vivo and in vitro approaches. For in vivo studies, experimental diabetes was induced in wild-type (WT) mice and 12/15-Lo (also known as Alox15) knockout mice (12/15-Lo -/- ); ER stress was then evaluated after 12-14 weeks of diabetes. We also tested the effect of intravitreal injection of 12-hydroxyeicosatetraenoic acid (HETE) on retinal ER stress in WT mice and in mice lacking the catalytic subunit of NADPH oxidase, encoded by Nox2 (also known as Cybb) (Nox2 -/- mice). In vitro studies were performed using human retinal endothelial cells (HRECs) treated with 15-HETE (0.1 μmol/l) or vehicle, with or without ER stress or NADPH oxidase inhibitors. This was followed by evaluation of ER stress response, NADPH oxidase expression/activity and the levels of phosphorylated vascular endothelial growth factor receptor-2 (p-VEGFR2) by western blotting and immunoprecipitation assays. Moreover, real-time imaging of intracellular calcium (Ca 2+ ) release in HRECs treated with or without 15-HETE was performed using confocal microscopy. Deletion of 12/15-Lo significantly attenuated diabetes-induced ER stress in mouse retina. In vitro, 15-HETE upregulated ER stress markers such as phosphorylated RNA-dependent protein kinase-like ER-regulated kinase (p-PERK), activating transcription factor 6 (ATF6) and protein disulfide isomerase (PDI) in HRECs. Inhibition of ER stress reduced 15-HETE-induced-leucocyte adhesion, VEGFR2 phosphorylation and NADPH oxidase expression/activity. However, inhibition of NADPH oxidase or deletion of Nox2 had no effect on ER stress induced by the 12/15-LO-derived metabolites both in vitro and in vivo. We also found that 15-HETE increases the intracellular calcium in HRECs. ER stress contributes to 12/15-LO-induced retinal inflammation in diabetic retinopathy via activation of NADPH oxidase and VEGFR2. Perturbation of calcium homeostasis in the retina might also play a role in linking 12/15-LO to retinal ER stress and subsequent microvascular dysfunction in diabetic retinopathy.

[Prevention and preventive therapy of age-related macular degeneration through the beneficial effect of treatment of endothelial dysfunction].

PubMed

Fischer, Tamás

2006-12-24

The beneficial effect achieved by the treatment of endothelial dysfunction in chronic cardiovascular diseases is already an evidence belonging to the basic treatment of the disease. Given the fact that the vascular system is uniform and consubstantial both physiologically, pathophysiologically and in terms of therapy, and that it plays a key role in age-related macular degeneration (AMD) - a disease leading to tragic loss of vision with its etiology and therapy being unknown -, endothelial dysfunction should be treated. The pleiotropic effects of ACE-inhibitors, AR-blockers and statins help to restitute the balance between vasodilators and vasoconstrictors in endothelial dysfunction caused by oxidative stress, the balance of growth factors and their inhibitors, pro- and anti-inflammatory substances and prothrombotic and fibrinolytic factors, inhibit the formation of oxidative stress and its harmful effects; while aspirin with its pleiotropic effects acting as an antiaggregation substance on platelets helps to set the endothelial layer back to its normal balance regarding its vasodilating, antithrombotic, anti-adhesive and anti-inflammatory functions. For the above reasons it is suggested that, as a part of long term primary and/or secondary prevention, the following groups of patients with AMD receive - taking into consideration all possible side effects - ACE-inhibitor and/or AR-blocker and statin and aspirin treatment: 1) those without maculopathy but being over the age of 50 and having risk factors inducing endothelial dysfunction; 2) those, who already developed AMD in one eye as a prevention in the second, unaffected eye; and 3) those patients who developed AMD in both eyes in order to ameliorate or merely slow the progression of the disease. Besides, it is advisory to inhibit AMD risk factors inducing oxidative stress with consecutive endothelial dysfunction.

Ticagrelor protects against AngII-induced endothelial dysfunction by alleviating endoplasmic reticulum stress.

PubMed

Wang, Xiaoyu; Han, Xuejie; Li, Minghui; Han, Yu; Zhang, Yun; Zhao, Shiqi; Li, Yue

2018-05-16

Ticagrelor has been reported to decrease cardiovascular mortality compared with clopidogrel. This benefit cannot be fully explained by the more efficient platelet inhibition. Many studies demonstrated that ticagrelor improved endothelial function, leaving the mechanism elusive though. The present study aims to investigate whether ticagrelor protects against endothelial dysfunction induced by angiotensinII (AngII) through alleviating endoplasmic reticulum (ER) stress. Male Sprague Dawley rats were infused with AngII or vehicle and administrated with ticagrelor or vehicle for 14 days. Reactive oxygen species (ROS) was detected. Aortas from normal mice were incubated with endoplasmic reticulum stress inducer tunicamycin with or without ticagrelor. Vasorecactivity was measured on wire myography. Rat aortic endothelial cells (RAECs) were pretreated with ticagrelor followed by AngII or tunicamycin. Endothelial nitric oxide synthase (eNOS) phosphorylation and ER stress markers were determined by western blotting. Impaired endothelial function, induction of ER stress, reduced eNOS phosphorylation and elevated ROS generation was restored by ticagrelor treatment in vivo. In addition, tunicamycin induced endothelial dysfunction was improved by ticagrelor. In vitro, the induction of ER stress and inhibited eNOS phosphorylation in REACs exposed to AngII as well as tunicamycin was reversed by co-culturing with ticagrelor. In conclusion, ticagrelor protects against AngII-induced endothelial dysfunction via alleviating ER stress. Copyright © 2017. Published by Elsevier Inc.

Targeting vascular (endothelial) dysfunction

PubMed Central

Steven, Sebastian; Weber, Alina; Shuvaev, Vladimir V.; Muzykantov, Vladimir R.; Laher, Ismail; Li, Huige; Lamas, Santiago

2016-01-01

Abstract Cardiovascular diseases are major contributors to global deaths and disability‐adjusted life years, with hypertension a significant risk factor for all causes of death. The endothelium that lines the inner wall of the vasculature regulates essential haemostatic functions, such as vascular tone, circulation of blood cells, inflammation and platelet activity. Endothelial dysfunction is an early predictor of atherosclerosis and future cardiovascular events. We review the prognostic value of obtaining measurements of endothelial function, the clinical techniques for its determination, the mechanisms leading to endothelial dysfunction and the therapeutic treatment of endothelial dysfunction. Since vascular oxidative stress and inflammation are major determinants of endothelial function, we have also addressed current antioxidant and anti‐inflammatory therapies. In the light of recent data that dispute the prognostic value of endothelial function in healthy human cohorts, we also discuss alternative diagnostic parameters such as vascular stiffness index and intima/media thickness ratio. We also suggest that assessing vascular function, including that of smooth muscle and even perivascular adipose tissue, may be an appropriate parameter for clinical investigations. Linked Articles This article is part of a themed section on Redox Biology and Oxidative Stress in Health and Disease. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.12/issuetoc PMID:27187006

Inhibition of Endothelial Progenitor Cells May Explain the High Cardiovascular Event Rate in Patients with Rheumatoid Arthritis.

PubMed

Adawi, Mohamad; Pastuck, Nina; Saaida, Golan; Sirchan, Rizak; Watad, Abdalla; Blum, Arnon

2018-05-16

Rheumatoid arthritis (RA) patients may suffer cardiovascular (CV) events much more than the general population, and CV disease is the leading cause of death in patients with RA. Our hypothesis was that impaired function of endothelial progenitor cells may contribute to endothelial dysfunction and the clinical CV events of patients with RA. 27 RA patients (9 males and 18 females) with an active disease and 13 healthy subjects who served as the control group (9 males and 4 females) were enrolled to this prospective study. The ability to grow in culture colony-forming units of endothelial progenitor cells (CFU-EPCs) was measured, as well as their endothelial function using high-resolution ultrasonography of the brachial artery, and levels of C reactive protein (CRP) in the serum. For statistical analysis we used the students T-test test. As a group, patients with RA were older (p < 0.0001), had severe endothelial dysfunction (<0.0001), with impaired ability to grow CFU-EPCs (<0.0001), and a higher inflammatory state (p = 0001). No difference was observed in BMI. All RA patients had an active disease (DAS28 3.9±0.9) for 9.2±6.5 years. The same differences were observed in both genders. Patients with RA had an impaired ability to grow endothelial progenitor cells and severe endothelial dysfunction. Inability to grow colonies of endothelial progenitor cells reflects the impaired regenerative capacity of patients with RA, and may explain the endothelial dysfunction and the high CV event rate among patients with RA.

Pulmonary Cerium Dioxide Nanoparticles Exposure Differentially Impairs Coronary and Mesenteric Arteriolar Reactivity

PubMed Central

Minarchick, Valerie C; Stapleton, Phoebe A; Porter, Dale W; Wolfarth, Michael G; Çiftyürek, Engin; Barger, Mark; Sabolsky, Edward M.; Nurkiewicz, Timothy R

2013-01-01

Cerium dioxide nanoparticles (CeO2 NPs) are an engineered nanomaterial that possesses unique catalytic, oxidative and reductive properties. Currently, CeO2 NPs are being used as a fuel catalyst but these properties are also utilized in the development of potential drug treatments for radiation and stroke protection. These uses of CeO2 NPs present a risk for human exposure; however, to date no studies have investigated the effects of CeO2 NPs on the microcirculation following pulmonary exposure. Previous studies in our laboratory with other nanomaterials have shown impairments in normal microvascular function after pulmonary exposures. Therefore, we predicted that CeO2 NP exposure would cause microvascular dysfunction that is dependent on the tissue bed and dose. Twenty-four hour post exposure to CeO2 NPs (0–400 μg), mesenteric and coronary arterioles were isolated and microvascular function was assessed. Our results provided evidence that pulmonary CeO2 NP exposure impairs endothelium-dependent and -independent arteriolar dilation in a dose-dependent manner. The CeO2 NP exposure dose which causes a 50% impairment in arteriolar function (EC50) was calculated and ranged from 15 – 100 μg depending on the chemical agonist and microvascular bed. Microvascular assessments with acetylcholine revealed a 33–75% reduction in function following exposure. Additionally, there was a greater sensitivity to CeO2 NP exposure in the mesenteric microvasculature due to the 40% decrease in the calculated EC50 compared to the coronary microvasculature EC50. CeO2 NP exposure increased mean arterial pressure in some groups. Taken together these observed microvascular changes may likely have detrimental effects on local blood flow regulation and contribute to cardiovascular dysfunction associated with particle exposure. PMID:23645470

Effect of deoxyribozymes targeting c-Jun on solid tumor growth and angiogenesis in rodents.

PubMed

Zhang, Guishui; Dass, Crispin R; Sumithran, Eric; Di Girolamo, Nick; Sun, Lun-Quan; Khachigian, Levon M

2004-05-05

The basic region-leucine zipper protein c-Jun has been linked to cell proliferation, transformation, and apoptosis. However, a direct role for c-Jun in angiogenesis has not been shown. We used human microvascular endothelial cells (HMEC-1) transfected with a DNAzyme targeting the c-Jun mRNA (Dz13), related oligonucleotides, or vehicle in in vitro models of microvascular endothelial cell proliferation, migration, chemoinvasion, and tubule formation, a rat model of corneal neovascularization, and a mouse model of solid tumor growth and vascular endothelial growth factor (VEGF)-induced angiogenesis. All statistical tests were two-sided. Compared with mock-transfected cells, HMEC-1 cells transfected with Dz13 expressed less c-Jun protein and possessed lower DNA-binding activity. Dz13 blocked endothelial cell proliferation, migration, chemoinvasion, and tubule formation. Dz13 inhibited the endothelial cell expression and proteolytic activity of MMP-2, a c-Jun-dependent gene. Dz13 inhibited VEGF-induced neovascularization in the rat cornea compared with vehicle control (Dz13 versus vehicle: 4.0 neovessels versus 30.7 neovessels, difference = 26.7 neovessels; P =.004; area occupied by new blood vessels for Dz13 versus vehicle: 0.35 mm2 versus 1.52 mm2, difference = 1.17 mm2; P =.005) as well as solid melanoma growth in mice (Dz13 versus vehicle at 14 days: 108 mm3 versus 283 mm3, difference = 175 mm3; P =.006) with greatly reduced vascular density (Dz13 versus vehicle: 30% versus 100%, difference = 70%; P<.001). DNAzymes targeting c-Jun may have therapeutic potential as inhibitors of tumor angiogenesis and growth.

Upcyte® Microvascular Endothelial Cells Repopulate Decellularized Scaffold

PubMed Central

Dally, Iris; Hartmann, Nadja; Münst, Bernhard; Braspenning, Joris; Walles, Heike

2013-01-01

A general problem in tissue engineering is the poor and insufficient blood supply to guarantee tissue cell survival as well as physiological tissue function. To address this limitation, we have developed an in vitro vascularization model in which a decellularized porcine small bowl segment, representing a capillary network within a collagen matrix (biological vascularized scaffold [BioVaSc]), is reseeded with microvascular endothelial cells (mvECs). However, since the supply of mvECs is limited, in general, and as these cells rapidly dedifferentiate, we have applied a novel technology, which allows the generation of large batches of quasi-primary cells with the ability to proliferate, whilst maintaining their differentiated functionality. These so called upcyte mvECs grew for an additional 15 population doublings (PDs) compared to primary cells. Upcyte mvECs retained endothelial characteristics, such as von Willebrandt Factor (vWF), CD31 and endothelial nitric oxide synthase (eNOS) expression, as well as positive Ulex europaeus agglutinin I staining. Upcyte mvECs also retained biological functionality such as tube formation, cell migration, and low density lipoprotein (LDL) uptake, which were still evident after PD27. Initial experiments using MTT and Live/Dead staining indicate that upcyte mvECs repopulate the BioVaSc Scaffold. As with conventional cultures, these cells also express key endothelial molecules (vWF, CD31, and eNOS) in a custom-made bioreactor system even after a prolonged period of 14 days. The combination of upcyte mvECs and the BioVaSc represents a novel and promising approach toward vascularizing bioreactor models which can better reflect organs, such as the liver. PMID:22799502

Measurement of endothelial dysfunction via peripheral arterial tonometry predicts vasculogenic erectile dysfunction

PubMed Central

Kovac, Jason R.; Gomez, Lissette; Smith, Ryan P.; Coward, Robert M.; Gonzales, Marshall A.; Khera, Mohit; Lamb, Dolores J.; Lipshultz, Larry I.

2014-01-01

Introduction Endothelial cell dysfunction is associated with cardiovascular disease and vasculogenic erectile dysfunction (ED). Measured via Peripheral Artery Tonometry (PAT), endothelial dysfunction in the penis is an independent predictor of future cardiovascular events. Aim Determine whether measurement of endothelial dysfunction differentiates men with vasculogenic ED identified by duplex ultrasound from those without. Methods A total of 142 men were retrospectively assessed using patient history, penile duplex ultrasonography (US) and PAT (EndoPAT 2000). ED was self reported and identified on history. Vasculogenic ED was identified in men who exhibited a peak systolic velocity (PSV) of ≤25 cm/s obtained 15 minutes following vasodilator injection. The reactive hyperemia index (RHI), a measurement of endothelial dysfunction in medium/small arteries and the Augmentation Index (AI), a measurement of arterial stiffness, were recorded via PAT. Results Penile duplex US separated men into those with ED (n=111) and without (n=31). The cohort with ED had a PSV of 21±1 cm/s (left cavernous artery) and 22±1 cm/s (Right). The control group without ED had values of 39±2 cm/s (Left) and 39±2 (Right). Given the potential for altered endothelial function in diabetes mellitus, we confirmed that hemoglobin A1c, urinary microalbumin, and vibration pulse threshold were not different in men with vasculogenic ED and those without. RHI in patients with ED (1.85±0.06) was significantly decreased compared to controls (2.15±0.2) (p<0.05). The AI was unchanged when examined in isolation, and when standardized to heart rate. Conclusions Measurement of endothelial function with EndoPAT differentiates men with vasculogenic ED from those without. RHI could be used as a non-invasive surrogate in the assessment of vasculogenic ED and to identify those patients with higher cardiovascular risk. PMID:24784889

Endothelial dysfunction in metabolic diseases: role of oxidation and possible therapeutic employment of N-acetylcysteine.

PubMed

Masha, A; Martina, V

2014-01-01

Several metabolic diseases present a high cardiovascular mortality due to endothelial dysfunction consequences. In the last years of the past century, it has come to light that the endothelial cells, previously considered as inert in what regards an eventual secretion activity, play a pivotal role in regulating different aspects of the vascular function (endothelial function). It was clearly demonstrated that the endothelium acts as a real active organ, owning endocrine, paracrine and autocrine modulation activities by means of which it is able to regulate the vascular homeostasis. The present review will investigate the relationship between some metabolic diseases and the endothelial dysfunction and in particular the mechanisms underlying the effects of metabolic pathologies on the endothelium. Furthermore, it will consider the possible therapeutic employment of the N-acetilcysteine in such conditions.

Peripheral vascular dysfunction in migraine: a review

PubMed Central

2013-01-01

Numerous studies have indicated an increased risk of vascular disease among migraineurs. Alterations in endothelial and arterial function, which predispose to atherosclerosis and cardiovascular diseases, have been suggested as an important link between migraine and vascular disease. However, the available evidence is inconsistent. We aimed to review and summarize the published evidence about the peripheral vascular dysfunction of migraineurs. We systematically searched in BIOSIS, the Cochrane database, Embase, Google scholar, ISI Web of Science, and Medline to identify articles, published up to April 2013, evaluating the endothelial and arterial function of migraineurs. Several lines of evidence for vascular dysfunction were reported in migraineurs. Findings regarding endothelial function are particularly controversial since studies variously indicated the presence of endothelial dysfunction in migraineurs, the absence of any difference in endothelial function between migraineurs and non-migraineurs, and even an enhanced endothelial function in migraineurs. Reports on arterial function are more consistent and suggest that functional properties of large arteries are altered in migraineurs. Peripheral vascular function, particularly arterial function, is a promising non-invasive indicator of the vascular health of subjects with migraine. However, further targeted research is needed to understand whether altered arterial function explains the increased risk of vascular disease among patients with migraine. PMID:24083826

Assessment of endothelial dysfunction: the role of symmetrical dimethylarginine and proinflammatory markers in chronic kidney disease and renal transplant recipients.

PubMed

Memon, Lidija; Spasojevic-Kalimanovska, Vesna; Bogavac-Stanojevic, Natasa; Kotur-Stevuljevic, Jelena; Simic-Ogrizovic, Sanja; Giga, Vojislav; Dopsaj, Violeta; Jelic-Ivanovic, Zorana; Spasic, Slavica

2013-01-01

The study was designed to evaluate associations between symmetric dimethylarginine (SDMA), inflammation, and superoxide anion (O2∙-) with endothelial function and to determine their potential for screening of endothelial dysfunction in patients with chronic kidney disease (CKD) and renal transplant (RT) recipients. We included 64 CKD and 52 RT patients. Patients were stratified according to brachial artery flow-mediated dilation (FMD). Logistic regression analysis showed that high SDMA and high sensitive C-reactive protein (hs-CRP) were associated with impaired FMD in CKD and RT patients, after adjustment for glomerular filtration rate. The ability of inflammation, SDMA, and O2∙- to detect impaired FMD was investigated by receiving operative characteristic analysis. Hs-CRP (area under the curves (AUC) = 0.754, P < 0.001), IL-6 (AUC = 0.699, P = 0.002), and SDMA (AUC = 0.689, P = 0.007) had the highest ability to detect impaired FMD. SDMA in combination with inflammatory parameters and/or O2∙- had better screening performance than SDMA alone. Our results indicate a strong predictable association between hs-CRP, SDMA, and endothelial dysfunction in CKD patients and RT recipients. The individual marker that showed the strongest discriminative ability for endothelial dysfunction is hs-CRP, but its usefulness as a discriminatory marker for efficient diagnosis of endothelial dysfunction should be examined in prospective studies.

Inhibition of Vascular c-Jun N-Terminal Kinase 2 Improves Obesity-Induced Endothelial Dysfunction After Roux-en-Y Gastric Bypass.

PubMed

Doytcheva, Petia; Bächler, Thomas; Tarasco, Erika; Marzolla, Vincenzo; Engeli, Michael; Pellegrini, Giovanni; Stivala, Simona; Rohrer, Lucia; Tona, Francesco; Camici, Giovanni G; Vanhoutte, Paul M; Matter, Christian M; Lutz, Thomas A; Lüscher, Thomas F; Osto, Elena

2017-11-14

Roux-en-Y gastric bypass (RYGB) reduces obesity-associated comorbidities and cardiovascular mortality. RYGB improves endothelial dysfunction, reducing c-Jun N-terminal kinase (JNK) vascular phosphorylation. JNK activation links obesity with insulin resistance and endothelial dysfunction. Herein, we examined whether JNK1 or JNK2 mediates obesity-induced endothelial dysfunction and if pharmacological JNK inhibition can mimic RYGB vascular benefits. After 7 weeks of a high-fat high-cholesterol diet, obese rats underwent RYGB or sham surgery; sham-operated ad libitum-fed rats received, for 8 days, either the control peptide D-TAT or the JNK peptide inhibitor D-JNKi-1 (20 mg/kg per day subcutaneous). JNK peptide inhibitor D-JNKi-1 treatment improved endothelial vasorelaxation in response to insulin and glucagon-like peptide-1, as observed after RYGB. Obesity increased aortic phosphorylation of JNK2, but not of JNK1. RYGB and JNK peptide inhibitor D-JNKi-1 treatment blunted aortic JNK2 phosphorylation via activation of glucagon-like peptide-1-mediated signaling. The inhibitory phosphorylation of insulin receptor substrate-1 was reduced, whereas the protein kinase B/endothelial NO synthase pathway was increased and oxidative stress was decreased, resulting in improved vascular NO bioavailability. Decreased aortic JNK2 phosphorylation after RYGB rapidly improves obesity-induced endothelial dysfunction. Pharmacological JNK inhibition mimics the endothelial protective effects of RYGB. These findings highlight the therapeutic potential of novel strategies targeting vascular JNK2 against the severe cardiovascular disease associated with obesity. © 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.

Mitochondria-targeted antioxidant (MitoQ) ameliorates age-related arterial endothelial dysfunction in mice.

PubMed

Gioscia-Ryan, Rachel A; LaRocca, Thomas J; Sindler, Amy L; Zigler, Melanie C; Murphy, Michael P; Seals, Douglas R

2014-06-15

Age-related arterial endothelial dysfunction, a key antecedent of the development of cardiovascular disease (CVD), is largely caused by a reduction in nitric oxide (NO) bioavailability as a consequence of oxidative stress. Mitochondria are a major source and target of vascular oxidative stress when dysregulated. Mitochondrial dysregulation is associated with primary ageing, but its role in age-related endothelial dysfunction is unknown. Our aim was to determine the efficacy of a mitochondria-targeted antioxidant, MitoQ, in ameliorating vascular endothelial dysfunction in old mice. Ex vivo carotid artery endothelium-dependent dilation (EDD) to increasing doses of acetylcholine was impaired by ∼30% in old (∼27 months) compared with young (∼8 months) mice as a result of reduced NO bioavailability (P < 0.05). Acute (ex vivo) and chronic (4 weeks in drinking water) administration of MitoQ completely restored EDD in older mice by improving NO bioavailability. There were no effects of age or MitoQ on endothelium-independent dilation to sodium nitroprusside. The improvements in endothelial function with MitoQ supplementation were associated with the normalization of age-related increases in total and mitochondria-derived arterial superoxide production and oxidative stress (nitrotyrosine abundance), as well as with increases in markers of vascular mitochondrial health, including antioxidant status. MitoQ also reversed the age-related increase in endothelial susceptibility to acute mitochondrial damage (rotenone-induced impairment in EDD). Our results suggest that mitochondria-derived oxidative stress is an important mechanism underlying the development of endothelial dysfunction in primary ageing. Mitochondria-targeted antioxidants such as MitoQ represent a promising novel strategy for the preservation of vascular endothelial function with advancing age and the prevention of age-related CVD. © 2014 The Authors. The Journal of Physiology © 2014 The Physiological Society.

Mitochondria-targeted antioxidant (MitoQ) ameliorates age-related arterial endothelial dysfunction in mice

PubMed Central

Gioscia-Ryan, Rachel A; LaRocca, Thomas J; Sindler, Amy L; Zigler, Melanie C; Murphy, Michael P; Seals, Douglas R

2014-01-01

Age-related arterial endothelial dysfunction, a key antecedent of the development of cardiovascular disease (CVD), is largely caused by a reduction in nitric oxide (NO) bioavailability as a consequence of oxidative stress. Mitochondria are a major source and target of vascular oxidative stress when dysregulated. Mitochondrial dysregulation is associated with primary ageing, but its role in age-related endothelial dysfunction is unknown. Our aim was to determine the efficacy of a mitochondria-targeted antioxidant, MitoQ, in ameliorating vascular endothelial dysfunction in old mice. Ex vivo carotid artery endothelium-dependent dilation (EDD) to increasing doses of acetylcholine was impaired by ∼30% in old (∼27 months) compared with young (∼8 months) mice as a result of reduced NO bioavailability (P < 0.05). Acute (ex vivo) and chronic (4 weeks in drinking water) administration of MitoQ completely restored EDD in older mice by improving NO bioavailability. There were no effects of age or MitoQ on endothelium-independent dilation to sodium nitroprusside. The improvements in endothelial function with MitoQ supplementation were associated with the normalization of age-related increases in total and mitochondria-derived arterial superoxide production and oxidative stress (nitrotyrosine abundance), as well as with increases in markers of vascular mitochondrial health, including antioxidant status. MitoQ also reversed the age-related increase in endothelial susceptibility to acute mitochondrial damage (rotenone-induced impairment in EDD). Our results suggest that mitochondria-derived oxidative stress is an important mechanism underlying the development of endothelial dysfunction in primary ageing. Mitochondria-targeted antioxidants such as MitoQ represent a promising novel strategy for the preservation of vascular endothelial function with advancing age and the prevention of age-related CVD. PMID:24665093

A novel method for measuring hydraulic conductivity at the human blood-nerve barrier in vitro.

PubMed

Helton, E Scott; Palladino, Steven; Ubogu, Eroboghene E

2017-01-01

Microvascular barrier permeability to water is an essential biophysical property required for the homeostatic maintenance of unique tissue microenvironments. This is of particular importance in peripheral nerves where strict control of ionic concentrations is needed for axonal signal transduction. Previous studies have associated inflammation, trauma, toxin exposure and metabolic disease with increases in water influx and hydrostatic pressure in peripheral nerves with resultant endoneurial edema that may impair axonal function. The regulation of water permeability across endoneurial microvessels that form the blood-nerve barrier (BNB) is poorly understood. Variations exist in apparatus and methods used to measure hydraulic conductivity. The objective of the study was to develop a simplified hydraulic conductivity system using commercially available components to evaluate the BNB. We determined the mean hydraulic conductivity of cultured confluent primary and immortalized human endoneurial endothelial cell layers as 2.00×10 -7 and 2.17×10 -7 cm/s/cm H₂O respectively, consistent with restrictive microvascular endothelial cells in vitro. We also determined the mean hydraulic conductivity of immortalized human brain microvascular endothelial cell layers, a commonly used blood-brain barrier (BBB) cell line, as 0.20×10 -7 cm/s/cm H₂O, implying a mean 10-fold higher resistance to transendothelial water flux in the brain compared to peripheral nerves. To our knowledge, this is the first reported measurement of human BNB and BBB hydraulic conductivities. This model represents an important tool to further characterize the human BNB and deduce the molecular determinants and signaling mechanisms responsible for BNB hydraulic conductivity in normal and disease states in vitro. Copyright © 2016 Elsevier Inc. All rights reserved.

Hyperuricemia induces endothelial dysfunction via mitochondrial Na+/Ca2+ exchanger-mediated mitochondrial calcium overload.

PubMed

Hong, Quan; Qi, Ka; Feng, Zhe; Huang, Zhiyong; Cui, Shaoyuan; Wang, Liyuan; Fu, Bo; Ding, Rui; Yang, Jurong; Chen, Xiangmei; Wu, Di

2012-05-01

Uric acid (UA) has proven to be a causal agent in endothelial dysfunction in which ROS production plays an important role. Calcium overload in mitochondria can promote the mitochondrial production of ROS. We hypothesize that calcium transduction in mitochondria contributes to UA-induced endothelial dysfunction. We first demonstrated that high concentrations of UA cause endothelial dysfunction, marked by a reduction in eNOS protein expression and NO release in vitro. We further found that a high concentration of UA increased levels of [Ca2+]mito, total intracellular ROS, H2O2, and mitochondrial O2·-, and Δψmito but not the [Ca2+]cyt level. When the mitochondrial calcium channels NCXmito and MCU were blocked by CGP-37157 and Ru360, respectively, the UA-induced increases in the levels of [Ca2+]mito and total intracellular ROS were significantly reduced. Mitochondrial levels of O2·- and Δψmito were reduced by inhibition of NCXmito but not of MCU. Moreover, inhibition of NCXmito, but not of MCU, blocked the UA-induced reductions in eNOS protein expression and NO release. The increased generation of mitochondrial O2·- induced by a high concentration of UA is triggered by mitochondrial calcium overload and ultimately leads to endothelial dysfunction. In this process, the activation of NCXmito is the major cause of the influx of calcium into mitochondria. Our results provide a new pathophysiological mechanism for UA-induced endothelial dysfunction and may offer a new therapeutic target for clinicians. Copyright © 2012 Elsevier Ltd. All rights reserved.

Ezetimibe in Combination With Statins Ameliorates Endothelial Dysfunction in Coronary Arteries After Stenting: The CuVIC Trial (Effect of Cholesterol Absorption Inhibitor Usage on Target Vessel Dysfunction After Coronary Stenting), a Multicenter Randomized Controlled Trial.

PubMed

Takase, Susumu; Matoba, Tetsuya; Nakashiro, Soichi; Mukai, Yasushi; Inoue, Shujiro; Oi, Keiji; Higo, Taiki; Katsuki, Shunsuke; Takemoto, Masao; Suematsu, Nobuhiro; Eshima, Kenichi; Miyata, Kenji; Yamamoto, Mitsutaka; Usui, Makoto; Sadamatsu, Kenji; Satoh, Shinji; Kadokami, Toshiaki; Hironaga, Kiyoshi; Ichi, Ikuyo; Todaka, Koji; Kishimoto, Junji; Egashira, Kensuke; Sunagawa, Kenji

2017-02-01

We sought to investigate whether treatment with ezetimibe in combination with statins improves coronary endothelial function in target vessels in coronary artery disease patients after coronary stenting. We conducted a multicenter, prospective, randomized, open-label, blinded-end point trial among 11 cardiovascular treatment centers. From 2011 to 2013, 260 coronary artery disease patients who underwent coronary stenting were randomly allocated to 2 arms (statin monotherapy, S versus ezetimibe [10 mg/d]+statin combinational therapy, E+S). We defined target vessel dysfunction as the primary composite outcome, which comprised target vessel failure during treatment and at the 6- to 8-month follow-up coronary angiography and coronary endothelial dysfunction determined via intracoronary acetylcholine testing performed in cases without target vessel failure at the follow-up coronary angiography. Coadministration of ezetimibe with statins further lowered low-density lipoprotein cholesterol levels (83±23 mg/dL in S versus 67±23 mg/dL in E+S; P<0.0001), with significant decreases in oxidized low-density lipoprotein and oxysterol levels. Among patients without target vessel failure, 46 out of 89 patients (52%) in the S arm and 34 out of 96 patients (35%) in the E+S arm were found to have coronary endothelial dysfunction (P=0.0256), and the incidence of target vessel dysfunction at follow-up was significantly decreased in the E+S arm (69/112 (62%) in S versus 47/109 (43%) in E+S; P=0.0059). A post hoc analysis of post-treatment low-density lipoprotein cholesterol-matched subgroups revealed that the incidence of both target vessel dysfunction and coronary endothelial dysfunction significantly decreased in the E+S arm, with significant reductions in oxysterol levels. The CuVIC trial (Effect of Cholesterol Absorption Inhibitor Usage on Target Vessel Dysfunction after Coronary Stenting) has shown that ezetimibe with statins, compared with statin monotherapy, improves functional prognoses, ameliorating endothelial dysfunction in stented coronary arteries, and was associated with larger decreases in oxysterol levels. © 2016 American Heart Association, Inc.

Nitric oxide-mediated cutaneous microvascular function is impaired in polycystic ovary sydrome but can be improved by exercise training.

PubMed

Sprung, V S; Cuthbertson, D J; Pugh, C J A; Daousi, C; Atkinson, G; Aziz, N F; Kemp, G J; Green, D J; Cable, N T; Jones, H

2013-03-15

Polycystic ovary syndrome (PCOS) is associated with cardiovascular disease. The contribution of the nitric oxide (NO) dilator system to cutaneous endothelial dysfunction is currently unknown in PCOS. Our aim was to examine whether women with PCOS demonstrate impaired cutaneous microvascular NO function and whether exercise training can ameliorate any impairment. Eleven women with PCOS (age, 29 ± 7 years; body mass index, 34 ± 6 kg m(-2)) were compared with six healthy obese control women (age, 29 ± 7 years; body mass index, 34 ± 5 kg m(-2)). Six women with PCOS (30 ± 7 years; 31 ± 6 kg m(-2)) then completed 16 weeks of exercise training. Laser Doppler flowmetry, combined with intradermal microdialysis of l-N(G)-monomethyl-l-arginine, a nitric oxide antagonist, in response to incremental local heating of the forearm was assessed in women with PCOS and control women, and again in women with PCOS following exercise training. Cardiorespiratory fitness, homeostasis model assessment for insulin resistance, hormone and lipid profiles were also assessed. Differences between women with PCOS and control women and changes with exercise were analysed using Student's unpaired t tests. Differences in the contribution of NO to cutaneous blood flow [expressed as a percentage of maximal cutaneous vasodilatation (CVCmax)] were analysed using general linear models. At 42°C heating, cutaneous NO-mediated vasodilatation was attenuated by 17.5%CVCmax (95% confidence interval, 33.3, 1.7; P = 0.03) in women with PCOS vs. control women. Exercise training improved cardiorespiratory fitness by 5.0 ml kg(-1) min(-1) (95% confidence interval, 0.9, 9.2; P = 0.03) and NO-mediated cutaneous vasodilatation at 42°C heating by 19.6% CVCmax (95% confidence interval, 4.3, 34.9; P = 0.02). Cutaneous microvascular NO function is impaired in women with PCOS compared with obese matched control women but can be improved with exercise training.

Nitric oxide-mediated cutaneous microvascular function is impaired in polycystic ovary sydrome but can be improved by exercise training

PubMed Central

Sprung, V S; Cuthbertson, D J; Pugh, C J A; Daousi, C; Atkinson, G; Aziz, N F; Kemp, G J; Green, D J; Cable, N T; Jones, H

2013-01-01

Polycystic ovary syndrome (PCOS) is associated with cardiovascular disease. The contribution of the nitric oxide (NO) dilator system to cutaneous endothelial dysfunction is currently unknown in PCOS. Our aim was to examine whether women with PCOS demonstrate impaired cutaneous microvascular NO function and whether exercise training can ameliorate any impairment. Eleven women with PCOS (age, 29 ± 7 years; body mass index, 34 ± 6 kg m−2) were compared with six healthy obese control women (age, 29 ± 7 years; body mass index, 34 ± 5 kg m−2). Six women with PCOS (30 ± 7 years; 31 ± 6 kg m−2) then completed 16 weeks of exercise training. Laser Doppler flowmetry, combined with intradermal microdialysis of l-NG-monomethyl-l-arginine, a nitric oxide antagonist, in response to incremental local heating of the forearm was assessed in women with PCOS and control women, and again in women with PCOS following exercise training. Cardiorespiratory fitness, homeostasis model assessment for insulin resistance, hormone and lipid profiles were also assessed. Differences between women with PCOS and control women and changes with exercise were analysed using Student's unpaired t tests. Differences in the contribution of NO to cutaneous blood flow [expressed as a percentage of maximal cutaneous vasodilatation (CVCmax)] were analysed using general linear models. At 42°C heating, cutaneous NO-mediated vasodilatation was attenuated by 17.5%CVCmax (95% confidence interval, 33.3, 1.7; P = 0.03) in women with PCOS vs. control women. Exercise training improved cardiorespiratory fitness by 5.0 ml kg−1 min−1 (95% confidence interval, 0.9, 9.2; P = 0.03) and NO-mediated cutaneous vasodilatation at 42°C heating by 19.6% CVCmax (95% confidence interval, 4.3, 34.9; P = 0.02). Cutaneous microvascular NO function is impaired in women with PCOS compared with obese matched control women but can be improved with exercise training. PMID:23318877

Cutaneous microvascular perfusion responses to insulin iontophoresis are differentially affected by insulin resistance after spinal cord injury.

PubMed

La Fountaine, Michael F; Cirnigliaro, Christopher M; Azarelo, Frank; Hobson, Joshua C; Tascione, Oriana; Swonger, Kirsten N; Dyson-Hudson, Trevor; Bauman, William A

2017-09-01

What is the central question of this study? What impact does insulin resistance have on cutaneous perfusion responses to insulin iontophoresis in vascular beds with markedly reduced or functionally ablated sympathetic nervous system vasomotor function resulting from spinal cord injury? What is the main finding and its importance? Persons with spinal cord injury have sublesional microvascular endothelial dysfunction, as indicated by a blunted cutaneous perfusion response to acetylcholine iontophoresis, and the presence of insulin resistance has a further confounding effect on endothelium-mediated changes to cutaneous perfusion in the lower extremities. Endothelium-mediated mechanisms that regulate skin blood flow might play an integral role in optimizing skin perfusion in vascular beds with sympathetic nervous system vasomotor impairment, such as in spinal cord injury (SCI). Insulin is a vasoactive hormone and second messenger of nitric oxide that facilitates endothelium-mediated dilatation. The effects of insulin resistance (IR) on sublesional cutaneous perfusion responses to insulin provocation have yet to be described in persons with SCI. Persons with SCI and an able-bodied (AB) cohort were divided into subgroups based upon fasting plasma insulin concentration cut-offs for IR (≥13.13 mIU ml -1 ) or insulin sensitivity (IS; <13.13 mIU ml -1 ), as follows: AB, IS (ABIS, n = 21); SCI, IS (SCIS, n = 21); AB, IR (ABIR, n = 9); and SCI, IR (SCIR, n = 11). Laser Doppler flowmetry characterized peak blood perfusion unit (BPU) responses (percentage change from baseline) to insulin, acetylcholine or placebo iontophoresis in the lower extremities; BPU responses were log 10 transformed to facilitate comparisons, and the net insulin response (NetIns) BPU response was calculated (insulin minus placebo BPU response). The NetIns was significantly greater in both IS groups compared with their corresponding IR group. The acetylcholine-mediated BPU responses in the SCI subgroups were significantly lower than those in the ABIS group. The proportional BPU responses of NetIns to acetylcholine in the IS cohorts (i.e. ABIS and SCIS) were significantly greater (P < 0.05) than that of each IR subgroup. The presence of IR has a confounding effect on sublesional microvascular endothelium-mediated cutaneous perfusion responses to provocation. Preservation of endothelial sensitivity to its agonists appears to be an important modifiable risk factor to optimize cutaneous perfusion in the lower extremities of persons with SCI. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.

Vaspin protects against LPS‑induced ARDS by inhibiting inflammation, apoptosis and reactive oxygen species generation in pulmonary endothelial cells via the Akt/GSK‑3β pathway.

PubMed

Qi, Di; Wang, Daoxin; Zhang, Chunrong; Tang, Xumao; He, Jing; Zhao, Yan; Deng, Wang; Deng, Xinyu

2017-12-01

Acute respiratory distress syndrome (ARDS) is characterized by uncontrolled extravasation of protein‑rich fluids, which is caused by disruption and dysfunction of the barrier of pulmonary endothelial cells (ECs). Visceral adipose tissue‑derived serine protease inhibitor (vaspin) is a novel adipokine with pleiotropic properties, which has been reported to exert beneficial effects against obesity‑associated systemic vascular diseases; however, its effects on ARDS remain unknown. In the present study, mice were subjected to systemic administration of adenoviral vector expressing vaspin (Ad‑vaspin) to examine its effects on lipopolysaccharide (LPS)‑induced ARDS in vivo. Histological analysis was then conducted, and cytokine [tumor necrosis factor (TNF)‑α, interleukin (IL)‑6 and IL‑10] levels, and intercellular cell adhesion molecule‑1 (ICAM‑1) and adherens junctions (AJs) expression were detected. In addition, human pulmonary microvascular ECs (HPMECs) were treated with recombinant human (rh)‑vaspin to further investigate its molecular basis and underlying mechanism. The mRNA expression levels of inflammatory cytokines (TNF‑α and IL‑6) and endothelial‑specific adhesion markers [vascular cell adhesion molecule‑1 and E‑selectin], activation of nuclear factor‑κB, and cell viability and apoptosis were then examined. Furthermore, the expression of AJs and organization of the cytoskeleton, as well as expression and activity of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and generation of reactive oxygen species (ROS) were determined. The results indicated that Ad‑vaspin protected against LPS‑induced ARDS by alleviating the pulmonary inflammatory response and pulmonary EC barrier dysfunction in mice, which was accompanied by activation of the protein kinase B (Akt)/glycogen synthase kinase (GSK)‑3β pathway. In addition, pretreatment of HPMECs with rh‑vaspin attenuated inflammation, apoptosis and ROS generation without alterations in AJs and cytoskeletal organization following LPS insult, which was accompanied by activation of the Akt/GSK3β pathway. In conclusion, the present study demonstrated that vaspin protects against LPS‑induced ARDS by reversing EC barrier dysfunction via the suppression of inflammation, apoptosis and ROS production in pulmonary ECs, at least partially via activation of the Akt/GSK3β pathway. These findings provide evidence of a causal link between vaspin and EC dysfunction in ARDS, and suggest a potential therapeutic intervention for patients with ARDS.

Targeting the Endoplasmic Reticulum Unfolded Protein Response to Counteract the Oxidative Stress-Induced Endothelial Dysfunction

PubMed Central

Moltedo, Ornella; Faraonio, Raffaella

2018-01-01

In endothelial cells, the tight control of the redox environment is essential for the maintenance of vascular homeostasis. The imbalance between ROS production and antioxidant response can induce endothelial dysfunction, the initial event of many cardiovascular diseases. Recent studies have revealed that the endoplasmic reticulum could be a new player in the promotion of the pro- or antioxidative pathways and that in such a modulation, the unfolded protein response (UPR) pathways play an essential role. The UPR consists of a set of conserved signalling pathways evolved to restore the proteostasis during protein misfolding within the endoplasmic reticulum. Although the first outcome of the UPR pathways is the promotion of an adaptive response, the persistent activation of UPR leads to increased oxidative stress and cell death. This molecular switch has been correlated to the onset or to the exacerbation of the endothelial dysfunction in cardiovascular diseases. In this review, we highlight the multiple chances of the UPR to induce or ameliorate oxidative disturbances and propose the UPR pathways as a new therapeutic target for the clinical management of endothelial dysfunction. PMID:29725497

Myeloperoxidase amplified high glucose-induced endothelial dysfunction in vasculature: Role of NADPH oxidase and hypochlorous acid.

PubMed

Tian, Rong; Ding, Yun; Peng, Yi-Yuan; Lu, Naihao

2017-03-11

Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-derived reactive oxygen species (ROS) such as superoxide and hydrogen peroxide (H 2 O 2 ), have emerged as important molecules in the pathogenesis of diabetic endothelial dysfunction. Additionally, neutrophils-derived myeloperoxidase (MPO) and MPO-catalyzed hypochlorous acid (HOCl) play important roles in the vascular injury. However, it is unknown whether MPO can use vascular-derived ROS to induce diabetic endothelial dysfunction. In the present study, we demonstrated that NADPH oxidase was the main source of ROS formation in high glucose-cultured human umbilical vein endothelial cells (HUVECs), and played a critical role in high glucose-induced endothelial dysfunction such as cell apoptosis, loss of cell viability and reduction of nitric oxide (NO). However, the addition of MPO could amplify the high glucose-induced endothelial dysfunction which was inhibited by the presence of apocynin (NADPH oxidase inhibitor), catalase (H 2 O 2 scavenger), or methionine (HOCl scavenger), demonstrating the contribution of NADPH oxidase-H 2 O 2 -MPO-HOCl pathway in the MPO/high glucose-induced vascular injury. In high glucose-incubated rat aortas, MPO also exacerbated the NADPH oxidase-induced impairment of endothelium-dependent relaxation. Consistent with these in vitro data, in diabetic rat aortas, both MPO expresion and NADPH oxidase activity were increased while the endothelial function was simultaneously impaired. The results suggested that vascular-bound MPO could amplify high glucose-induced vascular injury in diabetes. MPO-NADPH oxidase-HOCl may represent an important pathogenic pathway in diabetic vascular diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

Endothelial dysfunction in patients with obstructive sleep apnoea independent of metabolic syndrome.

PubMed

Amra, Babak; Karbasi, Elaheh; Hashemi, Mohammad; Hoffmann-Castendiek, Birgit; Golshan, Mohammad

2009-05-01

Obstructive sleep apnoea syndrome (OSAS), characterised by intermittent hypoxia/re-oxygenation, has been identified as an independent risk factor for cardiovascular diseases and endothelial dysfunction. Our aim was to investigate flow-mediated dilatation (FMD) in patients with obstructive sleep apnoea with and without metabolic syndrome. Fifty-two subjects with OSAS diagnosed by polysomnography were classified into 2 groups according to the presence and absence of the metabolic syndrome and also according to the severity: mild to moderate OSAS group and severe OSAS group. Endothelial function of the brachial artery was evaluated by using high-resolution vascular ultrasound. Endothelial-dependent dilatation (EDD) was assessed by establishing reactive hyperaemia and endothelial-independent dilatation (EID) was determined by using sublingual isosorbide dinitrate. Spearman correlation and regression analysis were performed. EDD was not significantly different in patients with OSAS and metabolic syndrome as compared with OSAS without metabolic syndrome (4.62 +/- 0.69 versus 4.49 +/- 0.93, P >0.05). Endothelial dysfunction in OSA may be independent of metabolic syndrome.

Effects of Flavonoid-Containing Beverages and EGCG on Endothelial Function

PubMed Central

Shenouda, Sherene M.; Vita, Joseph A.

2009-01-01

Abnormalities of the vascular endothelium contribute to all stages of atherosclerosis from lesion development to clinical cardiovascular disease events. Recognized risk factors, including diabetes mellitus, hypertension, dyslipidemia, cigarette smoking, and sedentary lifestyle are associated with endothelial dysfunction. A variety of pharmacological and behavioral interventions have been shown to reverse endothelial dysfunction in patients with cardiovascular disease. A large number of epidemiological studies suggest that dietary factors, including increased intake of flavonoid-containing foods and beverages, reduce cardiovascular risk, and recent studies have shown that such beverages have favorable effects on endothelial function. These studies have engendered interest in the development of dietary supplements or drugs that would allow for more convenient and higher dose administration of flavonoids and might prove useful for prevention or treatment of cardiovascular disease. In this paper, we will review the contribution of endothelial dysfunction to the pathogenesis and clinical expression of atherosclerosis and recent data linking flavonoid and EGCG consumption to improved endothelial function and reduced cardiovascular risk. PMID:17906190

Trafficking of adeno-associated virus vectors across a model of the blood-brain barrier; a comparative study of transcytosis and transduction using primary human brain endothelial cells.

PubMed

Merkel, Steven F; Andrews, Allison M; Lutton, Evan M; Mu, Dakai; Hudry, Eloise; Hyman, Bradley T; Maguire, Casey A; Ramirez, Servio H

2017-01-01

Developing therapies for central nervous system (CNS) diseases is exceedingly difficult because of the blood-brain barrier (BBB). Notably, emerging technologies may provide promising new options for the treatment of CNS disorders. Adeno-associated virus serotype 9 (AAV9) has been shown to transduce cells in the CNS following intravascular administration in rodents, cats, pigs, and non-human primates. These results suggest that AAV9 is capable of crossing the BBB. However, mechanisms that govern AAV9 transendothelial trafficking at the BBB remain unknown. Furthermore, possibilities that AAV9 may transduce brain endothelial cells or affect BBB integrity still require investigation. Using primary human brain microvascular endothelial cells as a model of the human BBB, we performed transduction and transendothelial trafficking assays comparing AAV9 to AAV2, a serotype that does not cross the BBB or transduce endothelial cells effectively in vivo. Results of our in vitro studies indicate that AAV9 penetrates brain microvascular endothelial cells barriers more effectively than AAV2, but has reduced transduction efficiency. In addition, our data suggest that (i) AAV9 penetrates endothelial barriers through an active, cell-mediated process, and (ii) AAV9 fails to disrupt indicators of BBB integrity such as transendothelial electrical resistance, tight junction protein expression/localization, and inflammatory activation status. Overall, this report shows how human brain endothelial cells configured in BBB models can be utilized for evaluating transendothelial movement and transduction kinetics of various AAV capsids. Importantly, the use of a human in vitro BBB model can provide import insight into the possible effects that candidate AVV gene therapy vectors may have on the status of BBB integrity. Read the Editorial Highlight for this article on page 192. © 2016 International Society for Neurochemistry.

Adipokine CTRP6 improves PPARγ activation to alleviate angiotensin II-induced hypertension and vascular endothelial dysfunction in spontaneously hypertensive rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chi, Liyi; Departments of Cardiology, The 451st Hospital of People's Liberation Army; Hu, Xiaojing

Angiotensin II (AngII) is the most important component of angiotensin, which has been regarded as a major contributor to the incidence of hypertension and vascular endothelial dysfunction. The adipocytokine C1q/TNF-related protein 6 (CTRP6) was recently reported to have multiple protective effects on cardiac and cardiovascular function. However, the exact role of CTRP6 in the progression of AngII induced hypertension and vascular endothelial function remains unclear. Here, we showed that serum CTRP6 content was significantly downregulated in SHRs, accompanied by a marked increase in arterial systolic pressure and serum AngII, CRP and ET-1 content. Then, pcDNA3.1-mediated CTRP6 delivery or CTRP6 siRNAmore » was injected into SHRs. CTRP6 overexpression caused a significant decrease in AngII expression and AngII-mediated hypertension and vascular endothelial inflammation. In contrast, CTRP6 knockdown had the opposite effect to CTRP6 overexpression. Moreover, we found that CTRP6 positively regulated the activation of the ERK1/2 signaling pathway and the expression of peroxisome proliferator-activated receptor γ (PPARγ), a recently proven negative regulator of AngII, in the brain and vascular endothelium of SHRs. Finally, CTRP6 was overexpressed in endothelial cells, and caused a significant increase in PPARγ activation and suppression in AngII-mediated vascular endothelial dysfunction and apoptosis. The effect of that could be rescued by the ERK inhibitor PD98059. In contrast, silencing CTRP6 suppressed PPARγ activation and exacerbated AngII-mediated vascular endothelial dysfunction and apoptosis. In conclusion, CTRP6 improves PPARγ activation and alleviates AngII-induced hypertension and vascular endothelial dysfunction. - Highlights: • Serum CTRP6 was significantly decreased in spontaneously hypertensive rats (SHRs). • CTRP6 positively regulated the activation of the ERK1/2 signaling pathway. • CTRP6 negatively regulates PPARγ mediated Angiotensin II (AngII) expression. • CTRP6 alleviates AngII-induced hypertension and vascular endothelial dysfunction.« less

Molecular Signatures of Tissue-Specific Microvascular Endothelial Cell Heterogeneity in Organ Maintenance and Regeneration

PubMed Central

Nolan, Daniel J.; Ginsberg, Michael; Israely, Edo; Palikuqi, Brisa; Poulos, Michael G.; James, Daylon; Ding, Bi-Sen; Schachterle, William; Liu, Ying; Rosenwaks, Zev; Butler, Jason M.; Xiang, Jenny; Rafii, Arash; Shido, Koji; Rabbany, Sina Y.; Elemento, Olivier; Rafii, Shahin

2013-01-01

SUMMARY Microvascular endothelial cells (ECs) within different tissues are endowed with distinct but as yet unrecognized structural, phenotypic, and functional attributes. We devised EC purification, cultivation, profiling, and transplantation models that establish tissue-specific molecular libraries of ECs devoid of lymphatic ECs or parenchymal cells. These libraries identify attributes that confer ECs with their organotypic features. We show that clusters of transcription factors, angiocrine growth factors, adhesion molecules, and chemokines are expressed in unique combinations by ECs of each organ. Furthermore, ECs respond distinctly in tissue regeneration models, hepatectomy, and myeloablation. To test the data set, we developed a transplantation model that employs generic ECs differentiated from embryonic stem cells. Transplanted generic ECs engraft into regenerating tissues and acquire features of organotypic ECs. Collectively, we demonstrate the utility of informational databases of ECs toward uncovering the extravascular and intrinsic signals that define EC heterogeneity. These factors could be exploited therapeutically to engineer tissue-specific ECs for regeneration. PMID:23871589

Tick-borne encephalitis virus infects human brain microvascular endothelial cells without compromising blood-brain barrier integrity.

PubMed

Palus, Martin; Vancova, Marie; Sirmarova, Jana; Elsterova, Jana; Perner, Jan; Ruzek, Daniel

2017-07-01

Alteration of the blood-brain barrier (BBB) is a hallmark of tick-borne encephalitis (TBE), a life-threating human viral neuroinfection. However, the mechanism of BBB breakdown during TBE, as well as TBE virus (TBEV) entry into the brain is unclear. Here, primary human microvascular endothelial cells (HBMECs) were infected with TBEV to study interactions with the BBB. Although the number of infected cells was relatively low in culture (<5%), the infection was persistent with high TBEV yields (>10 6 pfu/ml). Infection did not induce any significant changes in the expression of key tight junction proteins or upregulate the expression of cell adhesion molecules, and did not alter the highly organized intercellular junctions between HBMECs. In an in vitro BBB model, the virus crossed the BBB via a transcellular pathway without compromising the integrity of the cell monolayer. The results indicate that HBMECs may support TBEV entry into the brain without altering BBB integrity. Copyright © 2017 Elsevier Inc. All rights reserved.

Differentiation and characterization of human pluripotent stem cell-derived brain microvascular endothelial cells.

PubMed

Stebbins, Matthew J; Wilson, Hannah K; Canfield, Scott G; Qian, Tongcheng; Palecek, Sean P; Shusta, Eric V

2016-05-15

The blood-brain barrier (BBB) is a critical component of the central nervous system (CNS) that regulates the flux of material between the blood and the brain. Because of its barrier properties, the BBB creates a bottleneck to CNS drug delivery. Human in vitro BBB models offer a potential tool to screen pharmaceutical libraries for CNS penetration as well as for BBB modulators in development and disease, yet primary and immortalized models respectively lack scalability and robust phenotypes. Recently, in vitro BBB models derived from human pluripotent stem cells (hPSCs) have helped overcome these challenges by providing a scalable and renewable source of human brain microvascular endothelial cells (BMECs). We have demonstrated that hPSC-derived BMECs exhibit robust structural and functional characteristics reminiscent of the in vivo BBB. Here, we provide a detailed description of the methods required to differentiate and functionally characterize hPSC-derived BMECs to facilitate their widespread use in downstream applications. Copyright © 2015 Elsevier Inc. All rights reserved.

sAPP modulates iron efflux from brain microvascular endothelial cells by stabilizing the ferrous iron exporter ferroportin

PubMed Central

McCarthy, Ryan C; Park, Yun-Hee; Kosman, Daniel J

2014-01-01

A sequence within the E2 domain of soluble amyloid precursor protein (sAPP) stimulates iron efflux. This activity has been attributed to a ferroxidase activity suggested for this motif. We demonstrate that the stimulation of efflux supported by this peptide and by sAPPα is due to their stabilization of the ferrous iron exporter, ferroportin (Fpn), in the plasma membrane of human brain microvascular endothelial cells (hBMVEC). The peptide does not bind ferric iron explaining why it does not and thermodynamically cannot promote ferrous iron autoxidation. This peptide specifically pulls Fpn down from the plasma membrane of hBMVEC; based on these results, FTP, for ferroportin-targeting peptide, correctly identifies the function of this peptide. The data suggest that in stabilizing Fpn via the targeting due to the FTP sequence, sAPP will increase the flux of iron into the cerebral interstitium. This inference correlates with the observation of significant iron deposition in the amyloid plaques characteristic of Alzheimer’s disease. PMID:24867889

Escherichia coli K1 utilizes host macropinocytic pathways for invasion of brain microvascular endothelial cells.

PubMed

Loh, Lip Nam; McCarthy, Elizabeth M C; Narang, Priyanka; Khan, Naveed A; Ward, Theresa H

2017-11-01

Eukaryotic cells utilize multiple endocytic pathways for specific uptake of ligands or molecules, and these pathways are commonly hijacked by pathogens to enable host cell invasion. Escherichia coli K1, a pathogenic bacterium that causes neonatal meningitis, invades the endothelium of the blood-brain barrier, but the entry route remains unclear. Here, we demonstrate that the bacteria trigger an actin-mediated uptake route, stimulating fluid phase uptake, membrane ruffling and macropinocytosis. The route of uptake requires intact lipid rafts as shown by cholesterol depletion. Using a variety of perturbants we demonstrate that small Rho GTPases and their downstream effectors have a significant effect on bacterial invasion. Furthermore, clathrin-mediated endocytosis appears to play an indirect role in E. coli K1 uptake. The data suggest that the bacteria effect a complex interplay between the Rho GTPases to increase their chances of uptake by macropinocytosis into human brain microvascular endothelial cells. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Childhood obesity-related endothelial dysfunction: an update on pathophysiological mechanisms and diagnostic advancements.

PubMed

Bruyndonckx, Luc; Hoymans, Vicky Y; Lemmens, Katrien; Ramet, José; Vrints, Christiaan J

2016-06-01

Childhood obesity jeopardizes a healthy future for our society's children as it is associated with increased cardiovascular morbidity and mortality later on in life. Endothelial dysfunction, the first step in the development of atherosclerosis, is already present in obese children and may well represent a targetable risk factor. Technological advancements in recent years have facilitated noninvasive measurements of endothelial homeostasis in children. Thereby this topic ultimately starts to get the attention it deserves. In this paper, we aim to summarize the latest insights on endothelial dysfunction in childhood obesity. We discuss methodological advancements in peripheral endothelial function measurement and newly identified diagnostic markers of vascular homeostasis. Finally, future challenges and perspectives are set forth on how to efficiently tackle the catastrophic rise in cardiovascular morbidity and mortality that will be inflicted on obese children if they are not treated optimally.

Bioactivation of organic nitrates and the mechanism of nitrate tolerance.

PubMed

Klemenska, Emila; Beresewicz, Andrzej

2009-01-01

Organic nitrates, such as nitroglycerin, are commonly used in the therapy of cardiovascular disease. Long-term therapy with these drugs, however, results in the rapid development of nitrate tolerance, limiting their hemodynamic and anti-ischemic efficacy. In addition, nitrate tolerance is associated with the expression of potentially deleterious modifications such as increased oxidative stress, endothelial dysfunction, and sympathetic activation. In this review we discuss current concepts regarding the mechanisms of organic nitrate bioactivation, nitrate tolerance, and nitrate-mediated oxidative stress and endothelial dysfunction. We also examine how hydralazine may prevent nitrate tolerance and related endothelial dysfunction.

Sinusoidal Endothelial Dysfunction Precedes Inflammation and Fibrosis in a Model of NAFLD

PubMed Central

Pasarín, Marcos; La Mura, Vincenzo; Gracia-Sancho, Jorge; García-Calderó, Héctor; Rodríguez-Vilarrupla, Aina; García-Pagán, Juan Carlos; Bosch, Jaime; Abraldes, Juan G.

2012-01-01

Non-alcoholic fatty liver disease (NAFLD) is the hepatic manifestation of the metabolic syndrome. Most morbidity associated with the metabolic syndrome is related to vascular complications, in which endothelial dysfunction is a major pathogenic factor. However, whether NAFLD is associated with endothelial dysfunction within the hepatic vasculature is unknown. The aims of this study were to explore, in a model of diet-induced overweight that expresses most features of the metabolic syndrome, whether early NAFLD is associated with liver endothelial dysfunction. Wistar Kyoto rats were fed a cafeteria diet (CafD; 65% of fat, mostly saturated) or a control diet (CD) for 1 month. CafD rats developed features of the metabolic syndrome (overweight, arterial hypertension, hypertryglyceridemia, hyperglucemia and insulin resistance) and liver steatosis without inflammation or fibrosis. CafD rats had a significantly higher in vivo hepatic vascular resistance than CD. In liver perfusion livers from CafD rats had an increased portal perfusion pressure and decreased endothelium-dependent vasodilation. This was associated with a decreased Akt-dependent eNOS phosphorylation and NOS activity. In summary, we demonstrate in a rat model of the metabolic syndrome that shows features of NAFLD, that liver endothelial dysfunction occurs before the development of fibrosis or inflammation. PMID:22509248

Ameliorating Endothelial Mitochondrial Dysfunction Restores Coronary Function via Transient Receptor Potential Vanilloid 1-Mediated Protein Kinase A/Uncoupling Protein 2 Pathway.

PubMed

Xiong, Shiqiang; Wang, Peijian; Ma, Liqun; Gao, Peng; Gong, Liuping; Li, Li; Li, Qiang; Sun, Fang; Zhou, Xunmei; He, Hongbo; Chen, Jing; Yan, Zhencheng; Liu, Daoyan; Zhu, Zhiming

2016-02-01

Coronary heart disease arising from atherosclerosis is a leading cause of cardiogenic death worldwide. Mitochondria are the principal source of reactive oxygen species (ROS), and defective oxidative phosphorylation by the mitochondrial respiratory chain contributes to ROS generation. Uncoupling protein 2 (UCP2), an adaptive antioxidant defense factor, protects against mitochondrial ROS-induced endothelial dysfunction in atherosclerosis. The activation of transient receptor potential vanilloid 1 (TRPV1) attenuates vascular dysfunction. Therefore, whether TRPV1 activation antagonizes coronary lesions by alleviating endothelial mitochondrial dysfunction and enhancing the activity of the protein kinase A/UCP2 pathway warrants examination. ApoE(-/-), ApoE(-/-)/TRPV1(-/-), and ApoE(-/-)/UCP2(-/-) mice were fed standard chow, a high-fat diet (HFD), or the HFD plus 0.01% capsaicin. HFD intake profoundly impaired coronary vasodilatation and myocardial perfusion and shortened the survival duration of ApoE(-/-) mice. TRPV1 or UCP2 deficiency exacerbated HFD-induced coronary dysfunction and was associated with increased ROS generation and reduced nitric oxide production in the endothelium. The activation of TRPV1 by capsaicin upregulated UCP2 expression via protein kinase A phosphorylation, thereby alleviating endothelial mitochondrial dysfunction and inhibiting mitochondrial ROS generation. In vivo, dietary capsaicin supplementation enhanced coronary relaxation and prolonged the survival duration of HFD-fed ApoE(-/-) mice. These effects were not observed in ApoE(-/-) mice lacking the TRPV1 or UCP2 gene. The upregulation of protein kinase A /UCP2 via TRPV1 activation ameliorates coronary dysfunction and prolongs the lifespan of atherosclerotic mice by ameliorating endothelial mitochondrial dysfunction. Dietary capsaicin supplementation may represent a promising intervention for the primary prevention of coronary heart disease. © 2015 American Heart Association, Inc.

MicroRNA-27b plays a role in pulmonary arterial hypertension by modulating peroxisome proliferator-activated receptor γ dependent Hsp90-eNOS signaling and nitric oxide production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bi, Rui; Bao, Chunrong; Jiang, Lianyong

Pulmonary artery endothelial dysfunction is associated with pulmonary arterial hypertension (PAH). Based on recent studies showing that microRNA (miR)-27b is aberrantly expressed in PAH, we hypothesized that miR-27b may contribute to pulmonary endothelial dysfunction and vascular remodeling in PAH. The effect of miR-27b on pulmonary endothelial dysfunction and the underlying mechanism were investigated in human pulmonary artery endothelial cells (HPAECs) in vitro and in a monocrotaline (MCT)-induced model of PAH in vivo. miR-27b expression was upregulated in MCT-induced PAH and inversely correlated with the levels of peroxisome proliferator-activated receptor (PPAR)-γ, and miR-27b inhibition attenuated MCT-induced endothelial dysfunction and remodeling and prevented PAHmore » associated right ventricular hypertrophy and systolic pressure in rats. PPARγ was confirmed as a direct target of miR-27b in HPAECs and shown to mediate the effect of miR-27b on the disruption of endothelial nitric oxide synthase (eNOS) coupling to Hsp90 and the suppression of NO production associated with the PAH phenotype. We showed that miR-27b plays a role endothelial function and NO release and elucidated a potential mechanism by which miR-27b regulates Hsp90-eNOS and NO signaling by modulating PPARγ expression, providing potential therapeutic targets for the treatment of PAH. - Highlights: • miR-27b plays a role in endothelial function and NO release. • miR-27b inhibition ameliorates MCT-induced endothelial dysfunction and PAH. • miR-27b targets PPARγ in HPAECs. • miR-27b regulates PPARγ dependent Hsp90-eNOS and NO signaling.« less

Retinal microvascular damage and vasogenic edema produced by Clostridium perfringens type D epsilon toxin in rats.

PubMed

Finnie, John W; Manavis, Jim; Casson, Robert J; Chidlow, Glyn

2014-05-01

When the brain is exposed to large circulating levels of Clostridium perfringens type D epsilon toxin (EXT), microvascular damage with resulting severe, generalized, vasogenic edema seems to be principally responsible for the ensuing acute, and frequently fatal, neurologic disorder. However, although the blood-retinal barrier resembles in many respects the blood-brain barrier, retinal changes in livestock with acute epsilon intoxication have not, to the authors' knowledge, been previously reported. In rats given an acute dose of ETX, retinal microvascular endothelial injury led to widespread vasogenic edema as assessed immunohistochemically by marked plasma albumin extravasation. As laboratory rodents are a good model of the domestic livestock disease produced by ETX, it is probable that the latter sustain some visual deficit when exposed to large doses of this potent neurotoxin. © 2014 The Author(s).

Chronic Aerobic Exercise Associated to Dietary Modification Improve Endothelial Function and eNOS Expression in High Fat Fed Hamsters

PubMed Central

Boa, Beatriz C. S.; Souza, Maria das Graças C.; Leite, Richard D.; da Silva, Simone V.; Barja-Fidalgo, Thereza Christina; Kraemer-Aguiar, Luiz Guilherme; Bouskela, Eliete

2014-01-01

Obesity is epidemic in the western world and central adipose tissue deposition points to increased cardiovascular morbidity and mortality, independently of any association between obesity and other cardiovascular risk factors. Physical exercise has been used as non-pharmacological treatment to significantly reverse/attenuate obesity comorbidities. In this study we have investigated effects of exercise and/or dietary modification on microcirculatory function, body composition, serum glucose, iNOS and eNOS expression on 120 male hamsters treated for 12 weeks with high fat chow (HF, n = 30) starting on the 21st day of birth. From week 12 to 20, animals were randomly separated in HF (no treatment change), return to standard chow (HFSC, n = 30), high fat chow associated to an aerobic exercise training program (AET) (HFEX, n = 30) and return to standard chow+AET (HFSCEX, n = 30). Microvascular reactivity in response to acetylcholine and sodium nitroprusside and macromolecular permeability increase induced by 30 minutes ischemia followed by reperfusion were assessed on the cheek pouch preparation. Total body fat and aorta eNOS and iNOS expression by immunoblotting assay were evaluated on the experimental day. Compared to HFSC and HFSCEX groups, HF and HFEX ones presented increased visceral fat [(mean±SEM) (HF)4.9±1.5 g and (HFEX)4.7±0.9 g vs. (HFSC)*3.0±0.7 g and (HFSCEX)*1.9±0.4 g/100 g BW]; impaired endothelial-dependent vasodilatation [Ach 10−8 M (HF)87.9±2.7%; (HFSC)*116.7±5.9%; (HFEX)*109.1±4.6%; (HFSCEX)*105±2.8%; Ach10−6 M (HF)95.3±3.1%; (HFSC)*126±6.2%; (HFEX)*122.5±2.8%; (HFSCEX)*118.1±4.3% and Ach10−4 M (HF)109.5±4.8%; (HFSC)*149.6±6.6%; (HFEX)*143.5±5.4% and (HFSCEX)*139.4±5.2%], macromolecular permeability increase after ischemia/reperfusion [(HF)40.5±4.2; (HFSC)*19.0±1.6; (HFEX)*18.6±2.1 and (HFSCEX)* 21.5±3.7 leaks/cm2), decreased eNOS expression, increased leptin and glycaemic levels. Endothelial-independent microvascular reactivity was similar between groups, suggesting that only endothelial damage had occurred. Our results indicate that an aerobic routine and/or dietary modification may cause significant improvements to high fat fed animals, diminishing visceral depots, increasing eNOS expression and reducing microcirculatory dysfunction. PMID:25036223

Expression of psoriasis-associated fatty acid-binding protein in senescent human dermal microvascular endothelial cells.

PubMed

Ha, Moon Kyung; Chung, Kee Yang; Lee, Ju Hee; Bang, Dongsik; Park, Yoon Kee; Lee, Kwang Hoon

2004-09-01

Aging is associated with the progressive pathophysiologic modification of endothelial cells. In vitro endothelial cell senescence is accompanied by proliferative activity failure and by perturbations in gene and protein expressions. Moreover, this cellular senescence in culture has been proposed to reflect processes that occur in aging organisms. In order to observe the changing patterns of protein expression in senescent human dermal microvascular endothelial cells (HDMECs), proteins obtained from both early- and late-passaged HDMECs were separated by two-dimensional electrophoresis, visualized by silver staining, and quantified by image processing. Proteins of interest were extracted by in-gel digestion with trypsin and quantified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), by searching the National Center for Biotechnology Information protein-sequence database. More than 2000 spots were detected by 2D electrophoresis within a linear pH range of 3-10. Twenty-two major differentially expressed spots were observed in serially passaged HDMECs and identified with high confidence by MALDI-TOF-MS. One of these spots was found to be a 14-15 kDa psoriasis-associated fatty acid-binding protein (PA-FABP) with high affinity for long-chain fatty acids. The expression of PA-FABP was confirmed to be elevated in senescent HDMECs (passage 20) by fluorescence-activated cell sorting (FACS), confocal laser microscopy, and by immunohistochemistry in aged human skin tissue. Our results suggest that the overexpression of FABP in cultured senescent HDMECs is closely related to skin aging.

Contacting co-culture of human retinal microvascular endothelial cells alters barrier function of human embryonic stem cell derived retinal pigment epithelial cells.

PubMed

Skottman, H; Muranen, J; Lähdekorpi, H; Pajula, E; Mäkelä, K; Koivusalo, L; Koistinen, A; Uusitalo, H; Kaarniranta, K; Juuti-Uusitalo, K

2017-10-01

Here we evaluated the effects of human retinal microvascular endothelial cells (hREC) on mature human embryonic stem cell (hESC) derived retinal pigment epithelial (RPE) cells. The hESC-RPE cells (Regea08/017, Regea08/023 or Regea11/013) and hREC (ACBRI 181) were co-cultured on opposite sides of transparent membranes for up to six weeks. Thereafter barrier function, small molecule permeability, localization of RPE and endothelial cell marker proteins, cellular fine structure, and growth factor secretion of were evaluated. After co-culture, the RPE specific CRALBP and endothelial cell specific von Willebrand factor were appropriately localized. In addition, the general morphology, pigmentation, and fine structure of hESC-RPE cells were unaffected. Co-culture increased the barrier function of hESC-RPE cells, detected both with TEER measurements and cumulative permeability of FD4 - although the differences varied among the cell lines. Co-culturing significantly altered VEGF and PEDF secretion, but again the differences were cell line specific. The results of this study showed that co-culture with hREC affects hESC-RPE functionality. In addition, co-culture revealed drastic cell line specific differences, most notably in growth factor secretion. This model has the potential to be used as an in vitro outer blood-retinal barrier model for drug permeability testing. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Functional Polarity of Microvascular Brain Endothelial Cells Supported by Neurovascular Unit Computational Model of Large Neutral Amino Acid Homeostasis

PubMed Central

Taslimifar, Mehdi; Buoso, Stefano; Verrey, Francois; Kurtcuoglu, Vartan

2018-01-01

The homeostatic regulation of large neutral amino acid (LNAA) concentration in the brain interstitial fluid (ISF) is essential for proper brain function. LNAA passage into the brain is primarily mediated by the complex and dynamic interactions between various solute carrier (SLC) transporters expressed in the neurovascular unit (NVU), among which SLC7A5/LAT1 is considered to be the major contributor in microvascular brain endothelial cells (MBEC). The LAT1-mediated trans-endothelial transport of LNAAs, however, could not be characterized precisely by available in vitro and in vivo standard methods so far. To circumvent these limitations, we have incorporated published in vivo data of rat brain into a robust computational model of NVU-LNAA homeostasis, allowing us to evaluate hypotheses concerning LAT1-mediated trans-endothelial transport of LNAAs across the blood brain barrier (BBB). We show that accounting for functional polarity of MBECs with either asymmetric LAT1 distribution between membranes and/or intrinsic LAT1 asymmetry with low intraendothelial binding affinity is required to reproduce the experimentally measured brain ISF response to intraperitoneal (IP) L-tyrosine and L-phenylalanine injection. On the basis of these findings, we have also investigated the effect of IP administrated L-tyrosine and L-phenylalanine on the dynamics of LNAAs in MBECs, astrocytes and neurons. Finally, the computational model was shown to explain the trans-stimulation of LNAA uptake across the BBB observed upon ISF perfusion with a competitive LAT1 inhibitor. PMID:29593549

Connective Tissue Growth Factor (CTGF/CCN2) mediates angiogenic effect of S1P in human dermal microvascular endothelial cells

PubMed Central

MARKIEWICZ, MAGARET; NAKERAKANTI, SASHIDHAR S.; KAPANADZE, BAGRAT; GHATNEKAR, ANGELA; TROJANOWSKA, MARIA

2010-01-01

Objective The primary objective of this study was to examine the potential interaction between sphingosine-1-phosphate (S1P), a pleiotropic lipid mediator, and CTGF/CCN2 a secreted multimodular protein, in the process of endothelial cell migration. The second objective was to determine whether C- and N-terminal domains of CTGF/CCN2 have specific function in cell migration. Materials and Methods Migration of human dermal microvascular endothelial cells (HDMECs) was examined in monolayer wound healing “scratch” assay, while capillary-like tube formation was examined in 3 dimensional collagen co-culture assays. Results We observed that S1P stimulates HDMECs migration concomitant with upregulation of CTGF/CCN2 expression. Furthermore, the blockade of endogenous CTGF/CCN2 via siRNA abrogated S1P induced HDMECs migration and capillary-like tube formation. Full length CTGF induced cell migration and capillary-like tube formation with potency similar to that of S1P, while C-terminal domain of CTGF was slightly less effective. However; N-terminal domain had only a residual activity in inducing capillary-like tube formation. Conclusions This study revealed that CTGF/CCN2 is required for the S1P induced endothelial cell migration, which suggests that CTGF/CCN2 may be an important mediator of S1P induced physiological and pathological angiogenesis. Moreover, this study shows that the pro-migratory activity of CTGF/CCN2 is located in the C-terminal domain. PMID:21166920

Administration of tauroursodeoxycholic acid prevents endothelial dysfunction caused by an oral glucose load

PubMed Central

Walsh, Lauren K.; Restaino, Robert M.; Neuringer, Martha; Manrique, Camila; Padilla, Jaume

2017-01-01

Postprandial hyperglycemia leads to a transient impairment in endothelial function; however, the mechanisms remain largely unknown. Previous work in cell culture models demonstrate that high glucose results in endoplasmic reticulum (ER) stress and, in animal studies, ER stress has been implicated as a cause of endothelial dysfunction. Herein we tested the hypothesis that acute oral administration of tauroursodeoxycholic acid (TUDCA, 1500mg), a chemical chaperone known to alleviate ER stress, would prevent hyperglycemia-induced endothelial dysfunction. In 12 young healthy subjects (seven men, five women), brachial artery flow-mediated dilation (FMD) was assessed at baseline, 1 hour, and 2 hours post an oral glucose challenge. Subjects were tested on two separate visits in a single-blind randomized crossover design: after oral ingestion of TUDCA or placebo capsules. FMD was reduced from baseline during hyperglycemia under the placebo condition (−32% at 1 hr and −28% at 2 hr post oral glucose load; p<0.05 from baseline) but not under the TUDCA condition (−4% at 1 hr and +0.3% at 2 hr post oral glucose load; p>0.05 from baseline). Postprandial plasma glucose and insulin were not altered by TUDCA ingestion. Plasma oxidative stress markers 3-nitrotyrosine and TBARs remained unaltered throughout the oral glucose challenge in both conditions. These results suggest that hyperglycemia-induced endothelial dysfunction can be mitigated by oral administration of TUDCA, thus supporting the hypothesis that ER stress may contribute to endothelial dysfunction during postprandial hyperglycemia. PMID:27503949

Assessment of Endothelial Dysfunction: The Role of Symmetrical Dimethylarginine and Proinflammatory Markers in Chronic Kidney Disease and Renal Transplant Recipients

PubMed Central

Giga, Vojislav; Dopsaj, Violeta; Jelic-Ivanovic, Zorana

2013-01-01

Objectives. The study was designed to evaluate associations between symmetric dimethylarginine (SDMA), inflammation, and superoxide anion (O2∙−) with endothelial function and to determine their potential for screening of endothelial dysfunction in patients with chronic kidney disease (CKD) and renal transplant (RT) recipients. Materials and Methods. We included 64 CKD and 52 RT patients. Patients were stratified according to brachial artery flow-mediated dilation (FMD). Results. Logistic regression analysis showed that high SDMA and high sensitive C-reactive protein (hs-CRP) were associated with impaired FMD in CKD and RT patients, after adjustment for glomerular filtration rate. The ability of inflammation, SDMA, and O2∙− to detect impaired FMD was investigated by receiving operative characteristic analysis. Hs-CRP (area under the curves (AUC) = 0.754, P < 0.001), IL-6 (AUC = 0.699, P = 0.002), and SDMA (AUC = 0.689, P = 0.007) had the highest ability to detect impaired FMD. SDMA in combination with inflammatory parameters and/or O2∙− had better screening performance than SDMA alone. Conclusions. Our results indicate a strong predictable association between hs-CRP, SDMA, and endothelial dysfunction in CKD patients and RT recipients. The individual marker that showed the strongest discriminative ability for endothelial dysfunction is hs-CRP, but its usefulness as a discriminatory marker for efficient diagnosis of endothelial dysfunction should be examined in prospective studies. PMID:24167363

Age and diabetes related changes of the retinal capillaries: An ultrastructural and immunohistochemical study.

PubMed

Bianchi, Enrica; Ripandelli, Guido; Taurone, Samanta; Feher, Janos; Plateroti, Rocco; Kovacs, Illes; Magliulo, Giuseppe; Orlando, Maria Patrizia; Micera, Alessandra; Battaglione, Ezio; Artico, Marco

2016-03-01

Normal human aging and diabetes are associated with a gradual decrease of cerebral flow in the brain with changes in vascular architecture. Thickening of the capillary basement membrane and microvascular fibrosis are evident in the central nervous system of elderly and diabetic patients. Current findings assign a primary role to endothelial dysfunction as a cause of basement membrane (BM) thickening, while retinal alterations are considered to be a secondary cause of either ischemia or exudation. The aim of this study was to reveal any initial retinal alterations and variations in the BM of retinal capillaries during diabetes and aging as compared to healthy controls. Moreover, we investigated the potential role of vascular endothelial growth factor (VEGF) and pro-inflammatory cytokines in diabetic retina.Transmission electron microscopy (TEM) was performed on 46 enucleated human eyes with particular attention to alterations of the retinal capillary wall and Müller glial cells. Inflammatory cytokines expression in the retina was investigated by immunohistochemistry.Our electron microscopy findings demonstrated that thickening of the BM begins primarily at the level of the glial side of the retina during aging and diabetes. The Müller cells showed numerous cytoplasmic endosomes and highly electron-dense lysosomes which surrounded the retinal capillaries. Our study is the first to present morphological evidence that Müller cells start to deposit excessive BM material in retinal capillaries during aging and diabetes. Our results confirm the induction of pro-inflammatory cytokines TNF-α and IL-1β within the retina as a result of diabetes.These observations strongly suggest that inflammatory cytokines and changes in the metabolism of Müller glial cells rather than changes in of endothelial cells may play a primary role in the alteration of retinal capillaries BM during aging and diabetes. © The Author(s) 2015.

Age and diabetes related changes of the retinal capillaries: An ultrastructural and immunohistochemical study

PubMed Central

Bianchi, Enrica; Ripandelli, Guido; Taurone, Samanta; Feher, Janos; Plateroti, Rocco; Kovacs, Illes; Magliulo, Giuseppe; Orlando, Maria Patrizia; Micera, Alessandra; Battaglione, Ezio; Artico, Marco

2015-01-01

Normal human aging and diabetes are associated with a gradual decrease of cerebral flow in the brain with changes in vascular architecture. Thickening of the capillary basement membrane and microvascular fibrosis are evident in the central nervous system of elderly and diabetic patients. Current findings assign a primary role to endothelial dysfunction as a cause of basement membrane (BM) thickening, while retinal alterations are considered to be a secondary cause of either ischemia or exudation. The aim of this study was to reveal any initial retinal alterations and variations in the BM of retinal capillaries during diabetes and aging as compared to healthy controls. Moreover, we investigated the potential role of vascular endothelial growth factor (VEGF) and pro-inflammatory cytokines in diabetic retina. Transmission electron microscopy (TEM) was performed on 46 enucleated human eyes with particular attention to alterations of the retinal capillary wall and Müller glial cells. Inflammatory cytokines expression in the retina was investigated by immunohistochemistry. Our electron microscopy findings demonstrated that thickening of the BM begins primarily at the level of the glial side of the retina during aging and diabetes. The Müller cells showed numerous cytoplasmic endosomes and highly electron-dense lysosomes which surrounded the retinal capillaries. Our study is the first to present morphological evidence that Müller cells start to deposit excessive BM material in retinal capillaries during aging and diabetes. Our results confirm the induction of pro-inflammatory cytokines TNF-α and IL-1β within the retina as a result of diabetes. These observations strongly suggest that inflammatory cytokines and changes in the metabolism of Müller glial cells rather than changes in of endothelial cells may play a primary role in the alteration of retinal capillaries BM during aging and diabetes. PMID:26604209

Impaired ATP release from red blood cells promotes their adhesion to endothelial cells: A mechanism of hypoxemia after transfusion

PubMed Central

Zhu, Hongmei; Zennadi, Rahima; Xu, Bruce X.; Eu, Jerry P.; Torok, Jordan A.; Telen, Marilyn J.; McMahon, Timothy J.

2011-01-01

Objective Transfusion of red blood cells (RBCs) has been linked to disappointing clinical outcomes in the critically ill, but specific mechanisms of organ dysfunction after transfusion remain poorly understood. We tested the hypothesis that RBC storage impairs the ability of RBCs to release ATP and that impaired ATP-release was injurious in vivo, in part through increased RBC adhesion. Design Prospective, controlled, mechanistic study. Setting University research laboratory. Subjects Human and mouse blood donors; nude mouse transfusion recipients. Interventions Manipulation of ATP release, supplemental ATP, and antibodies to RBC and endothelial adhesion receptors were used in vitro and in vivo to probe the roles of released ATP and adhesion in responses to (transfused) RBCs. Measurements and main results The ability of stored RBCs to release ATP declined markedly within 14 days after collection, despite relatively stable levels of ATP within the RBCs. Inhibiting ATP release promoted the adhesion of stored RBCs to endothelial cells in vitro and RBC sequestration in the lungs of transfused mice in vivo. Unlike transfusion of fresh human RBCs, stored-RBC transfusion in mice decreased blood oxygenation and increased extravasation of RBCs into the lung’s alveolar airspaces. Similar findings were seen with transfusion of fresh RBCs treated with the ATP-release inhibitors glibenclamide and carbenoxolone. These findings were prevented by either co-infusion of an ATP analog or pre-transfusion incubation of the RBCs with an antibody against the erythrocyte adhesion receptor LW (Landsteiner-Wiener; ICAM-4). Conclusions The normal flow of RBCs in pulmonary microvessels depends in part on the release of anti-adhesive ATP from RBCs, and storage-induced deficiency in ATP release from transfused RBCs may promote or exacerbate microvascular pathophysiology in the lung, in part through increased RBC adhesion. PMID:21765360

The relationships among hyperuricemia, body mass index and impaired renal function in type 2 diabetic patients.

PubMed

Li, Yongmei; Fan, Xing; Li, Chunjun; Zhi, Xinyue; Peng, Liyuan; Han, Hongling; Sun, Bei

2018-03-28

Chronic kidney disease (CKD) is a common chronic microvascular complication and the major cause of death in diabetic patients. This study was conceived to explore the possible mechanisms of how hyperuricemia and obesity contribute to renal function impairment in type 2 diabetic (T2DM) patients. A cross-sectional study in 609 participants recruited from a T2DM population in North China was conducted. The multiplicative interaction between body mass index (BMI) and uric acid (UA) level was assessed using an interaction term in a logistic regression analysis. Our results indicate that male T2DM patients having higher BMI (OR 1.711, p = 0.038), blood urine nitrogen (BUN) (OR 1.100, p = 0.034), and 24-hour urinary micro-albumin levels (OR 1.004, p = 0.021) were much more likely to have high UA. Whereas, for female T2DM patients, the OR of BMI, BUN, and triglyceride were 1.169 (p = 0.001), 1.337 (p = 0.000), and 1.359 (p = 0.006), respectively. In this study population, obesity and elevated UA work together to increase the risk of renal injury. In vitro experiments indicate that reactive oxygen species (ROS) production increased with UA treatment in human renal glomerular endothelial cells (HRGECs), while endothelial nitric oxide synthase (eNOS) production level dropped. UA also increased monocyte chemotactic protein-1 (MCP-1) expression and nuclear factor kappa B (NF-κB) activation. Taken together, our results indicate that high concentrations of UA lead to endothelial dysfunction through the activation of the inflammatory response and induction of oxidative stress, even in non-obese T2DM patients.

Administration of nicotinamide riboside prevents oxidative stress and organ injury in sepsis.

PubMed

Hong, Guangliang; Zheng, Dong; Zhang, Lulu; Ni, Rui; Wang, Grace; Fan, Guo-Chang; Lu, Zhongqiu; Peng, Tianqing

2018-08-01

Sepsis-caused multiple organ failure remains the major cause of morbidity and mortality in intensive care units. Nicotinamide riboside (NR) is a precursor of nicotinamide adenine dinucleotide (NAD + ), which is important in regulating oxidative stress. This study investigated whether administration of NR prevented oxidative stress and organ injury in sepsis. Mouse sepsis models were induced by injection of lipopolysaccharides (LPS) or feces-injection-in-peritoneum. NR was given before sepsis onset. Cultured macrophages and endothelial cells were incubated with various agents. Administration of NR elevated the NAD + levels, and elicited a reduction of oxidative stress, inflammation and caspase-3 activity in lung and heart tissues, which correlated with attenuation of pulmonary microvascular permeability and myocardial dysfunction, leading to less mortality in sepsis models. These protective effects of NR were associated with decreased levels of plasma high mobility group box-1 (HMGB1) in septic mice. Consistently, pre-treatment of macrophages with NR increased NAD + content and reduced HMGB1 release upon LPS stimulation. NR also prevented reactive oxygen species (ROS) production and apoptosis in endothelial cells induced by a conditioned-medium collected from LPS-treated macrophages. Furthermore, inhibition of SIRT1 by EX527 offset the negative effects of NR on HMGB1 release in macrophages, and ROS and apoptosis in endothelial cells. Administration of NR prevents lung and heart injury, and improves the survival in sepsis, likely by inhibiting HMGB1 release and oxidative stress via the NAD + /SIRT1 signaling. Given NR has been used as a health supplement, it may be a useful agent to prevent organ injury in sepsis. Copyright © 2018 Elsevier Inc. All rights reserved.

Diabetes-induced microvascular complications at the level of the spinal cord; a contributing factor in diabetic neuropathic pain.

PubMed

Ved, N; Da Vitoria Lobo, M E; Bestall, S M; L Vidueira, C; Beazley-Long, N; Ballmer-Hofer, K; Hirashima, M; Bates, D O; Donaldson, L F; Hulse, R P

2018-05-17

Abnormalities of neurovascular interactions within the central nervous system of diabetic patients is associated with the onset of many neurological disease states. However, to date, the link between the neurovascular network within the spinal cord and regulation of nociception has not been investigated despite neuropathic pain being common in diabetes. We hypothesised that hyperglycaemia-induced endothelial degeneration in the spinal cord, due to suppression of VEGF-A/VEGFR2 signalling, induces diabetic neuropathic pain. Nociceptive pain behaviour was investigated in a chemically induced model of type 1 diabetes (streptozotocin induced, insulin supplemented; either vehicle or VEGF-A 165 b treated) and an inducible endothelial knockdown of VEGFR2 (tamoxifen induced). Diabetic animals developed mechanical allodynia and heat hyperalgesia. This was associated with a reduction in the number of blood vessels and reduction in Evans blue extravasation in the lumbar spinal cord of diabetic animals versus age-matched controls. Endothelial markers occludin, CD31 and VE-cadherin were downregulated in the spinal cord of the diabetic group versus controls, as well as a concurrent reduction of VEGF-A 165 b expression. In diabetic animals, VEGF-A 165 b treatment (biweekly intraperitoneal, 20 ng g -1 ) restored normal Evans blue extravasation and prevented vascular degeneration, diabetes-induced central neuron activation and neuropathic pain. Inducible knockdown of VEGFR2 (tamoxifen treated Tie2CreER T2 -vegfr2 flfl mice) led to a reduction in blood vessel network volume in the lumbar spinal cord and development of heat hyperalgesia. These findings indicate that hyperglycaemia leads to a reduction in the VEGF-A/VEGFR2 signalling cascade resulting in endothelial dysfunction in the spinal cord, which could be an undiscovered contributing factor to diabetic neuropathic pain. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Metabotropic glutamate receptor 5 mediates phosphorylation of vascular endothelial cadherin and nuclear localization of β-catenin in response to homocysteine.

PubMed

Beard, Richard S; Reynolds, Jason J; Bearden, Shawn E

2012-01-01

Elevated plasma homocysteine (Hcy) is an independent risk factor for vascular disease and stroke in part by causing generalized endothelial dysfunction. A receptor that is sensitive to Hcy and its intracellular signaling systems has not been identified. β-catenin is a pleiotropic regulator of transcription and cell function. Using a brain microvascular endothelial cell line (bEnd.3), we tested the hypothesis that Hcy causes receptor-dependent nuclear translocation of β-catenin. Hcy increased phosphorylation of Y731 on vascular endothelial cadherin (VE-cadherin), a site involved in coupling β-catenin to VE-cadherin. This was blocked by inhibition of either metabotropic glutamate receptor 5 (mGluR5) or ionotropic glutamate receptor (NMDAr) and by shRNA knockdown of mGluR5. Expression of these receptors was confirmed by flow cytometry, immunohistochemistry, and western blotting. Directed pharmacology with specific agonists elucidated a signaling cascade where Hcy activates mGluR5 which activates NMDAr with subsequent PKC activation and uncoupling of the VE-cadherin/β-catenin complex. Moreover, Hcy caused a shift in localization of β-catenin from membrane-bound VE-cadherin to the cell nucleus, where it bound DNA, including a regulatory region of the gene for claudin-5, leading to reduced expression of claudin-5. Nuclear localization, DNA binding of β-catenin, and reduced claudin-5 expression were blocked by inhibition of mGluR5. Knockdown of mGluR5 expression with shRNA also rescued claudin-5 expression from the effects of Hcy treatment. These data uniquely identify mGluR5 as a master switch that drives β-catenin nuclear localization in vascular endothelium and regulates cell-cell coupling in response to elevated Hcy levels. These studies dissect a pharmacological opportunity for developing new therapeutic strategies in HHcy. Copyright © 2012 Elsevier Inc. All rights reserved.

Cell Permeability, Migration, and Reactive Oxygen Species Induced by Multi-Walled Carbon Nanotubes in Human Microvascular Endothelial Cells

PubMed Central

Pacurari, M; Qian, Y; Fu, W; Schwegler-Berry, D; Ding, M; Castranova, V; Guo, NL

2011-01-01

Multi-walled carbon nanotubes (MWCNT) have elicited great interest in biomedical applications due to their extraordinary physical, chemical, and optical properties. Intravenous administration of MWCNT-based medical imaging agents and drugs in animal models was utilized. However, the potential harmful health effects of MWCNT administration in humans have not yet been elucidated. Furthermore, to date, there are no apparent reports regarding the precise mechanisms of translocation of MWCNT into target tissues and organs from blood circulation. This study demonstrates that exposure to MWCNT leads to an increase in cell permeability in human microvascular endothelial cells (HMVEC). The results obtained from this study also showed that the MWCNT-induced rise in endothelial permeability is mediated by reactive oxygen species (ROS) production and actin filament remodeling. In addition, it was found that MWCNT promoted cell migration in HMVEC. Mechanistically, MWCNT exposure elevated the levels of monocyte chemoattractant protein-1 (MCP-1) and intercellular adhesion molecule 1 (ICAM-1) in HMVEC. Taken together, these results provide new insights into the bioreactivity of MWCNT, which may have implications in the biomedical application of MWCNT in vascular targeting, imaging, and drug delivery. The results generated from this study also elucidate the potential adverse effects of MWCNT exposure on humans at the cellular level. PMID:22129238

Particulate matter from indoor environments of classroom induced higher cytotoxicity and leakiness in human microvascular endothelial cells in comparison with those collected from corridor.

PubMed

Chua, M L; Setyawati, M I; Li, H; Fang, C H Y; Gurusamy, S; Teoh, F T L; Leong, D T; George, S

2017-05-01

We investigated the physicochemical properties (size, shape, elemental composition, and endotoxin) of size resolved particulate matter (PM) collected from the indoor and corridor environments of classrooms. A comparative hazard profiling of these PM was conducted using human microvascular endothelial cells (HMVEC). Oxidative stress-dependent cytotoxicity responses were assessed using quantitative reverse transcriptase polymerase chain reaction (RT-PCR) and high content screening (HCS), and disruption of monolayer cell integrity was assessed using fluorescence microscopy and transwell assay. Scanning electron microscopy (SEM) coupled with energy-dispersive X-ray spectroscopy (EDX) analysis showed differences in the morphology and elemental composition of PM of different sizes and origins. While the total mass of PM collected from indoor environment was lower in comparison with those collected from the corridor, the endotoxin content was substantially higher in indoor PM (e.g., ninefold higher endotoxin level in indoor PM 8.1-20 ). The ability to induce oxidative stress-mediated cytotoxicity and leakiness in cell monolayer were higher for indoor PM compared to those collected from the corridor. In conclusion, this comparative analysis suggested that indoor PM is relatively more hazardous to the endothelial system possibly because of higher endotoxin content. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

N-n-butyl haloperidol iodide ameliorates hypoxia/reoxygenation injury through modulating the LKB1/AMPK/ROS pathway in cardiac microvascular endothelial cells.

PubMed

Lu, Binger; Wang, Bin; Zhong, Shuping; Zhang, Yanmei; Gao, Fenfei; Chen, Yicun; Zheng, Fuchun; Shi, Ganggang

2016-06-07

Endothelial cells are highly sensitive to hypoxia and contribute to myocardial ischemia/reperfusion injury. We have reported that N-n-butyl haloperidol iodide (F2) can attenuate hypoxia/reoxygenation (H/R) injury in cardiac microvascular endothelial cells (CMECs). However, the molecular mechanisms remain unclear. Neonatal rat CMECs were isolated and subjected to H/R. Pretreatment of F2 leads to a reduction in H/R injury, as evidenced by increased cell viability, decreased lactate dehydrogenase (LDH) leakage and apoptosis, together with enhanced AMP-activated protein kinase (AMPK) and liver kinase B1 (LKB1) phosphorylation in H/R ECs. Blockade of AMPK with compound C reversed F2-induced inhibition of H/R injury, as evidenced by decreased cell viability, increased LDH release and apoptosis. Moreover, compound C also blocked the ability of F2 to reduce H/R-induced reactive oxygen species (ROS) generation. Supplementation with the ROS scavenger N-acetyl-L-cysteine (NAC) reduced ROS levels, increased cell survival rate, and decreased both LDH release and apoptosis after H/R. In conclusion, our data indicate that F2 may mitigate H/R injury by stimulating LKB1/AMPK signaling pathway and subsequent suppression of ROS production in CMECs.

Endothelium-derived contracting factors mediate the Ang II-induced endothelial dysfunction in the rat aorta: preventive effect of red wine polyphenols.

PubMed

Kane, Modou O; Etienne-Selloum, Nelly; Madeira, Soccoro V F; Sarr, Mamadou; Walter, Allison; Dal-Ros, Stéphanie; Schott, Christa; Chataigneau, Thierry; Schini-Kerth, Valérie B

2010-04-01

Angiotensin II (Ang II)-induced hypertension is associated with vascular oxidative stress and an endothelial dysfunction. This study examined the role of reactive oxygen species (ROS) and endothelium-derived contracting factors in Ang II-induced endothelial dysfunction and whether these effects are prevented by red wine polyphenols (RWPs), a rich source of natural antioxidants. Rats were infused with Ang II for 14 days. RWPs were administered in the drinking water 1 week before and during the Ang II infusion. Arterial pressure was measured in conscious rats. Vascular reactivity was assessed in organ chambers and cyclooxygenase-1 (COX-1) and COX-2 expression by Western blot and immunofluorescence analyses. Ang II-induced hypertension was associated with blunted endothelium-dependent relaxations and induction of endothelium-dependent contractions in the presence of nitro-L-arginine in response to acetylcholine (Ach). These effects were not affected by the combination of membrane permeant analogs of superoxide dismutase and catalase but were abolished by the thromboxane A(2) (TP) receptor antagonist GR32191B and the COX-2 inhibitor NS-398. The COX-1 inhibitor SC-560 also prevented contractile responses to Ach. Ang II increased the expression of COX-1 and COX-2 in the aortic wall. RWPs prevented Ang II-induced hypertension, endothelial dysfunction, and upregulation of COX-1 and COX-2. Thus, Ang II-induced endothelial dysfunction cannot be explained by an acute formation of ROS reducing the bioavailability of nitric oxide but rather by COX-dependent formation of contracting factors acting on TP receptors. RWPs are able to prevent the Ang II-induced endothelial dysfunction mostly due to their antioxidant properties.

Impaired activity of adherens junctions contributes to endothelial dilator dysfunction in ageing rat arteries.

PubMed

Chang, Fumin; Flavahan, Sheila; Flavahan, Nicholas A

2017-08-01

Ageing-induced endothelial dysfunction contributes to organ dysfunction and progression of cardiovascular disease. VE-cadherin clustering at adherens junctions promotes protective endothelial functions, including endothelium-dependent dilatation. Ageing increased internalization and degradation of VE-cadherin, resulting in impaired activity of adherens junctions. Inhibition of VE-cadherin clustering at adherens junctions (function-blocking antibody; FBA) reduced endothelial dilatation in young arteries but did not affect the already impaired dilatation in old arteries. After junctional disruption with the FBA, dilatation was similar in young and old arteries. Src tyrosine kinase activity and tyrosine phosphorylation of VE-cadherin were increased in old arteries. Src inhibition increased VE-cadherin at adherens junctions and increased endothelial dilatation in old, but not young, arteries. Src inhibition did not increase dilatation in old arteries treated with the VE-cadherin FBA. Ageing impairs the activity of adherens junctions, which contributes to endothelial dilator dysfunction. Restoring the activity of adherens junctions could be of therapeutic benefit in vascular ageing. Endothelial dilator dysfunction contributes to pathological vascular ageing. Experiments assessed whether altered activity of endothelial adherens junctions (AJs) might contribute to this dysfunction. Aortas and tail arteries were isolated from young (3-4 months) and old (22-24 months) F344 rats. VE-cadherin immunofluorescent staining at endothelial AJs and AJ width were reduced in old compared to young arteries. A 140 kDa VE-cadherin species was present on the cell surface and in TTX-insoluble fractions, consistent with junctional localization. Levels of the 140 kDa VE-cadherin were decreased, whereas levels of a TTX-soluble 115 kDa VE-cadherin species were increased in old compared to young arteries. Acetylcholine caused endothelium-dependent dilatation that was decreased in old compared to young arteries. Disruption of VE-cadherin clustering at AJs (function-blocking antibody, FBA) inhibited dilatation to acetylcholine in young, but not old, arteries. After the FBA, there was no longer any difference in dilatation between old and young arteries. Src activity and tyrosine phosphorylation of VE-cadherin were increased in old compared to young arteries. In old arteries, Src inhibition (saracatinib) increased: (i) 140 kDa VE-cadherin in the TTX-insoluble fraction, (ii) VE-cadherin intensity at AJs, (iii) AJ width, and (iv) acetylcholine dilatation. In old arteries treated with the FBA, saracatinib no longer increased acetylcholine dilatation. Saracatinib did not affect dilatation in young arteries. Therefore, ageing impairs AJ activity, which appears to reflect Src-induced phosphorylation, internalization and degradation of VE-cadherin. Moreover, impaired AJ activity can account for the endothelial dilator dysfunction in old arteries. Restoring endothelial AJ activity may be a novel therapeutic approach to vascular ageing. © 2017 The Authors. The Journal of Physiology © 2017 The Physiological Society.

Endothelial binding of beta toxin to small intestinal mucosal endothelial cells in early stages of experimentally induced Clostridium perfringens type C enteritis in pigs.

PubMed

Schumacher, V L; Martel, A; Pasmans, F; Van Immerseel, F; Posthaus, H

2013-07-01

Beta toxin (CPB) is known to be an essential virulence factor in the development of lesions of Clostridium perfringens type C enteritis in different animal species. Its target cells and exact mechanism of toxicity have not yet been clearly defined. Here, we evaluate the suitability of a neonatal piglet jejunal loop model to investigate early lesions of C. perfringens type C enteritis. Immunohistochemically, CPB was detected at microvascular endothelial cells in intestinal villi during early and advanced stages of lesions induced by C. perfringens type C. This was first associated with capillary dilatation and subsequently with widespread hemorrhage in affected intestinal segments. CPB was, however, not demonstrated on intestinal epithelial cells. This indicates a tropism of CPB toward endothelial cells and suggests that CPB-induced endothelial damage plays an important role in the early stages of C. perfringens type C enteritis in pigs.

Tissue Factor Inflammatory Response Regulated by Promoter Genotype and p38 MAPK in Neonatal vs. Adult Microvascular Endothelial Cells

PubMed Central

Buzby, Jeffrey S.; Williams, Shirley A.; Imfeld, Karen L.; Kunicki, Thomas J.; Nugent, Diane J.

2014-01-01

Objective and design Variable tissue factor (TF) expression by human microvascular endothelial cells (HMVEC) may be regulated by two promoter haplotypes, distinguished by an 18 base pair deletion (D) or insertion (I) at -1208. We sought to determine the relationship between these haplotypes and interleukin-1 (IL-1α)-induced TF expression in neonatal versus adult HMVEC. Results IL-1-stimulated TF mRNA, protein, and activity were significantly higher in neonatal compared to adult D/D donors. IL-1-stimulated HMVEC from neonatal D/D donors expressed 3-fold higher levels of TF mRNA, 2-fold higher TF protein, and 4-fold increased TF activity compared to HMVEC from adult D/D donors. These results indicate that homozygosity for the D haplotype is characterized by increased response to IL-1 in neonates but not adults. IL-1 induced increased phosphorylation of p38 mitogen-activated protein kinase (MAPK), which was significantly greater in neonatal compared to adult HMVEC. Moreover, inhibition of the p38 MAPK pathway reduced IL-1-stimulated TF mRNA expression in D/D neonatal but not adult HMVEC. Conclusions Up-regulation of D/D neonatal HMVEC TF expression by IL-1 is mediated through the p38 MAPK pathway. This heightened response of D/D neonatal HMVEC to inflammatory stimuli may contribute to increased microvascular coagulopathies in susceptible newborn infants. PMID:24385191

Measurement of the filtration coefficient (Kfc) in the lung of Gallus domesticus and the effects of increased microvascular permeability.

PubMed

Weidner, W Jeffrey; Waddell, David S; Furlow, J David

2006-08-01

The filtration coefficient (Kfc) is a sensitive measure of microvascular hydraulic conductivity and has been reported for the alveolar lungs of many mammalian species, but not for the parabronchial avian lung. This study reports the Kfc in the isolated lungs of normal chickens and in the lungs of chickens given the edemogenic agents oleic acid (OA) or dimethyl amiloride (DMA). The control Kfc =0.04+/-0.01 ml min(-1) kPa(-1) g(-1). This parameter increased significantly following the administration of both OA (0.12+/-0.02 ml min(-1) kPa(-1) g(-1)) and DMA (0.07+/-0.01 ml min kPa(-1) g(-1)). As endothelial cadherins are thought to play a role in the dynamic response to acute lung injury, we utilized Western blot analysis to assess lung cadherin content and Northern blot analysis to assess pulmonary vascular endothelial (VE) cadherin expression following drug administration. Lung cadherin content decreases markedly following DMA, but not OA administration. VE cadherin expression increases as a result of DMA treatment, but is unchanged following OA. Our results suggest that the permeability characteristics of the avian lung are more closely consistent with those of the mammalian rather than the reptilian lung, and, that cadherins may play a significant role in the response to acute increases in avian pulmonary microvascular permeability.

Administration of sonic hedgehog protein induces angiogenesis and has therapeutic effects after stroke in rats.

PubMed

Chen, Sheng-Cai; Huang, Ming; He, Quan-Wei; Zhang, Yan; Opoku, Elvis Nana; Yang, Hang; Jin, Hui-Juan; Xia, Yuan-Peng; Hu, Bo

2017-06-03

The Sonic hedgehog (Shh) signaling pathway is recapitulated in response to ischemic injury. Here, we investigated the clinical implications of Shh protein in the ischemic stroke and explored the underlying mechanism. Intracerebroventricular injection of Shh, Cyclopamine, or anti-vascular endothelial growth factor (VEGF) was performed immediately after permanent middle cerebral artery occlusion (pMCAO) surgery and lasted for 7days (d). Phosphate-buffered saline (PBS) was used as control. Neurological deficits and infarct volume were examined 7d after pMCAO. Microvascular density with fluorescein-iso-thiocyanate (FITC) assay and double staining with CD31 and Ki-67 was measured at 7d. To observe in vitro angiogenesis, rat brain microvascular endothelial cells (RBMECs) were incubated under oxygen glucose deprivation (OGD) for 6h (h) and treated with Shh/anti-VEGF. We found that (1) Shh improved neurological scores and reduced infarct volume, which was blocked by Cyclopamine, (2) Shh improved the microvascular density and promoted angiogenesis and neuron survival in the ischemic boundary zone, (3) Shh enhanced VEGF expression and VEGF antibody could reverse angiogenic and protective effect of Shh in vivo and in vitro. These data demonstrate that the administration of Shh protein could protect brain from ischemic injury, in part by promoting angiogenic repair. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

Down-regulation of vascular PPAR-γ contributes to endothelial dysfunction in high-fat diet-induced obese mice exposed to chronic intermittent hypoxia.

PubMed

Zhang, Yanan; Zhang, Chunlian; Li, Haiou; Hou, Jingdong

2017-10-14

Obstructive sleep apnea (OSA), characterized by chronic intermittent hypoxia (CIH), is associated with endothelial dysfunction. The prevalence of OSA is linked to an epidemic of obesity. CIH has recently been reported to cause endothelial dysfunction in diet-induced obese animals by exaggerating oxidative stress and inflammation, but the underlying mechanism remains unclear. PPAR-γ, a ligand-inducible transcription factor that exerts anti-oxidant and anti-inflammatory effects, is down-regulated in the peripheral tissues in diet-induce obesity. We tested the hypothesis that down-regulation of vascular PPAR-γ in diet-induced obesity enhances inflammation and oxidative stress in response to CIH, resulting in endothelial dysfunction. Male C57BL/6 mice were fed either a high-fat diet (HFD) or a low-fat diet (LFD) and simultaneously exposed to CIH or intermittent air for 6 weeks. An additional HFD group received a combination of CIH and PPAR-γ agonist pioglitazone for 6 weeks. Endothelial-dependent vasodilation was impaired only in HFD group exposed to CIH, compared with other groups, but was restored by concomitant pioglitazone treatment. Molecular studies revealed that vascular PPAR-γ expression and activity were reduced in HFD groups, compared with LFD groups, but were reversed by pioglitazone treatment. In addition, CIH elevated vascular expression of NADPH oxidase 4 and dihydroethidium fluorescence, and increased expression of proinflammatory cytokines TNF-α and IL-1β in both LFD and HFD groups, but these increases was significantly greater in HFD group, along with decreased vascular eNOS activity. Pioglitazone treatment of HFD group prevented CIH-induced changes in above molecular markers. The results suggest that HFD-induced obesity down-regulates vascular PPAR-γ, which results in exaggerated oxidative stress and inflammation in response to CIH, contributing to endothelial dysfunction. This finding may provide new insights into the mechanisms by which OSA induces endothelial dysfunction and other cardiovascular disease in patients with obesity. Copyright © 2017 Elsevier Inc. All rights reserved.

Copper Transporter ATP7A Protects Against Endothelial Dysfunction in Type 1 Diabetic Mice by Regulating Extracellular Superoxide Dismutase

PubMed Central

Sudhahar, Varadarajan; Urao, Norifumi; Oshikawa, Jin; McKinney, Ronald D.; Llanos, Roxana M.; Mercer, Julian F.B.; Ushio-Fukai, Masuko; Fukai, Tohru

2013-01-01

Oxidative stress and endothelial dysfunction contribute to vascular complication in diabetes. Extracellular superoxide dismutase (SOD3) is one of the key antioxidant enzymes that obtains copper via copper transporter ATP7A. SOD3 is secreted from vascular smooth muscles cells (VSMCs) and anchors at the endothelial surface. The role of SOD3 and ATP7A in endothelial dysfunction in type 1 diabetes mellitus (T1DM) is entirely unknown. Here we show that the specific activity of SOD3, but not SOD1, is decreased, which is associated with increased O2•− production in aortas of streptozotocin-induced and genetically induced Ins2Akita T1DM mice. Exogenous copper partially rescued SOD3 activity in isolated T1DM vessels. Functionally, acetylcholine-induced, endothelium-dependent relaxation is impaired in T1DM mesenteric arteries, which is rescued by SOD mimetic tempol or gene transfer of SOD3. Mechanistically, ATP7A expression in T1DM vessels is dramatically decreased whereas other copper transport proteins are not altered. T1DM-induced endothelial dysfunction and decrease of SOD3 activity are rescued in transgenic mice overexpressing ATP7A. Furthermore, SOD3-deficient T1DM mice or ATP7A mutant T1DM mice augment endothelial dysfunction and vascular O2•− production versus T1DM mice. These effects are in part due to hypoinsulinemia in T1DM mice, since insulin treatment, but not high glucose, increases ATP7A expression in VSMCs and restores SOD3 activity in the organoid culture of T1DM vessels. In summary, a decrease in ATP7A protein expression contributes to impaired SOD3 activity, resulting in O2•− overproduction and endothelial dysfunction in blood vessels of T1DM. Thus, restoring copper transporter function is an essential therapeutic approach for oxidant stress–dependent vascular and metabolic diseases. PMID:23884884

Microvascular dysfunction with increased vascular leakage response in mice systemically exposed to arsenic.

PubMed

Chen, Shih-Chieh; Huang, Shin-Yin; Lu, Chi-Yu; Hsu, Ya-Hung; Wang, Dean-Chuan

2014-09-01

The mechanisms underlying cardiovascular disease induced by arsenic exposure are not completely understood. The objectives of this study were to investigate whether arsenic-fed mice have an increased vascular leakage response to vasoactive agents and whether enhanced type-2 protein phosphatase (PP2A) activity is involved in mustard oil-induced leakage. ICR mice were fed water or sodium arsenite (20 mg/kg) for 4 or 8 weeks. The leakage response to vasoactive agents was quantified using the Evans blue (EB) technique or vascular labeling with carbon particles. Increased EB leakage and high density of carbon-labeled microvessels were detected in arsenic-fed mice treated with mustard oil. Histamine induced significantly higher vascular leakage in arsenic-fed mice than in water-fed mice. Pretreatment with the PP2A inhibitor okadaic acid or the neurokinin 1 receptor (NK1R) blocker RP67580 significantly reduced mustard oil-induced vascular leakage in arsenic-fed mice. The protein levels of PP2Ac and NK1R were similar in both groups. PP2A activity was significantly higher in the arsenic-fed mice compared with the control group. These findings indicate that microvessels generally respond to vasoactive agents, and that the increased PP2A activity is involved in mustard oil-induced vascular leakage in arsenic-fed mice. Arsenic may initiate endothelial dysfunction, resulting in vascular leakage in response to vasoactive agents.

Obstructive Sleep Apnea is Linked to Depression and Cognitive Impairment: Evidence and Potential Mechanisms

PubMed Central

Kerner, Nancy A.; Roose, Steven P.

2017-01-01

Obstructive sleep apnea (OSA) is highly prevalent but very frequently undiagnosed. OSA is an independent risk factor for depression and cognitive impairment/dementia. Herein the authors review studies in the literature pertinent to the effects of OSA on the cerebral microvascular and neurovascular systems and present a model to describe the key pathophysiologic mechanisms that may underlie the associations, including hypoperfusion, endothelial dysfunction, and neuroinflammation. Intermittent hypoxia plays a critical role in initiating and amplifying these pathologic processes. Hypoperfusion and impaired cerebral vasomotor reactivity lead to the development or progression of cerebral small vessel disease (C-SVD). Hypoxemia exacerbates these processes, resulting in white matter lesions, white matter integrity abnormalities, and gray matter loss. Blood–brain barrier (BBB) hyperpermeability and neuroinflammation lead to altered synaptic plasticity, neuronal damage, and worsening C-SVD. Thus, OSA may initiate or amplify the pathologic processes of C-SVD and BBB dysfunction, resulting in the development or exacerbation of depressive symptoms and cognitive deficits. Given the evidence that adequate treatment of OSA with continuous positive airway pressure improves depression and neurocognitive functions, it is important to identify OSA when assessing patients with depression or cognitive impairment. Whether treatment of OSA changes the deteriorating trajectory of elderly patients with already-diagnosed vascular depression and cognitive impairment/dementia remains to be determined in randomized controlled trials. PMID:27139243

Myeloid-Derived Suppressor Cells Ameliorate Cyclosporine A-Induced Hypertension in Mice.

PubMed

Chiasson, Valorie L; Bounds, Kelsey R; Chatterjee, Piyali; Manandhar, Lochana; Pakanati, Abhinandan R; Hernandez, Marcos; Aziz, Bilal; Mitchell, Brett M

2018-01-01

The calcineurin inhibitor cyclosporine A (CsA) suppresses the immune system but promotes hypertension, vascular dysfunction, and renal damage. CsA decreases regulatory T cells and this contributes to the development of hypertension. However, CsA's effects on another important regulatory immune cell subset, myeloid-derived suppressor cells (MDSCs), is unknown. We hypothesized that augmenting MDSCs would ameliorate the CsA-induced hypertension and vascular and renal injury and dysfunction and that CsA reduces MDSCs in mice. Daily interleukin-33 treatment, which increased MDSC levels, completely prevented CsA-induced hypertension and vascular and renal toxicity. Adoptive transfer of MDSCs from control mice into CsA-treated mice after hypertension was established dose-dependently reduced blood pressure and vascular and glomerular injury. CsA treatment of aortas and kidneys isolated from control mice for 24 hours decreased relaxation responses and increased inflammation, respectively, and these effects were prevented by the presence of MDSCs. MDSCs also prevented the CsA-induced increase in fibronectin in microvascular and glomerular endothelial cells. Last, CsA dose-dependently reduced the number of MDSCs by inhibiting calcineurin and preventing cell proliferation, as other direct calcineurin signaling pathway inhibitors had the same dose-dependent effect. These data suggest that augmenting MDSCs can reduce the cardiovascular and renal toxicity and hypertension caused by CsA. © 2017 American Heart Association, Inc.

Coexistence of Eph receptor B1 and ephrin B2 in port-wine stain endothelial progenitor cells contributes to clinicopathological vasculature dilatation.

PubMed

Tan, W; Wang, J; Zhou, F; Gao, L; Yin, R; Liu, H; Sukanthanag, A; Wang, G; Mihm, M C; Chen, D-B; Nelson, J S

2017-12-01

Port-wine stain (PWS) is a vascular malformation characterized by progressive dilatation of postcapillary venules, but the molecular pathogenesis remains obscure. To illustrate that PWS endothelial cells (ECs) present a unique molecular phenotype that leads to pathoanatomical PWS vasculatures. Immunohistochemistry and transmission electron microscopy were used to characterize the ultrastructure and molecular phenotypes of PWS blood vessels. Primary culture of human dermal microvascular endothelial cells and in vitro tube formation assay were used for confirmative functional studies. Multiple clinicopathological features of PWS blood vessels during the development and progression of the disease were shown. There were no normal arterioles and venules observed phenotypically and morphologically in PWS skin; arterioles and venules both showed differentiation impairments, resulting in a reduction of arteriole-like vasculatures and defects in capillary loop formation in PWS lesions. PWS ECs showed stemness properties with expression of endothelial progenitor cell markers CD133 and CD166 in non-nodular lesions. They also expressed dual venous/arterial identities, Eph receptor B1 (EphB1) and ephrin B2 (EfnB2). Co-expression of EphB1 and EfnB2 in normal human dermal microvascular ECs led to the formation of PWS-like vasculatures in vitro, for example larger-diameter and thick-walled capillaries. PWS ECs are differentiation-impaired, late-stage endothelial progenitor cells with a specific phenotype of CD133 + /CD166 + /EphB1 + /EfnB2 + , which form immature venule-like pathoanatomical vasculatures. The disruption of normal EC-EC interactions by coexistence of EphB1 and EfnB2 contributes to progressive dilatation of PWS vasculatures. © 2017 British Association of Dermatologists.