Science.gov

Sample records for endothelial marker podoplanin

  1. Syk and Src Family Kinases Regulate C-type Lectin Receptor 2 (CLEC-2)-mediated Clustering of Podoplanin and Platelet Adhesion to Lymphatic Endothelial Cells*

    PubMed Central

    Pollitt, Alice Y.; Poulter, Natalie S.; Gitz, Eelo; Navarro-Nuñez, Leyre; Wang, Ying-Jie; Hughes, Craig E.; Thomas, Steven G.; Nieswandt, Bernhard; Douglas, Michael R.; Owen, Dylan M.; Jackson, David G.; Dustin, Michael L.; Watson, Steve P.

    2014-01-01

    The interaction of C-type lectin receptor 2 (CLEC-2) on platelets with Podoplanin on lymphatic endothelial cells initiates platelet signaling events that are necessary for prevention of blood-lymph mixing during development. In the present study, we show that CLEC-2 signaling via Src family and Syk tyrosine kinases promotes platelet adhesion to primary mouse lymphatic endothelial cells at low shear. Using supported lipid bilayers containing mobile Podoplanin, we further show that activation of Src and Syk in platelets promotes clustering of CLEC-2 and Podoplanin. Clusters of CLEC-2-bound Podoplanin migrate rapidly to the center of the platelet to form a single structure. Fluorescence lifetime imaging demonstrates that molecules within these clusters are within 10 nm of one another and that the clusters are disrupted by inhibition of Src and Syk family kinases. CLEC-2 clusters are also seen in platelets adhered to immobilized Podoplanin using direct stochastic optical reconstruction microscopy. These findings provide mechanistic insight by which CLEC-2 signaling promotes adhesion to Podoplanin and regulation of Podoplanin signaling, thereby contributing to lymphatic vasculature development. PMID:25368330

  2. Podoplanin (D2-40) is a reliable marker of urinary bladder myofibroblasts (telocytes).

    PubMed

    Povýšil, C; Kaňa, M; Zámečník, L; Vaľová, Z; Hanuš, T

    2014-01-01

    Podoplanin, D2-40, has been described in a variety of normal and neoplastic tissues. It is often used for highlighting lymphatics. We evaluated the expression of podoplanin in α-smooth muscle actinpositive myofibroblasts producing the suburothelial layer in tunica propria of the urinary bladder that have some similar features with telocytes. Our results showed that these cells demonstrate distinct D2-40 immunoreactivity from telocytes occurring in the renal pelvis and ureter. We observed positive reaction not only in bioptic specimens from women with interstitial cystitis, but also in a control group of women and men treated for pathological bladder lesion different from interstitial cystitis. It is interesting that identical staining reaction was observed in the ureters only exceptionally. In addition, we examined samples from myofibroblastic tumoriform lesions of soft tissue such as nodular fascitis and fibromatosis (desmoid) and we obtained negative results. It means that the so-called myofibroblasts of urinary bladder tunica propria have a unique immunophenotype that has probably not been described until now. Our findings suggest that D2-40 can be used as a complementary immunostainer to α-smooth muscle actin on urinary bladder biopsies from patients with interstitial cystitis. The role of D2-40 as an immunohistochemical marker is still being investigated. PMID:25629270

  3. Podoplanin maintains high endothelial venule integrity by interacting with platelet CLEC-2.

    PubMed

    Herzog, Brett H; Fu, Jianxin; Wilson, Stephen J; Hess, Paul R; Sen, Aslihan; McDaniel, J Michael; Pan, Yanfang; Sheng, Minjia; Yago, Tadayuki; Silasi-Mansat, Robert; McGee, Samuel; May, Frauke; Nieswandt, Bernhard; Morris, Andrew J; Lupu, Florea; Coughlin, Shaun R; McEver, Rodger P; Chen, Hong; Kahn, Mark L; Xia, Lijun

    2013-10-01

    Circulating lymphocytes continuously enter lymph nodes for immune surveillance through specialized blood vessels named high endothelial venules, a process that increases markedly during immune responses. How high endothelial venules (HEVs) permit lymphocyte transmigration while maintaining vascular integrity is unknown. Here we report a role for the transmembrane O-glycoprotein podoplanin (PDPN, also known as gp38 and T1α) in maintaining HEV barrier function. Mice with postnatal deletion of Pdpn lost HEV integrity and exhibited spontaneous bleeding in mucosal lymph nodes, and bleeding in the draining peripheral lymph nodes after immunization. Blocking lymphocyte homing rescued bleeding, indicating that PDPN is required to protect the barrier function of HEVs during lymphocyte trafficking. Further analyses demonstrated that PDPN expressed on fibroblastic reticular cells, which surround HEVs, functions as an activating ligand for platelet C-type lectin-like receptor 2 (CLEC-2, also known as CLEC1B). Mice lacking fibroblastic reticular cell PDPN or platelet CLEC-2 exhibited significantly reduced levels of VE-cadherin (also known as CDH5), which is essential for overall vascular integrity, on HEVs. Infusion of wild-type platelets restored HEV integrity in Clec-2-deficient mice. Activation of CLEC-2 induced release of sphingosine-1-phosphate from platelets, which promoted expression of VE-cadherin on HEVs ex vivo. Furthermore, draining peripheral lymph nodes of immunized mice lacking sphingosine-1-phosphate had impaired HEV integrity similar to Pdpn- and Clec-2-deficient mice. These data demonstrate that local sphingosine-1-phosphate release after PDPN-CLEC-2-mediated platelet activation is critical for HEV integrity during immune responses. PMID:23995678

  4. Podoplanin maintains high endothelial venule integrity by interacting with platelet CLEC-2

    PubMed Central

    Herzog, Brett H.; Fu, Jianxin; Wilson, Stephen J.; Hess, Paul R.; Sen, Aslihan; McDaniel, J. Michael; Pan, Yanfang; Sheng, Minjia; Yago, Tadayuki; Silasi-Mansat, Robert; McGee, Samuel; May, Frauke; Nieswandt, Bernhard; Morris, Andrew J.; Lupu, Florea; Coughlin, Shaun R.; McEver, Rodger P.; Chen, Hong; Kahn, Mark L.; Xia, Lijun

    2013-01-01

    Circulating lymphocytes continuously enter lymph nodes (LNs) for immune surveillance through specialised blood vessels named high endothelial venules (HEVs)1–5, a process that increases dramatically during immune responses. How HEVs permit lymphocyte transmigration while maintaining vascular integrity is unknown. Here, we report a role for the transmembrane O-glycoprotein podoplanin (PDPN, also known as gp38 and T1α)6–8 in maintaining HEV barrier function. Mice with postnatal deletion of PDPN lost HEV integrity and exhibited spontaneous bleeding in mucosal LNs, and bleeding in the draining peripheral LN after immunisation. Blocking lymphocyte homing rescued bleeding, indicating that PDPN is required to protect the barrier function of HEVs during lymphocyte trafficking. Further analyses demonstrated that PDPN expressed on fibroblastic reticular cells (FRCs)7, which surround HEVs, functions as an activating ligand for platelet C-type lectin-like receptor 2 (CLEC-2)9,10. Mice lacking FRC PDPN or platelet CLEC-2 exhibited significantly reduced levels of VE-cadherin (VE-cad), which is essential for overall vascular integrity11,12, on HEVs. Infusion of wild-type (WT) platelets restored HEV integrity in CLEC-2-deficient mice. Activation of CLEC-2 induced release of sphingosine-1-phosphate (S1P)13,14 from platelets, which promoted expression of VE-cad on HEVs ex vivo. Furthermore, draining peripheral LNs of immunised mice lacking S1P had impaired HEV integrity similar to PDPN- and CLEC-2-deficient mice. These data demonstrate that local S1P release after PDPN-CLEC-2-mediated platelet activation is critical for HEV integrity during immune responses. PMID:23995678

  5. Dual role of podoplanin in oral cancer development.

    PubMed

    Cîrligeriu, Laura; Cimpean, Anca Maria; Raica, Marius; Doroş, Caius Ioan

    2014-01-01

    Podoplanin plays a crucial role for normal and pathological tissue development. Known as a lymphatic endothelial marker, podoplanin has been found to be overexpressed in tumor cells of various cancers with a certified involvement in tumor progression, invasion and metastasis. Oral cancer includes a heterogeneous group of malignancies with unpredictable behaviour and sometimes poor prognosis. Based on these facts, development of new molecular markers with a more reliable impact on therapy and prognosis is required. The present study was designed to characterize podoplanin expression in tumor cells of lip, oral cavity, tongue and pharynx squamous cell carcinomas, together with lymphatic vessels distribution, morphology, density and their impact on tumor progression. Evaluation of podoplanin by D2-40 immunohistochemistry assessement on 56 cases of oral cancers, revealed two different expression patterns in tumor cells depending on their location. Peri-tumor and intra-tumor lymphatic vessels density, morphology and distribution were correlated with lymph node status but not with tumor stage. The highest number of lymphatic vessels was observed in grade 3 squamous cell carcinomas. Dual expression of podoplanin in tumor cells and lymphatics with particular patterns correlated with histopathology and lymph node status in oral cancer, representing the molecular basis for testing podoplanin as a potential target for anti D2-40 antibody based therapy. PMID:24815836

  6. Characterization of anti-podoplanin monoclonal antibodies: critical epitopes for neutralizing the interaction between podoplanin and CLEC-2.

    PubMed

    Ogasawara, Satoshi; Kaneko, Mika Kato; Price, Janet E; Kato, Yukinari

    2008-08-01

    Podoplanin (Aggrus) is a mucin-type sialoglycoprotein that is known as a useful marker for lymphatic endothelium and tumor-initiating cells (TICs). Interaction between podoplanin and C-type lectin-like receptor-2 (CLEC-2) is reported to be critical for podoplanin-induced platelet aggregation and cancer metastasis. Recently, several anti-human podoplanin antibodies have been created; however, these anti-podoplanin antibodies have not been well characterized. Five anti-podoplanin antibodies (NZ-1, D2-40, AB3, 18H5, and a rabbit polyclonal antibody) were investigated using ELISA, Western blot, and flow cytometry with synthesized podoplanin peptides and deletion mutants of recombinant podoplanin. The epitope of NZ-1 is platelet aggregation-stimulating (PLAG) domain-2/3; the epitope of D2-40, AB3, and 18H5 is PLAG1/2. The epitopes of D2-40 and AB3 are quite similar, although 18H5 is different from D2-40 and AB3. Using flow cytometric analysis, NZ-1 partially inhibited the interaction between podoplanin and CLEC-2, although other antibodies did not. In conclusion, the two most frequently used anti-podoplanin antibodies, D2-40 and AB3, have similar properties, although several studies have reported differences. NZ-1 neutralizes the interaction between podoplanin and CLEC-2, which may lead to the development of therapeutic antibodies against podoplanin-dependent cancer metastasis. PMID:18707544

  7. Lymphangiogenesis and expression of specific molecules as lymphatic endothelial cell markers.

    PubMed

    Kato, Seiji; Shimoda, Hiroshi; Ji, Rui-Cheng; Miura, Masahiro

    2006-06-01

    In recent years, several functional molecules specifically expressed and localized in lymphatic endothelial cells, such as 5'-nucleotidase, lymphatic vessel endothelial receptor-1, vascular endothelial growth factor receptor-3, podoplanin and Prox-1, have been identified. The discovery of the lymphatic endothelial cell markers facilitated detailed analysis of the nature and structural organization of the lymphatic vessels and their growth (lymphangiogenesis). As a result, over the past few years, advances have been made in understanding the cellular and molecular aspects of physiological lymphangiogenesis and tumor-induced lymphangiogenesis. The biology of lymphangiogenesis, particularly the mechanism of its regulation, is very important in understanding the formation of the lymphatic system as a biological regulation system transporting tissue fluid and wandering cells, including lymphocytes, and disease involving lymphangiogenesis. The understanding of the molecular mechanism of lymphangiogenesis and the elucidation of the development of normal and pathological tissues are expected to lead to the development of therapy for intractable diseases, such as malignant tumors and lymphedema. PMID:16800291

  8. Characterization of Monoclonal Antibody LpMab-3 Recognizing Sialylated Glycopeptide of Podoplanin

    PubMed Central

    Oki, Hiroharu; Ogasawara, Satoshi; Kaneko, Mika Kato; Takagi, Michiaki; Yamauchi, Masanori

    2015-01-01

    Podoplanin (PDPN/Aggrus/T1α/gp36/OTS-8), a type I transmembrane sialoglycoprotein, is involved in platelet aggregation, cell invasion, and cancer metastasis. Podoplanin expression in cancer cells or cancer-associated fibroblasts was reported to be involved in poor prognosis of several cancers. Furthermore, podoplanin is expressed in lymphatic endothelial cells or lung type I alveolar cells. Although many anti-podoplanin monoclonal antibodies (MAbs), such as NZ-1 and D2–40, have been established, almost all anti-podoplanin MAbs are produced against a platelet aggregation-inducing (PLAG) domain. In this study, we produced and characterized a novel anti-podoplanin monoclonal antibody, LpMab-3, the epitope of which is a sialylated glycopeptide of podoplanin. We identified the minimum epitope of LpMab-3 as Thr76–Glu81 of human podoplanin, which is different from PLAG domain, using Western blot analysis and flow cytometry. Immunohistochemical analysis showed that LpMab-3 is useful for detecting lung type I alveolar cells and lymphatic endothelial cells. Because LpMab-3 detects only sialylated podoplanin, it could be useful for uncovering the physiological function of sialylated human podoplanin. PMID:25723283

  9. A Cancer-specific Monoclonal Antibody Recognizes the Aberrantly Glycosylated Podoplanin

    PubMed Central

    Kato, Yukinari; Kaneko, Mika Kato

    2014-01-01

    Podoplanin (PDPN/Aggrus/T1α), a platelet aggregation-inducing mucin-like sialoglycoprotein, is highly expressed in many cancers and normal tissues. A neutralizing monoclonal antibody (mAb; NZ-1) can block the association between podoplanin and C-type lectin-like receptor-2 (CLEC-2) and inhibit podoplanin-induced cancer metastasis, but NZ-1 reacts with podoplanin-expressing normal cells such as lymphatic endothelial cells. In this study, we established a cancer-specific mAb (CasMab) against human podoplanin. Aberrantly glycosylated podoplanin including keratan sulfate or aberrant sialylation, which was expressed in LN229 glioblastoma cells, was used as an immunogen. The newly established LpMab-2 mAb recognized both an aberrant O-glycosylation and a Thr55-Leu64 peptide from human podoplanin. Because LpMab-2 reacted with podoplanin-expressing cancer cells but not with normal cells, as shown by flow cytometry and immunohistochemistry, it is an anti-podoplanin CasMab that is expected to be useful for molecular targeting therapy against podoplanin-expressing cancers. PMID:25080943

  10. Plasticity of Blood- and Lymphatic Endothelial Cells and Marker Identification

    PubMed Central

    Keuschnigg, Johannes; Karinen, Sirkku; Auvinen, Kaisa; Irjala, Heikki; Mpindi, John-Patrick; Kallioniemi, Olli; Hautaniemi, Sampsa; Jalkanen, Sirpa; Salmi, Marko

    2013-01-01

    The distinction between lymphatic and blood vessels is biologically fundamental. Here we wanted to rigorously analyze the universal applicability of vascular markers and characteristics of the two widely used vascular model systems human microvascular endothelial cell line-1 (HMEC-1) and telomerase-immortalized microvascular endothelial cell line (TIME). Therefore we studied the protein expression and functional properties of the endothelial cell lines HMEC-1 and TIME by flow cytometry and in vitro flow assays. We then performed microarray analyses of the gene expression in these two cell lines and compared them to primary endothelial cells. Using bioinformatics we then defined 39 new, more universal, endothelial-type specific markers from 47 primary endothelial microarray datasets and validated them using immunohistochemistry with normal and pathological tissues. We surprisingly found that both HMEC-1 and TIME are hybrid blood- and lymphatic cells. In addition, we discovered great discrepancies in the previous identifications of blood- and lymphatic endothelium-specific genes. Hence we identified and validated new, universally applicable vascular markers. Summarizing, the hybrid blood-lymphatic endothelial phenotype of HMEC-1 and TIME is indicative of plasticity in the gene expression of immortalized endothelial cell lines. Moreover, we identified new, stable, vessel-type specific markers for blood- and lymphatic endothelium, useful for basic research and clinical diagnostics. PMID:24058540

  11. Podoplanin expression in the development and progression of laryngeal squamous cell carcinomas

    PubMed Central

    2010-01-01

    Background Podoplanin expression is attracting interest as a marker for cancer diagnosis and prognosis. We therefore investigated the expression pattern and clinical significance of podoplanin during the development and progression of laryngeal carcinomas. Results Podoplanin expression was determined by immunohistochemistry in paraffin-embedded tissue specimens from 84 patients with laryngeal premalignancies and 53 patients with laryngeal squamous cell carcinomas. We found podoplanin expression extending from the basal to the suprabasal layer of the epithelium in 37 (44%) of 84 dysplastic lesions, whereas normal epithelium showed negligible expression. Patients carrying podoplanin-positive lesions had a higher laryngeal cancer incidence than those with negative expression reaching borderline statistical significance (51% versus 30%, P = 0.071). Podoplanin expression in laryngeal carcinomas exhibited two distinct patterns. 20 (38%) cases showed diffuse expression in most tumour cells and 33 (62%) focal expression at the proliferating periphery of tumour nests. High podoplanin expression was inversely correlated with T classification (P = 0.033), disease stage (P = 0.006), and pathological grade (P = 0.04). There was a trend, although not significant, towards reduced disease-specific survival for patients with low podoplanin levels (P = 0.31) and diffuse expression pattern (P = 0.08). Conclusions Podoplanin expression increases in the early stages of laryngeal tumourigenesis and it seems to be associated with a higher laryngeal cancer risk. Podoplanin expression in laryngeal squamous cell carcinomas, however, diminishes during tumour progression. Taken together, these data support a role for podoplanin expression in the initiation but not in the progression of laryngeal cancers. PMID:20196862

  12. Aberrant Lymphatic Endothelial Progenitors in Lymphatic Malformation Development

    PubMed Central

    Wu, June K.; Kitajewski, Christopher; Reiley, Maia; Keung, Connie H.; Monteagudo, Julie; Andrews, John P.; Liou, Peter; Thirumoorthi, Arul; Wong, Alvin

    2015-01-01

    Lymphatic malformations (LMs) are vascular anomalies thought to arise from dysregulated lymphangiogenesis. These lesions impose a significant burden of disease on affected individuals. LM pathobiology is poorly understood, hindering the development of effective treatments. In the present studies, immunostaining of LM tissues revealed that endothelial cells lining aberrant lymphatic vessels and cells in the surrounding stroma expressed the stem cell marker, CD133, and the lymphatic endothelial protein, podoplanin. Isolated patient-derived CD133+ LM cells expressed stem cell genes (NANOG, Oct4), circulating endothelial cell precursor proteins (CD90, CD146, c-Kit, VEGFR-2), and lymphatic endothelial proteins (podoplanin, VEGFR-3). Consistent with a progenitor cell identity, CD133+ LM cells were multipotent and could be differentiated into fat, bone, smooth muscle, and lymphatic endothelial cells in vitro. CD133+ cells were compared to CD133− cells isolated from LM fluids. CD133− LM cells had lower expression of stem cell genes, but expressed circulating endothelial precursor proteins and high levels of lymphatic endothelial proteins, VE-cadherin, CD31, podoplanin, VEGFR-3 and Prox1. CD133− LM cells were not multipotent, consistent with a differentiated lymphatic endothelial cell phenotype. In a mouse xenograft model, CD133+ LM cells differentiated into lymphatic endothelial cells that formed irregularly dilated lymphatic channels, phenocopying human LMs. In vivo, CD133+ LM cells acquired expression of differentiated lymphatic endothelial cell proteins, podoplanin, LYVE1, Prox1, and VEGFR-3, comparable to expression found in LM patient tissues. Taken together, these data identify a novel LM progenitor cell population that differentiates to form the abnormal lymphatic structures characteristic of these lesions, recapitulating the human LM phenotype. This LM progenitor cell population may contribute to the clinically refractory behavior of LMs. PMID:25719418

  13. Emerging Role of Endothelial and Inflammatory Markers in Preeclampsia

    PubMed Central

    Swellam, Menha; Samy, Nervana; Abdl Wahab, Susan; Ibrahim, Mohamed Saeed

    2009-01-01

    Objectives: Endothelial disturbance and excess inflammatory response are pathogenic mechanisms in pre-eclampsia (PE). Authors determine the clinical diagnostic role for thrombomodulin (TM), plasminogen activator inhibitor-1 (PAI-1) as endothelial markers and C-reactive protein (CRP), and interlukin-6 (IL-6) as inflammatory markers when tested independently or in combinations. Materials and methods: We conducted a retrospective study in a cohort of 185 women grouped as 80 women with PE, 55 normotensive pregnant and 50 healthy non-pregnant. Plasma levels of TM, PAI-1, CRP and IL-6 were examined using enzyme linked immunosorbent assays. Results: Median levels and the positivity rates for the investigated markers were higher in PE as compared to the other groups (P < 0.0001). Using linear regression analysis, the investigated markers were significantly correlated regarding healthy nonpregnant vs PE or normotensive pregnant vs PE. The sensitivity of PAI-1 was the highest (98%) among the tested biomarkers. Combination between the investigated markers revealed absolute sensitivity (100%) and reliable specificity especially when PAI-1 was combined with CRP at 83% specificity. Conclusions: Investigated endothelial and inflammatory markers revealed sensitive diagnostic test for PE. However, coupled combination between PAI-1 with CRP showed superior both sensitivity and specificity which represent a promising new approach for detection of PE. PMID:19597295

  14. Obstructive Sleep Apnoea Syndrome, Endothelial Function and Markers of Endothelialization. Changes after CPAP

    PubMed Central

    Sanchez Armengol, Angeles; Moreno-Luna, Rafael; Caballero-Eraso, Candela; Macher, Hada C.; Villar, Jose; Merino, Ana M; Castell, Javier; Capote, Francisco; Stiefel, Pablo

    2015-01-01

    Study objectives This study tries to assess the endothelial function in vivo using flow-mediated dilatation (FMD) and several biomarkers of endothelium formation/restoration and damage in patients with obstructive sleep apnoea (OSA) syndrome at baseline and after three months with CPAP therapy. Design Observational study, before and after CPAP therapy. Setting and Patients We studied 30 patients with apnoea/hypopnoea index (AHI) >15/h that were compared with themselves after three months of CPAP therapy. FMD was assessed non-invasively in vivo using the Laser-Doppler flowmetry. Circulating cell-free DNA (cf-DNA) and microparticles (MPs) were measured as markers of endothelial damage and the vascular endothelial growth factor (VEGF) was determined as a marker of endothelial restoration process. Measurements and results After three month with CPAP, FMD significantly increased (1072.26 ± 483.21 vs. 1604.38 ± 915.69 PU, p< 0.005) cf-DNA and MPs significantly decreased (187.93 ± 115.81 vs. 121.28 ± 78.98 pg/ml, p<0.01, and 69.60 ± 62.60 vs. 39.82 ± 22.14 U/μL, p<0.05, respectively) and VEGF levels increased (585.02 ± 246.06 vs. 641.11 ± 212.69 pg/ml, p<0.05). These changes were higher in patients with more severe disease. There was a relationship between markers of damage (r = -0.53, p<0.005) but not between markers of damage and restoration, thus suggesting that both types of markers should be measured together. Conclusions CPAP therapy improves FMD. This improvement may be related to an increase of endothelial restoration process and a decrease of endothelial damage. PMID:25815511

  15. Suprabasin as a novel tumor endothelial cell marker.

    PubMed

    Alam, Mohammad T; Nagao-Kitamoto, Hiroko; Ohga, Noritaka; Akiyama, Kosuke; Maishi, Nako; Kawamoto, Taisuke; Shinohara, Nobuo; Taketomi, Akinobu; Shindoh, Masanobu; Hida, Yasuhiro; Hida, Kyoko

    2014-12-01

    Recent studies have reported that stromal cells contribute to tumor progression. We previously demonstrated that tumor endothelial cells (TEC) characteristics were different from those of normal endothelial cells (NEC). Furthermore, we performed gene profile analysis in TEC and NEC, revealing that suprabasin (SBSN) was upregulated in TEC compared with NEC. However, its role in TEC is still unknown. Here we showed that SBSN expression was higher in isolated human and mouse TEC than in NEC. SBSN knockdown inhibited the migration and tube formation ability of TEC. We also showed that the AKT pathway was a downstream factor of SBSN. These findings suggest that SBSN is involved in the angiogenic potential of TEC and may be a novel TEC marker. PMID:25283635

  16. Podoplanin-Fc reduces lymphatic vessel formation in vitro and in vivo and causes disseminated intravascular coagulation when transgenically expressed in the skin

    PubMed Central

    Cueni, Leah N.; Chen, Lu; Zhang, Hui; Marino, Daniela; Huggenberger, Reto; Alitalo, Annamari; Bianchi, Roberta

    2010-01-01

    Podoplanin is a small transmembrane protein required for development and function of the lymphatic vascular system. To investigate the effects of interfering with its function, we produced an Fc fusion protein of its ectodomain. We found that podoplanin-Fc inhibited several functions of cultured lymphatic endothelial cells and also specifically suppressed lymphatic vessel growth, but not blood vessel growth, in mouse embryoid bodies in vitro and in mouse corneas in vivo. Using a keratin 14 expression cassette, we created transgenic mice that overexpressed podoplanin-Fc in the skin. No obvious outward phenotype was identified in these mice, but surprisingly, podoplanin-Fc—although produced specifically in the skin—entered the blood circulation and induced disseminated intravascular coagulation, characterized by microthrombi in most organs and by thrombocytopenia, occasionally leading to fatal hemorrhage. These findings reveal an important role of podoplanin in lymphatic vessel formation and indicate the potential of podoplanin-Fc as an inhibitor of lymphangiogenesis. These results also demonstrate the ability of podoplanin to induce platelet aggregation in vivo, which likely represents a major function of lymphatic endothelium. Finally, keratin 14 podoplanin-Fc mice represent a novel genetic animal model of disseminated intravascular coagulation. PMID:20716773

  17. 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. PMID:23489680

  18. The tumour-associated glycoprotein podoplanin is expressed in fibroblast-like synoviocytes of the hyperplastic synovial lining layer in rheumatoid arthritis

    PubMed Central

    2011-01-01

    Introduction Activated fibroblast-like synoviocytes (FLSs) in rheumatoid arthritis (RA) share many characteristics with tumour cells and are key mediators of synovial tissue transformation and joint destruction. The glycoprotein podoplanin is upregulated in the invasive front of several human cancers and has been associated with epithelial-mesenchymal transition, increased cell migration and tissue invasion. The aim of this study was to investigate whether podoplanin is expressed in areas of synovial transformation in RA and especially in promigratory RA-FLS. Methods Podoplanin expression in human synovial tissue from 18 RA patients and nine osteoarthritis (OA) patients was assessed by immunohistochemistry and confirmed by Western blot analysis. The expression was related to markers of synoviocytes and myofibroblasts detected by using confocal immunofluoresence microscopy. Expression of podoplanin, with or without the addition of proinflammatory cytokines and growth factors, in primary human FLS was evaluated by using flow cytometry. Results Podoplanin was highly expressed in cadherin-11-positive cells throughout the synovial lining layer in RA. The expression was most pronounced in areas with lining layer hyperplasia and high matrix metalloproteinase 9 expression, where it coincided with upregulation of α-smooth muscle actin (α-sma). The synovium in OA was predominantly podoplanin-negative. Podoplanin was expressed in 50% of cultured primary FLSs, and the expression was increased by interleukin 1β, tumour necrosis factor α and transforming growth factor β receptor 1. Conclusions Here we show that podoplanin is highly expressed in FLSs of the invading synovial tissue in RA. The concomitant upregulation of α-sma and podoplanin in a subpopulation of FLSs indicates a myofibroblast phenotype. Proinflammatory mediators increased the podoplanin expression in cultured RA-FLS. We conclude that podoplanin might be involved in the synovial tissue transformation and

  19. Podoplanin - a small glycoprotein with many faces.

    PubMed

    Ugorski, Maciej; Dziegiel, Piotr; Suchanski, Jaroslaw

    2016-01-01

    Podoplanin is a small membrane glycoprotein with a large number of O-glycoside chains and therefore it belongs to mucin-type proteins. It can be found on the surface of many types of normal cells originating from various germ layers. It is present primarily on the endothelium of lymphatic vessels, type I pneumocytes and glomerular podocytes. Increased levels of podoplanin or its neo-expression have been found in numerous types of human carcinomas, but it is especially common in squamous cell carcinomas, such as cervical, larynx, oral cavity, skin and lung cancer. This small sialomucin is also seen on the surface of cancer-associated fibroblasts (CAFs) in lung adenocarcinomas, as well as in breast and pancreatic tumors. In most cancers, a high level of podoplanin expression, both in cancer cells, as well as in CAFs, is correlated with an increased incidence of metastasis to lymph nodes and shorter survival time of patients. Little is known about the biological role of podoplanin, however research carried out on mice with a knock-out gene of this glycoprotein shows that the presence of podoplanin determines normal development of lungs, the lymphatic system and heart. Podoplanin on cancer cells and CAFs seems to play an important role in the development and progression of various cancers. However, its role in these processes is both unclear and controversial. In this review, the role of podoplanin in both physiological processes and carcinogenesis is discussed. PMID:27186410

  20. Podoplanin - a small glycoprotein with many faces

    PubMed Central

    Ugorski, Maciej; Dziegiel, Piotr; Suchanski, Jaroslaw

    2016-01-01

    Podoplanin is a small membrane glycoprotein with a large number of O-glycoside chains and therefore it belongs to mucin-type proteins. It can be found on the surface of many types of normal cells originating from various germ layers. It is present primarily on the endothelium of lymphatic vessels, type I pneumocytes and glomerular podocytes. Increased levels of podoplanin or its neo-expression have been found in numerous types of human carcinomas, but it is especially common in squamous cell carcinomas, such as cervical, larynx, oral cavity, skin and lung cancer. This small sialomucin is also seen on the surface of cancer-associated fibroblasts (CAFs) in lung adenocarcinomas, as well as in breast and pancreatic tumors. In most cancers, a high level of podoplanin expression, both in cancer cells, as well as in CAFs, is correlated with an increased incidence of metastasis to lymph nodes and shorter survival time of patients. Little is known about the biological role of podoplanin, however research carried out on mice with a knock-out gene of this glycoprotein shows that the presence of podoplanin determines normal development of lungs, the lymphatic system and heart. Podoplanin on cancer cells and CAFs seems to play an important role in the development and progression of various cancers. However, its role in these processes is both unclear and controversial. In this review, the role of podoplanin in both physiological processes and carcinogenesis is discussed. PMID:27186410

  1. Endothelial cell markers reflecting endothelial cell dysfunction in patients with mixed connective tissue disease

    PubMed Central

    2010-01-01

    Introduction The aim of the present study was to investigate the association between cardiovascular risk factors and endothelial dysfunction in patients with mixed connective tissue disease (MCTD) and to determine which biomarkers are associated with atherosclerotic complications, such as cardiovascular disease. Methods Fifty MCTD patients and 38 healthy age-matched and sex-matched controls were enrolled in this study. In order to describe endothelial dysfunction, we assessed flow-mediated dilation (FMD), nitrate-mediated dilation (NMD) and carotid artery intima-media thickness (IMT). We investigated FMD of the brachial artery after reactive hyperemia and NMD after sublingual nitroglycerin administration, while the IMT of the common carotid artery was determined by ultrasound. Anti-U1 ribonucleoprotein (anti-U1RNP) antibodies, anti-cardiolipin (anti-CL) antibodies, anti-endothelial cell antibody (AECA) and endothelial cell markers, such as soluble thrombomodulin (TM) and von Willebrand factor antigen (vWFAg), were assessed. Results The endothelium-dependent vasodilation (FMD) was significantly impaired in patients with MCTD, as compared with controls (%FMD: 4.7 ± 4.2% vs. 8.7 ± 5.0%; P < 0.001), while the percentage NMD did not differ (%NMD: 14.3 ± 6.6% vs. 17.1 ± 6.7%; P = 0.073). Mean carotid IMT values were higher in patients than in controls (IMT: MCTD, 0.64 ± 0.13 mm vs. controls, 0.53 ± 0.14 mm; P < 0.001). FMD negatively correlated with disease duration, the levels of apolipoprotein A1, the paraoxonase-1 activity, and systolic blood pressure in MCTD patients. The percentage FMD was significantly lower in MCTD patients with cardiovascular diseases (CVD), than in those without CVD (%FMD: 3.5 ± 2.9 vs. 5.8 ± 4.8, P < 0.0002), while percentage NMD did not differ between patients with and without CVDs. Serum levels of autoantibodies (anti-U1RNP, AECA and anti-CL) were significantly higher in MCTD patients and differed between MCTD patients with and

  2. Dimer conformation of soluble PECAM-1, an endothelial marker.

    PubMed

    Jiang, Longguang; Lin, Lin; Li, Rui; Yuan, Cai; Xu, Mingming; Huang, Joy H; Huang, Mingdong

    2016-08-01

    Platelet endothelial cell adhesion molecule 1 (PECAM-1) is a cell surface receptor widely distributed on endothelium and hematopoietic-derived cells, and maintain the integrity of the blood vessels. PECAM-1 is widely recognized as an endothelial cell marker. The homophilic interaction through its extracellular domain of PECAM-1 (soluble PECAM-1, or sPECAM-1) is important to its functions. However, structural details for such dimerization are not fully understood. Here we report the production of recombinant sPECAM-1 in large quantity by Drosophila expression system and the small-angle X-ray diffraction (SAXS) study. The recombinant sPECAM-1 was found to form one population of dimer, but not oligomer, and was able to bind to heparin immobilized on a chip in surface plasmon resonance imaging (SPRi) binding experiments. The results of SAXS demonstrated that sPECAM-1 formed a symmetric homodimer of Ω-shape in solution, and each protomer adopted an extended conformation. The dimer is mediated through the intermolecular interactions through the first N-terminal domain (D1) of sPECAM-1. This model provides new structural information for its homophilic interaction mechanism. PMID:27270333

  3. DIET QUALITY SCORES AND PLASMA CONCENTRATIONS OF MARKERS OF INFLAMMATION AND ENDOTHELIAL DYSFUNCTION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Endothelial dysfunction is one of the mechanisms linked to an increased risk of cardiovascular disease. We assessed the association between several diet-quality scores and plasma concentrations of markers of inflammation and endothelial dysfunction. Diet-quality scores on the Healthy Eating Index (H...

  4. Podoplanin-mediated TGF-β-induced epithelial-mesenchymal transition and its correlation with bHLH transcription factor DEC in TE-11 cells

    PubMed Central

    WU, YUNYAN; LIU, QIANG; YAN, XU; KATO, YUKIO; TANAKA, MAKIKO; INOKUCHI, SADAKI; YOSHIZAWA, TADASHI; MOROHASHI, SATOKO; KIJIMA, HIROSHI

    2016-01-01

    Podoplanin is reported involved in the collective cell invasion, another tumor invasion style which is distinct from the single cell invasion, so-called epithelial-mesenchymal transition (EMT). In this study, we investigated the correlation between podoplanin and EMT-related markers in esophageal squamous cell carcinoma (ESCC), and evaluated its linkage with the basic helix-loop-helix (bHLH) transcription factor differentiated embryonic chondrocyte (DEC) 1 and DEC2. Three ESCC cell lines and human squamous cell carcinoma A431 cells were subjected to western blot analyses for podoplanin and EMT markers, as well as the expression of DEC1 and DEC2. By RT-qPCR and western blotting, we found that TGF-β increased the expression of podoplanin and mensenchymal markers (e.g., N-cadherin and vimentin), while decreased the expression of epithelial markers (e.g., Claudin-4 and E-cadherin), accompanied by Smad2 phosphorylation and slug activation. Moreover, TGF-β has different effects on the expression of DEC1 and DEC2, that is, it upregulates DEC1, but downregulates DEC2. Capability of cell proliferation, invasion and migration were further analyzed using CCK-8 assay, Matrigel-invasion assay, and the wound-healing assay, respectively. The proliferation, invasion and migration ability were significantly lost in podoplanin-knockdown cells when compared with the scrambled siRNA group. In addition to these changes, the expression of Claudin-4, but not that of Claudin-1 or E-cadherin, was induced by the siRNA against podoplanin. On the contrary, overexpression of DEC1 and DEC2 exhibits opposite effects on podoplanin, but only slight effect on Claudin-4 was detected. These data indicated that podoplanin is significantly associated with EMT of TE-11 cells, and may be directly or indirectly regulated by bHLH transcription factors DEC1 and DEC2. PMID:27035755

  5. Podoplanin-mediated TGF-β-induced epithelial-mesenchymal transition and its correlation with bHLH transcription factor DEC in TE-11 cells.

    PubMed

    Wu, Yunyan; Liu, Qiang; Yan, Xu; Kato, Yukio; Tanaka, Makiko; Inokuchi, Sadaki; Yoshizawa, Tadashi; Morohashi, Satoko; Kijima, Hiroshi

    2016-06-01

    Podoplanin is reported involved in the collective cell invasion, another tumor invasion style which is distinct from the single cell invasion, so-called epithelial-mesenchymal transition (EMT). In this study, we investigated the correlation between podoplanin and EMT-related markers in esophageal squamous cell carcinoma (ESCC), and evaluated its linkage with the basic helix-loop-helix (bHLH) transcription factor differentiated embryonic chondrocyte (DEC) 1 and DEC2. Three ESCC cell lines and human squamous cell carcinoma A431 cells were subjected to western blot analyses for podoplanin and EMT markers, as well as the expression of DEC1 and DEC2. By RT-qPCR and western blotting, we found that TGF-β increased the expression of podoplanin and mensenchymal markers (e.g., N-cadherin and vimentin), while decreased the expression of epithelial markers (e.g., Claudin-4 and E-cadherin), accompanied by Smad2 phosphorylation and slug activation. Moreover, TGF-β has different effects on the expression of DEC1 and DEC2, that is, it upregulates DEC1, but downregulates DEC2. Capability of cell proliferation, invasion and migration were further analyzed using CCK-8 assay, Matrigel-invasion assay, and the wound-healing assay, respectively. The proliferation, invasion and migration ability were significantly lost in podoplanin-knockdown cells when compared with the scrambled siRNA group. In addition to these changes, the expression of Claudin-4, but not that of Claudin-1 or E-cadherin, was induced by the siRNA against podoplanin. On the contrary, overexpression of DEC1 and DEC2 exhibits opposite effects on podoplanin, but only slight effect on Claudin-4 was detected. These data indicated that podoplanin is significantly associated with EMT of TE-11 cells, and may be directly or indirectly regulated by bHLH transcription factors DEC1 and DEC2. PMID:27035755

  6. A microarray analysis of two distinct lymphatic endothelial cell populations

    PubMed Central

    Schweighofer, Bernhard; Rohringer, Sabrina; Pröll, Johannes; Holnthoner, Wolfgang

    2015-01-01

    We have recently identified lymphatic endothelial cells (LECs) to form two morphologically different populations, exhibiting significantly different surface protein expression levels of podoplanin, a major surface marker for this cell type. In vitro shockwave treatment (IVSWT) of LECs resulted in enrichment of the podoplaninhigh cell population and was accompanied by markedly increased cell proliferation, as well as 2D and 3D migration. Gene expression profiles of these distinct populations were established using Affymetrix microarray analyses. Here we provide additional details about our dataset (NCBI GEO accession number GSE62510) and describe how we analyzed the data to identify differently expressed genes in these two LEC populations. PMID:26484194

  7. Tumor endothelial markers define novel subsets of cancer-specific circulating endothelial cells associated with antitumor efficacy.

    PubMed

    Mehran, Reza; Nilsson, Monique; Khajavi, Mehrdad; Du, Zhiqiang; Cascone, Tina; Wu, Hua Kang; Cortes, Andrea; Xu, Li; Zurita, Amado; Schier, Robert; Riedel, Bernhard; El-Zein, Randa; Heymach, John V

    2014-05-15

    Circulating endothelial cells (CEC) are derived from multiple sources, including bone marrow (circulating endothelial progenitors; CEP), and established vasculature (mature CEC). Although CECs have shown promise as a biomarker for patients with cancer, their utility has been limited, in part, by the lack of specificity for tumor vasculature and the different nonmalignant causes that can impact CEC. Tumor endothelial markers (TEM) are antigens enriched in tumor versus nonmalignant endothelia. We hypothesized that TEMs may be detectable on CEC and that these circulating TEM(+) endothelial cells (CTEC) may be a more specific marker for cancer and tumor response than standard CEC. We found that tumor-bearing mice had a relative increase in numbers of circulating CTEC, specifically with increased levels of TEM7 and TEM8 expression. Following treatment with various vascular-targeting agents, we observed a decrease in CTEC that correlated with the reductions in tumor growth. We extended these findings to human clinical samples and observed that CTECs were present in patients with esophageal cancer and non-small cell lung cancer (N = 40), and their levels decreased after surgical resection. These results demonstrate that CTECs are detectable in preclinical cancer models and patients with cancer. Furthermore, they suggest that CTECs offer a novel cancer-associated marker that may be useful as a blood-based surrogate for assessing the presence of tumor vasculature and antiangiogenic drug activity. PMID:24626092

  8. Enhanced expression of Aggrus (T1alpha/podoplanin), a platelet-aggregation-inducing factor in lung squamous cell carcinoma.

    PubMed

    Kato, Yukinari; Kaneko, Mika; Sata, Makoto; Fujita, Naoya; Tsuruo, Takashi; Osawa, Motoki

    2005-01-01

    Aggrus (T1alpha/podoplanin, known as a specific marker for type I alveolar cells or lymphatic endothelial cells) is a transmembrane sialoglycoprotein that aggregates platelets. Previously, we showed that upregulated expression of Aggrus occurs in colorectal tumors or testicular tumors and could be associated with platelet-aggregating activity and metastatic ability. In testicular tumors, Aggrus is specifically expressed in seminoma. The present study investigates Aggrus expression in human primary lung cancer tissues of different types. Microarray analysis demonstrated that aggrus was significantly expressed in squamous cell carcinoma (10/15; 66.7%). Immunohistochemical analysis also showed that the incidence of positive staining in sections of squamous cell carcinoma (7/8; 87.5%) was higher than that in adenocarcinoma (2/13; 15.4%). Furthermore, Aggrus expression was detected in a squamous cell carcinoma cell line, NCI-H226, by real-time PCR. These findings indicated that overexpression of Aggrus occurred in squamous cell carcinoma of the lung. Therefore, Aggrus could be a useful diagnostic marker for squamous cell carcinoma of the lung. PMID:16006773

  9. Platelet Adhesion to Podoplanin Under Flow is Mediated by the Receptor CLEC-2 and Stabilised by Src/Syk-Dependent Platelet Signalling

    PubMed Central

    Pollitt, Alice Y.; Lowe, Kate; Latif, Arusa; Nash, Gerard B.

    2015-01-01

    Summary Platelet-specific deletion of CLEC-2, which signals through Src and Syk kinases, or global deletion of its ligand podoplanin results in blood-filled lymphatics during mouse development. Platelet-specific Syk deficiency phenocopies this defect, indicating that platelet activation is required for lymphatic development. In the present study, we investigated whether CLEC-2-podoplanin interactions could support platelet arrest from blood flow and whether platelet signalling is required for stable platelet adhesion to lymphatic endothelial cells (LECs) and recombinant podoplanin under flow. Perfusion of human or mouse blood over human LEC monolayers led to platelet adhesion and aggregation. Following αIIbβ3 blockade, individual platelets still adhered. Platelet binding occurred at venous but not arterial shear rates. There was no adhesion using CLEC-2-deficient blood or to vascular endothelial cells (which lack podoplanin). Perfusion of human blood over human Fc-podoplanin (hFcPDPN) in the presence of monoclonal antibody IV.3 to block FcγRIIA receptors led to platelet arrest at similar shear rates to those used on LECs. Src and Syk inhibitors significantly reduced global adhesion of human or mouse platelets to LECs and hFcPDPN. A similar result was seen using Syk-deficient mouse platelets. Reduced platelet adhesion was due to a decrease in the stability of binding. In conclusion, our data reveal that CLEC-2 is an adhesive receptor that supports platelet arrest to podoplanin under venous shear. Src/Syk-dependent signalling stabilises platelet adhesion to podoplanin, providing a possible molecular mechanism contributing to the lymphatic defects of Syk-deficient mice. PMID:25694214

  10. Markers of endothelial cell activation and injury in childhood haemolytic uraemic syndrome.

    PubMed

    Nevard, C H; Blann, A D; Jurd, K M; Haycock, G B; Hunt, B J

    1999-08-01

    Diarrhoea-associated haemolytic uraemic syndrome (D+ HUS) is usually caused by verotoxin-producing Escherichia coli. Histology shows endothelial swelling with localised thrombus. Activation of coagulation and fibrinolysis also occurs. These facts, combined with the knowledge that recovery usually follows within weeks, led us to hypothesise that verotoxin causes localised endothelial cell activation but not injury. Markers of endothelial cell activation and injury were measured serially in 30 children with acute D+ HUS, healthy children, and children receiving chronic dialysis. Interpretation of markers was complicated by the renal dysfunction characteristic of D+ HUS. Nevertheless there was no evidence for endothelial cell injury, as soluble tissue factor levels were not increased and soluble thrombomodulin levels were lower than dialysed controls (P<0.001). In the acute phase, soluble vascular cell adhesion molecule levels were raised above normal (P<0.001), but were lower than dialysed controls (P<0.001), and soluble E-selectin levels were not significantly increased compared with normal controls (P=0.2). Hence, there was no evidence for endothelial cell damage or endothelial cell activation by the time children reached hospital; but this study did not exclude the possibility that endothelial cell activation occurred prior to hospital admission. PMID:10452276

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  12. Lymphatic endothelial differentiation in pulmonary lymphangioleiomyomatosis cells.

    PubMed

    Davis, Jennifer M; Hyjek, Elizabeth; Husain, Aliya N; Shen, Le; Jones, Jennifer; Schuger, Lucia A

    2013-08-01

    Pulmonary lymphangioleiomyomatosis (LAM) is a rare, low-grade neoplasm affecting almost exclusively women of childbearing age. LAM belongs to the family of perivascular epithelioid cell tumors, characterized by spindle and epithelioid cells with smooth muscle and melanocytic differentiation. LAM cells infiltrate the lungs, producing multiple, bilateral lesions rich in lymphatic channels and forming cysts, leading to respiratory insufficiency. Here we used antibodies against four lymphatic endothelial markers-podoplanin (detected by D2-40), prospero homeobox 1 (PROX1), vascular endothelial growth factor receptor 3 (VEGFR-3), and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1)-to determine whether LAM cells show lymphatic differentiation. Twelve of 12 diagnostic biopsy specimens (early-stage LAM) and 19 of 19 explants (late-stage LAM) showed immunopositivity for D2-40 in most neoplastic cells. PROX1, VEGFR-3, and LYVE1 immunoreactivity varied from scarce in the early stage to abundant in the late stage. Lymphatic endothelial, smooth muscle, and melanocytic markers were partially co-localized. These findings indicate that lymphatic endothelial differentiation is a feature of LAM and provide evidence of a previously unidentified third lineage of differentiation in this neoplasm. This study has implications for the histological diagnosis of LAM, the origin of the neoplastic cells, and potential future treatment with drugs targeting lymphangiogenesis. PMID:23609227

  13. Identification of a biochemical marker for endothelial dysfunction using Raman spectroscopy.

    PubMed

    Rygula, A; Pacia, M Z; Mateuszuk, L; Kaczor, A; Kostogrys, R B; Chlopicki, S; Baranska, M

    2015-04-01

    In the present work, we propose the spectroscopic approach to identify biochemical alterations in endothelial dysfunction. The method is based on the quantification of the ratio of phenylalanine (Phe) to tyrosine (Tyr) contents in the endothelium. The synthesis of Tyr from Phe requires the presence of tetrahydrobiopterin (BH4) as a cofactor of phenylalanine hydroxylase (PAH). Limitation of BH4 availability in the endothelium is a hallmark endothelial nitric oxide synthase (eNOS) dysfunction that may also lead to PAH dysfunction and a fall in Tyr contents. Using Raman spectra, the ratio of marker bands of Tyr to Phe was calculated and the pathological state of the endothelium was detected. We provide evidence that Phe/Tyr ratio analysis by Raman spectroscopy discriminate endothelial dysfunction in ApoE/LDLR(-/-) mice as compared to control mice. PMID:25664353

  14. Protein markers and differentiation in culture for Schlemm's canal endothelial cells.

    PubMed

    Perkumas, K M; Stamer, W D

    2012-03-01

    The two cell types that populate the human conventional outflow pathway, Schlemm's canal (SC) and trabecular meshwork (TM) regulate intraocular pressure. In culture, SC and TM cells have been useful tools toward understanding their respective roles in conventional outflow homeostasis. Unfortunately, currently available protein markers that distinguish SC from TM cells are limited, motivating the present study. Antibodies that specifically recognize different vascular endothelial markers were used to probe lysates from mature cell monolayers subjected to SDS-PAGE followed by western blot analyses. Results show that SC and TM cells both expressed many of the endothelial candidate proteins investigated, such as Robo1/4, Tie2/TEK, VEGF-R1/R2, VCAM-1, eNOS and neuropilin-1. In contrast, all SC cell strains tested (n=11) expressed two proteins, fibulin-2 and vascular endothelial (VE) cadherin, not expressed by TM cells. To examine changes in VE-cadherin expression and cell-cell junction formation, indicated by transendothelial electrical resistance (TEER), SC cells were seeded onto filters at confluence and growth factors were withdrawn. Culturing cells in media containing adult bovine serum rather than fetal bovine serum resulted in a 75% mean increase in TEER and 67% corresponding average increase in VE-cadherin expression (p<0.05). While both TM and SC cells form monolayers, are contact inhibited, share some endothelial responsibilities and several endothelial protein markers, SC cells uniquely express at least two proteins which likely reflect a distinction in cellular responsibilities in vivo. One of these responsibilities, maintenance of the blood-aqueous barrier, can be modeled in culture upon withdrawal of growth factors from SC cell monolayers. PMID:22210126

  15. The use of novel lymphatic endothelial cell-specific immunohistochemical markers to differentiate cutaneous angiosarcomas in dogs.

    PubMed

    Halsey, C H C; Worley, D R; Curran, K; Charles, J B; Ehrhart, E J

    2016-09-01

    Lymphangiosarcomas are uncommon vascular neoplasms that arise from lymphatic endothelial cells (LECs). They efface and replace normal subcutaneous tissue and are characterised by arborising, vascular channels lined by a single layer of pleomorphic endothelial cells and a paucity of erythrocytes. Lymphangiosarcomas are architecturally similar to hemangiosarcomas, a common malignancy of vascular origin arising from blood vascular endothelial cells. Common immunohistochemical markers for vascular endothelium, such as Factor VIII-related antigen (F8RA) and CD31, have traditionally been used to confirm the diagnosis of tumours of vascular origin. However, these markers fail to differentiate between lymphangiosarcoma and hemangiosarcoma, which often show overlapping morphologic features, disparate clinical behaviour and require different treatment modalities. Here we describe the use of two novel LEC-specific markers, lymphatic vessel endothelial receptor-1 (LYVE-1) and prospero-related homeobox gene-1 (PROX-1), to further differentiate between vascular tumours of lymphatic (lymphangiosarcoma) and blood (hemangiosarcoma) endothelial cell origin in the dog. PMID:24593773

  16. Glutamate dependent NMDA receptor 2D is a novel angiogenic tumour endothelial marker in colorectal cancer

    PubMed Central

    Ward, Stephen; Heath, Victoria L.; Ismail, Tariq; Bicknell, Roy

    2016-01-01

    Current vascular-targeted therapies in colorectal cancer (CRC) have shown limited benefit. The lack of novel, specific treatment in CRC has been hampered by a dearth of specific endothelial markers. Microarray comparison of endothelial gene expression in patient-matched CRC and normal colon identified a panel of putative colorectal tumour endothelial markers. Of these the glutamate dependent NMDA receptor GRIN2D emerged as the most interesting target. GRIN2D expression was shown to be specific to colorectal cancer vessels by RTqPCR and IHC analysis. Its expression was additionally shown be predictive of improved survival in CRC. Targeted knockdown studies in vitro demonstrated a role for GRIN2D in endothelial function and angiogenesis. This effect was also shown in vivo as vaccination against the extracellular region of GRIN2D resulted in reduced vascularisation in the subcutaneous sponge angiogenesis assay. The utility of immunologically targeting GRIN2D in CRC was demonstrated by the vaccination approach inhibiting murine CRC tumour growth and vascularisation. GRIN2D represents a promising target for the future treatment of CRC. PMID:26943033

  17. Selective targeting of liver cancer with the endothelial marker CD146

    PubMed Central

    Thomann, Stefan; Longerich, Thomas; Bazhin, Alexandr V.; Mier, Walter; Schemmer, Peter; Ryschich, Eduard

    2014-01-01

    Hepatocellular carcinomas are well-vascularized tumors; the endothelial cells in these tumors have a specific phenotype. Our aim was to develop a new approach for tumor-specific drug delivery with monoclonal antibody targeting of endothelial ligands. CD146, a molecule expressed on the endothelial surface of hepatocellular carcinoma, was identified as a promising candidate for targeting. In the present study, endothelial cells immediately captured circulating anti-CD146 (ME-9F1) antibody, while antibody binding in tumors was significantly higher than in hepatic endothelium. Macroscopically, after intravenous injection, there were no differences in the mean accumulation of anti-CD146 antibody in tumor compared to liver tissue, due to a compensating higher blood vessel density in the liver tissue. Additional blockade of nontumoral epitopes and intra-arterial administration, improved selective antibody capture in the tumor microvasculature and largely prevented antibody distribution in the lung and liver. The potential practical use of this approach was demonstrated by imaging of radionuclide-labeled ME-9F1 antibody, which showed excellent tumor-selective uptake. Our results provide a promising principle for the use of endothelial markers for intratumoral drug delivery. Tumor endothelium–based access might offer new opportunities for the imaging and therapy of hepatocellular carcinoma and other liver malignancies. PMID:25238265

  18. Eculizumab reduces complement activation, inflammation, endothelial damage, thrombosis, and renal injury markers in aHUS

    PubMed Central

    Cofiell, Roxanne; Kukreja, Anjli; Bedard, Krystin; Yan, Yan; Mickle, Angela P.; Ogawa, Masayo; Bedrosian, Camille L.

    2015-01-01

    Atypical hemolytic uremic syndrome (aHUS) is a genetic, life-threatening disease characterized by uncontrolled complement activation, systemic thrombotic microangiopathy (TMA), and vital organ damage. We evaluated the effect of terminal complement blockade with the anti-C5 monoclonal antibody eculizumab on biomarkers of cellular processes involved in TMA in patients with aHUS longitudinally, during up to 1 year of treatment, compared with in healthy volunteers. Biomarker levels were elevated at baseline in most patients, regardless of mutational status, plasma exchange/infusion use, platelet count, or lactate dehydrogenase or haptoglobin levels. Eculizumab reduced terminal complement activation (C5a and sC5b-9) and renal injury markers (clusterin, cystatin-C, β2-microglobulin, and liver fatty acid binding protein-1) to healthy volunteer levels and reduced inflammation (soluble tumor necrosis factor receptor-1), coagulation (prothrombin fragment F1+2 and d-dimer), and endothelial damage (thrombomodulin) markers to near-normal levels. Alternative pathway activation (Ba) and endothelial activation markers (soluble vascular cell adhesion molecule-1) decreased but remained elevated, reflecting ongoing complement activation in aHUS despite complete terminal complement blockade. These results highlight links between terminal complement activation and inflammation, endothelial damage, thrombosis, and renal injury and underscore ongoing risk for systemic TMA and progression to organ damage. Further research regarding underlying complement dysregulation is warranted. This trial was registered at www.clinicaltrials.gov as #NCT01194973. PMID:25833956

  19. The Splicing Factor SRSF1 as a Marker for Endothelial Senescence.

    PubMed

    Blanco, Francisco Javier; Bernabéu, Carmelo

    2012-01-01

    Aging is the major risk factor per se for the development of cardiovascular diseases. The senescence of the endothelial cells (ECs) that line the lumen of blood vessels is the cellular basis for these age-dependent vascular pathologies, including atherosclerosis and hypertension. During their lifespan, ECs may reach a stage of senescence by two different pathways; a replicative one derived from their preprogrammed finite number of cell divisions; and one induced by stress stimuli. Also, certain physiological stimuli, such as transforming growth factor-β, are able to modulate cellular senescence. Currently, the cellular aging process is being widely studied to identify novel molecular markers whose changes correlate with senescence. This review focuses on the regulation of alternative splicing mediated by the serine-arginine splicing factor 1 (SRSF1, or ASF/SF2) during endothelial senescence, a process that is associated with a differential subcellular localization of SRSF1, which typically exhibits a scattered distribution throughout the cytoplasm. Based on its senescence-dependent involvement in alternative splicing, we postulate that SRSF1 is a key marker of EC senescence, regulating the expression of alternative isoforms of target genes such as endoglin (ENG), vascular endothelial growth factor A (VEGFA), tissue factor (T3), or lamin A (LMNA) that integrate in a common molecular senescence program. PMID:22470345

  20. ERG is a novel and reliable marker for endothelial cells in central nervous system tumors.

    PubMed

    Haber, Matthew A; Iranmahboob, Amir; Thomas, Cheddhi; Liu, Mengling; Najjar, Amanda; Zagzag, David

    2015-01-01

    ETS-related gene (ERG) is a transcription factor that has been linked to angiogenesis. Very little research has been done to assess ERG expression in central nervous system (CNS) tumors. We evaluated 57 CNS tumors, including glioblastomas (GBMs) and hemangioblastomas (HBs), as well as two arteriovenous malformations and four samples of normal brain tissue with immunohistochemistry using a specific ERG rabbit monoclonal antibody. In addition, immunostains for CD31, CD34, and α-smooth muscle actin (α-SMA) were performed on all samples. CD31 demonstrated variable and sometimes weak immunoreactivity for endothelial cells. Furthermore, in 1 case of a GBM, CD34 stained not only endothelial cells, but also tumor cells. In contrast, we observed that ERG was only expressed in the nuclei of endothelial cells, for example, in the hyperplastic vascular complexes that comprise the glomeruloid microvascular proliferation seen in GBMs. Conversely, α-SMA immunoreactivity was identified in the abluminal cells of these hyperplastic vessels. Quantitative evaluation with automated methodology and custom Matlab 2008b software was used to calculate percent staining of ERG in each case. We observed significantly higher quantitative expression of ERG in HBs than in other CNS tumors. Our results show that ERG is a novel, reliable, and specific marker for endothelial cells within CNS tumors that can be used to better study the process of neovascularization. PMID:25881913

  1. ERG is a novel and reliable marker for endothelial cells in central nervous system tumors

    PubMed Central

    Haber, Matthew A.; Iranmahboob, Amir; Thomas, Cheddhi; Liu, Mengling; Najjar, Amanda; Zagzag, David

    2015-01-01

    ETS-related gene (ERG) is a transcription factor that has been linked to angiogenesis. Very little research has been done to assess ERG expression in central nervous system (CNS) tumors. We evaluated 57 CNS tumors, including glioblastomas (GBMs) and hemangioblastomas (HBs), as well as two arteriovenous malformations and four samples of normal brain tissue with immunohistochemistry using a specific ERG rabbit monoclonal antibody. In addition, immunostains for CD31, CD34, and α-smooth muscle actin (α-SMA) were performed on all samples. CD31 demonstrated variable and sometimes weak immunoreactivity for endothelial cells. Furthermore, in 1 case of a GBM, CD34 stained not only endothelial cells, but also tumor cells. In contrast, we observed that ERG was only expressed in the nuclei of endothelial cells, for example, in the hyperplastic vascular complexes that comprise the glomeruloid microvascular proliferation seen in GBMs. Conversely, α-SMA immunoreactivity was identified in the abluminal cells of these hyperplastic vessels. Quantitative evaluation with automated methodology and custom Matlab 2008b software was used to calculate percent staining of ERG in each case. We observed significantly higher quantitative expression of ERG in HBs than in other CNS tumors. Our results show that ERG is a novel, reliable, and specific marker for endothelial cells within CNS tumors that can be used to better study the process of neovascularization. PMID:25881913

  2. Discovery of molecular markers to discriminate corneal endothelial cells in the human body.

    PubMed

    Yoshihara, Masahito; Ohmiya, Hiroko; Hara, Susumu; Kawasaki, Satoshi; Hayashizaki, Yoshihide; Itoh, Masayoshi; Kawaji, Hideya; Tsujikawa, Motokazu; Nishida, Kohji

    2015-01-01

    The corneal endothelium is a monolayer of hexagonal corneal endothelial cells (CECs) on the inner surface of the cornea. CECs are critical in maintaining corneal transparency through their barrier and pump functions. CECs in vivo have a limited capacity in proliferation, and loss of a significant number of CECs results in corneal edema called bullous keratopathy which can lead to severe visual loss. Corneal transplantation is the most effective method to treat corneal endothelial dysfunction, where it suffers from donor shortage. Therefore, regeneration of CECs from other cell types attracts increasing interests, and specific markers of CECs are crucial to identify actual CECs. However, the currently used markers are far from satisfactory because of their non-specific expression in other cell types. Here, we explored molecular markers to discriminate CECs from other cell types in the human body by integrating the published RNA-seq data of CECs and the FANTOM5 atlas representing diverse range of cell types based on expression patterns. We identified five genes, CLRN1, MRGPRX3, HTR1D, GRIP1 and ZP4 as novel markers of CECs, and the specificities of these genes were successfully confirmed by independent experiments at both the RNA and protein levels. Notably none of them have been documented in the context of CEC function. These markers could be useful for the purification of actual CECs, and also available for the evaluation of the products derived from other cell types. Our results demonstrate an effective approach to identify molecular markers for CECs and open the door for the regeneration of CECs in vitro. PMID:25807145

  3. Discovery of Molecular Markers to Discriminate Corneal Endothelial Cells in the Human Body

    PubMed Central

    Yoshihara, Masahito; Ohmiya, Hiroko; Hara, Susumu; Kawasaki, Satoshi; Hayashizaki, Yoshihide; Itoh, Masayoshi; Kawaji, Hideya; Tsujikawa, Motokazu; Nishida, Kohji

    2015-01-01

    The corneal endothelium is a monolayer of hexagonal corneal endothelial cells (CECs) on the inner surface of the cornea. CECs are critical in maintaining corneal transparency through their barrier and pump functions. CECs in vivo have a limited capacity in proliferation, and loss of a significant number of CECs results in corneal edema called bullous keratopathy which can lead to severe visual loss. Corneal transplantation is the most effective method to treat corneal endothelial dysfunction, where it suffers from donor shortage. Therefore, regeneration of CECs from other cell types attracts increasing interests, and specific markers of CECs are crucial to identify actual CECs. However, the currently used markers are far from satisfactory because of their non-specific expression in other cell types. Here, we explored molecular markers to discriminate CECs from other cell types in the human body by integrating the published RNA-seq data of CECs and the FANTOM5 atlas representing diverse range of cell types based on expression patterns. We identified five genes, CLRN1, MRGPRX3, HTR1D, GRIP1 and ZP4 as novel markers of CECs, and the specificities of these genes were successfully confirmed by independent experiments at both the RNA and protein levels. Notably none of them have been documented in the context of CEC function. These markers could be useful for the purification of actual CECs, and also available for the evaluation of the products derived from other cell types. Our results demonstrate an effective approach to identify molecular markers for CECs and open the door for the regeneration of CECs in vitro. PMID:25807145

  4. Effect of standard chemotherapy and antiangiogenic therapy on plasma markers and endothelial cells in colorectal cancer

    PubMed Central

    Ramcharan, K S; Lip, G Y H; Stonelake, P S; Blann, A D

    2014-01-01

    Introduction: The importance of the endothelium in angiogenesis and cancer is undisputed, and its integrity may be assessed by laboratory markers such as circulating endothelial cells (CECs), endothelial progenitor cells (EPCs), plasma von Willebrand factor (vWf), soluble E selectin, vascular endothelial growth factor (VEGF) and angiogenin. Antiantigenic therapy may be added to standard cytotoxic chemotherapy as a new treatment modality. We hypothesised that additional antiangiogenic therapy acts in a contrasting manner to that of standard chemotherapy on the laboratory markers. Methods: We recruited 68 patients with CRC, of whom 16 were treated with surgery alone, 32 were treated with surgery followed by standard chemotherapy (5-flurouracil), and 20 were treated with surgery followed by standard chemotherapy plus anti-VEGF therapy (Avastin). Peripheral blood was taken before surgery, and again 3 months and 6 months later. CD34+/CD45−/CD146+ CECs and CD34+/CD45−/CD309[KDR]+ EPCs were measured by flow cytometry, plasma markers by ELISA. Results: In each of the three groups, CECs and EPCs fell at 3 months but were back at pre-surgery levels at 6 months (P<0.05). VEGF was lower in both 3-and 6-month samples in the surgery-only and surgery plus standard chemotherapy groups (P<0.05), but in those on surgery followed by standard chemotherapy plus anti-VEGF therapy, low levels at 3 months (P<0.01) increased to pre-surgery levels at 6 months. In those having surgery and standard chemotherapy, soluble E selectin was lower, whereas angiogenin was higher at 6 months than at baseline (both P<0.05). Conclusions: We found disturbances in endotheliod cells regardless of treatment, whereas VEGF returned to levels before surgery in those on antiangiogenic therapy. These observations may have clinical and pathophysiological implications. PMID:25211664

  5. 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

    SciTech Connect

    Vallon, Mario; Rohde, Franziska; Janssen, Klaus-Peter; Essler, Markus

    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, 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.

  6. Identification of novel molecular markers through transcriptomic analysis in human fetal and adult corneal endothelial cells.

    PubMed

    Chen, Yinyin; Huang, Kevin; Nakatsu, Martin N; Xue, Zhigang; Deng, Sophie X; Fan, Guoping

    2013-04-01

    The corneal endothelium is composed of a monolayer of corneal endothelial cells (CECs), which is essential for maintaining corneal transparency. To better characterize CECs in different developmental stages, we profiled mRNA transcriptomes in human fetal and adult corneal endothelium with the goal to identify novel molecular markers in these cells. By comparing CECs with 12 other tissue types, we identified 245 and 284 signature genes that are highly expressed in fetal and adult CECs, respectively. Functionally, these genes are enriched in pathways characteristic of CECs, including inorganic anion transmembrane transporter, extracellular matrix structural constituent and cyclin-dependent protein kinase inhibitor activity. Importantly, several of these genes are disease target genes in hereditary corneal dystrophies, consistent with their functional significance in CEC physiology. We also identified stage-specific markers associated with CEC development, such as specific members in the transforming growth factor beta and Wnt signaling pathways only expressed in fetal, but not in adult CECs. Lastly, by the immunohistochemistry of ocular tissues, we demonstrated the unique protein localization for Wnt5a, S100A4, S100A6 and IER3, the four novel markers for fetal and adult CECs. The identification of a new panel of stage-specific markers for CECs would be very useful for characterizing CECs derived from stem cells or ex vivo expansion for cell replacement therapy. PMID:23257286

  7. Markers of Inflammation, Endothelial Activation and Autoimmunity in Adolescent Female Gymnasts

    PubMed Central

    Alshammari, Eyad; Shafi, Shahida; Nurmi-Lawton, Jaana; Burut, Dayangku Fatiha Pengiran; Lanham-New, Susan; Ferns, Gordon

    2010-01-01

    High levels of physical activity have been linked to benefits in cardiovascular and bone health by affecting, in part, changes in proinflammatory profile. Therefore, we have aimed to assess the effects of intensive training on markers of inflammation, endothelial activation and auto-immunity in the absence of the potential confounding effects of incident atherosclerosis. The subjects comprised 25 competitive gymnasts and 19 healthy sedentary adolescent females, aged 8-17 years. Serum soluble intercellular adhesion molecule 1 (sICAM-1), high sensitivity C-reactive protein (hsCRP), heat shock protein 27 (Hsp27) and Hsp27 antibody titres were measured by ELISAs in a sample of adolescent girls who were either physically active (competitive gymnasts) or sedentary. The association between age, body mass index (BMI), dietary intake, serum hsCRP, sICAM-1 and Hsp27 antigen and antibody titres were determined. The mean serum sICAM-1 concentrations were significantly higher in the gymnasts compared to the sedentary females (0.29 ± 0.02 versus 0.23 ± 0.01 mg·L-1, p < 0.01). In contrast serum hsCRP concentrations were substantially lower in the gymnasts compared to the sedentary adolescent females (0.49 ± 0.03 versus 1.38 ± 0.19 mg·L-1, p < 0.001). Differences remained significant after adjustment for anthropometric factors. We also found that serum Hsp27 antigen concentrations were determined by dietary saturated fat intake (p < 0.001), and antibody titres to Hsp27 were determined by dietary PUFA (p < 0.001) after adjustment for BMI. Our findings show that young female gymnasts have an altered profile of inflammatory markers and endothelial activation compared to their less physically active peers. Key points Results showed that adolescent young female gymnasts have an altered serum inflammatory markers and endothelial activation, compared to their less physically active peers. Physical activities improved immune system. Differences in these biochemical data kept

  8. VASCULAR ENDOTHELIAL GROWTH FACTOR RECEPTOR 2 (VEGFR2) AS A MARKER FOR MALIGNANT VASCULAR TUMORS AND MESOTHELIOMA – IMMUNOHISTOCHEMICAL STUDY OF 262 VASCULAR ENDOTHELIAL AND 1640 NONVASCULAR TUMORS

    PubMed Central

    Miettinen, Markku; Rikala, Maarit-Sarlomo; Rysz, Janusz; Lasota, Jerzy; Wang, Zeng-Feng

    2012-01-01

    Vascular endothelial growth factor receptor 2 (VEGFR2) is a primary responder to vascular endothelial growth factor signal, and thereby regulates endothelial migration and proliferation. This receptor is expressed in endothelial cells and some vascular tumors, but many reports also detail its expression in carcinomas and lymphomas. VEGFR2 is a potential cell type marker, and data on VEGFR2 expression may also have therapeutic significance in view of recent availability of VEGFR2 inhibitors. In this study we immunohistochemically examined 262 vascular endothelial and 1640 non-vascular tumors and selected non-neoplastic tissues with a VEGFR2-specific rabbit monoclonal antibody 55B11. In early human embryo, VEFGR2 was expressed in endothelia of developing capillaries, thoracic duct, great vessels, hepatic sinusoids, epidermis, and mesothelia. In late first trimester fetus peripheral soft tissues, VEGFR2 was restricted to capillary endothelia, chrondrocytes, and superficial portion of the epidermis. In normal adult tissues, it was restricted to endothelia and mesothelia. VEGFR2 was consistently expressed in angiosarcomas, Kaposi sarcomas, and retiform hemangioendotheliomas. It was detected only in half of epithelioid hemangioendotheliomas (15/27), usually focally. VEGFR2 was strongly expressed in most capillary hemangiomas and weakly or focally in cavernous, venous, and spindle cell hemangiomas, and lymphangiomas. Malignant epithelial mesothelioma was found to be a unique epithelial neoplasm with a strong and nearly consistent VEGFR2 expression, including membrane staining (35/38). Approximately 10% of squamous cell carcinomas and 23% of pulmonary adenocarcinomas contained focal positivity. The only non-endothelial mesenchymal tumors found VEGFR2-positive were biphasic synovial sarcoma (focal epithelial expression), and chordoma. All melanomas and lymphomas were negative. VEGFR2 is a promising marker for malignant vascular tumors and malignant epithelioid mesothelioma

  9. The effects of endothelial lipase gene (LIPG) variants on inflammation marker levels and atherosclerosis development.

    PubMed

    Dalan, Altay Burak; Toptaş, Bahar; Buğra, Zehra; Polat, Nihat; Yılmaz-Aydoğan, Hülya; Çimen, Arif; Isbir, Turgay

    2013-08-01

    Atherosclerosis is a major pathological process related with several important adverse vascular events including coronary artery disease, stroke, and peripheral arterial disease. Endothelial lipase is an enzyme the activity of which affects all of lipoproteins, whereas HDL is the main substrate. The purpose of our study was to investigate the effects of endothelial lipase gene polymorphism and inflammation markers (CRP, IL-1β, IL-6, IL-8 and TNF-α) in the atherosclerosis. 104 patients with atherosclerosis and 76 healthy individuals were included in the study. LIPG -584C/T polymorphism gene polymorphisms were assessed with PCR-RFLP method. The serum CRP levels were measured by turbidimetric method using a biochemistry autoanalyzer, whereas serum IL-1β, IL-6, IL-8, TNF-α levels were determined by enzyme-linked immunosorbent assay. In this study, we found that the frequencies of TC genotype are more prevalent in patients than controls. We found a statistically significant difference of IL-6 levels between patient and control group. Our findings suggest that T allele might play a potential role in the susceptibility to atherogenesis in the Turkish population. PMID:23673478

  10. Immunohistochemical Expression of VEGF and Podoplanin in Uterine Cervical Squamous Intraepithelial Lesions

    PubMed Central

    Belfort-Mattos, Patrícia Napoli; Focchi, Gustavo Rubino de Azevedo; Ribalta, Julisa Chamorro Lascasas; Megale De Lima, Tatiana; Nogueira Carvalho, Carmen Regina; Kesselring Tso, Fernanda; De Góis Speck, Neila Maria

    2016-01-01

    VEGF and podoplanin (PDPN) have been identified as angiogenesis and/or lymphangiogenesis regulators and might be essential to restrict tumor growth, progression, and metastasis. In the present study, we evaluate the association between the expression of these markers and CIN grade. Immunohistochemistry was performed in 234 uterine cervical samples using conventional histologic sections or TMA with the monoclonal antibodies to VEGF (C-1 clone) and podoplanin (D2-40 clone). Positive-staining rates of VEGF in 191 CIN specimens were significantly associated with histological grade (P < 0.001). Negative and/or focal immunostaining for PDPN were more frequent in CIN 3 (P = 0.016). We found that patients with CIN 3 more frequently had strong and more diffuse staining for VEGF and diminished staining for PDPN (P = 0.018). Strong and more diffuse VEGF immunoexpressions in CIN 2 and CIN 3 were detected when compared to CIN 1. Negative and/or focal PDPN immunoexpression appear to be more frequent in CIN 3. Moderate to strong VEGF expression may be a tendency among patients with high-grade lesions and diminished PDPN expression. PMID:27313335

  11. Long-term Exposure to Air Pollution and Markers of Inflammation, Coagulation, and Endothelial Activation

    PubMed Central

    Hajat, Anjum; Allison, Matthew; Diez-Roux, Ana V.; Jenny, Nancy Swords; Jorgensen, Neal W.; Szpiro, Adam A.; Vedal, Sverre; Kaufman, Joel D.

    2015-01-01

    Background Air pollution is associated with cardiovascular disease, and systemic inflammation may mediate this effect. We assessed associations between long- and short-term concentrations of air pollution and markers of inflammation, coagulation, and endothelial activation. Methods We studied participants from the Multi-Ethnic Study of Atherosclerosis from 2000 to 2012 with repeat measures of serum C-reactive protein (CRP), interleukin-6 (IL-6), fibrinogen, D-dimer, soluble E-selectin, and soluble Intercellular Adhesion Molecule-1. Annual average concentrations of ambient fine particulate matter (PM2.5), individual-level ambient PM2.5 (integrating indoor concentrations and time–location data), oxides of nitrogen (NOx), nitrogen dioxide (NO2), and black carbon were evaluated. Short-term concentrations of PM2.5 reflected the day of blood draw, day prior, and averages of prior 2-, 3-, 4-, and 5-day periods. Random-effects models were used for long-term exposures and fixed effects for short-term exposures. The sample size was between 9,000 and 10,000 observations for CRP, IL-6, fibrinogen, and D-dimer; approximately 2,100 for E-selectin; and 3,300 for soluble Intercellular Adhesion Molecule-1. Results After controlling for confounders, 5 µg/m3 increase in long-term ambient PM2.5 was associated with 6% higher IL-6 (95% confidence interval = 2%, 9%), and 40 parts per billion increase in long-term NOx was associated with 7% (95% confidence interval = 2%, 13%) higher level of D-dimer. PM2.5 measured at day of blood draw was associated with CRP, fibrinogen, and E-selectin. There were no other positive associations between blood markers and short- or long-term air pollution. Conclusions These data are consistent with the hypothesis that long-term exposure to air pollution is related to some markers of inflammation and fibrinolysis. PMID:25710246

  12. Tumor endothelial marker 1–specific DNA vaccination targets tumor vasculature

    PubMed Central

    Facciponte, John G.; Ugel, Stefano; De Sanctis, Francesco; Li, Chunsheng; Wang, Liping; Nair, Gautham; Sehgal, Sandy; Raj, Arjun; Matthaiou, Efthymia; Coukos, George; Facciabene, Andrea

    2014-01-01

    Tumor endothelial marker 1 (TEM1; also known as endosialin or CD248) is a protein found on tumor vasculature and in tumor stroma. Here, we tested whether TEM1 has potential as a therapeutic target for cancer immunotherapy by immunizing immunocompetent mice with Tem1 cDNA fused to the minimal domain of the C fragment of tetanus toxoid (referred to herein as Tem1-TT vaccine). Tem1-TT vaccination elicited CD8+ and/or CD4+ T cell responses against immunodominant TEM1 protein sequences. Prophylactic immunization of animals with Tem1-TT prevented or delayed tumor formation in several murine tumor models. Therapeutic vaccination of tumor-bearing mice reduced tumor vascularity, increased infiltration of CD3+ T cells into the tumor, and controlled progression of established tumors. Tem1-TT vaccination also elicited CD8+ cytotoxic T cell responses against murine tumor-specific antigens. Effective Tem1-TT vaccination did not affect angiogenesis-dependent physiological processes, including wound healing and reproduction. Based on these data and the widespread expression of TEM1 on the vasculature of different tumor types, we conclude that targeting TEM1 has therapeutic potential in cancer immunotherapy. PMID:24642465

  13. Relation between Endothelial Nitric Oxide Synthase Genotypes and Oxidative Stress Markers in Larynx Cancer

    PubMed Central

    Yanar, K.; Çakatay, U.; Aydın, S.; Verim, A.; Atukeren, P.; Özkan, N. E.; Karatoprak, K.; Cebe, T.; Turan, S.; Ozkök, E.; Korkmaz, G.; Cacına, C.; Küçükhüseyin, O.; Yaylım, İ.

    2016-01-01

    Nitric oxide synthase (eNOS/NOS3) is responsible for the endothelial synthesis of nitric oxide (NO•). G894T polymorphism leads to the amino acid substitution from Glu298Asp that causes lower NOS3 activity and basal NO• production in NOS3 894T (298Asp) allele carriers compared with the GG homozygotes. NO• acts as an antioxidant protecting against Fenton's reaction which generates highly reactive hydroxyl radicals. Allelic variation of NOS3 may influence an individual's risk of laryngeal cancer (LC). In the current study we have examined the possible relationship between NOS3 G894T genotypes and various systemic oxidative damage markers such as protein carbonyl, advanced oxidation protein products, Cu, Zn-superoxide dismutase, thiol group fractions, and lipid hydroperoxides in LC patients. Genotyping was carried out by PCR-RFLP. In LC patients with TT genotype, Cu, Zn-superoxide dismutase activities and nonprotein thiol levels were significantly higher than the controls. In patients with GT and GG genotype, high levels of lipid hydroperoxides showed statistical significance when compared to controls. Our results indicate a potential relationship among G894T polymorphism of NOS3, and impaired redox homeostasis. Further studies are required to determine the role of NOS3 gene polymorphism and impaired plasma redox homeostasis. PMID:26682008

  14. Carotid Endothelial VCAM-1 Is an Early Marker of Carotid Atherosclerosis and Predicts Coronary Artery Disease in Swine

    PubMed Central

    Masseau, I.; Bowles, D. K.

    2015-01-01

    Objective The aim was to determine if endothelial VCAM-1 (eVCAM-1) expression in the common carotid artery (CCA) would correlate with predictive markers of atherosclerotic disease, would precede reduction of markers of endothelial cell function and would predict coronary artery disease (CAD). Methods and results Carotid arterial segments (bifurcation, proximal and distal CCA) were harvested from 14 and 24 month-old male castrated familial hypercholesterolemic (FH) swine, a model of spontaneous atherosclerosis. Quantification of local expression of eVCAM-1, intimal macrophage accumulation, oxidative stress, intima-media (I/M) ratio, intima-media thickness (IMT), endothelial nitric oxide synthase (eNOS) and phosphorylated eNOS (p-eNOS) in selected regions of the carotids revealed a relationship between local inflammation and atheroscle-rotic plaque progression. Importantly, inflammation was not uniform throughout the CCA. Endo-thelial VCAM-1 expression was the greatest at the bifurcation and increased with age. Finally, eV-CAM-1 best estimated the severity of CAD compared to blood levels of glucose, hypercholestero-lemia, carotid IMT, and p-eNOS. Conclusion Our data suggested that eVCAM-1 was closely associated with atherosclerotic plaque progression and preceded impairment of EDD. Thus, this study supported the use of carotid VCAM-1 targeting agents to estimate the severity of CAD. PMID:26702331

  15. Effects of the contraceptive skin patch and subdermal contraceptive implant on markers of endothelial cell activation and inflammation.

    PubMed

    Hernandez-Juarez, Jesus; Sanchez-Serrano, Juan Carlos; Moreno-Hernandez, Manuel; Alvarado-Moreno, Jose Antonio; Hernandez-Lopez, Jose Rubicel; Isordia-Salas, Irma; Majluf-Cruz, Abraham

    2015-07-01

    Changes in blood coagulation factors may partially explain the association between hormonal contraceptives and thrombosis. Therefore, the likely effects of the contraceptive skin patch and subdermal contraceptive implant on levels of inflammatory markers and endothelial activation were analyzed. This was an observational, prospective, longitudinal, nonrandomized study composed of 80 women between 18 and 35 years of age who made the decision to use the contraceptive skin patch or subdermal contraceptive implant. vascular cell adhesion molecule-1 (VCAM-1), endothelial cell leukocyte adhesion molecule-1 (ELAM-1), von Willebrand factor (VWF), and plasminogen activator inhibitor type 1(PAI-1) as well as high-sensitivity C-reactive protein (hsCRP) were assayed before and after 4 months of use of the contraceptive method. VCAM-1, VWF, and PAI-1 remained unchanged in the contraceptive skin patch group; however, a significant increase in hsCRP (0.29-0.50 mg/dL; P =.012) and a significant decrease in ELAM-1 (44-25 ng/mL; P =.022) were observed. A significant diminution in VCAM-1 (463-362 ng/mL; P =.022) was also found in the subdermal contraceptive implant group. Our results strongly suggest that these contraceptive methods do not induce endothelial activation after 4 months of use. Increase in hsCRP levels was unrelated to changes in markers of endothelial activation. PMID:25655356

  16. Circulating biologic markers of endothelial dysfunction in cerebral small vessel disease: A review.

    PubMed

    Poggesi, Anna; Pasi, Marco; Pescini, Francesca; Pantoni, Leonardo; Inzitari, Domenico

    2016-01-01

    The term cerebral small vessel disease (SVD) refers to a group of pathologic processes with various etiologies that affect small arteries, arterioles, venules, and capillaries of the brain. Magnetic resonance imaging (MRI) correlates of SVD are lacunes, recent small subcortical infarcts, white-matter hyperintensities, enlarged perivascular spaces, microbleeds, and brain atrophy. Endothelial dysfunction is thought to have a role in the mechanisms leading to SVD-related brain changes, and the study of endothelial dysfunction has been proposed as an important step for a better comprehension of cerebral SVD. Among available methods to assess endothelial function in vivo, measurement of molecules of endothelial origin in peripheral blood is currently receiving selective attention. These molecules include products of endothelial cells that change when the endothelium is activated, as well as molecules that reflect endothelial damage and repair. This review examines the main molecular factors involved in both endothelial function and dysfunction, and the evidence linking endothelial dysfunction with cerebral SVD, and gives an overview of clinical studies that have investigated the possible association between endothelial circulating biomarkers and SVD-related brain changes. PMID:26058695

  17. The stem cell marker prominin-1/CD133 interacts with vascular endothelial growth factor and potentiates its action.

    PubMed

    Adini, Avner; Adini, Irit; Ghosh, Kaustabh; Benny, Ofra; Pravda, Elke; Hu, Ron; Luyindula, Dema; D'Amato, Robert J

    2013-04-01

    Prominin-1, a pentaspan transmembrane protein, is a unique cell surface marker commonly used to identify stem cells, including endothelial progenitor cells and cancer stem cells. However, recent studies have shown that prominin-1 expression is not restricted to stem cells but also occurs in modified forms in many mature adult human cells. Although prominin-1 has been studied extensively as a stem cell marker, its physiological function of the protein has not been elucidated. We investigated prominin-1 function in two cell lines, primary human endothelial cells and B16-F10 melanoma cells, both of which express high levels of prominin-1. We found that prominin-1 directly interacts with the angiogenic and tumor survival factor vascular endothelial growth factor (VEGF) in both the primary endothelial cells and the melanoma cells. Knocking down prominin-1 in the endothelial cells disrupted capillary formation in vitro and decreased angiogenesis in vivo. Similarly, tumors derived from prominin-1 knockdown melanoma cells had a reduced growth rate in vivo. Further, melanoma cells with knocked down prominin-1 had diminished ability to interact with VEGF, which was associated with decreased bcl-2 protein levels and increased apoptosis. In vitro studies with soluble prominin-1 showed that it stabilized dimer formation of VEGF164, but not VEGF121. Taken together, our findings support the notion that prominin-1 plays an active role in cell growth through its ability to interact and potentiate the anti-apoptotic and pro-angiogenic activities of VEGF. Additionally, prominin-1 promotes tumor growth by supporting angiogenesis and inhibiting tumor cell apoptosis. PMID:23150059

  18. Endothelial Markers May Link Kidney Function to Cardiovascular Events in Type 2 Diabetes

    PubMed Central

    Maier, Christina; Clodi, Martin; Neuhold, Stephanie; Resl, Michael; Elhenicky, Marie; Prager, Rudolf; Moertl, Deddo; Strunk, Guido; Luger, Anton; Struck, Joachim; Pacher, Richard; Hülsmann, Martin

    2009-01-01

    OBJECTIVE The increased cardiovascular risk in diabetes has been linked to endothelial and renal dysfunction. The aim of this study was to investigate the role of stable fragments of the precursors of adrenomedullin, endothelin-1, vasopressin, and atrial natriuretic peptide in progression of cardiovascular disease in patients with diabetes. RESEARCH DESIGN AND METHODS This was a prospective, observational study design with a composite end point (death or unexpected admission to hospital due to a cardiovascular event) on 781 patients with type 2 diabetes (54 events, median duration of observation 15 months). The four stable precursor peptides midregional adrenomedullin (MR-proADM), midregional proatrial natriuretic peptide (MR-proANP), COOH-terminal proendothelin-1 (CT-proET-1), and COOH-terminal provasopressin or copeptin (CT-proAVP) were determined at baseline, and their association to renal function and cardiovascular events was studied using stepwise linear and Cox logistic regression analysis and receiver operating characteristic analysis, respectively. RESULTS MR-proADM, CT-proET-1, CT-proAVP, and MR-proANP were all elevated in patients with future cardiovascular events and independently correlated to serum creatinine. MR-proADM and MR-proANP were significant predictors of a future cardiovascular event, with MR-proANP being the stronger (area under the curve 0.802 ± 0.034, sensitivity 0.833, specificity 0.576, positive predictive value 0.132, and negative predictive value 0.978 with a cutoff value of 75 pmol/l). CONCLUSIONS The four serum markers of vasoactive and natriuretic peptides are related to both kidney function and cardiovascular events, thus linking two major complications of diabetes, diabetic nephropathy and cardiovascular disease. PMID:19564455

  19. MMP-9 expression is associated with leukocytic but not endothelial markers in brain arteriovenous malformations.

    PubMed

    Chen, Yongmei; Fan, Yongfeng; Poon, K Y Trudy; Achrol, Achal S; Lawton, Michael T; Zhu, Yiqian; McCulloch, Charles E; Hashimoto, Tomoki; Lee, Chanhung; Barbaro, Nicholas M; Bollen, Andrew W; Yang, Guo-Yuan; Young, William L

    2006-01-01

    Brain arteriovenous malformations (BAVM) have high matrix metalloproteinase-9 (MMP-9) expression, the source of which is unclear. We hypothesized MMP-9 production might be due to inflammation in BAVM. Compared to control brain tissues (n = 5), BAVM tissue (n = 139) had a higher expression (by ELISA) of myeloperoxidase (MPO) (193 +/- 189 vs. 6 +/- 3, ng/mg, P < .001), MMP-9 (28 +/- 32 vs. 0.7 +/- 0.6, ng/mg, P < .001), and IL-6 (102 +/- 218 vs. 0.1 +/- 0.1, pg/mg, P < .001), but not eNOS (114 +/- 87 vs. 65 +/- 9, pg/mg, P = .09). MMP-9 expression in BAVM highly correlated with myeloperoxidase (R2 = .76, P < .001), as well as with IL-6 (R2 = .32, P < .001). In contrast, MMP-9 in BAVM poorly correlated with the endothelial marker, eNOS (R2 = .03, P = .05), and CD31 (R2 = .004, P = .57). Compared to non-embolized patients (n = 46), patients with pre-operative embolization (n = 93) had higher levels of myeloperoxidase (236 +/- 205 vs. 106 +/- 108, ng/mg, P < .001) and MMP-9 (33 +/- 35 vs. 16 +/- 20, ng/mg, P < .001), however the correlation between MMP-9 and myeloperoxidase was equally strong for both groups (R2 = .69, n = 93, P < .001, for both). MMP-9 expression correlated with the lipocalin-MMP-9 complex, suggesting neutrophils as the MMP-9 source. MPO co-localized with majority of MMP-9 signal by immunohistochemistry. Our data suggest that inflammation is a prominent feature of BAVM lesional phenotype, and neutrophils appear to be a major source of MMP-9 in these lesions. PMID:16720380

  20. Effect of repetitive SCUBA diving on humoral markers of endothelial and central nervous system integrity.

    PubMed

    Bilopavlovic, Nada; Marinovic, Jasna; Ljubkovic, Marko; Obad, Ante; Zanchi, Jaksa; Pollock, Neal W; Denoble, Petar; Dujic, Zeljko

    2013-07-01

    During SCUBA diving decompression, there is a significant gas bubble production in systemic veins, with rather frequent bubble crossover to arterial side even in asymptomatic divers. The aim of the current study was to investigate potential changes in humoral markers of endothelial and brain damage (endothelin-1, neuron-specific enolase and S-100β) after repetitive SCUBA diving with concomitant assessment of venous gas bubble production and subsequent arterialization. Sixteen male divers performed four open-water no-decompression dives to 18 msw (meters of sea water) lasting 49 min in consecutive days during which they performed moderate-level exercise. Before and after dives 1 and 4 blood was drawn, and bubble production and potential arterialization were echocardiographically evaluated. In addition, a control dive to 5 msw was performed with same duration, water temperature and exercise load. SCUBA diving to 18 msw caused significant bubble production with arterializations in six divers after dive 1 and in four divers after dive 4. Blood levels of endothelin-1 and neuron-specific enolase did not change after diving, but levels of S-100β were significantly elevated after both dives to 18 msw and a control dive. Creatine kinase activity following a control dive was also significantly increased. Although serum S-100β levels were increased after diving, concomitant increase of creatine kinase during control, almost bubble-free, dive suggests the extracranial release of S-100β, most likely from skeletal muscles. Therefore, despite the significant bubble production and sporadic arterialization after open-water dives to 18 msw, the current study found no signs of damage to neurons or the blood-brain barrier. PMID:23400567

  1. Generation of pure lymphatic endothelial cells from human pluripotent stem cells and their therapeutic effects on wound repair

    PubMed Central

    Lee, Shin-Jeong; Park, Changwon; Lee, Ji Yoon; Kim, Sangsung; Kwon, Pil Jae; Kim, Woansang; Jeon, Yong Heui; Lee, Eugine; Yoon, Young-sup

    2015-01-01

    Human pluripotent stem cells (hPSCs) have emerged as an important source for cell therapy. However, to date, no studies demonstrated generation of purified hPSC-derived lymphatic endothelial cells (LECs) and tested their therapeutic potential in disease models. Here we sought to differentiate hPSCs into the LEC lineage, purify them with LEC markers, and evaluate their therapeutic effects. We found that an OP9-assisted culture system reinforced by addition of VEGF-A, VEGF-C, and EGF most efficiently generated LECs, which were then isolated via FACS-sorting with LYVE-1 and PODOPLANIN. These hPSC-derived LYVE-1+PODOPLANIN+cells showed a pure committed LEC phenotype, formed new lymphatic vessels, and expressed lymphangiogenic factors at high levels. These hPSC-derived LECs enhanced wound healing through lymphangiogenesis and lymphvasculogenesis. Here we report, for the first time, that LECs can be selectively isolated from differentiating hPSCs, and that these cells are potent for lymphatic vessel formation in vivo and wound healing. This system and the purified hPSC-derived LECs can serve as a new platform for studying LEC development as well as for cell therapy. PMID:26066093

  2. A Case of Abnormal Lymphatic-Like Differentiation and Endothelial Progenitor Cell Activation in Neovascularization Associated with Hemi-Retinal Vein Occlusion

    PubMed Central

    Loukovaara, Sirpa; Gucciardo, Erika; Repo, Pauliina; Lohi, Jouko; Salven, Petri; Lehti, Kaisa

    2015-01-01

    Purpose Pathological vascular differentiation in retinal vein occlusion (RVO)-related neovessel formation remains poorly characterized. The role of intraocular lymphatic-like differentiation or endothelial progenitor cell activity has not been studied in this disease. Methods Vitrectomy was performed in an eye with hemi-RVO; the neovessel membrane located at the optic nerve head was removed and subjected to immunohistochemistry. Characterization of the neovascular tissue was performed using hematoxylin and eosin, α-smooth muscle actin, and the pan-endothelial cell (EC) adhesion molecule CD31. The expression of lymphatic EC markers was studied by lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), podoplanin (PDPN), and prospero-related homeobox protein 1 (Prox-1). Potential vascular stem/progenitor cells were identified by active cellular proliferation (Ki67) and expression of the stem cell marker CD117. Results The specimen contained blood vessels lined by ECs and surrounded by pericytes. Immunoreactivity for LYVE-1 and Prox-1 was detected, with Prox-1 being more widely expressed in the active Ki67-positive lumen-lining cells. PDPN expression was instead found in the cells residing in the extravascular tissue. Expression of the stem cell markers CD117 and Ki67 suggested vascular endothelial progenitor cell activity. Conclusions Intraocular lymphatic-like differentiation coupled with progenitor cell activation may be involved in the pathology of neovessel formation in ischemia-induced human hemi-RVO. PMID:26327908

  3. [Selected endothelial hemostatic markers in patients with peripheral arterial disease after endovascular revascularization and restenosis formation].

    PubMed

    Kotschy, Daniel; Kotschy, Maria; Socha, Paweł; Masłowski, Leszek; Kwapisz, Justyna; Żuk, Natalia; Dubis, Joanna; Karczewski, Maciej; Witkiewicz, Wojciech

    2015-01-01

    Surgical and endovascular revascularization of ischemic legs in patients with peripheral arterial disease (PAD) can damage the arterial wall (endothelial and smooth muscle cells). Hemostatic factors released during endothelial dysfunction can lead to restenosis. 1. Determination of selected endothelial hemostatic factors in PAD patients and a reference group. 2. Prospective observation of new restenosis appearance in PAD patients after endovascular revascularization. 3. Comparison of selected endothelial hemostatic factors between non-restenotic and restenotic PAD patients. 150 PAD patients after endovascular revascularization - 90 men and 60 women, aged 44-88 (mean 65.5) years - were examined. During one-year observation after the revascularization procedures in 38 PAD patients restenosis occurred, when blood samples were also collected. The reference group consisted of 53 healthy persons - 44 men and 9 women, aged 20-56 years. Blood was drawn in the morning into 3.2% sodium citrate at a ratio of 9:1. Tissue factor (TF), tissue factor pathway inhibitor (TFPI), thrombomodulin (TM), von Willebrand factor (vWF) and tissue plasminogen activator (t-PA) were measured in plasma with commercial tests using the enzyme immunoassay. In the plasma of PAD patients after revascularization, the concentrations of TF and vWF were significantly higher, TM lower, TFPI and t-PA similar compared to the reference group. Six months after revascularization the level of TF had increased and vWF had significantly decreased. The endothelial hemostatic factors before and after restenosis did not significantly differ except TF, which after restenosis was higher. Increased TF and vWF levels in PAD patients indicate arterial endothelial cell damage, by atherosclerotic and revascularization processes. In PAD patients with restenosis compared to these patients before restenosis the determined endothelial hemostatic factors, except TF level, did not significantly differ. Perhaps TF participates in

  4. 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

  5. Effects of Olive Oil on Markers of Inflammation and Endothelial Function—A Systematic Review and Meta-Analysis

    PubMed Central

    Schwingshackl, Lukas; Christoph, Marina; Hoffmann, Georg

    2015-01-01

    The aim of the present systematic review was to synthesize data from randomized controlled trials investigating the effects of olive oil on markers of inflammation or endothelial function. Literature search in electronic databases Cochrane Trial Register, EMBASE, and MEDLINE was performed. Thirty studies enrolling 3106 participants fulfilled the selection criteria. Pooled effects of different interventions were assessed as mean difference using a random effects model. Olive oil interventions (with daily consumption ranging approximately between 1 mg and 50 mg) resulted in a significantly more pronounced decrease in C-reactive protein (mean difference: −0.64 mg/L, (95% confidence interval (CI) −0.96 to −0.31), p < 0.0001, n = 15 trials) and interleukin-6 (mean difference: −0.29 (95% CI −0.7 to −0.02), p < 0.04, n = 7 trials) as compared to controls, respectively. Values of flow-mediated dilatation (given as absolute percentage) were significantly more increased in individuals subjected to olive oil interventions (mean difference: 0.76% (95% CI 0.27 to 1.24), p < 0.002, n = 8 trials). These results provide evidence that olive oil might exert beneficial effects on endothelial function as well as markers of inflammation and endothelial function, thus representing a key ingredient contributing to the cardiovascular-protective effects of a Mediterranean diet. However, due to the heterogeneous study designs (e.g., olive oil given as a supplement or as part of dietary pattern, variations in control diets), a conservative interpretation of the results is necessary. PMID:26378571

  6. Effects of Olive Oil on Markers of Inflammation and Endothelial Function-A Systematic Review and Meta-Analysis.

    PubMed

    Schwingshackl, Lukas; Christoph, Marina; Hoffmann, Georg

    2015-09-01

    The aim of the present systematic review was to synthesize data from randomized controlled trials investigating the effects of olive oil on markers of inflammation or endothelial function. Literature search in electronic databases Cochrane Trial Register, EMBASE, and MEDLINE was performed. Thirty studies enrolling 3106 participants fulfilled the selection criteria. Pooled effects of different interventions were assessed as mean difference using a random effects model. Olive oil interventions (with daily consumption ranging approximately between 1 mg and 50 mg) resulted in a significantly more pronounced decrease in C-reactive protein (mean difference: -0.64 mg/L, (95% confidence interval (CI) -0.96 to -0.31), p < 0.0001, n = 15 trials) and interleukin-6 (mean difference: -0.29 (95% CI -0.7 to -0.02), p < 0.04, n = 7 trials) as compared to controls, respectively. Values of flow-mediated dilatation (given as absolute percentage) were significantly more increased in individuals subjected to olive oil interventions (mean difference: 0.76% (95% CI 0.27 to 1.24), p < 0.002, n = 8 trials). These results provide evidence that olive oil might exert beneficial effects on endothelial function as well as markers of inflammation and endothelial function, thus representing a key ingredient contributing to the cardiovascular-protective effects of a Mediterranean diet. However, due to the heterogeneous study designs (e.g., olive oil given as a supplement or as part of dietary pattern, variations in control diets), a conservative interpretation of the results is necessary. PMID:26378571

  7. Markers of Thrombogenesis and Fibrinolysis and Their Relation to Inflammation and Endothelial Activation in Patients with Idiopathic Pulmonary Arterial Hypertension

    PubMed Central

    Kopeć, Grzegorz; Moertl, Deddo; Steiner, Sabine; Stępień, Ewa; Mikołajczyk, Tomasz; Podolec, Jakub; Waligóra, Marcin; Stępniewski, Jakub; Tomkiewicz-Pająk, Lidia; Guzik, Tomasz; Podolec, Piotr

    2013-01-01

    Background Chronic anticoagulation is a standard of care in idiopathic pulmonary arterial hypertension (IPAH). However, hemostatic abnormalities in this disease remain poorly understood. Therefore, we aimed to study markers of thrombogenesis and fibrinolysis in patients with IPAH. Methods We studied 27 consecutive patients (67% female) with IPAH aged 50.0 years (IQR: 41.0 - 65.0) and 16 controls without pulmonary hypertension. Prothrombin fragment 1+2 (F1+2) and thrombin-antithrombin (TAT) complexes were measured to assess thrombogenesis; tissue-type plasminogen activator (tPA) antigen and plasmin-anti-plasmin complex to characterize activation of fibrinolysis; plasminogen activator inhibitor 1 (PAI-1) to measure inhibition of fibrinolysis; and endothelin-1 (ET-1) and interleukin-6 (IL-6) to assess endothelial activation and systemic inflammation, respectively. In addition, in treatment-naive IPAH patients these markers were assessed after 3 months of PAH-specific therapies. Results TPA (10.1[6.8-15.8] vs 5.2[3.3-7.3] ng/ml, p<0.001), plasmin-anti-plasmin (91.5[60.3-94.2] vs 55.8[51.1-64.9] ng/ml, p<0.001), IL-6 (4.9[2.5-7.9] vs 2.1[1.3-3.8] pg/ml, p=0.001) and ET-1 (3.7 [3.3-4.5] vs 3.4[3.1-3.5], p= 0.03) were higher in patients with IPAH than in controls. In IPAH patients plasmin-anti-plasmin and tPA correlated positively with IL-6 (r=0.39, p=0.04 and r=0.63, p<0.001, respectively) and ET-1 (r=0.55, p=0.003 and r=0.59, p=0.001, respectively). No correlation was found between tPA or plasmin-anti-plasmin and markers of thrombogenesis. Plasmin-anti-plasmin decreased after 3 months of PAH specific therapy while the other markers remained unchanged. Conclusions In the present study we showed that markers of fibrynolysis were elevated in patients with IPAH however we did not find a clear evidence for increased thrombogenesis in this group of patients. Fibrinolysis, inflammation, and endothelial activation were closely interrelated in IPAH. PMID:24312667

  8. Cocaine Abstinence and Reduced Use Associated With Lowered Marker of Endothelial Dysfunction in African Americans: A Preliminary Study

    PubMed Central

    Lai, Hong; Stitzer, Maxine; Treisman, Glenn; Moore, Richard; Brinker, Jeffrey; Gerstenblith, Gary; Kickler, Thomas S.; Li, Ji; Chen, Shaoguang; Fishman, Elliot; Lai, Shenghan

    2016-01-01

    Objectives Clinical and epidemiological evidence suggests that cocaine use is associated with an increased risk of premature atherosclerosis. The objectives of this study were to explore (1) whether cocaine abstinence is associated with a reduced marker of endothelial dysfunction, (2) whether cocaine abstinence is associated with a slower coronary plaque progression, and (3) whether reduction in cocaine use is associated with a reduced marker of endothelial dysfunction in African American chronic cocaine users with contrast-enhanced coronary CT angiography-confirmed less than 50% coronary stenosis. Methods Between March and June 2014, a total of 57 African American cocaine users with contrast-enhanced CT angiography-confirmed less than 50% coronary stenosis in Baltimore, Maryland, were enrolled in a 6-month follow-up study to investigate whether cocaine abstinence or reduction in cocaine use is associated with decreased endothelin-1 (ET-1) levels and coronary plaque progression at the 6-month follow-up. A voucher-based incentive approach was used to systematically reinforce cocaine abstinence, and urine benzoylecgonine test was implemented to confirm cocaine use. Results Among the 57 participants, 44 were HIV-infected. The median of duration of cocaine use was 18 (interquartile range, 7–30) years. According to generalized estimating equation analyses, both cocaine abstinence and reduction in cocaine use in the 6 months were independently associated with decreased ET-1. The incidence of coronary plaque progression was 7.4/100 person-years and 23.1/100 person-years in those who were totally abstinent from cocaine and those who continued to use cocaine, respectively. However, the difference in the incidence between these 2 groups was not significant (exact P = 0.30). Conclusions The findings of this study revealed a possible association of cocaine abstinence/reduction with lowered ET levels, which suggests that such changes in cocaine use might be beneficial for

  9. Immuno-PET imaging of tumor endothelial marker 8 (TEM8).

    PubMed

    Kuo, Frank; Histed, Stephanie; Xu, Biying; Bhadrasetty, Veerendra; Szajek, Lawrence P; Williams, Mark R; Wong, Karen; Wu, Haitao; Lane, Kelly; Coble, Vincent; Vasalatiy, Olga; Griffiths, Gary L; Paik, Chang H; Elbuluk, Osama; Szot, Christopher; Chaudhary, Amit; St Croix, Brad; Choyke, Peter; Jagoda, Elaine M

    2014-11-01

    Tumor endothelial marker 8 (TEM8) is a cell surface receptor that is highly expressed in a variety of human tumors and promotes tumor angiogenesis and cell growth. Antibodies targeting TEM8 block tumor angiogenesis in a manner distinct from the VEGF receptor pathway. Development of a TEM8 imaging agent could aid in patient selection for specific antiangiogenic therapies and for response monitoring. In these studies, L2, a therapeutic anti-TEM8 monoclonal IgG antibody (L2mAb), was labeled with (89)Zr and evaluated in vitro and in vivo in TEM8 expressing cells and mouse xenografts (NCI-H460, DLD-1) as a potential TEM8 immuno-PET imaging agent. (89)Zr-df-L2mAb was synthesized using a desferioxamine-L2mAb conjugate (df-L2mAb); (125)I-L2mAb was labeled directly. In vitro binding studies were performed using human derived cell lines with high, moderate, and low/undetectable TEM8 expression. (89)Zr-df-L2mAb in vitro autoradiography studies and CD31 IHC staining were performed with cryosections from human tumor xenografts (NCI-H460, DLD-1, MKN-45, U87-MG, T-47D, and A-431). Confirmatory TEM8 Western blots were performed with the same tumor types and cells. (89)Zr-df-L2mAb biodistribution and PET imaging studies were performed in NCI-H460 and DLD-1 xenografts in nude mice. (125)I-L2mAb and (89)Zr-df-L2mAb exhibited specific and high affinity binding to TEM8 that was consistent with TEM8 expression levels. In NCI-H460 and DLD-1 mouse xenografts nontarget tissue uptake of (89)Zr-df-L2mAb was similar; the liver and spleen exhibited the highest uptake at all time points. (89)Zr-L2mAb was highly retained in NCI-H460 tumors with <10% losses from day 1 to day 3 with the highest tumor to muscle ratios (T:M) occurring at day 3. DLD-1 tumors exhibited similar pharmacokinetics, but tumor uptake and T:M ratios were reduced ∼2-fold in comparison to NCI-H460 at all time points. NCI-H460 and DLD-1 tumors were easily visualized in PET imaging studies despite low in vitro TEM8 expression

  10. Immuno-PET Imaging of Tumor Endothelial Marker 8 (TEM8)

    PubMed Central

    2015-01-01

    Tumor endothelial marker 8 (TEM8) is a cell surface receptor that is highly expressed in a variety of human tumors and promotes tumor angiogenesis and cell growth. Antibodies targeting TEM8 block tumor angiogenesis in a manner distinct from the VEGF receptor pathway. Development of a TEM8 imaging agent could aid in patient selection for specific antiangiogenic therapies and for response monitoring. In these studies, L2, a therapeutic anti-TEM8 monoclonal IgG antibody (L2mAb), was labeled with 89Zr and evaluated in vitro and in vivo in TEM8 expressing cells and mouse xenografts (NCI-H460, DLD-1) as a potential TEM8 immuno-PET imaging agent. 89Zr-df–L2mAb was synthesized using a desferioxamine–L2mAb conjugate (df–L2mAb); 125I-L2mAb was labeled directly. In vitro binding studies were performed using human derived cell lines with high, moderate, and low/undetectable TEM8 expression. 89Zr-df–L2mAb in vitro autoradiography studies and CD31 IHC staining were performed with cryosections from human tumor xenografts (NCI-H460, DLD-1, MKN-45, U87-MG, T-47D, and A-431). Confirmatory TEM8 Western blots were performed with the same tumor types and cells. 89Zr-df–L2mAb biodistribution and PET imaging studies were performed in NCI-H460 and DLD-1 xenografts in nude mice. 125I-L2mAb and 89Zr-df–L2mAb exhibited specific and high affinity binding to TEM8 that was consistent with TEM8 expression levels. In NCI-H460 and DLD-1 mouse xenografts nontarget tissue uptake of 89Zr-df–L2mAb was similar; the liver and spleen exhibited the highest uptake at all time points. 89Zr-L2mAb was highly retained in NCI-H460 tumors with <10% losses from day 1 to day 3 with the highest tumor to muscle ratios (T:M) occurring at day 3. DLD-1 tumors exhibited similar pharmacokinetics, but tumor uptake and T:M ratios were reduced ∼2-fold in comparison to NCI-H460 at all time points. NCI-H460 and DLD-1 tumors were easily visualized in PET imaging studies despite low in vitro TEM8 expression in

  11. Fine particles, genetic pathways, and markers of inflammation and endothelial dysfunction: Analysis on particulate species and sources.

    PubMed

    Dai, Lingzhen; Bind, Marie-Abele; Koutrakis, Petros; Coull, Brent A; Sparrow, David; Vokonas, Pantel S; Schwartz, Joel D

    2016-06-01

    Studies have found associations between PM2.5 and cardiovascular events. The role of different components of PM2.5 is not well understood. We used linear mixed-effects models with the adaptive LASSO penalty to select PM2.5 species and source(s), separately, that may be associated with markers of inflammation and endothelial dysfunction, with adjustment for age, obesity, smoking, statin use, diabetes mellitus, temperature, and season as fixed effects in a large longitudinal cohort of elderly men. We also analyzed these associations with source apportionment models and examined genetic pathway-air pollution interactions within three relevant pathways (oxidative stress, metal processing, and endothelial function). We found that independent of PM2.5 mass vanadium (V) was associated with intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). An IQR increase (3.2 ng/m(3)) in 2-day moving average V was associated with a 2.5% (95% CI: 1.2-3.8%) change in ICAM-1 and a 3.9% (95% CI: 2.2-5.7%) change in VCAM-1, respectively. In addition, an oil combustion source rich in V was linked to these adhesion molecules. People with higher allelic risk profiles related to oxidative stress may have greater associations (P-value of interaction=0.11). Our findings suggest that particles derived from oil combustion may be associated with inflammation and endothelial dysfunction, and it is likely that oxidative stress plays a role in the associations. PMID:26732377

  12. Label-Free and Sensitive Detection of Thrombomodulin, a Marker of Endothelial Cell Injury, Using Quartz Crystal Microbalance.

    PubMed

    Luo, Yiqun; Liu, Tong; Zhu, Jiaming; Kong, Liyan; Wang, Wen; Tan, Liang

    2015-11-17

    Thrombomodulin (TM), an integral glycoprotein on the surface of endothelial cells, can be released during endothelial cell injury and the levels of serum TM are regarded as an important parameter of activity in vasculitides in vivo. Quantitative detection of TM and investigation on the release of soluble thrombomodulin (sTM) by the injured HUVEC-C cells using quartz crystal microbalance (QCM) were achieved in this work. Anti-antibody (AAb) and bovine serum albumin (BSA) were bound on gold nanoparticles (GNPs) to construct BSA-GNPs-AAb nanocomposites and they were characterized by transmission electron microscope, UV-vis, and infrared spectrophotometry, respectively. The capture of the nanocomposites on the TM antibody modified electrode, which was tested by scanning electron microscope, could result in a great decrease of the resonant frequency (f0). This binding was effectively inhibited by the beforehand immobilized TM proteins on the electrode surface due to the strong steric hindrance effect. It led to the decrease of the frequency changing extent. The relative frequency-shift was found to be proportional to the logarithm of the TM concentration from 10 to 5000 ng mL(-1) with a detection limit of 2 ng mL(-1). By analyzing the growth medium used for cell incubation, the release of sTM by the injured HUVEC-C cells in the presence of H2O2 was confirmed. The sTM amount in the growth medium was increased with the enhancement of contact time of the cells with H2O2, proving that sTM may serve as a specific marker of endothelial cell injury. PMID:26507327

  13. Podoplanin and CLEC-2 drive cerebrovascular patterning and integrity during development.

    PubMed

    Lowe, Kate L; Finney, Brenda A; Deppermann, Carsten; Hägerling, René; Gazit, Salomé L; Frampton, Jon; Buckley, Christopher; Camerer, Eric; Nieswandt, Bernhard; Kiefer, Friedemann; Watson, Steve P

    2015-06-11

    Mice with a constitutive or platelet-specific deletion of the C-type-lectin-like receptor (CLEC-2) exhibit hemorrhaging in the brain at mid-gestation. We sought to investigate the basis of this defect, hypothesizing that it is mediated by the loss of CLEC-2 activation by its endogenous ligand, podoplanin, which is expressed on the developing neural tube. To induce deletion of podoplanin at the 2-cell stage, we generated a podoplanin(fl/fl) mouse crossed to a PGK-Cre mouse. Using 3-dimensional light-sheet microscopy, we observed cerebral vessels were tortuous and aberrantly patterned at embryonic (E) day 10.5 in podoplanin- and CLEC-2-deficient mice, preceding the formation of large hemorrhages throughout the fore-, mid-, and hindbrain by E11.5. Immunofluorescence and electron microscopy revealed defective pericyte recruitment and misconnections between the endothelium of developing blood vessels and surrounding pericytes and neuro-epithelial cells. Nestin-Cre-driven deletion of podoplanin on neural progenitors also caused widespread cerebral hemorrhaging. Hemorrhaging was also seen in the ventricles of embryos deficient in the platelet integrin subunit glycoprotein IIb or in embryos in which platelet α-granule and dense granule secretion is abolished. We propose a novel role for podoplanin on the neuro-epithelium, which interacts with CLEC-2 on platelets, mediating platelet adhesion, aggregation, and secretion to guide the maturation and integrity of the developing vasculature and prevent hemorrhage. PMID:25908104

  14. Down-regulation of tumor endothelial marker 8 suppresses cell proliferation mediated by ERK1/2 activity

    PubMed Central

    Cao, Chuangjie; Wang, Zhuo; Huang, Leilei; Bai, Lihong; Wang, Yuefeng; Liang, Yingjie; Dou, Chengyun; Wang, Liantang

    2016-01-01

    Tumor endothelial marker 8 (TEM8) was recently suggested as a putative anti-tumor target in several types of human cancer based on its selective overexpression in tumor versus normal endothelial cells. The objective of this study was to detect the potential functions of TEM8 in osteosarcoma. Overall, TEM8 was mainly located in cytoplasm and was up-regulated in osteosarcoma compared to benign bone lesions and adjacent non tumor tissue (ANT). High TEM8 expression group had a significant lower overall survival rate than that in the low TEM8 expression group. TEM8 knock-down by siRNA or shRNA results in significant reduction of osteosarcoma cell growth and proliferation both in vitro and in vivo. Ablation of TEM8 led to increasing of p21 and p27 and suppression of cyclin D1 mediated by Erk1/2 activity. These findings suggest that down-regulation of TEM8 play an important role in the inhibition of tumorigenesis and development of osteosarcoma. PMID:26996335

  15. Effects of Complementary Creatine Monohydrate and Physical Training on Inflammatory and Endothelial Dysfunction Markers Among Heart Failure Patients

    PubMed Central

    Hemati, Farajollah; Rahmani, Asghar; Asadollahi, Khairollah; Soleimannejad, Koroush; Khalighi, Zahra

    2016-01-01

    Background: Previous studies have reported endothelial dysfunction and inflammatory cytokine in heart failure patients (HF). Objectives: The purpose of this study was to determine the effects of creatine monohydrate and exercise on inflammatory and endothelial dysfunction markers among HF patients. Patients and Methods: One hundred patients were prospectively randomized into two groups: Intervention group which received 5 grams/day creatine monohydrate and exercised for 8 weeks; and control group which did not receive any interventions. Interleukine-6 (IL-6), high sensitivity C reactive protein (hs-CRP), P-selectin, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule 1 (VCAM-1) were measured at the start and end of the study for both groups. Results: In total, 100 patients including 50 controls and 50 intervention group (54% male, mean EF of 34.2 ± 10.5% and 52% male, mean EF of 35.6 ± 12.7%, respectively) were analyzed. The serum levels of hs-CRP and IL-6 increased at the end of the study in the control group compared to the baseline, (7.5 ± 1.5 mg/L vs. 6.9 ± 1.3 mg/L, P < 0.05 and 3.0 ± 0.75 ng/L vs. 2.55 ± 0.9 ng/L, P < 0.05, respectively). However, compared to the baseline, the level of both markers decreased at the end of the study in the intervention group (6.3 ± 1.6 mg/L vs.7.5 ± 1.5 mg/L, P < 0.05 and 2.1 ± 0.8 ng/L vs.2.5 ± 0.5 ng/L, P < 0.05). Also, P-selectin and ICAM-1 levels increased at the end of study (56.9 ± 1.8 ng/L vs. 51.9 ± 1.5 ng/L, P < 0.05 and 368.1 ± 25.4 µg/L vs. 353.1 ± 10.4 µg/L, P < 0.05 respectively). Inversely, the levels of these markers decreased in the intervention group, at the end of study (49.7 ± 1.9 ng/l vs. 51.4 ± 2.1 ng/l, P < 0.05 and 342.7 ± 16.5 µg/l vs. 350.4 ± 14.7 µg/l, P < 0.05, respectively). VCAM-1 level was not decreased significantly at the end of the study in the intervention group (570.5 ± 78.4 µg/L vs. 575.3 ± 86.5 µg/L, P > 0.05). Conclusions: Combination

  16. Effects of adenoidectomy on markers of endothelial function and inflammation in normal-weight and overweight prepubescent children with sleep apnea

    PubMed Central

    Kelishadi, Roya; Nilforoushan, Neshat; Okhovat, Ahmadreza; Amra, Babak; Poursafa, Parinaz; Rogha, Mehrdad

    2011-01-01

    BACKGROUND: This trial study aimed to assess the effects of adenoidectomy on the markers of endothelial function and inflammation in normal-weight and overweight prepubescent children with obstructive sleep apnea (OSA). METHODS: This trial study was conducted in Isfahan, Iran in 2009. The study population was comprised of 90 prepubescent children (45 normal-weight and 45 overweight children), aged between 4-10 years old, who volunteered for adenoidectomy and had OSA documented by validated questionnaire. The assessment included filling questionnaire, physical examination, and laboratory tests; it was conducted before the surgery and was repeated two weeks and six months after the surgery. RESULTS: Out of the 90 children evaluated, 83 completed the 2-week evaluation and 72 patients continued with the study for the 6-month follow up. Markers of endothelial function, i.e., serum adhesion molecules including endothelial leukocyte adhesion molecule (E-selectin), intercellular cell adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (sVCAM-1), and the markers of inflammation, i.e., interleukin-6, and high-sensitive C-reactive protein (hsCRP) decreased significantly in both normal-weight and overweight children after both two weeks and six months. After six months, the total and LDL-cholesterol showed a significant decrease in the overweight children. CONCLUSIONS: The findings of the study demonstrated that irrespective of the weight status, children with OSA had increased levels of the endothelial function and inflammation markers, which improved after OSA treatment by adenoidectomy. This might be a form of confirmatory evidence on the onset of atherogenesis from the early stages of the life, and the role of inflammation in the process. The reversibility of endothelial dysfunction after improvement of OSA underscores the importance of primordial and primary prevention of chronic diseases from the early stages of the life. PMID:22247723

  17. Venous Endothelial Marker COUP-TFII Regulates the Distinct Pathologic Potentials of Adult Arteries and Veins

    PubMed Central

    Cui, Xiaofeng; Lu, Yao Wei; Lee, Vivian; Kim, Diana; Dorsey, Taylor; Wang, Qingjie; Lee, Young; Vincent, Peter; Schwarz, John; Dai, Guohao

    2015-01-01

    Arteries and veins have very different susceptibility to certain vascular diseases such as atherosclerosis and vascular calcification. The molecular mechanisms of these differences are not fully understood. In this study, we discovered that COUP-TFII, a transcription factor critical for establishing the venous identity during embryonic vascular development, also regulates the pathophysiological functions of adult blood vessels, especially those directly related to vascular diseases. Specifically, we found that suppression of COUP-TFII in venous ECs switched its phenotype toward pro-atherogenic by up-regulating the expression of inflammatory genes and down-regulating anti-thrombotic genes. ECs with COUP-TFII knockdown also readily undergo endothelial-to-mesenchymal transition (EndoMT) and subsequent osteogenic differentiation with dramatically increased osteogenic transcriptional program and calcium deposition. Consistently, over-expression of COUP-TFII led to the completely opposite effects. In vivo validation of these pro-atherogenic and osteogenic genes also demonstrates a broad consistent differential expression pattern in mouse aorta vs. vena cava ECs, which cannot be explained by the difference in hemodynamic flow. These data reveal phenotypic modulation by different levels of COUP-TFII in arterial and venous ECs, and suggest COUP-TFII may play an important role in the different susceptibilities of arteries and veins to vascular diseases such as atherosclerosis and vascular calcification. PMID:26537113

  18. Co-culture with podoplanin+ cells protects leukemic blast cells with leukemia-associated antigens in the tumor microenvironment

    PubMed Central

    LEE, JI YOON; HAN, A-REUM; LEE, SUNG-EUN; MIN, WOO-SUNG; KIM, HEE-JE

    2016-01-01

    Podoplanin+ cells are indispensable in the tumor microenvironment. Increasing evidence suggests that podoplanin may support the growth and metastasis of solid tumors; however, to the best of our knowledge no studies have determined whether or not podoplanin serves a supportive role in acute myeloid leukemia (AML). The effects of co-culture with podoplanin+ cells on the cellular activities of the leukemic cells, such as apoptosis and cell proliferation, in addition to the expression of podoplanin in leukemic cells, were investigated. Due to the fact that genetic abnormalities are the primary cause of leukemogenesis, the overexpression of the fibromyalgia-like tyrosine kinase-3 gene in colony forming units was also examined following cell sorting. Podoplanin+ cells were found to play a protective role against apoptosis in leukemic cells and to promote cell proliferation. Tumor-associated antigens, including Wilms' tumor gene 1 and survivin, were increased when leukemic cells were co-cultured with podoplanin+ cells. In combination, the present results also suggest that podoplanin+ cells can function as stromal cells for blast cell retention in the AML tumor microenvironment. PMID:27035421

  19. Markers of Endothelial-to-Mesenchymal Transition: Evidence for Antibody-Endothelium Interaction during Antibody-Mediated Rejection in Kidney Recipients.

    PubMed

    Xu-Dubois, Yi-Chun; Peltier, Julie; Brocheriou, Isabelle; Suberbielle-Boissel, Caroline; Djamali, Arjang; Reese, Shannon; Mooney, Nuala; Keuylian, Zela; Lion, Julien; Ouali, Nacéra; Levy, Pierre P; Jouanneau, Chantal; Rondeau, Eric; Hertig, Alexandre

    2016-01-01

    Antibody-mediated rejection (ABMR) is a leading cause of allograft loss. Treatment efficacy depends on accurate diagnosis at an early stage. However, sensitive and reliable markers of antibody-endothelium interaction during ABMR are not available for routine use. Using immunohistochemistry, we retrospectively studied the diagnostic value of three markers of endothelial-to-mesenchymal transition (EndMT), fascin1, vimentin, and heat shock protein 47, for ABMR in 53 renal transplant biopsy specimens, including 20 ABMR specimens, 24 cell-mediated rejection specimens, and nine normal grafts. We validated our results in an independent set of 74 unselected biopsy specimens. Endothelial cells of the peritubular capillaries in grafts with ABMR expressed fascin1, vimentin, and heat shock protein 47 strongly, whereas those from normal renal grafts did not. The level of EndMT marker expression was significantly associated with current ABMR criteria, including capillaritis, glomerulitis, peritubular capillary C4d deposition, and donor-specific antibodies. These markers allowed us to identify C4d-negative ABMR and to predict late occurrence of disease. EndMT markers were more specific than capillaritis for the diagnosis and prognosis of ABMR and predicted late (up to 4 years after biopsy) renal graft dysfunction and proteinuria. In the independent set of 74 renal graft biopsy specimens, the EndMT markers for the diagnosis of ABMR had a sensitivity of 100% and a specificity of 85%. Fascin1 expression in peritubular capillaries was also induced in a rat model of ABMR. In conclusion, EndMT markers are a sensitive and reliable diagnostic tool for detecting endothelial activation during ABMR and predicting late loss of allograft function. PMID:25995444

  20. Podoplanin and CLEC-2 drive cerebrovascular patterning and integrity during development

    PubMed Central

    Lowe, Kate L.; Finney, Brenda A.; Deppermann, Carsten; Hägerling, René; Gazit, Salomé L.; Frampton, Jon; Buckley, Christopher; Camerer, Eric; Nieswandt, Bernhard; Kiefer, Friedemann

    2015-01-01

    Mice with a constitutive or platelet-specific deletion of the C-type-lectin-like receptor (CLEC-2) exhibit hemorrhaging in the brain at mid-gestation. We sought to investigate the basis of this defect, hypothesizing that it is mediated by the loss of CLEC-2 activation by its endogenous ligand, podoplanin, which is expressed on the developing neural tube. To induce deletion of podoplanin at the 2-cell stage, we generated a podoplaninfl/fl mouse crossed to a PGK-Cre mouse. Using 3-dimensional light-sheet microscopy, we observed cerebral vessels were tortuous and aberrantly patterned at embryonic (E) day 10.5 in podoplanin- and CLEC-2-deficient mice, preceding the formation of large hemorrhages throughout the fore-, mid-, and hindbrain by E11.5. Immunofluorescence and electron microscopy revealed defective pericyte recruitment and misconnections between the endothelium of developing blood vessels and surrounding pericytes and neuro-epithelial cells. Nestin-Cre-driven deletion of podoplanin on neural progenitors also caused widespread cerebral hemorrhaging. Hemorrhaging was also seen in the ventricles of embryos deficient in the platelet integrin subunit glycoprotein IIb or in embryos in which platelet α-granule and dense granule secretion is abolished. We propose a novel role for podoplanin on the neuro-epithelium, which interacts with CLEC-2 on platelets, mediating platelet adhesion, aggregation, and secretion to guide the maturation and integrity of the developing vasculature and prevent hemorrhage. PMID:25908104

  1. Targeting a novel domain in podoplanin for inhibiting platelet-mediated tumor metastasis

    PubMed Central

    Sekiguchi, Takaya; Takemoto, Ai; Takagi, Satoshi; Takatori, Kazuki; Sato, Shigeo; Takami, Miho; Fujita, Naoya

    2016-01-01

    Podoplanin/Aggrus is a sialoglycoprotein expressed in various cancers. We previously identified podoplanin as a key factor in tumor-induced platelet aggregation. Podoplanin-mediated platelet aggregation enhances tumor growth and metastasis by secreting growth factors and by forming tumor emboli in the microvasculature. Thus, precise analysis of the mechanisms of podoplanin-mediated platelet aggregation is critical for developing anti-tumor therapies. Here we report the discovery of a novel platelet aggregation-inducing domain, PLAG4 (81-EDLPT-85). PLAG4 has high homology to the previously reported PLAG3 and contributes to the binding of its platelet receptor CLEC-2. Mutant analyses indicated that PLAG4 exhibits a predominant platelet-aggregating function relative to PLAG3 and that conserved Glu81/Asp82/Thr85 residues in PLAG4 are indispensable for CLEC-2 binding. By establishing anti-PLAG4-neutralizing monoclonal antibodies, we confirmed its role in CLEC-2 binding, platelet aggregation, and tumor emboli formation. Our results suggest the requirement of simultaneous inhibition of PLAG3/4 for complete suppression of podoplanin-mediated tumor growth and metastasis. PMID:26684030

  2. Lymphatic endothelial regulation, lymphoedema, and lymph node metastasis.

    PubMed

    Karkkainen, Marika J; Alitalo, Kari

    2002-02-01

    Vascular endothelial growth factor receptor-3 (VEGFR-3) mediates lymphatic endothelial cell (LEC) growth, migration, and survival by binding VEGF-C and VEGF-D. Recent studies have revealed new regulators of the lymphatic endothelium, such as the transcription factor Prox1, and the cell surface proteins podoplanin and lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1). Furthermore, the isolation of LECs now allows detailed molecular studies of the factors regulating the lymphatic vasculature. These studies are aimed at targeting the lymphatic vasculature in the treatment of various diseases, such as tumour metastasis and lymphoedema. PMID:11969367

  3. Vascular endothelial growth factor as a predictive marker for POEMS syndrome treatment response: retrospective cohort study

    PubMed Central

    Misawa, S; Sato, Y; Katayama, K; Hanaoka, H; Sawai, S; Beppu, M; Nomura, F; Shibuya, K; Sekiguchi, Y; Iwai, Y; Watanabe, K; Amino, H; Ohwada, C; Takeuchi, M; Sakaida, E; Nakaseko, C; Kuwabara, S

    2015-01-01

    Objective POEMS (polyneuropathy, organomegaly, endocrinopathy, M-protein and skin changes) syndrome is a rare multisystem disease characterised by plasma cell dyscrasia and overproduction of vascular endothelial growth factor (VEGF). VEGF is assumed to be useful in monitoring disease activity, because VEGF levels usually decrease after treatment. However, there is no study to investigate whether the extent of decrease in VEGF correlates with clinical outcome. We tested the predictive efficacy of serum VEGF levels in POEMS syndrome. Method This was an institutional review board approved retrospective observational cohort study of 20 patients with POEMS monitored regularly for more than 12 months (median follow-up, 87 months) after treatment onset using our prospectively accumulated database of POEMS from 1999 to 2015. Patients were treated by autologous peripheral blood stem cell transplantation or thalidomide administration. Serum VEGF was measured by ELISA. Outcome measures included clinical and laboratory findings and relapse-free survival. Results Serum VEGF levels decreased rapidly after treatment, and stabilised by 6 months post treatment. Patients with normalised serum VEGF levels (<1040 pg/mL) at 6 months showed prolonged relapse-free survival (HR=12.81, 95% CI 2.691 to 90.96; p=0.0001) and greater later clinical improvement. The rate of serum VEGF reduction over the first 6 months post treatment correlated with increased grip strength, serum albumin levels, and compound muscle action potential amplitudes at 12 months. Conclusions Serum VEGF level at 6 months post treatment is a predicative biomarker for disease activity and prognosis in POEMS syndrome. Serum VEGF could be used as a surrogate endpoint for relapse-free survival or clinical or laboratory improvement of POEMS syndrome for clinical trials. PMID:26560063

  4. Diagnostic value of endothelial markers and HHV-8 staining in gastrointestinal Kaposi sarcoma and its difference in endoscopic tumor staging

    PubMed Central

    Nagata, Naoyoshi; Igari, Toru; Shimbo, Takuro; Sekine, Katsunori; Akiyama, Junichi; Hamada, Yohei; Yazaki, Hirohisa; Ohmagari, Norio; Teruya, Katsuji; Oka, Shinichi; Uemura, Naomi

    2013-01-01

    AIM: To clarify the diagnostic values of hematoxylin and eosin (HE), D2-40, CD31, CD34, and HHV-8 immunohistochemical (IHC) staining in gastrointestinal Kaposi’s sarcoma (GI-KS) in relation to endoscopic tumor staging. METHODS: Biopsy samples (n = 133) from 41 human immunodeficiency virus-infected patients were reviewed. GI-KS was defined as histologically negative for other GI diseases and as a positive clinical response to KS therapy. The receiver operating characteristic area under the curve (ROC-AUC) was compared in relation to lesion size, GI location, and macroscopic appearances on endoscopy. RESULTS: GI-KS was confirmed in 84 lesions (81.6%). Other endoscopic findings were polyps (n = 9), inflammation (n = 4), malignant lymphoma (n = 4), and condyloma (n = 2), which mimicked GI-KS on endoscopy. ROC-AUC of HE, D2-40, blood vessel markers, and HHV-8 showed results of 0.83, 0.89, 0.80, and 0.82, respectively. For IHC staining, the ROC-AUC of D2-40 was significantly higher (P < 0.05) than that of HE staining only. In the analysis of endoscopic appearance, the ROC-AUC of HE and IHC showed a tendency toward an increase in tumor staging (e.g., small to large, patches, and polypoid to SMT appearance). D2-40 was significantly (P < 0.05) advantageous in the upper GI tract and for polypoid appearance compared with HE staining. CONCLUSION: The diagnostic value of endothelial markers and HHV-8 staining was found to be high, and its accuracy tended to increase with endoscopic tumor staging. D2-40 will be useful for complementing HE staining in the diagnosis of GI-KS, especially in the upper GI tract and for polypoid appearance. PMID:23801862

  5. Tumour endothelial marker-8 in wound healing and its impact on the proliferation and migration of keratinocytes

    PubMed Central

    WANG, SHEILA C.; YE, LIN; SANDERS, ANDREW J.; RUGE, FIONA; HARDING, KEITH G.; JIANG, WEN G.

    2016-01-01

    Chronic wound management represents a significant burden on healthcare systems and negatively impacts on the quality of patient life. New strategies to understand and identify wounds that will not heal in a normal manner are required. Tumour endothelial marker-8 (TEM-8) has been implicated in the wound healing and angiogenesis processes. TEM-8 expression was examined at the transcript level in a cohort of acute (n=10) and chronic (n=14) wounds and in normal skin (n=10). Protein analysis of TEM-8 was also undertaken for this cohort using immunohistochemistry (IHC). TEM-8 impact on keratinocyte cell growth and migration was assessed following TEM-8 ribozyme transgene transfection of human HaCaT keratinocytes using cell growth and electric cell-substrate impedance sensing (ECIS)-based assays. Expression of TEM-8 was observed to be increased in acute wounds compared to chronic wounds and normal skin using quantitative polymerase chain reaction transcript analysis and IHC staining of wound tissues. Knockdown of TEM-8 in HaCaT cells, using two independent ribozyme transgenes, resulted in significant decreases in cell growth as well as reductions in the rate of migration assessed using an ECIS-based system. TEM-8 may be differentially expressed between wound types and loss of this molecule impacts HaCaT growth and migration, potentially implicating this molecule as a factor involved in successful progression of wound healing. PMID:26677171

  6. Effects of aerobic interval training and continuous training on cellular markers of endothelial integrity in coronary artery disease: a SAINTEX-CAD substudy.

    PubMed

    Van Craenenbroeck, Emeline M; Frederix, Geert; Pattyn, Nele; Beckers, Paul; Van Craenenbroeck, Amaryllis H; Gevaert, Andreas; Possemiers, Nadine; Cornelissen, Veronique; Goetschalckx, Kaatje; Vrints, Christiaan J; Vanhees, Luc; Hoymans, Vicky Y

    2015-12-01

    In this large multicenter trial, we aimed to assess the effect of aerobic exercise training in stable coronary artery disease (CAD) patients on cellular markers of endothelial integrity and to examine their relation with improvement of endothelial function. Two-hundred CAD patients (left ventricular ejection fraction > 40%, 90% male, mean age 58.4 ± 9.1 yr) were randomized on a 1:1 base to a supervised 12-wk rehabilitation program of either aerobic interval training or aerobic continuous training on a bicycle. At baseline and after 12 wk, numbers of circulating CD34(+)/KDR(+)/CD45dim endothelial progenitor cells (EPCs), CD31(+)/CD3(+)/CXCR4(+) angiogenic T cells, and CD31(+)/CD42b(-) endothelial microparticles (EMPs) were analyzed by flow cytometry. Endothelial function was assessed by flow-mediated dilation (FMD) of the brachial artery. After 12 wk of aerobic interval training or aerobic continuous training, numbers of circulating EPCs, angiogenic T cells, and EMPs were comparable with baseline levels. Whereas improvement in peak oxygen consumption was correlated to improvement in FMD (Pearson r = 0.17, P = 0.035), a direct correlation of baseline or posttraining EPCs, angiogenic T cells, and EMP levels with FMD was absent. Baseline EMPs related inversely to the magnitude of the increases in peak oxygen consumption (Spearman rho = -0.245, P = 0.027) and FMD (Spearman rho = -0.374, P = 0.001) following exercise training. In conclusion, endothelial function improvement in response to exercise training in patients with CAD did not relate to altered levels of EPCs and angiogenic T cells and/or a diminished shedding of EMPs into the circulation. EMP flow cytometry may be predictive of the increase in aerobic capacity and endothelial function. PMID:26453327

  7. Development of Blood and Lymphatic Endothelial Cells in Embryonic and Fetal Human Skin.

    PubMed

    Schuster, Christopher; Mildner, Michael; Botta, Albert; Nemec, Lucas; Rogojanu, Radu; Beer, Lucian; Fiala, Christian; Eppel, Wolfgang; Bauer, Wolfgang; Petzelbauer, Peter; Elbe-Bürger, Adelheid

    2015-09-01

    Blood and lymphatic vessels provide nutrients for the skin and fulfill important homeostatic functions, such as the regulation of immunologic processes. In this study, we investigated the development of blood and lymphatic endothelial cells in prenatal human skin in situ using multicolor immunofluorescence and analyzed angiogenic molecules by protein arrays of lysates and cell culture supernatants. We found that at 8 to 10 weeks of estimated gestational age, CD144(+) vessels predominantly express the venous endothelial cell marker PAL-E, whereas CD144(+)PAL-E(-) vessels compatible with arteries only appear at the end of the first trimester. Lymphatic progenitor cells at 8 weeks of estimated gestational age express CD31, CD144, Prox1, and temporary PAL-E. At that developmental stage not all lymphatic progenitor cells express podoplanin or Lyve-1, which are acquired with advancing gestational age in a stepwise fashion. Already in second-trimester human skin, the phenotype of blood and lymphatic vessels roughly resembles the one in adult skin. The expression pattern of angiogenic molecules in lysates and cell culture supernatants of prenatal skin did not reveal the expected bent to proangiogenic molecules, indicating a complex regulation of angiogenesis during ontogeny. In summary, this study provides enticing new insights into the development and phenotypic characteristics of the vascular system in human prenatal skin. PMID:26188132

  8. NOVEL CHARACTERIZATION OF bEnd.3 CELLS THAT EXPRESS LYMPHATIC VESSEL ENDOTHELIAL HYALURONAN RECEPTOR-1

    PubMed Central

    Yuen, D.; Leu, R.; Tse, J.; Wang, S.; Chen, L.L.; Chen, L.

    2015-01-01

    Murine bEnd.3 endothelioma cell line has been widely used in vascular research and here we report the novel finding that bEnd.3 cells express lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) and vascular endothelial growth factor receptor-3 (VEGFR-3). Moreover, these cells express progenitor cell markers of Sca-1 and CD133. Upon stimulation with tumor necrosis factor-alpha (TNF-α), the bEnd.3 cells demonstrate enhanced formation of capillary-type tubes, which express LYVE-1. As the bEnd.3 cell line is derived from murine endothelioma, we further examined human tissues of endothelioma and identified lymphatic vessels in the tumor samples which express both LYVE-1 and podoplanin. Moreover, a significantly higher number of lymphatic vessels were detected in the endothelioma samples compared with normal control. Taken together, this study not only redefines bEnd.3 cells for vascular research, but also indicates a broader category of human diseases that are associated with lymphatics, such as endothelioma. PMID:25282873

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

    PubMed

    Lanzinger, Stefanie; Breitner, Susanne; Neas, Lucas; Cascio, Wayne; Diaz-Sanchez, David; Hinderliter, Alan; Peters, Annette; Devlin, Robert B; Schneider, Alexandra

    2014-10-01

    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 investigate short-term effects of temperature and ozone on endothelial function in individuals having diabetes. Moreover, we investigated interactive effects between air temperature and air pollution on markers of endothelial function. Between November 2004 and December 2005 flow-mediated dilatation (FMD), nitroglycerin-mediated dilatation (NTGMD) and several blood markers representing endothelial function were measured using brachial artery ultrasound on four consecutive days in 22 individuals with type-2 diabetes mellitus in Chapel Hill, North Carolina (USA). Daily measurements of meteorological parameters, ozone and particulate matter with an aerodynamic diameter ≤2.5 µm (PM2.5) were obtained from fixed monitoring sites. We used additive mixed-models adjusting for time trend, day of the week, relative humidity and barometric pressure to assess temperature and ozone associations with endothelial function. A 1 °C decrease in the 24-h temperature average was associated with a decrease in mean FMD on the same day (-2.2% (95%-confidence interval:[-4.7;0.3%])) and with a delay of one and four days. A temperature decrement also led to an immediate (-1.7%[-3.3;-0.04]) decrease in NTGMD. Moreover, we observed an immediate (-14.6%[-26.3;-2.9%]) and a one day delayed (-13.5%[-27.0; 0.04%]) decrease in FMD in association with a 0.01 ppm increase in the maximum 8-h moving average of ozone. Temperature effects on FMD strengthened when PM2.5 and ozone concentrations were high. The associations were similar during winter and summer. We detected an association between temperature decreases and ozone increases on endothelial dysfunction in individuals having diabetes. We conclude that endothelial dysfunction

  10. 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

  11. Pathological lymphangiogenesis is modulated by galectin-8-dependent crosstalk between podoplanin and integrin-associated VEGFR-3

    PubMed Central

    Chen, Wei-Sheng; Cao, Zhiyi; Sugaya, Satoshi; Lopez, Maria J.; Sendra, Victor G.; Laver, Nora; Leffler, Hakon; Nilsson, Ulf J.; Fu, Jianxin; Song, Jianhua; Xia, Lijun; Hamrah, Pedram; Panjwani, Noorjahan

    2016-01-01

    Lymphangiogenesis plays a pivotal role in diverse pathological conditions. Here, we demonstrate that a carbohydrate-binding protein, galectin-8, promotes pathological lymphangiogenesis. Galectin-8 is markedly upregulated in inflamed human and mouse corneas, and galectin-8 inhibitors reduce inflammatory lymphangiogenesis. In the mouse model of corneal allogeneic transplantation, galectin-8-induced lymphangiogenesis is associated with an increased rate of corneal graft rejection. Further, in the murine model of herpes simplex virus keratitis, corneal pathology and lymphangiogenesis are ameliorated in Lgals8−/− mice. Mechanistically, VEGF-C-induced lymphangiogenesis is significantly reduced in the Lgals8−/− and Pdpn−/− mice; likewise, galectin-8-induced lymphangiogenesis is reduced in Pdpn−/− mice. Interestingly, knockdown of VEGFR-3 does not affect galectin-8-mediated lymphatic endothelial cell (LEC) sprouting. Instead, inhibiting integrins α1β1 and α5β1 curtails both galectin-8- and VEGF-C-mediated LEC sprouting. Together, this study uncovers a unique molecular mechanism of lymphangiogenesis in which galectin-8-dependent crosstalk among VEGF-C, podoplanin and integrin pathways plays a key role. PMID:27066737

  12. Markers of early endothelial dysfunction in intrauterine growth restriction-derived human umbilical vein endothelial cells revealed by 2D-DIGE and mass spectrometry analyses.

    PubMed

    Caniuguir, Andres; Krause, Bernardo J; Hernandez, Cherie; Uauy, Ricardo; Casanello, Paola

    2016-05-01

    Intrauterine growth restriction (IUGR) associates with fetal and placental vascular dysfunction, and increased cardiovascular risk later on life. We hypothesize that endothelial cells derived from IUGR umbilical veins present significant changes in the proteome which could be involved in the endothelial dysfunction associated to this conditions. To address this the proteome profile of human umbilical endothelial cells (HUVEC) isolated from control and IUGR pregnancies was compared by 2D-Differential In Gel Electrophoresis (DIGE) and further protein identification by MALDI-TOF MS. Using 2D-DIGE 124 spots were identified as differentially expressed between control and IUGR HUVEC, considering a cut-off of 2 fold change, which represented ∼10% of the total spots detected. Further identification by MALDI-TOF MS and in silico clustering of the proteins showed that those differentially expressed proteins between control and IUGR HUVEC were mainly related with cytoskeleton organization, proteasome degradation, oxidative stress response, mRNA processing, chaperones and vascular function. Finally Principal Component analysis of the identified proteins showed that differentially expressed proteins allow distinguishing between control and IUGR HUVEC based on their proteomic profile. This study demonstrates for the first time that IUGR-derived HUVEC maintained in primary culture conditions present an altered proteome profile, which could reflect an abnormal programming of endothelial function in this fetal condition. PMID:27208404

  13. Markers

    ERIC Educational Resources Information Center

    Healthy Schools Network, Inc., 2011

    2011-01-01

    Dry erase whiteboards come with toxic dry erase markers and toxic cleaning products. Dry erase markers labeled "nontoxic" are not free of toxic chemicals and can cause health problems. Children are especially vulnerable to environmental health hazards; moreover, schools commonly have problems with indoor air pollution, as they are more densely…

  14. Micro- and nano-topography to enhance proliferation and sustain functional markers of donor-derived primary human corneal endothelial cells.

    PubMed

    Muhammad, Rizwan; Peh, Gary S L; Adnan, Khadijah; Law, Jaslyn B K; Mehta, Jodhbir S; Yim, Evelyn K F

    2015-06-01

    One of the most common indications for corneal transplantation is corneal endothelium dysfunction, which can lead to corneal blindness. Due to a worldwide donor cornea shortage, alternative treatments are needed, but the development of new treatment strategies relies on the successful in vitro culture of primary human corneal endothelial cells (HCECs) because transformed cell lines and animal-derived corneal endothelial cells are not desirable for therapeutic applications. Primary HCECs are non-proliferative in vivo and challenging to expand in vitro while maintaining their characteristic cell morphology and critical markers. Biochemical cues such as growth factors and small molecules have been investigated to enhance the expansion of HCECs with a limited increase in proliferation. In this study, patterned tissue culture polystyrene (TCPS) was shown to significantly enhance the expansion of HCECs. The proliferation of HCECs increased up to 2.9-fold, and the expression amount and localization of cell-cell tight junction protein Zona Occludens-1 (ZO-1) was significantly enhanced when grown on 1 μm TCPS pillars. 250 nm pillars induced an optimal hexagonal morphology of HCEC cells. Furthermore, we demonstrated that the topographical effect on tight-junction expression and cell morphology could be maintained throughout each passage, and was effectively 'remembered' by the cells. Higher amount of tight-junction protein expression was maintained at cell junctions when topographic cues were removed in the successive seeding. This topographic memory suggested topography-exposed/induced cells would maintain the enhanced functional markers, which would be useful in cell-therapy based approaches to enable the in situ endothelial cell monolayer formation upon delivery. The development of patterned TCPS culture platforms could significantly benefit those researching human corneal endothelial cell cultivation for cell therapy, and tissue engineering applications. PMID:25796353

  15. Markers of endothelial cell activation and immune activation are increased in patients with severe leptospirosis and associated with disease severity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  16. Expression of Molecular Markers of Angiogenesis, Lymphangiogenesis, and Proliferation Depending on the Stage of Skin Melanoma.

    PubMed

    Bgatova, N P; Lomakin, A I; Fursov, S A; Kachesov, I V; Chepko, S A; Isakova, N B; Borodin, Yu I; Voytsitsky, V E; Konenkov, V I

    2016-08-01

    The expression of molecular markers characterizing activity of the tumor process and metastases (proliferation marker Ki-67, angiogenesis marker CD34, and lymphangiogenesis markers podoplanin and LYVE-1) was assessed by immunohictochemical method in the primary tumor specimens collected during surgery for cutaneous melanoma (40 patients). Proliferative activity of the tumor tissue and volume density of peritumoral blood and lymph vessels increased with increasing tumor malignancy, which could indicate the risk of metastases. PMID:27590758

  17. 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

    SciTech Connect

    Vormittag, Laurenz; Thurnher, Dietmar; Geleff, Silvana; Pammer, Johannes; Heiduschka, Gregor; Brunner, Markus; Grasl, Matthaeus Ch.; Erovic, Boban M.

    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. In 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.

  18. Dietary proteins improve endothelial function under fasting conditions but not in the postprandial state, with no effects on markers of low-grade inflammation.

    PubMed

    Teunissen-Beekman, Karianna F M; Dopheide, Janneke; Geleijnse, Johanna M; Bakker, Stephan J L; Brink, Elizabeth J; de Leeuw, Peter W; Schalkwijk, Casper G; van Baak, Marleen A

    2015-12-14

    Endothelial dysfunction (ED) and low-grade inflammation (LGI) have a role in the development of CVD. The two studies reported here explored the effects of dietary proteins and carbohydrates on markers of ED and LGI in overweight/obese individuals with untreated elevated blood pressure. In the first study, fifty-two participants consumed a protein mix or maltodextrin (3×20 g/d) for 4 weeks. Fasting levels and 12 h postprandial responses of markers of ED (soluble intercellular adhesion molecule 1 (sICAM), soluble vascular cell adhesion molecule 1 (sVCAM), soluble endothelial selectin and von Willebrand factor) and markers of LGI (serum amyloid A, C-reactive protein and sICAM) were evaluated before and after intervention. Biomarkers were also combined into mean Z-scores of ED and LGI. The second study compared 4 h postprandial responses of ED and LGI markers in forty-eight participants after ingestion of 0·6 g/kg pea protein, milk protein and egg-white protein. In addition, postprandial responses after maltodextrin intake were compared with a protein mix and sucrose. The first study showed significantly lower fasting ED Z-scores and sICAM after 4 weeks on the high-protein diet (P≤0·02). The postprandial studies found no clear differences of ED and LGI between test meals. However, postprandial sVCAM decreased more after the protein mix compared with maltodextrin in both studies (P≤0·04). In conclusion, dietary protein is beneficial for fasting ED, but not for fasting LGI, after 4 weeks of supplementation. On the basis of Z-scores, postprandial ED and LGI were not differentially affected by protein sources or carbohydrates. PMID:26400262

  19. Use of tritiated thymidine as a marker to compare the effects of matrix proteins on adult human vascular endothelial cell attachment: implications for seeding of vascular prostheses

    SciTech Connect

    Hasson, J.E.; Wiebe, D.H.; Sharefkin, J.B.; D'Amore, P.A.; Abbott, W.M.

    1986-11-01

    We have developed a technique to measure attachment of adult human vascular endothelial cells to test surfaces with tritiated thymidine used as a marker. With this technique, we measured attachment of adult human vascular endothelial cells to a series of extracellular matrix proteins, including fibronectin-coated (10 micrograms/cm/sup 2/), laminin-coated (10 micrograms/cm/sup 2/), and collagen-coated (1% gelatin) surfaces because of the role of these proteins in promoting cell attachment and growth. For a typical experiment, in the presence of serum, initial attachment (at 1 hour) was greatest on fibronectin-coated (63%) and gelatin-coated (60%) tissue culture plastic (polystyrene) and was least on laminin-coated (28%) or untreated polystyrene (18%). The data suggest that fibronectin, either alone, or with a more complex combination of extracellular components may need to be present on prosthetic surfaces to produce maximal cell attachment and subsequent growth to confluence in vivo. The described method of measuring attachment is independent of surface properties, ensures complete recovery of cells, and will allow systematic exploration of those properties that best support human endothelial cell attachment to vascular prosthetic surfaces.

  20. Anti-tumor angiogenesis effect of genetic fusion vaccine encoding murine beta-defensin 2 and tumor endothelial marker-8 in a CT-26 murine colorectal carcinoma model

    PubMed Central

    Liu, Ping; Xie, Ganfeng; Geng, Peiliang; Zheng, Chenhong; Li, Jianjun; Pan, Feng; Ruan, Zhihua; Liang, Houjie

    2015-01-01

    Tumor endothelial marker 8 (TEM8) is an endothelial-specific marker that is upregulated during tumor angiogenesis. We previously demonstrated that DNA-based vaccine encoding xenogeneic TEM8 can potentiate anti-angiogenesis immunotherapy of malignancy; nevertheless, it remains to be improved in minimizing immune tolerance. Recently, it has been reported that murine beta-defensin 2 (MBD2) is chemotactic for immature dendritic cells and plays a pivotal role in breaking immune tolerance. Herein, we constructed a genetic fusion vaccine encoding murine TEM8 and MBD2 to investigate whether the novel vaccine preferentially elicits therapeutic antitumor immune responses and suppresses cancerous angiogenesis in mouse models. The anti-angiogenesis effect was determined by microvessel density (MVD) using immunohistochemical staining. The efficacy of the fusion vaccine was primarily assessed by detecting cytotoxic T lymphocyte activity (51Cr-release assay). Enzyme-linked immunosorbent spot (ELISpot) assay was used to detect TEM8-specific INF-γ production, and the activity of CTL was further verified by a depletion of CD8+ T cells via anti-CD8 monoclonal antibody. Our results showed that the DNA fusion vaccine possessed an enhanced therapeutic antitumor immunity through anti-angiogenesis in BALB/c mice inoculated with CT26 cells, and this effect was generally attributed to stimulation of an antigen specific CD8+ T-cell response against mTEM8. In conclusion, our study demonstrated that the fusion vaccine based on mTEM8 and MBD2 induced autoimmunity against endothelial cells, resulting in deceleration of tumor growth, and could be potential therapeutical application in clinic. PMID:26064415

  1. Cell-Surface MMP-9 Protein Is a Novel Functional Marker to Identify and Separate Proangiogenic Cells from Early Endothelial Progenitor Cells Derived from CD133(+) Cells.

    PubMed

    Kanayasu-Toyoda, Toshie; Tanaka, Takeshi; Kikuchi, Yutaka; Uchida, Eriko; Matsuyama, Akifumi; Yamaguchi, Teruhide

    2016-05-01

    To develop cell therapies for ischemic diseases, endothelial progenitor cells (EPCs) have been expected to play a pivotal role in vascular regeneration. It is desirable to use a molecular marker that is related to the function of the cells. Here, a quantitative polymerase chain reaction array revealed that early EPCs derived from CD133(+) cells exhibited significant expression of MMP-9. Some populations of early EPCs expressed MMP-9 on the cell surface and others did not. We also attempted to separate the proangiogenic fraction from early EPCs derived from CD133(+) cells using a functional cell surface marker, and we then analyzed the MMP-9(+) and MMP-9(-) cell fractions. The MMP-9(+) cells not only revealed higher invasion ability but also produced a high amount of IL-8. Moreover, the stimulative effect of MMP-9(+) cells on angiogenesis in vitro and in vivo was prohibited by anti-IL-8 antibody. These data indicate that MMP-9 is one of the useful cell surface markers for the separation of angiogenic cells. Our treatment of early EPCs with hyaluronidase caused not only a downregulation of cell-surface MMP-9 but also a decrease in invasion ability, indicating that membrane-bound MMP-9, which is one of the useful markers for early EPCs, plays an important role in angiogenesis. Stem Cells 2016;34:1251-1262. PMID:26824798

  2. The Role of Podoplanin in the Biology of Differentiated Thyroid Cancers

    PubMed Central

    Rudzińska, Magdalena; Gaweł, Damian; Sikorska, Justyna; Karpińska, Kamila M.; Kiedrowski, Mirosław; Stępień, Tomasz; Marchlewska, Magdalena; Czarnocka, Barbara

    2014-01-01

    Podoplanin (PDPN), a mucin-type transmembrane glycoprotein specific to the lymphatic system is expressed in a variety of human cancers, and is regarded as a factor promoting tumor progression. The purpose of this study was to elucidate the molecular role of PDPN in the biology of thyroid cancer cells. PDPN expression was evaluated in primary thyroid carcinomas and thyroid carcinoma cell lines by RT-qPCR, Western blotting, IF and IHC. To examine the role of podoplanin in determining a cell's malignant potential (cellular migration, invasion, proliferation, adhesion, motility, apoptosis), a thyroid cancer cell line with silenced PDPN expression was used. We observed that PDPN was solely expressed in the cancer cells of 40% of papillary thyroid carcinoma (PTC) tissues. Moreover, PDPN mRNA and protein were highly expressed in PTC-derived TPC1 and BcPAP cell lines but were not detected in follicular thyroid cancer derived cell lines. PDPN knock-down significantly decreased cellular invasion, and modestly reduced cell migration, while proliferation and adhesion were not affected. Our results demonstrate that PDPN mediates the invasive properties of cells derived from papillary thyroid carcinomas, suggesting that podoplanin might promote PTC progression. PMID:24797369

  3. Epithelial deletion of podoplanin is dispensable for re-epithelialization of skin wounds.

    PubMed

    Baars, Sebastian; Bauer, Christine; Szabowski, Sibylle; Hartenstein, Bettina; Angel, Peter

    2015-10-01

    The mucin-like transmembrane protein podoplanin (PDPN) is prominently represented in tumor-associated gene expression signatures of numerous types of cancer including squamous cell carcinoma, and gain-of-function and knockdown approaches in tissue culture strongly suggested an important role of PDPN in cell proliferation, migration and adhesion. PDPN is absent during epidermal homeostasis but is highly expressed in basal keratinocytes during cutaneous wound healing. Enhanced motility of immortalized keratinocytes upon ectopic PDPN overexpression argues for wound healing defects upon podoplanin deficiency in keratinocytes; however, in vivo data that unequivocally define the impact of PDPN by functional studies in a physiologically relevant system are still missing. Here, we have applied an in vivo loss-of-function approach by generating a novel transgenic mouse line with keratinocyte-specific podoplanin deficiency. Performing cutaneous full-thickness excisional wounds to examine re-epithelialization capacity, unexpectedly, no defects were observed in wound healing properties of mutant mice. Similarly, PDPN-deficient primary keratinocytes showed no impairment in migration, adhesion or proliferation. Thus, PDPN function is not rate-limiting for re-epithelialization but may be functionally compensated by an as yet unknown protein. Our data also call for in vivo functional studies on PDPN in settings of skin tumor development and progression to clarify PDPN's role in skin pathology. PMID:26121181

  4. 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

  5. B-mode ultrasound-detected carotid artery lesions with and without acoustic shadowing and their association with markers of inflammation and endothelial activation: the atherosclerosis risk in communities study.

    PubMed

    Hunt, Kelly J; Pankow, James S; Offenbacher, Steven; Kritchevsky, Stephen B; Duncan, Bruce B; Shahar, Eyal; Sharrett, A Richey; Heiss, Gerardo

    2002-05-01

    In a cross-sectional study of 8695 men and women free of clinical CVD, aged 45-64 years at the 1987-1989 baseline Atherosclerosis Risk in Communities (ARIC) study exam, we examined the relationship between carotid artery lesions (CALs), with and without acoustic shadowing (AS) as an index of plaque mineralization, to systemic markers of inflammation and markers of endothelial function, including endothelial adhesion molecules. A three-level variable, based on the presence of extracranial CALs and AS, identified by B-mode ultrasound of six 1 cm arterial segments, defined the outcome. Among subjects without evidence of AS, after controlling for age, gender, ethnicity, study site, body mass index, hypertension, diabetes, and smoking status, CALs were associated with systemic markers of inflammation, including higher levels of fibrinogen [OR=1.24 (95% CI: 1.09, 1.40)] and white blood cell count [OR=1.37 (95% CI: 1.21, 1.56)]. Among subjects with a CAL, after controlling for the above risk factors as well as mean far wall intima-media thickness, AS was associated with higher levels of von Willebrand factor [OR=1.38 (95% CI: 1.10, 1.74)], a marker of endothelial activation. Associations with endothelial adhesion molecules were inconsistent. Further studies aimed at elucidating the mechanisms of arterial mineralization are warranted. PMID:11947908

  6. Insulin-like growth factor I messenger RNA and protein are expressed in the human lymph node and distinctly confined to subtypes of macrophages, antigen-presenting cells, lymphocytes and endothelial cells.

    PubMed

    Oberlin, Dominique; Fellbaum, Christian; Eppler, Elisabeth

    2009-11-01

    Insulin-like growth factor I (IGF-I) is a potent hormone that stimulates growth and differentiation and inhibits apoptosis in numerous tissues. Preliminary evidence suggests that IGF-I exerts differentiating, mitogenic and restoring activities in the immune system but the sites of synthesis of local IGF-I are unknown. Identification of these sites would allow the functional role of local IGF-I to be clarified. The presence of IGF-I in non-immune cells suggests that it acts as a trophic factor, while its occurrence in subtypes of lymphocytes or antigen-presenting cells indicates paracrine/autocrine direct regulatory involvement of IGF-I in the human immune response. The present study investigated the location of IGF-I messenger RNA and protein on archival human lymph node samples by in situ hybridization, immunohistochemistry and double immunofluorescence staining using an IGF-I probe and antisera specific for human IGF-I and CD3 (T lymphocytes), CD20 (B lymphocytes), CD68 (macrophages), CD21 (follicular dendritic cells), S100 (interdigitating dendritic cells) and podoplanin (fibroblastic reticular cells). Numerous cells within the B- and T-cell compartments expressed the IGF-I gene, and the majority of these cells were identified as macrophages. Solitary follicular dendritic cells exhibited IGF-I. A few T lymphocytes, and no B lymphocytes, contained IGF-I immunoreactive material. Furthermore, IGF-I immunoreactive cells outside the follicles that did not react with CD3, CD20, S100 or podoplanin markers were identified as high-endothelial venule (HEV) cells. From this we conclude that the main task of IGF-I in human non-tumoral lymph node may be autocrine and paracrine regulation of the differentiation, stimulation and survival of lymphocytes, antigen-presenting cells and macrophages and the differentiation and maintenance of HEV cells. PMID:20067534

  7. Insulin-like growth factor I messenger RNA and protein are expressed in the human lymph node and distinctly confined to subtypes of macrophages, antigen-presenting cells, lymphocytes and endothelial cells

    PubMed Central

    Oberlin, Dominique; Fellbaum, Christian; Eppler, Elisabeth

    2009-01-01

    Insulin-like growth factor I (IGF-I) is a potent hormone that stimulates growth and differentiation and inhibits apoptosis in numerous tissues. Preliminary evidence suggests that IGF-I exerts differentiating, mitogenic and restoring activities in the immune system but the sites of synthesis of local IGF-I are unknown. Identification of these sites would allow the functional role of local IGF-I to be clarified. The presence of IGF-I in non-immune cells suggests that it acts as a trophic factor, while its occurrence in subtypes of lymphocytes or antigen-presenting cells indicates paracrine/autocrine direct regulatory involvement of IGF-I in the human immune response. The present study investigated the location of IGF-I messenger RNA and protein on archival human lymph node samples by in situ hybridization, immunohistochemistry and double immunofluorescence staining using an IGF-I probe and antisera specific for human IGF-I and CD3 (T lymphocytes), CD20 (B lymphocytes), CD68 (macrophages), CD21 (follicular dendritic cells), S100 (interdigitating dendritic cells) and podoplanin (fibroblastic reticular cells). Numerous cells within the B- and T-cell compartments expressed the IGF-I gene, and the majority of these cells were identified as macrophages. Solitary follicular dendritic cells exhibited IGF-I. A few T lymphocytes, and no B lymphocytes, contained IGF-I immunoreactive material. Furthermore, IGF-I immunoreactive cells outside the follicles that did not react with CD3, CD20, S100 or podoplanin markers were identified as high-endothelial venule (HEV) cells. From this we conclude that the main task of IGF-I in human non-tumoral lymph node may be autocrine and paracrine regulation of the differentiation, stimulation and survival of lymphocytes, antigen-presenting cells and macrophages and the differentiation and maintenance of HEV cells. PMID:20067534

  8. Podoplanin is a component of extracellular vesicles that reprograms cell-derived exosomal proteins and modulates lymphatic vessel formation.

    PubMed

    Carrasco-Ramírez, Patricia; Greening, David W; Andrés, Germán; Gopal, Shashi K; Martín-Villar, Ester; Renart, Jaime; Simpson, Richard J; Quintanilla, Miguel

    2016-03-29

    Podoplanin (PDPN) is a transmembrane glycoprotein that plays crucial roles in embryonic development, the immune response, and malignant progression. Here, we report that cells ectopically or endogenously expressing PDPN release extracellular vesicles (EVs) that contain PDPN mRNA and protein. PDPN incorporates into membrane shed microvesicles (MVs) and endosomal-derived exosomes (EXOs), where it was found to colocalize with the canonical EV marker CD63 by immunoelectron microscopy. We have previously found that expression of PDPN in MDCK cells induces an epithelial-mesenchymal transition (EMT). Proteomic profiling of MDCK-PDPN cells compared to control cells shows that PDPN-induced EMT is associated with upregulation of oncogenic proteins and diminished expression of tumor suppressors. Proteomic analysis of exosomes reveals that MDCK-PDPN EXOs were enriched in protein cargos involved in cell adhesion, cytoskeletal remodeling, signal transduction and, importantly, intracellular trafficking and EV biogenesis. Indeed, expression of PDPN in MDCK cells stimulated both EXO and MV production, while knockdown of endogenous PDPN in human HN5 squamous carcinoma cells reduced EXO production and inhibited tumorigenesis. EXOs released from MDCK-PDPN and control cells both stimulated in vitro angiogenesis, but only EXOs containing PDPN were shown to promote lymphatic vessel formation. This effect was mediated by PDPN on the surface of EXOs, as demonstrated by a neutralizing specific monoclonal antibody. These results contribute to our understanding of PDPN-induced EMT in association to tumor progression, and suggest an important role for PDPN in EV biogenesis and/or release and for PDPN-EXOs in modulating lymphangiogenesis. PMID:26893367

  9. Podoplanin is a component of extracellular vesicles that reprograms cell-derived exosomal proteins and modulates lymphatic vessel formation

    PubMed Central

    Andrés, Germán; Gopal, Shashi K.; Martín-Villar, Ester; Renart, Jaime; Simpson, Richard J.; Quintanilla, Miguel

    2016-01-01

    Podoplanin (PDPN) is a transmembrane glycoprotein that plays crucial roles in embryonic development, the immune response, and malignant progression. Here, we report that cells ectopically or endogenously expressing PDPN release extracellular vesicles (EVs) that contain PDPN mRNA and protein. PDPN incorporates into membrane shed microvesicles (MVs) and endosomal-derived exosomes (EXOs), where it was found to colocalize with the canonical EV marker CD63 by immunoelectron microscopy. We have previously found that expression of PDPN in MDCK cells induces an epithelial-mesenchymal transition (EMT). Proteomic profiling of MDCK-PDPN cells compared to control cells shows that PDPN-induced EMT is associated with upregulation of oncogenic proteins and diminished expression of tumor suppressors. Proteomic analysis of exosomes reveals that MDCK-PDPN EXOs were enriched in protein cargos involved in cell adhesion, cytoskeletal remodeling, signal transduction and, importantly, intracellular trafficking and EV biogenesis. Indeed, expression of PDPN in MDCK cells stimulated both EXO and MV production, while knockdown of endogenous PDPN in human HN5 squamous carcinoma cells reduced EXO production and inhibited tumorigenesis. EXOs released from MDCK-PDPN and control cells both stimulated in vitro angiogenesis, but only EXOs containing PDPN were shown to promote lymphatic vessel formation. This effect was mediated by PDPN on the surface of EXOs, as demonstrated by a neutralizing specific monoclonal antibody. These results contribute to our understanding of PDPN-induced EMT in association to tumor progression, and suggest an important role for PDPN in EV biogenesis and/or release and for PDPN-EXOs in modulating lymphangiogenesis. PMID:26893367

  10. AKT hyper-phosphorylation associated with PI3K mutations in lymphatic endothelial cells from a patient with lymphatic malformation

    PubMed Central

    Boscolo, Elisa; Coma, Silvia; Luks, Valerie L.; Greene, Arin; Klagsbrun, Michael; Warman, Matthew L.; Bischoff, Joyce

    2014-01-01

    Lymphatic malformations (LM) are characterized by abnormal formation of lymphatic vessels and tissue overgrowth. The lymphatic vessels present in LM lesions may become blocked and enlarged as lymphatic fluid collects, forming a mass or cyst. Lesions are typically diagnosed during childhood, and are often disfiguring and life threatening. Available treatments consist of sclerotherapy, surgical removal and therapies to diminish complications. We isolated lymphatic endothelial cells (LM-LEC) from a surgically removed microcystic LM lesion. LM-LEC and normal human dermal-LEC (HD-LEC) expressed endothelial (CD31, VE-Cadherin) as well as lymphatic endothelial (Podoplanin, PROX1, LYVE1)-specific markers. Targeted gene sequencing analysis in patient-derived LM-LEC revealed the presence of two mutations in class I phosphoinositide 3-kinases (PI3K) genes. One is an inherited, premature stop codon in the PI3K regulatory subunit PIK3R3. The second is a somatic missense mutation in the PI3K catalytic subunit PIK3CA; this mutation has been found in association with overgrowth syndromes and cancer growth. LM-LEC exhibited angiogenic properties: both cellular proliferation and sprouting in collagen were significantly increased compared to HD-LEC. AKT-Thr308 was constitutively hyper-phosphorylated in LM-LEC. Treatment of LM-LEC with PI3-Kinase inhibitors Wortmannin and LY294 decreased cellular proliferation and prevented the phosphorylation of AKT-Thr308 in both HD-LEC and LM-LEC. Treatment with the mTOR inhibitor rapamycin also diminished cellular proliferation, sprouting and AKT phosphorylation, but only in LM-LEC. Our results implicate disrupted PI3K-AKT signaling in LEC isolated from a human lymphatic malformation lesion. PMID:25424831

  11. Evaluation of biochemical and clinical markers of endothelial dysfunction and their correlation with urinary albumin excretion in patients with type 1 diabetes mellitus.

    PubMed

    Polat, Sefika Burcak; Ugurlu, Nagihan; Aslan, Nabi; Cuhaci, Neslihan; Ersoy, Reyhan; Cakir, Bekir

    2016-04-01

    Objective Endothelial dysfunction (ED) plays an important role in the pathogenesis of diabetic nephropathy. The purpose of the study was to determine flow mediated endothelial dependent vasodilatation (FMD) measurements and serum soluble (s) endothelin-1 (ET-1), intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule (VCAM-1) levels in patients with type 1 diabetes mellitus (T1DM) with or without increased urinary albumin excretion (UAE) and compare them with the healthy controls. Subjects and methods Seventy three patients with T1DM were enrolled. Patients were divided into two subgroups according to microalbumin measurements in 24-hr urine collections. The diabetic patients without microalbuminuria (41 patients) were defined as Group I and those with microalbuminuria (32 patients) were defined as group II. A hundred age and sex matched healthy subjects participated as the control group (Group III). Serum sET-1, sICAM-1, sVCAM-1 levels and FMD measurements were determined in all participants. Results Median FMD measurement was significantly lower in the diabetic groups compared with the control group (6.6, 6.4 and 7.8% in Group I, II and III, respectively) (p < 0.05). FMD was negatively correlated with age (p = 0.042). Median serum sICAM-1 level was higher in the patient groups compared to the control group (p < 0.05). Median serum sVCAM-1 level was higher in the group of patients with increased albuminuria compared to the normoalbuinuric and the control group (p < 0.05). Serum sVCAM-1 level was found to be positively correlated with degree of urinary albumin excretion (p < 0.001). Conclusion We assume that sVCAM-1 may be used as a predictive marker for risk stratification for nephropathy development and progression. PMID:26886090

  12. 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

  13. 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...

  14. 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 Central

    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.

    2014-01-01

    Importance Modifying levels of factors associated with age-related macular degeneration (AMD) may decrease risk of 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 Longitudinal population-based cohort study. Setting Beaver Dam, Wisconsin. Participants 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 four follow-up examinations in 1993-1995, 1998-2000, 2003-2005, and 2008-2010. Exposures Serum markers of inflammation (high sensitivity C-reactive protein [hsCRP], tumor necrosis factor-α receptor 2 [TNF-αR2], interleukin-6 [IL-6], and white blood cell count), oxidative stress (8-isoprostane and total carbonyl content), and endothelial dysfunction (soluble vascular cell adhesion molecule-1 [sVCAM-1] and soluble intercellular adhesion molecule-1) were measured. Interactions with Complement Factor H (rs1061170) and Age-Related Maculopathy Susceptibility 2 (rs10490924), C3 (rs2230199) and C2/CFB (rs4151667) were examined using multiplicative models. AMD was assessed from fundus photographs. Main Outcome Measure 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, hsCRP (odds ratio [OR] comparing 4th to 1st quartile 2.18, P=0.005), TNF-αR2 (1.78, P=0.04), and IL-6 (1.78, P=0.03) were associated with the incidence of early AMD. Increased incidence of early AMD was associated with sVCAM-1 (OR per standard deviation on the log ng/mL scale 1.21, P=0.04). Conclusions and Relevance We found modest evidence of relationships of serum hsCRP, TNF-αR2, and IL-6 and sVCAM-1 to the 20

  15. Podoplanin expression in tumor-free resection margins of oral squamous cell carcinomas: an immunohistochemical and fractal analysis study.

    PubMed

    Margaritescu, C; Raica, M; Pirici, D; Simionescu, C; Mogoanta, L; Stinga, A C; Stinga, A S; Ribatti, D

    2010-06-01

    Podoplanin is involved in tumorigenesis and cancer progression in head and neck malignancies and its expression is not restricted to lymphatic vessel endothelium. The aim of this study was to establish podoplanin expression in the tumor-free resection margins of oral squamous cell carcinomas (OSCCs) and to evaluate the geometric complexity of the lymphatic vessels in oral mucosa by utilizing fractal analysis. As concerns the podoplanin expression in noncancerous tissue, forty tumor-free resection margins from OSCCs were investigated utilizing immunohistochemistry for D2-40 antibody and image densitometry analysis. Podoplanin expression was extremely low in basal cells, especially in resection margins of OSCCs developed in the lower lip regions. However, a highly variable D2-40 expression in tumor-free resection margins associated with hyperplastic or dysplastic lesions was identified. Moreover, podoplanin expression also extended to the basal layer of the lower lip skin appendages, the myoepithelial cells of acini and ducts of minor salivary glands, and other structures from the oral cavity. As concerns the study of the density and complexity of oral lymphatic vessels architecture by means of immunohistochemistry (D2-40, CD31 and Ki-67 antibodies) and fractal analysis, we demonstrated that in normal oral mucosa the geometry of the lymphatic vessels was less complex at the level of the lower lip compared to the anterior part of the oral floor mucosa or the tongue. A comparative analysis between the normal and pathological aspects revealed statistically significant differences between the fractal dimension (FD) of the vessels' outline, especially in the tongue. Fractal analysis proved an increasing lymphatic network complexity from normal to premalignant oral mucosal lesions, providing additional prognostic information in oral malignant tumors. PMID:20376776

  16. Upregulation of oxidative stress markers in human microvascular endothelial cells by complexes of serum albumin and digestion products of glycated casein.

    PubMed

    Deo, Permal; Glenn, Josephine V; Powell, Lesley A; Stitt, Alan W; Ames, Jennifer M

    2009-01-01

    The extent of absorption of dietary advanced glycation end products (AGEs) is not fully known. The possible physiological impact of these absorbed components on inflammatory processes has been studied little and was the aim of this investigation. Aqueous solutions of bovine casein and glucose were heated at 95 degrees C for 5 h to give AGE-casein (AGE-Cas). Simulated stomach and small intestine digestion of AGE-Cas and dialysis (molecular mass cutoff of membrane = 1 kDa) resulted in a low molecular mass (LMM) fraction of digestion products, which was used to prepare bovine serum albumin (BSA)-LMM-AGE-Cas complexes. Stimulation of human microvascular endothelial cells with BSA-LMM-AGE-Cas complexes significantly increased mRNA expression of the receptor of AGE (RAGE), galectin-3 (AGE-R3), tumor necrosis factor alpha, and a marker of the mitogen-activated protein kinase pathway (MAPK-1), as well as p65NF-kappaB activation. Cells treated with LMM digestion products of AGE-Cas significantly increased AGE-R3 mRNA expression. Intracellular reactive oxygen species production increased significantly in cells challenged with BSA-LMM-AGE-Cas and LMM-AGE-Cas. In conclusion, in an in vitro cell system, digested dietary AGEs complexed with serum albumin play a role in the regulation of RAGE and downstream inflammatory pathways. AGE-R3 may protect against these effects. PMID:19827132

  17. 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

  18. Detection of Autoantibodies to Vascular Endothelial Growth Factor Receptor-3 in Bile Duct Ligated Rats and Correlations with a Panel of Traditional Markers of Liver Diseases

    PubMed Central

    Duval, Florent; Cruz-Vega, Delia Elva; González-Gamboa, Ivonne; González-Garza, María Teresa; Ponz, Fernando; Sánchez, Flora; Alarcón-Galván, Gabriela; Moreno-Cuevas, Jorge E.

    2016-01-01

    There is a need for new noninvasive biomarkers (NIBMs) able to assess cholestasis and fibrosis in chronic cholestatic liver diseases (CCLDs). Tumorigenesis can arise from CCLDs. Therefore, autoantibodies to tumor-associated antigens (TAA) may be early produced in response to abnormal self-antigen expression caused by cholestatic injury. Vascular endothelial growth factor receptor-3 (VEGFR-3) has TAA potential since it is involved in cholangiocytes and lymphatic vessels proliferations during CCLDs. This study aims to detect autoantibodies directed at VEGFR-3 during bile duct ligation- (BDL-) induced cholestatic injury in rat sera and investigate whether they could be associated with traditional markers of liver damage, cholestasis, and fibrosis. An ELISA was performed to detect anti-VEGFR-3 autoantibodies in sera of rats with different degree of liver injury and results were correlated with aminotransferases, total bilirubin, and the relative fibrotic area. Mean absorbances of anti-VEGFR-3 autoantibodies were significantly increased from week one to week five after BDL. The highest correlation was observed with total bilirubin (R2 = 0.8450, P = 3.04e − 12). In conclusion, anti-VEGFR-3 autoantibodies are early produced during BDL-induced cholestatic injury, and they are closely related to cholestasis, suggesting the potential of anti-VEGFR-3 autoantibodies as NIBMs of cholestasis in CCLDs and justifying the need for further investigations in patients with CCLD. PMID:27212785

  19. Growth factors, their receptor expression and markers for proliferation of endothelial and neoplastic cells in human osteosarcoma.

    PubMed

    Bianchi, E; Artico, M; Di Cristofano, C; Leopizzi, M; Taurone, S; Pucci, M; Gobbi, P; Mignini, F; Petrozza, V; Pindinello, I; Conconi, M T; Della Rocca, C

    2013-01-01

    Osteosarcoma is the most common primary malignant tumour of the bone. Although new therapies continue to be reported, osteosarcoma-related morbidity and mortality remain high. Modern medicine has greatly increased knowledge of the physiopathology of this neoplasm. Novel targets for drug development may be identified through an understanding of the normal molecular processes that are deeply modified in pathological conditions. The aim of the present study is to investigate, by immunohistochemistry, the localisation of different growth factors and of the proliferative marker Ki-67 in order to determine whether these factors are involved in the transformation of osteogenic cells and in the development of human osteosarcoma. We observed a general positivity for NGF - TrKA - NT3 - TrKC - VEGF in the cytoplasm of neoplastic cells and a strong expression for NT4 in the nuclear compartment. TGF-beta was strongly expressed in the extracellular matrix and vascular endothelium. BDNF and TrKB showed a strong immunolabeling in the extracellular matrix. Ki-67/MIB-1 was moderately expressed in the nucleus of neoplastic cells. We believe that these growth factors may be considered potential therapeutic targets in the treatment of osteosarcoma, although proof of this hypothesis requires further investigation. PMID:24067459

  20. Interdependencies among Selected Pro-Inflammatory Markers of Endothelial Dysfunction, C-Peptide, Anti-Inflammatory Interleukin-10 and Glucose Metabolism Disturbance in Obese Women

    PubMed Central

    Janowska, Joanna; Chudek, Jerzy; Olszanecka-Glinianowicz, Magdalena; Semik-Grabarczyk, Elżbieta; Zahorska-Markiewicz, Barbara

    2016-01-01

    Background: Currently increasing importance is attributed to the inflammatory process as a crucial factor responsible for the progressive damage to vascular walls and progression of atherosclerosis in obese people. We have studied the relationship between clinical and biochemical parameters and C-peptide and anti-inflammatory IL-10, as well as selected markers of inflammation and endothelial dysfunction such as: CCL2, CRP, sICAM-1, sVCAM-1 and E-selectin in obese women with various degree of glucose metabolism disturbance. Material and methods: The studied group consisted of 61 obese women, and 20 normal weight, healthy volunteers. Obese patients were spited in subgroups based on the degree of glucose metabolism disorder. Serum samples were analyzed using ELISA kits. Results: Increased concentrations of sICAM-1, sVCAM-1, E-selectin, CCL2 and CRP were found in all obese groups compared to the normal weight subjects. In patients with Type 2 diabetes mellitus (T2DM) parameters characterizing the degree of obesity significantly positively correlated with levels of CRP and CCL2. Significant relationships were found between levels of glucose and sICAM-1and also E-selectin and HOMA-IR. C-peptide levels are positively associated with CCL2, E-selectin, triglycerides levels, and inversely with IL-10 levels in newly diagnosed T2DM group (p<0.05). Concentrations of IL-10 correlated negatively with E-selectin, CCL2, C-peptide levels, and HOMA-IR in T2DM group (p<0.05). Conclusion: Disturbed lipid and carbohydrate metabolism are manifested by enhanced inflammation and endothelial dysfunction in patients with simply obesity. These disturbances are associates with an increase of adhesion molecules. The results suggest the probable active participation of higher concentrations of C-peptide in the intensification of inflammatory and atherogenic processes in obese patients with type 2 diabetes. In patients with obesity and type 2 diabetes, altered serum concentrations of Il-10 seems

  1. 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

  2. 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

  3. Increased Vascular Endothelial Growth Factor in the Ventricular Cerebrospinal Fluid as a Predictive Marker for Subsequent Ventriculoperitoneal Shunt Infection : A Comparison Study among Hydrocephalic Patients

    PubMed Central

    Lee, Jeong-Hyun; Back, Dong-Bin; Cha, Yoo-Hyun; Kang, Shin-Hyuk; Suh, Jung-Keun

    2012-01-01

    Objective The aim of this study is to determine the association between the cerebrospinal fluid (CSF) biomarkers and inflammation, and the predictive value of these CSF biomarkers for subsequent shunt associated infection. Methods We obtained CSF samples from the patients with hydrocephalus during ventriculoperitoneal (VP) shunt operations. Twenty-two patients were enrolled for this study and divided into 3 groups: subarachnoid hemorrhage (SAH)-induced hydrocephalus, idiopathic normal pressure hydrocephalus (INPH) and hydrocephalus with a subsequent shunt infection. We analyzed the transforming growth factor-β1, tumor necrosis factor-α, vascular endothelial growth factor (VEGF) and total tau in the CSF by performing enzyme-linked immunosorbent assay. The subsequent development of shunt infection was confirmed by the clinical presentations, the CSF parameters and CSF culture from the shunt devices. Results The mean VEGF concentration (±standard deviation) in the CSF of the SAH-induced hydrocephalus, INPH and shunt infection groups was 236±138, 237±80 and 627±391 pg/mL, respectively. There was a significant difference among the three groups (p=0.01). Between the SAH-induced hydrocephalus and infection groups and between the INPH and infection groups, there was a significant difference of the VEGF levels (p<0.01). However, the other marker levels did not differ among them. Conclusion The present study showed that only the CSF VEGF levels are associated with the subsequent development of shunt infection. Our results suggest that increased CSF VEGF could provide a good condition for bacteria that are introduced at the time of surgery to grow in the brain, rather than reflecting a sequel of bacterial infection before VP shunt. PMID:22949960

  4. 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

  5. LpMab-12 Established by CasMab Technology Specifically Detects Sialylated O-Glycan on Thr52 of Platelet Aggregation-Stimulating Domain of Human Podoplanin

    PubMed Central

    Kato, Yukinari; Ogasawara, Satoshi; Oki, Hiroharu; Goichberg, Polina; Honma, Ryusuke; Fujii, Yuki; Kaneko, Mika K.

    2016-01-01

    Podoplanin (PDPN), also known as Aggrus, possesses three tandem repeat of platelet aggregation-stimulating (PLAG) domains in its N-terminus. Among the PLAG domains, sialylated O-glycan on Thr52 of PLAG3 is essential for the binding to C-type lectin-like receptor-2 (CLEC-2) and the platelet-aggregating activity of human PDPN (hPDPN). Although various anti-hPDPN monoclonal antibodies (mAbs) have been generated, no specific mAb has been reported to target the epitope containing glycosylated Thr52. We recently established CasMab technology to develop mAbs against glycosylated membrane proteins. Herein, we report the development of a novel anti-glycopeptide mAb (GpMab), LpMab-12. LpMab-12 detected endogenous hPDPN by flow cytometry. Immunohistochemical analyses also showed that hPDPN-expressing lymphatic endothelial and cancer cells were clearly labeled by LpMab-12. The minimal epitope of LpMab-12 was identified as Asp49–Pro53 of hPDPN. Furthermore, LpMab-12 reacted with the synthetic glycopeptide of hPDPN, corresponding to 38–54 amino acids (hpp3854: 38-EGGVAMPGAEDDVVTPG-54), which carries α2–6 sialylated N-acetyl-D-galactosamine (GalNAc) on Thr52. LpMab-12 did not recognize non-sialylated GalNAc-attached glycopeptide, indicating that sialylated GalNAc on Thr52 is necessary for the binding of LpMab-12 to hPDPN. Thus, LpMab-12 could serve as a new diagnostic tool for determining whether hPDPN possesses the sialylation on Thr52, a site-specific post-translational modification critical for the hPDPN association with CLEC-2. PMID:27031228

  6. Mutation of Threonine 34 in Mouse Podoplanin-Fc Reduces CLEC-2 Binding and Toxicity in Vivo While Retaining Anti-lymphangiogenic Activity*

    PubMed Central

    Bianchi, Roberta; Fischer, Eliane; Yuen, Don; Ernst, Ellen; Ochsenbein, Alexandra M.; Chen, Lu; Otto, Vivianne I.; Detmar, Michael

    2014-01-01

    The lymphatic system plays an important role in cancer metastasis and inhibition of lymphangiogenesis could be valuable in fighting cancer dissemination. Podoplanin (Pdpn) is a small, transmembrane glycoprotein expressed on the surface of lymphatic endothelial cells (LEC). During mouse development, binding of Pdpn to the C-type lectin-like receptor 2 (CLEC-2) on platelets is critical for the separation of the lymphatic and blood vascular systems. Competitive inhibition of Pdpn functions with a soluble form of the protein, Pdpn-Fc, leads to reduced lymphangiogenesis in vitro and in vivo. However, the transgenic overexpression of human Pdpn-Fc in mouse skin causes disseminated intravascular coagulation due to platelet activation via CLEC-2. In the present study, we produced and characterized a mutant form of mouse Pdpn-Fc, in which threonine 34, which is considered essential for CLEC-2 binding, was mutated to alanine (PdpnT34A-Fc). Indeed, PdpnT34A-Fc displayed a 30-fold reduced binding affinity for CLEC-2 compared with Pdpn-Fc. This also translated into fewer side effects due to platelet activation in vivo. Mice showed less prolonged bleeding time and fewer embolized vessels in the liver, when PdpnT34A-Fc was injected intravenously. However, PdpnT34A-Fc was still as active as wild-type Pdpn-Fc in inhibiting lymphangiogenesis in vitro and also inhibited lymphangiogenesis in vivo. These data suggest that the function of Pdpn in lymphangiogenesis does not depend on threonine 34 in the CLEC-2 binding domain and that PdpnT34A-Fc might be an improved inhibitor of lymphangiogenesis with fewer toxic side effects. PMID:24907275

  7. Short communication: Effect of supplementation with Lactobacillus casei Shirota on insulin sensitivity, β-cell function, and markers of endothelial function and inflammation in subjects with metabolic syndrome--a pilot study.

    PubMed

    Tripolt, N J; Leber, B; Blattl, D; Eder, M; Wonisch, W; Scharnagl, H; Stojakovic, T; Obermayer-Pietsch, B; Wascher, T C; Pieber, T R; Stadlbauer, V; Sourij, H

    2013-01-01

    Based on animal studies, intake of probiotic bacteria was suggested to improve insulin sensitivity by reducing endotoxinemia and inflammation. The objective of this study was to determine the effects of supplementation with the probiotic strain Lactobacillus casei Shirota (LcS) over 12 wk on insulin sensitivity, β-cell function, inflammation, and endothelial dysfunction parameters in subjects with metabolic syndrome. In a randomized-controlled study, 30 subjects with metabolic syndrome either received Lactobacillus casei Shirota 3 times daily for 12 wk or served as controls with standard medical therapy. Fasting blood samples were taken and a 75-g oral glucose tolerance test was performed to derive indices for insulin sensitivity and β-cell function. In addition, parameters to assess endothelial function and inflammation markers were determined. Even though the insulin sensitivity index significantly improved after 3 mo of probiotic supplementation (0.058±0.021 vs. 0.038±0.025), the change was not significantly different compared with the control group. No improvements were seen in additional indices of insulin sensitivity (quantitative insulin sensitivity check index, insulin sensitivity by oral glucose tolerance test, and homeostasis model assessment for insulin resistance) and β-cell function (first and second phase insulin secretion, and homeostasis model assessment for β-cell function). Probiotic supplementation resulted in a significant reduction in soluble vascular cell adhesion molecule-1 (sVCAM-1) level (1,614±343 vs. 1,418±265 ng/mL). No significant changes in parameters used to assess low-grade inflammation or endothelial dysfunction were observed. Intake of LcS for 12 wk in subjects with metabolic syndrome did not improve insulin sensitivity, β-cell function, endothelial function, or inflammation markers in this trial. PMID:23164226

  8. [Endothelial function test].

    PubMed

    Tomiyama, Hirofumi

    2015-11-01

    Endothelial dysfunction is thought to have pivotal roles for the development of hypertension, initiation/progression of hypertensive organ damages, and prognosis. In clinical setting, flow-mediated vasodilatation (FMD) of brachial artery is used as a marker of endothelial function. However, well-trained sonographer is needed to conduct FMD measurement, and therefore, FMD has not been fully standardized (i.e., the reference value of FMD has not been established). Even so, FMD predicts future cardiovascular events. Lifestyle modifications (i.e., smoking cessation, exercise, or weight loss) and antihypertensive medication provide beneficial effects on endothelial function. Thus, FMD have a potential as a useful surrogate marker for the management of hypertension. PMID:26619655

  9. Expression of Aggrus/podoplanin in bladder cancer and its role in pulmonary metastasis

    PubMed Central

    Takagi, Satoshi; Oh-hara, Tomoko; Sato, Shigeo; Gong, Bo; Takami, Miho; Fujita, Naoya

    2014-01-01

    Platelet aggregation-inducing factor Aggrus, also known as podoplanin, is associated with tumor malignancy by promoting hematogenous metastasis. Aggrus overexpression has been reported in some tumor tissues including lung, esophagus, head and neck and brain. We here found the frequent upregulation of aggrus mRNA in urinary bladder cancers using cancer tissue panels from various organs. Immunohistochemical analysis confirmed Aggrus protein expression in urinary bladder cancers and suggested a positive correlation between Aggrus expression and metastatic tendency in bladder cancers. Endogenous expression of Aggrus protein on the cell surface was found in the mouse bladder cancer MBT-2 cell line and human bladder cancer SCaBER cell lines. Knockdown of Aggrus expression in MBT-2 cells decreased their ability to induce platelet aggregation and form pulmonary metastasis in syngeneic mouse models. Knockdown of Aggrus expression in the human bladder cancer SCaBER cells also attenuated their ability to induce platelet aggregation and form pulmonary metastasis in mice. Moreover, pulmonary metastasis of SCaBER cells was prevented by prior administration of our generated anti-Aggrus neutralizing monoclonal antibodies by attenuating their retention in lung. These results indicate that Aggrus plays an important role in bladder cancer metastasis. Thus, anti-Aggrus neutralizing antibodies would be useful for the prevention of hematogenous metastasis of Aggrus-positive bladder cancer. PMID:24222607

  10. Platelets Promote Tumor Growth and Metastasis via Direct Interaction between Aggrus/Podoplanin and CLEC-2

    PubMed Central

    Takagi, Satoshi; Sato, Shigeo; Oh-hara, Tomoko; Takami, Miho; Koike, Sumie; Mishima, Yuji; Hatake, Kiyohiko; Fujita, Naoya

    2013-01-01

    The platelet aggregation-inducing factor Aggrus, also known as podoplanin, is frequently upregulated in several types of tumors and enhances hematogenous metastasis by interacting with and activating the platelet receptor CLEC-2. Thus, Aggrus–CLEC-2 binding could be a therapeutic molecular mechanism for cancer therapy. We generated a new anti-human Aggrus monoclonal antibody, MS-1, that suppressed Aggrus–CLEC-2 binding, Aggrus-induced platelet aggregation, and Aggrus-mediated tumor metastasis. Interestingly, the MS-1 monoclonal antibody attenuated the growth of Aggrus-positive tumors in vivo. Moreover, the humanized chimeric MS-1 antibody, ChMS-1, also exhibited strong antitumor activity against Aggrus-positive lung squamous cell carcinoma xenografted into NOD-SCID mice compromising antibody-dependent cellular cytotoxic and complement-dependent cytotoxic activities. Because Aggrus knockdown suppressed platelet-induced proliferation in vitro and tumor growth of the lung squamous cell carcinoma in vivo, Aggrus may be involved in not only tumor metastasis but also tumor growth by promoting platelet-tumor interaction, platelet activation, and secretion of platelet-derived factors in vivo. Our results indicate that molecular target drugs inhibiting specific platelet–tumor interactions can be developed as antitumor drugs that suppress both metastasis and proliferation of tumors such as lung squamous cell carcinoma. PMID:23991201

  11. [Stable expression of recombinant human podoplanin in Chinese hamster ovary (CHO) cells].

    PubMed

    Qu, Le; Zhao, Xingpeng; Fu, Jianxin; Xia, Lijun; Dai, Lan; Ruan, Changgeng; Zhao, Yiming

    2016-01-01

    Objective To construct podoplanin (PDPN) eukaryotic expression plasmid PDPN-pEGFP-N1, establish Chinese hamster ovary (CHO) cell line stably expressing recombinant human PDPN and investigate its biological activity. Methods PDPN cDNA was cloned from HEK293 cells by reverse transcription PCR and recombinant DNA technology and inserted into plasmid pEGFP-N1 labeled by enhanced green fluorescent protein (EGFP). The recombinant vector was identified by PCR, restriction enzyme digestion and DNA sequencing, and then transfected into CHO cells. Recombinant PDPN-EGFP was observed by fluorescent microscopy and CHO cell line with the high expression of PDPN-EGFP was selected by flow cytometry. Recombinant PDPN was detected by Western blotting and the biological activity of the cell line was determined by platelet aggregation assay. Results DNA sequencing and restriction enzyme digestion proved that the gene of PDPN was inserted successfully into pEGFP-N1 plasmid. After stable transfection of the recombinant plasmid into CHO cells, CHO with EGFP could be seen under a fluorescent microscope. The CHO cell line with the high expression of recombinant PDPN-EGFP was obtained after sorting by flow cytometry. Western blotting showed that the recombinant PDPN was expressed on the cell surface. The over-expressing PDPN-EGFP CHO cells were able to induce human platelet aggregation. Conclusion The CHO cell line with the stable and high expression of recombinant PDPN-EGFP has been constructed successfully, and it could induce platelet aggregation. PMID:26728373

  12. Up-regulation of podoplanin involves in neuronal apoptosis in LPS-induced neuroinflammation.

    PubMed

    Song, Yan; Shen, Jianhong; Lin, Yuchang; Shen, Jiabing; Wu, Xinming; Yan, Yaohua; Zhou, Li; Zhang, Haiyan; Zhou, Ying; Cao, Maohong; Liu, Yonghua

    2014-08-01

    Podoplanin (PDPN) is a mucin-type transmembrane sialoglycoprotein expressed in multiple tissues in adult animals, including the brain, lungs, kidney, and lymphoid organs. Studies of this molecule have demonstrated its great importance in tumor metastasis, platelet aggregation, and lymphatic vessel formation. However, information regarding its regulation and possible function in the central nervous system is still limited. In this study, we performed a neuroinflammatory model by lipopolysaccharide (LPS) lateral ventral injection in adult rats and detected increased expression of PDPN in the brain cortex. Immunofluorescence indicated that PDPN was located in the neurons, but not astrocytes. Moreover, there was a concomitant up-regulation of active caspase-3, cyclin D1, and CDK4 in vivo and vitro studies. In addition, the expression of these three proteins in cortical primary neurons was decreased after knocking down PDPN by siRNA. Collectively, all these results suggested that the up-regulation of PDPN might be involved in neuronal apoptosis in neuroinflammation after LPS injection. PMID:24821010

  13. 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. Cancer Immunol Res; 4(3); 259-68. ©2016 AACR. PMID:26822025

  14. Angiomodulin, a marker of cancer vasculature, is upregulated by vascular endothelial growth factor and increases vascular permeability as a ligand of integrin αvβ3

    PubMed Central

    Komiya, Eriko; Sato, Hiroki; Watanabe, Naoko; Ise, Marii; Higashi, Shouichi; Miyagi, Yohei; Miyazaki, Kaoru

    2014-01-01

    Angiomodulin (AGM) is a member of insulin-like growth factor binding protein (IGFBP) superfamily and often called IGFBP-rP1 or IGFBP-7. AGM was originally identified as a tumor-derived cell adhesion factor, which was highly accumulated in blood vessels of human cancer tissues. AGM is also overexpressed in cancer-associated fibroblasts (CAFs) and activates fibroblasts. However, some studies have shown tumor-suppressing activity of AGM. To understand the roles of AGM in cancer progression, we here investigated the expression of AGM in benign and invasive breast cancers and its functions in cancer vasculature. Immunohistochemical analysis showed that AGM was highly expressed in cancer vasculature even in ductal carcinoma in situ (DCIS) as compared to normal vasculature, while its expression in CAFs was more prominent in invasive carcinomas than DCIS. In vitro analyses showed that AGM was strongly induced by vascular endothelial cell growth factor (VEGF) in vascular endothelial cells. Although AGM stimulated neither the growth nor migration of endothelial cells, it supported efficient adhesion of endothelial cells. Integrin αvβ3 was identified as a novel major receptor for AGM in vascular endothelial cells. AGM retracted endothelial cells by inducing actin stress fibers and loosened their VE-cadherin-mediated intercellular junction. Consequently, AGM increased vascular permeability both in vitro and in vivo. Furthermore, AGM and integrin αvβ3 were highly expressed and colocalized in cancer vasculature. These results suggest that AGM cooperates with VEGF to induce the aberrant functions of cancer vasculature as a ligand of integrin αvβ3. PMID:24737780

  15. An exquisite cross-control mechanism among endothelial cell fate regulators directs the plasticity and heterogeneity of lymphatic endothelial cells

    PubMed Central

    Kang, Jinjoo; Yoo, Jaehyuk; Lee, Sunju; Tang, Wanli; Aguilar, Berenice; Ramu, Swapnika; Choi, Inho; Otu, Hasan H.; Shin, Jay W.; Dotto, G. Paolo; Koh, Chester J.; Detmar, Michael

    2010-01-01

    Arteriovenous-lymphatic endothelial cell fates are specified by the master regulators, namely, Notch, COUP-TFII, and Prox1. Whereas Notch is expressed in the arteries and COUP-TFII in the veins, the lymphatics express all 3 cell fate regulators. Previous studies show that lymphatic endothelial cell (LEC) fate is highly plastic and reversible, raising a new concept that all 3 endothelial cell fates may coreside in LECs and a subtle alteration can result in a reprogramming of LEC fate. We provide a molecular basis verifying this concept by identifying a cross-control mechanism among these cell fate regulators. We found that Notch signal down-regulates Prox1 and COUP-TFII through Hey1 and Hey2 and that activated Notch receptor suppresses the lymphatic phenotypes and induces the arterial cell fate. On the contrary, Prox1 and COUP-TFII attenuate vascular endothelial growth factor signaling, known to induce Notch, by repressing vascular endothelial growth factor receptor-2 and neuropilin-1. We show that previously reported podoplanin-based LEC heterogeneity is associated with differential expression of Notch1 in human cutaneous lymphatics. We propose that the expression of the 3 cell fate regulators is controlled by an exquisite feedback mechanism working in LECs and that LEC fate is a consequence of the Prox1-directed lymphatic equilibrium among the cell fate regulators. PMID:20351309

  16. Evaluation of immunohistochemical markers of lymphatic and blood vessels in canine mammary tumours.

    PubMed

    Sleeckx, N; Van Brantegem, L; Fransen, E; Van den Eynden, G; Casteleyn, C; Veldhuis Kroeze, E; Van Ginneken, C

    2013-05-01

    Canine mammary tumours (CMTs) are the most common neoplasms in intact female dogs. Bitches with spontaneously arising CMTs represent a promising animal model for human breast cancer research. The aim of the present study was to develop an immunohistochemical protocol for the identification of blood and lymphatic vessels in CMTs. Antibodies specific for human lymphatic vessels (prox-1, lyve-1, podoplanin and D2-40) and blood vessels (von Willebrand factor [vWf], CD31 and CD34) were utilized. Serial sections of 18 samples (eight samples of normal canine mammary tissue, five benign and five malignant CMTs) were examined. Antibodies specific for podoplanin, D2-40 and CD34 showed no immunoreactivity with canine tissue. Prox-1 and CD31 were determined to be the most suitable markers for lymphatic and blood vessels, respectively. PMID:23123127

  17. 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

  18. Relationships of Circulating Carotenoid Concentrations with Several Markers of Inflammation, Oxidative Stress, and Endothelial Dysfunction: The Coronary Artery Risk Development in Young Adults (CARDIA)/Young Adult Longitudinal Trends in Antioxidants (YALTA) Study

    PubMed Central

    Hozawa, Atsushi; Jacobs, David R.; Steffes, Michael W.; Gross, Myron D.; Steffen, Lyn M.; Lee, Duk-Hee

    2008-01-01

    Background Serum carotenoid concentrations relate inversely to cardiovascular disease incidence. To clarify the effect of carotenoids on atherosclerotic risk factors, we examined the association of circulating carotenoids with inflammation, oxidative stress, endothelial dysfunction, and smoking. Methods Black and white men and women in the Coronary Artery Risk Development in Young Adults study, ages 18 to 30 years at recruitment (1985–1986) from 4 US cities, were investigated over 15 years. We included 2048 to 4580 participants in analyses of the sum of serum α-carotene, β-carotene, zeaxanthin/lutein, and β-cryptoxanthin concentrations and of lycopene at year 0 and at year 7. Results The year 0 sum of 4 carotenoids was inversely associated (all P <0.05) with year 0 leukocyte count (slope per sum carotenoid SD, −0.17); year 7 fibrinogen (slope, −0.10); year 7 and year 15 C-reactive protein (slope, −0.12 and −0.09); and year 15 F2-isoprostanes (slope, −13.0), soluble P-selectin (slope, −0.48), and soluble intercellular adhesion molecule-1 (sICAM1; slope, −5.1). Leukocyte counts and sICAM1 and F2-isoprostane concentrations had stronger associations in smokers than in nonsmokers, and sICAM1 concentrations were higher in the highest carotenoid quartile in smokers than in the lowest carotenoid quartile in nonsmokers. Superoxide dismutase was positively associated with the sum of 4 carotenoids (slope, 0.12; P <0.01). Lycopene was inversely associated only with sICAM1. The year 7 carotenoid associations with these markers were mostly similar to those at year 0. Conclusions Circulating serum carotenoids were associated, some interactively with smoking, in apparently beneficial directions with markers of inflammation, oxidative stress, and endothelial dysfunction. PMID:17234732

  19. The endothelial cell markers von Willebrand Factor (vWF), CD31 and CD34 are lost in glomerulonephritis and no longer correlate with the morphological indices of glomerular sclerosis, interstitial fibrosis, activity and chronicity.

    PubMed

    Gluhovschi, Cristina; Gluhovschi, Gheorghe; Potencz, Elena; Herman, Diana; Trandafirescu, Virginia; Petrica, Ligia; Velciov, Silvia; Bozdog, Gheorghe; Bob, Flaviu; Vernic, Corina; Cioca, Daniel

    2010-01-01

    Endothelial cells (ECs) are active participants of an inflammatory process in glomeruli. EC damage has been shown to play an important role in the progression of glomerulonephritis (GN). The degree of glomerular and peritubular capillary loss in models of progressive renal disease correlates with the severity of glomerulosclerosis and interstitial fibrosis. The aim of our study was to analyze the association of vWF, CD31 and CD34 immunoreactivity with the morphological indices of glomerular sclerosis, interstitial fibrosis, activity and chronicity in GN. A cross-sectional study of 22 patients with GN was conducted. Conventional stains (hematoxylin-eosin, periodic acid Schiff and Trichrome Gömöri stains) and immunohistochemistry (vWF, CD31 and CD34) were employed on kidney biopsies. Activity and chronicity of GN, as well as glomerular segmental sclerosis and interstitial fibrosis, were evaluated according to a scoring system initially used for lupus nephritis and antineutrophil-cytoplasmic-antibody-associated vasculitis. Immunohistochemistry was assessed using a semi-quantitative score. Statistical analysis was performed using EpiInfo 6.04. The mean patient age was 46.68+/-14.09; 14 patients were male, and eight were female. Performing Spearman's rank correlation test, no correlation was found between each marker and glomerular segmental sclerosis, interstitial fibrosis, activity and chronicity, which suggests a loss of these markers and microvasculature involvement. PMID:20675279

  20. Plasma haemostatic markers, endothelial function and ambulatory blood pressure changes with home versus hospital cardiac rehabilitation: the Birmingham Rehabilitation Uptake Maximisation Study

    PubMed Central

    Lee, K W; Blann, A D; Jolly, K; Lip, G Y H

    2006-01-01

    Background Cardiac rehabilitation is an accepted therapeutic intervention in patients after myocardial infarction or coronary revascularisation. The effects of cardiac rehabilitation programmes, whether home based or hospital based, on haemostatic indices (as reflected by fibrinogen, plasma viscosity, fibrin D‐dimer (an index of thrombogenesis), von Willebrand factor (vWf, an index of endothelial damage/dysfunction), soluble P‐selectin (an index of platelet activation)), vasomotor function (using flow‐mediated dilatation (FMD)) and ambulatory blood pressure (ABP) in patients with coronary heart disease are unknown. Methods 81 patients (66 men, mean (SD) 59 (11) years) after myocardial infarction or coronary revascularisation were randomised to comprehensive hospital‐based (n = 40) or home‐based (n = 41) cardiac rehabilitation. Plasma levels of vWf, D‐dimer, fibrinogen, soluble P‐selectin and plasma viscosity, as well as FMD and 24‐h ABP, were measured at baseline and after 3 months of cardiac rehabilitation. Results In patients who completed cardiac rehabilitation, levels of vWf, fibrinogen and D‐dimer were significantly lower and FMD improved (all p⩽0.001), whereas levels were unchanged in controls. Significant reductions were also observed in 24‐h mean systolic blood pressure, diastolic blood pressure and mean aortic pressure after completion of cardiac rehabilitation (all p<0.05). No significant differences were observed between the hospital‐based and home‐based cardiac rehabilitation programmes on these indices. Conclusions Cardiac rehabilitation improves haemostasis, endothelial function and ABP in patients with coronary heart disease, with no significant differences between home‐based and hospital‐based cardiac rehabilitation programmes. These effects may contribute to the beneficial effects of cardiac rehabilitation programmes on CV outcomes. PMID:16807272

  1. Effects of high-intensity interval training and moderate-intensity continuous training on endothelial function and cardiometabolic risk markers in obese adults.

    PubMed

    Sawyer, Brandon J; Tucker, Wesley J; Bhammar, Dharini M; Ryder, Justin R; Sweazea, Karen L; Gaesser, Glenn A

    2016-07-01

    We hypothesized that high-intensity interval training (HIIT) would be more effective than moderate-intensity continuous training (MICT) at improving endothelial function and maximum oxygen uptake (V̇o2 max) in obese adults. Eighteen participants [35.1 ± 8.1 (SD) yr; body mass index = 36.0 ± 5.0 kg/m(2)] were randomized to 8 wk (3 sessions/wk) of either HIIT [10 × 1 min, 90-95% maximum heart rate (HRmax), 1-min active recovery] or MICT (30 min, 70-75% HRmax). Brachial artery flow-mediated dilation (FMD) increased after HIIT (5.13 ± 2.80% vs. 8.98 ± 2.86%, P = 0.02) but not after MICT (5.23 ± 2.82% vs. 3.05 ± 2.76%, P = 0.16). Resting artery diameter increased after MICT (3.68 ± 0.58 mm vs. 3.86 ± 0.58 mm, P = 0.02) but not after HIIT (4.04 ± 0.70 mm vs. 4.09 ± 0.70 mm; P = 0.63). There was a significant (P = 0.02) group × time interaction in low flow-mediated constriction (L-FMC) between MICT (0.63 ± 2.00% vs. -2.79 ± 3.20%; P = 0.03) and HIIT (-1.04 ± 4.09% vs. 1.74 ± 3.46%; P = 0.29). V̇o2 max increased (P < 0.01) similarly after HIIT (2.19 ± 0.65 l/min vs. 2.64 ± 0.88 l/min) and MICT (2.24 ± 0.48 l/min vs. 2.55 ± 0.61 l/min). Biomarkers of cardiovascular risk and endothelial function were unchanged. HIIT and MICT produced different vascular adaptations in obese adults, with HIIT improving FMD and MICT increasing resting artery diameter and enhancing L-FMC. HIIT required 27.5% less total exercise time and ∼25% less energy expenditure than MICT. PMID:27255523

  2. Endothelial Lessons.

    PubMed

    Vanhoutte, Paul M

    2016-01-01

    This essay focuses on nine important lessons learned during more than thirty years of endothelial research. They include: the danger of hiding behind a word, the confusion generated by abbreviations, the need to define the physiological role of the response studied, the local role of endothelium- dependent responses, the strength of pharmacological analyses, endothelial dysfunction as consequence and cause of disease, the importance of rigorous protocols, the primacy of in vivo studies and the importance of serendipity. PMID:26638800

  3. Evaluation of Vascular Endothelial Growth Factor as a Prognostic Marker for Local Relapse in Early-Stage Breast Cancer Patients Treated With Breast-Conserving Therapy

    SciTech Connect

    Moran, Meena S.; Yang Qifeng; Goyal, Sharad; Harris, Lyndsay; Chung, Gina; Haffty, Bruce G.

    2011-12-01

    Purpose: Vascular endothelial growth factor (VEGF) is an important protein involved in the process of angiogenesis that has been found to correlate with relapse-free and overall survival in breast cancer, predominantly in locally advanced and metastatic disease. A paucity of data is available on the prognostic implications of VEGF in early-stage breast cancer; specifically, its prognostic value for local relapse after breast-conserving therapy (BCT) is largely unknown. The purpose of our study was to assess VEGF expression in a cohort of early-stage breast cancer patients treated with BCT and to correlate the clinical and pathologic features and outcomes with overexpression of VEGF. Methods and Materials: After obtaining institutional review board approval, the paraffin specimens of 368 patients with early-stage breast cancer treated with BCT between 1975 and 2005 were constructed into tissue microarrays with twofold redundancy. The tissue microarrays were stained for VEGF and read by a trained pathologist, who was unaware of the clinical details, as positive or negative according the standard guidelines. The clinical and pathologic data, long-term outcomes, and results of VEGF staining were analyzed. Results: The median follow-up for the entire cohort was 6.5 years. VEGF expression was positive in 56 (15%) of the 368 patients. Although VEGF expression did not correlate with age at diagnosis, tumor size, nodal status, histologic type, family history, estrogen receptor/progesterone receptor status, or HER-2 status, a trend was seen toward increased VEGF expression in the black cohort (26% black vs. 13% white, p = .068). Within the margin-negative cohort, VEGF did not predict for local relapse-free survival (RFS) (96% vs. 95%), nodal RFS (100% vs. 100%), distant metastasis-free survival (91% vs. 92%), overall survival (92% vs. 97%), respectively (all p >.05). Subset analysis revealed that VEGF was highly predictive of local RFS in node-positive, margin

  4. Methylglyoxal Induced Basophilic Spindle Cells with Podoplanin at the Surface of Peritoneum in Rat Peritoneal Dialysis Model

    PubMed Central

    Hirahara, Ichiro; Sato, Hideki; Imai, Toshimi; Onishi, Akira; Morishita, Yoshiyuki; Muto, Shigeaki; Kusano, Eiji; Nagata, Daisuke

    2015-01-01

    Peritoneal dialysis (PD) is a common treatment for patients with reduced or absent renal function. Long-term PD leads to peritoneal injury with structural changes and functional decline. At worst, peritoneal injury leads to encapsulating peritoneal sclerosis (EPS), which is a serious complication of PD. In order to carry out PD safely, it is important to define the mechanism of progression of peritoneal injury and EPS. We prepared rat models of peritoneal injury by intraperitoneal administration of glucose degradation products, such as methylglyoxal (MGO) or formaldehyde (FA), chlorhexidine gluconate (CG), and talc. In rats treated with MGO, peritoneal fibrous thickening with the appearance of basophilic spindle cells with podoplanin, cytokeratin, and α-smooth muscle actin at the surface of the peritoneum was observed. These cells may have been derived from mesothelial cells by epithelial-to-mesenchymal transition. In FA- or CG-treated rats, the peritoneum was thickened, and mesothelial cells were absent at the surface of the peritoneum. The CG- or MGO-treated rats presented with a so-called abdominal cocoon. In the talc-treated rats, extensive peritoneal adhesion and peritoneal thickening were observed. MGO-induced peritoneal injury model may reflect human histopathology and be suitable to analyze the mechanism of progression of peritoneal injury and EPS. PMID:26064894

  5. Podoplanin-positive periarteriolar stromal cells promote megakaryocyte growth and proplatelet formation in mice by CLEC-2.

    PubMed

    Tamura, Shogo; Suzuki-Inoue, Katsue; Tsukiji, Nagaharu; Shirai, Toshiaki; Sasaki, Tomoyuki; Osada, Makoto; Satoh, Kaneo; Ozaki, Yukio

    2016-03-31

    Megakaryopoiesis is the hierarchical differentiation of hematopoietic stem cells into megakaryocytes. Differentiating megakaryocytes undergo maturation characterized by endomitosis and produce numerous platelets through proplatelet formation. C-type lectin-like receptor 2 (CLEC-2) is a podoplanin (PDPN) receptor mainly expressed on platelets and megakaryocytes. Deletion of platelet/megakaryocyte CLEC-2 causes thrombocytopenia in mice; however, its contribution to megakaryopoiesis remains unknown. Here, we show that megakaryopoiesis is promoted through the CLEC-2/PDPN interaction in the vicinity of arterioles in the bone marrow (BM). We have also identified PDPN-expressing BM arteriolar stromal cells, tentatively termed as BM fibroblastic reticular cell (FRC)-like cells. Platelet/megakaryocyte-specific CLEC-2 conditional knockout (cKO) mice showed a decrease in the number of immature megakaryocytes. CLEC-2 wild-type megakaryocyte expansion was augmented in vitro by the addition of recombinant PDPN, but not cKO megakaryocytes. Moreover, megakaryocyte colonies were colocalized with periarteriolar BM FRC-like cells in the BM. Coculture of megakaryocytes with BM FRC-like cells augmented megakaryocyte expansion, which was dependent upon the CLEC-2/PDPN interaction. Furthermore, we found that the CLEC-2/PDPN interaction induces BM FRC-like cells to secrete chemokine (C-C motif) ligand 5 (CCL5) to facilitate proplatelet formation. These observations indicate that a reciprocal interaction between CLEC-2 on megakaryocytes and PDPN on BM FRC-like cells contributes to the periarteriolar megakaryopoietic microenvironment in mouse BM. PMID:26796360

  6. Plant lectin can target receptors containing sialic acid, exemplified by podoplanin, to inhibit transformed cell growth and migration.

    PubMed

    Ochoa-Alvarez, Jhon Alberto; Krishnan, Harini; Shen, Yongquan; Acharya, Nimish K; Han, Min; McNulty, Dean E; Hasegawa, Hitoki; Hyodo, Toshinori; Senga, Takeshi; Geng, Jian-Guo; Kosciuk, Mary; Shin, Seung S; Goydos, James S; Temiakov, Dmitry; Nagele, Robert G; Goldberg, Gary S

    2012-01-01

    Cancer is a leading cause of death of men and women worldwide. Tumor cell motility contributes to metastatic invasion that causes the vast majority of cancer deaths. Extracellular receptors modified by α2,3-sialic acids that promote this motility can serve as ideal chemotherapeutic targets. For example, the extracellular domain of the mucin receptor podoplanin (PDPN) is highly O-glycosylated with α2,3-sialic acid linked to galactose. PDPN is activated by endogenous ligands to induce tumor cell motility and metastasis. Dietary lectins that target proteins containing α2,3-sialic acid inhibit tumor cell growth. However, anti-cancer lectins that have been examined thus far target receptors that have not been identified. We report here that a lectin from the seeds of Maackia amurensis (MASL) with affinity for O-linked carbohydrate chains containing sialic acid targets PDPN to inhibit transformed cell growth and motility at nanomolar concentrations. Interestingly, the biological activity of this lectin survives gastrointestinal proteolysis and enters the cardiovascular system to inhibit melanoma cell growth, migration, and tumorigenesis. These studies demonstrate how lectins may be used to help develop dietary agents that target specific receptors to combat malignant cell growth. PMID:22844530

  7. Antibody and lectin target podoplanin to inhibit oral squamous carcinoma cell migration and viability by distinct mechanisms

    PubMed Central

    Ochoa-Alvarez, Jhon A.; Krishnan, Harini; Pastorino, John G.; Nevel, Evan; Kephart, David; Lee, Joseph J.; Retzbach, Edward P.; Shen, Yongquan; Fatahzadeh, Mahnaz; Baredes, Soly; Kalyoussef, Evelyne; Honma, Masaru; Adelson, Martin E.; Kaneko, Mika K.; Kato, Yukinari; Young, Mary Ann; Deluca-Rapone, Lisa; Shienbaum, Alan J.; Yin, Kingsley; Jensen, Lasse D.; Goldberg, Gary S.

    2015-01-01

    Podoplanin (PDPN) is a unique transmembrane receptor that promotes tumor cell motility. Indeed, PDPN may serve as a chemotherapeutic target for primary and metastatic cancer cells, particularly oral squamous cell carcinoma (OSCC) cells that cause most oral cancers. Here, we studied how a monoclonal antibody (NZ-1) and lectin (MASL) that target PDPN affect human OSCC cell motility and viability. Both reagents inhibited the migration of PDPN expressing OSCC cells at nanomolar concentrations before inhibiting cell viability at micromolar concentrations. In addition, both reagents induced mitochondrial membrane permeability transition to kill OSCC cells that express PDPN by caspase independent nonapoptotic necrosis. Furthermore, MASL displayed a surprisingly robust ability to target PDPN on OSCC cells within minutes of exposure, and significantly inhibited human OSCC dissemination in zebrafish embryos. Moreover, we report that human OSCC cells formed tumors that expressed PDPN in mice, and induced PDPN expression in infiltrating host murine cancer associated fibroblasts. Taken together, these data suggest that antibodies and lectins may be utilized to combat OSCC and other cancers that express PDPN. PMID:25826087

  8. Methylglyoxal Induced Basophilic Spindle Cells with Podoplanin at the Surface of Peritoneum in Rat Peritoneal Dialysis Model.

    PubMed

    Hirahara, Ichiro; Sato, Hideki; Imai, Toshimi; Onishi, Akira; Morishita, Yoshiyuki; Muto, Shigeaki; Kusano, Eiji; Nagata, Daisuke

    2015-01-01

    Peritoneal dialysis (PD) is a common treatment for patients with reduced or absent renal function. Long-term PD leads to peritoneal injury with structural changes and functional decline. At worst, peritoneal injury leads to encapsulating peritoneal sclerosis (EPS), which is a serious complication of PD. In order to carry out PD safely, it is important to define the mechanism of progression of peritoneal injury and EPS. We prepared rat models of peritoneal injury by intraperitoneal administration of glucose degradation products, such as methylglyoxal (MGO) or formaldehyde (FA), chlorhexidine gluconate (CG), and talc. In rats treated with MGO, peritoneal fibrous thickening with the appearance of basophilic spindle cells with podoplanin, cytokeratin, and α-smooth muscle actin at the surface of the peritoneum was observed. These cells may have been derived from mesothelial cells by epithelial-to-mesenchymal transition. In FA- or CG-treated rats, the peritoneum was thickened, and mesothelial cells were absent at the surface of the peritoneum. The CG- or MGO-treated rats presented with a so-called abdominal cocoon. In the talc-treated rats, extensive peritoneal adhesion and peritoneal thickening were observed. MGO-induced peritoneal injury model may reflect human histopathology and be suitable to analyze the mechanism of progression of peritoneal injury and EPS. PMID:26064894

  9. Plant Lectin Can Target Receptors Containing Sialic Acid, Exemplified by Podoplanin, to Inhibit Transformed Cell Growth and Migration

    PubMed Central

    Shen, Yongquan; Acharya, Nimish K.; Han, Min; McNulty, Dean E.; Hasegawa, Hitoki; Hyodo, Toshinori; Senga, Takeshi; Geng, Jian-Guo; Kosciuk, Mary; Shin, Seung S.; Goydos, James S.; Temiakov, Dmitry; Nagele, Robert G.; Goldberg, Gary S.

    2012-01-01

    Cancer is a leading cause of death of men and women worldwide. Tumor cell motility contributes to metastatic invasion that causes the vast majority of cancer deaths. Extracellular receptors modified by α2,3-sialic acids that promote this motility can serve as ideal chemotherapeutic targets. For example, the extracellular domain of the mucin receptor podoplanin (PDPN) is highly O-glycosylated with α2,3-sialic acid linked to galactose. PDPN is activated by endogenous ligands to induce tumor cell motility and metastasis. Dietary lectins that target proteins containing α2,3-sialic acid inhibit tumor cell growth. However, anti-cancer lectins that have been examined thus far target receptors that have not been identified. We report here that a lectin from the seeds of Maackia amurensis (MASL) with affinity for O-linked carbohydrate chains containing sialic acid targets PDPN to inhibit transformed cell growth and motility at nanomolar concentrations. Interestingly, the biological activity of this lectin survives gastrointestinal proteolysis and enters the cardiovascular system to inhibit melanoma cell growth, migration, and tumorigenesis. These studies demonstrate how lectins may be used to help develop dietary agents that target specific receptors to combat malignant cell growth. PMID:22844530

  10. Aging-associated oxidized albumin promotes cellular senescence and endothelial damage

    PubMed Central

    Luna, Carlos; Alique, Matilde; Navalmoral, Estefanía; Noci, Maria-Victoria; Bohorquez-Magro, Lourdes; Carracedo, Julia; Ramírez, Rafael

    2016-01-01

    Increased levels of oxidized proteins with aging have been considered a cardiovascular risk factor. However, it is unclear whether oxidized albumin, which is the most abundant serum protein, induces endothelial damage. The results of this study indicated that with aging processes, the levels of oxidized proteins as well as endothelial microparticles release increased, a novel marker of endothelial damage. Among these, oxidized albumin seems to play a principal role. Through in vitro studies, endothelial cells cultured with oxidized albumin exhibited an increment of endothelial damage markers such as adhesion molecules and apoptosis levels. In addition, albumin oxidation increased the amount of endothelial microparticles that were released. Moreover, endothelial cells with increased oxidative stress undergo senescence. In addition, endothelial cells cultured with oxidized albumin shown a reduction in endothelial cell migration measured by wound healing. As a result, we provide the first evidence that oxidized albumin induces endothelial injury which then contributes to the increase of cardiovascular disease in the elderly subjects. PMID:27042026

  11. Soluble fms-like tyrosine kinase-1 and endothelial adhesion molecules (intercellular cell adhesion molecule-1 and vascular cell adhesion molecule-1) as predictive markers for blood pressure reduction after renal sympathetic denervation.

    PubMed

    Dörr, Oliver; Liebetrau, Christoph; Möllmann, Helge; Gaede, Luise; Troidl, Christian; Rixe, Johannes; Hamm, Christian; Nef, Holger

    2014-05-01

    Renal sympathetic denervation (RSD) is a treatment option for patients with resistant arterial hypertension, but in some patients it is not successful. Predictive parameters on the success of RSD remain unknown. The angiogenic factors soluble fms-like tyrosine kinase-1 (sFLT-1), intercellular cell adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) are known to be associated with endothelial dysfunction, vascular remodeling, and hypertension. We evaluated whether sFLT-1, ICAM-1, and VCAM-1 are predictive markers for blood pressure reduction after RSD. Consecutive patients (n=55) undergoing renal denervation were included. Venous serum samples for measurement of sFlt-1, ICAM-1, and VCAM-1 were collected before and 6 months after RSD. A therapeutic response was defined as an office systolic blood pressure reduction of >10 mm Hg 6 months after RSD. A significant mean office systolic blood pressure reduction of 31.2 mm Hg was observed in 46 patients 6 months after RSD. Nine patients were classified as nonresponders, with a mean systolic blood pressure reduction of 4.6 mm Hg. At baseline, sFLT-1 levels were significantly higher in responders than in nonresponders (P<0.001) as were ICAM-1 (P<0.001) and VCAM-1 levels (P<0.01). The areas under the curve for sFLT-1, ICAM-1, and VCAM-1 were 0.82 (interquartile range, 0.718-0.921; P<0.001), 0.754 (0.654-0.854; P<0.001), and 0.684 (0.564-804; P=0.01), respectively, demonstrating prediction of an RSD response. Responders showed significantly higher serum levels of sFLT-1, ICAM-1, and VCAM-1 at baseline compared with nonresponders. Thus, this study identified for the first time potential biomarkers with a predictive value indicating a responder or nonresponder before renal denervation. PMID:24470464

  12. Endothelial Dysfunction in Type 2 Diabetes Mellitus.

    PubMed

    Dhananjayan, R; Koundinya, K S Srivani; Malati, T; Kutala, Vijay Kumar

    2016-10-01

    Endothelial dysfunction is an imbalance in the production of vasodilator factors and when this balance is disrupted, it predisposes the vasculature towards pro-thrombotic and pro-atherogenic effects. This results in vasoconstriction, leukocyte adherence, platelet activation, mitogenesis, pro-oxidation, impaired coagulation and nitric oxide production, vascular inflammation, atherosclerosis and thrombosis. Endothelial dysfunction is focussed as it is a potential contributor to the pathogenesis of vascular disease in diabetes mellitus. Under physiological conditions, there is a balanced release of endothelial-derived relaxing and contracting factors, but this delicate balance is altered in diabetes mellitus and atherosclerosis, thereby contributing to further progression of vascular and end-organ damage. This review focuses on endothelial dysfunction in atherosclerosis, insulin resistance, metabolic syndrome, oxidative stress associated with diabetes mellitus, markers and genetics that are implicated in endothelial dysfunction. PMID:27605734

  13. Podoplanin, novel 43-kd membrane protein of glomerular epithelial cells, is down-regulated in puromycin nephrosis.

    PubMed Central

    Breiteneder-Geleff, S.; Matsui, K.; Soleiman, A.; Meraner, P.; Poczewski, H.; Kalt, R.; Schaffner, G.; Kerjaschki, D.

    1997-01-01

    Puromycin aminonucleoside nephrosis (PAN), a rat model of human minimal change nephropathy, is characterized by extensive flattening of glomerular epithelial cell (podocyte) foot processes and by severe proteinuria. For comparison of expression of glomerular membrane proteins of normal and PAN rats, a membrane protein fraction of isolated rat glomeruli was prepared and monoclonal antibodies were raised against it. An IgG-secreting clone designated LF3 was selected that specifically immunolabeled podocytes of normal but not of PAN rats. The antigen of LF3 IgG was identified as a 43-kd glycoprotein. Molecular cloning of its cDNA was performed in a delta gt11 expression library prepared from mRNA of isolated rat glomeruli. The predicted amino acid sequence indicated a 166-amino-acid integral membrane protein with a single membrane-spanning domain, two potential phosphorylation sites in its short cytoplasmic tail, and six potential O-glycosylation sites in the large ectodomain. High amino acid sequence identities were found to membrane glycoproteins of rat lung and bone and mouse thymus epithelial cells as well as to a phorbol-ester-induced protein in a mouse osteoblast cell line and to a canine influenza C virus receptor. In PAN, expression of this 43-kd protein was selectively reduced to < 30%, as determined by quantitative immunogold electron microscopy, immunoblotting, and Northern blotting. These data provide evidence that transcription of the 43-kd transmembrane podocyte glycoprotein is specifically down-regulated in PAN. To indicate that this protein could be associated with transformation of arborized foot processes to flat feet (Latin, pes planus) we have called it podoplanin. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 10 Figure 12 Figure 13 Figure 14 Figure 15 PMID:9327748

  14. Transforming growth factor-β, insulin-like growth factor I/insulin-like growth factor I receptor and vascular endothelial growth factor-A: Prognostic and predictive markers in triple-negative and non-triple-negative breast cancer

    PubMed Central

    BAHHNASSY, ABEER; MOHANAD, MARWA; SHAARAWY, SABRY; ISMAIL, MANAL F.; EL-BASTAWISY, AHMED; ASHMAWY, ABEER M.; ZEKRI, ABDEL-RAHMAN

    2015-01-01

    In the current study, the prognostic and predictive values of serum transforming growth factor-β1 (TGF-β1), insulin-like growth factor I (IGF-I)/IGF-I receptor (IGF-IR) and vascular endothelial growth factor-A (VEGF-A) were evaluated in triple-negative and non-triple-negative breast cancer (TNBC and non-TNBC). The aim was to identify a group of serological biomarkers and to identify possible candidates for targeted therapy in patients with TNBC and non-TNBC. Protein levels of TGF-β1, IGF-I/IGF-IR and VEGF-A in the serum were measured in 43 TNBC, 53 non-TNBC and 20 normal control participants using quantitative ELISA assays. Results were correlated against standard prognostic factors, response to treatment and survival. TNBC was identified to be associated with poor prognosis and serum levels of VEGF-A and IGF/IGF-IR were significantly higher in the TNBC group compared with the non-TNBC group. IGF-IR and VEGF-A overexpression was observed to be correlated with TGF-β1 expression and all of the markers investigated were associated with metastasis and disease progression. In the multivariate analysis, VEGF-A, IGF-I and IGF-IR were observed to be independent predictors for overall survival, whereas TGF-β1 and lymph node status were identified as independent predictors for disease-free survival. The overall response rate was significantly lower in patients with TNBC and those with high levels of TGF-β1, IGF-I/IGF-IR and VEGF-A. In view of the present results, it was concluded that TGF-β1, IGF-I/IGF-IR and VEGF-A overexpression is associated with the presence of aggressive tumors, which exhibit an increased probability of metastasis, a poor response to treatment and reduced survival rate. This indicates that VEGF-A, IGF-IR and IGF-I have the potential to be used as surrogate biomarkers and are promising candidates for targeted therapy, particularly in patients with TNBC. PMID:25824321

  15. Lack of vimentin impairs endothelial differentiation of embryonic stem cells.

    PubMed

    Boraas, Liana C; Ahsan, Tabassum

    2016-01-01

    The cytoskeletal filament vimentin is inherent to the endothelial phenotype and is critical for the proper function of endothelial cells in adult mice. It is unclear, however, if the presence of vimentin is necessary during differentiation to the endothelial phenotype. Here we evaluated gene and protein expression of differentiating wild type embryonic stem cells (WT ESCs) and vimentin knockout embryonic stem cells (VIM -/- ESCs) using embryoid bodies (EBs) formed from both cell types. Over seven days of differentiation VIM -/- EBs had altered morphology compared to WT EBs, with a rippled outer surface and a smaller size due to decreased proliferation. Gene expression of pluripotency markers decreased similarly for EBs of both cell types; however, VIM -/- EBs had impaired differentiation towards the endothelial phenotype. This was quantified with decreased expression of markers along the specification pathway, specifically the early mesodermal marker Brachy-T, the lateral plate mesodermal marker FLK1, and the endothelial-specific markers TIE2, PECAM, and VE-CADHERIN. Taken together, these results indicate that the absence of vimentin impairs spontaneous differentiation of ESCs to the endothelial phenotype in vitro. PMID:27480130

  16. Lack of vimentin impairs endothelial differentiation of embryonic stem cells

    PubMed Central

    Boraas, Liana C.; Ahsan, Tabassum

    2016-01-01

    The cytoskeletal filament vimentin is inherent to the endothelial phenotype and is critical for the proper function of endothelial cells in adult mice. It is unclear, however, if the presence of vimentin is necessary during differentiation to the endothelial phenotype. Here we evaluated gene and protein expression of differentiating wild type embryonic stem cells (WT ESCs) and vimentin knockout embryonic stem cells (VIM −/− ESCs) using embryoid bodies (EBs) formed from both cell types. Over seven days of differentiation VIM −/− EBs had altered morphology compared to WT EBs, with a rippled outer surface and a smaller size due to decreased proliferation. Gene expression of pluripotency markers decreased similarly for EBs of both cell types; however, VIM −/− EBs had impaired differentiation towards the endothelial phenotype. This was quantified with decreased expression of markers along the specification pathway, specifically the early mesodermal marker Brachy-T, the lateral plate mesodermal marker FLK1, and the endothelial-specific markers TIE2, PECAM, and VE-CADHERIN. Taken together, these results indicate that the absence of vimentin impairs spontaneous differentiation of ESCs to the endothelial phenotype in vitro. PMID:27480130

  17. Endothelial Dysfunction in Chronic Inflammatory Diseases

    PubMed Central

    Steyers, Curtis M.; Miller, Francis J.

    2014-01-01

    Chronic inflammatory diseases are associated with accelerated atherosclerosis and increased risk of cardiovascular diseases (CVD). As the pathogenesis of atherosclerosis is increasingly recognized as an inflammatory process, similarities between atherosclerosis and systemic inflammatory diseases such as rheumatoid arthritis, inflammatory bowel diseases, lupus, psoriasis, spondyloarthritis and others have become a topic of interest. Endothelial dysfunction represents a key step in the initiation and maintenance of atherosclerosis and may serve as a marker for future risk of cardiovascular events. Patients with chronic inflammatory diseases manifest endothelial dysfunction, often early in the course of the disease. Therefore, mechanisms linking systemic inflammatory diseases and atherosclerosis may be best understood at the level of the endothelium. Multiple factors, including circulating inflammatory cytokines, TNF-α (tumor necrosis factor-α), reactive oxygen species, oxidized LDL (low density lipoprotein), autoantibodies and traditional risk factors directly and indirectly activate endothelial cells, leading to impaired vascular relaxation, increased leukocyte adhesion, increased endothelial permeability and generation of a pro-thrombotic state. Pharmacologic agents directed against TNF-α-mediated inflammation may decrease the risk of endothelial dysfunction and cardiovascular disease in these patients. Understanding the precise mechanisms driving endothelial dysfunction in patients with systemic inflammatory diseases may help elucidate the pathogenesis of atherosclerosis in the general population. PMID:24968272

  18. Marker development

    SciTech Connect

    Adams, M.R.

    1987-05-01

    This report is to discuss the marker development for radioactive waste disposal sites. The markers must be designed to last 10,000 years, and place no undue burdens on the future generations. Barriers cannot be constructed that preclude human intrusion. Design specifications for surface markers will be discussed, also marker pictograms will also be covered.

  19. Endothelial progenitor cells: identity defined?

    PubMed Central

    Timmermans, Frank; Plum, Jean; Yöder, Mervin C; Ingram, David A; Vandekerckhove, Bart; Case, Jamie

    2009-01-01

    Abstract In the past decade, researchers have gained important insights on the role of bone marrow (BM)-derived cells in adult neovascularization. A subset of BM-derived cells, called endothelial progenitor cells (EPCs), has been of particular interest, as these cells were suggested to home to sites of neovascularization and neoendothelialization and differentiate into endothelial cells (ECs) in situ, a process referred to as postnatal vasculogenesis. Therefore, EPCs were proposed as a potential regenerative tool for treating human vascular disease and a possible target to restrict vessel growth in tumour pathology. However, conflicting results have been reported in the field, and the identification, characterization, and exact role of EPCs in vascular biology is still a subject of much discussion. The focus of this review is on the controversial issues in the field of EPCs which are related to the lack of a unique EPC marker, identification challenges related to the paucity of EPCs in the circulation, and the important phenotypical and functional overlap between EPCs, haematopoietic cells and mature ECs. We also discuss our recent findings on the origin of endothelial outgrowth cells (EOCs), showing that this in vitro defined EC population does not originate from circulating CD133+ cells or CD45+ haematopoietic cells. PMID:19067770

  20. Effect of cinnamon, cardamom, saffron and ginger consumption on blood pressure and a marker of endothelial function in patients with type 2 diabetes mellitus: A randomized controlled clinical trial.

    PubMed

    Azimi, Paria; Ghiasvand, Reza; Feizi, Awat; Hosseinzadeh, Javad; Bahreynian, Maryam; Hariri, Mitra; Khosravi-Boroujeni, Hossein

    2016-06-01

    Herbal medicines with high amounts of phytochemicals have been shown to have beneficial effects on blood pressure (BP), endothelial function and anthropometric measures. This study aimed to determine the effect of herbal treatment on BP, endothelial function and anthropometric measures in patients with type 2 diabetes mellitus (T2DM). This clinical trial included 204 T2DM patients randomly assigned to four intervention groups receiving 3 g cinnamon, 3 g cardamom, 1 g saffron or 3 g ginger with three glasses of black tea, and one control group consuming only three glasses of tea without any herbals, for 8 weeks. Intercellular adhesion molecule-1 (ICAM-1), systolic and diastolic BP and anthropometric measures were collected at baseline and after 8 weeks. No significant difference was found between various medicinal plants in terms of influencing BP, serum soluble (s)ICAM-1 concentrations and anthropometric measures. However, in within-group comparison saffron and ginger intakes significantly reduced sICAM-1 concentrations (340.9 ± 14.4 vs 339.69 ± 14.4 ng/ml, p = 0.01, and 391.78 ± 16.0 vs 390.97 ± 15.8 ng/ml, p = 0.009, respectively) and ginger intake affected systolic BP (143.06 ± 0.2 vs 142.07 ± 0.2 mmHg, p = 0.02). Although administration of these herbal medicines as supplementary remedies could affect BP and sICAM-1 concentrations, there was no significant difference between the plants in terms of influencing anthropometric measures, BP and endothelial function. PMID:26758574

  1. Bone Markers

    MedlinePlus

    ... Alkaline Phosphatase; Osteocalcin; P1NP; Procollagen Type 1 N-Terminal Propeptide Formal name: Biochemical Markers of Bone Remodeling ... tests for evaluating bone turnover: C-telopeptide (C-terminal telopeptide of type 1 collagen (CTx)) – a marker ...

  2. Islet Endothelial Cells Derived From Mouse Embryonic Stem Cells.

    PubMed

    Jain, Neha; Lee, Eun Jung

    2016-01-01

    The islet endothelium comprises a specialized population of islet endothelial cells (IECs) expressing unique markers such as nephrin and α-1 antitrypsin (AAT) that are not found in endothelial cells in surrounding tissues. However, due to difficulties in isolating and maintaining a pure population of these cells, the information on these islet-specific cells is currently very limited. Interestingly, we have identified a large subpopulation of endothelial cells exhibiting IEC phenotype, while deriving insulin-producing cells from mouse embryonic stem cells (mESCs). These cells were identified by the uptake of low-density lipoprotein (LDL) and were successfully isolated and subsequently expanded in endothelial cell culture medium. Further analysis demonstrated that the mouse embryonic stem cell-derived endothelial cells (mESC-ECs) not only express classical endothelial markers, such as platelet endothelial cell adhesion molecule (PECAM1), thrombomodulin, intercellular adhesion molecule-1 (ICAM-1), and endothelial nitric oxide synthase (eNOS) but also IEC-specific markers such as nephrin and AAT. Moreover, mESC-ECs secrete basement membrane proteins such as collagen type IV, laminin, and fibronectin in culture and form tubular networks on a layer of Matrigel, demonstrating angiogenic activity. Further, mESC-ECs not only express eNOS, but also its eNOS expression is glucose dependent, which is another characteristic phenotype of IECs. With the ability to obtain highly purified IECs derived from pluripotent stem cells, it is possible to closely examine the function of these cells and their interaction with pancreatic β-cells during development and maturation in vitro. Further characterization of tissue-specific endothelial cell properties may enhance our ability to formulate new therapeutic angiogenic approaches for diabetes. PMID:25751085

  3. Focally regulated endothelial proliferation and cell death in human synovium.

    PubMed Central

    Walsh, D. A.; Wade, M.; Mapp, P. I.; Blake, D. R.

    1998-01-01

    Angiogenesis and vascular insufficiency each may support the chronic synovial inflammation of rheumatoid arthritis. We have shown by quantitative immunohistochemistry and terminal uridyl deoxynucleotide nick end labeling that endothelial proliferation and cell death indices were each increased in synovia from patients with rheumatoid arthritis compared with osteoarthritic and noninflamed controls, whereas endothelial fractional areas did not differ significantly among disease groups. Markers of proliferation were associated with foci immunoreactive for vascular endothelial growth factor and integrin alpha(v)beta3, whereas cell death was observed in foci in which immunoreactivities for these factors were weak or absent. No association was found with thrombospondin immunoreactivity. The balance between angiogenesis and vascular regression in rheumatoid synovitis may be determined by the focal expression of angiogenic and endothelial survival factors. Increased endothelial cell turnover may contribute to microvascular dysfunction and thereby facilitate persistent synovitis. Images Figure 1 Figure 3 Figure 4 PMID:9502411

  4. Endothelial Dysfunction and Inflammation: Immunity in Rheumatoid Arthritis

    PubMed Central

    Yang, XueZhi; Chang, Yan; Wei, Wei

    2016-01-01

    Inflammation, as a feature of rheumatoid arthritis (RA), leads to the activation of endothelial cells (ECs). Activated ECs induce atherosclerosis through an increased expression of leukocyte adhesion molecules. Endothelial dysfunction (ED) is recognized as a failure of endothelial repair mechanisms. It is also an early preclinical marker of atherosclerosis and is commonly found in RA patients. RA is now established as an independent cardiovascular risk factor, while mechanistic determinants of ED in RA are still poorly understood. An expanding body of study has shown that EC at a site of RA is both active participant and regulator of inflammatory process. Over the last decade, a role for endothelial dysfunction in RA associated with cardiovascular disease (CVD) has been hypothesized. At the same time, several maintenance drugs targeting this phenomenon have been tested, which has promising results. Assessment of endothelial function may be a useful tool to identify and monitor RA patients. PMID:27122657

  5. Clinical Significance of Endothelial Dysfunction in Essential Hypertension.

    PubMed

    Gkaliagkousi, Eugenia; Gavriilaki, Eleni; Triantafyllou, Areti; Douma, Stella

    2015-11-01

    The endothelium is recognized as a major determinant of vascular physiology and pathophysiology. Over the last few decades, a plethora of studies have implicated endothelial dysfunction in the progression of atherosclerosis and the subclinical target organ damage observed in essential hypertension. However, the clinical significance of diagnosing endothelial dysfunction in patients with essential hypertension remains under investigation. Although a number of vascular and non-vascular markers of endothelial dysfunction have been proposed, there is an ongoing quest for a marker in the clinical setting that is optimal, inexpensive, and reproducible. In addition, endothelial dysfunction emerges as a promising therapeutic target of agents that are readily available in clinical practice. In this context, a better understanding of its role in essential hypertension becomes of great importance. Here, we aim to investigate the clinical significance of endothelial dysfunction in essential hypertension by accumulating novel data on (a) early diagnosis using robust markers with prognostic value in cardiovascular risk prediction, (b) the association of endothelial dysfunction with subclinical vascular organ damage, and (c) potential therapeutic targets. PMID:26371063

  6. Vascular endothelial-cadherin downregulation as a feature of endothelial transdifferentiation in monocrotaline-induced pulmonary hypertension.

    PubMed

    Nikitopoulou, Ioanna; Orfanos, Stylianos E; Kotanidou, Anastasia; Maltabe, Violetta; Manitsopoulos, Nikolaos; Karras, Panagiotis; Kouklis, Panos; Armaganidis, Apostolos; Maniatis, Nikolaos A

    2016-08-01

    Increased pulmonary vascular resistance in pulmonary hypertension (PH) is caused by vasoconstriction and obstruction of small pulmonary arteries by proliferating vascular cells. In analogy to cancer, subsets of proliferating cells may be derived from endothelial cells transitioning into a mesenchymal phenotype. To understand phenotypic shifts transpiring within endothelial cells in PH, we injected rats with alkaloid monocrotaline to induce PH and measured lung tissue levels of endothelial-specific protein and critical differentiation marker vascular endothelial (VE)-cadherin. VE-cadherin expression by immonoblotting declined significantly 24 h and 15 days postinjection to rebound to baseline at 30 days. There was a concomitant increase in transcriptional repressors Snail and Slug, along with a reduction in VE-cadherin mRNA. Mesenchymal markers α-smooth muscle actin and vimentin were upregulated by immunohistochemistry and immunoblotting, and α-smooth muscle actin was colocalized with endothelial marker platelet endothelial cell adhesion molecule-1 by confocal microscopy. Apoptosis was limited in this model, especially in the 24-h time point. In addition, monocrotaline resulted in activation of protein kinase B/Akt, endothelial nitric oxide synthase (eNOS), nuclear factor (NF)-κB, and increased lung tissue nitrotyrosine staining. To understand the etiological relationship between nitrosative stress and VE-cadherin suppression, we incubated cultured rat lung endothelial cells with endothelin-1, a vasoconstrictor and pro-proliferative agent in pulmonary arterial hypertension. This resulted in activation of eNOS, NF-κB, and Akt, in addition to induction of Snail, downregulation of VE-cadherin, and synthesis of vimentin. These effects were blocked by eNOS inhibitor N(ω)-nitro-l-arginine methyl ester. We propose that transcriptional repression of VE-cadherin by nitrosative stress is involved in endothelial-mesenchymal transdifferentiation in experimental PH. PMID

  7. Identification of a novel platelet antagonist that binds to CLEC-2 and suppresses podoplanin-induced platelet aggregation and cancer metastasis

    PubMed Central

    Chang, Yu-Tsui; Lu, Meng-Hong; Huang, Tur-Fu; Chong, Kowit-Yu; Liao, Hsiang-Ruei; Cheng, Ju-Chien; Tseng, Ching-Ping

    2015-01-01

    Podoplanin (PDPN) enhances tumor metastases by eliciting tumor cell-induced platelet aggregation (TCIPA) through activation of platelet C-type lectin-like receptor 2 (CLEC-2). A novel and non-cytotoxic 5-nitrobenzoate compound 2CP was synthesized that specifically inhibited the PDPN/CLEC-2 interaction and TCIPA with no effect on platelet aggregation stimulated by other platelet agonists. 2CP possessed anti-cancer metastatic activity in vivo and augmented the therapeutic efficacy of cisplatin in the experimental animal model without causing a bleeding risk. Analysis of the molecular action of 2CP further revealed that Akt1/PDK1 and PKCμ were two alternative CLEC-2 signaling pathways mediating PDPN-induced platelet activation. 2CP directly bound to CLEC-2 and, by competing with the same binding pocket of PDPN in CLEC-2, inhibited PDPN-mediated platelet activation. This study provides evidence that 2CP is the first defined platelet antagonist with CLEC-2 binding activity. The augmentation in the therapeutic efficacy of cisplatin by 2CP suggests that a combination of a chemotherapeutic agent and a drug with anti-TCIPA activity such as 2CP may prove clinically effective. PMID:26528756

  8. Identification of a novel platelet antagonist that binds to CLEC-2 and suppresses podoplanin-induced platelet aggregation and cancer metastasis.

    PubMed

    Chang, Yao-Wen; Hsieh, Pei-Wen; Chang, Yu-Tsui; Lu, Meng-Hong; Huang, Tur-Fu; Chong, Kowit-Yu; Liao, Hsiang-Ruei; Cheng, Ju-Chien; Tseng, Ching-Ping

    2015-12-15

    Podoplanin (PDPN) enhances tumor metastases by eliciting tumor cell-induced platelet aggregation (TCIPA) through activation of platelet C-type lectin-like receptor 2 (CLEC-2). A novel and non-cytotoxic 5-nitrobenzoate compound 2CP was synthesized that specifically inhibited the PDPN/CLEC-2 interaction and TCIPA with no effect on platelet aggregation stimulated by other platelet agonists. 2CP possessed anti-cancer metastatic activity in vivo and augmented the therapeutic efficacy of cisplatin in the experimental animal model without causing a bleeding risk. Analysis of the molecular action of 2CP further revealed that Akt1/PDK1 and PKCμ were two alternative CLEC-2 signaling pathways mediating PDPN-induced platelet activation. 2CP directly bound to CLEC-2 and, by competing with the same binding pocket of PDPN in CLEC-2, inhibited PDPN-mediated platelet activation. This study provides evidence that 2CP is the first defined platelet antagonist with CLEC-2 binding activity. The augmentation in the therapeutic efficacy of cisplatin by 2CP suggests that a combination of a chemotherapeutic agent and a drug with anti-TCIPA activity such as 2CP may prove clinically effective. PMID:26528756

  9. Tumor and Endothelial Cell Hybrids Participate in Glioblastoma Vasculature

    PubMed Central

    El Hallani, Soufiane; Colin, Carole; El Houfi, Younas; Boisselier, Blandine; Marie, Yannick; Ravassard, Philippe; Labussière, Marianne; Mokhtari, Karima; Thomas, Jean-Léon; Delattre, Jean-Yves; Eichmann, Anne; Sanson, Marc

    2014-01-01

    Background. Recently antiangiogenic therapy with bevacizumab has shown a high but transient efficacy in glioblastoma (GBM). Indeed, GBM is one of the most angiogenic human tumors and endothelial proliferation is a hallmark of the disease. We therefore hypothesized that tumor cells may participate in endothelial proliferation of GBM. Materials and Methods. We used EGFR FISH Probe to detect EGFR amplification and anti-CD31, CD105, VE-cadherin, and vWF to identify endothelial cells. Endothelial and GBM cells were grown separately, labeled with GFP and DsRed lentiviruses, and then cocultured with or without contact. Results. In a subset of GBM tissues, we found that several tumor endothelial cells carry EGFR amplification, characteristic of GBM tumor cells. This observation was reproduced in vitro: when tumor stem cells derived from GBM were grown in the presence of human endothelial cells, a fraction of them acquired endothelial markers (CD31, CD105, VE-cadherin, and vWF). By transduction with GFP and DsRed expressing lentiviral vectors, we demonstrate that this phenomenon is due to cell fusion and not transdifferentiation. Conclusion. A fraction of GBM stem cells thus has the capacity to fuse with endothelial cells and the resulting hybrids may participate in tumor microvascular proliferation and in treatment resistance. PMID:24868550

  10. Involvement of marrow-derived endothelial cells in vascularization.

    PubMed

    Larrivée, B; Karsan, A

    2007-01-01

    Until recently, the adult neovasculature was thought to arise only through angiogenesis, the mechanism by which new blood vessels form from preexisting vessels through endothelial cell migration and proliferation. However, recent studies have provided evidence that postnatal neovasculature can also arise though vasculogenesis, a process by which endothelial progenitor cells are recruited and differentiate into mature endothelial cells to form new blood vessels. Evidence for the existence of endothelial progenitors has come from studies demonstrating the ability of bone marrow-derived cells to incorporate into adult vasculature. However, the exact nature of endothelial progenitor cells remains controversial. Because of the lack of definitive markers of endothelial progenitors, the in vivo contribution of progenitor cells to physiological and pathological neovascularization remains unclear. Early studies reported that endothelial progenitor cells actively integrate into the adult vasculature and are critical in the development of many types of vascular-dependent disorders such as neoplastic progression. Moreover, it has been suggested that endothelial progenitor cells can be used as a therapeutic strategy aimed at promoting vascular growth in a variety of ischemic diseases. However, increasing numbers of studies have reported no clear contribution of endothelial progenitors in physiological or pathological angiogenesis. In this chapter, we discuss the origin of the endothelial progenitor cell in the embryo and adult, and we discuss the cell's link to the primitive hematopoietic stem cell. We also review the potential significance of endothelial progenitor cells in the formation of a postnatal vascular network and discuss the factors that may account for the current lack of consensus of the scientific community on this important issue. PMID:17554506

  11. Human Pulmonary Endothelial Cells in Culture

    PubMed Central

    Johnson, Alice R.

    1980-01-01

    Endothelial cells were cultured from various different human vessels, including aortas, pulmonary, ovarian, and umbilical arteries, and pulmonary, ovarian, and umbilical veins. The cultured cells were identified as endothelial cells by the presence of Factor VIII antigen and antiotensin I converting enzyme (kininase II). They retained these markers for at least five passages in culture, and some cells had them for seven passages or more. Endothelial cells from the various vessels were compared with respect to their ability to metabolize angiotensins I and II and bradykinin. Cells from arteries had three to five times the angiotensin I converting enzyme activity as cells from veins. The activity of angiotensinase A (aspartyl aminopeptidase) had a similar distribution, and cells from arteries were consistently more active than cells from veins. Cultures of endothelial cells from pulmonary and umbilical vessels formed prostacyclin in response to mechanical stimulation. Media from cell monolayers that were subjected to a change of medium and gentle agitation inhibited aggregation of human platelets. This inhibitory activity was generated within 2-5 min, and it was not formed by cells that were treated with indomethacin or tranylcypromine. Addition of prostaglandin (PG)H2 to indomethacin-treated cells restored the ability to form the inhibitor, but cells treated with tranylcypromine were not responsive to PGH2. In experiments where [14C]arachidonic acid was added to the cells before stimulation, the major metabolite identified by thin-layer chromatography was 6-keto PGF1α. Thus, it appears that pulmonary endothelial cells, as well as umbilical cord cells, can form prostacyclin. In experiments comparing the ability of arterial and venous cells to form prostacyclin, arterial cells were more active than venous cells. These studies of cells from various human vessels suggest that the vascular origin of cultured endothelial cells determines how they metabolize vasoactive

  12. Novel markers of endothelial dysfunction and inflammation in Behçet's disease patients with ocular involvement: epicardial fat thickness, carotid intima media thickness, serum ADMA level, and neutrophil-to-lymphocyte ratio.

    PubMed

    Yuksel, Murat; Yildiz, Abdulkadir; Oylumlu, Mustafa; Turkcu, Fatih Mehmet; Bilik, Mehmet Zihni; Ekinci, Aysun; Elbey, Bilal; Tekbas, Ebru; Alan, Sait

    2016-03-01

    The etiology of Behçet's disease (BD) has not been fully elucidated. However, immunological and environmental factors, endothelial dysfunction (ED), and genetic susceptibility have been proposed to play a role. In this study, we aimed to evaluate epicardial fat thickness (EFT) together with serum asymmetric dimethylarginine (ADMA), carotid intima media thickness (CIMT), and neutrophil-to-lymphocyte ratio (NLR) in BD patients with ocular involvement. Thirty-six ocular BD patients (17 active and 19 inactive ocular involvement), and 35 age and sex-matched healthy controls were enrolled to this cross-sectional study. All patients underwent examinations with transthoracic echocardiography and carotid Doppler ultrasound. Serum ADMA levels, CIMT, EFT, and NLR were compared between groups, and their association with disease activity was evaluated. Behçet's disease patients had higher WBC counts, neutrophil counts, NLR, CIMT, EFT values, and serum ADMA levels than do healthy controls. The other biochemical, hematological, and echocardiographic parameters were comparable between the two groups. Behçet's disease duration was positively correlated with EFT and CIMT. Multivariate logistic regression analysis revealed that increased serum ADMA concentration and CIMT are independently associated with BD. Neutrophil counts, NLR, and serum ADMA level were higher, and lymphocyte count was lower in patients with active ocular BD compared to those of inactive ocular BD group. Carotid intima media thickness, serum ADMA level, EFT, and NLR were increased in ocular BD patients compared to healthy subjects. In addition, both serum ADMA level and NLR were associated with disease activity of ocular involvement. Increase in disease duration was associated with increase in CIMT and EFT which suggests that anatomical changes occur in time during the disease course. Increased CIMT, serum ADMA level, EFT, and NLR may provide new clues about the role of ED and inflammation in the

  13. Increased Detection of Lymphatic Vessel Invasion by D2-40 (Podoplanin) in Early Breast Cancer: Possible Influence on Patient Selection for Accelerated Partial Breast Irradiation

    SciTech Connect

    Debald, Manuel; Poelcher, Martin; Flucke, Uta; Walgenbach-Bruenagel, Gisela

    2010-07-15

    Purpose: Several international trials are currently investigating accelerated partial breast irradiation (APBI) for patients with early-stage breast cancer. According to existing guidelines, patients with lymphatic vessel invasion (LVI) do not qualify for APBI. D2-40 (podoplanin) significantly increases the frequency of LVI detection compared with conventional hematoxylin and eosin (HE) staining in early-stage breast cancer. Our purpose was to retrospectively assess the hypothetical change in management from APBI to whole breast radiotherapy with the application of D2-40. Patients and Methods: Immunostaining with D2-40 was performed on 254 invasive breast tumors of 247 patients. The following criteria were used to determine the eligibility for APBI: invasive ductal adenocarcinoma of {<=}3 cm, negative axillary node status (N0), and unifocal disease. Of the 247 patients, 74 with available information concerning LVI, as detected by D2-40 immunostaining and routine HE staining, formed our study population. Results: Using D2-40, our results demonstrated a significantly greater detection rate (p = .031) of LVI compared with routine HE staining. LVI was correctly identified by D2-40 (D2-40-positive LVI) in 10 (13.5%) of 74 tumors. On routine HE staining, 4 tumors (5.4%) were classified as HE-positive LVI. Doublestaining of these specimens with D2-40 unmasked false-positive LVI status in 2 (50%) of the 4 tumors. According to the current recommendations for APBI, immunostaining with D2-40 would have changed the clinical management from APBI to whole breast radiotherapy in 8 (10.8%) of 74 patients and from whole breast radiotherapy to APBI in 2 patients (2.7%). Conclusion: These data support the implementation of D2-40 immunostaining in the routine workup to determine a patient's eligibility for APBI.

  14. The chimeric antibody chLpMab-7 targeting human podoplanin suppresses pulmonary metastasis via ADCC and CDC rather than via its neutralizing activity

    PubMed Central

    Ogasawara, Satoshi; Fujii, Yuki; Oki, Hiroharu; Fukayama, Masashi; Nishioka, Yasuhiko; Kaneko, Mika K.

    2015-01-01

    Podoplanin (PDPN/Aggrus/T1α) binds to C-type lectin-like receptor-2 (CLEC-2) and induces platelet aggregation. PDPN is associated with malignant progression, tumor metastasis, and poor prognosis in several types of cancer. Although many anti-human PDPN (hPDPN) monoclonal antibodies (mAbs), such as D2-40 and NZ-1, have been established, these epitopes are limited to the platelet aggregation-stimulating (PLAG) domain (amino acids 29-54) of hPDPN. Recently, we developed a novel mouse anti-hPDPN mAb, LpMab-7, which is more sensitive than D2-40 and NZ-1, using the Cancer-specific mAb (CasMab) method. The epitope of LpMab-7 was shown to be entirely different from that of NZ-1, a neutralizing mAb against the PLAG domain according to an inhibition assay and lectin microarray analysis. In the present study, we produced a mouse-human chimeric anti-hPDPN mAb, chLpMab-7. ChLpMab-7 showed high antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). Furthermore, chLpMab-7 inhibited the growth of hPDPN-expressing tumors in vivo. Although chLpMab-7 recognizes a non-PLAG domain of hPDPN, it suppressed the hematogenous metastasis of hPDPN-expressing tumors. These results indicated that chLpMab-7 suppressed tumor development and hematogenous metastasis in a neutralization-independent manner. In conclusion, hPDPN shows promise as a target in the development of a novel antibody-based therapy. PMID:26416352

  15. Chronic Inflammatory Diseases and Endothelial Dysfunction

    PubMed Central

    Castellon, Xavier; Bogdanova, Vera

    2016-01-01

    Chronic inflammatory diseases are associated with increases in cardiovascular diseases (CVD) and subclinical atherosclerosis as well as early-stage endothelial dysfunction screening using the FMD method (Flow Mediated Dilation). This phenomenon, referred to as accelerated pathological remodeling of arterial wall, could be attributed to traditional risk factors associated with atherosclerosis. Several new non-invasive techniques have been used to study arterial wall’s structural and functional alterations. These techniques (based of Radio Frequency, RF) allow for an assessment of artery age through calculations of intima-media thickness (RF- QIMT), pulse wave rate (RF- QAS) and endothelial dysfunction degree (FMD). The inflammatory and autoimmune diseases should now be considered as new cardiovascular risk factors, result of the major consequences of oxidative stress and RAS (Renin Angiotensin System) imbalance associated with the deleterious effect of known risk factors that lead to the alteration of the arterial wall. Inflammation plays a key role in all stages of the formation of vascular lesions maintained and exacerbated by the risk factors. The consequence of chronic inflammation is endothelial dysfunction that sets in and we can define it as an integrated marker of the damage to arterial walls by classic risk factors. The atherosclerosis, which develops among these patients, is the main cause for cardiovascular morbi-mortality and uncontrolled chronic biological inflammation, which quickly favors endothelial dysfunction. These inflammatory and autoimmune diseases should now be considered as new cardiovascular risk factors. PMID:26815098

  16. Assessment of Endothelial Dysfunction in Childhood Obesity and Clinical Use

    PubMed Central

    Hoymans, Vicky Y.; Van Craenenbroeck, Amaryllis H.; Vissers, Dirk K.; Vrints, Christiaan J.; Conraads, Viviane M.

    2013-01-01

    The association of obesity with noncommunicable diseases, such as cardiovascular complications and diabetes, is considered a major threat to the management of health care worldwide. Epidemiological findings show that childhood obesity is rapidly rising in Western society, as well as in developing countries. This pandemic is not without consequences and can affect the risk of future cardiovascular disease in these children. Childhood obesity is associated with endothelial dysfunction, the first yet still reversible step towards atherosclerosis. Advanced research techniques have added further insight on how childhood obesity and associated comorbidities lead to endothelial dysfunction. Techniques used to measure endothelial function were further brought to perfection, and novel biomarkers, including endothelial progenitor cells, were discovered. The aim of this paper is to provide a critical overview on both in vivo as well as in vitro markers for endothelial integrity. Additionally, an in-depth description of the mechanisms that disrupt the delicate balance between endothelial damage and repair will be given. Finally, the effects of lifestyle interventions and pharmacotherapy on endothelial dysfunction will be reviewed. PMID:23691262

  17. Endothelial cells derived from human embryonic stem cells

    NASA Astrophysics Data System (ADS)

    Levenberg, Shulamit; Golub, Justin S.; Amit, Michal; Itskovitz-Eldor, Joseph; Langer, Robert

    2002-04-01

    Human embryonic stem cells have the potential to differentiate into various cell types and, thus, may be useful as a source of cells for transplantation or tissue engineering. We describe here the differentiation steps of human embryonic stem cells into endothelial cells forming vascular-like structures. The human embryonic-derived endothelial cells were isolated by using platelet endothelial cell-adhesion molecule-1 (PECAM1) antibodies, their behavior was characterized in vitro and in vivo, and their potential in tissue engineering was examined. We show that the isolated embryonic PECAM1+ cells, grown in culture, display characteristics similar to vessel endothelium. The cells express endothelial cell markers in a pattern similar to human umbilical vein endothelial cells, their junctions are correctly organized, and they have high metabolism of acetylated low-density lipoprotein. In addition, the cells are able to differentiate and form tube-like structures when cultured on matrigel. In vivo, when transplanted into SCID mice, the cells appeared to form microvessels containing mouse blood cells. With further studies, these cells could provide a source of human endothelial cells that could be beneficial for potential applications such as engineering new blood vessels, endothelial cell transplantation into the heart for myocardial regeneration, and induction of angiogenesis for treatment of regional ischemia.

  18. Endothelin and endothelial dysfunction.

    PubMed

    Masaki, Tomoh; Sawamura, Tatsuya

    2006-03-01

    Nitric oxide (NO) and endothelin (ET) produced in endothelial cells are leading molecules which regulate vascular function. Failure of the physiological balance between these two molecules is usually referred to as endothelial dysfunction. ET was initially identified as a potent vasoconstrictive peptide. Three ET isoforms and two ET receptors have been identified. One of the isoforms, ET-1, plays a significant role in many cardiovascular diseases. On the other hand, oxidized low-density lipoprotein (oxLDL) is known to induce endothelial dysfunction. The endothelial receptor for oxLDL was cloned, and named lectin-like oxidized receptor-1 (LOX-1). Activation of LOX-1 generates reactive oxygen species (ROS), and acivates a transcriptional factor, nuclear factor κB (NFκB), resulting in down-regulation of NO and up-regulation of ET-1. LOX-1 might be a key molecule in the generation of endothelial dysfunction. In endothelial dysfunction, ET-1 is an aggravating factor of cardiovascular diseases. PMID:25792766

  19. Evolution of endothelial keratoplasty.

    PubMed

    Price, Francis W; Price, Marianne O

    2013-11-01

    Endothelial keratoplasty has evolved into a popular alternative to penetrating keratoplasty (PK) for the treatment of endothelial dysfunction. Although the earliest iterations were challenging and were not widely adopted, the iteration known as Descemet stripping endothelial keratoplasty (DSEK) has gained widespread acceptance. DSEK combines a simplified technique for stripping dysfunctional endothelium from the host cornea and microkeratome dissection of the donor tissue, a step now commonly completed in advance by eye bank technicians. Studies show that a newer endothelial keratoplasty iteration, known as Descemet membrane endothelial keratoplasty (DMEK), provides an even faster and better visual recovery than DSEK does. In addition, DMEK significantly reduces the risk of immunologic graft rejection episodes compared with that in DSEK or in PK. Although the DMEK donor tissue, consisting of the bare endothelium and Descemet membrane without any stroma, is more challenging to prepare and position in the recipient eye, recent improvements in instrumentation and surgical techniques are increasing the ease and the reliability of the procedure. DSEK successfully mitigates 2 of the main liabilities of PK: ocular surface complications and structural problems (including induced astigmatism and perpetually weak wounds), whereas DMEK further mitigates the 2 principal remaining liabilities of PK: immunologic graft reactions and secondary glaucoma from prolonged topical corticosteroid use. PMID:24104929

  20. PDGFRβ Is a Novel Marker of Stromal Activation in Oral Squamous Cell Carcinomas

    PubMed Central

    Han, Rong; Haines, Paul; Gallagher, George; Noonan, Vikki; Kukuruzinska, Maria; Monti, Stefano; Trojanowska, Maria

    2016-01-01

    Carcinoma associated fibroblasts (CAFs) form the main constituents of tumor stroma and play an important role in tumor growth and invasion. The presence of CAFs is a strong predictor of poor prognosis of head and neck squamous cell carcinoma. Despite significant progress in determining the role of CAFs in tumor progression, the mechanisms contributing to their activation remain poorly characterized, in part due to fibroblast heterogeneity and the scarcity of reliable fibroblast surface markers. To search for such markers in oral squamous cell carcinoma (OSCC), we applied a novel approach that uses RNA-sequencing data derived from the cancer genome atlas (TCGA). Specifically, our strategy allowed for an unbiased identification of genes whose expression was closely associated with a set of bona fide stroma-specific transcripts, namely the interstitial collagens COL1A1, COL1A2, and COL3A1. Among the top hits were genes involved in cellular matrix remodeling and tumor invasion and migration, including platelet-derived growth factor receptor beta (PDGFRβ), which was found to be the highest-ranking receptor protein genome-wide. Similar analyses performed on ten additional TCGA cancer datasets revealed that other tumor types shared CAF markers with OSCC, including PDGFRβ, which was found to significantly correlate with the reference collagen expression in ten of the 11 cancer types tested. Subsequent immunostaining of OSCC specimens demonstrated that PDGFRβ was abundantly expressed in stromal fibroblasts of all tested cases (12/12), while it was absent in tumor cells, with greater specificity than other known markers such as alpha smooth muscle actin or podoplanin (3/11). Overall, this study identified PDGFRβ as a novel marker of stromal activation in OSCC, and further characterized a list of promising candidate CAF markers that may be relevant to other carcinomas. Our novel approach provides for a fast and accurate method to identify CAF markers without the need for

  1. PDGFRβ Is a Novel Marker of Stromal Activation in Oral Squamous Cell Carcinomas.

    PubMed

    Kartha, Vinay K; Stawski, Lukasz; Han, Rong; Haines, Paul; Gallagher, George; Noonan, Vikki; Kukuruzinska, Maria; Monti, Stefano; Trojanowska, Maria

    2016-01-01

    Carcinoma associated fibroblasts (CAFs) form the main constituents of tumor stroma and play an important role in tumor growth and invasion. The presence of CAFs is a strong predictor of poor prognosis of head and neck squamous cell carcinoma. Despite significant progress in determining the role of CAFs in tumor progression, the mechanisms contributing to their activation remain poorly characterized, in part due to fibroblast heterogeneity and the scarcity of reliable fibroblast surface markers. To search for such markers in oral squamous cell carcinoma (OSCC), we applied a novel approach that uses RNA-sequencing data derived from the cancer genome atlas (TCGA). Specifically, our strategy allowed for an unbiased identification of genes whose expression was closely associated with a set of bona fide stroma-specific transcripts, namely the interstitial collagens COL1A1, COL1A2, and COL3A1. Among the top hits were genes involved in cellular matrix remodeling and tumor invasion and migration, including platelet-derived growth factor receptor beta (PDGFRβ), which was found to be the highest-ranking receptor protein genome-wide. Similar analyses performed on ten additional TCGA cancer datasets revealed that other tumor types shared CAF markers with OSCC, including PDGFRβ, which was found to significantly correlate with the reference collagen expression in ten of the 11 cancer types tested. Subsequent immunostaining of OSCC specimens demonstrated that PDGFRβ was abundantly expressed in stromal fibroblasts of all tested cases (12/12), while it was absent in tumor cells, with greater specificity than other known markers such as alpha smooth muscle actin or podoplanin (3/11). Overall, this study identified PDGFRβ as a novel marker of stromal activation in OSCC, and further characterized a list of promising candidate CAF markers that may be relevant to other carcinomas. Our novel approach provides for a fast and accurate method to identify CAF markers without the need for

  2. Endothelial activation, lymphangiogenesis, and humoral rejection of kidney transplants.

    PubMed

    Phillips, Sharon; Kapp, Meghan; Crowe, Deborah; Garces, Jorge; Fogo, Agnes B; Giannico, Giovanna A

    2016-05-01

    Antibody-mediated rejection (ABMR) is implicated in 45% of renal allograft failure and 57% of late allograft dysfunction. Peritubular capillary C4d is a specific but insensitive marker of ABMR. The 2013 Banff Conference ABMR revised criteria included C4d-negative ABMR with evidence of endothelial-antibody interaction. We hypothesized that endothelial activation and lymphangiogenesis are increased with C4d-negative ABMR and correlate with intragraft T-regulatory cells and T-helper 17. Seventy-four renal transplant biopsies were selected to include (a) ABMR with C4d Banff scores ≥2 (n = 35), (b) variable microvascular injury and C4d score 0-1 (n = 24), and (c) variable microvascular injury and C4d score = 0 (n = 15). Controls included normal preimplantation donor kidneys (n = 5). Immunohistochemistry for endothelial activation (P- and E-selectins [SEL]), lymphangiogenesis (D2-40), T-regulatory cells (FOXP3), and T-helper 17 (STAT3) was performed. Microvessel and inflammatory infiltrate density was assessed morphometrically in interstitium and peritubular capillaries. All transplants had significantly higher microvessel and lymph vessel density compared with normal. Increased expression of markers of endothelial activation predicted transplant glomerulopathy (P-SEL, P = .003). Increased P-SEL and D2-40 were associated with longer interval from transplant to biopsy (P = .005). All 3 markers were associated with increased interstitial fibrosis, tubular atrophy, and graft failure (P-SEL, P < .001; E-SEL, P = .0011; D2-40, P = .012). There was no association with the intragraft FOXP3/STAT3 ratio. We conclude that endothelial activation and lymphangiogenesis could represent a late response to injury leading to fibrosis and progression of kidney damage, and are independent of the intragraft FOXP3/STAT3 ratio. Our findings support the therapeutic potential of specifically targeting endothelial activation. PMID:27067786

  3. Mechanisms regulating endothelial permeability

    PubMed Central

    Sukriti, Sukriti; Tauseef, Mohammad; Yazbeck, Pascal

    2014-01-01

    Abstract The endothelial monolayer partitioning underlying tissue from blood components in the vessel wall maintains tissue fluid balance and host defense through dynamically opening intercellular junctions. Edemagenic agonists disrupt endothelial barrier function by signaling the opening of the intercellular junctions leading to the formation of protein-rich edema in the interstitial tissue, a hallmark of tissue inflammation that, if left untreated, causes fatal diseases, such as acute respiratory distress syndrome. In this review, we discuss how intercellular junctions are maintained under normal conditions and after stimulation of endothelium with edemagenic agonists. We have focused on reviewing the new concepts dealing with the alteration of adherens junctions after inflammatory stimulus. PMID:25610592

  4. Expression and endocytosis of VEGF and its receptors in human colonic vascular endothelial cells.

    PubMed

    Wang, Dongfang; Lehman, Richard E; Donner, David B; Matli, Mary R; Warren, Robert S; Welton, Mark L

    2002-06-01

    Normal human colonic microvascular endothelial cells (HUCMEC) have been isolated from surgical specimens by their adherence to Ulex europaeus agglutinin bound to magnetic dynabeads that bind alpha-L-fucosyl residues on the endothelial cell membrane. Immunocytochemistry demonstrated the presence of a range of endothelial-specific markers on HUCMEC, including the von Willebrand factor, Ulex europaeus agglutinin, and platelet endothelial cell adhesion molecule-1. The growing cells form monolayers with the characteristic cobblestone morphology of endothelial cells and eventually form tube-like structures. HUCMEC produce vascular endothelial growth factor (VEGF) and express the receptors, kinase insert domain-containing receptor (KDR) and fms-like tyrosine kinase, through which VEGF mediates its actions in the endothelium. VEGF induces the tyrosine phosphorylation of KDR and a proliferative response from HUCMEC comparable to that elicited from human umbilical vein endothelial cells (HUVEC). On binding to HUCMEC or HUVEC, (125)I-labeled VEGF internalizes or dissociates to the medium. Once internalized, (125)I-labeled VEGF is degraded and no evidence of ligand recycling was observed. However, significantly less VEGF is internalized, and more is released to the medium from HUCMEC than HUVEC. Angiogenesis results from the proliferation and migration of microvascular, not large-vessel, endothelial cells. The demonstration that microvascular endothelial cells degrade less and release more VEGF to the medium than large-vessel endothelial cells identifies a mechanism permissive of the role of microvascular cells in angiogenesis. PMID:12016135

  5. Tumor-derived circulating endothelial cell clusters in colorectal cancer.

    PubMed

    Cima, Igor; Kong, Say Li; Sengupta, Debarka; Tan, Iain B; Phyo, Wai Min; Lee, Daniel; Hu, Min; Iliescu, Ciprian; Alexander, Irina; Goh, Wei Lin; Rahmani, Mehran; Suhaimi, Nur-Afidah Mohamed; Vo, Jess H; Tai, Joyce A; Tan, Joanna H; Chua, Clarinda; Ten, Rachel; Lim, Wan Jun; Chew, Min Hoe; Hauser, Charlotte A E; van Dam, Rob M; Lim, Wei-Yen; Prabhakar, Shyam; Lim, Bing; Koh, Poh Koon; Robson, Paul; Ying, Jackie Y; Hillmer, Axel M; Tan, Min-Han

    2016-06-29

    Clusters of tumor cells are often observed in the blood of cancer patients. These structures have been described as malignant entities for more than 50 years, although their comprehensive characterization is lacking. Contrary to current consensus, we demonstrate that a discrete population of circulating cell clusters isolated from the blood of colorectal cancer patients are not cancerous but consist of tumor-derived endothelial cells. These clusters express both epithelial and mesenchymal markers, consistent with previous reports on circulating tumor cell (CTC) phenotyping. However, unlike CTCs, they do not mirror the genetic variations of matched tumors. Transcriptomic analysis of single clusters revealed that these structures exhibit an endothelial phenotype and can be traced back to the tumor endothelium. Further results show that tumor-derived endothelial clusters do not form by coagulation or by outgrowth of single circulating endothelial cells, supporting a direct release of clusters from the tumor vasculature. The isolation and enumeration of these benign clusters distinguished healthy volunteers from treatment-naïve as well as pathological early-stage (≤IIA) colorectal cancer patients with high accuracy, suggesting that tumor-derived circulating endothelial cell clusters could be used as a means of noninvasive screening for colorectal cancer. In contrast to CTCs, tumor-derived endothelial cell clusters may also provide important information about the underlying tumor vasculature at the time of diagnosis, during treatment, and throughout the course of the disease. PMID:27358499

  6. Asymmetric Dimethylarginine, Endothelial Dysfunction and Renal Disease

    PubMed Central

    Aldámiz-Echevarría, Luis; Andrade, Fernando

    2012-01-01

    l-Arginine (Arg) is oxidized to l-citrulline and nitric oxide (NO) by the action of endothelial nitric oxide synthase (NOS). In contrast, protein-incorporated Arg residues can be methylated with subsequent proteolysis giving rise to methylarginine compounds, such as asymmetric dimethylarginine (ADMA) that competes with Arg for binding to NOS. Most ADMA is degraded by dimethylarginine dimethyaminohydrolase (DDAH), distributed widely throughout the body and regulates ADMA levels and, therefore, NO synthesis. In recent years, several studies have suggested that increased ADMA levels are a marker of atherosclerotic change, and can be used to assess cardiovascular risk, consistent with ADMA being predominantly absorbed by endothelial cells. NO is an important messenger molecule involved in numerous biological processes, and its activity is essential to understand both pathogenic and therapeutic mechanisms in kidney disease and renal transplantation. NO production is reduced in renal patients because of their elevated ADMA levels with associated reduced DDAH activity. These factors contribute to endothelial dysfunction, oxidative stress and the progression of renal damage, but there are treatments that may effectively reduce ADMA levels in patients with kidney disease. Available data on ADMA levels in controls and renal patients, both in adults and children, also are summarized in this review. PMID:23109853

  7. Endothelial microparticles carrying hedgehog-interacting protein induce continuous endothelial damage in the pathogenesis of acute graft-versus-host disease.

    PubMed

    Nie, Di-Min; Wu, Qiu-Ling; Zheng, Peng; Chen, Ping; Zhang, Ran; Li, Bei-Bei; Fang, Jun; Xia, Ling-Hui; Hong, Mei

    2016-05-15

    Accumulating evidence suggests that endothelial microparticles (EMPs), a marker of endothelial damage, are elevated in acute graft-versus-host disease (aGVHD), and that endothelial damage is implicated in the pathogenesis of aGVHD, but the mechanisms remain elusive. In this study, we detected the plasma EMP levels and endothelial damage in patients and mice with aGVHD in vivo and then examined the effects of EMPs derived from injured endothelial cells (ECs) on endothelial damage and the role of hedgehog-interacting protein (HHIP) carried by EMPs in these effects in vitro. Our results showed that EMPs were persistently increased in the early posttransplantation phase in patients and mice with aGVHD. Meanwhile, endothelial damage was continuous in aGVHD mice, but was temporary in non-aGVHD mice after transplantation. In vitro, EMPs induced endothelial damage, including increased EC apoptosis, enhanced reactive oxygen species, decreased nitric oxide production and impaired angiogenic activity. Enhanced expression of HHIP, an antagonist for the Sonic hedgehog (SHH) signaling pathway, was observed in patients and mice with aGVHD and EMPs from injured ECs. The endothelial damage induced by EMPs was reversed when the HHIP incorporated into EMPs was silenced with an HHIP small interfering RNA or inhibited with the SHH pathway agonist, Smoothened agonist. This work supports a feasible vicious cycle in which EMPs generated during endothelial injury, in turn, aggravate endothelial damage by carrying HHIP into target ECs, contributing to the continuously deteriorating endothelial damage in the development of aGVHD. EMPs harboring HHIP would represent a potential therapeutic target for aGVHD. PMID:27009877

  8. CD133 positive progenitor endothelial cell lines from human cord blood.

    PubMed

    Paprocka, Maria; Krawczenko, Agnieszka; Dus, Danuta; Kantor, Aneta; Carreau, Aude; Grillon, Catherine; Kieda, Claudine

    2011-08-01

    Endothelial progenitor cells (EPCs) modulate postnatal vascularization and contribute to vessel regeneration in adults. Stem cells and progenitor cells were found in umbilical cord blood, bone marrow, and mobilized peripheral blood cells, from where they were isolated and cultured. However, the yield of progenitor cells is usually not sufficient for clinical application and the quality of progenitor cells varies. The aim of the study was the immortalization of early progenitor cells with high proliferative potential, capable to differentiate to EPCs and, further, toward endothelial cells. Two cell lines, namely HEPC-CB.1 and HEPC-CB.2 (human endothelial progenitor cells-cord blood) were isolated. As assessed by specific antibody labeling and flow cytometric analysis, they express a panel of stem cell markers: CD133, CD13, CD271, CD90 and also endothelial cell markers: CD202b, CD309 (VEGFR2), CD146, CD105, and CD143 but they do not present markers of finally differentiated endothelial cells: CD31, vWf, nor CD45 which is a specific hematopoietic cell marker. Using the multiplex Cytometric Bead Assay, the simultaneous production of proangiogenic cytokines IL8, angiogenin, and VEGF was demonstrated in normoxia and was shown to be increased by hypoxia. Both cell lines, similarly as mature endothelial cells, underwent in vitro pre-angiogenic process, formed pseudovessel structures and present an accelerated angiogenesis in hypoxic conditions. To date, these are the first CD133 positive established cell lines from human cord blood cells. PMID:21710642

  9. Wine and endothelial function.

    PubMed

    Caimi, G; Carollo, C; Lo Presti, R

    2003-01-01

    In recent years many studies have focused on the well-known relationship between wine consumption and cardiovascular risk. Wine exerts its protective effects through various changes in lipoprotein profile, coagulation and fibrinolytic cascades, platelet aggregation, oxidative mechanisms and endothelial function. The last has earned more attention for its implications in atherogenesis. Endothelium regulates vascular tone by a delicate balancing among vasorelaxing (nitric oxide [NO]) and vasoconstrincting (endothelins) factors produced by endothelium in response to various stimuli. In rat models, wine and other grape derivatives exerted an endothelium-dependent vasorelaxing capacity especially associated with the NO-stimulating activity of their polyphenol components. In experimental conditions, reservatrol (a stilbene polyphenol) protected hearts and kidneys from ischemia-reperfusion injury through antioxidant activity and upregulation of NO production. Wine polyphenols are also able to induce the expression of genes involved in the NO pathway within the arterial wall. The effects of wine on endothelial function in humans are not yet clearly understood. A favorable action of red wine or dealcoholized wine extract or purple grape juice on endothelial function has been observed by several authors, but discrimination between ethanol and polyphenol effects is controversial. It is, however likely that regular and prolonged moderate wine drinking positively affects endothelial function. The beneficial effects of wine on cardiovascular health are greater if wine is associated with a healthy diet. The most recent nutritional and epidemiologic studies show that the ideal diet closely resembles the Mediterranean diet. PMID:15134380

  10. In Vivo Vascularization of Endothelial Cells Derived from Bone Marrow Mesenchymal Stem Cells in SCID Mouse Model

    PubMed Central

    Allameh, Abdolamir; Jazayeri, Maryam; Adelipour, Maryam

    2016-01-01

    Objective In vivo and in vitro stem cell differentiation into endothelial cells is a promising area of research for tissue engineering and cell therapy. Materials and Methods We induced human mesenchymal stem cells (MSCs) to differentiate to endothelial cells that had the ability to form capillaries on an extracellular matrix (ECM) gel. Thereafter, the differentiated endothelial cells at early stage were characterized by expression of specific markers such as von Willebrand factor (vWF), vascular endothelial growth factor (VEGF) receptor 2, and CD31. In this experimental model, the endothelial cells were transplanted into the groins of severe combined immunodeficiency (SCID) mice. After 30 days, we obtained tissue biopsies from the transplantation sites. Biopsies were processed for histopathological and double immunohistochemistry (DIHC) staining. Results Endothelial cells at the early stage of differentiation expressed endothelial markers. Hematoxylin and eosin (H&E) staining, in addition to DIHC demonstrated homing of the endothelial cells that underwent vascularization in the injected site. Conclusion The data clearly showed that endothelial cells at the early stage of differentiation underwent neovascularization in vivo in SCID mice. Endothelial cells at their early stage of differentiation have been proven to be efficient for treatment of diseases with impaired vasculogenesis. PMID:27540522

  11. [Transplantation of corneal endothelial cells].

    PubMed

    Amano, Shiro

    2002-12-01

    endothelial cells. Cultured rabbit corneal endothelial cells that endocytosed iron were injected into the anterior chamber of rabbits whose corneal endothelium was cryo-injured, and were pulled to Descemet's membrane by putting a magnet on the eyelid. In these rabbits, corneal edema decreased more quickly than in the control group and no intraocular pressure rise was observed during 8 weeks after the operation, suggesting that the direct delivery of cultured HCECs into the anterior chamber can be an alternative method of choice. The following obstacles should be addressed to make the transplantation of cultured corneal endothelial cells clinically applicable. 1. To reconstruct a cornea that is the same as or superior to the normal cornea, more innovation is necessary in the method of culturing and seeding HCECs. We should consider utilizing HCECs obtained from fetuses after clearing ethical issues. Moreover, we need to develop a method to enhance the cell density and the cell functions. 2. Porcine corneal stroma is promising as a carrier of HCECs instead of human corneal stroma, which is in very limited supply. The usefulness of porcine corneal stroma acellularized to prevent retrovirus infection should be evaluated. 3. To make the self immature cells applicable to corneal transplantation, we should elucidate the corneal endothelial cell specific markers and the factors that are necessary to induce self immature cells to become corneal endothelial cells. 4. The direct delivery of cultured HCECs into the anterior chamber can be an alternative method of choice when its long-term safety is confirmed. PMID:12610838

  12. Glycated serum albumin stimulates expression of endothelial cell specific molecule-1 in human umbilical vein endothelial cells: Implication in diabetes mediated endothelial dysfunction.

    PubMed

    Nirala, Bikesh K; Perumal, Vivekanandan; Gohil, Nivedita K

    2015-07-01

    Pro-inflammatory conditions induced by products of protein glycation in diabetes substantially enhance the risk of endothelial dysfunction and related vascular complications. Endothelial cell specific molecule-1 (ESM-1) or endocan has been demonstrated as a potential biomarker in cancer and sepsis. Its role in diabetes-induced pathologies remains unknown. The expression of ESM-1 gene is under cytokine regulation, indicating its role in endothelium-dependent pathological disorders. In this study, we investigated the effect of advanced glycated human serum albumin (AGE-HSA) on the production of ESM-1. We show that AGE-HSA exerts a modulating role on the expression of ESM-1 in human umbilical vein endothelial cells. It up-regulates expression of ESM-1 protein in a dose-dependent manner which correlates with its messenger RNA (mRNA) transcription. RAGE and galectin-3, both AGE receptors, show antagonistic action on its expression. While gene silencing of RAGE has down-regulatory effect, that of galectin-3 has up-regulatory effect on AGE-induced expression of ESM-1. Inhibition of MAPKKK and JNK pathways did not alter the expression. In contrast, phosphatidylinositol 3 kinase (PI3K) inhibition significantly up-regulated ESM-1 expression. In conclusion, these results suggest that AGE-induced activation of human umbilical vein endothelial cells promotes formation of endocan which is an endothelial dysfunction marker and may be related to vascular disease in diabetes. PMID:25963575

  13. Myofibroblasts in proliferative diabetic retinopathy can originate from infiltrating fibrocytes and through endothelial-to-mesenchymal transition (EndoMT).

    PubMed

    Abu El-Asrar, Ahmed M; De Hertogh, Gert; van den Eynde, Kathleen; Alam, Kaiser; Van Raemdonck, Katrien; Opdenakker, Ghislain; Van Damme, Jo; Geboes, Karel; Struyf, Sofie

    2015-03-01

    Myofibroblasts expressing α-smooth muscle actin (α-SMA) are the key cellular mediator of fibrosis. Fibrovascular epiretinal membranes from patients with proliferative diabetic retinopathy (PDR) are characterized by the accumulation of a large number of myofibroblasts. We explored the hypothesis that proliferating endothelial cells via endothelial-to-mesenchymal transition (EndoMT) and/or bone marrow-derived circulating fibrocytes contribute to the myofibroblast population present in PDR epiretinal membranes. Epiretinal membranes from 14 patients with PDR were studied by immunohistochemistry. All membranes contained neovessels expressing the endothelial cell marker CD31. CD31(+) endothelial cells co-expressed the fibroblast/myofibroblast markers fibroblast-specific protein-1 (FSP-1) and α-SMA, indicative for the occurrence of endoMT. In the stroma, cells expressing FSP-1, α-SMA, the leukocyte common antigen CD45, and the myelomonocytic marker CD11b were detected. Double labeling showed co-localization of CD45 with FSP-1 and α-SMA and co-localization of CD11b with α-SMA and matrix metalloproteinase-9, demonstrating the presence of infiltrating fibrocytes. In addition, we investigated the phenotypic changes that take place in human retinal microvascular endothelial cells following exposure to transforming growth factor-β1 (TGF-β1), connective tissue growth factor (CTGF) and the proinflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α). Retinal microvascular endothelial cells changed morphology upon cytokine exposure, lost the expression of endothelial cell markers (endothelial nitric oxide synthase and vascular endothelial-cadherin) and started to express mesenchymal markers (calponin, snail, transgelin and FSP-1). These results suggest that endothelial cells as well as circulating fibrocytes may differentiate into myofibroblasts in the diabetic eye and contribute to pathologic fibrosis in PDR. PMID:25637870

  14. Effect of Polyelectrolyte Film Stiffness on Endothelial Cells During Endothelial-to-Mesenchymal Transition.

    PubMed

    Zhang, He; Chang, Hao; Wang, Li-mei; Ren, Ke-feng; Martins, M Cristina L; Barbosa, Mário A; Ji, Jian

    2015-11-01

    Endothelial-to-mesenchymal transition (EndMT), during which endothelial cells (ECs) transdifferentiate into mesenchymal phenotype, plays a key role in the development of vascular implant complications such as endothelium dysfunction and in-stent restenosis. Substrate stiffness has been confirmed as a key factor to influence EC behaviors; however, so far, the relationship between substrate stiffness and EndMT has been rarely studied. Here, ECs were cultured on the (poly(L-lysine)/hyaluronate acid) (PLL/HA) multilayer films with controlled stiffness for 2 weeks, and their EndMT behaviors were studied. We demonstrated that ECs lost their markers (vWf and CD31) in a stiffness-dependent manner even without supplement of growth factors, and the softer film favored the maintaining of EC phenotype. Further, induced by transforming growth factor β1 (TGF-β1), ECs underwent EndMT, as characterized by losing their typical cobblestone morphology and markers and gaining smooth muscle cell markers (α-smooth muscle actin and calponin). Interestingly, stronger EndMT was observed when ECs were cultured on the stiffer film. Collectively, our findings suggest that substrate stiffness has significant effects on EndMT, and a softer substrate is beneficial to ECs by keeping their phenotype and inhibiting EndMT, which presents a new strategy for surface design of vascular implant materials. PMID:26477358

  15. (-)-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. PMID:26993496

  16. Endothelial dysfunction and antioxidants.

    PubMed

    Duvall, W Lane

    2005-03-01

    The vascular endothelium plays a crucial role in the physiology of blood vessels and the pathological processes of atherosclerotic disease and acute coronary syndromes. Endothelial dysfunction is the core problem; it is an impairment of endothelium-dependent vasorelaxation caused by a loss of nitric oxide activity in the vessel wall, which results in impairment in the regulation of vascular homeostasis. Further understanding of its mechanisms of action and possible therapeutic targets will be of great importance. The group of antioxidant vitamins, A, C and E, would seem uniquely situated to reduce cardiovascular events by improving endothelial function by reducing the concentration of reactive oxygen species in the vessel wall and by preventing oxidative modification of low-density lipoprotein. Unfortunately, despite extensive studies in both observational and randomized trials, the weight of evidence points to little or no benefit from antioxidant therapy. PMID:15770336

  17. Prognostic value of endothelial dysfunction in type 1 diabetes mellitus

    PubMed Central

    Ladeia, Ana Marice; Sampaio, Raphael Ribeiro; Hita, Maiara Costa; Adan, Luis F

    2014-01-01

    Patients with diabetes mellitus are at high risk of developing atherosclerosis, associated with higher rates of micro and macro vascular involvement such as coronary artery disease and renal disease. The role of hyperglycemia to induce synthesis of reactive oxygen species by the oxidation of glucose, leading to an increased production of advanced glycosylation end products, as well as inflammation and oxidative stress has been proposed as a possible mechanism in the pathogenesis of endothelial dysfunction (ED). The interaction between C-peptide - the connecting segment of pro-insulin-and nitric oxide in vasodilation is also discussed. Therefore, endothelial dysfunction has been identified as an early marker of vascular disorder in type 1 and type 2 diabetes mellitus. In some other diseases, ED has been considered an independent predictor of vascular disease, regardless of the method used. Studies have demonstrated the importance of endothelial dysfunction as an useful tool for identifying the risk of vascular complications in patients with type 1 diabetes mellitus, particularly as regards to renal impairment. The aim of this review is to clarify the prognostic value of endothelial dysfunction as a marker of vascular disease in these subjects. PMID:25317238

  18. In vitro differentiation of porcine aortic vascular precursor cells to endothelial and vascular smooth muscle cells.

    PubMed

    Zaniboni, Andrea; Bernardini, Chiara; Bertocchi, Martina; Zannoni, Augusta; Bianchi, Francesca; Avallone, Giancarlo; Mangano, Chiara; Sarli, Giuseppe; Calzà, Laura; Bacci, Maria Laura; Forni, Monica

    2015-09-01

    Recent findings suggest that progenitor and multipotent mesenchymal stromal cells (MSCs) are associated with vascular niches. Cells displaying mesenchymal properties and differentiating to whole components of a functional blood vessel, including endothelial and smooth muscle cells, can be defined as vascular stem cells (VSCs). Recently, we isolated a population of porcine aortic vascular precursor cells (pAVPCs), which have MSC- and pericyte-like properties. The aim of the present work was to investigate whether pAVPCs possess VSC-like properties and assess their differentiation potential toward endothelial and smooth muscle lineages. pAVPCs, maintained in a specific pericyte growth medium, were cultured in high-glucose DMEM + 10% FBS (long-term medium, LTM) or in human endothelial serum-free medium + 5% FBS and 50 ng/ml of hVEGF (endothelial differentiation medium, EDM). After 21 days of culture in LTM, pAVPCs showed an elongated fibroblast-like morphology, and they seem to organize in cord-like structures. qPCR analysis of smooth muscle markers [α-smooth muscle actin (α-SMA), calponin, and smooth muscle myosin (SMM) heavy chain] showed a significant increment of the transcripts, and immunofluorescence analysis confirmed the presence of α-SMA and SMM proteins. After 21 days of culture in EDM, pAVPCs displayed an endothelial cell-like morphology and revealed the upregulation of the expression of endothelial markers (CD31, vascular endothelial-cadherin, von Willebrand factor, and endothelial nitric oxide synthase) showing the CD31-typical pattern. In conclusion, pAVPCs could be defined as a VSC-like population considering that, if they are maintained in a specific pericyte medium, they express MSC markers, and they have, in addition to the classical mesenchymal trilineage differentiation potential, the capacity to differentiate in vitro toward the smooth muscle and the endothelial cell phenotypes. PMID:26135800

  19. Local and Circulating Endothelial Cells Undergo Endothelial to Mesenchymal Transition (EndMT) in Response to Musculoskeletal Injury.

    PubMed

    Agarwal, Shailesh; Loder, Shawn; Cholok, David; Peterson, Joshua; Li, John; Fireman, David; Breuler, Christopher; Hsieh, Hsiao Sung; Ranganathan, Kavitha; Hwang, Charles; Drake, James; Li, Shuli; Chan, Charles K; Longaker, Michael T; Levi, Benjamin

    2016-01-01

    Endothelial-to-mesenchymal transition (EndMT) has been implicated in a variety of aberrant wound healing conditions. However, unambiguous evidence of EndMT has been elusive due to limitations of in vitro experimental designs and animal models. In vitro experiments cannot account for the myriad ligands and cells which regulate differentiation, and in vivo tissue injury models may induce lineage-independent endothelial marker expression in mesenchymal cells. By using an inducible Cre model to mark mesenchymal cells (Scx-creERT/tdTomato + ) prior to injury, we demonstrate that musculoskeletal injury induces expression of CD31, VeCadherin, or Tie2 in mesenchymal cells. VeCadherin and Tie2 were expressed in non-endothelial cells (CD31-) present in marrow from uninjured adult mice, thereby limiting the specificity of these markers in inducible models (e.g. VeCadherin- or Tie2-creERT). However, cell transplantation assays confirmed that endothelial cells (ΔVeCadherin/CD31+/CD45-) isolated from uninjured hindlimb muscle tissue undergo in vivo EndMT when transplanted directly into the wound without intervening cell culture using PDGFRα, Osterix (OSX), SOX9, and Aggrecan (ACAN) as mesenchymal markers. These in vivo findings support EndMT in the presence of myriad ligands and cell types, using cell transplantation assays which can be applied for other pathologies implicated in EndMT including tissue fibrosis and atherosclerosis. Additionally, endothelial cell recruitment and trafficking are potential therapeutic targets to prevent EndMT. PMID:27616463

  20. Phenotypic and Functional Changes of Endothelial and Smooth Muscle Cells in Thoracic Aortic Aneurysms

    PubMed Central

    Malashicheva, Anna; Kostina, Daria; Kostina, Aleksandra; Irtyuga, Olga; Voronkina, Irina; Smagina, Larisa; Ignatieva, Elena; Gavriliuk, Natalia; Uspensky, Vladimir; Moiseeva, Olga; Vaage, Jarle; Kostareva, Anna

    2016-01-01

    Thoracic aortic aneurysm develops as a result of complex series of events that alter the cellular structure and the composition of the extracellular matrix of the aortic wall. The purpose of the present work was to study the cellular functions of endothelial and smooth muscle cells from the patients with aneurysms of the thoracic aorta. We studied endothelial and smooth muscle cells from aneurysms in patients with bicuspid aortic valve and with tricuspid aortic valve. The expression of key markers of endothelial (CD31, vWF, and VE-cadherin) and smooth muscle (SMA, SM22α, calponin, and vimentin) cells as well extracellular matrix and MMP activity was studied as well as and apoptosis and cell proliferation. Expression of functional markers of endothelial and smooth muscle cells was reduced in patient cells. Cellular proliferation, migration, and synthesis of extracellular matrix proteins are attenuated in the cells of the patients. We show for the first time that aortic endothelial cell phenotype is changed in the thoracic aortic aneurysms compared to normal aortic wall. In conclusion both endothelial and smooth muscle cells from aneurysms of the ascending aorta have downregulated specific cellular markers and altered functional properties, such as growth rate, apoptosis induction, and extracellular matrix synthesis. PMID:26904289

  1. Phenotypic and Functional Changes of Endothelial and Smooth Muscle Cells in Thoracic Aortic Aneurysms.

    PubMed

    Malashicheva, Anna; Kostina, Daria; Kostina, Aleksandra; Irtyuga, Olga; Voronkina, Irina; Smagina, Larisa; Ignatieva, Elena; Gavriliuk, Natalia; Uspensky, Vladimir; Moiseeva, Olga; Vaage, Jarle; Kostareva, Anna

    2016-01-01

    Thoracic aortic aneurysm develops as a result of complex series of events that alter the cellular structure and the composition of the extracellular matrix of the aortic wall. The purpose of the present work was to study the cellular functions of endothelial and smooth muscle cells from the patients with aneurysms of the thoracic aorta. We studied endothelial and smooth muscle cells from aneurysms in patients with bicuspid aortic valve and with tricuspid aortic valve. The expression of key markers of endothelial (CD31, vWF, and VE-cadherin) and smooth muscle (SMA, SM22α, calponin, and vimentin) cells as well extracellular matrix and MMP activity was studied as well as and apoptosis and cell proliferation. Expression of functional markers of endothelial and smooth muscle cells was reduced in patient cells. Cellular proliferation, migration, and synthesis of extracellular matrix proteins are attenuated in the cells of the patients. We show for the first time that aortic endothelial cell phenotype is changed in the thoracic aortic aneurysms compared to normal aortic wall. In conclusion both endothelial and smooth muscle cells from aneurysms of the ascending aorta have downregulated specific cellular markers and altered functional properties, such as growth rate, apoptosis induction, and extracellular matrix synthesis. PMID:26904289

  2. Endothelial dysfunction and tendinopathy: how far have we come?

    PubMed

    Papalia, R; Moro, L; Franceschi, F; Albo, E; D'Adamio, S; Di Martino, A; Vadalà, G; Faldini, C; Denaro, V

    2013-12-01

    Symptomatic tendon tears are one of the most important causes of pain and joint dysfunction. Among the intrinsic causes, vascularization recently gained a major role. Endothelial function is indeed a key factor, as well as vascular tone and thrombotic factors, in the regulation of vascular homeostasis and the composition of vascular wall. In this review, we studied systematically whether there is a relationship between endothelial dysfunction and tendinopathy. A literature search was performed using the isolated or combined keywords endothelial dysfunction and tendon,' 'nitric oxide (NO) and tendinopathy,' and 'endothelial dysfunction in tendon healing.' We identified 21 published studies. Of the selected studies, 9 were in vivo studies, 2 focusing on animals and 7 on humans, while 12 reported about in vitro evaluations, where 7 were carried out on humans and 5 on animals. The evidence about a direct relationship between tendinopathy and endothelial dysfunction is still poor. As recent studies have shown, there is no significant improvement in clinical and functional assessments after treatment with NO in patients suffering from tendinopathy in different locations. No significant differences were identified in the outcomes reported for experiment group when compared with controls treated with conventional surgical procedures or rehabilitation programs. Nitric oxide could be a marker to quantify the response of the endothelium to mechanical stress or hypoxia indicating the final balance between vasodilatating and vasoconstricting factors and their effects, but more ad stronger evidence is still needed to fully support this practice. PMID:23907599

  3. Effects of shear stress on endothelial progenitor cells.

    PubMed

    Obi, Syotaro; Yamamoto, Kimiko; Ando, Joji

    2014-10-01

    Endothelial progenitor cells (EPCs) are adult stem cells that play a central role in neovascularization. EPCs are mobilized from bone marrow into peripheral blood, attach to existing endothelial cells, and then transmigrate across the endothelium into tissues, where they proliferate, differentiate, and form new blood vessels. In the process, EPCs are exposed to shear stress, a biomechanical force generated by flowing blood and tissue fluid flow. When cultured EPCs are exposed to controlled levels of shear stress in a flow-loading device, their bioactivities in terms of proliferation, anti-apoptosis, migration, production of bioactive substances, anti-thrombosis, and tube formation increase markedly. Expression of endothelial marker genes and proteins by EPCs also increases in response to shear stress, and they differentiate into mature endothelial cells. Great advances have been made in elucidating the mechanisms by which mature endothelial cells sense and respond to shear stress, but not in EPCs. Further study of EPC responses to shear stress will be necessary to better understand the physiological and pathophysiological roles of EPCs and to apply EPCs to new therapies in the field of regenerative medicine. PMID:25992410

  4. Efficient Generation of Human Embryonic Stem Cell-Derived Corneal Endothelial Cells by Directed Differentiation

    PubMed Central

    McCabe, Kathryn L.; Kunzevitzky, Noelia J.; Chiswell, Brian P.; Xia, Xin; Goldberg, Jeffrey L.; Lanza, Robert

    2015-01-01

    Aim To generate human embryonic stem cell derived corneal endothelial cells (hESC-CECs) for transplantation in patients with corneal endothelial dystrophies. Materials and Methods Feeder-free hESC-CECs were generated by a directed differentiation protocol. hESC-CECs were characterized by morphology, expression of corneal endothelial markers, and microarray analysis of gene expression. Results hESC-CECs were nearly identical morphologically to primary human corneal endothelial cells, expressed Zona Occludens 1 (ZO-1) and Na+/K+ATPaseα1 (ATPA1) on the apical surface in monolayer culture, and produced the key proteins of Descemet’s membrane, Collagen VIIIα1 and VIIIα2 (COL8A1 and 8A2). Quantitative PCR analysis revealed expression of all corneal endothelial pump transcripts. hESC-CECs were 96% similar to primary human adult CECs by microarray analysis. Conclusion hESC-CECs are morphologically similar, express corneal endothelial cell markers and express a nearly identical complement of genes compared to human adult corneal endothelial cells. hESC-CECs may be a suitable alternative to donor-derived corneal endothelium. PMID:26689688

  5. Endothelial RIG-I activation impairs endothelial function

    SciTech Connect

    Asdonk, Tobias; Nickenig, Georg; Zimmer, Sebastian

    2012-03-30

    Highlights: Black-Right-Pointing-Pointer RIG-I activation impairs endothelial function in vivo. Black-Right-Pointing-Pointer RIG-I activation alters HCAEC biology in vitro. Black-Right-Pointing-Pointer EPC function is affected by RIG-I stimulation in vitro. -- Abstract: Background: Endothelial dysfunction is a crucial part of the chronic inflammatory atherosclerotic process and is mediated by innate and acquired immune mechanisms. Recent studies suggest that pattern recognition receptors (PRR) specialized in immunorecognition of nucleic acids may play an important role in endothelial biology in a proatherogenic manner. Here, we analyzed the impact of endothelial retinoic acid inducible gene I (RIG-I) activation upon vascular endothelial biology. Methods and results: Wild type mice were injected intravenously with 32.5 {mu}g of the RIG-ligand 3pRNA (RNA with triphosphate at the 5 Prime end) or polyA control every other day for 7 days. In 3pRNA-treated mice, endothelium-depended vasodilation was significantly impaired, vascular oxidative stress significantly increased and circulating endothelial microparticle (EMP) numbers significantly elevated compared to controls. To gain further insight in RIG-I dependent endothelial biology, cultured human coronary endothelial cells (HCAEC) and endothelial progenitor cells (EPC) were stimulated in vitro with 3pRNA. Both cells types express RIG-I and react with receptor upregulation upon stimulation. Reactive oxygen species (ROS) formation is enhanced in both cell types, whereas apoptosis and proliferation is not significantly affected in HCAEC. Importantly, HCAEC release significant amounts of proinflammatory cytokines in response to RIG-I stimulation. Conclusion: This study shows that activation of the cytoplasmatic nucleic acid receptor RIG-I leads to endothelial dysfunction. RIG-I induced endothelial damage could therefore be an important pathway in atherogenesis.

  6. Interaction of recombinant octameric hemoglobin with endothelial cells.

    PubMed

    Gaucher, Caroline; Domingues-Hamdi, Élisa; Prin-Mathieu, Christine; Menu, Patrick; Baudin-Creuza, Véronique

    2015-02-01

    Hemoglobin-based oxygen carriers (HBOCs) may generate oxidative stress, vasoconstriction and inflammation. To reduce these undesirable vasoactive properties, we increased hemoglobin (Hb) molecular size by genetic engineering with octameric Hb, recombinant (r) HbβG83C. We investigate the potential side effects of rHbβG83C on endothelial cells. The rHbβG83C has no impact on cell viability, and induces a huge repression of endothelial nitric oxide synthase gene transcription, a marker of vasomotion. No induction of Intermolecular-Adhesion Molecule 1 and E-selectin (inflammatory markers) transcription was seen. In the presence of rHbβG83C, the transcription of heme oxygenase-1 (oxidative stress marker) is weakly increased compared to the two other HBOCs (references) or Voluven (control). This genetically engineered octameric Hb, based on a human Hb βG83C mutant, leads to little impact at the level of endothelial cell inflammatory response and thus appears as an interesting molecule for HBOC development. PMID:25543885

  7. In Vitro Endothelialization Test of Biomaterials Using Immortalized Endothelial Cells

    PubMed Central

    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. PMID:27348615

  8. The Novel Methods for Analysis of Exosomes Released from Endothelial Cells and Endothelial Progenitor Cells

    PubMed Central

    Wang, Jinju; Guo, Runmin; Yang, Yi; Jacobs, Bradley; Chen, Suhong; Iwuchukwu, Ifeanyi; Gaines, Kenneth J.; Chen, Yanfang; Simman, Richard; Lv, Guiyuan; Wu, Keng; Bihl, Ji C.

    2016-01-01

    Exosomes (EXs) are cell-derived vesicles that mediate cell-cell communication and could serve as biomarkers. Here we described novel methods for purification and phenotyping of EXs released from endothelial cells (ECs) and endothelial progenitor cells (EPCs) by combining microbeads and fluorescence quantum dots (Q-dots®) techniques. EXs from the culture medium of ECs and EPCs were isolated and detected with cell-specific antibody conjugated microbeads and second antibody conjugated Q-dots by using nanoparticle tracking analysis (NTA) system. The sensitivities of the cell origin markers for ECs (CD105, CD144) and EPCs (CD34, KDR) were evaluated. The sensitivity and specificity were determined by using positive and negative markers for EXs (CD63), platelets (CD41), erythrocytes (CD235a), and microvesicles (Annexin V). Moreover, the methods were further validated in particle-free plasma and patient samples. Results showed that anti-CD105/anti-CD144 and anti-CD34/anti-KDR had the highest sensitivity and specificity for isolating and detecting EC-EXs and EPC-EXs, respectively. The methods had the overall recovery rate of over 70% and were able to detect the dynamical changes of circulating EC-EXs and EPC-EXs in acute ischemic stroke. In conclusion, we have developed sensitive and specific microbeads/Q-dots fluorescence NTA methods for EC-EX and EPC-EX isolation and detection, which will facilitate the functional study and biomarker discovery. PMID:27118976

  9. Na(+), K(+)-ATPase dysfunction causes cerebrovascular endothelial cell degeneration in rat prefrontal cortex slice cultures.

    PubMed

    Kurauchi, Yuki; Hisatsune, Akinori; Seki, Takahiro; Katsuki, Hiroshi

    2016-08-01

    Cerebrovascular endothelial cell dysfunction resulting in imbalance of cerebral blood flow contributes to the onset of psychiatric disorders such as depression, schizophrenia and bipolar disorder. Although decrease in Na(+), K(+)-ATPase activity has been reported in the patients with schizophrenia and bipolar disorder, the contribution of Na(+), K(+)-ATPase to endothelial cell dysfunction remains poorly understood. Here, by using rat neonatal prefrontal cortex slice cultures, we demonstrated that pharmacological inhibition of Na(+), K(+)-ATPase by ouabain induced endothelial cell injury. Treatment with ouabain significantly decreased immunoreactive area of rat endothelial cell antigen-1 (RECA-1), a marker of endothelial cells, in a time-dependent manner. Ouabain also decreased Bcl-2/Bax ratio and phosphorylation level of glycogen synthase kinase 3β (GSK3β) (Ser9), which were prevented by lithium carbonate. On the other hand, ouabain-induced endothelial cell injury was exacerbated by concomitant treatment with LY294002, an inhibitor of phosphoinositide 3- (PI3-) kinase. We also found that xestospongin C, an inhibitor of inositol triphosphate (IP3) receptor, but not SEA0400, an inhibitor of Na(+), Ca(2+) exchanger (NCX), protected endothelial cells from cytotoxicity of ouabain. These results suggest that cerebrovascular endothelial cell degeneration induced by Na(+), K(+)-ATPase inhibition resulting in Ca(2+) release from endoplasmic reticulum (ER) and activation of GSK3β signaling underlies pathogenesis of these psychiatric disorders. PMID:27208492

  10. Meeting report: ISN forefronts in nephrology on endothelial biology and renal disease: from bench to prevention.

    PubMed

    Goligorsky, M S; Rabelink, T

    2006-07-01

    This ISN-sponsored Forefront in Nephrology meeting, which has brought together 120 scientists from 21 countries, has been concerned with various aspects of endothelial function and dysfunction and their contribution to progression of chronic kidney disease and/or its cardiovascular complications. The following themes were discussed in great depth: (1) phenotypical changes in the vascular endothelium - permeability, senescence, and apoptosis; (2) regulation of endothelial nitric oxide (NO) synthase function - caveolar and shear stress mechanisms, epigenetic regulation, S-nitrosylation, and Rho-kinase regulation; (3) oxidative stress and hypoxia-induced changes; (4) organellar dysfunction - lysosomes, mitochondria, and endoplasmic reticulum; (5) NO-independent mechanisms of vasomotion - epoxides, heme oxygenase-1 and carbon monoxide, thromboxane, tumor necrosis factor-alpha, and uric acid; (6) endothelial crosstalk with podocytes, monocytes, smooth muscle cells, and platelets; (7) candidate clinical biomarkers of endothelial dysfunction - functional testing of macro- and micro-vascular functions, surrogate markers, circulating detached endothelial cells, and endothelial precursor cells; and culminated in Round Table discussion on the diagnosis of endothelial dysfunction and its treatment options. In conclusion, this meeting has focused on several key problems of endothelial cell pathobiology relevant to chronic kidney disease. PMID:16775602

  11. [Hypertension, endothelial dysfunction and cardiovascular risk].

    PubMed

    Nitenberg, A

    2006-10-01

    Increased blood pressure induces functional and structural changes of the vascular endothelium. Depression of endothelium-dependant vasodilatation is an early manifestation of endothelial dysfunction due to hypertension. It can be demonstrated by pharmacological or physiological tests. Decreased availability of nitric oxide (NO) is a major determinant of the depression of vasodilatation. It may be caused by a reduction in the activity of NO-endothelial synthase (NOSe) related to: 1) a deficit in substrate (L-arginine), 2) an inhibition by asymmetrical dimethylarginine, 3) a deficit in the cofactor tetrahydrobiopterin (BH4). However, the increase in oxidative stress, a producer of superoxide radicals which combine with NO to form peroxynitrates (ONOO-), is the determining factor. It is related to activation of membranous NAD(P)H oxidases initiated by the stimulation of activating mecanosensors of protein C kinase. The message is amplified by oxidation of BH4 which transforms the NOSe into a producer of superoxide radicals. A cascade of auto-amplification loops leading to atherosclerosis and its complications is then triggered. The superoxide radicals and the peroxynitrates oxidise the LDL-cholesterol. They activate the nuclear factor-kappaB which controls the genes stimulating the expression of many proteins: angiotensinogen and AT1 receptors which stimulate the sympathetic system, receptors of oxidised LDL, adhesion and migration factors (ICAM-1, VCAM-1, E-selectin and MCP-1), pro-inflammatory cytokins (interleukines and TNF-alpha), growth factors (MAP kinases), plasminogen activator inhibitor 1. The monocytes and smooth muscle cells produce metalloproteinases and pro-inflammatory cytokins which destabilise the atheromatous plaque and favourise vascular remodelling. Inshort, the endothelial dysfunction due to hypertension plays a role in a complex physiopathological process and is a marker of future cardiovascular events. PMID:17100143

  12. Tumor endothelial cells with distinct patterns of TGFβ-driven endothelial-to-mesenchymal transition

    PubMed Central

    Xiao, Lin; Kim, Dae Joong; Davis, Clayton L.; McCann, James V.; Dunleavey, James M.; Vanderlinden, Alissa; Xu, Nuo; Pattenden, Samantha G.; Frye, Stephen V.; Xu, Xia; Onaitis, Mark; Monaghan-Benson, Elizabeth; Burridge, Keith; Dudley, Andrew C.

    2015-01-01

    Endothelial-to-mesenchymal transition (EndMT) occurs during development and underlies the pathophysiology of multiple diseases. In tumors, unscheduled EndMT generates cancer-associated myofibroblasts that fuel inflammation and fibrosis, and may contribute to vascular dysfunction that promotes tumor progression. We report that freshly isolated subpopulations of tumor-specific endothelial cells (TEC) from a spontaneous mammary tumor model undergo distinct forms of EndMT in response to TGFβ stimulation. Whereas some TEC strikingly up-regulate alpha smooth muscle actin (SMA), a principal marker of EndMT and activated myofibroblasts, counterpart normal mammary gland endothelial cells (NEC) showed little change in SMA expression after TGFβ treatment. Compared with NEC, SMA+ TEC were 40 % less motile in wound healing assays and formed more stable vascular-like networks in vitro when challenged with TGFβ. Lineage tracing using ZsGreenCdh5-Cre reporter mice confirmed that only a fraction of vessels in breast tumors contain SMA+ TEC, suggesting that not all endothelial cells (EC) respond identically to TGFβ in vivo. Indeed, examination of 84 TGFβ-regulated target genes revealed entirely different genetic signatures in TGFβ-stimulated NEC and TEC cultures. Finally, we found that basic FGF (bFGF) exerts potent inhibitory effects on many TGFβ-regulated genes but operates in tandem with TGFβ to up-regulate others. EC challenged with TGFβ secrete bFGF which blocks SMA expression in secondary cultures suggesting a cell-autonomous or lateral-inhibitory mechanism for impeding mesenchymal differentiation. Together, our results suggest that TGFβ-driven EndMT produces a spectrum of EC phenotypes with different functions that could underlie the plasticity and heterogeneity of the tumor vasculature. PMID:25634211

  13. 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 (

  14. Endothelial cell permeability to water and antipyrine

    SciTech Connect

    Garrick, R.A.

    1986-03-05

    The endothelium provides a structural barrier between plasma constituents and the tissues. The permeability characteristics of the the endothelial cells regulate the transcellular movement of materials across this barrier while other movement is paracellular. In this study the permeability of the endothelial cells to tritiated water (/sup 3/HHO) and /sup 14/C-labeled antipyrine (AP) was investigated. The cells were isolated non-enzymatically from calf pulmonary artery and were maintained in culture and used between the seventh and fifteenth passage. The cells were removed from the T-flasks with a rubber policeman, titurated with a 22g needle and centrifuged. The cells were mixed with an extracellular marker, drawn into polyethylene tubing and packed by centrifugation for use in the linear diffusion technique. All measurements were made at 37 C. The diffusion coefficients for /sup 3/HHO through the packed cells (D), the intracellular material (D/sub 2/), and the extracellular material (D/sub 1/) were 0.682, 0.932 and 2.45 x 10/sup -5/ cm/sup 2/ s/sup -1/ and for AP were 0.273, 0.355 and 1.13 x 10/sup -5/ cm/sup 2/ s/sup -1/ respectively. The permeability coefficient calculated by the series-parallel pathway model for /sup 3/HHO was higher than that for AP and for both /sup 3/HHO and AP were lower than those calculated for isolated lung cells and erythrocytes.

  15. Impaired endothelial proliferation and mesenchymal transition contribute to vascular rarefaction following acute kidney injury.

    PubMed

    Basile, David P; Friedrich, Jessica L; Spahic, Jasmina; Knipe, Nicole; Mang, Henry; Leonard, Ellen C; Changizi-Ashtiyani, Saeed; Bacallao, Robert L; Molitoris, Bruce A; Sutton, Timothy A

    2011-03-01

    Acute kidney injury induces the loss of renal microvessels, but the fate of endothelial cells and the mechanism of potential vascular endothelial growth factor (VEGF)-mediated protection is unknown. Cumulative cell proliferation was analyzed in the kidney of Sprague-Dawley rats following ischemia-reperfusion (I/R) injury by repetitive administration of BrdU (twice daily) and colocalization in endothelial cells with CD31 or cablin. Proliferating endothelial cells were undetectable for up to 2 days following I/R and accounted for only ∼1% of BrdU-positive cells after 7 days. VEGF-121 preserved vascular loss following I/R but did not affect proliferation of endothelial, perivascular cells or tubular cells. Endothelial mesenchymal transition states were identified by localizing endothelial markers (CD31, cablin, or infused tomato lectin) with the fibroblast marker S100A4. Such structures were prominent within 6 h and sustained for at least 7 days following I/R. A Tie-2-cre transgenic crossed with a yellow fluorescent protein (YFP) reporter mouse was used to trace the fate of endothelial cells and demonstrated interstititial expansion of YFP-positive cells colocalizing with S100A4 and smooth muscle actin following I/R. The interstitial expansion of YFP cells was attenuated by VEGF-121. Multiphoton imaging of transgenic mice revealed the alteration of YFP-positive vascular cells associated with blood vessels characterized by limited perfusion in vivo. Taken together, these data indicate that vascular dropout post-AKI results from endothelial phenotypic transition combined with an impaired regenerative capacity, which may contribute to progressive chronic kidney disease. PMID:21123492

  16. Mechanisms of Tubulogenesis and Endothelial Phenotype Expression by MSCs

    PubMed Central

    Rytlewski, Julie A; Aldon, M Alejandra; Lewis, Evan W; Suggs, Laura J

    2015-01-01

    Stem cell-based therapies are a promising new avenue for treating ischemic disease and chronic wounds. Mesenchymal stem cells (MSCs) have a proven ability to augment the neovascularization processes necessary for wound healing and are widely popular as an autologous source of progenitor cells. Our lab has previously reported on PEGylated fibrin as a unique hydrogel that promotes spontaneous tubulogenesis of encapsulated MSCs without exogenous factors. However, the mechanisms underlying this process have remained unknown. To better understand the therapeutic value of PEGylated fibrin delivery of MSCs, we sought to clarify the relationship between biomaterial properties and cell behavior. Here we find that fibrin PEGylation does not dramatically alter the macroscopic mechanical properties of the fibrin-based matrix (less than 10% difference). It does, however, dramatically reduce the rate of diffusion through the gel matrix. PEGylated fibrin enhances the tubulogenic growth of encapsulated MSCs demonstrating fluid-filled lumens by interconnected MSCs. Image analysis gave a value of 4320±1770µm total network length versus 618±443µm for unmodified fibrin. PEGylation promotes the endothelial phenotype of encapsulated MSCs—compared to unmodified fibrin—as evidenced by higher levels of endothelial markers (von Willebrand factor, 2.2-fold; vascular endothelial cadherin, 1.8-fold) and vascular endothelial growth factor (VEGF, up to 1.8-fold). Prospective analysis of underlying molecular pathways demonstrated that this endothelial-like MSC behavior is sensitively modulated by hypoxic stress, but not VEGF supplementation as evidenced by a significant increase in VEGF and MMP-2 secretion per cell under hypoxia. Further gain-of-function studies under hypoxic stress demonstrated that hypoxia culture of MSCs in unmodified fibrin could increase both vWF and VE-cadherin levels to values that were not significantly different than cells cultured in PEGylated fibrin. This

  17. Mechanisms of tubulogenesis and endothelial phenotype expression by MSCs.

    PubMed

    Rytlewski, Julie A; Alejandra Aldon, M; Lewis, Evan W; Suggs, Laura J

    2015-05-01

    Stem cell-based therapies are a promising new avenue for treating ischemic disease and chronic wounds. Mesenchymal stem cells (MSCs) have a proven ability to augment the neovascularization processes necessary for wound healing and are widely popular as an autologous source of progenitor cells. Our lab has previously reported on PEGylated fibrin as a unique hydrogel that promotes spontaneous tubulogenesis of encapsulated MSCs without exogenous factors. However, the mechanisms underlying this process have remained unknown. To better understand the therapeutic value of PEGylated fibrin delivery of MSCs, we sought to clarify the relationship between biomaterial properties and cell behavior. Here we find that fibrin PEGylation does not dramatically alter the macroscopic mechanical properties of the fibrin-based matrix (less than 10% difference). It does, however, dramatically reduce the rate of diffusion through the gel matrix. PEGylated fibrin enhances the tubulogenic growth of encapsulated MSCs demonstrating fluid-filled lumens by interconnected MSCs. Image analysis gave a value of 4320 ± 1770 μm total network length versus 618 ± 443 μm for unmodified fibrin. PEGylation promotes the endothelial phenotype of encapsulated MSCs--compared to unmodified fibrin--as evidenced by higher levels of endothelial markers (von Willebrand factor, 2.2-fold; vascular endothelial cadherin, 1.8-fold) and vascular endothelial growth factor (VEGF, up to 1.8-fold). Prospective analysis of underlying molecular pathways demonstrated that this endothelial-like MSC behavior is sensitively modulated by hypoxic stress, but not VEGF supplementation as evidenced by a significant increase in VEGF and MMP-2 secretion per cell under hypoxia. Further gain-of-function studies under hypoxic stress demonstrated that hypoxia culture of MSCs in unmodified fibrin could increase both vWF and VE-cadherin levels to values that were not significantly different than cells cultured in PEGylated fibrin. This

  18. Regulation of human endothelial progenitor cell maturation by polyurethane nanocomposites.

    PubMed

    Hung, Huey-Shan; Yang, Yi-Chun; Lin, Yu-Chun; Lin, Shinn-Zong; Kao, Wei-Chien; Hsieh, Hsien-Hsu; Chu, Mei-Yun; Fu, Ru-Huei; Hsu, Shan-hui

    2014-08-01

    The mobilization and homing of endothelial progenitor cells (EPCs) are critical to the development of an antithrombotic cardiovascular prosthesis. Polyurethane (PU) with superior elasticity may provide a mechanical environment resembling that of the natural vascular tissues. The topographical cues of PU were maximized by making nanocomposites with a small amount of gold nanoparticles (AuNPs). The nanocomposites of PU-AuNPs ("PU-Au") with a favorable response of endothelial cells were previously established. In the current study, the effect of PU and PU-Au nanocomposites on the behavior of human peripheral blood EPCs was investigated in vitro and in vivo. It was found that PU-Au promoted EPCs to become differentiated endothelial cells in vitro, confirmed by the increased expressions of CD31 and VEGF-R2 surface markers. The increased maturation of EPCs was significantly more remarkable on PU-Au, probably through the stromal derived factor 1α (SDF-1α)/CXCR4 signaling pathway. In vivo experiments showed that EPCs seeded on PU-Au coated catheters effectively reduced thrombosis by differentiation into endothelial cells. Surface endothelialization with CD31 and CD34 expression as well as intimal formation with α-SMA expression was significantly accelerated in the group receiving EPC-seeded PU-Au catheters. Moreover, the analysis of collagen deposition revealed a reduction of fibrosis in the group receiving EPC-seeded PU-Au catheters as compared to the other groups. These results suggest that EPCs engineered with a proper elastic substrate may provide unique endothelialization and antithrombogenic properties that benefit vascular tissue regeneration. PMID:24836305

  19. Examining Endothelial Function and Platelet Reactivity in Patients with Depression before and after SSRI Therapy.

    PubMed

    Dawood, Tye; Barton, David A; Lambert, Elisabeth A; Eikelis, Nina; Lambert, Gavin W

    2016-01-01

    Although it is recognized that patients with major depressive disorder (MDD) are at increased risk of developing cardiovascular disease (CVD) the mechanisms responsible remain unknown. Endothelial dysfunction is one of the first signs of CVD. Using two techniques, flow-mediated dilatation in response to reactive hyperemia and laser Doppler velocimetry with iontophoresis, we examined endothelial function in the forearm before and after serotonin-specific reuptake inhibitor (SSRI) treatment in 31 patients with MDD. Measurement of intercellular adhesion molecule-1, vascular cell adhesion molecule-1, soluble P-selectin, and noradrenaline in plasma was also performed. Prior to treatment, markers of endothelial and vascular function and platelet reactivity were within the normal range. Following SSRI therapy (95 ± 5 days) symptoms of depression were reduced (paired difference between pre- and post-treatment Hamilton rating -18 ± 1, P < 0.001) with 19 patients recovered and 4 remitted. There occurred no significant change in markers of endothelial or vascular function following SSRI therapy. The improvement in Hamilton depression rating in response to therapy could be independently predicted by the baseline arterial plasma noradrenaline concentration (r (2) = 0.36, P = 0.003). In this cohort of patients with MDD, SSRI therapy did not influence endothelial function or markers of vascular or platelet reactivity. Patient response to SSRI therapy could be predicted by the initial circulating level of noradrenaline, with noradrenaline levels being lower in responders. PMID:26924994

  20. Examining Endothelial Function and Platelet Reactivity in Patients with Depression before and after SSRI Therapy

    PubMed Central

    Dawood, Tye; Barton, David A.; Lambert, Elisabeth A.; Eikelis, Nina; Lambert, Gavin W.

    2016-01-01

    Although it is recognized that patients with major depressive disorder (MDD) are at increased risk of developing cardiovascular disease (CVD) the mechanisms responsible remain unknown. Endothelial dysfunction is one of the first signs of CVD. Using two techniques, flow-mediated dilatation in response to reactive hyperemia and laser Doppler velocimetry with iontophoresis, we examined endothelial function in the forearm before and after serotonin-specific reuptake inhibitor (SSRI) treatment in 31 patients with MDD. Measurement of intercellular adhesion molecule-1, vascular cell adhesion molecule-1, soluble P-selectin, and noradrenaline in plasma was also performed. Prior to treatment, markers of endothelial and vascular function and platelet reactivity were within the normal range. Following SSRI therapy (95 ± 5 days) symptoms of depression were reduced (paired difference between pre- and post-treatment Hamilton rating −18 ± 1, P < 0.001) with 19 patients recovered and 4 remitted. There occurred no significant change in markers of endothelial or vascular function following SSRI therapy. The improvement in Hamilton depression rating in response to therapy could be independently predicted by the baseline arterial plasma noradrenaline concentration (r2 = 0.36, P = 0.003). In this cohort of patients with MDD, SSRI therapy did not influence endothelial function or markers of vascular or platelet reactivity. Patient response to SSRI therapy could be predicted by the initial circulating level of noradrenaline, with noradrenaline levels being lower in responders. PMID:26924994

  1. Tailoring Material Properties of Cardiac Matrix Hydrogels to Induce Endothelial Differentiation of Human Mesenchymal Stem Cells

    PubMed Central

    Jeffords, Megan E.; Wu, Jinglei; Shah, Mickey; Hong, Yi; Zhang, Ge

    2015-01-01

    Cardiac matrix hydrogel has shown great promise as an injectable biomaterial due to the possession of cardiac-specific extracellular matrix composition. A cardiac matrix hydrogel facilitating neovascularization will further improve its therapeutic outcomes in cardiac repair. In this study, we explored the feasibility of tailoring material properties of cardiac matrix hydrogels using a natural compound, genipin, to promote endothelial differentiation of stem cells. Our results demonstrated that the genipin crosslinking could increase the mechanical properties of the cardiac matrix hydrogel to a stiffness range promoting endothelial differentiation of human mesenchymal stem cells (hMSCs). It also decreased the swelling ratio and prolonged degradation without altering gelation time. Human mesenchymal stem cells cultured on the genipin crosslinked cardiac matrix hydrogels showed great viability. After 1-day culture, hMSCs demonstrated down-regulation of early endothelial marker expression and up-regulation of mature endothelial marker expression. Especially for 1 mM genipin crosslinked cardiac matrix hydrogels, hMSCs showed particularly significant expression of mature endothelial cell marker vWF. These attractive results indicate the potential of using genipin crosslinked cardiac matrix hydrogels to promote rapid vascularization for cardiac infarction treatment through minimally invasive therapy. PMID:25946697

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

    SciTech Connect

    Matsuda, Kohei; Ohga, Noritaka; Hida, Yasuhiro; Muraki, Chikara; Tsuchiya, Kunihiko; Kurosu, Takuro; Akino, Tomoshige; Shih, Shou-Ching; and others

    2010-04-16

    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, we 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.

  3. LPS-mediated endothelial activation in pulmonary endothelial cells: role of Nox2-dependent IKK-β phosphorylation

    PubMed Central

    Menden, Heather; Tate, Everett; Hogg, Neil

    2013-01-01

    Lipopolysaccharide (LPS)-mediated endothelial activation contributes to lung inflammation and alveolar remodeling seen in premature infants with bronchopulmonary dysplasia (BPD). The mechanisms underlying LPS-mediated oxidative stress and proinflammatory signaling in human pulmonary microvascular endothelial cells (HPMEC) remain unclear. We hypothesized that NADPH oxidase (Nox) mediates LPS-induced endothelial activation in HPMEC by regulating phosphorylation of Toll-like receptor (TLR) pathway proteins. LPS-induced expression of intercellular adhesion molecule 1 (ICAM-1) was associated with increased 2-OH-E+ (marker for superoxide formation) levels and was attenuated by apocynin and the Nox inhibitor, VAS2870. LPS triggered membrane translocation of p67phox, suggesting activation of Nox2. Silencing Nox2, but not Nox4, suppressed LPS-induced ICAM-1 expression in HPMEC. Immunoprecipitation studies showed that inhibitor of κ-B kinase-β (IKK-β) serine phosphorylation induced by LPS was inhibited by Nox2 silencing. We examined whether Nox2-dependent, LPS-mediated IKK-β phosphorylation was regulated by protein phosphatase 2A (PP2A) or TGF-β associated kinase-1 (TAK1) in HPMEC. LPS increased PP2A activity in HPMEC, and inhibition of PP2A did not alter LPS-mediated ICAM-1 expression but attenuated IKK-β phosphorylation. TAK1 inhibition decreased LPS-induced ICAM-1 expression in HPMEC, and Nox2 silencing attenuated LPS-mediated TAK1 phosphorylation (Thr184/187). We demonstrate that Nox2 regulates LPS-mediated endothelial activation in pulmonary endothelial cells by modulating phosphorylation of key kinases in the TLR signaling cascade. Our data support a novel mechanism by which Nox-dependent signaling regulates proinflammatory signaling in pulmonary endothelial cells. Inhibition of vascular Nox may potentially limit lung injury and alveolar remodeling caused by infections in BPD. PMID:23333803

  4. 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

  5. Solid tumor therapy by selectively targeting stromal endothelial cells.

    PubMed

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

    2016-07-12

    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

  6. Evolution of Neuronal and Endothelial Transcriptomes in Primates

    PubMed Central

    Giger, Thomas; Khaitovich, Philipp; Somel, Mehmet; Lorenc, Anna; Lizano, Esther; Harris, Laura W.; Ryan, Margaret M.; Lan, Martin; Wayland, Matthew T.; Bahn, Sabine; Pääbo, Svante

    2010-01-01

    The study of gene expression evolution in vertebrates has hitherto focused on the analysis of transcriptomes in tissues of different species. However, because a tissue is made up of different cell types, and cell types differ with respect to their transcriptomes, the analysis of tissues offers a composite picture of transcriptome evolution. The isolation of individual cells from tissue sections opens up the opportunity to study gene expression evolution at the cell type level. We have stained neurons and endothelial cells in human brains by antibodies against cell type-specific marker proteins, isolated the cells using laser capture microdissection, and identified genes preferentially expressed in the two cell types. We analyze these two classes of genes with respect to their expression in 62 different human tissues, with respect to their expression in 44 human “postmortem” brains from different developmental stages and with respect to between-species brain expression differences. We find that genes preferentially expressed in neurons differ less across tissues and developmental stages than genes preferentially expressed in endothelial cells. We also observe less expression differences within primate species for neuronal transcriptomes. In stark contrast, we see more gene expression differences between humans, chimpanzees, and rhesus macaques relative to within-species differences in genes expressed preferentially in neurons than in genes expressed in endothelial cells. This suggests that neuronal and endothelial transcriptomes evolve at different rates within brain tissue. PMID:20624733

  7. Endothelial dysfunction in cirrhosis: Role of inflammation and oxidative stress

    PubMed Central

    Vairappan, Balasubramaniyan

    2015-01-01

    This review describes the recent developments in the pathobiology of endothelial dysfunction (ED) in the context of cirrhosis with portal hypertension and defines novel strategies and potential targets for therapy. ED has prognostic implications by predicting unfavourable early hepatic events and mortality in patients with portal hypertension and advanced liver diseases. ED characterised by an impaired bioactivity of nitric oxide (NO) within the hepatic circulation and is mainly due to decreased bioavailability of NO and accelerated degradation of NO with reactive oxygen species. Furthermore, elevated inflammatory markers also inhibit NO synthesis and causes ED in cirrhotic liver. Therefore, improvement of NO availability in the hepatic circulation can be beneficial for the improvement of endothelial dysfunction and associated portal hypertension in patients with cirrhosis. Furthermore, therapeutic agents that are identified in increasing NO bioavailability through improvement of hepatic endothelial nitric oxide synthase (eNOS) activity and reduction in hepatic asymmetric dimethylarginine, an endogenous modulator of eNOS and a key mediator of elevated intrahepatic vascular tone in cirrhosis would be interesting therapeutic approaches in patients with endothelial dysfunction and portal hypertension in advanced liver diseases. PMID:25848469

  8. Senescence-Induced Oxidative Stress Causes Endothelial Dysfunction.

    PubMed

    Bhayadia, Raj; Schmidt, Bernhard M W; Melk, Anette; Hömme, Meike

    2016-02-01

    Age is a risk factor for cardiovascular disease, suggesting a causal relationship between age-related changes and vascular damage. Endothelial dysfunction is an early pathophysiological hallmark in the development of cardiovascular disease. Senescence, the cellular equivalent of aging, was proposed to be involved in endothelial dysfunction, but functional data showing a causal relationship are missing.Endothelium-dependent vasodilation was measured in aortic rings ex vivo. We investigated aortas from aged C57Bl/6 mice (24-28 months), in which p16 (INK4a) and p19 (ARF) expression, markers of stress-induced senescence, were significantly induced compared to young controls (4-6 months). To reflect telomere shortening in human aging, we investigated aortas from telomerase deficient (Terc(-/-)) mice of generation 3 (G3). Endothelium-dependent vasodilation in aged wildtype and in Terc(-/-) G3 mice was impaired. A combination of the superoxide dismutase mimetic 1-Oxyl-2,2,6, 6-tetramethyl-4-hydroxypiperidine (TEMPOL) and the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor apocynin significantly improved endothelium-dependent vasodilation in aged wildtype and Terc(-/-) G3 mice compared to untreated controls. We show that both, aging and senescence induced by telomere shortening, cause endothelial dysfunction that can be restored by antioxidants, indicating a role for oxidative stress. The observation that cellular senescence is a direct signalling event leading to endothelial dysfunction holds the potential to develop new targets for the prevention of cardiovascular disease. PMID:25735595

  9. ELIMINATION OF VITAMIN D RECEPTOR IN VASCULAR ENDOTHELIAL CELLS ALTERS VASCULAR FUNCTION

    PubMed Central

    Ni, Wei; Watts, Stephanie W.; Ng, Michael; Chen, Songcang; Glenn, Denis J.; Gardner, David G.

    2014-01-01

    Vitamin D deficiency has been associated with cardiovascular dysfunction. We evaluated the role of the vitamin D receptor (VDR) in vascular endothelial function, a marker of cardiovascular health, at baseline and in the presence of angiotensin II, using an endothelial-specific knockout of the murine VDR gene. In the absence of endothelial VDR, acetylcholine-induced aortic relaxation was significantly impaired (maximal relaxation, endothelial-specific VDR knockout =58% vs. control=73%, p<0.05). This was accompanied by a reduction in eNOS expression and phospho-vasodilator-stimulated phosphoprotein levels in aortae from the endothelial-specific VDR knockout vs. control mice. While blood pressure levels at baseline were comparable at 12 and 24 weeks of age, the endothelial VDR knockout mice demonstrated increased sensitivity to the hypertensive effects of angiotensin II compared to control mice (after 1-week infusion: knockout = 155±15 mmHg vs. control = 133±7 mmHg, p<0.01; after 2-week infusion: knockout = 164±9 mmHg vs. control = 152±13 mmHg, p<0.05). By the end of two weeks, angiotensin II infusion-induced, hypertrophy-sensitive myocardial gene expression was higher in endothelial-specific VDR knockout mice (fold change compared to saline-infused control mice, ANP: knockout mice = 3.12 vs. control= 1.7, p<0.05; BNP: knockout mice= 4.72 vs. control= 2.68, p<0.05). These results suggest that endothelial VDR plays an important role in endothelial cell function and blood pressure control and imply a potential role for VDR agonists in the management of cardiovascular disease associated with endothelial dysfunction. PMID:25201890

  10. Generation of vascular endothelial and smooth muscle cells from human pluripotent stem cells

    PubMed Central

    Patsch, Christoph; Challet-Meylan, Ludivine; Thoma, Eva C.; Urich, Eduard; Heckel, Tobias; O’Sullivan, John F; Grainger, Stephanie J; Kapp, Friedrich G.; Sun, Lin; Christensen, Klaus; Xia, Yulei; Florido, Mary H. C.; He, Wei; Pan, Wei; Prummer, Michael; Warren, Curtis R.; Jakob-Roetne, Roland; Certa, Ulrich; Jagasia, Ravi; Freskgård, Per-Ola; Adatto, Isaac; Kling, Dorothee; Huang, Paul; Zon, Leonard I; Chaikof, Elliot L.; Gerszten, Robert E.; Graf, Martin; Iacone, Roberto; Cowan, Chad A.

    2015-01-01

    The use of human pluripotent stem cells for in vitro disease modeling and clinical applications requires protocols that convert these cells into relevant adult cell types. Here, we report the rapid and efficient differentiation of human pluripotent stem cells into vascular endothelial and smooth muscle cells. We found that GSK3 inhibition and BMP4 treatment rapidly committed pluripotent cells to a mesodermal fate and subsequent exposure to VEGF or PDGF-BB resulted in the differentiation of either endothelial or vascular smooth muscle cells, respectively. Both protocols produced mature cells with efficiencies over 80% within six days. Upon purification to 99% via surface markers, endothelial cells maintained their identity, as assessed by marker gene expression, and showed relevant in vitro and in vivo functionality. Global transcriptional and metabolomic analyses confirmed that the cells closely resembled their in vivo counterparts. Our results suggest that these cells could be used to faithfully model human disease. PMID:26214132

  11. Generation of vascular endothelial and smooth muscle cells from human pluripotent stem cells.

    PubMed

    Patsch, Christoph; Challet-Meylan, Ludivine; Thoma, Eva C; Urich, Eduard; Heckel, Tobias; O'Sullivan, John F; Grainger, Stephanie J; Kapp, Friedrich G; Sun, Lin; Christensen, Klaus; Xia, Yulei; Florido, Mary H C; He, Wei; Pan, Wei; Prummer, Michael; Warren, Curtis R; Jakob-Roetne, Roland; Certa, Ulrich; Jagasia, Ravi; Freskgård, Per-Ola; Adatto, Isaac; Kling, Dorothee; Huang, Paul; Zon, Leonard I; Chaikof, Elliot L; Gerszten, Robert E; Graf, Martin; Iacone, Roberto; Cowan, Chad A

    2015-08-01

    The use of human pluripotent stem cells for in vitro disease modelling and clinical applications requires protocols that convert these cells into relevant adult cell types. Here, we report the rapid and efficient differentiation of human pluripotent stem cells into vascular endothelial and smooth muscle cells. We found that GSK3 inhibition and BMP4 treatment rapidly committed pluripotent cells to a mesodermal fate and subsequent exposure to VEGF-A or PDGF-BB resulted in the differentiation of either endothelial or vascular smooth muscle cells, respectively. Both protocols produced mature cells with efficiencies exceeding 80% within six days. On purification to 99% via surface markers, endothelial cells maintained their identity, as assessed by marker gene expression, and showed relevant in vitro and in vivo functionality. Global transcriptional and metabolomic analyses confirmed that the cells closely resembled their in vivo counterparts. Our results suggest that these cells could be used to faithfully model human disease. PMID:26214132

  12. Diabetes Causes Bone Marrow Endothelial Barrier Dysfunction by Activation of the RhoA–Rho-Associated Kinase Signaling Pathway

    PubMed Central

    Mangialardi, Giuseppe; Katare, Rajesh; Oikawa, Atsuhiko; Meloni, Marco; Reni, Carlotta; Emanueli, Costanza; Madeddu, Paolo

    2013-01-01

    Objective Diabetes mellitus causes bone marrow (BM) microangiopathy. This study aimed to investigate the mechanisms responsible for BM endothelial dysfunction in diabetes mellitus. Methods and Results The analysis of differentially expressed transcripts in BM endothelial cells (BMECs) from type-1 diabetic and nondiabetic mice showed an effect of diabetes mellitus on signaling pathways controlling cell death, migration, and cytoskeletal rearrangement. Type-1 diabetic-BMECs displayed high reactive oxygen species levels, increased expression and activity of RhoA and its associated protein kinases Rho-associated kinase 1/Rho-associated kinase 2, and reduced Akt phosphorylation/activity. Likewise, diabetes mellitus impaired Akt-related BMEC functions, such as migration, network formation, and angiocrine factor-releasing activity, and increased vascular permeability. Moreover, high glucose disrupted BMEC contacts through Src tyrosine kinase phosphorylation of vascular endothelial cadherin. These alterations were prevented by constitutively active Akt (myristoylated Akt), Rho-associated kinase inhibitor Y-27632, and Src inhibitors. Insulin replacement restored BMEC abundance, as assessed by flow cytometry analysis of the endothelial marker MECA32, and endothelial barrier function in BM of type-1 diabetic mice. Conclusion Redox-dependent activation of RhoA/Rho-associated kinase and Src/vascular endothelial cadherin signaling pathways, together with Akt inactivation, contribute to endothelial dysfunction in diabetic BM. Metabolic control is crucial for maintenance of endothelial cell homeostasis and endothelial barrier function in BM of diabetic mice. PMID:23307872

  13. HDL in children with CKD promotes endothelial dysfunction and an abnormal vascular phenotype.

    PubMed

    Shroff, Rukshana; Speer, Thimoteus; Colin, Sophie; Charakida, Marietta; Zewinger, Stephen; Staels, Bart; Chinetti-Gbaguidi, Giulia; Hettrich, Inga; Rohrer, Lucia; O'Neill, Francis; McLoughlin, Eve; Long, David; Shanahan, Catherine M; Landmesser, Ulf; Fliser, Danilo; Deanfield, John E

    2014-11-01

    Endothelial dysfunction begins in early CKD and contributes to cardiovascular mortality. HDL is considered antiatherogenic, but may have adverse vascular effects in cardiovascular disease, diabetes, and inflammatory conditions. The effect of renal failure on HDL properties is unknown. We studied the endothelial effects of HDL isolated from 82 children with CKD stages 2-5 (HDL(CKD)), who were free of underlying inflammatory diseases, diabetes, or active infections. Compared with HDL from healthy children, HDL(CKD) strongly inhibited nitric oxide production, promoted superoxide production, and increased vascular cell adhesion molecule-1 expression in human aortic endothelial cells, and reduced cholesterol efflux from macrophages. The effects on endothelial cells correlated with CKD grade, with the most profound changes induced by HDL from patients on dialysis, and partial recovery observed with HDL isolated after kidney transplantation. Furthermore, the in vitro effects on endothelial cells associated with increased aortic pulse wave velocity, carotid intima-media thickness, and circulating markers of endothelial dysfunction in patients. Symmetric dimethylarginine levels were increased in serum and fractions of HDL from children with CKD. In a longitudinal follow-up of eight children undergoing kidney transplantation, HDL-induced production of endothelial nitric oxide, superoxide, and vascular cell adhesion molecule-1 in vitro improved significantly at 3 months after transplantation, but did not reach normal levels. These results suggest that in children with CKD without concomitant disease affecting HDL function, HDL dysfunction begins in early CKD, progressing as renal function declines, and is partially reversed after kidney transplantation. PMID:24854267

  14. 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. PMID:25953848

  15. Endothelial cells from bovine adrenal medulla develop capillary-like growth patterns in culture.

    PubMed Central

    Banerjee, D K; Ornberg, R L; Youdim, M B; Heldman, E; Pollard, H B

    1985-01-01

    The endocrine barrier between chromaffin cells and the blood stream in the adrenal medulla is made of capillary endothelial cells. We have now succeeded in isolating endothelial cells from adrenal medullary tissue, which are probably derived from this barrier. These cells grow on plastic surfaces in the absence of special growth factors or collagen overlays and differentiate into organized structures quite similar to true capillaries. The cells contain factor VIII:R, a marker for endothelial cells, and form intercellular junctions characteristic of capillary endothelial cells. They also synthesize and secrete basal lamina structures and engage in transcytosis, a characteristic ultrastructural and functional combination of exocytosis and endocytosis across the thin endothelial cell processes. These endothelial cells can take up and deaminate catecholamines by A-type monoamine oxidase, an enzyme functionally distinct from the B-type monoamine oxidase found in chromaffin cells. These data indicate that the chromaffin cell and its endothelial cell neighbor may constitute the functional unit of catecholamine metabolism in the adrenal medulla. Images PMID:3927288

  16. 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

  17. A Methodology for Concomitant Isolation of Intimal and Adventitial Endothelial Cells from the Human Thoracic Aorta

    PubMed Central

    Leclercq, Anne; Veillat, Véronique; Loriot, Sandrine; Spuul, Pirjo; Madonna, Francesco; Roques, Xavier; Génot, Elisabeth

    2015-01-01

    Background Aortic diseases are diverse and involve a multiplicity of biological systems in the vascular wall. Aortic dissection, which is usually preceded by aortic aneurysm, is a leading cause of morbidity and mortality in modern societies. Although the endothelium is now known to play an important role in vascular diseases, its contribution to aneurysmal aortic lesions remains largely unknown. The aim of this study was to define a reliable methodology for the isolation of aortic intimal and adventitial endothelial cells in order to throw light on issues relevant to endothelial cell biology in aneurysmal diseases. Methodology/Principal Findings We set up protocols to isolate endothelial cells from both the intima and the adventitia of human aneurysmal aortic vessel segments. Throughout the procedure, analysis of cell morphology and endothelial markers allowed us to select an endothelial fraction which after two rounds of expansion yielded a population of >90% pure endothelial cells. These cells have the features and functionalities of freshly isolated cells and can be used for biochemical studies. The technique was successfully used for aortic vessel segments of 20 patients and 3 healthy donors. Conclusions/Significance This simple and highly reproducible method allows the simultaneous preparation of reasonably pure primary cultures of intimal and adventitial human endothelial cells, thus providing a reliable source for investigating their biology and involvement in both thoracic aneurysms and other aortic diseases. PMID:26599408

  18. Endothelial mitochondria and heart disease.

    PubMed

    Davidson, Sean Michael

    2010-10-01

    The endothelium is vital to the proper functioning in the heart, in particular due to its production of nitric oxide (NO) which regulates vascular tone. Damage to the endothelium contributes to the development of atherosclerosis, and hence to possible myocardial infarction and subsequent heart failure. Like most cells, endothelial cells contain mitochondria, despite their having relatively little dependence on oxidative phosphorylation for ATP production. However, endothelial mitochondria are centrally involved in maintaining the fine regulatory balance between mitochondrial calcium concentration, reactive oxygen species (ROS) production, and NO. This raises the question of whether damage to endothelial mitochondria would have repercussions in terms of the development of heart disease. In fact, increasingly nuanced techniques enabling restricted transgenic expression of antioxidant proteins in mice has demonstrated that mitochondrial ROS do contribute to endothelial damage. New pharmaceutical approaches designed to target protective molecules such as ROS scavengers to the mitochondria promise to be effective in preventing heart disease. As well as protecting cardiomyocytes, these drugs may have the added benefit of preventing damage to the endothelial mitochondria. However, much remains to be done in understanding the contribution that mitochondria make to endothelial function. PMID:20558442

  19. NHERF-2 maintains endothelial homeostasis

    PubMed Central

    Bhattacharya, Resham; Wang, Enfeng; Dutta, Shamit K.; Vohra, Pawan K.; E, Guangqi; Prakash, Y. S.

    2012-01-01

    The Na+/H+ exchanger regulatory factor-2 (NHERF-2) is an integral component of almost all endothelial cells (ECs), yet its endothelial function is not known. Here, we found that NHERF-2, is a key regulator of endothelial homeostasis because NHERF-2–silenced ECs proliferate at a much higher rate even in the absence of mitogens such as VEGF compared with control ECs. We further show that the hyperproliferation phenotype of NHERF-2–silenced EC is because of an accelerated cell cycle that is probably caused by a combination of the following factors: increased cytoplasmic calcium, increased expression of c-Myc, increased expression of cyclin D1, and reduced expression of p27. Using an experimental mouse model of human hemangioma, we found that the endothelial neoplasms derived from NHERF-2–silenced cells were much larger in volume than those derived from control cells. Thus, NHERF-2 is a negative regulator of endothelial proliferation and may have important roles in endothelial homeostasis and vascular modeling. PMID:22343917

  20. Discourse Markers across Language.

    ERIC Educational Resources Information Center

    Fraser, Bruce

    This paper discusses discourse markers (e.g., "and, so, anyway") and offers an overview of their characteristics and occurrence, using English for illustration. The role of discourse markers is to signal speaker comment on the current utterance. The discourse marker is not part of the sentence's propositional content. While absence of markers does…

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

    SciTech Connect

    Mathieu, Celine . E-mail: marc-andre.mouthon@cea.fr

    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. Endothelial 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.

  2. Endothelial cellular senescence is inhibited by liver X receptor activation with an additional mechanism for its atheroprotection in diabetes

    PubMed Central

    Hayashi, Toshio; Kotani, Hitoshi; Yamaguchi, Tomoe; Taguchi, Kumiko; Iida, Mayu; Ina, Koichiro; Maeda, Morihiko; Kuzuya, Masafumi; Hattori, Yuichi; Ignarro, Louis J.

    2014-01-01

    Senescence of vascular endothelial cells leads to endothelial dysfunction and contributes to the progression of atherosclerosis. Liver X receptors (LXRs) are nuclear receptors whose activation protects against atherosclerosis by transcriptional regulation of genes important in promoting cholesterol efflux and inhibiting inflammation. Here we found that LXR activation with specific ligands reduced the increase in senescence-associated (SA) β-gal activity, a senescence marker, and reversed the decrease in telomerase activity, a replicative senescence marker, in human endothelial cells under high glucose. This effect of LXR activation was associated with reduced reactive oxygen species and increased endothelial NO synthase activity. A series of experiments that used siRNAs indicated that LXRβ mediates the prevention of endothelial cellular senescence, and that sterol regulatory element binding protein-1, which was up-regulated as a direct LXRβ target gene, may act as a brake of endothelial cellular senescence. Although oral administration of the LXR ligand led to severe fatty liver in diabetic rats, concomitant therapy with metformin avoided the development of hepatic steatosis. However, the preventive effect of the LXR ligand on SA β-gal–stained cells in diabetic aortic endothelium was preserved even if metformin was coadministered. Taken together, our studies demonstrate that an additional mechanism, such as the regulation of endothelial cellular senescence, is related to the antiatherogenic properties of LXRs, and concomitant treatment with metformin may provide a clinically useful therapeutic strategy to alleviate an LXR activation-mediated adverse effects on liver triglyceride metabolism. PMID:24398515

  3. A novel method of differential gene expression analysis using multiple cDNA libraries applied to the identification of tumour endothelial genes

    PubMed Central

    Herbert, John MJ; Stekel, Dov; Sanderson, Sharon; Heath, Victoria L; Bicknell, Roy

    2008-01-01

    Background In this study, differential gene expression analysis using complementary DNA (cDNA) libraries has been improved. Firstly by the introduction of an accurate method of assigning Expressed Sequence Tags (ESTs) to genes and secondly, by using a novel likelihood ratio statistical scoring of differential gene expression between two pools of cDNA libraries. These methods were applied to the latest available cell line and bulk tissue cDNA libraries in a two-step screen to predict novel tumour endothelial markers. Initially, endothelial cell lines were in silico subtracted from non-endothelial cell lines to identify endothelial genes. Subsequently, a second bulk tumour versus normal tissue subtraction was employed to predict tumour endothelial markers. Results From an endothelial cDNA library analysis, 431 genes were significantly up regulated in endothelial cells with a False Discovery Rate adjusted q-value of 0.01 or less and 104 of these were expressed only in endothelial cells. Combining the cDNA library data with the latest Serial Analysis of Gene Expression (SAGE) library data derived a complete list of 459 genes preferentially expressed in endothelium. 27 genes were predicted tumour endothelial markers in multiple tissues based on the second bulk tissue screen. Conclusion This approach represents a significant advance on earlier work in its ability to accurately assign an EST to a gene, statistically measure differential expression between two pools of cDNA libraries and predict putative tumour endothelial markers before entering the laboratory. These methods are of value and available to researchers that are interested in the analysis of transcriptomic data. PMID:18394197

  4. Oxidized plasma albumin promotes platelet-endothelial crosstalk and endothelial tissue factor expression

    PubMed Central

    Pasterk, Lisa; Lemesch, Sandra; Leber, Bettina; Trieb, Markus; Curcic, Sanja; Stadlbauer, Vanessa; Schuligoi, Rufina; Schicho, Rudolf; Heinemann, Akos; Marsche, Gunther

    2016-01-01

    Plasma advanced oxidation protein products (AOPPs), a class of pro-inflammatory pathogenic mediators, accumulate in subjects with chronic kidney disease. Whether AOPPs contribute to coagulation abnormalities, which are frequently seen in uremic patients, is unknown. Here we report that AOPPs activate platelets via a CD36-mediated signaling pathway. Activation of signaling pathways by AOPP-platelet interaction resulted in the expression of several platelet activation markers and rapidly induced the expression of CD40 ligand, triggering platelet adhesion to endothelial cells and promoting endothelial tissue factor expression. AOPPs and serum tissue factor levels were considerably increased in end stage renal disease patients on hemodialysis and a significant correlation of AOPPs and serum tissue factor was found. Interestingly, serum levels of AOPPs and tissue factor were substantially lower in stable kidney transplant patients when compared with hemodialysis patients. Given that CD36 is known to transduce the effects of oxidized lipids into platelet hyperactivity, our findings reveal previously unknown pro-thrombotic activities of oxidized plasma albumin via a CD36 dependent pathway. PMID:26905525

  5. Dietary flavanol intervention lowers the levels of endothelial microparticles in coronary artery disease patients.

    PubMed

    Horn, Patrick; Amabile, Nicolas; Angeli, Franca S; Sansone, Roberto; Stegemann, Berthold; Kelm, Malte; Springer, Matthew L; Yeghiazarians, Yerem; Schroeter, Hagen; Heiss, Christian

    2014-04-14

    Current evidence suggests that regenerative v. degenerative endothelial responses can be integrated in a clinical endothelial phenotype, reflecting the net result between damage from risk factors and endogenous repair capacity. We have previously shown that a cocoa flavanol (CF) intervention can improve endothelial function and increase the regenerative capacity of the endothelium by mobilising circulating angiogenic cells in patients with coronary artery disease (CAD). The aim of the present study was to investigate whether CF can lower the levels of circulating endothelial microparticles (EMP), markers of endothelial integrity, along with improvements in endothelial function. The levels of EMP in the frozen plasma samples of CAD patients were measured along with endothelial function (flow-mediated vasodilation, FMD); n 16, FMD data published previously), and these data were compared with those of young (n 12) and age-matched (n 12) healthy control subjects. The CAD patients exhibited significantly increased levels of EMP along with impaired FMD when compared with the healthy control subjects. The levels of CD144⁺ and CD31⁺/41⁻ EMP were inversely correlated with FMD (r -0.67, P=0.01 and r -0.59, P=0.01, respectively). In these CAD patients, the levels of EMP were measured after they had consumed a drink containing 375 mg of CF (high-CF intervention, HiFI) or 9 mg of CF (macro- and micronutrient-matched low-CF control, LoFl) twice daily over a 30-d period in a randomised, double-blind, cross-over study. After 1 month of HiFI, the levels of CD31⁺/41⁻ and CD144⁺ EMP decreased (-25 and -23%, respectively), but not after LoFl. Our data show that flavanols lower the levels of EMP along with higher endothelial function, lending evidence to the novel concept that flavanols may improve endothelial integrity. PMID:24286443

  6. Gene expression programs of mouse endothelial cells in kidney development and disease.

    PubMed

    Brunskill, Eric W; Potter, S Steven

    2010-01-01

    Endothelial cells are remarkably heterogeneous in both morphology and function, and they play critical roles in the formation of multiple organ systems. In addition endothelial cell dysfunction can contribute to disease processes, including diabetic nephropathy, which is a leading cause of end stage renal disease. In this report we define the comprehensive gene expression programs of multiple types of kidney endothelial cells, and analyze the differences that distinguish them. Endothelial cells were purified from Tie2-GFP mice by cell dissociation and fluorescent activated cell sorting. Microarrays were then used to provide a global, quantitative and sensitive measure of gene expression levels. We examined renal endothelial cells from the embryo and from the adult glomerulus, cortex and medulla compartments, as well as the glomerular endothelial cells of the db/db mutant mouse, which represents a model for human diabetic nephropathy. The results identified the growth factors, receptors and transcription factors expressed by these multiple endothelial cell types. Biological processes and molecular pathways were characterized in exquisite detail. Cell type specific gene expression patterns were defined, finding novel molecular markers and providing a better understanding of compartmental distinctions. Further, analysis of enriched, evolutionarily conserved transcription factor binding sites in the promoters of co-activated genes begins to define the genetic regulatory network of renal endothelial cell formation. Finally, the gene expression differences associated with diabetic nephropathy were defined, providing a global view of both the pathogenic and protective pathways activated. These studies provide a rich resource to facilitate further investigations of endothelial cell functions in kidney development, adult compartments, and disease. PMID:20706631

  7. Gene Expression Programs of Mouse Endothelial Cells in Kidney Development and Disease

    PubMed Central

    Brunskill, Eric W.; Potter, S. Steven

    2010-01-01

    Endothelial cells are remarkably heterogeneous in both morphology and function, and they play critical roles in the formation of multiple organ systems. In addition endothelial cell dysfunction can contribute to disease processes, including diabetic nephropathy, which is a leading cause of end stage renal disease. In this report we define the comprehensive gene expression programs of multiple types of kidney endothelial cells, and analyze the differences that distinguish them. Endothelial cells were purified from Tie2-GFP mice by cell dissociation and fluorescent activated cell sorting. Microarrays were then used to provide a global, quantitative and sensitive measure of gene expression levels. We examined renal endothelial cells from the embryo and from the adult glomerulus, cortex and medulla compartments, as well as the glomerular endothelial cells of the db/db mutant mouse, which represents a model for human diabetic nephropathy. The results identified the growth factors, receptors and transcription factors expressed by these multiple endothelial cell types. Biological processes and molecular pathways were characterized in exquisite detail. Cell type specific gene expression patterns were defined, finding novel molecular markers and providing a better understanding of compartmental distinctions. Further, analysis of enriched, evolutionarily conserved transcription factor binding sites in the promoters of co-activated genes begins to define the genetic regulatory network of renal endothelial cell formation. Finally, the gene expression differences associated with diabetic nephropathy were defined, providing a global view of both the pathogenic and protective pathways activated. These studies provide a rich resource to facilitate further investigations of endothelial cell functions in kidney development, adult compartments, and disease. PMID:20706631

  8. Radial keratotomy associated endothelial degeneration

    PubMed Central

    Moshirfar, Majid; Ollerton, Andrew; Semnani, Rodmehr T; Hsu, Maylon

    2012-01-01

    Purpose To describe the presentation and clinical course of eyes with a history of radial keratotomy (RK) and varying degrees of endothelial degeneration. Methods Retrospective case series were used. Results Thirteen eyes (seven patients) were identified with clinical findings of significant guttata and a prior history of RK. The mean age of presentation for cornea evaluation was 54.3 years (range: 38–72 years), averaging 18.7 years (range: 11–33 years) after RK. The presentation of guttata varied in degree from moderate to severe. Best corrected visual acuity (BCVA) ranged from 20/25 to 20/80. All patients had a history of bilateral RK, except one patient who did not develop any guttata in the eye without prior RK. No patients reported a family history of Fuch’s Dystrophy. One patient underwent a penetrating keratoplasty in one eye and a Descemet’s stripping automated endothelial keratoplasty (DSAEK) in the other eye. Conclusions RK may induce a spectrum of endothelial degeneration. In elderly patients, the findings of guttata may signify comorbid Fuch’s dystrophy in which RK incisions could potentially hasten endothelial decomposition. In these select patients with stable cornea topography and prior RK, DSAEK may successfully treat RK endothelial degeneration. PMID:22347792

  9. How cholesterol regulates endothelial biomechanics

    PubMed Central

    Hong, Zhongkui; Staiculescu, Marius C.; Hampel, Paul; Levitan, Irena; Forgacs, Gabor

    2012-01-01

    As endothelial cells form the barrier between blood flow and surrounding tissue, many of their functions depend on mechanical integrity, in particular those of the plasma membrane. As component and organizer of the plasma membrane, cholesterol is a regulator of cellular mechanical properties. Disruption of cholesterol balance leads to impairment of endothelial functions and eventually to disease. The mechanical properties of the membrane are strongly affected by the cytoskeleton. As Phosphatidylinositol-4,5-bisphosphate (PIP2) is a key mediator between the membrane and cytoskeleton, it also affects cellular biomechanical properties. Typically, PIP2 is concentrated in cholesterol-rich microdomains, such as caveolae and lipid rafts, which are particularly abundant in the endothelial plasma membrane. We investigated the connection between cholesterol and PIP2 by extracting membrane tethers from bovine aortic endothelial cells (BAEC) at different cholesterol levels and PIP2 conditions. Our results suggest that in BAEC the role of PIP2, as a mediator of membrane-cytoskeleton adhesion, is regulated by cholesterol. Our findings confirm the specific role of cholesterol in endothelial cells and may have implications for cholesterol-dependent vascular pathologies. PMID:23162471

  10. Endothelial antioxidant compound prolonged the endothelial antiapoptotic effects registered after tadalafil treatment in patients with arterial erectile dysfunction.

    PubMed

    La Vignera, Sandro; Condorelli, Rosita; Vicari, Enzo; D'Agata, Rosario; Calogero, Aldo E

    2012-01-01

    This study evaluated the effects of a chronic treatment with tadalafil, a specific phosphodiesterase V inhibitor, on endothelial apoptosis through changes in the serum concentration of endothelial microparticles (EMP). EMPs were arbitrarily chosen as a marker of endothelial apoptosis, and the changes in their concentration were monitored before and after treatment. Additionally, administration of endothelial antioxidant compound (EAC) during the follow-up, after discontinuation of tadalafil, was evaluated to determine whether this treatment improved the potential effects of tadalafil on the endothelium. Seventy-five patients with arterial erectile dysfunction were evaluated at baseline and after administration of tadalafil (5 mg once daily for 90 days). The International Index of Erectile Function questionnaire was administered, and penile dynamic Doppler and flow-cytometric (serum concentrations of EMPs) analyses were performed before (T0) and after treatment. Time points after tadalafil discontinuation: T1, after 1 week; T2, after 3 months; and T3, after 6 months. Three different schemes of follow-up were evaluated: group A, follow-up with EAC administration, after tadalafil discontinuation, for 6 months; group B, follow-up without other treatment; and group C, follow-up with placebo during the follow-up, after tadalafil cessation. The events CD45(neg)/CD144(pos)/annexinV(pos) were defined EMPs. Patients treated with tadalafil showed a significant decrease in serum EMPs 1 week after discontinuing tadalafil (16.4% ± 3.6% vs 7.1% ± 3.3%). This effect was maintained for up to 3 months in the group without other treatment during follow-up and was maintained for up to 6 months in the group treated with EAC during follow-up. Chronic treatment with tadalafil reduces endothelial apoptosis in patients with arterial erectile dysfunction. Further, EAC treatment prolongs and stabilizes the duration of antiapoptotic effects on the endothelium that are initially promoted by

  11. Association of Kidney Function with Changes in the Endothelial Surface Layer

    PubMed Central

    Dane, Martijn J.C.; Khairoun, Meriem; Lee, Dae Hyun; van den Berg, Bernard M.; Eskens, Bart J.M.; Boels, Margien G.S.; van Teeffelen, Jurgen W.G.E.; Rops, Angelique L.W.M.M.; van der Vlag, Johan; van Zonneveld, Anton Jan; Reinders, Marlies E.J.; Vink, Hans; Rabelink, Ton J.

    2014-01-01

    Background and objectives ESRD is accompanied by endothelial dysfunction. Because the endothelial glycocalyx (endothelial surface layer) governs interactions between flowing blood and the vessel wall, perturbation could influence disease progression. This study used a novel noninvasive sidestream–darkfield imaging method, which measures the accessibility of red blood cells to the endothelial surface layer in the microcirculation (perfused boundary region), to investigate whether renal function is associated with endothelial surface layer dimensions. Design, setting, participants, & measurements Perfused boundary region was measured in control participants (n=10), patients with ESRD (n=23), participants with normal kidney function after successful living donor kidney transplantation (n=12), and patients who developed interstitial fibrosis/tubular atrophy after kidney transplantation (n=10). In addition, the endothelial activation marker angiopoietin-2 and shed endothelial surface layer components syndecan-1 and soluble thrombomodulin were measured using ELISA. Results Compared with healthy controls (1.82±0.16 µm), ESRD patients had a larger perfused boundary region (+0.23; 95% confidence interval, 0.46 to <0.01; P<0.05), which signifies loss of endothelial surface layer dimensions. This large perfused boundary region was accompanied by higher circulating levels of syndecan-1 (+57.71; 95% confidence interval, 17.38 to 98.04; P<0.01) and soluble thrombomodulin (+12.88; 95% confidence interval, 0.29 to 25.46; P<0.001). After successful transplantation, the perfused boundary region was indistinguishable from healthy controls (without elevated levels of soluble thrombomodulin or syndecan-1). In contrast, however, patients who developed interstitial fibrosis and tubular atrophy showed a large perfused boundary region (+0.36; 95% confidence interval, 0.09 to 0.63; P<0.01) and higher levels of endothelial activation markers. In addition, a significant correlation

  12. Traumatic corneal endothelial rings from homemade explosives.

    PubMed

    Ng, Soo Khai; Rudkin, Adam K; Galanopoulos, Anna

    2013-08-01

    Traumatic corneal endothelial rings are remarkably rare ocular findings that may result from blast injury. We present a unique case of bilateral traumatic corneal endothelial rings secondary to blast injury from homemade explosives. PMID:23474743

  13. Ceramic subsurface marker prototypes

    SciTech Connect

    Lukens, C.E.

    1985-05-02

    The client submitted 5 sets of porcelain and stoneware subsurface (radioactive site) marker prototypes (31 markers each set). The following were determined: compressive strength, thermal shock resistance, thermal crazing resistance, alkali resistance, color retention, and chemical resistance.

  14. Changes in endothelial cell proliferation and vascular permeability after systemic lipopolysaccharide administration in the subfornical organ.

    PubMed

    Morita-Takemura, Shoko; Nakahara, Kazuki; Tatsumi, Kouko; Okuda, Hiroaki; Tanaka, Tatsuhide; Isonishi, Ayami; Wanaka, Akio

    2016-09-15

    The subfornical organ (SFO) has highly permeable fenestrated vasculature and is a key site for immune-to-brain communications. Recently, we showed the occurrence of continuous angiogenesis in the SFO. In the present study, we found that systemic administration of bacterial lipopolysaccharide (LPS) reduced the vascular permeability and endothelial cell proliferation. In LPS-administered mice, the SFO vasculature showed a significant decrease in the immunoreactivity of plasmalemma vesicle associated protein-1, a marker of endothelial fenestral diaphragms. These data suggest that vasculature undergoes structural change to decrease vascular permeability in response to systemic LPS administration. PMID:27609286

  15. Liver Sinusoidal Endothelial Cells Escape Senescence by Loss of p19ARF

    PubMed Central

    Koudelkova, Petra; Weber, Gerhard; Mikulits, Wolfgang

    2015-01-01

    Liver sinusoidal endothelial cells (LSECs) represent a highly differentiated cell type that lines hepatic sinusoids. LSECs form a discontinuous endothelium due to fenestrations under physiological conditions, which are reduced upon chronic liver injury. Cultivation of rodent LSECs associates with a rapid onset of stress-induced senescence a few days post isolation, which limits genetic and biochemical studies ex vivo. Here we show the establishment of LSECs isolated from p19ARF-/- mice which undergo more than 50 cell doublings in the absence of senescence. Isolated p19ARF-/- LSECs display a cobblestone-like morphology and show the ability of tube formation. Analysis of DNA content revealed a stable diploid phenotype after long-term passaging without a gain of aneuploidy. Notably, p19ARF-/- LSECs express the endothelial markers CD31, vascular endothelial growth factor receptor (VEGFR)-2, VE-cadherin, von Willebrand factor, stabilin-2 and CD146 suggesting that these cells harbor and maintain an endothelial phenotype. In line, treatment with small molecule inhibitors against VEGFR-2 caused cell death, demonstrating the sustained ability of p19ARF-/- LSECs to respond to anti-angiogenic therapeutics. From these data we conclude that loss of p19ARF overcomes senescence of LSECs, allowing immortalization of cells without losing endothelial characteristics. Thus, p19ARF-/- LSECs provide a novel cellular model to study endothelial cell biology. PMID:26528722

  16. 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. PMID:22609942

  17. The Soluble VEGF Receptor sFlt1 Contributes to Endothelial Dysfunction in CKD

    PubMed Central

    Marco, Giovana S. Di; Reuter, Stefan; Hillebrand, Uta; Amler, Susanne; König, Maximilian; Larger, Etienne; Oberleithner, Hans; Brand, Eva; Pavenstädt, Hermann

    2009-01-01

    Endothelial dysfunction contributes to the increased cardiovascular risk that accompanies CKD. We hypothesized that the soluble VEGF receptor 1 (sFlt-1), a VEGF antagonist, plays a role in endothelial dysfunction and decreased angiogenesis in CKD. We enrolled 130 patients with CKD stages 3 to 5 and 56 age- and gender-matched control patients. Plasma sFlt-1 levels were higher in patients with CKD and, after multivariate regression analyses, exclusively associated with renal function and levels of vWF, a marker of endothelial dysfunction. Compared with serum from control patients, both recombinant sFlt-1 and serum from patients with CKD had antiangiogenic activity in the chick chorioallantoic membrane (CAM) assay, induced endothelial cell apoptosis in vitro, and decreased nitric oxide generation in two different endothelial cell lines. Pretreating the sera with an antibody against sFlt-1 abrogated all of these effects. Furthermore, we observed increased sFlt1 levels in 5/6-nephrectomized rats compared with sham-operated animals. Finally, using real-time PCR and ELISA, we identified monocytes as a possible source of increased sFlt-1 in patients with CKD. Our findings show that excess sFlt-1 associates with endothelial dysfunction in CKD and suggest that increased sFlt-1 may predict cardiovascular risk in CKD. PMID:19608702

  18. Methylglyoxal-Induced Endothelial Cell Loss and Inflammation Contribute to the Development of Diabetic Cardiomyopathy.

    PubMed

    Vulesevic, Branka; McNeill, Brian; Giacco, Ferdinando; Maeda, Kay; Blackburn, Nick J R; Brownlee, Michael; Milne, Ross W; Suuronen, Erik J

    2016-06-01

    The mechanisms for the development of diabetic cardiomyopathy remain largely unknown. Methylglyoxal (MG) can accumulate and promote inflammation and vascular damage in diabetes. We examined if overexpression of the MG-metabolizing enzyme glyoxalase 1 (GLO1) in macrophages and the vasculature could reduce MG-induced inflammation and prevent ventricular dysfunction in diabetes. Hyperglycemia increased circulating inflammatory markers in wild-type (WT) but not in GLO1-overexpressing mice. Endothelial cell number was reduced in WT-diabetic hearts compared with nondiabetic controls, whereas GLO1 overexpression preserved capillary density. Neuregulin production, endothelial nitric oxide synthase dimerization, and Bcl-2 expression in endothelial cells was maintained in the hearts of GLO1-diabetic mice and corresponded to less myocardial cell death compared with the WT-diabetic group. Lower receptor for advanced glycation end products and tumor necrosis factor-α (TNF-α) levels were also observed in GLO1-diabetic versus WT-diabetic mice. Over a period of 8 weeks of hyperglycemia, GLO1 overexpression delayed and limited the loss of cardiac function. In vitro, MG and TNF-α were shown to synergize in promoting endothelial cell death, which was associated with increased angiopoietin 2 expression and reduced Bcl-2 expression. These results suggest that MG in diabetes increases inflammation, leading to endothelial cell loss. This contributes to the development of diabetic cardiomyopathy and identifies MG-induced endothelial inflammation as a target for therapy. PMID:26956489

  19. Retinal Endothelial Cell Apoptosis Stimulates Recruitment of Endothelial Progenitor Cells

    PubMed Central

    Bhatwadekar, Ashay D.; Glenn, Josephine V.; Curtis, Tim M.; Grant, Maria B.; Stitt, Alan W.; Gardiner, Tom A.

    2013-01-01

    Purpose Bone marrow–derived endothelial progenitor cells (EPCs) contribute to vascular repair although it is uncertain how local endothelial cell apoptosis influences their reparative function. This study was conducted to determine how the presence of apoptotic bodies at sites of endothelial damage may influence participation of EPCs in retinal microvascular repair. Methods Microlesions of apoptotic cell death were created in monolayers of retinal microvascular endothelial cells (RMECs) by using the photodynamic drug verteporfin. The adhesion of early-EPCs to these lesions was studied before detachment of the apoptotic cells or after their removal from the wound site. Apoptotic bodies were fed to normal RMECs and mRNA levels for adhesion molecules were analyzed. Results Endothelial lesions where apoptotic bodies were left attached at the wound site showed a fivefold enhancement in EPC recruitment (P < 0.05) compared with lesions where the apoptotic cells had been removed. In intact RMEC monolayers exposed to apoptotic bodies, expression of ICAM, VCAM, and E-selectin was upregulated by 5- to 15-fold (P < 0.05– 0.001). EPCs showed a characteristic chemotactic response (P < 0.05) to conditioned medium obtained from apoptotic bodies, whereas analysis of the medium showed significantly increased levels of VEGF, IL-8, IL-6, and TNF-α when compared to control medium; SDF-1 remained unchanged. Conclusions The data indicate that apoptotic bodies derived from retinal capillary endothelium mediate release of proangiogenic cytokines and chemokines and induce adhesion molecule expression in a manner that facilitates EPC recruitment. PMID:19474402

  20. A boost of BMP4 accelerates the commitment of human embryonic stem cells to the endothelial lineage.

    PubMed

    Goldman, Orit; Feraud, Olivier; Boyer-Di Ponio, Julie; Driancourt, Catherine; Clay, Denis; Le Bousse-Kerdiles, Marie-Caroline; Bennaceur-Griscelli, Annelise; Uzan, Georges

    2009-08-01

    Embryoid bodies (EBs) generated during differentiation of human embryonic stem cells (hESCs) contain vascular-like structures, suggesting that commitment of mesoderm progenitors into endothelial cells occurs spontaneously. We showed that bone morphogenetic protein 4 (BMP4), an inducer of mesoderm, accelerates the peak expression of CD133/kinase insert domain-containing receptor (KDR) and CD144/KDR. Because the CD133(+)KDR(+) population could represent endothelial progenitors, we sorted them at day 7 and cultured them in endothelial medium. These cells were, however, unable to differentiate into endothelial cells. Under standard conditions, the CD144(+)KDR(+) population represents up to 10% of the total cells at day 12. In culture, these cells, if sorted, give rise to a homogeneous population with a morphology typical of endothelial cells and express endothelial markers. These endothelial cells derived from the day 12 sorted population were functional, as assessed by different in vitro assays. When EBs were stimulated by BMP4, the CD144(+)KDR(+) peak was shifted to day 7. Most of these cells, however, were CD31(-), becoming CD31(+) in culture. They then expressed von Willebrand factor and were functional. This suggests that, initially, the BMP4-boosted day 7, CD144(+)KDR(+)CD31(-) population represents immature endothelial cells that differentiate into mature endothelial cells in culture. The expression of OCT3/4, a marker of immaturity for hESCs decreases during EB differentiation, decreasing faster following BMP4 induction. We also show that BMP4 inhibits the global expression of GATA2 and RUNX1, two transcription factors involved in hemangioblast formation, at day 7 and day 12. PMID:19544443

  1. Heterogeneity of endothelial cell phenotype within and amongst conduit vessels of the swine vasculature.

    PubMed

    Simmons, Grant H; Padilla, Jaume; Laughlin, M Harold

    2012-09-01

    The purpose of this study was to investigate the extent of endothelial cell phenotypic heterogeneity throughout the swine vasculature, with a focus on the conduit vessels of the arterial and venous circulations. We tested the hypothesis that atheroprone arteries exhibit higher expression of markers of inflammation and oxidative stress than do veins and atheroresistant arteries. The study sample included tissues from 79 castrated, male swine. Immediately after the animals were killed, endothelial cells were mechanically scraped from isolated segments of the thoracic and abdominal aorta, carotid, brachial, femoral and renal arteries, and the vein regionally associated with each of these vessels, as well as the internal mammary and right coronary arteries. Cells were also taken from two regions of the aortic arch contrasted by atheroprone versus atherosusceptible haemodynamics. Endothelial cell phenotype was assessed by either immunoblotting or quantitative real-time PCR for a host of both pro- and anti-atherogenic markers (e.g. endothelial nitric oxide synthase, p67phox, cyclo-oxygenase-1 and superoxide dismutase 1). Marked heterogeneity across the vasculature was observed in the expression of both pro- and anti-atherogenic markers, at both the protein and transcriptional levels. In particular, the coronary vascular endothelium expressed higher levels of the oxidative stress marker p67phox (P < 0.05 versus other arteries). In addition, differential expression of endothelial nitric oxide synthase and KLF4 was evident between atheroprone and atherosusceptible regions of the aorta, while expression of endothelial nitric oxide synthase, KLF2, KLF4 and cyclo-oxygenase-1 was lower in both areas of the aortic arch compared with the internal mammary artery. Conduit arteries typically expressed higher levels of both pro- and anti-atherogenic markers relative to their associated veins. We show, for the first time, that endothelial cell phenotype is variable within vessels

  2. Activation of the TGFβ pathway impairs endothelial to haematopoietic transition

    PubMed Central

    Vargel, Özge; Zhang, Yang; Kosim, Kinga; Ganter, Kerstin; Foehr, Sophia; Mardenborough, Yannicka; Shvartsman, Maya; Enright, Anton J.; Krijgsveld, Jeroen; Lancrin, Christophe

    2016-01-01

    The endothelial to haematopoietic transition (EHT) is a key developmental process where a drastic change of endothelial cell morphology leads to the formation of blood stem and progenitor cells during embryogenesis. As TGFβ signalling triggers a similar event during embryonic development called epithelial to mesenchymal transition (EMT), we hypothesised that TGFβ activity could play a similar role in EHT as well. We used the mouse embryonic stem cell differentiation system for in vitro recapitulation of EHT and performed gain and loss of function analyses of the TGFβ pathway. Quantitative proteomics analysis showed that TGFβ treatment during EHT increased the secretion of several proteins linked to the vascular lineage. Live cell imaging showed that TGFβ blocked the formation of round blood cells. Using gene expression profiling we demonstrated that the TGFβ signalling activation decreased haematopoietic genes expression and increased the transcription of endothelial and extracellular matrix genes as well as EMT markers. Finally we found that the expression of the transcription factor Sox17 was up-regulated upon TGFβ signalling activation and showed that its overexpression was enough to block blood cell formation. In conclusion we showed that triggering the TGFβ pathway does not enhance EHT as we hypothesised but instead impairs it. PMID:26891705

  3. Therapeutically targeting mitochondrial redox signalling alleviates endothelial dysfunction in preeclampsia.

    PubMed

    McCarthy, Cathal; Kenny, Louise C

    2016-01-01

    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. PMID:27604418

  4. Endothelial microparticles mediate inflammation-induced vascular calcification.

    PubMed

    Buendía, Paula; Montes de Oca, Addy; Madueño, Juan Antonio; Merino, Ana; Martín-Malo, Alejandro; Aljama, Pedro; Ramírez, Rafael; Rodríguez, Mariano; Carracedo, Julia

    2015-01-01

    Stimulation of endothelial cells (ECs) with TNF-α causes an increase in the expression of bone morphogenetic protein-2 (BMP-2) and the production of endothelial microparticles (EMPs). BMP-2 is known to produce osteogenic differentiation of vascular smooth muscle cells (VSMCs). It was found that EMPs from TNF-α-stimulated endothelial cells (HUVECs) contained a significant amount of BMP-2 and were able to enhance VSMC osteogenesis and calcification. Calcium content was greater in VSMCs exposed to EMPs from TNF-α-treated HUVECs than EMPs from nontreated HUVECs (3.56 ± 0.57 vs. 1.48 ± 0.56 µg/mg protein; P < 0.05). The increase in calcification was accompanied by up-regulation of Cbfa1 (osteogenic transcription factor) and down-regulation of SM22α (VSMC lineage marker). Inhibition of BMP-2 by small interfering RNA reduced the VSMC calcification induced by EMPs from TNF-α-treated HUVECs. Similar osteogenic capability was observed in EMPs from both patients with chronic kidney disease and senescent cells, which also presented a high level of BMP-2 expression. Labeling of EMPs with CellTracker shows that EMPs are phagocytized by VSMCs under all conditions (with or without high phosphate, control, and EMPs from TNF-α-treated HUVECs). Our data suggest that EC damage results in the release of EMPs with a high content of calcium and BMP-2 that are able to induce calcification and osteogenic differentiation of VSMCs. PMID:25342130

  5. Activation of the TGFβ pathway impairs endothelial to haematopoietic transition.

    PubMed

    Vargel, Özge; Zhang, Yang; Kosim, Kinga; Ganter, Kerstin; Foehr, Sophia; Mardenborough, Yannicka; Shvartsman, Maya; Enright, Anton J; Krijgsveld, Jeroen; Lancrin, Christophe

    2016-01-01

    The endothelial to haematopoietic transition (EHT) is a key developmental process where a drastic change of endothelial cell morphology leads to the formation of blood stem and progenitor cells during embryogenesis. As TGFβ signalling triggers a similar event during embryonic development called epithelial to mesenchymal transition (EMT), we hypothesised that TGFβ activity could play a similar role in EHT as well. We used the mouse embryonic stem cell differentiation system for in vitro recapitulation of EHT and performed gain and loss of function analyses of the TGFβ pathway. Quantitative proteomics analysis showed that TGFβ treatment during EHT increased the secretion of several proteins linked to the vascular lineage. Live cell imaging showed that TGFβ blocked the formation of round blood cells. Using gene expression profiling we demonstrated that the TGFβ signalling activation decreased haematopoietic genes expression and increased the transcription of endothelial and extracellular matrix genes as well as EMT markers. Finally we found that the expression of the transcription factor Sox17 was up-regulated upon TGFβ signalling activation and showed that its overexpression was enough to block blood cell formation. In conclusion we showed that triggering the TGFβ pathway does not enhance EHT as we hypothesised but instead impairs it. PMID:26891705

  6. Therapeutically targeting mitochondrial redox signalling alleviates endothelial dysfunction in preeclampsia

    PubMed Central

    McCarthy, Cathal; Kenny, Louise C.

    2016-01-01

    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. PMID:27604418

  7. Circulating Endothelial Microparticles: A Key Hallmark of Atherosclerosis Progression

    PubMed Central

    Panth, Nisha; Kim, Dong-Wook

    2016-01-01

    The levels of circulating microparticles (MPs) are raised in various cardiovascular diseases. Their increased level in plasma is regarded as a biomarker of alteration in vascular function. The prominent MPs present in blood are endothelial microparticles (EMPs) described as complex submicron (0.1 to 1.0 μm) vesicles like structure, released in response to endothelium cell activation or apoptosis. EMPs possess both physiological and pathological effects and may promote oxidative stress and vascular inflammation. EMPs release is triggered by inducer like angiotensin II, lipopolysaccharide, and hydrogen peroxide leading to the progression of atherosclerosis. However, there are multiple physiological pathways for EMPs generation like NADPH oxidase derived endothelial ROS formation, Rho kinase pathway, and mitogen-activated protein kinases. Endothelial dysfunction is a key initiating event in atherosclerotic plaque formation. Atheroemboli, resulting from ruptured carotid plaques, is a major cause of stroke. Increasing evidence suggests that EMPs play an important role in the pathogenesis of cardiovascular disease, acting as a marker of damage, either exacerbating disease progression or triggering a repair response. In this regard, it has been suggested that EMPs have the potential to act as biomarkers of disease status. This review aims to provide updated information of EMPs in relation to atherosclerosis pathogenesis. PMID:27066292

  8. Analyses of Endothelial Cells and Endothelial Progenitor Cells Released Microvesicles by Using Microbead and Q-dot Based Nanoparticle Tracking Analysis

    PubMed Central

    Wang, Jinju; Zhong, Yun; Ma, Xiaotang; Xiao, Xiang; Cheng, Chuanfang; Chen, Yusen; Iwuchukwu, Ifeanyi; Gaines, Kenneth J.; Bin Zhao; Liu, Shiming; Travers, Jeffrey B.; Bihl, Ji C.; Chen, Yanfang

    2016-01-01

    Accurate analysis of specific microvesicles (MVs) from biofluids is critical and challenging. Here we described novel methods to purify and detect MVs shed from endothelial cells (ECs) and endothelial progenitor cells (EPCs) by combining microbeads with fluorescence quantum dots (Q-dots) coupled nanoparticle tracking analysis (NTA). In the in vitro screening systems, we demonstrated that 1) anti-CD105 (EC marker) and anti-CD34 (EPC marker) conjugated-microbeads had the highest sensitivity and specificity for isolating respective MVs, which were confirmed with negative controls, CD41 and CD235a; 2) anti-CD144 (EC marker) and anti-KDR (EPC marker) conjugated-Q-dots exhibited the best sensitivity and specificity for their respective MV NTA detection, which were confirmed with positive control, anti-Annexin V (MV universal marker). The methods were further validated by their ability to efficiently recover the known amount of EC-MVs and EPC-MVs from particle-depleted plasma, and to detect the dynamical changes of plasma MVs in ischemic stroke patients, as compared with traditional flow cytometry. These novel methods provide ideal approaches for functional analysis and biomarker discovery of ECs- and EPCs- derived MVs. PMID:27094208

  9. Analysis of endothelial barrier function in vitro.

    PubMed

    Wang, Yuping; Alexander, J Steven

    2011-01-01

    Increased microvascular solute permeability underlies many forms of pathophysiological conditions, including inflammation. Endothelial monolayer cultures provide an excellent model system which allows systemic and mechanistic study of endothelial barrier function and paracellular permeability in vitro. The endothelial-specific complexus adherens junction protein VE-cadherin and their intracellular complex form pericellular structures along the cell borders which are critical to regulate endothelial barrier function by controlling pericellular permeability of vasculature. Here, we describe methods for both visualizing and quantifying junctional permeability and barrier changes in endothelial monolayers in vitro. PMID:21874457

  10. Endothelial cells in dengue hemorrhagic fever.

    PubMed

    Srikiatkhachorn, Anon; Kelley, James F

    2014-09-01

    Therapies to prevent or reverse endothelial dysfunction and vascular leak found in dengue hemorrhagic fever (DHF) have not been identified. In this review we summarize dengue viruses and the spectrum of human disease and highlight evidence of endothelial cell dysfunction in DHF based on studies in patients and mouse and tissue culture models. Evidence suggests that both virus antigen and host immune response, can cause endothelial cell dysfunction and weaken endothelial barrier integrity. We suggest possible therapeutic interventions and highlight how therapies targeting altered endothelial function might be evaluated in animal models and in patients with DHF. PMID:25025934

  11. Generation of functional endothelial-like cells from adult mouse germline-derived pluripotent stem cells.

    PubMed

    Kim, Julee; Eligehausen, Sarah; Stehling, Martin; Nikol, Sigrid; Ko, Kinarm; Waltenberger, Johannes; Klocke, Rainer

    2014-01-10

    Functional endothelial cells and their progenitors are required for vascular development, adequate vascular function, vascular repair and for cell-based therapies of ischemic diseases. Currently, cell therapy is limited by the low abundance of patient-derived cells and by the functional impairment of autologous endothelial progenitor cells (EPCs). In the present study, murine germline-derived pluripotent stem (gPS) cells were evaluated as a potential source for functional endothelial-like cells. Cells displaying an endothelial cell-like morphology were obtained from gPS cell-derived embryoid bodies using a combination of fluorescence-activated cell sorting (FACS)-based selection of CD31-positive cells and their subsequent cultivation on OP9 stromal cells in the presence of VEGF-A. Real-time reverse transcriptase polymerase chain reaction, FACS analysis and immunofluorescence staining showed that the gPS cell-derived endothelial-like cells (gPS-ECs) expressed endothelial cell-specific markers including von Willebrand Factor, Tie2, VEGFR2/Flk1, intercellular adhesion molecule 2 and vascular endothelial-cadherin. The high expression of ephrin B2, as compared to Eph B4 and VEGFR3, suggests an arterial rather than a venous or lymphatic differentiation. Their capability to take up Dil-conjugated acetylated low-density lipoprotein and to form capillary-like networks on matrigel confirmed their functionality. We conclude that gPS cells could be a novel source of endothelial cells potentially suitable for regenerative cell-based therapies for ischemic diseases. PMID:24333870

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

    SciTech Connect

    Muenzel, Daniela; Lehle, Karla Haubner, Frank; Schmid, Christof; Birnbaum, Dietrich E.; Preuner, Juergen G.

    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 even 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)

  13. The Essential Autophagy Gene ATG7 Modulates Organ Fibrosis via Regulation of Endothelial-to-Mesenchymal Transition*

    PubMed Central

    Singh, Krishna K.; Lovren, Fina; Pan, Yi; Quan, Adrian; Ramadan, Azza; Matkar, Pratiek N.; Ehsan, Mehroz; Sandhu, Paul; Mantella, Laura E.; Gupta, Nandini; Teoh, Hwee; Parotto, Matteo; Tabuchi, Arata; Kuebler, Wolfgang M.; Al-Omran, Mohammed; Finkel, Toren; Verma, Subodh

    2015-01-01

    Pulmonary fibrosis is a progressive disease characterized by fibroblast proliferation and excess deposition of collagen and other extracellular matrix components. Although the origin of fibroblasts is multifactorial, recent data implicate endothelial-to-mesenchymal transition as an important source of fibroblasts. We report herein that loss of the essential autophagy gene ATG7 in endothelial cells (ECs) leads to impaired autophagic flux accompanied by marked changes in EC architecture, loss of endothelial, and gain of mesenchymal markers consistent with endothelial-to-mesenchymal transition. Loss of ATG7 also up-regulates TGFβ signaling and key pro-fibrotic genes in vitro. In vivo, EC-specific ATG7 knock-out mice exhibit a basal reduction in endothelial-specific markers and demonstrate an increased susceptibility to bleomycin-induced pulmonary fibrosis and collagen accumulation. Our findings help define the role of endothelial autophagy as a potential therapeutic target to limit organ fibrosis, a condition for which presently there are no effective available treatments. PMID:25527499

  14. Endothelial Plasticity: Shifting Phenotypes through Force Feedback

    PubMed Central

    Krenning, Guido; Barauna, Valerio G.; Krieger, José E.; Harmsen, Martin C.; Moonen, Jan-Renier A. J.

    2016-01-01

    The endothelial lining of the vasculature is exposed to a large variety of biochemical and hemodynamic stimuli with different gradients throughout the vascular network. Adequate adaptation requires endothelial cells to be highly plastic, which is reflected by the remarkable heterogeneity of endothelial cells in tissues and organs. Hemodynamic forces such as fluid shear stress and cyclic strain are strong modulators of the endothelial phenotype and function. Although endothelial plasticity is essential during development and adult physiology, proatherogenic stimuli can induce adverse plasticity which contributes to disease. Endothelial-to-mesenchymal transition (EndMT), the hallmark of endothelial plasticity, was long thought to be restricted to embryonic development but has emerged as a pathologic process in a plethora of diseases. In this perspective we argue how shear stress and cyclic strain can modulate EndMT and discuss how this is reflected in atherosclerosis and pulmonary arterial hypertension. PMID:26904133

  15. Itinerary of high density lipoproteins in endothelial cells.

    PubMed

    Perisa, Damir; Rohrer, Lucia; Kaech, Andres; von Eckardstein, Arnold

    2016-02-01

    High density lipoprotein (HDL) and its main protein component apolipoprotein A-I (ApoA-I) have multiple anti-atherogenic functions. Some of them are exerted within the vessel wall, so that HDL needs to pass the endothelial barrier. To elucidate their itinerary through endothelial cells (ECs), we labelled ApoA-I and HDL either fluorescently or with 1.4 nm nanogold and investigated their cellular localization by using immunofluorescent microscopy (IFM) and electron microscopy (EM). HDL as well as ApoA-I is taken up by ECs into the same route of intracellular trafficking. Time kinetics and pulse chase experiments revealed that HDL is trafficked through different vesicles. HDL partially co-localized with LDL, albumin, and transferrin. HDL did not co-localize with clathrin and caveolin-1. Fluorescent HDL was recovered at small proportions in early endosomes and endosome to trans-golgi network vesicles but not at all in recycling endosomes, in late endosomes or lysosomes. EM identified HDL mainly in large filled vesicles which however upon IFM did not colocalize with markers of multivesicular bodies or autophagosomes. The uptake or cellular distribution of HDL was altered upon pharmacological interference with cytochalasine D, colchicine and dynasore. Blockage of fluid phase uptake with Amiloride or EIPA did not reduce the uptake of HDL. Neither did we observe any co-localization of HDL with dextran as the marker of fluid phase uptake. In conclusion, HDL and ApoA-I are internalized and trafficked by endothelial cells through a non-classical endocytic route. PMID:26577406

  16. Immunohistochemical expression of vascular endothelial growth factor and vascular endothelial growth factor receptor-2 in canine simple mammary gland adenocarcinomas

    PubMed Central

    Al-Dissi, Ahmad N.; Haines, Deborah M.; Singh, Baljit; Kidney, Beverly A.

    2010-01-01

    The expression of 5 markers associated with angiogenesis, proliferation, and apoptosis was studied in 26 canine simple mammary gland adenocarcinomas (SMGAs). The adenocarcinomas were graded histologically, and tissue sections were immunohistochemically stained for the expression of vascular endothelial growth factor (VEGF), VEGF receptor-2 (VEGFR-2), intra-tumor microvessel density, and tumor proliferation (PI) using antibodies against VEGF, VEGFR-2, von Willebrand factor, and Ki-67 antigen, respectively. Apoptotic indices (AI) were determined by an apoptosis assay. Markers VEGF and VEGFR-2 were detected in 96% and 100% of SMGAs, respectively. A high correlation between histologic grade and PI (r = 0.73), a moderate correlation between VEGF and histologic grade (r = 0.33), and between VEGF and PI (r = 0.42) were found. There was a significant difference in median PI among the 3 histologic grade groups (r < 0.05). Vascular endothelial growth factor may stimulate tumor cell proliferation through an autocrine loop, since VEGF and VEGFR-2 were expressed in most tumors. PMID:21197202

  17. Therapeutic transdifferentiation of human fibroblasts into endothelial cells using forced expression of lineage-specific transcription factors

    PubMed Central

    Wong, Wing Tak; Cooke, John P

    2016-01-01

    Transdifferentiation is the direct conversion from one somatic cell type into another desired somatic cell type. This reprogramming method offers an attractive approach for regenerative medicine. Here, we demonstrate that neonatal fibroblasts can be transdifferentiated into endothelial cells using only four endothelial transcription factors, namely, ETV2, FLI1, GATA2, and KLF4. We observed a significant up-regulation of endothelial genes including KDR, CD31, CD144, and vWF in human neonatal foreskin (BJ) fibroblasts infected with the lentiviral construct encoding the open reading frame of the four transcription factors. We observed morphological changes in BJ fibroblasts from the fibroblastic spindle shape into a more endothelial-like cobblestone structures. Fluorescence-activated cell sorting analysis revealed that ~16% of the infected cells with the lentiviral constructs encoding 4F expressed CD31. The sorted cells were allowed to expand for 2 weeks and these cells were immunostained and found to express endothelial markers CD31. The induced endothelial cells also incorporated fluorescence-labeled acetylated low-density lipoprotein and efficiently formed capillary-like networks when seeded on Matrigel. These results suggested that the induced endothelial cells were functional in vitro. Taken together, we successfully demonstrated the direct conversion of human neonatal fibroblasts into endothelial cells by transduction of lentiviral constructs encoding endothelial lineage-specific transcription factors ETV2, FLI1, GATA2, and KLF4. The directed differentiation of fibroblasts into endothelial cells may have significant utility in diseases characterized by fibrosis and loss of microvasculature. PMID:27081470

  18. Endothelial dysfunction: a comprehensive appraisal

    PubMed Central

    Esper, Ricardo J; Nordaby, Roberto A; Vilariño, Jorge O; Paragano, Antonio; Cacharrón, José L; Machado, Rogelio A

    2006-01-01

    The endothelium is a thin monocelular layer that covers all the inner surface of the blood vessels, separating the circulating blood from the tissues. It is not an inactive organ, quite the opposite. It works as a receptor-efector organ and responds to each physical or chemical stimulus with the release of the correct substance with which it may maintain vasomotor balance and vascular-tissue homeostasis. It has the property of producing, independently, both agonistic and antagonistic substances that help to keep homeostasis and its function is not only autocrine, but also paracrine and endocrine. In this way it modulates the vascular smooth muscle cells producing relaxation or contraction, and therefore vasodilatation or vasoconstriction. The endothelium regulating homeostasis by controlling the production of prothrombotic and antithrombotic components, and fibrynolitics and antifibrynolitics. Also intervenes in cell proliferation and migration, in leukocyte adhesion and activation and in immunological and inflammatory processes. Cardiovascular risk factors cause oxidative stress that alters the endothelial cells capacity and leads to the so called endothelial "dysfunction" reducing its capacity to maintain homeostasis and leads to the development of pathological inflammatory processes and vascular disease. There are different techniques to evaluate the endothelium functional capacity, that depend on the amount of NO produced and the vasodilatation effect. The percentage of vasodilatation with respect to the basal value represents the endothelial functional capacity. Taking into account that shear stress is one of the most important stimulants for the synthesis and release of NO, the non-invasive technique most often used is the transient flow-modulate "endothelium-dependent" post-ischemic vasodilatation, performed on conductance arteries such as the brachial, radial or femoral arteries. This vasodilatation is compared with the vasodilatation produced by drugs that

  19. 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. PMID:26782021

  20. [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. PMID:26596115

  1. Endothelial Cells Derived From Nuclear Reprogramming

    PubMed Central

    Wong, Wing Tak; Huang, Ngan F.; Botham, Crystal M.; Sayed, Nazish; Cooke, John P.

    2012-01-01

    The endothelium plays a pivotal role in vascular homeostasis, regulating the tone of the vascular wall, and its interaction with circulating blood elements. Alterations in endothelial functions facilitate the infiltration of inflammatory cells and permit vascular smooth muscle proliferation and platelet aggregation. Therefore, endothelial dysfunction is an early event in disease processes including atherosclerosis, and because of its critical role in vascular health the endothelium is worthy of the intense focus it has received. However, there are limitations to studying human endothelial function in vivo, or human vascular segments ex vivo. Thus, methods for endothelial cell culture have been developed and refined. More recently, methods to derive endothelial cells from pluripotent cells have extended the scientific range of human endothelial cell studies. Pluripotent stem cells may be generated, expanded and then differentiated into endothelial cells for in vitro studies. Constructs for molecular imaging can also be employed to facilitate tracking these cells in vivo. Furthermore, one can generate patient-specific endothelial cells to study the effects of genetic or epigenetic alterations on endothelial behavior. Finally, there is the opportunity to apply these cells for vascular therapy. This review focuses on the generation of endothelial cells from stem cells; their characterization by genetic, histological and functional studies; and their translational applications. PMID:23104878

  2. Mussel-inspired immobilization of vascular endothelial growth factor (VEGF) for enhanced endothelialization of vascular grafts.

    PubMed

    Shin, Young Min; Lee, Yu Bin; Kim, Seok Joo; Kang, Jae Kyeong; Park, Jong-Chul; Jang, Wonhee; Shin, Heungsoo

    2012-07-01

    Most polymeric vascular prosthetic materials have low patency rate for replacement of small diameter vessels (<5 mm), mainly due to failure to generate healthy endothelium. In this study, we present polydopamine-mediated immobilization of growth factors on the surface of polymeric materials as a versatile tool to modify surface characteristics of vascular grafts potentially for accelerated endothelialization. Polydopamine was deposited on the surface of biocompatible poly(L-lactide-co-ε-caprolactone) (PLCL) elastomer, on which vascular endothelial growth factor (VEGF) was subsequently immobilized by simple dipping. Surface characteristics and composition were investigated by using scanning electron microscopy, atomic force microscopy, and X-ray photoelectron spectroscopy. Immobilization of VEGF on the polydopamine-deposited PLCL films was effective (19.8 ± 0.4 and 197.4 ± 19.7 ng/cm(2) for DPv20 and DPv200 films, respectively), and biotin-mediated labeling of immobilized VEGF revealed that the fluorescence intensity increased as a function of the concentration of VEGF solution. The effect of VEGF on adhesion of HUVECs was marginal, which may have been masked by polydopamine layer that also enhanced cell adhesion. However, VEGF-immobilized substrate significantly enhanced proliferation of HUVECs for over 7 days of in vitro culture and also improved their migration. In addition, immobilized VEGF supported robust cell to cell interactions with strong expression of CD 31 marker. The same process was effective for immobilization of basic fibroblast growth factor, demonstrating the robustness of polydopamine layer for secondary ligation of growth factors as a simple and novel surface modification strategy for vascular graft materials. PMID:22617001

  3. In vitro effects of waterpipe smoke condensate on endothelial cell function: A potential risk factor for vascular disease

    PubMed Central

    Rammah, Mayyasa; Dandachi, Farah; Salman, Rola; Shihadeh, Alan; El-Sabban, Marwan

    2013-01-01

    Aim Despite its increasing popularity, little is known about the health effects of waterpipe smoking (WPS), particularly on the cardiovascular system. To investigate the role of WPS as a risk factor for vascular disease, we evaluated its effect on endothelial cell function, which is an early event in vascular disease pathogenesis. We assessed the changes in cell viability, ROS generation, inflammatory and vasodilatory markers and in vitro angiogenesis of human aortic endothelial cells in response to waterpipe smoke condensate exposure. Methods and results Mainstream waterpipe smoke condensate (WSC) was generated using a standard laboratory machine protocol. Compared to control, WSC induced cell cycle arrest, apoptosis, and oxidative stress in human primary endothelial cells. In addition, we assayed for impaired endothelium-dependent vasodilation and induced inflammation by studying the effect of WPS on the content and activity of AMPK, eNOS proteins and NF-κB p65 ser536 phosphorylation, respectively. WSC inhibited AMPK/eNOS phosphorylation and induced phosphorylation of p65. Moreover, we evaluated endothelial cells repair mechanism related properties that include migration/invasion and in vitro tube formation upon treatment with WSC. WSC reduced the motility and inhibited angiogenic potential of HAEC cells. Conclusions WPS induced endothelial cell dysfunction as evident by exerting oxidative stress, inflammation, and impaired endothelial vasodilatory function and repair mechanisms. All together these data provide evidence for the potential contribution of WPS to endothelial dysfunction and thus to vascular disease. PMID:23454654

  4. Long-Term Exposure to AZT, but not d4T, Increases Endothelial Cell Oxidative Stress and Mitochondrial Dysfunction

    PubMed Central

    Kline, Erik R.; Bassit, Leda; Hernandez-Santiago, Brenda I.; Detorio, Mervi A.; Liang, Bill; Kleinhenz, Dean J.; Walp, Erik R.; Dikalov, Sergey; Jones, Dean P.; Schinazi, Raymond F.

    2009-01-01

    Nucleoside reverse transcriptase inhibitors (NRTIs), such as zidovudine (AZT) and stavudine (d4T), cause toxicities to numerous tissues, including the liver and vasculature. While much is known about hepatic NRTI toxicity, the mechanism of toxicity in endothelial cells is incompletely understood. Human aortic endothelial and HepG2 liver cells were exposed to 1 μM AZT or d4T for up to 5 weeks. Markers of oxidative stress, mitochondrial function, NRTI phosphorylation, mitochondrial DNA (mtDNA) levels, and cytotoxicity were monitored over time. In endothelial cells, AZT significantly oxidized glutathione redox potential, increased total cellular and mitochondrial-specific superoxide, decreased mitochondrial membrane potential, increased lactate release, and caused cell death from weeks 3 through 5. Toxicity occurred in the absence of di- and tri-phosphorylated AZT and mtDNA depletion. These data show that oxidative stress and mitochondrial dysfunction in endothelial cells occur with a physiologically relevant concentration of AZT, and require long-term exposure to develop. In contrast, d4T did not induce endothelial oxidative stress, mitochondrial dysfunction, or cytotoxicity despite the presence of d4T-triphosphate. Both drugs depleted mtDNA in HepG2 cells without causing cell death. Endothelial cells are more susceptible to AZT-induced toxicity than HepG2 cells, and AZT caused greater endothelial dysfunction than d4T because of its pro-oxidative effects. PMID:19067249

  5. Engineering an Endothelialized Vascular Graft: A Rational Approach to Study Design in a Non-Human Primate Model

    PubMed Central

    Anderson, Deirdre E. J.; Glynn, Jeremy J.; Song, Howard K.; Hinds, Monica T.

    2014-01-01

    After many years of research, small diameter, synthetic vascular grafts still lack the necessary biologic integration to perform ideally in clinical settings. Endothelialization of vascular grafts has the potential to improve synthetic graft function, and endothelial outgrowth cells (EOCs) are a promising autologous cell source. Yet no work has established the link between endothelial cell functions and outcomes of implanted endothelialized grafts. This work utilized steady flow, oscillatory flow, and tumor necrosis factor stimulation to alter EOC phenotype and enable the formulation of a model to predict endothelialized graft performance. To accomplish this, EOC in vitro expression of coagulation and inflammatory markers was quantified. In parallel, in non-human primate (baboon) models, the platelet and fibrinogen accumulation on endothelialized grafts were quantified in an ex vivo shunt, or the tissue ingrowth on implanted grafts were characterized after 1mth. Oscillatory flow stimulation of EOCs increased in vitro coagulation markers and ex vivo platelet accumulation. Steady flow preconditioning did not affect platelet accumulation or intimal hyperplasia relative to static samples. To determine whether in vitro markers predict implant performance, a linear regression model of the in vitro data was fit to platelet accumulation data—correlating the markers with the thromboprotective performance of the EOCs. The model was tested against implant intimal hyperplasia data and found to correlate strongly with the parallel in vitro analyses. This research defines the effects of flow preconditioning on EOC regulation of coagulation in clinical vascular grafts through parallel in vitro, ex vivo, and in vivo analyses, and contributes to the translatability of in vitro tests to in vivo clinical graft performance. PMID:25526637

  6. Flow-Mediated Endothelial Mechanotransduction

    PubMed Central

    Davies, Peter F.

    2011-01-01

    Mechanical forces associated with blood flow play important roles in the acute control of vascular tone, the regulation of arterial structure and remodeling, and the localization of atherosclerotic lesions. Major regulation of the blood vessel responses occurs by the action of hemodynamic shear stresses on the endothelium. The transmission of hemodynamic forces throughout the endothelium and the mechanotransduction mechanisms that lead to biophysical, biochemical, and gene regulatory responses of endothelial cells to hemodynamic shear stresses are reviewed. PMID:7624393

  7. Innovative Flow Cytometry Allows Accurate Identification of Rare Circulating Cells Involved in Endothelial Dysfunction

    PubMed Central

    Boraldi, Federica; Bartolomeo, Angelica; De Biasi, Sara; Orlando, Stefania; Costa, Sonia; Cossarizza, Andrea; Quaglino, Daniela

    2016-01-01

    Introduction Although rare, circulating endothelial and progenitor cells could be considered as markers of endothelial damage and repair potential, possibly predicting the severity of cardiovascular manifestations. A number of studies highlighted the role of these cells in age-related diseases, including those characterized by ectopic calcification. Nevertheless, their use in clinical practice is still controversial, mainly due to difficulties in finding reproducible and accurate methods for their determination. Methods Circulating mature cells (CMC, CD45-, CD34+, CD133-) and circulating progenitor cells (CPC, CD45dim, CD34bright, CD133+) were investigated by polychromatic high-speed flow cytometry to detect the expression of endothelial (CD309+) or osteogenic (BAP+) differentiation markers in healthy subjects and in patients affected by peripheral vascular manifestations associated with ectopic calcification. Results This study shows that: 1) polychromatic flow cytometry represents a valuable tool to accurately identify rare cells; 2) the balance of CD309+ on CMC/CD309+ on CPC is altered in patients affected by peripheral vascular manifestations, suggesting the occurrence of vascular damage and low repair potential; 3) the increase of circulating cells exhibiting a shift towards an osteoblast-like phenotype (BAP+) is observed in the presence of ectopic calcification. Conclusion Differences between healthy subjects and patients with ectopic calcification indicate that this approach may be useful to better evaluate endothelial dysfunction in a clinical context. PMID:27560136

  8. Endothelial dysfunction and oxidative stress in polycystic kidney disease.

    PubMed

    Klawitter, Jelena; Reed-Gitomer, Berenice Y; McFann, Kim; Pennington, Alexander; Klawitter, Jost; Abebe, Kaleab Z; Klepacki, Jacek; Cadnapaphornchai, Melissa A; Brosnahan, Godela; Chonchol, Michel; Christians, Uwe; Schrier, Robert W

    2014-12-01

    Cardiovascular disease (CVD) is the leading cause of premature mortality in ADPKD patients. The aim was to identify potential serum biomarkers associated with the severity of ADPKD. Serum samples from a homogenous group of 61 HALT study A ADPKD patients [early disease group with estimated glomerular filtration rate (eGFR) >60 ml·min(-1)·1.73 m(-2)] were compared with samples from 49 patients from the HALT study B group with moderately advanced disease (eGFR 25-60 ml·min(-1)·1.73 m(-2)). Targeted tandem-mass spectrometry analysis of markers of endothelial dysfunction and oxidative stress was performed and correlated with eGFR and total kidney volume normalized to the body surface area (TKV/BSA). ADPKD patients with eGFR >60 ml·min(-1)·1.73 m(-2) showed higher levels of CVD risk markers asymmetric and symmetric dimethylarginine (ADMA and SDMA), homocysteine, and S-adenosylhomocysteine (SAH) compared with the healthy controls. Upon adjustments for age, sex, systolic blood pressure, and creatinine, SDMA, homocysteine, and SAH remained negatively correlated with eGFR. Resulting cellular methylation power [S-adenosylmethionine (SAM)/SAH ratio] correlated with the reduction of renal function and increase in TKV. Concentrations of prostaglandins (PGs), including oxidative stress marker 8-isoprostane, as well as PGF2α, PGD₂, and PGE₂, were markedly elevated in patients with ADPKD compared with healthy controls. Upon adjustments for age, sex, systolic blood pressure, and creatinine, increased PGD₂ and PGF₂α were associated with reduced eGFR, whereas 8-isoprostane and again PGF₂α were associated with an increase in TKV/BSA. Endothelial dysfunction and oxidative stress are evident early in ADPKD patients, even in those with preserved kidney function. The identified pathways may provide potential therapeutic targets for slowing down the disease progression. PMID:25234311

  9. [Interpretation of hemostatic and fibrinolytic markers].

    PubMed

    Asakura, Hidesaku

    2011-10-01

    Blood dose not normally coagulate in the blood vessels covered with endothelial cells, because these cells contain some substances responsible for antithrombotic action such as thrombomodulin, heparin-like substance, prostacyclin, nitric oxide and tissue plasminogen activator. Most important role of blood coagulation is hemostasis. Blood can coagulate in two ways: intrinsic coagulation pathway and extrinsic coagulation pathway that is activated by negatively charged substances and FVIIa-tissue-factor (TF) complex, respectively. Prothrombin time(PT) can represent extrinsic pathway, while activated partial thromboplastin time (APTT) can represent intrinsic pathway. PT is prolonged in such diseases as vitamin K deficiency, hepatic failure and warfarin intake, while APTT is prolonged such diseases as hemophilia A & B, von Willebrand disease and lupus anticoagulant. Cross mixing test is very useful to assess prolonged clotting time. FDP means fibrin/fibrinogen degradation products and D-dimer is the smallest products of fibrin degradation. These markers are often used to diagnose disseminated intravascular coagulation (DIC) and deep vein thrombosis (DVT). Thrombin-antithrombin complex (TAT) and plasmin-alpha2 plasmin inhibitor (PIC) can be used to evaluate the extent of coagulation and fibrinolysis activation, respectively. These two markers is essential for classify the pathophysiology of DIC: DIC with suppressed fibrinolysis, enhanced fibrinolysis or balanced fibrinolysis. In conclusion, exact interpretation of hemostatic and fibrinolytic markers is one of the most important abilities in clinical situation. PMID:22184880

  10. Expression of Endothelial Nitric Oxide Synthase and Endothelin-1 in Skin Tissue from Amputated Limbs of Patients with Complex Regional Pain Syndrome

    PubMed Central

    Groeneweg, J. George; Antonissen, Claudia Heijmans; Huygen, Frank J. P. M.; Zijlstra, Freek J.

    2008-01-01

    Background and Objectives. Impaired microcirculation during the chronic stage of complex regional pain syndrome (CRPS) is related to increased vasoconstriction, tissue hypoxia, and metabolic tissue acidosis in the affected limb. Endothelial dysfunction is suggested to be the main cause of diminished blood flow. The aim of this study was to examine the distribution of endothelial nitric oxide synthase (eNOS) and endothelin-1(ET-1) relative to vascular density represented by the endothelial marker CD31-immunoreactivity in the skin tissue of patients with chronic CRPS. Methods. We performed immunohistochemical staining on sections of skin specimens obtained from the amputated limbs (one arm and one leg) of two patients with CRPS. Results. In comparison to proximal specimens we found an increased number of migrated endothelial cells as well as an increase of eNOS activity in distal dermis specimens. Conclusions. We found indications that endothelial dysfunction plays a role in chronic CRPS. PMID:18663383

  11. Endothelial CD276 (B7-H3) expression is increased in human malignancies and distinguishes between normal and tumour-derived circulating endothelial cells

    PubMed Central

    Kraan, J; van den Broek, P; Verhoef, C; Grunhagen, D J; Taal, W; Gratama, J W; Sleijfer, S

    2014-01-01

    Background: Mature circulating endothelial cells (CEC) are surrogate markers of endothelial damage. CEC measured in patients with advanced cancer are thought not only to derive from damaged normal vasculature (n-CEC), but also from damaged (t-CEC). Therefore, assays that allow the discrimination between these two putative types of CEC are thought to improve the specificity of the enumeration of CEC in cancer. Methods: Identification of tumour-associated endothelial markers (TEM) by comparing antigen expression on normal vs t-CEC and assess the presence of t-CEC in peripheral blood of cancer patients by incorporating TEM in our novel flow cytometry-based CEC detection assay. Results: No difference in antigen expression between normal and malignant endothelial cells (ECs) was found for CD54, CD109, CD137, CD141, CD144 and CXCR7. In contrast, overexpression for CD105, CD146, CD276 and CD309 was observed in tumour ECs compared with normal ECs. CD276 was most differentially expressed and chosen as a marker for further investigation. CD276-expressing CEC were significantly higher in 15 patients with advanced colorectal cancer (median 9 (range 1–293 cell per 4 ml); P<0.005), in 83 patients with a glioblastoma multiforme (median 10 (range 0–804); P<0.0001) and in 14 patients with advanced breast cancer (median 14 (range 0–390) P<0.05) as compared with 24 healthy individuals (median 3 (range 0–11)). Of all patients with malignancies, 58% had CD276+ CEC counts above the ULN (8 cell per 4 ml). Conclusions: The present study shows that CD276 can be used to discriminate ECs from malignant tissue from ECs from normal tissue. In addition, CD276+ CEC do occur in higher frequencies in patients with advanced cancer. PMID:24892449

  12. Polyphenol-rich Cranberry Juice has a neutral effect on endothelial function but decreases the fraction of osteocalcin expressing endothelial progenitor cells

    PubMed Central

    Flammer, Andreas J.; Martin, Elizabeth; Gossl, Mario; Widmer, R Jay; Lennon, Ryan; Sexton, Jasmine A.; Loeffler, Darrell; Khosla, Sundeep; Lerman, Lilach O.; Lerman, Amir

    2014-01-01

    Purpose Cranberry juice (CJ) contains a remarkably high concentration of polyphenols, considered to be beneficial for cardiovascular and bone health. The current double-blind, randomized study was designed to test whether daily consumption of double-strength Ocean Spray light CJ (2×230ml) over 4 months has beneficial effects on vascular function and on endothelial progenitor cells (EPCs), EPCs carrying the osteoblastic marker osteocalcin in particular. Methods 84 participants (49.5±16.2yrs.) with peripheral endothelial dysfunction and cardiovascular risk factors were enrolled in this double-blind, randomized, controlled trial (69 completed the four month protocol - 32 in the CJ group and 37 in the placebo group - respectively). Vascular responses to reactive hyperemia were measured non-invasively by peripheral arterial tonometry (EndoPAT). Peripheral blood mononuclear cells were stained for EPC markers, as well as osteocalcin, and counted by flow-cytometry. Results Baseline characteristics were similar in both groups. The effect of CJ on peripheral endothelial function and on circulating EPC counts (CD34+/CD133+/KDR+) did not change during the study. A high percentage of EPCs expressed osteocalcin (59.4±35.7%). CJ, as compared to placebo, induced a decrease in the fraction of EPCs expressing osteocalcin (-8.64±48.98 and 19.13±46.11%, respectively, p=0.019). Systemic levels of the adhesion marker ICAM correlated significantly with the number of EPCs expressing osteocalcin. Conclusions The study demonstrated that long term supplementation of polyphenol-rich CJ did not improve peripheral endothelial function. However, the decrease in the fraction of osteocalcin+ EPCs suggests a potential beneficial effect of polyphenol-rich CJ. PMID:22382203

  13. Novel regulators of endothelial barrier function

    PubMed Central

    Ravindran, Krishnan; Kuebler, Wolfgang M.

    2014-01-01

    Endothelial barrier function is an essential and tightly regulated process that ensures proper compartmentalization of the vascular and interstitial space, while allowing for the diffusive exchange of small molecules and the controlled trafficking of macromolecules and immune cells. Failure to control endothelial barrier integrity results in excessive leakage of fluid and proteins from the vasculature that can rapidly become fatal in scenarios such as sepsis or the acute respiratory distress syndrome. Here, we highlight recent advances in our understanding on the regulation of endothelial permeability, with a specific focus on the endothelial glycocalyx and endothelial scaffolds, regulatory intracellular signaling cascades, as well as triggers and mediators that either disrupt or enhance endothelial barrier integrity, and provide our perspective as to areas of seeming controversy and knowledge gaps, respectively. PMID:25381026

  14. Replication of human endothelial cells in culture.

    PubMed

    Lewis, L J; Hoak, J C; Maca, R D; Fry, G L

    1973-08-01

    Investigative studies dealing with the properties and functions of endothelial cells have been hampered because there has been little or no success in the isolation, growth, and passage of individual cells in large numbers. We have developed a system whereby pure cultures of endothelial cells derived from umbilical veins can be subcultured for at least five serial passages. Many facets of endothelial function and interaction can be evaluated with the use of this new adaptive system of isolation and culture. PMID:4718112

  15. Calcineurin inhibitors cyclosporine A and tacrolimus induce vascular inflammation and endothelial activation through TLR4 signaling.

    PubMed

    Rodrigues-Diez, Raquel; González-Guerrero, Cristian; Ocaña-Salceda, Carlos; Rodrigues-Diez, Raúl R; Egido, Jesús; Ortiz, Alberto; Ruiz-Ortega, Marta; Ramos, Adrián M

    2016-01-01

    The introduction of the calcineurin inhibitors (CNIs) cyclosporine and tacrolimus greatly reduced the rate of allograft rejection, although their chronic use is marred by a range of side effects, among them vascular toxicity. In transplant patients, it is proved that innate immunity promotes vascular injury triggered by ischemia-reperfusion damage, atherosclerosis and hypertension. We hypothesized that activation of the innate immunity and inflammation may contribute to CNI toxicity, therefore we investigated whether TLR4 mediates toxic responses of CNIs in the vasculature. Cyclosporine and tacrolimus increased the production of proinflammatory cytokines and endothelial activation markers in cultured murine endothelial and vascular smooth muscle cells as well as in ex vivo cultures of murine aortas. CNI-induced proinflammatory events were prevented by pharmacological inhibition of TLR4. Moreover, CNIs were unable to induce inflammation and endothelial activation in aortas from TLR4(-/-) mice. CNI-induced cytokine and adhesion molecules synthesis in endothelial cells occurred even in the absence of calcineurin, although its expression was required for maximal effect through upregulation of TLR4 signaling. CNI-induced TLR4 activity increased O2(-)/ROS production and NF-κB-regulated synthesis of proinflammatory factors in cultured as well as aortic endothelial and VSMCs. These data provide new insight into the mechanisms associated with CNI vascular inflammation. PMID:27295076

  16. Phosphorylation inactivation of endothelial nitric oxide synthesis in pulmonary arterial hypertension.

    PubMed

    Ghosh, Sudakshina; Gupta, Manveen; Xu, Weiling; Mavrakis, Deloris A; Janocha, Allison J; Comhair, Suzy A A; Haque, Mohammad Mahfuzul; Stuehr, Dennis J; Yu, Jun; Polgar, Peter; Naga Prasad, Sathyamangla V; Erzurum, Serpil C

    2016-06-01

    The impairment of vasodilator nitric oxide (NO) production is well accepted as a typical marker of endothelial dysfunction in vascular diseases, including in the pathophysiology of pulmonary arterial hypertension (PAH), but the molecular mechanisms accounting for loss of NO production are unknown. We hypothesized that low NO production by pulmonary arterial endothelial cells in PAH is due to inactivation of NO synthase (eNOS) by aberrant phosphorylation of the protein. To test the hypothesis, we evaluated eNOS levels, dimerization, and phosphorylation in the vascular endothelial cells and lungs of patients with PAH compared with controls. In mechanistic studies, eNOS activity in endothelial cells in PAH lungs was found to be inhibited due to phosphorylation at T495. Evidence pointed to greater phosphorylation/activation of protein kinase C (PKC) α and its greater association with eNOS as the source of greater phosphorylation at T495. The presence of greater amounts of pT495-eNOS in plexiform lesions in lungs of patients with PAH confirmed the pathobiological mechanism in vivo. Transfection of the activating mutation of eNOS (T495A/S1177D) restored NO production in PAH cells. Pharmacological blockade of PKC activity by β-blocker also restored NO formation by PAH cells, identifying one mechanism by which β-blockers may benefit PAH and cardiovascular diseases through recovery of endothelial functions. PMID:27130529

  17. Effect of Vitamins C and E on Endothelial Function in Type 1 Diabetes Mellitus

    PubMed Central

    Cazeau, Rachel-Marie; Huang, Hong; Bauer, John A.; Hoffman, Robert P.

    2016-01-01

    Background/Objectives. Endothelial dysfunction due to hyperglycemia-induced oxidative damage is an important predictor of future cardiovascular risk in patients with type 1 diabetes mellitus (T1DM) and is present in adolescent T1DM. We hypothesized that combined treatment with the antioxidant vitamins C and E might improve endothelial function (EF) and other biochemical risk factors in adolescents with T1DM. Subjects/Methods. Open-label antioxidant supplementation was given for six weeks with endpoint measurements collected at baseline and study completion. Endpoints measured included EF and plasma measurements of biochemical endothelial risk. Results. Two males and 7 females were studied. Mean age was 12.9 ± 0.9 yrs; mean T1DM duration was 5.5 ± 2.5 yrs; mean BMI was 22.1 ± 3.8 kg/m2; and mean hemoglobin A1c was 9.3 ± 1.1%. No differences were found for EF, high sensitivity CRP, total antioxidant capacity, adiponectin, or endothelial progenitor cells (EPCs) between before and after combined vitamin C and E therapy. Conclusions. Our negative study results do not support previous findings of decreased oxidative damage, improved endothelial function, and increased vascular repair capacity with antioxidant therapy. Longer term studies may be needed to determine the effects, if any, of combined antioxidant therapy on EPCs, EF, and markers of micro- and macrovascular complications in T1DM. PMID:26783536

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

    PubMed

    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

  19. 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

  20. Calcineurin inhibitors cyclosporine A and tacrolimus induce vascular inflammation and endothelial activation through TLR4 signaling

    PubMed Central

    Rodrigues-Diez, Raquel; González-Guerrero, Cristian; Ocaña-Salceda, Carlos; Rodrigues-Diez, Raúl R.; Egido, Jesús; Ortiz, Alberto; Ruiz-Ortega, Marta; Ramos, Adrián M.

    2016-01-01

    The introduction of the calcineurin inhibitors (CNIs) cyclosporine and tacrolimus greatly reduced the rate of allograft rejection, although their chronic use is marred by a range of side effects, among them vascular toxicity. In transplant patients, it is proved that innate immunity promotes vascular injury triggered by ischemia-reperfusion damage, atherosclerosis and hypertension. We hypothesized that activation of the innate immunity and inflammation may contribute to CNI toxicity, therefore we investigated whether TLR4 mediates toxic responses of CNIs in the vasculature. Cyclosporine and tacrolimus increased the production of proinflammatory cytokines and endothelial activation markers in cultured murine endothelial and vascular smooth muscle cells as well as in ex vivo cultures of murine aortas. CNI-induced proinflammatory events were prevented by pharmacological inhibition of TLR4. Moreover, CNIs were unable to induce inflammation and endothelial activation in aortas from TLR4−/− mice. CNI-induced cytokine and adhesion molecules synthesis in endothelial cells occurred even in the absence of calcineurin, although its expression was required for maximal effect through upregulation of TLR4 signaling. CNI-induced TLR4 activity increased O2−/ROS production and NF-κB-regulated synthesis of proinflammatory factors in cultured as well as aortic endothelial and VSMCs. These data provide new insight into the mechanisms associated with CNI vascular inflammation. PMID:27295076

  1. Does obstructive sleep apnea cause endothelial dysfunction? A critical review of the literature.

    PubMed

    Hoyos, Camilla M; Melehan, Kerri L; Liu, Peter Y; Grunstein, Ronald R; Phillips, Craig L

    2015-04-01

    Endothelial dysfunction is characterized by impaired endothelium-dependent vasodilatation and is an independent predictor of adverse cardiovascular consequences. The ease with which endothelial function can be assessed has led to it becoming a useful marker of cardiovascular diseases in research studies. Obstructive sleep apnea (OSA) has been independently associated with endothelial dysfunction which may explain the increased risk for cardiovascular and all-cause mortality in this population. One possible mechanism for the development of endothelial dysfunction in OSA is through the cyclical pattern of hypoxia and re-oxygenation. This creates a haemostatic imbalance in which nitric oxide bio-availability is reduced and pro-inflammatory and pro-thrombotic forces prevail. Furthermore the repair capacity of the endothelium to protect itself against this increased damage is diminished. All of these pathways contribute to vascular disease which ultimately gives rise to adverse cardiovascular consequences. This review aims to provide a critical appraisal of the cross-sectional and interventional studies which have investigated micro- and macro-vascular endothelial dysfunction in OSA with emphasis on randomised controlled studies. PMID:25088969

  2. Endothelial Cell Growth and Differentiation on Collagen-Immobilized Polycaprolactone Nanowire Surfaces.

    PubMed

    Leszczak, Victoria; Baskett, Dominique A; Popat, Ketul C

    2015-06-01

    The success of cardiovascular implants is associated with the development of an endothelium on material surface, critical to the prevention of intimal hyperplasia, calcification and thrombosis. A thorough understanding of the interaction between vascular endothelial cells and the biomaterial involved is essential in order to have a successful application which promotes healing and regeneration through integration with native tissue. In this study, we have developed collagen immobilized nanostructured surfaces with controlled arrays of high aspect ratio nanowires for the growth and maintenance of human microvascular endothelial cells (HMVECs). The nanowire surfaces were fabricated from polycaprolactone using a novel nanotemplating technique, and were immobilized with collagen utilizing an aminolysis method. The collagen immobilized nanowire surfaces were characterized using contact angle measurements, scanning electron microscopy and X-ray photoelectron spectroscopy. Human microvascular endothelial cells were used to evaluate the efficacy of the collagen immobilized nanowire surfaces to promote cell adhesion, proliferation, viability and differentiation. The results presented here indicate significantly higher cellular adhesion, proliferation and viability on nanowire and collagen immobilized surfaces as compared to the control surface. Further, HMVECs have a more elongated body and low shape factor on nanostructured surfaces. The differentiation potential of collagen immobilized nanowire surfaces was also evaluated by immunostaining and western blotting for key endothelial cell markers that are expressed when human microvascular endothelial cells are differentiated. Results indicate that expression of VE-cadherin is increased on collagen immobilized surfaces while the expression of von Willebrand factor is statistically similar on all surfaces. PMID:26353596

  3. Donor origin of circulating endothelial progenitors after allogeneic bone marrow transplantation.

    PubMed

    Ikpeazu, C; Davidson, M K; Halteman, D; Browning, P J; Brandt, S J

    2000-01-01

    Endothelial cell precursors circulate in blood and express antigens found on hematopoietic stem cells, suggesting that such precursors might be subject to transplantation. To investigate, we obtained adherence-depleted peripheral blood mononuclear cells from 3 individuals who had received a sex-mismatched allogeneic bone marrow transplant (BMT) and cultured the cells on fibronectin-coated plates with endothelial growth factors. The phenotype of the spindle-shaped cells that emerged in culture was characterized by immunofluorescent staining, and the origin of the cells was determined using a polymerase chain reaction (PCR)-based assay for polymorphic short tandem repeats (STRs). The cells manifested a number of endothelial characteristics-such as von Wlllebrand factor, CD31, and Flk-1/KDR expression; Bandeiraea simplicifolia lectin 1 binding; and acetylated low-density lipoprotein uptake-but lacked expression of certain markers of activation or differentiation, including intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and the epitope for the anti-endothelial cell antibody P1H12. For each patient and at all time points studied (ranging from 5 to 52 months after transplantation), STR-PCR analysis showed that cultured cells and nucleated blood cells came exclusively from the bone marrow donor. These results demonstrate that circulating endothelial progenitors are both transplantable and capable of long-term repopulation of human allogeneic BMT recipients. PMID:10905767

  4. Cardiomyocytes induce endothelial cells to trans-differentiate into cardiac muscle: implications for myocardium regeneration.

    PubMed

    Condorelli, G; Borello, U; De Angelis, L; Latronico, M; Sirabella, D; Coletta, M; Galli, R; Balconi, G; Follenzi, A; Frati, G; Cusella De Angelis, M G; Gioglio, L; Amuchastegui, S; Adorini, L; Naldini, L; Vescovi, A; Dejana, E; Cossu, G

    2001-09-11

    The concept of tissue-restricted differentiation of postnatal stem cells has been challenged by recent evidence showing pluripotency for hematopoietic, mesenchymal, and neural stem cells. Furthermore, rare but well documented examples exist of already differentiated cells in developing mammals that change fate and trans-differentiate into another cell type. Here, we report that endothelial cells, either freshly isolated from embryonic vessels or established as homogeneous cells in culture, differentiate into beating cardiomyocytes and express cardiac markers when cocultured with neonatal rat cardiomyocytes or when injected into postischemic adult mouse heart. Human umbilical vein endothelial cells also differentiate into cardiomyocytes under similar experimental conditions and transiently coexpress von Willebrand factor and sarcomeric myosin. In contrast, neural stem cells, which efficiently differentiate into skeletal muscle, differentiate into cardiomyocytes at a low rate. Fibroblast growth factor 2 and bone morphogenetic protein 4, which activate cardiac differentiation in embryonic cells, do not activate cardiogenesis in endothelial cells or stimulate trans-differentiation in coculture, suggesting that different signaling molecules are responsible for cardiac induction during embryogenesis and in successive periods of development. The fact that endothelial cells can generate cardiomyocytes sheds additional light on the plasticity of endothelial cells during development and opens perspectives for cell autologous replacement therapies. PMID:11535818

  5. Cardiomyocytes induce endothelial cells to trans-differentiate into cardiac muscle: Implications for myocardium regeneration

    PubMed Central

    Condorelli, G.; Borello, U.; De Angelis, L.; Latronico, M.; Sirabella, D.; Coletta, M.; Galli, R.; Balconi, G.; Follenzi, A.; Frati, G.; Cusella De Angelis, M. G.; Gioglio, L.; Amuchastegui, S.; Adorini, L.; Naldini, L.; Vescovi, A.; Dejana, E.; Cossu, G.

    2001-01-01

    The concept of tissue-restricted differentiation of postnatal stem cells has been challenged by recent evidence showing pluripotency for hematopoietic, mesenchymal, and neural stem cells. Furthermore, rare but well documented examples exist of already differentiated cells in developing mammals that change fate and trans-differentiate into another cell type. Here, we report that endothelial cells, either freshly isolated from embryonic vessels or established as homogenous cells in culture, differentiate into beating cardiomyocytes and express cardiac markers when cocultured with neonatal rat cardiomyocytes or when injected into postischemic adult mouse heart. Human umbilical vein endothelial cells also differentiate into cardiomyocytes under similar experimental conditions and transiently coexpress von Willebrand factor and sarcomeric myosin. In contrast, neural stem cells, which efficiently differentiate into skeletal muscle, differentiate into cardiomyocytes at a low rate. Fibroblast growth factor 2 and bone morphogenetic protein 4, which activate cardiac differentiation in embryonic cells, do not activate cardiogenesis in endothelial cells or stimulate trans-differentiation in coculture, suggesting that different signaling molecules are responsible for cardiac induction during embryogenesis and in successive periods of development. The fact that endothelial cells can generate cardiomyocytes sheds additional light on the plasticity of endothelial cells during development and opens perspectives for cell autologous replacement therapies. PMID:11535818

  6. Soluble adhesion molecules correlate with surface expression in an in vitro model of endothelial activation.

    PubMed

    Kjaergaard, Anders G; Dige, Anders; Krog, Jan; Tønnesen, Else; Wogensen, Lise

    2013-10-01

    Endothelial activation is a pivotal event in the development and progression of inflammation. Central to endothelial activation is the up-regulation of cellular adhesion molecules (CAMs) including E-selectin (CD62E), ICAM-1 (CD54), VCAM-1 (CD106) and PECAM-1 (CD31). These CAMs are also found in soluble forms (sCAMs). In this in vitro study of endothelial activation, we examined whether the levels of sCAMs correlate with the endothelial surface expression of CAMs in a dose-dependent and time-dependent manner. Such a correlation would support the use of sCAMs as surrogate markers for endothelial activation in inflammatory conditions. Human umbilical vein endothelial cells (HUVEC) were cultured with various concentrations of TNF-α for 8 hr and at a fixed concentration of TNF-α for various durations. The levels of soluble and surface-bound E-selectin, ICAM-1, VCAM-1 and PECAM-1 were quantified by flow cytometry. TNF-α stimulation increased CAM and sCAM expression in a dose-dependent and time-dependent manner. There was a significant positive correlation between the levels of ICAM-1 and sICAM-1 and between the levels of VCAM and sVCAM-1 in both the dose-response and time-response experiments. A positive correlation between the levels of E-selectin and sE-selectin was observed in the time-response experiment. This study supports the use of sCAMs as potential biomarkers of endothelial activation. In particular, the use of sICAM-1, sVCAM-1 and sE-selectin seems promising. PMID:23724832

  7. Plasticity of endothelial cells during arterial-venous differentiation in the avian embryo.

    PubMed

    Moyon, D; Pardanaud, L; Yuan, L; Bréant, C; Eichmann, A

    2001-09-01

    Remodeling of the primary vascular system of the embryo into arteries and veins has long been thought to depend largely on the influence of hemodynamic forces. This view was recently challenged by the discovery of several molecules specifically expressed by arterial or venous endothelial cells. We here analysed the expression of neuropilin-1 and TIE2, two transmembrane receptors known to play a role in vascular development. In birds, neuropilin-1 was expressed by arterial endothelium and wall cells, but absent from veins. TIE2 was strongly expressed in embryonic veins, but only weakly transcribed in most arteries. To examine whether endothelial cells are committed to an arterial or venous fate once they express these specific receptors, we constructed quail-chick chimeras. The dorsal aorta, carotid artery and the cardinal and jugular veins were isolated together with the vessel wall from quail embryos between embryonic day 2 to 15 and grafted into the coelom of chick hosts. Until embryonic day 7, all grafts yielded endothelial cells that colonized both host arteries and veins. After embryonic day 7, endothelial plasticity was progressively lost and from embryonic day 11 grafts of arteries yielded endothelial cells that colonized only chick arteries and rarely reached the host veins, while grafts of jugular veins colonized mainly host veins. When isolated from the vessel wall, quail aortic endothelial cells from embryonic day 11 embryos were able to colonize both host arteries and veins. Our results show that despite the expression of arterial or venous markers the endothelium remains plastic with regard to arterial-venous differentiation until late in embryonic development and point to a role for the vessel wall in endothelial plasticity and vessel identity. PMID:11546752

  8. 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

  9. Progenitor endothelial cell involvement in Alzheimer's disease

    SciTech Connect

    Budinger, Thomas F.

    2003-05-01

    There is compelling evidence that endothelial cells of the brain and periphery are dysfunctional in Alzheimer's Disease. There is evidence for a fundamental defect in, or abnormal aging of, endothelial progenitor cells in atherosclerosis. The possibility that endothelial cell defects are a primary cause for Alzheimer's Disease or other dementias can be researched by molecular and cell biology studies as well as cell trafficking studies using recently demonstrated molecular imaging methods. The evidence for abnormal endothelial function and the methods to explore this hypothesis are presented.

  10. Endothelial function and coronary artery disease.

    PubMed

    Kinlay, S; Libby, P; Ganz, P

    2001-08-01

    The endothelium produces a number of vasodilator and vasoconstrictor substances that not only regulate vasomotor tone, but also the recruitment and activity of inflammatory cells and the propensity towards thrombosis. Endothelial vasomotor function is a convenient way to assess these other functions, and is related to the long-term risk of cardiovascular disease. Lipids (particularly low density lipoprotein cholesterol) and oxidant stress play a major role in impairing these functions, by reducing the bioavailability of nitric oxide and activating pro-inflammatory signalling pathways such as nuclear factor kappa B. Biomechanical forces on the endothelium, including low shear stress from disturbed blood flow, also activate the endothelium increasing vasomotor dysfunction and promoting inflammation by upregulating pro-atherogenic genes. In contrast, normal laminar shear stress promotes the expression of genes that may protect against atherosclerosis. The sub-cellular structure of endothelial cells includes caveolae that play an integral part in regulating the activity of endothelial nitric oxide synthase. Low density lipoprotein cholesterol and oxidant stress impair caveolae structure and function and adversely affect endothelial function. Lipid-independent pathways of endothelial cell activation are increasingly recognized, and may provide new therapeutic targets. Endothelial vasoconstrictors, such as endothelin, antagonize endothelium-derived vasodilators and contribute to endothelial dysfunction. Some but not all studies have linked certain genetic polymorphisms of the nitric oxide synthase enzyme to vascular disease and impaired endothelial function. Such genetic heterogeneity may nonetheless offer new insights into the variability of endothelial function. PMID:11507322

  11. Vascular endothelial growth factor C promotes survival of retinal vascular endothelial cells via vascular endothelial growth factor receptor‐2

    PubMed Central

    Zhao, Bojun; Smith, Gill; Cai, Jun; Ma, Aihua; Boulton, Mike

    2007-01-01

    Aim To determine vascular endothelial growth factor C (VEGF‐C) expression in retinal endothelial cells, its antiapoptotic potential and its putative role in diabetic retinopathy. Method Cultured retinal endothelial cells and pericytes were exposed to tumour necrosis factor (TNF)α and VEGF‐C expression determined by reverse transcriptase‐polymerase chain reaction. Secreted VEGF‐C protein levels in conditioned media from endothelial cells were examined by western blotting analysis. The ability of VEGF‐C to prevent apoptosis induced by TNFα or hyperglycaemia in endothelial cells was assessed by flow cytometry. The expression of VEGF‐C in diabetic retinopathy was studied by immunohistochemistry of retinal tissue. Result VEGF‐C was expressed by both vascular endothelial cells and pericytes. TNFα up regulated both VEGF‐C and vascular endothelial growth factor receptor‐2 (VEGFR)‐2 expression in endothelial cells in a dose‐dependent manner, but had no effect on VEGFR‐3. Flow cytometry results showed that VEGF‐C prevented endothelial cell apoptosis induced by TNFα and hyperglycaemia and that the antiapoptotic effect was mainly via VEGFR‐2. In pericytes, the expression of VEGF‐C mRNA remained stable on exogenous TNFα treatment. VEGF‐C immunostaining was increased in retinal vessels in specimens with diabetes compared with retinal specimens from controls without diabetes. Conclusion In retinal endothelial cells, TNFα stimulates the expression of VEGF‐C, which in turn protects endothelial cells from apoptosis induced by TNFα or hyperglycaemia via VEGFR‐2 and thus helps sustain retinal neovascularisation. PMID:16943230

  12. 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. PMID:24710004

  13. Endotoxin Induces Fibrosis in Vascular Endothelial Cells through a Mechanism Dependent on Transient Receptor Protein Melastatin 7 Activity

    PubMed Central

    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 endothelial­to­mesenchymal 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. PMID:24710004

  14. Human liver endothelial cells, but not macrovascular or microvascular endothelial cells, engraft in the mouse liver.

    PubMed

    Filali, Ebtisam El; Hiralall, Johan K; van Veen, Henk A; Stolz, Donna B; Seppen, Jurgen

    2013-01-01

    Liver cell transplantation has had limited clinical success so far, partly due to poor engraftment of hepatocytes. Instead of hepatocytes. other cell types, such as endothelial cells, could be used in ex vivo liver gene therapy. The goal of the present study was to compare the grafting and repopulation capacity of human endothelial cells derived from various tissues. Human endothelial cells were isolated from adult and fetal livers using anti-human CD31 antibody-conjugated magnetic beads. Human macrovascular endothelial cells were obtained from umbilical vein. Human microvascular endothelial cells were isolated from adipose tissue. Cells were characterized using flow cytometry. Liver engraftment and repopulation of endothelial cells was studied after intrasplenic transplantation in monocrotaline-treated immunodeficient mice. Following transplantation, human liver endothelial cells engrafted throughout the mouse liver. With immunoscanning electron microscopy, fenestrae in engrafted human liver endothelial cells were identified, a characteristic feature of liver sinusoidal endothelial cells. In contrast, CD31-negative liver cells, human macrovascular and microvascular endothelial cells were not capable of repopulating mouse liver. Characterization of human liver, macrovascular, and microvascular endothelial cells demonstrated expression of CD31, CD34, and CD146 but not CD45. Our study shows that only human liver endothelial cells, but not macro- and microvascular endothelial cells, have the unique capacity to engraft and repopulate the mouse liver. These results indicate that mature endothelial cells cannot transdifferentiate in vivo and thus do not exhibit phenotypic plasticity. Our results have set a basis for further research to the potential of human liver endothelial cells in liver-directed cell and gene therapy. PMID:23044355

  15. Ectodomain Shedding of Lymphatic Vessel Endothelial Hyaluronan Receptor 1 (LYVE-1) Is Induced by Vascular Endothelial Growth Factor A (VEGF-A).

    PubMed

    Nishida-Fukuda, Hisayo; Araki, Ryoichi; Shudou, Masachika; Okazaki, Hidenori; Tomono, Yasuko; Nakayama, Hironao; Fukuda, Shinji; Sakaue, Tomohisa; Shirakata, Yuji; Sayama, Koji; Hashimoto, Koji; Detmar, Michael; Higashiyama, Shigeki; Hirakawa, Satoshi

    2016-05-13

    Lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), a type I transmembrane glycoprotein, is known as one of the most specific lymphatic vessel markers in the skin. In this study, we found that the ectodomain of LYVE-1 undergoes proteolytic cleavage, and this process produces soluble LYVE-1. We further identified the cleavage site for ectodomain shedding and generated an uncleavable mutant of LYVE-1. In lymphatic endothelial cells, ectodomain shedding of LYVE-1 was induced by vascular endothelial growth factor (VEGF)-A, an important factor for angiogenesis and lymphangiogenesis under pathological conditions. VEGF-A-induced LYVE-1 ectodomain shedding was mediated via the extracellular signal-regulated kinase (ERK) and a disintegrin and metalloproteinase (ADAM) 17. Wild-type LYVE-1, but not uncleavable LYVE-1, promoted migration of lymphatic endothelial cells in response to VEGF-A. Immunostaining analyses in human psoriasis skin lesions and VEGF-A transgenic mouse skin suggested that the ectodomain shedding of LYVE-1 occurred in lymphatic vessels undergoing chronic inflammation. These results indicate that the ectodomain shedding of LYVE-1 might be involved in promoting pathological lymphangiogenesis. PMID:26966180

  16. 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.

  17. Atherogenic Cytokines Regulate VEGF-A-Induced Differentiation of Bone Marrow-Derived Mesenchymal Stem Cells into Endothelial Cells.

    PubMed

    Ikhapoh, Izuagie Attairu; Pelham, Christopher J; Agrawal, Devendra K

    2015-01-01

    Coronary artery stenting or angioplasty procedures frequently result in long-term endothelial dysfunction or loss and complications including arterial thrombosis and myocardial infarction. Stem cell-based therapies have been proposed to support endothelial regeneration. Mesenchymal stem cells (MSCs) differentiate into endothelial cells (ECs) in the presence of VEGF-A in vitro. Application of VEGF-A and MSC-derived ECs at the interventional site is a complex clinical challenge. In this study, we examined the effect of atherogenic cytokines (IL-6, TNFα, and Ang II) on EC differentiation and function. MSCs (CD44(+), CD73(+), CD90(+), CD14(-), and CD45(-)) were isolated from the bone marrow of Yucatan microswine. Naïve MSCs cultured in differentiation media containing VEGF-A (50 ng/mL) demonstrated increased expression of EC-specific markers (vWF, PECAM-1, and VE-cadherin), VEGFR-2 and Sox18, and enhanced endothelial tube formation. IL-6 or TNFα caused a dose-dependent attenuation of EC marker expression in VEGF-A-stimulated MSCs. In contrast, Ang II enhanced EC marker expression in VEGF-A-stimulated MSCs. Addition of Ang II to VEGF-A and IL-6 or TNFα was sufficient to rescue the EC phenotype. Thus, Ang II promotes but IL-6 and TNFα inhibit VEGF-A-induced differentiation of MSCs into ECs. These findings have important clinical implications for therapies intended to increase cardiac vascularity and reendothelialize coronary arteries following intervention. PMID:26106428

  18. Vascular endothelial dysfunction and pharmacological treatment

    PubMed Central

    Su, Jin Bo

    2015-01-01

    The endothelium exerts multiple actions involving regulation of vascular permeability and tone, coagulation and fibrinolysis, inflammatory and immunological reactions and cell growth. Alterations of one or more such actions may cause vascular endothelial dysfunction. Different risk factors such as hypercholesterolemia, homocystinemia, hyperglycemia, hypertension, smoking, inflammation, and aging contribute to the development of endothelial dysfunction. Mechanisms underlying endothelial dysfunction are multiple, including impaired endothelium-derived vasodilators, enhanced endothelium-derived vasoconstrictors, over production of reactive oxygen species and reactive nitrogen species, activation of inflammatory and immune reactions, and imbalance of coagulation and fibrinolysis. Endothelial dysfunction occurs in many cardiovascular diseases, which involves different mechanisms, depending on specific risk factors affecting the disease. Among these mechanisms, a reduction in nitric oxide (NO) bioavailability plays a central role in the development of endothelial dysfunction because NO exerts diverse physiological actions, including vasodilation, anti-inflammation, antiplatelet, antiproliferation and antimigration. Experimental and clinical studies have demonstrated that a variety of currently used or investigational drugs, such as angiotensin-converting enzyme inhibitors, angiotensin AT1 receptors blockers, angiotensin-(1-7), antioxidants, beta-blockers, calcium channel blockers, endothelial NO synthase enhancers, phosphodiesterase 5 inhibitors, sphingosine-1-phosphate and statins, exert endothelial protective effects. Due to the difference in mechanisms of action, these drugs need to be used according to specific mechanisms underlying endothelial dysfunction of the disease. PMID:26635921

  19. How mental stress affects endothelial function.

    PubMed

    Toda, Noboru; Nakanishi-Toda, Megumi

    2011-12-01

    Mental stress is an important factor contributing to recognized mechanisms underlying cardiovascular events. Among these, stress-related endothelial dysfunction is an early risk factor that predicts future development of severe cardiovascular disorders. Acute mental stress by a variety of tests impairs endothelial function in humans, although the opposite results have been reported by some investigators. Chronic stress always deteriorates endothelial function in humans and experimental animals. Stress hormones, such as glucocorticoids and pro-inflammatory cytokines, and endothelin-1 liberated in response to mental stress participate in endothelial dysfunction possibly via downregulation of endothelial nitric oxide synthase (eNOS) expression, eNOS inactivation, decreased nitric oxide (NO) actions, and increased NO degradation, together with vasoconstriction counteracting against NO-induced vasodilatation. Catecholamines do not directly affect endothelial function but impair its function when blood pressure elevation by the amines is sustained. Endogenous opioids favorably affect endothelial function, which counteract deteriorating effects of other stress hormones and mediators. Inhibition of cortisol and endothelin-1 production, prevention of pro-inflammatory mediator accumulation, hypnotics, mirthful laughter, humor orientation, and lifestyle modification would contribute to the prevention and treatment for stress-related endothelial dysfunction and future serious cardiovascular disease. PMID:21947555

  20. Vascular endothelial dysfunction and pharmacological treatment.

    PubMed

    Su, Jin Bo

    2015-11-26

    The endothelium exerts multiple actions involving regulation of vascular permeability and tone, coagulation and fibrinolysis, inflammatory and immunological reactions and cell growth. Alterations of one or more such actions may cause vascular endothelial dysfunction. Different risk factors such as hypercholesterolemia, homocystinemia, hyperglycemia, hypertension, smoking, inflammation, and aging contribute to the development of endothelial dysfunction. Mechanisms underlying endothelial dysfunction are multiple, including impaired endothelium-derived vasodilators, enhanced endothelium-derived vasoconstrictors, over production of reactive oxygen species and reactive nitrogen species, activation of inflammatory and immune reactions, and imbalance of coagulation and fibrinolysis. Endothelial dysfunction occurs in many cardiovascular diseases, which involves different mechanisms, depending on specific risk factors affecting the disease. Among these mechanisms, a reduction in nitric oxide (NO) bioavailability plays a central role in the development of endothelial dysfunction because NO exerts diverse physiological actions, including vasodilation, anti-inflammation, antiplatelet, antiproliferation and antimigration. Experimental and clinical studies have demonstrated that a variety of currently used or investigational drugs, such as angiotensin-converting enzyme inhibitors, angiotensin AT1 receptors blockers, angiotensin-(1-7), antioxidants, beta-blockers, calcium channel blockers, endothelial NO synthase enhancers, phosphodiesterase 5 inhibitors, sphingosine-1-phosphate and statins, exert endothelial protective effects. Due to the difference in mechanisms of action, these drugs need to be used according to specific mechanisms underlying endothelial dysfunction of the disease. PMID:26635921

  1. Signaling hierarchy regulating human endothelial cell development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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 stud...

  2. Quantitation of Endothelial Cell Adhesiveness In Vitro

    PubMed Central

    Lowe, Donna J.; Raj, Kenneth

    2015-01-01

    One of the cardinal processes of inflammation is the infiltration of immune cells from the lumen of the blood vessel to the surrounding tissue. This occurs when endothelial cells, which line blood vessels, become adhesive to circulating immune cells such as monocytes. In vitro measurement of this adhesiveness has until now been done by quantifying the total number of monocytes that adhere to an endothelial layer either as a direct count or by indirect measurement of the fluorescence of adherent monocytes. While such measurements do indicate the average adhesiveness of the endothelial cell population, they are confounded by a number of factors, such as cell number, and do not reveal the proportion of endothelial cells that are actually adhesive. Here we describe and demonstrate a method which allows the enumeration of adhesive cells within a tested population of endothelial monolayer. Endothelial cells are grown on glass coverslips and following desired treatment are challenged with monocytes (that may be fluorescently labeled). After incubation, a rinsing procedure, involving multiple rounds of immersion and draining, the cells are fixed. Adhesive endothelial cells, which are surrounded by monocytes are readily identified and enumerated, giving an adhesion index that reveals the actual proportion of endothelial cells within the population that are adhesive. PMID:26132714

  3. An integrative microfluidically supported in vitro model of an endothelial barrier combined with cortical spheroids simulates effects of neuroinflammation in neocortex development.

    PubMed

    Raasch, Martin; Rennert, Knut; Jahn, Tobias; Gärtner, Claudia; Schönfelder, Gilbert; Huber, Otmar; Seiler, Andrea E M; Mosig, Alexander S

    2016-07-01

    The development of therapeutic substances to treat diseases of the central nervous system is hampered by the tightness and selectivity of the blood-brain barrier. Moreover, testing of potential drugs is time-consuming and cost-intensive. Here, we established a new microfluidically supported, biochip-based model of the brain endothelial barrier in combination with brain cortical spheroids suitable to detect effects of neuroinflammation upon disruption of the endothelial layer in response to inflammatory signals. Unilateral perfusion of the endothelial cell layer with a cytokine mix comprising tumor necrosis factor, IL-1β, IFNγ, and lipopolysaccharide resulted in a loss of endothelial von Willebrand factor and VE-cadherin expression accompanied with an increased leakage of the endothelial layer and diminished endothelial cell viability. In addition, cytokine treatment caused a loss of neocortex differentiation markers Tbr1, Tbr2, and Pax6 in the cortical spheroids concomitant with reduced cell viability and spheroid integrity. From these observations, we conclude that our endothelial barrier/cortex model is suitable to specifically reflect cytokine-induced effects on barrier integrity and to uncover damage and impairment of cortical tissue development and viability. With all its limitations, the model represents a novel tool to study cross-communication between the brain endothelial barrier and underlying cortical tissue that can be utilized for toxicity and drug screening studies focusing on inflammation and neocortex formation. PMID:27478526

  4. [MUSCULOSKELETAL MARKERS, ARTHROPATY, TRAUMAS].

    PubMed

    Caldarini, Carla; Zavaroni, Federica; Benassi, Valentina

    2015-01-01

    The bone tissue remodeling due to strong physical/working activity is defined as ergonomic markers or MSM (Muscoloskeletal Stress Markers) (Capasso et al. 1999) and MOS (Markers of Occupational Stress). Among them we can find: enthesopaties, arthropaties, non metrical stress and traumas markers. In the present study, the analysis of these traits has been used to clarify habitual activity patterns of four imperial populations from Suburbium: Castel Malnome, Casal Bertone area Q, Via Padre Semeria e Quarto Cappello del Prete. The very high prevalence of activity-induced stress lesions occurred among the individuals of Castel Malnome and Casal Bertone area Q suggests that these groups were involved in strenuous occupations such as, respectively: the processing and storage of salt and the dyeing of textiles and hides discernible from the archaeological context. For the individuals of Via Padre Semeria and Quarto Cappello del Prete the alterations, instead, could be compatibles with agricultural work. PMID:27348990

  5. [Biological markers of alcoholism].

    PubMed

    Marcos Martín, M; Pastor Encinas, I; Laso Guzmán, F J

    2005-09-01

    Diagnosis of alcoholism is very important, given its high prevalence and possibility of influencing the disease course. For this reason, the so-called biological markers of alcoholism are useful. These are analytic parameters that alter in the presence of excessive alcohol consumption. The two most relevant markers are the gamma-glutamyltranspeptidase and carbohydrate deficient transferrin. With this clinical comment, we aim to contribute to the knowledge of these tests and promote its use in the clinical practice. PMID:16194480

  6. 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

  7. Vascular Endothelial Growth Factor Receptor -2 in Breast Cancer

    PubMed Central

    Guo, Shanchun; Colbert, Laronna S.; Fuller, Miles; Zhang, Yuanyuan; Gonzalez-Perez, Ruben R.

    2010-01-01

    Investigations over the last decade have established the essential role of growth factors and their receptors during angiogenesis and carcinogenesis. The vascular endothelial growth factor receptor (VEGFR) family in mammals contains three members, VEGFR-1 (Flt-1), VEGFR-2 (KDR/Flk-1) and VEGFR-3 (Flt-4), which are transmembrane tyrosine kinase receptors that regulate the formation of blood and lymphatic vessels. In the early 1990s, the above VEGFR were structurally characterized by cDNA cloning. Among these three receptors, VEGFR-2 is generally recognized to have a principal role in mediating VEGF-induced responses. VEGFR-2 is considered as the earliest marker for endothelial cell development. Importantly, VEGFR-2 directly regulates tumor angiogenesis. Therefore, several inhibitors of VEGFR-2 have been developed and many of them are now in clinical trials. In addition to targeting endothelial cells, the VEGF/VEGFR-2 system works as an essential autocrine/paracrine process for cancer cell proliferation and survival. Recent studies mark the continuous and increased interest in this related, but distinct, function of VEGF/VEGFR-2 in cancer cells: the autocrine/paracrine loop. Several mechanisms regulate VEGFR-2 levels and modulate its role in tumor angiogenesis and physiologic functions, i.e.: cellular localization/trafficking, regulation of cis-elements of promoter, epigenetic regulation and signaling from Notch, cytokines/growth factors and estrogen, etc. In this review, we will focus on updated information regarding VEGFR-2 research with respect to the molecular mechanisms of VEGFR-2 regulation in human breast cancer. Investigations in the activation, function, and regulation of VEGFR-2 in breast cancer will allow the development of new pharmacological strategies aimed at directly targeting cancer cell proliferation and survival. PMID:20462514

  8. Endothelial Transmigration by Trypanosoma cruzi

    PubMed Central

    Coates, Bria M.; Sullivan, David P.; Makanji, Ming Y.; Du, Nga Y.; Olson, Cheryl L.; Muller, William A.; Engman, David M.; Epting, Conrad L.

    2013-01-01

    Chagas heart disease, the leading cause of heart failure in Latin America, results from infection with the parasite Trypanosoma cruzi. Although T. cruzi disseminates intravascularly, how the parasite contends with the endothelial barrier to escape the bloodstream and infect tissues has not been described. Understanding the interaction between T. cruzi and the vascular endothelium, likely a key step in parasite dissemination, could inform future therapies to interrupt disease pathogenesis. We adapted systems useful in the study of leukocyte transmigration to investigate both the occurrence of parasite transmigration and its determinants in vitro. Here we provide the first evidence that T. cruzi can rapidly migrate across endothelial cells by a mechanism that is distinct from productive infection and does not disrupt monolayer integrity or alter permeability. Our results show that this process is facilitated by a known modulator of cellular infection and vascular permeability, bradykinin, and can be augmented by the chemokine CCL2. These represent novel findings in our understanding of parasite dissemination, and may help identify new therapeutic strategies to limit the dissemination of the parasite. PMID:24312535

  9. Endothelial cell micropatterning: Methods, effects, and applications

    PubMed Central

    Anderson, Deirdre E.J.; Hinds, Monica T.

    2012-01-01

    The effects of flow on endothelial cells have been widely examined for the ability of fluid shear stress to alter cell morphology and function; however, the effects of endothelial cell morphology without flow have only recently been observed. An increase in lithographic techniques in cell culture spurred a corresponding increase in research aiming to confine cell morphology. These studies lead to a better understanding of how morphology and cytoskeletal configuration affect the structure and function of the cells. This review examines endothelial cell micropatterning research by exploring both the many alternative methods used to alter endothelial cell morphology and the resulting changes in cellular shape and phenotype. Micropatterning induced changes in endothelial cell proliferation, apoptosis, cytoskeletal organization, mechanical properties, and cell functionality. Finally, the ways these cellular manipulation techniques have been applied to biomedical engineering research, including angiogenesis, cell migration, and tissue engineering, is discussed. PMID:21761242

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

    SciTech Connect

    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. - 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.

  11. 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. PMID:27472163

  12. Early systemic sclerosis: marker autoantibodies and videocapillaroscopy patterns are each associated with distinct clinical, functional and cellular activation markers

    PubMed Central

    2013-01-01

    Introduction Early systemic sclerosis (SSc) is characterized by Raynaud's phenomenon together with scleroderma marker autoantibodies and/or a scleroderma pattern at capillaroscopy and no other distinctive feature of SSc. Patients presenting with marker autoantibodies plus a capillaroscopic scleroderma pattern seem to evolve into definite SSc more frequently than patients with either feature. Whether early SSc patients with only marker autoantibodies or capillaroscopic positivity differ in any aspect at presentation is unclear. Methods Seventy-one consecutive early SSc patients were investigated for preclinical cardiopulmonary alterations. Out of these, 44 patients and 25 controls affected by osteoarthritis or primary fibromyalgia syndrome were also investigated for serum markers of fibroblast (carboxyterminal propeptide of collagen I), endothelial (soluble E-selectin) and T-cell (soluble IL-2 receptor alpha) activation. Results Thirty-two of the 71 patients (45.1%) had both a marker autoantibody and a capillaroscopic scleroderma pattern (subset 1), 16 patients (22.5%) had only a marker autoantibody (subset 2), and 23 patients (32.4%) had only a capillaroscopic scleroderma pattern (subset 3). Patients with marker autoantibodies (n = 48, 67.6%) had a higher prevalence of impaired diffusing lung capacity for carbon monoxide (P = 0.0217) and increased serum levels of carboxyterminal propeptide of collagen I (P = 0.0037), regardless of capillaroscopic alterations. Patients with a capillaroscopic scleroderma pattern (n = 55, 77.5%) had a higher prevalence of puffy fingers (P = 0.0001) and increased serum levels of soluble E-selectin (P = 0.0003) regardless of marker autoantibodies. Conclusion These results suggest that the autoantibody and microvascular patterns in early SSc may each be related to different clinical-preclinical features and circulating activation markers at presentation. Longitudinal studies are warranted to investigate whether these subsets undergo a

  13. Endothelial Progenitor Cells for Diagnosis and Prognosis in Cardiovascular Disease

    PubMed Central

    Cairo, Valentina; D'Ascola, Angela; Scuruchi, Michele; Basile, Giorgio; Mandraffino, Giuseppe

    2016-01-01

    Objective. To identify, evaluate, and synthesize evidence on the predictive power of circulating endothelial progenitor cells (EPCs) in cardiovascular disease, through a systematic review of quantitative studies. Data Sources. MEDLINE was searched using keywords related to “endothelial progenitor cells” and “endothelium” and, for the different categories, respectively, “smoking”; “blood pressure”; “diabetes mellitus” or “insulin resistance”; “dyslipidemia”; “aging” or “elderly”; “angina pectoris” or “myocardial infarction”; “stroke” or “cerebrovascular disease”; “homocysteine”; “C-reactive protein”; “vitamin D”. Study Selection. Database hits were evaluated against explicit inclusion criteria. From 927 database hits, 43 quantitative studies were included. Data Syntheses. EPC count has been suggested for cardiovascular risk estimation in the clinical practice, since it is currently accepted that EPCs can work as proangiogenic support cells, maintaining their importance as regenerative/reparative potential, and also as prognostic markers. Conclusions. EPCs showed an important role in identifying cardiovascular risk conditions, and to suggest their evaluation as predictor of outcomes appears to be reasonable in different defined clinical settings. Due to their capability of proliferation, circulation, and the development of functional progeny, great interest has been directed to therapeutic use of progenitor cells in atherosclerotic diseases. This trial is registered with registration number: Prospero CRD42015023717. PMID:26839569

  14. Hydrogen prevents corneal endothelial damage in phacoemulsification cataract surgery.

    PubMed

    Igarashi, Tsutomu; Ohsawa, Ikuroh; Kobayashi, Maika; Igarashi, Toru; Suzuki, Hisaharu; Iketani, Masumi; Takahashi, Hiroshi

    2016-01-01

    In phacoemulsification, ultrasound induces hydroxyl radical (·OH) formation, damaging corneal endothelium. Whether H2 can prevent such oxidative damage in phacoemulsification was examined by in vitro and in vivo studies. H2 was dissolved in a commercial irrigating solution. The effects of H2 against ·OH generation were first confirmed in vitro by electron-spin resonance (ESR) and hydroxyphenyl fluorescein (HPF). ESR showed a significantly decreased signal magnitude, and fluorescence intensity by oxidized HPF was significantly less in the H2-dissolved solution. The effects of H2 in phacoemulsification were evaluated in rabbits, comparing H2-dissolved and control solutions. Five hours after the procedure, the whole cornea was excised and subjected to image analysis for corneal edema, real-time semiquantitative PCR (qPCR) for heme oxygenase (HO)-1, catalase (CAT), superoxide dismutase 1 (SOD1), and SOD2 mRNA, and immunohistochemistry. Corneal edema was significantly less and the increases in anti-oxidative HO-1, CAT and SOD2 mRNA expressions were significantly suppressed in the H2 group. In addition, corneal endothelial cell expressions of two oxidative stress markers, 4-HNE and 8-OHdG, were significantly lower in the H2 group. In conclusion, H2 dissolved in the ocular irrigating solution protected corneal endothelial cells from phacoemulsification-induced oxidative stress and damage. PMID:27498755

  15. Hydrogen prevents corneal endothelial damage in phacoemulsification cataract surgery

    PubMed Central

    Igarashi, Tsutomu; Ohsawa, Ikuroh; Kobayashi, Maika; Igarashi, Toru; Suzuki, Hisaharu; Iketani, Masumi; Takahashi, Hiroshi

    2016-01-01

    In phacoemulsification, ultrasound induces hydroxyl radical (·OH) formation, damaging corneal endothelium. Whether H2 can prevent such oxidative damage in phacoemulsification was examined by in vitro and in vivo studies. H2 was dissolved in a commercial irrigating solution. The effects of H2 against ·OH generation were first confirmed in vitro by electron-spin resonance (ESR) and hydroxyphenyl fluorescein (HPF). ESR showed a significantly decreased signal magnitude, and fluorescence intensity by oxidized HPF was significantly less in the H2-dissolved solution. The effects of H2 in phacoemulsification were evaluated in rabbits, comparing H2-dissolved and control solutions. Five hours after the procedure, the whole cornea was excised and subjected to image analysis for corneal edema, real-time semiquantitative PCR (qPCR) for heme oxygenase (HO)-1, catalase (CAT), superoxide dismutase 1 (SOD1), and SOD2 mRNA, and immunohistochemistry. Corneal edema was significantly less and the increases in anti-oxidative HO-1, CAT and SOD2 mRNA expressions were significantly suppressed in the H2 group. In addition, corneal endothelial cell expressions of two oxidative stress markers, 4-HNE and 8-OHdG, were significantly lower in the H2 group. In conclusion, H2 dissolved in the ocular irrigating solution protected corneal endothelial cells from phacoemulsification-induced oxidative stress and damage. PMID:27498755

  16. In Vitro Expansion of Corneal Endothelial Cells on Biomimetic Substrates

    PubMed Central

    Palchesko, Rachelle N.; Lathrop, Kira L.; Funderburgh, James L.; Feinberg, Adam W.

    2015-01-01

    Corneal endothelial (CE) cells do not divide in vivo, leading to edema, corneal clouding and vision loss when the density drops below a critical level. The endothelium can be replaced by transplanting allogeneic tissue; however, access to donated tissue is limited worldwide resulting in critical need for new sources of corneal grafts. In vitro expansion of CE cells is a potential solution, but is challenging due to limited proliferation and loss of phenotype in vitro via endothelial to mesenchymal transformation (EMT) and senescence. We hypothesized that a bioengineered substrate recapitulating chemo-mechanical properties of Descemet's membrane would improve the in vitro expansion of CE cells while maintaining phenotype. Results show that bovine CE cells cultured on a polydimethylsiloxane surface with elastic modulus of 50 kPa and collagen IV coating achieved >3000-fold expansion. Cells grew in higher-density monolayers with polygonal morphology and ZO-1 localization at cell-cell junctions in contrast to control cells on polystyrene that lost these phenotypic markers coupled with increased α-smooth muscle actin expression and fibronectin fibril assembly. In total, these results demonstrate that a biomimetic substrate presenting native basement membrane ECM proteins and mechanical environment may be a key element in bioengineering functional CE layers for potential therapeutic applications. PMID:25609008

  17. Endothelial microparticles as conveyors of information in atherosclerotic disease.

    PubMed

    Schiro, A; Wilkinson, F L; Weston, R; Smyth, J V; Serracino-Inglott, F; Alexander, M Y

    2014-06-01

    Endothelial microparticles (EMPs) are complex submicron membrane-shed vesicles released into the circulation following endothelium cell activation or apoptosis. They are classified as either physiological or pathological, with anticoagulant or pro-inflammatory effects respectively. Endothelial dysfunction caused by inflammation is a key initiating event in atherosclerotic plaque formation. Athero-emboli, resulting from ruptured carotid plaques are a major cause of stroke. Current clinical techniques for arterial assessment, angiography and carotid ultrasound, give accurate information about stenosis but limited evidence on plaque composition, inflammation or vulnerability; as a result, patients with asymptomatic, or fragile carotid lesions, may not be identified and treated effectively. There is a need to discover novel biomarkers and develop more efficient diagnostic approaches in order to stratify patients at most risk of stroke, who would benefit from interventional surgery. Increasing evidence suggests that EMPs play an important role in the pathogenesis of cardiovascular disease, acting as a marker of damage, either exacerbating disease progression or triggering a repair response. In this regard, it has been suggested that EMPs have the potential to act as biomarkers of disease status. In this review, we will present the evidence to support this hypothesis and propose a novel concept for the development of a diagnostic device that could be implemented in the clinic. PMID:24721189

  18. Chronic Sleep Fragmentation Induces Endothelial Dysfunction and Structural Vascular Changes in Mice

    PubMed Central

    Carreras, Alba; Zhang, Shelley X.; Peris, Eduard; Qiao, Zhuanhong; Gileles-Hillel, Alex; Li, Richard C.; Wang, Yang; Gozal, David

    2014-01-01

    Study Objectives: Sleep fragmentation (SF) is a common occurrence and constitutes a major characteristic of obstructive sleep apnea (OSA). SF has been implicated in multiple OSA-related morbidities, but it is unclear whether SF underlies any of the cardiovascular morbidities of OSA. We hypothesized that long-term SF exposures may lead to endothelial dysfunction and altered vessel wall structure. Methods and Results: Adult male C57BL/6J mice were fed normal chow and exposed to daylight SF or control sleep (CTL) for 20 weeks. Telemetric blood pressure and endothelial function were assessed weekly using a modified laser-Doppler hyperemic test. Atherosclerotic plaques, elastic fiber disruption, lumen area, wall thickness, foam cells, and macrophage recruitment, as well as expression of senescence-associated markers were examined in excised aortas. Increased latencies to reach baseline perfusion levels during the post-occlusive period emerged in SF mice with increased systemic BP values starting at 8 weeks of SF and persisting thereafter. No obvious atherosclerotic plaques emerged, but marked elastic fiber disruption and fiber disorganization were apparent in SF-exposed mice, along with increases in the number of foam cells and macrophages in the aorta wall. Senescence markers showed reduced TERT and cyclin A and increased p16INK4a expression, with higher IL-6 plasma levels in SF-exposed mice. Conclusions: Long-term sleep fragmentation induces vascular endothelial dysfunction and mild blood pressure increases. Sleep fragmentation also leads to morphologic vessel changes characterized by elastic fiber disruption and disorganization, increased recruitment of inflammatory cells, and altered expression of senescence markers, thereby supporting a role for sleep fragmentation in the cardiovascular morbidity of OSA. Citation: Carreras A, Zhang SX, Peris E, Qiao Z, Gileles-Hillel A, Li RC, Wang Y, Gozal D. Chronic sleep fragmentation induces endothelial dysfunction and

  19. Flow cytometric analysis of circulating endothelial cells and endothelial progenitors for clinical purposes in oncology: A critical evaluation

    PubMed Central

    DANOVA, MARCO; COMOLLI, GIUDITTA; MANZONI, MARIANGELA; TORCHIO, MARTINA; MAZZINI, GIULIANO

    2016-01-01

    Malignant tumors are characterized by uncontrolled cell growth and metastatic spread, with a pivotal importance of the phenomenon of angiogenesis. For this reason, research has focused on the development of agents targeting the vascular component of the tumor microenvironment and regulating the angiogenic switch. As a result, the therapeutic inhibition of angiogenesis has become an important component of anticancer treatment, however, its utility is partly limited by the lack of an established methodology to assess its efficacy in vivo. Circulating endothelial cells (CECs), which are rare in healthy subjects and significantly increased in different tumor types, represent a promising tool for monitoring the tumor clinical outcome and the treatment response. A cell population circulating into the blood also able to form endothelial colonies in vitro and to promote vasculogenesis is represented by endothelial progenitor cells (EPCs). The number of both of these cell types is extremely low and they cannot be identified using a single marker, therefore, in absence of a definite consensus on their phenotype, require discrimination using combinations of antigens. Multiparameter flow cytometry (FCM) is ideal for rapid processing of high numbers of cells per second and is commonly utilized to quantify CECs and EPCs, however, remains technically challenging since there is as yet no standardized protocol for the identification and enumeration of these rare events. Methodology in studies on CECs and/or EPCs as clinical biomarkers in oncology is heterogeneous and data have been obtained from different studies leading to conflicting conclusions. The present review presented a critical review of the issues that limit the comparability of results of the most significant studies employing FCM for CEC and/or EPC detection in patients with cancer. PMID:27284422

  20. Influenza Virus Infection Induces Platelet-Endothelial Adhesion Which Contributes to Lung Injury.

    PubMed

    Sugiyama, Michael G; Gamage, Asela; Zyla, Roman; Armstrong, Susan M; Advani, Suzanne; Advani, Andrew; Wang, Changsen; Lee, Warren L

    2016-02-01

    Lung injury after influenza infection is characterized by increased permeability of the lung microvasculature, culminating in acute respiratory failure. Platelets interact with activated endothelial cells and have been implicated in the pathogenesis of some forms of acute lung injury. Autopsy studies have revealed pulmonary microthrombi after influenza infection, and epidemiological studies suggest that influenza vaccination is protective against pulmonary thromboembolism; however, the effect of influenza infection on platelet-endothelial interactions is unclear. We demonstrate that endothelial infection with both laboratory and clinical strains of influenza virus increased the adhesion of human platelets to primary human lung microvascular endothelial cells. Platelets adhered to infected cells as well as to neighboring cells, suggesting a paracrine effect. Influenza infection caused the upregulation of von Willebrand factor and ICAM-1, but blocking these receptors did not prevent platelet-endothelial adhesion. Instead, platelet adhesion was inhibited by both RGDS peptide and a blocking antibody to platelet integrin α5β1, implicating endothelial fibronectin. Concordantly, lung histology from infected mice revealed viral dose-dependent colocalization of viral nucleoprotein and the endothelial marker PECAM-1, while platelet adhesion and fibronectin deposition also were observed in the lungs of influenza-infected mice. Inhibition of platelets using acetylsalicylic acid significantly improved survival, a finding confirmed using a second antiplatelet agent. Thus, influenza infection induces platelet-lung endothelial adhesion via fibronectin, contributing to mortality from acute lung injury. The inhibition of platelets may constitute a practical adjunctive strategy to the treatment of severe infections with influenza.IMPORTANCE There is growing appreciation of the involvement of the lung endothelium in the pathogenesis of severe infections with influenza virus. We have

  1. Influenza Virus Infection Induces Platelet-Endothelial Adhesion Which Contributes to Lung Injury

    PubMed Central

    Sugiyama, Michael G.; Gamage, Asela; Zyla, Roman; Armstrong, Susan M.; Advani, Suzanne; Advani, Andrew; Wang, Changsen

    2015-01-01

    ABSTRACT Lung injury after influenza infection is characterized by increased permeability of the lung microvasculature, culminating in acute respiratory failure. Platelets interact with activated endothelial cells and have been implicated in the pathogenesis of some forms of acute lung injury. Autopsy studies have revealed pulmonary microthrombi after influenza infection, and epidemiological studies suggest that influenza vaccination is protective against pulmonary thromboembolism; however, the effect of influenza infection on platelet-endothelial interactions is unclear. We demonstrate that endothelial infection with both laboratory and clinical strains of influenza virus increased the adhesion of human platelets to primary human lung microvascular endothelial cells. Platelets adhered to infected cells as well as to neighboring cells, suggesting a paracrine effect. Influenza infection caused the upregulation of von Willebrand factor and ICAM-1, but blocking these receptors did not prevent platelet-endothelial adhesion. Instead, platelet adhesion was inhibited by both RGDS peptide and a blocking antibody to platelet integrin α5β1, implicating endothelial fibronectin. Concordantly, lung histology from infected mice revealed viral dose-dependent colocalization of viral nucleoprotein and the endothelial marker PECAM-1, while platelet adhesion and fibronectin deposition also were observed in the lungs of influenza-infected mice. Inhibition of platelets using acetylsalicylic acid significantly improved survival, a finding confirmed using a second antiplatelet agent. Thus, influenza infection induces platelet-lung endothelial adhesion via fibronectin, contributing to mortality from acute lung injury. The inhibition of platelets may constitute a practical adjunctive strategy to the treatment of severe infections with influenza. IMPORTANCE There is growing appreciation of the involvement of the lung endothelium in the pathogenesis of severe infections with influenza

  2. The expression of endothelial cell surface antigens by AIDS-associated Kaposi's sarcoma. Evidence for a vascular endothelial cell origin.

    PubMed Central

    Rutgers, J. L.; Wieczorek, R.; Bonetti, F.; Kaplan, K. L.; Posnett, D. N.; Friedman-Kien, A. E.; Knowles, D. M.

    1986-01-01

    The authors investigated 19 cases of Kaposi's sarcoma (KS) obtained from patients with the acquired immune deficiency syndrome (AIDS) for their expression of Factor VIII-related antigen (FVIIIRAg), HLA-DR (Ia) antigens, OKM1, and three distinctive vascular, but not lymphatic, endothelial-cell-associated antigens, E92, OKM5, and HCl. Antigen expression was demonstrated by immunoperoxidase staining of cryostat sections. FVIIIRAg is strongly expressed by the cells lining the vascular spaces (VCs) but is absent, weakly or focally, and variably expressed by the spindle cell (SC) component of KS. The VC component of each KS lesion examined strongly expressed E92, moderately expressed HCl, and weakly expressed OKM5. In contrast, the entire SC component of each KS lesion studied strongly expressed E92 and OKM5 and weakly expressed HCl. Neither the VCs nor the SCs expressed OKM1. These studies provide strong and compelling evidence for the vascular endothelial cell histogenesis of both the vascular and spindle cell components of KS, demonstrate the intertumor and intratumor phenotypic heterogeneity of KS, and suggest that monoclonal antibodies OKM5 and anti-E92 are the best currently available immunohistochemical markers for identifying the spindle cell component of AIDS-associated KS in cryostat sections. Images Figure 1 PMID:3953772

  3. Effects of sodium and potassium supplementation on endothelial function: a fully controlled dietary intervention study.

    PubMed

    Gijsbers, Lieke; Dower, James I; Schalkwijk, Casper G; Kusters, Yvo H A M; Bakker, Stephan J L; Hollman, Peter C H; Geleijnse, Johanna M

    2015-11-14

    High Na and low K intakes have adverse effects on blood pressure, which increases the risk for CVD. The role of endothelial dysfunction and inflammation in this pathophysiological process is not yet clear. In a randomised placebo-controlled cross-over study in untreated (pre)hypertensives, we examined the effects of Na and K supplementation on endothelial function and inflammation. During the study period, subjects were provided with a diet that contained 2·4 g/d of Na and 2·3 g/d of K for a 10 460 kJ (2500 kcal) intake. After 1-week run-in, subjects received capsules with supplemental Na (3·0 g/d), supplemental K (2·8 g/d) or placebo, for 4 weeks each, in random order. After each intervention, circulating biomarkers of endothelial function and inflammation were measured. Brachial artery flow-mediated dilation (FMD) and skin microvascular vasomotion were assessed in sub-groups of twenty-two to twenty-four subjects. Of thirty-seven randomised subjects, thirty-six completed the study. Following Na supplementation, serum endothelin-1 was increased by 0·24 pg/ml (95 % CI 0·03, 0·45), but no change was seen in other endothelial or inflammatory biomarkers. FMD and microvascular vasomotion were unaffected by Na supplementation. K supplementation reduced IL-8 levels by 0·28 pg/ml (95 % CI 0·03, 0·53), without affecting other circulating biomarkers. FMD was 1·16 % (95% CI 0·37, 1·96) higher after K supplementation than after placebo. Microvascular vasomotion was unaffected. In conclusion, a 4-week increase in Na intake increased endothelin-1, but had no effect on other endothelial or inflammatory markers. Increased K intake had a beneficial effect on FMD and possibly IL-8, without affecting other circulating endothelial or inflammatory biomarkers. PMID:26343780

  4. Endothelial Dysfunction: Associations with Exposure to Ambient Fine Particles in Diabetic Individuals

    PubMed Central

    Schneider, Alexandra; Neas, Lucas; Herbst, Margaret C.; Case, Martin; Williams, Ronald W.; Cascio, Wayne; Hinderliter, Alan; Holguin, Fernando; Buse, John B.; Dungan, Kathleen; Styner, Maya; Peters, Annette; Devlin, Robert B.

    2008-01-01

    Background Exposure to fine airborne particulate matter [≤2.5 μm in aerodynamic diameter (PM2.5)] has been associated with cardiovascular and hematologic effects, especially in older people with cardiovascular disease. Some epidemiologic studies suggest that adults with diabetes also may be a particularly susceptible population. Objectives The purpose of this study was to analyze the short-term effects of ambient PM2.5 on markers of endothelial function in diabetic volunteers. Methods We conducted a prospective panel study in 22 people with type 2 diabetes mellitus in Chapel Hill, North Carolina (USA), from November 2004 to December 2005. We acquired daily measurements of PM2.5 and meteorologic data at central monitoring sites. On 4 consecutive days, we measured endothelial function by brachial artery ultrasound in all participants and by pulsewave measurements in a subgroup. Data were analyzed using additive mixed models with a random participant effect and adjusted for season, day of the week, and meteorology. Results Flow-mediated dilatation decreased in association with PM2.5 during the first 24 hr, whereas small-artery elasticity index decreased with a delay of 1 and 3 days. These PM2.5-associated decrements in endothelial function were greater among participants with a high body mass index, high glycosylated hemoglobin A1c, low adiponectin, or the null polymorphism of glutathione S-transferase M1. However, high levels of myeloperoxidase on the examination day led to strongest effects on endothelial dysfunction. Conclusions These data demonstrate that PM2.5 exposure may cause immediate endothelial dysfunction. Clinical characteristics associated with insulin resistance were associated with enhanced effects of PM on endothelial function. In addition, participants with greater oxidative potential seem to be more susceptible. PMID:19079718

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

    PubMed

    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; Anders, Hans-Joachim

    2016-06-01

    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

  6. Dynamics of circulating microparticles in chronic kidney disease and transplantation: Is it really reliable marker?

    PubMed Central

    Dursun, Ismail; Yel, Sibel; Unsur, Emel

    2015-01-01

    The deterioration of endothelial structure plays a very important role in the development of vascular diseases. It is believed that endothelial dysfunction starts in the early stage of kidney disease and is a risk factor of an unfavorable cardiovascular prognosis. Because a direct assessment of biological states in endothelial cells is not applicable, the measurement of endothelial microparticles (EMPs) detached from endothelium during activation or apoptosis is thought to be a marker of early vascular disease and endothelial dysfunction in children with chronic kidney disease (CKD). Few studies have shown increased circulating EMPs and its relationship with cardiovascular risk factors in patients with CKD. MPs contain membrane proteins and cytosolic material derived from the cell from which they originate. EMPs having CD144, CD 146, CD31+/CD41-, CD51 and CD105 may be used to evaluate the vascular endothelial cell damage and determine asymptomatic patients who might be at higher risk of developing cardiovascular disease in CKD and renal transplant. PMID:26722654

  7. Blood cells and endothelial barrier function.

    PubMed

    Rodrigues, Stephen F; Granger, D Neil

    2015-01-01

    The barrier properties of endothelial cells are critical for the maintenance of water and protein balance between the intravascular and extravascular compartments. An impairment of endothelial barrier function has been implicated in the genesis and/or progression of a variety of pathological conditions, including pulmonary edema, ischemic stroke, neurodegenerative disorders, angioedema, sepsis and cancer. The altered barrier function in these conditions is often linked to the release of soluble mediators from resident cells (e.g., mast cells, macrophages) and/or recruited blood cells. The interaction of the mediators with receptors expressed on the surface of endothelial cells diminishes barrier function either by altering the expression of adhesive proteins in the inter-endothelial junctions, by altering the organization of the cytoskeleton, or both. Reactive oxygen species (ROS), proteolytic enzymes (e.g., matrix metalloproteinase, elastase), oncostatin M, and VEGF are part of a long list of mediators that have been implicated in endothelial barrier failure. In this review, we address the role of blood borne cells, including, neutrophils, lymphocytes, monocytes, and platelets, in the regulation of endothelial barrier function in health and disease. Attention is also devoted to new targets for therapeutic intervention in disease states with morbidity and mortality related to endothelial barrier dysfunction. PMID:25838983

  8. Blood cells and endothelial barrier function

    PubMed Central

    Rodrigues, Stephen F; Granger, D Neil

    2015-01-01

    Abstract The barrier properties of endothelial cells are critical for the maintenance of water and protein balance between the intravascular and extravascular compartments. An impairment of endothelial barrier function has been implicated in the genesis and/or progression of a variety of pathological conditions, including pulmonary edema, ischemic stroke, neurodegenerative disorders, angioedema, sepsis and cancer. The altered barrier function in these conditions is often linked to the release of soluble mediators from resident cells (e.g., mast cells, macrophages) and/or recruited blood cells. The interaction of the mediators with receptors expressed on the surface of endothelial cells diminishes barrier function either by altering the expression of adhesive proteins in the inter-endothelial junctions, by altering the organization of the cytoskeleton, or both. Reactive oxygen species (ROS), proteolytic enzymes (e.g., matrix metalloproteinase, elastase), oncostatin M, and VEGF are part of a long list of mediators that have been implicated in endothelial barrier failure. In this review, we address the role of blood borne cells, including, neutrophils, lymphocytes, monocytes, and platelets, in the regulation of endothelial barrier function in health and disease. Attention is also devoted to new targets for therapeutic intervention in disease states with morbidity and mortality related to endothelial barrier dysfunction. PMID:25838983

  9. Fibrinogen induces endothelial cell permeability

    PubMed Central

    Tyagi, Neetu; Roberts, Andrew M.; Dean, William L.; Tyagi, Suresh C.

    2010-01-01

    Many cardiovascular and cerebrovascular disorders are accompanied by an increased blood content of fibrinogen (Fg), a high molecular weight plasma adhesion protein. Fg is a biomarker of inflammation and its degradation products have been associated with microvascular leakage. We tested the hypothesis that at pathologically high levels, Fg increases endothelial cell (EC) permeability through extracellular signal regulated kinase (ERK) signaling and by inducing F-actin formation. In cultured ECs, Fg binding to intercellular adhesion molecule-1 and to α5β1 integrin, caused phosphorylation of ERK. Subsequently, F-actin formation increased and coincided with formation of gaps between ECs, which corresponded with increased permeability of ECs to albumin. Our data suggest that formation of F-actin and gaps may be the mechanism for increased albumin leakage through the EC monolayer. The present study indicates that elevated un-degraded Fg may be a factor causing microvascular permeability that typically accompanies cardiovascular and cerebrovascular disorders. PMID:17849175

  10. Dextran induces differentiation of circulating endothelial progenitor cells

    PubMed Central

    Obi, Syotaro; Masuda, Haruchika; Akimaru, Hiroshi; Shizuno, Tomoko; Yamamoto, Kimiko; Ando, Joji; Asahara, Takayuki

    2014-01-01

    Abstract Endothelial progenitor cells (EPCs) have been demonstrated to be effective for the treatment of cardiovascular diseases. However, the differentiation process from circulation to adhesion has not been clarified because circulating EPCs rarely attached to dishes in EPC cultures previously. Here we investigated whether immature circulating EPCs differentiate into mature adhesive EPCs in response to dextran. When floating‐circulating EPCs derived from ex vivo expanded human cord blood were cultured with 5% and 10% dextran, they attached to fibronectin‐coated dishes and grew exponentially. The bioactivities of adhesion, proliferation, migration, tube formation, and differentiated type of EPC colony formation increased in EPCs exposed to dextran. The surface protein expression rate of the endothelial markers vascular endothelial growth factor (VEGF)‐R1/2, VE‐cadherin, Tie2, ICAM1, VCAM1, and integrin αv/β3 increased in EPCs exposed to dextran. The mRNA levels of VEGF‐R1/2, VE‐cadherin, Tie2, endothelial nitric oxide synthase, MMP9, and VEGF increased in EPCs treated with dextran. Those of endothelium‐related transcription factors ID1/2, FOXM1, HEY1, SMAD1, FOSL1, NFkB1, NRF2, HIF1A, EPAS1 increased in dextran‐treated EPCs; however, those of hematopoietic‐ and antiangiogenic‐related transcription factors TAL1, RUNX1, c‐MYB, GATA1/2, ERG, FOXH1, HHEX, SMAD2/3 decreased in dextran‐exposed EPCs. Inhibitor analysis showed that PI3K/Akt, ERK1/2, JNK, and p38 signal transduction pathways are involved in the differentiation in response to dextran. In conclusion, dextran induces differentiation of circulating EPCs in terms of adhesion, migration, proliferation, and vasculogenesis. The differentiation mechanism in response to dextran is regulated by multiple signal transductions including PI3K/Akt, ERK1/2, JNK, and p38. These findings indicate that dextran is an effective treatment for EPCs in regenerative medicines. PMID:24760515

  11. Supplementation with orange and blackcurrant juice, but not vitamin E, improves inflammatory markers in patients with peripheral arterial disease.

    PubMed

    Dalgård, Christine; Nielsen, Flemming; Morrow, Jason D; Enghusen-Poulsen, Henrik; Jonung, Torbjörn; Hørder, Mogens; de Maat, Moniek P M

    2009-01-01

    Inflammation and endothelial activation are associated with an increased risk of CVD and epidemiological evidence suggests an association between levels of markers of inflammation or endothelial activation and the intake of fruit. Also, vitamin E, a fat-soluble antioxidant, has anti-inflammatory properties. We performed a randomised 2 x 2 factorial, crossover trial to determine the effect of orange and blackcurrant juice (500 ml/d) and vitamin E (15 mg RRR-alpha-tocopherol/d) supplementation on markers of inflammation and endothelial activation in forty-eight patients with peripheral arterial disease. Patients were randomly allocated to two dietary supplements from the four possible combinations of juice and vitamin E: juice+vitamin E; juice+placebo; reference beverage (sugar drink)+vitamin E; and reference beverage+placebo. The supplementations were given for 28 d, separated by a 4-week wash-out period. Analysis of main effects showed that juice decreased C-reactive protein (CRP) by 11% and fibrinogen by 3% while the reference drink increased CRP by 13% and fibrinogen by 2% (P<0.008 and P<0.002, respectively). No significant differences were measured for IL-6 and the endothelial activation markers von Willebrand factor, tissue-plasminogen activator and plasmin activator inhibitor-1. Vitamin E supplementation had no significant effects on the various markers. We observed no significant interaction between juice and vitamin E. In this study, orange and blackcurrant juice reduced markers of inflammation, but not markers of endothelial activation, in patients with peripheral arterial disease, relative to sugar drinks. PMID:18507878

  12. Interleukin-3 greatly expands non-adherent endothelial forming cells with pro-angiogenic properties.

    PubMed

    Moldenhauer, Lachlan M; Cockshell, Michaelia P; Frost, Lachlan; Parham, Kate A; Tvorogov, Denis; Tan, Lih Y; Ebert, Lisa M; Tooley, Katie; Worthley, Stephen; Lopez, Angel F; Bonder, Claudine S

    2015-05-01

    Circulating endothelial progenitor cells (EPCs) provide revascularisation for cardiovascular disease and the expansion of these cells opens up the possibility of their use as a cell therapy. Herein we show that interleukin-3 (IL3) strongly expands a population of human non-adherent endothelial forming cells (EXnaEFCs) with low immunogenicity as well as pro-angiogenic capabilities in vivo, making their therapeutic utilisation a realistic option. Non-adherent CD133(+) EFCs isolated from human umbilical cord blood and cultured under different conditions were maximally expanded by day 12 in the presence of IL3 at which time a 350-fold increase in cell number was obtained. Cell surface marker phenotyping confirmed expression of the hematopoietic progenitor cell markers CD133, CD117 and CD34, vascular cell markers VEGFR2 and CD31, dim expression of CD45 and absence of myeloid markers CD14 and CD11b. Functional experiments revealed that EXnaEFCs exhibited classical properties of endothelial cells (ECs), namely binding of Ulex europaeus lectin, up-take of acetylated-low density lipoprotein and contribution to EC tube formation in vitro. These EXnaEFCs demonstrated a pro-angiogenic phenotype within two independent in vivo rodent models. Firstly, a Matrigel plug assay showed increased vascularisation in mice. Secondly, a rat model of acute myocardial infarction demonstrated reduced heart damage as determined by lower levels of serum creatinine and a modest increase in heart functionality. Taken together, these studies show IL3 as a potent growth factor for human CD133(+) cell expansion with clear pro-angiogenic properties (in vitro and in vivo) and thus may provide clinical utility for humans in the future. PMID:25900163

  13. Informativeness of microsatellite markers.

    PubMed

    Reyes-Valdés, M Humberto

    2013-01-01

    Simple sequence repeats (SSR) are extensively used as genetic markers for studies of diversity, genetic mapping, and cultivar discrimination. The informativeness of a given SSR locus or a loci group depends on the number of alleles, their frequency distribution, as well as the kind of application. Here I describe several methods for calculating marker informativeness, all of them suitable for SSR polymorphisms, proposed by several authors and synthesized in an Information Theory framework. Additionally, free access software resources are described as well as their application through worked examples. PMID:23546797

  14. Skin Autofluorescence Is Associated with Endothelial Dysfunction in Uremic Subjects on Hemodialysis

    PubMed Central

    Wang, Chun-Cheng; Wang, Yao-Chang; Wang, Guei-Jane; Shen, Ming-Yi; Chang, Yen-Lin; Liou, Show-Yih; Chen, Hung-Chih; Chang, Chiz-Tzung

    2016-01-01

    Background Elevated levels of advanced glycation end products (AGEs) within tissues may contribute to endothelial dysfunction, an early indicator of atherosclerosis. We aimed to investigate whether levels of skin AGEs could be a useful marker to predict endothelial dysfunction in uremic subjects on hemodialysis. Methods and Results One hundred and nineteen uremic patients on hemodialysis and 57 control subjects with moderate-to-high cardiovascular risk factors and without chronic kidney disease (CKD) were enrolled. We used ultrasound to measure flow-mediated vasodilation (FMD). An AGE reader measured skin autoflurorescence (AF). We then compared differences in FMD and skin AF values between the two groups. The uremic subjects had significantly higher levels of skin AF (3.47±0.76 AU vs. 2.21±0.45 arbitrary units; P<0.01) and significantly lower levels of FMD (4.79%±1.88% vs. 7.19%±2.17%; P<0.01) than the non-CKD subjects. After adjusting for all potential covariates, we found that skin AF level independently predicted FMD in both the hemodialysis and the non-CKD groups. In the hemodialysis group, skin AF ≥ 3.05 arbitrary units predicted abnormal FMD at a sensitivity of 87.9% and a specificity of 78.6% (P<0.01). Conclusions Skin AF could be a useful marker to predict endothelial dysfunction in uremic subjects on hemodialysis. PMID:26809145

  15. Deeper Penetration of Erythrocytes into the Endothelial Glycocalyx Is Associated with Impaired Microvascular Perfusion

    PubMed Central

    Lee, Dae Hyun; Dane, Martijn J. C.; van den Berg, Bernard M.; Boels, Margien G. S.; van Teeffelen, Jurgen W.; de Mutsert, Renée; den Heijer, Martin; Rosendaal, Frits R.; van der Vlag, Johan; van Zonneveld, Anton Jan; Vink, Hans; Rabelink, Ton J.

    2014-01-01

    Changes in endothelial glycocalyx are one of the earliest changes in development of cardiovascular disease. The endothelial glycocalyx is both an important biological modifier of interactions between flowing blood and the vessel wall, and a determinant of organ perfusion. We hypothesize that deeper penetration of erythrocytes into the glycocalyx is associated with reduced microvascular perfusion. The population-based prospective cohort study (the Netherlands Epidemiology of Obesity [NEO] study) includes 6,673 middle-aged individuals (oversampling of overweight and obese individuals). Within this cohort, we have imaged the sublingual microvasculature of 915 participants using sidestream darkfield (SDF) imaging together with a recently developed automated acquisition and analysis approach. Presence of RBC (as a marker of microvascular perfusion) and perfused boundary region (PBR), a marker for endothelial glycocalyx barrier properties for RBC accessibility, were assessed in vessels between 5 and 25 µm RBC column width. A wide range of variability in PBR measurements, with a mean PBR of 2.14 µm (range: 1.43–2.86 µm), was observed. Linear regression analysis showed a marked association between PBR and microvascular perfusion, reflected by RBC filling percentage (regression coefficient β: −0.034; 95% confidence interval: −0.037 to −0.031). We conclude that microvascular beds with a thick (“healthy”) glycocalyx (low PBR), reflects efficient perfusion of the microvascular bed. In contrast, a thin (“risk”) glycocalyx (high PBR) is associated with a less efficient and defective microvascular perfusion. PMID:24816787

  16. Hyaluronidase 1 Deficiency Preserves Endothelial Function and Glycocalyx Integrity in Early Streptozotocin-Induced Diabetes.

    PubMed

    Dogné, Sophie; Rath, Géraldine; Jouret, François; Caron, Nathalie; Dessy, Chantal; Flamion, Bruno

    2016-09-01

    Hyaluronic acid (HA) is a major component of the glycocalyx involved in the vascular wall and endothelial glomerular permeability barrier. Endocytosed hyaluronidase HYAL1 is known to degrade HA into small fragments in different cell types, including endothelial cells. In diabetes, the size and permeability of the glycocalyx are altered. In addition, patients with type 1 diabetes present increased plasma levels of both HA and HYAL1. To investigate the potential implication of HYAL1 in the development of diabetes-induced endothelium dysfunction, we measured endothelial markers, endothelium-dependent vasodilation, arteriolar glycocalyx size, and glomerular barrier properties in wild-type and HYAL1 knockout (KO) mice with or without streptozotocin (STZ)-induced diabetes. We observed that 4 weeks after STZ injections, the lack of HYAL1 1) prevents diabetes-induced increases in soluble P-selectin concentrations and limits the impact of the disease on endothelium-dependent hyperpolarization (EDH)-mediated vasorelaxation; 2) increases glycocalyx thickness and maintains glycocalyx structure and HA content during diabetes; and 3) prevents diabetes-induced glomerular barrier dysfunction assessed using the urinary albumin-to-creatinine ratio and urinary ratio of 70- to 40-kDa dextran. Our findings suggest that HYAL1 contributes to endothelial and glycocalyx dysfunction induced by diabetes. HYAL1 inhibitors could be explored as a new therapeutic approach to prevent vascular complications in diabetes. PMID:27246914

  17. Low Shear Stress Inhibited Endothelial Cell Autophagy Through TET2 Downregulation.

    PubMed

    Yang, Qin; Li, Xiaohong; Li, Rongqing; Peng, Juan; Wang, Zuo; Jiang, Zhisheng; Tang, Xiaoqing; Peng, Zhao; Wang, Yu; Wei, Dangheng

    2016-07-01

    Low shear stress plays a crucial role in the initiation and progression of atherosclerotic lesions. However, the detailed mechanisms of these processes remain unclear. In this study, the effect of low shear stress on endothelial cell autophagy and its potential mechanism were investigated. Results showed autophagy dysfunction and ten-eleven translocation 2 (TET2) protein downregulation during atherosclerotic lesion progression. Autophagic markers BECLIN 1 and LC3II/LC3I under low shear stress (5 dyne/cm(2)) obviously decreased compared with those under physiological shear stress (15 dyne/cm(2)), whereas autophagic substrate p62 increased. TET2 expression was also downregulated under low shear stress. Endothelial cell autophagy was improved with TET2 overexpression but was impaired by TET2 siRNA treatment. Moreover, TET2 overexpression upregulated the expression of endothelial cell nitric oxide synthase (eNOS) and downregulated the expression of endothelin-1 (ET-1). TET2 siRNA further attenuated eNOS expression and stimulated ET-1 expression. Overall, the results showed that low shear stress downregulated endothelial cell autophagy by impaired TET2 expression, which might contribute to the atherogenic process. PMID:26493943

  18. Black tea protects against hypertension-associated endothelial dysfunction through alleviation of endoplasmic reticulum stress

    PubMed Central

    San Cheang, Wai; Yuen Ngai, Ching; Yen Tam, Ye; Yu Tian, Xiao; Tak Wong, Wing; Zhang, Yang; Wai Lau, Chi; Chen, Zhen Yu; Bian, Zhao-Xiang; Huang, Yu; Ping Leung, Fung

    2015-01-01

    Hypertensive patients have been found to be associated with elevated levels of homocysteine, known as hyperhomocysteinemia. Homocysteine (Hcy) can induce endoplasmic reticulum (ER) stress in endothelial cells. This study aims to investigate whether black tea (BT) protects against hypertension-associated endothelial dysfunction through alleviation of ER stress. Rat aortae and cultured rat aortic endothelial cells were treated with Hcy, BT extract, and theaflavin-3,3’-digallate (TF3). Male Sprague Dawley rats were infused with angiotensin II (Ang II) to induce hypertension and orally administrated with BT extract at 15 mg/kg/day for 2 weeks. Hcy impaired endothelium-dependent relaxations of rat aortae and led to ER stress in endothelial cells, which were ameliorated by co-incubation of BT extract and TF3. The blood pressure of Ang II-infused rats and plasma Hcy level were normalized by BT consumption. Impaired endothelium-dependent relaxations in renal arteries, carotid arteries and aortae, and flow-mediated dilatations in third-order mesenteric resistance arteries were improved. Elevations of ER stress markers and ROS level, plus down-regulation of Hcy metabolic enzymes in aortae from Ang II-infused rats were prevented by BT treatment. Our data reveal the novel cardiovascular benefits of BT in ameliorating vascular dysfunctions, providing insight into developing BT into beneficial dietary supplements in hypertensive patients. PMID:25976123

  19. PCB 77 dechlorination products modulate pro-inflammatory events in vascular endothelial cells

    PubMed Central

    Eske, Katryn; Newsome, Bradley; Han, Sung Gu; Murphy, Margaret; Bhattacharyya, Dibakar

    2013-01-01

    Persistent organic pollutants such as polychlorinated biphenyls (PCBs) are associated with detrimental health outcomes including cardiovascular diseases. Remediation of these compounds is a critical component of environmental policy. Although remediation efforts aim to completely remove toxicants, little is known about the effects of potential remediation byproducts. We previously published that Fe/Pd nanoparticles effectively dechlorinate PCB 77 to biphenyl, thus eliminating PCB-induced endothelial dysfunction using primary vascular endothelial cells. Herein, we analyzed the toxic effects of PCB congener mixtures (representative mixtures of commercial PCBs based on previous dechlorination data) produced at multiple time points during the dechlorination of PCB 77 to biphenyl. Compared with pure PCB 77, exposing endothelial cells to lower chlorinated PCB byproducts led to improved cellular viability, decreased superoxide production, and decreased nuclear factor kappa B activation based on duration of remediation. Presence of the parent compound, PCB 77, led to significant increases in mRNA and protein inflammatory marker expression. These data implicate that PCB dechlorination reduces biological toxicity to vascular endothelial cells. PMID:23504249

  20. Molecular basis for globotriaosylceramide regulation and enzyme uptake in immortalized aortic endothelial cells from Fabry mice.

    PubMed

    Meng, Xing-Li; Day, Taniqua S; McNeill, Nathan; Ashcraft, Paula; Frischmuth, Thomas; Cheng, Seng H; Liu, Zhi-Ping; Shen, Jin-Song; Schiffmann, Raphael

    2016-05-01

    Fabry disease is caused by deficient activity of α-galactosidase A and subsequent intracellular accumulation of glycosphingolipids, mainly globotriaosylceramide (Gb3). Vascular endothelial cells may play important roles in disease pathogenesis, and are one of the main target cell types in therapeutic interventions. In this study, we generated immortalized aortic endothelial cell lines from a mouse model of Fabry disease. These cells retained endothelial cell-specific markers and functions. Gb3 expression level in one of these clones (referred to as FMEC2) was highly susceptible to culture media, and appeared to be regulated by glucosylceramide synthase. Results also showed that Gb3 could be upregulated by hydrocortisone. FMEC2 express the mannose 6-phosphate receptor and sortilin but not the mannose receptor. Uptake studies suggested that sortilin plays a role in the binding and internalization of mammalian cell-produced α-galactosidase A. Moss-aGal (a plant-made enzyme) was endocytosed by FMEC2 via a receptor other than the aforementioned receptors. In conclusion, this study suggests that glucosylceramide synthase and hydrocortisone may play important roles in modulating Gb3 levels in Fabry mouse aortic endothelial cells, and that endocytosis of recombinant α-galactosidase A involves a combination of multiple receptors depending on the properties of the enzyme. PMID:26960552

  1. Pancreatic-carcinoma-cell-derived pro-angiogenic factors can induce endothelial-cell differentiation of a subset of circulating CD34+ progenitors

    PubMed Central

    2013-01-01

    Background CD34+ progenitor cells comprise both hematopoietic and endothelial progenitor cells. Recent studies suggest that circulating endothelial progenitor cells are recruited into the angiogenic vascular system of several cancers, including pancreatic carcinoma, and that they correlate with clinical progress. However, whether endothelial progenitor cell mobilization occurs in response to cytokine release by tumor cells is still unclear. Methods The chemotactic- and/or differentiating-activities of the poorly-differentiated pancreatic carcinoma cell line PT45, and of the immortal H6c7 cell line, a line of near-normal pancreatic duct epithelial cells, on endothelial progenitor cells were investigated in vitro using circulating CD34+ as model. Results The study showed that Vascular Endothelial Growth Factor produced by PT45 cells and, at lesser extent, by H6c7 cells, predominantly chemoattract peripheral blood CD34+ expressing the type 2 relative receptor. Addition of PT45-conditioned medium to CD34+ cells, cultured under conditions supporting myeloid cell development, diverted the differentiation of a subset of these progenitor cells into cells expressing endothelial cell markers, such as CD146, CD105, VE-cadherin and von Willebrand Factor-related antigen. Moreover, these endothelial-like cells formed capillary networks in vitro, chiefly through the release of Angiopoietin-1 by PT45 cells. Conclusions The results demonstrate that pancreatic-carcinoma cells potentially attract circulating endothelial progenitor cells to the tumor site, by releasing high levels of pro-angiogenic factors such as Vascular Endothelial Growth Factor and Angiopoietin-1, and may direct the differentiation of these cell subsets of the CD34+ cell population into endothelial cells; the latter cells may become a component of the newly-formed vessels, contributing to angiogenesis-mediated tumor growth and metastasis. PMID:24341512

  2. The relationship between oxidised LDL, endothelial progenitor cells and coronary endothelial function in patients with CHD

    PubMed Central

    Watt, Jonathan; Kennedy, Simon; Ahmed, Nadeem; Hayhurst, James; McClure, John D; Berry, Colin; Wadsworth, Roger M; Oldroyd, Keith G

    2016-01-01

    Objective The balance between coronary endothelial dysfunction and repair is influenced by many protective and deleterious factors circulating in the blood. We studied the relationship between oxidised low-density lipoprotein (oxLDL), circulating endothelial progenitor cells (EPCs) and coronary endothelial function in patients with stable coronary heart disease (CHD). Methods 33 patients with stable CHD were studied. Plasma oxLDL was measured using ELISA, coronary endothelial function was assessed using intracoronary acetylcholine infusion and EPCs were quantified using flow cytometry for CD34+/KDR+ cells. Results Plasma oxLDL correlated positively with the number of EPCs in the blood (r=0.46, p=0.02). There was a positive correlation between the number of circulating EPCs and coronary endothelial function (r=0.42, p=0.04). There was no significant correlation between oxLDL and coronary endothelial function. Conclusions Plasma levels of oxLDL are associated with increased circulating EPCs in the blood of patients with CHD, which may reflect a host-repair response to endothelial injury. Patients with stable CHD had a high prevalence of coronary endothelial dysfunction, which was associated with lower numbers of circulating EPCs, suggesting a mechanistic link between endothelial dysfunction and the pathogenesis of atherosclerosis. PMID:26848395

  3. Plasmodium falciparum Histidine-Rich Protein II Compromises Brain Endothelial Barriers and May Promote Cerebral Malaria Pathogenesis

    PubMed Central

    Pal, Priya; Daniels, Brian P.; Oskman, Anna; Diamond, Michael S.; Klein, Robyn S.

    2016-01-01

    ABSTRACT Cerebral malaria (CM) is a disease of the vascular endothelium caused by Plasmodium falciparum. It is characterized by parasite sequestration, inflammatory cytokine production, and vascular leakage. A distinguishing feature of P. falciparum infection is parasite production and secretion of histidine-rich protein II (HRPII). Plasma HRPII is a diagnostic and prognostic marker for falciparum malaria. We demonstrate that disruption of a human cerebral microvascular endothelial barrier by P. falciparum-infected erythrocytes depends on expression of HRPII. Purified recombinant or native HRPII can recapitulate these effects. HRPII action occurs via activation of the inflammasome, resulting in decreased integrity of tight junctions and increased endothelial permeability. We propose that HRPII is a virulence factor that may contribute to cerebral malaria by compromising endothelial barrier integrity within the central nervous system. PMID:27273825

  4. Changes of soluble CD40 ligand in the progression of acute myocardial infarction associate to endothelial nitric oxide synthase polymorphisms and vascular endothelial growth factor but not to platelet CD62P expression.

    PubMed

    Napoleão, Patrícia; Monteiro, Maria do Céu; Cabral, Luís B P; Criado, Maria Begoña; Ramos, Catarina; Selas, Mafalda; Viegas-Crespo, Ana Maria; Saldanha, Carlota; Carmo, Miguel Mota; Ferreira, Rui Cruz; Pinheiro, Teresa

    2015-12-01

    Reported in vitro data implicated soluble CD40 ligand (sCD40L) in endothelial dysfunction and angiogenesis. However, whether sCD40L could exert that influence in endothelial dysfunction and angiogenesis after injury in acute myocardial infarction (AMI) patients remains unclear. In the present study, we evaluated the association of sCD40L with markers of platelet activation, endothelial, and vascular function during a recovery period early after AMI. To achieve this goal, the time changes of soluble, platelet-bound, and microparticle-bound CD40L levels over 1 month were assessed in AMI patients and correlated with endothelial nitric oxide synthase (eNOS) polymorphisms, vascular endothelial growth factor (VEGF) concentrations, and platelet expression of P-selectin (CD62P). The association of soluble form, platelet-bound, and microparticle-bound CD40L with CD62P expression on platelets, a marker of platelet activation, was also assessed to evaluate the role of CD40L in the thrombosis, whereas the association with eNOS and VEGF was to evaluate the role of CD40L in vascular dysfunction. This work shows for the first time that time changes of sCD40L over 1 month after myocardial infarct onset were associated with G894T eNOS polymorphism and with the VEGF concentrations, but not to the platelet CD62P expression. These results indicate that, in terms of AMI pathophysiology, the sCD40L cannot be consider just as being involved in thrombosis and inflammation but also as having a relevant role in vascular and endothelial dysfunction. PMID:26279254

  5. Novel heart valve prosthesis with self-endothelialization potential made of modified polyhedral oligomeric silsesquioxane-nanocomposite material.

    PubMed

    Ghanbari, Hossein; Radenkovic, Dina; Marashi, Sayed Mahdi; Parsno, Shirin; Roohpour, Nima; Burriesci, Gaetano; Seifalian, Alexander M

    2016-06-01

    In the cardiovascular system, the endothelial layer provides a natural antithrombogenic surface on the inner portion of the heart and associated vessels. For a synthetic material therefore, the ability to attract and retain endothelial or endothelial progenitor cells (EPCs), ultimately creating a single endothelial layer on its surface, is of prime importance. The authors have developed a nanocomposite polymer, based on a combination of polyhedral oligomeric silsesquioxane nanoparticles and polycarbonate urea urethane (POSS-PCU), which is biocompatible and has been used in human for the world's first synthetic trachea, tear duct, and bypass graft. In this study, the authors modified the surface of this casted nanocomposite by grafting fibronectin derived bioactive peptides [glycine-arginine-glycine-aspartic acid-glycine (GRGDG) and lauric acid conjugated GRGDG (GRGDG-LA)] to enhance the endothelialization for using heart valves leaflets from circulating EPCs. Human peripheral blood mononuclear cells were separated using Ficoll-Paque centrifugation, with harvested EPCs purified using CD34 microbead labeling and magnetic-activated cell sorting. Cells were seeded onto 96 well plates coated with POSS-PCU, GRGDG/GRGDG-LA modified POSS-PCU and PCU polymers, for a period of 21 days. Cells were studied under light, confocal, and scanning electron microscope (SEM). Fluorescence-activated cell sorting was used to analyze cell surface markers. Cell attachment and proliferation was observed in all POSS-PCU samples, significantly higher than the activity seen within the control PCU polymers (p < 0.05). Microscopic examination revealed clonal expansion and morphological changes in cells seeded on POSS-PCU. The cells expressed increasing levels of mature endothelial cell markers over time with a concurrent reduction in hematopoietic stem cell marker expression. SEM showed a mixed population of morphologically differentiated endothelial cells and EPCs. These results support

  6. Soluble endoglin, hypercholesterolemia and endothelial dysfunction.

    PubMed

    Rathouska, Jana; Jezkova, Katerina; Nemeckova, Ivana; Nachtigal, Petr

    2015-12-01

    A soluble form of endoglin (sEng) is known to be an extracellular domain of the full-length membrane endoglin, which is elevated during various pathological conditions related to vascular endothelium. In the current review, we tried to summarize a possible role of soluble endoglin in cardiovascular pathologies, focusing on its relation to endothelial dysfunction and cholesterol levels. We discussed sEng as a proposed biomarker of cardiovascular disease progression, cardiovascular disease treatment and endothelial dysfunction. We also addressed a potential interaction of sEng with TGF-β/eNOS or BMP-9 signaling. We suggest soluble endoglin levels to be monitored, because they reflect the progression/treatment efficacy of cardiovascular diseases related to endothelial dysfunction and hypercholesterolemia. A possible role of soluble endoglin as an inducer of endothelial dysfunction however remains to be elucidated. PMID:26520890

  7. Apicobasal polarity of brain endothelial cells.

    PubMed

    Worzfeld, Thomas; Schwaninger, Markus

    2016-02-01

    Normal brain homeostasis depends on the integrity of the blood-brain barrier that controls the access of nutrients, humoral factors, and immune cells to the CNS. The blood-brain barrier is composed mainly of brain endothelial cells. Forming the interface between two compartments, they are highly polarized. Apical/luminal and basolateral/abluminal membranes differ in their lipid and (glyco-)protein composition, allowing brain endothelial cells to secrete or transport soluble factors in a polarized manner and to maintain blood flow. Here, we summarize the basic concepts of apicobasal cell polarity in brain endothelial cells. To address potential molecular mechanisms underlying apicobasal polarity in brain endothelial cells, we draw on investigations in epithelial cells and discuss how polarity may go awry in neurological diseases. PMID:26661193

  8. Genetics Home Reference: Fuchs endothelial dystrophy

    MedlinePlus

    ... a protein that is part of type VIII collagen. Type VIII collagen is largely found within the cornea, surrounding the endothelial cells. Specifically, type VIII collagen is a major component of a tissue at ...

  9. Endothelial glucocorticoid receptor suppresses atherogenesis- Brief Report

    PubMed Central

    Zhang, Xinbo; Rotllan, Noemi; Feng, Yan; Zhou, Han; Fernández-Hernando, Carlos; Yu, Jun; Sessa, William C.

    2015-01-01

    Objective The purpose of this study was to determine the role of the endothelial glucocorticoid receptor in the pathogenesis of atherosclerosis. Approach and Results Control mice and mice lacking the endothelial glucocorticoid receptor were bred onto an Apoe knockout background and subjected to high-fat diet feeding for 12 weeks. Assessment of body weight and total cholesterol and triglycerides before and after the diet revealed no differences between the two groups of mice. However, mice lacking the endothelial glucocorticoid receptor developed more severe atherosclerotic lesions in the aorta, brachiocephalic artery and aortic sinus as well as a heightened inflammatory milieu as evidence by increased macrophage recruitment in the lesions. Conclusions These data suggest the endothelial glucocorticoid receptor is important for tonic inhibition of inflammation and limitation of atherosclerosis progression in this model. PMID:25810297

  10. Specular Microscopic Features of Corneal Endothelial Vacuolation

    PubMed Central

    Kanavi, Mozhgan Rezaei; Javadi, Mohammad-Ali; Chamani, Tahereh

    2011-01-01

    Purpose To introduce a specular microscopic reference image for endothelial vacuolation in donated corneas. Methods Two corneas from a donor with diffuse, round to oval dark areas at the endothelial level on slit lamp biomicroscopy and one normal-appearing donor cornea underwent specular microscopy, histopathologic evaluation and transmission electron microscopy. Results Specular microscopy of the two corneas with abnormal-looking endothelium revealed large numbers of dark, round to oval structures within the endothelium in favor of endothelial vacuolation. Light microscopy disclosed variable sized cyst-like structures within the cytoplasm. Transmission electron microscopy showed electron-lucent and relatively large-sized intracytoplasmic vacuoles. These features were not observed in the endothelium of the normal cornea. Conclusion The specular microscopic features of endothelial vacuolation in donated corneas were confirmed by light microscopy and transmission electron microscopy, therefore the specular image may be proposed as a reference to eye banks. PMID:22454699

  11. The endothelial glycocalyx: composition, functions, and visualization

    PubMed Central

    Reitsma, Sietze; Slaaf, Dick W.; Vink, Hans; van Zandvoort, Marc A. M. J.

    2007-01-01

    This review aims at presenting state-of-the-art knowledge on the composition and functions of the endothelial glycocalyx. The endothelial glycocalyx is a network of membrane-bound proteoglycans and glycoproteins, covering the endothelium luminally. Both endothelium- and plasma-derived soluble molecules integrate into this mesh. Over the past decade, insight has been gained into the role of the glycocalyx in vascular physiology and pathology, including mechanotransduction, hemostasis, signaling, and blood cell–vessel wall interactions. The contribution of the glycocalyx to diabetes, ischemia/reperfusion, and atherosclerosis is also reviewed. Experimental data from the micro- and macrocirculation alludes at a vasculoprotective role for the glycocalyx. Assessing this possible role of the endothelial glycocalyx requires reliable visualization of this delicate layer, which is a great challenge. An overview is given of the various ways in which the endothelial glycocalyx has been visualized up to now, including first data from two-photon microscopic imaging. PMID:17256154

  12. Fuchs endothelial corneal dystrophy: current perspectives

    PubMed Central

    Vedana, Gustavo; Villarreal, Guadalupe; Jun, Albert S

    2016-01-01

    Fuchs endothelial corneal dystrophy (FECD) is the most common corneal dystrophy and frequently results in vision loss. Hallmarks of the disease include loss of corneal endothelial cells and formation of excrescences of Descemet’s membrane. Later stages involve all layers of the cornea. Impairment of endothelial barrier and pump function and cell death from oxidative and unfolded protein stress contribute to disease progression. The genetic basis of FECD includes numerous genes and chromosomal loci, although alterations in the transcription factor 4 gene are associated with the majority of cases. Definitive treatment of FECD is corneal transplantation. In this paper, we highlight advances that have been made in understanding FECD’s clinical features, pathophysiology, and genetics. We also discuss recent advances in endothelial keratoplasty and potential future treatments. PMID:26937169

  13. Endothelial Cell Stimulation by Candida albicans

    PubMed Central

    Phan, Quynh T.; Filler, Scott G.

    2013-01-01

    The opportunistic fungal pathogen, Candida albicans, enters the bloodstream and causes hematogenously disseminated infection in hospitalized patients. During the initiation of a hematogenously disseminated infection, endothelial cells are one of the first host cells to come in contact with C. albicans. Endothelial cells can significantly influence the local host response to C. albicans by expressing leukocyte adhesion molecules and pro-inflammatory cytokines. Thus, it is of interest to investigate the response of endothelial cells to C. albicans in vitro. We describe the use of real-time PCR and enzyme immunoassays to measure the effects of C. albicans on the endothelial cell production of E-selectin and tumor necrosis factor α in vitro. PMID:19089392

  14. Strategy Markers in Writing.

    ERIC Educational Resources Information Center

    Sindermann, Gerda; Horsella, Maria

    1989-01-01

    Part of a research project is presented that aims at identifying the difficulties tertiary level students encounter in reading scientific texts in a foreign language and the strategies they apply to overcome them. Strategy markers are identified and listed, and are then analyzed to interpret the linguistic difficulty and the strategy used.…

  15. The Swift Turbidity Marker

    ERIC Educational Resources Information Center

    Omar, Ahmad Fairuz; MatJafri, Mohd Zubir

    2011-01-01

    The Swift Turbidity Marker is an optical instrument developed to measure the level of water turbidity. The components and configuration selected for the system are based on common turbidity meter design concepts but use a simplified methodology to produce rapid turbidity measurements. This work is aimed at high school physics students and is the…

  16. Effects of industrially produced trans fat on markers of systemic inflammation: evidence from a randomized trial in women

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  17. Endothelial microparticles: Sophisticated vesicles modulating vascular function

    PubMed Central

    Curtis, Anne M; Edelberg, Jay; Jonas, Rebecca; Rogers, Wade T; Moore, Jonni S; Syed, Wajihuddin; Mohler, Emile R

    2015-01-01

    Endothelial microparticles (EMPs) belong to a family of extracellular vesicles that are dynamic, mobile, biological effectors capable of mediating vascular physiology and function. The release of EMPs can impart autocrine and paracrine effects on target cells through surface interaction, cellular fusion, and, possibly, the delivery of intra-vesicular cargo. A greater understanding of the formation, composition, and function of EMPs will broaden our understanding of endothelial communication and may expose new pathways amenable for therapeutic manipulation. PMID:23892447

  18. 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

  19. Developing a biohybrid lung - sufficient endothelialization of poly-4-methly-1-pentene gas exchange hollow-fiber membranes.

    PubMed

    Wiegmann, Bettina; von Seggern, Heide; Höffler, Klaus; Korossis, Sotirios; Dipresa, Daniele; Pflaum, Michael; Schmeckebier, Sabrina; Seume, Jörg; Haverich, Axel

    2016-07-01

    Working towards establishing a biohybrid lung with optimized hemocompatibility, this study analyzed the feasibility of establishing flow-resistant endothelium on heparin/albumin coated poly-4-methly-1-pentene hollow fiber gas exchange membranes (PMP-HFs). The seeding efficiency and proliferation of human cord blood derived endothelial cells (HCBEC) on PMP-HFs were analyzed under static conditions by WST-8 cell proliferation assay and fluorescence microscopy. The HCBEC monolayer integrity under different flow conditions was also assessed. Endothelial-specific phenotype verification, expression activation levels and thrombogenic state markers were quantified by real-time RT-PCR for cell-to-PMP-HF contact under static and dynamic conditions. The results demonstrated the feasibility of establishing a viable, confluent, and flow-resistant endothelial monolayer on the blood-contact surface of PMP-HFs, which maintained a physiological response to TNFα-stimulation and flow conditions. The endothelial phenotype, expression levels of adhesion molecules and thrombogenic state markers were unaffected by cell-to-PMP-HFs contact. These results represent a significant step towards establishing a biohybrid lung. PMID:26919566

  20. Paeonol protects against endoplasmic reticulum stress-induced endothelial dysfunction via AMPK/PPARδ signaling pathway.

    PubMed

    Choy, Ker-Woon; Mustafa, Mohd Rais; Lau, Yeh Siang; Liu, Jian; Murugan, Dharmani; Lau, Chi Wai; Wang, Li; Zhao, Lei; Huang, Yu

    2016-09-15

    Endoplasmic reticulum (ER) stress in endothelial cells often leads to endothelial dysfunction which underlies the pathogenesis of cardiovascular diseases. Paeonol, a major phenolic component extracted from Moutan Cortex, possesses various medicinal benefits which have been used extensively in traditional Chinese medicine. The present study investigated the protective mechanism of paeonol against tunicamycin-induced ER stress in isolated mouse aortas and human umbilical vein endothelial cells (HUVECs). Vascular reactivity in aorta was measured using a wire myograph. The effects of paeonol on protein expression of ER stress markers, reactive oxygen species (ROS) production, nitric oxide (NO) bioavailability and peroxisome proliferator-activated receptor δ (PPARδ) activity in the vascular wall were assessed by Western blot, dihydroethidium fluorescence (DHE) or lucigenin enhanced-chemiluminescence, 4-amino-5-methylamino-2',7'-difluorofluorescein (DAF-FM DA) and dual luciferase reporter assay, respectively. Ex vivo treatment with paeonol (0.1μM) for 16h reversed the impaired endothelium-dependent relaxations in C57BJ/6J and PPARδ wild type (WT) mouse aortas following incubation with tunicamycin (0.5μg/mL). Elevated ER stress markers, oxidative stress and reduction of NO bioavailability induced by tunicamycin in HUVECs, C57BJ/6J and PPARδ WT mouse aortas were reversed by paeonol treatment. These beneficial effects of paeonol were diminished in PPARδ knockout (KO) mouse aortas. Paeonol increased the expression of 5' adenosine monophosphate-activated protein kinase (AMPK) and PPARδ expression and activity while restoring the decreased phosphorylation of eNOS. The present study delineates that paeonol protects against tunicamycin-induced vascular endothelial dysfunction by inhibition of ER stress and oxidative stress, thus elevating NO bioavailability via the AMPK/PPARδ signaling pathway. PMID:27449753

  1. Blocking Notch in endothelial cells prevents arteriovenous fistula failure despite CKD.

    PubMed

    Wang, Yun; Liang, Anlin; Luo, Jinlong; Liang, Ming; Han, Guofeng; Mitch, William E; Cheng, Jizhong

    2014-04-01

    Neointima formation causes the failure of 60% of arteriovenous fistulas (AVFs) within 2 years. Neointima-forming mechanisms are controversial but possibly linked to excess proinflammatory responses and dysregulated Notch signaling. To identify how AVFs fail, we anastomosed the carotid artery to the internal jugular vein in normal and uremic mice and compared these findings with those in failed AVFs from patients with ESRD. Endothelial cells (ECs) of AVFs in uremic mice or patients expressed mesenchymal markers (FSP-1 and/or α-SMA) and exhibited increased expression and nuclear localization of Notch intracellular domain compared with ECs of AVFs in pair-fed control mice. Furthermore, expression of VE-Cadherin decreased, whereas expression of Notch1 and -4, Notch ligands, the downstream transcription factor of Notch, RBP-Jκ, and Notch target genes increased in ECs of AVFs in uremic mice. In cultured ECs, ectopic expression of Notch ligand or treatment with TGF-β1 triggered the expression of mesenchymal markers and induced endothelial cell barrier dysfunction, both of which were blocked by Notch inhibition or RBP-Jκ knockout. Furthermore, Notch-induced defects in barrier function, invasion of inflammatory cells, and neointima formation were suppressed in mice with heterozygous knockdown of endothelial-specific RBP-Jκ. These results suggest that increased TGF-β1, a complication of uremia, activates Notch in endothelial cells of AVFs, leading to accelerated neointima formation and AVF failure. Suppression of Notch activation could be a strategy for improving AFV function in uremia. PMID:24480830

  2. CD34+ Cells Represent Highly Functional Endothelial Progenitor Cells in Murine Bone Marrow

    PubMed Central

    Yang, Junjie; Ii, Masaaki; Kamei, Naosuke; Alev, Cantas; Kwon, Sang-Mo; Kawamoto, Atsuhiko; Akimaru, Hiroshi; Masuda, Haruchika; Sawa, Yoshiki; Asahara, Takayuki

    2011-01-01

    Background Endothelial progenitor cells (EPCs) were shown to have angiogenic potential contributing to neovascularization. However, a clear definition of mouse EPCs by cell surface markers still remains elusive. We hypothesized that CD34 could be used for identification and isolation of functional EPCs from mouse bone marrow. Methodology/Principal Findings CD34+ cells, c-Kit+/Sca-1+/Lin− (KSL) cells, c-Kit+/Lin− (KL) cells and Sca-1+/Lin− (SL) cells were isolated from mouse bone marrow mononuclear cells (BMMNCs) using fluorescent activated cell sorting. EPC colony forming capacity and differentiation capacity into endothelial lineage were examined in the cells. Although CD34+ cells showed the lowest EPC colony forming activity, CD34+ cells exhibited under endothelial culture conditions a more adherent phenotype compared with the others, demonstrating the highest mRNA expression levels of endothelial markers vWF, VE-cadherin, and Flk-1. Furthermore, a dramatic increase in immediate recruitment of cells to the myocardium following myocardial infarction and systemic cell injection was observed for CD34+ cells comparing with others, which could be explained by the highest mRNA expression levels of key homing-related molecules Integrin β2 and CXCR4 in CD34+ cells. Cell retention and incorporation into the vasculature of the ischemic myocardium was also markedly increased in the CD34+ cell-injected group, giving a possible explanation for significant reduction in fibrosis area, significant increase in neovascularization and the best cardiac functional recovery in this group in comparison with the others. Conclusion These findings suggest that mouse CD34+ cells may represent a functional EPC population in bone marrow, which could benefit the investigation of therapeutic EPC biology. PMID:21655289

  3. Foramen Ovale Closure Is a Process of Endothelial-to-Mesenchymal Transition Leading to Fibrosis

    PubMed Central

    Elliott, Graeme C.; Gurtu, Rockesh; McCollum, Charles; Newman, William G.; Wang, Tao

    2014-01-01

    Patent foramen ovale (PFO) is an atrial septal deformity present in around 25% of the general population. PFO is associated with major causes of morbidity, including stroke and migraine. PFO appears to be heritable but genes involved in the closure of foramen ovale have not been identified. The aim of this study is to determine molecular pathways and genes that are responsible to the postnatal closure of the foramen ovale. Using Sprague-Dawley rat hearts as a model we analysed the dynamic histological changes and gene expressions at the foramen ovale region between embryonic day 20 and postnatal day 7. We observed a gradual loss of the endothelial marker PECAM1, an upregulation of the mesenchymal marker vimentin and α-smooth muscle actin, the elevation of the transcription factor Snail, and an increase of fibroblast activation protein (FAP) in the foramen ovale region as well as the deposition of collagen-rich connective tissues at the closed foramen ovale, suggesting endothelial-to-mesenchymal transition (EndMT) occurring during foramen ovale closure which leads to fibrosis. In addition, Notch1 and Notch3 receptors, Notch ligand Jagged1 and Notch effector HRT1 were highly expressed in the endocardium of the foramen ovale region during EndMT. Activation of Notch3 alone in an endothelial cell culture model was able to drive EndMT and transform endothelial cells to mesenchymal phenotype. Our data demonstrate for the first time that FO closure is a process of EndMT-mediated fibrosis, and Notch signalling is an important player participating in this process. Elucidation of the molecular mechanisms of the closure of foramen ovale informs the pathogenesis of PFO and may provide potential options for screening and prevention of PFO related conditions. PMID:25215881

  4. Hematopoietic stem cells develop in the absence of endothelial cadherin 5 expression.

    PubMed

    Anderson, Heidi; Patch, Taylor C; Reddy, Pavankumar N G; Hagedorn, Elliott J; Kim, Peter G; Soltis, Kathleen A; Chen, Michael J; Tamplin, Owen J; Frye, Maike; MacLean, Glenn A; Hübner, Kathleen; Bauer, Daniel E; Kanki, John P; Vogin, Guillaume; Huston, Nicholas C; Nguyen, Minh; Fujiwara, Yuko; Paw, Barry H; Vestweber, Dietmar; Zon, Leonard I; Orkin, Stuart H; Daley, George Q; Shah, Dhvanit I

    2015-12-24

    Rare endothelial cells in the aorta-gonad-mesonephros (AGM) transition into hematopoietic stem cells (HSCs) during embryonic development. Lineage tracing experiments indicate that HSCs emerge from cadherin 5 (Cdh5; vascular endothelial-cadherin)(+) endothelial precursors, and isolated populations of Cdh5(+) cells from mouse embryos and embryonic stem cells can be differentiated into hematopoietic cells. Cdh5 has also been widely implicated as a marker of AGM-derived hemogenic endothelial cells. Because Cdh5(-/-) mice embryos die before the first HSCs emerge, it is unknown whether Cdh5 has a direct role in HSC emergence. Our previous genetic screen yielded malbec (mlb(bw306)), a zebrafish mutant for cdh5, with normal embryonic and definitive blood. Using time-lapse confocal imaging, parabiotic surgical pairing of zebrafish embryos, and blastula transplantation assays, we show that HSCs emerge, migrate, engraft, and differentiate in the absence of cdh5 expression. By tracing Cdh5(-/-)green fluorescent protein (GFP)(+/+) cells in chimeric mice, we demonstrated that Cdh5(-/-)GFP(+/+) HSCs emerging from embryonic day 10.5 and 11.5 (E10.5 and E11.5) AGM or derived from E13.5 fetal liver not only differentiate into hematopoietic colonies but also engraft and reconstitute multilineage adult blood. We also developed a conditional mouse Cdh5 knockout (Cdh5(flox/flox):Scl-Cre-ER(T)) and demonstrated that multipotent hematopoietic colonies form despite the absence of Cdh5. These data establish that Cdh5, a marker of hemogenic endothelium in the AGM, is dispensable for the transition of hemogenic endothelium to HSCs. PMID:26385351

  5. Vitamin D improves endothelial dysfunction and restores myeloid angiogenic cell function via reduced CXCL-10 expression in systemic lupus erythematosus.

    PubMed

    Reynolds, John A; Haque, Sahena; Williamson, Kate; Ray, David W; Alexander, M Yvonne; Bruce, Ian N

    2016-01-01

    Patients with systemic lupus erythematosus (SLE) have accelerated cardiovascular disease and dysfunctional endothelial repair mechanisms. Myeloid angiogenic cells (MACs), derived from circulating monocytes, augment vascular repair by paracrine secretion of pro-angiogenic factors. We observed that SLE MACs are dysfunctional and secrete pro-inflammatory cytokines. We also found that the vitamin D receptor was transiently expressed during MAC differentiation and that in vitro, calcitriol increased differentiation of monocytes into MACs in both SLE and in a model using the prototypic SLE cytokine, interferon-alpha. The active form of vitamin D (calcitriol) restored the SLE MAC phenotype towards that of healthy subjects with reduced IL-6 secretion, and normalised surface marker expression. Calcitriol also augmented the angiogenic capacity of MACs via the down-regulation of CXCL-10. In SLE patients treated with cholecalciferol for 12 weeks, the improvement in endothelial function correlated with increase in serum 25(OH)D concentrations independently of disease activity. We also show that MACs were able to positively modulate eNOS expression in human endothelial cells in vitro, an effect further enhanced by calcitriol treatment of SLE MACs. The results demonstrate that vitamin D can positively modify endothelial repair mechanisms and thus endothelial function in a population with significant cardiovascular risk. PMID:26930567

  6. Role of Epac1, an Exchange Factor for Rap GTPases, in Endothelial Microtubule Dynamics and Barrier Function

    PubMed Central

    Sehrawat, Seema; Cullere, Xavier; Patel, Sunita; Italiano, Joseph

    2008-01-01

    Rap1 GTPase activation by its cAMP responsive nucleotide exchange factor Epac present in endothelial cells increases endothelial cell barrier function with an associated increase in cortical actin. Here, Epac1 was shown to be responsible for these actin changes and to colocalize with microtubules in human umbilical vein endothelial cells. Importantly, Epac activation with a cAMP analogue, 8-pCPT-2′O-Me-cAMP resulted in a net increase in the length of microtubules. This did not require cell–cell interactions or Rap GTPase activation, and it was attributed to microtubule growth as assessed by time-lapse microscopy of human umbilical vein endothelial cell expressing fluorophore-linked microtubule plus-end marker end-binding protein 3. An intact microtubule network was required for Epac-mediated changes in cortical actin and barrier enhancement, but it was not required for Rap activation. Finally, Epac activation reversed microtubule-dependent increases in vascular permeability induced by tumor necrosis factor-α and transforming growth factor-β. Thus, Epac can directly promote microtubule growth in endothelial cells. This, together with Rap activation leads to an increase in cortical actin, which has functional significance for vascular permeability. PMID:18172027

  7. Vitamin D improves endothelial dysfunction and restores myeloid angiogenic cell function via reduced CXCL-10 expression in systemic lupus erythematosus

    PubMed Central

    Reynolds, John A.; Haque, Sahena; Williamson, Kate; Ray, David W.; Alexander, M. Yvonne; Bruce, Ian N.

    2016-01-01

    Patients with systemic lupus erythematosus (SLE) have accelerated cardiovascular disease and dysfunctional endothelial repair mechanisms. Myeloid angiogenic cells (MACs), derived from circulating monocytes, augment vascular repair by paracrine secretion of pro-angiogenic factors. We observed that SLE MACs are dysfunctional and secrete pro-inflammatory cytokines. We also found that the vitamin D receptor was transiently expressed during MAC differentiation and that in vitro, calcitriol increased differentiation of monocytes into MACs in both SLE and in a model using the prototypic SLE cytokine, interferon-alpha. The active form of vitamin D (calcitriol) restored the SLE MAC phenotype towards that of healthy subjects with reduced IL-6 secretion, and normalised surface marker expression. Calcitriol also augmented the angiogenic capacity of MACs via the down-regulation of CXCL-10. In SLE patients treated with cholecalciferol for 12 weeks, the improvement in endothelial function correlated with increase in serum 25(OH)D concentrations independently of disease activity. We also show that MACs were able to positively modulate eNOS expression in human endothelial cells in vitro, an effect further enhanced by calcitriol treatment of SLE MACs. The results demonstrate that vitamin D can positively modify endothelial repair mechanisms and thus endothelial function in a population with significant cardiovascular risk. PMID:26930567

  8. IFNα Serum Levels Are Associated with Endothelial Progenitor Cells Imbalance and Disease Features in Rheumatoid Arthritis Patients

    PubMed Central

    Rodríguez-Carrio, Javier; de Paz, Banesa; López, Patricia; Prado, Catuxa; Alperi-López, Mercedes; Ballina-García, Francisco Javier; Suárez, Ana

    2014-01-01

    Introduction IFNα has been largely implicated in the ethiopathogenesis of autoimmune diseases but only recently it has been linked to endothelial damage and accelerated atherosclerosis in autoimmunity. In addition, proinflammatory conditions are supposed to be implicated in the cardiovascular status of these patients. Since a role for IFNα in endothelial damage and impaired Endothelial Progenitor Cell (EPC) number and function has been reported in other diseases, we aimed to evaluate the potential associations of IFNα serum levels on EPC populations and cytokine profiles in Rheumatoid Arthritis (RA) patients. Methods pre-EPC, EPC and mature EPC (mEPC) populations were quantified by flow cytometry analyzing their differential CD34, CD133 and VEGFR2 expression in blood samples from 120 RA patients, 52 healthy controls (HC), and 83 systemic lupus erythematosus (SLE) patients as disease control. Cytokine serum levels were measured by immunoassays and clinical and immunological data, including cardiovascular (CV) events and CV risk factors, were retrospectively obtained by reviewing clinical records. Results Long-standing, but not recent onset RA patients displayed a significant depletion of all endothelial progenitor populations, unless high IFNα levels were present. In fact, the IFNhigh RA patient group (n = 40, 33%), showed increased EPC levels, comparable to SLE patients. In addition, high IFNα serum levels were associated with higher disease activity (DAS28), presence of autoantibodies, higher levels of IL-1β, IL-6, IL-10 and MIP-1α, lower amounts of TGF-β, and increased mEPC/EPC ratio, thus suggesting higher rates of endothelial damage and an endothelial repair failure. Finally, the relationship between high IFNα levels and occurrence of CV events observed in RA patients seems to support this hypothesis. Conclusions IFNα serum marker could be used to identify a group of RA patients with increased disease activity, EPC imbalance, enhanced

  9. Lipoprotein marker for hypertriglyceridemia

    DOEpatents

    Cubicciotti, Roger S.; Karu, Alexander E.; Krauss, Ronald M.

    1986-01-01

    Methods and compositions are provided for the detection of a particular low density lipoprotein which has been found to be a marker for patients suffering from type IV hypertriglyceridemia. A monoclonal antibody capable of specifically binding to a characteristic epitopic site on this LDL subspecies can be utilized in a wide variety of immunoassays. Hybridoma cell line SPL.IVA5A1 was deposited at the American Type Culture Collection on Mar. 29, 1984, and granted accession no. HB 8535.

  10. Aging and vascular endothelial function in humans

    PubMed Central

    SEALS, Douglas R.; JABLONSKI, Kristen L.; DONATO, Anthony J.

    2012-01-01

    Advancing age is the major risk factor for the development of CVD (cardiovascular diseases). This is attributable, in part, to the development of vascular endothelial dysfunction, as indicated by reduced peripheral artery EDD (endothelium-dependent dilation) in response to chemical [typically ACh (acetylcholine)] or mechanical (intravascular shear) stimuli. Reduced bioavailability of the endothelium-synthesized dilating molecule NO (nitric oxide) as a result of oxidative stress is the key mechanism mediating reduced EDD with aging. Vascular oxidative stress increases with age as a consequence of greater production of reactive oxygen species (e.g. superoxide) without a compensatory increase in antioxidant defences. Sources of increased superoxide production include up-regulation of the oxidant enzyme NADPH oxidase, uncoupling of the normally NO-producing enzyme, eNOS (endothelial NO synthase) (due to reduced availability of the cofactor tetrahydrobiopterin) and increased mitochondrial synthesis during oxidative phosphorylation. Increased bioactivity of the potent endothelial-derived constricting factor ET-1 (endothelin-1), reduced endothelial production of/responsiveness to dilatory prostaglandins, the development of vascular inflammation, formation of AGEs (advanced glycation end-products), an increased rate of endothelial apoptosis and reduced expression of oestrogen receptor α (in postmenopausal females) also probably contribute to impaired EDD with aging. Several lifestyle and biological factors modulate vascular endothelial function with aging, including regular aerobic exercise, dietary factors (e.g. processed compared with non-processed foods), body weight/fatness, vitamin D status, menopause/oestrogen deficiency and a number of conventional and non-conventional risk factors for CVD. Given the number of older adults now and in the future, more information is needed on effective strategies for the prevention and treatment of vascular endothelial aging. PMID

  11. Autonomic Neuropathy and Endothelial Dysfunction in Patients With Impaired Glucose Tolerance or Type 2 Diabetes Mellitus.

    PubMed

    Tiftikcioglu, Bedile Irem; Bilgin, Sule; Duksal, Tarik; Kose, Sukran; Zorlu, Yasar

    2016-04-01

    Autonomic neuropathy is one of the most common complications of diabetes mellitus (DM). The etiology of autonomic impairment is not well-understood, yet. There is need for studies to investigate the cause-effect relationships of inflammation and/or endothelial dysfunction and diabetic autonomic neuropathy. Only a few reports have mentioned autonomic neuropathy in individuals with impaired glucose tolerance (IGT), previously. Furthermore, the association between the plasma markers of endothelial dysfunction (von Willebrand factor (vWF), soluble E-selectin) and autonomic neuropathy in patients with IGT or DM has not been studied before. In this study, we aimed to investigate the correlation between plasma markers of endothelial dysfunction and autonomic neuropathy in patients with IGT or type 2 DM (T2DM).In this case-control study, 25 IGT patients, 25 T2DM patients with autonomic symptoms, and 30 controls were included. Demographical data, HbA1c, vWF, and soluble E-selectin (sE-selectin) levels were analyzed. Sympathetic skin response (SSR) and heart rate variability (HRV) were used as the indicator of autonomic activity.Plasma levels of HbA1c, vWF, and sE-selectin were higher in patients with IGT than the controls; patients with T2DM had higher levels than both the controls and the patients with IGT. SSR measures were similar among the groups. However, higher number of T2DM patients had absent plantar SSR than controls. HRV analysis at rest revealed lower standard deviation of R-R interval, coefficient of variation of R-R interval, low-frequency (LF) power and total power in patients with IGT and T2DM than the controls. In addition, HRV analysis at deep breathing showed lower high-frequency (HF) power in IGT group. LF:HF ratio was lower in both patient groups at rest. No strong correlation was found between the levels of HbA1c, vWF, sE-selectin, HRV, and SSR measures.Our results support that endothelial dysfunction is evident in individuals with IGT or T2DM and HRV

  12. Post-Exercise Ankle-Brachial Pressure Index Demonstrates Altered Endothelial Function in the Elderly

    PubMed Central

    Sato, Shinji; Masami, Kozue; Otsuki, Shingo; Tanaka, Shiro; Nakayama, Noriko; Makita, Shigeru; Koshiyama, Hiroyuki; Nohara, Ryuji

    2011-01-01

    Background: The ankle-brachial pressure index (ABI), the ratio of the systolic blood pressure of the ankle to the systolic brachial pressure, is commonly measured at rest, but ABI values post-exercise enhance the sensitivity of the test and can be used to identify atherosclerotic vascular damage. However, it has not been established whether or not enhanced post-exercise ABI is also associated with endothelial dysfunction. We hypothesized that a decrease in post-exercise ABI is related to impaired endothelial function. Purpose: To investigate alterations in post-exercise ABI values and endothelial dysfunction in the elderly. Methods: The study population comprised 35 men and women aged 51–77 years (mean age: 66 years). Patients with peripheral arterial disease or a history of heart failure were excluded. The ABI was estimated at rest and immediately after exercise. The exercise protocol comprised 2.5 min of active pedal flexion exercises at a speed of 60 times/min. Endothelial function was assessed by measuring flow-mediated vasodilation (FMD) in the brachial artery using ultrasound imaging. Results: No correlation was found between FMD and the ABI at rest. However, a weak correlation was found between FMD and post-exercise ABI (r = 0.46, P = 0.06). A strong correlation was observed between FMD and a decrease in post-exercise ABI compared to baseline readings (r = −0.52, P = 0.01). Multiple linear regression analysis was used to generate a prediction equation for FMD using the percentage decrease in post-exercise ABI. Significant correlations were observed between the ultrasound imaging-measured FMD and the predicted FMD (R2 = 0.27, P = 0.001). Conclusions: Post-exercise ABI appears to be a simple surrogate marker for endothelial function in the elderly, although larger studies are required for validation. PMID:23885186

  13. IL-4 Causes Hyperpermeability of Vascular Endothelial Cells through Wnt5A Signaling

    PubMed Central

    Skaria, Tom; Burgener, Julia; Bachli, Esther; Schoedon, Gabriele

    2016-01-01

    Microvascular leakage due to endothelial barrier dysfunction is a prominent feature of T helper 2 (Th2) cytokine mediated allergic inflammation. Interleukin-4 (IL-4) is a potent Th2 cytokine, known to impair the barrier function of endothelial cells. However, the effectors mediating IL-4 induced cytoskeleton remodeling and consequent endothelial barrier dysfunction remain poorly defined. Here we have used whole genome transcriptome profiling and gene ontology analyses to identify the genes and processes regulated by IL-4 signaling in human coronary artery endothelial cells (HCAEC). The study revealed Wnt5A as an effector that can mediate actin cytoskeleton remodeling in IL-4 activated HCAEC through the regulation of LIM kinase (LIMK) and Cofilin (CFL). Following IL-4 treatment, LIMK and CFL were phosphorylated, thereby indicating the possibility of actin stress fiber formation. Imaging of actin showed the formation of stress fibers in IL-4 treated live HCAEC. Stress fiber formation was notably decreased in the presence of Wnt inhibitory factor 1 (WIF1). Non-invasive impedance measurements demonstrated that IL-4 increased the permeability and impaired the barrier function of HCAEC monolayers. Silencing Wnt5A significantly reduced permeability and improved the barrier function of HCAEC monolayers upon IL-4 treatment. Our study identifies Wnt5A as a novel marker of IL-4 activated vascular endothelium and demonstrates a critical role for Wnt5A in mediating IL-4 induced endothelial barrier dysfunction. Wnt5A could be a potential therapeutic target for reducing microvascular leakage and edema formation in Th2 driven inflammatory diseases. PMID:27214384

  14. 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

  15. [Molecular markers of carcinogenesis in the diagnostics of cervical cancer].

    PubMed

    Bedkowska, Grazyna Ewa; Ławicki, Sławomir; Szmitkowski, Maciej

    2009-01-01

    Cervical carcinoma is the most frequent disease of the reproductive organ and is the second most common cancer in women after breast cancer. As it is characterized by high mortality, new diagnostic methods are needed, for example tumor markers, enabling earlier diagnosis and rapid detection of recurrence after therapy. Different tumor markers may be useful in the diagnostics of cervical cancer, for example squamous cell carcinoma antigen (SCC-Ag), tissue polypeptide antigen (TPA), and CYFRA 21-1, as well as some cytokines such as vascular endothelial growth factor (VEGF), granulocyte colony-stimulating factor, and macrophage colony-stimulating factor (M-CSF). About 150 genes connected with the carcinogenesis of cervical carcinoma have been identified. This paper is devoted to evaluating the diagnostic usefulness of molecular markers of carcinogenesis, especially P53, Bcl-2, Brn-3a, and MCM, and comparing the results with those of typical tumor markers or cytokines useful in diagnosing this type of cancer. It was shown that telomerase and Brn-3a proteins demonstrate usefulness in screening examination, P53 in monitoring the effectiveness of therapy, and Bcl-2 as a survival prognostic factor. In summary, it is evident that molecular makers of carcinogenesis are helpful in the diagnostics of cervical cancer, but further investigation and confirmation by a prospective study is necessary. PMID:19252468

  16. Endothelial damage in major depression patients is modulated by SSRI treatment, as demonstrated by circulating biomarkers and an in vitro cell model.

    PubMed

    Lopez-Vilchez, I; Diaz-Ricart, M; Navarro, V; Torramade, S; Zamorano-Leon, J; Lopez-Farre, A; Galan, A M; Gasto, C; Escolar, G

    2016-01-01

    There is a link between depression, cardiovascular events and inflammation. We have explored this connection through endothelial dysfunction, using in vivo and in vitro approaches. We evaluated circulating biomarkers of endothelial dysfunction in patients with major depression at their diagnosis (MD-0) and during antidepressant treatment with the selective serotonin reuptake inhibitor escitalopram, for 8 and 24 weeks (MD-8 and MD-24). Results were always compared with matched healthy controls (CON). We measured in vivo circulating endothelial cells (CECs) and endothelial progenitor cells (EPCs) in blood samples, and assessed plasma levels of soluble von Willebrand factor (VWF) and vascular cell adhesion molecule-1 (VCAM-1). CEC counts, soluble VWF and VCAM-1 were statistically elevated in MD-0 (P<0.01 versus CON) and gradually decreased during treatment. Conversely, EPC levels were lower in MD-0, tending to increase throughout treatment. In vitro studies were performed in human endothelial cells cultured in the presence of sera from each study group. Elevated expression of the inflammation marker intercellular adhesion molecule-1 and oxidative stress, with lower presence of endothelial nitric oxide synthase and higher reactive oxygen species production, were found in cells exposed to MD-0 sera (P<0.05 versus CON). These results were normalized in cells exposed to MD-24 sera. Thrombogenicity of extracellular matrices generated by these cells, measured as expression of VWF, tissue factor and platelet reactivity, showed non-significant differences. We provide a model of cultured endothelial cells reproducing endothelial dysfunction in naive patients with major depression, demonstrating endothelial damage and inflammation at diagnosis, and recovering with selective serotonin reuptake inhibitor treatment for 24 weeks. PMID:27598970

  17. Circulating endothelial cells and their progenitors in acute myeloid leukemia

    PubMed Central

    Zahran, Asmaa Mohammed; Aly, Sanaa Shaker; Altayeb, Hanan Ahmed; Ali, Arwa Mohammed

    2016-01-01

    Acute myeloid leukemia (AML) is an aggressive hematological malignancy characterized by the accumulation of immature myeloid progenitor cells in the bone marrow. Studies are required to investigate the prognostic and predictive value of surrogate biomarkers. Given the importance of angiogenesis in oncology in terms of pathogenesis as well as being a target for treatment, circulating endothelial cells (CECs) and endothelial progenitor cells (EPCs) are promising candidates to serve as such markers. The aim of the present study was to quantify CECs and EPCs in patients with AML at initial diagnosis and following induction chemotherapy, and to correlate these findings with the response to treatment in AML patients. The present study included 40 patients with de novo AML and 20 age- and gender-matched healthy controls. CECs and EPCs were evaluated by flow cytometry at initial diagnosis and after induction chemotherapy (3+7 protocol for AML other than M3 and all-trans-retinoic acid plus anthracycline for M3 disease). CECs and EPCs were significantly higher in AML patients at diagnosis and after induction chemotherapy than in controls. After induction chemotherapy, CECs and EPCs were significantly decreased compared with the levels at initial diagnosis. Patients who achieved complete response (n=28) had lower initial CEC and EPC levels compared with patients who did not respond to treatment. These results suggest that CEC levels are higher in AML patients and may correlate with disease status and treatment response. Further investigations are required to better determine the predictive value and implication of these cells in AML management. PMID:27602121

  18. Protection against Recurrent Stroke with Resveratrol: Endothelial Protection

    PubMed Central

    Clark, Darren; Tuor, Ursula I.; Thompson, Roger; Institoris, Adam; Kulynych, Angela; Zhang, Xu; Kinniburgh, David W.; Bari, Ferenc; Busija, David W.; Barber, Philip A.

    2012-01-01

    Despite increased risk of a recurrent stroke following a minor stroke, information is minimal regarding the interaction between injurious mild cerebral ischemic episodes and the possible treatments which might be effective. The aim of the current study was to investigate recurrent ischemic stroke and whether resveratrol, a nutritive polyphenol with promising cardio- and neuro- protective properties, could ameliorate the associated brain damage. Experiments in adult rats demonstrated that a mild ischemic stroke followed by a second mild cerebral ischemia exacerbated brain damage, and, daily oral resveratrol treatment after the first ischemic insult reduced ischemic cell death with the recurrent insult (P<0.002). Further investigation demonstrated reduction of both inflammatory changes and markers of oxidative stress in resveratrol treated animals. The protection observed with resveratrol treatment could not be explained by systemic effects of resveratrol treatment including effects either on blood pressure or body temperature measured telemetrically. Investigation of resveratrol effects on the blood-brain barrier in vivo demonstrated that resveratrol treatment reduced blood-brain barrier disruption and edema following recurrent stroke without affecting regional cerebral blood flow. Investigation of the mechanism in primary cell culture studies demonstrated that resveratrol treatment significantly protected endothelial cells against an in vitro ‘ischemia’ resulting in improved viability against oxygen and glucose deprivation (39.6±6.6% and 81.3±9.5% in vehicle and resveratrol treated cells, respectively). An inhibition of nitric oxide synthesis did not prevent the improved cell viability following oxygen glucose deprivation but SIRT-1 inhibition with sirtinol partially blocked the protection (P<0.001) suggesting endothelial protection is to some extent SIRT-1 dependent. Collectively, the results support that oral resveratrol treatment provides a low risk

  19. Effect of selected flavones on cancer and endothelial cells.

    PubMed

    Pilátová, Martina; Stupáková, Viktória; Varinská, Lenka; Sarisský, Marek; Mirossay, Ladislav; Mirossay, Andrej; Gál, Peter; Kraus, Vladimír; Dianisková, Katarína; Mojzis, Ján

    2010-06-01

    In our study we used quercetin (3,3 ,4 ,5,7-pentahydroxyflavone) as the reference standard to compare antiproliferative and antiangiogenic effects of chrysin (5,7-dihydroxyflavone) and 3-hydroxyflavone. Our data indicates that chrysin and 3-hydroxyflavone showed significantly higher cytotoxic effect than reference standard quercetin. These tested agents significantly decreased cell survival with the efficacy of 65-85% at the concentration 100 micromol/l for HUVEC, lung carcinoma and leukemic cells being the most sensitive. Cell cycle analysis indicates that quercetin and 3-hydroxyflavone might affect the cell cycle of Jurkat cells by a similar or the same mechanism of action which lead to G2/M arrest as well as to an increase in sub-G0/G1 fraction. Treatment of Jurkat cells with chrysin resulted only increase in the fraction of cells with sub-G0/G1 DNA content, which is considered to be a marker of apoptotic cell death. Apoptosis was confirmed by DNA fragmentation and by staining with annexin V. All three tested flavones inhibited endothelial cell migration after 24 h of incubation at a concentration 100 micromol/l. At a lower concentration (10 micromol/l) only quercetin significantly inhibited migration of endothelial cells. Furthermore, in our experiments decreased secretion of matrix metalloproteinases (MMP-2 and MMP-9) was observed after a 72 h treatment with quercetin. No decrease in secretion of MMP-2 and MMP-9 was seen after chrysin and 3-hydroxyflavone treatment. On the other hand, our results showed that none of three flavonoids blocked microcapillary tube formation. Further studies are necessary to investigate the mechanism of action and to find out the relationship between the structure, character and position of substituents of natural substances and their biological activities. PMID:20577025

  20. PECAM-1: regulator of endothelial junctional integrity.

    PubMed

    Privratsky, Jamie R; Newman, Peter J

    2014-03-01

    PECAM-1 (also known as CD31) is a cellular adhesion and signaling receptor comprising six extracellular immunoglobulin (Ig)-like homology domains, a short transmembrane domain and a 118 amino acid cytoplasmic domain that becomes serine and tyrosine phosphorylated upon cellular activation. PECAM-1 expression is restricted to blood and vascular cells. In circulating platelets and leukocytes, PECAM-1 functions largely as an inhibitory receptor that, via regulated sequential phosphorylation of its cytoplasmic domain, limits cellular activation responses. PECAM-1 is also highly expressed at endothelial cell intercellular junctions, where it functions as a mechanosensor, as a regulator of leukocyte trafficking and in the maintenance of endothelial cell junctional integrity. In this review, we will describe (1) the functional domains of PECAM-1 and how they contribute to its barrier-enhancing properties, (2) how the physical properties of PECAM-1 influence its subcellular localization and its ability to influence endothelial cell barrier function, (3) various stimuli that initiate PECAM-1 signaling and/or function at the endothelial junction and (4) cross-talk of PECAM-1 with other junctional molecules, which can influence endothelial cell function. PMID:24435645

  1. Molecular Bases of Corneal Endothelial Dystrophies

    PubMed Central

    Schmedt, Thore; Silva, Mariana Mazzini; Ziaei, Alireza; Jurkunas, Ula

    2011-01-01

    The phrase “corneal endothelial dystrophies” embraces a group of bilateral corneal conditions that are characterized by a non-inflammatory and progressive degradation of corneal endothelium. Corneal endothelial cells exhibit a high pump site density and, along with barrier function, are responsible for maintaining the cornea in its natural state of relative dehydration. Gradual loss of endothelial cells leads to an insufficient water outflow, resulting in corneal edema and loss of vision. Since the pathologic mechanisms remain largely unknown, the only current treatment option is surgical transplantation when vision is severely impaired. In the past decade, important steps have been taken to understand how endothelial degeneration progresses on the molecular level. Studies of affected multigenerational families and sporadic cases identified genes and chromosomal loci, and revealed either Mendelian or complex disorder inheritance patterns. Mutations have been detected in genes that carry important structural, metabolic, cytoprotective, and regulatory functions in corneal endothelium. In addition to genetic predisposition, environmental factors like oxidative stress were found to be involved in the pathogenesis of endotheliopathies. This review summarizes and crosslinks the recent progress on deciphering the molecular bases of corneal endothelial dystrophies. PMID:21855542

  2. Sphingosine-1-Phosphate Signaling in Endothelial Disorders.

    PubMed

    Sanchez, Teresa

    2016-06-01

    Numerous preclinical studies indicate that sustained endothelial activation significantly contributes to tissue edema, perpetuates the inflammatory response, and exacerbates tissue injury ultimately resulting in organ failure. However, no specific therapies aimed at restoring endothelial function are available as yet. Sphingosine-1-phosphate (S1P) is emerging as a potent modulator of endothelial function and endothelial responses to injury. Recent studies indicate that S1PR are attractive targets to treat not only disorders of the arterial endothelium but also microvascular dysfunction caused by ischemic or inflammatory injury. In this article, we will review the current knowledge of the role of S1P and its receptors in endothelial function in health and disease, and we will discuss the therapeutic potential of targeting S1PR not only for disorders of the arterial endothelium but also the microvasculature. The therapeutic targeting of S1PR in the endothelium could help to bridge the gap between biomedical research in vascular biology and clinical practice. PMID:27115142

  3. Endothelial Cell Response to Fusobacterium nucleatum.

    PubMed

    Mendes, Reila Tainá; Nguyen, Daniel; Stephens, Danielle; Pamuk, Ferda; Fernandes, Daniel; Van Dyke, Thomas E; Kantarci, Alpdogan

    2016-07-01

    Vascular response is an essential aspect of an effective immune response to periodontal disease pathogens, as new blood vessel formation contributes to wound healing and inflammation. Gaining a greater understanding of the factors that affect vascular response may then contribute to future breakthroughs in dental medicine. In this study, we have characterized the endothelial cell response to the common bacterium Fusobacterium nucleatum, an important bridging species that facilitates the activity of late colonizers of the dental biofilm. Endothelial cells were infected with Fusobacterium nucleatum (strain 25586) for periods of 4, 12, 24, or 48 h. Cell proliferation and tube formation were analyzed, and expression of adhesion molecules (CD31 and CD34) and vascular endothelial growth factor (VEGF) receptors 1 and 2 was measured by fluorescence-activated cell sorter (FACS) analysis. Data indicate that F. nucleatum impaired endothelial cell proliferation and tube formation. The findings suggest that the modified endothelial cell response acts as a mechanism promoting the pathogenic progression of periodontal diseases and may potentially suggest the involvement of periodontopathogens in systemic diseases associated with periodontal inflammation. PMID:27185790

  4. Immunostaining of macrophages, endothelial cells and smooth muscle cells in the atherosclerotic mouse aorta

    PubMed Central

    Menon, Prashanthi; Fisher, Edward A.

    2016-01-01

    The atherosclerotic mouse aorta consists of a heterogeneous population of cells, including macrophages, endothelial cells (EC) and smooth muscle cells (SMC), that play critical roles in cardiovascular disease. Identification of these vascular cells in the vessel wall is important to understanding their function in pathological conditions. Immunohistochemistry is an invaluable technique used to detect the presence of cells in different tissues. Here, we describe immunohistochemical techniques commonly used for the detection of the vascular cells in the atherosclerotic mouse aorta using cell specific markers. PMID:26445786

  5. Increase of serum vascular endothelial growth factors in wet beriberi: two case reports.

    PubMed

    Imai, Noboru; Kubota, Mai; Saitou, Mayu; Yagi, Nobuyasu; Serizawa, Masahiro; Kobari, Masahiro

    2012-01-01

    Beriberi is a disease caused by thiamine deficiency resulting in peripheral neuropathy and myocardial dysfunction. Increases in vascular endothelial growth factor (VEGF) are seen in polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and skin changes, called POEMS syndrome. We present herein two cases of wet beriberi accompanied by a moderate increase in VEGF level. Serum VEGF decreased after treatment in both cases. Our experience with these cases suggests that beriberi should be considered in the differential diagnosis of polyneuropathy with a moderate increase in serum VEGF, and that the serum VEGF level may be a therapeutic marker for beriberi. PMID:22504253

  6. Development of Endothelial-Specific Single Inducible Lentiviral Vectors for Genetic Engineering of Endothelial Progenitor Cells

    PubMed Central

    Yang, Guanghua; Kramer, M. Gabriela; Fernandez-Ruiz, Veronica; Kawa, Milosz P.; Huang, Xin; Liu, Zhongmin; Prieto, Jesus; Qian, Cheng

    2015-01-01

    Endothelial progenitor cells (EPC) are able to migrate to tumor vasculature. These cells, if genetically modified, can be used as vehicles to deliver toxic material to, or express anticancer proteins in tumor. To test this hypothesis, we developed several single, endothelial-specific, and doxycycline-inducible self-inactivating (SIN) lentiviral vectors. Two distinct expression cassettes were inserted into a SIN-vector: one controlled by an endothelial lineage-specific, murine vascular endothelial cadherin (mVEcad) promoter for the expression of a transactivator, rtTA2S-M2; and the other driven by an inducible promoter, TREalb, for a firefly luciferase reporter gene. We compared the expression levels of luciferase in different vector constructs, containing either the same or opposite orientation with respect to the vector sequence. The results showed that the vector with these two expression cassettes placed in opposite directions was optimal, characterized by a robust induction of the transgene expression (17.7- to 73-fold) in the presence of doxycycline in several endothelial cell lines, but without leakiness when uninduced. In conclusion, an endothelial lineage-specific single inducible SIN lentiviral vector has been developed. Such a lentiviral vector can be used to endow endothelial progenitor cells with anti-tumor properties. PMID:26612671

  7. Endothelial α3β1-Integrin Represses Pathological Angiogenesis and Sustains Endothelial-VEGF

    PubMed Central

    da Silva, Rita Graça; Tavora, Bernardo; Robinson, Stephen D.; Reynolds, Louise E.; Szekeres, Charles; Lamar, John; Batista, Sílvia; Kostourou, Vassiliki; Germain, Mitchel A.; Reynolds, Andrew R.; Jones, Dylan T.; Watson, Alan R.; Jones, Janet L.; Harris, Adrian; Hart, Ian R.; Iruela-Arispe, M. Luisa; DiPersio, C. Michael; Kreidberg, Jordan A.; Hodivala-Dilke, Kairbaan M.

    2010-01-01

    Integrin α3β1 is a major receptor for laminin. The expression levels of laminins-8 and -10 in the basement membrane surrounding blood vessels are known to change during tumor angiogenesis. Although some studies have suggested that certain ligands of α3β1 can affect angiogenesis either positively or negatively, either a direct in vivo role for α3β1 in this process or its mechanism of action in endothelial cells during angiogenesis is still unknown. Because the global genetic ablation of α3-integrin results in an early lethal phenotype, we have generated conditional-knockout mice where α3 is deleted specifically in endothelial cells (ec-α3−/−). Here we show that ec-α3−/− mice are viable, fertile, and display enhanced tumor growth, elevated tumor angiogenesis, augmented hypoxia-induced retinal angiogenesis, and increased vascular endothelial growth factor (VEGF)-mediated neovascularization ex vivo and in vivo. Furthermore, our data provide a novel method by which an integrin may regulate angiogenesis. We show that α3β1 is a positive regulator of endothelial-VEGF and that, surprisingly, the VEGF produced by endothelial cells can actually repress VEGF-receptor 2 (Flk-1) expression. These data, therefore, identify directly that endothelial α3β1 negatively regulates pathological angiogenesis and implicate an unexpected role for low levels of endothelial-VEGF as an activator of neovascularization. PMID:20639457

  8. KRIT1 Protein Depletion Modifies Endothelial Cell Behavior via Increased Vascular Endothelial Growth Factor (VEGF) Signaling*

    PubMed Central

    DiStefano, Peter V.; Kuebel, Julia M.; Sarelius, Ingrid H.; Glading, Angela J.

    2014-01-01

    Disruption of endothelial cell-cell contact is a key event in many cardiovascular diseases and a characteristic of pathologically activated vascular endothelium. The CCM (cerebral cavernous malformation) family of proteins (KRIT1 (Krev-interaction trapped 1), PDCD10, and CCM2) are critical regulators of endothelial cell-cell contact and vascular homeostasis. Here we show novel regulation of vascular endothelial growth factor (VEGF) signaling in KRIT1-depleted endothelial cells. Loss of KRIT1 and PDCD10, but not CCM2, increases nuclear β-catenin signaling and up-regulates VEGF-A protein expression. In KRIT1-depleted cells, increased VEGF-A levels led to increased VEGF receptor 2 (VEGFR2) activation and subsequent alteration of cytoskeletal organization, migration, and barrier function and to in vivo endothelial permeability in KRIT1-deficient animals. VEGFR2 activation also increases β-catenin phosphorylation but is only partially responsible for KRIT1 depletion-dependent disruption of cell-cell contacts. Thus, VEGF signaling contributes to modifying endothelial function in KRIT1-deficient cells and microvessel permeability in Krit1+/− mice; however, VEGF signaling is likely not the only contributor to disrupted endothelial cell-cell contacts in the absence of KRIT1. PMID:25320085

  9. Development of Endothelial-Specific Single Inducible Lentiviral Vectors for Genetic Engineering of Endothelial Progenitor Cells.

    PubMed

    Yang, Guanghua; Kramer, M Gabriela; Fernandez-Ruiz, Veronica; Kawa, Milosz P; Huang, Xin; Liu, Zhongmin; Prieto, Jesus; Qian, Cheng

    2015-01-01

    Endothelial progenitor cells (EPC) are able to migrate to tumor vasculature. These cells, if genetically modified, can be used as vehicles to deliver toxic material to, or express anticancer proteins in tumor. To test this hypothesis, we developed several single, endothelial-specific, and doxycycline-inducible self-inactivating (SIN) lentiviral vectors. Two distinct expression cassettes were inserted into a SIN-vector: one controlled by an endothelial lineage-specific, murine vascular endothelial cadherin (mVEcad) promoter for the expression of a transactivator, rtTA2S-M2; and the other driven by an inducible promoter, TREalb, for a firefly luciferase reporter gene. We compared the expression levels of luciferase in different vector constructs, containing either the same or opposite orientation with respect to the vector sequence. The results showed that the vector with these two expression cassettes placed in opposite directions was optimal, characterized by a robust induction of the transgene expression (17.7- to 73-fold) in the presence of doxycycline in several endothelial cell lines, but without leakiness when uninduced. In conclusion, an endothelial lineage-specific single inducible SIN lentiviral vector has been developed. Such a lentiviral vector can be used to endow endothelial progenitor cells with anti-tumor properties. PMID:26612671

  10. Digital imaging of diabetic endothelial cells

    NASA Astrophysics Data System (ADS)

    Paltauf-Doburzynska, Jolanta; Kohlwein, Sepp D.; Graier, Wolfgang F.

    2001-05-01

    Endothelial cells release factors that regulate dilatation and contraction of the vessels. They play an important role in modulating both the inflammatory response and vasomotor abnormalities that occur in coronary artery diseases. This endothelial function is associated with changes of intracellular Ca2+ concentration. For this study we used spatially and temporally resolved measurements of local Ca2+ concentration in human endothelial cells cultured in high glucose containing medium. Deconvolution techniques procedure allowed determination of intracellular Ca2+ concentration and its distribution into cellular compartments. We also used a confocal microscope for visualization of intracellular compartments (endoplasmatic reticulum, mitochondria) under normal and pathological conditions. We showed that the interrupted connection between superficial compartments and membrane channels is already the beginning of the cell damage in diabetes.

  11. The phacoemulsification procedure. II. Corneal endothelial changes.

    PubMed

    Polack, F M; Sugar, A

    1976-06-01

    The effect of phacoemulsification, with the Cavitron-Kelman instrument, on the corneal endothelium of rabbit and cats was studied by scanning electron microscopy and nitroblue tetrazolium staining. The various steps of the procedure were examined separately. Irrigation of the anterior chamber of the eye with balanced salt solution (Plasma-Lyte) for ten minutes caused no cell damage. Ultrasound and irrigation alone for four to six minutes caused increased permeability to NBT. Edema of endothelial cells and cell junction disruption occurred after eight minutes of anterior chamber irrigation with Plasma-Lyte. Uncomplicated phacoemulsification produced moderate cellular edema with scattered loss of endothelial cells. Destruction of endothelial cells was frequent after phacoemulsification, it appeared to be due to lens nucleus manipulation in the anterior chamber, instrumentation, and needle contact. From two to five days postoperatively, intercellular edema, altered cell morphology, and mosaic pattern were seen. However, it gradually recovered and seven to ten days later the endothelium appeared normal. PMID:931690

  12. Corneal endothelial changes in superficial epithelial keratopathy.

    PubMed

    Brooks, A M; Grant, G; Gillies, W E

    1986-05-01

    A series of five cases is described in which superficial punctate keratopathy was associated with endothelial cell changes. The most striking change was the presence of dark areas or blebs, usually two to four cell diameters in extent. There was also distortion and crumpling of the corneal endothelium, mild pleomorphism and polymegathism of the endothelial cells, with a reduced cell count in some cases. These blebs have previously been reported in hard and soft contact lens wearers and are due to intercellular oedema with separation of endothelial cells from Descemet's membrane. Anoxia and interference with osmosis have been proposed as possible mechanisms for the production of these blebs, but our cases demonstrate that fine disruption of the corneal epithelium can affect the integrity of the corneal endothelium and may lead to significant damage over a long period of time. PMID:3801208

  13. Isolation and culture of pulmonary endothelial cells.

    PubMed

    Ryan, U S

    1984-06-01

    Methods for isolation, identification and culture of pulmonary endothelial cells are now routine. In the past, methods of isolation have used proteolytic enzymes to detach cells; thereafter, traditional methods for cell passaging have used trypsin/EDTA mixtures. Cells isolated and passaged using proteolytic enzymes have been useful in establishing the field and in verifying certain endothelial properties. However, there is a growing awareness of the role of endothelial cells in processing vasoactive substances, in responding to hormones and other agonists and in cell-cell interactions with other cell types of the vascular wall, with blood cells and with cellular products. Consequently, a new requirement has arisen for cells in vitro that maintain the differentiated properties of their counterparts in vivo. The deleterious effects of trypsin and other proteolytic enzymes commonly used in cell culture on surface structures of endothelial cells such as enzymes, receptors and junctional proteins, as well as on extracellular layers such as the glycocalyx or "endothelial fuzz," have led to the development of methods that avoid use of proteolytic enzymes at both the isolation step and during subsequent subculture. This chapter describes traditional methods for isolating pulmonary endothelial cells but emphasizes newer approaches using mechanical harvest and scale-up using microcarriers. The new methods allow maintenance of long-term, large-scale cultures of cells that retain the full complement of surface properties and that maintain the cobblestone monolayer morphology and differentiated functional properties. Methods for identification of isolated cells are therefore also considered as methods for validation of cultures during their in vitro lifespan. PMID:6090112

  14. In Brief: Endothelial-to-mesenchymal transition.

    PubMed

    Sanchez-Duffhues, Gonzalo; Orlova, Valeria; Ten Dijke, Peter

    2016-02-01

    Recent evidence has highlighted the role of endothelial-to-mesenchymal transition (EndoMT) in the onset and progression of a number of human pathologies. EndoMT involves the loss of an endothelial signature to generate unspecialized mesenchymal-like cells, susceptible to being re-differentiated into mesodermal cell types, including osteoblasts, chondrocytes, and adipocytes. Therefore, modulation of the molecular pathways controlling EndoMT can be considered as a therapeutic approach for particular human diseases. In addition, EndoMT may be harnessed for tissue engineering by producing multipotent mesenchymal cells able to differentiate into mutiple cell types. PMID:26446982

  15. Crossing the endothelial barrier during metastasis.

    PubMed

    Reymond, Nicolas; d'Água, Bárbara Borda; Ridley, Anne J

    2013-12-01

    During metastasis, cancer cells disseminate to other parts of the body by entering the bloodstream in a process that is called intravasation. They then extravasate at metastatic sites by attaching to endothelial cells that line blood vessels and crossing the vessel walls of tissues or organs. This Review describes how cancer cells cross the endothelial barrier during extravasation and how different receptors, signalling pathways and circulating cells such as leukocytes and platelets contribute to this process. Identification of the mechanisms that underlie cancer cell extravasation could lead to the development of new therapies to reduce metastasis. PMID:24263189

  16. TRAIL-Death Receptor 4 Signaling via Lysosome Fusion and Membrane Raft Clustering In Coronary Arterial Endothelial Cells: Evidence from ASM Knockout Mice

    PubMed Central

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

    2012-01-01

    Tumor necrosis factor (TNF)-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 and 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 (MRs) clustering and 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 its co-localization with DR4 in coronary arterial endothelial cells (CAECs) isolated from wild-type (Smpd1+/+) mice. Further, 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 O2−· production in CAECs and abolished TRAIL-induced impairment on endothelium-dependent vasodilation in small resistance arteries. By measuring fluorescence resonance energy transfer (FRET), we found that Lamp-1 (lysosome membrane marker protein) and ganglioside GM1 (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 and fusion with membrane and formation of MR redox signaling platforms, which may

  17. Characterization of Umbilical Cord Blood–Derived Late Outgrowth Endothelial Progenitor Cells Exposed to Laminar Shear Stress

    PubMed Central

    Brown, Melissa A.; Wallace, Charles S.; Angelos, Mathew

    2009-01-01

    Endothelial progenitor cells isolated from umbilical cord blood (CB-EPCs) represent a promising source of endothelial cells for synthetic vascular grafts and tissue-engineered blood vessels since they are readily attainable, can be easily isolated, and possess a high proliferation potential. The objective of this study was to compare the functional behavior of late outgrowth CB-EPCs with human aortic endothelial cells (HAECs). CB-EPCs and HAECs were cultured on either smooth muscle cells in a coculture model of a tissue-engineered blood vessels or fibronectin adsorbed to Teflon-AF™–coated glass slides. Late outgrowth CB-EPCs expressed endothelial cell–specific markers and were negative for the monocytic marker CD14. CB-EPCs have higher proliferation rates than HAECs, but are slightly smaller in size. CB-EPCs remained adherent under supraphysiological shear stresses, oriented and elongated in the direction of flow, and expressed similar numbers of α5β1 and αvβ3 integrins and antithrombotic genes compared to HAECs. There were some differences in mRNA levels of E-selectin and vascular cell adhesion molecule 1 between CB-EPCs and HAECs; however, protein levels were similar on the two cell types, and CB-EPCs did not support adhesion of monocytes in the absence of tumor necrosis factor-α stimulation. Although CB-EPCs expressed significantly less endothelial nitric oxide synthase protein after exposure to flow than HAECs, nitric oxide levels induced by flow were not significantly different. These results suggest that late outgrowth CB-EPCs are functionally similar to HAECs under flow conditions and are a promising cell source for cardiovascular therapies. PMID:19480571

  18. Sevelamer revisited: pleiotropic effects on endothelial and cardiovascular risk factors in chronic kidney disease and end-stage renal disease

    PubMed Central

    2013-01-01

    Endothelial dysfunction underlies multiple cardiovascular consequences of chronic kidney disease (CKD) and antecedent diabetes or hypertension. Endothelial insults in CKD or end-stage renal disease (ESRD) patients include uremic toxins, serum uric acid, hyperphosphatemia, reactive oxygen species, and advanced glycation endproducts (AGEs). Sevelamer carbonate, a calcium-free intestinally nonabsorbed polymer, is approved for hyperphosphatemic dialysis patients in the US and hyperphosphatemic stage 3–5 CKD patients in many other countries. Sevelamer has been observed investigationally to reduce absorption of AGEs, bacterial toxins, and bile acids, suggesting that it may reduce inflammatory, oxidative, and atherogenic stimuli in addition to its on-label action of lowering serum phosphate. Some studies also suggest that noncalcium binders may contribute less to vascular calcification than calcium-based binders. Exploratory sevelamer carbonate use in patients with stages 2–4 diabetic CKD significantly reduced HbA1c, AGEs, fibroblast growth factor (FGF)-23, and total and low-density lipoprotein (LDL) cholesterol versus calcium carbonate; inflammatory markers decreased and defenses against AGEs increased. Sevelamer has also been observed to reduce circulating FGF-23, potentially reducing risk of left ventricular hypertrophy. Sevelamer but not calcium-based binders in exploratory studies increases flow-mediated vasodilation, a marker of improved endothelial function, in patients with CKD. In contrast, lanthanum carbonate and calcium carbonate effects on FMV did not differ in hemodialysis recipients. The recent INDEPENDENT-CKD randomized trial compared sevelamer versus calcium carbonate in predialysis CKD patients (investigational in the US, on-label in European participants); sevelamer reduced 36-month mortality and the composite endpoint of mortality or dialysis inception. Similarly, INDEPENDENT-HD in incident dialysis patients showed improved survival with 24 months

  19. 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. PMID:26934950

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

    PubMed Central

    IGARASHI, YASUYUKI; CHOSA, NAOYUKI; SAWADA, SHUNSUKE; KONDO, HISATOMO; YAEGASHI, TAKASHI; ISHISAKI, AKIRA

    2016-01-01

    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. PMID:26934950

  1. Shear Stress Induces Differentiation of Endothelial Lineage Cells to Protect Neonatal Brain from Hypoxic-Ischemic Injury through NRP1 and VEGFR2 Signaling.

    PubMed

    Huang, Chia-Wei; Huang, Chao-Ching; Chen, Yuh-Ling; Fan, Shih-Chen; Hsueh, Yuan-Yu; Ho, Chien-Jung; Wu, Chia-Ching

    2015-01-01

    Neonatal hypoxic-ischemic (HI) brain injuries disrupt the integrity of neurovascular structure and lead to lifelong neurological deficit. The devastating damage can be ameliorated by preserving the endothelial network, but the source for therapeutic cells is limited. We aim to evaluate the beneficial effect of mechanical shear stress in the differentiation of endothelial lineage cells (ELCs) from adipose-derived stem cells (ASCs) and the possible intracellular signals to protect HI injury using cell-based therapy in the neonatal rats. The ASCs expressed early endothelial markers after biochemical stimulation of endothelial growth medium. The ELCs with full endothelial characteristics were accomplished after a subsequential shear stress application for 24 hours. When comparing the therapeutic potential of ASCs and ELCs, the ELCs treatment significantly reduced the infarction area and preserved neurovascular architecture in HI injured brain. The transplanted ELCs can migrate and engraft into the brain tissue, especially in vessels, where they promoted the angiogenesis. The activation of Akt by neuropilin 1 (NRP1) and vascular endothelial growth factor receptor 2 (VEGFR2) was important for ELC migration and following in vivo therapeutic outcomes. Therefore, the current study demonstrated importance of mechanical factor in stem cell differentiation and showed promising protection of brain from HI injury using ELCs treatment. PMID:26509169

  2. Shear Stress Induces Differentiation of Endothelial Lineage Cells to Protect Neonatal Brain from Hypoxic-Ischemic Injury through NRP1 and VEGFR2 Signaling

    PubMed Central

    Huang, Chia-Wei; Huang, Chao-Ching; Chen, Yuh-Ling; Fan, Shih-Chen; Hsueh, Yuan-Yu; Ho, Chien-Jung; Wu, Chia-Ching

    2015-01-01

    Neonatal hypoxic-ischemic (HI) brain injuries disrupt the integrity of neurovascular structure and lead to lifelong neurological deficit. The devastating damage can be ameliorated by preserving the endothelial network, but the source for therapeutic cells is limited. We aim to evaluate the beneficial effect of mechanical shear stress in the differentiation of endothelial lineage cells (ELCs) from adipose-derived stem cells (ASCs) and the possible intracellular signals to protect HI injury using cell-based therapy in the neonatal rats. The ASCs expressed early endothelial markers after biochemical stimulation of endothelial growth medium. The ELCs with full endothelial characteristics were accomplished after a subsequential shear stress application for 24 hours. When comparing the therapeutic potential of ASCs and ELCs, the ELCs treatment significantly reduced the infarction area and preserved neurovascular architecture in HI injured brain. The transplanted ELCs can migrate and engraft into the brain tissue, especially in vessels, where they promoted the angiogenesis. The activation of Akt by neuropilin 1 (NRP1) and vascular endothelial growth factor receptor 2 (VEGFR2) was important for ELC migration and following in vivo therapeutic outcomes. Therefore, the current study demonstrated importance of mechanical factor in stem cell differentiation and showed promising protection of brain from HI injury using ELCs treatment. PMID:26509169

  3. Rapamycin inhibits re-endothelialization after percutaneous coronary intervention by impeding the proliferation and migration of endothelial cells and inducing apoptosis of endothelial progenitor cells.

    PubMed

    Liu, Hai-Tao; Li, Fei; Wang, Wen-Yong; Li, Xiao-Jing; Liu, Yi-Meng; Wang, Rui-An; Guo, Wen-Yi; Wang, Hai-Chang

    2010-01-01

    Endothelial-cell function is important in the healing of damaged endothelium after percutaneous coronary artery damage. In 3 different animal models, we sought to determine whether rapamycin (sirolimus) affects the proliferation and migration of endothelial cells and endothelial progenitor cells. First, after we implanted stents in dogs, we found that re-endothelialization was impeded more by drug-eluting stents than by bare-metal stents, 30 days after percutaneous coronary intervention. Second, in vitro in rats, we found that 1-100 ng/mL of rapamycin time- and dose-dependently inhibited proliferation over 72 hr (with effects evident as early as 24 hr) and also dose-dependently induced endothelial progenitor-cell apoptosis. Finally, in vivo in rats, we observed that vascular endothelial growth factor expression was decreased after 5 days of rapamycin treatment. We conclude that rapamycin impedes re-endothelialization after drug-eluting stent implantation by inhibiting the proliferation and migration of coronary endothelial cells, inducing endothelial progenitor-cell apoptosis, and decreasing vascular endothelial growth factor expression in the circulation. PMID:20401293

  4. Induction of Hepatic and Endothelial Differentiation by Perfusion in a Three-Dimensional Cell Culture Model of Human Fetal Liver

    PubMed Central

    Pekor, Christopher; Gerlach, Jörg C.; Nettleship, Ian

    2015-01-01

    The development of functional engineered tissue constructs depends on high cell densities and appropriate vascularization. In this study we implemented a four-compartment three-dimensional perfusion bioreactor culture model for studying the effects of medium perfusion on endothelial, hepatic, and hematopoietic cell populations of primary human fetal liver in an in vivo-like environment. Human fetal liver cells were cultured in bioreactors configured to provide either perfusion or diffusion conditions. Metabolic activities of the cultures were monitored daily by measuring glucose consumption and lactate production. Cell viability during culture was analyzed by lactate dehydrogenase activity. Hepatic functionality was determined by the release of albumin and alpha-fetoprotein (AFP) in culture medium samples. After 4 days of culture, cells were analyzed for the expression of a variety of endothelial, hepatic, and hematopoietic genes, as well as the surface marker expression of CD31 and CD34 in flow cytometry. We found that medium perfusion increased the gene expression of endothelial markers such as CD31, von Willebrand factor (vWF), CD140b, CD309, and CD144 while decreasing the gene expression of the erythrocyte-surface marker CD235a. Hepatic differentiation was promoted under perfusion conditions as demonstrated by lower AFP and higher albumin secretion compared with cultures not exposed to medium perfusion. Additionally, cultures exposed to medium perfusion gave higher rates of glucose consumption and lactate production, indicating increased metabolic activity. In conclusion, high-density bioreactors configured to provide constant medium perfusion significantly induced hepatic and endothelial cell differentiation and provided improved conditions for the culture of human fetal liver cells compared with cultures without perfusion. PMID:25559936

  5. Induction of Hepatic and Endothelial Differentiation by Perfusion in a Three-Dimensional Cell Culture Model of Human Fetal Liver.

    PubMed

    Pekor, Christopher; Gerlach, Jörg C; Nettleship, Ian; Schmelzer, Eva

    2015-07-01

    The development of functional engineered tissue constructs depends on high cell densities and appropriate vascularization. In this study we implemented a four-compartment three-dimensional perfusion bioreactor culture model for studying the effects of medium perfusion on endothelial, hepatic, and hematopoietic cell populations of primary human fetal liver in an in vivo-like environment. Human fetal liver cells were cultured in bioreactors configured to provide either perfusion or diffusion conditions. Metabolic activities of the cultures were monitored daily by measuring glucose consumption and lactate production. Cell viability during culture was analyzed by lactate dehydrogenase activity. Hepatic functionality was determined by the release of albumin and alpha-fetoprotein (AFP) in culture medium samples. After 4 days of culture, cells were analyzed for the expression of a variety of endothelial, hepatic, and hematopoietic genes, as well as the surface marker expression of CD31 and CD34 in flow cytometry. We found that medium perfusion increased the gene expression of endothelial markers such as CD31, von Willebrand factor (vWF), CD140b, CD309, and CD144 while decreasing the gene expression of the erythrocyte-surface marker CD235a. Hepatic differentiation was promoted under perfusion conditions as demonstrated by lower AFP and higher albumin secretion compared with cultures not exposed to medium perfusion. Additionally, cultures exposed to medium perfusion gave higher rates of glucose consumption and lactate production, indicating increased metabolic activity. In conclusion, high-density bioreactors configured to provide constant medium perfusion significantly induced hepatic and endothelial cell differentiation and provided improved conditions for the culture of human fetal liver cells compared with cultures without perfusion. PMID:25559936

  6. Platelet endothelial cell adhesion molecule-1 and mechanotransduction in vascular endothelial cells.

    PubMed

    Fujiwara, K

    2006-04-01

    Endothelial cells are known to respond to mechanical forces such as fluid shear stress and cyclic stretch, but elucidating the mechanism for mechanosensing has been difficult. Experimental data indicate that there are probably several sensing mechanisms. We have recently proposed a novel mechanoresponse mechanism that involves platelet endothelial cell adhesion molecule-1 (PECAM-1). When endothelial cells are stimulated by fluid shear stress, PECAM-1 is tyrosine phosphorylated and activates the extracellular signal-regulated kinase 1 and 2 (ERK1/2) signalling cascade. The same signalling events occurred when we applied pulling force directly on PECAM-1 on the endothelial cell surface using magnetic beads coated with antibodies against the external domain of PECAM-1. These results appear to indicate that PECAM-1 is a mechanotransduction molecule. To our knowledge, this is the first mammalian molecule that is shown to respond to mechanical force directly exerted to it. PMID:16594905

  7. 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

  8. Protective effects of Astragalus polysaccharides against endothelial dysfunction in hypertrophic rats induced by isoproterenol.

    PubMed

    Han, Ronghui; Tang, Futian; Lu, Meili; Xu, Chonghua; Hu, Jin; Mei, Meng; Wang, Hongxin

    2016-09-01

    Astragalus polysaccharide (APS) is an important bioactive component extracted from Chinese herb Astragalus membranaceus. It has been widely used in treatment of cardiovascular diseases. We have previously reported that APS could inhibit isoproterenol-induced cardiac hypertrophy. The present study was designed to evaluate the protective effect of APS on vascular endothelia in cardiac hypertrophy rats induced by isoproterenol (ISO). ISO (10mg×kg(-1)) was intraperitoneally injected once daily for 2weeks to induce cardiac hypertrophy. APS (400 and 800mg×kg(-1)) was intragastrically injected once daily along with ISO. The results showed that combination with APS significantly ameliorates the endothelial dysfunction while attenuates cardiac hypertrophy induced by ISO. We found that administration with APS could attenuate the increase in number of circulating endothelial cell (CEC). APS also decreases the superoxide anion generation and the protein expression of p65 and the levels of TNF-α and IL-6; while increases the cGMP levels, an activity marker for nitric oxide (NO) in aortas. In addition, APS improves the relaxation dysfunction in isolated aortic rings and increases the protein expression of IκBα and Cu/Zn-SOD in aortas. In conclusion, our results suggested that APS had a protective effect against endothelial dysfunction in hypertrophic rats induced by ISO. The underlining mechanisms may be contributed to the anti-inflammatory effects and the improvement of the imbalance between reactive oxygen species (ROS) and NO. PMID:27344039

  9. Endothelial to mesenchymal transition is common in atherosclerotic lesions and is associated with plaque instability.

    PubMed

    Evrard, Solene M; Lecce, Laura; Michelis, Katherine C; Nomura-Kitabayashi, Aya; Pandey, Gaurav; Purushothaman, K-Raman; d'Escamard, Valentina; Li, Jennifer R; Hadri, Lahouaria; Fujitani, Kenji; Moreno, Pedro R; Benard, Ludovic; Rimmele, Pauline; Cohain, Ariella; Mecham, Brigham; Randolph, Gwendalyn J; Nabel, Elizabeth G; Hajjar, Roger; Fuster, Valentin; Boehm, Manfred; Kovacic, Jason C

    2016-01-01

    Endothelial to mesenchymal transition (EndMT) plays a major role during development, and also contributes to several adult cardiovascular diseases. Importantly, mesenchymal cells including fibroblasts are prominent in atherosclerosis, with key functions including regulation of: inflammation, matrix and collagen production, and plaque structural integrity. However, little is known about the origins of atherosclerosis-associated fibroblasts. Here we show using endothelial-specific lineage-tracking that EndMT-derived fibroblast-like cells are common in atherosclerotic lesions, with EndMT-derived cells expressing a range of fibroblast-specific markers. In vitro modelling confirms that EndMT is driven by TGF-β signalling, oxidative stress and hypoxia; all hallmarks of atherosclerosis. 'Transitioning' cells are readily detected in human plaques co-expressing endothelial and fibroblast/mesenchymal proteins, indicative of EndMT. The extent of EndMT correlates with an unstable plaque phenotype, which appears driven by altered collagen-MMP production in EndMT-derived cells. We conclude that EndMT contributes to atherosclerotic patho-biology and is associated with complex plaques that may be related to clinical events. PMID:27340017

  10. Endothelial to mesenchymal transition is common in atherosclerotic lesions and is associated with plaque instability

    PubMed Central

    Evrard, Solene M.; Lecce, Laura; Michelis, Katherine C.; Nomura-Kitabayashi, Aya; Pandey, Gaurav; Purushothaman, K-Raman; d'Escamard, Valentina; Li, Jennifer R.; Hadri, Lahouaria; Fujitani, Kenji; Moreno, Pedro R.; Benard, Ludovic; Rimmele, Pauline; Cohain, Ariella; Mecham, Brigham; Randolph, Gwendalyn J.; Nabel, Elizabeth G.; Hajjar, Roger; Fuster, Valentin; Boehm, Manfred; Kovacic, Jason C.

    2016-01-01

    Endothelial to mesenchymal transition (EndMT) plays a major role during development, and also contributes to several adult cardiovascular diseases. Importantly, mesenchymal cells including fibroblasts are prominent in atherosclerosis, with key functions including regulation of: inflammation, matrix and collagen production, and plaque structural integrity. However, little is known about the origins of atherosclerosis-associated fibroblasts. Here we show using endothelial-specific lineage-tracking that EndMT-derived fibroblast-like cells are common in atherosclerotic lesions, with EndMT-derived cells expressing a range of fibroblast-specific markers. In vitro modelling confirms that EndMT is driven by TGF-β signalling, oxidative stress and hypoxia; all hallmarks of atherosclerosis. ‘Transitioning' cells are readily detected in human plaques co-expressing endothelial and fibroblast/mesenchymal proteins, indicative of EndMT. The extent of EndMT correlates with an unstable plaque phenotype, which appears driven by altered collagen-MMP production in EndMT-derived cells. We conclude that EndMT contributes to atherosclerotic patho-biology and is associated with complex plaques that may be related to clinical events. PMID:27340017

  11. The relationship between circulating TRAIL and endothelial dysfunction in subclinical hypothyroidism.

    PubMed

    Xiang, Guangda; Yue, Ling; Zhang, Junxia; Xiang, Lin; Dong, Jing

    2015-05-01

    Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is associated with atherosclerosis. Subclinical hypothyroidism (sHT) is associated with the increased prevalence of atherosclerotic lesions and cardiovascular events. Therefore, we hypothesized that circulating TRAIL levels are associated with endothelial dysfunction in sHT patients. Two hundred and four patients with newly diagnosed sHT and 52 healthy subjects were recruited. Circulating TRAIL concentration was measured by an ELISA, and flow-mediated dilation (FMD) of brachial artery was measured using high-resolution ultrasound. The mean value of circulating TRAIL in newly diagnosed sHT patients was 67.2 pg/ml, which was lower than that in controls (78.5 pg/ml, p < 0.001). By dividing the distribution of FMD levels into quartiles, TRAIL levels were increased gradually with the increase of FMD levels (p < 0.001). Multivariate regression analysis demonstrated that serum TRAIL levels were independently associated with FMD (p = 0.007). By logistic regression analysis, the odds ratio for lower FMD levels was reduced by 12.1 % per 1 pg/ml increase in serum TRAIL concentration after adjustment for multivariate metabolic factors [OR (95 % CI); 0.879 (0.721-0.973)]. Circulating TRAIL level decreased in newly diagnosed sHT patients and is positively associated with endothelial function, suggesting that circulating TRAIL level may be a protective marker of endothelial function in sHT patients. PMID:25294014

  12. Fever temperature enhances mechanisms of survival of Streptococcus agalactiae within human endothelial cells.

    PubMed

    Freitas Lione, Viviane Oliveira; Bittencourt Dos Santos, Michelle Hanthequeste; Ulisses Carvalho, Técia Maria; Hirata, Raphael; Mattos-Guaraldi, Ana Luiza; Arruda Mortara, Renato; Nagao, Prescilla Emy

    2010-10-01

    Group B streptococci (GBS) are the most common cause of pneumonia and sepsis during the neonatal period. However, the pathogenesis of invasive infection is poorly understood. We investigated the ability of GBS grown at 37 degrees C and 40 degrees C to adhere and invade human umbilical vein endothelial cells (HUVECs) at different periods of incubation (0, 0.5, 1, 2, 18 and 24 h). All strains tested, except strain 88641-vagina survived for 24 h in the intracellular environment at 40 degrees C. For serotype III grown at 40 degrees C, both strains (80340-vagina and 90356-liquor) showed increased adherence and intracellular survival when compared to bacteria grown at 37 degrees C (P<0.01). GBS serotype V strains (88641-vagina and 90186-blood) showed ability to survive inside HUVECs until 2 and 24 h post-infection at 40 degrees C and 37 degrees C, respectively (P<0.01). Influence of growth temperature in bacterial interaction with endothelial cells was partially dependent of serotypes and the clinical origin of strains. Serotypes III and V strains grown at both temperatures remained viable within acidic endothelial vacuoles which acquired Rab7 and LAMP-1 endosomal markers. The data emphasize the influence of temperature on cellular events of phagocytosis and pathogenesis of GBS diseases. PMID:20818490

  13. Dose-dependent surface endothelialization and biocompatibility of polyurethane noble metal nanocomposites.

    PubMed

    Hess, Christian; Schwenke, Andreas; Wagener, Philipp; Franzka, Steffen; Laszlo Sajti, Csaba; Pflaum, Michael; Wiegmann, Bettina; Haverich, Axel; Barcikowski, Stephan

    2014-06-01

    Surface pre-endothelialization is a promising approach to improve the hemocompatibility of implants, medical devices, and artificial organs. To promote the adhesive property of thermoplastic polyurethane (TPU) for endothelial cells (ECs), up to 1 wt % of gold (Au) or platinum (Pt) nanoparticles, fabricated by pulsed laser ablation in polymer solution, were embedded into the polymer matrix. The analysis of these nanocomposites showed a homogenous dispersion of the nanoparticles, with average diameters of 7 nm for Au or 9 nm for Pt. A dose-dependent effect was found when ECs were seeded onto nanocomposites comprising different nanoparticle concentrations, resulting in a fivefold improvement of proliferation at 0.1 wt % nanoparticle load. This effect was associated with a nanoparticle concentration-dependent hydrophilicity and negative charge of the nanocomposite. In dynamic flow tests, nanocomposites containing 0.1 wt % Au or Pt nanoparticles allowed for the generation of a confluent and resistant EC layer. Real-time polymerase chain reaction quantification of specific markers for EC activation indicated that ECs cultivated on nanocomposites remain in an inactivated, nonthrombogenic and noninflammatory state; however, maintain the ability to trigger an inflammatory response upon stimulation. These findings were confirmed by a platelet and leukocyte adhesion assay. The results of this study suggest the possible applicability of TPU nanocomposites, containing 0.1 wt % Au or Pt nanoparticles, for the generation of pre-endothelialized surfaces of medical devices. PMID:23852964

  14. Micropatterned 3-Dimensional Hydrogel System to Study Human Endothelial-Mesenchymal Stem Cell Interactions

    PubMed Central

    Trkov, Sasa; Eng, George; di Liddo, Rosa; Parnigotto, Pier Paolo; Vunjak-Novakovic, Gordana

    2009-01-01

    The creation of vascularized engineered tissues of clinically relevant size is a major challenge of tissue engineering. While it is known that endothelial and mural vascular cells are integral to the formation of stable blood vessels, the specific cell type and optimal conditions for engineered vascular networks are poorly understood. To this end, we investigated the vasculogenic potential of human mesenchymal stem cell (MSC) populations derived from three different sources: (i) bone marrow aspirates, (ii) perivascular cells from umbilical cord vein, and (iii) perivascular cells from umbilical cord artery. Cell populations were isolated and identified as MSCs according to their phenotypes and differentiation potential. Human umbilical vein endothelial cells (HUVEC) were used as a standard for endothelial cells. A novel co-culture system was developed to study cell-cell interactions in a spatially controlled three-dimensional (3D) fibrin hydrogel model. Using microfluidic patterning, it was possible to localize hydrogel-encapsulated HUVECs and MSCs within separate channels spaced at 500, 1000 or 2000 μm. All three MSC populations had similar expression profiles of mesenchymal cell markers, and similar capacity for osteogenic and adipogenic differentiation. However, bone marrow-derived MSCs (but not umbilical vein or artery derived MSCs) showed strong distance-dependent migration toward HUVECs and supported the formation of stable vascular networks resembling capillary-like vasculature. The presented approach provides a simple and robust model to study cell-cell communication of relevance to engineering vascularized tissues. PMID:19998330

  15. Human pulmonary artery endothelial cells in the model of mucopolysaccharidosis VI present a prohypertensive phenotype

    PubMed Central

    Golda, Adam; Jurecka, Agnieszka; Gajda, Karolina; Tylki-Szymańska, Anna; Lalik, Anna

    2015-01-01

    Background Mucopolysaccharidosis type VI (MPS VI) is an autosomal recessive lysosomal disorder caused by a deficient activity of N-acetylgalactosamine-4-sulfatase (ARSB). Pulmonary hypertension (PH) occurs in MPS VI patients and is a marker of bad prognosis. Malfunction of endothelium, which regulates vascular tonus and stimulates angiogenesis, can contribute to the occurrence of PH in MPS VI. Aim The aim of the study was to establish a human MPS VI cellular model of pulmonary artery endothelial cells (HPAECs) and evaluate how it affects factors that may trigger PH such as proliferation, apoptosis, expression of endothelial nitric oxide synthase (eNOS), natriuretic peptide type C (NPPC), and vascular endothelial growth factor A (VEGFA). Results Increasing concentrations of dermatan sulfate (DS) reduce the viability of the cells in both ARSB deficiency and controls, but hardly influence apoptosis. The expression of eNOS in HPAECs is reduced up to two thirds in the presence of DS. NPPC shows a biphasic expression reaction with an increase at 50 μg/mL DS and reduction at 0 and 100 μg/mL DS. The expression of VEGFA decreases with increasing DS concentrations and absence of elastin, and increases with increasing DS in the presence of elastin. Conclusion Our data suggest that MPS VI endothelium presents a prohypertensive phenotype due to the reduction of endothelium's proliferation ability and expression of vasorelaxing factors. PMID:26937388

  16. Secretion of SerpinB2 from endothelial cells activated with inflammatory stimuli.

    PubMed

    Boncela, Joanna; Przygodzka, Patrycja; Wyroba, Elzbieta; Papiewska-Pajak, Izabela; Cierniewski, Czeslaw S

    2013-05-01

    Due to the lack of an N-terminal signal peptide, SerpinB2 (plasminogen activator inhibitor type 2) accumulates in cells and only a small percentage of it is secreted. The extracellular concentration of SerpinB2 significantly increases during inflammation. In the present study we investigated the mechanism with which SerpinB2 can be secreted from endothelial cells activated with LPS. We evaluated the intracellular distribution of SerpinB2 by double immunogold labeling followed by a high resolution electron microscopy analysis. We found that SerpinB2 gathers in the vesicular structures and in the endothelial cell periphery. These vesicles stained positive for the trans-Golgi network marker TGN46, which is consistent with their formation by the endoplasmatic reticulum (ER) and Golgi-dependent pathways. SerpinB2 was delivered to the plasma membrane, apparently together with TGN46 in the same vesicles, which after fusion with the membranes released cargo. Secretion of SerpinB2 was partially inhibited by brefeldin A. The secreted SerpinB2 was predominantly in its nonglycosylated 43kDa form as evaluated by Western immunoblotting. Our data suggest that increased expression of SerpinB2 by an inflammatory stimulus is sufficient to generate structures that resemble secretory vesicles. These vesicles may represent the mechanism by which high local concentrations of SerpinB2 are released at inflammation sites from endothelial cells. PMID:23474086

  17. Novel mechanisms of endothelial mechano-transduction

    PubMed Central

    Abe, Jun-ichi; Berk, Bradford C

    2014-01-01

    Atherosclerosis is a focal disease that develops preferentially where non-laminar, disturbed blood flow (d-flow) occurs such as branches, bifurcations, and curvatures of large arteries. Endothelial cells sense and respond differently to d-flow compared to steady laminar flow (s-flow). D-flow that occurs in so-called athero-prone areas activates pro-inflammatory and apoptotic signaling, and this results in endothelial dysfunction and leads to subsequent development of atherosclerosis. In contrast, s-flow as “athero-protective flow” promotes expression of many anti-inflammatory genes such as Kruppel-like factor 2 (KLF2) and endothelial nitric oxide synthase (eNOS) and inhibits endothelial inflammation and athrogenesis. Here we will discuss that d-flow and s-flow induce pro- and anti-atherogenic events via flow type-specific “mechanotransduction” pathways. We will focus on five mechano-sensitive pathways: MEK5 (MAPK/ERK kinase 5)-ERK5-KLF2 signaling, ERK5-PPAR (peroxisome proliferator-activated receptor) signaling, and mechano-signaling pathways involving SUMOylation, protein kinase C-ζ, (PKCζ), and p90 ribosomal S6 kinase (p90RSK). We believe that clarifying regulation mechanisms between these two flow types will provide new insights into therapeutic approaches for the prevention and treatment of atherosclerosis. PMID:25301843

  18. Lymphatic endothelial lineage assemblage during corneal lymphangiogenesis.

    PubMed

    Connor, Alicia L; Kelley, Philip M; Tempero, Richard M

    2016-03-01

    Postnatal inflammatory lymphangiogenesis presumably requires precise regulatory processes to properly assemble proliferating lymphatic endothelial cells (LECs). The specific mechanisms that regulate the assembly of LECs during new lymphatic vessel synthesis are unclear. Dynamic endothelial shuffling and rearrangement has been proposed as a mechanism of blood vessel growth. We developed genetic lineage-tracing strategies using an inductive transgenic technology to track the fate of entire tandem dimer tomato-positive (tdT) lymphatic vessels or small, in some cases clonal, populations of LECs. We coupled this platform with a suture-induced mouse model of corneal lymphangiogenesis and used different analytic microscopy techniques including serial live imaging to study the spatial properties of proliferating tdT(+) LEC progenies. LEC precursors and their progeny expanded from the corneal limbal lymphatic vessel and were assembled contiguously to comprise a subunit within a new lymphatic vessel. VE-cadherin blockade induced morphologic abnormalities in newly synthesized lymphatic vessels, but did not disrupt the tdT(+) lymphatic endothelial lineage assembly. Analysis of this static and dynamic data based largely on direct in vivo observations supports a model of lymphatic endothelial lineage assemblage during corneal inflammatory lymphangiogenesis. PMID:26658452

  19. Endothelial progenitor cells in hematologic malignancies

    PubMed Central

    Saulle, Ernestina; Castelli, Germana; Pelosi, Elvira

    2016-01-01

    Studies carried out in the last years have improved the understanding of the cellular and molecular mechanisms controlling angiogenesis during adult life in normal and pathological conditions. Some of these studies have led to the identification of some progenitor cells that sustain angiogenesis through indirect, paracrine mechanisms (hematopoietic angiogenic cells) and through direct mechanisms, i.e., through their capacity to generate a progeny of phenotypically and functionally competent endothelial cells [endothelial colony forming cells (ECFCs)]. The contribution of these progenitors to angiogenetic processes under physiological and pathological conditions is intensively investigated. Angiogenetic mechanisms are stimulated in various hematological malignancies, including chronic myeloid leukemia (CML), acute myeloid leukemia (AML), myelodysplastic syndromes and multiple myeloma, resulting in an increased angiogenesis that contributes to disease progression. In some of these conditions there is preliminary evidence that some endothelial cells could derive from the malignant clone, thus leading to the speculation that the leukemic cell derives from the malignant transformation of a hemangioblastic progenitor, i.e., of a cell capable of differentiation to the hematopoietic and to the endothelial cell lineages. Our understanding of the mechanisms underlying increased angiogenesis in these malignancies not only contributed to a better knowledge of the mechanisms responsible for tumor progression, but also offered the way for the discovery of new therapeutic targets. PMID:27583252

  20. Regulation of endothelial cell differentiation and specification

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The circulatory system is the first organ system to develop in the vertebrate embryo and is critical throughout gestation for the delivery of oxygen and nutrients to, as well as removal of metabolic waste products from, growing tissues. Endothelial cells, which constitute the luminal layer of all bl...