The brain-tumor related protein podoplanin regulates synaptic plasticity and hippocampus-dependent learning and memory

PubMed Central

Cicvaric, Ana; Yang, Jiaye; Krieger, Sigurd; Khan, Deeba; Kim, Eun-Jung; Dominguez-Rodriguez, Manuel; Cabatic, Maureen; Molz, Barbara; Acevedo Aguilar, Juan Pablo; Milicevic, Radoslav; Smani, Tarik; Breuss, Johannes M.; Kerjaschki, Dontscho; Pollak, Daniela D.; Uhrin, Pavel; Monje, Francisco J.

2016-01-01

Abstract Introduction: Podoplanin is a cell-surface glycoprotein constitutively expressed in the brain and implicated in human brain tumorigenesis. The intrinsic function of podoplanin in brain neurons remains however uncharacterized. Materials and methods: Using an established podoplanin-knockout mouse model and electrophysiological, biochemical, and behavioral approaches, we investigated the brain neuronal role of podoplanin. Results: Ex-vivo electrophysiology showed that podoplanin deletion impairs dentate gyrus synaptic strengthening. In vivo, podoplanin deletion selectively impaired hippocampus-dependent spatial learning and memory without affecting amygdala-dependent cued fear conditioning. In vitro, neuronal overexpression of podoplanin promoted synaptic activity and neuritic outgrowth whereas podoplanin-deficient neurons exhibited stunted outgrowth and lower levels of p-Ezrin, TrkA, and CREB in response to nerve growth factor (NGF). Surface Plasmon Resonance data further indicated a physical interaction between podoplanin and NGF. Discussion: This work proposes podoplanin as a novel component of the neuronal machinery underlying neuritogenesis, synaptic plasticity, and hippocampus-dependent memory functions. The existence of a relevant cross-talk between podoplanin and the NGF/TrkA signaling pathway is also for the first time proposed here, thus providing a novel molecular complex as a target for future multidisciplinary studies of the brain function in the physiology and the pathology.Key messagesPodoplanin, a protein linked to the promotion of human brain tumors, is required in vivo for proper hippocampus-dependent learning and memory functions.Deletion of podoplanin selectively impairs activity-dependent synaptic strengthening at the neurogenic dentate-gyrus and hampers neuritogenesis and phospho Ezrin, TrkA and CREB protein levels upon NGF stimulation.Surface plasmon resonance data indicates a physical interaction between podoplanin and NGF. On these

The brain-tumor related protein podoplanin regulates synaptic plasticity and hippocampus-dependent learning and memory.

PubMed

Cicvaric, Ana; Yang, Jiaye; Krieger, Sigurd; Khan, Deeba; Kim, Eun-Jung; Dominguez-Rodriguez, Manuel; Cabatic, Maureen; Molz, Barbara; Acevedo Aguilar, Juan Pablo; Milicevic, Radoslav; Smani, Tarik; Breuss, Johannes M; Kerjaschki, Dontscho; Pollak, Daniela D; Uhrin, Pavel; Monje, Francisco J

2016-12-01

Podoplanin is a cell-surface glycoprotein constitutively expressed in the brain and implicated in human brain tumorigenesis. The intrinsic function of podoplanin in brain neurons remains however uncharacterized. Using an established podoplanin-knockout mouse model and electrophysiological, biochemical, and behavioral approaches, we investigated the brain neuronal role of podoplanin. Ex-vivo electrophysiology showed that podoplanin deletion impairs dentate gyrus synaptic strengthening. In vivo, podoplanin deletion selectively impaired hippocampus-dependent spatial learning and memory without affecting amygdala-dependent cued fear conditioning. In vitro, neuronal overexpression of podoplanin promoted synaptic activity and neuritic outgrowth whereas podoplanin-deficient neurons exhibited stunted outgrowth and lower levels of p-Ezrin, TrkA, and CREB in response to nerve growth factor (NGF). Surface Plasmon Resonance data further indicated a physical interaction between podoplanin and NGF. This work proposes podoplanin as a novel component of the neuronal machinery underlying neuritogenesis, synaptic plasticity, and hippocampus-dependent memory functions. The existence of a relevant cross-talk between podoplanin and the NGF/TrkA signaling pathway is also for the first time proposed here, thus providing a novel molecular complex as a target for future multidisciplinary studies of the brain function in the physiology and the pathology. Key messages Podoplanin, a protein linked to the promotion of human brain tumors, is required in vivo for proper hippocampus-dependent learning and memory functions. Deletion of podoplanin selectively impairs activity-dependent synaptic strengthening at the neurogenic dentate-gyrus and hampers neuritogenesis and phospho Ezrin, TrkA and CREB protein levels upon NGF stimulation. Surface plasmon resonance data indicates a physical interaction between podoplanin and NGF. On these grounds, a relevant cross-talk between podoplanin and NGF as well

Evaluation of anti-podoplanin rat monoclonal antibody NZ-1 for targeting malignant gliomas.

PubMed

Kato, Yukinari; Vaidyanathan, Ganesan; Kaneko, Mika Kato; Mishima, Kazuhiko; Srivastava, Nidhi; Chandramohan, Vidyalakshmi; Pegram, Charles; Keir, Stephen T; Kuan, Chien-Tsun; Bigner, Darell D; Zalutsky, Michael R

2010-10-01

Podoplanin/aggrus is a mucin-like sialoglycoprotein that is highly expressed in malignant gliomas. Podoplanin has been reported to be a novel marker to enrich tumor-initiating cells, which are thought to resist conventional therapies and to be responsible for cancer relapse. The purpose of this study was to determine whether an anti-podoplanin antibody is suitable to target radionuclides to malignant gliomas. The binding affinity of an anti-podoplanin antibody, NZ-1 (rat IgG(2a)), was determined by surface plasmon resonance and Scatchard analysis. NZ-1 was radioiodinated with (125)I using Iodogen [(125)I-NZ-1(Iodogen)] or N-succinimidyl 4-guanidinomethyl 3-[(131)I]iodobenzoate ([(131)I]SGMIB-NZ-1), and paired-label internalization assays of NZ-1 were performed. The tissue distribution of (125)I-NZ-1(Iodogen) and that of [(131)I]SGMIB-NZ-1 were then compared in athymic mice bearing glioblastoma xenografts. The dissociation constant (K(D)) of NZ-1 was determined to be 1.2 × 10(-10) M by surface plasmon resonance and 9.8 × 10(-10) M for D397MG glioblastoma cells by Scatchard analysis. Paired-label internalization assays in LN319 glioblastoma cells indicated that [(131)I]SGMIB-NZ-1 resulted in higher intracellular retention of radioactivity (26.3 ± 0.8% of initially bound radioactivity at 8 h) compared to that from the (125)I-NZ-1(Iodogen) (10.0 ± 0.1% of initially bound radioactivity at 8 h). Likewise, tumor uptake of [(131)I]SGMIB-NZ-1 (39.9 ± 8.8 %ID/g at 24 h) in athymic mice bearing D2159MG xenografts in vivo was significantly higher than that of (125)I-NZ-1(Iodogen) (29.7 ± 6.1 %ID/g at 24 h). The overall results suggest that an anti-podoplanin antibody NZ-1 warrants further evaluation for antibody-based therapy against glioblastoma. Copyright © 2010 Elsevier Inc. All rights reserved.

Chimeric Anti-Human Podoplanin Antibody NZ-12 of Lambda Light Chain Exerts Higher Antibody-Dependent Cellular Cytotoxicity and Complement-Dependent Cytotoxicity Compared with NZ-8 of Kappa Light Chain.

PubMed

Kaneko, Mika K; Abe, Shinji; Ogasawara, Satoshi; Fujii, Yuki; Yamada, Shinji; Murata, Takeshi; Uchida, Hiroaki; Tahara, Hideaki; Nishioka, Yasuhiko; Kato, Yukinari

2017-02-01

Podoplanin (PDPN), a type I transmembrane 36-kDa glycoprotein, is expressed not only in normal cells, such as renal epithelial cells (podocytes), lymphatic endothelial cells, and pulmonary type I alveolar cells, but also in cancer cells, including brain tumors and lung squamous cell carcinomas. Podoplanin activates platelet aggregation by binding to C-type lectin-like receptor-2 (CLEC-2) on platelets, and the podoplanin/CLEC-2 interaction facilitates blood/lymphatic vessel separation. We previously produced neutralizing anti-human podoplanin monoclonal antibody (mAb), clone NZ-1 (rat IgG 2a , lambda), which neutralizes the podoplanin/CLEC-2 interaction and inhibits platelet aggregation and cancer metastasis. Human-rat chimeric antibody, NZ-8, was previously developed using variable regions of NZ-1 and human constant regions of heavy chain (IgG 1 ) and light chain (kappa chain). Although NZ-8 showed high antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) against human podoplanin-expressing cancer cells, the binding affinity of NZ-8 was lower than that of NZ-1. Herein, we produced a novel human-rat chimeric antibody, NZ-12, the constant regions of which consist of IgG 1 heavy chain and lambda light chain. Using flow cytometry, we demonstrated that the binding affinity of NZ-12 was much higher than that of NZ-8. Furthermore, ADCC and CDC activities of NZ-12 were significantly increased against glioblastoma cell lines (LN319 and D397) and lung cancer cell line (PC-10). These results suggested that NZ-12 could become a promising therapeutic antibody against podoplanin-expressing brain tumors and lung cancers.

Lymphangiogenesis assessed using three methods is related to tumour grade, breast cancer subtype and expression of basal marker.

PubMed

Niemiec, Joanna; Adamczyk, Agnieszka; Ambicka, Aleksandra; Mucha-Małecka, Anna; Wysocki, Wojciech; Mituś, Jerzy; Ryś, Janusz

2012-11-01

Lymphangiogenesis is a potential indicator of cancer patients' survival. However, there is no standardisation of methodologies applied to the assessment of lymphatic vessel density. In 156 invasive ductal breast cancers (T  1/N+/M0), lymphatic and blood vessels were visualised using podoplanin and CD34, respectively. Based on these markers expression, four parameters were assessed: (i) distribution of podoplanin-stained vessels (DPV) - the percentage of fields with at least one lymphatic vessel (a simple method proposed by us), (ii) lymphatic vessel density (LVD), (iii) LVD to microvessel density ratio (LVD/MVD) and (iv) the expression of podoplanin in cancer-associated fibroblasts. Next, we estimated relations between the above-mentioned parameters and: (i) breast cancer subtype, (ii) tumour grade, and (iii) basal markers expression. We found that intensive lymphangiogenesis, assessed using all studied methods, is positively related to high tumour grade, triple negative or HER2 subtype and expression of basal markers. Whereas, the absence of podoplanin expression in fibroblasts of cancer stroma is related to luminal A subtype, low tumour grade or lack of basal markers expression. Distribution of podoplanin-stained vessels, assessed by a simple method proposed by us (indicating the percentage of fields with at least one lymphatic vessel), might be used instead of the "hot-spot" method.

Spatial distribution of osteopontin, CD44v6 and podoplanin in the lining epithelium of odontogenic keratocyst, and their biological relevance.

PubMed

Kechik, Khamisah Awang; Siar, Chong Huat

2018-02-01

The odontogenic keratocyst (OKC) remains the most challenging jaw cyst to treat because of its locally-aggressive behaviour and high recurrence potential. Emerging evidence suggests that osteopontin, its receptors CD44v6 and integrin α v , and podoplanin, have a role in the local invasiveness of this cyst. However the spatial distribution characteristics of these pro-invasive markers in the lining epithelium of OKC, and their association with the clinicopathologic parameters of OKC are largely unexplored. This study sought to address these issues in comparison with dentigerous cysts (DCs) and radicular cysts (RCs) and to evaluate their biological relevance. A sample consisting of 20 OKC cases, 10 DCs and 10 RCs was subjected to immunohistochemical staining for osteopontin, CD44v6 and integrin α v , and podoplanin, and semiquantitative analysis was performed. All factors (except integrin α v ) were detected heterogeneously in the constitutive layers of the lining epithelium in all three cyst types. Key observations were significant upregulation of CD44v6 and podoplanin in OKC compared to DCs and RCs, suggesting that these protein molecules may play crucial roles in promoting local invasiveness in OKC (P<0.05). Osteopontin underexpression and distribution patterns were indistinctive among all three cysts indicating its limited role as pro-invasive factor. Clinical parameters showed no significant correlations with all protein factors investigated. Present findings suggest that an osteopontin low CD44v6 high and podoplanin high immunoprofile most probably represent epithelial signatures of OKC and are markers of local invasiveness in this cyst. Copyright © 2017 Elsevier Inc. All rights reserved.

Human endothelial precursor cells express tumor endothelial marker 1/endosialin/CD248.

PubMed

Bagley, Rebecca G; Rouleau, Cecile; St Martin, Thia; Boutin, Paula; Weber, William; Ruzek, Melanie; Honma, Nakayuki; Nacht, Mariana; Shankara, Srinivas; Kataoka, Shiro; Ishida, Isao; Roberts, Bruce L; Teicher, Beverly A

2008-08-01

Angiogenesis occurs during normal physiologic processes as well as under pathologic conditions such as tumor growth. Serial analysis of gene expression profiling revealed genes [tumor endothelial markers (TEM)] that are overexpressed in tumor endothelial cells compared with normal adult endothelial cells. Because blood vessel development of malignant tumors under certain conditions may include endothelial precursor cells (EPC) recruited from bone marrow, we investigated TEM expression in EPC. The expression of TEM1 or endosialin (CD248) and other TEM has been discovered in a population of vascular endothelial growth factor receptor 2+/CD31+/CD45-/VE-cadherin+ EPC derived from human CD133+/CD34+ cells. EPC share some properties with fully differentiated endothelial cells from normal tissue, yet reverse transcription-PCR and flow cytometry reveal that EPC express higher levels of endosialin at the molecular and protein levels. The elevated expression of endosialin in EPC versus mature endothelial cells suggests that endosialin is involved in the earlier stages of tumor angiogenesis. Anti-endosialin antibodies inhibited EPC migration and tube formation in vitro. In vivo, immunohistochemistry indicated that human EPC continued to express endosialin protein in a Matrigel plug angiogenesis assay established in nude mice. Anti-endosialin antibodies delivered systemically at 25 mg/kg were also able to inhibit circulating murine EPC in nude mice bearing s.c. SKNAS tumors. EPC and bone marrow-derived cells have been shown previously to incorporate into malignant blood vessels in some instances, yet they remain controversial in the field. The data presented here on endothelial genes that are up-regulated in tumor vasculature and in EPC support the hypothesis that the angiogenesis process in cancer can involve EPC.

Tumor Endothelial Marker Imaging in Melanomas Using Dual-Tracer Fluorescence Molecular Imaging

PubMed Central

Tichauer, Kenneth M.; Deharvengt, Sophie J.; Samkoe, Kimberley S.; Gunn, Jason R.; Bosenberg, Marcus W.; Turk, Mary-Jo; Hasan, Tayyaba; Stan, Radu V.; Pogue, Brian W.

2014-01-01

Purpose Cancer-specific endothelial markers available for intravascular binding are promising targets for new molecular therapies. In this study, a molecular imaging approach of quantifying endothelial marker concentrations (EMCI) is developed and tested in highly light-absorbing melanomas. The approach involves injection of targeted imaging tracer in conjunction with an untargeted tracer, which is used to account for nonspecific uptake and tissue optical property effects on measured targeted tracer concentrations. Procedures Theoretical simulations and a mouse melanoma model experiment were used to test out the EMCI approach. The tracers used in the melanoma experiments were fluorescently labeled anti-Plvap/PV1 antibody (plasmalemma vesicle associated protein Plvap/PV1 is a transmembrane protein marker exposed on the luminal surface of endothelial cells in tumor vasculature) and a fluorescent isotype control antibody, the uptakes of which were measured on a planar fluorescence imaging system. Results The EMCI model was found to be robust to experimental noise under reversible and irreversible binding conditions and was capable of predicting expected overexpression of PV1 in melanomas compared to healthy skin despite a 5-time higher measured fluorescence in healthy skin compared to melanoma: attributable to substantial light attenuation from melanin in the tumors. Conclusions This study demonstrates the potential of EMCI to quantify endothelial marker concentrations in vivo, an accomplishment that is currently unavailable through any other methods, either in vivo or ex vivo. PMID:24217944

Endothelial markers are associated with pancreatic necrosis and overall prognosis in acute pancreatitis: A preliminary cohort study.

PubMed

Chen, Yizhe; Ke, Lu; Meng, Lei; Yang, Qi; Tong, Zhihui; Pan, Yiyuan; Li, Weiqin; Li, Jieshou

Endothelial injury is believed to play an important role in the evolution of pancreatic microcirculatory dysfunction and pancreatic necrosis (PN) in patients with acute pancreatitis (AP). The aim of this study was to investigate the role of three endothelial markers (von Willebrand factor, vWF; E-selectin; endothelial protein C receptor, EPCR) in the early phase of AP, especially the relationship between endothelial markers and PN. From March 2015 to March 2016, 57 AP patients admitted within 72 h of symptom onset in our hospital were included for this study. Blood samples were taken on admission and the clinical characteristics and outcomes of these patients were recorded. The levels of vWF, E-selectin and EPCR were measured using ELISA for analysis and compared with other severity markers of AP. All the three markers were significantly different in healthy control, mild, moderate and severe AP patients. Moreover, the endothelial markers, especially vWF, also showed significant difference in patients with different extent of PN, as well as those with or without MODS. Additionally, the levels of endothelial markers correlated well with other commonly used markers of AP severity. Elevated endothelium-related mediators (vWF, E-selectin and EPCR) appear to participate in the development of PN and may be a potential indicator of overall prognosis. Our results may help clinicians better understand the pathophysiological process of the development of PN. Copyright © 2016 IAP and EPC. Published by Elsevier B.V. All rights reserved.

CD144+ endothelial microparticles as a marker of endothelial injury in neonatal ABO blood group incompatibility.

PubMed

Awad, Hisham A E; Tantawy, Azza A G; El-Farrash, Rania A; Ismail, Eman A; Youssif, Noha M

2014-04-01

ABO antigens are expressed on the surfaces of red blood cells and the vascular endothelium. We studied circulating endothelial microparticles (EMP) in ABO haemolytic disease of the newborn (ABO HDN) as a marker of endothelial activation to test a hypothesis of possible endothelial injury in neonates with ABO HDN, and its relation with the occurrence and severity of haemolysis. Forty-five neonates with ABO HDN were compared with 20 neonates with Rhesus incompatibility (Rh HDN; haemolytic controls) and 20 healthy neonates with matched mother and infant blood groups (healthy controls). Laboratory investigations were done for markers of haemolysis and von Willebrand factor antigen (vWF Ag). EMP (CD144(+)) levels were measured before and after therapy (exchange transfusion and/or phototherapy). vWF Ag and pre-therapy EMP levels were higher in infants with ABO HDN or Rh HDN than in healthy controls, and were significantly higher in babies with ABO HDN than in those with Rh HDN (p<0.05). In ABO HDN, pre-therapy EMP levels were higher in patients with severe hyperbilirubinaemia than in those with mild and moderate disease or those with Rh HDN (p<0.001). Post-therapy EMP levels were lower than pre-therapy levels in both the ABO HDN and Rh HDN groups; however, the decline in EMP levels was particularly evident after exchange transfusion in ABO neonates with severe hyperbilirubinaemia (p<0.001). Multiple regression analysis revealed that the concentrations of haemoglobin, lactate dehydrogenase and indirect bilirubin were independently correlated with pre-therapy EMP levels in ABO HDN. Elevated EMP levels in ABO HDN may reflect an IgG-mediated endothelial injury parallel to the IgG-mediated erythrocyte destruction and could serve as a surrogate marker of vascular dysfunction and disease severity in neonates with this condition.

Endothelial marker-expressing stromal cells are critical for kidney formation.

PubMed

Mukherjee, Elina; Maringer, Katherine; Papke, Emily; Bushnell, Daniel; Schaefer, Caitlin; Kramann, Rafael; Ho, Jacqueline; Humphreys, Benjamin D; Bates, Carlton; Sims-Lucas, Sunder

2017-09-01

Kidneys are highly vascularized and contain many distinct vascular beds. However, the origins of renal endothelial cells and roles of the developing endothelia in the formation of the kidney are unclear. We have shown that the Foxd1-positive renal stroma gives rise to endothelial marker-expressing progenitors that are incorporated within a subset of peritubular capillaries; however, the significance of these cells is unclear. The purpose of this study was to determine whether deletion of Flk1 in the Foxd1 stroma was important for renal development. To that end, we conditionally deleted Flk1 (critical for endothelial cell development) in the renal stroma by breeding-floxed Flk1 mice ( Flk1 fl/fl ) with Foxd1cre mice to generate Foxd1cre; Flk1 fl/fl ( Flk1 ST-/- ) mice. We then performed FACsorting, histological, morphometric, and metabolic analyses of Flk1 ST-/- vs. control mice. We confirmed decreased expression of endothelial markers in the renal stroma of Flk1 ST-/- kidneys via flow sorting and immunostaining, and upon interrogation of embryonic and postnatal Flk1 ST-/- mice, we found they had dilated peritubular capillaries. Three-dimensional reconstructions showed reduced ureteric branching and fewer nephrons in developing Flk1 ST-/- kidneys vs. Juvenile Flk1 ST-/- kidneys displayed renal papillary hypoplasia and a paucity of collecting ducts. Twenty-four-hour urine collections revealed that postnatal Flk1 ST-/- mice had urinary-concentrating defects. Thus, while lineage-tracing revealed that the renal cortical stroma gave rise to a small subset of endothelial progenitors, these Flk1-expressing stromal cells are critical for patterning the peritubular capillaries. Also, loss of Flk1 in the renal stroma leads to nonautonomous-patterning defects in ureteric lineages. Copyright © 2017 the American Physiological Society.

ChLpMab-23: Cancer-Specific Human-Mouse Chimeric Anti-Podoplanin Antibody Exhibits Antitumor Activity via Antibody-Dependent Cellular Cytotoxicity.

PubMed

Kaneko, Mika K; Nakamura, Takuro; Kunita, Akiko; Fukayama, Masashi; Abe, Shinji; Nishioka, Yasuhiko; Yamada, Shinji; Yanaka, Miyuki; Saidoh, Noriko; Yoshida, Kanae; Fujii, Yuki; Ogasawara, Satoshi; Kato, Yukinari

2017-06-01

Podoplanin is expressed in many cancers, including oral cancers and brain tumors. The interaction between podoplanin and its receptor C-type lectin-like receptor 2 (CLEC-2) has been reported to be involved in cancer metastasis and tumor malignancy. We previously established many monoclonal antibodies (mAbs) against human podoplanin using the cancer-specific mAb (CasMab) technology. LpMab-23 (IgG 1 , kappa), one of the mouse anti-podoplanin mAbs, was shown to be a CasMab. However, we have not shown the usefulness of LpMab-23 for antibody therapy against podoplanin-expressing cancers. In this study, we first determined the minimum epitope of LpMab-23 and revealed that Gly54-Leu64 peptide, especially Gly54, Thr55, Ser56, Glu57, Asp58, Arg59, Tyr60, and Leu64 of podoplanin, is a critical epitope of LpMab-23. We further produced human-mouse chimeric LpMab-23 (chLpMab-23) and investigated whether chLpMab-23 exerts antibody-dependent cellular cytotoxicity (ADCC) and antitumor activity. In flow cytometry, chLpMab-23 showed high sensitivity against a podoplanin-expressing glioblastoma cell line, LN319, and an oral cancer cell line, HSC-2. chLpMab-23 also showed ADCC activity against podoplanin-expressing CHO cells (CHO/podoplanin). In xenograft models with HSC-2 and CHO/podoplanin, chLpMab-23 exerts antitumor activity using human natural killer cells, indicating that chLpMab-23 could be useful for antibody therapy against podoplanin-expressing cancers.

The expression of podoplanin protein is a diagnostic marker to distinguish the early infiltration of esophageal squamous cell carcinoma.

PubMed

Chen, Guangyong; Xu, Rui; Yue, Bing; Mei, Xue; Li, Peng; Zhou, Xiaoge; Huang, Shoufang; Gong, Liping; Zhang, Shutian

2017-03-21

The esophageal squamous cell carcinoma (ESCC) is usually develped from low-grade intraepithelial neoplasia (LGIEN) and high-grade intraepithelial neoplasia (HGIEN) to infiltrative squamous cell carcinoma. Till now, it remains hard to screen for infiltration at earlier stages, especially the differentiation between HGEIN and early infiltrative carcinoma. The purpose of this study is to determine a role of podoplanin in differentiating between HGEIN and early infiltrative squamous cell carcinoma. Totally 133 patients pathologically diagnosed with early ESCC and/or precancerous lesions were enrolled.The EnVision two-step IHC staining technique was applied using the monoclonal mouse anti-human Podoplanin antibody (clone number: D2-40). The expressions of PDPN protein on the basal layer of squamous epithelium lesions could be divided into three different patterns: complete type, incomplete (non-continuous) type, or missing type. A diagnosis of HGEIN can be made if the basal layer showed non-continuous or complete expression of PDPN and a diagnosis of early infiltration can be made if the expression of PDPN is completely missing. Our study confirmed that PDPN was a potential biomarker to identify the presence of early infiltrative squamous cell carcinoma.

The expression of podoplanin protein is a diagnostic marker to distinguish the early infiltration of esophageal squamous cell carcinoma

PubMed Central

Chen, Guangyong; Xu, Rui; Yue, Bing; Mei, Xue; Li, Peng; Zhou, Xiaoge; Huang, Shoufang; Gong, Liping; Zhang, Shutian

2017-01-01

The esophageal squamous cell carcinoma (ESCC) is usually develped from low-grade intraepithelial neoplasia (LGIEN) and high-grade intraepithelial neoplasia (HGIEN) to infiltrative squamous cell carcinoma. Till now, it remains hard to screen for infiltration at earlier stages, especially the differentiation between HGEIN and early infiltrative carcinoma. The purpose of this study is to determine a role of podoplanin in differentiating between HGEIN and early infiltrative squamous cell carcinoma. Totally 133 patients pathologically diagnosed with early ESCC and/or precancerous lesions were enrolled.The EnVision two-step IHC staining technique was applied using the monoclonal mouse anti-human Podoplanin antibody (clone number: D2-40). The expressions of PDPN protein on the basal layer of squamous epithelium lesions could be divided into three different patterns: complete type, incomplete (non-continuous) type, or missing type. A diagnosis of HGEIN can be made if the basal layer showed non-continuous or complete expression of PDPN and a diagnosis of early infiltration can be made if the expression of PDPN is completely missing. Our study confirmed that PDPN was a potential biomarker to identify the presence of early infiltrative squamous cell carcinoma. PMID:28086225

[Undifferentiated cutaneous angiosarcoma of the head: identification by the endothelial marker Ulex europaeus agglutinin I].

PubMed

Bork, K; Fries, J; Hoede, N; Korting, G W; Dienes, P

1985-06-01

Cutaneous angiosarcoma of the head is a rare tumor of the elderly and can occur in an undifferentiated form without any clinical or histological signs of the vascular origin of this tumor. In these cases, the tumor can be identified by using endothelial cell markers, such as factor-VIII-related antigen and ulex europaeus agglutinin I, in an immunofluorescence technique or a peroxidase-antiperoxidase method. A 78-year-old patient is described who died within 18 months from such a tumor, which was diagnosed using the endothelial cell marker, ulex europaeus agglutinin I.

PMab-52: Specific and Sensitive Monoclonal Antibody Against Cat Podoplanin for Immunohistochemistry.

PubMed

Yamada, Shinji; Itai, Shunsuke; Nakamura, Takuro; Yanaka, Miyuki; Saidoh, Noriko; Chang, Yao-Wen; Handa, Saori; Harada, Hiroyuki; Kagawa, Yumiko; Ichii, Osamu; Konnai, Satoru; Kaneko, Mika K; Kato, Yukinari

2017-10-01

Podoplanin (PDPN) is expressed in several normal tissues, such as lymphatic endothelial cells, podocytes of renal glomerulus, and type I alveolar cells of lung. PDPN activates platelet aggregation by binding to C-type lectin-like receptor-2 (CLEC-2) on platelet. Although monoclonal antibodies (mAbs) against human PDPN, mouse PDPN, rat PDPN, rabbit PDPN, dog PDPN, and bovine PDPN have been established, anticat PDPN (cPDPN) mAbs have not been developed. In this study, we immunized mice with Chinese hamster ovary (CHO)-K1 cell lines expressing cPDPN, and developed anti-cPDPN mAbs. One of the clones, PMab-52 (IgM, kappa), detected cPDPN specifically in flow cytometry and Western blot analysis. PMab-52 is also useful for detecting feline squamous cell carcinoma cells in immunohistochemical analysis. PMab-52 is expected to be useful for investigating the function of cPDPN in feline carcinomas.

Tumour endothelial marker-1 is expressed in canine Haemangiopericytomas.

PubMed

Fujii, Y; Tsuchiya, T; Morita, R; Kimura, M; Suzuki, K; Machida, N; Mitsumori, K; Shibutani, M

2013-01-01

The aim of this study was to characterize immunohistochemically 18 cases of canine haemangiopericytoma (CHP) using two new candidate markers for pericytes, tumour endothelial marker (TEM)-1 and new glue (NG)-2, as well as the conventional mesenchymal cellular markers, vimentin, α-smooth muscle actin (α-SMA), desmin and von Willebrand factor (vWF). Because pericytes may have the same origin as endothelial or smooth muscle cells or the same differentiation potential as myofibroblasts, 17 cases of leiomyosarcoma (LMS), 20 cases of haemangiosarcoma (HS) and three cases of myofibroblastic sarcoma (MFS) were also examined. Expression of TEM-1 by >10% of the neoplastic population was observed in 94.4% (17/18) of haemangiopericytomas, 23.5% (4/17) of LMSs, 30.0% (6/20) of HSs and 66.7% (2/3) of MFSs. NG-2 expression by >10% of the neoplastic population was observed in 16.7% (3/18) of haemangiopericytomas, 52.9% (9/17) of LMSs, 0% (0/20) of HSs and 33.3% (1/3) of MFSs. Vimentin was expressed by all of tumours. In haemangiopericytoma, the incidence of positive immunoreactivity in >10% of the neoplastic population was 5.6% (1/18) for both α-SMA and desmin and 0% (0/18) for vWF. Considering the phenotypic features of cells expressing TEM-1, CHPs are thought to originate from immature vascular mural cells sharing their phenotype with myofibroblasts. NG-2 expression may be a phenotype of smooth muscle cells rather than pericytes in dogs. Copyright © 2012 Elsevier Ltd. All rights reserved.

Stromal cell markers are differentially expressed in the synovial tissue of patients with early arthritis.

PubMed

Choi, Ivy Y; Karpus, Olga N; Turner, Jason D; Hardie, Debbie; Marshall, Jennifer L; de Hair, Maria J H; Maijer, Karen I; Tak, Paul P; Raza, Karim; Hamann, Jörg; Buckley, Christopher D; Gerlag, Danielle M; Filer, Andrew

2017-01-01

Previous studies have shown increased expression of stromal markers in synovial tissue (ST) of patients with established rheumatoid arthritis (RA). Here, ST expression of stromal markers in early arthritis in relationship to diagnosis and prognostic outcome was studied. ST from 56 patients included in two different early arthritis cohorts and 7 non-inflammatory controls was analysed using immunofluorescence to detect stromal markers CD55, CD248, fibroblast activation protein (FAP) and podoplanin. Diagnostic classification (gout, psoriatic arthritis, unclassified arthritis (UA), parvovirus associated arthritis, reactive arthritis and RA), disease outcome (resolving vs persistent) and clinical variables were determined at baseline and after follow-up, and related to the expression of stromal markers. We observed expression of all stromal markers in ST of early arthritis patients, independent of diagnosis or prognostic outcome. Synovial expression of FAP was significantly higher in patients developing early RA compared to other diagnostic groups and non-inflammatory controls. In RA FAP protein was expressed in both lining and sublining layers. Podoplanin expression was higher in all early inflammatory arthritis patients than controls, but did not differentiate diagnostic outcomes. Stromal marker expression was not associated with prognostic outcomes of disease persistence or resolution. There was no association with clinical or sonographic variables. Stromal cell markers CD55, CD248, FAP and podoplanin are expressed in ST in the earliest stage of arthritis. Baseline expression of FAP is higher in early synovitis patients who fulfil classification criteria for RA over time. These results suggest that significant fibroblast activation occurs in RA in the early window of disease.

Combination of isocitrate dehydrogenase 1 (IDH1) mutation and podoplanin expression in brain tumors identifies patients at high or low risk of venous thromboembolism.

PubMed

Mir Seyed Nazari, P; Riedl, J; Preusser, M; Posch, F; Thaler, J; Marosi, C; Birner, P; Ricken, G; Hainfellner, J A; Pabinger, I; Ay, C

2018-06-01

Essentials Risk stratification for venous thromboembolism (VTE) in patients with brain tumors is challenging. Patients with IDH1 wildtype and high podoplanin expression have a 6-month VTE risk of 18.2%. Patients with IDH1 mutation and no podoplanin expression have a 6-month VTE risk of 0%. IDH1 mutation and podoplanin overexpression in primary brain tumors appear to be exclusive. Background Venous thromboembolism (VTE) is a frequent complication in primary brain tumor patients. Independent studies revealed that podoplanin expression in brain tumors is associated with increased VTE risk, whereas the isocitrate dehydrogenase 1 (IDH1) mutation is associated with very low VTE risk. Objectives To investigate the interrelation between intratumoral podoplanin expression and IDH1 mutation, and their mutual impact on VTE development. Patients/Methods In a prospective cohort study, intratumoral IDH1 R132H mutation and podoplanin were determined in brain tumor specimens (mainly glioma) by immunohistochemistry. The primary endpoint of the study was symptomatic VTE during a 2-year follow-up. Results All brain tumors that expressed podoplanin to a medium-high extent showed also an IDH1 wild-type status. A score based on IDH1 status and podoplanin expression levels allowed prediction of the risk of VTE. Patients with wild-type IDH1 brain tumors and high podoplanin expression had a significantly increased VTE risk compared with those with mutant IDH1 tumors and no podoplanin expression (6-month risk 18.2% vs. 0%). Conclusions IDH1 mutation and podoplanin overexpression seem to be exclusive. Although brain tumor patients with IDH1 mutation are at very low risk of VTE, the risk of VTE in patients with IDH1 wild-type tumors is strongly linked to podoplanin expression levels. © 2018 International Society on Thrombosis and Haemostasis.

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

Urine albumin to creatinine ratio: A marker of early endothelial dysfunction in youth

USDA-ARS?s Scientific Manuscript database

The urine albumin-to-creatinine ratio (UACR) is a useful predictor of cardiovascular (CV) events in adults. Its relationship to vascular function in children is not clear. We investigated whether UACR was related to insulin resistance and endothelial function, a marker of subclinical atherosclerosis...

[Circulating endothelial cells--markers of blood vessel lesions in patients with diffuse liver disease].

PubMed

Dynnik, O B

2006-01-01

The increased level of the circulating endothelial cells (CEC) is in direct dependance with a degree of an endothelial trauma. We defined CEC in blood (cells x 104/L) of 67 adults and children with acute and chronic viral hepatitis (A and B) and healthy volontiars. The author obtained reliable results of increased CEC in all groups consisting of patients with diffuse liver deseases (17,4+/-6,5 and 19,8+/-8,4) in comparison with control groups (3,8+/-1,9 and 3,8+/-1,2) thus CEC can be of practical value as a marker of microvessel lesions of the liver. Endotheliocytemia testifies to be a factor during endothelial trauma in pathogenesis of diffuse liver disease.

Comparative Expression of CD34, Intercellular Adhesion Molecule-1, and Podoplanin and the Presence of Mast Cells in Periapical Granulomas, Cysts, and Residual Cysts.

PubMed

Lopes, Cristiane Barbosa; Armada, Luciana; Pires, Fábio Ramôa

2018-07-01

The aim of the present study was to compare the immunoexpression of CD34, intercellular adhesion molecule-1 (ICAM-1), and podoplanin and the presence of mast cells with clinical, demographic, radiologic, and histologic features from periapical granulomas, periapical cysts, and residual cysts. Thirty-one lesions (5 granulomas, 15 periapical cysts, and 11 residual cysts) were selected. Histologic sections in silanized slides were used for the immunohistochemical reactions. The analysis of the images was performed by using an optical microscope, and data were analyzed with 5% significance (P < .05). Cysts presented atrophic and hyperplastic epithelium in 11 cases (35.5%) and 15 cases (48.8%), respectively (P > .05). The intensity of the inflammatory infiltrate was similar when comparing the 3 groups (P > .05). CD34 and podoplanin expression and the presence of mast cells were similar when comparing the 3 groups; ICAM-1 expression was more intense in granulomas than cysts (P < .05). There were no statistically significant differences associated with the expression of the evaluated markers according to the intensity of the inflammatory infiltrate. There were no differences in the expression of CD34 and podoplanin and in the presence of mast cells when the 3 groups were compared. ICAM-1 expression was more common in periapical granulomas. Copyright © 2018 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Predicting the severity of systemic inflammatory response syndrome (SIRS)-associated coagulopathy with hemostatic molecular markers and vascular endothelial injury markers.

PubMed

Iba, Toshiaki; Gando, Satoshi; Murata, Atsuo; Kushimoto, Shigeki; Saitoh, Daizoh; Eguchi, Yutaka; Ohtomo, Yasuhiro; Okamoto, Kohji; Koseki, Kazuhide; Mayumi, Toshihiko; Ikeda, Toshiaki; Ishhikura, Hiroyasu; Ueyama, Masashi; Ogura, Yuji; Endo, Shigeatsu; Shimazaki, Shuji

2007-11-01

The changes in biomarkers of coagulation or fibrinolysis, anticoagulation, inflammation, and endothelial damage occur in patients with systemic inflammatory response syndrome (SIRS). The purpose of this study is to assess the prognostic value of these markers in patients with SIRS-associated hypercoagulopathy. Sixty-six SIRS patients with a platelet count less than 15.0 x 10(4)/mm3 in three university hospital intensive care units were enrolled in this prospective, comparative study. Blood samples were obtained on day 0 and day 2. Twelve hemostatic, inflammatory, and vascular endothelial indices were measured and the data were compared between the severe group (patients with a total maximum Sequential Organ Failure Assessment score of 10 or more and nonsurvivors; n = 25) and the less-severe group (Sequential Organ Failure Assessment score <10; n = 41). Significant changes between the groups were observed in platelet count, fibrin or fibrinogen degradation products, interleukin-6, soluble thrombomodulin, antithrombin (AT) activity, and protein C activity, both on day 0 and on day 2. In contrast, the d-dimer, soluble fibrin, plasmin-[alpha]2-antiplasmin complex, and E-selectin levels were higher in the severe group only on day 2. No significant difference was seen regarding the thrombin-AT complex and total plasminogen activator inhibitor on both days. A comparison of the areas under the receiver operating characteristic curve revealed the AT activity to be the best predictor of a progression of organ dysfunction. The changes in some hemostatic molecular markers and vascular endothelial markers were conspicuous in patients with organ dysfunction. The AT activity is considered to be the most useful predictor of organ dysfunction.

Coagulopathy in patients with acute pulmonary embolism: a pilot study of whole blood coagulation and markers of endothelial damage.

PubMed

Lehnert, Per; Johansson, Pär I; Ostrowski, Sisse R; Møller, Christian H; Bang, Lia E; Olsen, Peter Skov; Carlsen, Jørn

2017-02-01

Whole blood coagulation and markers of endothelial damage were studied in patients with acute pulmonary embolism (PE), and evaluated in relation to PE severity. Twenty-five patients were enrolled prospectively each having viscoelastical analysis of whole blood done using thrombelastography (TEG) and Multiplate aggregometry. Fourteen of these patients were investigated for endothelial damage by ELISA measurements of Syndecan-1 (endothelial glycocalyx degradation), soluble endothelial Selectin (endothelial cell activation), soluble Thrombomodulin (endothelial cell injury) and Histone Complexed DNA fragments (endothelial cytotoxic histones). The mean values of TEG and Multiplate parameters were all within the reference levels, but a significant difference between patients with high and intermediate risk PE was observed for Ly30 (lytic activity) 1.5% [0-10] vs. 0.2% [0-2.2] p = .04, and ADP (platelet reactivity) 92 U [20-145] vs. 59 U [20-111] p = .03. A similar difference was indicated for functional fibrinogen 21 mm [17-29] vs. 18 mm [3-23] p = .05. Analysis of endothelial markers identified a significant difference in circulating levels between high and intermediate risk PE patients for Syndecan-1 118.6 ng/mL [76-133] vs. 36.3 ng/mL [11.8-102.9] p = .008. In conclusion, patients with acute PE had normal whole blood coagulation, but high risk PE patients had signs of increased activity of the haemostatic system and significantly increased level of endothelial glycocalyx degradation.

Facial nerve hemangiomas: vascular tumors or malformations?

PubMed

Benoit, Margo McKenna; North, Paula E; McKenna, Michael J; Mihm, Martin C; Johnson, Matthew M; Cunningham, Michael J

2010-01-01

To reclassify facial nerve hemangiomas in the context of presently accepted vascular lesion nomenclature by examining histology and immunohistochemical markers. Cohort analysis of patients diagnosed with a facial nerve hemangioma between 1990 and 2008. Collaborative analysis at a specialty hospital and a major academic hospital. Seven subjects were identified on composite review of office charts, a pathology database spanning both institutions, and an encrypted patient registry. Clinical data were compiled, and hematoxylin-eosin-stained specimens were reviewed. For six patients, archived pathological tissue was available for immunohistochemical evaluation of markers specific for infantile hemangioma (glucose transporter protein isoform 1 [GLUT1] and Lewis Y antigen) and for lymphatic endothelial cells (podoplanin). All patients clinically presented with slowly progressive facial weakness at a mean age of 45 years without prior symptomatology. Hemotoxylin-eosin-stained histopathological slides showed irregularly shaped, dilated lesional vessels with flattened endothelial cells, scant smooth muscle, and no internal elastic lamina. Both podoplanin staining for lymphatic endothelial cells and GLUT1 and LewisY antigen staining for infantile hemangioma endothelial cells were negative in lesional vessels in all specimens for which immunohistochemical analysis was performed. Lesions of the geniculate ganglion historically referred to as "hemangiomas" do not demonstrate clinical, histopathological, or immunohistochemical features consistent with a benign vascular tumor, but instead are consistent with venous malformation. We propose that these lesions be classified as "venous vascular malformations of the facial nerve." This nomenclature should more accurately predict clinical behavior and guide therapeutic interventions.

Isolation and Characterization of Human Lung Lymphatic Endothelial Cells

PubMed Central

Lorusso, Bruno; Falco, Angela; Madeddu, Denise; Frati, Caterina; Cavalli, Stefano; Graiani, Gallia; Gervasi, Andrea; Rinaldi, Laura; Lagrasta, Costanza; Maselli, Davide; Gnetti, Letizia; Silini, Enrico M.; Quaini, Eugenio; Ampollini, Luca; Carbognani, Paolo; Quaini, Federico

2015-01-01

Characterization of lymphatic endothelial cells from the respiratory system may be crucial to investigate the role of the lymphatic system in the normal and diseased lung. We describe a simple and inexpensive method to harvest, isolate, and expand lymphatic endothelial cells from the human lung (HL-LECs). Fifty-five samples of healthy lung selected from patients undergoing lobectomy were studied. A two-step purification tool, based on paramagnetic sorting with monoclonal antibodies to CD31 and Podoplanin, was employed to select a pure population of HL-LECs. The purity of HL-LECs was assessed by morphologic criteria, immunocytochemistry, flow cytometry, and functional assays. Interestingly, these cells retain in vitro several receptor tyrosine kinases (RTKs) implicated in cell survival and proliferation. HL-LECs represent a clinically relevant cellular substrate to study lymphatic biology, lymphoangiogenesis, interaction with microbial agents, wound healing, and anticancer therapy. PMID:26137493

Detection of lymphovascular invasion by D2-40 (podoplanin) immunoexpression in endometrial cancer.

PubMed

Weber, Sarah K; Sauerwald, Axel; Pölcher, Martin; Braun, Michael; Debald, Manuel; Serce, Nuran Bektas; Kuhn, Walther; Brunagel-Walgenbach, Giesela; Rudlowski, Christian

2012-10-01

Lymph node involvement is a major feature in tumor spread of endometrial cancer and predicts prognosis. Therefore, evaluation of lymph vessel invasion (LVI) in tumor tissue as a predictor for lymph node metastasis is of great importance. Immunostaining of D2-40 (podoplanin), a specific marker for lymphatic endothelial cells, might be able to increase the detection rate of LVI compared with conventional hematoxylin-eosin (H-E) staining. The aim of this retrospective study was to analyze the eligibility of D2-40-based LVI evaluation for the prediction of lymph node metastases and patients' outcome. Immunohistochemical staining with D2-40 monoclonal antibodies was performed on paraffin-embedded tissue sections of 182 patients with primary endometrioid adenocarcinoma treated in 1 gynecologic cancer center. Tumors were screened for the presence of LVI. Correlations with clinicopathological features and clinical outcome were assessed. Immunostaining of D2-40 significantly increased the frequency LVI detection compared with conventional H-E staining. Lymph vessel invasion was identified by D2-40 in 53 (29.1%) of 182 tumors compared with 34 (18.3%) of 182 carcinomas by routine H-E staining (P = 0.001). D2-40 LVI was detectable in 81.0% (17/21) of nodal-positive tumors and significantly predicted lymph node metastasis (P = 0.001). Furthermore, D2-40 LVI was an independent prognostic factor for patients overall survival considering tumor stage, lymph node involvement, and tumor differentiation (P < 0.01). D2-40-negative tumors confined to the inner half of the myometrium showed an excellent outcome (5-year overall survival, 97.8%). D2-40-based LVI assessment improves the histopathological detection of lymphovascular invasion in endometrial cancer. Furthermore, LVI is of prognostic value and predicts lymph node metastasis. D2-40 LVI detection might help to select endometrial cancer patients who will benefit from a lymphadenectomy.

Endothelial stress products and coagulation markers in patients with multiple myeloma treated with lenalidomide plus dexamethasone: an observational study

PubMed Central

Rosovsky, Rachel; Hong, Fangxin; Tocco, Deanna; Connell, Brendan; Mitsiades, Constantine; Schlossman, Robert; Ghobrial, Irene; Lockridge, Leslie; Warren, Diane; Bradwin, Gary; Doyle, Mary; Munshi, Nikhil; Soiffer, Robert J.; Anderson, Kenneth C.; Weller, Edie; Richardson, Paul

2014-01-01

Summary In this prospective study of patients with relapsed and/or refractory multiple myeloma (MM) treated with lenalidomide and dexamethasone, relationships between markers of endothelial stress and drug administration and incidence of venous thromboembolism (VTE) were assessed. Of 33 enrolled patients, laboratory and treatment data were available for 32 patients. Of these, 23 received pulsed dexamethasone (40 mg/day on days 1–4, 9–12, and 17–21 of each 28-day cycle) and 9 received weekly dexamethasone (40 mg/day on days 1, 8, 15, and 21 of each cycle). The overall incidence of VTE was 9%. A decreasing trend in markers values was observed with intercellular adhesion molecule (P = 0·05), fibrinogen (P = 0·008), plasminogen activator inhibitor-1 (P < 0·001), homocysteine (P = 0·002), and P-selectin (P < 0·001) during therapy. Compared with weekly dexamethasone, pulsed dexamethasone was associated with significantly greater variation in mean adjusted relative values of fibrinogen, P-selectin, and vascular endothelial growth factor (P < 0·001 for all comparisons), although there was no apparent association with VTE incidence. Lenalidomide plus dexamethasone affects endothelial stress marker levels in patients with advanced MM. The higher variation seen with pulsed dexamethasone suggests greater endothelial stress with this approach. PMID:23240658

Podoplanin-positive cancer-associated fibroblast recruitment within cancer stroma is associated with a higher number of single nucleotide variants in cancer cells in lung adenocarcinoma.

PubMed

Nakasone, Shoko; Mimaki, Sachiyo; Ichikawa, Tomohiro; Aokage, Keiju; Miyoshi, Tomohiro; Sugano, Masato; Kojima, Motohiro; Fujii, Satoshi; Kuwata, Takeshi; Ochiai, Atsushi; Tsuboi, Masahiro; Goto, Koichi; Tsuchihara, Katsuya; Ishii, Genichiro

2018-05-01

Podoplanin-positive cancer-associated fibroblasts (CAFs) play an essential role in tumor progression. However, it is still unclear whether specific genomic alterations of cancer cells are required to recruit podoplanin-positive CAFs. The aim of this study was to investigate the relationship between the mutation status of lung adenocarcinoma cells and the presence of podoplanin-positive CAFs. Ninety-seven lung adenocarcinomas for which whole exome sequencing data were available were enrolled. First, we analyzed the clinicopathological features of the cases, and then, evaluated the relationship between genetic features of cancer cells (major driver mutations and the number of single nucleotide variants, SNVs) and the presence of podoplanin-positive CAFs. The presence of podoplanin-positive CAFs was associated with smoking history, solid predominant subtype, and lymph node metastasis. We could not find any significant correlations between major genetic mutations (EGFR, KRAS, TP53, MET, ERBB2, BRAF, and PIC3CA) in cancer cells and the presence of podoplanin-positive CAFs. However, cases with podoplanin-positive CAFs had a significantly higher number of SNVs in cancer cells than the podoplanin-negative CAFs cases (median 84 vs 37, respectively; p = 0.001). This was also detected in a non-smoker subgroup (p = 0.037). Multivariate analyses revealed that the number of SNVs in cancer cells was the only statistically significant independent predictor for the presence of podoplanin-positive CAFs (p = 0.044). In lung adenocarcinoma, the presence of podoplanin-positive CAFs was associated with higher numbers of SNVs in cancer cells, suggesting a relationship between accumulations of SNVs in cancer cells and the generation of a tumor-promoting microenvironment.

Co-Expression of Bmi-1 and Podoplanin Predicts Overall Survival in Patients With Squamous Cell Carcinoma of the Head and Neck Treated With Radio(chemo)therapy

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

Vormittag, Laurenz; Thurnher, Dietmar; Geleff, Silvana

2009-03-01

Purpose: This study was conducted to determine the expression of Bmi-1 and podoplanin in healthy oral mucosa and in untreated tumor tissues samples of patients with squamous cell carcinomas of the head and neck. All patients were treated by primary radio(chemo)therapy. Methods and Materials: The expression of Bmi-1 and podoplanin was immunohistochemically evaluated in 12 normal oral mucosa and 63 tumor specimens and correlated with patients' clinical data. Results: In healthy mucosa expression of Bmi-1 and podoplanin was restricted to the basal cell layer. Expression of both proteins was found in 79% and 86% of our tumor samples, respectively. Inmore » 17 and 8 samples, Bmi-1 and podoplanin were co-expressed at the invasive border or diffuse in the bulk of the tumor, respectively. Univariate analysis showed that the co-expression of Bmi-1 and podoplanin correlated to decreased overall survival (p = 0.044). Moreover, multivariate testing identified high expression of podoplanin (p = 0.044), co-expression of Bmi-1 and podoplanin (p = 0.007) and lack of response to therapy (p < 0.0001) as predictors of shortened overall survival in patients treated with primary radio(chemo)therapy. Conclusions: Bmi-1 and podoplanin are expressed at the invasive front of squamous cell carcinomas of the head and neck. Co-expression of Bmi-1 and podoplanin predicts significantly overall survival of patients treated with primary radio(chemo)therapy.« less

Antitumor effect of novel anti-podoplanin antibody NZ-12 against malignant pleural mesothelioma in an orthotopic xenograft model.

PubMed

Abe, Shinji; Kaneko, Mika Kato; Tsuchihashi, Yuki; Izumi, Toshihiro; Ogasawara, Satoshi; Okada, Naoto; Sato, Chiemi; Tobiume, Makoto; Otsuka, Kenji; Miyamoto, Licht; Tsuchiya, Koichiro; Kawazoe, Kazuyoshi; Kato, Yukinari; Nishioka, Yasuhiko

2016-09-01

Podoplanin (aggrus) is highly expressed in several types of cancers, including malignant pleural mesothelioma (MPM). Previously, we developed a rat anti-human podoplanin mAb, NZ-1, and a rat-human chimeric anti-human podoplanin antibody, NZ-8, derived from NZ-1, which induced antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity against podoplanin-positive MPM cell lines. In this study, we showed the antitumor effect of NZ-1, NZ-8, and NZ-12, a novel rat-human chimeric anti-human podoplanin antibody derived from NZ-1, in an MPM orthotopic xenograft SCID mouse model. Treatment with NZ-1 and rat NK (CD161a(+) ) cells inhibited the growth of tumors and the production of pleural effusion in NCI-H290/PDPN or NCI-H226 orthotopic xenograft mouse models. NZ-8 and human natural killer (NK) (CD56(+) ) cells also inhibited tumor growth and pleural effusion in MPM orthotopic xenograft mice. Furthermore, NZ-12 induced potent ADCC mediated by human MNC, compared with either NZ-1 or NZ-8. Antitumor effects were observed following treatment with NZ-12 and human NK (CD56(+) ) cells in MPM orthotopic xenograft mice. In addition, combined immunotherapy using the ADCC activity of NZ-12 mediated by human NK (CD56(+) ) cells with pemetrexed, led to enhanced antitumor effects in MPM orthotopic xenograft mice. These results strongly suggest that combination therapy with podoplanin-targeting immunotherapy using both NZ-12 and pemetrexed might provide an efficacious therapeutic strategy for the treatment of MPM. © 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

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.

[Placental atherosclerosis and markers of endothelial dysfunction in infants born to mothers with gestational diabetes].

PubMed

López Morales, Cruz Mónica; Brito Zurita, Olga Rosa; González Heredia, Ricardo; Cruz López, Miguel; Méndez Padrón, Araceli; Matute Briseño, Juan Antonio

2016-08-05

The pathophysiology of gestational diabetes itself causes hyperstimulation of adipose tissue and of the placenta cells increasing the production of inflammatory cytokines, which cause changes in the tissues exposed such as the placenta and foetus. Therefore, the objective of this study was to compare metabolic markers and endothelial dysfunction in umbilical cord blood, as well as to determine the presence of atherosclerosis in the placentas of newborn infants of patients with gestational diabetes and in patients with normally progressing pregnancies. An analytical cross-sectional study was carried out in 84 patients, obtaining data such as age, smoking and weight gain in pregnancy; the gestational age of the newborns was determined by Capurro, and their weight and destination subsequent to birth, the placentas were also collected in order to look for atherosclerosis through histological studies and glucose, insulin, VLDL-C, HDL-C, triglycerides, cholesterol, fibrinogen, PCR and markers of endothelial dysfunction (adiponectin, VCAM-1, ICAM-1 and IL-6) were determined in blood samples obtained from the umbilical cord. Placental atherosclerosis presented in 28.94% of the group with gestational diabetes compared to 10.52% of the group with normally progressing pregnancies (P=.044); differences were found in glucose, cholesterol, triglycerides, fibrinogen, HOMA-IR, PCR-us, HDL-C, not in VLDL-C. Twenty-one point five percent of the newborns of the gestational diabetes patients required hospitalization, against 5.2% in the control group, Pregnancies that involve diabetes have higher proportion of atherosclerosis, hospitalization of the newborn, insulin resistance, as well as elevation of markers associated with inflammation and endothelial dysfunction in umbilical cord blood. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

Tumor endothelial marker 5 expression in endothelial cells during capillary morphogenesis is induced by the small GTPase Rac and mediates contact inhibition of cell proliferation

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

Vallon, Mario, E-mail: m.vallon@arcor.de; Rohde, Franziska; Janssen, Klaus-Peter

2010-02-01

Tumor endothelial marker (TEM) 5 is an adhesion G-protein-coupled receptor upregulated in endothelial cells during tumor and physiologic angiogenesis. So far, the mechanisms leading to upregulation of TEM5 and its function during angiogenesis have not been identified. Here, we report that TEM5 expression in endothelial cells is induced during capillary-like network formation on Matrigel, during capillary morphogenesis in a three-dimensional collagen I matrix, and upon confluence on a two-dimensional matrix. TEM5 expression was not induced by a variety of soluble angiogenic factors, including VEGF and bFGF, in subconfluent endothelial cells. TEM5 upregulation was blocked by toxin B from Clostridium difficile,more » an inhibitor of the small GTPases Rho, Rac, and Cdc42. The Rho inhibitor C3 transferase from Clostridium botulinum did not affect TEM5 expression, whereas the Rac inhibitor NSC23766 suppressed TEM5 upregulation. An excess of the soluble TEM5 extracellular domain or an inhibitory monoclonal TEM5 antibody blocked contact inhibition of endothelial cell proliferation resulting in multilayered islands within the endothelial monolayer and increased vessel density during capillary formation. Based on our results we conclude that TEM5 expression during capillary morphogenesis is induced by the small GTPase Rac and mediates contact inhibition of proliferation in endothelial cells.« less

Plasma markers of inflammation and hemostatic and endothelial activity in naturally overweight and obese dogs.

PubMed

Barić Rafaj, R; Kuleš, J; Marinculić, A; Tvarijonaviciute, A; Ceron, J; Mihaljević, Ž; Tumpa, A; Mrljak, V

2017-01-06

Obesity is one of the most prevalent health problems in the canine population. While haemostatic parameters and markers of endothelial function have been evaluated in various disease conditions in dogs, there are no studies of these markers in canine obesity. This study was designed to evaluate the effect of naturally gained weight excess and obesity on inflammatory, hemostatic and endothelial biomarkers in dogs. A total of 37 overweight and obese dogs were compared with 28 normal weight dogs. Overweight and obese dogs had significantly elevated concentrations of serum interleukin-6 (IL-6) and C-reactive protein (hsCRP). Number of platelets, activity of factor X and factor VII were significantly higher, while activated partial thromboplastine time (aPTT) and soluble plasminogen activator receptor (suPAR) were significantly decreased. Statistical analysis of high mobility group box - 1 protein (HMGB-1), soluble intercellular adhesive molecule -1 (sICAM-1) and plasminogen activator inhibitor type 1 (PAI-1) concentrations did not show significant differences between the total overweight and obese group and the normal weight group of dogs. Analytical changes in the dogs in our study reflects that weight excess in dogs can be associated with a chronic low degree of inflammation and a hypercoagulable state, where primary and secondary hemostasis are both affected. However obesity is not associated with impairment of endothelial function in dogs.

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.

Markers of endothelial dysfunction and severity of hypoxaemia in the Eisenmenger syndrome.

PubMed

de P S Soares, Rosangela; Maeda, Nair Y; Bydlowski, Sérgio P; Lopes, Antonio Augusto

2005-10-01

Endothelial dysfunction has been reported in hypoxaemic patients with the Eisenmenger syndrome, but a direct correlation between levels of endothelial markers and the severity of hypoxaemia has not been explored. With this in mind, we compared the levels in the plasma of tissue-type plasminogen activator, thrombomodulin, and von Willebrand factor in 25 patients with the Eisenmenger syndrome. They had a median age of 31 years, and were divided into 2 groups according to their recent clinical history. Thus, 18 patients were stable, being in functional class II or III, seen as outpatients, and having peripheral saturations of oxygen of 89 plus or minus 5 percent. In contrast, 7 patients were unstable, showing episodes of symptoms placing them in functional class IV, requiring care in hospital, and manifesting saturations of oxygen of 77 plus or minus 5 percent. We were able to follow 12 patients, 8 who were stable and 4 unstable, for 24 months. At baseline, levels of von Willebrand factor were higher in the unstable patients when compared to those who were stable, at 142 plus or minus 29 and 110 plus or minus 25 units per decilitre, respectively (p equal to 0.013). This correlated positively with oxygen desaturation (p less than 0.020). The structural abnormalities also correlated positively with the magnitude of hypoxaemia (p less than 0.020). Levels remained higher in the unstable patients throughout the period of follow-up (p equal to 0.006). Tissue-type plasminogen activator was also increased, at 14.3 plus or minus 8.4 versus 6.5 plus or minus 2.7 nanograms per millilitre in controls (p less than 0.001), whereas thrombomodulin was decreased, with values of 14.4 versus 34.6 nanograms per millilitre in controls (p for median values of less than 0.001). There was no correlation with saturations of oxygen. We conclude that measurement of von Willebrand factor, as compared with tissue-type plasminogen activator and thrombomodulin, will prove a better marker of

Comparison of skin microvascular reactivity with hemostatic markers of endothelial dysfunction and damage in type 2 diabetes

PubMed Central

Beer, Sandra; Feihl, François; Ruiz, Juan; Juhan-Vague, Irène; Aillaud, Marie-Françoise; Wetzel, Sandrine Golay; Liaudet, Lucas; Gaillard, Rolf C; Waeber, Bernard

2008-01-01

Aim: Patients with non-insulin-dependent diabetes mellitus (NIDDM) are at increased cardiovascular risk due to an accelerated atherosclerotic process. The present study aimed to compare skin microvascular function, pulse wave velocity (PWV), and a variety of hemostatic markers of endothelium injury [von Willebrand factor (vWF), plasminogen activator inhibitor-1 (PAI-1), tissue plasminogen activator (t-PA), tissue factor pathway inhibitor (TFPI), and the soluble form of thrombomodulin (s-TM)] in patients with NIDDM. Methods: 54 patients with NIDDM and 38 sex- and age-matched controls were studied. 27 diabetics had no overt micro- and/or macrovascular complications, while the remainder had either or both. The forearm skin blood flow was assessed by laser-Doppler imaging, which allowed the measurement of the response to iontophoretically applied acetylcholine (endothelium-dependent vasodilation) and sodium nitroprusside (endothelium-independent vasodilation), as well as the reactive hyperemia triggered by the transient occlusion of the circulation. Results: Both endothelial and non-endothelial reactivity were significantly blunted in diabetics, regardless of the presence or the absence of vascular complications. Plasma vWF, TFPI and s-TM levels were significantly increased compared with controls only in patients exhibiting vascular complications. Concentrations of t-PA and PAI-1 were significantly increased in the two groups of diabetics versus controls. Conclusion: In NIDDM, both endothelium-dependent and -independent microvascular skin reactivity are impaired, whether or not underlying vascular complications exist. It also appears that microvascular endothelial dysfunction is not necessarily associated in NIDDM with increased circulating levels of hemostatic markers of endothelial damage known to reflect a hypercoagulable state. PMID:19337558

Increased endothelial and macrophage markers are associated with disease severity and mortality in scrub typhus.

PubMed

Otterdal, Kari; Janardhanan, Jeshina; Astrup, Elisabeth; Ueland, Thor; Prakash, John A J; Lekva, Tove; Abraham, O C; Thomas, Kurien; Damås, Jan Kristian; Mathews, Prasad; Mathai, Dilip; Aukrust, Pål; Varghese, George M

2014-11-01

Scrub typhus is endemic in the Asia-Pacific region. Mortality is high even with treatment, and further knowledge of the immune response during this infection is needed. This study was aimed at comparing plasma levels of monocyte/macrophage and endothelial related inflammatory markers in patients and controls in South India and to explore a possible correlation to disease severity and clinical outcome. Plasma levels of ALCAM, VCAM-1, sCD163, sCD14, YKL-40 and MIF were measured in scrub typhus patients (n = 129), healthy controls (n = 31) and in infectious disease controls (n = 31), both in the acute phase and after recovery, by enzyme immunoassays. Patients had markedly elevated levels of all mediators in the acute phase, differing from both healthy and infectious disease controls. During follow-up levels of ALCAM, VCAM-1, sCD14 and YKL-40 remained elevated compared to levels in healthy controls. High plasma ALCAM, VCAM-1, sCD163, sCD14, and MIF, and in particular YKL-40 were all associated with disease severity and ALCAM, sCD163, MIF and especially YKL-40, were associated with mortality. Our findings show that scrub typhus is characterized by elevated levels of monocyte/macrophage and endothelial related markers. These inflammatory markers, and in particular YKL-40, may contribute to disease severity and clinical outcome. Copyright © 2014 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

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

Impact of vitamin D supplementation on endothelial and inflammatory markers in adults: A systematic review.

PubMed

Agbalalah, Tari; Hughes, Stephen F; Freeborn, Ellen J; Mushtaq, Sohail

2017-10-01

This systematic review aims to evaluate randomised controlled trials (RCTs) investigating the effect of vitamin D supplementation on endothelial function and inflammation in adults. An electronic search of published randomised controlled trials, using Cochrane, Pubmed and Medline databases was conducted, with the search terms related to vitamin D and endothelial function. Inclusion criteria were RCTs in adult humans with a measure of vitamin D status using serum/plasma 25(OH)D and studies which administered the intervention through the oral route. Among the 1107 studies retrieved, 29 studies met the full inclusion criteria for this systematic review. Overall, 8 studies reported significant improvements in the endothelial/inflammatory biomarkers/parameters measured. However, in 2 out of the 8 studies, improvements were reported at interim time points, but improvements were absent post-intervention. The remaining 21 trial studies did not show significant improvements in the markers of interest measured. Evidence from the studies included in this systematic review did not demonstrate that vitamin D supplementation in adults, results in an improvement in circulating inflammatory and endothelial function biomarkers/parameters. This systematic review does not therefore support the use of vitamin D supplementation as a therapeutic or preventative measure for CVD in this respect. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

A Real-Time Near-Infrared Fluorescence Imaging Method for the Detection of Oral Cancers in Mice Using an Indocyanine Green-Labeled Podoplanin Antibody.

PubMed

Ito, Akihiro; Ohta, Mitsuhiko; Kato, Yukinari; Inada, Shunko; Kato, Toshio; Nakata, Susumu; Yatabe, Yasushi; Goto, Mitsuo; Kaneda, Norio; Kurita, Kenichi; Nakanishi, Hayao; Yoshida, Kenji

2018-01-01

Podoplanin is distinctively overexpressed in oral squamous cell carcinoma than oral benign neoplasms and plays a crucial role in the pathogenesis and metastasis of oral squamous cell carcinoma but its diagnostic application is quite limited. Here, we report a new near-infrared fluorescence imaging method using an indocyanine green (ICG)-labeled anti-podoplanin antibody and a desktop/a handheld ICG detection device for the visualization of oral squamous cell carcinoma-xenografted tumors in nude mice. Both near-infrared imaging methods using a desktop (in vivo imaging system: IVIS) and a handheld device (photodynamic eye: PDE) successfully detected oral squamous cell carcinoma tumors in nude mice in a podoplanin expression-dependent manner with comparable sensitivity. Of these 2 devices, only near-infrared imaging methods using a handheld device visualized oral squamous cell carcinoma xenografts in mice in real time. Furthermore, near-infrared imaging methods using the handheld device (PDE) could detect smaller podoplanin-positive oral squamous cell carcinoma tumors than a non-near-infrared, autofluorescence-based imaging method. Based on these results, a near-infrared imaging method using an ICG-labeled anti-podoplanin antibody and a handheld detection device (PDE) allows the sensitive, semiquantitative, and real-time imaging of oral squamous cell carcinoma tumors and therefore represents a useful tool for the detection and subsequent monitoring of malignant oral neoplasms in both preclinical and some clinical settings.

A Real-Time Near-Infrared Fluorescence Imaging Method for the Detection of Oral Cancers in Mice Using an Indocyanine Green–Labeled Podoplanin Antibody

PubMed Central

Ito, Akihiro; Ohta, Mitsuhiko; Kato, Yukinari; Inada, Shunko; Kato, Toshio; Nakata, Susumu; Yatabe, Yasushi; Goto, Mitsuo; Kaneda, Norio; Kurita, Kenichi; Nakanishi, Hayao; Yoshida, Kenji

2018-01-01

Podoplanin is distinctively overexpressed in oral squamous cell carcinoma than oral benign neoplasms and plays a crucial role in the pathogenesis and metastasis of oral squamous cell carcinoma but its diagnostic application is quite limited. Here, we report a new near-infrared fluorescence imaging method using an indocyanine green (ICG)–labeled anti-podoplanin antibody and a desktop/a handheld ICG detection device for the visualization of oral squamous cell carcinoma–xenografted tumors in nude mice. Both near-infrared imaging methods using a desktop (in vivo imaging system: IVIS) and a handheld device (photodynamic eye: PDE) successfully detected oral squamous cell carcinoma tumors in nude mice in a podoplanin expression–dependent manner with comparable sensitivity. Of these 2 devices, only near-infrared imaging methods using a handheld device visualized oral squamous cell carcinoma xenografts in mice in real time. Furthermore, near-infrared imaging methods using the handheld device (PDE) could detect smaller podoplanin-positive oral squamous cell carcinoma tumors than a non-near-infrared, autofluorescence-based imaging method. Based on these results, a near-infrared imaging method using an ICG-labeled anti-podoplanin antibody and a handheld detection device (PDE) allows the sensitive, semiquantitative, and real-time imaging of oral squamous cell carcinoma tumors and therefore represents a useful tool for the detection and subsequent monitoring of malignant oral neoplasms in both preclinical and some clinical settings. PMID:29649929

Platelet CLEC-2 protects against lung injury via effects of its ligand podoplanin on inflammatory alveolar macrophages in the mouse

PubMed Central

Rayes, Julie; Wichaiyo, Surasak; Haining, Elizabeth J.; Lowe, Kate; Grygielska, Beata; Laloo, Ryan; Flodby, Per; Borok, Zea; Crandall, Edward D.; Thickett, David R.; Watson, Steve P.

2017-01-01

There is no therapeutic intervention proven to prevent acute respiratory distress syndrome (ARDS). Novel mechanistic insights into the pathophysiology of ARDS are therefore required. Platelets are implicated in regulating many of the pathogenic processes that occur during ARDS; however, the mechanisms remain elusive. The platelet receptor CLEC-2 has been shown to regulate vascular integrity at sites of acute inflammation. Therefore the purpose of this study was to establish the role of CLEC-2 and its ligand podoplanin in a mouse model of ARDS. Platelet-specific CLEC-2-deficient, as well as alveolar epithelial type I cell (AECI)-specific or hematopoietic-specific podoplanin deficient, mice were established using cre-loxP strategies. Combining these with intratracheal (IT) instillations of lipopolysaccharide (LPS), we demonstrate that arterial oxygen saturation decline in response to IT-LPS in platelet-specific CLEC-2-deficient mice is significantly augmented. An increase in bronchoalveolar lavage (BAL) neutrophils and protein was also observed 48 h post-IT-LPS, with significant increases in pro-inflammatory chemokines detected in BAL of platelet-specific CLEC-2-deficient animals. Deletion of podoplanin from hematopoietic cells but not AECIs also reduces lung function and increases pro-inflammatory chemokine expression following IT-LPS. Furthermore, we demonstrate that following IT-LPS, platelets are present in BAL in aggregates with neutrophils, which allows for CLEC-2 interaction with podoplanin expressed on BAL inflammatory alveolar macrophages. Taken together, these data suggest that the platelet CLEC-2-podoplanin signaling axis regulates the severity of lung inflammation in mice and is a possible novel target for therapeutic intervention in patients at risk of developing ARDS. PMID:28839100

Garlic extract favorably modifies markers of endothelial function in obese patients -randomized double blind placebo-controlled nutritional intervention.

PubMed

Szulińska, Monika; Kręgielska-Narożna, Matylda; Świątek, Joanna; Styś, Paulina; Kuźnar-Kamińska, Barbara; Jakubowski, Hieronim; Walkowiak, Jarosław; Bogdański, Paweł

2018-06-01

Garlic exerts a range of effects relevant to human health. However, its influence on the endothelium in obese individuals remains unknown. We aimed to determine the effects of garlic extract (GE) on arterial stiffness and markers of endothelial function. Ninety-two subjects were enrolled in this study. The participants were randomly assigned to receive 400 mg of GE or placebo daily for 3 months. The arterial stiffness index (SI) and markers of endothelial function such as high-sensitivity C-reactive protein (hsCRP), cholesterol (total, LDL, HDL), triglycerides, and plasminogen activator inhibitor 1 (PAI-1), as well as total antioxidant status (TAS) were quantified at baseline and the end of study. At the end of study SI (p = 0.01), hsCRP (p < 0.001, PAI-1 (p < 0.001), LDL cholesterol (p < 0.001), and TAS (p < 0.01) were reduced in the GE-supplemented group, but not in the placebo group. This randomized, double-blind, placebo-controlled trial demonstrates that supplementation with GE favorably modifies endothelial biomarkers associated with cardiovascular risk and suggests that GE can be used to suppress chronic inflammation in obese individuals. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Citrus Polyphenol Hesperidin Stimulates Production of Nitric Oxide in Endothelial Cells while Improving Endothelial Function and Reducing Inflammatory Markers in Patients with Metabolic Syndrome

PubMed Central

Rizza, Stefano; Muniyappa, Ranganath; Iantorno, Micaela; Kim, Jeong-a; Chen, Hui; Pullikotil, Philomena; Senese, Nicoletta; Tesauro, Manfredi; Lauro, Davide; Cardillo, Carmine

2011-01-01

Context: Hesperidin, a citrus flavonoid, and its metabolite hesperetin may have vascular actions relevant to their health benefits. Molecular and physiological mechanisms of hesperetin actions are unknown. Objective: We tested whether hesperetin stimulates production of nitric oxide (NO) from vascular endothelium and evaluated endothelial function in subjects with metabolic syndrome on oral hesperidin therapy. Design, Setting, and Interventions: Cellular mechanisms of action of hesperetin were evaluated in bovine aortic endothelial cells (BAEC) in primary culture. A randomized, placebo-controlled, double-blind, crossover trial examined whether oral hesperidin administration (500 mg once daily for 3 wk) improves endothelial function in individuals with metabolic syndrome (n = 24). Main Outcome Measure: We measured the difference in brachial artery flow-mediated dilation between placebo and hesperidin treatment periods. Results: Treatment of BAEC with hesperetin acutely stimulated phosphorylation of Src, Akt, AMP kinase, and endothelial NO synthase to produce NO; this required generation of H2O2. Increased adhesion of monocytes to BAEC and expression of vascular cell adhesion molecule-1 in response to TNF-α treatment was reduced by pretreatment with hesperetin. In the clinical study, when compared with placebo, hesperidin treatment increased flow-mediated dilation (10.26 ± 1.19 vs. 7.78 ± 0.76%; P = 0.02) and reduced concentrations of circulating inflammatory biomarkers (high-sensitivity C-reactive protein, serum amyloid A protein, soluble E-selectin). Conclusions: Novel mechanisms for hesperetin action in endothelial cells inform effects of oral hesperidin treatment to improve endothelial dysfunction and reduce circulating markers of inflammation in our exploratory clinical trial. Hesperetin has vasculoprotective actions that may explain beneficial cardiovascular effects of citrus consumption. PMID:21346065

Occludin as a functional marker of vascular endothelial cells on tube-forming activity.

PubMed

Kanayasu-Toyoda, Toshie; Ishii-Watabe, Akiko; Kikuchi, Yutaka; Kitagawa, Hiroko; Suzuki, Hiroko; Tamura, Hiroomi; Tada, Minoru; Suzuki, Takuo; Mizuguchi, Hiroyuki; Yamaguchi, Teruhide

2018-02-01

Cell therapy using endothelial progenitor cells (EPCs) is a promising strategy for the treatment of ischemic diseases. Two types of EPCs have been identified: early EPCs and late EPCs. Late EPCs are able to form tube structure by themselves, and have a high proliferative ability. The functional marker(s) of late EPCs, which relate to their therapeutic potential, have not been fully elucidated. Here we compared the gene expression profiles of several human cord blood derived late EPC lines which exhibit different tube formation activity, and we observed that the expression of occludin (OCLN) in these lines correlated with the tube formation ability, suggesting that OCLN is a candidate functional marker of late EPCs. When OCLN was knocked down by transfecting siRNA, the tube formation on Matrigel, the S phase + G 2 /M phase in the cell cycle, and the spheroid-based sprouting of late EPCs were markedly reduced, suggesting the critical role of OCLN in tube formation, sprouting, and proliferation. These results indicated that OCLN plays a novel role in neovascularization and angiogenesis. © 2017 Wiley Periodicals, Inc.

Circulating endothelial cells as marker of endothelial damage in male hypogonadism.

PubMed

Milardi, Domenico; Grande, Giuseppe; Giampietro, Antonella; Vendittelli, Francesca; Palumbo, Sara; Tartaglione, Linda; Marana, Riccardo; Pontecorvi, Alfredo; de Marinis, Laura; Zuppi, Cecilia; Capoluongo, Ettore

2012-01-01

Testosterone deficiency has become a frequently diagnosed condition in today's society affected by epidemic obesity, and is associated with cardiovascular risk. Recent studies have established the importance of altered vascular endothelium function in cardiovascular disease. The damage to the endothelium might also cause endothelial cell detachment, resulting in increased numbers of circulating endothelial cells (CEC) within the bloodstream. To evaluate whether hypogonadism could modify CEC count in peripheral bloodstream, we investigated peripheral blood CEC count using the CellSearch System, a semiautomatic method to accurately and reliably enumerate CECs, which are sorted based on a CD146(+), CD105(+), DAPI(+), CD45(-) phenotype, in a population of 20 patients with hypogonadism. The control group comprised 10 age- and sex-matched healthy participants. CEC count per milliliter was significantly increased in patients with hypogonadism vs the control group. In the group with hypogonadism, an inverse exponential correlation was present between testosterone levels and CEC count per milliliter. A direct linear correlation was present between waist circumference and CECs and between body mass index and CECs. The regression analysis showed that testosterone was the significant independent determinant of CECs. Our results underline that male hypogonadism is associated with endothelial dysfunction. The correlation between CEC and waist circumference underlines that visceral obesity may be synergically implicated in this regulation. Future studies are required to unveil the mechanisms involved in the pathogenesis of testosterone-induced endothelial disfunction, which may provide novel therapeutic targets to be incorporated in the management of hypogonadism.

Dermal Stem Cells Can Differentiate Down an Endothelial Lineage

PubMed Central

Bell, Emma; Richardson, Gavin D.; Jahoda, Colin A.; Gledhill, Karl; Phillips, Helen M.; Henderson, Deborah; Owens, W. Andrew

2012-01-01

In this study, we have demonstrated that cells of neural crest origin located in the dermal papilla (DP) exhibit endothelial marker expression and a functional activity. When grown in endothelial growth media, DP primary cultures upregulate expression of vascular endothelial growth factor receptor 1 (FLT1) mRNA and downregulate expression of the dermal stem cell marker α-smooth muscle actin. DP cells have demonstrated functional characteristics of endothelial cells, including the ability to form capillary-like structures on Matrigel, increase uptake of low-density lipoprotein and upregulate ICAM1 (CD54) in response to tumour necrosis factor alpha (TNF-α) stimulation. We confirmed that these observations were not due to contaminating endothelial cells, by using DP clones. We have also used the WNT1cre/ROSA26R and WNT1cre/YFP lineage-tracing mouse models to identify a population of neural crest-derived cells in DP cultures that express the endothelial marker PECAM (CD31); these cells also form capillary-like structures on Matrigel. Importantly, cells of neural crest origin that express markers of endothelial and mesenchymal lineages exist within the dermal sheath of the vibrissae follicle. PMID:22571645

Predictive Role of Coagulation, Fibrinolytic, and Endothelial Markers in Patients with Atrial Fibrillation, Stroke, and Thromboembolism: A Meta-Analysis, Meta-Regression, and Systematic Review.

PubMed

Weymann, Alexander; Sabashnikov, Anton; Ali-Hasan-Al-Saegh, Sadeq; Popov, Aron-Frederik; Jalil Mirhosseini, Seyed; Baker, William L; Lotfaliani, Mohammadreza; Liu, Tong; Dehghan, Hamidreza; Yavuz, Senol; de Oliveira Sá, Michel Pompeu Barros; Jang, Jae-Sik; Zeriouh, Mohamed; Meng, Lei; D'Ascenzo, Fabrizio; Deshmukh, Abhishek J; Biondi-Zoccai, Guiseppe; Dohmen, Pascal M; Calkins, Hugh; Cardiac Surgery And Cardiology-Group Imcsc-Group, Integrated Meta-Analysis Of Cardiac

2017-03-31

BACKGROUND The pathophysiological mechanism associated with the higher prothrombotic tendency in atrial fibrillation (AF) is complex and multifactorial. However, the role of prothrombotic markers in AF remains inconclusive. MATERIAL AND METHODS We conducted a meta-analysis of observational studies evaluating the association of coagulation activation, fibrinolytic, and endothelial function with occurrence of AF and clinical adverse events. A comprehensive subgroup analysis and meta-regression was performed to explore potential sources of heterogeneity. RESULTS A literature search of major databases retrieved 1703 studies. After screening, a total of 71 studies were identified. Pooled analysis showed the association of coagulation markers (D-dimer (weighted mean difference (WMD) =197.67 and p<0.001), fibrinogen (WMD=0.43 and p<0.001), prothrombin fragment 1-2 (WMD=0.53 and p<0.001), antithrombin III (WMD=23.90 and p=0.004), thrombin-antithrombin (WMD=5.47 and p=0.004));  fibrinolytic markers (tissue-type plasminogen activator (t-PA) (WMD=2.13 and p<0.001), plasminogen activator inhibitor (WMD=11.44 and p<0.001), fibrinopeptide-A (WMD=4.13 and p=0.01)); and  endothelial markers (von Willebrand factor (WMD=27.01 and p<0.001) and soluble thrombomodulin (WMD=3.92 and p<0.001)) with AF. CONCLUSIONS The levels of coagulation, fibrinolytic, and endothelial markers have been reported to be significantly higher in AF patients than in SR patients.

Predictive Role of Coagulation, Fibrinolytic, and Endothelial Markers in Patients with Atrial Fibrillation, Stroke, and Thromboembolism: A Meta-Analysis, Meta-Regression, and Systematic Review

PubMed Central

Weymann, Alexander; Sabashnikov, Anton; Ali-Hasan-Al-Saegh, Sadeq; Popov, Aron-Frederik; Mirhosseini, Seyed Jalil; Baker, William L.; Lotfaliani, Mohammadreza; Liu, Tong; Dehghan, Hamidreza; Yavuz, Senol; de Oliveira Sá, Michel Pompeu Barros; Jang, Jae-Sik; Zeriouh, Mohamed; Meng, Lei; D’Ascenzo, Fabrizio; Deshmukh, Abhishek J.; Biondi-Zoccai, Giuseppe; Dohmen, Pascal M.; Calkins, Hugh

2017-01-01

Background The pathophysiological mechanism associated with the higher prothrombotic tendency in atrial fibrillation (AF) is complex and multifactorial. However, the role of prothrombotic markers in AF remains inconclusive. Material/Methods We conducted a meta-analysis of observational studies evaluating the association of coagulation activation, fibrinolytic, and endothelial function with occurrence of AF and clinical adverse events. A comprehensive subgroup analysis and meta-regression was performed to explore potential sources of heterogeneity. Results A literature search of major databases retrieved 1703 studies. After screening, a total of 71 studies were identified. Pooled analysis showed the association of coagulation markers (D-dimer (weighted mean difference (WMD)=197.67 and p<0.001), fibrinogen (WMD=0.43 and p<0.001), prothrombin fragment 1–2 (WMD=0.53 and p<0.001), antithrombin III (WMD=23.90 and p=0.004), thrombin-antithrombin (WMD=5.47 and p=0.004)); fibrinolytic markers (tissue-type plasminogen activator (t-PA) (WMD=2.13 and p<0.001), plasminogen activator inhibitor (WMD=11.44 and p<0.001), fibrinopeptide-A (WMD=4.13 and p=0.01)); and endothelial markers (von Willebrand factor (WMD=27.01 and p<0.001) and soluble thrombomodulin (WMD=3.92 and p<0.001)) with AF. Conclusions The levels of coagulation, fibrinolytic, and endothelial markers have been reported to be significantly higher in AF patients than in SR patients. PMID:28360407

Chronic exposure to biomass fuel smoke and markers of endothelial inflammation

PubMed Central

Caravedo, Maria A; Herrera, Phabiola M; Mongilardi, Nicole; de Ferrari, Aldo; Davila-Roman, Victor G; Gilman, Robert H; Wise, Robert A; Miele, Catherine H; Miranda, J Jaime; Checkley, William

2016-01-01

Indoor smoke exposure may affect cardiovascular disease (CVD) risk via lung-mediated inflammation, oxidative stress, and endothelial inflammation. We sought to explore the association between indoor smoke exposure from burning biomass fuels and a selected group of markers for endothelial inflammation. We compared serum concentrations of amyloid A protein, E-selectin, soluble ICAM-1 and VCAM-1, von Willebrand factor (VWF), and high sensitivity C-reactive protein (hs-CRP) in 228 biomass exposed vs. 228 non-exposed participants living in Puno, Peru. Average age was 56 years (SD=13), average BMI was 26.5 kg/m2 (SD=4.4), 48% were male, 59.4% completed high school and 2% reported a physician diagnosis of CVD. In unadjusted analysis, serum levels of soluble ICAM-1 (330 vs. 302 ng/mL; p<0.001), soluble VCAM-1 (403 vs. 362 ng/mL; p<0.001), and E-selectin (54.2 vs. 52.7 ng/mL; p=0.05) were increased in biomass exposed vs. non-exposed participants, respectively; whereas serum levels of vWF (1148 vs. 1311 mU/mL; p<0.001) and hs-CRP (2.56 vs. 3.12 mg/L; p<0.001) were decreased, respectively. In adjusted analyses, chronic exposure to biomass fuels remained positively associated with serum levels of soluble ICAM-1 (p=0.03) and VCAM-1 (p=0.05) and E-selectin (p=0.05), and remained negatively associated with serum levels of vWF (p=0.02) and hs-CRP (p<0.001). Daily exposure to biomass fuel smoke was associated with important differences in specific biomarkers of endothelial inflammation and may help explain accelerated atherosclerosis among those who are chronically exposed. PMID:26476302

Asymmetric Dimethylarginine as a Surrogate Marker of Endothelial Dysfunction and Cardiovascular Risk in Patients with Systemic Rheumatic Diseases

PubMed Central

Dimitroulas, Theodoros; Sandoo, Aamer; Kitas, George D.

2012-01-01

The last few decades have witnessed an increased life expectancy of patients suffering with systemic rheumatic diseases, mainly due to improved management, advanced therapies and preventative measures. However, autoimmune disorders are associated with significantly enhanced cardiovascular morbidity and mortality not fully explained by traditional cardiovascular disease (CVD) risk factors. It has been suggested that interactions between high-grade systemic inflammation and the vasculature lead to endothelial dysfunction and atherosclerosis, which may account for the excess risk for CVD events in this population. Diminished nitric oxide synthesis—due to down regulation of endothelial nitric oxide synthase—appears to play a prominent role in the imbalance between vasoactive factors, the consequent impairment of the endothelial hemostasis and the early development of atherosclerosis. Asymmetric dimethylarginine (ADMA) is one of the most potent endogenous inhibitors of the three isoforms of nitric oxide synthase and it is a newly discovered risk factor in the setting of diseases associated with endothelial dysfunction and adverse cardiovascular events. In the context of systemic inflammatory disorders there is increasing evidence that ADMA contributes to the vascular changes and to endothelial cell abnormalities, as several studies have revealed derangement of nitric oxide/ADMA pathway in different disease subsets. In this article we discuss the role of endothelial dysfunction in patients with rheumatic diseases, with a specific focus on the nitric oxide/ADMA system and we provide an overview on the literature pertaining to ADMA as a surrogate marker of subclinical vascular disease. PMID:23202900

Expression of podoplanin in Warthin tumours.

PubMed

Hansen, Torsten; Kirkpatrick, C James

2010-12-01

Warthin tumour is the second most common benign tumour of the parotid gland. This study was designed to investigate the lymphatic vessels in Warthin tumours in an effort to understand better its pathogenesis. Tissue specimens of 31 patients (19 men and 11 women; mean age 57 years, median size of the tumours 2.86 cm) were analysed by means of immunohistochemistry applying the monoclonal antibody D2-40. We found numerous D2-40-positive sinus-like vessels particularly at the inner layer of the capsule. Since subcapsular sinuses are a major morphological feature of lymph nodes in general, the finding of podoplanin expression in the large majority of subcapsular vessels in Warthin tumours confirms the view that this tumour has its origin in regional lymph nodes.

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.

The effect of diet and exercise on markers of endothelial function in overweight and obese women with polycystic ovary syndrome.

PubMed

Thomson, R L; Brinkworth, G D; Noakes, M; Clifton, P M; Norman, R J; Buckley, J D

2012-07-01

Women with polycystic ovary syndrome (PCOS) present with vascular abnormalities, including elevated markers of endothelial dysfunction. There is limited evidence for the effect of lifestyle modification and weight loss on these markers. The aim of this study was to determine if 20 weeks of a high-protein energy-restricted diet with or without exercise in women with PCOS could improve endothelial function. This is a secondary analysis of a subset of 50 overweight/obese women with PCOS (age: 30.3 ± 6.3 years; BMI: 36.5 ± 5.7 kg/m(2)) from a previous study. Participants were randomly assigned by computer generation to one of three 20-week interventions: diet only (DO; n = 14, ≈ 6000 kJ/day), diet and aerobic exercise (DA; n = 16, ≈ 6000 kJ/day and five walking sessions/week) and diet and combined aerobic-resistance exercise (DC; n = 20, ≈ 6000 kJ/day, three walking and two strength sessions/week). At Weeks 0 and 20, weight, markers of endothelial function [vascular cell adhesion molecule-1 (sVCAM-1), inter-cellular adhesion molecule-1 (sICAM-1), plasminogen activator inhibitor-1 (PAI-1) and asymmetric dimethylarginine (ADMA)], insulin resistance and hormonal profile were assessed. All three treatments resulted in significant weight loss (DO 7.9 ± 1.2%, DA 11.0 ± 1.6%, DC 8.8 ± 1.1; P < 0.001 for time; P = 0.6 time × treatment). sVCAM-1, sICAM-1 and PAI-1 levels decreased with weight loss (P≤ 0.01), with no differences between treatments (P ≥ 0.4). ADMA levels did not change significantly (P = 0.06). Testosterone, sex hormone-binding globulin and the free androgen index (FAI) and insulin resistance also improved (P < 0.001) with no differences between treatments (P ≥ 0.2). Reductions in sVCAM-1 were correlated to reductions in testosterone (r = 0.32, P = 0.03) and FAI (r = 0.33, P = 0.02) as well as weight loss (r= 0.44, P = 0.002). Weight loss was also associated with reductions in sICAM-1 (r= 0.37, P = 0.008). Exercise training provided no

Markers of endothelial and hemostatic activation and progression of cerebral white matter hyperintensities: longitudinal results of the Austrian Stroke Prevention Study.

PubMed

Markus, Hugh S; Hunt, Beverley; Palmer, Kiran; Enzinger, Christian; Schmidt, Helena; Schmidt, Reinhold

2005-07-01

The pathogenesis of cerebral small vessel disease (SVD) is poorly understood, but endothelial activation and dysfunction may play a causal role. Cross-sectional studies have found increased circulating markers of endothelial activation, but this study design cannot exclude causality from secondary elevations. Confluent white matter hyperintensities (WMHs) on magnetic resonance imaging (MRI) appear to represent asymptomatic cerebral SVD. In a prospective study, we determined whether circulating markers of endothelial activation predicted progression of WMH. In the community-based Austrian Stroke Prevention Study, MRI was performed at baseline in 296 subjects and repeated at 3 and 6 years. The following were measured on baseline plasma samples: intercellular adhesion molecule (ICAM), thrombomodulin, tissue factor plasma inhibitor, prothrombin fragments 1 and 2, and D-dimers. ICAM was associated with age- and gender-adjusted WMH lesion progression at both 3 and 6 years, respectively; (odds ratio [OR], 1.007; 95% confidence interval [CI], 1.002 to 1.012; P=0.004; and OR, 1.004; 95% CI, 1.000 to 1.009 per ng/mL; P=0.057). After multivariate analysis controlling for other cardiovascular risk factors and C-reactive protein, 3-year OR was 1.010 (95% CI, 1.004 to 1.017; P=0.001) and 6-year OR was 1.008 (1.002 to 1.014 per ng/mL; P=0.006). Baseline log lesion volume was a strong independent predictor of progression but associations remained after controlling for this (3-year OR, 1.011; 95% CI, 1.002 to 1.020; P=0.013; and 6-year OR, 1.009; 95% CI, 1.000 to 1.017; P=0.039 per ng/mL). There was no association between WMH progression and other markers. ICAM levels are related to progression of WMH on MRI. The prospective study design increases the likelihood that this association is causal and supports a role of endothelial cell activation in disease pathogenesis. In contrast, we found no evidence for coagulation activation being important.

Endothelial cell O-glycan deficiency causes blood/lymphatic misconnections and consequent fatty liver disease in mice

PubMed Central

Fu, Jianxin; Gerhardt, Holger; McDaniel, J. Michael; Xia, Baoyun; Liu, Xiaowei; Ivanciu, Lacramioara; Ny, Annelii; Hermans, Karlien; Silasi-Mansat, Robert; McGee, Samuel; Nye, Emma; Ju, Tongzhong; Ramirez, Maria I.; Carmeliet, Peter; Cummings, Richard D.; Lupu, Florea; Xia, Lijun

2008-01-01

Mucin-type O-glycans (O-glycans) are highly expressed in vascular ECs. However, it is not known whether they are important for vascular development. To investigate the roles of EC O-glycans, we generated mice lacking T-synthase, a glycosyltransferase encoded by the gene C1galt1 that is critical for the biosynthesis of core 1–derived O-glycans, in ECs and hematopoietic cells (termed here EHC T-syn–/– mice). EHC T-syn–/– mice exhibited embryonic and neonatal lethality associated with disorganized and blood-filled lymphatic vessels. Bone marrow transplantation and EC C1galt1 transgene rescue demonstrated that lymphangiogenesis specifically requires EC O-glycans, and intestinal lymphatic microvessels in EHC T-syn–/– mice expressed a mosaic of blood and lymphatic EC markers. The level of O-glycoprotein podoplanin was significantly reduced in EHC T-syn–/– lymphatics, and podoplanin-deficient mice developed blood-filled lymphatics resembling EHC T-syn–/– defects. In addition, postnatal inactivation of C1galt1 caused blood/lymphatic vessel misconnections that were similar to the vascular defects in the EHC T-syn–/– mice. One consequence of eliminating T-synthase in ECs and hematopoietic cells was that the EHC T-syn–/– pups developed fatty liver disease, because of direct chylomicron deposition via misconnected portal vein and intestinal lymphatic systems. Our studies therefore demonstrate that EC O-glycans control the separation of blood and lymphatic vessels during embryonic and postnatal development, in part by regulating podoplanin expression. PMID:18924607

Endothelial cell markers in vascular neoplasms: an immunohistochemical study comparing factor VIII-related antigen, blood group specific antigens, 6-keto-PGF1 alpha, and Ulex europaeus 1 lectin.

PubMed

Little, D; Said, J W; Siegel, R J; Fealy, M; Fishbein, M C

1986-06-01

Markers for endothelial cells including Ulex europaeus 1 lectin, blood group A, B, and H, and the prostaglandin metabolite 6-keto-PGF1 alpha were evaluated in paraffin secretions from formalin-fixed benign and malignant vascular neoplasms using a variety of immunohistochemical techniques, and results compared with staining for factor VIII-related antigen. Staining for Ulex appeared more sensitive than factor VIII-related antigen in identifying poorly differentiated neoplasms including haemangiosarcomas and spindle cell proliferations in Kaposi's sarcoma. Staining for blood group related antigens correlated with blood group in all cases. Ulex europaeus 1 lectin was the only marker for endothelial cells in lymphangiomas.

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

Clinical significance of nailfold capillaroscopy in systemic lupus erythematosus: correlation with endothelial cell activation markers and disease activity.

PubMed

Kuryliszyn-Moskal, A; Ciolkiewicz, M; Klimiuk, P A; Sierakowski, S

2009-01-01

To evaluate whether nailfold capillaroscopy (NC) changes are associated with the main serum endothelial cell activation markers and the disease activity of systemic lupus erythematosus (SLE). Serum levels of vascular endothelial growth factor (VEGF), endothelin-1 (ET-1), soluble E-selectin (sE-selectin), and soluble thrombomodulin (sTM) were determined by an enzyme-linked immunosorbent assay (ELISA) in 80 SLE patients and 33 healthy controls. Nailfold capillary abnormalities were seen in 74 out of 80 (92.5%) SLE patients. A normal capillaroscopic pattern or mild changes were found in 33 (41.25%) and moderate/severe abnormalities in 47 (58.75%) of all SLE patients. In SLE patients a capillaroscopic score >1 was more frequently associated with the presence of internal organ involvement (p < 0.001) as well as with immunosuppressive therapy (p < 0.01). Significant differences were found in VEGF (p < 0.001), ET-1 (p < 0.001), sE-selectin (p < 0.01), and sTM (p < 0.001) serum concentrations between SLE patients with a capillaroscopic score > 1 and controls. SLE patients with severe/moderate capillaroscopic abnormalities showed significantly higher VEGF serum levels than patients with mild changes (p < 0.001). Moreover, there was a significant positive correlation between the severity of capillaroscopic changes and the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) (p < 0.005) as well as between capillaroscopic score and VEGF serum levels (p < 0.001). Our findings confirm the usefulness of NC as a non-invasive technique for the evaluation of microvascular involvement in SLE patients. A relationship between changes in NC, endothelial cell activation markers and clinical features of SLE suggest an important role for microvascular abnormalities in clinical manifestation of the disease.

Blood markers of coagulation, fibrinolysis, endothelial dysfunction and inflammation in lacunar stroke versus non-lacunar stroke and non-stroke: systematic review and meta-analysis.

PubMed

Wiseman, Stewart; Marlborough, Fergal; Doubal, Fergus; Webb, David J; Wardlaw, Joanna

2014-01-01

The cause of cerebral small vessel disease is not fully understood, yet it is important, accounting for about 25% of all strokes. It also increases the risk of having another stroke and contributes to about 40% of dementias. Various processes have been implicated, including microatheroma, endothelial dysfunction and inflammation. A previous review investigated endothelial dysfunction in lacunar stroke versus mostly non-stroke controls while another looked at markers of inflammation and endothelial damage in ischaemic stroke in general. We have focused on blood markers between clinically evident lacunar stroke and other subtypes of ischaemic stroke, thereby controlling for stroke in general. We systematically assessed the literature for studies comparing blood markers of coagulation, fibrinolysis, endothelial dysfunction and inflammation in lacunar stroke versus non-stroke controls or other ischaemic stroke subtypes. We assessed the quality of included papers and meta-analysed results. We split the analysis on time of blood draw in relation to the stroke. We identified 1,468 full papers of which 42 were eligible for inclusion, including 4,816 ischaemic strokes, of which 2,196 were lacunar and 2,500 non-stroke controls. Most studies subtyped stroke using TOAST. The definition of lacunar stroke varied between studies. Markers of coagulation/fibrinolysis (tissue plasminogen activator (tPA), plasminogen activator inhibitor (PAI), fibrinogen, D-dimer) were higher in lacunar stroke versus non-stroke although fibrinogen was no different to non-stroke in the acute phase. tPA and PAI were no different between lacunar and non-lacunar stroke. Fibrinogen and D-dimer were significantly lower in lacunar stroke compared to other ischaemic strokes, both acutely and chronically. Markers of endothelial dysfunction (homocysteine, von Willebrand Factor (vWF), E-selectin, P-selectin, intercellular adhesion molecule-1 (ICAM), vascular cellular adhesion molecule-1 (VCAM)) were higher or

Increased endothelial microparticles and oxidative stress at extreme altitude.

PubMed

Pichler Hefti, Jacqueline; Leichtle, Alexander; Stutz, Monika; Hefti, Urs; Geiser, Thomas; Huber, Andreas R; Merz, Tobias M

2016-04-01

Hypoxia and oxidative stress affect endothelial function. Endothelial microparticles (MP) are established measures of endothelial dysfunction and influence vascular reactivity. To evaluate the effects of hypoxia and antioxidant supplementation on endothelial MP profiles, a double-blind, placebo-controlled trial, during a high altitude expedition was performed. 29 participants were randomly assigned to a treatment group (n = 14), receiving vitamin E, C, A, and N-acetylcysteine daily, and a control group (n = 15), receiving placebo. Blood samples were obtained at 490 m (baseline), 3530, 4590, and 6210 m. A sensitive tandem mass spectrometry method was used to measure 8-iso-prostaglandin F2α and hydroxyoctadecadienoic acids as markers of oxidative stress. Assessment of MP profiles including endothelial activation markers (CD62+MP and CD144+MP) and cell apoptosis markers (phosphatidylserine+MP and CD31+MP) was performed using a standardized flow cytometry-based protocol. 15 subjects reached all altitudes and were included in the final analysis. Oxidative stress increased significantly at altitude. No statistically significant changes were observed comparing baseline to altitude measurements of phosphatidylserine expressing MP (p = 0.1718) and CD31+MP (p = 0.1305). Compared to baseline measurements, a significant increase in CD62+MP (p = 0.0079) and of CD144+MP was detected (p = 0.0315) at high altitudes. No significant difference in any MP level or oxidative stress markers were found between the treatment and the control group. Hypobaric hypoxia is associated with increased oxidative stress and induces a significant increase in CD62+ and CD144+MP, whereas phosphatidylserine+MP and CD31+MP remain unchanged. This indicates that endothelial activation rather than an apoptosis is the primary factor of hypoxia induced endothelial dysfunction.

Prognostic value of S100/CD31 and S100/podoplanin double immunostaining in mucosal malignant melanoma of the head and neck.

PubMed

Wermker, Kai; Brauckmann, Till; Klein, Martin; Haßfeld, Stefan; Schulze, Hans-Joachim; Hallermann, Christian

2015-09-01

In uncommon mucosal melanomas of the head and neck established prognostic factors are rare and controversially discussed. The purpose of this study was to evaluate outcome and value of S100/podoplanin and S100/CD31 double immunostaining in head and neck mucosal melanomas. Retrospectively, patients with head and neck mucosal melanomas treated between 1973 and 2008 were analyzed. S100/podoplanin and S100/CD31 immunostaining were performed to detect lymph vessel invasion (LVI) and blood vessel invasion (BVI). Predictive parameters for disease-specific survival (DSS) were identified using univariate and multivariate statistics. Forty-two patients with head and neck mucosal melanoma were included. Three-year, 5-year, and 10-year DSS rates were 59%, 44%, and 20%, respectively. Age above 70 years, occurrence of distant metastasis, LVI, and BVI were significantly associated with shorter DSS time (p < .05), whereas localization at the conjunctiva showed better outcome. S100/podoplanin and S100/CD31 double immunostaining detect reliable LVI and BVI in head and neck mucosal melanoma and both are associated significantly with worse prognosis. © 2014 Wiley Periodicals, Inc.

Endocan--the new endothelial activation marker independently associated with soluble endothelial adhesion molecules in uraemic patients with cardiovascular disease.

PubMed

Pawlak, Krystyna; Mysliwiec, Michal; Pawlak, Dariusz

2015-04-01

Endocan is a new marker of endothelial cell activation that mediates adhesion of leukocytes into endothelium. Soluble intercellular (sICAM-1) and vascular cellular (sVCAM-1) adhesion molecules play an important role in the prevalence of cardiovascular disease (CVD) in chronic kidney disease (CKD) patients. The aim of this study is to investigate whether endocan could affect the concentrations of sICAM-1 and sVCAM-1 in CKD patients, particularly in those with CVD. We evaluated plasma endocan, sICAM-1, sVCAM-1 and the markers of inflammation: high sensitivity C-reactive protein (hs CRP), interleukin-6, tumor necrosis factor-α (TNF-α) and their interrelationships in 53 CKD patients (both with and without CVD) and 29 healthy controls. Endocan, sICAM-1, sVCAM-1 and inflammatory markers were significantly higher in CKD patients than in controls, and patients with CVD had levels significantly higher (except interleukin-6 and TNF-α) than those without CVD. The presence of CVD, ferritin, TNF-α and SBP were the independent predictors of endocan levels in the whole CKD group. In this group, the weak relationship was between endocan and sICAM-1 and sVCAM-1, but age was the only independent predictor of these adhesion molecules. The strong association between endocan and sICAM-1 and sVCAM-1 was exclusively observed in subgroup with CVD, and the low % of lymphocytes followed by increased endocan was identified as the independent variables significantly associated with these soluble molecule levels. This study shows that plasma endocan is significantly increased and independently associated with sICAM-1 and sVCAM-1 levels in CKD patients with cardiovascular complications. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Evaluation of the prognostic role of tumor cell podoplanin expression in locally advanced squamous cell carcinoma of the head and neck.

PubMed

Huttenlocher, Stefan; Seibold, Nina D; Gebhard, Maximilian P; Noack, Frank; Thorns, Christoph; Hasselbacher, Katrin; Wollenberg, Barbara; Schild, Steven E; Rades, Dirk

2014-10-01

To investigate the potential prognostic role of tumor cell podoplanin expression in patients treated with resection followed by irradiation or chemoradiotherapy for locally advanced squamous cell carcinoma of the head and neck (SCCHN). Podoplanin expression (≤10 % versus > 10 %) and 12 other factors were evaluated in 160 patients for their association with locoregional control (LRC), metastases-free (MFS) and overall survival (OS). Other factors were age, gender, Eastern Cooperative Oncology Group (ECOG) performance status, preradiotherapy (pre-RT) hemoglobin level, tumor site, histological grading, T category, N category, American Joint Committee on Cancer (AJCC) stage, human papillomavirus (HPV) status, extent of resection and concurrent chemotherapy. In multivariate analysis, ECOG performance status 0-1 (risk ratio, RR: 3.01; 95 % confidence interval, CI: 1.42-7.14; p = 0.003), pre-RT hemoglobin levels ≥ 7.45 mmol/l (12 g/dl; RR: 2.03; 95 % CI: 1.04-3.94; p = 0.038), oropharyngeal cancer (RR: 1.25; 95 % CI: 1.01-1.55; p = 0.038) and T category T1-2 (RR: 1.81; 95 % CI: 1.24-2.79; p = 0.002) were significantly associated with improved LRC. T category T1-2 (RR: 1.90; 95 % CI: 1.25-3.06; p = 0.002) and N category N0-2a (RR: 5.22; 95 % CI: 1.96-18.09; p < 0.001) were significantly associated with better MFS. Pre-RT hemoglobin levels ≥ 7.45 mmol/l (RR: 2.44; 95 % CI: 1.27-4.74; p = 0.007), T category T1-2 (RR: 1.97; 95 % CI: 1.36-3.04; p < 0.001) and N category N0-2a (RR: 2.87; 95 % CI: 1.37-6.61; p = 0.005) were significantly associated with improved OS. Podoplanin expression ≤ 10 % showed a trend towards improved OS on both univariate (p = 0.050) and multivariate analysis (RR: 1.86; 95 % CI: 0.96-3.59; p = 0.07). Treatment outcomes were significantly associated with performance status, pre-RT hemoglobin level, tumor site and tumor stage. Tumor cell expression of podoplanin ≤ 10�

Hematopoietic stem cell loss and hematopoietic failure in severe aplastic anemia is driven by macrophages and aberrant podoplanin expression.

PubMed

McCabe, Amanda; Smith, Julianne N P; Costello, Angelica; Maloney, Jackson; Katikaneni, Divya; MacNamara, Katherine C

2018-05-17

Severe aplastic anemia results from profound hematopoietic stem cell loss. T cells and interferon gamma have long been associated with severe aplastic anemia, yet the underlying mechanisms driving hematopoietic stem cell loss remain unknown. Using a mouse model of severe aplastic anemia, we demonstrate that interferon gamma-dependent hematopoietic stem cell loss required macrophages. Interferon gamma was necessary for bone marrow macrophage persistence, despite loss of other myeloid cells and hematopoietic stem cells. Depleting macrophages or abrogating interferon gamma signaling specifically in macrophages did not impair T cell activation or interferon gamma production in the bone marrow but rescued hematopoietic stem cells and reduced mortality. Thus, macrophages are not required for induction of interferon gamma in severe aplastic anemia and rather act as sensors of interferon gamma. Macrophage depletion rescued thrombocytopenia, increased bone marrow megakaryocytes, preserved platelet-primed stem cells, and increased the platelet-repopulating capacity of transplanted hematopoietic stem cells. In addition to the hematopoietic effects, severe aplastic anemia induced loss of non-hematopoietic stromal populations, including podoplanin-positive stromal cells. However, a subset of podoplanin-positive macrophages was increased during disease, and blockade of podoplanin in mice was sufficient to rescue disease. Our data further our understanding of disease pathogenesis demonstrating a novel role for macrophages as sensors of interferon gamma, thus illustrating an important role for the microenvironment in pathogenesis of severe aplastic anemia. Copyright © 2018, Ferrata Storti Foundation.

Effect of teriparatide treatment on endothelial function, glucose metabolism and inflammation markers in patients with postmenopausal osteoporosis.

PubMed

Celer, Ozgen; Akalın, Aysen; Oztunali, Cigdem

2016-10-01

Teriparatide, an anabolic agent used in the treatment of postmenopausal osteoporosis, can induce effects similar to primary hyperparathyroidism. Our objective was to evaluate the effects of teriparatide on endothelial functions, glucose metabolism and inflammation markers in patients diagnosed with postmenopausal osteoporosis. This was a single-centre, single-arm, 6-month prospective study. Twenty-three postmenopausal women over 65 years old with a lumbar spine or femoral neck T-score of -4·0 or lower and having at least two compression fractures in thoracic or lumbar spine were studied. Low-dose intermittent teriparatide (20 μg/day) was supplemented with calcium carbonate (1000 mg elemental calcium) and 880 IU cholecalciferol for 6 months. The biochemical parameters for glucose metabolism, inflammation and atherosclerosis were determined. For the assessment of vascular endothelial function, carotid intima-media thickness (CIMT), brachial artery intima-media thickness (BIMT), per cent change in flow-mediated dilation (FMD%) and nitroglycerine-induced dilations (NID%) were measured on ultrasonography. The fasting plasma glucose, homoeostatic model assessment of insulin resistance, fibrinogen, homocysteine and high-density lipoprotein cholesterol increased significantly with teriparatide treatment (P < 0·05 for all). Baseline CIMT and BIMT did not change significantly with 6 months of teriparatide treatment (P > 0·05); however, FMD% and NID% showed significant decrease after treatment (P < 0·01 for both). Intermittent teriparatide treatment may adversely affect some parameters of glucose metabolism, inflammation and endothelial function. On the basis of our findings, further large-scale and controlled studies are needed to clarify the exact effect of teriparatide treatment on glucose metabolism, inflammation and endothelial function. © 2016 John Wiley & Sons Ltd.

Triglycerides as an early pathophysiological marker of endothelial dysfunction in nondiabetic women with a previous history of gestational diabetes.

PubMed

Sokup, Alina; Góralczyk, Barbara; Góralczyk, Krzysztof; Rość, Danuta

2012-02-01

To investigate whether baseline triglyceride levels are associated with early glucose dysregulation and/or cardiovascular risk in women with a previous history of gestational diabetes. Prospective postpregnancy cohort study. Polish university hospitals. Participants included 125 women with previous gestational diabetes and 40 women with normal glucose regulation during pregnancy. All women were studied 2-24 months (mean 12 ± 10 months) after the index pregnancy. Women with previous gestational diabetes were divided into tertiles in accordance with baseline triglyceride levels. We assessed glucose regulation (oral glucose tolerance test), insulin resistance (homeostasis model assessment), markers of endothelial dysfunction (soluble: intercellular adhesion molecule-1, vascular cell adhesion molecule-1, E-selectin, tissue plasminogen activator antigen, von Willebrand factor antigen), fibrinolysis (plasminogen activator inhibitor antigen), inflammation (high-sensitivity C-reactive protein) and lipid levels. Women with previous gestational diabetes (78% normal glucose regulation, 22% impaired glucose tolerance) had a high cardiometabolic risk profile compared with control women (100% normal glucose regulation). Baseline triglycerides >0.83 mmol/l were associated with a higher prevalence of impaired glucose tolerance, higher high-sensitivity C-reactive protein and triglyceride/high-density lipoprotein-cholesterol ratio. Triglycerides >1.22 mmol/l were associated with higher body fat indexes, higher insulin resistance, higher levels of endothelial dysfunction biomarkers, higher plasminogen activator inhibitor antigen and dyslipidemia. Only E-selectin was independently associated with triglyceride levels. Baseline triglyceride levels are a cardiovascular risk marker as well as a pathophysiological parameter independently associated with endothelial dysfunction in nondiabetic women with previous gestational diabetes at 2-24 months after an index pregnancy. Normalization of

Endothelial progenitor cells in chronic obstructive pulmonary disease and emphysema

PubMed Central

Tracy, Russell P.; Parikh, Megha A.; Hoffman, Eric A.; Shimbo, Daichi; Austin, John H. M.; Smith, Benjamin M.; Hueper, Katja; Vogel-Claussen, Jens; Lima, Joao; Gomes, Antoinette; Watson, Karol; Kawut, Steven; Barr, R. Graham

2017-01-01

Endothelial injury is implicated in the pathogenesis of COPD and emphysema; however the role of endothelial progenitor cells (EPCs), a marker of endothelial cell repair, and circulating endothelial cells (CECs), a marker of endothelial cell injury, in COPD and its subphenotypes is unresolved. We hypothesized that endothelial progenitor cell populations would be decreased in COPD and emphysema and that circulating endothelial cells would be increased. Associations with other subphenotypes were examined. The Multi-Ethnic Study of Atherosclerosis COPD Study recruited smokers with COPD and controls age 50–79 years without clinical cardiovascular disease. Endothelial progenitor cell populations (CD34+KDR+ and CD34+KDR+CD133+ cells) and circulating endothelial cells (CD45dimCD31+CD146+CD133-) were measured by flow cytometry. COPD was defined by standard spirometric criteria. Emphysema was assessed qualitatively and quantitatively on CT. Full pulmonary function testing and expiratory CTs were measured in a subset. Among 257 participants, both endothelial progenitor cell populations, and particularly CD34+KDR+ endothelial progenitor cells, were reduced in COPD. The CD34+KDR+CD133+ endothelial progenitor cells were associated inversely with emphysema extent. Both endothelial progenitor cell populations were associated inversely with extent of panlobular emphysema and positively with diffusing capacity. Circulating endothelial cells were not significantly altered in COPD but were inversely associated with pulmonary microvascular blood flow on MRI. There was no consistent association of endothelial progenitor cells or circulating endothelial cells with measures of gas trapping. These data provide evidence that endothelial repair is impaired in COPD and suggest that this pathological process is specific to emphysema. PMID:28291826

Mesenchymal Stem/Multipotent Stromal Cells from Human Decidua Basalis Reduce Endothelial Cell Activation.

PubMed

Alshabibi, Manal A; Al Huqail, Al Joharah; Khatlani, Tanvir; Abomaray, Fawaz M; Alaskar, Ahmed S; Alawad, Abdullah O; Kalionis, Bill; Abumaree, Mohamed Hassan

2017-09-15

Recently, we reported the isolation and characterization of mesenchymal stem cells from the decidua basalis of human placenta (DBMSCs). These cells express a unique combination of molecules involved in many important cellular functions, which make them good candidates for cell-based therapies. The endothelium is a highly specialized, metabolically active interface between blood and the underlying tissues. Inflammatory factors stimulate the endothelium to undergo a change to a proinflammatory and procoagulant state (ie, endothelial cell activation). An initial response to endothelial cell activation is monocyte adhesion. Activation typically involves increased proliferation and enhanced expression of adhesion and inflammatory markers by endothelial cells. Sustained endothelial cell activation leads to a type of damage to the body associated with inflammatory diseases, such as atherosclerosis. In this study, we examined the ability of DBMSCs to protect endothelial cells from activation through monocyte adhesion, by modulating endothelial proliferation, migration, adhesion, and inflammatory marker expression. Endothelial cells were cocultured with DBMSCs, monocytes, monocyte-pretreated with DBMSCs and DBMSC-pretreated with monocytes were also evaluated. Monocyte adhesion to endothelial cells was examined following treatment with DBMSCs. Expression of endothelial cell adhesion and inflammatory markers was also analyzed. The interaction between DBMSCs and monocytes reduced endothelial cell proliferation and monocyte adhesion to endothelial cells. In contrast, endothelial cell migration increased in response to DBMSCs and monocytes. Endothelial cell expression of adhesion and inflammatory molecules was reduced by DBMSCs and DBMSC-pretreated with monocytes. The mechanism of reduced endothelial proliferation involved enhanced phosphorylation of the tumor suppressor protein p53. Our study shows for the first time that DBMSCs protect endothelial cells from activation by

Effect of a prolonged endurance marathon on vascular endothelial and inflammation markers in runners with exercise-induced hypertension.

PubMed

Jee, Haemi; Park, Jaehyun; Oh, Jae-Gun; Lee, Yoon-Hee; Shin, Kyung-A; Kim, Young-Joo

2013-06-01

The aim of this study was to observe the changes in endothelial and inflammatory markers in middle-aged male runners with exercise-induced hypertension (EIH) at baseline and at 100-km, 200-km, and 308-km checkpoints during a prolonged endurance ultramarathon. Among a total of 62 ultramarathon volunteers, 8 with systolic blood pressure higher than 210 mm Hg and 8 with normal systolic blood pressure were selected for this study. The subjects were designated to EIH and control (CON) groups. Blood was collected for the analysis of soluble vascular cell adhesion molecule-1, soluble E-selectin, leukocytes, creatine kinase, and high-sensitivity C-reactive protein. Soluble vascular cell adhesion molecule-1 showed a significantly greater increase in the EIH group than in the CON group at 100 km and 200 km. Soluble E-selectin also showed a significantly greater increase in the EIH group than in the CON group at 100 km. Leukocytes significantly increased in the EIH group than in the CON group at 308 km. Creatine kinase and high-sensitivity C-reactive protein showed no group differences. Leukocytes, creatine kinase, and high-sensitivity C-reactive protein showed delayed-onset increases in both groups. Increased exercise intensity may stimulate greater endothelial responses independent of the inflammatory markers in EIH. The loss of a protective effect may be greater in those with EIH than in CONs. Acknowledging and prescribing proper exercise intensity may be critical in preventing possible vascular-related complications in runners with EIH.

Association between markers of endothelial dysfunction and early signs of renal dysfunction in pediatric obesity and type 1 diabetes.

PubMed

Marcovecchio, M L; de Giorgis, T; Di Giovanni, I; Chiavaroli, V; Chiarelli, F; Mohn, A

2017-06-01

To evaluate whether circulating markers of endothelial dysfunction, such as intercellular adhesion molecule-1 (ICAM-1) and myeloperoxidase (MPO), are increased in youth with obesity and in those with type 1 diabetes (T1D) at similar levels, and whether their levels are associated with markers of renal function. A total of 60 obese youth [M/F: 30/30, age: 12.5 ± 2.8 yr; body mass index (BMI) z-score: 2.26 ± 0.46], 30 with T1D (M/F: 15/15; age: 12.9 ± 2.4 yr; BMI z-score: 0.45 ± 0.77), and 30 healthy controls (M/F: 15/15, age: 12.4 ± 3.3 yr, BMI z-score: -0.25 ± 0.56) were recruited. Anthropometric measurements were assessed and a blood sample was collected to measure ICAM-1, MPO, creatinine, cystatin C and lipid levels. A 24-h urine collection was obtained for assessing albumin excretion rate (AER). Levels of ICAM-1 and MPO were significantly higher in obese [ICAM-1: 0.606 (0.460-1.033) µg/mL; MPO: 136.6 (69.7-220.8) ng/mL] and T1D children [ICAM-1: 0.729 (0.507-0.990) µg/mL; MPO: 139.5 (51.0-321.3) ng/mL] compared with control children [ICAM-1: 0.395 (0.272-0.596) µg/mL MPO: 41.3 (39.7-106.9) ng/mL], whereas no significant difference was found between T1D and obese children. BMI z-score was significantly associated with ICAM-1 (β = 0.21, p = 0.02) and MPO (β = 0.41, p < 0.001). A statistically significant association was also found between ICAM-1 and markers of renal function (AER: β = 0.21, p = 0.03; e-GFR: β = 0.19, p = 0.04), after adjusting for BMI. Obese children have increased markers of endothelial dysfunction and early signs of renal damage, similarly to children with T1D, confirming obesity to be a cardiovascular risk factor as T1D. The association between ICAM-1 with e-GFR and AER confirm the known the association between general endothelial and renal dysfunction. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Central Role of eNOS in the Maintenance of Endothelial Homeostasis

PubMed Central

Rodriguez-Mateos, Ana; Kelm, Malte

2015-01-01

Abstract Significance: Disruption of endothelial function is considered a key event in the development and progression of atherosclerosis. Endothelial nitric oxide synthase (eNOS) is a central regulator of cellular function that is important to maintain endothelial homeostasis. Recent Advances: Endothelial homeostasis encompasses acute responses such as adaption of flow to tissue's demand and more sustained responses to injury such as re-endothelialization and sprouting of endothelial cells (ECs) and attraction of circulating angiogenic cells (CAC), both of which support repair of damaged endothelium. The balance and the intensity of endothelial damage and repair might be reflected by changes in circulating endothelial microparticles (EMP) and CAC. Flow-mediated vasodilation (FMD) is a generally accepted clinical read-out of NO-dependent vasodilation, whereas EMP are upcoming prognostically validated markers of endothelial injury and CAC are reflective of the regenerative capacity with both expressing a functional eNOS. These markers can be integrated in a clinical endothelial phenotype, reflecting the net result between damage from risk factors and endogenous repair capacity with NO representing a central signaling molecule. Critical Issues: Improvements of reproducibility and observer independence of FMD measurements and definitions of relevant EMP and CAC subpopulations warrant further research. Future Directions: Endothelial homeostasis may be a clinical therapeutic target for cardiovascular health maintenance. Antioxid. Redox Signal. 22, 1230–1242. PMID:25330054

Compressive elasticity of three-dimensional nanofiber matrix directs mesenchymal stem cell differentiation to vascular cells with endothelial or smooth muscle cell markers.

PubMed

Wingate, K; Bonani, W; Tan, Y; Bryant, S J; Tan, W

2012-04-01

The importance of mesenchymal stem cells (MSC) in vascular regeneration is becoming increasingly recognized. However, few in vitro studies have been performed to identify the effects of environmental elasticity on the differentiation of MSC into vascular cell types. Electrospinning and photopolymerization techniques were used to fabricate a three-dimensional (3-D) polyethylene glycol dimethacrylate nanofiber hydrogel matrix with tunable elasticity for use as a cellular substrate. Compression testing demonstrated that the elastic modulus of the hydrated 3-D matrices ranged from 2 to 15 kPa, similar to the in vivo elasticity of the intima basement membrane and media layer. MSC seeded on rigid matrices (8-15 kPa) showed an increase in cell area compared with those seeded on soft matrices (2-5 kPa). Furthermore, the matrix elasticity guided the cells to express different vascular-specific phenotypes with high differentiation efficiency. Around 95% of MSC seeded on the 3-D matrices with an elasticity of 3 kPa showed Flk-1 endothelial markers within 24h, while only 20% of MSC seeded on the matrices with elasticity >8 kPa demonstrated Flk-1 marker. In contrast, ∼80% of MSC seeded on 3-D matrices with elasticity >8 kPa demonstrated smooth muscle α-actin marker within 24h, while fewer than 10% of MSC seeded on 3-D matrices with elasticity <5 kPa showed α-actin markers. The ability to control MSC differentiation into either endothelial or smooth muscle-like cells based purely on the local elasticity of the substrate could be a powerful tool for vascular tissue regeneration. Copyright © 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

The role of shear stress and altered tissue properties on endothelial to mesenchymal transformation and tumor-endothelial cell interaction.

PubMed

Mina, Sara G; Huang, Peter; Murray, Bruce T; Mahler, Gretchen J

2017-07-01

Tumor development is influenced by stromal cells in aspects including invasion, growth, angiogenesis, and metastasis. Activated fibroblasts are one group of stromal cells involved in cancer metastasis, and one source of activated fibroblasts is endothelial to mesenchymal transformation (EndMT). EndMT begins when the endothelial cells delaminate from the cell monolayer, lose cell-cell contacts, lose endothelial markers such as vascular endothelial-cadherin (VE-cadherin), gain mesenchymal markers like alpha-smooth muscle actin (α-SMA), and acquire mesenchymal cell-like properties. A three-dimensional (3D) culture microfluidic device was developed for investigating the role of steady low shear stress (1 dyne/cm 2 ) and altered extracellular matrix (ECM) composition and stiffness on EndMT. Shear stresses resulting from fluid flow within tumor tissue are relevant to both cancer metastasis and treatment effectiveness. Low and oscillatory shear stress rates have been shown to enhance the invasion of metastatic cancer cells through specific changes in actin and tubulin remodeling. The 3D ECM within the device was composed of type I collagen and glycosaminoglycans (GAGs), hyaluronic acid and chondroitin sulfate. An increase in collagen and GAGs has been observed in the solid tumor microenvironment and has been correlated with poor prognosis in many different cancer types. In this study, it was found that ECM composition and low shear stress upregulated EndMT, including upregulation of mesenchymal-like markers (α-SMA and Snail) and downregulated endothelial marker protein and gene expression (VE-cadherin). Furthermore, this novel model was utilized to investigate the role of EndMT in breast cancer cell proliferation and migration. Cancer cell spheroids were embedded within the 3D ECM of the microfluidic device. The results using this device show for the first time that the breast cancer spheroid size is dependent on shear stress and that the cancer cell migration rate

Endothelial markers in malignant vascular tumours of the liver: superiority of QB-END/10 over von Willebrand factor and Ulex europaeus agglutinin 1.

PubMed

Anthony, P P; Ramani, P

1991-01-01

A new monoclonal antibody, QB-END/10, raised against the CD34 antigen in human endothelial cell membranes and haemopoietic progenitor cells, was studied for its usefulness as a marker of neoplastic vascular cells in 21 angiosarcomas and seven malignant haemangioendotheliomas of the liver. QB-END/10 was both more sensitive and more specific than Von Willebrand factor (VWF) and Ulex europaeus 1 agglutinin (UEA-1) in labelling endothelial cells and it did not cross react with epithelia as UEA-1 often does. Staining was uniformly strong and clear in all histological variants of these two tumours. QB-END/10 should prove particularly useful in the differential diagnosis of malignant vascular tumours of the liver.

Endothelial markers in malignant vascular tumours of the liver: superiority of QB-END/10 over von Willebrand factor and Ulex europaeus agglutinin 1.

PubMed Central

Anthony, P P; Ramani, P

1991-01-01

A new monoclonal antibody, QB-END/10, raised against the CD34 antigen in human endothelial cell membranes and haemopoietic progenitor cells, was studied for its usefulness as a marker of neoplastic vascular cells in 21 angiosarcomas and seven malignant haemangioendotheliomas of the liver. QB-END/10 was both more sensitive and more specific than Von Willebrand factor (VWF) and Ulex europaeus 1 agglutinin (UEA-1) in labelling endothelial cells and it did not cross react with epithelia as UEA-1 often does. Staining was uniformly strong and clear in all histological variants of these two tumours. QB-END/10 should prove particularly useful in the differential diagnosis of malignant vascular tumours of the liver. Images PMID:1705261

Intensity and Pattern of Enhancement on CESM: Prognostic Significance and its Relation to Expression of Podoplanin in Tumor Stroma - A Preliminary Report.

PubMed

Luczynska, Elzbieta; Niemiec, Joanna; Heinze, Sylwia; Adamczyk, Agnieszka; Ambicka, Aleksandra; Marcyniuk, Paulina; Rudnicki, Wojciech; Mitus, Jerzy W; Dyczek, Sonia; Rys, Janusz; Sas-Korczynska, Beata

2018-02-01

It is possible that the degree of enhancement on contrast-enhanced spectral mammography (CESM), a new diagnostic method, might provide prognostic information for breast cancer patients. Therefore, in a group of 82 breast cancer patients, we analyzed the prognostic significance of degree and pattern of enhancement on CESM as well as its relation to: (a) breast cancer immunophenotype (based on ER/PR/HER2 status) (b) podoplanin expression in cancer stroma (lymphatic vessel density plus podoplanin-positivity of cancer-associated fibroblasts), and (c) other histological parameters. For each tumor the intensity of enhancement on CESM was qualitatively assessed as strong or weak/medium, while the pattern - as homogenous and heterogenous. Herein we report, for the first time, that strong and heterogenous enhancement on CESM was related to unfavorable disease-free survival of breast cancer patients (p=0.005). Moreover, the strong enhancement was more frequent in large and node-positive tumors (pT>1, pN>0) (p=0.002), as well as in carcinomas with podoplanin-sparse stroma (p=0.008). Intensity and pattern of enhancement on CESM might provide (together with the results of other diagnostic imaging methods) not only the confirmation of presence or absence of tumor, but also prognostic information. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Recombinant anti-podoplanin (NZ-1) immunotoxin for the treatment of malignant brain tumors

PubMed Central

Chandramohan, Vidyalakshmi; Bao, Xuhui; Kaneko, Mika Kato; Kato, Yukinari; Keir, Stephen T.; Szafranski, Scott E.; Kuan, Chien-Tsun; Pastan, Ira H.; Bigner, Darell D.

2013-01-01

Current study demonstrates the glioma tumor antigen podoplanin to be present at very high levels (>90%) in both glioblastoma (D2159MG, D08-0308MG, and D08-0493MG) and medulloblastoma (D283MED, D425MED, and DAOY) xenografts and cell line. We constructed a novel recombinant single-chain antibody variable region fragment (scFv), NZ-1, specific for podoplanin from the NZ-1 hybridoma. NZ-1-scFv was then fused to Pseudomonas exotoxin A, carrying a C-terminal KDEL peptide (NZ-1-PE38KDEL). The immunotoxin was further stabilized by a disulfide (ds) bond between the heavy-chain and light-chain variable regions as the construct NZ-1-(scdsFv)-PE38KDEL. NZ-1-(scdsFv)-PE38KDEL exhibited significant reactivity to glioblastoma and medulloblastoma cells. The affinity of NZ-1-(scdsFv), NZ-1-(scdsFv)-PE38KDEL and NZ-1 antibody, for podoplanin peptide was 2.1×10−8 M, 8.0×10−8 M, and 3.9×10−10 M, respectively. In a protein stability assay, NZ-1-(scdsFv)-PE38KDEL retained 33-98% of its activity while that of NZ-1-PE38KDEL declined to 13% of its initial levels after incubation at 37°C for 3 days. In vitro cytotoxicity of the NZ-1-(scdsFv)-PE38KDEL was measured in cells isolated from glioblastoma xenografts, D2159MG, D08-0308MG, D08-0493MG, and in the medulloblastoma D283MED, D425MED, and DOAY xenografts and cell line. The NZ-1-(scdsFv)-PE38KDEL immunotoxin was highly cytotoxic, with an IC50 in the range of 1.6–29 ng/mL. Significantly, NZ-1-(scdsFv)-PE38KDEL demonstrated tumor-growth delay, averaging 24 days (P<0.001) and 21 days (P<0.001) in D2159MG and D283MED in vivo tumor models, respectively. Crucially, in the D425MED intracranial tumor model, NZ-1-(scdsFv)-PE38KDEL caused a 41% increase in survival (P≤0.001). In preclinical studies, NZ-1-(scdsFv)-PE38KDEL exhibited significant potential as a targeting agent for malignant brain tumors. PMID:23115013

Recombinant anti-podoplanin (NZ-1) immunotoxin for the treatment of malignant brain tumors.

PubMed

Chandramohan, Vidyalakshmi; Bao, Xuhui; Kato Kaneko, Mika; Kato, Yukinari; Keir, Stephen T; Szafranski, Scott E; Kuan, Chien-Tsun; Pastan, Ira H; Bigner, Darell D

2013-05-15

Our study demonstrates the glioma tumor antigen podoplanin to be present at very high levels (>90%) in both glioblastoma (D2159MG, D08-0308MG and D08-0493MG) and medulloblastoma (D283MED, D425MED and DAOY) xenografts and cell line. We constructed a novel recombinant single-chain antibody variable region fragment (scFv), NZ-1, specific for podoplanin from the NZ-1 hybridoma. NZ-1-scFv was then fused to Pseudomonas exotoxin A, carrying a C-terminal KDEL peptide (NZ-1-PE38KDEL). The immunotoxin (IT) was further stabilized by a disulfide (ds) bond between the heavy-chain and light-chain variable regions as the construct NZ-1-(scdsFv)-PE38KDEL. NZ-1-(scdsFv)-PE38KDEL exhibited significant reactivity to glioblastoma and medulloblastoma cells. The affinity of NZ-1-(scdsFv), NZ-1-(scdsFv)-PE38KDEL and NZ-1 antibody for podoplanin peptide was 2.1 × 10(-8) M, 8.0 × 10(-8) M and 3.9 × 10(-10) M, respectively. In a protein stability assay, NZ-1-(scdsFv)-PE38KDEL retained 33-98% of its activity, whereas that of NZ-1-PE38KDEL declined to 13% of its initial levels after incubation at 37°C for 3 days. In vitro cytotoxicity of the NZ-1-(scdsFv)-PE38KDEL was measured in cells isolated from glioblastoma xenografts, D2159MG, D08-0308MG and D08-0493MG, and in the medulloblastoma D283MED, D425MED and DOAY xenografts and cell line. The NZ-1-(scdsFv)-PE38KDEL IT was highly cytotoxic, with an 50% inhibitory concentration in the range of 1.6-29 ng/ml. Significantly, NZ-1-(scdsFv)-PE38KDEL demonstrated tumor growth delay, averaging 24 days (p < 0.001) and 21 days (p < 0.001) in D2159MG and D283MED in vivo tumor models, respectively. Crucially, in the D425MED intracranial tumor model, NZ-1-(scdsFv)-PE38KDEL caused a 41% increase in survival (p ≤ 0.001). In preclinical studies, NZ-1-(scdsFv)-PE38KDEL exhibited significant potential as a targeting agent for malignant brain tumors. Copyright © 2012 UICC.

Nitric oxide releasing hydrogel promotes endothelial differentiation of mouse embryonic stem cells.

PubMed

Nie, Yan; Zhang, Kaiyue; Zhang, Shuaiqiang; Wang, Dan; Han, Zhibo; Che, Yongzhe; Kong, Deling; Zhao, Qiang; Han, Zhongchao; He, Zuo-Xiang; Liu, Na; Ma, Fengxia; Li, Zongjin

2017-11-01

Transplantation of endothelial cells (ECs) holds great promise for treating various kinds of ischemic diseases. However, the major challenge in ECs-based therapy in clinical applications is to provide high quality and enough amounts of cells. In this study, we developed a simple and efficient system to direct endothelial differentiation of mouse embryonic stem cells (ESCs) using a controllable chitosan nitric oxide (NO)-releasing hydrogel (CS-NO). ESCs were plated onto the hydrogel culture system, and the expressions of differentiation markers were measured. We found that the expression of Flk-1 (early ECs marker) and VE-cadherin (mature ECs marker) increased obviously under the controlled NO releasing environment. Moreover, the Flk-1 upregulation was accompanied by the activation of the phospho-inositide-3 kinase (PI3K)/Akt signaling. We also found that in the presence of the PI3K inhibitor (LY294002), the endothelial commitment of ESCs was abolished, indicating the importance of Akt phosphorylation in the endothelial differentiation of ESCs. Interestingly, in the absence of NO, the activation of Akt phosphorylation alone by using AKT activator (SC-79) did not profoundly promote the endothelial differentiation of ESCs, suggesting an interdependent relationship between NO and the Akt phosphorylation in driving endothelial fate specification of ESCs. Taken together, we demonstrated that NO releasing in a continuous and controlled manner is a simple and efficient method for directing the endothelial differentiation of ESCs without adding growth factors. Fascinating data continues to show that artificial stem cell niche not only serve as a physical supporting scaffold for stem cells proliferation, but also as a novel platform for directing stem cell differentiation. Because of the lack of proper microenvironment for generating therapeutic endothelial cells (ECs) in vitro, the source of ECs for transplantation is the major limitation in ECs-based therapy to clinical

Isolation and Characterization of Rat Pituitary Endothelial Cells

PubMed Central

Chaturvedi, Kirti; Sarkar, Dipak K.

2010-01-01

Most previous studies that determined the effect of estradiol on angiogenesis used endothelial cells from nonpituitary sources. Because pituitary tumor tissue receives its blood supply via portal and arterial circulation, it is important to use pituitary-derived endothelial cells in studying pituitary angiogenesis. We have developed a magnetic separation technique to isolate endothelial cells from pituitary tissues and have characterized these cells in primary cultures. Endothelial cells of the pituitary showed the existence of endothelial cell marker, CD31, and of von Willebrand factor protein. These cells in cultures also showed immunore-activity of estrogen receptors alpha and beta. The angiogenic factors, vascular endothelial growth factor and basic fibroblast growth factor, significantly increased proliferation and migration of the pituitary-derived endothelial cells in primary cultures. These results suggest that a magnetic separation technique can be used for enrichment of pituitary-derived endothelial cells for determination of cellular mechanisms governing the vascularization in the pituitary. PMID:17028416

Thrombomodulin, von Willebrand factor and E-selectin as plasma markers of endothelial damage/dysfunction and activation in pregnancy induced hypertension.

PubMed

Nadar, Sunil K; Al Yemeni, Eman; Blann, Andrew D; Lip, Gregory Y H

2004-01-01

Endothelial disturbance (whether activation, dysfunction or damage) is a likely pathogenic mechanism in pre-eclampsia and pregnancy-induced hypertension (PIH). We set out to determine which of three plasma markers of endothelial disturbance, indicating endothelial activation (E-selectin) or damage/dysfunction (von Willebrand factor (vWf), soluble thrombomodulin), would provide the best discriminator of PIH compared to normotensive pregnancy. Cross-sectional study of 36 consecutive women with PIH (age 31+/-6 years) and 36 consecutive women with normotensive pregnancies (age 29+/-5 years) of similar parity. Plasma levels of vWf, E-selectin and thrombomodulin were measured using ELISA. As expected, women with PIH had significantly higher levels of plasma vWf (by 19%, p=0.003), E-selectin (by 40%, p<0.001) and thrombomodulin (by 61%, p=0.01) than normotensive women. However, on stepwise multiple regression analysis, only thrombomodulin was an independent significant predictor of the presence of PIH (p=0.023). We conclude that although vWf, E-selectin and thrombomodulin are all raised in PIH, only thrombomodulin was independently associated with PIH. This molecule could potentially be useful in monitoring and in providing clues in aetiology and pathophysiology, and may have implications for the clinical complications associated with PIH.

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.

Debulking surgery for elephantiasis nostras with large ectatic podoplanin-negative lymphatic vessels in patients with lipo-lymphedema.

PubMed

Wollina, Uwe; Heinig, Birgit; Schönlebe, Jaqueline; Nowak, Andreas

2014-01-01

Elephantiasis nostras is a rare complication in advanced lipo-lymphedema. While lipedema can be treated by liposuction and lymphedema by decongestive lymphatic therapy, elephantiasis nostras may need debulking surgery. We present 2 cases of advanced lipo-lymphedema complicated by elephantiasis nostras. After tumescent microcannular laser-assisted liposuction both patients underwent a debulking surgery with a modification of Auchincloss-Kim's technique. Histologic examination of the tissue specimen was performed. The surgical treatment was well tolerated and primary healing was uneventful. After primary wound healing and ambulation of the patients, a delayed ulceration with lymphorrhea developed. It was treated by surgical necrectomy and vacuum-assisted closure leading to complete healing. Mobility of the leg was much improved. Histologic examination revealed massive ectatic lymphatic vessels nonreactive for podoplanin. Debulking surgery can be an adjuvant technique for elephantiasis nostras in advanced lipo-lymphedema. Although delayed postoperative wound healing problems were observed, necrectomy and vacuum-assisted closure achieved a complete healing. Histologic data suggest that the ectatic lymphatic vessels in these patients resemble finding in podoplanin knockout mice. The findings would explain the limitations of decongestive lymphatic therapy and tumescent liposuction in such patients and their predisposition to relapsing erysipelas.

The CD11a and Endothelial Protein C Receptor Marker Combination Simplifies and Improves the Purification of Mouse Hematopoietic Stem Cells

PubMed Central

Karimzadeh, Alborz; Scarfone, Vanessa M.; Varady, Erika; Chao, Connie; Grathwohl, Karin; Fathman, John W.; Fruman, David A.; Serwold, Thomas

2018-01-01

Abstract Hematopoietic stem cells (HSCs) are the self‐renewing multipotent progenitors to all blood cell types. Identification and isolation of HSCs for study has depended on the expression of combinations of surface markers on HSCs that reliably distinguish them from other cell types. However, the increasing number of markers required to isolate HSCs has made it tedious, expensive, and difficult for newcomers, suggesting the need for a simpler panel of HSC markers. We previously showed that phenotypic HSCs could be separated based on expression of CD11a and that only the CD11a negative fraction contained true HSCs. Here, we show that CD11a and another HSC marker, endothelial protein C receptor (EPCR), can be used to effectively identify and purify HSCs. We introduce a new two‐color HSC sorting method that can highly enrich for HSCs with efficiencies comparable to the gold standard combination of CD150 and CD48. Our results demonstrate that adding CD11a and EPCR to the HSC biologist's toolkit improves the purity of and simplifies isolation of HSCs. stem cells translational medicine 2018;7:468–476 PMID:29543389

Isolated tumor endothelial cells maintain specific character during long-term culture

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

Matsuda, Kohei; Oral Pathology and Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586; Oral and Maxillofacial Surgery, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586

Tumor angiogenesis is necessary for solid tumor progression and metastasis. Increasing evidence indicates that tumor endothelial cells (TECs) are more relevant to the study of tumor angiogenesis than normal endothelial cells (NECs) because their morphologies and gene expression are different from NECs. However, it is challenging to isolate and culture large numbers of pure ECs from tumor tissue since the percentage of ECs is only about 1-2% and tumor cells and fibroblasts easily overgrow them. In addition, there has been concern that isolated TECs may lose their special phenotype once they are dissociated from tumor cells. In this study, wemore » have successfully purified murine TECs from four different human tumor xenografts and NECs from murine dermal tissue. Isolated ECs expressed endothelial markers, such as CD31, VE-cadherin (CD144), and endoglin (CD105), for more than 3 months after isolation. TECs maintained tumor endothelial-specific markers, such as tumor endothelial marker 8 (TEM8) and aminopeptidase N (APN), as in tumor blood vessels in vivo. In addition, TECs were more proliferative and motile than NECs. TECs showed a higher response to VEGF and higher expression of VEGF receptors-1 and -2 than NECs did. Stem cell antigen-1 was up-regulated in all four TECs, suggesting that they have a kind of stemness. Cultured TECs maintain distinct biological differences from NECs as in vivo. In conclusion, it was suggested that TECs are relevant material for tumor angiogenesis research.« less

Debulking Surgery for Elephantiasis Nostras With Large Ectatic Podoplanin-Negative Lymphatic Vessels in Patients With Lipo-Lymphedema

PubMed Central

Wollina, Uwe; Heinig, Birgit; Schönlebe, Jaqueline; Nowak, Andreas

2014-01-01

Objective: Elephantiasis nostras is a rare complication in advanced lipo-lymphedema. While lipedema can be treated by liposuction and lymphedema by decongestive lymphatic therapy, elephantiasis nostras may need debulking surgery. Methods: We present 2 cases of advanced lipo-lymphedema complicated by elephantiasis nostras. After tumescent microcannular laser-assisted liposuction both patients underwent a debulking surgery with a modification of Auchincloss-Kim's technique. Histologic examination of the tissue specimen was performed. Results: The surgical treatment was well tolerated and primary healing was uneventful. After primary wound healing and ambulation of the patients, a delayed ulceration with lymphorrhea developed. It was treated by surgical necrectomy and vacuum-assisted closure leading to complete healing. Mobility of the leg was much improved. Histologic examination revealed massive ectatic lymphatic vessels nonreactive for podoplanin. Conclusions: Debulking surgery can be an adjuvant technique for elephantiasis nostras in advanced lipo-lymphedema. Although delayed postoperative wound healing problems were observed, necrectomy and vacuum-assisted closure achieved a complete healing. Histologic data suggest that the ectatic lymphatic vessels in these patients resemble finding in podoplanin knockout mice. The findings would explain the limitations of decongestive lymphatic therapy and tumescent liposuction in such patients and their predisposition to relapsing erysipelas. PMID:24741382

Circadian variability of fibrinolytic markers and endothelial function in patients with obstructive sleep apnea.

PubMed

Bagai, Kanika; Muldowney, James A S; Song, Yanna; Wang, Lily; Bagai, Jayant; Artibee, Kay J; Vaughan, Douglas E; Malow, Beth A

2014-02-01

Obstructive sleep apnea (OSA) is strongly associated with cardiovascular disease, including stroke and acute coronary syndromes. Plasminogen activator inhibitor-1 (PAI-1), the principal inhibitor of tissue-type plasminogen activator (t-PA), has a pronounced circadian rhythm and is elevated in both OSA and cardiovascular disease and may be an important link between the two conditions. Endothelial dysfunction is one of the underlying pathophysiological mechanisms of cardiovascular disease, and may be altered in OSA. Our primary aim was to compare circadian variability of PAI-1 and t-PA in patients with OSA and normal controls by determining the amplitude (peak level) and mesor (rhythm adjusted mean) of PAI-1 and t-PA in serial blood samples over a 24-h period. The secondary aim was to measure markers of endothelial function (brachial and radial artery flow) in patients with OSA compared with normal controls. Cross-sectional cohort study. Subjects age 18 y or older, with a body mass index of 25-45 kg/m(2), with or without evidence of untreated OSA. Plasma samples were collected every 2 h, in OSA patients and matched controls, over a 24-h period. PAI-1 and t-PA antigen and activity were measured. The presence or absence of OSA (apnea-hypopnea index of 5 or greater) was confirmed by overnight polysomnography. Endothelial function was measured via brachial artery flow mediated vasodilatation and computerized arterial pulse waveform analysis. The rhythm-adjusted mean levels of PAI-1 antigen levels in the OSA group (21.8 ng/mL, 95% confidence level [CI], 18 to 25.7) were significantly higher as compared to the non-OSA group (16 ng/mL, 95% CI, 12.2 to 19.8; P = 0.03). The rhythm-adjusted mean levels of PAI-1 activity levels in the OSA group (23.9 IU/mL, 95% CI, 21.4 to 26.5) were also significantly higher than in the non-OSA group (17.2 IU/ mL, 95% CI, 14.6 to 19.9; P < 0.001).There were strong correlations between amplitude of PAI-1 activity and severity of OSA as measured

Primitive erythrocytes are generated from hemogenic endothelial cells.

PubMed

Stefanska, Monika; Batta, Kiran; Patel, Rahima; Florkowska, Magdalena; Kouskoff, Valerie; Lacaud, Georges

2017-07-25

Primitive erythroblasts are the first blood cells generated during embryonic hematopoiesis. Tracking their emergence both in vivo and in vitro has remained challenging due to the lack of specific cell surface markers. To selectively investigate primitive erythropoiesis, we have engineered a new transgenic embryonic stem (ES) cell line, where eGFP expression is driven by the regulatory sequences of the embryonic βH1 hemoglobin gene expressed specifically in primitive erythroid cells. Using this ES cell line, we observed that the first primitive erythroblasts are detected in vitro around day 1.5 of blast colony differentiation, within the cell population positive for the early hematopoietic progenitor marker CD41. Moreover, we establish that these eGFP + cells emerge from a hemogenic endothelial cell population similarly to their definitive hematopoietic counterparts. We further generated a corresponding βH1-eGFP transgenic mouse model and demonstrated the presence of a primitive erythroid primed hemogenic endothelial cell population in the developing embryo. Taken together, our findings demonstrate that both in vivo and in vitro primitive erythrocytes are generated from hemogenic endothelial cells.

In vitro differentiation of human tooth germ stem cells into endothelial- and epithelial-like cells.

PubMed

Doğan, Ayşegül; Demirci, Selami; Şahin, Fikrettin

2015-01-01

Current clinical techniques in dental practice include stem cell and tissue engineering applications. Dental stem cells are promising primary cell source for mainly tooth tissue engineering. Interaction of mesenchymal stem cell with epithelial and endothelial cells is strictly required for an intact tooth morphogenesis. Therefore, it is important to investigate whether human tooth germ stem cells (hTGSCs) derived from wisdom tooth are suitable for endothelial and epithelial cell transformation in dental tissue regeneration approaches. Differentiation into endothelial and epithelial cell lineages were mimicked under defined conditions, confirmed by real time PCR, western blotting and immunocytochemical analysis by qualitative and quantitative methods. HUVECs and HaCaT cells were used as positive controls for the endothelial and epithelial differentiation assays, respectively. Immunocytochemical and western blotting analysis revealed that terminally differentiated cells expressed cell-lineage markers including CD31, VEGFR2, VE-Cadherin, vWF (endothelial cell markers), and cytokeratin (CK)-17, CK-19, EpCaM, vimentin (epithelial cell markers) in significant levels with respect to undifferentiated control cells. Moreover, high expression levels of VEGFR1, VEGFR2, VEGF, CK-18, and CK-19 genes were detected in differentiated endothelial and epithelial-like cells. Endothelial-like cells derived from hTGSCs were cultured on Matrigel, tube-like structure formations were followed as an indication for functional endothelial differentiation. hTGSCs successfully differentiate into various cell types with a broad range of functional abilities using an in vitro approach. These findings suggest that hTGSCs may serve a potential stem cell source for tissue engineering and cell therapy of epithelial and endothelial tissue. © 2014 International Federation for Cell Biology.

The CD11a and Endothelial Protein C Receptor Marker Combination Simplifies and Improves the Purification of Mouse Hematopoietic Stem Cells.

PubMed

Karimzadeh, Alborz; Scarfone, Vanessa M; Varady, Erika; Chao, Connie; Grathwohl, Karin; Fathman, John W; Fruman, David A; Serwold, Thomas; Inlay, Matthew A

2018-06-01

Hematopoietic stem cells (HSCs) are the self-renewing multipotent progenitors to all blood cell types. Identification and isolation of HSCs for study has depended on the expression of combinations of surface markers on HSCs that reliably distinguish them from other cell types. However, the increasing number of markers required to isolate HSCs has made it tedious, expensive, and difficult for newcomers, suggesting the need for a simpler panel of HSC markers. We previously showed that phenotypic HSCs could be separated based on expression of CD11a and that only the CD11a negative fraction contained true HSCs. Here, we show that CD11a and another HSC marker, endothelial protein C receptor (EPCR), can be used to effectively identify and purify HSCs. We introduce a new two-color HSC sorting method that can highly enrich for HSCs with efficiencies comparable to the gold standard combination of CD150 and CD48. Our results demonstrate that adding CD11a and EPCR to the HSC biologist's toolkit improves the purity of and simplifies isolation of HSCs. Stem Cells Translational Medicine 2018;7:468-476. © 2018 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Lymphatic endothelial cell line (CH3) from a recurrent retroperitoneal lymphangioma.

PubMed

Way, D; Hendrix, M; Witte, M; Witte, C; Nagle, R; Davis, J

1987-09-01

An endothelial cell line derived from a massive recurrent chyle-containing retroperitoneal lymphangioma was isolated in monolayer culture. Scanning and transmission electron microscopy and immunohistochemistry confirmed a close resemblance to blood vascular endothelium with typical cobblestone morphology, positive immunofluorescence staining for endothelial marker Factor VIII-associated antigen and fibronectin, and prominent Weibel-Palade bodies. The endothelial cells also exhibited other ultrastructural features characteristic of lymphatic endothelium, including sparse microvillous surface projections, overlapping intercellular junctions, and abundant intermediate filaments. This endothelial cell line represents a new source of proliferating lymphatic endothelium for future study, including structural and functional comparison to blood vascular endothelium.

Endothelial-dependent flow-mediated dilation in African Americans with masked-hypertension.

PubMed

Veerabhadrappa, Praveen; Diaz, Keith M; Feairheller, Deborah L; Sturgeon, Katie M; Williamson, Sheara T; Crabbe, Deborah L; Kashem, Abul M; Brown, Michael D

2011-10-01

Office-blood pressure (BP) measurements alone overlook a significant number of individuals with masked-hypertension (office-BP: 120/80-139/89 mm Hg and 24-h ambulatory BP monitoring (ABPM) daytime ≥135/85 mm Hg or night-time ≥120/70 mm Hg). Diminished endothelial function contributes to the pathogenesis of hypertension. To better understand the pathophysiology involved in the increased cardiovascular (CV) disease risk associated with masked-hypertension, we estimated the occurrence, assessed the endothelial function, compared plasma levels of inflammatory markers, white blood cell count (WBC count), tumor necrosis factor-α (TNF-α), and high sensitivity C-reactive protein (hsCRP) and examined the possible relationship between endothelial function and inflammatory markers in apparently healthy prehypertensive (office-BP: 120/80-139/89 mm Hg) African Americans. Fifty African Americans who were sedentary, nondiabetic, nonsmoking, devoid of CV disease were recruited. Office-BP was measured according to JNC-7 guidelines to identify prehypertensives in whom ABPM was then assessed. Fasting plasma samples were assayed for inflammatory markers. Brachial artery flow-mediated dilation (FMD) at rest and during reactive hyperemia was measured in a subset of prehypertensives. Subjects in the masked-hypertension sub-group had a higher hsCRP (P = 0.04) and diminished endothelial function (P = 0.03) compared to the true-prehypertensive sub-group (office-BP: 120/80-139/89 mm Hg and ABPM: daytime <135/85 mm Hg or night-time <120/70 mm Hg). Regression analysis showed that endothelial function was inversely related to hsCRP amongst the masked-hypertensive sub-group (R(2) = 0.160; P = 0.04). Masked-hypertension was identified in 58% of African Americans which suggests that a masking phenomenon may exist in a sub-group of prehypertensives who also seem to have a diminished endothelial function that could be mediated by an elevated subclinical inflammation leading to the increased CV

Circulating Endothelial Cells in Patients with Heart Failure and Left Ventricular Dysfunction

PubMed Central

Martínez-Sales, Vicenta; Sánchez-Lázaro, Ignacio; Vila, Virtudes; Almenar, Luis; Contreras, Teresa; Reganon, Edelmiro

2011-01-01

Introduction and Aims: Acute and chronic heart failure may manifest different degrees of endothelial damage and angiogenesis. Circulating endothelial cells (CEC) have been identified as marker of vascular damage. The aim of our study was to evaluate the evolution of the CEC at different stages of patients with heart failure. We also investigated a potential correlation between CEC and markers of vascular damage and angiogenesis. Methods: We studied 32 heart failure patients at hospital admission (acute phase) and at revision after 3 months (stable phase) and 32 controls. Circulating markers of endothelial damage (CEC; von Willebrand factor, vWF and soluble E-selectin, sEsel) and angiogenesis (vascular endothelial growth factor, VEGF and thrombospondin-1) were quantified. Results: Levels of CEC, vWF, sEsel and VEGF are significantly higher in heart failure patients than in controls. Levels of CEC (36.9 ± 15.3 vs. 21.5 ± 10.0 cells/ml; p < 0.001), vWF (325 ± 101 vs. 231 ± 82%; p < 0.001) and VEGF (26.3 ± 15.2 vs. 21.9 ± 11.9 ng/ml; p < 0.001) are significantly higher in the acute phase than in the stable phase of heart failure. CEC levels correlate with vWF and VEGF. Results show than 100% of patients in acute phase and 37.5% in stable phase have levels of CEC higher than the 99th percentile of the distribution of controls (16 cells/ml). Therefore, increases in CEC represent a relative risk of 9.5 for heart failure patients suffering from acute phase. Conclusions: CEC, in addition to being elevated in heart failure, correlate with vWF levels, providing further support for CEC as markers of endothelial damage. Levels of CEC are associated with the acute phase of heart failure and could be used as a marker of the worsening in heart failure. PMID:21897001

SILAC-based proteomics of human primary endothelial cell morphogenesis unveils tumor angiogenic markers.

PubMed

Zanivan, Sara; Maione, Federica; Hein, Marco Y; Hernández-Fernaud, Juan Ramon; Ostasiewicz, Pawel; Giraudo, Enrico; Mann, Matthias

2013-12-01

Proteomics has been successfully used for cell culture on dishes, but more complex cellular systems have proven to be challenging and so far poorly approached with proteomics. Because of the complexity of the angiogenic program, we still do not have a complete understanding of the molecular mechanisms involved in this process, and there have been no in depth quantitative proteomic studies. Plating endothelial cells on matrigel recapitulates aspects of vessel growth, and here we investigate this mechanism by using a spike-in SILAC quantitative proteomic approach. By comparing proteomic changes in primary human endothelial cells morphogenesis on matrigel to general adhesion mechanisms in cells spreading on culture dish, we pinpoint pathways and proteins modulated by endothelial cells. The cell-extracellular matrix adhesion proteome depends on the adhesion substrate, and a detailed proteomic profile of the extracellular matrix secreted by endothelial cells identified CLEC14A as a matrix component, which binds to MMRN2. We verify deregulated levels of these proteins during tumor angiogenesis in models of multistage carcinogenesis. This is the most in depth quantitative proteomic study of endothelial cell morphogenesis, which shows the potential of applying high accuracy quantitative proteomics to in vitro models of vessel growth to shed new light on mechanisms that accompany pathological angiogenesis. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium with the data set identifier PXD000359.

Podoplanin emerges as a functionally relevant oral cancer biomarker and therapeutic target.

PubMed

Retzbach, Edward P; Sheehan, Stephanie A; Nevel, Evan M; Batra, Amber; Phi, Tran; Nguyen, Angels T P; Kato, Yukinari; Baredes, Soly; Fatahzadeh, Mahnaz; Shienbaum, Alan J; Goldberg, Gary S

2018-03-01

Oral cancer has become one of the most aggressive types of cancer, killing 140,000 people worldwide every year. Current treatments for oral cancer include surgery and radiation therapies. These procedures can be very effective; however, they can also drastically decrease the quality of life for survivors. New chemotherapeutic treatments are needed to more effectively combat oral cancer. The transmembrane receptor podoplanin (PDPN) has emerged as a functionally relevant oral cancer biomarker and chemotherapeutic target. PDPN expression promotes tumor cell migration leading to oral cancer invasion and metastasis. Here, we describe the role of PDPN in oral squamous cell carcinoma progression, and how it may be exploited to prevent and treat oral cancer. Copyright © 2018 Elsevier Ltd. All rights reserved.

Advanced glycation end products of bovine serum albumin-induced endothelial-to-mesenchymal transition in cultured human and monkey endothelial cells via protein kinase B signaling cascades.

PubMed

Ma, Jianli; Liu, Ting; Dong, Xiaoguang

2010-12-09

Advanced glycation end products of BSA (AGE-BSA) participate in the pathogenesis of diabetic vascular disease. However, the role of AGE-BSA in diabetic retinopathy, especially in retinal neovascularization, remains incomplete. This study aimed to determine the contributions of AGE-BSA in the endothelial-to-mesenchymal transition (EnMT) of cultured human and monkey endothelial cell lines and the mechanism that may be related with the transition. Monkey choroid-retinal endothelial cells (RF/6A) and human umbilical vein endothelial cells (HUVEC) were cultured in Dulbecco's modified Eagle's Medium (DMEM) and Ham's F12 medium containing 200 mg/l AGE-BSA. The expression of VE-cadherin, β-catenin, vimentin, N-cadherin, and protein kinase B (AKT2) was observed by immunocytochemistry and flow cytometry. Cell motility was determined by migration assays; the endothelial function of the formatting tube was measured by tube formation assays, while the change in the polarity was measured using resistance instruments. The characteristics of EnMT included loss of endothelial markers of VE-cadherin and β-catenin, which were replaced by mesenchymal markers of vimentin and N-cadherin, enhanced migration and tube formation, and diminished polarity. AGE-BSA contributed to upregulation of the protein expression of VE-cadherin and β-catenin and downregulation of protein expression of vimentin and N-cadherin, leading to enhanced migration and tube formation and diminished polarity. During this process, expression of AKT2 was upregulated. AGE-BSA can induce EnMT of cultured human and monkey endothelial cells. The signal pathway involving AKT2 may play a role in this process.

Short-term effects of air pollution, markers of endothelial activation, and coagulation to predict major adverse cardiovascular events in patients with acute coronary syndrome: insights from AIRACOS study.

PubMed

Dominguez-Rodriguez, Alberto; Abreu-Gonzalez, Pedro; Rodríguez, Sergio; Avanzas, Pablo; Juarez-Prera, Ruben A

2017-07-01

The aim of this study was to determine whether markers of inflammation and coagulation are associated with short-term particulate matter exposure and predict major adverse cardiovascular events at 360 d in patients with acute coronary syndrome (ACS). We included 307 consecutive patients, and assessed the average concentrations of data on atmospheric pollution in ambient air and meteorological variables from 1 d up to 7 d prior to admission. In patients with ACS, the markers of endothelial activation and coagulation, but not black carbon exposure, are associated with major adverse cardiovascular events at one-year follow-up.

Peripheral blood "endothelial progenitor cells" are derived from monocyte/macrophages and secrete angiogenic growth factors.

PubMed

Rehman, Jalees; Li, Jingling; Orschell, Christie M; March, Keith L

2003-03-04

Endothelial progenitor cells (EPCs) have been isolated from peripheral blood and can enhance angiogenesis after infusion into host animals. It is not known whether the proangiogenic effects are a result of such events as endothelial differentiation and subsequent proliferation of EPCs or secondary to secretion of angiogenic growth factors. Human EPCs were isolated as previously described, and their phenotypes were confirmed by uptake of acetylated LDL and binding of ulex-lectin. EPC proliferation and surface marker expression were analyzed by flow cytometry, and conditioned medium was assayed for growth factors. The majority of EPCs expressed monocyte/macrophage markers such as CD14 (95.7+/-0.3%), Mac-1 (57.6+/-13.5%), and CD11c (90.8+/-4.9%). A much lower percentage of cells expressed the specific endothelial marker VE-cadherin (5.2+/-0.7%) or stem/progenitor-cell markers AC133 (0.16+/-0.05%) and c-kit (1.3+/-0.7%). Compared with circulating monocytes, cultured EPCs showed upregulation of monocyte activation and macrophage differentiation markers. EPCs did not demonstrate any significant proliferation but did secrete the angiogenic growth factors vascular endothelial growth factor, hepatocyte growth factor, granulocyte colony-stimulating factor, and granulocyte-macrophage colony-stimulating factor. Our findings suggest that acetylated LDL(+)ulex-lectin(+) cells, commonly referred to as EPCs, do not proliferate but release potent proangiogenic growth factors. The majority of acetylated LDL(+)ulex-lectin(+) cells are derived from monocyte/macrophages. The findings of low proliferation and endothelial differentiation suggest that their angiogenic effects are most likely mediated by growth factor secretion. These findings may allow for development of novel angiogenic therapies relying on secreted growth factors or on recruitment of endogenous monocytes/macrophages to sites of ischemia.

The relationship of exposure to air pollutants in pregnancy with surrogate markers of endothelial dysfunction in umbilical cord.

PubMed

Poursafa, Parinaz; Baradaran-Mahdavi, Sadegh; Moradi, Bita; Haghjooy Javanmard, Shaghayegh; Tajadini, Mohammadhasan; Mehrabian, Ferdous; Kelishadi, Roya

2016-04-01

This study aims to investigate the association of exposure to ambient air pollution during pregnancy with cord blood concentrations of surrogate markers of endothelial dysfunction. This population-based cohort was conducted from March 2014 to March 2015 among 250 mother-neonate pairs in urban areas of Isfahan, the second large and air-polluted city in Iran. We analyzed the association between the ambient carbon monoxide (CO), ozone (O3), nitrogen dioxide (NO2), sulfur dioxide (SO2), particular matter 10 (PM10), and air quality index (AQI) with cord blood levels of endothelin-1, vascular adhesion molecule (VCAM), and intercellular adhesion molecule (ICAM). Multiple regression analysis was conducted after adjustment for potential confounding factors and covariates. The regression coefficient (beta), standard error of the estimate (SE), and 95% confidence intervals for each regression coefficient (95% CI) are reported. Data of 233 mother-neonate pairs were complete, and included in the analysis. Multiple regression analyses showed that AQI, CO and O3 had significant correlation with cord blood ICAM-1 [Beta (SE), 95%CI: 2.93 (0.72), 1.33,5.54; 2.28(1.44), 1.56,5.12; and 2.02(0.01), 1.03,2.04, respectively] as well as with VCAM-1 [2.78(0.91), 1.69,4.57; 2.47(1.47), 1.43,5.37; and 2.01(0.01),1.07,2.04, respectively]. AQI, PM10, and SO2 were significantly associated with Endothelin-1 concentrations [Beta (SE), 95%CI: 10.16(5.08),7.61,14.28; 9.70(3.46), 2.88,16.52; and 1.07(0.02), 1.03,2.11, respectively]. The significant associations of air pollutants with markers of endothelial dysfunction during fetal period may provide another evidence on the adverse health effects of air pollutants on early stages of atherosclerosis from fetal period. Our findings underscore the importance of considering environmental factors in primordial prevention of chronic diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

Role of endothelial-to-mesenchymal transition in the pathogenesis of central nervous system hemangioblastomas.

PubMed

Takada, Shigeki; Hojo, Masato; Takebe, Noriyoshi; Tanigaki, Kenji; Miyamoto, Susumu

2018-06-07

Hemangioblastomas (HBs) are benign vascular tumors of the central nervous system and histologically contain abundant microvessels. Therefore, they clinically exhibit vascular malformation-like characteristics. It has been described that endothelial-to-mesenchymal transition (EndMT) contributes to the pathogenesis of cerebral cavernous malformations. However, it remains unknown whether EndMT contributes to the pathogenesis of central nervous system HBs. The aim of our study was to investigate whether EndMT occurs in central nervous system HBs. Ten central nervous system HBs were immunohistochemically investigated. CD31 (an endothelial marker) and EndMT markers, such as α-smooth muscle actin (a mesenchymal marker) and CD44 (a mesenchymal stem cell marker), were expressed in the endothelial layer of microvessels in all cases. These findings suggest that endothelial cells (ECs) of microvessels in central nervous system HBs have acquired mesenchymal and stem-cell-like characteristics and undergone EndMT. In all cases, both ephrin-B2 and EphB4, which are not detected in adult normal brain vessels, were expressed in the endothelial layer of microvessels. These data suggest that ECs of microvessels in central nervous system HBs are immature or malformed cells and have both arterial and venous characteristics. This is the first report showing the possibility that EndMT contributes to the pathogenesis of central nervous system HBs. It is likely that ECs of microvessels in central nervous system HBs are immature or malformed cells and have both arterial and venous characteristics. EndMT is expected to be a new therapeutic target in central nervous system HBs. Copyright © 2018 Elsevier Inc. All rights reserved.

Novel Monoclonal Antibody LpMab-17 Developed by CasMab Technology Distinguishes Human Podoplanin from Monkey Podoplanin.

PubMed

Kato, Yukinari; Ogasawara, Satoshi; Oki, Hiroharu; Honma, Ryusuke; Takagi, Michiaki; Fujii, Yuki; Nakamura, Takuro; Saidoh, Noriko; Kanno, Hazuki; Umetsu, Mitsuo; Kamata, Satoshi; Kubo, Hiroshi; Yamada, Mitsuhiro; Sawa, Yoshihiko; Morita, Kei-Ichi; Harada, Hiroyuki; Suzuki, Hiroyoshi; Kaneko, Mika Kato

2016-04-01

Podoplanin (PDPN) is a type-I transmembrane sialoglycoprotein, which possesses a platelet aggregation-stimulating (PLAG) domain in its N-terminus. Among the three PLAG domains, O-glycan on Thr52 of PLAG3 is critical for the binding with C-type lectin-like receptor-2 (CLEC-2) and is essential for platelet-aggregating activity of PDPN. Although many anti-PDPN monoclonal antibodies (mAbs) have been established, almost all mAbs bind to PLAG domains. We recently established CasMab technology to produce mAbs against membranous proteins. Using CasMab technology, we produced a novel anti-PDPN mAb, LpMab-17, which binds to non-PLAG domains. LpMab-17 clearly detected endogenous PDPN of cancer cells and normal cells in Western-blot, flow cytometry, and immunohistochemistry. LpMab-17 recognized glycan-deficient PDPN in flow cytometry, indicating that the interaction between LpMab-17 and PDPN is independent of its glycosylation. The minimum epitope of LpMab-17 was identified as Gly77-Asp82 of PDPN using enzyme-linked immunosorbent assay. Of interest, LpMab-17 did not bind to monkey PDPN, whereas the homology is 94% between human PDPN and monkey PDPN, indicating that the epitope of LpMab-17 is unique compared with the other anti-PDPN mAbs. The combination of different epitope-possessing mAbs could be advantageous for the PDPN-targeting diagnosis or therapy.

Comparative effects of enzogenol and vitamin C supplementation versus vitamin C alone on endothelial function and biochemical markers of oxidative stress and inflammation in chronic smokers.

PubMed

Young, Joanna M; Shand, Brett I; McGregor, Patrice M; Scott, Russell S; Frampton, Christopher M

2006-01-01

Chronic smoking is associated with endothelial dysfunction and inflammation, with oxidative stress contributing to both these processes. In this study, we investigated the effect of combined antioxidant treatment with Enzogenol, a flavonoid extract from the bark of Pinus radiata and vitamin C, over and above vitamin C alone, on endothelial function, plasma markers of inflammation and oxidative stress, blood pressure (BP) and anthropometrics. Forty-four chronic smokers without established cardiovascular disease were assigned randomly to receive either 480 mg Enzogenol and 60 mg vitamin C, or 60 mg vitamin C alone daily for 12 weeks. Endothelial function in the brachial artery was assessed by flow-mediated vasodilation (FMD). FMD improved in both treatment groups (p < 0.001), with no significant difference between the two groups (p = 0.84). In the group receiving Enzogenol and vitamin C, protein carbonyl levels were significantly reduced compared to the group taking vitamin C alone (p = 0.03). Enzogenol and vitamin C resulted in a significant reduction in fibrinogen levels in heavy smokers compared with vitamin C alone (p < 0.009). These findings demonstrated that co-supplementation with Enzogenol and vitamin C in smokers conferred no additional beneficial effect on macrovascular endothelial function over and above that seen in the vitamin C alone group. However, Enzogenol did demonstrate additional favourable effects on protein oxidative damage and fibrinogen levels.

Osteocalcin expression by circulating endothelial progenitor cells in patients with coronary atherosclerosis.

PubMed

Gössl, Mario; Mödder, Ulrike I; Atkinson, Elizabeth J; Lerman, Amir; Khosla, Sundeep

2008-10-14

This study was designed to test whether patients with coronary atherosclerosis have increases in circulating endothelial progenitor cells (EPCs) expressing an osteogenic phenotype. Increasing evidence indicates a link between bone and the vasculature, and bone marrow and circulating osteogenic cells have been identified by staining for the osteoblastic marker, osteocalcin (OCN). Endothelial progenitor cells contribute to vascular repair, but repair of vascular injury may result in calcification. Using cell surface markers (CD34, CD133, kinase insert domain receptor [KDR]) to identify EPCs, we examined whether patients with coronary atherosclerosis had increases in the percentage of EPCs expressing OCN. We studied 72 patients undergoing invasive coronary assessment: control patients (normal coronary arteries and no endothelial dysfunction, n = 21) versus 2 groups with coronary atherosclerosis-early coronary atherosclerosis (normal coronary arteries but with endothelial dysfunction, n = 22) and late coronary atherosclerosis (severe, multivessel coronary artery disease, n = 29). Peripheral blood mononuclear cells were analyzed using flow cytometry. Compared with control patients, patients with early or late coronary atherosclerosis had significant increases (approximately 2-fold) in the percentage of CD34+/KDR+ and CD34+/CD133+/KDR+ cells costaining for OCN. Even larger increases were noted in the early and late coronary atherosclerosis patients in the percentage of CD34+/CD133-/KDR+ cells costaining for OCN (5- and 2-fold, p < 0.001 and 0.05, respectively). A higher percentage of EPCs express OCN in patients with coronary atherosclerosis compared with subjects with normal endothelial function and no structural coronary artery disease. These findings have potential implications for the mechanisms of vascular calcification and for the development of novel markers for coronary atherosclerosis.

Kalirin and CHD7: novel endothelial dysfunction indicators in circulating extracellular vesicles from hypertensive patients with albuminuria

PubMed Central

de la Cuesta, Fernando; Baldan-Martin, Montserrat; Moreno-Luna, Rafael; Alvarez-Llamas, Gloria; Gonzalez-Calero, Laura; Mourino-Alvarez, Laura; Sastre-Oliva, Tamara; López, Juan A.; Vázquez, Jesús; Ruiz-Hurtado, Gema; Segura, Julian; Vivanco, Fernando; Ruilope, Luis M.; Barderas, Maria G.

2017-01-01

Despite of the great advances in anti-hypertensive therapies, many patients under Renin-Angiotensin- System (RAS) suppression develop albuminuria, which is a clear indicator of therapeutic inefficiency. Hence, indicators of vascular function are needed to assess patients’ condition and help deciding future therapies. Proteomic analysis of circulating extracellular vesicles (EVs) showed two proteins, kalirin and chromodomain-helicase-DNA-binding protein 7 (CHD7), increased in albuminuric patients. A positive correlation of both with the expression of the endothelial activation marker E-selectin was found in EVs. In vitro analysis using TNFα-treated adult human endothelial cells proved their involvement in endothelial cell activation. Hence, we propose protein levels of kalirin and CHD7 in circulating EVs as novel endothelial dysfunction markers to monitor vascular condition in hypertensive patients with albuminuria. PMID:28152519

circHECTD1 promotes the silica-induced pulmonary endothelial-mesenchymal transition via HECTD1.

PubMed

Fang, Shencun; Guo, Huifang; Cheng, Yusi; Zhou, Zewei; Zhang, Wei; Han, Bing; Luo, Wei; Wang, Jing; Xie, Weiping; Chao, Jie

2018-03-14

Excessive proliferation and migration of fibroblasts contribute to pulmonary fibrosis in silicosis, and both epithelial cells and endothelial cells participate in the accumulation of fibroblasts via the epithelial-mesenchymal transition (EMT) and the endothelial-mesenchymal transition (EndMT), respectively. A mouse endothelial cell line (MML1) was exposed to silicon dioxide (SiO 2 , 50 μg/cm 2 ), and immunofluorescence and western blot analyses were performed to evaluate levels of specific endothelial and mesenchymal markers and to elucidate the mechanisms by which SiO 2 induces the EndMT. Functional changes were evaluated by analyzing cell migration and proliferation. The mRNA and circular RNA (circRNA) levels were measured using qPCR and fluorescent in situ hybridization (FISH). Lung tissue samples from both Tie2-GFP mice exposed to SiO 2 and silicosis patients were applied to confirm the observations from in vitro experiments. Based on the results from the current study, SiO 2 increased the expression of mesenchymal markers (type I collagen (COL1A1), type III collagen (COL3A1) and alpha smooth muscle actin (α-SMA/Acta2)) and decreased the expression of endothelial markers (vascular endothelial cadherin (VE-Cad/Cdh 5) and platelet endothelial cell adhesion molecule-1 (PECAM1)), indicating the occurrence of the EndMT in response to SiO 2 exposure both in vivo and in vitro. SiO 2 concomitantly increased circHECTD1 expression, which, in turn, inhibited HECTD1 protein expression. SiO 2 -induced increases in cell proliferation, migration, and changes in marker levels were restored by either a small interfering RNA (siRNA) targeting circHECTD1 or overexpression of HECTD1 via the CRISPR/Cas9 system, confirming the involvement of the circHECTD1/HECTD1 pathway in the EndMT. Moreover, tissue samples from SiO 2 -exposed mice and silicosis patients confirmed the EndMT and change in HECTD1 expression. Our findings reveal a potentially new function for the circHECTD1/HECTD

T1alpha/podoplanin shows raft-associated distribution in mouse lung alveolar epithelial E10 cells.

PubMed

Barth, Kathrin; Bläsche, Robert; Kasper, Michael

2010-01-01

T1alpha/(podoplanin) is abundantly expressed in the alveolar epithelial type I cells (ATI) of rodent and human lungs. Caveolin-1 is a classical primary structural protein of plasmalemal invaginations, so-called caveolae, which represent specialized lipid rafts, and which are particularly abundant in ATI cells. The biological functions of T1alpha in the alveolar epithelium are unknown. Here we report on the characteristics of raft domains in the microplicae/microvillar protrusions of ATI cells, which contain T1alpha. Detergent resistant membranes (DRMs) from cell lysates of the mouse epithelial ATI-like cell line E10 were prepared using different detergents followed by flotation in a sucrose gradient and tested by Western and dot blots with raft markers (caveolin-1, GM1) and nonraft markers (transferrin receptor, PDI and beta-Cop). Immunocytochemistry was employed for the localization of T1alpha in E10 cells and in situ in rat lungs. Our biochemical results showed that the solubility or insolubility of T1alpha and caveolin-1 differs in Triton X-100 and Lubrol WX, two distinct non-ionic detergents. Caveolin-1 was unsoluble in both detergents, whereas T1alpha was Triton X-100 soluble but Lubrol WX insoluble. Immunofluorescence double stainings revealed that both proteins were colocalized with GM1, while caveolin-1 and T1alpha were not colocalized in the plasma membrane. Cholesterol depletion modified the segregation of T1alpha in Lubrol WX DRMs. Cellular processes in ultrathin sections of cultured mouse E10 cells were immunogold positive. Immunoelectron microscopy (postembedding) of rat lung tissue revealed the preferential localization of T1alpha on apical microvillar protrusions of ATI cells. We conclude that T1alpha and caveolin-1 are located in distinct plasma membrane microdomains, which differ in their protein-lipid interactions. The raft-associated distribution of T1alpha may have an impact on a specific, not yet clarified function of this protein in the

Solid tumor therapy by selectively targeting stromal endothelial cells

PubMed Central

Liu, Shihui; Liu, Jie; Ma, Qian; Cao, Liu; Fattah, Rasem J.; Yu, Zuxi; Bugge, Thomas H.; Finkel, Toren; Leppla, Stephen H.

2016-01-01

Engineered tumor-targeted anthrax lethal toxin proteins have been shown to strongly suppress growth of solid tumors in mice. These toxins work through the native toxin receptors tumor endothelium marker-8 and capillary morphogenesis protein-2 (CMG2), which, in other contexts, have been described as markers of tumor endothelium. We found that neither receptor is required for tumor growth. We further demonstrate that tumor cells, which are resistant to the toxin when grown in vitro, become highly sensitive when implanted in mice. Using a range of tissue-specific loss-of-function and gain-of-function genetic models, we determined that this in vivo toxin sensitivity requires CMG2 expression on host-derived tumor endothelial cells. Notably, engineered toxins were shown to suppress the proliferation of isolated tumor endothelial cells. Finally, we demonstrate that administering an immunosuppressive regimen allows animals to receive multiple toxin dosages and thereby produces a strong and durable antitumor effect. The ability to give repeated doses of toxins, coupled with the specific targeting of tumor endothelial cells, suggests that our strategy should be efficacious for a wide range of solid tumors. PMID:27357689

[Definitions destruction of endothelial cells as a marker of endothelial dysfunction in aging men].

PubMed

Пустовойт, Ганна Л; Ярмола, Тетяна І; Мохначов, Олександр В; Ткаченко, Лідія А; Супруненко, Сергій М

Ukraine occupies the 143 place in the world in life expectancy and the first place in terms of mortality. The main cause of death - cardiovascular diseases - 58%. Recent studies show the important and independent role of endothelium in the development of cardiovascular disease. examination of the endothelium destruction in aging men by determining the level of surface specific antigens of endothelial microparticles. 88 men age from 45 to 76 years. 50 people of the main group had a history of the second type diabetes mellitus (DM-2) combined with arterial hypertension (AH). The control group included 38 men without aforementioned diseases. Also, men were divided into two groups by age: 45-59 years and over 60. in the main subgroup I endothelin level was higher than the control subgroup I: 2,07 ± 0,6 and 1,27 ± 0,25 (p <0.05). In main subgroup II endothelin level was also significantly higher compared with the specified index in control subgroup II: 3,91 ± 0,7 and 1,79 ± 0,27 (p <0.05). Among patients of the main subgroup II endothelin level (3,88 ± 0,7 and 2,04 ± 0,6 (p <0.05)), and triglycerides (2,77 ± 0,08 vs. 1.99 ± 0.05 (p <0.05)) was higher compared with the I subgroup. age androgen deficiency is accompanied by lipid metabolism, development of endothelial dysfunction, insulin resistance, diabetes and hypertension. Reduction of the cardiovascular risk includes measures aimed at normalizing hormonal balance and lipid metabolism in aging men with DM-2 and hypertension.

[Definitions destruction of endothelial cells as a marker of endothelial dysfunction in aging men].

PubMed

Пустовойт, Ганна Л; Ярмола, Тетяна І; Мохначов, Олександр В; Ткаченко, Лідія А; Супруненко, Сергій М

2016-01-01

Ukraine occupies the 143 place in the world in life expectancy and the first place in terms of mortality. The main cause of death - cardiovascular diseases - 58%. Recent studies show the important and independent role of endothelium in the development of cardiovascular disease. examination of the endothelium destruction in aging men by determining the level of surface specific antigens of endothelial microparticles. 88 men age from 45 to 76 years. 50 people of the main group had a history of the second type diabetes mellitus (DM-2) combined with arterial hypertension (AH). The control group included 38 men without aforementioned diseases. Also, men were divided into two groups by age: 45-59 years and over 60. in the main subgroup I endothelin level was higher than the control subgroup I: 2,07 ± 0,6 and 1,27 ± 0,25 (p <0.05). In main subgroup II endothelin level was also significantly higher compared with the specified index in control subgroup II: 3,91 ± 0,7 and 1,79 ± 0,27 (p <0.05). Among patients of the main subgroup II endothelin level (3,88 ± 0,7 and 2,04 ± 0,6 (p <0.05)), and triglycerides (2,77 ± 0,08 vs. 1.99 ± 0.05 (p <0.05)) was higher compared with the I subgroup. age androgen deficiency is accompanied by lipid metabolism, development of endothelial dysfunction, insulin resistance, diabetes and hypertension. Reduction of the cardiovascular risk includes measures aimed at normalizing hormonal balance and lipid metabolism in aging men with DM-2 and hypertension.

Endothelial-to-Osteoblast Conversion Generates Osteoblastic Metastasis of Prostate Cancer.

PubMed

Lin, Song-Chang; Lee, Yu-Chen; Yu, Guoyu; Cheng, Chien-Jui; Zhou, Xin; Chu, Khoi; Murshed, Monzur; Le, Nhat-Tu; Baseler, Laura; Abe, Jun-Ichi; Fujiwara, Keigi; deCrombrugghe, Benoit; Logothetis, Christopher J; Gallick, Gary E; Yu-Lee, Li-Yuan; Maity, Sankar N; Lin, Sue-Hwa

2017-06-05

Prostate cancer (PCa) bone metastasis is frequently associated with bone-forming lesions, but the source of the osteoblastic lesions remains unclear. We show that the tumor-induced bone derives partly from tumor-associated endothelial cells that have undergone endothelial-to-osteoblast (EC-to-OSB) conversion. The tumor-associated osteoblasts in PCa bone metastasis specimens and patient-derived xenografts (PDXs) were found to co-express endothelial marker Tie-2. BMP4, identified in PDX-conditioned medium, promoted EC-to-OSB conversion of 2H11 endothelial cells. BMP4 overexpression in non-osteogenic C4-2b PCa cells led to ectopic bone formation under subcutaneous implantation. Tumor-induced bone was reduced in trigenic mice (Tie2 cre /Osx f/f /SCID) with endothelial-specific deletion of osteoblast cell-fate determinant OSX compared with bigenic mice (Osx f/f /SCID). Thus, tumor-induced EC-to-OSB conversion is one mechanism that leads to osteoblastic bone metastasis of PCa. Copyright © 2017 Elsevier Inc. All rights reserved.

Endothelial biomarkers in human sepsis: pathogenesis and prognosis for ARDS

PubMed Central

Hendrickson, Carolyn M.; Matthay, Michael A.

2018-01-01

Experimental models of sepsis in small and large animals and a variety of in vitro preparations have established several basic mechanisms that drive endothelial injury. This review is focused on what can be learned from the results of clinical studies of plasma biomarkers of endothelial injury and inflammation in patients with sepsis. There is excellent evidence that elevated plasma levels of several biomarkers of endothelial injury, including von Willebrand factor antigen (VWF), angiopoietin-2 (Ang-2), and soluble fms-like tyrosine kinase 1 (sFLT-1), and biomarkers of inflammation, especially interleukin-8 (IL-8) and soluble tumor necrosis factor receptor (sTNFr), identify sepsis patients with a higher mortality. There are also some data that elevated levels of endothelial biomarkers can identify which patients with non-pulmonary sepsis will develop acute respiratory distress syndrome (ARDS). If ARDS patients are divided among those with indirect versus direct lung injury, then there is an association of elevated levels of endothelial biomarkers in indirect injury and markers of inflammation and alveolar epithelial injury in patients with direct lung injury. New research suggests that the combination of biologic and clinical markers may make it possible to segregate patients with ARDS into hypo- versus hyper-inflammatory phenotypes that may have implications for therapeutic responses to fluid therapy. Taken together, the studies reviewed here support a primary role of the microcirculation in the pathogenesis and prognosis of ARDS after sepsis. Biological differences identified by molecular patterns could explain heterogeneity of treatment effects that are not explained by clinical factors alone. PMID:29575977

Somatic GNAQ Mutation is Enriched in Brain Endothelial Cells in Sturge-Weber Syndrome.

PubMed

Huang, Lan; Couto, Javier A; Pinto, Anna; Alexandrescu, Sanda; Madsen, Joseph R; Greene, Arin K; Sahin, Mustafa; Bischoff, Joyce

2017-02-01

Sturge-Weber syndrome (SWS) is a rare congenital neurocutaneous disorder characterized by facial and extracraniofacial capillary malformations and capillary-venule malformations in the leptomeninges. A somatic mosaic mutation in GNAQ (c.548G>A; p.R183Q) was found in SWS brain and skin capillary malformations. Our laboratory showed endothelial cells in skin capillary malformations are enriched for the GNAQ mutation. The purpose of this study is to determine whether the GNAQ mutation is also enriched in endothelial cells in affected SWS brain. Two human SWS brain specimens were fractionated by fluorescence-activated cell sorting into hematopoietic (CD45), endothelial (CD31, VE-Cadherin, and vascular endothelial growth factor receptor 2), and perivascular (platelet-derived growth factor receptor beta) cells and cells negative for all markers. The sorted cell populations were analyzed for GNAQ p.R183Q mutation by droplet digital polymerase chain reaction. SWS patient-derived brain endothelial cells were selected by anti-CD31-coated magnetic beads and cultured in endothelial growth medium in vitro. The GNAQ p.R183Q mutation was present in brain endothelial cells in two SWS specimens, with mutant allelic frequencies of 34.7% and 24.0%. Cells negative for all markers also harbored the GNAQ mutation. The mutant allelic frequencies in these unidentified cells were 9.2% and 8.4%. SWS patient-derived brain endothelial cells with mutant allelic frequencies of 14.7% and 21% survived and proliferated in vitro. Our study provides evidence that GNAQ p.R183Q mutation is enriched in endothelial cells in SWS brain lesions and thereby reveals endothelial cells as a source of aberrant Gαq signaling. This will help to understand the pathophysiology of SWS, to discover biomarkers for predicting cerebral involvement, and to develop therapeutic targets to prevent neurological impairments in SWS. Copyright © 2016 Elsevier Inc. All rights reserved.

N-Acetylcysteine Increases Corneal Endothelial Cell Survival in a Mouse Model of Fuchs Endothelial Corneal Dystrophy

PubMed Central

Kim, Eun Chul; Meng, Huan; Jun, Albert S.

2014-01-01

The present study evaluated survival effects of N-acetylcysteine (NAC) on cultured corneal endothelial cells exposed to oxidative and endoplasmic reticulum (ER) stress and in a mouse model of early-onset Fuchs endothelial corneal dystrophy (FECD). Cultured bovine corneal endothelial cell viability against oxidative and ER stress was determined by CellTiter-Glo® luminescent reagent. Two-month-old homozygous knock-in Col8a2L450W/L450W mutant (L450W) and C57/Bl6 wild-type (WT) animals were divided into two groups of 15 mice. Group I received 7 mg/mL NAC in drinking water and Group II received control water for 7 months. Endothelial cell density and morphology were evaluated with confocal microscopy. Antioxidant gene (iNos) and ER stress/unfolded protein response gene (Grp78 and Chop) mRNA levels and protein expression were measured in corneal endothelium by real time PCR and Western blotting. Cell viability of H2O2 and thapsigargin exposed cells pre-treated with NAC was significantly increased compared to untreated controls (pitalic>0.01). Corneal endothelial cell density (CD) was higher (p=0.001) and percent polymegathism was lower (p=0.04) in NAC treated L450W mice than in untreated L450W mice. NAC treated L450W endothelium showed significant upregulation of iNos, whereas Grp78 and Chop were downregulated compared to untreated L450W endothelium by real time PCR and Western blotting. NAC increases survival in cultured corneal endothelial cells exposed against ER and oxidative stress. Systemic NAC ingestion increases corneal endothelial cell survival which is associated with increased antioxidant and decreased ER stress markers in a mouse model of early-onset FECD. Our study presents in vivo evidence of a novel potential medical treatment for FECD. PMID:24952277

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

Endovascular treatment of chronic cerebro spinal venous insufficiency in patients with multiple sclerosis modifies circulating markers of endothelial dysfunction and coagulation activation: a prospective study.

PubMed

Napolitano, Mariasanta; Bruno, Aldo; Mastrangelo, Diego; De Vizia, Marcella; Bernardo, Benedetto; Rosa, Buonagura; De Lucia, Domenico

2014-10-01

We performed a monocentric observational prospective study to evaluate coagulation activation and endothelial dysfunction parameters in patients with multiple sclerosis undergoing endovascular treatment for cerebro-spinal-venous insufficiency. Between February 2011 and July 2012, 144 endovascular procedures in 110 patients with multiple sclerosis and chronical cerebro-spinal venous insufficiency were performed and they were prospectively analyzed. Each patient was included in the study according to previously published criteria, assessed by the investigators before enrollment. Endothelial dysfunction and coagulation activation parameters were determined before the procedure and during follow-up at 1, 3, 6, 9, 12, 15 and 18 months after treatment, respectively. After the endovascular procedure, patients were treated with standard therapies, with the addition of mesoglycan. Fifty-five percent of patients experienced a favorable outcome of multiple sclerosis within 1 month after treatment, 25% regressed in the following 3 months, 24.9% did not experience any benefit. In only 0.1% patients, acute recurrence was observed and it was treated with high-dose immunosuppressive therapy. No major complications were observed. Coagulation activation and endothelial dysfunction parameters were shown to be reduced at 1 month and stable up to 12-month follow-up, and they were furthermore associated with a good clinical outcome. Endovascular procedures performed by a qualified staff are well tolerated; they can be associated with other currently adopted treatments. Correlations between inflammation, coagulation activation and neurodegenerative disorders are here supported by the observed variations in plasma levels of markers of coagulation activation and endothelial dysfunction.

Pseudoangiomatous Stromal Hyperplasia in Core Needle Biopsies of Breast Specimens.

PubMed

Kelten Talu, Canan; Boyaci, Ceren; Leblebici, Cem; Hacihasanoglu, Ezgi; Bozkurt, Erol Rustu

2017-02-01

Pseudoangiomatous stromal hyperplasia (PASH) is a benign lesion of myofibroblasts that is composed of a network of slit-like channels that resemble vascular spaces. The aims of this study were to document the frequency of PASH in core needle biopsy specimens (CNBS) of the breast, to describe which histopathologic findings coexist with PASH and to examine any endothelial cell differentiation. We reevaluated hematoxylin and eosin-stained sections of all CNBS that were obtained during a 1-year period. First, we performed CD34 and CD31 immunostainings to highlight the areas of PASH, then performed D2-40/podoplanin (lymphatic endothelial marker) and Fli-1 (vascular endothelial cell marker) immunostains. The total number of CNBS was 412. Areas of PASH were noted in 37 of the 412 cases (9%), with a mean age of 38.5 years. The lesions that were described in association with PASH were "benign breast parenchyma with stromal fibrosis" (17/37; 46%), "fibroepithelial tumors" (17/37; 46%), "columnar cell changes (CCC)" (2/37; 5%), and "invasive carcinoma" (1/37; 3%). There were 2 cases of CCC within the foci of PASH (direct contact with PASH), and 8 additional cases of CCC that coexisted in the same specimen but were not in direct contact. There was no staining for D2-40 or Fli-1 within PASH foci. PASH lesions occurred with a frequency of 9% in CNBS and were mostly in association with benign breast lesions in premenopausal women. CCC was determined as an accompanying epithelial lesion within or near PASH areas. No obvious immunopositivity compatible with endothelial cell differentiation was revealed.

Podoplanin immunopositive lymphatic vessels at the implant interface in a rat model of osteoporotic fractures.

PubMed

Lips, Katrin Susanne; Kauschke, Vivien; Hartmann, Sonja; Thormann, Ulrich; Ray, Seemun; Kampschulte, Marian; Langheinrich, Alexander; Schumacher, Matthias; Gelinsky, Michael; Heinemann, Sascha; Hanke, Thomas; Kautz, Armin R; Schnabelrauch, Matthias; Schnettler, Reinhard; Heiss, Christian; Alt, Volker; Kilian, Olaf

2013-01-01

Insertion of bone substitution materials accelerates healing of osteoporotic fractures. Biodegradable materials are preferred for application in osteoporotic patients to avoid a second surgery for implant replacement. Degraded implant fragments are often absorbed by macrophages that are removed from the fracture side via passage through veins or lymphatic vessels. We investigated if lymphatic vessels occur in osteoporotic bone defects and whether they are regulated by the use of different materials. To address this issue osteoporosis was induced in rats using the classical method of bilateral ovariectomy and additional calcium and vitamin deficient diet. In addition, wedge-shaped defects of 3, 4, or 5 mm were generated in the distal metaphyseal area of femur via osteotomy. The 4 mm defects were subsequently used for implantation studies where bone substitution materials of calcium phosphate cement, composites of collagen and silica, and iron foams with interconnecting pores were inserted. Different materials were partly additionally functionalized by strontium or bisphosphonate whose positive effects in osteoporosis treatment are well known. The lymphatic vessels were identified by immunohistochemistry using an antibody against podoplanin. Podoplanin immunopositive lymphatic vessels were detected in the granulation tissue filling the fracture gap, surrounding the implant and growing into the iron foam through its interconnected pores. Significant more lymphatic capillaries were counted at the implant interface of composite, strontium and bisphosphonate functionalized iron foam. A significant increase was also observed in the number of lymphatics situated in the pores of strontium coated iron foam. In conclusion, our results indicate the occurrence of lymphatic vessels in osteoporotic bone. Our results show that lymphatic vessels are localized at the implant interface and in the fracture gap where they might be involved in the removal of lymphocytes, macrophages

Podoplanin Immunopositive Lymphatic Vessels at the Implant Interface in a Rat Model of Osteoporotic Fractures

PubMed Central

Lips, Katrin Susanne; Kauschke, Vivien; Hartmann, Sonja; Thormann, Ulrich; Ray, Seemun; Kampschulte, Marian; Langheinrich, Alexander; Schumacher, Matthias; Gelinsky, Michael; Heinemann, Sascha; Hanke, Thomas; Kautz, Armin R.; Schnabelrauch, Matthias; Schnettler, Reinhard; Heiss, Christian; Alt, Volker; Kilian, Olaf

2013-01-01

Insertion of bone substitution materials accelerates healing of osteoporotic fractures. Biodegradable materials are preferred for application in osteoporotic patients to avoid a second surgery for implant replacement. Degraded implant fragments are often absorbed by macrophages that are removed from the fracture side via passage through veins or lymphatic vessels. We investigated if lymphatic vessels occur in osteoporotic bone defects and whether they are regulated by the use of different materials. To address this issue osteoporosis was induced in rats using the classical method of bilateral ovariectomy and additional calcium and vitamin deficient diet. In addition, wedge-shaped defects of 3, 4, or 5 mm were generated in the distal metaphyseal area of femur via osteotomy. The 4 mm defects were subsequently used for implantation studies where bone substitution materials of calcium phosphate cement, composites of collagen and silica, and iron foams with interconnecting pores were inserted. Different materials were partly additionally functionalized by strontium or bisphosphonate whose positive effects in osteoporosis treatment are well known. The lymphatic vessels were identified by immunohistochemistry using an antibody against podoplanin. Podoplanin immunopositive lymphatic vessels were detected in the granulation tissue filling the fracture gap, surrounding the implant and growing into the iron foam through its interconnected pores. Significant more lymphatic capillaries were counted at the implant interface of composite, strontium and bisphosphonate functionalized iron foam. A significant increase was also observed in the number of lymphatics situated in the pores of strontium coated iron foam. In conclusion, our results indicate the occurrence of lymphatic vessels in osteoporotic bone. Our results show that lymphatic vessels are localized at the implant interface and in the fracture gap where they might be involved in the removal of lymphocytes, macrophages

Flavorings in Tobacco Products Induce Endothelial Cell Dysfunction.

PubMed

Fetterman, Jessica L; Weisbrod, Robert M; Feng, Bihua; Bastin, Reena; Tuttle, Shawn T; Holbrook, Monica; Baker, Gregory; Robertson, Rose Marie; Conklin, Daniel J; Bhatnagar, Aruni; Hamburg, Naomi M

2018-06-14

Use of alternative tobacco products including electronic cigarettes is rapidly rising. The wide variety of flavored tobacco products available is of great appeal to smokers and youth. The flavorings added to tobacco products have been deemed safe for ingestion, but the cardiovascular health effects are unknown. The purpose of this study was to examine the effect of 9 flavors on vascular endothelial cell function. Freshly isolated endothelial cells from participants who use nonmenthol- or menthol-flavored tobacco cigarettes showed impaired A23187-stimulated nitric oxide production compared with endothelial cells from nonsmoking participants. Treatment of endothelial cells isolated from nonsmoking participants with either menthol (0.01 mmol/L) or eugenol (0.01 mmol/L) decreased A23187-stimulated nitric oxide production. To further evaluate the effects of flavoring compounds on endothelial cell phenotype, commercially available human aortic endothelial cells were incubated with vanillin, menthol, cinnamaldehyde, eugenol, dimethylpyrazine, diacetyl, isoamyl acetate, eucalyptol, and acetylpyrazine (0.1-100 mmol/L) for 90 minutes. Cell death, reactive oxygen species production, expression of the proinflammatory marker IL-6 (interleukin-6), and nitric oxide production were measured. Cell death and reactive oxygen species production were induced only at high concentrations unlikely to be achieved in vivo. Lower concentrations of selected flavors (vanillin, menthol, cinnamaldehyde, eugenol, and acetylpyridine) induced both inflammation and impaired A23187-stimulated nitric oxide production consistent with endothelial dysfunction. Our data suggest that short-term exposure of endothelial cells to flavoring compounds used in tobacco products have adverse effects on endothelial cell phenotype that may have relevance to cardiovascular toxicity. © 2018 American Heart Association, Inc.

Endothelial Dysfunction in Rheumatoid Arthritis: Mechanistic Insights and Correlation with Circulating Markers of Systemic Inflammation.

PubMed

Totoson, Perle; Maguin-Gaté, Katy; Nappey, Maude; Wendling, Daniel; Demougeot, Céline

2016-01-01

To determine mechanisms involved in endothelial dysfunction (ED) during the course of arthritis and to investigate the link between cytokines, chemokines and osteoprotegerin. Experiments were conducted on aortic rings at day 4 (preclinical), day 11 (onset of disease), day 33 (acute disease) and day 90 (chronic disease) after adjuvant-induced arthritis (AIA) in Lewis rats. At day 4, the unique vascular abnormality was a reduced norepinephrine-induced constriction. At day 11, endothelial function assessed by the relaxation to acetylcholine was normal despite increased cyclo-oxygenase-2 activity (COX-2) and overproduction of superoxide anions that was compensated by increased nitric oxide synthase (NOS) activity. At day 33, ED apparition coincides with the normalization of NOS activity. At day 90, ED was only observed in rats with a persisting imbalance between endothelial NOS and COX-2 pathways and higher plasma levels of IL-1β and TNFα. Plasma levels of IL-1β, TNFα and MIP-1α negatively correlated with Ach-induced relaxation throughout the course of AIA. Our data identified increased endothelial NOS activity as an important compensatory response that opposes the ED in the early arthritis. Thereafter, a cross-talk between endothelial COX-2/NOS pathways appears as an important element for the occurrence of ED. Our results encourage determining the clinical value of IL-1β, TNFα and MIP-1α as biomarkers of ED in RA.

Serum Markers of Endothelial Dysfunction and Inflammation Increase in Hypertension with Prediabetes Mellitus.

PubMed

Huang, Zhouqing; Chen, Chen; Li, Sheng; Kong, Fanqi; Shan, Peiren; Huang, Weijian

2016-06-01

The aim of this study was to examine endothelial dysfunction and inflammation in hypertension and prediabetes by studying adhesion molecules and inflammatory factors. This study included 133 outpatients. Participants were categorized into three groups based on the presence or absence of hypertension and prediabetes: control subjects without prediabetes and hypertension (N group, n = 39); patients with hypertension only (H group, n = 34); and patients with hypertension and prediabetes (HD group, n = 60). Hypertension was diagnosed according to JNC7 criteria. Prediabetes was defined according to 2010 American Diabetes Association criteria. Plasma was isolated from overnight fasting blood samples for enzyme-linked immunosorbent assay (ELISA) analysis of concentrations of intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor-α (TNF-α), P-selectin, and interleukin-6 (IL-6) as indicators of endothelial function and inflammation. We found that the H and HD groups showed significantly higher levels of all four biomarkers compared with the N group (all p < 0.01). The HD group also showed significantly higher levels of ICAM-1 (p = 0.042) and TNF-α (p < 0.01) compared with the H group; no significant differences in P-selectin (p = 0.59) and IL-6 (p = 0.70) levels were observed among these groups. Prediabetes and hypertension induce endothelial dysfunction and inflammation by elevating levels of soluble adhesion molecules and inflammatory cytokines. The comorbidity of these diseases may exacerbate inflammation and endothelial dysfunction by enhancing the expression of ICAM-1 and TNF-α.

Endothelial dysfunction, vascular disease and stroke: the ARTICO study.

PubMed

Roquer, J; Segura, T; Serena, J; Castillo, J

2009-01-01

Endothelial dysfunction is a fundamental step in the atherosclerotic disease process. Its presence is a risk factor for the development of clinical events, and may represent a marker of atherothrombotic burden. Also, endothelial dysfunction contributes to enhanced plaque vulnerability, may trigger plaque rupture, and favors thrombus formation. The assessment of endothelial vasomotion is a useful marker of atherosclerotic vascular disease. There are different methods to assess endothelial function: endothelium-dependent vasodilatation brachial flow-mediated dilation, cerebrovascular reactivity to L-arginine, and the determination of some biomarkers such as microalbuminuria, platelet function, and C-reactive protein. Endothelial dysfunction has been observed in stroke patients and has been related to stroke physiopathology, stroke subtypes, clinical severity and outcome. Resting ankle-brachial index (ABI) is also considered an indicator of generalized atherosclerosis, and a low ABI is associated with an increase in stroke incidence in the elderly. Despite all these data, there are no studies analyzing the predictive value of ABI for new cardiovascular events in patients after suffering an acute ischemic stroke. ARTICO is an ongoing prospective, observational, multicenter study being performed in 50 Spanish hospitals. The aim of the ARTICO study is to evaluate the prognostic value of a pathological ABI (

Impact of diabetic serum on endothelial cells: An in-vitro-analysis of endothelial dysfunction in diabetes mellitus type 2

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

Muenzel, Daniela; Lehle, Karla; Haubner, Frank

2007-10-19

Diabetic endothelial dysfunction was characterized by altered levels of adhesion molecules and cytokines. Aim of our study was to evaluate the effects of diabetic serum on cell-growth and proinflammatory markers in human saphenous vein endothelial cells (HSVEC) from diabetic and non-diabetic patients. Diabetic serum showed (1) complementary proliferative activity for non-diabetic and diabetic HSVEC, (2) unchanged surface expression of adhesion molecules, and (3) elevated levels of sICAM-1 in HSVEC of all donors. The concentration of sVCAM-1 was increased only in diabetic cells. The proinflammatory state of diabetic HSVEC characterized by increased levels of cytokines was compensated. We concluded that evenmore » under normoglycemic conditions the serum itself contains critical factors leading to abnormal regulation of inflammation in diabetics. We introduced an in vitro model of diabetes representing the endothelial situation at the beginning of diabetes (non-diabetic cells/diabetic serum) as well as the diabetic chronic state (diabetic cells/diabetic serum)« less

Transgene delivery to endothelial cultures derived from porcine carotid artery ex vivo.

PubMed

Andoh, J; Sawyer, B; Szewczyk, K; Nortley, M; Rossetti, T; Loftus, I M; Yáñez-Muñoz, R J; Hainsworth, A H

2013-10-01

Carotid artery disease is a widespread cause of morbidity and mortality. Porcine models of vascular disease are well established in vivo, but existing endothelial systems in vitro (e.g. human umbilical vein endothelial cells, rat aortic endothelial cultures) poorly reflect carotid endothelium. A reliable in vitro assay would improve design of in vivo experiments and allow reduction and refinement of animal use. This study aimed (1) to develop ex vivo endothelial cultures from porcine carotid and (2) to test whether these were suitable for lentivector-mediated transgene delivery. Surplus carotid arteries were harvested from young adult female Large White pigs within 10 min post-mortem. Small sectors of carotid artery wall (approximately 4 mm×4 mm squares) were immobilised in a stable gel matrix. Cultures were exposed to HIV-derived lentivector (LV) encoding a reporter transgene or the equivalent integration-deficient vector (IDLV). After 7-14 days in vitro, cultures were fixed and labelled histochemically. Thread-like multicellular outgrowths were observed that were positive for endothelial cell markers (CD31, VEGFR2, von Willebrand factor). A minority of cells co-labelled for smooth muscle markers. Sensitivity to cytotoxic agents (paclitaxel, cycloheximide, staurosporine) was comparable to that in cell cultures, indicating that the gel matrix permits diffusive access of small pharmacological molecules. Transgene-expressing cells were more abundant following exposure to LV than IDLV (4.7, 0.1% of cells, respectively). In conclusion, ex vivo adult porcine carotid artery produced endothelial cell outgrowths that were effectively transduced by LV. This system will facilitate translation of novel therapies to clinical trials, with reduction and refinement of in vivo experiments.

Characterisation of human induced pluripotent stem cell-derived endothelial cells under shear stress using an easy-to-use microfluidic cell culture system.

PubMed

Ohtani-Kaneko, Rsituko; Sato, Kenjiro; Tsutiya, Atsuhiro; Nakagawa, Yuka; Hashizume, Kazutoshi; Tazawa, Hidekatsu

2017-10-09

Induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) can contribute to elucidating the pathogenesis of heart and vascular diseases and developing their treatments. Their precise characteristics in fluid flow however remain unclear. Therefore, the aim of the present study is to characterise these features. We cultured three types of ECs in a microfluidic culture system: commercially available human iPS-ECs, human umbilical vein endothelial cells (HUVECs) and human umbilical artery endothelial cells (HUAECs). We then examined the mRNA expression levels of endothelial marker gene cluster of differentiation 31 (CD31), fit-related receptor tyrosine kinase (Flk-1), and the smooth muscle marker gene smooth muscle alpha-actin, and investigated changes in plasminogen activator inhibitor-1 (PAI-1) secretion and intracellular F-actin arrangement following heat stress. We also compared expressions of the arterial and venous marker genes ephrinB2 and EphB4, and the endothelial gap junction genes connexin (Cx) 37, 40, and 43 under fluidic shear stress to determine their arterial or venous characteristics. We found that iPS-ECs had similar endothelial marker gene expressions and exhibited similar increases in PAI-1 secretion under heat stress as HUVECs and HUAECs. In addition, F-actin arrangement in iPSC-ECs also responded to heat stress, as previously reported. However, they had different expression patterns of arterial and venous marker genes and Cx genes under different fluidic shear stress levels, showing that iPSC-ECs exhibit different characteristics from arterial and venous ECs. This microfluidic culture system equipped with variable shear stress control will provide an easy-to-use assay tool to examine characteristics of iPS-ECs generated by different protocols in various laboratories and contribute to basic and applied biomedical researches on iPS-ECs.

Early detection of endothelial injury and dysfunction in conjunction with correction of hemodynamic maladjustment can effectively restore renal function in type 2 diabetic nephropathy.

PubMed

Futrakul, Narisa; Butthep, Punnee; Vongthavarawat, Varaphon; Futrakul, Prasit; Sirisalipoch, Sasitorn; Chaivatanarat, Tawatchai; Suwanwalaikorn, Sompongse

2006-01-01

This paper was aimed to investigate (1) the early marker of endothelial injury in type 2 diabetes, (2) the intrarenal hemodynamics and renal function, and (3) the therapeutic strategy aiming to restore renal function. Fifty patients (35 normoalbuminuric and 15 albuminuric type 2 diabetes) were examined. Blood was collected for determination of circulating vascular endothelial cells (CEC) and the serum was prepared for determination of transforming growth factor beta (TGFbeta), ratio of CEC/TGFbeta, and soluble vascular cell adhesion molecule. Intrarenal hemodynamics and renal function were also assessed. The results showed that increased number of circulating EC, elevated TGFbeta and depleted ratio of CEC/TGFbeta were significantly observed. Intrarenal hemodynamic study revealed a hemodynamic maladjustment characterized by preferential constriction of the efferent arteriole, intraglomerular hypertension and reduction in peritubular capillary flow. It was concluded that early marker of endothelial injury is reflected by increasing number of CEC. Such markers correlate with the glomerular endothelial dysfunction associated with hemodynamic maladjustment. Early detection of endothelial injury and appropriate correction of hemodynamic maladjustment by multidrug vasodilators can effectively restore renal function in type 2 diabetic nephropathy.

Calcium dobesilate may alleviate diabetes-induced endothelial dysfunction and inflammation

PubMed Central

Zhou, Yijun; Yuan, Jiangzi; Qi, Chaojun; Shao, Xinghua; Mou, Shan; Ni, Zhaohui

2017-01-01

Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease. However, the pathogenesis of DKD remains unclear, and no effective treatments for the disease are available. Thus, there is an urgent need to elucidate the pathogenic mechanisms of DKD and to develop more effective therapies for this disease. Human umbilical vein endothelial cells (HUVECs) were cultured using different D-glucose concentrations to determine the effect of high glucose (HG) on the cells. Alternatively, HUVECs were incubated with 100 µmol/l calcium dobesilate (CaD) to detect its effects. The authors subsequently measured HUVEC proliferation via cell counting kit-8 assays. In addition, HUVEC angiogenesis was investigated via migration assays and fluorescein isothiocyanate (FITC)-labelled bovine serum albumin (BSA) permeability assays. The content or distribution of markers of endothelial dysfunction [vascular endothelial growth factor (VEGF), VEGF receptor (R) and endocan) or inflammation [intercellular adhesion molecule (ICAM)-1, monocyte chemotactic protein (MCP)-1 and pentraxin-related protein (PTX3)] was evaluated via reverse transcription-quantitative polymerase chain reaction and western blotting. HG treatment induced increased in VEGF, VEGFR, endocan, ICAM-1, MCP-1 and PTX3 mRNA and protein expression in HUVECs. HG treatment for 24 to 48 h increased cell proliferation in a time-dependent manner, but the cell proliferation rate was decreased at 72 h of HG treatment. Conversely, CaD inhibited abnormal cell proliferation. HG treatment also significantly enhanced HVUEC migration compared to the control treatment. In contrast, CaD treatment partially inhibited HUVEC migration compared to HG exposure. HG-treated HUVECs exhibited increased FITC-BSA permeability compared to control cells cultured in medium alone; however, CaD application prevented the HG-induced increase in FITC-BSA permeability and suppressed HG-induced overexpression of endothelial markers (VEGF, VEGFR-2

Weight loss improves biomarkers endothelial function and systemic inflammation in obese postmenopausal Saudi women.

PubMed

Abd El-Kader, Shehab Mahmoud; Saiem Al-Dahr, Mohammed H

2016-06-01

Although postmenopausal associated disorders are important public health problems worldwide, to date limited studies evaluated the endothelial function and systemic inflammation response to weight loss in obese postmenopausal women. This study was done to evaluate the endothelial function and systemic inflammation response to weight loss in obese postmenopausal Saudi women. Eighty postmenopausal obese Saudi women (mean age 52.64±6.13 year) participated in two groups: Group (A) received aerobic exercise on treadmill and diet whereas, group (B) received no intervention. Markers of inflammation and endothelial function were measured before and after 3 months at the end of the study. The values of body mass index(BMI), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), C-reactive protein (CRP), inter-cellular adhesion molecule (ICAM-1), vascular cell adhesion molecule (VCAM-1) and plasminogen activator inhibitor-1 activity (PAI-1:Ac) were significantly decreased in group (A), while changes were not significant in group (B). Also, there were significant differences between mean levels of the investigated parameters in group (A) and group (B) after treatment. Weight loss ameliorates inflammatory cytokines and markers of endothelial function in obese postmenopausal Saudi women.

Effect of Saxagliptin on Circulating Endothelial Progenitor Cells and Endothelial Function in Newly Diagnosed Type 2 Diabetic Patients.

PubMed

Li, Fang; Chen, Jiachao; Leng, Fei; Lu, Zhiqiang; Ling, Yan

2017-06-01

Endothelial dysfunction is associated with the risk of cardiovascular complications in diabetic patients. Endothelial progenitor cells (EPCs) and flow-mediated dilation (FMD) are common markers of endothelial function. In this study, we aim to investigate whether the DPP-4 inhibitor saxagliptin modulate EPCs number and FMD in newly diagnosed, treatment-naive type 2 diabetic patients. This was a controlled, randomized, open-label clinical trial. Saxagliptin group and metformin group consumed either saxagliptin 5 mg per day or metformin 1 500 mg per day respectively for 12 weeks. Changes of FMD and EPCs number after 12-week intervention were the primary endpoints. 31 patients were initially enrolled and randomized to saxagliptin group (n=16) and metformin group (n=15). 27 patients completed the trial (saxagliptin group n=14 and metformin group n=13), and 4 patients dropped out during the study. FMD and EPCs number increased significantly in both saxagliptin group and metformin group, and there was no significant difference between groups. 2-h postprandial plasma glucose, HbA1c and diastolic blood pressure improved significantly in both groups, and there was no significant difference between groups. Saxagliptin and metformin had comparable beneficial effects on endothelial function. © Georg Thieme Verlag KG Stuttgart · New York.

Endothelial microparticles and vascular parameters in subjects with and without arterial hypertension and coronary artery disease.

PubMed

Sansone, Roberto; Baaken, Maximilian; Horn, Patrick; Schuler, Dominik; Westenfeld, Ralf; Amabile, Nicolas; Kelm, Malte; Heiss, Christian

2018-08-01

Endothelial microparticles (EMPs) are markers of endothelial injury and activation. The role of EMPs in arterial hypertension is not well understood and EMPs are increased both in arterial hypertension and coronary artery disease (CAD). The data presented here show EMPs as defined by CD31 + /41 - , CD62e + , and CD144 + surface markers and vascular hemodynamic parameters including office and central blood pressure, heart rate, aortic augmentation index, pulse wave velocity, flow-mediated dilation, nitroglycerin-mediated dilation, brachial artery diameter, hyperemic wall shear stress, and laser Doppler perfusion of the cutaneous microcirculation of normotensives and hypertensives with and without CAD.

N-Acetylcysteine increases corneal endothelial cell survival in a mouse model of Fuchs endothelial corneal dystrophy.

PubMed

Kim, Eun Chul; Meng, Huan; Jun, Albert S

2014-10-01

The present study evaluated survival effects of N-acetylcysteine (NAC) on cultured corneal endothelial cells exposed to oxidative and endoplasmic reticulum (ER) stress and in a mouse model of early-onset Fuchs endothelial corneal dystrophy (FECD). Cultured bovine corneal endothelial cell viability against oxidative and ER stress was determined by CellTiter-Glo(®) luminescent reagent. Two-month-old homozygous knock-in Col8a2(L450W/L450W) mutant (L450W) and C57/Bl6 wild-type (WT) animals were divided into two groups of 15 mice. Group I received 7 mg/mL NAC in drinking water and Group II received control water for 7 months. Endothelial cell density and morphology were evaluated with confocal microscopy. Antioxidant gene (iNos) and ER stress/unfolded protein response gene (Grp78 and Chop) mRNA levels and protein expression were measured in corneal endothelium by real time PCR and Western blotting. Cell viability of H2O2 and thapsigargin exposed cells pre-treated with NAC was significantly increased compared to untreated controls (p < 0.01). Corneal endothelial cell density (CD) was higher (p = 0.001) and percent polymegathism was lower (p = 0.04) in NAC treated L450W mice than in untreated L450W mice. NAC treated L450W endothelium showed significant upregulation of iNos, whereas Grp78 and Chop were downregulated compared to untreated L450W endothelium by real time PCR and Western blotting. NAC increases survival in cultured corneal endothelial cells exposed against ER and oxidative stress. Systemic NAC ingestion increases corneal endothelial cell survival which is associated with increased antioxidant and decreased ER stress markers in a mouse model of early-onset FECD. Our study presents in vivo evidence of a novel potential medical treatment for FECD. Copyright © 2014 Elsevier Ltd. All rights reserved.

Endotoxin-Induced Endothelial Fibrosis Is Dependent on Expression of Transforming Growth Factors β1 and β2

PubMed Central

Echeverría, César; Montorfano, Ignacio; Tapia, Pablo; Riedel, Claudia; Cabello-Verrugio, Claudio

2014-01-01

During endotoxemia-induced inflammatory disease, bacterial endotoxins circulate in the bloodstream and interact with endothelial cells (ECs), inducing dysfunction of the ECs. We previously reported that endotoxins induce the conversion of ECs into activated fibroblasts. Through endotoxin-induced endothelial fibrosis, ECs change their morphology and their protein expression pattern, thereby suppressing endothelial markers and upregulating fibrotic proteins. The most commonly used fibrotic inducers are transforming growth factor β1 (TGF-β1) and TGF-β2. However, whether TGF-β1 and TGF-β2 participate in endotoxin-induced endothelial fibrosis remains unknown. We have shown that the endotoxin-induced endothelial fibrosis process is dependent on the TGF-β receptor, ALK5, and the activation of Smad3, a protein that is activated by ALK5 activation, thus suggesting that endotoxin elicits TGF-β production to mediate endotoxin-induced endothelial fibrosis. Therefore, we investigated the dependence of endotoxin-induced endothelial fibrosis on the expression of TGF-β1 and TGF-β2. Endotoxin-treated ECs induced the expression and secretion of TGF-β1 and TGF-β2. TGF-β1 and TGF-β2 downregulation inhibited the endotoxin-induced changes in the endothelial marker VE-cadherin and in the fibrotic proteins α-SMA and fibronectin. Thus, endotoxin induces the production of TGF-β1 and TGF-β2 as a mechanism to promote endotoxin-induced endothelial fibrosis. To the best of our knowledge, this is the first report showing that endotoxin induces endothelial fibrosis via TGF-β secretion, which represents an emerging source of vascular dysfunction. These findings contribute to understanding the molecular mechanism of endotoxin-induced endothelial fibrosis, which could be useful in the treatment of inflammatory diseases. PMID:24935972

Endotoxin-induced endothelial fibrosis is dependent on expression of transforming growth factors β1 and β2.

PubMed

Echeverría, César; Montorfano, Ignacio; Tapia, Pablo; Riedel, Claudia; Cabello-Verrugio, Claudio; Simon, Felipe

2014-09-01

During endotoxemia-induced inflammatory disease, bacterial endotoxins circulate in the bloodstream and interact with endothelial cells (ECs), inducing dysfunction of the ECs. We previously reported that endotoxins induce the conversion of ECs into activated fibroblasts. Through endotoxin-induced endothelial fibrosis, ECs change their morphology and their protein expression pattern, thereby suppressing endothelial markers and upregulating fibrotic proteins. The most commonly used fibrotic inducers are transforming growth factor β1 (TGF-β1) and TGF-β2. However, whether TGF-β1 and TGF-β2 participate in endotoxin-induced endothelial fibrosis remains unknown. We have shown that the endotoxin-induced endothelial fibrosis process is dependent on the TGF-β receptor, ALK5, and the activation of Smad3, a protein that is activated by ALK5 activation, thus suggesting that endotoxin elicits TGF-β production to mediate endotoxin-induced endothelial fibrosis. Therefore, we investigated the dependence of endotoxin-induced endothelial fibrosis on the expression of TGF-β1 and TGF-β2. Endotoxin-treated ECs induced the expression and secretion of TGF-β1 and TGF-β2. TGF-β1 and TGF-β2 downregulation inhibited the endotoxin-induced changes in the endothelial marker VE-cadherin and in the fibrotic proteins α-SMA and fibronectin. Thus, endotoxin induces the production of TGF-β1 and TGF-β2 as a mechanism to promote endotoxin-induced endothelial fibrosis. To the best of our knowledge, this is the first report showing that endotoxin induces endothelial fibrosis via TGF-β secretion, which represents an emerging source of vascular dysfunction. These findings contribute to understanding the molecular mechanism of endotoxin-induced endothelial fibrosis, which could be useful in the treatment of inflammatory diseases. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Glioblastoma stem cell differentiation into endothelial cells evidenced through live-cell imaging.

PubMed

Mei, Xin; Chen, Yin-Sheng; Chen, Fu-Rong; Xi, Shao-Yan; Chen, Zhong-Ping

2017-08-01

Glioblastoma cell-initiated vascularization is an alternative angiogenesis called vasculogenic mimicry. However, current knowledge on the mechanism of de novo vessel formation from glioblastoma stem cells (GSCs) is limited. Sixty-four glioblastoma samples from patients and 10 fluorescent glioma xenograft samples were examined by immunofluorescence staining for endothelial marker (CD34 and CD31) and glial cell marker (glial fibrillary acidic protein [GFAP]) expression. GSCs were then isolated from human glioblastoma tissue and CD133+/Sox2+ red fluorescent protein-containing (RFP)-GSC-1 cells were established. The ability of these cells to form vascular structures was examined by live-cell imaging of 3D cultures. CD34-GFAP or CD31-GFAP coexpressing glioblastoma-derived endothelial cells (GDEC) were found in 30 of 64 (46.9%) of clinical glioblastoma samples. In those 30 samples, GDEC were found to form vessel structures in 21 (70%) samples. Among 21 samples with GDEC vessels, the CD34+ GDEC vessels and CD31+ GDEC vessels accounted for about 14.16% and 18.08% of total vessels, respectively. In the xenograft samples, CD34+ GDEC were found in 7 out of 10 mice, and 4 out of 7 mice had CD34+ GDEC vessels. CD31+ GDEC were also found in 7 mice, and 4 mice had CD31+ GDEC vessels (10 mice in total). Through live-cell imaging, we observed gradual CD34 expression when cultured with vascular endothelial growth factor in some glioma cells, and a dynamic increase in endothelial marker expression in RFP-GSC-1 in vitro was recorded. Cells expressed CD34 (9.46%) after 6 hours in culture. The results demonstrated that GSCs may differentiate into endothelial cells and promote angiogenesis in glioblastomas. © The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com

Inflammation, Endothelial Dysfunction and Arterial Stiffness as Therapeutic Targets in Cardiovascular Medicine.

PubMed

Della Corte, Vittoriano; Tuttolomondo, Antonino; Pecoraro, Rosaria; Di Raimondo, Domenico; Vassallo, Valerio; Pinto, Antonio

2016-01-01

In the last decades, many factors thought to be associated with the atherosclerotic process and cardiovascular events have been studied, and some of these have been shown to correlate with clinical outcome, such as arterial stiffness, endothelial dysfunction and immunoinflammatory markers. Arterial stiffness is an important surrogate marker that describes the capability of an artery to expand and contract in response to pressure changes. It can be assessed with different techniques, such as the evaluation of PWV and AIx. It is related to central systolic pressure and it is an independent predictor of cardiovascular morbidity and mortality in hypertensive patients, type 2 diabetes, end-stage renal disease and in elderly populations. The endothelium has emerged as the key regulator of vascular homeostasis, in fact, it has not merely a barrier function but also acts as an active signal transducer for circulating influences that modify the vessel wall phenotype. When its function is lost, it predisposes the vasculature to vasoconstriction, leukocyte adherence, platelet activation, thrombosis and atherosclerosis. Non-invasive methods were developed to evaluate endothelial function, such as the assesment of FMD, L-FMC and RHI. Moreover in the last years, a large number of studies have clarified the role of inflammation and the underlying cellular and molecular mechanisms that contribute to atherogenesis. For clinical purposes, the most promising inflammatory biomarker appears to be CRP and a variety of population-based studies have showed that baseline CRP levels predict future cardiovascular events. Each of the markers listed above has its importance from the pathophysiological and clinical point of view, and those can also be good therapeutic targets. However, it must be stressed that assessments of these vascular markers are not mutually exclusive, but rather complementary and those can offer different views of the same pathology. The purpose of this review is to

Identification of myogenic-endothelial progenitor cells in the interstitial spaces of skeletal muscle.

PubMed

Tamaki, Tetsuro; Akatsuka, Akira; Ando, Kiyoshi; Nakamura, Yoshihiko; Matsuzawa, Hideyuki; Hotta, Tomomitsu; Roy, Roland R; Edgerton, V Reggie

2002-05-13

Putative myogenic and endothelial (myo-endothelial) cell progenitors were identified in the interstitial spaces of murine skeletal muscle by immunohistochemistry and immunoelectron microscopy using CD34 antigen. Enzymatically isolated cells were characterized by fluorescence-activated cell sorting on the basis of cell surface antigen expression, and were sorted as a CD34+ and CD45- fraction. Cells in this fraction were approximately 94% positive for Sca-1, and mostly negative (<3% positive) for CD14, 31, 49, 144, c-kit, and FLK-1. The CD34+/45- cells formed colonies in clonal cell cultures and colony-forming units displayed the potential to differentiate into adipocytes, endothelial, and myogenic cells. The CD34+/45- cells fully differentiated into vascular endothelial cells and skeletal muscle fibers in vivo after transplantation. Immediately after sorting, CD34+/45- cells expressed only c-met mRNA, and did not express any other myogenic cell-related markers such as MyoD, myf-5, myf-6, myogenin, M-cadherin, Pax-3, and Pax-7. However, after 3 d of culture, these cells expressed mRNA for all myogenic markers. CD34+/45- cells were distinct from satellite cells, as they expressed Bcrp1/ABCG2 gene mRNA (Zhou et al., 2001). These findings suggest that myo-endothelial progenitors reside in the interstitial spaces of mammalian skeletal muscles, and that they can potentially contribute to postnatal skeletal muscle growth.

Endothelial progenitor cells bind and inhibit platelet function and thrombus formation.

PubMed

Abou-Saleh, Haissam; Yacoub, Daniel; Théorêt, Jean-François; Gillis, Marc-Antoine; Neagoe, Paul-Eduard; Labarthe, Benoit; Théroux, Pierre; Sirois, Martin G; Tabrizian, Maryam; Thorin, Eric; Merhi, Yahye

2009-12-01

Interactions of endothelial progenitor cells (EPCs) with vascular and blood cells contribute to vascular homeostasis. Although platelets promote the homing of EPCs to sites of vascular injury and their differentiation into endothelial cells, the functional consequences of such interactions on platelets remain unknown. Herein, we addressed the interactions between EPCs and platelets and their impact on platelet function and thrombus formation. Cultured on fibronectin in conditioned media, human peripheral blood mononuclear cells differentiated, within 10 days of culture, into EPCs, which uptake acetylated low-density lipoprotein, bind ulex-lectin, lack monocyte/leukocyte markers (CD14, P-selectin glycoprotein ligand-1, L-selectin), express progenitor/endothelial markers (CD34, vascular endothelial growth factor receptor-2, von Willebrand factor, and vascular endothelial cadherin), and proliferate in culture. These EPCs bound activated platelets via CD62P and inhibited its translocation, glycoprotein IIb/IIIa activation, aggregation, and adhesion to collagen, mainly via prostacyclin secretion. Indeed, this was associated with upregulation of cyclooxygenase-2 and inducible nitric oxide synthase. However, the effects on platelets in vitro were reversed by cyclooxygenase and cyclooxygenase-2 inhibition but not by nitric oxide or inducible nitric oxide synthase inhibition. Moreover, in a ferric chloride-induced murine arterial thrombosis model, injection of EPCs led to their incorporation into sites of injury and impaired thrombus formation, leading to an incomplete occlusion with 50% residual flow. Peripheral blood mononuclear cell-derived EPCs bind platelets via CD62P and inhibit platelet activation, aggregation, adhesion to collagen, and thrombus formation, predominantly via upregulation of cyclooxygenase-2 and secretion of prostacyclin. These findings add new insights into the biology of EPCs and define their potential roles in regulating platelet function and

Biomaterials trigger endothelial cell activation when co-incubated with human whole blood.

PubMed

Herklotz, Manuela; Hanke, Jasmin; Hänsel, Stefanie; Drichel, Juliane; Marx, Monique; Maitz, Manfred F; Werner, Carsten

2016-10-01

Endothelial cell activation resulting from biomaterial contact or biomaterial-induced blood activation may in turn also affect hemostasis and inflammatory processes in the blood. Current in vitro hemocompatibility assays typically ignore these modulating effects of the endothelium. This study describes a co-incubation system of human whole blood, biomaterial and endothelial cells (ECs) that was developed to overcome this limitation. First, human endothelial cells were characterized in terms of their expression of coagulation- and inflammation-relevant markers in response to various activators. Subsequently, their capacity to regulate hemostasis as well as complement and granulocyte activation was monitored in a hemocompatibility assay. After blood contact, quiescent ECs exhibited anticoagulant and anti-inflammatory properties. When they were co-incubated with surfaces exhibiting pro-coagulant or pro-inflammatory characteristics, the ECs down-regulated coagulation but not complement or leukocyte activation. Analysis of intracellular levels of the endothelial activation markers E-selectin and tissue factor showed that co-incubation with model surfaces and blood significantly increased the activation state of ECs. Finally, the coagulation- and inflammation-modulating properties of the ECs were tested after blood/biomaterial exposure. Pre-activation of ECs by biomaterials in the blood induced a pro-coagulant and pro-inflammatory state of the ECs, wherein the pro-coagulant response was higher for biomaterial/blood pre-activated ECs than for TNF-α-pre-activated cells. This work provides evidence that biomaterials, even without directly contacting the endothelium, affect the endothelial activation state with and have consequences for plasmatic and cellular reactions in the blood. Copyright © 2016 Elsevier Ltd. All rights reserved.

Role of glutathione biosynthesis in endothelial dysfunction and fibrosis.

PubMed

Espinosa-Díez, Cristina; Miguel, Verónica; Vallejo, Susana; Sánchez, Francisco J; Sandoval, Elena; Blanco, Eva; Cannata, Pablo; Peiró, Concepción; Sánchez-Ferrer, Carlos F; Lamas, Santiago

2018-04-01

Glutathione (GSH) biosynthesis is essential for cellular redox homeostasis and antioxidant defense. The rate-limiting step requires glutamate-cysteine ligase (GCL), which is composed of the catalytic (GCLc) and the modulatory (GCLm) subunits. To evaluate the contribution of GCLc to endothelial function we generated an endothelial-specific Gclc haplo-insufficient mouse model (Gclc e/+ mice). In murine lung endothelial cells (MLEC) derived from these mice we observed a 50% reduction in GCLc levels compared to lung fibroblasts from the same mice. MLEC obtained from haplo-insufficient mice showed significant reduction in GSH levels as well as increased basal and stimulated ROS levels, reduced phosphorylation of eNOS (Ser 1177) and increased eNOS S-glutathionylation, compared to MLEC from wild type (WT) mice. Studies in mesenteric arteries demonstrated impaired endothelium-dependent vasodilation in Gclc(e/+) male mice, which was corrected by pre-incubation with GSH-ethyl-ester and BH 4 . To study the contribution of endothelial GSH synthesis to renal fibrosis we employed the unilateral ureteral obstruction model in WT and Gclc(e/+) mice. We observed that obstructed kidneys from Gclc(e/+) mice exhibited increased deposition of fibrotic markers and reduced Nrf2 levels. We conclude that the preservation of endothelial GSH biosynthesis is not only critical for endothelial function but also in anti-fibrotic responses. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Calcium supplementation prevents endothelial cell activation: possible relevance to preeclampsia.

PubMed

Chen, Qi; Tong, Mancy; Wu, Man; Stone, Peter R; Snowise, Saul; Chamley, Lawrence W

2013-09-01

Preeclampsia is a leading cause of maternal and fetal mortality and morbidity. A hallmark of preeclampsia is endothelial cell dysfunction/activation in response to 'toxins' from the placenta. Necrotic trophoblastic debris (NTD) is one possible placental toxin and other activators of endothelial cells include inflammatory cytokines. Calcium supplementation appears to protect 'at-risk' women from developing preeclampsia but how is unclear. Placental explants were cultured with interleukin-6 (IL-6) in varied concentrations of calcium. The resultant trophoblastic debris was exposed to endothelial cells. Endothelial cells were exposed to activators including NTD, IL-6, and preeclamptic sera in the presence of varied concentrations of calcium and activation monitored by quantifying cell surface markers by ELISA. Raising the levels of calcium did not prevent the IL-6-induced shedding of NTD from placental explants but did prevent the activation of endothelial cells in response to IL-6, preeclamptic sera, or NTD. Reducing the level of calcium directly induced the activation of endothelial cells. Inhibiting nitric oxide synthetase ablated the ability of high calcium levels to protect endothelial cell activation. The activity of endothelial cell nitric oxide synthetase was blocked with L-N-nitroarginine methyl ester. Our results demonstrate calcium levels do not affect the shedding of trophoblastic debris but are important to endothelial cell activation and supplemental calcium may reverse the activation of the endothelium in preeclamptic women. These results may in part explain the benefits of calcium supplementation in the reduction of risk for developing preeclampsia and provide in-vitro mechanistic support for the use of calcium supplementation in at-risk women.

Effect of exercise intensity on postprandial lipemia, markers of oxidative stress, and endothelial function after a high-fat meal.

PubMed

Lopes Krüger, Renata; Costa Teixeira, Bruno; Boufleur Farinha, Juliano; Cauduro Oliveira Macedo, Rodrigo; Pinto Boeno, Francesco; Rech, Anderson; Lopez, Pedro; Silveira Pinto, Ronei; Reischak-Oliveira, Alvaro

2016-12-01

The aim of this study was to compare the effects of 2 different exercise intensities on postprandial lipemia, oxidative stress markers, and endothelial function after a high-fat meal (HFM). Eleven young men completed 2-day trials in 3 conditions: rest, moderate-intensity exercise (MI-Exercise) and heavy-intensity exercise (HI-Exercise). Subjects performed an exercise bout or no exercise (Rest) on the evening of day 1. On the morning of day 2, an HFM was provided. Blood was sampled at fasting (0 h) and every hour from 1 to 5 h during the postprandial period for triacylglycerol (TAG), thiobarbituric acid reactive substance (TBARS), and nitrite/nitrate (NOx) concentrations. Flow-mediated dilatation (FMD) was also analyzed. TAG concentrations were reduced in exercise conditions compared with Rest during the postprandial period (P < 0.004). TAG incremental area under the curve (iAUC) was smaller after HI-Exercise compared with Rest (P = 0.012). TBARS concentrations were reduced in MI-Exercise compared with Rest (P < 0.041). FMD was higher in exercise conditions than Rest at 0 h (P < 0.02) and NOx concentrations were enhanced in MI-Exercise compared with Rest at 0 h (P < 0.01). These results suggest that acute exercise can reduce lipemia after an HFM. However, HI-Exercise showed to be more effective in reducing iAUC TAG, which might suggest higher protection against postprandial TAG enhancement. Conversely, MI-Exercise can be beneficial to attenuate the susceptibility of oxidative damage induced by an HFM and to increase endothelial function in the fasted state compared with Rest.

Inducible Knockdown of Endothelial Protein Tyrosine Phosphatase-1B Promotes Neointima Formation in Obese Mice by Enhancing Endothelial Senescence.

PubMed

Jäger, Marianne; Hubert, Astrid; Gogiraju, Rajinikanth; Bochenek, Magdalena L; Münzel, Thomas; Schäfer, Katrin

2018-02-01

Protein tyrosine phosphatase-1B (PTP1B) is a negative regulator of receptor tyrosine kinase signaling. In this study, we determined the importance of PTP1B expressed in endothelial cells for the vascular response to arterial injury in obesity. Morphometric analysis of vascular lesions generated by 10% ferric chloride (FeCl 3 ) revealed that tamoxifen-inducible endothelial PTP1B deletion (Tie2.ER T2 -Cre × PTP1B fl/fl ; End.PTP1B knockout, KO) significantly increased neointima formation, and reduced numbers of (endothelial lectin-positive) luminal cells in End.PTP1B-KO mice suggested impaired lesion re-endothelialization. Significantly higher numbers of proliferating cell nuclear antigen (PCNA)-positive proliferating cells as well as smooth muscle actin (SMA)-positive or vascular cell adhesion molecule-1 (VCAM1)-positive activated smooth muscle cells or vimentin-positive myofibroblasts were detected in neointimal lesions of End.PTP1B-KO mice, whereas F4/80-positive macrophage numbers did not differ. Activated receptor tyrosine kinase and transforming growth factor-beta (TGFβ) signaling and oxidative stress markers were also significantly more abundant in End.PTP1B-KO mouse lesions. Genetic knockdown or pharmacological inhibition of PTP1B in endothelial cells resulted in increased expression of caveolin-1 and oxidative stress, and distinct morphological changes, elevated numbers of senescence-associated β-galactosidase-positive cells, and increased expression of tumor suppressor protein 53 (p53) or the cell cycle inhibitor cyclin-dependent kinase inhibitor-2A (p16INK4A) suggested senescence, all of which could be attenuated by small interfering RNA (siRNA)-mediated downregulation of caveolin-1. In vitro, senescence could be prevented and impaired re-endothelialization restored by preincubation with the antioxidant Trolox. Our results reveal a previously unknown role of PTP1B in endothelial cells and provide mechanistic insights how PTP1B deletion or inhibition

Extraembryonic origin of circulating endothelial cells.

PubMed

Pardanaud, Luc; Eichmann, Anne

2011-01-01

Circulating endothelial cells (CEC) are contained in the bone marrow and peripheral blood of adult humans and participate to the revascularization of ischemic tissues. These cells represent attractive targets for cell or gene therapy aimed at improving ischemic revascularization or inhibition of tumor angiogenesis. The embryonic origin of CEC has not been addressed previously. Here we use quail-chick chimeras to study CEC origin and participation to the developing vasculature. CEC are traced with different markers, in particular the QH1 antibody recognizing only quail endothelial cells. Using yolk-sac chimeras, where quail embryos are grafted onto chick yolk sacs and vice-versa, we show that CEC are generated in the yolk sac. These cells are mobilized during wound healing, demonstrating their participation to angiogenic repair processes. Furthermore, we found that the allantois is also able to give rise to CEC in situ. In contrast to the yolk sac and allantois, the embryo proper does not produce CEC. Our results show that CEC exclusively originate from extra-embryonic territories made with splanchnopleural mesoderm and endoderm, while definitive hematopoietic stem cells and endothelial cells are of intra-embryonic origin.

Corona sign: manifestation of peripheral corneal epithelial edema as a possible marker of the progression of corneal endothelial dysfunction.

PubMed

Inoue, Tomoyuki; Hara, Yuko; Kobayashi, Takeshi; Zheng, Xiaodong; Suzuki, Takashi; Shiraishi, Atsushi; Ohashi, Yuichi

2016-09-01

To describe a characteristic form of the corona sign and its clinical relevance to the degree of corneal endothelial decompensation and investigate the underlying mechanism using a rabbit model. These observational cases include 31 patients undergoing penetrating keratoplasty (PKP) and 15 patients undergoing Descemet stripping automated endothelial keratoplasty (DSAEK) with special attention to the circumferentially developed corneal epithelial edema. We also conducted a laboratory observation of horizontal water flow in the rabbit cornea. We consistently observed the corona sign at the superior periphery during the initial stage of corneal endothelial decompensation after PKP. With progressive corneal endothelial cellular loss, the epithelial edema gradually expanded circumferentially in the periphery. The endothelial cellular density associated with the corona sign significantly (P < 0.01) decreased compared with that without the sign. The endothelial cellular density decreased significantly (P < 0.05) in cases with a circumferential corona sign compared with a superior corona sign. After DSAEK, however, the corneal epithelial edema subsided from the center but persisted peripherally as a corona sign in all cases. By 3 months postoperatively, the epithelial edema was confined to the superior periphery along with uneventful corneal endothelial healing. Rabbit experiments showed that total corneal endothelial decompensation decreased the horizontal intracorneal water migration (Inoue-Ohashi phenomenon) in the corneal periphery and induced peripheral corneal edema. The slit-lamp microscopic findings of the corona-like epithelial edema in the peripheral cornea are associated with the stage of corneal endothelial function. To support this, the developmental mechanism of the corona sign was demonstrated experimentally.

Origins of endothelial and osteogenic cells in the subcutaneous collagen gel implant.

PubMed

Bilic-Curcic, I; Kalajzic, Z; Wang, L; Rowe, D W

2005-11-01

The interdependent relationship between vascular endothelial cells and osteoblasts during bone formation and fracture healing has been long appreciated. This paper reports a heterotopic implant model using FGF-2-expanded bone marrow stromal cells (BMSC) derived from Tie2eGFP (endothelial marker) and pOBCol3.6GFPcyan or topaz (early osteoblast marker) transgenic mice to appreciate the host/donor relationships of cells participating in the process of heterotopic bone formation. The study included various combinations of Tie2eGFP and pOBCol3.6GFPcyan and topaz transgenics as BMSC or whole bone marrow (WBM) donors and also as recipients. Rat tail collagen was used as a carrier of donor cells and implantation was done in lethally irradiated mice rescued with WBM injection. Development of ossicles in the implants was followed weekly during the 4- to 5-week long post-implantation period. By 4-5 weeks after total body irradiation (TBI) and implantation, a well-formed bone spicule had developed that was invested with bone marrow. Experiments showed absolute dominance of donor-derived cells in the formation of endothelial-lined vessels inside the implants as well as the marrow stromal-derived osteogenic cells. Host-derived fibroblasts and osteogenic cells were confined to the fibrous capsule surrounding the implant. In addition, cells lining the endosteal surface of newly formed marrow space carrying a pOBCol3.6GFP marker were observed that were contributed by WBM donor cells and the host. Thus, FGF-2-expanded BMSC appear to be a source of endothelial and osteogenic progenitor cells capable of eliciting heterotopic bone formation independent of cells from the host. This model should be useful for understanding the interactions between these two cell types that control osteogenic differentiation in vivo.

Circulating endothelial progenitor cells in obese children and adolescents.

PubMed

Pires, António; Martins, Paula; Paiva, Artur; Pereira, Ana Margarida; Marques, Margarida; Castela, Eduardo; Sena, Cristina; Seiça, Raquel

2015-01-01

This study aimed to investigate the relationship between circulating endothelial progenitor cell count and endothelial activation in a pediatric population with obesity. Observational and transversal study, including 120 children and adolescents with primary obesity of both sexes, aged 6-17 years, who were recruited at this Cardiovascular Risk Clinic. The control group was made up of 41 children and adolescents with normal body mass index. The variables analyzed were: age, gender, body mass index, systolic and diastolic blood pressure, high-sensitivity C-reactive protein, lipid profile, leptin, adiponectin, homeostasis model assessment-insulin resistance, monocyte chemoattractant protein-1, E-selectin, asymmetric dimethylarginine and circulating progenitor endothelial cell count. Insulin resistance was correlated to asymmetric dimethylarginine (ρ=0.340; p=0.003), which was directly, but weakly correlated to E-selectin (ρ=0.252; p=0.046). High sensitivity C-reactive protein was not found to be correlated to markers of endothelial activation. Systolic blood pressure was directly correlated to body mass index (ρ=0.471; p<0.001) and the homeostasis model assessment-insulin resistance (ρ=0.230; p=0.012), and inversely correlated to adiponectin (ρ=-0.331; p<0.001) and high-density lipoprotein cholesterol (ρ=-0.319; p<0.001). Circulating endothelial progenitor cell count was directly, but weakly correlated, to body mass index (r=0.211; p=0.016), leptin (ρ=0.245; p=0.006), triglyceride levels (r=0.241; p=0.031), and E-selectin (ρ=0.297; p=0.004). Circulating endothelial progenitor cell count is elevated in obese children and adolescents with evidence of endothelial activation, suggesting that, during infancy, endothelial repairing mechanisms are present in the context of endothelial activation. Copyright © 2015 Sociedade Brasileira de Pediatria. Published by Elsevier Editora Ltda. All rights reserved.

Characterization of a Distinct Population of Circulating Human Non-Adherent Endothelial Forming Cells and Their Recruitment via Intercellular Adhesion Molecule-3

PubMed Central

Thompson, Emma J.; Barrett, Jeffrey M.; Tooley, Katie; Sen, Shaundeep; Sun, Wai Yan; Grose, Randall; Nicholson, Ian; Levina, Vitalina; Cooke, Ira; Talbo, Gert; Lopez, Angel F.; Bonder, Claudine S.

2012-01-01

Circulating vascular progenitor cells contribute to the pathological vasculogenesis of cancer whilst on the other hand offer much promise in therapeutic revascularization in post-occlusion intervention in cardiovascular disease. However, their characterization has been hampered by the many variables to produce them as well as their described phenotypic and functional heterogeneity. Herein we have isolated, enriched for and then characterized a human umbilical cord blood derived CD133+ population of non-adherent endothelial forming cells (naEFCs) which expressed the hematopoietic progenitor cell markers (CD133, CD34, CD117, CD90 and CD38) together with mature endothelial cell markers (VEGFR2, CD144 and CD31). These cells also expressed low levels of CD45 but did not express the lymphoid markers (CD3, CD4, CD8) or myeloid markers (CD11b and CD14) which distinguishes them from ‘early’ endothelial progenitor cells (EPCs). Functional studies demonstrated that these naEFCs (i) bound Ulex europaeus lectin, (ii) demonstrated acetylated-low density lipoprotein uptake, (iii) increased vascular cell adhesion molecule (VCAM-1) surface expression in response to tumor necrosis factor and (iv) in co-culture with mature endothelial cells increased the number of tubes, tubule branching and loops in a 3-dimensional in vitro matrix. More importantly, naEFCs placed in vivo generated new lumen containing vasculature lined by CD144 expressing human endothelial cells (ECs). Extensive genomic and proteomic analyses of the naEFCs showed that intercellular adhesion molecule (ICAM)-3 is expressed on their cell surface but not on mature endothelial cells. Furthermore, functional analysis demonstrated that ICAM-3 mediated the rolling and adhesive events of the naEFCs under shear stress. We suggest that the distinct population of naEFCs identified and characterized here represents a new valuable therapeutic target to control aberrant vasculogenesis. PMID:23144795

The Methods and Mechanisms to Differentiate Endothelial-Like Cells and Smooth Muscle Cells from Mesenchymal Stem Cells for Vascularization in Vaginal Reconstruction.

PubMed

Zhang, Hua; Zhang, Jingkun; Huang, Xianghua; Li, Yanan

2018-06-01

Endothelial cells and smooth muscle cells (SMCs) are important aspects of vascularization in vaginal reconstruction. Research has confirmed that mesenchymal stem cells could differentiate into endothelial-like cells and SMCs. But the methods were more complicated and the mechanism was unknown. In the current study, we induced the bone mesenchymal stem cells (BMSCs) to differentiate into endothelial-like cells and SMCs in vitro by differentiation medium and investigated the effect of Wnt/β-catenin signaling on the differentiation process of BMSCs. Results showed that the hypoxic environment combined with VEGF and bFGF could induce increased expression of endothelial-like cells markers VEGFR1, VEGFR2, and vWF. The SMCs derived from BMSCs induced by TGF-β1 and PDGF-AB significantly expressed SMC markers SMMHC11 and α-SMA. The data also showed that activation of Wnt/β-catenin signaling could promote the differentiation of BMSCs into endothelial-like cells and SMCs. Thus, we established endothelial-like cells and SMCs in vitro by more simple methods, presented the important role of hypoxic environment on the differentiation of BMSCs into endothelial-like cells, and confirmed that the Wnt/β-catenin signaling pathway has a positive impact on the differentiation of BMSCs into endothelial-like cells and SMCs. This is important for vascular reconstruction.

[Association of age, inflammatory markers and subclinical atherosclerosis in subjects free from cardiovascular disease].

PubMed

Páramo, José A; Orbe, Josune; Beloqui, Oscar; Colina, Inmaculada; Benito, Alberto; Rodríguez, José A; Díez, Javier

2008-09-27

We assessed whether an independent association between inflammatory markers and age-related subclinical atherosclerosis could be found in subjects free from cardiovascular disease. Metabolic parameters, inflammatory and endothelial markers, such as high-sensitivity C-reactive protein, interleukin-6, fibrinogen and von Willebrand factor, as well as the carotid intima-media thickness were assessed in 890 asymptomatic subjects (mean age: 55 years; range: 20-80 years; 80% men) with cardiovascular risk factors. Subjects in the upper quartile (age 61-80 years) showed a significant increase of traditional risk factors, particularly arterial pressure and glucose levels (p < 0.01) as compared with lower quartiles. We also found a significant increase in the levels on inflammatory and endothelial markers (p < 0.001) and intima-media thickness (p < 0.001) in older adults. In the multivarate analysis, after adjustment for cardiovascular risk factors, intima-media thickness was independently associated with inflammation and endothelial dysfunction in older adults (p < 0.01). Besides age, systemic inflammation and vascular damage are associated with subclinical atherosclerosis in asymptomatic subjects. The age-related inflammatory profile may predispose to cardiovascular complications.

Endothelial Progenitor Cells Bind and Inhibit Platelet Function and Thrombus Formation

PubMed Central

Abou-Saleh, Haissam; Yacoub, Daniel; Théorêt, Jean-François; Gillis, Marc-Antoine; Neagoe, Paul-Eduard; Labarthe, Benoit; Théroux, Pierre; Sirois, Martin G.; Tabrizian, Maryam; Thorin, Eric; Merhi, Yahye

2013-01-01

Background Interactions of endothelial progenitor cells (EPCs) with vascular and blood cells contribute to vascular homeostasis. Although platelets promote the homing of EPCs to sites of vascular injury and their differentiation into endothelial cells, the functional consequences of such interactions on platelets remain unknown. Herein, we addressed the interactions between EPCs and platelets and their impact on platelet function and thrombus formation. Methods and Results Cultured on fibronectin in conditioned media, human peripheral blood mononuclear cells differentiated, within 10 days of culture, into EPCs, which uptake acetylated low-density lipoprotein, bind ulex-lectin, lack monocyte/leukocyte markers (CD14, P-selectin glycoprotein ligand-1, L-selectin), express progenitor/endothelial markers (CD34, vascular endothelial growth factor receptor-2, von Willebrand factor, and vascular endothelial cadherin), and proliferate in culture. These EPCs bound activated platelets via CD62P and inhibited its translocation, glycoprotein IIb/IIIa activation, aggregation, and adhesion to collagen, mainly via prostacyclin secretion. Indeed, this was associated with upregulation of cyclooxygenase-2 and inducible nitric oxide synthase. However, the effects on platelets in vitro were reversed by cyclooxygenase and cyclooxygenase-2 inhibition but not by nitric oxide or inducible nitric oxide synthase inhibition. Moreover, in a ferric chloride–induced murine arterial thrombosis model, injection of EPCs led to their incorporation into sites of injury and impaired thrombus formation, leading to an incomplete occlusion with 50% residual flow. Conclusions Peripheral blood mononuclear cell– derived EPCs bind platelets via CD62P and inhibit platelet activation, aggregation, adhesion to collagen, and thrombus formation, predominantly via upregulation of cyclooxygenase-2 and secretion of prostacyclin. These findings add new insights into the biology of EPCs and define their potential

Thrombomodulin as a marker for vascular tumors. Comparative study with factor VIII and Ulex europaeus I lectin.

PubMed

Yonezawa, S; Maruyama, I; Sakae, K; Igata, A; Majerus, P W; Sato, E

1987-10-01

Thrombomodulin (TM) is a newly described endothelial cell-associated protein that functions as a potent natural anticoagulant by converting thrombin from a procoagulant protease to an anticoagulant. Various vascular tumors were characterized with immunoperoxidase staining with the use of a polyclonal anti-TM serum. The staining patterns of TM were compared with those of Factor VIII-related antigen (FVIII-RAG) and Ulex europaeus agglutinin-I (UEA-I), which have been used as markers for endothelial cells. The results showed that TM is a specific and a highly sensitive marker for angiosarcomas in comparison with FVIII-RAG or UEA-I. In contrast, UEA-I is more sensitive for benign vascular tumors than TM or FVIII-RAG. The other mesenchymal tumors of nonvascular origin showed negative staining for three endothelial markers. These results indicate that TM is a new specific and sensitive tool for the diagnosis of angiosarcomas.

Low oxygen tension enhances endothelial fate of human pluripotent stem cells.

PubMed

Kusuma, Sravanti; Peijnenburg, Elizabeth; Patel, Parth; Gerecht, Sharon

2014-04-01

A critical regulator of the developing or regenerating vasculature is low oxygen tension. Precise elucidation of the role of low oxygen environments on endothelial commitment from human pluripotent stem cells necessitates controlled in vitro differentiation environments. We used a feeder-free, 2-dimensional differentiation system in which we could monitor accurately dissolved oxygen levels during human pluripotent stem cell differentiation toward early vascular cells (EVCs). We found that oxygen uptake rate of differentiating human pluripotent stem cells is lower in 5% O2 compared with atmospheric conditions. EVCs differentiated in 5% O2 had an increased vascular endothelial cadherin expression with clusters of vascular endothelial cadherin+ cells surrounded by platelet-derived growth factor β+ cells. When we assessed the temporal effects of low oxygen differentiation environments, we determined that low oxygen environments during the early stages of EVC differentiation enhance endothelial lineage commitment. EVCs differentiated in 5% O2 exhibited an increased expression of vascular endothelial cadherin and CD31 along with their localization to the membrane, enhanced lectin binding and acetylated low-density lipoprotein uptake, rapid cord-like structure formation, and increased expression of arterial endothelial cell markers. Inhibition of reactive oxygen species generation during the early stages of differentiation abrogated the endothelial inductive effects of the low oxygen environments. Low oxygen tension during early stages of EVC derivation induces endothelial commitment and maturation through the accumulation of reactive oxygen species, highlighting the importance of regulating oxygen tensions during human pluripotent stem cell-vascular differentiation.

Arginase-I enhances vascular endothelial inflammation and senescence through eNOS-uncoupling.

PubMed

Zhu, Cuicui; Yu, Yi; Montani, Jean-Pierre; Ming, Xiu-Fen; Yang, Zhihong

2017-02-02

Augmented arginase-II (Arg-II) is implicated in endothelial senescence and inflammation through a mutual positive regulatory circuit with S6K1. This study was conducted to investigate whether Arg-I, another isoform of arginase that has been also reported to play a role in vascular endothelial dysfunction, promotes endothelial senescence through similar mechanisms. The non-senescent human endothelial cells from umbilical veins (passage 2 to 4) were transduced with empty recombinant adenovirus vector (rAd/CMV) as control or rAd/CMV-Arg-I to overexpress Arg-I. Overexpressing Arg-I promoted eNOS-uncoupling, enhanced senescence markers including p53-S15, p21 and senescence-associated β-galactosidase (SA-β-gal) staining, and increased inflammatory vascular adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) as well as monocyte adhesion to endothelial cells without activating S6K1. All the effects of Arg-I were inhibited by the anti-oxidant N-acetylcysteine (NAC). Our study demonstrates that Arg-I promotes endothelial senescence and inflammatory responses through eNOS-uncoupling unrelated to activation of the S6K1 pathway.

VEGF-C and TGF-β reciprocally regulate mesenchymal stem cell commitment to differentiation into lymphatic endothelial or osteoblastic phenotypes.

PubMed

Igarashi, Yasuyuki; Chosa, Naoyuki; Sawada, Shunsuke; Kondo, Hisatomo; Yaegashi, Takashi; Ishisaki, Akira

2016-04-01

The direction of mesenchymal stem cell (MSC) differentiation is regulated by stimulation with various growth factors and cytokines. We recently established MSC lines, [transforming growth factor-β (TGF-β)-responsive SG‑2 cells, bone morphogenetic protein (BMP)-responsive SG‑3 cells, and TGF-β/BMP-non-responsive SG‑5 cells], derived from the bone marrow of green fluorescent protein-transgenic mice. In this study, to compare gene expression profiles in these MSC lines, we used DNA microarray analysis to characterize the specific gene expression profiles observed in the TGF-β-responsive SG‑2 cells. Among the genes that were highly expressed in the SG‑2 cells, we focused on vascular endothelial growth factor (VEGF) receptor 3 (VEGFR3), the gene product of FMS-like tyrosine kinase 4 (Flt4). We found that VEGF-C, a specific ligand of VEGFR3, significantly induced the cell proliferative activity, migratory ability (as shown by Transwell migration assay), as well as the phosphorylation of extracellular signal-regulated kinase (ERK)1/2 in the SG‑2 cells. Additionally, VEGF-C significantly increased the expression of prospero homeobox 1 (Prox1) and lymphatic vessel endothelial hyaluronan receptor 1 (Lyve1), which are lymphatic endothelial cell markers, and decreased the expression of osteogenic differentiation marker genes in these cells. By contrast, TGF-β significantly increased the expression of early-phase osteogenic differentiation marker genes in the SG‑2 cells and markedly decreased the expression of lymphatic endothelial cell markers. The findings of our study strongly suggest the following: i) that VEGF-C promotes the proliferative activity and migratory ability of MSCs; and ii) VEGF-C and TGF-β reciprocally regulate MSC commitment to differentiation into lymphatic endothelial or osteoblastic phenotypes, respectively. Our findings provide new insight into the molecular mechanisms underlying the regenerative ability of MSCs.

Physical exercise, fitness and dietary pattern and their relationship with circadian blood pressure pattern, augmentation index and endothelial dysfunction biological markers: EVIDENT study protocol

PubMed Central

2010-01-01

Background Healthy lifestyles may help to delay arterial aging. The purpose of this study is to analyze the relationship of physical activity and dietary pattern to the circadian pattern of blood pressure, central and peripheral blood pressure, pulse wave velocity, carotid intima-media thickness and biological markers of endothelial dysfunction in active and sedentary individuals without arteriosclerotic disease. Methods/Design Design: A cross-sectional multicenter study with six research groups. Subjects: From subjects of the PEPAF project cohort, in which 1,163 who were sedentary became active, 1,942 were sedentary and 2,346 were active. By stratified random sampling, 1,500 subjects will be included, 250 in each group. Primary measurements: We will evaluate height, weight, abdominal circumference, clinical and ambulatory blood pressure with the Radial Pulse Wave Acquisition Device (BPro), central blood pressure and augmentation index with Pulse Wave Application Software (A-Pulse) and SphymgoCor System Px (Pulse Wave Analysis), pulse wave velocity (PWV) with SphymgoCor System Px (Pulse Wave Velocity), nutritional pattern with a food intake frequency questionnaire, physical activity with the 7-day PAR questionnaire and accelerometer (Actigraph GT3X), physical fitness with the cycle ergometer (PWC-170), carotid intima-media thickness by ultrasound (Micromax), and endothelial dysfunction biological markers (endoglin and osteoprotegerin). Discussion Determining that sustained physical activity and the change from sedentary to active as well as a healthy diet improve circadian pattern, arterial elasticity and carotid intima-media thickness may help to propose lifestyle intervention programs. These interventions could improve the cardiovascular risk profile in some parameters not routinely assessed with traditional risk scales. From the results of this study, interventional approaches could be obtained to delay vascular aging that combine physical exercise and diet

Endothelial cells genetically selected from differentiating mouse embryonic stem cells incorporate at sites of neovascularization in vivo.

PubMed

Marchetti, Sandrine; Gimond, Clotilde; Iljin, Kristiina; Bourcier, Christine; Alitalo, Kari; Pouysségur, Jacques; Pagès, Gilles

2002-05-15

Large scale purification of endothelial cells is of great interest as it could improve tissue transplantation, reperfusion of ischemic tissues and treatment of pathologies in which an endothelial cell dysfunction exists. In this study, we describe a novel genetic approach that selects for endothelial cells from differentiating embryonic stem (ES) cells. Our strategy is based on the establishment of ES-cell clones that carry an integrated puromycin resistance gene under the control of a vascular endothelium-specific promoter, tie-1. Using EGFP as a reporter gene, we first confirmed the endothelial specificity of the tie-1 promoter in the embryoid body model and in cells differentiated in 2D cultures. Subsequently, tie-1-EGFP ES cells were used as recipients for the tie-1-driven puror transgene. The resulting stable clones were expanded and differentiated for seven days in the presence of VEGF before puromycin selection. As expected, puromycin-resistant cells were positive for EGFP and also expressed several endothelial markers, including CD31, CD34, VEGFR-1, VEGFR-2, Tie-1, VE-cadherin and ICAM-2. Release from the puromycin selection resulted in the appearance of alpha-smooth muscle actin-positive cells. Such cells became more numerous when the population was cultured on laminin-1 or in the presence of TGF-beta1, two known inducers of smooth muscle cell differentiation. The hypothesis that endothelial cells or their progenitors may differentiate towards a smooth muscle cell phenotype was further supported by the presence of cells expressing both CD31 and alpha-smooth muscle actin markers. Finally, we show that purified endothelial cells can incorporate into the neovasculature of transplanted tumors in nude mice. Taken together, these results suggest that application of endothelial lineage selection to differentiating ES cells may become a useful approach for future pro-angiogenic and endothelial cell replacement therapies.

Circulating endothelial cells in acute ischaemic stroke.

PubMed

Nadar, Sunil K; Lip, Gregory Y H; Lee, Kaeng W; Blann, Andrew D

2005-10-01

Increased numbers of CD146-bearing circulating endothelial cells (CECs) in the peripheral blood probably represent the most direct evidence of endothelial cell damage. As acute ischaemic strokes are associated with endothelial abnormalities, we hypothesised that these CECs are raised in acute stroke, and that they would correlate with the other indices of endothelial perturbation, i.e. plasma von Willebrand factor (vWf) and soluble E-selectin. We studied 29 hypertensive patients (19 male; mean age 63 years) who presented with an acute stroke and compared them with 30 high risk hypertensive patients (21 male; mean age 62 years) and 30 normotensive controls (16 male; mean age 58 years). CECs were estimated by CD146 immunobead capture, vWf and soluble E-selectin by ELISA. Patients with an acute ischaemic stroke had significantly higher numbers of CECs/ml of blood (p<0.001) plasma vWf (p=0.008) soluble E-selectin (p=0.002) and higher systolic blood pressure (SBP) as compared to the other groups. The number of CECs significantly correlated with soluble E-selectin (r=0.432, p<0.001) and vWf (r=0.349, p=0.001) but not with SBP (r=0.198, p=0.069). However, in multivariate analysis, only disease group (i.e. health, hypertension or stroke) was associated with increased CECs. Acute ischaemic stroke is associated with increased numbers of CECs. The latter correlate well with established plasma markers of endothelial dysfunction or damage, thus unequivocally confirming severe vasculopathy in this condition. However, the greatest influence on CECs numbers was clinical group.

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.

Endothelial function and insulin resistance in polycystic ovary syndrome: the effects of medical therapy.

PubMed

Teede, Helena J; Meyer, Caroline; Hutchison, Samantha K; Zoungas, Sophia; McGrath, Barry P; Moran, Lisa J

2010-01-01

To assess the interaction between insulin resistance and endothelial function and the optimal treatment strategy addressing cardiovascular risk in polycystic ovary syndrome. Randomized controlled trial. Controlled clinical study. Overweight age- and body mass index-matched women with polycystic ovary syndrome. Six months metformin (1 g two times per day, n = 36) or oral contraceptive pill (OCP) (35 microg ethinyl E(2)-2 mg cytoproterone acetate, n = 30). Fasting and oral glucose tolerance test glucose and insulin levels, endothelial function (flow-mediated dilation, asymmetric dimethylarginine, plasminogen activator inhibitor-1, von Willebrand factor), inflammatory markers (high-sensitivity C-reactive protein), lipids, and hyperandrogenism. The OCP increased levels of glucose and insulin on oral glucose tolerance test, high-sensitivity C-reactive protein, triglycerides, and sex-hormone binding globulin and decreased levels of low-density lipoprotein cholesterol and T. Metformin decreased levels of fasting insulin, oral glucose tolerance test insulin, high-density lipoprotein cholesterol, and high-sensitivity C-reactive protein. Flow-mediated dilation increased only with metformin (+2.2% +/- 4.8%), whereas asymmetric dimethylarginine decreased equivalently for OCP and metformin (-0.3 +/- 0.1 vs. -0.1 +/- 0.1 mmol/L). Greater decreases in plasminogen activator inhibitor-1 occurred for the OCP than for metformin (-1.8 +/- 1.6 vs. -0.7 +/- 1.7 U/mL). In polycystic ovary syndrome, metformin improves insulin resistance, inflammatory markers, and endothelial function. The OCP worsens insulin resistance and glucose homeostasis, inflammatory markers, and triglycerides and has neutral or positive endothelial effects. The effect of the OCP on cardiovascular risk in polycystic ovary syndrome is unclear. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Orbital Angiogenesis and Lymphangiogenesis in Thyroid Eye Disease: An Analysis of Vascular Growth Factors with Clinical Correlation.

PubMed

Wong, Lindsay L; Lee, Nahyoung Grace; Amarnani, Dhanesh; Choi, Catherine J; Bielenberg, Diane R; Freitag, Suzanne K; D'Amore, Patricia A; Kim, Leo A

2016-09-01

The human orbit is an environment that is vulnerable to inflammation and edema in the setting of autoimmune thyroid disease. Our study investigated the tenet that orbital adipose tissue lacks lymphatic vessels and analyzed the clinicopathologic differences between patients with acute and chronic thyroid eye disease (TED). The underlying molecular mediators of blood and lymphatic vessel formation within the orbital fat also were evaluated. Retrospective cohort study. The study included fat specimens from 26 orbits of 15 patients with TED undergoing orbital decompression. Orbital fat specimens from patients without TED as well as cadaveric orbital fat served as controls. Tissue specimens were processed as formalin-fixed, paraffin-embedded sections or frozen cryosections for immunohistochemistry. Total RNA was extracted and analyzed via quantitative (real-time) reverse-transcription polymerase chain reaction. Clinicopathologic correlation was made by determining the clinical activity score (CAS) of each patient with TED. Samples were examined for vascular and lymphatic markers including podoplanin, lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), and cluster of differentiation 31 (CD31) by immunohistochemistry, as well as for mRNA levels of vascular endothelial growth factor (VEGF), VEGF receptors, semaphorin 3F, neuropilin 1, neuropilin 2, podoplanin, and LYVE-1 by quantitative (real-time) reverse-transcription polymerase chain reaction. Clinicopathologic correlation revealed increased staining of CD31-positive blood vessels in patients with acute TED with a CAS more than 4, as well as rare staining of podoplanin-positive lymphatic vessels within acutely inflamed orbital fat tissue. Additionally, quantitative (real-time) reverse-transcription polymerase chain reaction analysis demonstrated increased expression of VEGF receptor (VEGFR) 2 as well as VEGF signaling molecules VEGF-A, VEGF-C, and VEGF-D. In acute TED, compared with chronic TED and control

Endothelial function in youth: A biomarker modulated by adiposity-related insulin resistance

USDA-ARS?s Scientific Manuscript database

To investigate the physical and metabolic determinants of endothelial dysfunction, an early marker of subclinical atherosclerosis, in normal weight and overweight adolescents with and without type 2 diabetes mellitus. A cross-sectional study of 81 adolescents: 21 normal weight, 25 overweight with no...

Tetrahydrocurcumin ameliorates homocysteine-mediated mitochondrial remodeling in brain endothelial cells.

PubMed

Vacek, Jonathan C; Behera, Jyotirmaya; George, Akash K; Kamat, Pradip K; Kalani, Anuradha; Tyagi, Neetu

2018-04-01

Homocysteine (Hcy) causes endothelial dysfunction by inducing oxidative stress in most neurodegenerative disorders. This dysfunction is highly correlated with mitochondrial dynamics such as fusion and fission. However, there are no strategies to prevent Hcy-induced mitochondrial remodeling. Tetrahydrocurcumin (THC) is an anti-inflammatory and anti-oxidant compound. We hypothesized that THC may ameliorates Hcy-induced mitochondria remodeling in mouse brain endothelial cells (bEnd3) cells. bEnd3 cells were exposed to Hcy treatment in the presence or absence of THC. Cell viability and autophagic cell death were measured with MTT and MDC staining assay. Reactive oxygen species (ROS) production was determined using DCFH-DA staining by confocal microscopy. Autophagy flux was assessed using a conventional GFP-microtubule-associated protein 1 light chain 3 (LC3) dot assay. Interaction of phagophore marker LC-3 with mitochondrial receptor NIX was observed by confocal imaging. Mitochondrial fusion and fission were evaluated by western blot and RT-PCR. Our results demonstrated that Hcy resulted in cell toxicity in a dose-dependent manner and supplementation of THC prevented the detrimental effects of Hcy on cell survival. Furthermore, Hcy also upregulated fission marker (DRP-1), fusion marker (Mfn2), and autophagy marker (LC-3). Finally, we observed that Hcy activated mitochondrial specific phagophore marker (LC-3) and co-localized with the mitochondrial receptor NIX, as viewed by confocal microscopy. Pretreatment of bEnd3 with THC (15 μM) ameliorated Hcy-induced oxidative damage, mitochondrial fission/fusion, and mitophagy. Our studies strongly suggest that THC has beneficial effects on mitochondrial remodeling and could be developed as a potential therapeutic agent against hyperhomocysteinemia (HHcy) induced mitochondrial dysfunction. © 2017 Wiley Periodicals, Inc.

Soya isoflavone-enriched cereal bars affect markers of endothelial function in postmenopausal women.

PubMed

Hallund, J; Bügel, S; Tholstrup, T; Ferrari, M; Talbot, D; Hall, W L; Reimann, M; Williams, C M; Wiinberg, N

2006-06-01

Soya isoflavones are thought to be cardioprotective due to their structural similarity to oestrogen. In order to investigate the effect of soya isoflavones on markers of endothelial function we conducted a randomised, double-blind, placebo-controlled, cross-over study with thirty healthy postmenopausal women. The women consumed cereal bars, with or without soya isoflavones (50 mg/d), for 8 weeks, separated by an 8-week washout period. Systemic arterial compliance (SAC), isobaric arterial compliance (IAC), flow-mediated endothelium-dependent vasodilation (FMD) and nitroglycerine-mediated endothelium-independent vasodilation (NMD) were measured at the beginning of the study and after each intervention period. Blood pressure (BP) and plasma concentrations of nitrite and nitrate (NOx) and endothelin-1 (ET-1) were measured at the beginning and end of each intervention period. NMD was 13.4 (SEM 2.0)% at baseline and 15.5 (SEM 1.1) % after isoflavone treatment compared with 12.4 (SEM 1.0)% after placebo treatment (P=0.03). NOx increased from 27.7 (SEM 2.7) to 31.1 (SEM 3.2) microM after isoflavones treatment compared with 25.4 (SEM 1.5) to 20.4 (SEM 1.1) microM after placebo treatment (P=0.003) and a significant increase in the NOx:ET-1 ratio (P=0.005) was observed after the isoflavone treatment compared with placebo. A significant difference in SAC after the isoflavone and placebo treatment was observed (P=0.04). No significant difference was found in FMD, IAC, BP and ET-1. In conclusion, 8 weeks' consumption of cereals bars enriched with 50 mg soya isoflavones/d increased plasma NOx concentrations and improved endothelium-independent vasodilation in healthy postmenopausal women.

Coculture with endothelial cells reduces the population of cycling LeX neural precursors but increases that of quiescent cells with a side population phenotype

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

Mathieu, Celine; Fouchet, Pierre; Gauthier, Laurent R.

2006-04-01

Neural stem cell proliferation and differentiation are regulated by external cues from their microenvironment. As endothelial cells are closely associated with neural stem cell in brain germinal zones, we investigated whether endothelial cells may interfere with neurogenesis. Neural precursor cells (NPC) from telencephalon of EGFP mouse embryos were cocultured in direct contact with endothelial cells. Endothelial cells did not modify the overall proliferation and apoptosis of neural cells, albeit they transiently delayed spontaneous apoptosis. These effects appeared to be specific to endothelial cells since a decrease in proliferation and a raise in apoptosis were observed in cocultures with fibroblasts. Endothelialmore » cells stimulated the differentiation of NPC into astrocytes and into neurons, whereas they reduced differentiation into oligodendrocytes in comparison to adherent cultures on polyornithine. Determination of NPC clonogenicity and quantification of LeX expression, a marker for NPC, showed that endothelial cells decreased the number of cycling NPC. On the other hand, the presence of endothelial cells increased the number of neural cells having 'side population' phenotype, another marker reported on NPC, which we have shown to contain quiescent cells. Thus, we show that endothelial cells may regulate neurogenesis by acting at different level of NPC differentiation, proliferation and quiescence.« less

Ezetimibe inhibits platelet activation and uPAR expression on endothelial cells.

PubMed

Becher, Tobias; Schulze, Torsten J; Schmitt, Melanie; Trinkmann, Frederik; El-Battrawy, Ibrahim; Akin, Ibrahim; Kälsch, Thorsten; Borggrefe, Martin; Stach, Ksenija

2017-01-15

Lipid lowering therapy constitutes the basis of cardiovascular disease therapy. The purpose of this study was to investigate effects of ezetimibe, a selective inhibitor of intestinal cholesterol absorption, on platelets and endothelial cells in an in vitro endothelial cell model. After a 24h incubation period with ezetimibe (concentrations 1, 50, 100 and 1000ng/ml), human umbilical vein endothelial cells (HUVEC) were stimulated for 1h with lipopolysaccharide (LPS) and were then incubated in direct contact with activated platelets. Following this, the expression of CD40L and CD62P on platelets, and the expression of ICAM-1, VCAM-1, uPAR, and MT1-MMP on endothelial cells were measured by flow cytometry. Supernatants were analysed by enzyme linked immunosorbent assay for soluble MCP-1, IL-6 and MMP-1. The increased expression of uPAR on endothelial cells by proinflammatory stimulation with LPS and by direct endothelial contact with activated platelets was significantly reduced through pre-incubation with 100ng/ml and 1000ng/ml ezetimibe (p<0.05). Platelets directly incubated with ezetimibe but without endothelial cell contact showed significantly reduced CD62P and CD40L surface expression (p<0.05). Ezetimibe had no significant effects on HUVEC expression of MT1-MMP, ICAM-1 and VCAM-1 and on CD40L expression on platelets in direct contact with endothelial cells. Levels of soluble IL-6 in HUVEC supernatants were significantly lower after pre-incubation with ezetimibe. In this in vitro analysis, ezetimibe directly attenuates platelet activation and has significant endothelial cell mediated effects on selected markers of atherosclerosis. Copyright © 2016. Published by Elsevier Ireland Ltd.

Maternal endothelial damage as a disorder shared by early preeclampsia, late preeclampsia and intrauterine growth restriction.

PubMed

Kwiatkowski, Sebastian; Dołegowska, Barbara; Kwiatkowska, Ewa; Rzepka, Rafał; Marczuk, Natalia; Loj, Beata; Torbè, Andrzej

2017-10-26

Preeclampsia (PE) and intrauterine growth restriction (IUGR) are separate disease entities that have frequently been reported as sharing the same pathogenesis. In both of them, angiogenesis disorders and generalized endothelial damage with an accompanying inflammation are the dominant symptoms. In this study, we attempted to prove that both these processes demonstrate the same profile in early PE, late PE and IUGR patients, while the only difference is in the degree of exacerbation of the lesions. In 167 patients divided into four groups, three of those with early PE, late PE and IUGR and one control group, fms-like tyrosine kinase-1 (sFlt-1), placental growth factor (PlGF), high sensitive c-reactive protein (hsCRP) and fibronectin were determined. The behavior of these parameters in each of the groups was studied, and correlations between them were sought for. Higher concentrations of sFlt-1, hsCRP and fibronectin and a lower concentration of PlGF were found in the study groups compared to the control group. Significant correlations were observed between the factors concerned. The higher values of disordered angiogenesis markers, endothelial damage markers and inflammatory markers both in the PE and the intrauterine growth restriction (IUGR) groups suggest the existence of shared disorders in the development of these pathologies. The correlations between disordered angiogenesis markers and endothelial damage markers argue in favor of a mutual relationship between these two processes in the development of pathologies evolving as secondary to placental ischemia. The results obtained confirm that the lesion profiles are the same in both PE and IUGR patients, which can be utilized in developing common diagnostic criteria.

Plasma levels of endothelial and B-cell-derived microparticles are restored by fingolimod treatment in multiple sclerosis patients.

PubMed

Zinger, Anna; Latham, Sharissa L; Combes, Valery; Byrne, Scott; Barnett, Michael H; Hawke, Simon; Grau, Georges E

2016-12-01

No molecular marker can monitor disease progression and treatment efficacy in multiple sclerosis (MS). Circulating microparticles represent a potential snapshot of disease activity at the blood brain barrier. To profile plasma microparticles by flow cytometry in MS and determine how fingolimod could impact endothelial microparticles production. In non-treated MS patients compared to healthy and fingolimod-treated patients, endothelial microparticles were higher, while B-cell-microparticle numbers were lower. Fingolimod dramatically reduced tumour necrosis factor (TNF)-induced endothelial microparticle release in vitro. Fingolimod restored dysregulated endothelial and B-cell-microparticle numbers, which could serve as a biomarker in MS. © The Author(s), 2016.

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

Circulating microparticles, protein C, free protein S and endothelial vascular markers in children with sickle cell anaemia

PubMed Central

Piccin, Andrea; Murphy, Ciaran; Eakins, Elva; Kunde, Jan; Corvetta, Daisy; Di Pierro, Angela; Negri, Giovanni; Guido, Mazzoleni; Sainati, Laura; Mc Mahon, Corrina; Smith, Owen Patrick; Murphy, William

2015-01-01

Introduction Circulating microparticles (MP) have been described in sickle cell anaemia (SCA); however, their interaction with endothelial markers remains unclear. We investigated the relationship between MP, protein C (PC), free protein S (PS), nitric oxide (NO), endothelin-1 (ET-1) and adrenomedullin (ADM) in a large cohort of paediatric patients. Method A total of 111 children of African ethnicity with SCA: 51 in steady state; 15 in crises; 30 on hydroxyurea (HU) therapy; 15 on transfusion; 17 controls (HbAA) of similar age/ethnicity. MP were analysed by flow cytometry using: Annexin V (AV), CD61, CD42a, CD62P, CD235a, CD14, CD142 (tissue factor), CD201 (endothelial PC receptor), CD62E, CD36 (TSP-1), CD47 (TSP-1 receptor), CD31 (PECAM), CD144 (VE-cadherin). Protein C, free PS, NO, pro-ADM and C-terminal ET-1 were also measured. Results Total MP AV was lower in crisis (1.26×106 ml−1; 0.56–2.44×106) and steady state (1.35×106 ml−1; 0.71–3.0×106) compared to transfusion (4.33×106 ml−1; 1.6–9.2×106, p<0.01). Protein C levels were significantly lower in crisis (median 0.52 IU ml−1; interquartile range 0.43–0.62) compared with all other groups: HbAA (0.72 IU ml−1; 0.66–0.82, p<0.001); HU (0.67 IU ml−1; 0.58–0.77, p<0.001); steady state (0.63 IU ml−1; 0.54–0.70, p<0.05) and transfusion (0.60 IU ml−1; 0.54–0.70, p<0.05). In addition, levels were significantly reduced in steady state (0.63 IU ml−1; 0.54–0.70) compared with HbAA (0.72 IU ml−1; 0.66–0.80, p<0.01). PS levels were significantly higher in HbAA (0.85 IU ml−1; 0.72–0.97) compared with crisis (0.49 IU ml−1; 0.42–0.64, p<0.001), HU (0.65 IU ml−1; 0.56–0.74, p<0.01) and transfusion (0.59 IU ml−1; 0.47–0.71, p<0.01). There was also a significant difference in crisis patients compared with steady state (0.49 IU ml−1; 0.42–0.64 vs. 0.68 IU ml−1; 0.58–0.79, p<0.05). There was high correlation (R>0.9, p<0.05) between total numbers of AV-positive MP

Astrocytes Can Adopt Endothelial Cell Fates in a p53-Dependent Manner.

PubMed

Brumm, Andrew J; Nunez, Stefanie; Doroudchi, Mehdi M; Kawaguchi, Riki; Duan, Jinhzu; Pellegrini, Matteo; Lam, Larry; Carmichael, S Thomas; Deb, Arjun; Hinman, Jason D

2017-08-01

Astrocytes respond to a variety of CNS injuries by cellular enlargement, process outgrowth, and upregulation of extracellular matrix proteins that function to prevent expansion of the injured region. This astrocytic response, though critical to the acute injury response, results in the formation of a glial scar that inhibits neural repair. Scar-forming cells (fibroblasts) in the heart can undergo mesenchymal-endothelial transition into endothelial cell fates following cardiac injury in a process dependent on p53 that can be modulated to augment cardiac repair. Here, we sought to determine whether astrocytes, as the primary scar-forming cell of the CNS, are able to undergo a similar cellular phenotypic transition and adopt endothelial cell fates. Serum deprivation of differentiated astrocytes resulted in a change in cellular morphology and upregulation of endothelial cell marker genes. In a tube formation assay, serum-deprived astrocytes showed a substantial increase in vessel-like morphology that was comparable to human umbilical vein endothelial cells and dependent on p53. RNA sequencing of serum-deprived astrocytes demonstrated an expression profile that mimicked an endothelial rather than astrocyte transcriptome and identified p53 and angiogenic pathways as specifically upregulated. Inhibition of p53 with genetic or pharmacologic strategies inhibited astrocyte-endothelial transition. Astrocyte-endothelial cell transition could also be modulated by miR-194, a microRNA downstream of p53 that affects expression of genes regulating angiogenesis. Together, these studies demonstrate that differentiated astrocytes retain a stimulus-dependent mechanism for cellular transition into an endothelial phenotype that may modulate formation of the glial scar and promote injury-induced angiogenesis.

Simultaneous isolation of vascular endothelial cells and mesenchymal stem cells from the human umbilical cord.

PubMed

Kadam, Sachin S; Tiwari, Shubha; Bhonde, Ramesh R

2009-01-01

The umbilical cord represents the link between mother and fetus during pregnancy. This cord is usually discarded as a biological waste after the child's birth; however, its importance as a "store house" of stem cells has been explored recently. We developed a method of simultaneous isolation of endothelial cells (ECs) from the vein and mesenchymal stem cells from umbilical cord Wharton's jelly of the same cord. The isolation protocol has been simplified, modified, and improvised with respect to choice of enzyme and enzyme mixture, digestion time, cell yield, cell growth, and culture medium. Isolated human umbilical vascular ECs (hUVECs) were positive for von-Willibrand factor, a classical endothelial marker, and could form capillary-like structures when seeded on Matrigel, thus proving their functionality. The isolated human umbilical cord mesenchymal stem cells (hUCMSCs) were found positive for CD44, CD90, CD 73, and CD117 and were found negative for CD33, CD34, CD45, and CD105 surface markers; they were also positive for cytoskeleton markers of smooth muscle actin and vimentin. The hUCMSCs showed multilineage differentiation potential and differentiated into adipogenic, chondrogenic, osteogenic, and neuronal lineages under influence of lineage specific differentiation medium. Thus, isolating endothelial cells as well as mesenchymal cells from the same umbilical cord could lead to complete utilization of the available tissue for the tissue engineering and cell therapy.

Biomimetic, ultrathin and elastic hydrogels regulate human neutrophil extravasation across endothelial-pericyte bilayers.

PubMed

Lauridsen, Holly M; Gonzalez, Anjelica L

2017-01-01

The vascular basement membrane-a thin, elastic layer of extracellular matrix separating and encasing vascular cells-provides biological and mechanical cues to endothelial cells, pericytes, and migrating leukocytes. In contrast, experimental scaffolds typically used to replicate basement membranes are stiff and bio-inert. Here, we present thin, porated polyethylene glycol hydrogels to replicate human vascular basement membranes. Like commercial transwells, our hydrogels are approximately 10μm thick, but like basement membranes, the hydrogels presented here are elastic (E: 50-80kPa) and contain a dense network of small pores. Moreover, the inclusion of bioactive domains introduces receptor-mediated biochemical signaling. We compare elastic hydrogels to common culture substrates (E: >2GPa) for human endothelial cell and pericyte monolayers and bilayers to replicate postcapillary venules in vitro. Our data demonstrate that substrate elasticity facilitates differences in vascular phenotype, supporting expression of vascular markers that are increasingly replicative of venules. Endothelial cells differentially express vascular markers, like EphB4, and leukocyte adhesion molecules, such as ICAM-1, with decreased mechanical stiffness. With porated PEG hydrogels we demonstrate the ability to evaluate and observe leukocyte recruitment across endothelial cell and pericyte monolayers and bilayers, reporting that basement membrane scaffolds can significantly alter the rate of vascular migration in experimental systems. Overall, this study demonstrates the creation and utility of a new and accessible method to recapture the mechanical and biological complexity of human basement membranes in vitro.

Ulex europaeus I lectin as a marker for vascular endothelium in human tissues.

PubMed

Holthöfer, H; Virtanen, I; Kariniemi, A L; Hormia, M; Linder, E; Miettinen, A

1982-07-01

Ulex europaeus I agglutinin, a lectin specific for some alpha-L-fucose-containing glycocompounds, was used in fluorescence microscopy to stain cryostat sections of human tissues. The endothelium of vessels of all sizes was stained ubiquitously in all tissues studied as judged by double staining with a known endothelial marker, antibodies against human clotting factor VIII. Cultured human umbilical vein endothelial cells, but not fibroblasts, also bound Ulex lectin. The staining was not affected by the blood group type of the tissue donor. In some tissues Ulex lectin presented additional binding to epithelial structures. Also, this was independent on the blood group or the ability of the tissue donor to secrete soluble blood group substances. Lotus tetragonolobus agglutinin, another lectin specific for some alpha-L-fucose-containing moieties failed to react with endothelial cells. Our results suggest that Ulex europaeus I agglutinin is a good histologic marker for endothelium in human tissues.

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.

Characterization of nanoparticle uptake by endothelial cells.

PubMed

Davda, Jasmine; Labhasetwar, Vinod

2002-02-21

Endothelium is an important target for drug or gene therapy because of its important role in the biological system. In this paper, we have characterized nanoparticle uptake by endothelial cells in cell culture. Nanoparticles were formulated using poly DL-lactide-co-glycolide polymer containing bovine serum albumin as a model protein and 6-coumarin as a fluorescent marker. It was observed that the cellular uptake of nanoparticles depends on the time of incubation and the concentration of nanoparticles in the medium. The uptake of nanoparticles was rapid with confocal microscopy demonstrating their localization mostly in the cytoplasm. The mitogenic study demonstrated biocompatability of nanoparticles with the cells. The study thus demonstrates that nanoparticles could be used for localizing therapeutic agents or gene into endothelial cells. Nanoparticles localized in the endothelium could provide prolonged drug effects because of their sustained release characterics, and also could protect the encapsulated agent from enzymatic degradation.

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.

De Novo Lipogenesis Maintains Vascular Homeostasis through Endothelial Nitric-oxide Synthase (eNOS) Palmitoylation*♦

PubMed Central

Wei, Xiaochao; Schneider, Jochen G.; Shenouda, Sherene M.; Lee, Ada; Towler, Dwight A.; Chakravarthy, Manu V.; Vita, Joseph A.; Semenkovich, Clay F.

2011-01-01

Endothelial dysfunction leads to lethal vascular complications in diabetes and related metabolic disorders. Here, we demonstrate that de novo lipogenesis, an insulin-dependent process driven by the multifunctional enzyme fatty-acid synthase (FAS), maintains endothelial function by targeting endothelial nitric-oxide synthase (eNOS) to the plasma membrane. In mice with endothelial inactivation of FAS (FASTie mice), eNOS membrane content and activity were decreased. eNOS and FAS were physically associated; eNOS palmitoylation was decreased in FAS-deficient cells, and incorporation of labeled carbon into eNOS-associated palmitate was FAS-dependent. FASTie mice manifested a proinflammatory state reflected as increases in vascular permeability, endothelial inflammatory markers, leukocyte migration, and susceptibility to LPS-induced death that was reversed with an NO donor. FAS-deficient endothelial cells showed deficient migratory capacity, and angiogenesis was decreased in FASTie mice subjected to hindlimb ischemia. Insulin induced FAS in endothelial cells freshly isolated from humans, and eNOS palmitoylation was decreased in mice with insulin-deficient or insulin-resistant diabetes. Thus, disrupting eNOS bioavailability through impaired lipogenesis identifies a novel mechanism coordinating nutritional status and tissue repair that may contribute to diabetic vascular disease. PMID:21098489

Vascular endothelial growth factor increases fenestral permeability in hepatic sinusoidal endothelial cells.

PubMed

Yokomori, Hiroaki; Oda, Masaya; Yoshimura, Kazunori; Nagai, Toshihiro; Ogi, Mariko; Nomura, Masahiko; Ishii, Hiromasa

2003-12-01

Vascular endothelial growth factor (VEGF) is an important regulator of vasculogenesis and vascular permeability. Hepatic sinusoidal endothelial cells (SECs) possess sieve-like pores that form an anastomosing labyrinth structure by the deeply invaginated plasma membrane. Caveolin is the principal structural protein in caveolae. In this study, we examined the role of VEGF on the fenestration and permeability of SECs and the relation with caveolin-1. SECs isolated from rat livers by collagenase infusion method were cultured for 24 h with (10 or 100 ng/ml) or without VEGF. The cells were then examined by transmission and scanning electron microscopy (EM). The expression of caveolin was investigated by confocal immunofluorescence, immunogold EM, and Western blot. Endocytosis and intracellular traffic was studied using horseradish peroxidase (HRP) reaction as a marker of fluid phase transport in SECs. Both transmission and scanning EM showed an increased number of sinusoidal endothelial fenestrae (SEF) in SECs cultured with VEGF. By confocal immunofluorescence, SECs cultured with VEGF displayed prominent caveolin-l-positive aggregates in the cytoplasm, especially surrounding the nucleus region. Immunogold EM depicted increased caveolin-1 reactivity on vesicles and vacuoles of VEGF-treated SECs compared with VEGF-nontreated cells. However, there was no change in the level of caveolin-1 protein expression on Western blot. After HRP injection, an increase of electron-dense tracer filled the SEF in cells treated with VEGF. Our results suggested that VEGF induced fenestration in SECs, accompanied by an increased number of caveolae-like vesicles. Increased caveolin-1 might be associated with vesicle formation but not with fenestration. Increased fenestration may augment hepatic sinusoidal permeability and transendothelial transport.

Lack of inhibitory effects of the anti-fibrotic drug imatinib on endothelial cell functions in vitro and in vivo.

PubMed

Venalis, Paulius; Maurer, Britta; Akhmetshina, Alfiya; Busch, Nicole; Dees, Clara; Stürzl, Michael; Zwerina, Jochen; Jüngel, Astrid; Gay, Steffen; Schett, Georg; Distler, Oliver; Distler, Jörg H W

2009-10-01

Systemic sclerosis (SSc) is a systemic autoimmune disease that is characterized by microangiopathy with progressive loss of capillaries and tissue fibrosis. Imatinib exerts potent anti-fibrotic effects and is currently evaluated in clinical trials. The aim of the present study was to exclude that the anti-fibrotic effects of imatinib are complicated by inhibitory effects on endothelial cell functions, which might augment vascular disease in SSc. Endothelial cells and mice were treated with pharmacologically relevant concentrations of imatinib. The expression of markers of vascular activation was assessed with real-time PCR. Proliferation was analysed with the cell counting experiments and the MTT assay. Apoptosis was quantified with caspase 3 assays, annexin V in vitro and with TUNEL staining in vivo. Migration was studied with scratch and transwell assays. Tube forming was investigated with the matrigel assay. Imatinib did not alter the expression of markers of vascular activation. Imatinib did not increase the percentage of annexin V positive cells or the activity of caspase 3. No reduction in proliferation or metabolic activity of endothelial cells was observed. Imatinib did not affect migration of endothelial cells and did not reduce the formation of capillary tubes. Consistent with the in vitro data, no difference in the number of apoptotic endothelial cells was observed in vivo in mice treated with imatinib. Imatinib does not inhibit activation, viability, proliferation, migration or tube forming of endothelial cells in vitro and in vivo. Thus, treatment with imatinib might not augment further endothelial cell damage in SSc.

Lack of inhibitory effects of the anti-fibrotic drug imatinib on endothelial cell functions in vitro and in vivo

PubMed Central

Venalis, Paulius; Maurer, Britta; Akhmetshina, Alfiya; Busch, Nicole; Dees, Clara; Stürzl, Michael; Zwerina, Jochen; Jüngel, Astrid; Gay, Steffen; Schett, Georg; Distler, Oliver; Distler, Jörg HW

2009-01-01

Systemic sclerosis (SSc) is a systemic autoimmune disease that is characterized by microangiopathy with progressive loss of capillaries and tissue fibrosis. Imatinib exerts potent anti-fibrotic effects and is currently evaluated in clinical trials. The aim of the present study was to exclude that the anti-fibrotic effects of imatinib are complicated by inhibitory effects on endothelial cell functions, which might augment vascular disease in SSc. Endothelial cells and mice were treated with pharmacologically relevant concentrations of imatinib. The expression of markers of vascular activation was assessed with real-time PCR. Proliferation was analysed with the cell counting experiments and the MTT assay. Apoptosis was quantified with caspase 3 assays, annexin V in vitro and with TUNEL staining in vivo. Migration was studied with scratch and transwell assays. Tube forming was investigated with the matrigel assay. Imatinib did not alter the expression of markers of vascular activation. Imatinib did not increase the percentage of annexin V positive cells or the activity of caspase 3. No reduction in proliferation or metabolic activity of endothelial cells was observed. Imatinib did not affect migration of endothelial cells and did not reduce the formation of capillary tubes. Consistent with the in vitro data, no difference in the number of apoptotic endothelial cells was observed in vivo in mice treated with imatinib. Imatinib does not inhibit activation, viability, proliferation, migration or tube forming of endothelial cells in vitro and in vivo. Thus, treatment with imatinib might not augment further endothelial cell damage in SSc. PMID:18774958

Influence of irradiation on release of endothelial microparticles (EMP) in vitro.

PubMed

Neuber, Christin; Pufe, Johanna; Pietzsch, Jens

2015-01-01

Survivors of Hodgkin's disease as well as of breast and lung cancer are at risk of radiation-associated cardiovascular disease. Recent studies demonstrated a correlation between cardiovascular risk factors and circulating endothelial microparticles (EMP) and thereby suggest increased EMP levels in circulation to be an early biomarker of endothelial dysfunction and cardiovascular risk. This prompted us to analyze the amount of EMP released by human aortic endothelial cells (HAEC) after exposure to different doses of X-ray (0.4, 2, 4, 6, and 20 Gy) using antibodies against the endothelial cell markers CD31, CD144, and CD146 by flow cytometry. In this pilot experiment only CD146 proved appropriate for quantification of HAEC-derived EMP. Exposure of HAEC to different doses of X-ray did not significantly influence formation of CD146-positive EMP. However, low doses (0.4 Gy) tended to decrease EMP formation, whereas higher doses (2 or 4 Gy) slightly increased release of CD146-positive EMP. By contrast, inflammatory activation of HAEC by TPA significantly increased EMP release about 15-fold (P <  0.01). In conclusion, under the present experimental conditions EMP did not prove a suitable biomarker for radiation-induced endothelial dysfunction in vitro.

Periodontal treatment improves endothelial dysfunction in patients with severe periodontitis.

PubMed

Seinost, Gerald; Wimmer, Gernot; Skerget, Martina; Thaller, Erik; Brodmann, Marianne; Gasser, Robert; Bratschko, Rudolf O; Pilger, Ernst

2005-06-01

Because epidemiological studies provide evidence that periodontal infections are associated with an increased risk of progression of cardiovascular and cerebrovascular disease, we postulated that endothelial dysfunction, a critical element in the pathogenesis of atherosclerosis, would be present in patients with periodontal disease. We tested endothelial function in 30 patients with severe periodontitis and 31 control subjects using flow-mediated dilation (FMD) of the brachial artery. The groups were matched for age, sex, and cardiovascular risk factors. Three months after periodontal treatment, including both mechanical and pharmacological therapy, endothelial function was reassessed by brachial artery FMD. Markers of systemic inflammation were measured at baseline and at follow up. Flow-mediated dilation was significantly lower in patients with periodontitis than in control subjects (6.1% +/- 4.4% vs 8.5% +/- 3.4%, P = .002). Successful periodontal treatment resulted in a significant improvement in FMD (9.8% +/- 5.7%; P = .003 compared to baseline) accompanied by a significant decrease in C-reactive protein concentrations (1.1 +/- 1.9 vs 0.8 +/- 0.8 at baseline, P = .026). Endothelium-independent nitro-induced vasodilation did not differ between the study groups at baseline or after periodontal therapy. These results indicate that treatment of severe periodontitis reverses endothelial dysfunction. Whether improved endothelial function will translate into a beneficial effect on atherogenesis and cardiovascular events needs further investigation.

Intravital imaging of a pulmonary endothelial surface layer in a murine sepsis model.

PubMed

Park, Inwon; Choe, Kibaek; Seo, Howon; Hwang, Yoonha; Song, Eunjoo; Ahn, Jinhyo; Hwan Jo, You; Kim, Pilhan

2018-05-01

Direct intravital imaging of an endothelial surface layer (ESL) in pulmonary microcirculation could be a valuable approach to investigate the role of a vascular endothelial barrier in various pathological conditions. Despite its importance as a marker of endothelial cell damage and impairment of the vascular system, in vivo visualization of ESL has remained a challenging technical issue. In this work, we implemented a pulmonary microcirculation imaging system integrated to a custom-design video-rate laser scanning confocal microscopy platform. Using the system, a real-time cellular-level microscopic imaging of the lung was successfully performed, which facilitated a clear identification of individual flowing erythrocytes in pulmonary capillaries. Subcellular level pulmonary ESL was identified in vivo by fluorescence angiography using a dextran conjugated fluorophore to label blood plasma and the red blood cell (RBC) exclusion imaging analysis. Degradation of ESL width was directly evaluated in a murine sepsis model in vivo , suggesting an impairment of pulmonary vascular endothelium and endothelial barrier dysfunction.

Intravital imaging of a pulmonary endothelial surface layer in a murine sepsis model

PubMed Central

Park, Inwon; Choe, Kibaek; Seo, Howon; Hwang, Yoonha; Song, Eunjoo; Ahn, Jinhyo; Hwan Jo, You; Kim, Pilhan

2018-01-01

Direct intravital imaging of an endothelial surface layer (ESL) in pulmonary microcirculation could be a valuable approach to investigate the role of a vascular endothelial barrier in various pathological conditions. Despite its importance as a marker of endothelial cell damage and impairment of the vascular system, in vivo visualization of ESL has remained a challenging technical issue. In this work, we implemented a pulmonary microcirculation imaging system integrated to a custom-design video-rate laser scanning confocal microscopy platform. Using the system, a real-time cellular-level microscopic imaging of the lung was successfully performed, which facilitated a clear identification of individual flowing erythrocytes in pulmonary capillaries. Subcellular level pulmonary ESL was identified in vivo by fluorescence angiography using a dextran conjugated fluorophore to label blood plasma and the red blood cell (RBC) exclusion imaging analysis. Degradation of ESL width was directly evaluated in a murine sepsis model in vivo, suggesting an impairment of pulmonary vascular endothelium and endothelial barrier dysfunction. PMID:29760995

The Lack of Utility of Circulating Biomarkers of Inflammation and Endothelial Dysfunction for Type 2 Diabetes Risk Prediction Among Postmenopausal Women

PubMed Central

Chao, Chun; Song, Yiqing; Cook, Nancy; Tseng, Chi-Hong; Manson, JoAnn E.; Eaton, Charles; Margolis, Karen L.; Rodriguez, Beatriz; Phillips, Lawrence S.; Tinker, Lesley F.; Liu, Simin

2011-01-01

Background Recent studies have linked plasma markers of inflammation and endothelial dysfunction to type 2 diabetes mellitus (DM) development. However, the utility of these novel biomarkers for type 2 DM risk prediction remains uncertain. Methods The Women’s Health Initiative Observational Study (WHIOS), a prospective cohort, and a nested case-control study within the WHIOS of 1584 incident type 2 DM cases and 2198 matched controls were used to evaluate the utility of plasma markers of inflammation and endothelial dysfunction for type 2 DM risk prediction. Between September 1994 and December 1998, 93 676 women aged 50 to 79 years were enrolled in the WHIOS. Fasting plasma levels of glucose, insulin, white blood cells, tumor necrosis factor receptor 2, interleukin 6, high-sensitivity C-reactive protein, E-selectin, soluble intercellular adhesion molecule 1, and vascular cell adhesion molecule 1 were measured using blood samples collected at baseline. A series of prediction models including traditional risk factors and novel plasma markers were evaluated on the basis of global model fit, model discrimination, net reclassification improvement, and positive and negative predictive values. Results Although white blood cell count and levels of interleukin 6, high-sensitivity C-reactive protein, and soluble intercellular adhesion molecule 1 significantly enhanced model fit, none of the inflammatory and endothelial dysfunction markers improved the ability of model discrimination (area under the receiver operating characteristic curve, 0.93 vs 0.93), net reclassification, or predictive values (positive, 0.22 vs 0.24; negative, 0.99 vs 0.99 [using 15% 6-year type 2 DM risk as the cutoff]) compared with traditional risk factors. Similar results were obtained in ethnic-specific analyses. Conclusion Beyond traditional risk factors, measurement of plasma markers of systemic inflammation and endothelial dysfunction contribute relatively little additional value in clinical type

CAR T Cells Targeting Podoplanin Reduce Orthotopic Glioblastomas in Mouse Brains.

PubMed

Shiina, Satoshi; Ohno, Masasuke; Ohka, Fumiharu; Kuramitsu, Shunichiro; Yamamichi, Akane; Kato, Akira; Motomura, Kazuya; Tanahashi, Kuniaki; Yamamoto, Takashi; Watanabe, Reiko; Ito, Ichiro; Senga, Takeshi; Hamaguchi, Michinari; Wakabayashi, Toshihiko; Kaneko, Mika K; Kato, Yukinari; Chandramohan, Vidyalakshmi; Bigner, Darell D; Natsume, Atsushi

2016-03-01

Glioblastoma (GBM) is the most common and lethal primary malignant brain tumor in adults with a 5-year overall survival rate of less than 10%. Podoplanin (PDPN) is a type I transmembrane mucin-like glycoprotein, expressed in the lymphatic endothelium. Several solid tumors overexpress PDPN, including the mesenchymal type of GBM, which has been reported to present the worst prognosis among GBM subtypes. Chimeric antigen receptor (CAR)-transduced T cells can recognize predefined tumor surface antigens independent of MHC restriction, which is often downregulated in gliomas. We constructed a lentiviral vector expressing a third-generation CAR comprising a PDPN-specific antibody (NZ-1-based single-chain variable fragment) with CD28, 4-1BB, and CD3ζ intracellular domains. CAR-transduced peripheral blood monocytes were immunologically evaluated by calcein-mediated cytotoxic assay, ELISA, tumor size, and overall survival. The generated CAR T cells were specific and effective against PDPN-positive GBM cells in vitro. Systemic injection of the CAR T cells into an immunodeficient mouse model inhibited the growth of intracranial glioma xenografts in vivo. CAR T-cell therapy that targets PDPN would be a promising adoptive immunotherapy to treat mesenchymal GBM. ©2016 American Association for Cancer Research.

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.

Dual targeting of therapeutics to endothelial cells: collaborative enhancement of delivery and effect.

PubMed

Greineder, Colin F; Brenza, Jacob B; Carnemolla, Ronald; Zaitsev, Sergei; Hood, Elizabeth D; Pan, Daniel C; Ding, Bi-Sen; Esmon, Charles T; Chacko, Ann Marie; Muzykantov, Vladimir R

2015-08-01

Anchoring pharmacologic agents to the vascular lumen has the potential to modulate critical processes at the blood-tissue interface, avoiding many of the off-target effects of systemically circulating agents. We report a novel strategy for endothelial dual targeting of therapeutics, which both enhances drug delivery and enables targeted agents to partner enzymatically to generate enhanced biologic effect. Based on the recent discovery that paired antibodies directed to adjacent epitopes of platelet endothelial cell adhesion molecule (PECAM)-1 stimulate each other's binding, we fused single-chain fragments (scFv) of paired anti-mouse PECAM-1 antibodies to recombinant murine thrombomodulin (TM) and endothelial protein C receptor (EPCR), endothelial membrane proteins that partner in activation of protein C (PC). scFv/TM and scFv/EPCR bound to mouse endothelial PECAM-1 with high affinity (EC50 1.5 and 3.8 nM, respectively), and codelivery induced a 5-fold increase in PC activation not seen when TM and EPCR are anchored to distinct cell adhesion molecules. In a mouse model of acute lung injury, dual targeting reduces both the expression of lung inflammatory markers and trans-endothelial protein leak by as much as 40%, as compared to either agent alone. These findings provide proof of principle for endothelial dual targeting, an approach with numerous potential biomedical applications. © FASEB.

Isoflavone supplementation and endothelial function in menopausal women.

PubMed

Hale, Georgina; Paul-Labrador, Maura; Dwyer, James H; Merz, C Noel Bairey

2002-06-01

Despite strong observational evidence for a beneficial role of oestrogen in cardiovascular disease, recent trial results suggest that hormone replacement therapy (HRT) may have adverse effects in menopausal women with established coronary heart disease. Isoflavones are oestrogen analogues found in plants with oestrogen-like properties and, because of a favourable side-effect profile, may be ideal alternatives to HRT with respect to cardiovascular benefits. Endothelial function is a marker of cardiovascular health. We aimed to determine the effect of isoflavones on endothelial function using the brachial artery reactivity test. Twenty-nine healthy menopausal women underwent entry and exit brachial artery reactivity testing following randomization to 2 weeks of an oral soy isoflavone concentrate containing 80 mg of soy isoflavones (Archer Daniel Midland Inc., IL, USA) or placebo. At study exit, there was no difference between placebo and isoflavone groups with respect to flow-mediated dilation (%FMD(max)), change (entry to exit) in %FMD(max) or response to nitroglycerine (%TNG). Subgroup analyses assessing lipid and oestrogen effects did not produce any significant results. These results suggest that short-term oral isoflavone supplements do not improve endothelial function in healthy menopausal women.

Marathon running increases circulating endothelial- and thrombocyte-derived microparticles.

PubMed

Schwarz, Viktoria; Düsing, Philip; Liman, Thomas; Werner, Christian; Herm, Juliane; Bachelier, Katrin; Krüll, Matthias; Brechtel, Lars; Jungehulsing, Gerhard J; Haverkamp, Wilhelm; Böhm, Michael; Endres, Matthias; Haeusler, Karl Georg; Laufs, Ulrich

2018-02-01

Background Acute vascular effects of high intensity physical activity are incompletely characterized. Circulating microparticles are cellular markers for vascular activation and damage. Methods Microparticles were analysed in 99 marathon runners (49 ± 6 years, 22% female) of the prospective Berlin Beat of Running study. Blood samples were taken within three days before, immediately after and within two days after the marathon run. Endothelial-derived microparticles were labelled with CD144, CD31 and CD62E, platelet-derived microparticles with CD62P and CD42b, leukocyte-derived microparticles with CD45 and monocyte-derived microparticles with CD14. Results Marathon running induced leukocytosis (5.9 ± 0.1 to 14.8 ± 0.3 10 9 /l, p < 0.0001) and increased platelet counts (239 ± 4.6 to 281 ± 5.9 10 9 /l, p < 0.0001) immediately after the marathon. Blood monocytes increased and lymphocytes decreased after the run ( p < 0.0001). Endothelial-derived microparticles were acutely increased ( p = 0.008) due to a 23% increase of apoptotic endothelial-derived microparticles ( p = 0.007) and returned to baseline within two days after the marathon. Thrombocyte-derived microparticles acutely increased by 38% accompanied by an increase in activated and apoptotic thrombocyte-derived microparticles ( p ≤ 0.0001) each. Both monocyte- and leukocyte-derived microparticles were decreased immediately after marathon run ( p < 0.0001) and remained below baseline until day 2. Troponin T increased from 12 to 32 ng/l ( p < 0.0001) immediately after the run and returned to baseline after two days. Conclusion Circulating apoptotic endothelial- and thrombocyte-derived microparticles increased after marathon running consistent with an acute pro-thrombotic and pro-inflammatory state. Exercise-induced vascular damage reflected by microparticles could indicate potential mechanisms of post-exertional cardiovascular complications. Further

A Decrease in Glomerular Endothelial Cells and Endothelial-mesenchymal Transition during Glomerulosclerosis in the Tensin2-deficient Mice (ICGN strain).

PubMed

Kato, Takashi; Mizuno, Shinya; Ito, Akihiko

2014-01-01

The ICR-derived glomerulonephritis (ICGN) mouse is a unique model of nephrotic syndrome, and albuminuria becomes evident in a neonatal stage, due to a genetic mutation of tensin2. We previously provided evidence that an apparent decrease in nephrin, caused by tensin2-deficiencient states, leads to podocytopathy, albuminuria and eventually, chronic renal failure. In general, glomerular endothelial cells (ECs) function as a barrier through tight attachment of glomerular basement membrane to podocytes, while decreased ECs can worsen renal failure. Nevertheless, it is still unknown whether glomerular ECs are altered under the tensin-2-deficient states during the manifestation of chronic renal failure. Herein, we examined the changes of glomerular ECs, with focus on the expression of PECAM-1 and VE-cadherin (EC-specific markers), or of α-SMA (myofibroblast marker) in this mouse model by histological methods. Compared with the non-nephrotic (+/nep) mice, the nephrotic (nep/nep) mice exhibited the reduced expression of PECAM-1, or of VE-cadherin, in glomerular area. Notably, some glomerular ECs showed the positive stainings for both PECAM-1 and α-SMA, suggesting endothelial-to-mesenchymal transition (EndoMT) during progression of glomerular sclerosis. This is the first report showing that a decrease in glomerular ECs, at least in part, via EndoMT is involved in tensin2-deficient pathological conditions.

Endocannabinoid receptor blockade increases vascular endothelial growth factor and inflammatory markers in obese women with polycystic ovary syndrome.

PubMed

Sathyapalan, Thozhukat; Javed, Zeeshan; Kilpatrick, Eric S; Coady, Anne-Marie; Atkin, Stephen L

2017-03-01

Animal studies suggest that cannabinoid receptor-1 (CB-1) blockade reduces inflammation and neovascularization by decreasing vascular endothelial growth factor (VEGF) levels associated with a reduction in inflammatory markers, thereby potentially reducing cardiovascular risk. To determine the impact of CB1 antagonism by rimonabant on VEGF and inflammatory markers in obese PCOS women. Randomized, open-labelled parallel study. Endocrinology outpatient clinic in a referral centre. Twenty patients with PCOS (PCOS) and biochemical hyperandrogenaemia with a body mass index of ≥30 kg/m 2 were recruited. Patients were randomized to 1·5 g daily of metformin or 20 mg daily of rimonabant. Post hoc review to detect VEGF and pro-inflammatory cytokines TNF-α, IL-1β, IL-1ra, IL-2, IL6, IL-8, IL-10 and MCP-1 before and after 12 weeks of treatment. After 12 weeks of rimonabant treatment, there was a significant increase in VEGF (99·2 ± 17·6 vs 116·2 ± 15·8 pg/ml, P < 0·01) and IL-8 (7·4 ± 11·0 vs 18·1 ± 13·2 pg/ml, P < 0·05) but not after metformin (VEGF P = 0·7; IL-8 P = 0·9). There was no significant difference in the pro-inflammatory cytokines TNF-α, IL-1β, IL-1ra, IL-2, IL6, IL-8, IL-10 and MCP-1 following either treatment. This study suggests that rimonabant CB-I blockade paradoxically raised VEGF and the cytokine IL-8 in obese women with PCOS that may have offset the potential benefit associated with weight loss. © 2016 John Wiley & Sons Ltd.

A Cell Culture Approach to Optimized Human Corneal Endothelial Cell Function

PubMed Central

Bartakova, Alena; Kuzmenko, Olga; Alvarez-Delfin, Karen; Kunzevitzky, Noelia J.; Goldberg, Jeffrey L.

2018-01-01

Purpose Cell-based therapies to replace corneal endothelium depend on culture methods to optimize human corneal endothelial cell (HCEC) function and minimize endothelial-mesenchymal transition (EnMT). Here we explore contribution of low-mitogenic media on stabilization of phenotypes in vitro that mimic those of HCECs in vivo. Methods HCECs were isolated from cadaveric donor corneas and expanded in vitro, comparing continuous presence of exogenous growth factors (“proliferative media”) to media without those factors (“stabilizing media”). Identity based on canonical morphology and expression of surface marker CD56, and function based on formation of tight junction barriers measured by trans-endothelial electrical resistance assays (TEER) were assessed. Results Primary HCECs cultured in proliferative media underwent EnMT after three to four passages, becoming increasingly fibroblastic. Stabilizing the cells before each passage by switching them to a media low in mitogenic growth factors and serum preserved canonical morphology and yielded a higher number of cells. HCECs cultured in stabilizing media increased both expression of the identity marker CD56 and also tight junction monolayer integrity compared to cells cultured without stabilization. Conclusions HCECs isolated from donor corneas and expanded in vitro with a low-mitogenic media stabilizing step before each passage demonstrate more canonical structural and functional features and defer EnMT, increasing the number of passages and total canonical cell yield. This approach may facilitate development of HCEC-based cell therapies. PMID:29625488

Circulating endothelial cells are increased in chronic myeloid leukemia blast crisis.

PubMed

Godoy, C R T; Levy, D; Giampaoli, V; Chamone, D A F; Bydlowski, S P; Pereira, J

2015-06-01

We measured circulating endothelial precursor cells (EPCs), activated circulating endothelial cells (aCECs), and mature circulating endothelial cells (mCECs) using four-color multiparametric flow cytometry in the peripheral blood of 84 chronic myeloid leukemia (CML) patients and 65 healthy controls; and vascular endothelial growth factor (VEGF) by quantitative real-time PCR in 50 CML patients and 32 healthy controls. Because of an increase in mCECs, the median percentage of CECs in CML blast crisis (0.0146%) was significantly higher than in healthy subjects (0.0059%, P<0.01) and in the accelerated phase (0.0059%, P=0.01). There were no significant differences in the percentages of CECs in chronic- or active-phase patients and healthy subjects (P>0.05). In addition, VEGF gene expression was significantly higher in all phases of CML: 0.245 in blast crisis, 0.320 in the active phase, and 0.330 in chronic phase patients than it was in healthy subjects (0.145). In conclusion, CML in blast crisis had increased levels of CECs and VEGF gene expression, which may serve as markers of disease progression and may become targets for the management of CML.

Endothelial Progenitor Cells and Kidney Diseases.

PubMed

Ozkok, Abdullah; Yildiz, Alaattin

2018-05-10

Endothelial progenitor cells (EPC) are bone marrow derived or tissue-resident cells that play major roles in the maintenance of vascular integrity and repair of endothelial damage. Although EPCs may be capable of directly engrafting and regenerating the endothelium, the most important effects of EPCs seem to be depended on paracrine effects. In recent studies, specific microvesicles and mRNAs have been found to mediate the pro-angiogenic and regenerative effects of EPCs on endothelium. EPC counts have important prognostic implications in cardiovascular diseases (CVD). Uremia and inflammation are associated with lower EPC counts which probably contribute to increased CVD risks in patients with chronic kidney disease. Beneficial effects of the EPC therapies have been shown in studies performed on different models of CVD and kidney diseases such as acute and chronic kidney diseases and glomerulonephritis. However, lack of a clear definition and specific marker of EPCs is the most important problem causing difficulties in interpretation of the results of the studies investigating EPCs. © 2018 The Author(s). Published by S. Karger AG, Basel.

Expansion and cryopreservation of porcine and human corneal endothelial cells.

PubMed

Marquez-Curtis, Leah A; McGann, Locksley E; Elliott, Janet A W

2017-08-01

Impairment of the corneal endothelium causes blindness that afflicts millions worldwide and constitutes the most often cited indication for corneal transplants. The scarcity of donor corneas has prompted the alternative use of tissue-engineered grafts which requires the ex vivo expansion and cryopreservation of corneal endothelial cells. The aims of this study are to culture and identify the conditions that will yield viable and functional corneal endothelial cells after cryopreservation. Previously, using human umbilical vein endothelial cells (HUVECs), we employed a systematic approach to optimize the post-thaw recovery of cells with high membrane integrity and functionality. Here, we investigated whether improved protocols for HUVECs translate to the cryopreservation of corneal endothelial cells, despite the differences in function and embryonic origin of these cell types. First, we isolated endothelial cells from pig corneas and then applied an interrupted slow cooling protocol in the presence of dimethyl sulfoxide (Me 2 SO), with or without hydroxyethyl starch (HES). Next, we isolated and expanded endothelial cells from human corneas and applied the best protocol verified using porcine cells. We found that slow cooling at 1 °C/min in the presence of 5% Me 2 SO and 6% HES, followed by rapid thawing after liquid nitrogen storage, yields membrane-intact cells that could form monolayers expressing the tight junction marker ZO-1 and cytoskeleton F-actin, and could form tubes in reconstituted basement membrane matrix. Thus, we show that a cryopreservation protocol optimized for HUVECs can be applied successfully to corneal endothelial cells, and this could provide a means to address the need for off-the-shelf cryopreserved cells for corneal tissue engineering and regenerative medicine. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Gene expression profiles of brain endothelial cells during embryonic development at bulk and single-cell levels.

PubMed

Hupe, Mike; Li, Minerva Xueting; Kneitz, Susanne; Davydova, Daria; Yokota, Chika; Kele-Olovsson, Julianna; Hot, Belma; Stenman, Jan M; Gessler, Manfred

2017-07-11

The blood-brain barrier is a dynamic interface that separates the brain from the circulatory system, and it is formed by highly specialized endothelial cells. To explore the molecular mechanisms defining the unique nature of vascular development and differentiation in the brain, we generated high-resolution gene expression profiles of mouse embryonic brain endothelial cells using translating ribosome affinity purification and single-cell RNA sequencing. We compared the brain vascular translatome with the vascular translatomes of other organs and analyzed the vascular translatomes of the brain at different time points during embryonic development. Because canonical Wnt signaling is implicated in the formation of the blood-brain barrier, we also compared the brain endothelial translatome of wild-type mice with that of mice lacking the transcriptional cofactor β-catenin ( Ctnnb1 ). Our analysis revealed extensive molecular changes during the embryonic development of the brain endothelium. We identified genes encoding brain endothelium-specific transcription factors ( Foxf2 , Foxl2 , Foxq1 , Lef1 , Ppard , Zfp551 , and Zic3 ) that are associated with maturation of the blood-brain barrier and act downstream of the Wnt-β-catenin signaling pathway. Profiling of individual brain endothelial cells revealed substantial heterogeneity in the population. Nevertheless, the high abundance of Foxf2 , Foxq1 , Ppard , or Zic3 transcripts correlated with the increased expression of genes encoding markers of brain endothelial cell differentiation. Expression of Foxf2 and Zic3 in human umbilical vein endothelial cells induced the production of blood-brain barrier differentiation markers. This comprehensive data set may help to improve the engineering of in vitro blood-brain barrier models. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

The effect of bisphosphonates on the endothelial differentiation of mesenchymal stem cells

PubMed Central

Sharma, Dileep; Hamlet, Stephen Mark; Petcu, Eugen Bogdan; Ivanovski, Saso

2016-01-01

The contribution of the local stem cell niche to providing an adequate vascular framework during healing cannot be overemphasized. Bisphosphonates (BPs) are known to have a direct effect on the local vasculature, but their effect on progenitor cell differentiation is unknown. This in vitro study evaluated the effect(s) of various BPs on the differentiation of human placental mesenchymal stem cells (pMSCs) along the endothelial lineage and their subsequent functional and morphogenic capabilities. pMSC multipotency was confirmed by successful differentiation into cells of both the osteogenic and endothelial lineages, as demonstrated by positive Alizarin Red S staining and Ac-LDL uptake. pMSC differentiation in the presence of non-cytotoxic BP concentrations showed that nitrogen containing BPs had a significant inhibitory effect on cell migration and endothelial marker gene expression, as well as compromised endothelial differentiation as demonstrated using von Willebrand factor immunofluorescence staining and tube formation assay. This in vitro study demonstrated that at non-cytotoxic levels, nitrogen-containing BPs inhibit differentiation of pMSCs into cells of an endothelial lineage and affect the downstream functional capability of these cells supporting a multi-modal effect of BPs on angiogenesis as pathogenic mechanism contributing to bone healing disorders such as bisphosphonate related osteonecrosis of the jaws (BRONJ). PMID:26857282

Automated Deep Learning-Based System to Identify Endothelial Cells Derived from Induced Pluripotent Stem Cells.

PubMed

Kusumoto, Dai; Lachmann, Mark; Kunihiro, Takeshi; Yuasa, Shinsuke; Kishino, Yoshikazu; Kimura, Mai; Katsuki, Toshiomi; Itoh, Shogo; Seki, Tomohisa; Fukuda, Keiichi

2018-06-05

Deep learning technology is rapidly advancing and is now used to solve complex problems. Here, we used deep learning in convolutional neural networks to establish an automated method to identify endothelial cells derived from induced pluripotent stem cells (iPSCs), without the need for immunostaining or lineage tracing. Networks were trained to predict whether phase-contrast images contain endothelial cells based on morphology only. Predictions were validated by comparison to immunofluorescence staining for CD31, a marker of endothelial cells. Method parameters were then automatically and iteratively optimized to increase prediction accuracy. We found that prediction accuracy was correlated with network depth and pixel size of images to be analyzed. Finally, K-fold cross-validation confirmed that optimized convolutional neural networks can identify endothelial cells with high performance, based only on morphology. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Oral cancer/endothelial cell fusion experiences nuclear fusion and acquisition of enhanced survival potential

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

Song, Kai; The State Key Laboratory Breeding Base of Basic Science of Stomatology; Song, Yong

Most previous studies have linked cancer–macrophage fusion with tumor progression and metastasis. However, the characteristics of hybrid cells derived from oral cancer and endothelial cells and their involvement in cancer remained unknown. Double-immunofluorescent staining and fluorescent in situ hybridization (FISH) were performed to confirm spontaneous cell fusion between eGFP-labeled human umbilical vein endothelial cells (HUVECs) and RFP-labeled SCC9, and to detect the expression of vementin and cytokeratin 18 in the hybrids. The property of chemo-resistance of such hybrids was examined by TUNEL assay. The hybrid cells in xenografted tumor were identified by FISH and GFP/RFP dual-immunofluoresence staining. We showed thatmore » SCC9 cells spontaneously fused with cocultured endothelial cells, and the resultant hybrid cells maintained the division and proliferation activity after re-plating and thawing. Such hybrids expressed markers of both parental cells and became more resistant to chemotherapeutic drug cisplatin as compared to the parental SCC9 cells. Our in vivo data indicated that the hybrid cells contributed to tumor composition by using of immunostaining and FISH analysis, even though the hybrid cells and SCC9 cells were mixed with 1:10,000, according to the FACS data. Our study suggested that the fusion events between oral cancer and endothelial cells undergo nuclear fusion and acquire a new property of drug resistance and consequently enhanced survival potential. These experimental findings provide further supportive evidence for the theory that cell fusion is involved in cancer progression. - Highlights: • The fusion events between oral cancer and endothelial cells undergo nuclear fusion. • The resulting hybrid cells acquire a new property of drug resistance. • The resulting hybrid cells express the markers of both parental cells (i.e. vimentin and cytokeratin 18). • The hybrid cells contribute to tumor repopulation in vivo.« less

Cross talk between primary human renal tubular cells and endothelial cells in cocultures.

PubMed

Tasnim, Farah; Zink, Daniele

2012-04-15

Interactions between renal tubular epithelial cells and adjacent endothelial cells are essential for normal renal functions but also play important roles in renal disease and repair. Here, we investigated cocultures of human primary renal proximal tubular cells (HPTC) and human primary endothelial cells to address the cross talk between these cell types. HPTC showed improved proliferation, marker gene expression, and enzyme activity in cocultures. Also, the long-term maintenance of epithelia formed by HPTC was improved, which was due to the secretion of transforming growth factor-β1 and its antagonist α2-macroglobulin. HPTC induced endothelial cells to secrete increased amounts of these factors, which balanced each other functionally and only displayed in combination the observed positive effects. In addition, in the presence of HPTC endothelial cells expressed increased amounts of hepatocyte growth factor and vascular endothelial growth factor, which have well-characterized effects on renal tubular epithelial cells as well as on endothelial cells. Together, the results showed that HPTC stimulated endothelial cells to express a functionally balanced combination of various factors, which in turn improved the performance of HPTC. The results give new insights into the cross talk between renal epithelial and endothelial cells and suggest that cocultures could be also useful models for the analysis of cellular communication in renal disease and repair. Furthermore, the characterization of defined microenvironments, which positively affect HPTC, will be helpful for improving the performance of this cell type in in vitro applications including in vitro toxicology and kidney tissue engineering.

Dual targeting of therapeutics to endothelial cells: collaborative enhancement of delivery and effect

PubMed Central

Greineder, Colin F.; Brenza, Jacob B.; Carnemolla, Ronald; Zaitsev, Sergei; Hood, Elizabeth D.; Pan, Daniel C.; Ding, Bi-Sen; Esmon, Charles T.; Chacko, Ann Marie; Muzykantov, Vladimir R.

2015-01-01

Anchoring pharmacologic agents to the vascular lumen has the potential to modulate critical processes at the blood–tissue interface, avoiding many of the off-target effects of systemically circulating agents. We report a novel strategy for endothelial dual targeting of therapeutics, which both enhances drug delivery and enables targeted agents to partner enzymatically to generate enhanced biologic effect. Based on the recent discovery that paired antibodies directed to adjacent epitopes of platelet endothelial cell adhesion molecule (PECAM)-1 stimulate each other’s binding, we fused single-chain fragments (scFv) of paired anti-mouse PECAM-1 antibodies to recombinant murine thrombomodulin (TM) and endothelial protein C receptor (EPCR), endothelial membrane proteins that partner in activation of protein C (PC). scFv/TM and scFv/EPCR bound to mouse endothelial PECAM-1 with high affinity (EC50 1.5 and 3.8 nM, respectively), and codelivery induced a 5-fold increase in PC activation not seen when TM and EPCR are anchored to distinct cell adhesion molecules. In a mouse model of acute lung injury, dual targeting reduces both the expression of lung inflammatory markers and trans-endothelial protein leak by as much as 40%, as compared to either agent alone. These findings provide proof of principle for endothelial dual targeting, an approach with numerous potential biomedical applications.—Greineder, C. F., Brenza, J. B., Carnemolla, R., Zaitsev, S., Hood, E. D., Pan, D. C., Ding, B.-S., Esmon, C. T., Chacko, A. M., Muzykantov, V. R. Dual targeting of therapeutics to endothelial cells: collaborative enhancement of delivery and effect. PMID:25953848

Induction of vascular endothelial phenotype and cellular proliferation from human cord blood stem cells cultured in simulated microgravity

NASA Astrophysics Data System (ADS)

Chiu, Brian; Z-M Wan, Jim; Abley, Doris; Akabutu, John

2005-05-01

Recent studies have demonstrated that stem cells derived from adult hematopoietic tissues are capable of trans-differentiation into non-hematopoietic cells, and that the culture in microgravity ( μg) may modulate the proliferation and differentiation. We investigated the application of μg to human umbilical cord blood stem cells (CBSC) in the induction of vascular endothelial phenotype expression and cellular proliferation. CD34+ mononuclear cells were isolated from waste human umbilical cord blood samples and cultured in simulated μg for 14 days. The cells were seeded in rotary wall vessels (RWV) with or without microcarrier beads (MCB) and vascular endothelial growth factor was added during culture. Controls consisted of culture in 1 G. The cell cultures in RWV were examined by inverted microscopy. Cell counts, endothelial cell and leukocyte markers performed by flow-cytometry and FACS scan were assayed at days 1, 4, 7 and at the termination of the experiments. Culture in RWV revealed significantly increased cellular proliferation with three-dimensional (3D) tissue-like aggregates. At day 4, CD34+ cells cultured in RWV bioreactor without MCB developed vascular tubular assemblies and exhibited endothelial phenotypic markers. These data suggest that CD34+ human umbilical cord blood progenitors are capable of trans-differentiation into vascular endothelial cell phenotype and assemble into 3D tissue structures. Culture of CBSC in simulated μg may be potentially beneficial in the fields of stem cell biology and somatic cell therapy.

(-)-Epicatechin-induced recovery of mitochondria from simulated diabetes: Potential role of endothelial nitric oxide synthase.

PubMed

Ramírez-Sánchez, Israel; Rodríguez, Alonso; Moreno-Ulloa, Aldo; Ceballos, Guillermo; Villarreal, Francisco

2016-05-01

(-)-Epicatechin increases indicators associated with mitochondrial biogenesis in endothelial cells and myocardium. We investigated endothelial nitric oxide synthase involvement on (-)-epicatechin-induced increases in indicators associated with mitochondrial biogenesis in human coronary artery endothelial cells cultured in normal-glucose and high-glucose media, as well as to restore indicators of cardiac mitochondria from the effects of simulated diabetes. Here, we demonstrate the role of endothelial nitric oxide synthase on (-)-epicatechin-induced increases in mitochondrial proteins, transcription factors and sirtuin 1 under normal-glucose conditions. In simulated diabetes endothelial nitric oxide synthase function, mitochondrial function-associated and biogenesis-associated indicators were adversely impacted by high glucose, effects that were reverted by (-)-epicatechin. As an animal model of type 2 diabetes, 2-month old C57BL/6 mice were fed a high-fat diet for 16 weeks. Fasting and fed blood glucose levels were increased and NO plasma levels decreased. High-fat-diet-fed mice myocardium revealed endothelial nitric oxide synthase dysfunction, reduced mitochondrial activity and markers of mitochondrial biogenesis. The administration of 1 mg/kg (-)-epicatechin for 15 days by oral gavage shifted these endpoints towards control mice values. Results suggest that endothelial nitric oxide synthase mediates (-)-epicatechin-induced increases of indicators associated with mitochondrial biogenesis in endothelial cells. (-)-Epicatechin also counteracts the negative effects that high glucose or simulated type 2 diabetes has on endothelial nitric oxide synthase function. © The Author(s) 2016.

Maggot debridement therapy promotes diabetic foot wound healing by up-regulating endothelial cell activity.

PubMed

Sun, Xinjuan; Chen, Jin'an; Zhang, Jie; Wang, Wei; Sun, Jinshan; Wang, Aiping

2016-03-01

To determine the role of maggot debridement therapy (MDT) on diabetic foot wound healing, we compared growth related factors in wounds before and after treatment. Furthermore, we utilized human umbilical vein endothelial cells (HUVECs) to explore responses to maggot excretions/secretions on markers of angiogenesis and proliferation. The results showed that there was neo-granulation and angiogenesis in diabetic foot wounds after MDT. Moreover, significant elevation in CD34 and CD68 levels was also observed in treated wounds. In vitro, ES increased HUVEC proliferation, improved tube formation, and increased expression of vascular endothelial growth factor receptor 2 in a dose dependent manner. These results demonstrate that MDT and maggot ES can promote diabetic foot wound healing by up-regulating endothelial cell activity. Copyright © 2016. Published by Elsevier Inc.

Epidermal growth factor-like domain 7 is a marker of the endothelial lineage and active angiogenesis.

PubMed

Bambino, Kathryn; Lacko, Lauretta A; Hajjar, Katherine A; Stuhlmann, Heidi

2014-07-01

Epidermal growth factor-like domain 7 (Egfl7) expression in the developing embryo is largely restricted to sites of mesodermal progenitors of angioblasts/hemangioblasts and the vascular endothelium. We hypothesize that Egfl7 marks the endothelial lineage during embryonic development, and can be used to define the emergence of endothelial progenitor cells, as well as to visualize newly-forming vasculature in the embryo and during the processes of physiologic and pathologic angiogenesis in the adult. We have generated a transgenic mouse strain that expresses enhanced green fluorescent protein (eGFP) under the control of a minimal Egfl7 regulatory sequence (Egfl7:eGFP). Expression of the transgene recapitulated that of endogenous Egfl7 at sites of vasculogenesis and angiogenesis in the allantois, yolk sac, and in the embryo proper. The transgene was not expressed in the quiescent endothelium of most adult organs. However, the uterus and ovary, which undergo vascular growth and remodeling throughout the estrus cycle, expressed high levels of Egfl7:eGFP. Importantly, expression of the Egfl7:eGFP transgene was induced in adult neovasculature. We also found that increased Egfl7 expression contributed to pathologic revascularization in the mouse retina. To our knowledge, this is the first mouse model that enables monitoring of endothelial cells at sites of active vasculogenesis and angiogenesis. This model also facilitated the isolation and characterization of EGFL7(+) endothelial cell populations by fluorescence activated cell sorting (FACS). Together, our results demonstrate that the Egfl7:eGFP reporter mouse is a valuable tool that can be used to elucidate the mechanisms by which blood vessels form during development and under pathologic circumstances. © 2014 Wiley Periodicals, Inc.

The predictive value of markers of fibrinolysis and endothelial dysfunction in the post thrombotic syndrome. A systematic review.

PubMed

Rabinovich, Anat; Cohen, Jacqueline M; Kahn, Susan R

2014-06-01

The post thrombotic syndrome (PTS) develops in 20-40% of deep venous thrombosis (DVT) patients. Risk factors for PTS have not been well elucidated. Identification of risk factors would facilitate individualised risk assessment for PTS. We conducted a systematic review to determine whether biomarkers of fibrinolysis or endothelial dysfunction can predict the risk for PTS among DVT patients. Studies were identified by searching the electronic databases PubMed, EMBASE, Scopus and Web of science. We included studies published between 1990 and 2013, measured biomarker levels in adult DVT patients, and reported rates of PTS development. Fourteen studies were included: 11 investigated the association between D-dimer and PTS; three examined fibrinogen; two measured von Willebrand factor; one measured plasminogen activator inhibitor-1; one assessed ADAMTS-13 (A Disintegrin and Metalloprotease with Thrombospondin type 1 repeats) and one measured factor XIII activity. Studies varied with regards to inclusion criteria, definition of PTS, time point and method of biomarker measurement. We were unable to meta-analyse results due to marked clinical heterogeneity. Descriptively, a significant association with PTS was found for D-dimer in four studies and factor XIII in one study. Further prospective research is needed to elucidate whether these markers might be useful to predict PTS development.

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

Circulating Endothelial Cells and Endothelial Function predict Major Adverse Cardiac Events and Early Adverse Left Ventricular Remodeling in Patients with ST-Segment Elevation Myocardial Infarction

PubMed Central

Magdy, Abdel Hamid; Bakhoum, Sameh; Sharaf, Yasser; Sabry, Dina; El-Gengehe, Ahmed T; Abdel-Latif, Ahmed

2016-01-01

Endothelial progenitor cells (EPCs) and circulating endothelial cells (CECs) are mobilized from the bone marrow and increase in the early phase after ST-elevation myocardial infarction (STEMI). The aim of this study was to assess the prognostic significance of CECs and indices of endothelial dysfunction in patients with STEMI. In 78 patients with acute STEMI, characterization of CD34+/VEGFR2+ CECs, and indices of endothelial damage/dysfunction such as brachial artery flow mediated dilatation (FMD) were determined. Blood samples for CECs assessment and quantification were obtained within 24 hours of admission and FMD was assessed during the index hospitalization. At 30 days follow up, the primary composite end point of major cardiac adverse events (MACE) consisting of all-cause mortality, recurrent non-fatal MI, or heart failure and the secondary endpoint of early adverse left ventricular (LV) remodeling were analyzed. The 17 patients (22%) who developed MACE had significantly higher CEC level (P = 0.004), vWF level (P =0.028), and significantly lower FMD (P = 0.006) compared to the remaining patients. Logistic regression analysis showed that CECs level and LV ejection fraction were independent predictors of MACE. The areas under the receiver operating characteristic curves (ROC) for CEC level, FMD, and the logistic model with both markers were 0.73, 0.75, and 0.82 respectively for prediction of the MACE. The 16 patients who developed the secondary endpoint had significantly higher CEC level compared to remaining patients (p =0.038). In conclusion, increased circulating endothelial cells and endothelial dysfunction predicted the occurrence of major adverse cardiac events and adverse cardiac remodeling in patients with STEMI. PMID:26864952

In Vitro Impact of Conditioned Medium From Demineralized Freeze-Dried Bone on Human Umbilical Endothelial Cells.

PubMed

Harnik, Branko; Miron, Richard J; Buser, Daniel; Gruber, Reinhard

2017-03-01

Angiogenesis is essential for the consolidation of bone allografts. The underlying molecular mechanism, however, remains unclear. Soluble factors released from demineralized freeze-dried bone target mesenchymal cells; however, their effect on endothelial cells has not been investigated so far. The aim of the present study was therefore to examine the effect of conditioned medium from demineralized freeze-dried bone on human umbilical endothelial cells in vitro. Conditioned medium was first prepared from demineralized freeze-dried bone following 24 hours incubation at room temperature to produce demineralized bone conditioned media. Thereafter, conditioned medium was used to stimulate human umbilical vein endothelial cells in vitro by determining the cell response based on viability, proliferation, expression of apoptotic genes, a Boyden chamber to determine cell migration, and the formation of branches. The authors report here that conditioned medium decreased viability and proliferation of endothelial cells. Neither of the apoptotic marker genes was significantly altered when endothelial cells were exposed to conditioned medium. The Boyden chamber revealed that endothelial cells migrate toward conditioned medium. Moreover, conditioned medium moderately stimulated the formation of branches. These findings support the concept that conditioned medium from demineralized freeze-dried bone targets endothelial cells by decreasing their proliferation and enhancing their motility under these in vitro conditions.

Suppression of endothelial cell adhesion by XJP-1, a new phenolic compound derived from banana peel.

PubMed

Fu, Rong; Yan, Tianhua; Wang, Qiujuan; Guo, Qinglong; Yao, Hequan; Wu, Xiaoming; Li, Yang

2012-01-01

The adhesion of monocytes to activated vascular endothelial cells is a critical event in the initiation of atherosclerosis. Adhesion is mediated by oxidized low-density lipoprotein (ox-LDL) which up-regulates inflammatory markers on endothelial cells. Here we report that (±) 7, 8-dihydroxy-3-methyl-isochromanone-4 (XJP-1), an inhibitor of ox-LDL-induced adhesion of monocytes to endothelial cells blocks cellular functions which are associated with adhesion. We show that XJP-1 down-regulates ox-LDL-induced over-expression of adhesion molecules (ICAM-1 and VCAM-1) in a dose-dependent manner in human umbilical vein endothelial cells (HUVECs), attenuates ox-LDL-induced up-regulation of low-density lipoprotein receptor (LOX)-1, decreases generation of reactive oxygen species (ROS), blocks translocation of nuclear factor-kappa B (NF-κB) activity, and prevents activation of c-Jun N-terminal kinase (JNK)/p38 pathways in endothelial cells. These findings suggest that XJP-1 may attenuate ox-LDL-induced endothelial adhesion of monocytes by blocking expression of adhesion molecules through suppressing ROS/NF-κB, JNK and p38 pathways. Copyright © 2012 Elsevier Inc. All rights reserved.

High-Mobility Group Box 1 From Hypoxic Trophoblasts Promotes Endothelial Microparticle Production and Thrombophilia in Preeclampsia.

PubMed

Hu, Yae; Yan, Ruhong; Zhang, Ce; Zhou, Zhichao; Liu, Meng; Wang, Can; Zhang, Hong; Dong, Liang; Zhou, Tiantian; Wu, Yi; Dong, Ningzheng; Wu, Qingyu

2018-04-12

Thrombophilia is a major complication in preeclampsia, a disease associated with placental hypoxia and trophoblast inflammation. Preeclampsia women are known to have increased circulating microparticles that are procoagulant, but the underlying mechanisms remain unclear. In this study, we sought to understand the mechanism connecting placental hypoxia, circulating microparticles, and thrombophilia. We analyzed protein markers on plasma microparticles from preeclampsia women and found that the increased circulating microparticles were mostly from endothelial cells. In proteomic studies, we identified HMGB1 (high-mobility group box 1), a proinflammatory protein, as a key factor from hypoxic trophoblasts in stimulating microparticle production in human umbilical vein endothelial cells. Immunodepletion or inhibition of HMGB1 in the conditioned medium from hypoxic human trophoblasts abolished the endothelial microparticle-stimulating activity. Conversely, recombinant HMGB1 stimulated microparticle production in cultured human umbilical vein endothelial cells. The microparticles from recombinant HMGB1-stimulated human umbilical vein endothelial cells promoted blood coagulation and neutrophil activation in vitro. Injection of recombinant HMGB1 in pregnant mice increased plasma endothelial microparticles and promoted blood coagulation. In preeclampsia women, elevated placental HMGB1 expression was detected and high levels of plasma HMGB1 correlated with increased plasma endothelial microparticles. Our results indicate that placental hypoxia-induced HMGB1 expression and release from trophoblasts are important mechanism underlying increased circulating endothelial microparticles and thrombophilia in preeclampsia. © 2018 American Heart Association, Inc.

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

Placental oxidative stress and maternal endothelial function in pregnant women with normotensive fetal growth restriction.

PubMed

Yoshida, Atsumi; Watanabe, Kazushi; Iwasaki, Ai; Kimura, Chiharu; Matsushita, Hiroshi; Wakatsuki, Akihiko

2018-04-01

The purpose of this study was to investigate the relationship between placental oxidative stress and maternal endothelial function in pregnant women with normotensive fetal growth restriction (FGR). We examined serum concentrations of oxygen free radicals (d-ROMs), maternal angiogenic factor (PlGF), and sFlt-1, placental oxidative DNA damage, and maternal endothelial function in 17 women with early-onset preeclampsia (PE), 18 with late-onset PE, 14 with normotensive FGR, and 21 controls. Flow-mediated vasodilation (FMD) was assessed as a marker of maternal endothelial function. Immunohistochemical analysis was performed to measure the proportion of placental trophoblast cell nuclei staining positive for 8-hydroxy-2'-deoxyguanosine (8-OHdG), a marker of oxidative DNA damage. Maternal serum d-ROM, sFlt-1 concentrations, and FMD did not significantly differ between the control and normotensive FGR groups. The proportion of nuclei staining positive for 8-OHdG was significantly higher in the normotensive FGR group relative to the control group. Our findings demonstrate that, despite the presence of placental oxidative DNA damage as observed in PE patients, pregnant women with normotensive FGR show no increase in the concentrations of sFlt-1 and d-ROMs, or a decrease in FMD.

Storage and regulated secretion of factor VIII in blood outgrowth endothelial cells

PubMed Central

van den Biggelaar, Maartje; Bouwens, Eveline A.M.; Kootstra, Neeltje A.; Hebbel, Robert P.; Voorberg, Jan; Mertens, Koen

2009-01-01

Background Gene therapy provides an attractive alternative for protein replacement therapy in hemophilia A patients. Recent studies have shown the potential benefit of directing factor (F)VIII gene delivery to cells that also express its natural carrier protein von Willebrand factor (VWF). In this study, we explored the feasibility of blood outgrowth endothelial cells as a cellular FVIII delivery device with particular reference to long-term production levels, intracellular storage in Weibel-Palade bodies and agonist-induced regulated secretion. Design and Methods Human blood outgrowth endothelial cells were isolated from peripheral blood collected from healthy donors, transduced at passage 5 using a lentiviral vector encoding human B-domain deleted FVIII-GFP and characterized by flow cytometry and confocal microscopy. Results Blood outgrowth endothelial cells displayed typical endothelial morphology and expressed the endothelial-specific marker VWF. Following transduction with a lentivirus encoding FVIII-GFP, 80% of transduced blood outgrowth endothelial cells expressed FVIII-GFP. Levels of FVIII-GFP positive cells declined slowly upon prolonged culturing. Transduced blood outgrowth endothelial cells expressed 1.6±1.0 pmol/1×106 cells/24h FVIII. Morphological analysis demonstrated that FVIII-GFP was stored in Weibel-Palade bodies together with VWF and P-selectin. FVIII levels were only slightly increased following agonist-induced stimulation, whereas a 6- to 8-fold increase of VWF levels was observed. Subcellular fractionation revealed that 15–22% of FVIII antigen was present within the dense fraction containing Weibel-Palade bodies. Conclusions We conclude that blood outgrowth endothelial cells, by virtue of their ability to store a significant portion of synthesized FVIII-GFP in Weibel-Palade bodies, provide an attractive cellular on-demand delivery device for gene therapy of hemophilia A. PMID:19336741

Metformin improves circulating endothelial cells and endothelial progenitor cells in type 1 diabetes: MERIT study.

PubMed

Ahmed, Fahad W; Rider, Rachel; Glanville, Michael; Narayanan, Kilimangalam; Razvi, Salman; Weaver, Jolanta U

2016-08-26

Type 1 diabetes is associated with increased cardiovascular disease (CVD). Decreased endothelial progenitor cells (EPCs) number plays a pivotal role in reduced endothelial repair and development of CVD. We aimed to determine if cardioprotective effect of metformin is mediated by increasing circulating endothelial progenitor cells (cEPCs), pro-angiogenic cells (PACs) and decreasing circulating endothelial cells (cECs) count whilst maintaining unchanged glycemic control. This study was an open label and parallel standard treatment study. Twenty-three type 1 diabetes patients without overt CVD were treated with metformin for 8 weeks (treatment group-TG). They were matched with nine type 1 diabetes patients on standard treatment (SG) and 23 age- and sex-matched healthy volunteers (HC). Insulin dose was adjusted to keep unchanged glycaemic control. cEPCs and cECs counts were determined by flow cytometry using surface markers CD45(dim)CD34(+)VEGFR-2(+) and CD45(dim)CD133(-)CD34(+)CD144(+) respectively. Peripheral blood mononuclear cells were cultured to assess changes in PACs number, function and colony forming units (CFU-Hill's colonies). At baseline TG had lower cEPCs, PACs, CFU-Hills' colonies and PACs adhesion versus HC (p < 0.001-all variables) and higher cECs versus HC (p = 0.03). Metformin improved cEPCs, PACs, CFU-Hill's colonies number, cECs and PACs adhesion (p < 0.05-all variables) to levels seen in HC whilst HbA1c (one-way ANOVA p = 0.78) and glucose variability (average glucose, blood glucose standard deviation, mean amplitude of glycaemic excursion, continuous overall net glycaemic action and area under curve) remained unchanged. No changes were seen in any variables in SG. There was an inverse correlation between CFU-Hill's colonies with cECs. Metformin has potential cardio-protective effect through improving cEPCs, CFU-Hill's colonies, cECs, PACs count and function independently of hypoglycaemic effect. This finding needs to be confirmed by long

Isolation and characterization of conditionally immortalized mouse glomerular endothelial cell lines.

PubMed

Rops, Angelique L; van der Vlag, Johan; Jacobs, Cor W; Dijkman, Henry B; Lensen, Joost F; Wijnhoven, Tessa J; van den Heuvel, Lambert P; van Kuppevelt, Toin H; Berden, Jo H

2004-12-01

The culture and establishment of glomerular cell lines has proven to be an important tool for the understanding of glomerular cell functions in glomerular physiology and pathology. Especially, the recent establishment of a conditionally immortalized visceral epithelial cell line has greatly boosted the research on podocyte biology. Glomeruli were isolated from H-2Kb-tsA58 transgenic mice that contain a gene encoding a temperature-sensitive variant of the SV40 large tumor antigen, facilitating proliferative growth at 33 degrees C and differentiation at 37 degrees C. Glomerular endothelial cells were isolated from glomerular outgrowth by magnetic beads loaded with CD31, CD105, GSL I-B4, and ULEX. Clonal cell lines were characterized by immunofluorescence staining with antibodies/lectins specific for markers of endothelial cells, podocytes, and mesangial cells. Putative glomerular endothelial cell lines were analyzed for (1) cytokine-induced expression of adhesion molecules; (2) tube formation on Matrigel coating; and (3) the presence of fenestrae. As judged by immunostaining for Wilms tumor-1, smooth muscle actin (SMA), podocalyxin, and von Willebrand factor (vWF), we obtained putative endothelial, podocyte and mesangial cell lines. The mouse glomerular endothelial cell clone #1 (mGEnC-1) was positive for vWF, podocalyxin, CD31, CD105, VE-cadherin, GSL I-B4, and ULEX, internalized acetylated-low-density lipoprotein (LDL), and showed increased expression of adhesion molecules after activation with proinflammatory cytokines. Furthermore, mGEnC-1 formed tubes and contained nondiaphragmed fenestrae. The mGEnC-1 represents a conditionally immortalized cell line with various characteristics of differentiated glomerular endothelial cells when cultured at 37 degrees C. Most important, mGEnC-1 contains nondiaphragmed fenestrae, which is a unique feature of glomerular endothelial cells.

Endothelial induced EMT in breast epithelial cells with stem cell properties.

PubMed

Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla; Fridriksdottir, Agla J R; Ringnér, Markus; Villadsen, Rene; Borg, Ake; Agnarsson, Bjarni A; Petersen, Ole William; Magnusson, Magnus K; Gudjonsson, Thorarinn

2011-01-01

Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M) derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad) to N-Cadherin (N-Cad) and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44(high)/CD24(low) ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer.

Endothelial Induced EMT in Breast Epithelial Cells with Stem Cell Properties

PubMed Central

Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla; Fridriksdottir, Agla J. R.; Ringnér, Markus; Villadsen, Rene; Borg, Ake; Agnarsson, Bjarni A.; Petersen, Ole William; Magnusson, Magnus K.; Gudjonsson, Thorarinn

2011-01-01

Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M) derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad) to N-Cadherin (N-Cad) and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44high/CD24low ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer. PMID:21915264

Tissue distribution of mesenchymal stem cell marker Stro-1.

PubMed

Lin, Guiting; Liu, Gang; Banie, Lia; Wang, Guifang; Ning, Hongxiu; Lue, Tom F; Lin, Ching-Shwun

2011-10-01

Stro-1 is the best-known mesenchymal stem cell marker. However, despite its bone marrow origin, its localization in bone marrow has never been demonstrated. By immunofluorescence staining, we show here that ∼ 0.74% of nucleated bone marrow cells expressed Stro-1. We also found that ∼ 8.7% of CD34-expressing cells expressed Stro-1, and more than 20% of Stro-1-expressing cells did not express CD34. In adipose tissue Stro-1 expression was identified in the endothelium of arterioles and capillaries. Stro-1 was also localized in the endothelium of some but not all adipose tissue veins. Endothelial expression of Stro-1 was also identified in blood vessels in penis and in leg muscles, but not in other tested tissues. In these other tissues, Stro-1 was scantly expressed near but not in blood vessels. These variable and endothelial expression patterns of Stro-1 point to a need to re-examine published data that relied on Stro-1 as a mesenchymal stem cell marker.

Translational evidence of prothrombotic and inflammatory endothelial damage in Cushing syndrome after remission.

PubMed

Aranda, Gloria; Fernandez-Ruiz, Rebeca; Palomo, Marta; Romo, Mónica; Mora, Mireia; Halperin, Irene; Casals, Gregori; Enseñat, Joaquim; Vidal, Oscar; Diaz-Ricart, Maribel; Hanzu, Felicia A

2018-03-01

Sustained evidence from observational studies indicates that after remission of Cushing syndrome (CS) a cardiovascular risk phenotype persists. Here, we performed a translational study in active CS and CS in remission (RCS) to evaluate the subclinical cardiometabolic burden and to explore the direct pro-inflammatory and prothrombotic potential of their sera on the endothelium in an in vitro translational atherothrombotic cell model. Cross sectional study. The groups were (n = 9/group): I. RCS; II. Active CS (ACS) and III. Controls (CTR), all matched for age, body mass index, sex, without other hormonal deficits. We evaluated in vivo: cardiometabolic profile; endothelial markers (sVCAM-1, NO); endothelial dysfunction (FMD); intima-media thickness and body composition (DEXA). In vitro endothelial cells (EC) were exposed to sera taken from the different subjects to evaluate inflammatory EC response (tisVCAM) and thrombogenicity of the generated extracellular matrix (ECM): von Willebrand factor (VWF) and platelet reactivity. Three of the 9 RCS subjects were on glucocorticoid replacement therapy (GC-RT). Patients on GC-RT had a shorter period of time in stable remission. In vivo analysis ACS showed typically metabolic features, while cardiometabolic markers reached statistical significance for RCS only for Hs-CRP (P < .01). In vitro:EC exposed to ACS and RCS sera displayed increased tisVCAM-1 (P < .01 for ACS and P < .05 for RCS vs CTR), VWF (P < .01 for ACS and P < .05 for RCS vs CTR) and platelet adhesion on ECM (P < .01 for ACC and P < .05 for RCS vs CTR). No statistically significant differences were observed between GC-RT RSC and RCS without GC-RT. The sera of premenopausal women with CS in remission, without atherothrombotic disease, contain circulatory endothelial deleterious factors with a direct thrombogenic and pro-inflammatory endothelial effect that could increase cardiovascular risk. © 2017 John Wiley & Sons Ltd.

Dietary patterns are associated with biochemical markers of inflammation and endothelial activation in the Multi-Ethnic Study of Atherosclerosis (MESA)2

PubMed Central

Nettleton, Jennifer A; Steffen, Lyn M; Mayer-Davis, Elizabeth J; Jenny, Nancy S; Jiang, Rui; Herrington, David M; Jacobs, David R

2010-01-01

Background Dietary patterns may influence cardiovascular disease risk through effects on inflammation and endothelial activation. Objective We examined relations between dietary patterns and markers of inflammation and endothelial activation. Design At baseline, diet (food-frequency questionnaire) and concentrations of C-reactive protein (CRP), interleukin 6 (IL-6), homocysteine, soluble intercellular adhesion molecule-1 (sICAM-1), and soluble E selectin were assessed in 5089 nondiabetic participants in the Multi-Ethnic Study of Atherosclerosis. Results Four dietary patterns were derived by using factor analysis. The fats and processed meats pattern (fats, oils, processed meats, fried potatoes, salty snacks, and desserts) was positively associated with CRP (P for trend < 0.001), IL-6 (P for trend < 0.001), and homocysteine (P for trend = 0.002). The beans, tomatoes, and refined grains pattern (beans, tomatoes, refined grains, and high-fat dairy products) was positively related to sICAM-1 (P for trend = 0.007). In contrast, the whole grains and fruit pattern (whole grains, fruit, nuts, and green leafy vegetables) was inversely associated with CRP, IL-6, homocysteine (P for trend ≤ 0.001), and sICAM-1 (P for trend = 0.034), and the vegetables and fish pattern (fish and dark-yellow, cruciferous, and other vegetables) was inversely related to IL-6 (P for trend = 0.009). CRP, IL-6, and homocysteine relations across the fats and processed meats and whole grains and fruit patterns were independent of demographics and lifestyle factors and were not modified by race-ethnicity. CRP and homocysteine relations were independent of waist circumference. Conclusions These results corroborate previous findings that empirically derived dietary patterns are associated with inflammation and show that these relations in an ethnically diverse population with unique dietary habits are similar to findings in more homogeneous populations. PMID:16762949

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

Inflammation, coagulation, endothelial dysfunction and oxidative stress in prediabetes--Biomarkers as a possible tool for early disease detection for rural screening.

PubMed

Maschirow, L; Khalaf, K; Al-Aubaidy, H A; Jelinek, H F

2015-06-01

This study aims to increase understanding of the connection between oxidative stress and inflammation in diabetes disease progression to provide a basis for investigating improved diagnostic possibilities, treatment and prevention of prediabetes. Differences in the level of biochemical markers of oxidative stress (erythrocyte GSH/GSSG and urinary 8-isoprostane), inflammation (CRP, IL-6), endothelial dysfunction (plasma homocysteine, urinary 8-hydroxy-2-deoxy-guanosine) and coagulation/fibrinolysis (C5a, D-Dimer) were determined in prediabetes and control subjects. While no difference was found in the 8-isoprostane levels between the two groups, the erythrocyte GSH/GSSG ratio was significantly reduced in the prediabetes group compared to control, indicating increased oxidative stress in the prediabetic state. Both urinary 8-OHdG and surprisingly also plasma homocysteine were significantly elevated in the prediabetes group, indicating endothelial dysfunction. The inflammation markers were slightly elevated in the prediabetic subjects and the same trend was found for the coagulation/fibrinolysis markers C5a and D-Dimer. These results were however not significant. The small elevation of blood glucose levels in the prediabetic state may have a detectable influence on endothelial function as indicated by changes to 8-OHdG, indicating an increased DNA-damage and homocysteine release from endothelial cells. Increased oxidative stress as indicated by the reduced GSH/GSSG ratio is likely to be the link between the moderate hyperglycaemia in prediabetes and pathological changes in endothelial function, which in the long-term may promote atherogenesis and result in the development of cardiovascular disease. Early detection of prediabetes is essential to avoid diabetes development and the associated complications like cardiovascular disease. The GSH/GSSG ratio and biomarkers like urinary 8-OHdG and plasma homocysteine offer a possible tool for the assessment of prediabetes in

Femtosecond laser cutting of endothelial grafts: comparison of endothelial and epithelial applanation.

PubMed

Bernard, Aurélien; He, Zhiguo; Gauthier, Anne Sophie; Trone, Marie Caroline; Baubeau, Emmanuel; Forest, Fabien; Dumollard, Jean Marc; Peocʼh, Michel; Thuret, Gilles; Gain, Philippe

2015-02-01

Stromal surface quality of endothelial lamellae cut for endothelial keratoplasty with a femtosecond laser (FSL) with epithelial applanation remains disappointing. Applanation of the endothelial side of the cornea, mounted inverted on an artificial chamber, has therefore been proposed to improve cut quality. We compared lamellar quality after FSL cutting using epithelial versus endothelial applanation. Lamellae were cut with an FSL from organ-cultured corneas. After randomization, 7 were cut with epithelial applanation and 7 with endothelial applanation. Lamellae of 50-, 75-, and 100-μm thickness were targeted. Thickness was measured by optical coherence tomography before and immediately after cutting. Viable endothelial cell density was quantified immediately after cutting using triple labeling with Hoechst/ethidium/calcein-AM coupled with image analysis with ImageJ. The stromal surface was evaluated by 9 masked observers using semiquantitative scoring of scanning electronic microscopy images. Histology of 2 samples was also analyzed before lamellar detachment. Precision (difference in target/actual thickness) and thickness regularity [coefficient of variation (CV) of 10 measurements] were significantly better with endothelial applanation (precision: 18 μm; range, 10-30; CV: 11%; range, 8-12) than with epithelial applanation (precision: 84 μm; range, 54-107; P = 0.002; CV: 24%; range, 13-47; P = 0.001). Endothelial applanation provided thinner lamellae. However, viable endothelial cell density was significantly lower after endothelial applanation (1183 cells/mm2; range, 787-1725 versus 1688 cells/mm2; range, 1288-2025; P = 0.018). FSL cutting of endothelial lamellae using endothelial applanation provides thinner more regular grafts with more predictable thickness than with conventional epithelial applanation but strongly reduces the pool of viable endothelial cells.

Endothelial and circulating progenitor cells in hematological diseases and allogeneic hematopoietic stem cell transplantation.

PubMed

Ruggeri, Annalisa; Paviglianiti, Annalisa; Volt, Fernanda; Kenzey, Chantal; Rafii, Hanadi; Rocha, Vanderson; Gluckman, Eliane

2017-10-12

Circulating endothelial cells (CECs), originated form endothelial progenitors (EPCs) are mature cells which are not associated with vessel walls, and that are detached from the endothelium. Normally, they are present in insignificant amounts in the peripheral blood of healthy individuals. On the other hand, elevated CECs and EPCs levels have been reported in the peripheral blood of patients with different types of cancers and some other diseases. Consequently, CECs and EPCs represent a potential biomarker in several clinical conditions involving endothelial turnover and remodeling, such as hematological diseases. These cells may be involved in disease progression and the neoplastic angiogenesis process. Moreover, CESs and EPCs are probably involved in endothelial damage that is a marker of several complications following allogeneic hematopoietic stem cell transplantation. This review aims to provide an overview on the characterization of CECs and EPCs, describe isolation methods and to identify the potential role of these cells in hematological diseases and hematopoietic stem cell transplantation. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

CMTM3 (CKLF-Like Marvel Transmembrane Domain 3) Mediates Angiogenesis by Regulating Cell Surface Availability of VE-Cadherin in Endothelial Adherens Junctions.

PubMed

Chrifi, Ihsan; Louzao-Martinez, Laura; Brandt, Maarten; van Dijk, Christian G M; Burgisser, Petra; Zhu, Changbin; Kros, Johan M; Duncker, Dirk J; Cheng, Caroline

2017-06-01

Decrease in VE-cadherin adherens junctions reduces vascular stability, whereas disruption of adherens junctions is a requirement for neovessel sprouting during angiogenesis. Endocytosis plays a key role in regulating junctional strength by altering bioavailability of cell surface proteins, including VE-cadherin. Identification of new mediators of endothelial endocytosis could enhance our understanding of angiogenesis. Here, we assessed the function of CMTM3 (CKLF-like MARVEL transmembrane domain 3), which we have previously identified as highly expressed in Flk1 + endothelial progenitor cells during embryonic development. Using a 3-dimensional coculture of human umbilical vein endothelial cells-GFP (green fluorescent protein) and pericytes-RFP (red fluorescent protein), we demonstrated that siRNA-mediated CMTM3 silencing in human umbilical vein endothelial cells impairs angiogenesis. In vivo CMTM3 inhibition by morpholino injection in developing zebrafish larvae confirmed that CMTM3 expression is required for vascular sprouting. CMTM3 knockdown in human umbilical vein endothelial cells does not affect proliferation or migration. Intracellular staining demonstrated that CMTM3 colocalizes with early endosome markers EEA1 (early endosome marker 1) and Clathrin + vesicles and with cytosolic VE-cadherin in human umbilical vein endothelial cells. Adenovirus-mediated CMTM3 overexpression enhances endothelial endocytosis, shown by an increase in Clathrin + , EEA1 + , Rab11 + , Rab5 + , and Rab7 + vesicles. CMTM3 overexpression enhances, whereas CMTM3 knockdown decreases internalization of cell surface VE-cadherin in vitro. CMTM3 promotes loss of endothelial barrier function in thrombin-induced responses, shown by transendothelial electric resistance measurements in vitro. In this study, we have identified a new regulatory function for CMTM3 in angiogenesis. CMTM3 is involved in VE-cadherin turnover and is a regulator of the cell surface pool of VE-cadherin. Therefore, CMTM

Aspirin in type 2 diabetes, a randomised controlled study: effect of different doses on inflammation, oxidative stress, insulin resistance and endothelial function.

PubMed

Raghavan, R P; Laight, D W; Cummings, M H

2014-02-01

The effect of aspirin upon platelet function is well documented although experimental studies suggest that aspirin may also affect oxidative stress, vascular inflammation, endothelial dysfunction and dysglycaemia. The optimal dose of aspirin for cardiovascular protection in type 2 diabetes is still debated. We examined the effects of different doses of aspirin upon these novel markers of cardiovascular risk and any association between aspirin-mediated changes in these markers. Subjects with type 2 diabetes attended for baseline evaluation including BMI, glycaemic and lipid markers, endothelial function (photoplethysmography), insulin resistance (HOMA), inflammation (sVCAM-1 and Hs-CRP) and markers of oxidative stress [total anti-oxidant status (TAOS and FRAP), whole blood total glutathione (GSH) assays]. Subjects then received in random, sequential, blinded fashion aspirin 75 mg day(-1) , aspirin 300 mg day(-1) , aspirin 3.6 g day(-1) or placebo for 2 weeks with a 2-week washout. The above investigations were repeated after each intervention. Aspirin-related changes compared with placebo were analysed using repeated measures ANOVA. Subjects = 17 (M - 12; F - 5), mean age - 57.4 ± 9.1 years (mean ± 1 SD), HbA1c - 63 ± 13 mmol mol(-1) (7.9 ± 1.2%), total cholesterol 4.57 ± 1.01 mmol l(-1) . At baseline TAOS value was 59.3 ± 9.7 μM AEAC (Ascorbate Equivalent Anti-oxidant Concentration), glutathione 302.2 ± 183.3 mmol l(-1) and FRAP 0.86 ± 0.23 mM FeII. None of the aspirin doses had a significant impact upon BMI, blood pressure, lipid parameters, insulin sensitivity (HOMA), FRAP, TAOS, GSH, endothelial function, glycaemic control (fructosamine) or inflammation (sVCAM-1 and HsCRP). Aspirin exhibited no significant dose-dependent effect on markers of vascular inflammation, oxidative stress, insulin resistance and endothelial function (photoplethysmography) when used in type 2 diabetes over a 2-week period. ( NCT00898950, EUDRACT:2004-001418-14). © 2013 John

Study of anti-endothelial cell antibodies in SLE patients with acute ischemic stroke.

PubMed

Cojocaru, Inimioara Mihaela; Cojocaru, M; Butnaru, Ludmila; Miu, Gabriela; Tănăsescu, R

2010-01-01

Endothelial dysfunction is the predominant manifestation of SLE. Anti-endothelial cell antibodies (AECA) are a heterogeneous group of autoantibodies directed against different antigens in endothelial cells. The objective of this study was to assess the possible correlation between the presence of AECA and ischemic stroke manifestations in SLE. The AECA titers in serum from 34 patients with SLE and acute ischemic stroke (8 men and 26 women, mean age 38.37 +/- 3.25 years) and in 32 controls (11 men and 21 women, mean age 37.52 +/- 3.86 years) were tested. The method used was ELISA. The data were expressed as mean +/- SD from indicated number of patients. Comparison between patients and controls was expressed as relative risk with its 95% confidence interval (RR[95% CI]), where lower limit > 1.0 was considered significant. All p values were determined by Fisher's exact test. A value of p < 0.05 was considered statistically significant. AECA were positive in 31 out of 34 patients, mean value 19.2 +/- 16.3 U/mL and in 8 out of 32 controls, mean value 5.5 +/- 2.6 U/mL (RR 7.154 [95% CI 2.801 to 18.274]), p < 0.0001. Patients with SLE and acute ischemic stroke tended to have higher mean values of AECA. AECA play a pivotal role in the pathogenesis of neurologic complications of SLE. AECA titers in SLE patients with acute ischemic stroke support a role for AECA as potential diagnostic marker possibly associated to cerebral manifestations of SLE patients. Further study is needed in order to clarify if AECA presence is related to systemic diseases severity and to situate their importance among other markers of endothelial dysfunction.

Prospective Surface Marker-Based Isolation and Expansion of Fetal Endothelial Colony-Forming Cells From Human Term Placenta

PubMed Central

Patel, Jatin; Seppanen, Elke; Chong, Mark S.K.; Yeo, Julie S.L.; Teo, Erin Y.L.; Chan, Jerry K.Y.; Fisk, Nicholas M.

2013-01-01

The term placenta is a highly vascularized tissue and is usually discarded upon birth. Our objective was to isolate clinically relevant quantities of fetal endothelial colony-forming cells (ECFCs) from human term placenta and to compare them to the well-established donor-matched umbilical cord blood (UCB)-derived ECFCs. A sorting strategy was devised to enrich for CD45−CD34+CD31Lo cells prior to primary plating to obtain pure placental ECFCs (PL-ECFCs) upon culture. UCB-ECFCs were derived using a well-described assay. PL-ECFCs were fetal in origin and expressed the same cell surface markers as UCB-ECFCs. Most importantly, a single term placenta could yield as many ECFCs as 27 UCB donors. PL-ECFCs and UCB-ECFCs had similar in vitro and in vivo vessel forming capacities and restored mouse hind limb ischemia in similar proportions. Gene expression profiles were only minimally divergent between PL-ECFCs and UCB-ECFCs, probably reflecting a vascular source versus a circulating source. Finally, PL-ECFCs and UCB-ECFCs displayed similar hierarchies between high and low proliferative colonies. We report a robust strategy to isolate ECFCs from human term placentas based on their cell surface expression. This yielded much larger quantities of ECFCs than UCB, but the cells were comparable in immunophenotype, gene expression, and in vivo functional ability. We conclude that PL-ECFCs have significant bio-banking and clinical translatability potential. PMID:24106336

Revisited microanatomy of the corneal endothelial periphery: new evidence for continuous centripetal migration of endothelial cells in humans.

PubMed

He, Zhiguo; Campolmi, Nelly; Gain, Philippe; Ha Thi, Binh Minh; Dumollard, Jean-Marc; Duband, Sébastien; Peoc'h, Michel; Piselli, Simone; Garraud, Olivier; Thuret, Gilles

2012-11-01

The control of corneal transparency depends on the integrity of its endothelial monolayer, which is considered nonregenerative in adult humans. In pathological situations, endothelial cell (EC) loss, not offset by mitosis, can lead to irreversible corneal edema and blindness. However, the hypothesis of a slow, clinically insufficient regeneration starting from the corneal periphery remains debatable. The authors have re-evaluated the microanatomy of the endothelium in order to identify structures likely to support this homeostasis model. Whole endothelia of 88 human corneas (not stored, and stored in organ culture) with mean donor age of 80 ± 12 years were analyzed using an original flat-mounting technique. In 61% of corneas, cells located at the extreme periphery (last 200 μm of the endothelium) were organized in small clusters with two to three cell layers around Hassall-Henle bodies. In 68% of corneas, peripheral ECs formed centripetal rows 830 ± 295 μm long, with Descemet membrane furrows visible by scanning electron microscopy. EC density was significantly higher in zones with cell rows. When immunostained, ECs in the extreme periphery exhibited lesser differentiation (ZO-1, Actin, Na/K ATPase, CoxIV) than ECs in the center of the cornea but preferentially expressed stem cell markers (Nestin, Telomerase, and occasionally breast cancer resistance protein) and, in rare cases, the proliferation marker Ki67. Stored corneas had fewer cell clusters but more Ki67-positive ECs. We identified a novel anatomic organization in the periphery of the human corneal endothelium, suggesting a continuous slow centripetal migration, throughout life, of ECs from specific niches. Copyright © 2012 AlphaMed Press.

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.

Microfluidically supported biochip design for culture of endothelial cell layers with improved perfusion conditions.

PubMed

Raasch, Martin; Rennert, Knut; Jahn, Tobias; Peters, Sven; Henkel, Thomas; Huber, Otmar; Schulz, Ingo; Becker, Holger; Lorkowski, Stefan; Funke, Harald; Mosig, Alexander

2015-03-02

Hemodynamic forces generated by the blood flow are of central importance for the function of endothelial cells (ECs), which form a biologically active cellular monolayer in blood vessels and serve as a selective barrier for macromolecular permeability. Mechanical stimulation of the endothelial monolayer induces morphological remodeling in its cytoskeleton. For in vitro studies on EC biology culture devices are desirable that simulate conditions of flow in blood vessels and allow flow-based adhesion/permeability assays under optimal perfusion conditions. With this aim we designed a biochip comprising a perfusable membrane that serves as cell culture platform multi-organ-tissue-flow (MOTiF biochip). This biochip allows an effective supply with nutrition medium, discharge of catabolic cell metabolites and defined application of shear stress to ECs under laminar flow conditions. To characterize EC layers cultured in the MOTiF biochip we investigated cell viability, expression of EC marker proteins and cell adhesion molecules of ECs dynamically cultured under low and high shear stress, and compared them with an endothelial culture in established two-dimensionally perfused flow chambers and under static conditions. We show that ECs cultured in the MOTiF biochip form a tight EC monolayer with increased cellular density, enhanced cell layer thickness, presumably as the result of a rapid and effective adaption to shear stress by remodeling of the cytoskeleton. Moreover, endothelial layers in the MOTiF biochip express higher amounts of EC marker proteins von-Willebrand-factor and PECAM-1. EC layers were highly responsive to stimulation with TNFα as detected at the level of ICAM-1, VCAM-1 and E-selectin expression and modulation of endothelial permeability in response to TNFα/IFNγ treatment under flow conditions. Compared to static and two-dimensionally perfused cell culture condition we consider MOTiF biochips as a valuable tool for studying EC biology in vitro under

Endurance capacity is not correlated with endothelial function in male university students.

PubMed

Wang, Yan; Zeng, Xian-bo; Yao, Feng-juan; Wu, Fang; Su, Chen; Fan, Zhen-guo; Zhu, Zhu; Tao, Jun; Huang, Yi-jun

2014-01-01

Endurance capacity, assessed by 1000-meter (1000 m) run of male university students, is an indicator of cardiovascular fitness in Chinese students physical fitness surveillance. Although cardiovascular fitness is related to endothelial function closely in patients with cardiovascular diseases, it remains unclear whether endurance capacity correlates with endothelial function, especially with circulating endothelial microparticles (EMPs), a new sensitive marker of endothelial dysfunction in young students. The present study aimed to investigate the relationship between endurance capacity and endothelial function in male university students. Forty-seven healthy male university students (mean age, 20.1 ± 0.6 years; mean height, 172.4 ± 6.3 cm; and mean weight, 60.0 ± 8.2 kg) were recruited in this study. The measurement procedure of 1000 m run time was followed to Chinese national students Constitutional Health Criterion. Endothelium function was assessed by flow-mediated vasodilation (FMD) in the brachial artery measured by ultrasonic imaging, and the level of circulating EMPs was measured by flow cytometry. Cardiovascular fitness indicator--maximal oxygen uptake (VO2max)--was also measured on a cycle ergometer using a portable gas analyzer. 1000 m run time was correlated with VO2max (r  =  -0.399, p<0.05). However, there were no correlations between VO2max and FMD or levels of circulating CD31+/CD42- microparticles. Similarly, no correlations were found between 1000 m run time and FMD, and levels of circulating CD31+/CD42- microparticles in these male university students (p>0.05). The correlations between endurance capacity or cardiovascular fitness and endothelial function were not found in healthy Chinese male university students. These results suggest that endurance capacity may not reflect endothelial function in healthy young adults with well preserved FMD and low level of circulating CD31+/CD42-EMPs.

Endurance Capacity Is Not Correlated with Endothelial Function in Male University Students

PubMed Central

Wu, Fang; Su, Chen; Fan, Zhen-guo; Zhu, Zhu; Tao, Jun; Huang, Yi-jun

2014-01-01

Background Endurance capacity, assessed by 1000-meter (1000 m) run of male university students, is an indicator of cardiovascular fitness in Chinese students physical fitness surveillance. Although cardiovascular fitness is related to endothelial function closely in patients with cardiovascular diseases, it remains unclear whether endurance capacity correlates with endothelial function, especially with circulating endothelial microparticles (EMPs), a new sensitive marker of endothelial dysfunction in young students. The present study aimed to investigate the relationship between endurance capacity and endothelial function in male university students. Methods Forty-seven healthy male university students (mean age, 20.1±0.6 years; mean height, 172.4±6.3 cm; and mean weight, 60.0±8.2 kg) were recruited in this study. The measurement procedure of 1000 m run time was followed to Chinese national students Constitutional Health Criterion. Endothelium function was assessed by flow-mediated vasodilation (FMD) in the brachial artery measured by ultrasonic imaging, and the level of circulating EMPs was measured by flow cytometry. Cardiovascular fitness indicator - maximal oxygen uptake (VO2 max) - was also measured on a cycle ergometer using a portable gas analyzer. Results 1000 m run time was correlated with VO2max (r = −0.399, p<0.05). However, there were no correlations between VO2max and FMD or levels of circulating CD31+/CD42- microparticles. Similarly, no correlations were found between 1000 m run time and FMD, and levels of circulating CD31+/CD42- microparticles in these male university students (p>0.05). Conclusion The correlations between endurance capacity or cardiovascular fitness and endothelial function were not found in healthy Chinese male university students. These results suggest that endurance capacity may not reflect endothelial function in healthy young adults with well preserved FMD and low level of circulating CD31+/CD42-EMPs. PMID:25101975

The endothelial glycocalyx

PubMed Central

Yang, Yimu; Schmidt, Eric P.

2013-01-01

Once thought to be a structure of small size and uncertain significance, the endothelial glycocalyx is now known to be an important regulator of endothelial function. Studies of the systemic vasculature have demonstrated that the glycocalyx forms a substantial in vivo endothelial surface layer (ESL) critical to inflammation, barrier function and mechanotransduction. The pulmonary ESL is significantly thicker than the systemic ESL, suggesting unique physiologic function. We have recently demonstrated that the pulmonary ESL regulates exposure of endothelial surface adhesion molecules, thereby serving as a barrier to neutrophil adhesion and extravasation. While the pulmonary ESL is not a critical structural component of the endothelial barrier to fluid and protein, it serves a major role in the mechanotransduction of vascular pressure, with impact on the active regulation of endothelial permeability. It is likely that the ESL serves numerous additional functions in vascular physiology, representing a fertile area for future investigation. PMID:24073386

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

Dynamic Changes of Smooth Muscle and Endothelial Markers in the Early Healing Process of Dacron Vascular Grafts in the Dog, Using RT-PCR.

PubMed

Ishida; Wu; Shi; Fujita; Sauvage; Hammond; Wijelath

2000-03-01

Previous studies of neointima formation on Dacron vascular grafts mainly focused on the late stages using immunohistochemistry staining for von Willebrand factor (vWF) and smooth muscle (SM) alpha-actin. However, it is impossible to use immunohistochemistry to study the early events of neointima formation, because graft samples lack sufficient cellular material. Therefore, we used reverse transcriptase-polymerase chain reaction (RT-PCR) to demonstrate dynamic changes of SM and endothelial markers during the early stages of neointima formation. Preclotted Dacron grafts were implanted in the descending thoracic aorta of 14 mongrel dogs. Specimens were retrieved at 1-4 weeks. Total RNAs were extracted from mid-portion of graft flow surfaces, and RT-PCR for vWF, SM myosin heavy chain (MHC), and SM alpha-actin were performed and expressed as a ratio to the ribosome s17 signal. SM MHC and vWF mRNA expression was low at 1-2 weeks but elevated at 3-4 weeks (P < 0.05). However, SM alpha-actin mRNA levels were expressed consistently throughout the study period. At 3-4 weeks, vWF mRNA expression was inversely correlated to thrombus formation on the graft flow surface. Increased expressions of SM MHC and vWF mRNA corresponded to the formation of neointima and an endothelial layer at the later stages. However, SM alpha-actin mRNA expression did not vary during the healing process. The application of RT-PCR should permit further studies of gene regulation in the early vascular graft healing process in vivo. This model can also be used to study the molecular events that are involved in SM cell differentiation.

Morphology and vasoactive hormone profiles from endothelial cells derived from stem cells of different sources.

PubMed

Reed, Daniel M; Foldes, Gabor; Kirkby, Nicholas S; Ahmetaj-Shala, Blerina; Mataragka, Stefania; Mohamed, Nura A; Francis, Catherine; Gara, Edit; Harding, Sian E; Mitchell, Jane A

2014-12-12

Endothelial cells form a highly specialised lining of all blood vessels where they provide an anti-thrombotic surface on the luminal side and protect the underlying vascular smooth muscle on the abluminal side. Specialised functions of endothelial cells include their unique ability to release vasoactive hormones and to morphologically adapt to complex shear stress. Stem cell derived-endothelial cells have a growing number of applications and will be critical in any organ regeneration programme. Generally endothelial cells are identified in stem cell studies by well-recognised markers such as CD31. However, the ability of stem cell-derived endothelial cells to release vasoactive hormones and align with shear stress has not been studied extensively. With this in mind, we have compared directly the ability of endothelial cells derived from a range of stem cell sources, including embryonic stem cells (hESC-EC) and adult progenitors in blood (blood out growth endothelial cells, BOEC) with those cultured from mature vessels, to release the vasoconstrictor peptide endothelin (ET)-1, the cardioprotective hormone prostacyclin, and to respond morphologically to conditions of complex shear stress. All endothelial cell types, except hESC-EC, released high and comparable levels of ET-1 and prostacyclin. Under static culture conditions all endothelial cell types, except for hESC-EC, had the typical cobblestone morphology whilst hESC-EC had an elongated phenotype. When cells were grown under shear stress endothelial cells from vessels (human aorta) or BOEC elongated and aligned in the direction of shear. By contrast hESC-EC did not align in the direction of shear stress. These observations show key differences in endothelial cells derived from embryonic stem cells versus those from blood progenitor cells, and that BOEC are more similar than hESC-EC to endothelial cells from vessels. This may be advantageous in some settings particularly where an in vitro test bed is required

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.

[The role of endothelial cells and endothelial precursor cells in angiogenesis].

PubMed

Poreba, Małgorzata; Usnarska-Zubkiewicz, Lidia; Kuliczkowski, Kazimierz

2006-01-01

Endothelium plays a key role in maintenance of vascular homeostasis in human organism. According to new data endothelial cells and hematopoietic cells have a common precursor in prenatal life--a hemangioblast, which explains the fact of sharing the same determinants on the surface of both type of cells. Circulating endothelial precursors were identified in adults and this suggests that hemangioblasts may be present not only during embriogenesis. In some clinical situations the increased numbers of endothelial cells and endothelial precursors were noted, and especially in patients with neoplastic diseases, which is probably the result of increased angiogenesis. Endothelial precursors are thought to be the promice for therapeutic purposes in future--to increase local angiogenesis.

Polymeric stent materials dysregulate macrophage and endothelial cell functions: implications for coronary artery stent

PubMed Central

Wang, Xintong; Zachman, Angela L.; Chun, Young Wook; Shen, Fang-Wen; Hwang, Yu-Shik; Sung, Hak-Joon

2014-01-01

Background Biodegradable polymers have been applied as bulk or coating materials for coronary artery stents. The degradation of polymers, however, could induce endothelial dysfunction and aggravate neointimal formation. Here we use polymeric microparticles to simulate and demonstrate the effects of degraded stent materials on phagocytic activity, cell death and dysfunction of macrophages and endothelial cells. Methods Microparticles made of low molecular weight polyesters were incubated with human macrophages and coronary artery endothelial cells (ECs). Microparticle-induced phagocytosis, cytotoxicity, apoptosis, cytokine release and surface marker expression were determined by immunostaining or ELISA. Elastase expression was analyzed by ELISA and the elastase-mediated polymer degradation was assessed by mass spectrometry. Results We demonstrated poly(D,L-lactic acid) (PLLA) and polycaprolactone (PCL) microparticles induced cytotoxicity in macrophages and ECs, partially through cell apoptosis. The particle treatment alleviated EC phagocytosis, as opposed to macrophages, but enhanced the expression of vascular cell adhesion molecule-1 (VCAM) along with decreased nitric oxide production, indicating ECs were activated and lost their capacity to maintain homeostasis. The activation of both cell types induced release of elastase or elastase-like protease, which further accelerated polymer degradation. Conclusions This study revealed that low molecule weight PLLA and PCL microparticles increased cytotoxicity and dysregulated endothelial cell function, which in turn enhanced elastase release and polymer degradation. These indicate polymer or polymer-coated stents impose a risk of endothelial dysfunction after deployment which can potentially lead to delayed endothelialization, neointimal hyperplasia and late thrombosis. PMID:24820736

Effects of long- and short-term darbepoetin-α treatment on oxidative stress, inflammation and endothelial injury in ApoE knockout mice.

PubMed

Özdemir, Evrim Dursun; Hanikoglu, Aysegul; Cort, Aysegul; Ozben, Beste; Suleymanlar, Gultekin; Ozben, Tomris

2017-07-01

Atherosclerosis and atherosclerosis-related complications are the main cause of death in the world. Vascular injury in response to inflammation and enhanced oxidant stress promotes endothelial dysfunction and leads to atherosclerotic lesions. Low-dose treatment with darbepoetin-α may be a potential therapeutic tool for endothelial injury and atherosclerosis. In order to study the effect of darbepoetin-α on endothelial injury and atherosclerosis, we used ApoE-/- mice as the atherosclerotic mice model. We monitored atherosclerosis and plaque formation histochemically in ApoE knockout mice at early and late stages of atherosclerosis. Darbepoetin-α was injected intraperitoneally at a dose of 0.1 μg/kg to ApoE-/- mice. The results of 2 ApoE-/- mice groups injected with darbepoetin-α (early and late stages of atherosclerosis) were compared to the results of the corresponding saline injected ApoE-/- mice groups and the control (C57BL/6) mice. Lipid profile (total cholesterol, triglyceride), inflammation (CRP, IL-6, histamine), endothelial injury (ICAM-1, selectin) and oxidative stress markers (lipid peroxidation, protein oxidation) were significantly increased in 4 atherosclerotic groups compared to the control group. Short-term darbepoetin-α had no marked effects on indicators of inflammation and endothelial injury in the ApoE knockout mice groups compared to the ApoE knockout mice not treated with darbepoetin-α, however, darbepoetin-α significantly decreased 8-isoprostane and protein carbonyl content. Long term darbepoetin-α treatment reduced oxidative stress in ApoE-/- mice. This study contributes to understanding and elucidating the biochemical changes occurring during early and late stages of atherosclerosis development regarding lipid profile, inflammation, endothelial injury and oxidative stress markers.

Short communication: initiation of an abacavir-containing regimen in HIV-infected adults is associated with a smaller decrease in inflammation and endothelial activation markers compared to non-abacavir-containing regimens.

PubMed

Hileman, Corrilynn O; Wohl, David A; Tisch, Daniel J; Debanne, Sara M; McComsey, Grace A

2012-12-01

Abacavir has been associated with myocardial infarction in several studies. This may be related to inflammation and endothelial cell activation. We compared changes in inflammation and endothelial activation markers between antiretroviral-naive adults initiating zidovudine, lamivudine, abacavir, and nonnucleoside reverse transcriptase inhibitor (NNRTI) or this regimen without abacavir. Changes in soluble tumor necrosis factor receptors-I, -II (sTNFR-I, -II), high sensitivity C-reactive protein, and soluble vascular cell adhesion molecule-1 (sVCAM-1) from baseline (pre-ART) to a second time point about 24 weeks after initiating antiretroviral therapy (ART) were compared between groups using multivariable linear regression. A total of 37 met eligibility criteria; 12 received abacavir. The median (interquartile range) age was 37 years (27-45). Most were men (32/37), African-American (15/37), or white (15/37). The median nadir CD4(+) and baseline HIV-1 RNA were 230 cells/mm(3) (180-301) and 82,642 copies/ml (34,400-204,703). In all, 15/30 smoked, 7/37 had hypertension, 1/37 had diabetes, and 1/37 had hyperlipidemia. None had coronary or renal disease. Changes in CD4(+) and HIV-1 RNA level and timing of stored samples with regard to ART initiation were not different between groups. In univariable analysis, log transformed percent change in sTNFR-I (p=0.05) and -II (p=0.04) showed significant between-group differences and trended toward significance for sVCAM-1 (p=0.08). These markers decreased less in the abacavir group. After adjustment for confounders, significantly less decrease for sTNFR-II and sVCAM-1 was seen for those receiving the abacavir-containing regimen. When taken with an NNRTI, abacavir induced a smaller decrease in inflammation biomarkers in this cohort, suggesting a possible proinflammatory effect of this nucleoside analogue.

TRAIL death receptor 4 signaling via lysosome fusion and membrane raft clustering in coronary arterial endothelial cells: evidence from ASM knockout mice.

PubMed

Li, Xiang; Han, Wei-Qing; Boini, Krishna M; Xia, Min; Zhang, Yang; Li, Pin-Lan

2013-01-01

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and its receptor, death receptor 4 (DR4), have been implicated in the development of endothelial dysfunction and atherosclerosis. However, the signaling mechanism mediating DR4 activation leading to endothelial injury remains unclear. We recently demonstrated that ceramide production via hydrolysis of membrane sphingomyelin by acid sphingomyelinase (ASM) results in membrane raft (MR) clustering and the formation of important redox signaling platforms, which play a crucial role in amplifying redox signaling in endothelial cells leading to endothelial dysfunction. The present study aims to investigate whether TRAIL triggers MR clustering via lysosome fusion and ASM activation, thereby conducting transmembrane redox signaling and changing endothelial function. Using confocal microscopy, we found that TRAIL induced MR clustering and co-localized with DR4 in coronary arterial endothelial cells (CAECs) isolated from wild-type (Smpd1 (+/+)) mice. Furthermore, TRAIL triggered ASM translocation, ceramide production, and NADPH oxidase aggregation in MR clusters in Smpd1 ( +/+ ) CAECs, whereas these observations were not found in Smpd1 (-/-) CAECs. Moreover, ASM deficiency reduced TRAIL-induced O(2) (-[Symbol: see text]) production in CAECs and abolished TRAIL-induced impairment on endothelium-dependent vasodilation in small resistance arteries. By measuring fluorescence resonance energy transfer, we found that Lamp-1 (lysosome membrane marker protein) and ganglioside G(M1) (MR marker) were trafficking together in Smpd1 (+/+) CAECs, which was absent in Smpd1 (-/-) CAECs. Consistently, fluorescence imaging of living cells with specific lysosome probes demonstrated that TRAIL-induced lysosome fusion with membrane was also absent in Smpd1 (-/-) CAECs. Taken together, these results suggest that ASM is essential for TRAIL-induced lysosomal trafficking, membrane fusion and formation of MR redox signaling platforms

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

Skeletal Myogenic Progenitors Originating from Embryonic Dorsal Aorta Coexpress Endothelial and Myogenic Markers and Contribute to Postnatal Muscle Growth and Regeneration

PubMed Central

De Angelis, Luciana; Berghella, Libera; Coletta, Marcello; Lattanzi, Laura; Zanchi, Malvina; Gabriella, M.; Ponzetto, Carola; Cossu, Giulio

1999-01-01

Skeletal muscle in vertebrates is derived from somites, epithelial structures of the paraxial mesoderm, yet many unrelated reports describe the occasional appearance of myogenic cells from tissues of nonsomite origin, suggesting either transdifferentiation or the persistence of a multipotent progenitor. Here, we show that clonable skeletal myogenic cells are present in the embryonic dorsal aorta of mouse embryos. This finding is based on a detailed clonal analysis of different tissue anlagen at various developmental stages. In vitro, these myogenic cells show the same morphology as satellite cells derived from adult skeletal muscle, and express a number of myogenic and endothelial markers. Surprisingly, the latter are also expressed by adult satellite cells. Furthermore, it is possible to clone myogenic cells from limbs of mutant c-Met−/− embryos, which lack appendicular muscles, but have a normal vascular system. Upon transplantation, aorta-derived myogenic cells participate in postnatal muscle growth and regeneration, and fuse with resident satellite cells. The potential of the vascular system to generate skeletal muscle cells may explain observations of nonsomite skeletal myogenesis and raises the possibility that a subset of satellite cells may derive from the vascular system. PMID:10562287

MMP‐2 and MMP‐14 Silencing Inhibits VEGFR2 Cleavage and Induces the Differentiation of Porcine Adipose‐Derived Mesenchymal Stem Cells to Endothelial Cells

PubMed Central

Almalki, Sami G.; Llamas Valle, Yovani

2017-01-01

Abstract The molecular mechanisms that control the ability of adipose‐derived mesenchymal stem cells (AMSCs) to remodel three‐dimensional extracellular matrix barriers during differentiation are not clearly understood. Herein, we studied the expression of matrix metalloproteinases (MMPs) during the differentiation of AMSCs to endothelial cells (ECs) in vitro. MSCs were isolated from porcine abdominal adipose tissue, and characterized by immunopositivity to CD44, CD90, CD105, and immunonegativity to CD14 and CD45. Plasticity of AMSCs was confirmed by multilineage differentiation. The mRNA transcripts for MMPs and Tissue Inhibitor of Metalloproteinases (TIMPs), and protein expression of EC markers were analyzed. The enzyme activity and protein expression were analyzed by gelatin zymography, enzyme‐linked immunosorbent assay (ELISA), and Western blot. The differentiation of AMSCs to ECs was confirmed by mRNA and protein expressions of the endothelial markers. The mRNA transcripts for MMP‐2 and MMP‐14 were significantly increased during the differentiation of MSCs into ECs. Findings revealed an elevated MMP‐14 and MMP‐2 expression, and MMP2 enzyme activity. Silencing of MMP‐2 and MMP‐14 significantly increased the expression of EC markers, formation of capillary tubes, and acetylated‐low‐density lipoprotein uptake, and decreased the cleavage of vascular endothelial growth factor receptor type 2 (VEGFR2). Inhibition of VEGFR2 significantly decreased the expression of EC markers. These novel findings demonstrate that the upregulation of MMP2 and MMP14 has an inhibitory effect on the differentiation of AMSCs to ECs, and silencing these MMPs inhibit the cleavage of VEGFR2 and stimulate the differentiation of AMSCs to ECs. These findings provide a potential mechanism for the regulatory role of MMP‐2 and MMP‐14 in the re‐endothelialization of coronary arteries following intervention. Stem Cells Translational Medicine 2017;6:1385–1398 PMID

In Vitro Endothelialization of Biodegradable Vascular Grafts Via Endothelial Progenitor Cell Seeding and Maturation in a Tubular Perfusion System Bioreactor.

PubMed

Melchiorri, Anthony J; Bracaglia, Laura G; Kimerer, Lucas K; Hibino, Narutoshi; Fisher, John P

2016-07-01

A critical challenge to the success of biodegradable vascular grafts is the establishment of a healthy endothelium. To establish this monolayer of endothelial cells (ECs), a variety of techniques have been developed, including cell seeding. Vascular grafts may be seeded with relevant cell types and allowed to mature before implantation. Due to the low proliferative ability of adult ECs and issues with donor site morbidity, there has been increasing interest in using endothelial progenitor cells (EPCs) for vascular healing procedures. In this work, we combined the proliferative and differentiation capabilities of a commercial cell line of early EPCs with an established bioreactor system to support the maturation of cell-seeded vascular grafts. All components of the vascular graft and bioreactor setup are commercially available and allow for complete customization of the scaffold and culturing system. This bioreactor setup enables the control of flow through the graft, imparting fluid shear stress on EPCs and affecting cellular proliferation and differentiation. Grafts cultured with EPCs in the bioreactor system demonstrated greatly increased cell populations and neotissue formation compared with grafts seeded and cultured in a static system. Increased expression of markers for mature endothelial tissues were also observed in bioreactor-cultured EPC-seeded grafts. These findings suggest the distinct advantages of a customizable bioreactor setup for the proliferation and maturation of EPCs. Such a strategy may be beneficial for utilizing EPCs in vascular tissue engineering applications.

Homocysteine impaired endothelial function through compromised vascular endothelial growth factor/Akt/endothelial nitric oxide synthase signalling.

PubMed

Yan, Ting-Ting; Li, Qian; Zhang, Xuan-Hong; Wu, Wei-Kang; Sun, Juan; Li, Lin; Zhang, Quan; Tan, Hong-Mei

2010-11-01

1. Hyperhomocysteinaemia (HHcy) is associated with endothelial dysfunction and has been recognized as a risk factor of cardiovascular disease. The present study aimed to investigate the effect of homocysteine (Hcy) on endothelial function in vivo and in vitro, and the underlying signalling pathways. 2. The HHcy animal model was established by intragastric administration with l-methionine in rats. Plasma Hcy and nitric oxide (NO) concentration were measured by fluorescence immunoassay or nitrate reductase method, respectively. Vasorelaxation in response to acetylcholine and sodium nitroprusside were carried out on aortic rings. Human umbilical vein endothelial cells (HUVEC) were treated with indicated concentrations of Hcy in the in vitro experiments. Intracellular NO level and NO concentration in culture medium were assayed. The alterations of possible signalling proteins were detected by western blot analysis. 3. l-methionine administration induced a significant increase in plasma Hcy and decrease in plasma NO. Endothelium-dependent relaxation of aortic rings in response to acetylcholine was impaired in l-methionine-administrated rats. The in vitro study showed that Hcy reduced both intracellular and culture medium NO levels. Furthermore, Hcy decreased phosphorylation of endothelial nitric oxide synthase (eNOS) at serine-1177 and phosphorylation of Akt at serine-473. Hcy-induced dephosphorylation of eNOS at Ser-1177 was partially reversed by insulin (Akt activator) and GF109203X (PKC inhibitor). Furthermore, Hcy reduced vascular endothelial growth factor (VEGF) expression in a dose-dependent manner. 4. In conclusion, Hcy impaired endothelial function through compromised VEGF/Akt/endothelial nitric oxide synthase signalling. These findings will be beneficial for further understanding the role of Hcy in cardiovascular disease. © 2010 Blackwell Publishing Asia Pty Ltd.

Association Between Inflammatory Markers and Progression to Kidney Dysfunction: Examining Different Assessment Windows in Patients With Type 1 Diabetes.

PubMed

Baker, Nathaniel L; Hunt, Kelly J; Stevens, Danielle R; Jarai, Gabor; Rosen, Glenn D; Klein, Richard L; Virella, Gabriel; Lopes-Virella, Maria F

2018-01-01

To determine whether biomarkers of inflammation and endothelial dysfunction are associated with the development of kidney dysfunction and the time frame of their association. Biomarkers were measured at four time points during 28 years of treatment and follow-up in patients with type 1 diabetes in the Diabetes Control and Complications Trial/Epidemiology of Diabetes Interventions and Complications (DCCT/EDIC) cohort. In addition to traditional biomarkers of inflammation (C-reactive protein and fibrinogen), we measured interleukin-6 (IL-6) and soluble tumor necrosis factor receptors 1 and 2 (sTNFR-1/2), markers of endothelial dysfunction (soluble intracellular adhesion molecule-1, vascular cell adhesion molecule-1, and E-selectin [sE-selectin]), and fibrinolysis (total and active plasminogen activator inhibitor-1 [PAI-1]). Renal outcomes were defined as progression to incident chronic kidney disease (stage 3 or more severe) or macroalbuminuria (albumin excretion rate ≥300 mg/24 h). Prospective multivariate event-time analyses were used to determine the association of each biomarker with each subsequent event within prespecified intervals (3-year and 10-year windows). Multivariate event-time models indicated that several markers of inflammation (sTNFR-1/2), endothelial dysfunction (sE-selectin), and clotting/fibrinolysis (fibrinogen and PAI-1) are significantly associated with subsequent development of kidney dysfunction. Although some markers showed variations in the associations between the follow-up windows examined, the results indicate that biomarkers (sTNFR-1/2, sE-selectin, PAI-1, and fibrinogen) are associated with progression to chronic kidney disease in both the 3-year and the 10-year windows. Plasma markers of inflammation, endothelial dysfunction, and clotting/fibrinolysis are associated with progression to kidney dysfunction in type 1 diabetes during both short-term and long-term follow-up. © 2017 by the American Diabetes Association.

A small molecule-based strategy for endothelial differentiation and three-dimensional morphogenesis from human embryonic stem cells.

PubMed

Geng, Yijie; Feng, Bradley

2016-07-01

The emerging models of human embryonic stem cell (hESC) self-organizing organoids provide a valuable in vitro platform for studying self-organizing processes that presumably mimic in vivo human developmental events. Here we report that through a chemical screen, we identified two novel and structurally similar small molecules BIR1 and BIR2 which robustly induced the self-organization of a balloon-shaped three-dimensional structure when applied to two-dimensional adherent hESC cultures in the absence of growth factors. Gene expression analyses and functional assays demonstrated an endothelial identity of this balloon-like structure, while cell surface marker analyses revealed a VE-cadherin(+)CD31(+)CD34(+)KDR(+)CD43(-) putative endothelial progenitor population. Furthermore, molecular marker labeling and morphological examinations characterized several other distinct DiI-Ac-LDL(+) multi-cellular modules and a VEGFR3(+) sprouting structure in the balloon cultures that likely represented intermediate structures of balloon-formation.

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

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

The impact of decreases in air temperature and increases in ozone on markers of endothelial function in individuals having type-2 diabetes

EPA Science Inventory

Several studies have reported an association between air pollution and endothelial dysfunction, especially in individuals having diabetes. However, very few studies have examined the impact of air temperature on endothelial function. The objective of this analysis was to investig...

Moderate endurance exercise in patients with sickle cell anaemia: effects on oxidative stress and endothelial activation.

PubMed

Faes, Camille; Balayssac-Siransy, Edwige; Connes, Philippe; Hivert, Ludovic; Danho, Clotaire; Bogui, Pascal; Martin, Cyril; Pialoux, Vincent

2014-01-01

Very few studies have investigated the effects of exercise on the biological parameters involved in vaso-occlusive events in sickle cell anaemia (SCA). The aim of this study was to test how a mild-moderate endurance exercise modulates oxidative stress, nitric oxide bioavailability and endothelial activation in SCA patients and healthy individuals. Eleven patients with SCA and 15 healthy subjects completed a 20-min duration submaximal cycling exercise at ≈45 Watts. Plasma markers of oxidative stress, antioxidant activity, endothelial activation and nitric oxide bioavailability were investigated before and after the exercise. Nitric oxide levels, anti-oxidant capacity, soluble (s)E-selectin and sP-selectin did not change in response to this exercise. Except for the malondialdehyde levels, which increased in the two groups, the other markers of oxidative stress remained unchanged in both groups in response to exercise. Soluble vascular cell adhesion molecule 1 levels were increased at the end of exercise in both groups. sL-selectin decreased and soluble intercellular adhesion molecule 1 increased with exercise in SCA patients only. The present data suggest that patients with SCA may undertake mild-moderate physical activities without any acute clinical complications, but care should be taken because oxidative stress and endothelial activation significantly increased in some patients. © 2013 John Wiley & Sons Ltd.

Orbital angiogenesis and lymphangiogenesis in thyroid eye disease: an analysis of vascular growth factors with clinical correlation

PubMed Central

Wong, Lindsay L.; Lee, Nahyoung Grace; Amarnani, Dhanesh; Choi, Catherine J.; Bielenberg, Diane R.; Freitag, Suzanne K.; D’Amore, Patricia A.; Kim, Leo A.

2017-01-01

Purpose The human orbit is an environment that is vulnerable to inflammation and edema in the setting of autoimmune thyroid disease. Our study investigated the tenet that orbital adipose tissue lacks lymphatic vessels and analyzed the clinicopathologic differences between patients with acute and chronic thyroid eye disease (TED). The underlying molecular mediators of blood and lymphatic vessel formation within the orbital fat were also evaluated. Design Retrospective cohort study Participants The study included fat specimens from 26 orbits of 15 patients with TED undergoing orbital decompression. Orbital fat specimens from patients without TED as well as cadaveric orbital fat served as controls. Methods Tissue specimens were processed as formalin-fixed paraffin-embedded sections (FFPE) or frozen cryosections for immunohistochemistry. Total RNA was extracted and analyzed via quantitative (real-time) reverse transcription polymerase chain reaction (qRT-PCR). Clinicopathological correlation was made by determining the Clinical Activity Score (CAS) of each patient with TED. Main Outcome Measures Samples were examined for vascular and lymphatic markers including podoplanin, LYVE-1, and CD31 by immunohistochemistry, as well as for mRNA levels of VEGF, VEGF receptors, SEMA-3F, NRP-1, NRP-2, podoplanin and LYVE-1 by qRT-PCR. Results Clinicopathological correlation revealed increased staining of CD31-positive blood vessels in patients with acute TED with CAS > 4, as well as rare staining of podoplanin-positive lymphatic vessels within acutely inflamed orbital fat tissue. Additionally, qRT-PCR analysis demonstrated increased expression of vascular endothelial growth factor receptor 2 (VEGFR-2) as well as VEGF signaling molecules: VEGF-A, VEGF-C, and VEGF-D. Conclusions In acute TED, compared to chronic TED and control orbital fat, there is increased blood vessel density suggesting neovascularization and rare lymphatic vessels suggestive of limited lymphangiogenesis. This pro

Therapeutically targeting mitochondrial redox signalling alleviates endothelial dysfunction in preeclampsia.

PubMed

McCarthy, Cathal; Kenny, Louise C

2016-09-08

Aberrant placentation generating placental oxidative stress is proposed to play a critical role in the pathophysiology of preeclampsia. Unfortunately, therapeutic trials of antioxidants have been uniformly disappointing. There is provisional evidence implicating mitochondrial dysfunction as a source of oxidative stress in preeclampsia. Here we provide evidence that mitochondrial reactive oxygen species mediates endothelial dysfunction and establish that directly targeting mitochondrial scavenging may provide a protective role. Human umbilical vein endothelial cells exposed to 3% plasma from women with pregnancies complicated by preeclampsia resulted in a significant decrease in mitochondrial function with a subsequent significant increase in mitochondrial superoxide generation compared to cells exposed to plasma from women with uncomplicated pregnancies. Real-time PCR analysis showed increased expression of inflammatory markers TNF-α, TLR-9 and ICAM-1 respectively in endothelial cells treated with preeclampsia plasma. MitoTempo is a mitochondrial-targeted antioxidant, pre-treatment of cells with MitoTempo protected against hydrogen peroxide-induced cell death. Furthermore MitoTempo significantly reduced mitochondrial superoxide production in cells exposed to preeclampsia plasma by normalising mitochondrial metabolism. MitoTempo significantly altered the inflammatory profile of plasma treated cells. These novel data support a functional role for mitochondrial redox signaling in modulating the pathogenesis of preeclampsia and identifies mitochondrial-targeted antioxidants as potential therapeutic candidates.

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

Endothelial Nitric Oxide Pathways in the Pathophysiology of Dengue: A Prospective Observational Study.

PubMed

Yacoub, Sophie; Lam, Phung Khanh; Huynh, Trieu Trung; Nguyen Ho, Hong Hanh; Dong Thi, Hoai Tam; Van, Nguyen Thu; Lien, Le Thi; Ha, Quyen Nguyen Than; Le, Duyen Huynh Thi; Mongkolspaya, Juthathip; Culshaw, Abigail; Yeo, Tsin Wen; Wertheim, Heiman; Simmons, Cameron; Screaton, Gavin; Wills, Bridget

2017-10-16

Dengue can cause increased vascular permeability that may lead to hypovolemic shock. Endothelial dysfunction may underlie this; however, the association of endothelial nitric oxide (NO) pathways with disease severity is unknown. We performed a prospective observational study in 2 Vietnamese hospitals, assessing patients presenting early (<72 hours of fever) and patients hospitalized with warning signs or severe dengue. The reactive hyperemic index (RHI), which measures endothelium-dependent vasodilation and is a surrogate marker of endothelial function and NO bioavailability, was evaluated using peripheral artery tonometry (EndoPAT), and plasma levels of l-arginine, arginase-1, and asymmetric dimethylarginine were measured at serial time-points. The main outcome of interest was plasma leakage severity. Three hundred fourteen patients were enrolled; median age of the participants was 21(interquartile range, 13-30) years. No difference was found in the endothelial parameters between dengue and other febrile illness. Considering dengue patients, the RHI was significantly lower for patients with severe plasma leakage compared to those with no leakage (1.46 vs 2.00; P < .001), over acute time-points, apparent already in the early febrile phase (1.29 vs 1.75; P = .012). RHI correlated negatively with arginase-1 and positively with l-arginine (P = .001). Endothelial dysfunction/NO bioavailability is associated with worse plasma leakage, occurs early in dengue illness and correlates with hypoargininemia and high arginase-1 levels. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

Endothelial Nitric Oxide Pathways in the Pathophysiology of Dengue: A Prospective Observational Study

PubMed Central

Yacoub, Sophie; Lam, Phung Khanh; Huynh, Trieu Trung; Nguyen Ho, Hong Hanh; Dong Thi, Hoai Tam; Van, Nguyen Thu; Lien, Le Thi; Ha, Quyen Nguyen Than; Le, Duyen Huynh Thi; Mongkolspaya, Juthathip; Culshaw, Abigail; Yeo, Tsin Wen; Wertheim, Heiman; Simmons, Cameron; Screaton, Gavin; Wills, Bridget

2017-01-01

Abstract Background Dengue can cause increased vascular permeability that may lead to hypovolemic shock. Endothelial dysfunction may underlie this; however, the association of endothelial nitric oxide (NO) pathways with disease severity is unknown. Methods We performed a prospective observational study in 2 Vietnamese hospitals, assessing patients presenting early (<72 hours of fever) and patients hospitalized with warning signs or severe dengue. The reactive hyperemic index (RHI), which measures endothelium-dependent vasodilation and is a surrogate marker of endothelial function and NO bioavailability, was evaluated using peripheral artery tonometry (EndoPAT), and plasma levels of l-arginine, arginase-1, and asymmetric dimethylarginine were measured at serial time-points. The main outcome of interest was plasma leakage severity. Results Three hundred fourteen patients were enrolled; median age of the participants was 21(interquartile range, 13–30) years. No difference was found in the endothelial parameters between dengue and other febrile illness. Considering dengue patients, the RHI was significantly lower for patients with severe plasma leakage compared to those with no leakage (1.46 vs 2.00; P < .001), over acute time-points, apparent already in the early febrile phase (1.29 vs 1.75; P = .012). RHI correlated negatively with arginase-1 and positively with l-arginine (P = .001). Conclusions Endothelial dysfunction/NO bioavailability is associated with worse plasma leakage, occurs early in dengue illness and correlates with hypoargininemia and high arginase-1 levels. PMID:28673038

MCP-1-Induced Protein Promotes Endothelial-Like and Angiogenic Properties in Human Bone Marrow Monocytic Cells

PubMed Central

Wang, Kangkai; Zhelyabovska, Olga; Saad, Yasser; Kolattukudy, Pappachan E.

2013-01-01

Monocytic cells enhance neovascularization by releasing proangiogenic mediators and/or by transdifferentiating into endothelial-like cells. However, the mechanisms that govern this transdifferentiation process are largely unknown. Recently, monocyte chemotactic protein-1 (MCP-1)-induced protein (MCPIP) has been identified as a novel CCCH-type zinc-finger protein expressed primarily in monocytic cells. Here, we analyzed whether MCPIP might exert angiogenic effects by promoting differentiation of monocytic cells into endothelial cell (EC)-like phenotype. The expression of MCPIP increased during MCP-1-induced transdifferentiation in human bone marrow mononuclear cells (BMNCs). Knockdown of MCPIP with small interfering RNA (siRNA) abolished MCP-1-induced expression of EC markers Flk-1 and Tie-2 in human BMNCs. BMNCs transfected with MCPIP expression vector displayed EC-like morphology accompanied by downregulation of monocytic markers CD14 and CD11b, upregulation of EC markers Flk-1 and Tie-2, induction of cadherin (cdh)-12 and -19, activation of endoplasmic reticulum (ER) stress, and autophagy. Knockdown of cdh-12 or cdh-19 markedly inhibited MCPIP-induced enhancement of cell attachment and EC-marker expression. Inhibition of ER stress by tauroursodeoxycholate abolished MCPIP-induced expression of EC markers. Inhibition of autophagy by knockdown of Beclin-1 with siRNA or by an autophagy inhibitor 3′-methyladenine inhibited MCPIP-induced expression of EC markers. Expression of MCPIP in BMNCs enhanced uptake of acetylated low-density lipoprotein (acLDL), formation of EC-colony, incorporation of cells into capillary-like structure on Matrigel, and exhibited increased neovascularization in the ischemic hindlimb in mice. These results demonstrate that MCPIP may be an important regulator of inflammatory angiogenesis and provide novel mechanistic insights into the link between MCP-1 and cardiovascular diseases. PMID:24008336

In vitro biomimetic platforms featuring a perfusion system and 3D spheroid culture promote the construction of tissue-engineered corneal endothelial layers.

PubMed

Li, Shanyi; Han, Yuting; Lei, Hao; Zeng, Yingxin; Cui, Zekai; Zeng, Qiaolang; Zhu, Deliang; Lian, Ruiling; Zhang, Jun; Chen, Zhe; Chen, Jiansu

2017-04-10

Corneal endothelial cells (CECs) are very important for the maintenance of corneal transparency. However, in vitro, CECs display limited proliferation and loss of phenotype via endothelial to mesenchymal transformation (EMT) and cellular senescence. In this study, we demonstrate that continuous supplementary nutrition using a perfusion culture bioreactor and three-dimensional (3D) spheroid culture can be used to improve CEC expansion in culture and to construct a tissue-engineered CEC layer. Compared with static culture, perfusion-derived CECs exhibited an increased proliferative ability as well as formed close cell-cell contact junctions and numerous surface microvilli. We also demonstrated that the CEC spheroid culture significantly down-regulated gene expression of the proliferation marker Ki67 and EMT-related markers Vimentin and α-SMA, whereas the gene expression level of the CEC marker ATP1A1 was significantly up-regulated. Furthermore, use of the perfusion system in conjunction with a spheroid culture on decellularized corneal scaffolds and collagen sheets promoted the generation of CEC monolayers as well as neo-synthesized ECM formation. This study also confirmed that a CEC spheroid culture on a curved collagen sheet with controlled physiological intraocular pressure could generate a CEC monolayer. Thus, our results show that the use of a perfusion system and 3D spheroid culture can promote CEC expansion and the construction of tissue-engineered corneal endothelial layers in vitro.

Comparison of cardiovascular disease risk markers in HIV-infected patients receiving abacavir and tenofovir: the nucleoside inflammation, coagulation and endothelial function (NICE) study

PubMed Central

Wohl, David A; Arnoczy, Gretchen; Fichtenbaum, Carl J; Campbell, Thomas; Taiwo, Babafemi; Hicks, Charles; McComsey, Grace A; Koletar, Susan; Sax, Paul; Tebas, Pablo; Ha, Belinda; Massengale, Kelly; Walsh, Kendall; Stein, James H

2015-01-01

Background The association between abacavir (ABC) and cardiovascular disease (CVD) risk in HIV-infected individuals is unclear. Putative mechanisms for an effect of ABC on CVD risk including endothelial dysfunction have been proposed; however, a biological mechanism has not been established. Methods This was a cross-sectional study of HIV-infected subjects with HIV RNA levels <400 copies/ml, who were randomly assigned to ABC or tenofovir (TDF) as initial therapy during a prior clinical trial. A small cohort of subjects on zidovudine (AZT; not randomly assigned) were studied to explore long-term exposure to this agent. All underwent brachial artery ultrasound for flow-mediated dilation (FMD), and D-dimer, high-sensitivity C-reactive protein (hsCRP), interleukin-6 (IL-6) and fasting lipids were measured. Between-arm differences were evaluated by multivariable linear or logistic regression modelling. Results There were 148 subjects (46 on ABC, 72 on TDF and 30 on AZT). Demographic characteristics were balanced across the groups except, as expected, AZT-treated participants were older, had higher CD4+ T-cell counts, and longer antiretroviral therapy duration. After adjusting for age, brachial artery diameter, and treatment duration, FMD was similar in those on ABC (3.9%) and TDF (5.4%; P=0.181). FMD was higher in those on AZT (6.1%; P<0.005). Levels of IL-6, hsCRP and detectable D-dimer were similar between groups. Conclusions Among individuals assigned to ABC or TDF in randomized clinical trials there were no significant differences in FMD or markers of inflammation and coagulation. Whether ABC contributes to risk of CVD remains unclear, but our results suggest that endothelial dysfunction, heightened inflammation, and altered coagulation are unlikely to be mechanisms by which the drug could increase CVD risk above that seen with TDF. PMID:23985706

Serum podocalyxin is significantly increased in early-onset preeclampsia and may represent a novel marker of maternal endothelial cell dysfunction.

PubMed

Chen, Qi; Wang, Yao; Li, Ying; Zhao, Min; Nie, Guiying

2017-11-01

Podocalyxin is a glomerular podocyte protein and increased in urine of preeclampsia. However, podocalyxin is also expressed in endothelial cells of other organs. Here we investigated whether podocalyxin is detectable in pregnant serum and whether the levels are altered in preeclampsia. Podocalyxin was determined by ELISA in sera collected from normal pregnancy across gestation (n = 44) and from preeclamptic pregnancies at diagnosis (n = 34) with gestation-age-matched controls (n = 68). Immunohistochemistry examined podocalyxin in placentas and in 32 human tissues on a tissue array. Human umbilical vein endothelial cells (HUVECs) were treated with interleukin (IL)-6 and podocalyxin was analysed by ELISA and western blotting. Podocalyxin was detected in serum of normal pregnancy, with levels increasing progressively with advancing gestation. Podocalyxin serum levels were significantly elevated in preeclampsia, especially the early-onset subtype. Within the placenta, blood vessels but not trophoblasts expressed podocalyxin, and preeclampsia didn't differ from controls. Endothelial cells in all 32 human organs examined, as well as HUVECs, expressed podocalyxin. Its levels increased in the conditioned media but decreased in the lysates when HUVECs were treated with IL-6. Podocalyxin likely derived from maternal endothelial cells is present in pregnant serum and significantly increased in early-onset preeclampsia. Podocalyxin release was stimulated by IL-6 in HUVECs.

Inflammatory Markers and Preeclampsia: A Systematic Review.

PubMed

Black, Kathleen Darrah; Horowitz, June Andrews

Preeclampsia (PE), a serious and variable pregnancy complication affecting 5%-10% of the obstetric population, has an undetermined etiology, yet inflammation is concomitant with its development, particularly in relation to endothelial dysfunction. The purpose of this systematic review was to examine the published evidence concerning an association between PE and inflammatory markers for their usefulness in the prediction or early identification of women with PE in antepartum clinical settings. In this systematic review, we used the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. The Cumulative Index for Nursing and Allied Health and MEDLINE/OVID were the electronic databases used for identifying published articles. We placed no time limit on the publication year. The search generated 798 articles. After removing duplicates, screening abstracts, and conducting full-text reviews, we retained 73 articles and examined 57 unique markers. This review shows that C-reactive protein and the cytokines, specifically the proinflammatory markers IL-6, IL-8, and tumor necrosis factor alpha, garner the most support as potential inflammatory markers for clinical surveillance of PE, particularly during the second and third trimesters. Based on this review, we cannot recommend any single inflammatory marker for routine clinical use to predict/identify PE onset or progression. Research is recommended to examine a combination panel of these four inflammatory markers both with and without clinical risk factors toward the goal of translation to practice.

Establishment of a translational endothelial cell model using directed differentiation of induced pluripotent stem cells from Cynomolgus monkey.

PubMed

Thoma, Eva C; Heckel, Tobias; Keller, David; Giroud, Nicolas; Leonard, Brian; Christensen, Klaus; Roth, Adrian; Bertinetti-Lapatki, Cristina; Graf, Martin; Patsch, Christoph

2016-10-25

Due to their broad differentiation potential, pluripotent stem cells (PSCs) offer a promising approach for generating relevant cellular models for various applications. While human PSC-based cellular models are already advanced, similar systems for non-human primates (NHPs) are still lacking. However, as NHPs are the most appropriate animals for evaluating the safety of many novel pharmaceuticals, the availability of in vitro systems would be extremely useful to bridge the gap between cellular and animal models. Here, we present a NHP in vitro endothelial cell system using induced pluripotent stem cells (IPSCs) from Cynomolgus monkey (Macaca fascicularis). Based on an adapted protocol for human IPSCs, we directly differentiated macaque IPSCs into endothelial cells under chemically defined conditions. The resulting endothelial cells can be enriched using immuno-magnetic cell sorting and display endothelial marker expression and function. RNA sequencing revealed that the differentiation process closely resembled vasculogenesis. Moreover, we showed that endothelial cells derived from macaque and human IPSCs are highly similar with respect to gene expression patterns and key endothelial functions, such as inflammatory responses. These data demonstrate the power of IPSC differentiation technology to generate defined cell types for use as translational in vitro models to compare cell type-specific responses across species.

Generation and characterization of a spontaneously immortalized endothelial cell line from mice microcirculation.

PubMed

Loiola, Rodrigo A; Torres, Tathiany C; Aburaya, Carla M; Landgraf, Maristella A; Landgraf, Richardt G; Bosco Pesquero, João; Fernandes, Liliam

2013-05-01

Endothelial cells from microvasculature are directly involved in a large number of vascular diseases; however, culture of these cells is problematic, since most methodologies employ proteolytic enzymes or mechanical techniques, leading to cell damage and contamination of endothelial cultures with other cellular types. Besides, primary cultured cells have a short life span in vitro and undergo replicative senescence after 3-4 passages, limiting long-term studies. In the present work we report the generation of a spontaneously immortalized endothelial culture obtained from mice pulmonary capillaries. Firstly, primary (third passage) and immortalized (100th) cultures were established. Further, monoclonal populations were obtained by serial dilutions from immortalized cultures. Cells were analyzed according to: (1) morphological appearance, (2) expression of specific endothelial markers by fluorescent staining [von Willebrand Factor (vWF), endothelial nitric oxide synthase (eNOS), angiotensin converting enzyme (ACE) and Ulex europaeus (UEA-1)] and by flow cytometry (endoglin, VE-cadherin and VCAM-1), and (3) release of nitric oxide (NO), assessed by the specific fluorescent dye DAF-2 DA, and prostacyclin (PGI2), quantified by enzyme immune assay. In both cultures cells grew in monolayers and presented cobblestone appearance at confluence. Positive staining for vWF, eNOS, ACE and UEA-1 was detected in cloned as well as in early-passage cultured cells. Similarly, cultures presented equal expressions of endoglin, VE-cadherin and VCAM-1. Values of NO and PGI2 levels did not differ between cultures. From these results we confirm that the described spontaneously immortalized endothelial cell line is capable of unlimited growth and retains typical morphological and functional properties exhibited by primary cultured cells. Therefore, the endothelial cell line described in the present study can become a suitable tool in the field of endothelium research and can be useful for

Decursin inhibited proliferation and angiogenesis of endothelial cells to suppress diabetic retinopathy via VEGFR2.

PubMed

Yang, Ying; Yang, Ke; Li, Yiping; Li, Xianli; Sun, Qiangming; Meng, Hua; Zeng, Ying; Hu, Yong; Zhang, Ying

2013-09-25

Diabetes induces pathologic proliferation and angiogenesis in the retina that leads to catastrophic loss of vision. Decursin is a novel therapeutic that targets the vascular endothelial growth factor (VEGF) receptor (VEGFR) with putative anti-proliferative and anti-angiogenic activities. Thereby we utilized human retinal microvascular endothelial cells (HRMEC) and human umbilical vein endothelial cells (HUVEC) under conditions of excess glucose to explore dose-dependent responses of decursin on markers of migration, angiogenesis, and proliferation. Decursin dose-dependently inhibited tube formation, VEGFR-2 expression, along with relative metabolic activity and 5-bromo-2'-deoxy-uridine (BrdU) activity in both cell lines. We then correlated our findings to the streptozotocin-induced rat model of diabetes. Following three months of decursin treatment VEGFR-2 expression was significantly inhibited. Our data would suggest that decursin may be a potent anti-angiogenic and anti-proliferative agent targeting the VEGFR-2 signaling pathway, which significantly inhibits diabetic retinal neovascularization. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

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.

Effect of cerium oxide nanoparticles on inflammation in vascular endothelial cells

PubMed Central

Gojova, Andrea; Lee, Jun-Tae; Jung, Heejung S.; Guo, Bing; Barakat, Abdul I.; Kennedy, Ian M.

2010-01-01

Because vascular endothelial cell inflammation is critical in the development of cardiovascular pathology, we hypothesized that direct exposure of human aortic endothelial cells (HAECs) to ultrafine particles induces an inflammatory response. To test the hypothesis, we incubated HAECs for 4 h with different concentrations (0.001–50 μg/ml) of CeO2 nanoparticles and subsequently measured mRNA levels of the three inflammatory markers intercellular adhesion molecule 1 (ICAM-1), interleukin (IL)-8, and monocyte chemotactic protein (MCP-1) using real-time polymerase chain reaction (PCR). Ceria nanoparticles caused very little inflammatory response in HAECs, even at the highest dose. This material is apparently rather benign in comparison with Y2O3 and ZnO nanoparticles that we have studied previously. These results suggest that inflammation in HAECs following acute exposure to metal oxide nanoparticles depends strongly on particle composition. PMID:19558244

Cross-sectional and longitudinal associations between serum uric acid and endothelial function in subjects with treated hypertension.

PubMed

Tanaka, Atsushi; Kawaguchi, Atsushi; Tomiyama, Hirofumi; Ishizu, Tomoko; Matsumoto, Chisa; Higashi, Yukihito; Takase, Bonpei; Suzuki, Toru; Ueda, Shinichiro; Yamazaki, Tsutomu; Furumoto, Tomoo; Kario, Kazuomi; Inoue, Teruo; Koba, Shinji; Takemoto, Yasuhiko; Hano, Takuzo; Sata, Masataka; Ishibashi, Yutaka; Maemura, Koji; Ohya, Yusuke; Furukawa, Taiji; Ito, Hiroshi; Yamashina, Akira; Node, Koichi

2018-06-06

The endothelial dysfunction-arterial stiffness-atherosclerosis continuum plays an important pathophysiological role in hypertension. The aim of this study was to investigate the cross-sectional association between serum uric acid (SUA) and vascular markers related to this continuum, and to assess the longitudinal association between SUA and endothelial function that represents the initial step of the continuum. We evaluated the baseline associations between SUA levels and vascular markers that included flow-mediated vasodilatation (FMD), brachial-ankle pulse wave velocity (baPWV), and common carotid artery intima-media thickness (CCA-IMT) in 648 subjects receiving antihypertensive treatment. The longitudinal association between baseline SUA levels and FMD measured at 1.5 and 3 yr of follow-up was also investigated. At baseline, modest, but significant correlations were observed between SUA and FMD in females (r = -0.171), baPWV in males with SUA >368.78 μmol/L (r = -0.122) and in females with a SUA level ≤ 362.83 μmol/L (r = 0.217), mean CCA-IMT in females with a SUA level ≤ 333.09 μmol/L (r = 0.139), and max CCA-IMT in females with SUA level ≤ 333.09 μmol/L (r = 0.138). A longitudinal association between SUA and FMD was less observed in males. In females, the baseline SUA was associated significantly with FMD values at 1.5 yr (r = -0.211), and SUA levels >237.92 μmol/L were associated significantly and independently with FMD values at 3 yr (r = -0.166). Lower SUA levels were associated with better vascular markers of the continuum, especially in females. Furthermore, we observed a longitudinal association between SUA and endothelial function, suggesting SUA level may be a potential marker of the continuum in hypertension. Copyright © 2017 Elsevier B.V. All rights reserved.

Angiogenesis in mucous retention cyst: a human in vivo-like model of endothelial cell differentiation in mucous substrate.

PubMed

Swelam, Wael; Ida-Yonemochi, Hiroko; Saku, Takashi

2005-01-01

Mucous retention cysts contain a mucous pool in the lumina, in which pure angiogenic processes are occasionally observed. By using this unique human material, our aim was to understand the in vivo angiogenic process. Fifteen surgical tissue samples of mucous retention cysts of the lip were examined for expression of vascular endothelial markers and extracellular matrix molecules by immunohistochemistry and in situ hybridization (ISH). Endothelial cells forming new vascular channels showed immunopositivities for CD31, CD34, vascular endothelial growth factor (VEGF), and von Willebrand factor (vWF). These newly formed capillaries were surrounded by tenascin-positive matrices and further by a dense infiltration of CD68-positive cells with foamy to epitheloid appearances. Some of these cells were simultaneously positive for CD34, VEGF, and one of its receptors, Flk-1, and they showed definite mRNA as well as protein signals for tenascin. In addition, these cells often tended to be aligned, which suggested tubule formation. The results suggest that monocyte/macrophage lineage cells are a major source for endothelial cells at least in mucous retention cysts and that tenascin produced by those cells plays an important role in differentiation of endothelial cells.

The effects of vitamin E and omega-3 PUFAs on endothelial function among adolescents with metabolic syndrome.

PubMed

Ahmadi, Alireza; Gharipour, Mojgan; Arabzadeh, Gholamreza; Moin, Payam; Hashemipour, Mahin; Kelishadi, Roya

2014-01-01

The present study aims to explore the effects of vitamin E and omega-3 on endothelial function indicators among adolescents with metabolic syndrome. In a randomized, double blind, and placebo-controlled trial, 90 young individuals, aged 10 to 18 years, with metabolic syndrome were randomly assigned to receive either vitamin E tablets (400 IU/day) or omega-3 tablets (2.4 gr/day) or placebo. For assessing endothelial functional state, the serum level of vascular endothelial growth factor (VEGF) was measured by ELISA test. The use of omega-3 supplementation for eight weeks led to significant increase in serum HDL level compared with the group treated with vitamin E or placebo group. In this regard, no significant correlations were found between the change in VEGF and baseline levels of other markers including anthropometric indices and serum lipids. Omega-3 could significantly reduce VEGF with the presence of other baseline variables (Beta = -12.55; P = 0.012). The administration of omega-3 can effectively improve endothelial function in adolescents with metabolic syndrome by reducing the level of serum VEGF, as a major index for atherosclerosis progression and endothelial destabilization. Omega-3 can be proposed as a VEGF antagonist for improving endothelial function in metabolic syndrome. The clinical implications of our findings should be assessed in future studies.

The apolipoprotein B/A1 ratio is associated with reactive oxygen metabolites and endothelial dysfunction in statin-treated patients with coronary artery disease.

PubMed

Emoto, Takuo; Sawada, Takahiro; Morimoto, Natsumi; Tenjin, Takako; Wakimoto, Taku; Ikeda, Fumie; Sato, Chiaki; Terashita, Daisuke; Mizoguchi, Taiji; Mizuguchi, Takao; Okamoto, Hiroshi; Matsuo, Yosuke; Kim, Sushi-Ku; Takarada, Akira; Yokoyama, Mitsuhiro

2013-01-01

The prognostic significance of the apolipoprotein B/A1 (ApoB/A1) ratio in statintreated patients with coronary artery disease (CAD) is unknown. We aimed to evaluate the association of the ApoB/A1 ratio with oxidative stress and endothelial dysfunction in these patients. We enrolled 62 consecutive statin-treated patients who underwent percutaneous coronary intervention (PCI). Their lipid profiles, diacron-reactive oxygen metabolites (d-ROMs), as a marker of oxidative stress, flow-mediated dilatation (FMD), as a marker of vascular endothelial function, and C-reactive protein (CRP) levels, as a marker of inflammation, were measured. Our study population comprised 44 men and 18 women (mean age, 70.5 ± 2.5 years). The ApoB/A1 ratio was positively correlated with the results of the d-ROMs test (p=0.004, r=0.36) and CRP level (p=0.02, r=0.30) and negatively correlated with the %FMD (p=0.005, r=-0.40). A multivariate logistic regression analysis showed that the most powerful predictive factor for the d-ROMs was the ApoB/A1 ratio (p=0.026). We therefore divided patients into two groups according to the cutoff point reported by the INTERHEART study: a low ApoB/A1 ratio (<0.641, n=26) and a high ApoB/A1 ratio (>0.641, n=36). The patients with a high ApoB/A1 ratio had higher levels of d-ROMs and CRP, and tended to have a lower %FMD. The ApoB/A1 ratio was associated with the d-ROMs, a marker of oxidative stress, endothelial dysfunction and inflammation, and could be useful as a residual atherosclerotic risk marker to help prevent CAD in statin-treated patients.

Endothelial-regenerating cells: an expanding universe.

PubMed

Steinmetz, Martin; Nickenig, Georg; Werner, Nikos

2010-03-01

Atherosclerosis is the most common cause for cardiovascular diseases and is based on endothelial dysfunction. A growing body of evidence suggests the contribution of bone marrow-derived endothelial progenitor cells, monocytic cells, and mature endothelial cells to vessel formation and endothelial rejuvenation. To this day, various subsets of these endothelial-regenerating cells have been identified according to cellular origin, phenotype, and properties in vivo and in vitro. However, the definition and biology, especially of endothelial progenitor cells, is complex and under heavy debate. In this review, we focus on current definitions of endothelial progenitor cells, highlight the clinical relevance of endothelial-regenerating cells, and provide new insights into cell-cell interactions involved in endothelial cell rejuvenation.

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

A sensitive ELISA for measuring the adhesion of leukocytic cells to human endothelial cells.

PubMed

Krakauer, T

1994-12-28

A new, sensitive ELISA using monoclonal antibodies reactive with surface molecules specific for various leukocytes was devised to measure the attachment of these cells to cultured monolayers of human umbilical vein endothelial cells. Preparations of peripheral blood mononuclear cells, a human monocytic cell line (THP-1) and a human lymphoblastic T cell line (MOLT-4) were used to test the sensitivity of this method and compare it with the conventional 51Cr-radiolabeled cell assay. The extent of adhesion to endothelial cells was assayed by measuring the optical density produced by a complex of peroxidase-labeled streptavidin, biotin-conjugated F(ab')2 anti-mouse Ig and monoclonal antibody on fixed leukocytic cells that had adhered to endothelial cells. This method is fast and sensitive, eliminates the use of radioisotopes, and, because the detection uses a specific marker on the cell of interest, can be used in preparations of unseparated mixtures of cells. As this is a microassay, using relatively small number of cells and reagents, the methodology can be applied to screen a large number of therapeutic agents that may regulate adhesion. Using this method, the anti-inflammatory corticosteroid, dexamethasone, was found to inhibit the adhesion of THP-1 and MOLT-4 cells to cytokine-activated endothelial cells.

Cooperative control of blood compatibility and re-endothelialization by immobilized heparin and substrate topography.

PubMed

Ding, Yonghui; Yang, Meng; Yang, Zhilu; Luo, Rifang; Lu, Xiong; Huang, Nan; Huang, Pingbo; Leng, Yang

2015-03-01

A wide variety of environmental cues provided by the extracellular matrix, including biophysical and biochemical cues, are responsible for vascular cell behavior and function. In particular, substrate topography and surface chemistry have been shown to regulate blood and vascular compatibility individually. The combined impact of chemical and topographic cues on blood and vascular compatibility, and the interplay between these two types of cues, are subjects that are currently being explored. In the present study, a facile polydopamine-mediated approach is introduced for immobilization of heparin on topographically patterned substrates, and the combined effects of these cues on blood compatibility and re-endothelialization are systematically investigated. The results show that immobilized heparin and substrate topography cooperatively modulate anti-coagulation activity, endothelial cell (EC) attachment, proliferation, focal adhesion formation and endothelial marker expression. Meanwhile, the substrate topography is the primary determinant of cell alignment and elongation, driving in vivo-like endothelial organization. Importantly, combining immobilized heparin with substrate topography empowers substantially greater competitive ability of ECs over smooth muscle cells than each cue individually. Moreover, a model is proposed to elucidate the cooperative interplay between immobilized heparin and substrate topography in regulating cell behavior. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Isolation of a circulating CD45−, CD34dim cell population and validation of their endothelial phenotype

PubMed Central

Tropea, Margaret M.; Harper, Bonnie J. A.; Graninger, Grace M.; Phillips, Terry M.; Ferreyra, Gabriela; Mostowski, Howard S.; Danner, Robert L.; Suffredini, Anthony F.; Solomon, Michael A.

2016-01-01

Summary Accurately detecting circulating endothelial cells (CECs) is important since their enumeration has been proposed as a biomarker to measure injury to the vascular endothelium. However, there is no single methodology for determining CECs in blood, making comparison across studies difficult. Many methods for detecting CECs rely on characteristic cell surface markers and cell viability indicators, but lack secondary validation. Here, a CEC population in healthy adult human subjects was identified by flow cytometry as CD45−, CD34dim that is comparable to a previously described CD45−, CD31bright population. In addition, nuclear staining with 7-aminoactinomycin D (7-AAD) was employed as a standard technique to exclude dead cells. Unexpectedly, the CD45−, CD34dim, 7-AAD− CECs lacked surface detectable CD146, a commonly used marker of CECs. Furthermore, light microscopy revealed this cell population to be composed primarily of large cells without a clearly defined nucleus. Nevertheless, immunostains still demonstrated the presence of the lectin Ulex europaeus and van Willebrand factor. Ultramicro analytical immunochemistry assays for the endothelial cell proteins CD31, CD34, CD62E, CD105, CD141, CD144 and vWF indicated these cells possess an endothelial phenotype. However, only a small amount of RNA, which was mostly degraded, could be isolated from these cells. Thus the majority of CECs in healthy individuals as defined by CD45−, CD34dim, and 7-AAD− have shed their CD146 surface marker and are senescent cells without an identifiable nucleus and lacking RNA of sufficient quantity and quality for transcriptomal analysis. This study highlights the importance of secondary validation of CEC identification. PMID:25057108

Deterioration of endothelial function of micro- and macrocirculation in patients with diabetes type 1 and 2.

PubMed

Besic, Hana; Jeraj, Luka; Spirkoska, Ana; Jezovnik, Mateja K; Poredoš, Pavel

2017-08-01

Vascular complications are an important cause of morbidity in patients with diabetes mellitus (DM). Endothelial dysfunction is an early marker of atherosclerosis and has already been shown in macrocirculation of diabetic patients; however, data on endothelial function of microcirculation is scarce. Our aim was to evaluate endothelial function in macro- and microcirculation and their interrelationship in patients with type 1 and 2 DM. The study included 30 patients with type 1 DM, 30 patients with type 2 DM and 25 healthy controls. The endothelial function of large arteries was studied measuring flow-mediated dilation (FMD). Peripheral arterial tonometry was used for investigation of the endothelial function of microcirculation, measuring Reactive Hyperemia Index (RHI) and Augmentation Index (AI). In comparison to controls, both DM groups had decreased FMD: type 1 (4.0±5.0% vs. 10.0±7.8%, P=0.005) and type 2 (5.0±0.6% vs. 10.0±7.8%, P=0.007). However, only type 2 DM group had a lower RHI (1.71±0.44 vs. 2.05±0.54, P=0.017) in comparison to controls. Patients with type 1 and 2 DM had deteriorated functional capability of macrocirculation. However, endothelial dysfunction of microcirculation was present only in type 2 DM patients. Type 2 DM patients were also at higher risk for atherosclerosis because of the more frequent presence of risk factors.

Endotoxin induces fibrosis in vascular endothelial cells through a mechanism dependent on transient receptor protein melastatin 7 activity.

PubMed

Echeverría, Cesar; Montorfano, Ignacio; Hermosilla, Tamara; Armisén, Ricardo; Velásquez, Luis A; Cabello-Verrugio, Claudio; Varela, Diego; Simon, Felipe

2014-01-01

The pathogenesis of systemic inflammatory diseases, including endotoxemia-derived sepsis syndrome, is characterized by endothelial dysfunction. It has been demonstrated that the endotoxin lipopolysaccharide (LPS) induces the conversion of endothelial cells (ECs) into activated fibroblasts through endothelial-to-mesenchymal transition mechanism. Fibrogenesis is highly dependent on intracellular Ca2+ concentration increases through the participation of calcium channels. However, the specific molecular identity of the calcium channel that mediates the Ca2+ influx during endotoxin-induced endothelial fibrosis is still unknown. Transient receptor potential melastatin 7 (TRPM7) is a calcium channel that is expressed in many cell types, including ECs. TRPM7 is involved in a number of crucial processes such as the conversion of fibroblasts into activated fibroblasts, or myofibroblasts, being responsible for the development of several characteristics of them. However, the role of the TRPM7 ion channel in endotoxin-induced endothelial fibrosis is unknown. Thus, our aim was to study whether the TRPM7 calcium channel participates in endotoxin-induced endothelial fibrosis. Using primary cultures of ECs, we demonstrated that TRPM7 is a crucial protein involved in endotoxin-induced endothelial fibrosis. Suppression of TRPM7 expression protected ECs from the fibrogenic process stimulated by endotoxin. Downregulation of TRPM7 prevented the endotoxin-induced endothelial markers decrease and fibrotic genes increase in ECs. In addition, TRPM7 downregulation abolished the endotoxin-induced increase in ECM proteins in ECs. Furthermore, we showed that intracellular Ca2+ levels were greatly increased upon LPS challenge in a mechanism dependent on TRPM7 expression. These results demonstrate that TRPM7 is a key protein involved in the mechanism underlying endotoxin-induced endothelial fibrosis.

Endothelial to mesenchymal transition in the cardiovascular system.

PubMed

Gong, Hui; Lyu, Xing; Wang, Qiong; Hu, Min; Zhang, Xiangyu

2017-09-01

Endothelial to mesenchymal transition (EndMT) is a special type of epithelial to mesenchymal transition. It is a process that is characterized by the loss of features of endothelial cells and acquisition of specific markers of mesenchymal cells. A variety of stimuli, such as inflammation, growth factors, and hypoxia, regulate EndMT through various signaling pathways and intracellular transcription factors. It has been demonstrated that epigenetic modifications are also involved in this process. Recent studies have identified the essential role of EndMT in the cardiovascular system. EndMT contributes to steps in cardiovascular development, such as cardiac valve formation and septation, as well as the pathogenesis of various cardiovascular disorders, such as congenital heart disease, myocardial fibrosis, myocardial infarction and pulmonary arterial hypertension. Thus, comprehensive understanding of the underlying mechanisms of EndMT will provide novel therapeutic strategies to overcome congenital heart disease due to abnormal development and other cardiovascular diseases. This review will focus on summarizing the currently understood signaling pathways and epigenetic modifications involved in the regulation of EndMT and the role of EndMT in pathophysiological conditions of the cardiovascular system. Copyright © 2017. Published by Elsevier Inc.

Endothelial ischemia-reperfusion injury in humans: association with age and habitual exercise.

PubMed

Devan, Allison E; Umpierre, Daniel; Harrison, Michelle L; Lin, Hsin-Fu; Tarumi, Takashi; Renzi, Christopher P; Dhindsa, Mandeep; Hunter, Stacy D; Tanaka, Hirofumi

2011-03-01

Advancing age is a major risk factor for coronary artery disease. Endothelial dysfunction accompanied by increased oxidative stress and inflammation with aging may predispose older arteries to greater ischemia-reperfusion (I/R) injury. Because coronary artery ischemia cannot be induced safely, the effects of age and habitual endurance exercise on endothelial I/R injury have not been determined in humans. Using the brachial artery as a surrogate model of the coronary arteries, endothelial function, assessed by brachial artery flow-mediated dilation (FMD), was measured before and after 20 min of continuous forearm occlusion in young sedentary (n = 10, 24 ± 2 yr) and middle-aged (n = 9, 48 ± 2 yr) sedentary adults to gain insight into the effects of primary aging on endothelial I/R injury. Young (n = 9, 25 ± 1 yr) and middle-aged endurance-trained (n = 9, 50 ± 2 yr) adults were also studied to determine whether habitual exercise provides protection from I/R injury. Fifteen minutes after ischemic injury, FMD decreased significantly by 37% in young sedentary, 35% in young endurance-trained, 68% in middle-aged sedentary, and 50% in middle-aged endurance-trained subjects. FMD returned to baseline levels within 30 min in young sedentary and endurance-trained subjects but remained depressed in middle-aged sedentary and endurance-trained subjects. Circulating markers of antioxidant capacity and inflammation were not related to FMD. In conclusion, advancing age is associated with a greater magnitude and delayed recovery from endothelial I/R injury in humans. Habitual endurance exercise may provide partial protection to the endothelium against this form of I/R injury with advancing age.

Oral Mucosa Harbors a High Frequency of Endothelial Cells: A Novel Postnatal Cell Source for Angiogenic Regeneration.

PubMed

Zhou, Jian; Rogers, Jason H; Lee, Scott H; Sun, DongMing; Yao, Hai; Mao, Jeremy J; Kong, Kimi Y

2017-01-15

Endothelial progenitor cells/endothelial cells (EPCs/ECs) have great potential to treat pathological conditions such as cardiac infarction, muscle ischemia, and bone fractures, but isolation of EPC/ECs from existing cell sources is challenging due to their low EC frequency. We have isolated endothelial progenitor (EP)-like cells from rat oral mucosa and characterized their yield, immunophenotype, growth, and in vivo angiogenic potential. The frequency of EP-like cells derived from oral mucosa is thousands of folds higher than EPCs derived from donor-match bone marrow samples. EP-like cells from oral mucosa were positive for EC markers CD31, VE-Cadherin, and VEGFR2. Oral mucosa-derived EP-like cells displayed robust uptake of acetylated low-density lipoprotein and formed stable capillary networks in Matrigel. Subcutaneously implanted oral mucosa-derived EP-like cells anastomosed with host blood vessels, implicating their ability to elicit angiogenesis. Similar to endothelial colony-forming cells, EP-like cells from oral mucosa have a significantly higher proliferative rate than human umbilical vein endothelial cells. These findings identify a putative EPC source that is easily accessible in the oral cavity, potentially from discarded tissue specimens, and yet with robust yield and potency for angiogenesis in tissue and organ regeneration.

Oral Mucosa Harbors a High Frequency of Endothelial Cells: A Novel Postnatal Cell Source for Angiogenic Regeneration

PubMed Central

Zhou, Jian; Rogers, Jason H.; Lee, Scott H.; Sun, DongMing; Yao, Hai; Mao, Jeremy J.

2017-01-01

Endothelial progenitor cells/endothelial cells (EPCs/ECs) have great potential to treat pathological conditions such as cardiac infarction, muscle ischemia, and bone fractures, but isolation of EPC/ECs from existing cell sources is challenging due to their low EC frequency. We have isolated endothelial progenitor (EP)-like cells from rat oral mucosa and characterized their yield, immunophenotype, growth, and in vivo angiogenic potential. The frequency of EP-like cells derived from oral mucosa is thousands of folds higher than EPCs derived from donor-match bone marrow samples. EP-like cells from oral mucosa were positive for EC markers CD31, VE-Cadherin, and VEGFR2. Oral mucosa-derived EP-like cells displayed robust uptake of acetylated low-density lipoprotein and formed stable capillary networks in Matrigel. Subcutaneously implanted oral mucosa-derived EP-like cells anastomosed with host blood vessels, implicating their ability to elicit angiogenesis. Similar to endothelial colony-forming cells, EP-like cells from oral mucosa have a significantly higher proliferative rate than human umbilical vein endothelial cells. These findings identify a putative EPC source that is easily accessible in the oral cavity, potentially from discarded tissue specimens, and yet with robust yield and potency for angiogenesis in tissue and organ regeneration. PMID:27832737

Cathepsin S Cleavage of Protease-Activated Receptor-2 on Endothelial Cells Promotes Microvascular Diabetes Complications

PubMed Central

Kumar VR, Santhosh; Darisipudi, Murthy N.; Steiger, Stefanie; Devarapu, Satish Kumar; Tato, Maia; Kukarni, Onkar P.; Mulay, Shrikant R.; Thomasova, Dana; Popper, Bastian; Demleitner, Jana; Zuchtriegel, Gabriele; Reichel, Christoph; Cohen, Clemens D.; Lindenmeyer, Maja T.; Liapis, Helen; Moll, Solange; Reid, Emma; Stitt, Alan W.; Schott, Brigitte; Gruner, Sabine; Haap, Wolfgang; Ebeling, Martin; Hartmann, Guido

2016-01-01

Endothelial dysfunction is a central pathomechanism in diabetes-associated complications. We hypothesized a pathogenic role in this dysfunction of cathepsin S (Cat-S), a cysteine protease that degrades elastic fibers and activates the protease-activated receptor-2 (PAR2) on endothelial cells. We found that injection of mice with recombinant Cat-S induced albuminuria and glomerular endothelial cell injury in a PAR2-dependent manner. In vivo microscopy confirmed a role for intrinsic Cat-S/PAR2 in ischemia–induced microvascular permeability. In vitro transcriptome analysis and experiments using siRNA or specific Cat-S and PAR2 antagonists revealed that Cat-S specifically impaired the integrity and barrier function of glomerular endothelial cells selectively through PAR2. In human and mouse type 2 diabetic nephropathy, only CD68+ intrarenal monocytes expressed Cat-S mRNA, whereas Cat-S protein was present along endothelial cells and inside proximal tubular epithelial cells also. In contrast, the cysteine protease inhibitor cystatin C was expressed only in tubules. Delayed treatment of type 2 diabetic db/db mice with Cat-S or PAR2 inhibitors attenuated albuminuria and glomerulosclerosis (indicators of diabetic nephropathy) and attenuated albumin leakage into the retina and other structural markers of diabetic retinopathy. These data identify Cat-S as a monocyte/macrophage–derived circulating PAR2 agonist and mediator of endothelial dysfunction–related microvascular diabetes complications. Thus, Cat-S or PAR2 inhibition might be a novel strategy to prevent microvascular disease in diabetes and other diseases. PMID:26567242

Angiogenic properties of endometrial mesenchymal stromal cells in endothelial co-culture: an in vitro model of endometriosis.

PubMed

Canosa, S; Moggio, A; Brossa, A; Pittatore, G; Marchino, G L; Leoncini, S; Benedetto, C; Revelli, A; Bussolati, B

2017-03-01

Can endometrial mesenchymal stromal cells (E-MSCs) differentiate into endothelial cells in an in vitro co-culture system with human umbilical vein endothelial cells (HUVECs)? E-MSCs can acquire endothelial markers and function in a direct co-culture system with HUVECs. E-MSCs have been identified in the human endometrium as well as in endometriotic lesions. E-MSCs appear to be involved in formation of the endometrial stromal vascular tissue and the support of tissue growth and vascularization. The use of anti-angiogenic drugs appears as a possible therapeutic strategy against endometriosis. This is an in vitro study comprising patients receiving surgical treatment of ovarian endometriosis (n = 9). E-MSCs were isolated from eutopic and ectopic endometrial tissue and were characterized for the expression of mesenchymal and endothelial markers by FACS analysis and Real-Time PCR. CD31 acquisition was evaluated by FACS analysis and immunofluorescence after a 48 h-direct co-culture with green fluorescent protein +-HUVECs. A tube-forming assay was set up in order to analyze the functional potential of their interaction. Finally, co-cultures were treated with the anti-angiogenic agent Cabergoline. A subpopulation of E-MSCs acquired CD31 expression and integrated into tube-like structures when directly in contact with HUVECs, as observed by both FACS analysis and immunofluorescence. The isolation of CD31+ E-MSCs revealed significant increases in CD31, vascular endothelial growth factor receptor 2, TEK receptor tyrosine kinase and vascular endothelial-Cadherin mRNA expression levels with respect to basal and to CD31neg cells (P < 0.05). On the other hand, the expression of mesenchymal genes such as c-Myc, Vimentin, neuronal-Cadherin and sushi domain containing 2 remained unchanged. Cabergoline treatment induced a significant reduction of the E-MSC angiogenic potential (P < 0.05 versus control). Not applicable. Further studies are necessary to investigate the cellular and

Role of Nitric Oxide Signaling in Endothelial Differentiation of Embryonic Stem Cells

PubMed Central

Huang, Ngan F.; Fleissner, Felix; Sun, John

2010-01-01

Signaling pathways that govern embryonic stem cell (ESCs) differentiation are not well characterized. Nitric oxide (NO) is a potent vasodilator that modulates other signaling pathways in part by activating soluble guanylyl cyclase (sGC) to produce cyclic guanosine monophosphate (cGMP). Because of its importance in endothelial cell (EC) growth in the adult, we hypothesized that NO may play a critical role in EC development. Accordingly, we assessed the role of NO in ESC differentiation into ECs. Murine ESCs differentiated in the presence of NO synthase (NOS) inhibitor NG-nitroarginine methyl ester (l-NAME) for up to 11 days were not significantly different from vehicle-treated cells in EC markers. However, by 14 days, l-NAME-treated cells manifested modest reduction in EC markers CD144, FLK1, and endothelial NOS. ESC-derived ECs generated in the presence of l-NAME exhibited reduced tube-like formation in Matrigel. To understand the discrepancy between early and late effects of l-NAME, we assessed the NOS machinery and observed low mRNA expression of NOS and sGC subunits in ESCs, compared to differentiating cells after 14 days. In response to NO donors or activation of NOS or sGC, cellular cGMP levels were undetectable in undifferentiated ESCs, at low levels on day 7, and robustly increased in day 14 cells. Production of cGMP upon NOS activation at day 14 was inhibited by l-NAME, confirming endogenous NO dependence. Our data suggest that NOS elements are present in ESCs but inactive until later stages of differentiation, during which period NOS inhibition reduces expression of EC markers and impairs angiogenic function. PMID:20064011

Facilitated Engraftment of Isolated Islets Coated With Expanded Vascular Endothelial Cells for Islet Transplantation.

PubMed

Barba-Gutierrez, D Alonso; Daneri-Navarro, A; Villagomez-Mendez, J Jesus Alejandro; Kanamune, J; Robles-Murillo, A Karina; Sanchez-Enriquez, S; Villafan-Bernal, J Rafael; Rivas-Carrillo, J D

2016-03-01

Diabetes is complex disease, which involves primary metabolic changes followed by immunological and vascular pathophysiological adjustments. However, it is mostly characterized by an unbalanced decreased number of the β-cells unable to maintain the metabolic requirements and failure to further regenerate newly functional pancreatic islets. The objective of this study was to analyze the properties of the endothelial cells to facilitate the islet cells engraftment after islet transplantation. We devised a co-cultured engineer system to coat isolated islets with vascular endothelial cells. To assess the cell integration of cell-engineered islets, we stained them for endothelial marker CD31 and nuclei counterstained with DAPI dye. We comparatively performed islet transplantations into streptozotocin-induced diabetic mice and recovered the islet grafts for morphometric analyses on days 3, 7, 10, and 30. Blood glucose levels were measured continuously after islet transplantation to monitor the functional engraftment and capacity to achieve metabolic control. Cell-engineered islets showed a well-defined rounded shape after co-culture when compared with native isolated islets. Furthermore, the number of CD31-positive cells layered on the islet surface showed a direct proportion with engraftment capacities and less TUNEL-positive cells on days 3 and 7 after transplantation. We observed that vascular endothelial cells could be functional integrated into isolated islets. We also found that islets that are coated with vascular endothelial cells increased their capacity to engraft. These findings indicate that islets coated with endothelial cells have a greater capacity of engraftment and thus establish a definitely vascular network to support the metabolic requirements. Copyright © 2016 Elsevier Inc. All rights reserved.

Association of Abacavir and Impaired Endothelial Function in Treated and Suppressed HIV-Infected Patients

PubMed Central

Hsue, Priscilla Y.; Hunt, Peter W.; Wu, Yuaner; Schnell, Amanda; Ho, Jennifer E.; Hatano, Hiroyu; Xie, Yu; Martin, Jeffrey N.; Ganz, Peter; Deeks, Steven G.

2009-01-01

Background HIV-infected patients have accelerated atherosclerosis. Abacavir has been associated with increased risk of cardiovascular events, for reasons that remain to be elucidated. As endothelial dysfunction is central to the pathogenesis of atherosclerosis, we tested the hypothesis that current treatment with abacavir is associated with impaired endothelial function. Methods We studied a cohort of 61 antiretroviral-treated patients who had undetectable plasma HIV RNA levels. Endothelial function was assessed by measuring flow-mediated vasodilation (FMD) of the brachial artery. We compared FMD in patients treated with or without abacavir, while adjusting for traditional risk factors and HIV-specific characteristics. Results The median age was 50 years (IQR 45–57). The median duration of HIV infection was 18 years, and the median CD4 cell count was 369 cells/mm3. Thirty subjects (49%) were receiving abacavir. Overall, the median FMD in the HIV-infected patients was low (3.5%; IQR 2.3–5.6%). The FMD was lower in the abacavir-treated patients than those not on abacavir (2.8% vs. 4.9%, p=0.01). After adjustment for traditional risk factors, HIV specific factors, and baseline brachial artery diameter, current abacavir use was independently associated with lower FMD (p=0.017). Duration of therapy and CD4 count were not associated with reduced FMD. Conclusions Endothelial function, a central mechanism in atherosclerosis and a marker of cardiovascular risk, is impaired among antiretroviral-treated patients with undetectable viral loads. Current use of abacavir was independently associated with impaired endothelial function. This finding suggests that abnormal endothelial function may underlie the clinically observed increased risk in myocardial infarction among abacavir-treated patients. PMID:19542863

Preconditioning with Endoplasmic Reticulum Stress Ameliorates Endothelial Cell Inflammation

PubMed Central

Leonard, Antony; Paton, Adrienne W.; El-Quadi, Monaliza; Paton, James C.; Fazal, Fabeha

2014-01-01

Endoplasmic Reticulum (ER) stress, caused by disturbance in ER homeostasis, has been implicated in several pathological conditions such as ischemic injury, neurodegenerative disorders, metabolic diseases and more recently in inflammatory conditions. Our present study aims at understanding the role of ER stress in endothelial cell (EC) inflammation, a critical event in the pathogenesis of acute lung injury (ALI). We found that preconditioning human pulmonary artery endothelial cells (HPAEC) to ER stress either by depleting ER chaperone and signaling regulator BiP using siRNA, or specifically cleaving (inactivating) BiP using subtilase cytotoxin (SubAB), alleviates EC inflammation. The two approaches adopted to abrogate BiP function induced ATF4 protein expression and the phosphorylation of eIF2α, both markers of ER stress, which in turn resulted in blunting the activation of NF-κB, and restoring endothelial barrier integrity. Pretreatment of HPAEC with BiP siRNA inhibited thrombin-induced IκBα degradation and its resulting downstream signaling pathway involving NF-κB nuclear translocation, DNA binding, phosphorylation at serine536, transcriptional activation and subsequent expression of adhesion molecules. However, TNFα-mediated NF-κB signaling was unaffected upon BiP knockdown. In an alternative approach, SubAB-mediated inactivation of NF-κB was independent of IκBα degradation. Mechanistic analysis revealed that pretreatment of EC with SubAB interfered with the binding of the liberated NF-κB to the DNA, thereby resulting in reduced expression of adhesion molecules, cytokines and chemokines. In addition, both knockdown and inactivation of BiP stimulated actin cytoskeletal reorganization resulting in restoration of endothelial permeability. Together our studies indicate that BiP plays a central role in EC inflammation and injury via its action on NF-κB activation and regulation of vascular permeability. PMID:25356743

Preconditioning with endoplasmic reticulum stress ameliorates endothelial cell inflammation.

PubMed

Leonard, Antony; Paton, Adrienne W; El-Quadi, Monaliza; Paton, James C; Fazal, Fabeha

2014-01-01

Endoplasmic Reticulum (ER) stress, caused by disturbance in ER homeostasis, has been implicated in several pathological conditions such as ischemic injury, neurodegenerative disorders, metabolic diseases and more recently in inflammatory conditions. Our present study aims at understanding the role of ER stress in endothelial cell (EC) inflammation, a critical event in the pathogenesis of acute lung injury (ALI). We found that preconditioning human pulmonary artery endothelial cells (HPAEC) to ER stress either by depleting ER chaperone and signaling regulator BiP using siRNA, or specifically cleaving (inactivating) BiP using subtilase cytotoxin (SubAB), alleviates EC inflammation. The two approaches adopted to abrogate BiP function induced ATF4 protein expression and the phosphorylation of eIF2α, both markers of ER stress, which in turn resulted in blunting the activation of NF-κB, and restoring endothelial barrier integrity. Pretreatment of HPAEC with BiP siRNA inhibited thrombin-induced IκBα degradation and its resulting downstream signaling pathway involving NF-κB nuclear translocation, DNA binding, phosphorylation at serine536, transcriptional activation and subsequent expression of adhesion molecules. However, TNFα-mediated NF-κB signaling was unaffected upon BiP knockdown. In an alternative approach, SubAB-mediated inactivation of NF-κB was independent of IκBα degradation. Mechanistic analysis revealed that pretreatment of EC with SubAB interfered with the binding of the liberated NF-κB to the DNA, thereby resulting in reduced expression of adhesion molecules, cytokines and chemokines. In addition, both knockdown and inactivation of BiP stimulated actin cytoskeletal reorganization resulting in restoration of endothelial permeability. Together our studies indicate that BiP plays a central role in EC inflammation and injury via its action on NF-κB activation and regulation of vascular permeability.

Breast Angiosarcoma: Case Series and Expression of Vascular Endothelial Growth Factor

PubMed Central

Brar, Rondeep; West, Robert; Witten, Daniela; Raman, Bhargav; Jacobs, Charlotte; Ganjoo, Kristen

2009-01-01

Purpose Angiosarcoma of the breast is a rare, malignant tumor for which little is known regarding prognostic indicators and optimal therapeutic regimens. To address this issue, we performed a retrospective analysis of breast angiosarcoma cases seen at Stanford University along with immunohistochemical analysis for markers of angiogenesis. Methods Breast angiosarcoma cases seen between 1980 and 2008 were examined. Viable tissue blocks were analyzed for expression of vascular endothelial growth factor and its receptors. Results A total of 16 cases were identified. Data was collected regarding epidemiology, treatment, response rates, disease-free survival, and the use of various imaging modalities. Five tissue blocks remained viable for immunohistochemical analysis. Vascular endothelial growth factor-A was positively expressed in 3 of these samples. Conclusion Angiosarcoma of the breast is an aggressive malignancy with a propensity for both local recurrence and distant metastases. Angiogenesis inhibition may represent a novel therapeutic modality in this rare, vascular malignancy. PMID:20737044

Kisspeptin-10 induces endothelial cellular senescence and impaired endothelial cell growth.

PubMed

Usui, Sayaka; Iso, Yoshitaka; Sasai, Masahiro; Mizukami, Takuya; Mori, Hiroyoshi; Watanabe, Takuya; Shioda, Seiji; Suzuki, Hiroshi

2014-07-01

The KPs (kisspeptins) are a family of multifunctional peptides with established roles in cancer metastasis, puberty and vasoconstriction. The effects of KPs on endothelial cells have yet to be determined. The aim of the present study was to investigate the effects of KP-10 on endothelial cell growth and the mechanisms underlying those effects. The administration of recombinant KP-10 into the hindlimbs of rats with ischaemia significantly impaired blood flow recovery, as shown by laser Doppler, and capillary growth, as shown using histology, compared with the controls. HUVECs (human umbilical vein endothelial cells) express the KP receptor and were treated with KP-10 in culture studies. KP-10 inhibited endothelial cell tube formation and proliferation in a significant and dose-dependent manner. The HUVECs treated with KP exhibited the senescent phenotype, as determined using a senescence-associated β-galactosidase assay, cell morphology analysis, and decreased Sirt1 (sirtuin 1) expression and increased p53 expression shown by Western blot analysis. Intriguingly, a pharmacological Rho kinase inhibitor, Y-27632, was found to increase the proliferation of HUVECs and to reduce the number of senescent phenotype cells affected by KP-10. In conclusion, KP-10 suppressed endothelial cells growth both in vivo and in vitro in the present study. The adverse effect of KP on endothelial cells was attributable, at least in part, to the induction of cellular senescence.

Lymphatic Endothelial Cells Produce M-CSF, Causing Massive Bone Loss in Mice.

PubMed

Wang, Wensheng; Wang, Hua; Zhou, Xichao; Li, Xing; Sun, Wen; Dellinger, Michael; Boyce, Brendan F; Xing, Lianping

2017-05-01

Gorham-Stout disease (GSD) is a rare bone disorder characterized by aggressive osteolysis associated with lymphatic vessel invasion within bone marrow cavities. The etiology of GSD is not known, and there is no effective therapy or animal model for the disease. Here, we investigated if lymphatic endothelial cells (LECs) affect osteoclasts (OCs) to cause a GSD osteolytic phenotype in mice. We examined the effect of a mouse LEC line on osteoclastogenesis in co-cultures. LECs significantly increased receptor activator of NF-κB ligand (RANKL)-mediated OC formation and bone resorption. LECs expressed high levels of macrophage colony-stimulating factor (M-CSF), but not RANKL, interleukin-6 (IL-6), and tumor necrosis factor (TNF). LEC-mediated OC formation and bone resorption were blocked by an M-CSF neutralizing antibody or Ki20227, an inhibitor of the M-CSF receptor, c-Fms. We injected LECs into the tibias of wild-type (WT) mice and observed massive osteolysis on X-ray and micro-CT scans. Histology showed that LEC-injected tibias had significant trabecular and cortical bone loss and increased OC numbers. M-CSF protein levels were significantly higher in serum and bone marrow plasma of mice given intra-tibial LEC injections. Immunofluorescence staining showed extensive replacement of bone and marrow by podoplanin+ LECs. Treatment of LEC-injected mice with Ki20227 significantly decreased tibial bone destruction. In addition, lymphatic vessels in a GSD bone sample were stained positively for M-CSF. Thus, LECs cause bone destruction in vivo in mice by secreting M-CSF, which promotes OC formation and activation. Blocking M-CSF signaling may represent a new therapeutic approach for treatment of patients with GSD. Furthermore, tibial injection of LECs is a useful mouse model to study GSD. © 2017 American Society for Bone and Mineral Research. © 2017 American Society for Bone and Mineral Research.

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

By Different Cellular Mechanisms, Lymphatic Vessels Sprout by Endothelial Cell Recruitment Whereas Blood Vessels Grow by Vascular Expansion

NASA Technical Reports Server (NTRS)

Parsons-Wingerter, Patricia; McKay, Terri L.; Leontiev, Dmitry; Condrich, Terence K.; DiCorleto, Paul E.

2005-01-01

The development of effective vascular therapies requires the understanding of all modes of vessel formation contributing to vasculogenesis, angiogenesis (here termed hemangiogenesis) and lymphangiogenesis. We show that lymphangiogenesis proceeds by blind-ended vessel sprouting via recruitment of isolated endothelial progenitor cells to the tips of growing vessels, whereas hemangiogenesis occurs by non-sprouting vessel expansion from the capillary network, during middevelopment in the quail chorioallantoic membrane (CAM). Blood vessels expanded out of capillaries that displayed transient expression of alpha smooth muscle actin (alphaSMA), accompanied by mural recruitment of migratory progenitor cells expressing SMA. Lymphatics and blood vessels were identified by confocal/fluorescence microscopy of vascular endothelial growth factor (VEGF) receptors VEGFR-1 and VEGFR-2, alphaSMA (expressed on CAM blood vessels but not on lymphatics), homeobox transcription factor Prox-1 (specific to CAM lymphatic endothelium), and the quail hematopoetic/vascular marker, QH-1. Expression of VEGFR-1 was highly restricted to blood vessels (primarily capillaries). VEGFR-2 was expressed intensely in isolated hematopoietic cells, lymphatic vessels and moderately in blood vessels. Prox-1 was absent from endothelial progenitor cells prior to lymphatic recruitment. Although vascular endothelial growth factor-165 (VEGF(sub 165)) is a key regulator of numerous cellular processes in hemangiogenesis and vasculogenesis, the role of VEGF(sub 165) in lymphangiogenesis is less clear. Exogenous VEGF(sub 165) increased blood vessel density without changing endogenous modes of vascular/lymphatic vessel formation or marker expression patterns. However, VEGF(sub 165) did increase the frequency of blood vascular anastomoses and strongly induced the antimaturational dissociation of lymphatics from blood vessels, with frequent formation of homogeneous lymphatic networks.

Effectiveness of Vascular Markers (Immunohistochemical Stains) in Soft Tissue Sarcomas.

PubMed

Naeem, Namra; Mushtaq, Sajid; Akhter, Noreen; Hussain, Mudassar; Hassan, Usman

2018-05-01

To ascertain the effectiveness of IHC markers of vascular origin like CD31, CD34, FLI1 and ERG in vascular soft tissue sarcomas including angiosarcomas, Kaposi sarcomas, epithelioid hemangioendothelioma and a non-vascular soft tissue sarcoma (Epithelioid sarcoma). Descriptive study. Shaukat Khanum Memorial Cancer Hospital and Research Centre, Lahore, from 2011 to 2017. Diagnosed cases of angiosarcomas (n=48), epithelioid hemangioendothelioma (n=9), Kaposi sarcoma (n=9) and epithelioid sarcoma (n=20) were selected. Immunohistochemical staining as performed on formalin fixed paraffin embedded sections. The sections were stained for the following markers: CD34 (VENTANA clone Q Bend 10), CD31 (Leica clone 1 A 10), FLI1 (CELL MARQUE clone MRQ-1) and ERG (CELL MARQUE clone EP111). A complete panel of CD34, CD31 and ERG was applied on 8/48 cases of angiosarcomas with triple positivity in 6 cases. Eight cases showed positivity for only CD31 and ERG and 2 cases showed positivity for only ERG. A complete panel of CD34, CD31 and ERG was applied on 3/9 cases of epithelioid hemangioendothelioma with positivity for all markers in 2 cases. Combined positivity for ERG and CD34 was seen in 2 cases and on 4 cases only CD31 immunohistochemical was solely applied with 100% positivity. FLI1 was not applied on any case. Among 9 cases of Kaposi sarcoma, ERG, CD34 and CD31 in combination were applied on only 1 case with triple positivity. Remaining cases show positivity for either CD34, CD31 or FLI1. Majority of cases of epithelioid sarcomas were diagnosed on the basis of cytokeratin and CD34 positivity with loss of INI1. The other vascular markers showed negativity in all cases. Among these four markers, ERG immunohistochemical stain is highly effective for endothelial differentiation due to its specific nuclear staining pattern in normal blood vessel endothelial cells (internal control) as well as neoplastic cells of vascular tumors and lack of background staining.

Enhanced autophagy in pulmonary endothelial cells on exposure to HIV-Tat and morphine: Role in HIV-related pulmonary arterial hypertension

PubMed Central

Dalvi, Pranjali; Sharma, Himanshu; Chinnappan, Mahendran; Sanderson, Miles; Allen, Julie; Zeng, Ruoxi; Choi, Augustine; O'Brien-Ladner, Amy; Dhillon, Navneet K.

2016-01-01

ABSTRACT Intravenous drug use is one of the major risk factors for HIV-infection in HIV-related pulmonary arterial hypertension patients. We previously demonstrated exaggerated pulmonary vascular remodeling with enhanced apoptosis followed by increased proliferation of pulmonary endothelial cells on simultaneous exposure to both opioids and HIV protein(s). Here we hypothesize that the exacerbation of autophagy may be involved in the switching of endothelial cells from an early apoptotic state to later hyper-proliferative state. Treatment of human pulmonary microvascular endothelial cells (HPMECs) with both the HIV-protein Tat and morphine resulted in an oxidative stress-dependent increase in the expression of various markers of autophagy and formation of autophagosomes when compared to either Tat or morphine monotreatments as demonstrated by western blot, transmission electron microscopy and immunofluorescence. Autophagy flux experiments suggested increased formation rather than decreased clearance of autolysosomes. Inhibition of autophagy resulted in a significant increase in apoptosis and reduction in proliferation of HPMECs with combined morphine and Tat (M+T) treatment compared to monotreatments whereas stimulation of autophagy resulted in opposite effects. Significant increases in the expression of autophagy markers as well as the number of autophagosomes and autolysosomes was observed in the lungs of SIV-infected macaques and HIV-infected humans exposed to opioids. Overall our findings indicate that morphine in combination with viral protein(s) results in the induction of autophagy in pulmonary endothelial cells that may lead to an increase in severity of angio-proliferative remodeling of the pulmonary vasculature on simian and human immunodeficiency virus infection in the presence of opioids. PMID:27723373

Enhanced autophagy in pulmonary endothelial cells on exposure to HIV-Tat and morphine: Role in HIV-related pulmonary arterial hypertension.

PubMed

Dalvi, Pranjali; Sharma, Himanshu; Chinnappan, Mahendran; Sanderson, Miles; Allen, Julie; Zeng, Ruoxi; Choi, Augustine; O'Brien-Ladner, Amy; Dhillon, Navneet K

2016-12-01

Intravenous drug use is one of the major risk factors for HIV-infection in HIV-related pulmonary arterial hypertension patients. We previously demonstrated exaggerated pulmonary vascular remodeling with enhanced apoptosis followed by increased proliferation of pulmonary endothelial cells on simultaneous exposure to both opioids and HIV protein(s). Here we hypothesize that the exacerbation of autophagy may be involved in the switching of endothelial cells from an early apoptotic state to later hyper-proliferative state. Treatment of human pulmonary microvascular endothelial cells (HPMECs) with both the HIV-protein Tat and morphine resulted in an oxidative stress-dependent increase in the expression of various markers of autophagy and formation of autophagosomes when compared to either Tat or morphine monotreatments as demonstrated by western blot, transmission electron microscopy and immunofluorescence. Autophagy flux experiments suggested increased formation rather than decreased clearance of autolysosomes. Inhibition of autophagy resulted in a significant increase in apoptosis and reduction in proliferation of HPMECs with combined morphine and Tat (M+T) treatment compared to monotreatments whereas stimulation of autophagy resulted in opposite effects. Significant increases in the expression of autophagy markers as well as the number of autophagosomes and autolysosomes was observed in the lungs of SIV-infected macaques and HIV-infected humans exposed to opioids. Overall our findings indicate that morphine in combination with viral protein(s) results in the induction of autophagy in pulmonary endothelial cells that may lead to an increase in severity of angio-proliferative remodeling of the pulmonary vasculature on simian and human immunodeficiency virus infection in the presence of opioids.

Non-invasive endothelial function assessment in patients with neurofibromatosis type 1: a cross-sectional study

PubMed Central

2013-01-01

Background Neurofibromatosis type 1 (NF1) is a multi-systemic disease caused by neurofibromin deficiency. The reduced life expectancy of patients with NF1 has been attributed to NF1-associated malignant neoplasms. However, an analysis of death certificates in the USA suggests that vascular disease could be an important cause of early death among these patients. Endothelial dysfunction (ED) is related to vasculopathy and is an early marker of subclinical atherosclerosis. Since neurofibromin has already been demonstrated to affect endothelial cell function, ED may be associated with NF1. The purpose of this study was to assess endothelial function in patients with NF1 using a non-invasive method. Methods NF1 patients and healthy control subjects, aged 18 to 35 years, were included. Subjects were excluded if they had any risk factor for vascular disease or any other condition known to affect endothelial function. Endothelial function was assessed using reactive hyperemia-peripheral arterial tone (RH-PAT) technology. ED was defined as a reactive hyperemia index (RHI) lower than 1.35. Results Four of the 29 (13.8%) NF1 patients and 1 of the 30 (3.3%) healthy volunteers had ED (p = 0.153). RHI medians and interquartile intervals were 1.8 (1.58-2.43) for the NF1 group and 2.02 (1.74 – 2.49) for the control group (p = 0.361). Conclusion The prevalence of ED was similar in NF1 patients and healthy controls. PMID:23497412

Endothelial microparticle-mediated transfer of MicroRNA-126 promotes vascular endothelial cell repair via SPRED1 and is abrogated in glucose-damaged endothelial microparticles.

PubMed

Jansen, Felix; Yang, Xiaoyan; Hoelscher, Marion; Cattelan, Arianna; Schmitz, Theresa; Proebsting, Sebastian; Wenzel, Daniela; Vosen, Sarah; Franklin, Bernardo S; Fleischmann, Bernd K; Nickenig, Georg; Werner, Nikos

2013-10-29

Repair of the endothelium after vascular injury is crucial for preserving endothelial integrity and preventing the development of vascular disease. The underlying mechanisms of endothelial cell repair are largely unknown. We sought to investigate whether endothelial microparticles (EMPs), released from apoptotic endothelial cells (ECs), influence EC repair. Systemic treatment of mice with EMPs after electric denudation of the endothelium accelerated reendothelialization in vivo. In vitro experiments revealed that EMP uptake in ECs promotes EC migration and proliferation, both critical steps in endothelial repair. To dissect the underlying mechanisms, Taqman microRNA array was performed, and microRNA (miR)-126 was identified as the predominantly expressed miR in EMPs. The following experiments demonstrated that miR-126 was transported into recipient human coronary artery endothelial cells by EMPs and functionally regulated the target protein sprouty-related, EVH1 domain-containing protein 1 (SPRED1). Knockdown of miR-126 in EMPs abrogated EMP-mediated effects on human coronary artery endothelial cell migration and proliferation in vitro and reendothelialization in vivo. Interestingly, after simulating diabetic conditions, EMPs derived from glucose-treated ECs contained significantly lower amounts of miR-126 and showed reduced endothelial repair capacity in vitro and in vivo. Finally, expression analysis of miR-126 in circulating microparticles from 176 patients with stable coronary artery disease with and without diabetes mellitus revealed a significantly reduced miR-126 expression in circulating microparticles from diabetic patients. Endothelial microparticles promote vascular endothelial repair by delivering functional miR-126 into recipient cells. In pathological hyperglycemic conditions, EMP-mediated miR-126-induced EC repair is altered.

Expression of Vascular Notch Ligand Delta-Like 4 and Inflammatory Markers in Breast Cancer

PubMed Central

Jubb, Adrian M.; Soilleux, Elizabeth J.; Turley, Helen; Steers, Graham; Parker, Andrew; Low, Irene; Blades, Jennifer; Li, Ji-Liang; Allen, Paul; Leek, Russell; Noguera-Troise, Irene; Gatter, Kevin C.; Thurston, Gavin; Harris, Adrian L.

2010-01-01

Delta-like ligand 4 (Dll4) is a Notch ligand that is predominantly expressed in the endothelium. Evidence from xenografts suggests that inhibiting Dll4 may overcome resistance to antivascular endothelial growth factor therapy. The aims of this study were to characterize the expression of Dll4 in breast cancer and assess whether it is associated with inflammatory markers and prognosis. We examined 296 breast adenocarcinomas and 38 ductal carcinoma in situ tissues that were represented in tissue microarrays. Additional whole sections representing 10 breast adenocarcinomas, 10 normal breast tissues, and 16 angiosarcomas were included. Immunohistochemistry was then performed by using validated antibodies against Dll4, CD68, CD14, Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Non-integrin (DC-SIGN), CD123, neutrophil elastase, CD31, and carbonic anhydrase 9. Dll4 was selectively expressed by intratumoral endothelial cells in 73% to 100% of breast adenocarcinomas, 18% of in situ ductal carcinomas, and all lactating breast cases, but not normal nonlactating breast. High intensity of endothelial Dll4 expression was a statistically significant adverse prognostic factor in univariate (P = 0.002 and P = 0.01) and multivariate analyses (P = 0.03 and P = 0.04) of overall survival and relapse-free survival, respectively. Among the inflammatory markers, only CD68 and DC-SIGN were significant prognostic factors in univariate (but not multivariate) analyses of overall survival (P = 0.01 and 0.002, respectively). In summary, Dll4 was expressed by endothelium associated with breast cancer cells. In these retrospective subset analyses, endothelial Dll4 expression was a statistically significant multivariate prognostic factor. PMID:20167860

N‐Acetylcysteine, a glutathione precursor, reverts vascular dysfunction and endothelial epigenetic programming in intrauterine growth restricted guinea pigs

PubMed Central

Herrera, Emilio A.; Cifuentes‐Zúñiga, Francisca; Figueroa, Esteban; Villanueva, Cristian; Hernández, Cherie; Alegría, René; Arroyo‐Jousse, Viviana; Peñaloza, Estefania; Farías, Marcelo; Uauy, Ricardo; Casanello, Paola

2016-01-01

Key points Intrauterine growth restriction (IUGR) is associated with vascular dysfunction, oxidative stress and signs of endothelial epigenetic programming of the umbilical vessels.There is no evidence that this epigenetic programming is occurring on systemic fetal arteries.In IUGR guinea pigs we studied the functional and epigenetic programming of endothelial nitric oxide synthase (eNOS) (Nos3 gene) in umbilical and systemic fetal arteries, addressing the role of oxidative stress in this process by maternal treatment with N‐acetylcysteine (NAC) during the second half of gestation.The present study suggests that IUGR endothelial cells have common molecular markers of programming in umbilical and systemic arteries. Notably, maternal treatment with NAC restores fetal growth by increasing placental efficiency and reverting the functional and epigenetic programming of eNOS in arterial endothelium in IUGR guinea pigs. Abstract In humans, intrauterine growth restriction (IUGR) is associated with vascular dysfunction, oxidative stress and signs of endothelial programming in umbilical vessels. We aimed to determine the effects of maternal antioxidant treatment with N‐acetylcysteine (NAC) on fetal endothelial function and endothelial nitric oxide synthase (eNOS) programming in IUGR guinea pigs. IUGR was induced by implanting ameroid constrictors on uterine arteries of pregnant guinea pigs at mid gestation, half of the sows receiving NAC in the drinking water (from day 34 until term). Fetal biometry and placental vascular resistance were followed by ultrasound throughout gestation. At term, umbilical arteries and fetal aortae were isolated to assess endothelial function by wire‐myography. Primary cultures of endothelial cells (ECs) from fetal aorta, femoral and umbilical arteries were used to determine eNOS mRNA levels by quantitative PCR and analyse DNA methylation in the Nos3 promoter by pyrosequencing. Doppler ultrasound measurements showed that NAC reduced

N-Acetylcysteine, a glutathione precursor, reverts vascular dysfunction and endothelial epigenetic programming in intrauterine growth restricted guinea pigs.

PubMed

Herrera, Emilio A; Cifuentes-Zúñiga, Francisca; Figueroa, Esteban; Villanueva, Cristian; Hernández, Cherie; Alegría, René; Arroyo-Jousse, Viviana; Peñaloza, Estefania; Farías, Marcelo; Uauy, Ricardo; Casanello, Paola; Krause, Bernardo J

2017-02-15

Intrauterine growth restriction (IUGR) is associated with vascular dysfunction, oxidative stress and signs of endothelial epigenetic programming of the umbilical vessels. There is no evidence that this epigenetic programming is occurring on systemic fetal arteries. In IUGR guinea pigs we studied the functional and epigenetic programming of endothelial nitric oxide synthase (eNOS) (Nos3 gene) in umbilical and systemic fetal arteries, addressing the role of oxidative stress in this process by maternal treatment with N-acetylcysteine (NAC) during the second half of gestation. The present study suggests that IUGR endothelial cells have common molecular markers of programming in umbilical and systemic arteries. Notably, maternal treatment with NAC restores fetal growth by increasing placental efficiency and reverting the functional and epigenetic programming of eNOS in arterial endothelium in IUGR guinea pigs. In humans, intrauterine growth restriction (IUGR) is associated with vascular dysfunction, oxidative stress and signs of endothelial programming in umbilical vessels. We aimed to determine the effects of maternal antioxidant treatment with N-acetylcysteine (NAC) on fetal endothelial function and endothelial nitric oxide synthase (eNOS) programming in IUGR guinea pigs. IUGR was induced by implanting ameroid constrictors on uterine arteries of pregnant guinea pigs at mid gestation, half of the sows receiving NAC in the drinking water (from day 34 until term). Fetal biometry and placental vascular resistance were followed by ultrasound throughout gestation. At term, umbilical arteries and fetal aortae were isolated to assess endothelial function by wire-myography. Primary cultures of endothelial cells (ECs) from fetal aorta, femoral and umbilical arteries were used to determine eNOS mRNA levels by quantitative PCR and analyse DNA methylation in the Nos3 promoter by pyrosequencing. Doppler ultrasound measurements showed that NAC reduced placental vascular resistance

Endothelial cell density to predict endothelial graft failure after penetrating keratoplasty.

PubMed

Lass, Jonathan H; Sugar, Alan; Benetz, Beth Ann; Beck, Roy W; Dontchev, Mariya; Gal, Robin L; Kollman, Craig; Gross, Robert; Heck, Ellen; Holland, Edward J; Mannis, Mark J; Raber, Irving; Stark, Walter; Stulting, R Doyle

2010-01-01

To determine whether preoperative and/or postoperative central endothelial cell density (ECD) and its rate of decline postoperatively are predictive of graft failure caused by endothelial decompensation following penetrating keratoplasty to treat a moderate-risk condition, principally, Fuchs dystrophy or pseudophakic corneal edema. In a subset of Cornea Donor Study participants, a central reading center determined preoperative and postoperative ECD from available specular images for 17 grafts that failed because of endothelial decompensation and 483 grafts that did not fail. Preoperative ECD was not predictive of graft failure caused by endothelial decompensation (P = .91). However, the 6-month ECD was predictive of subsequent failure (P < .001). Among those that had not failed within the first 6 months, the 5-year cumulative incidence (+/-95% confidence interval) of failure was 13% (+/-12%) for the 33 participants with a 6-month ECD of less than 1700 cells/mm(2) vs 2% (+/-3%) for the 137 participants with a 6-month ECD of 2500 cells/mm(2) or higher. After 5 years' follow-up, 40 of 277 participants (14%) with a clear graft had an ECD below 500 cells/mm(2). Preoperative ECD is unrelated to graft failure from endothelial decompensation, whereas there is a strong correlation of ECD at 6 months with graft failure from endothelial decompensation. A graft can remain clear after 5 years even when the ECD is below 500 cells/mm(2).

Assessment of vascular and endothelial dysfunction in nutritional studies.

PubMed

Ray, S; Miglio, C; Eden, T; Del Rio, D

2014-09-01

Vascular and endothelial dysfunction (VED) is emerging as a potential set of early markers of cardiovascular disease risk and tests for its measurement have been widely used in clinical research. The aim of this viewpoint is to describe and discuss the current usage of these measures in well-designed nutritional trials, using the potential relationship between fruit juice intake and VED as example. A search was conducted using the NHS evidence portal including studies published in English between January 1980 and October 2013. Only 10 suitable studies were selected, which investigated the effect of fruit juice intake on VED, among which 4 interventions used flow-mediated dilatation, 2 arterial stiffness, 2 a combination of arterial stiffness and flow-mediated dilatation, 2 carotid intimal media thickness and 1 iontophoresis with laser Doppler. Despite minimal effects reported on classical CVD markers, such as lipids, 8 out of the 10 identified studies reported an effect on endothelial function following juice consumption, indicating that VED tests can be effectively used in human dietary interventions to identify relationships between bioactive compounds from fruit and CVD risk. However, paucity of available data, scarcity of compound bioavailability and metabolism information, strong heterogeneity among experimental methodologies and a number of limitations to study designs, still limit the interpretation of the results obtained through these measures. Future, well-designed studies with greater attention to consider use of VED measures are needed to strengthen the utility of VED tests in nutrition research such as those investigating the impact of polyphenol-rich juices and CVD risk. Copyright © 2014 Elsevier B.V. All rights reserved.

Endothelial MMP14 is required for endothelial-dependent growth support of human airway basal cells

PubMed Central

Ding, Bi-Sen; Gomi, Kazunori; Rafii, Shahin; Crystal, Ronald G.; Walters, Matthew S.

2015-01-01

ABSTRACT Human airway basal cells are the stem (or progenitor) population of the airway epithelium, and play a central role in anchoring the epithelium to the basement membrane. The anatomic position of basal cells allows for potential paracrine signaling between them and the underlying non-epithelial stromal cells. In support of this, we have previously demonstrated that endothelial cells support growth of basal cells during co-culture through vascular endothelial growth factor A (VEGFA)-mediated signaling. Building on these findings, we found, by RNA sequencing analysis, that basal cells expressed multiple fibroblast growth factor (FGF) ligands (FGF2, FGF5, FGF11 and FGF13) and that only FGF2 and FGF5 were capable of functioning in a paracrine manner to activate classical FGF receptor (FGFR) signaling. Antibody-mediated blocking of FGFR1 during basal-cell–endothelial-cell co-culture significantly reduced the endothelial-cell-dependent basal cell growth. Stimulation of endothelial cells with basal-cell-derived growth factors induced endothelial cell expression of matrix metallopeptidase 14 (MMP14), and short hairpin RNA (shRNA)-mediated knockdown of endothelial cell MMP14 significantly reduced the endothelial-cell-dependent growth of basal cells. Overall, these data characterize a new growth-factor-mediated reciprocal ‘crosstalk’ between human airway basal cells and endothelial cells that regulates proliferation of basal cells. PMID:26116571

Human Herpesvirus-8-Transformed Endothelial Cells Have Functionally Activated Vascular Endothelial Growth Factor/Vascular Endothelial Growth Factor Receptor

PubMed Central

Masood, Rizwan; Cesarman, Ethel; Smith, D. Lynne; Gill, Parkash S.; Flore, Ornella

2002-01-01

Kaposi’s sarcoma is a vascular tumor commonly associated with human immunodeficiency virus (HIV)-1 and human herpesvirus (HHV-8) also known as Kaposi’s sarcoma-associated herpesvirus. The principal features of this tumor are abnormal proliferation of vascular structures lined with spindle-shaped endothelial cells. HHV-8 may transform a subpopulation of endothelial cells in vitro via viral and cellular gene expression. We hypothesized that among the cellular genes, vascular endothelial growth factors (VEGFs) and their cognate receptors may be involved in viral-mediated transformation. We have shown that HHV-8-transformed endothelial cells (EC-HHV-8) express higher levels of VEGF, VEGF-C, VEGF-D, and PlGF in addition to VEGF receptors-1, -2, and -3. Furthermore, antibodies to VEGF receptor-2 inhibited cell proliferation and viability. Similarly, inhibition of VEGF gene expression with antisense oligonucleotides inhibited EC-HHV-8 cell proliferation/viability. The growth and viability of primary endothelial cells and a fibroblast cell line however were unaffected by either the VEGF receptor-2 antibody or the VEGF antisense oligodeoxynucleotides. VEGF and VEGF receptors are thus induced in EC-HHV-8 and participate in the transformation. Inhibitors of VEGF may thus modulate the disease process during development and progression. PMID:11786394

Pulmonary endothelial pavement patterns.

PubMed Central

Kibria, G; Heath, D; Smith, P; Biggar, R

1980-01-01

The appearance of the endothelial pavement pattern was studied in the pulmonary trunk, pulmonary veins, aorta, and inferior vena cava of the rat by means of silver staining of the cell borders. The endothelial cell in each of the four blood vessels was found to have its own distinctive shape, fusiform and pointed in the direction of blood flow in the case of the aorta and larger and more rectangular in the pulmonary trunk and pulmonary veins. Detailed quantitation of the dimensions and surface area of the endothelial cells in each blood vessel was carried out by a photographic technique. Pulmonary hypertension was induced in one group of rats by feeding them on Crotalaria spectabilis seeds. The endothelial pavement pattern in their pulmonary trunks became disrupted with many of the cells assuming a fusiform shape reminiscent of aortic endothelium. Many small, new endothelial cells formed in the pulmonary trunk suggesting division of cells to line the enlarging blood vessels. In contrast the endothelial cells of the inferior vena cava merely increased in size to cope with the dilatation of this vein. Images PMID:7385090

Recombinant tissue plasminogen activator enhances microparticle release from mouse brain-derived endothelial cells through plasmin.

PubMed

Garraud, Marie; Khacef, Kahina; Vion, Anne-Clémence; Leconte, Claire; Yin, Min; Renard, Jean-Marie; Marchand-Leroux, Catherine; Boulanger, Chantal M; Margaill, Isabelle; Beray-Berthat, Virginie

2016-11-15

Thrombolysis with recombinant tissue plasminogen activator (rt-PA) is currently the only approved pharmacological strategy for acute ischemic stroke. However, rt-PA exhibits vascular toxicity mainly due to endothelial damage. To investigate the mechanisms underlying rt-PA-induced endothelial alterations, we assessed the role of rt-PA in the generation of endothelial microparticles (EMPs), emerging biological markers and effectors of endothelial dysfunction. The mouse brain-derived endothelial cell line bEnd.3 was used. Cells were treated with rt-PA at 20, 40 or 80μg/ml for 15 or 24h, and EMPs were quantified in the culture media using Annexin-V staining coupled with flow cytometry. Rt-PA enhanced EMP release from bEnd.3 cells with a maximal increase at the 40μg/ml dose for 24h (+78% compared to controls). Using tranexamic acid and aprotinin we demonstrated that plasmin is responsible for rt-PA-induced EMP release. The p38 MAPK inhibitor SB203580 and the poly(ADP-ribose)polymerase (PARP) inhibitor PJ34 also reduced rt-PA-induced EMP production, suggesting that p38 MAPK and PARP are downstream intracellular effectors of rt-PA/plasmin. Rt-PA also altered through plasmin the morphology and the confluence of bEnd.3 cells. By contrast, these changes did not implicate p38 MAPK and PARP. This study demonstrates that rt-PA induces the production of microparticles by cerebral endothelial cells, through plasmin, p38 MAPK and PARP pathways. Determining the phenotype of these EMPs to clarify their role on the endothelium in ischemic conditions could thus be of particular interest. Copyright © 2016 Elsevier B.V. All rights reserved.

Endothelial- and Platelet-Derived Microparticles Are Generated During Liver Resection in Humans.

PubMed

Banz, Yara; Item, Gian-Marco; Vogt, Andreas; Rieben, Robert; Candinas, Daniel; Beldi, Guido

2016-01-01

Cell-derived plasma microparticles (<1.5 μm) originating from various cell types have the potential to regulate thrombogenesis and inflammatory responses. The aim of this study was to test the hypothesis that microparticles generated during hepatic surgery co-regulate postoperative procoagulant and proinflammatory events. In 30 patients undergoing liver resection, plasma microparticles were isolated, quantitated, and characterized as endothelial (CD31+, CD41-), platelet (CD41+), or leukocyte (CD11b+) origin by flow cytometry and their procoagulant and proinflammatory activity was measured by immunoassays. During liver resection, the total numbers of microparticles increased with significantly more Annexin V-positive, endothelial and platelet-derived microparticles following extended hepatectomy compared to standard and minor liver resections. After liver resection, microparticle tissue factor and procoagulant activity increased along with overall coagulation as assessed by thrombelastography. Levels of leukocyte-derived microparticles specifically increased in patients with systemic inflammation as assessed by C-reactive protein but are independent of the extent of liver resection. Endothelial and platelet-derived microparticles are specifically elevated during liver resection, accompanied by increased procoagulant activity. Leukocyte-derived microparticles are a potential marker for systemic inflammation. Plasma microparticles may represent a specific response to surgical stress and may be an important mediator of postoperative coagulation and inflammation.

Reprogramming of Adult Peripheral Blood Cells into Human Induced Pluripotent Stem Cells as a Safe and Accessible Source of Endothelial Cells.

PubMed

Simara, Pavel; Tesarova, Lenka; Rehakova, Daniela; Farkas, Simon; Salingova, Barbara; Kutalkova, Katerina; Vavreckova, Eva; Matula, Pavel; Matula, Petr; Veverkova, Lenka; Koutna, Irena

2018-01-01

New approaches in regenerative medicine and vasculogenesis have generated a demand for sufficient numbers of human endothelial cells (ECs). ECs and their progenitors reside on the interior surface of blood and lymphatic vessels or circulate in peripheral blood; however, their numbers are limited, and they are difficult to expand after isolation. Recent advances in human induced pluripotent stem cell (hiPSC) research have opened possible avenues to generate unlimited numbers of ECs from easily accessible cell sources, such as the peripheral blood. In this study, we reprogrammed peripheral blood mononuclear cells, human umbilical vein endothelial cells (HUVECs), and human saphenous vein endothelial cells (HSVECs) into hiPSCs and differentiated them into ECs. The phenotype profiles, functionality, and genome stability of all hiPSC-derived ECs were assessed and compared with HUVECs and HSVECs. hiPSC-derived ECs resembled their natural EC counterparts, as shown by the expression of the endothelial surface markers CD31 and CD144 and the results of the functional analysis. Higher expression of endothelial progenitor markers CD34 and kinase insert domain receptor (KDR) was measured in hiPSC-derived ECs. An analysis of phosphorylated histone H2AX (γH2AX) foci revealed that an increased number of DNA double-strand breaks upon reprogramming into pluripotent cells. However, differentiation into ECs restored a normal number of γH2AX foci. Our hiPSCs retained a normal karyotype, with the exception of the HSVEC-derived hiPSC line, which displayed mosaicism due to a gain of chromosome 1. Peripheral blood from adult donors is a suitable source for the unlimited production of patient-specific ECs through the hiPSC interstage. hiPSC-derived ECs are fully functional and comparable to natural ECs. The protocol is eligible for clinical applications in regenerative medicine, if the genomic stability of the pluripotent cell stage is closely monitored.

Association between anthropometry, cardiometabolic risk factors, & early life factors & adult measures of endothelial function: Results from the New Delhi Birth Cohort.

PubMed

Huffman, Mark D; Khalil, Anita; Osmond, Clive; Fall, Caroline H D; Tandon, Nikhil; Lakshmy, Ramakrishnan; Ramji, Siddharth; Gera, Tarun; Prabhakaran, Poornima; Dey Biswas, S K; Reddy, K Srinath; Bhargava, Santosh K; Sachdev, Harshpal S; Prabhakaran, Dorairaj

2015-12-01

Abnormal endothelial function represents a preclinical marker of atherosclerosis. This study was conducted to evaluate associations between anthropometry, cardiometabolic risk factors, and early life factors and adult measures of endothelial function in a young urban Indian cohort free of clinical cardiovascular disease. Absolute changes in brachial artery diameter following cuff inflation and sublingual nitroglycerin (400 µg) were recorded to evaluate endothelium-dependent and -independent measures of endothelial function in 600 participants (362 men; 238 women) from the New Delhi Birth Cohort (2006-2009). Data on anthropometry, cardiometabolic risk factors, medical history, socio-economic position, and lifestyle habits were collected. Height and weight were recorded at birth, two and 11 yr of age. Age- and sex-adjusted linear regression models were developed to evaluate these associations. The mean age of participants was 36±1 yr. Twenty two per cent men and 29 per cent women were obese (BMI th > 30 kg/m [2] ). Mean systolic blood pressure (SBP) was 131±14 and 119±13 mmHg, and diabetes prevalence was 12 and 8 per cent for men and women, respectively. Brachial artery diameter was higher for men compared with women both before (3.48±0.37 and 2.95±0.35 cm) and after hyperaemia (3.87±0.37 vs. 3.37±0.35 cm). A similar difference was seen before and after nitroglycerin. Markers of increased adiposity, smoking, SBP, and metabolic syndrome, but not early life anthropometry, were inversely associated with endothelial function after adjustment for age and sex. The analysis of the current prospective data from a young urban Indian cohort showed that cardiometabolic risk factors, but not early life anthropometry, were associated with worse endothelial function.

Association between anthropometry, cardiometabolic risk factors, & early life factors & adult measures of endothelial function: Results from the New Delhi Birth Cohort

PubMed Central

Huffman, Mark D.; Khalil, Anita; Osmond, Clive; Fall, Caroline H. D.; Tandon, Nikhil; Lakshmy, Ramakrishnan; Ramji, Siddharth; Gera, Tarun; Prabhakaran, Poornima; Dey Biswas, S. K.; Reddy, K. Srinath; Bhargava, Santosh K.; Sachdev, Harshpal S.; Prabhakaran, Dorairaj

2015-01-01

Background & objectives: Abnormal endothelial function represents a preclinical marker of atherosclerosis. This study was conducted to evaluate associations between anthropometry, cardiometabolic risk factors, and early life factors and adult measures of endothelial function in a young urban Indian cohort free of clinical cardiovascular disease. Methods: Absolute changes in brachial artery diameter following cuff inflation and sublingual nitroglycerin (400 µg) were recorded to evaluate endothelium-dependent and -independent measures of endothelial function in 600 participants (362 men; 238 women) from the New Delhi Birth Cohort (2006-2009). Data on anthropometry, cardiometabolic risk factors, medical history, socio-economic position, and lifestyle habits were collected. Height and weight were recorded at birth, two and 11 yr of age. Age- and sex-adjusted linear regression models were developed to evaluate these associations. Results: The mean age of participants was 36±1 yr. Twenty two per cent men and 29 per cent women were obese (BMI > 30 kg/m2). Mean systolic blood pressure (SBP) was 131±14 and 119±13 mmHg, and diabetes prevalence was 12 and 8 per cent for men and women, respectively. Brachial artery diameter was higher for men compared with women both before (3.48±0.37 and 2.95±0.35 cm) and after hyperaemia (3.87±0.37 vs. 3.37±0.35 cm). A similar difference was seen before and after nitroglycerin. Markers of increased adiposity, smoking, SBP, and metabolic syndrome, but not early life anthropometry, were inversely associated with endothelial function after adjustment for age and sex. Interpretation & conclusions: The analysis of the current prospective data from a young urban Indian cohort showed that cardiometabolic risk factors, but not early life anthropometry, were associated with worse endothelial function. PMID:26831418

Observations on the microvasculature of bone defects filled with biodegradable nanoparticulate hydroxyapatite.

PubMed

Kilian, Olaf; Wenisch, Sabine; Karnati, Srikanth; Baumgart-Vogt, Eveline; Hild, Anne; Fuhrmann, Rosemarie; Jonuleit, Tarja; Dingeldein, Elvira; Schnettler, Reinhard; Franke, Ralf-Peter

2008-01-01

The microvascularization of metaphyseal bone defects filled with nanoparticulate, biodegradable hydroxyapatite biomaterial with and without platelet factors enrichment was investigated in a minipig model. Results from morphological analysis and PECAM-1 immunohistochemistry showed the formation of new blood vessels into the bone defects by sprouting and intussusception of pre-existing ones. However, no significant differences were observed in the microvascularization of the different biomaterials applied (pure versus platelet factors-enriched hydroxyapatite), concerning the number of vessels and their morphological structure at day 20 after operation. The appearance of VEGFR-2 positive endothelial progenitor cells in the connective tissue between hydroxyapatite particles was also found to be independent from platelet factors enrichment of the hydroxyapatite bone substitute. In both groups formation of lymphatic vessels was detected with a podoplanin antibody. No differences were noted between HA/PLF- and HA/PLF+ implants with respect to the podoplanin expression level, the staining pattern or number of lymphatic vessels. In conclusion, the present study demonstrates different mechanisms of blood and lymphatic vessel formation in hydroxyapatite implants in minipigs.

Suppression of transient receptor potential melastatin 4 expression promotes conversion of endothelial cells into fibroblasts via transforming growth factor/activin receptor-like kinase 5 pathway.

PubMed

Echeverría, Cesar; Montorfano, Ignacio; Cabello-Verrugio, Claudio; Armisén, Ricardo; Varela, Diego; Simon, Felipe

2015-05-01

To study whether transient receptor potential melastatin 4 (TRPM4) participates in endothelial fibrosis and to investigate the underlying mechanism. Primary human endothelial cells were used and pharmacological and short interfering RNA-based approaches were used to test the transforming growth factor beta (TGF-β)/activin receptor-like kinase 5 (ALK5) pathway participation and contribution of TRPM7 ion channel. Suppression of TRPM4 expression leads to decreased endothelial protein expression and increased expression of fibrotic and extracellular matrix markers. Furthermore, TRPM4 downregulation increases intracellular Ca levels as a potential condition for fibrosis. The underlying mechanism of endothelial fibrosis shows that inhibition of TRPM4 expression induces TGF-β1 and TGF-β2 expression, which act through their receptor, ALK5, and the nuclear translocation of the profibrotic transcription factor smad4. TRPM4 acts to maintain endothelial features and its loss promotes fibrotic conversion via TGF-β production. The regulation of TRPM4 levels could be a target for preserving endothelial function during inflammatory diseases.

Omega-3 Fatty Acid Supplementation Improves Endothelial Function in Primary Antiphospholipid Syndrome: A Small-Scale Randomized Double-Blind Placebo-Controlled Trial.

PubMed

Felau, Sheylla M; Sales, Lucas P; Solis, Marina Y; Hayashi, Ana Paula; Roschel, Hamilton; Sá-Pinto, Ana Lúcia; Andrade, Danieli Castro Oliveira De; Katayama, Keyla Y; Irigoyen, Maria Claudia; Consolim-Colombo, Fernanda; Bonfa, Eloisa; Gualano, Bruno; Benatti, Fabiana B

2018-01-01

Endothelial cells are thought to play a central role in the pathogenesis of antiphospholipid syndrome (APS). Omega-3 polyunsaturated fatty acid (n-3 PUFA) supplementation has been shown to improve endothelial function in a number of diseases; thus, it could be of high clinical relevance in APS. The aim of this study was to evaluate the efficacy of n-3 PUFA supplementation on endothelial function (primary outcome) of patients with primary APS (PAPS). A 16-week randomized clinical trial was conducted with 22 adult women with PAPS. Patients were randomly assigned (1:1) to receive placebo (PL, n  = 11) or n-3 PUFA (ω-3, n  = 11) supplementation. Before (pre) and after (post) 16 weeks of the intervention, patients were assessed for endothelial function (peripheral artery tonometry) (primary outcome). Patients were also assessed for systemic markers of endothelial cell activation, inflammatory markers, dietary intake, international normalized ratio (INR), and adverse effects. At post, ω-3 group presented significant increases in endothelial function estimates reactive hyperemia index (RHI) and logarithmic transformation of RHI (LnRHI) when compared with PL (+13 vs. -12%, p  = 0.06, ES = 0.9; and +23 vs. -22%, p  = 0.02, ES = 1.0). No changes were observed for e-selectin, vascular adhesion molecule-1, and fibrinogen levels ( p  > 0.05). In addition, ω-3 group showed decreased circulating levels of interleukin-10 (-4 vs. +45%, p  = 0.04, ES = -0.9) and tumor necrosis factor (-13 vs. +0.3%, p  = 0.04, ES = -0.95) and a tendency toward a lower intercellular adhesion molecule-1 response (+3 vs. +48%, p  = 0.1, ES = -0.7) at post when compared with PL. No changes in dietary intake, INR, or self-reported adverse effects were observed. In conclusion, 16 weeks of n-3 PUFA supplementation improved endothelial function in patients with well-controlled PAPS. These results support a role of n-3 PUFA supplementation as an

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.

Cell Homogeneity Indispensable for Regenerative Medicine by Cultured Human Corneal Endothelial Cells.

PubMed

Hamuro, Junji; Toda, Munetoyo; Asada, Kazuko; Hiraga, Asako; Schlötzer-Schrehardt, Ursula; Montoya, Monty; Sotozono, Chie; Ueno, Morio; Kinoshita, Shigeru

2016-09-01

To identify the subpopulation (SP) among heterogeneous cultured human corneal endothelial cells (cHCECs) devoid of cell-state transition applicable for cell-based therapy. Subpopulation presence in cHCECs was confirmed via surface CD-marker expression level by flow cytometry. CD markers effective for distinguishing distinct SPs were selected by analyzing those on established cHCECs with a small cell area and high cell density. Contrasting features among three typical cHCEC SPs was confirmed by PCR array for extracellular matrix (ECM). Combined analysis of CD markers was performed to identify the SP (effector cells) applicable for therapy. ZO-1 and Na+/K+ ATPase, CD200, and HLA expression were compared among heterogeneous SPs. Flow cytometry analysis identified the effector cell expressing CD166+CD105-CD44-∼+/-CD26-CD24-, but CD200-, and the presence of other SPs with CD166+ CD105-CD44+++ (CD26 and CD24, either + or -) was confirmed. PCR array revealed three distinct ECM expression profiles. Some SPs expressed ZO-1 and Na+/K+ ATPase at comparable levels with effector cells, while only one SP expressed CD200, but not on effector cells. Human leukocyte antigen expression was most reduced in the effector SP. The proportion of effector cells (E-ratio) inversely paralleled donor age and decreased during prolonged culture passages. The presence of Rho-associated protein kinase (ROCK) inhibitor increased the E-ratio in cHCECs. The average area of effector cells was approximately 200∼220 μm2, and the density of cHCECs exceeded 2500 cells/mm2. A specified cultured effector cell population sharing the surface phenotypes with mature HCECs in corneal tissues may serve as an alternative to donor corneas for the treatment of corneal endothelial dysfunction.

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

Curcumin modulates endothelial permeability and monocyte transendothelial migration by affecting endothelial cell dynamics.

PubMed

Monfoulet, Laurent-Emmanuel; Mercier, Sylvie; Bayle, Dominique; Tamaian, Radu; Barber-Chamoux, Nicolas; Morand, Christine; Milenkovic, Dragan

2017-11-01

Curcumin is a phenolic compound that exhibits beneficial properties for cardiometabolic health. We previously showed that curcumin reduced the infiltration of immune cells into the vascular wall and prevented atherosclerosis development in mice. This study aimed to investigate the effect of curcumin on monocyte adhesion and transendothelial migration (TEM) and to decipher the underlying mechanisms of these actions. Human umbilical vein endothelial cells (HUVECs) were exposed to curcumin (0.5-1μM) for 3h prior to their activation by Tumor Necrosis Factor alpha (TNF-α). Endothelial permeability, monocyte adhesion and transendothelial migration assays were conducted under static condition and shear stress that mimics blood flow. We further investigated the impact of curcumin on signaling pathways and on the expression of genes using macroarrays. Pre-exposure of endothelial cells to curcumin reduced monocyte adhesion and their transendothelial migration in both static and shear stress conditions. Curcumin also prevented changes in both endothelial permeability and the area of HUVECs when induced by TNF-α. We showed that curcumin modulated the expression of 15 genes involved in the control of cytoskeleton and endothelial junction dynamic. Finally, we showed that curcumin inhibited NF-κB signaling likely through an antagonist interplay with several kinases as suggested by molecular docking analysis. Our findings demonstrate the ability of curcumin to reduce monocyte TEM through a multimodal regulation of the endothelial cell dynamics with a potential benefit on the vascular endothelial function barrier. Copyright © 2017 Elsevier Inc. All rights reserved.

Edaravone Protects against Methylglyoxal-Induced Barrier Damage in Human Brain Endothelial Cells

PubMed Central

Tóth, Andrea E.; Walter, Fruzsina R.; Bocsik, Alexandra; Sántha, Petra; Veszelka, Szilvia; Nagy, Lajos; Puskás, László G.; Couraud, Pierre-Olivier; Takata, Fuyuko; Dohgu, Shinya; Kataoka, Yasufumi; Deli, Mária A.

2014-01-01

Background Elevated level of reactive carbonyl species, such as methylglyoxal, triggers carbonyl stress and activates a series of inflammatory responses leading to accelerated vascular damage. Edaravone is the active substance of a Japanese medicine, which aids neurological recovery following acute brain ischemia and subsequent cerebral infarction. Our aim was to test whether edaravone can exert a protective effect on the barrier properties of human brain endothelial cells (hCMEC/D3 cell line) treated with methylglyoxal. Methodology Cell viability was monitored in real-time by impedance-based cell electronic sensing. The barrier function of the monolayer was characterized by measurement of resistance and flux of permeability markers, and visualized by immunohistochemistry for claudin-5 and β-catenin. Cell morphology was also examined by holographic phase imaging. Principal Findings Methylglyoxal exerted a time- and dose-dependent toxicity on cultured human brain endothelial cells: a concentration of 600 µM resulted in about 50% toxicity, significantly reduced the integrity and increased the permeability of the barrier. The cell morphology also changed dramatically: the area of cells decreased, their optical height significantly increased. Edaravone (3 mM) provided a complete protection against the toxic effect of methylglyoxal. Co-administration of edaravone restored cell viability, barrier integrity and functions of brain endothelial cells. Similar protection was obtained by the well-known antiglycating molecule, aminoguanidine, our reference compound. Conclusion These results indicate for the first time that edaravone is protective in carbonyl stress induced barrier damage. Our data may contribute to the development of compounds to treat brain endothelial dysfunction in carbonyl stress related diseases. PMID:25033388

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.

Is reversal of endothelial dysfunction by tea related to flavonoid metabolism?

PubMed

Hodgson, Jonathan M; Puddey, Ian B; Burke, Valerie; Croft, Kevin D

2006-01-01

Dietary flavonoids can improve endothelial function, but the response varies between individuals. Wide variability is also seen in flavonoid O-methylation, a major pathway of flavonoid metabolism. The O-methylation of flavonoids could alter activity, and thus influence any effect on endothelial function. The objective of the current analysis was to investigate whether variability in the endothelial function response to ingestion of tea, a rich source of flavonoids, is related to the degree of O-methylation of flavonoids. This relationship was investigated in two studies in which endothelium-dependent flow-mediated dilatation (FMD) of the brachial artery was assessed and urinary 4-O-methylgallic acid (4OMGA) excretion was used as a marker of the O-methylation of tea-derived flavonoids. In the first study, amongst participants consuming five cups of tea per day for 4 weeks, the degree of increase in 4OMGA excretion was inversely associated with the change in FMD responses (r -078, P=0.008). In the second study, there was a significant difference in the FMD responses to acute ingestion of three cups of tea between individuals with a low (median) 4OMGA response (1.94 (sem 0.79) % and -0.25 (sem 0.53) %, respectively; P=0.03). That is, any improvement in FMD following ingestion of tea may be enhanced in individuals who O-methylate less of the absorbed flavonoids. The present results are consistent with the suggestion that differences in flavonoid metabolism may influence their effect on endothelial function. Thus, differences in flavonoid metabolism could be related to the level of benefit of dietary flavonoids on the risk of CVD.

Endothelial progenitor cells (EPCs) in ageing and age-related diseases: How currently available treatment modalities affect EPC biology, atherosclerosis, and cardiovascular outcomes.

PubMed

Altabas, Velimir; Altabas, Karmela; Kirigin, Lora

2016-10-01

Endothelial progenitor cells (EPCs) are mononuclear cells that circulate in the blood and are derived from different tissues, expressing cell surface markers that are similar to mature endothelial cells. The discovery of EPCs has lead to new insights in vascular repair and atherosclerosis and also a new theory for ageing. EPCs from the bone marrow and some other organs aid in vascular repair by migrating to distant vessels where they differentiate into mature endothelial cells and replace old and injured endothelial cells. The ability of EPCs to repair vascular damage depends on their number and functionality. Currently marketed drugs used in a variety of diseases can modulate these characteristics. In this review, the effect of currently available treatment options for cardiovascular and metabolic disorders on EPC biology will be discussed. The various EPC-based therapies that will be discussed include lipid-lowering agents, antihypertensive agents, antidiabetic drugs, phosphodiesteraze inhibitors, hormones, as well as EPC capturing stents. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

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 OSA on hypoxic and inflammatory markers during CPAP withdrawal: Further evidence from three randomized control trials.

PubMed

Turnbull, Chris D; Rossi, Valentina A; Santer, Peter; Schwarz, Esther I; Stradling, John R; Petousi, Nayia; Kohler, Malcolm

2017-05-01

Obstructive sleep apnoea (OSA) is associated with cardiovascular disease. Intermittent hypoxia, endothelial dysfunction and adipose tissue-mediated inflammation have all been linked to cardiovascular disease in OSA. We therefore explored the effect of OSA on relevant associated blood markers: adrenomedullin (ADM), endocan, endothelin-1 (ET-1), resistin and vascular endothelial growth factor (VEGF). Patients with OSA, established on and compliant with continuous positive airways pressure (CPAP) therapy for >1 year were included from three randomized controlled trials, conducted at two centres. Patients were randomized to either continued therapeutic CPAP or sham CPAP (CPAP withdrawal) for 2 weeks. Blood markers were measured at baseline and at 14 days and the treatment effect between sham CPAP and therapeutic CPAP was analysed. A total of 109 patients were studied (therapeutic CPAP n = 54, sham CPAP n = 55). Sham CPAP was associated with a return of OSA (between-group difference in oxygen desaturation index (ODI) 36.0/h, 95% CI 29.9-42.2, P < 0.001). Sham CPAP was associated with a reduction in ADM levels at 14 days (-26.0 pg/mL, 95% CI -47.8 to -4.3, P = 0.02), compared to therapeutic CPAP. Return of OSA was not associated with changes in endocan, ET-1, resistin or VEGF. Whilst CPAP withdrawal was associated with return of OSA, it was associated with an unexpected significant reduction in the vasodilator ADM and not with expected increases in hypoxia-induced markers, markers of endothelial function or resistin. We propose that the vascular effects occurring in OSA may be brought about by other mechanisms, perhaps partly through a reduction in ADM. © 2016 Asian Pacific Society of Respirology.

Implantation of healthy matrix-embedded endothelial cells rescues dysfunctional endothelium and ischemic tissue in liver engraftment

PubMed Central

Melgar-Lesmes, Pedro; Balcells, Mercedes; Edelman, Elazer R.

2017-01-01

Objective Liver transplantation is limited by ischemic injury which promotes endothelial cell and hepatocyte dysfunction and eventually organ failure. We sought to understand how endothelial state determines liver recover after hepatectomy and engraftment. Design Matrix-embedded endothelial cells (MEECs) with retained healthy phenotype or control acellular matrices were implanted in direct contact with the remaining median lobe of donor mice undergoing partial hepatectomy (70%), or in the interface between the remaining median lobe and an autograft or isograft from the left lobe in hepatectomized recipient mice. Hepatic vascular architecture, DNA fragmentation and apoptosis in the median lobe and grafts, serum markers of liver damage and phenotype of macrophage and lymphocyte subsets in the liver after engraftment were analyzed 7 days post-op. Results Healthy MEECs create a functional vascular splice in donor and recipient liver after 70% hepatectomy in mouse protecting these livers from ischemic injury, hepatic congestion and inflammation. Macrophages recruited adjacent to the vascular nodes into the implants switched to an anti-inflammatory and regenerative profile M2. MEECs improved liver function and the rate of liver regeneration and prevented apoptosis in donor liver lobes, autologous grafts, and allogeneic engraftment. Conclusions Implants with healthy endothelial cells rescue liver donor and recipient endothelium and parenchyma from ischemic injury after major hepatectomy and engraftment. This study highlights endothelial-hepatocyte crosstalk in hepatic repair and provides a promising new approach to improve regenerative medicine outcomes and liver transplantation. PMID:26851165

Impaired Endothelial Repair Capacity of Early Endothelial Progenitor Cells in Hypertensive Patients With Primary Hyperaldosteronemia: Role of 5,6,7,8-Tetrahydrobiopterin Oxidation and Endothelial Nitric Oxide Synthase Uncoupling.

PubMed

Chen, Long; Ding, Mei-Lin; Wu, Fang; He, Wen; Li, Jin; Zhang, Xiao-Yu; Xie, Wen-Li; Duan, Sheng-Zhong; Xia, Wen-Hao; Tao, Jun

2016-02-01

Although hyperaldosteronemia exerts detrimental impacts on vascular endothelium in addition to elevating blood pressure, the effects and molecular mechanisms of hyperaldosteronemia on early endothelial progenitor cell (EPC)-mediated endothelial repair after arterial damage are yet to be determined. The aim of this study was to investigate the endothelial repair capacity of early EPCs from hypertensive patients with primary hyperaldosteronemia (PHA). In vivo endothelial repair capacity of early EPCs from PHAs (n=20), age- and blood pressure-matched essential hypertension patients (n=20), and age-matched healthy subjects (n=20) was evaluated by transplantation into a nude mouse carotid endothelial denudation model. Endothelial function was evaluated by flow-mediated dilation of brachial artery in human subjects. In vivo endothelial repair capacity of early EPCs and flow-mediated dilation were impaired both in PHAs and in essential hypertension patients when compared with age-matched healthy subjects; however, the early EPC in vivo endothelial repair capacity and flow-mediated dilation of PHAs were impaired more severely than essential hypertension patients. Oral spironolactone improved early EPC in vivo endothelial repair capacity and flow-mediated dilation of PHAs. Increased oxidative stress, oxidative 5,6,7,8-tetrahydrobiopterin degradation, endothelial nitric oxide synthase uncoupling and decreased nitric oxide production were found in early EPCs from PHAs. Nicotinamide adenine dinucleotide phosphate oxidase subunit p47(phox) knockdown or 5,6,7,8-tetrahydrobiopterin supplementation attenuated endothelial nitric oxide synthase uncoupling and enhanced in vivo endothelial repair capacity of early EPCs from PHAs. In conclusion, PHAs exhibited more impaired endothelial repair capacity of early EPCs than did essential hypertension patients independent of blood pressure, which was associated with mineralocorticoid receptor-dependent oxidative stress and subsequently 5

High-polyphenol chocolate reduces endothelial dysfunction and oxidative stress during acute transient hyperglycaemia in Type 2 diabetes: a pilot randomized controlled trial.

PubMed

Mellor, D D; Madden, L A; Smith, K A; Kilpatrick, E S; Atkin, S L

2013-04-01

To investigate the effects of high-polyphenol chocolate upon endothelial function and oxidative stress in Type 2 diabetes mellitus during acute transient hyperglycaemia induced following a 75-g oral glucose challenge. Ten subjects with Type 2 diabetes underwent a double-blinded randomized controlled crossover study. A 75-g oral glucose load was used to induce hyperglycaemia, which was administered to participants 60 min after they had ingested either low (control) or high-polyphenol chocolate. Participants undertook testing at weekly intervals, following an initial cocoa-free period. Endothelial function was assessed by both functional [reactive hyperaemia peripheral artery tonometry (EndoPAT-2000) and serum markers (including intercellular adhesion molecule 1, P-selectin and P-selectin glycoprotein ligand 1]. Urinary 15-F2t-isoprostane adjusted for creatinine was used as an oxidative stress marker. Measurements were made at baseline and 2 h post-ingestion of the glucose load. Prior consumption of high-polyphenol chocolate before a glucose load improved endothelial function (1.7 ± 0.1 vs. 2.3 ± 0.1%, P = 0.01), whereas prior consumption of control chocolate resulted in a significant increase in intercellular adhesion molecule 1 (321.1 ± 7.6 vs. 373.6 ± 10.5 ng/ml, P = 0.04) and 15-F2t-isoprostane (116.8 ± 5.7 vs. 207.1 ± 5.7 mg/mol, P = 0.02). Analysis of percentage changes from baseline comparing control and high-polyphenol chocolate showed a significant improvement for high-polyphenol chocolate in both measures of endothelial function (P < 0.05) and for urinary 15-F2t-isoprostane (P = 0.04). High-polyphenol chocolate protected against acute hyperglycaemia-induced endothelial dysfunction and oxidative stress in individuals with Type 2 diabetes mellitus. © 2012 The Authors. Diabetic Medicine © 2012 Diabetes UK.

Markers of inflammation, oxidative stress, and endothelial dysfunction and the 20-year cumulative incidence of early age-related macular degeneration: the Beaver Dam Eye Study.

PubMed

Klein, Ronald; Myers, Chelsea E; Cruickshanks, Karen J; Gangnon, Ronald E; Danforth, Lorraine G; Sivakumaran, Theru A; Iyengar, Sudha K; Tsai, Michael Y; Klein, Barbara E K

2014-04-01

IMPORTANCE Modifying levels of factors associated with age-related macular degeneration (AMD) may decrease the risk for visual impairment in older persons. OBJECTIVE To examine the relationships of markers of inflammation, oxidative stress, and endothelial dysfunction to the 20-year cumulative incidence of early AMD. DESIGN, SETTING, AND PARTICIPANTS This longitudinal population-based cohort study involved a random sample of 975 persons in the Beaver Dam Eye Study without signs of AMD who participated in the baseline examination in 1988-1990 and up to 4 follow-up examinations in 1993-1995, 1998-2000, 2003-2005, and 2008-2010. EXPOSURES Serum markers of inflammation (high-sensitivity C-reactive protein, tumor necrosis factor-α receptor 2, interleukin-6, and white blood cell count), oxidative stress (8-isoprostane and total carbonyl content), and endothelial dysfunction (soluble vascular cell adhesion molecule-1 and soluble intercellular adhesion molecule-1) were measured. Interactions with complement factor H (rs1061170), age-related maculopathy susceptibility 2 (rs10490924), complement component 3 (rs2230199), and complement component 2/complement factor B (rs4151667) were examined using multiplicative models. Age-related macular degeneration was assessed from fundus photographs. MAIN OUTCOMES AND MEASURES Early AMD defined by the presence of any size drusen and the presence of pigmentary abnormalities or by the presence of large-sized drusen (≥125-μm diameter) in the absence of late AMD. RESULTS The 20-year cumulative incidence of early AMD was 23.0%. Adjusting for age, sex, and other risk factors, high-sensitivity C-reactive protein (odds ratio comparing fourth with first quartile, 2.18; P = .005), tumor necrosis factor-α receptor 2 (odds ratio, 1.78; P = .04), and interleukin-6 (odds ratio, 1.78; P = .03) were associated with the incidence of early AMD. Increased incidence of early AMD was associated with soluble vascular cell adhesion molecule

Isolation and characterization of vascular endothelial cells derived from fetal tooth buds of miniature swine.

PubMed

Nasu, Masanori; Nakahara, Taka; Tominaga, Noriko; Tamaki, Yuichi; Ide, Yoshiaki; Tachibana, Toshiaki; Ishikawa, Hiroshi

2013-03-01

The aim of the present study was to isolate endothelial cells from tooth buds (unerupted deciduous teeth) of miniature swine. Mandibular molar tooth buds harvested from swine fetuses at fetal days 90-110 were cultured in growth medium supplemented with 15% fetal bovine serum in 100-mm culture dishes until the primary cells outgrown from the tooth buds reached confluence. A morphologically defined set of pavement-shaped primary cells were picked up manually with filter paper containing trypsin/ethylenediamine tetraacetic acid solution and transferred to a separate dish. A characterization of the cellular characteristics and a functional analysis of the cultured cells at passages 3 to 5 were performed using immunofluorescence, a reverse transcriptase polymerase chain reaction assay, a tube formation assay, and transmission electron microscopy. The isolated cells grew in a pavement arrangement and showed the characteristics of contact inhibition upon reaching confluence. The population doubling time was ~48 h at passage 3. As shown by immunocytostaining and western blotting with specific antibodies, the cells produced the endothelial marker proteins such as vascular endothelial cadherin, von Willebrand factor, and vascular endothelial growth factor receptor-2. Observation with time-lapse images showed that small groups of cells aggregated and adhered to each other to form tube-like structures. Moreover, as revealed through transmission electron microscopy, these adherent cells had formed junctional complexes. These endothelial cells from the tooth buds of miniature swine are available as cell lines for studies on tube formation and use in regenerative medical science.

Mitochondria and Endothelial Function

PubMed Central

Kluge, Matthew A.; Fetterman, Jessica L.; Vita, Joseph A.

2013-01-01

In contrast to their role in other cell types with higher energy demands, mitochondria in endothelial cells primarily function in signaling cellular responses to environmental cues. This article provides an overview of key aspects of mitochondrial biology in endothelial cells, including subcellular location, biogenesis, dynamics, autophagy, ROS production and signaling, calcium homeostasis, regulated cell death, and heme biosynthesis. In each section, we introduce key concepts and then review studies showing the importance of that mechanism to endothelial control of vasomotor tone, angiogenesis, and inflammatory activation. We particularly highlight the small number of clinical and translational studies that have investigated each mechanism in human subjects. Finally, we review interventions that target different aspects of mitochondrial function and their effects on endothelial function. The ultimate goal of such research is the identification of new approaches for therapy. The reviewed studies make it clear that mitochondria are important in endothelial physiology and pathophysiology. A great deal of work will be needed, however, before mitochondria-directed therapies are available for the prevention and treatment of cardiovascular disease. PMID:23580773

Systemic inflammation, endothelial dysfunction, and activation in clinically healthy children exposed to air pollutants.

PubMed

Calderón-Garcidueñas, L; Villarreal-Calderon, R; Valencia-Salazar, G; Henríquez-Roldán, C; Gutiérrez-Castrellón, P; Torres-Jardón, R; Osnaya-Brizuela, N; Romero, L; Torres-Jardón, R; Solt, A; Reed, W

2008-03-01

Mexico City children are chronically exposed to significant concentrations of air pollutants and exhibit chronic respiratory-tract inflammation. Epidemiological, controlled human exposures, laboratory-based animal models, and in vitro/in vivo studies have shown that inflammatory, endothelial dysfunction, and endothelial damage mediators are upregulated upon exposure to particulate matter (PM). Endothelial dysfunction is a critical event in cardiovascular disease. The focus of this work was to investigate whether exposure to ambient air pollution including PM(2.5) produces systemic inflammation and endothelial injury in healthy children. We measured markers of endothelial activation, and inflammatory mediators in 52 children age 8.6+/-0.1 yr, residents of Mexico City (n: 28) or of Polotitlán (n: 24), a city with low levels of pollutants. Mexico City children had significant increases in inflammatory mediators and vasoconstrictors, including tumor necrosis factor (TNF)alpha, prostaglandin (PG) E2, C-reactive protein, interleukin-1beta, and endothelin-1. There was a significant anti-inflammatory response, and a downregulation of vascular adhesion molecule-1, intercellular adhesion molecule-1 and -2, and selectins sE and sL. Results from linear regression found TNF a positively associated with 24- and 48-h cumulative levels of PM(2.5), while the 7-d PM(2.5) value was negatively associated with the numbers of white blood cells in peripheral blood in highly exposed children. Systemic subclinical inflammation, increased endothelin- 1, and significant downregulation of soluble adhesion molecules are seen in Mexico City children. Children chronically exposed to fine PM above the standard could be at risk of developing cardiovascular diseases, atherosclerosis, stroke, and other systemic effects later in life.

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

Endothelial Microparticles From Acute Coronary Syndrome Patients Induce Premature Coronary Artery Endothelial Cell Aging and Thrombogenicity: Role of the Ang II/AT1 Receptor/NADPH Oxidase-Mediated Activation of MAPKs and PI3-Kinase Pathways.

PubMed

Abbas, Malak; Jesel, Laurence; Auger, Cyril; Amoura, Lamia; Messas, Nathan; Manin, Guillaume; Rumig, Cordula; León-González, Antonio J; Ribeiro, Thais P; Silva, Grazielle C; Abou-Merhi, Raghida; Hamade, Eva; Hecker, Markus; Georg, Yannick; Chakfe, Nabil; Ohlmann, Patrick; Schini-Kerth, Valérie B; Toti, Florence; Morel, Olivier

2017-01-17

Microparticles (MPs) have emerged as a surrogate marker of endothelial dysfunction and cardiovascular risk. This study examined the potential of MPs from senescent endothelial cells (ECs) or from patients with acute coronary syndrome (ACS) to promote premature EC aging and thrombogenicity. Primary porcine coronary ECs were isolated from the left circumflex coronary artery. MPs were prepared from ECs and venous blood from patients with ACS (n=30) and from healthy volunteers (n=4) by sequential centrifugation. The level of endothelial senescence was assessed as senescence-associated β-galactosidase activity using flow cytometry, oxidative stress using the redox-sensitive probe dihydroethidium, tissue factor activity using an enzymatic Tenase assay, the level of target protein expression by Western blot analysis, platelet aggregation using an aggregometer, and shear stress using a cone-and-plate viscometer. Senescence, as assessed by senescence-associated β-galactosidase activity, was induced by the passaging of porcine coronary artery ECs from passage P1 to P4, and was associated with a progressive shedding of procoagulant MPs. Exposure of P1 ECs to MPs shed from senescent P3 cells or circulating MPs from ACS patients induced increased senescence-associated β-galactosidase activity, oxidative stress, early phosphorylation of mitogen-activated protein kinases and Akt, and upregulation of p53, p21, and p16. Ex vivo, the prosenescent effect of circulating MPs from ACS patients was evidenced only under conditions of low shear stress. Depletion of endothelial-derived MPs from ACS patients reduced the induction of senescence. Prosenescent MPs promoted EC thrombogenicity through tissue factor upregulation, shedding of procoagulant MPs, endothelial nitric oxide synthase downregulation, and reduced nitric oxide-mediated inhibition of platelet aggregation. These MPs exhibited angiotensin-converting enzyme activity and upregulated AT1 receptors and angiotensin

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

Endothelial dysfunction: methods of assessment and application to hypertension.

PubMed

Nadar, Sunil; Blann, Andrew D; Lip, Gregory Y H

2004-01-01

Interest in the endothelium has been growing in recent decades and the traditional belief that it provides an inert interface between blood and the vessel wall is no longer the case. It is now clear that the endothelium produces a large number of substances that influence blood flow, and it is in turn affected by changes in the blood and the pressure of blood flow. Nitric oxide and endothelins are the major regulators of the vascular tone, and thereby the blood pressure. Historically speaking, concepts such as endothelial cell damage and injury were described in the 1960s and 1970s. More recently, terms such as endothelial cell activation and dysfunction have also been introduced. Although similar in some respects or part of a continuum, these terms differ in the actual effects on the endothelium, and hence differentiation is important. In hypertension, the delicate balance between the vasodilators and the vasoconstrictors is upset, with disturbance in the nitric oxide pathways that lead to a predominance of the vasoconstrictors. This in turn leads to many other changes that take place in the endothelium, setting up a vicious cycle that maintains the high blood pressure. Therefore, accurate assessment of vascular function is important in linking pathophysiology with clinical disease, such as hypertension. Indeed, there are several methods currently employed experimentally to assess endothelial dysfunction. However, the most widely studied and accepted tests are the estimation of plasma markers such as von Willebrand factor, E-selectin and thrombomodulin, and studies of forearm circulation in response to hypoxia induced stress ('flow mediated dilatation', FMD) or intra arterially administered drugs such as acetyl choline. The present document examines these topics. Whilst acknowledging the debt owed to animal models in the study of hypertension, we shall focus on work where primary study is in homo sapiens. A greater appreciation of how endothelial assessments are

Effects of industrially produced trans fat on markers of systemic inflammation: evidence from a randomized trial in women

USDA-ARS?s Scientific Manuscript database

Background: Intake of industrial trans fatty acids (TFA) has been positively associated to systemic markers of low-grade inflammation and endothelial dysfunction in cross-sectional studies, but results from intervention studies are inconclusive. Objective: To examine the effect of a high intake of T...

Effects of Swedish massage therapy on blood pressure, heart rate, and inflammatory markers in hypertensive women.

PubMed

Supa'at, Izreen; Zakaria, Zaiton; Maskon, Oteh; Aminuddin, Amilia; Nordin, Nor Anita Megat Mohd

2013-01-01

Swedish Massage Therapy (SMT) is known for its therapeutic relaxation effects. Hypertension is associated with stress and elevated endothelial inflammatory markers. This randomized control trial measured the effects of whole body SMT (massage group) or resting (control group) an hour weekly for four weeks on hypertensive women. Blood pressure (BP) and heart rate (HR) were measured before and after each intervention and endothelial inflammatory markers: vascular endothelial adhesion molecules 1 (VCAM-1) and intracellular adhesion molecules 1 (ICAM-1) were measured at baseline and after the last intervention. Massage group (n=8) showed significant systolic BP (SBP) reduction of 12 mmHg (P=0.01) and diastolic BP (DBP) reduction of 5 mmHg (P=0.01) after four sessions with no significant difference between groups. Reductions in HR were also seen in massage group after sessions 1, 3, and 4 with significant difference between groups. VCAM-1 showed significant reduction after four sessions: the massage group showed reduction of 998.05 ng/mL (P=0.03) and the control group of 375.70 ng/mL (P=0.01) with no significant differences between groups. There were no changes in ICAM-1. In conclusion, SMT or resting an hour weekly has effects on reducing BP, HR, and VCAM-1 in hypertensive women.

Substrate effects on endothelial cell adherence rates.

PubMed

Scott, W J; Mann, P

1990-01-01

Endothelial cell attachment to a synthetic substrate was studied using an in vitro model system. Attachment rate was defined as the number of tritium-labeled endothelial cells attached to a synthetic substrate after 30 minutes. The surface of the synthetic substrate was chemically modified with either laminin or fibronectin. Labeled endothelial cells attached more rapidly to synthetic substrate, chemically modified with biomolecules, as compared with the untreated substrate controls. Unlabeled endothelial cells were grown to confluency on a second set of modified and untreated substrates. The cells were removed with 1% Triton, and the rate of re-endothelialization with tritium-labeled endothelial cells was determined. The rate was 11-13 times that of the same cells on untreated substrate. These data confirm that biomolecules increase the attachment rate of endothelial cells to synthetic substrate, and also suggest that endothelial cells may secrete a Triton-insoluble product (Sigma, St. Louis, MO) into subendothelial matrix that increases re-endothelialization.

Mesenchymal-endothelial-transition contributes to cardiac neovascularization

PubMed Central

Ubil, Eric; Duan, Jinzhu; Pillai, Indulekha C.L.; Rosa-Garrido, Manuel; Wu, Yong; Bargiacchi, Francesca; Lu, Yan; Stanbouly, Seta; Huang, Jie; Rojas, Mauricio; Vondriska, Thomas M.; Stefani, Enrico; Deb, Arjun

2014-01-01

Endothelial cells contribute to a subset of cardiac fibroblasts by undergoing endothelial-to-mesenchymal-transition, but whether cardiac fibroblasts can adopt an endothelial cell fate and directly contribute to neovascularization after cardiac injury is not known. Here, using genetic fate map techniques, we demonstrate that cardiac fibroblasts rapidly adopt an endothelial cell like phenotype after acute ischemic cardiac injury. Fibroblast derived endothelial cells exhibit anatomical and functional characteristics of native endothelial cells. We show that the transcription factor p53 regulates such a switch in cardiac fibroblast fate. Loss of p53 in cardiac fibroblasts severely decreases the formation of fibroblast derived endothelial cells, reduces post infarct vascular density and worsens cardiac function. Conversely, stimulation of the p53 pathway in cardiac fibroblasts augments mesenchymal to endothelial transition, enhances vascularity and improves cardiac function. These observations demonstrate that mesenchymal-to-endothelial-transition contributes to neovascularization of the injured heart and represents a potential therapeutic target for enhancing cardiac repair. PMID:25317562

[Endothelial dysfunction and nonspecific immune reactions in development and progression of osteoarthrosis in women engaged into manual work].

PubMed

Maliutina, N N; Nevzorova, M S

2015-01-01

The article considers mechanisms of development and progression of osteoarthrosis as an occupationally conditioned disease in women of manual work. Women working in physical overstrain conditions are under occupational risk with dysfunction of many body systems. The authors set a hypothesis on association of endothelial dysfunction markers dysbalance and structural remodelling of cartilage matrix as a proof of degenerative changes.

Visit-to-visit blood pressure variability as a prognostic marker in patients with cardiovascular and cerebrovascular diseases--relationships and comparisons with vascular markers of atherosclerosis.

PubMed

Lau, Kui Kai; Wong, Yuen Kwun; Chan, Yap Hang; Teo, Kay Cheong; Chan, Koon Ho; Wai Li, Leonard Sheung; Cheung, Raymond Tak Fai; Siu, Chung Wah; Ho, Shu Leong; Tse, Hung Fat

2014-07-01

Visit-to-visit blood pressure variability (BPV) is a simple surrogate marker for the development of atherosclerotic diseases, cardiovascular and all-cause mortality. Nevertheless, the relative prognostic value of BPV in comparison with other established vascular assessments remain uncertain. We prospectively followed-up 656 high-risk patients with diabetes or established cardiovascular or cerebrovascular diseases for the occurrence of major adverse cardiovascular events (MACEs). Baseline brachial endothelial function, carotid intima-media thickness (IMT) and plaque burden, ankle-brachial index and arterial stiffness were determined. Visit-to-visit BPV were recorded during a mean 18 ± 9 outpatient clinic visits. After a mean 81 ± 12 month's follow-up, 123 patients (19%) developed MACEs. Patients who developed a MACE had significantly higher systolic BPV, more severe endothelial function, arterial stiffness and systemic atherosclerotic burden compared to patients who did not develop a MACE (all P<0.01). BPV significantly correlated with all of the vascular assessments (P<0.01). A high carotid IMT had the greatest prognostic value in predicting development of a MACE (area under receiver operating characteristic curve (AUC) 0.69 ± 0.03, P<0.01). A high BPV also had moderate prognostic value in prediction of MACE (AUC 0.65 ± 0.03, P<0.01). After adjustment of confounding factors, a high BPV remained a significant independent predictor of MACE (hazards ratio 1.67, 95% confidence interval 1.14-2.43, P<0.01). Compared with established surrogate markers of atherosclerosis, visit-to-visit BPV provides similar prognostic information and may represent a new and simple marker for adverse outcomes in patients with vascular diseases. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Far-infrared protects vascular endothelial cells from advanced glycation end products-induced injury via PLZF-mediated autophagy in diabetic mice

PubMed Central

Chen, Cheng-Hsien; Chen, Tso-Hsiao; Wu, Mei-Yi; Chou, Tz-Chong; Chen, Jia-Rung; Wei, Meng-Jun; Lee, San-Liang; Hong, Li-Yu; Zheng, Cai-Mei; Chiu, I-Jen; Lin, Yuh-Feng; Hsu, Ching-Min; Hsu, Yung-Ho

2017-01-01

The accumulation of advanced glycation end products (AGEs) in diabetic patients induces vascular endothelial injury. Promyelocytic leukemia zinc finger protein (PLZF) is a transcription factor that can be activated by low-temperature far-infrared (FIR) irradiation to exert beneficial effects on the vascular endothelium. In the present study, we investigated the influence of FIR-induced PLZF activation on AGE-induced endothelial injury both in vitro and in vivo. FIR irradiation inhibited AGE-induced apoptosis in human umbilical vein endothelial cells (HUVECs). PLZF activation increased the expression of phosphatidylinositol-3 kinases (PI3K), which are important kinases in the autophagic signaling pathway. FIR-induced PLZF activation led to autophagy in HUVEC, which was mediated through the upregulation of PI3K. Immunofluorescence staining showed that AGEs were engulfed by HUVECs and localized to lysosomes. FIR-induced autophagy promoted AGEs degradation in HUVECs. In nicotinamide/streptozotocin-induced diabetic mice, FIR therapy reduced serum AGEs and AGEs deposition at the vascular endothelium. FIR therapy also reduced diabetes-induced inflammatory markers in the vascular endothelium and improved vascular endothelial function. These protective effects of FIR therapy were not found in PLZF-knockout mice. Our data suggest that FIR-induced PLZF activation in vascular endothelial cells protects the vascular endothelium in diabetic mice from AGE-induced injury. PMID:28071754

Endothelial glycocalyx: permeability barrier and mechanosensor.

PubMed

Curry, F E; Adamson, R H

2012-04-01

Endothelial cells are covered with a polysaccharide rich layer more than 400 nm thick, mechanical properties of which limit access of circulating plasma components to endothelial cell membranes. The barrier properties of this endothelial surface layer are deduced from the rate of tracer penetration into the layer and the mechanics of red and white cell movement through capillary microvessels. This review compares the mechanosensor and permeability properties of an inner layer (100-150 nm, close to the endothelial membrane) characterized as a quasi-periodic structure which accounts for key aspects of transvascular exchange and vascular permeability with those of the whole endothelial surface layers. We conclude that many of the barrier properties of the whole surface layer are not representative of the primary fiber matrix forming the molecular filter determining transvascular exchange. The differences between the properties of the whole layer and the inner glycocalyx structures likely reflect dynamic aspects of the endothelial surface layer including tracer binding to specific components, synthesis and degradation of key components, activation of signaling pathways in the endothelial cells when components of the surface layer are lost or degraded, and the spatial distribution of adhesion proteins in microdomains of the endothelial cell membrane.

The natural compound codonolactone impairs tumor induced angiogenesis by downregulating BMP signaling in endothelial cells.

PubMed

Wang, Shan; Cai, Rui; Ma, Junchao; Liu, Ting; Ke, Xiaoqin; Lu, Hong; Fu, Jianjiang

2015-10-15

Angiogenesis, the recruitment of new blood vessels, was demonstrated that is an essential component of the growth of a tumor beyond a certain size and the metastatic pathway. The potential use of angiogenesis-based agents, such as those involving natural and synthetic inhibitors as anticancer drugs is currently under intense investigation. In this study, the anti-angiogenic properties of codonolactone (CLT), a sesquiterpene lactone from Atractylodes lancea, were examined in endothelial cells. Our published study reported that CLT shows significant anti-metastatic properties in vitro and in vivo. In order to determine whether angiogenic-involved mechanisms contribute to the anti-metastatic effects of CLT, we checked the anti-angiogenic properties of CLT and its potential mechanisms. Human umbilical vein endothelial cells (HUVECs) and EA.hy 926 cells were involved in this study. Immunofluorescence assay for cells and immunohistochemistry assay for tissues were used to check the expression of angiogenic markers. In vitro migration and invasion of endothelial cells treated with and without CLT were analyzed. Protein expressions were measured by Western blot analysis. For MMPs activity assay, fluorescence resonance energy transfer-based MMPs activity assay and gelatin zymography assay were involved in this study. Here we demonstrated that CLT exhibited inhibition on cancer cell induced angiogenesis in vivo, and direct inhibited migration and invasion of endothelial cells in vitro. Moreover, we observed that the down-regulation of MMPs and VEGF-VEGFR2 was involved in the anti-angiogenic effects of CLT. Data from Western blotting showed that, in endothelial cells, CLT reduced Runx2 activation and BMP signaling. Our findings demonstrated that CLT impaired the development of angiogenesis both in vitro and in vivo by direct inhibition on endothelial cells. These inhibitory effects were depended on its ability to interference with BMP signaling in endothelial cells, which may

Sirtuin1 protects endothelial Caveolin-1 expression and preserves endothelial function via suppressing miR-204 and endoplasmic reticulum stress.

PubMed

Kassan, M; Vikram, A; Kim, Y R; Li, Q; Kassan, A; Patel, H H; Kumar, S; Gabani, M; Liu, J; Jacobs, J S; Irani, K

2017-02-09

Sirtuin1 (Sirt1) is a class III histone deacetylase that regulates a variety of physiological processes, including endothelial function. Caveolin1 (Cav1) is also an important determinant of endothelial function. We asked if Sirt1 governs endothelial Cav1 and endothelial function by regulating miR-204 expression and endoplasmic reticulum (ER) stress. Knockdown of Sirt1 in endothelial cells, and in vivo deletion of endothelial Sirt1, induced endothelial ER stress and miR-204 expression, reduced Cav1, and impaired endothelium-dependent vasorelaxation. All of these effects were reversed by a miR-204 inhibitor (miR-204 I) or with overexpression of Cav1. A miR-204 mimic (miR-204 M) decreased Cav1 in endothelial cells. In addition, high-fat diet (HFD) feeding induced vascular miR-204 and reduced endothelial Cav1. MiR-204-I protected against HFD-induced downregulation of endothelial Cav1. Moreover, pharmacologic induction of ER stress with tunicamycin downregulated endothelial Cav1 and impaired endothelium-dependent vasorelaxation that was rescued by overexpressing Cav1. In conclusion, Sirt1 preserves Cav1-dependent endothelial function by mitigating miR-204-mediated vascular ER stress.

c-Met–mediated endothelial plasticity drives aberrant vascularization and chemoresistance in glioblastoma

PubMed Central

Huang, Menggui; Liu, Tianrun; Ma, Peihong; Mitteer, R. Alan; Zhang, Zhenting; Kim, Hyun Jun; Yeo, Eujin; Zhang, Duo; Cai, Peiqiang; Li, Chunsheng; Zhang, Lin; Zhao, Botao; Roccograndi, Laura; O’Rourke, Donald M.; Dahmane, Nadia; Gong, Yanqing; Koumenis, Constantinos

2016-01-01

Aberrant vascularization is a hallmark of cancer progression and treatment resistance. Here, we have shown that endothelial cell (EC) plasticity drives aberrant vascularization and chemoresistance in glioblastoma multiforme (GBM). By utilizing human patient specimens, as well as allograft and genetic murine GBM models, we revealed that a robust endothelial plasticity in GBM allows acquisition of fibroblast transformation (also known as endothelial mesenchymal transition [Endo-MT]), which is characterized by EC expression of fibroblast markers, and determined that a prominent population of GBM-associated fibroblast-like cells have EC origin. Tumor ECs acquired the mesenchymal gene signature without the loss of EC functions, leading to enhanced cell proliferation and migration, as well as vessel permeability. Furthermore, we identified a c-Met/ETS-1/matrix metalloproteinase–14 (MMP-14) axis that controls VE-cadherin degradation, Endo-MT, and vascular abnormality. Pharmacological c-Met inhibition induced vessel normalization in patient tumor–derived ECs. Finally, EC-specific KO of Met inhibited vascular transformation, normalized blood vessels, and reduced intratumoral hypoxia, culminating in suppressed tumor growth and prolonged survival in GBM-bearing mice after temozolomide treatment. Together, these findings illustrate a mechanism that controls aberrant tumor vascularization and suggest that targeting Endo-MT may offer selective and efficient strategies for antivascular and vessel normalization therapies in GBM, and possibly other malignant tumors. PMID:27043280

Activation of the endoplasmic reticulum stress response by the amyloid-beta 1-40 peptide in brain endothelial cells.

PubMed

Fonseca, Ana Catarina R G; Ferreiro, Elisabete; Oliveira, Catarina R; Cardoso, Sandra M; Pereira, Cláudia F

2013-12-01

Neurovascular dysfunction arising from endothelial cell damage is an early pathogenic event that contributes to the neurodegenerative process occurring in Alzheimer's disease (AD). Since the mechanisms underlying endothelial dysfunction are not fully elucidated, this study was aimed to explore the hypothesis that brain endothelial cell death is induced upon the sustained activation of the endoplasmic reticulum (ER) stress response by amyloid-beta (Aβ) peptide, which deposits in the cerebral vessels in many AD patients and transgenic mice. Incubation of rat brain endothelial cells (RBE4 cell line) with Aβ1-40 increased the levels of several markers of ER stress-induced unfolded protein response (UPR), in a time-dependent manner, and affected the Ca(2+) homeostasis due to the release of Ca(2+) from this intracellular store. Finally, Aβ1-40 was shown to activate both mitochondria-dependent and -independent apoptotic cell death pathways. Enhanced release of cytochrome c from mitochondria and activation of the downstream caspase-9 were observed in cells treated with Aβ1-40 concomitantly with caspase-12 activation. Furthermore, Aβ1-40 activated the apoptosis effectors' caspase-3 and promoted the translocation of apoptosis-inducing factor (AIF) to the nucleus demonstrating the involvement of caspase-dependent and -independent mechanisms during Aβ-induced endothelial cell death. In conclusion, our data demonstrate that ER stress plays a significant role in Aβ1-40-induced apoptotic cell death in brain endothelial cells suggesting that ER stress-targeted therapeutic strategies might be useful in AD to counteract vascular defects and ultimately neurodegeneration. © 2013.

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

Early outgrowth cells versus endothelial colony forming cells functions in platelet aggregation.

PubMed

Bou Khzam, Lara; Bouchereau, Olivier; Boulahya, Rahma; Hachem, Ahmed; Zaid, Younes; Abou-Saleh, Haissam; Merhi, Yahye

2015-11-09

Endothelial progenitor cells (EPCs) have been implicated in neoangiogenesis, endothelial repair and cell-based therapies for cardiovascular diseases. We have previously shown that the recruitment of EPCs to sites of vascular lesions is facilitated by platelets where EPCs, in turn, modulate platelet function and thrombosis. However, EPCs encompass a heterogeneous population of progenitor cells that may exert different effects on platelet function. Recent evidence suggests the existence of two EPC subtypes: early outgrowth cells (EOCs) and endothelial colony-forming cells (ECFCs). We aimed at characterizing these two EPC subtypes and at identifying their role in platelet aggregation. EOCs and ECFCs were generated from human peripheral blood mononuclear cells (PBMCs) seeded in conditioned media on fibronectin and collagen, respectively. The morphological, phenotypical and functional characteristics of EOCs and ECFCs were assessed by optical and confocal laser scanning microscopes, cell surface markers expression, and Matrigel tube formation. The impact of EOCs and ECFCs on platelet aggregation was monitored in collagen-induced optical aggregometry and compared with PBMCs and human umbilical vein endothelial cells (HUVECs). The levels of the anti-platelet agents' nitric oxide (NO) and prostacyclin (PGI2) released from cultured cells as well as the expression of their respective producing enzymes NO synthases (NOS) and cyclooxygenases (COX) were also assessed. We showed that EOCs display a monocytic-like phenotype whereas ECFCs have an endothelial-like phenotype. We demonstrated that both EOCs and ECFCs and their supernatants inhibited platelet aggregation; however ECFCs were more efficient than EOCs. This could be related to the release of significantly higher amounts of NO and PGI2 from ECFCs, in comparison to EOCs. Indeed, ECFCs, like HUVECs, constitutively express the endothelial (eNOS)-and inducible (iNOS)-NOS isoforms, and COX-1 and weakly express COX-2, whereas

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

Screening and Characterization of Drugs That Protect Corneal Endothelial Cells Against Unfolded Protein Response and Oxidative Stress.

PubMed

Kim, Eun Chul; Toyono, Tetsuya; Berlinicke, Cynthia A; Zack, Donald J; Jurkunas, Ula; Usui, Tomohiko; Jun, Albert S

2017-02-01

To screen for and characterize compounds that protect corneal endothelial cells against unfolded protein response (UPR) and oxidative stress. Bovine corneal endothelial cells (BCECs) were treated for 48 hours with 640 compounds from a Food and Drug Administration (FDA)-approved drug library and then challenged with thapsigargin or H2O2 to induce UPR or oxidative stress, respectively. Cell viability was measured using the CellTiter-Glo survival assay. Selected "hits" were subjected to further dose-response testing, and their ability to modulate expression of UPR and oxidative stress markers was assessed by RT-PCR, Western blot, and measurement of protein carbonyl and 8-hydroxydeoxyguanosine (8-OHdG) adducts in immortalized human corneal endothelial cells (iHCECs). Forty-one drugs at 20 μM and 55 drugs at 100 μM increased survival of H2O2-challenged cells, and 8 drugs at 20 μM and 2 drugs at 100 μM increased survival of thapsigargin-challenged cells, compared with untreated control cells. Nicergoline, ergothioneine, nimesulide, oxotremorine, and mefenamic acid increased survival of both H2O2- and thapsigargin-challenged cells. Oxotremorine altered DNA damage inducible 3 (CHOP) gene expression, glucose-regulated protein 78 kDa (GRP78) and activating transcription factor 4 (ATF4) protein expression, and protein carbonyl and 8-OHdG levels. Mefenamic acid altered GRP78 protein expression and protein carbonyl and 8-OHdG levels. Oxotremorine and mefenamic acid are potential survival factors for corneal endothelial cells under UPR and oxidative stress. The described assay can be further expanded to screen additional drugs for potential therapeutic effect in corneal endothelial diseases such as Fuchs' endothelial corneal dystrophy.

Tumor microenvironment conditions alter Akt and Na+/H+ exchanger NHE1 expression in endothelial cells more than hypoxia alone: implications for endothelial cell function in cancer.

PubMed

Pedersen, A K; Mendes Lopes de Melo, J; Mørup, N; Tritsaris, K; Pedersen, S F

2017-08-14

migration. TME effects on HIF-1α, VEGF, Akt, translation, proliferation or apoptosis markers were unaffected by NHE1 knockdown/inhibition. NHE1 and Akt are downregulated by TME conditions, more potently than by hypoxia alone. This inhibits endothelial cell migration and growth in a manner likely modulated by the cancer cell secretome.

[Circuit resistance training improved endothelial dysfunction in obese aged women].

PubMed

Rosety, Ignacio; Pery, María Teresa; Rosety, Jesús; García, Natalia; Rodríguez-Pareja, María Antonia; Brenes-Martín, Francisco; Díaz, Antonio; Rosety-Rodríguez, Manuel; Ordoñez, Francisco Javier; Rosety, Miguel Ángel

2016-02-16

It is widely accepted that obesity is associated with endothelial dysfunction. In a recent paper, we have also found circuit resistance training may reduce visceral fat in obese aged women. Accordingly, the current study was conducted to ascertain the effects of circuit resistance training on markers of endothelial dysfunction in this population group. In the present interventional study, a total of 48 obese aged women were recruited from the community. Twenty-four of them were randomly assigned to perform a 12-week resistance circuit training programme, 3-days per week. This training was circularly performed in 6 stations: arm curl, leg extension, seated row, leg curl, triceps extension and leg press. The Jamar handgrip electronic dynamometer was used to assess maximal handgrip strength of the dominant hand. Lastly, serum samples were analysed using an immunoassay (ELISA) for endothelin-1, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). When compared to baseline, resistance training significantly reduced serum levels of endothelin-1 (2.28 ± 0.7 vs. 1.98 ± 1.1 pg/ml; p = 0.019; d = 0.67) and ICAM-1 (290 ± 69 vs. 255 ± 76 ng/ml; p = 0.004; d = 0.92) in the experimental group. No significant changes in any of the tested outcomes were found in the control group. A short-term circuit resistance program improved endothelial dysfunction in aged obese women. Further studies on this topic are still required to consolidate this approach in clinical application.

Impact of HIV and Type 2 diabetes on Gut Microbiota Diversity, Tryptophan Catabolism and Endothelial Dysfunction.

PubMed

Hoel, Hedda; Hove-Skovsgaard, Malene; Hov, Johannes R; Gaardbo, Julie Christine; Holm, Kristian; Kummen, Martin; Rudi, Knut; Nwosu, Felix; Valeur, Jørgen; Gelpi, Marco; Seljeflot, Ingebjørg; Ueland, Per Magne; Gerstoft, Jan; Ullum, Henrik; Aukrust, Pål; Nielsen, Susanne Dam; Trøseid, Marius

2018-04-30

HIV infection and type 2 diabetes are associated with altered gut microbiota, chronic inflammation, and increased cardiovascular risk. We aimed to investigate the combined effect of these diseases on gut microbiota composition and related metabolites, and a potential relation to endothelial dysfunction in individuals with HIV-infection only (n = 23), diabetes only (n = 16) or both conditions (n = 21), as well as controls (n = 24). Fecal microbiota was analyzed by Illumina sequencing of the 16 S rRNA gene. Markers of endothelial dysfunction (asymmetric dimethylarginine [ADMA]), tryptophan catabolism (kynurenine/tryptophan [KT]-ratio), and inflammation (neopterin) were measured by liquid chromatography-tandem mass spectrometry. The combination of HIV and type 2 diabetes was associated with reduced gut microbiota diversity, increased plasma KT-ratio and neopterin. Microbial genes related to tryptophan metabolism correlated with KT-ratio and low alpha diversity, in particular in HIV-infected with T2D. In multivariate analyses, KT-ratio associated with ADMA (β = 4.58 [95% CI 2.53-6.63], p < 0.001), whereas microbiota composition per se was not associated with endothelial dysfunction. Our results indicate that tryptophan catabolism may be related to endothelial dysfunction, with a potentially detrimental interaction between HIV and diabetes. The potential contribution of gut microbiota and the impact for cardiovascular risk should be further explored in prospective studies powered for clinical end points.

Ethnicity, plasma phospholipid fatty acid composition and inflammatory/endothelial activation biomarkers in the Multi-Ethnic Study of Atherosclerosis (MESA).

PubMed

Steffen, B T; Steffen, L M; Tracy, R; Siscovick, D; Jacobs, D; Liu, K; He, K; Hanson, N Q; Nettleton, J A; Tsai, M Y

2012-05-01

It has been recognized that certain long-chain polyunsaturated fatty acids (LC-PUFAs) are involved in inflammation and its resolution. It has also been shown that ethnicity may be a factor in affecting systemic inflammation, and limited evidence suggests it may influence plasma LC-PUFA composition. Given the links among these three factors, we aim to determine ethnicity-based differences in plasma LC-PUFA composition among White, Black, Hispanic and Chinese participants, and whether such differences contribute to variations in markers of inflammation and endothelial activation in a sub-cohort of the Multi-Ethnic Study of Atherosclerosis (MESA). Plasma phospholipid LC-PUFAs levels (%) were determined in 2848 MESA participants using gas chromatography-flame ionization detection. Enzyme immunoassays determined inflammatory markers levels for high-sensitivity C-reactive protein (n=2848), interleukin-6 (n=2796), soluble tumor necrosis factor-α receptor type 1 (n=998), and endothelial activation markers soluble intercellular adhesion molecule-1 (n=1192) and soluble E-selectin (n=998). The modifying influence of ethnicity was tested by linear regression analysis. Chinese adults were found to have the highest mean levels of plasma eicosapentaenoic acid (EPA, 1.24%) and docosahexaenoic acid (DHA, 4.95%), and the lowest mean levels of γ-linolenic (0.10%), dihomo-γ-linolenic (DGLA, 2.96%) and arachidonic (10.72%) acids compared with the other ethnicities (all P ≤ 0.01). In contrast, Hispanics had the lowest mean levels of plasma EPA (0.70%) and DHA (3.49%), and the highest levels of DGLA (3.59%; all P ≤ 0.01). Significant differences in EPA and DHA among ethnicities were attenuated following adjustment for dietary non-fried fish and fish oil supplementation. Ethnicity did not modify the associations of LC-PUFAs with markers of inflammation or endothelial activation (all P (interaction)>0.05). The absence of a modifying effect of ethnicity indicates that the putative

A sliding technique to load thin endothelial donor lamella onto Busin glide for Descemet-stripping automated endothelial keratoplasty

PubMed Central

Kobayashi, Akira; Yokogawa, Hideaki; Sugiyama, Kazuhisa

2012-01-01

We describe a modified technique for loading donor corneal endothelial lamella onto a Busin glide® without causing wrinkles, as part of the procedure of Descemet-stripping automated endothelial keratoplasty. Briefly, after punching out a composite of the donor-endothelial lamella and a microkeratome-dissected cap, several drops of dispersive ophthalmic viscosurgical device are placed onto the endothelial surface. The Busin glide surface is then wetted with several drops of balanced salt solution. After the composite is transferred onto the Busin glide, hydrodissection of the potential space between the donor-endothelial lamella and the microkeratome-dissected cap is carefully performed to enable smooth detachment of these two lamellae. Whereas simply dragging the donor-endothelial lamella directly onto the glide can cause wrinkling or folding of the donor lamella, this technique enables smooth detachment of the composite without wrinkle or fold formation, and results in less endothelial cell damage. PMID:22927732

High glucose increases the formation and pro-oxidative activity of endothelial microparticles.

PubMed

Burger, Dylan; Turner, Maddison; Xiao, Fengxia; Munkonda, Mercedes N; Akbari, Shareef; Burns, Kevin D

2017-09-01

Individuals with diabetes exhibit increases in circulating endothelial microparticles (eMPs, also referred to as endothelial microvesicles), which are associated with endothelial dysfunction and a heightened risk of cardiovascular complications. We have shown that eMPs are markers and mediators of vascular injury although their role in diabetes is unclear. We hypothesised that the composition and biological activity of eMPs are altered in response to high glucose exposure. We assessed the effects of high glucose on eMP formation, composition and signalling in cultured HUVECs. eMPs were isolated from the media of HUVECs cultured under conditions of normal glucose (eMP NG ), high glucose (eMP HG ) or osmotic control of L-glucose (eMP LG ). eMP size, concentration and surface charge were assessed by nanoparticle tracking analysis and flow cytometry. eMP protein composition was assessed by liquid chromatography-tandem mass spectrometry, and eMP-mediated effects on coagulation, reactive oxygen species (ROS) production and vessel function were assessed. Exposure of HUVECs to high glucose for 24 h caused a threefold increase in eMP formation, increased mean particle size (269 ± 18 nm vs 226 ± 11 nm) and decreased surface charge. Compared with eMP NG or eMP LG , eMP HG possessed approximately threefold greater pro-coagulant activity, stimulated HUVEC ROS production to a greater extent (~250% of eMP NG ) and were more potent inhibitors of endothelial-dependent relaxation. Proteomic analysis of eMPs identified 1212 independent proteins of which 68 were exclusively found in eMP HG . Gene ontology analysis revealed that eMP HG -exclusive proteins were associated with signalling pathways related to blood coagulation, cell signalling and immune cell activation. Our results indicate that elevated glucose is a potent stimulus for eMP formation that also alters their molecular composition leading to increased bioactivity. Such effects may contribute to progressive

Elevated plasma free fatty acids increase cardiovascular risk by inducing plasma biomarkers of endothelial activation, myeloperoxidase and PAI-1 in healthy subjects.

PubMed

Mathew, Manoj; Tay, Eric; Cusi, Kenneth

2010-02-16

CVD in obesity and T2DM are associated with endothelial activation, elevated plasma vascular inflammation markers and a prothrombotic state. We examined the contribution of FFA to these abnormalities following a 48-hour physiological increase in plasma FFA to levels of obesity and diabetes in a group of healthy subjects. 40 non-diabetic subjects (age = 38 +/- 3 yr, BMI = 28 +/- 1 kg/m2, FPG = 95 +/- 1 mg/dl, HbA1c = 5.3 +/- 0.1%) were admitted twice and received a 48-hour infusion of normal saline or low-dose lipid. Plasma was drawn for intracellular (ICAM-1) and vascular (VCAM-1) adhesion molecules-1, E-selectin (sE-S), myeloperoxidase (MPO) and total plasminogen inhibitor-1 (tPAI-1). Insulin sensitivity was measured by a hyperglycemic clamp (M/I). Lipid infusion increased plasma FFA to levels observed in obesity and T2DM and reduced insulin sensitivity by 27% (p = 0.01). Elevated plasma FFA increased plasma markers of endothelial activation ICAM-1 (138 +/- 10 vs. 186 +/- 25 ng/ml), VCAM-1 (1066 +/- 67 vs. 1204 +/- 65 ng/ml) and sE-S (20 +/- 1 vs. 24 +/- 1 ng/ml) between 13-35% and by > or = 2-fold plasma levels of myeloperoxidase (7.5 +/- 0.9 to 15 +/- 25 ng/ml), an inflammatory marker of future CVD, and tPAI-1 (9.7 +/- 0.6 to 22.5 +/- 1.5 ng/ml), an indicator of a prothrombotic state (all p < or = 0.01). The FFA-induced increase was independent from the degree of adiposity, being of similar magnitude in lean, overweight and obese subjects. An increase in plasma FFA within the physiological range observed in obesity and T2DM induces markers of endothelial activation, vascular inflammation and thrombosis in healthy subjects. This suggests that even transient (48-hour) and modest increases in plasma FFA may initiate early vascular abnormalities that promote atherosclerosis and CVD.

A biphasic endothelial stress-survival mechanism regulates the cellular response to vascular endothelial growth factor A

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

Latham, Antony M.; Odell, Adam F.; Mughal, Nadeem A.

2012-11-01

Vascular endothelial growth factor A (VEGF-A) is an essential cytokine that regulates endothelial function and angiogenesis. VEGF-A binding to endothelial receptor tyrosine kinases such as VEGFR1 and VEGFR2 triggers cellular responses including survival, proliferation and new blood vessel sprouting. Increased levels of a soluble VEGFR1 splice variant (sFlt-1) correlate with endothelial dysfunction in pathologies such as pre-eclampsia; however the cellular mechanism(s) underlying the regulation and function of sFlt-1 are unclear. Here, we demonstrate the existence of a biphasic stress response in endothelial cells, using serum deprivation as a model of endothelial dysfunction. The early phase is characterized by a highmore » VEGFR2:sFlt-1 ratio, which is reversed in the late phase. A functional consequence is a short-term increase in VEGF-A-stimulated intracellular signaling. In the late phase, sFlt-1 is secreted and deposited at the extracellular matrix. We hypothesized that under stress, increased endothelial sFlt-1 levels reduce VEGF-A bioavailability: VEGF-A treatment induces sFlt-1 expression at the cell surface and VEGF-A silencing inhibits sFlt-1 anchorage to the extracellular matrix. Treatment with recombinant sFlt-1 inhibits VEGF-A-stimulated in vitro angiogenesis and sFlt-1 silencing enhances this process. In this response, increased VEGFR2 levels are regulated by the phosphatidylinositol-3-kinase and PKB/Akt signaling pathways and increased sFlt-1 levels by the ERK1/2 signaling pathway. We conclude that during serum withdrawal, cellular sensing of environmental stress modulates sFlt-1 and VEGFR2 levels, regulating VEGF-A bioavailability and ensuring cell survival takes precedence over cell proliferation and migration. These findings may underpin an important mechanism contributing to endothelial dysfunction in pathological states. -- Highlights: Black-Right-Pointing-Pointer Endothelial cells mount a stress response under conditions of low serum. Black

Endothelial Glycocalyx as a Shield Against Diabetic Vascular Complications: Involvement of Hyaluronan and Hyaluronidases.

PubMed

Dogné, Sophie; Flamion, Bruno; Caron, Nathalie

2018-07-01

The endothelial glycocalyx (EG), which covers the apical surface of the endothelial cells and floats into the lumen of the vessels, is a key player in vascular integrity and cardiovascular homeostasis. The EG is composed of PGs (proteoglycans), glycoproteins, glycolipids, and glycosaminoglycans, in particular hyaluronan (HA). HA seems to be implicated in most of the functions described for EG such as creating a space between blood and the endothelium, controlling vessel permeability, restricting leukocyte and platelet adhesion, and allowing an appropriate endothelial response to flow variation through mechanosensing. The amount of HA in the EG may be regulated by HYAL (hyaluronidase) 1, the most active somatic hyaluronidase. HYAL1 seems enriched in endothelial cells through endocytosis from the bloodstream. The role of the other main somatic hyaluronidase, HYAL2, in the EG is uncertain. Damage to the EG, accompanied by shedding of one or more of its components, is an early sign of various pathologies including diabetes mellitus. Shedding increases the blood or plasma concentration of several EG components, such as HA, heparan sulfate, and syndecan. The plasma levels of these molecules can then be used as sensitive markers of EG degradation. This has been shown in type 1 and type 2 diabetic patients. Recent experimental studies suggest that preserving the size and amount of EG HA in the face of diabetic insults could be a useful novel therapeutic strategy to slow diabetic complications. One way to achieve this goal, as suggested by a murine model of HYAL1 deficiency, may be to inhibit the function of HYAL1. The same approach may succeed in other pathological situations involving endothelial dysfunction and EG damage. © 2018 American Heart Association, Inc.

Signaling hierarchy regulating human endothelial cell development.

PubMed

Kelly, Melissa A; Hirschi, Karen K

2009-05-01

Our present knowledge of the regulation of mammalian endothelial cell differentiation has been largely derived from studies of mouse embryonic development. However, unique mechanisms and hierarchy of signals that govern human endothelial cell development are unknown and, thus, explored in these studies. Using human embryonic stem cells as a model system, we were able to reproducibly and robustly generate differentiated endothelial cells via coculture on OP9 marrow stromal cells. We found that, in contrast to studies in the mouse, bFGF and VEGF had no specific effects on the initiation of human vasculogenesis. However, exogenous Ihh promoted endothelial cell differentiation, as evidenced by increased production of cells with cobblestone morphology that coexpress multiple endothelial-specific genes and proteins, form lumens, and exhibit DiI-AcLDL uptake. Inhibition of BMP signaling using Noggin or BMP4, specifically, using neutralizing antibodies suppressed endothelial cell formation; whereas, addition of rhBMP4 to cells treated with the hedgehog inhibitor cyclopamine rescued endothelial cell development. Our studies revealed that Ihh promoted human endothelial cell differentiation from pluripotent hES cells via BMP signaling, providing novel insights applicable to modulating human endothelial cell formation and vascular regeneration for human clinical therapies.

Reactive hyperemia index (RHI) and cognitive performance indexes are associated with histologic markers of liver disease in subjects with non-alcoholic fatty liver disease (NAFLD): a case control study.

PubMed

Tuttolomondo, Antonino; Petta, Salvatore; Casuccio, Alessandra; Maida, Carlo; Corte, Vittoriano Della; Daidone, Mario; Di Raimondo, Domenico; Pecoraro, Rosaria; Fonte, Roberto; Cirrincione, Anna; Zafonte, Rita; Cabibi, Daniela; Cammà, Calogero; Di Marco, Vito; Licata, Anna; Magliozzo, Franco; Marchesini, Giulio; Merlino, Giovanni; Craxì, Antonio; Pinto, Antonio

2018-02-16

No study evaluated vascular health markers in subjects with non-alcoholic fatty liver disease (NAFLD) through a combined analysis of reactive hyperemia peripheral arterial tonometry (RH-PAT) and arterial stiffness indexes. We aimed to assess whether NAFLD and its histological severity are associated with impairment of arterial stiffness and RH-PAT indexes in a mixed cohort of patients with biopsy-proven NAFLD. The Kleiner classification was used to grade NAFLD grade. Pulse wave velocity (PWV) and augmentation index (Aix) were used as markers of arterial stiffness, whereas endothelial function was assessed using reactive hyperemia index (RHI). The mini-mental state examination (MMSE) was administered to test cognitive performance. 80 consecutive patients with biopsy-proven NAFLD and 83 controls without fatty liver disease. NAFLD subjects showed significantly lower mean RHI, higher mean arterial stiffness indexes and lower mean MMSE score. Multivariable analysis after correction for BMI, dyslipidaemia, hypertension, sex, diabetes, age and cardiovascular disease showed that BMI, diastolic blood pressure and RHI are significantly associated to NAFLD. Simple linear regression analysis showed among non-alcoholic steatohepatitis (NASH) subjects a significant negative relationship between ballooning grade and MMSE and a significant positive association between Kleiner steatosis grade and augmentation index. Future research will be addressed to evaluate the relationship between inflammatory markers and arterial stiffness and endothelial function indexes in NAFLD subjects. These study will evaluate association between cardiovascular event incidence and arterial stiffness, endothelial and cognitive markers, and they will address the beneficial effects of cardiovascular drugs such as statins and ACE inhibitors on these surrogate markers in NAFLD subjects.

Reduced Ang2 expression in aging endothelial cells

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

Hohensinner, P.J., E-mail: philipp.hohensinner@meduniwien.ac.at; Ebenbauer, B.; Ludwig Boltzmann Cluster for Cardiovascular Research, Vienna

Aging endothelial cells are characterized by increased cell size, reduced telomere length and increased expression of proinflammatory cytokines. In addition, we describe here that aging reduces the migratory distance of endothelial cells. Furthermore, we observe an increase of the quiescence protein Ang1 and a decrease of the endothelial activation protein Ang2 upon aging. Supplementing Ang2 to aged endothelial cells restored their migratory capacity. We conclude that aging shifts the balance of the Ang1/Ang2 network favouring a quiescent state. Activation of endothelial cells in aging might be necessary to enhance wound healing capacities. -- Highlights: •Endothelial cells display signs of agingmore » before reaching proliferative senescence. •Aging endothelial cells express more angiopoietin 1 and less angiopoietin 2 than young endothelial cells. •Migratory capacity is reduced in aging endothelial cells.« less

Transient CD15-positive endothelial phenotype in the human placenta correlates with physiological and pathological fetoplacental immaturity.

PubMed

Seidmann, L; Suhan, T; Unger, R; Gerein, V; Kirkpatrick, C J

2014-09-01

Placental growth and villous maturation are critical parameters of placental function at the end of pregnancy. A failure in these processes leads to the development of placental dysfunction, as well as fetal and neonatal mortality and morbidity. The aim of the study was to determine the relevant diagnostic markers associated with pathological placental development. Forty tissue samples from normal placentas of different gestational age and 68 pathological term placentas with defective villous maturation (GDM, idiopathic IUFD, preeclamsia, HELLP syndrome) comprised the comparative immunohistochemical study (CD15, CD45 and CD34). Positive immunohistochemical reactions were quantitatively assessed in the chorionic plate and vessels of the villi of different histological type. Physiologically immature placentas of the first and second trimester and pathologically immature term placentas were characterized by marked endothelial CD15-immunostaining. A significant loss of CD15-positive endothelium of the placentas was associated with a physiological and accelerated villous maturity. A spatio-temporal correlation was shown for CD15+ endothelial cells (ECs) and the number of CD45+ stromal cells (SCs). A negative temporal correlation was shown for CD15+ ECs and CD15+ myelomonocytes in the fetal blood. CD34 expression in the ECs was stable during the pregnancy. A correlation between a transient CD15-positive endothelial phenotype and a physiological and pathological fetoplacental immaturity was demonstrated. Physiological and accelerated placental maturation was accompanied by a significant disappearance of CD15-positive endothelium. We propose that "immature" CD15+ endothelium is an important diagnostic marker of the physiological and pathological fetoplacental immaturity. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Endothelial Progenitor Cells as a Sole Source for Ex Vivo Seeding of Tissue-Engineered Heart Valves

PubMed Central

Mettler, Bret A.; Engelmayr, George C.; Aikawa, Elena; Bischoff, Joyce; Martin, David P.; Exarhopoulos, Alexis; Moses, Marsha A.; Schoen, Frederick J.; Sacks, Michael S.

2010-01-01

Purposes: We investigated whether circulating endothelial progenitor cells (EPCs) can be used as a cell source for the creation of a tissue-engineered heart valve (TEHV). Methods: Trileaflet valved conduits were fabricated using nonwoven polyglycolic acid/poly-4-hydroxybutyrate polymer. Ovine peripheral blood EPCs were dynamically seeded onto a valved conduit and incubated for 7, 14, and 21 days. Results: Before seeding, EPCs were shown to express CD31+, eNOS+, and VE-Cadherin+ but not α-smooth muscle actin. Histological analysis demonstrated relatively homogenous cellular ingrowth throughout the valved conduit. TEHV constructs revealed the presence of endothelial cell (EC) markers and α-smooth muscle actin+ cells comparable with native valves. Protein levels were comparable with native valves and exceeded those in unseeded controls. EPC-TEHV demonstrated a temporal pattern of matrix metalloproteinases-2/9 expression and tissue inhibitors of metalloproteinase activities comparable to that of native valves. Mechanical properties of EPC-TEHV demonstrated significantly greater stiffness than that of the unseeded scaffolds and native valves. Conclusions: Circulating EPC appears to have the potential to provide both interstitial and endothelial functions and could potentially serve as a single-cell source for construction of autologous heart valves. PMID:19698056

Podoplanin-positive Cancer-associated Stromal Fibroblasts in Primary Tumor and Synchronous Lymph Node Metastases of HER2-overexpressing Breast Carcinomas.

PubMed

Niemiec, Joanna; Adamczyk, Agnieszka; Harazin-Lechowska, Agnieszka; Ambicka, Aleksandra; Grela-Wojewoda, Aleksandra; Majchrzyk, Kaja; Kruczak, Anna; Sas-Korczyńska, Beata; Ryś, Janusz

2018-04-01

We compared the status of stromal podoplanin-positive cancer-associated fibroblasts (ppCAFs) between primary tumors and paired synchronous lymph node metastases (LNMs) and analyzed the prognostic significance of tumoral ppCAFs in 203 patients with human epidermal growth factor receptor 2-positive breast carcinoma. ppCAFs were found in 167/203 and in 35/87 tumors and LNM, respectively. ppCAFs were most frequently found in tumors and corresponding LNM (n=52, 59.8%; p=0.001). However, for all LNMs (n=12) without ppCAFs, their paired tumors also lacked ppCAFs. In both tumors and LNMs, ppCAFs were α-smooth muscle actin-positive and cluster of differentiation 21 protein-negative, suggesting them not to be resident lymph node cells. Moreover, in our series, the presence of ppCAFs in tumors was borderline related to poor disease-free survival (p=0.058). These results speak in favor of a hypothesis suggesting ppCAFs accompany metastatic cancer cells migrating from tumor to LNMs. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Endocan, a putative endothelial cell marker, is elevated in preeclampsia, decreased in acute pyelonephritis, and unchanged in other obstetrical syndromes

PubMed Central

Adekola, Henry; Romero, Roberto; Chaemsaithong, Piya; Korzeniewski, Steven J.; Dong, Zhong; Yeo, Lami; Hassan, Sonia S.; Chaiworapongsa, Tinnakorn

2015-01-01

Abstract Objective: Endocan, a dermatan sulphate proteoglycan produced by endothelial cells, is considered a biomarker for endothelial cell activation/dysfunction. Preeclampsia is characterized by systemic vascular inflammation, and endothelial cell activation/dysfunction. Therefore, the objectives of this study were to determine whether: (1) plasma endocan concentrations in preeclampsia differ from those in uncomplicated pregnancies; (2) changes in plasma endocan concentration relate to the severity of preeclampsia, and whether these changes are specific or observed in other obstetrical syndromes such as small-for-gestational age (SGA), fetal death (FD), preterm labor (PTL) or preterm prelabor rupture of membranes (PROM); (3) a correlation exists between plasma concentration of endocan and angiogenic (placental growth factor or PlGF)/anti-angiogenic factors (soluble vascular endothelial growth factor receptor or sVEGFR-1, and soluble endoglin or sEng) among pregnancies complicated by preeclampsia; and (4) plasma endocan concentrations in patients with preeclampsia and acute pyelonephritis (both conditions in which there is endothelial cell activation) differ. Method: This cross-sectional study included the following groups: (1) uncomplicated pregnancy (n = 130); (2) preeclampsia (n = 102); (3) pregnant women without preeclampsia who delivered an SGA neonate (n = 51); (4) FD (n = 49); (5) acute pyelonephritis (AP; n = 35); (6) spontaneous PTL (n = 75); and (7) preterm PROM (n = 64). Plasma endocan concentrations were determined in all groups, and PIGF, sEng and VEGFR-1 plasma concentrations were measured by ELISA in the preeclampsia group. Results: (1) Women with preeclampsia had a significantly higher median plasma endocan concentration than those with uncomplicated pregnancies (p = 0.004); (2) among women with preeclampsia, the median plasma endocan concentration did not differ significantly according to disease severity (p = 0

Endothelial microparticles (EMP) for the assessment of endothelial function: an in vitro and in vivo study on possible interference of plasma lipids.

PubMed

van Ierssel, Sabrina H; Hoymans, Vicky Y; Van Craenenbroeck, Emeline M; Van Tendeloo, Viggo F; Vrints, Christiaan J; Jorens, Philippe G; Conraads, Viviane M

2012-01-01

Circulating endothelial microparticles (EMP) reflect the condition of the endothelium and are of increasing interest in cardiovascular and inflammatory diseases. Recently, increased numbers of EMP following oral fat intake, possibly due to acute endothelial injury, have been reported. On the other hand, the direct interference of lipids with the detection of EMP has been suggested. This study aimed to investigate the effect of lipid-rich solutions, commonly administered in clinical practice, on the detection, both in vitro and in vivo, of EMP. For the in vitro assessment, several lipid-rich solutions were added to whole blood of healthy subjects (n = 8) and patients with coronary heart disease (n = 5). EMP (CD31+/CD42b-) were detected in platelet poor plasma by flow cytometry. For the in vivo study, healthy volunteers were evaluated on 3 different study-days: baseline evaluation, following lipid infusion and after a NaCl infusion. EMP quantification, lipid measurements and peripheral arterial tonometry were performed on each day. Both in vitro addition and in vivo administration of lipids significantly decreased EMP (from 198.6 to 53.0 and from 272.6 to 90.6/µl PPP, respectively, p = 0.001 and p = 0.012). The EMP number correlated inversely with the concentration of triglycerides, both in vitro and in vivo (r = -0.707 and -0.589, p<0.001 and p = 0.021, respectively). The validity of EMP as a marker of endothelial function is supported by their inverse relationship with the reactive hyperemia index (r = -0.758, p = 0.011). This inverse relation was confounded by the intravenous administration of lipids. The confounding effect of high circulating levels of lipids, commonly found in patients that receive intravenous lipid-based solutions, should be taken into account when flow cytometry is used to quantify EMP.

Lymphangioma circumscriptum, angiokeratoma, or superficial vascular ectasia with epithelial hyperplasia?

PubMed

Katsoulas, Nikolaos; Tosios, Konstantinos I; Argyris, Prokopios; Koutlas, Ioannis G; Sklavounou, Alexandra

2014-08-01

We report a case of lymphangioma circumscriptum (cavernous lymphangioma with epithelial hyperplasia) in a 12-year-old girl, presenting as a papillary tumor on the right dorsal side of her tongue. Microscopic examination found cavernous vascular channels lined by a single layer of CD31(+), podoplanin-positive, CD34(-) endothelial cells that occupied the papillary lamina propria and were accompanied by epithelial hyperplasia. A review of the literature on oral vascular tumors with epithelial hyperplasia, namely, lymphangioma circumscriptum and angiokeratoma, provided information that draws into question the use of these terms. Copyright © 2014 Elsevier Inc. All rights reserved.

Oral cancer/endothelial cell fusion experiences nuclear fusion and acquisition of enhanced survival potential.

PubMed

Song, Kai; Song, Yong; Zhao, Xiao-Ping; Shen, Hui; Wang, Meng; Yan, Ting-Lin; Liu, Ke; Shang, Zheng-Jun

2014-10-15

Most previous studies have linked cancer-macrophage fusion with tumor progression and metastasis. However, the characteristics of hybrid cells derived from oral cancer and endothelial cells and their involvement in cancer remained unknown. Double-immunofluorescent staining and fluorescent in situ hybridization (FISH) were performed to confirm spontaneous cell fusion between eGFP-labeled human umbilical vein endothelial cells (HUVECs) and RFP-labeled SCC9, and to detect the expression of vementin and cytokeratin 18 in the hybrids. The property of chemo-resistance of such hybrids was examined by TUNEL assay. The hybrid cells in xenografted tumor were identified by FISH and GFP/RFP dual-immunofluoresence staining. We showed that SCC9 cells spontaneously fused with cocultured endothelial cells, and the resultant hybrid cells maintained the division and proliferation activity after re-plating and thawing. Such hybrids expressed markers of both parental cells and became more resistant to chemotherapeutic drug cisplatin as compared to the parental SCC9 cells. Our in vivo data indicated that the hybrid cells contributed to tumor composition by using of immunostaining and FISH analysis, even though the hybrid cells and SCC9 cells were mixed with 1:10,000, according to the FACS data. Our study suggested that the fusion events between oral cancer and endothelial cells undergo nuclear fusion and acquire a new property of drug resistance and consequently enhanced survival potential. These experimental findings provide further supportive evidence for the theory that cell fusion is involved in cancer progression. Copyright © 2014 Elsevier Inc. All rights reserved.

Endothelial Heparan Sulfate 6-O-Sulfation Levels Regulate Angiogenic Responses of Endothelial Cells to Fibroblast Growth Factor 2 and Vascular Endothelial Growth Factor*

PubMed Central

Ferreras, Cristina; Rushton, Graham; Cole, Claire L.; Babur, Muhammad; Telfer, Brian A.; van Kuppevelt, Toin H.; Gardiner, John M.; Williams, Kaye J.; Jayson, Gordon C.; Avizienyte, Egle

2012-01-01

Fibroblast growth factor 2 (FGF2) and vascular endothelial growth factor 165 (VEGF165) are potent pro-angiogenic growth factors that play a pivotal role in tumor angiogenesis. The activity of these growth factors is regulated by heparan sulfate (HS), which is essential for the formation of FGF2/FGF receptor (FGFR) and VEGF165/VEGF receptor signaling complexes. However, the structural characteristics of HS that determine activation or inhibition of such complexes are only partially defined. Here we show that ovarian tumor endothelium displays high levels of HS sequences that harbor glucosamine 6-O-sulfates when compared with normal ovarian vasculature where these sequences are also detected in perivascular area. Reduced HS 6-O-sulfotransferase 1 (HS6ST-1) or 6-O-sulfotransferase 2 (HS6ST-2) expression in endothelial cells impacts upon the prevalence of HS 6-O-sulfate moieties in HS sequences, which consist of repeating short, highly sulfated S domains interspersed by transitional N-acetylated/N-sulfated domains. 1–40% reduction in 6-O-sulfates significantly compromises FGF2- and VEGF165-induced endothelial cell sprouting and tube formation in vitro and FGF2-dependent angiogenesis in vivo. Moreover, HS on wild-type neighboring endothelial or smooth muscle cells fails to restore endothelial cell sprouting and tube formation. The affinity of FGF2 for HS with reduced 6-O-sulfation is preserved, although FGFR1 activation is inhibited correlating with reduced receptor internalization. These data show that 6-O-sulfate moieties in endothelial HS are of major importance in regulating FGF2- and VEGF165-dependent endothelial cell functions in vitro and in vivo and highlight HS6ST-1 and HS6ST-2 as potential targets of novel antiangiogenic agents. PMID:22927437

Endothelial nitric oxide synthase is dynamically expressed during bone marrow stem cell differentiation into endothelial cells.

PubMed

Liu, Zhenguo; Jiang, Yuehua; Hao, Hong; Gupta, Kalpna; Xu, Jian; Chu, Ling; McFalls, Edward; Zweier, Jay; Verfaillie, Catherine; Bache, Robert J

2007-09-01

This study was designed to investigate the developmental expression of endothelial nitric oxide synthase (eNOS) during stem cell differentiation into endothelial cells and to examine the functional status of the newly differentiated endothelial cells. Mouse adult multipotent progenitor cells (MAPCs) were used as the source of stem cells and were induced to differentiate into endothelial cells with vascular endothelial growth factor (VEGF) in serum-free medium. Expression of eNOS in the cells during differentiation was evaluated with real-time PCR, nitric oxide synthase (NOS) activity, and Western blot analysis. It was found that eNOS, but no other NOS, was present in undifferentiated MAPCs. eNOS expression disappeared in the cells immediately after induction of differentiation. However, eNOS expression reoccurred at day 7 during differentiation. Increasing eNOS mRNA, protein content, and activity were observed in the cells at days 14 and 21 during differentiation. The differentiated endothelial cells formed dense capillary networks on growth factor-reduced Matrigel. VEGF-stimulated phosphorylation of extracellular signal-regulated kinase (ERK)-1 and ERK-2 occurred in these cells, which was inhibited by NOS inhibitor N(G)-nitro-L-arginine methyl ester. In conclusion, these data demonstrate that eNOS is present in MAPCs and is dynamically expressed during the differentiation of MAPCs into endothelial cells in vitro.

Soluble fms-Like Tyrosine Kinase 1 as a Link Between Angiogenesis and Endothelial Dysfunction in Pediatric Patients With β-Thalassemia Intermedia.

PubMed

Tantawy, Azza Abdel Gawad; Adly, Amira Abdel Moneam; Ismail, Eman Abdel Rahman; Youssef, Omneya Ibrahim; Ali, Mohamed ElSayed

2017-11-01

Endothelial damage has been implicated in the pathogenesis of vascular complications in β-thalassemia intermedia (β-TI). Soluble fms-like tyrosine kinase 1 (sFLT-1) is a member of the vascular endothelial growth factor receptor (VEGFR) family. Soluble fms-like tyrosine kinase 1 is an antiangiogenic protein that induces endothelial dysfunction by adhering to and inhibiting VEGF and placenta growth factor. The aim of this study was to assess the level of sFLT-1 in 35 children and adolescents with β-TI, correlating it with markers of hemolysis and iron overload as well as cardiopulmonary complications. Patients were studied focusing on the history of cardiac disease, splenectomy, transfusion, chelation/hydroxyurea therapy, serum ferritin, and sFLT-1 levels. Echocardiography and measurement of carotid intima-media thickness (CIMT) were done for all participants. Soluble fms-like tyrosine kinase 1 was significantly higher in TI patients compared to the control group (median [interquartile range], 110 [80-155] pg/mL versus 70 [60-90] pg/mL; P < .001). Splenectomized patients and those who had pulmonary hypertension risk or heart disease had higher sFLT-1 levels than those without ( P < .001). The sFLT-1 cutoff value that differentiates patients with and without pulmonary hypertension risk or heart disease was determined. Soluble fms-like tyrosine kinase 1 was lower among patients who received chelation therapy and/or hydroxyurea. Significant positive relations were observed between sFLT-1 and lactate dehydrogenase, serum ferritin, liver iron concentration, tricuspid regurgitant jet velocity, and CIMT. We suggest that sFLT-1 represents a link between angiogenesis, endothelial dysfunction, and subclinical atherosclerosis. Measurement of sFLT-1 as a marker of vascular dysfunction in β-TI may provide utility for early identification of patients at increased risk of cardiopulmonary complications.

Screening and Characterization of Drugs That Protect Corneal Endothelial Cells Against Unfolded Protein Response and Oxidative Stress

PubMed Central

Kim, Eun Chul; Toyono, Tetsuya; Berlinicke, Cynthia A.; Zack, Donald J.; Jurkunas, Ula; Usui, Tomohiko; Jun, Albert S.

2017-01-01

Purpose To screen for and characterize compounds that protect corneal endothelial cells against unfolded protein response (UPR) and oxidative stress. Methods Bovine corneal endothelial cells (BCECs) were treated for 48 hours with 640 compounds from a Food and Drug Administration (FDA)-approved drug library and then challenged with thapsigargin or H2O2 to induce UPR or oxidative stress, respectively. Cell viability was measured using the CellTiter-Glo survival assay. Selected “hits” were subjected to further dose-response testing, and their ability to modulate expression of UPR and oxidative stress markers was assessed by RT-PCR, Western blot, and measurement of protein carbonyl and 8-hydroxydeoxyguanosine (8-OHdG) adducts in immortalized human corneal endothelial cells (iHCECs). Results Forty-one drugs at 20 μM and 55 drugs at 100 μM increased survival of H2O2-challenged cells, and 8 drugs at 20 μM and 2 drugs at 100 μM increased survival of thapsigargin-challenged cells, compared with untreated control cells. Nicergoline, ergothioneine, nimesulide, oxotremorine, and mefenamic acid increased survival of both H2O2- and thapsigargin-challenged cells. Oxotremorine altered DNA damage inducible 3 (CHOP) gene expression, glucose-regulated protein 78 kDa (GRP78) and activating transcription factor 4 (ATF4) protein expression, and protein carbonyl and 8-OHdG levels. Mefenamic acid altered GRP78 protein expression and protein carbonyl and 8-OHdG levels. Conclusions Oxotremorine and mefenamic acid are potential survival factors for corneal endothelial cells under UPR and oxidative stress. The described assay can be further expanded to screen additional drugs for potential therapeutic effect in corneal endothelial diseases such as Fuchs' endothelial corneal dystrophy. PMID:28159976

Expression of synaptopodin in endothelial cells exposed to laminar shear stress and its role in endothelial wound healing.

PubMed

Mun, Gyeong In; Park, Soojin; Kremerskothen, Joachim; Boo, Yong Chool

2014-03-18

We examined the hypothesis that certain actin binding proteins might be upregulated by laminar shear stress (LSS) and could contribute to endothelial wound healing. Analysis of mRNA expression profiles of human umbilical vein endothelial cells under static and LSS-exposed conditions provided a list of LSS-induced actin binding proteins including synaptopodin (SYNPO) whose endothelial expression has not been previously reported. Additional studies demonstrated that SYNPO is a key mediator of endothelial wound healing because small interfering RNA-mediated suppression of SYNPO attenuated wound closure under LSS whereas overexpression of exogenous SYNPO enhanced endothelial wound closure in the absence of LSS. This study suggests that LSS-induced actin binding proteins including SYNPO may play a critical role in the endothelial wound healing stimulated by LSS. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

[Coronary disease extension determines mobilization of endothelial progenitor cells and cytokines after a first myocardial infarction with ST elevation].

PubMed

Jiménez-Navarro, Manuel F; González, Francisco Jesús; Caballero-Borrego, Juan; Marchal, Juan Antonio; Rodríguez-Losada, Noela; Carrillo, Esmeralda; García-Pinilla, José Manuel; Hernández-García, José M; Pérez-González, Rita; Ramírez, Gemma; Aránega, Antonia; de Teresa Galván, Eduardo

2011-12-01

Multivessel coronary disease is still a postinfarction prognostic marker despite new forms of reperfusion, such as primary angioplasty. The aim of this study was to determine the time sequence of various sets of endothelial progenitor cells and angiogenic cytokines (vascular endothelial growth factor, hepatocyte growth factor) according to the degree of extension of the postinfarction coronary disease. We studied the release kinetics in 32 patients admitted for a first myocardial infarction with ST elevation, grouped according to whether they had single or multivessel disease, and 26 controls. The patients had a higher number of endothelial progenitor cells and angiogenic cytokines than the controls at all 3 measurements (admission, day 3, and day 7) of the following subsets: CD34, CD34+CD133+, CD34+KDR+, and CD34+CD133+KDR+CD45+(weak); this latter was higher on day 7. The levels of these cell subsets were all higher in the patients with single-vessel disease and at all 3 measurements. The vascular endothelial growth factor levels were raised during the first week and the hepatocyte growth factor showed an early peak on admission for infarction. No significant differences were seen in the cytokines according to coronary disease extension. Although the release kinetics of different subsets of endothelial progenitor cells in patients with a first acute myocardial infarction with ST elevation was similar in those with single vessel disease to those with multivessel disease, the number of circulating endothelial progenitor cells was greater in the patients with single vessel disease. The vascular endothelial growth factor levels were raised during the first postinfarction week and the hepatocyte growth factor were higher on admission. Copyright © 2011 Sociedad Española de Cardiología. Published by Elsevier Espana. All rights reserved.

Heat Shock Protein 70 Prevents Hyperoxia-Induced Disruption of Lung Endothelial Barrier via Caspase-Dependent and AIF-Dependent Pathways

PubMed Central

Kondrikov, Dmitry; Fulton, David; Dong, Zheng; Su, Yunchao

2015-01-01

Exposure of pulmonary artery endothelial cells (PAECs) to hyperoxia results in a compromise in endothelial monolayer integrity, an increase in caspase-3 activity, and nuclear translocation of apoptosis-inducing factor (AIF), a marker of caspase-independent apoptosis. In an endeavor to identify proteins involved in hyperoxic endothelial injury, we found that the protein expression of heat-shock protein 70 (Hsp70) was increased in hyperoxic PAECs. The hyperoxia-induced Hsp70 protein expression is from hspA1B gene. Neither inhibition nor overexpression of Hsp70 affected the first phase barrier disruption of endothelial monolayer. Nevertheless, inhibition of Hsp70 by using the Hsp70 inhibitor KNK437 or knock down Hsp70 using siRNA exaggerated and overexpression of Hsp70 prevented the second phase disruption of lung endothelial integrity. Moreover, inhibition of Hsp70 exacerbated and overexpression of Hsp70 prevented hyperoxia-induced apoptosis, caspase-3 activation, and increase in nuclear AIF protein level in PAECs. Furthermore, we found that Hsp70 interacted with AIF in the cytosol in hyperoxic PAECs. Inhibition of Hsp70/AIF association by KNK437 correlated with increased nuclear AIF level and apoptosis in KNK437-treated PAECs. Finally, the ROS scavenger NAC prevented the hyperoxia-induced increase in Hsp70 expression and reduced the interaction of Hsp70 with AIF in hyperoxic PAECs. Together, these data indicate that increased expression of Hsp70 plays a protective role against hyperoxia-induced lung endothelial barrier disruption through caspase-dependent and AIF-dependent apoptotic pathways. Association of Hsp70 with AIF prevents AIF nuclear translocation, contributing to the protective effect of Hsp70 on hyperoxia-induced endothelial apoptosis. The hyperoxia-induced increase in Hsp70 expression and Hsp70/AIF interaction is contributed to ROS formation. PMID:26066050

Acute effects of high-fat meals enriched with walnuts or olive oil on postprandial endothelial function.

PubMed

Cortés, Berenice; Núñez, Isabel; Cofán, Montserrat; Gilabert, Rosa; Pérez-Heras, Ana; Casals, Elena; Deulofeu, Ramón; Ros, Emilio

2006-10-17

We sought to investigate whether the addition of walnuts or olive oil to a fatty meal have differential effects on postprandial vasoactivity, lipoproteins, markers of oxidation and endothelial activation, and plasma asymmetric dimethylarginine (ADMA). Compared with a Mediterranean diet, a walnut diet has been shown to improve endothelial function in hypercholesterolemic patients. We hypothesized that walnuts would reverse postprandial endothelial dysfunction associated with consumption of a fatty meal. We randomized in a crossover design 12 healthy subjects and 12 patients with hypercholesterolemia to 2 high-fat meal sequences to which 25 g olive oil or 40 g walnuts had been added. Both test meals contained 80 g fat and 35% saturated fatty acids, and consumption of each meal was separated by 1 week. Venipunctures and ultrasound measurements of brachial artery endothelial function were performed after fasting and 4 h after test meals. In both study groups, flow-mediated dilation (FMD) was worse after the olive oil meal than after the walnut meal (p = 0.006, time-period interaction). Fasting, but not postprandial, triglyceride concentrations correlated inversely with FMD (r = -0.324; p = 0.024). Flow-independent dilation and plasma ADMA concentrations were unchanged, and the concentration of oxidized low-density lipoproteins decreased (p = 0.051) after either meal. The plasma concentrations of soluble inflammatory cytokines and adhesion molecules decreased (p < 0.01) independently of meal type, except for E-selectin, which decreased more (p = 0.033) after the walnut meal. Adding walnuts to a high-fat meal acutely improves FMD independently of changes in oxidation, inflammation, or ADMA. Both walnuts and olive oil preserve the protective phenotype of endothelial cells.

Extracellular signal-regulated kinase activation and endothelin-1 production in human endothelial cells exposed to vibration

PubMed Central

White, Charles R; Haidekker, Mark A; Stevens, Hazel Y; Frangos, John A

2004-01-01

Hand–arm vibration syndrome is a vascular disease of occupational origin and a form of secondary Raynaud's phenomenon. Chronic exposure to hand-held vibrating tools may cause endothelial injury. This study investigates the biomechanical forces involved in the transduction of fluid vibration in the endothelium. Human endothelial cells were exposed to direct vibration and rapid low-volume fluid oscillation. Rapid low-volume fluid oscillation was used to simulate the effects of vibration by generating defined temporal gradients in fluid shear stress across an endothelial monolayer. Extracellular signal-regulated kinase (ERK1/2) phosphorylation and endothelin-1 (ET-1) release were monitored as specific biochemical markers for temporal gradients and endothelial response, respectively. Both vibrational methods were found to phosphorylate ERK1/2 in a similar pattern. At a fixed frequency of fluid oscillation where the duration of each pulse cycle remained constant, ERK1/2 phosphorylation increased with the increasing magnitude of the applied temporal gradient. However, when the frequency of flow oscillation was increased (thus decreasing the duration of each pulse cycle), ERK1/2 phosphorylation was attenuated across all temporal gradient flow profiles. Fluid oscillation significantly stimulated ET-1 release compared to steady flow, and endothelin-1 was also attenuated with the increase in oscillation frequency. Taken together, these results show that both the absolute magnitude of the temporal gradient and the frequency/duration of each pulse cycle play a role in the biomechanical transduction of fluid vibrational forces in endothelial cells. Furthermore, this study reports for the first time a link between the ERK1/2 signal transduction pathway and transmission of vibrational forces in the endothelium. PMID:14724194

Sustained apnea induces endothelial activation.

PubMed

Eichhorn, Lars; Dolscheid-Pommerich, Ramona; Erdfelder, Felix; Ayub, Muhammad Ajmal; Schmitz, Theresa; Werner, Nikos; Jansen, Felix

2017-09-01

Apnea diving has gained worldwide popularity, even though the pathophysiological consequences of this challenging sport on the human body are poorly investigated and understood. This study aims to assess the influence of sustained apnea in healthy volunteers on circulating microparticles (MPs) and microRNAs (miRs), which are established biomarkers reflecting vascular function. Short intermittent hypoxia due to voluntary breath-holding affects circulating levels of endothelial cell-derived MPs (EMPs) and endothelial cell-derived miRs. Under dry laboratory conditions, 10 trained apneic divers performed maximal breath-hold. Venous blood samples were taken, once before and at 4 defined points in time after apnea. Samples were analyzed for circulating EMPs and endothelial miRs. Average apnea time was 329 seconds (±103), and SpO 2 at the end of apnea was 79% (±12). Apnea was associated with a time-dependent increase of circulating endothelial cell-derived EMPs and endothelial miRs. Levels of circulating EMPs in the bloodstream reached a peak 4 hours after the apnea period and returned to baseline levels after 24 hours. Circulating miR-126 levels were elevated at all time points after a single voluntary maximal apnea, whereas miR-26 levels were elevated significantly only after 30 minutes and 4 hours. Also miR-21 and miR-92 levels increased, but did not reach the level of significance. Even a single maximal breath-hold induces acute endothelial activation and should be performed with great caution by subjects with preexisting vascular diseases. Voluntary apnea might be used as a model to simulate changes in endothelial function caused by hypoxia in humans. © 2017 Wiley Periodicals, Inc.

Neutrophil-endothelial cell interactions on endothelial monolayers grown on micropore filters.

PubMed Central

Taylor, R F; Price, T H; Schwartz, S M; Dale, D C

1981-01-01

We have developed a technique for growing endothelial monolayers on micropore filters. These monolayers demonstrate confluence by phase and electron microscopy and provide a functional barrier to passage of radiolabeled albumin. Neutrophils readily penetrate the monolayer in response to chemotaxin, whereas there is little movement in the absence of chemotaxin. This system offers unique advantages over available chemotaxis assays and may have wider applications in the study of endothelial function. Images PMID:7007441

Markers of endothelial cell activation and immune activation are increased in patients with severe leptospirosis and associated with disease severity

USDA-ARS?s Scientific Manuscript database

Objectives: Previous studies concluded that haemorrhage is one of the most accurate prognostic factors of mortality in leptospirosis. Therefore, endothelial cell activation was investigated in relation to disease severity in severe leptospirosis. Methods: Prospective cohort study of severe leptospi...

Endothelial cell-derived exosomes protect SH-SY5Y nerve cells against ischemia/reperfusion injury.

PubMed

Xiao, Bing; Chai, Yi; Lv, Shigang; Ye, Minhua; Wu, Miaojing; Xie, Liyuan; Fan, Yanghua; Zhu, Xingen; Gao, Ziyun

2017-10-01

Cerebral ischemia is a leading cause of death and disability. A previous study indicated that remote ischemic postconditioning (RIP) in the treatment of cerebral ischemia reduces ischemia/reperfusion (I/R) injury. However, the underlying mechanism is not well understood. In the present study, the authors hypothesized that the protective effect of RIP on neurological damage is mediated by exosomes that are released by endothelial cells in femoral arteries. To test this, right middle cerebral artery occlusion/reperfusion with RIP was performed in rats. In addition, an I/R injury cell model was tested that included human umbilical vein endothelial cells (HUVECs) and SH-SY5Y cells. Both the in vivo and in vitro models were examined for injury. Markers of exosomes (CD63, HSP70 and TSG101) were assessed by immunohistochemistry, western blot analysis and flow cytometry. Exosomes were extracted from both animal serum and HUVEC culture medium and identified by electron microscopy. They investigated the role of endothelial cell-derived exosomes in the proliferation, apoptosis, cell cycle, migration and invasion of I/R-injured SH-SY5Y cells. In addition, apoptosis-related molecules caspase-3, Bax and Bcl-2 were detected. RIP was determined to increase the number of exosomes and the expression levels of CD63, HSP70 and TSG101 in plasma, but not in brain hippocampal tissue. The size of exosomes released after I/R in HUVECs was similar to the size of exosomes released in rats subjected to RIP. Endothelial cell-derived exosomes partly suppressed the I/R-induced cell cycle arrest and apoptosis, and inhibited cell proliferation, migration and invasion in SH-SY5Y nerve cells. Endothelial cell-derived exosomes directly protect nerve cells against I/R injury, and are responsible for the protective role of RIP in I/R.

The Notch ligand Delta-like 4 negatively regulates endothelial tip cell formation and vessel branching

PubMed Central

Suchting, Steven; Freitas, Catarina; le Noble, Ferdinand; Benedito, Rui; Bréant, Christiane; Duarte, Antonio; Eichmann, Anne

2007-01-01

Delta-like 4 (Dll4) is a transmembrane ligand for Notch receptors that is expressed in arterial blood vessels and sprouting endothelial cells. Here we show that Dll4 regulates vessel branching during development by inhibiting endothelial tip cell formation. Heterozygous deletion of dll4 or pharmacological inhibition of Notch signaling using γ-secretase inhibitor revealed a striking vascular phenotype, with greatly increased numbers of filopodia-extending endothelial tip cells and increased expression of tip cell marker genes compared with controls. Filopodia extension in dll4+/− retinal vessels required the vascular growth factor VEGF and was inhibited when VEGF signaling was blocked. Although VEGF expression was not significantly altered in dll4+/− retinas, dll4+/− vessels showed increased expression of VEGF receptor 2 and decreased expression of VEGF receptor 1 compared with wild-type, suggesting they could be more responsive to VEGF stimulation. In addition, expression of dll4 in wild-type tip cells was itself decreased when VEGF signaling was blocked, indicating that dll4 may act downstream of VEGF as a “brake” on VEGF-mediated angiogenic sprouting. Taken together, these data reveal Dll4 as a negative regulator of vascular sprouting and vessel branching that is required for normal vascular network formation during development. PMID:17296941

The Notch ligand Delta-like 4 negatively regulates endothelial tip cell formation and vessel branching.

PubMed

Suchting, Steven; Freitas, Catarina; le Noble, Ferdinand; Benedito, Rui; Bréant, Christiane; Duarte, Antonio; Eichmann, Anne

2007-02-27

Delta-like 4 (Dll4) is a transmembrane ligand for Notch receptors that is expressed in arterial blood vessels and sprouting endothelial cells. Here we show that Dll4 regulates vessel branching during development by inhibiting endothelial tip cell formation. Heterozygous deletion of dll4 or pharmacological inhibition of Notch signaling using gamma-secretase inhibitor revealed a striking vascular phenotype, with greatly increased numbers of filopodia-extending endothelial tip cells and increased expression of tip cell marker genes compared with controls. Filopodia extension in dll4(+/-) retinal vessels required the vascular growth factor VEGF and was inhibited when VEGF signaling was blocked. Although VEGF expression was not significantly altered in dll4(+/-) retinas, dll4(+/-) vessels showed increased expression of VEGF receptor 2 and decreased expression of VEGF receptor 1 compared with wild-type, suggesting they could be more responsive to VEGF stimulation. In addition, expression of dll4 in wild-type tip cells was itself decreased when VEGF signaling was blocked, indicating that dll4 may act downstream of VEGF as a "brake" on VEGF-mediated angiogenic sprouting. Taken together, these data reveal Dll4 as a negative regulator of vascular sprouting and vessel branching that is required for normal vascular network formation during development.

Endothelial keratoplasty with infant donor tissue

PubMed Central

Kobayashi, Akira; Yokogawa, Hideaki; Yamazaki, Natsuko; Masaki, Toshinori; Sugiyama, Kazuhisa

2014-01-01

Here we report a case of endothelial keratoplasty with infant donor tissue obtained after brain death. A 52-year-old man with endothelial dysfunction of unknown cause in the right eye underwent non-Descemet stripping automated endothelial keratoplasty (nDSAEK) with tissue from an infant donor (2 years). Intraoperative and postoperative complications were recorded. Best corrected visual acuity and donor central endothelial cell density were recorded preoperatively and postoperatively. Infant donor tissue preparation with a microkeratome set at 300 μm was successful; the donor tissue was extremely elastic and soft compared with adult tissue. The central endothelial cell density of the infant donor tissue was as high as 4,291 cells/mm2. No complications were observed during donor tissue (8.0 mm in diameter) insertion with the double-glide technique (Busin glide with intraocular lens sheet glide) or any of the other procedures. Best corrected visual acuity improved from 1.7 logMAR (logarithm of the minimum angle of resolution; 0.02 decimal visual acuity) preoperatively to 0.2 logMAR (0.6) after 6 months and 0.1 logMAR (0.8) after 1 year. The central endothelial cell density after 6 months was 4,098 cells/mm2 (representing a 4.5% cell loss from preoperative donor cell measurements), and the central endothelial cell density after 1 year was 4,032 cells/mm2 (6.0% decrease). Infant donor tissue may be preferably used for DSAEK/nDASEK, since it may not be suitable for penetrating keratoplasty or Descemet membrane endothelial keratoplasty. PMID:25246761

The research on endothelial function in women and men at risk for cardiovascular disease (REWARD) study: methodology

PubMed Central

2011-01-01

Background Endothelial function has been shown to be a highly sensitive marker for the overall cardiovascular risk of an individual. Furthermore, there is evidence of important sex differences in endothelial function that may underlie the differential presentation of cardiovascular disease (CVD) in women relative to men. As such, measuring endothelial function may have sex-specific prognostic value for the prediction of CVD events, thus improving risk stratification for the overall prediction of CVD in both men and women. The primary objective of this study is to assess the clinical utility of the forearm hyperaemic reactivity (FHR) test (a proxy measure of endothelial function) for the prediction of CVD events in men vs. women using a novel, noninvasive nuclear medicine -based approach. It is hypothesised that: 1) endothelial dysfunction will be a significant predictor of 5-year CVD events independent of baseline stress test results, clinical, demographic, and psychological variables in both men and women; and 2) endothelial dysfunction will be a better predictor of 5-year CVD events in women compared to men. Methods/Design A total of 1972 patients (812 men and 1160 women) undergoing a dipyridamole stress testing were recruited. Medical history, CVD risk factors, health behaviours, psychological status, and gender identity were assessed via structured interview or self-report questionnaires at baseline. In addition, FHR was assessed, as well as levels of sex hormones via blood draw. Patients will be followed for 5 years to assess major CVD events (cardiac mortality, non-fatal MI, revascularization procedures, and cerebrovascular events). Discussion This is the first study to determine the extent and nature of any sex differences in the ability of endothelial function to predict CVD events. We believe the results of this study will provide data that will better inform the choice of diagnostic tests in men and women and bring the quality of risk stratification in

Habitually exercising older men do not demonstrate age-associated vascular endothelial oxidative stress

PubMed Central

Pierce, Gary L.; Donato, Anthony J.; LaRocca, Thomas J.; Eskurza, Iratxe; Silver, Annemarie E.; Seals, Douglas R.

2011-01-01

SUMMARY We tested the hypothesis that older men who perform habitual aerobic exercise do not demonstrate age-associated vascular endothelial oxidative stress compared with their sedentary peers. Older exercising men (n=13, 62±2 y) had higher (P<0.05) physical activity (79±7 vs. 30±6 MET h/wk) and maximal exercise oxygen consumption (42±1 vs. 29±1 ml/kg/min) vs. sedentary men (n=28, 63±1 y). Brachial artery flow-mediated dilation, a measure of vascular endothelial function, was greater (P<0.05) in the exercising vs. sedentary older men (6.3±0.5 vs. 4.9±0.4 %Δ) and not different than young controls (n=20, 25±1 y, 7.1 ± 0.5 %Δ). In vascular endothelial cells sampled from the brachial artery, nitrotyrosine, a marker of oxidative stress, was 51% lower in the exercising vs. sedentary older men (0.38±0.06 vs. 0.77±0.10 AU). This was associated with lower endothelial expression of the oxidant enzyme nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase (p47phox subunit, 0.33±0.05 vs. 0.61±0.09 AU) and the redox-sensitive transcription factor nuclear factor kappa B (NFκB) (p65 subunit, 0.36±0.05 vs. 0.72±0.09 AU). Expression of the antioxidant enzyme manganese superoxide dismutase (SOD) (0.57±0.13 vs. 0.30±0.04 AU) and activity of endothelium-bound extracellular SOD were greater (6.4±0.5 vs. 5.0±0.6 U/ml/min) in the exercising men (both P<0.05), but differences no longer were significant after correcting for adiposity and circulating metabolic factors. Overall, values for the young controls differed with those for the sedentary, but not the exercising older men. Older men who exercise regularly do not demonstrate vascular endothelial oxidative stress and this may be a key molecular mechanism underlying their reduced risk of cardiovascular diseases. PMID:21943306

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.

Functional Definition of Progenitors Versus Mature Endothelial Cells Reveals Key SoxF-Dependent Differentiation Process.

PubMed

Patel, Jatin; Seppanen, Elke J; Rodero, Mathieu P; Wong, Ho Yi; Donovan, Prudence; Neufeld, Zoltan; Fisk, Nicholas M; Francois, Mathias; Khosrotehrani, Kiarash

2017-02-21

During adult life, blood vessel formation is thought to occur via angiogenic processes involving branching from existing vessels. An alternate proposal suggests that neovessels form from endothelial progenitors able to assemble the intimal layers. We here aimed to define vessel-resident endothelial progenitors in vivo in a variety of tissues in physiological and pathological situations such as normal aorta, lungs, and wound healing, tumors, and placenta, as well. Based on protein expression levels of common endothelial markers using flow cytometry, 3 subpopulations of endothelial cells could be identified among VE-Cadherin+ and CD45- cells. Lineage tracing by using Cdh5cre ERt2 /Rosa-YFP reporter strategy demonstrated that the CD31-/loVEGFR2lo/intracellular endothelial population was indeed an endovascular progenitor (EVP) of an intermediate CD31intVEGFR2lo/intracellular transit amplifying (TA) and a definitive differentiated (D) CD31hiVEGFR2hi/extracellular population. EVP cells arose from vascular-resident beds that could not be transferred by bone marrow transplantation. Furthermore, EVP displayed progenitor-like status with a high proportion of cells in a quiescent cell cycle phase as assessed in wounds, tumors, and aorta. Only EVP cells and not TA and D cells had self-renewal capacity as demonstrated by colony-forming capacity in limiting dilution and by transplantation in Matrigel plugs in recipient mice. RNA sequencing revealed prominent gene expression differences between EVP and D cells. In particular, EVP cells highly expressed genes related to progenitor function including Sox9 , Il33 , Egfr , and Pdfgrα. Conversely, D cells highly expressed genes related to differentiated endothelium including Ets1&2 , Gata2 , Cd31 , Vwf , and Notch . The RNA sequencing also pointed to an essential role of the Sox18 transcription factor. The role of SOX18 in the differentiation process was validated by using lineage-tracing experiments based on S ox18Cre ERt2 /Rosa

Carbohydrates and Endothelial Function: Is a Low-Carbohydrate Diet or a Low-Glycemic Index Diet Favourable for Vascular Health?

PubMed Central

Jovanovski, Elena; Zurbau, Andreea

2015-01-01

Low-carbohydrate diets have become increasingly popular in both media and clinical research settings. Although they may improve some metabolic markers, their effects on arterial function remain unclear. Endothelial dysfunction is the well-established response to cardiovascular risk factors and a pivotal feature that precedes atherosclerotic diseases. It has been demonstrated that a high carbohydrate-induced hyperglycemia and subsequent oxidative stress acutely worsen the efficacy of the endothelial vasodilatory system. Thus, in theory, a carbohydrate restricted diet may preserve the integrity of the arterial system. This review attempts to provide insight on whether low-carbohydrate diets have a favorable or detrimental impact on vascular function, or it is perhaps the quality of carbohydrate that should direct dietary recommendations. Research to date suggests that diets low in carbohydrate amount may negatively impact vascular endothelial function. Conversely, it appears that maintaining recommended carbohydrate intake with utilization of low glycemic index foods generates a more favorable vascular profile. Understanding these relationships will aid in deciphering the diverging role of modulating quantity and quality of carbohydrates on cardiovascular risk. PMID:25954727

Carbohydrates and endothelial function: is a low-carbohydrate diet or a low-glycemic index diet favourable for vascular health?

PubMed

Jovanovski, Elena; Zurbau, Andreea; Vuksan, Vladimir

2015-04-01

Low-carbohydrate diets have become increasingly popular in both media and clinical research settings. Although they may improve some metabolic markers, their effects on arterial function remain unclear. Endothelial dysfunction is the well-established response to cardiovascular risk factors and a pivotal feature that precedes atherosclerotic diseases. It has been demonstrated that a high carbohydrate-induced hyperglycemia and subsequent oxidative stress acutely worsen the efficacy of the endothelial vasodilatory system. Thus, in theory, a carbohydrate restricted diet may preserve the integrity of the arterial system. This review attempts to provide insight on whether low-carbohydrate diets have a favorable or detrimental impact on vascular function, or it is perhaps the quality of carbohydrate that should direct dietary recommendations. Research to date suggests that diets low in carbohydrate amount may negatively impact vascular endothelial function. Conversely, it appears that maintaining recommended carbohydrate intake with utilization of low glycemic index foods generates a more favorable vascular profile. Understanding these relationships will aid in deciphering the diverging role of modulating quantity and quality of carbohydrates on cardiovascular risk.

Anthrax Toxin Receptor 1 / Tumor Endothelial Marker 8: Mutation of Conserved Inserted Domain Residues Overrides Cytosolic Control of Protective Antigen Binding†

PubMed Central

Ramey, Jordan D.; Villareal, Valerie A.; Ng, Charles; Ward, Sabrina; Xiong, Jian-Ping; Clubb, Robert T.; Bradley, Kenneth A.

2010-01-01

Anthrax toxin receptor 1 (ANTXR1) / tumor endothelial marker 8 (TEM8) is one of two known proteinaceous cell surface anthrax toxin receptors. A metal ion dependent adhesion site (MIDAS) present in the integrin-like inserted (I) domain of ANTXR1 mediates the binding of the anthrax toxin subunit, protective antigen (PA). Here we provide evidence that single point mutations in the I domain can override regulation of ANTXR1 ligand-binding activity mediated by intracellular signals. A previously reported MIDAS-mutant of ANTXR1 (T118A) was found to retain normal metal ion binding and secondary structure but failed to bind PA, consistent with a locked inactive state. Conversely, mutation of a conserved I domain phenylalanine residue to a tryptophan (F205W) increased the proportion of cell-surface ANTXR1 that bound PA, consistent with a locked active state. Interestingly, the KD and total amount of PA bound by the isolated ANTXR1 I domain was not affected by the F205W mutation, indicating that ANTXR1 is preferentially found in the active state in the absence of inside-out signaling. Circular dichroism (CD) spectroscopy and 1H-15N heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) revealed that structural changes between T118A, F205W and WT I domains were minor despite a greater than 103-fold difference in their abilities to bind toxin. Regulation of toxin binding has important implications for the design of toxin inhibitors and for the targeting of ANTXR1 for anti-tumor therapies. PMID:20690680

Endothelial Cell Implantation and Survival within Experimental Gliomas

NASA Astrophysics Data System (ADS)

Lal, Bachchu; Indurti, Ravi R.; Couraud, Pierre-Olivier; Goldstein, Gary W.; Laterra, John

1994-10-01

The delivery of therapeutic genes to primary brain neoplasms opens new opportunities for treating these frequently fatal tumors. Efficient gene delivery to tissues remains an important obstacle to therapy, and this problem has unique characteristics in brain tumors due to the blood-brain and blood-tumor barriers. The presence of endothelial mitogens and vessel proliferation within solid tumors suggests that genetically modified endothelial cells might efficiently transplant to brain tumors. Rat brain endothelial cells immortalized with the adenovirus E1A gene and further modified to express the β-galactosidase reporter were examined for their ability to survive implantation to experimental rat gliomas. Rats received 9L, F98, or C6 glioma cells in combination with endothelial cells intracranially to caudate/putamen or subcutaneously to flank. Implanted endothelial cells were identified by β-galactosidase histochemistry or by polymerase chain reaction in all tumors up to 35 days postimplantation, the latest time examined. Implanted endothelial cells appeared to cooperate in tumor vessel formation and expressed the brain-specific endothelial glucose transporter type 1 as identified by immunohistochemistry. The proliferation of implanted endothelial cells was supported by their increased number within tumors between postimplantation days 14 and 21 (P = 0.015) and by their expression of the proliferation antigen Ki67. These findings establish that genetically modified endothelial cells can be stably engrafted to growing gliomas and suggest that endothelial cell implantation may provide a means of delivering therapeutic genes to brain neoplasms and other solid tumors. In addition, endothelial implantation to brain may be useful for defining mechanisms of brain-specific endothelial differentiation.

Dark Endothelial Spots After Descemet Membrane Endothelial Keratoplasty May Appear as Recurrent Fuchs Dystrophy or Herald Graft Failure or Rejection.

PubMed

Zygoura, Vasiliki; Baydoun, Lamis; Monnereau, Claire; Satué, Maria; Oellerich, Silke; Melles, Gerrit R J

2017-12-01

To evaluate the clinical significance of dark spots in the donor endothelial cell layer as observed with specular microscopy, in patients who underwent Descemet membrane endothelial keratoplasty (DMEK) for Fuchs endothelial dystrophy (FED). Specular microscopy images of 83 consecutive eyes up to 7 years after DMEK were retrospectively reviewed in a masked fashion for the presence of dark spots and morphologic changes in the endothelial cell layer and processed for endothelial cell density (ECD) measurements. A normal endothelial cell layer was found in 52/83 eyes (62.7%) (group 0). In the remaining 31/83 eyes, various dark discolorations with or without altered endothelial cell morphology were categorized into 4 groups. Dark spots were classified as artifacts in 10/83 (12.0%) eyes (group I) and as "superimposed" dots in 10/83 (12.0%) eyes (group II), that is, optical irregularities slightly anterior to a healthy endothelial cell layer. In 11/83 (13.3%) eyes, endothelial stress was characterized by dark grayish discolorations and/or nuclear activation (group III). Most of the latter eyes also had a significant ECD decrease; 3 of these eyes later developed secondary graft failure, of which one was preceded by allograft rejection. None of the eyes showed recurrent guttae typical for FED (group IV). Dark endothelial spots after DMEK for FED may not represent a recurrent disease, but tissue irregularities just anterior to the graft. However, if associated with changes in endothelial cell morphology, nuclear activation and/or ECD decrease, dark discolorations may reflect "cellular stress" heralding secondary graft failure or (subclinical) allograft rejection.

Elevated circulating soluble thrombomodulin activity, tissue factor activity and circulating procoagulant phospholipids: new and useful markers for pre-eclampsia?