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Sample records for endothelial cells synthesize

  1. Endothelial cells are able to synthesize and release catecholamines both in vitro and in vivo.

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    Sorriento, Daniela; Santulli, Gaetano; Del Giudice, Carmine; Anastasio, Antonio; Trimarco, Bruno; Iaccarino, Guido

    2012-07-01

    Recently it has been demonstrated that catecholamines are produced and used by macrophages and mediate immune response. The aim of this study is to verify whether endothelial cells (ECs), which are of myeloid origin, can produce catecholamines. We demonstrated that genes coding for tyrosine hydroxylase, Dopa decarboxylase, dopamine β hydroxylase (DβH), and phenylethanolamine-N-methyl transferase, enzymes involved in the synthesis of catecholamines, are all expressed in basal conditions in bovine aorta ECs, and their expression is enhanced in response to hypoxia. Moreover, hypoxia enhances catecholamine release. To evaluate the signal transduction pathway that regulates catecholamine synthesis in ECs, we overexpressed in bovine aorta ECs either protein kinase A (PKA) or the transcription factor cAMP response element binding, because PKA/cAMP response element binding activation induces tyrosine hydroxylase transcription and activity in response to stress. Both cAMP response element binding and PKA overexpression enhance DβH and phenylethanolamine-N-methyl transferase gene expression and catecholamine release, whereas H89, inhibitor of PKA, exerts the opposite effect, evidencing the role of PKA/cAMP response element binding transduction pathway in the regulation of catecholamine release in bovine aorta ECs. We then evaluated by immunohistochemistry the expression of tyrosine hydroxylase, Dopa decarboxylase, DβH, and phenylethanolamine-N-methyl transferase in femoral arteries from hindlimbs of C57Bl/6 mice 3 days after removal of the common femoral artery to induce chronic ischemia. Ischemia evokes tyrosine hydroxylase, Dopa decarboxylase, DβH, and phenylethanolamine-N-methyl transferase expression in the endothelium. Finally, the pharmacological inhibition of catecholamine release by fusaric acid, an inhibitor of DβH, reduces the ability of ECs to form network-like structures on Matrigel matrix. In conclusion, our study demonstrates for the first time that ECs

  2. Endothelial-regenerating cells: an expanding universe.

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    Steinmetz, Martin; Nickenig, Georg; Werner, Nikos

    2010-03-01

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

  3. Targeting Endothelial Cells with Multifunctional GaN/Fe Nanoparticles

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    Braniste, Tudor; Tiginyanu, Ion; Horvath, Tibor; Raevschi, Simion; Andrée, Birgit; Cebotari, Serghei; Boyle, Erin C.; Haverich, Axel; Hilfiker, Andres

    2017-08-01

    In this paper, we report on the interaction of multifunctional nanoparticles with living endothelial cells. The nanoparticles were synthesized using direct growth of gallium nitride on zinc oxide nanoparticles alloyed with iron oxide followed by core decomposition in hydrogen flow at high temperature. Using transmission electron microscopy, we demonstrate that porcine aortic endothelial cells take up GaN-based nanoparticles suspended in the growth medium. The nanoparticles are deposited in vesicles and the endothelial cells show no sign of cellular damage. Intracellular inert nanoparticles are used as guiding elements for controlled transportation or designed spatial distribution of cells in external magnetic fields.

  4. Endothelial cells, fibroblasts and vasculitis.

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    Buckley, Christopher D; Rainger, G Ed; Nash, Gerard B; Raza, Karim

    2005-07-01

    One of the most important questions in vasculitis research is not why inflammation of blood vessels occurs but why it persists, often in a site-specific manner. In this review we illustrate how stromal cells, such as fibroblasts and pericytes, might play an important role in regulating the site at which vasculitis occurs. Smooth muscle cells and fibroblasts directly influence the behaviour of overlying vascular cells, amplifying the response of the endothelium to proinflammatory agents such as TNF-alpha and allowing enhanced and inappropriate leucocyte recruitment. An abnormal local vascular stromal environment can therefore influence local endothelial function and drive the persistence of local vascular inflammation. However, such local vascular inflammation can have distant effects on the systemic vascular system, leading to widespread endothelial cell dysfunction. Vascular endothelial dysfunction is common in a range of immune-mediated inflammatory diseases, is seen in multiple vascular beds, and is reversible following the induction of disease remission. The mechanisms that drive such systemic vascular endothelial dysfunction are unclear but factors such as TNF-alpha and CRP may play a role. Persistence of such widespread endothelial dysfunction in systemic vasculitis appears to have long-term consequences, leading to the acceleration of atherosclerosis and premature ischaemic heart disease. It may also underlie the accelerated atherosclerosis seen in other immune-mediated rheumatic diseases, such as rheumatoid arthritis.

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

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    Ken Kono

    Full Text Available 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.

  6. Endothelial cell loss after autologous rotational keratoplasty.

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    Birnbaum, Florian; Reinhard, Thomas; Böhringer, Daniel; Sundmacher, Rainer

    2005-01-01

    To investigate whether it may be possible to ascertain the influence of immunological factors on chronic endothelial cell loss by comparing chronic endothelial cell loss after autologous rotational penetrating keratoplasty and after homologous penetrating keratoplasty. For six patients who had undergone autologous rotational penetrating keratoplasty the relative annual loss of endothelial cells was calculated by means of an exponential regression analysis. The findings were compared with those in a homogeneous historical control group (53 patients undergoing homologous penetrating keratoplasty for keratoconus). After autologous rotational keratoplasty relative annual loss of endothelial cells was 1.1%+/-2.6% (mean +/- standard deviation). Relative annual loss of endothelial cells in the control-group was 16.7%+/-20.8%. The results of the study lead to the assumption that immunological influences might be the main cause for chronic endothelial cell loss after homologous penetrating keratoplasty.

  7. [Transplantation of corneal endothelial cells].

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    Amano, Shiro

    2002-12-01

    Though conventional corneal transplantation has achieved great success, it still has several drawbacks including limited availability of donor corneas, recurrent allograft rejection, and subsequent graft failure in certain cases. Reconstructing clinically usable corneas by applying the technology of regenerative medicine can offer a solution to these problems, as well as making corneal transplantation a non-emergency surgery and enabling the usage of banked corneal cells. In the present study, we focused on corneal endothelium that is critical for corneal transparency and investigated the reconstruction of cornea utilizing cultured human corneal endothelial cells (HCECs). We succeeded in steadily culturing HCECs by using culture dishes pre-coated with extracellular matrix produced by calf corneal endothelial cells and culture media that contained basic fibroblast growth factor and fetal bovine serum. We performed the following analysis utilizing these cultured HCECs. The older the donor was, the more frequently large senescent cells appeared in the passaged HCECs. The telomeres of HCECs were measured as terminal restriction fragments (TRF) by Southern blotting. HCECs, in vivo from donors in their seventies had a long TRFs of over 12 kilobases. Passaging shortened the TRFs but there was no difference in TRFs among donors of various ages. These results indicated that shortening of telomere length is not related to senescence of HCECs. We investigated the role of advanced glycation end products (AGEs) in the senescence of in vivo HCECs. The results indicated that AGE-protein in the aqueous humor is endocytosed into HCECs via AGE receptors expressed on the surface of HCECs and damages HCECs by producing reactive oxygen species and inducing apoptosis, suggesting that AGEs, at least partly, cause the senescence of HECEs. HCECs were cultured using adult human serum instead of bovine serum to get rid of bovine material that can be infected with prions. Primary and passage

  8. Circulating Endothelial Cells and Endothelial Progenitor Cells in Pediatric Sepsis.

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    Zahran, Asmaa Mohamad; Elsayh, Khalid Ibrahim; Mohamad, Ismail Lotfy; Hassan, Gamal Mohamad; Abdou, Madleen Adel A

    2016-03-01

    The aim of the study was to measure the number of circulating endothelial cells (CECs) and circulating endothelial progenitor cells (CEPs) in pediatric patients with sepsis and correlating it with the severity of the disease and its outcome. The study included 19 children with sepsis, 26 with complicated sepsis, and 30 healthy controls. The patients were investigated within 48 hours of pediatric intensive care unit admission together with flow cytometric detection of CECs and CEPs. The levels of both CECs and CEPs were significantly higher in patient with sepsis and complicated sepsis than the controls. The levels of CECs were higher in patients with complicated sepsis, whereas the levels of CEPs were lower in patients with complicated sepsis. Comparing the survival and nonsurvival septic patients, the levels of CEPs were significantly higher in the survival than in nonsurvival patients, whereas the levels of CECs were significantly lower in the survival than in nonsurvival patients. Serum albumin was higher in survival than in nonsurvival patients. Estimation of CECs and CEPs and their correlation with other parameters such as serum albumen could add important information regarding prognosis in septic pediatric patients.

  9. PPAR Gamma and Angiogenesis: Endothelial Cells Perspective

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    Jerzy Kotlinowski

    2016-01-01

    Full Text Available We summarize the current knowledge concerning PPARγ function in angiogenesis. We discuss the mechanisms of action for PPARγ and its role in vasculature development and homeostasis, focusing on endothelial cells, endothelial progenitor cells, and bone marrow-derived proangiogenic cells.

  10. Molecular expression in transfected corneal endothelial cells

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    Wang, Fan; Miao, Zhuang; Lu, Chengwei; Hao, Jilong

    2017-10-01

    To investigate the capability of human corneal endothelial cells serving as immunological cells. Expression of HLA-DP, -DQ, -DR, CD40, CD80, and CD86 was determined by immunohistochemical methods. Meanwhile, purified peripheral blood mononuclear cells were cocultured with human corneal endothelial cells which were pre-treated with and without -IFN respectively, activation of lymphocytes was determined by FACS analysis. In coculture system, T lymphocyte was activated by corneal endothelial cells, HLA-DP, -DQ, -DR and CD40 expression were increased by - IFN induction. Costimulatory molecular CD80 was shown on the endothelial cells. Human corneal endothelial cells were assumed to be involved in the corneal transplantation rejection process as potential antigen presenting cells.

  11. Estetrol modulates endothelial nitric oxide synthesis in human endothelial cells

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    Maria Magdalena eMontt-Guevara

    2015-07-01

    Full Text Available Estetrol (E4 is a natural human estrogen that is present at high concentrations during pregnancy. E4 has been reported to act as an endogenous estrogen receptor modulator, exerting estrogenic actions on the endometrium or the central nervous system but presenting antagonistic effects on the breast. Due to these characteristics, E4 is currently being developed for a number of clinical applications, including contraception and menopausal hormone therapy. Endothelial nitric oxide (NO is a key player for vascular function and disease during pregnancy and throughout ageing in women. Endothelial NO is an established target of estrogens that enhance its formation in human endothelial cells. We here addressed the effects of E4 on the activity and expression of the endothelial nitric oxide synthase (eNOS in cultured human umbilical vein endothelial cells (HUVEC. E4 stimulated the activation of eNOS and NO secretion in HUVEC. E4 was significantly less effective compared to E2 and a peculiar concentration-dependent effect was found, with higher amounts of E4 being less effective than lower concentrations. When E2 was combined with E4, an interesting pattern was noted. E4 antagonized NO synthesis induced by pregnancy-like E2 concentrations. However, E4 did not impede the modest induction of NO synthesis associated with postmenopausal-like E2 levels. These results support the hypothesis that E4 may be a regulator of NO synthesis in endothelial cells and raise questions on its peculiar signaling in this context. Our results may be useful to interpret the role of E4 during human pregnancy and possibly to help develop this interesting steroid for clinical use.

  12. Apicobasal polarity of brain endothelial cells.

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    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. © The Author(s) 2015.

  13. Endothelial progenitor cell dysfunction in diabetes mellitus

    NARCIS (Netherlands)

    Loomans, Cindy Johanna Maria

    2007-01-01

    Postnatally, Endothelial Progenitor Cells are needed to maintain the integrity of the endothelium (re-endothelialization) and to augment wound healing or vascularize hypoxic areas (neovascularization). Complex networks of different signals and regulators have been identified to be involved in these

  14. Cataract phacoemulsification and corneal endothelial cell damage

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    Ni Zhu

    2013-07-01

    Full Text Available Phacoemulsification with small incision, reduced number of inflammation cells, and better postoperative recovery has been recognized as the world's most popular option for cataract surgery. Modern cataract surgery is developing gradually from sight rehabilitating to refractive surgery with better vision acuity. Being the most important part of the eye refractive system, maintenance of the cornea's transparency relies heavily upon the healthy endothelial cells. It is well known that there will be endothelial cell loss after phacoemulsification and the damage of the endothelial cells may lead to corneal swellings and opacity, or even the corneal descompensation, which often severely influenced the postoperative vision recovery. This is a review of phacoemulsification and the risk factors of corneal endothelial damage pre-and postoperation.

  15. Dysfunctional Endothelial Progenitor Cells in Metabolic Syndrome

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    Devaraj, Sridevi; Jialal, Ishwarlal

    2012-01-01

    The metabolic syndrome (MetS) is highly prevalent and confers an increased risk of diabetes and cardiovascular disease. A key early event in atherosclerosis is endothelial dysfunction. Numerous groups have reported endothelial dysfunction in MetS. However, the measurement of endothelial function is far from optimum. There has been much interest recently in a subtype of progenitor cells, termed endothelial progenitor cells (EPCs), that can circulate, proliferate, and dfferentiate into mature endothelial cells. EPCs can be characterized by the assessment of surface markers, CD34 and vascular endothelial growth factor receptor-2, VEGFR-2 (KDR). The CD34+KDR+ phenotype has been demonstrated to be an independent predictor of cardiovascular outcomes. MetS patients without diabetes or cardiovascular diseases have decreased EPC number and functionality as evidenced by decreased numbers of colony forming units, decreased adhesion and migration, and decreased tubule formation. Strategies that have been shown to upregulate and enhance EPC number and functionality include statins, angiotensin converting enzyme inhibitors, angiotensin receptor blockers, and peroxisome-proliferator-activating-receptor gamma agonists. Mechanisms by which they affect EPC number and functionality need to be studied. Thus, EPC number and/or functionality could emerge as novel cellular biomarkers of endothelial dysfunction and cardiovascular disease risk in MetS. PMID:21941528

  16. Heterologous corneal endothelial cell transplantation--human corneal endothelial cell transplantation in Lewis rats.

    OpenAIRE

    Tchah, H.

    1992-01-01

    A heterologous corneal endothelial transplantation was attempted using human endothelial cells and a Lewis rat penetrating keratoplasty model. Cultured human endothelial cells were seeded to a Lewis rat cornea, which was denuded of its endothelium. When grafted into the syngeneic Lewis rat, the graft remained clear for at least five days, and then became opaque and edematous because of immune rejection reaction. In contrast, corneas denuded of their endothelium became opaque and edematous imm...

  17. Endothelial cells derived from human embryonic stem cells

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    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. Reduced Ang2 expression in aging endothelial cells

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    Hohensinner, P.J., E-mail: philipp.hohensinner@meduniwien.ac.at [Department of Internal Medicine II, Medical University of Vienna, Vienna (Austria); Ebenbauer, B. [Department of Internal Medicine II, Medical University of Vienna, Vienna (Austria); Ludwig Boltzmann Cluster for Cardiovascular Research, Vienna (Austria); Kaun, C.; Maurer, G. [Department of Internal Medicine II, Medical University of Vienna, Vienna (Austria); Huber, K. [Ludwig Boltzmann Cluster for Cardiovascular Research, Vienna (Austria); 3rd Medical Department, Wilhelminenhospital, Vienna (Austria); Sigmund Freud University, Medical Faculty, Vienna (Austria); Wojta, J. [Department of Internal Medicine II, Medical University of Vienna, Vienna (Austria); Ludwig Boltzmann Cluster for Cardiovascular Research, Vienna (Austria); Core Facilities, Medical University of Vienna, Vienna (Austria)

    2016-06-03

    Aging endothelial cells are characterized by increased cell size, reduced telomere length and increased expression of proinflammatory cytokines. In addition, we describe here that aging reduces the migratory distance of endothelial cells. Furthermore, we observe an increase of the quiescence protein Ang1 and a decrease of the endothelial activation protein Ang2 upon aging. Supplementing Ang2 to aged endothelial cells restored their migratory capacity. We conclude that aging shifts the balance of the Ang1/Ang2 network favouring a quiescent state. Activation of endothelial cells in aging might be necessary to enhance wound healing capacities. -- Highlights: •Endothelial cells display signs of aging before reaching proliferative senescence. •Aging endothelial cells express more angiopoietin 1 and less angiopoietin 2 than young endothelial cells. •Migratory capacity is reduced in aging endothelial cells.

  19. Bacteria and endothelial cells: a toxic relationship.

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    Lubkin, Ashira; Torres, Victor J

    2017-02-01

    Pathogenic bacteria use the bloodstream as a highway for getting around the body, and thus have to find ways to enter and exit through the endothelium. Many bacteria approach this problem by producing toxins that can breach the endothelial barrier through diverse creative mechanisms, including directly killing endothelial cells (ECs), weakening the cytoskeleton within ECs, and breaking the junctions between ECs. Toxins can also modulate the immune response by influencing endothelial biology, and can modulate endothelial function by influencing the response of leukocytes. Understanding these interactions, in both the in vitro and in vivo contexts, is of critical importance for designing new therapies for sepsis and other severe bacterial diseases. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Endothelial progenitor cells in chronic obstructive pulmonary disease and emphysema

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

    2017-01-01

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

  1. Endothelial progenitor cells in chronic obstructive pulmonary disease and emphysema.

    Science.gov (United States)

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

    2017-01-01

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

  2. Adenovirus vectors can induce activation of endothelial cells: CD40 ...

    African Journals Online (AJOL)

    Adenovirus vectors can induce activation of endothelial cells: CD40-CD40L interactions partly participate in the endothelial cells activation induced by adenovirus vectors in an NF-kappaB-dependent manner.

  3. Identification of epigenetically silenced genes in tumor endothelial cells

    NARCIS (Netherlands)

    Hellebrekers, Debby M. E. I.; Melotte, Veerle; Vire, Emmanuelle; Langenkamp, Elise; Molema, Grietje; Fuks, Francois; Herman, James G.; Van Criekinge, Wim; Griffioen, Arjan W.; van Engeland, Manon

    2007-01-01

    Tumor angiogenesis requires intricate regulation of gene expression in endothelial cells. We recently showed that DNA methyltransferase (DNMT) and histone deacetylase (HDAC) inhibitors directly repress endothelial cell growth and tumor angiogenesis, suggesting that epigenetic modifications mediated

  4. Spreading endothelial cell dysfunction in response to necrotic trophoblasts. Soluble factors released from endothelial cells that have phagocytosed necrotic shed trophoblasts reduce the proliferation of additional endothelial cells.

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    Chen, Q; Ding, J X; Liu, B; Stone, P; Feng, Y J; Chamley, L

    2010-11-01

    The pathogenesis of preeclampsia is not clear but the disease is characterised by systemic endothelial cell dysfunction that is considered to be triggered by a placental factor. Necrotic trophoblastic debris that is deported in the maternal blood is one possible placental trigger for preeclampsia. Syncytial knots were first associated with preeclampsia over 100 years ago. However, syncytial knots are very large and most are trapped in the pulmonary capillaries making it difficult to envisage how they could lead to widespread systemic endothelial cell dysfunction. This study was undertaken to examine whether conditioned medium from endothelial cells that have phagocytosed necrotic trophoblastic debris could adversely affect the proliferation or survival of fresh endothelial cells. Trophoblastic cellular debris, harvested from placental explants was added to endothelial cell monolayers directly or after induction of necrosis by freeze-thawing. Conditioned medium from the endothelial cell cultures was exposed to fresh endothelial cells and their proliferation measured by Alamar Blue, and CyQUANTNF cell proliferation assays. Endothelial cell death was examined by a fluorogenic caspase-3 activity assay and LDH release. Conditioned medium from endothelial cells that had phagocytosed necrotic but not apoptotic trophoblastic debris significantly inhibited the proliferation of fresh endothelial cells but did not induce their death. The conditioned medium also reduced cell-surface endoglin expression by fresh endothelial cells. These results confirm that phagocytosis of necrotic trophoblastic debris by endothelial cells results in the secretion of soluble factors that might explain how necrotic trophoblastic debris trapped in the pulmonary capillaries could induce systemic endothelial cell dysfunction. Copyright © 2010 Elsevier Ltd. All rights reserved.

  5. Endothelial progenitor cell biology in ankylosing spondylitis.

    Science.gov (United States)

    Verma, Inderjeet; Syngle, Ashit; Krishan, Pawan

    2015-03-01

    Endothelial progenitor cells (EPCs) are unique populations which have reparative potential in overcoming endothelial damage and reducing cardiovascular risk. Patients with ankylosing spondylitis (AS) have increased risk of cardiovascular morbidity and mortality. The aim of this study was to investigate the endothelial progenitor cell population in AS patients and its potential relationships with disease variables. Endothelial progenitor cells were measured in peripheral blood samples from 20 AS and 20 healthy controls by flow cytometry on the basis of CD34 and CD133 expression. Disease activity was evaluated by using Bath Ankylosing Spondylitis Disease Activity Index (BASDAI). Functional ability was monitored by using Bath Ankylosing Spondylitis Functional Index (BASFI). EPCs were depleted in AS patients as compared to healthy controls (CD34(+) /CD133(+) : 0.027 ± 0.010% vs. 0.044 ± 0.011%, P < 0.001). EPC depletions were significantly associated with disease duration (r = -0.52, P = 0.01), BASDAI (r = -0.45, P = 0.04) and C-reactive protein (r = -0.5, P = 0.01). This is the first study to demonstrate endothelial progenitor cell depletion in AS patients. EPC depletions inversely correlate with disease duration, disease activity and inflammation, suggesting the pivotal role of inflammation in depletion of EPCs. EPC would possibly also serve as a therapeutic target for preventing cardiovascular disease in AS. © 2014 Asia Pacific League of Associations for Rheumatology and Wiley Publishing Asia Pty Ltd.

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

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    Caterina Oriana Aragona

    2016-01-01

    Full Text Available 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.

  7. Endothelial cell oxidative stress and signal transduction

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    ROCIO FONCEA

    2000-01-01

    Full Text Available Endothelial dysfunction (ED is an early event in atherosclerotic disease, preceding clinical manifestations and complications. Increased reactive oxygen species (ROS have been implicated as important mechanisms that contribute to ED, and ROS’s may function as intracellular messengers that modulate signaling pathways. Several intracellular signal events stimulated by ROS have been defined, including the identification of two members of the mitogen activated protein kinase family (ERK1/2 and big MAP kinase, BMK1, tyrosine kinases (Src and Syk and different isoenzymes of PKC as redox-sensitive kinases. ROS regulation of signal transduction components include the modification in the activity of transcriptional factors such as NFkB and others that result in changes in gene expression and modifications in cellular responses. In order to understand the intracellular mechanisms induced by ROS in endothelial cells (EC, we are studying the response of human umbilical cord vein endothelial cells to increased ROS generation by different pro-atherogenic stimuli. Our results show that Homocysteine (Hcy and oxidized LDL (oxLDL enhance the activity and expression of oxidative stress markers, such as NFkB and heme oxygenase 1. These results suggest that these pro-atherogenic stimuli increase oxidative stress in EC, and thus explain the loss of endothelial function associated with the atherogenic process

  8. Oligodendrocyte precursor cells synthesize neuromodulatory factors.

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    Dominik Sakry

    Full Text Available NG2 protein-expressing oligodendrocyte progenitor cells (OPC are a persisting and major glial cell population in the adult mammalian brain. Direct synaptic innervation of OPC by neurons throughout the brain together with their ability to sense neuronal network activity raises the question of additional physiological roles of OPC, supplementary to generating myelinating oligodendrocytes. In this study we investigated whether OPC express neuromodulatory factors, typically synthesized by other CNS cell types. Our results show that OPC express two well-characterized neuromodulatory proteins: Prostaglandin D2 synthase (PTGDS and neuronal Pentraxin 2 (Nptx2/Narp. Expression levels of the enzyme PTGDS are influenced in cultured OPC by the NG2 intracellular region which can be released by cleavage and localizes to glial nuclei upon transfection. Furthermore PTGDS mRNA levels are reduced in OPC from NG2-KO mouse brain compared to WT cells after isolation by cell sorting and direct analysis. These results show that OPC can contribute to the expression of these proteins within the CNS and suggest PTGDS expression as a downstream target of NG2 signaling.

  9. Syncytin is involved in breast cancer-endothelial cell fusions

    DEFF Research Database (Denmark)

    Bjerregaard, B; Holck, Susanne; Christensen, Ib Jarle

    2006-01-01

    Cancer cells can fuse spontaneously with normal host cells, including endothelial cells, and such fusions may strongly modulate the biological behaviour of tumors. However, the underlying mechanisms are unknown. We now show that human breast cancer cell lines and 63 out of 165 (38%) breast cancer...... and inhibits fusions between breast cancer cells and endothelial cells. Moreover, a syncytin inhibitory peptide also inhibits fusions between cancer and endothelial cells. These results are the first to show that syncytin is expressed by human cancer cells and is involved in cancer-endothelial cell fusions....

  10. Oxidative stress induced pulmonary endothelial cell proliferation is ...

    African Journals Online (AJOL)

    Cellular hyper-proliferation, endothelial dysfunction and oxidative stress are hallmarks of the pathobiology of pulmonary hypertension. Indeed, pulmonary endothelial cells proliferation is susceptible to redox state modulation. Some studies suggest that superoxide stimulates endothelial cell proliferation while others have ...

  11. Endothelial cells, fibroblasts and vasculitis

    OpenAIRE

    Buckley, Christopher D.; Rainger, G.Ed; Nash, Gerard B; Raza, Karim

    2005-01-01

    One of the most important questions in vasculitis research is not why inflammation of blood vessels occurs but why it persists, often in a site-specific manner. In this review we illustrate how stromal cells, such as fibroblasts and pericytes, might play an important role in regulating the site at which vasculitis occurs. Smooth muscle cells and fibroblasts directly influence the behaviour of overlying vascular cells, amplifying the response of the endothelium to proinflammatory agents such a...

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

    NARCIS (Netherlands)

    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

  13. Arecoline is cytotoxic for human endothelial cells.

    Science.gov (United States)

    Ullah, Mafaz; Cox, Stephen; Kelly, Elizabeth; Boadle, Ross; Zoellner, Hans

    2014-11-01

    Oral submucous fibrosis is a pre-malignant fibrotic condition caused by areca nut use and involves reduced mucosal vascularity. Arecoline is the principal areca nut alkaloid and is cytotoxic for epithelium and fibroblasts. Endothelial cell cycle arrest is reported on exposure to arecoline, as is cytotoxicity for endothelial-lung carcinoma hybrid cells. We here describe cytotoxicity for primary human endothelial cultures from seven separate donors. Human umbilical vein endothelial cells were exposed to increasing concentrations of arecoline and examined by: phase-contrast microscopy, haemocytometer counts, transmission electron microscopy, lactate dehydrogenase release and the methyl-thiazol-tetrazolium assay. Vacuolation and detachment of endothelium were observed at and above arecoline concentrations of 333 μg/ml or more. Ultrastructural features of cellular stress were seen after 24-h treatment with 111 μg/ml arecoline and included reduced ribosomal studding of endoplasmic reticulum, increased autophagolysosomal structures, increased vacuolation and reduced mitochondrial cristae with slight swelling. Similar changes were seen at 4 h with arecoline at 333 μg/ml or above, but with more severe mitochondrial changes including increased electron density of mitochondrial matrix and greater cristal swelling, while by 24 h, these cells were frankly necrotic. Haemocytometer counts were paralleled by both lactate dehydrogenase release and the methyl-thiazol-tetrazolium assays. Arecoline is cytotoxic via necrosis for endothelium, while biochemical assays indicate no appreciable cellular leakage before death and detachment, as well as no clear effect on mitochondrial function in viable cells. Arecoline toxicity may thus contribute to reduced vascularity in oral submucous fibrosis. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Transition of mesenchymal stem/stromal cells to endothelial cells

    NARCIS (Netherlands)

    M. Crisan (Mihaela)

    2013-01-01

    textabstractMesenchymal stem/stromal cells (MSCs) are heterogeneous. A fraction of these cells constitute multipotent cells that can self-renew and mainly give rise to mesodermal lineage cells such as adipocytes, osteocytes and chondrocytes. The ability of MSCs to differentiate into endothelial

  15. Soluble endothelial cell molecules and circulating endothelial cells in patients with venous thromboembolism.

    Science.gov (United States)

    Torres, Cláudia; Matos, Rui; Morais, Sara; Campos, Manuel; Lima, Margarida

    2017-12-01

    : To evaluate the plasma levels of soluble endothelial cell molecules in patients with venous thromboembolism (VTE) out of the acute phase as compared with healthy individuals. We also investigated the possible associations of the soluble endothelial cell molecules among them, as well as with other clinical and laboratory data, including the numbers of circulating endothelial cells (CEC), circulating endothelial progenitor cells (CEP), and CEC expressing activation-related [cluster of differentiation (CD)54 and CD62E] and procoagulant (CD142) markers. In total, 15 patients with VTE and 20 normal individuals were studied. The CEC and CEP were quantified and characterized by flow cytometry. The soluble molecules studied included P-selectin, E-selectin, intercellular cell adhesion molecule 1, vascular cell adhesion molecule 1 and tissue factor (ELISA), and von Willebrand factor antigen (immunoturbidimetry). VTE patients had significantly higher levels of vascular cell adhesion molecule 1 and von Willebrand factor antigen and lower levels of soluble E-selectin than controls. They also showed significantly higher numbers of CEC, as of activated/procoagulant CEC and lower numbers of CEP, compared with controls. We did not find any correlation between the levels of soluble molecules and the numbers of endothelial cell in circulation, but there was with several clinical and laboratory data in VTE patients. Our results would suggest that in VTE patients, the endothelium remains activated and in some hypercoagulable state. The levels of soluble endothelial cell molecules did not seem to be directly related to the numbers of CEC and CEP neither reflected the number of activated CEC, which may be because of the different function that surface and soluble molecules may have.

  16. Cell-cell junctional mechanotransduction in endothelial remodeling.

    Science.gov (United States)

    Dorland, Yvonne L; Huveneers, Stephan

    2017-01-01

    The vasculature is one of the most dynamic tissues that encounter numerous mechanical cues derived from pulsatile blood flow, blood pressure, activity of smooth muscle cells in the vessel wall, and transmigration of immune cells. The inner layer of blood and lymphatic vessels is covered by the endothelium, a monolayer of cells which separates blood from tissue, an important function that it fulfills even under the dynamic circumstances of the vascular microenvironment. In addition, remodeling of the endothelial barrier during angiogenesis and trafficking of immune cells is achieved by specific modulation of cell-cell adhesion structures between the endothelial cells. In recent years, there have been many new discoveries in the field of cellular mechanotransduction which controls the formation and destabilization of the vascular barrier. Force-induced adaptation at endothelial cell-cell adhesion structures is a crucial node in these processes that challenge the vascular barrier. One of the key examples of a force-induced molecular event is the recruitment of vinculin to the VE-cadherin complex upon pulling forces at cell-cell junctions. Here, we highlight recent advances in the current understanding of mechanotransduction responses at, and derived from, endothelial cell-cell junctions. We further discuss their importance for vascular barrier function and remodeling in development, inflammation, and vascular disease.

  17. Receptor-Mediated Transport of Insulin across Endothelial Cells

    Science.gov (United States)

    King, George L.; Johnson, Sandra M.

    1985-03-01

    Hormones such as insulin are transported from the interior to the exterior of blood vessels. Whether endothelial cells, which line the inner walls of blood vessels have a role in this transport of hormones is not clear, but it is known that endothelial cells can internalize and release insulin rapidly with little degradation. The transport of iodine-125-labeled insulin was measured directly through the use of dual chambers separated by a horizontal monolayer of cultured bovine aortic endothelial cells. In this setting, endothelial cells took up and released the labeled insulin, thereby transporting it across the cells. The transport of insulin across the endothelial cells was temperature sensitive and was inhibited by unlabeled insulin and by antibody to insulin receptor in proportion to the ability of these substances to inhibit insulin binding to its receptor. More than 80 percent of the transported insulin was intact. These data suggest that insulin is rapidly transported across endothelial cells by a receptor-mediated process.

  18. Optical Investigations of Endothelial Cell Motility

    DEFF Research Database (Denmark)

    Rossen, Ninna Struck

    A monolayer of endothelial cells lines the entire circulatory system and create a barrier between the circulatory system and the tissues. To create and maintain an intact barrier, the individual cells have to connect tightly with their neighbors, which causes a highly correlated motion between...... the cells within the monolayer. The cells have to maintain this barrier while apoptotic cells are being replaced and even while new blood vessels are being created. Meanwhile they are constantly exposed to a shear stress from the ow of blood through the vessels. These extreme micro-environmental conditions....... Below is a summary of the thesis' general outline to give an overview of the thesis and provide the structure of how the di erent projects described in individual chapters relate to each other. Each chapter describes a separate project and will contain a small introduction that address the issues...

  19. Production of soluble Neprilysin by endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Kuruppu, Sanjaya, E-mail: Sanjaya.Kuruppu@monash.edu [Department of Biochemistry and Molecular Biology, Building 77, Monash University, Wellington Rd, Clayton, Vic 3800 (Australia); Rajapakse, Niwanthi W. [Department of Physiology, Building 13F, Monash University, Wellington Rd, Clayton, Vic 3800 (Australia); Minond, Dmitriy [Torrey Pines Institute for Molecular Studies, 11350 SW Village Parkway, Port Saint Lucie, FL 34987 (United States); Smith, A. Ian [Department of Biochemistry and Molecular Biology, Building 77, Monash University, Wellington Rd, Clayton, Vic 3800 (Australia)

    2014-04-04

    Highlights: • A soluble full-length form of Neprilysin exists in media of endothelial cells. • Exosomal release is the key mechanism for the production of soluble Neprilysin. • Inhibition of ADAM-17 by specific inhibitors reduce Neprilysin release. • Exosome mediated release of Neprilysin is dependent on ADAM-17 activity. - Abstract: A non-membrane bound form of Neprilysin (NEP) with catalytic activity has the potential to cleave substrates throughout the circulation, thus leading to systemic effects of NEP. We used the endothelial cell line Ea.hy926 to identify the possible role of exosomes and A Disintegrin and Metalloprotease 17 (ADAM-17) in the production of non-membrane bound NEP. Using a bradykinin based quenched fluorescent substrate (40 μM) assay, we determined the activity of recombinant human NEP (rhNEP; 12 ng), and NEP in the media of endothelial cells (10% v/v; after 24 h incubation with cells) to be 9.35 ± 0.70 and 6.54 ± 0.41 μmols of substrate cleaved over 3 h, respectively. The presence of NEP in the media was also confirmed by Western blotting. At present there are no commercially available inhibitors specific for ADAM-17. We therefore synthesised two inhibitors TPI2155-14 and TPI2155-17, specific for ADAM-17 with IC{sub 50} values of 5.36 and 4.32 μM, respectively. Treatment of cells with TPI2155-14 (15 μM) and TPI2155-17 (4.3 μM) resulted in a significant decrease in NEP activity in media (62.37 ± 1.43 and 38.30 ± 4.70, respectively as a % of control; P < 0.0001), implicating a possible role for ADAM-17 in NEP release. However, centrifuging media (100,000g for 1 h at 4 °C) removed all NEP activity from the supernatant indicating the likely role of exosomes in the release of NEP. Our data therefore indicated for the first time that NEP is released from endothelial cells via exosomes, and that this process is dependent on ADAM-17.

  20. Endothelial Progenitor Cells Enter the Aging Arena.

    Directory of Open Access Journals (Sweden)

    Kate eWilliamson

    2012-02-01

    Full Text Available Age is a significant risk factor for the development of vascular diseases, such as atherosclerosis. Although pharmacological treatments, including statins and anti-hypertensive drugs, have improved the prognosis for patients with cardiovascular disease, it remains a leading cause of mortality in those aged 65 years and over. Furthermore, given the increased life expectancy of the population in developed countries, there is a clear need for alternative treatment strategies. Consequently, the relationship between aging and progenitor cell-mediated repair is of great interest. Endothelial progenitor cells (EPCs play an integral role in the cellular repair mechanisms for endothelial regeneration and maintenance. However, EPCs are subject to age-associated changes that diminish their number in circulation and function, thereby enhancing vascular disease risk. A great deal of research is aimed at developing strategies to harness the regenerative capacity of these cells.In this review, we discuss the current understanding of the cells termed ‘EPCs’, examine the impact of age on EPC-mediated repair and identify therapeutic targets with potential for attenuating the age-related decline in vascular health via beneficial actions on EPCs.

  1. Differentiation state determines neural effects on microvascular endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Muffley, Lara A., E-mail: muffley@u.washington.edu [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Pan, Shin-Chen, E-mail: pansc@mail.ncku.edu.tw [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Smith, Andria N., E-mail: gnaunderwater@gmail.com [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Ga, Maricar, E-mail: marga16@uw.edu [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Hocking, Anne M., E-mail: ahocking@u.washington.edu [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Gibran, Nicole S., E-mail: nicoleg@u.washington.edu [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States)

    2012-10-01

    Growing evidence indicates that nerves and capillaries interact paracrinely in uninjured skin and cutaneous wounds. Although mature neurons are the predominant neural cell in the skin, neural progenitor cells have also been detected in uninjured adult skin. The aim of this study was to characterize differential paracrine effects of neural progenitor cells and mature sensory neurons on dermal microvascular endothelial cells. Our results suggest that neural progenitor cells and mature sensory neurons have unique secretory profiles and distinct effects on dermal microvascular endothelial cell proliferation, migration, and nitric oxide production. Neural progenitor cells and dorsal root ganglion neurons secrete different proteins related to angiogenesis. Specific to neural progenitor cells were dipeptidyl peptidase-4, IGFBP-2, pentraxin-3, serpin f1, TIMP-1, TIMP-4 and VEGF. In contrast, endostatin, FGF-1, MCP-1 and thrombospondin-2 were specific to dorsal root ganglion neurons. Microvascular endothelial cell proliferation was inhibited by dorsal root ganglion neurons but unaffected by neural progenitor cells. In contrast, microvascular endothelial cell migration in a scratch wound assay was inhibited by neural progenitor cells and unaffected by dorsal root ganglion neurons. In addition, nitric oxide production by microvascular endothelial cells was increased by dorsal root ganglion neurons but unaffected by neural progenitor cells. -- Highlights: Black-Right-Pointing-Pointer Dorsal root ganglion neurons, not neural progenitor cells, regulate microvascular endothelial cell proliferation. Black-Right-Pointing-Pointer Neural progenitor cells, not dorsal root ganglion neurons, regulate microvascular endothelial cell migration. Black-Right-Pointing-Pointer Neural progenitor cells and dorsal root ganglion neurons do not effect microvascular endothelial tube formation. Black-Right-Pointing-Pointer Dorsal root ganglion neurons, not neural progenitor cells, regulate

  2. Endothelial Cell Implantation and Survival within Experimental Gliomas

    Science.gov (United States)

    Lal, Bachchu; Indurti, Ravi R.; Couraud, Pierre-Olivier; Goldstein, Gary W.; Laterra, John

    1994-10-01

    The delivery of therapeutic genes to primary brain neoplasms opens new opportunities for treating these frequently fatal tumors. Efficient gene delivery to tissues remains an important obstacle to therapy, and this problem has unique characteristics in brain tumors due to the blood-brain and blood-tumor barriers. The presence of endothelial mitogens and vessel proliferation within solid tumors suggests that genetically modified endothelial cells might efficiently transplant to brain tumors. Rat brain endothelial cells immortalized with the adenovirus E1A gene and further modified to express the β-galactosidase reporter were examined for their ability to survive implantation to experimental rat gliomas. Rats received 9L, F98, or C6 glioma cells in combination with endothelial cells intracranially to caudate/putamen or subcutaneously to flank. Implanted endothelial cells were identified by β-galactosidase histochemistry or by polymerase chain reaction in all tumors up to 35 days postimplantation, the latest time examined. Implanted endothelial cells appeared to cooperate in tumor vessel formation and expressed the brain-specific endothelial glucose transporter type 1 as identified by immunohistochemistry. The proliferation of implanted endothelial cells was supported by their increased number within tumors between postimplantation days 14 and 21 (P = 0.015) and by their expression of the proliferation antigen Ki67. These findings establish that genetically modified endothelial cells can be stably engrafted to growing gliomas and suggest that endothelial cell implantation may provide a means of delivering therapeutic genes to brain neoplasms and other solid tumors. In addition, endothelial implantation to brain may be useful for defining mechanisms of brain-specific endothelial differentiation.

  3. Endothelial progenitor cells, cardiovascular risk factors and lifestyle modifications.

    Science.gov (United States)

    Di Stefano, Rossella; Felice, Francesca; Feriani, Roberto; Balbarini, Alberto

    2013-04-01

    Endothelial progenitor cells (EPCs) contribute substantially to preservation of a structurally and functionally intact endothelium. EPCs home in to the sites of endothelial injury and ischemia, where they proliferate, differentiate and integrate into the endothelial layer or exert a paracrine function by producing vascular growth factors. This review will focus on successful lifestyle interventions that aim to maintain vascular health through beneficial actions on cell populations with vasculogenic potential. The results of the studies proving the role of healthy lifestyle are particularly emphasized.

  4. Chemical functionalization of bioceramics to enhance endothelial cells adhesion for tissue engineering.

    Science.gov (United States)

    Borcard, Françoise; Staedler, Davide; Comas, Horacio; Juillerat, Franziska Krauss; Sturzenegger, Philip N; Heuberger, Roman; Gonzenbach, Urs T; Juillerat-Jeanneret, Lucienne; Gerber-Lemaire, Sandrine

    2012-09-27

    To control the selective adhesion of human endothelial cells and human serum proteins to bioceramics of different compositions, a multifunctional ligand containing a cyclic arginine-glycine-aspartate (RGD) peptide, a tetraethylene glycol spacer, and a gallate moiety was designed, synthesized, and characterized. The binding of this ligand to alumina-based, hydroxyapatite-based, and calcium phosphate-based bioceramics was demonstrated. The conjugation of this ligand to the bioceramics induced a decrease in the nonselective and integrin-selective binding of human serum proteins, whereas the binding and adhesion of human endothelial cells was enhanced, dependent on the particular bioceramics.

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

    Science.gov (United States)

    Suschek, C; Kolb, H; Kolb-Bachofen, V

    1997-12-01

    1. Dobesilate is used for normalizing vascular dysfunction in a number of diseases. In search for an effect on endothelial NO production, macrovascular endothelial cells from rat aorta, microvascular endothelial cells from rat exocrine pancreatic tissue, and capillary endothelial cells from rat islets, were cultured in the presence or absence of Mg-Dobesilate. The activity of constitutive nitric oxide synthase (ecNOS) in resident cells as well as of inducible nitric oxide synthase (iNOS) in cytokine-activated cells was measured indirectly by recording the citrulline concentrations in culture supernatants. 2. In each of the different endothelial cells Mg-Dobesilate incubation (0.25-1 mM) for 24 h led to a significant and concentration-dependent increase in ecNOS-activities. With cytokine-activated endothelial cell cultures only moderate effects were seen with little or no concentration-dependency. Addition of the NOS-inhibitor N(G)-monomethyl-L-arginine led to a significant suppression of citrulline formation in all cultures as an evidence for the enzyme specificity of these effects. 3. iNOS- and ecNOS-specific reverse transcription and semi-quantitative polymerase chain reaction (RT-PCR) with RNA from resident or cytokine-activated endothelial cells gave no evidence for an increase in NOS-specific mRNA after Mg-Dobesilate-treatment. Furthermore, Dobesilate-mediated enhancement of NO synthesis in resting endothelial cells was not due to iNOS induction in these cells, as no iNOS-specific signal was found by RT-PCR.

  6. Radioprotection of mouse CNS endothelial cells in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Lyubimova, N.; Coultas, P.; Martin, R. [Peter McCallum Cancer Institute, Melbourne, VIC (Australia)

    1996-12-31

    Full text: Radioprotection using the minor groove binding DNA ligand Hoechst 33342 has been demonstrated in vitro, and more recently in vivo, in mouse lung. Intravenous administration was used for the lung studies, and both endothelial and alveolar epithelial cells-showed good up-take. Radiation damage to the endothelial cell population has also been postulated as important in late developing radionecrosis of spinal cord and brain. Endothelial cell density in brain can be readily determined by a fluorescent-histochemical technique. Treatment with a monoamine oxidase inhibitor and subsequent injection with L-DOPA results in an accumulation of dopamine (DA) in CNS endothelial cells. DA is converted to a fluorophore by exposure to paraformaldehyde, and cell numbers assayed by fluorescence microscopy. Earlier studies used this technique to monitor post-irradiation changes in endothelial cell density in rodent brain and showed the loss, within 24 hours, of a sensitive subpopulation comprising about 15% of the endothelial cells. Ten minutes after intravenous injection of Hoechst 33342 (80mg/kg) the ligand is confined by its limited penetration to the endothelial cells in mouse brain. When we irradiated at this time, there was protection against early endothelial cell loss. Ablation of the sensitive subpopulation in unprotected mice takes place over a dose range of 1 to 3 Gy {gamma}-rays, but doses between 12 to 20 Gy are required in the presence of ligand. This protection equates to a very high dose modification factor of about 7 and possibly reflects a suppression of apoptosis in the sensitive endothelial subpopulation. The extent to which there is enhanced survival in the endothelial population as a whole and how the observed protection affects late CNS necrosis development has yet to be determined. However present results clearly show potential for the use of DNA-binding radioprotectors with limited penetration for investigations into the relative significance of

  7. Comparison of Endothelial Cell Loss by Specular Microscopy ...

    African Journals Online (AJOL)

    Aim: To compare the endothelial cell loss between phacoemulsification and manual small‑incision cataract surgery (SICS). Endothelial cell loss was also compared in phacoemulsification group by temporal clear corneal incision (CCI) and by superior scleral incision (SI) technique. Materials and Methods: A total of 200 ...

  8. Endothelial cell cultures as a tool in biomaterial research

    NARCIS (Netherlands)

    Kirkpatrick, CJ; Otto, M; van Kooten, T; Krump, [No Value; Kriegsmann, J; Bittinger, F

    1999-01-01

    Progress in biocompatibility and tissue engineering would today be inconceivable without the aid of in vitro techniques. Endothelial cell cultures represent a valuable tool not just in haemocompatibility testing, but also in the concept of designing hybrid organs. In the past endothelial cells (EC)

  9. Endothelial protein C receptor in renal tubular epithelial cells and ...

    African Journals Online (AJOL)

    The endothelial protein C receptor (EPCR) plays an important role within the protein C pathway in regulating coagulation and inflammation. It was reported that EPCR was expressed in large vessels, placenta, heart, liver and lung endothelial cell. However, there are a few studies concerned about renal epithelial cells.

  10. Increased endothelial cell-leukocyte interaction in murine schistosomiasis: possible priming of endothelial cells by the disease.

    Directory of Open Access Journals (Sweden)

    Suellen D S Oliveira

    Full Text Available BACKGROUND AND AIMS: Schistosomiasis is an intravascular parasitic disease associated with inflammation. Endothelial cells control leukocyte transmigration and vascular permeability being modulated by pro-inflammatory mediators. Recent data have shown that endothelial cells primed in vivo in the course of a disease keep the information in culture. Herein, we evaluated the impact of schistosomiasis on endothelial cell-regulated events in vivo and in vitro. METHODOLOGY AND PRINCIPAL FINDINGS: The experimental groups consisted of Schistosoma mansoni-infected and age-matched control mice. In vivo infection caused a marked influx of leukocytes and an increased protein leakage in the peritoneal cavity, characterizing an inflamed vascular and cellular profile. In vitro leukocyte-mesenteric endothelial cell adhesion was higher in cultured cells from infected mice as compared to controls, either in the basal condition or after treatment with the pro-inflammatory cytokine tumor necrosis factor (TNF. Nitric oxide (NO donation reduced leukocyte adhesion to endothelial cells from control and infected groups; however, in the later group the effect was more pronounced, probably due to a reduced NO production. Inhibition of control endothelial NO synthase (eNOS increased leukocyte adhesion to a level similar to the one observed in the infected group. Besides, the adhesion of control leukocytes to endothelial cells from infected animals is similar to the result of infected animals, confirming that schistosomiasis alters endothelial cells function. Furthermore, NO production as well as the expression of eNOS were reduced in cultured endothelial cells from infected animals. On the other hand, the expression of its repressor protein, namely caveolin-1, was similar in both control and infected groups. CONCLUSION/SIGNIFICANCE: Schistosomiasis increases vascular permeability and endothelial cell-leukocyte interaction in vivo and in vitro. These effects are partially

  11. Obstructive sleep apnea and endothelial progenitor cells

    Directory of Open Access Journals (Sweden)

    Wang Q

    2013-10-01

    Full Text Available Qing Wang,1,* Qi Wu,2,* Jing Feng,3,4 Xin Sun5 1The Second Respiratory Department of the First People's Hospital of Kunming, Yunnan, People's Republic of China; 2Tianjin Haihe Hospital, Tianjin, People's Republic of China; 3Respiratory Department of Tianjin Medical University General Hospital, Tianjin, People's Republic of China; 4Division of Pulmonary and Critical Care Medicine, Duke University Medical Center, Durham, NC, USA; 5Respiratory Department of Tianjin Haihe Hospital, Tianjin, People's Republic of China *These authors contributed equally to this work Background: Obstructive sleep apnea (OSA occurs in 4% of middle-aged men and 2% of middle-aged women in the general population, and the prevalence is even higher in specific patient groups. OSA is an independent risk factor for a variety of cardiovascular diseases. Endothelial injury could be the pivotal determinant in the development of cardiovascular pathology in OSA. Endothelial damage ultimately represents a dynamic balance between the magnitude of injury and the capacity for repair. Bone marrow–derived endothelial progenitor cells (EPCs within adult peripheral blood present a possible means of vascular maintenance that could home to sites of injury and restore endothelial integrity and normal function. Methods: We summarized pathogenetic mechanisms of OSA and searched for available studies on numbers and functions of EPCs in patients with OSA to explore the potential links between the numbers and functions of EPCs and OSA. In particular, we tried to elucidate the molecular mechanisms of the effects of OSA on EPCs. Conclusion: Intermittent hypoxia cycles and sleep fragmentation are major pathophysiologic characters of OSA. Intermittent hypoxia acts as a trigger of oxidative stress, systemic inflammation, and sympathetic activation. Sleep fragmentation is associated with a burst of sympathetic activation and systemic inflammation. In most studies, a reduction in circulating EPCs has

  12. Protective effects of vascular endothelial growth factor in cultured brain endothelial cells against hypoglycemia.

    Science.gov (United States)

    Zhao, Fei; Deng, Jiangshan; Yu, Xiaoyan; Li, Dawei; Shi, Hong; Zhao, Yuwu

    2015-08-01

    Hypoglycemia is a common and serious problem among patients with type 1 diabetes receiving treatment with insulin. Clinical studies have demonstrated that hypoglycemic edema is involved in the initiation of hypoglycemic brain damage. However, the mechanisms of this edema are poorly understood. Vascular endothelial growth factor (VEGF), a potent regulator of blood vessel function, has been observed an important candidate hormone induced by hypoglycemia to protect neurons by restoring plasma glucose. Whether VEGF has a protective effect against hypoglycemia-induced damage in brain endothelial cells is still unknown. To investigate the effects of hypoglycemia on cerebral microvascular endothelial cells and assess the protective effect of exogenous VEGF on endothelial cells during hypoglycemia, confluent monolayers of the brain endothelial cell line bEnd.3 were treated with normal (5.5 mM glucose), hypoglycemic (0, 0.5, 1 mM glucose) medium or hypoglycemic medium in the presence of VEGF. The results clearly showed that hypoglycemia significantly downregulated the expression of claudin-5 in bEnd.3 cells, without affecting ZO-1 and occludin expression and distribution. Besides, transendothelial permeability significantly increased under hypoglycemic conditions compared to that under control conditions. Moreover, the hypoglycemic medium in presence of VEGF decreased endothelial permeability via the inhibition of claudin-5 degradation and improved hypoglycemia-induced cell toxicity. Furthermore, Glucose transporter-1 (Glut-1) and apoptosis regulator Bcl-2 expression were significantly upregulated. Taken together, hypoglycemia can significantly increase paraendocellular permeability by downregulating claudin-5 expression. We further showed that VEGF protected brain endothelial cells against hypoglycemia by enhancing glucose passage, reducing endothelial cell death, and ameliorating paraendocellular permeability.

  13. BIGH3 protein and macrophages in retinal endothelial cell apoptosis.

    Science.gov (United States)

    Mondragon, Albert A; Betts-Obregon, Brandi S; Moritz, Robert J; Parvathaneni, Kalpana; Navarro, Mary M; Kim, Hong Seok; Lee, Chi Fung; LeBaron, Richard G; Asmis, Reto; Tsin, Andrew T

    2015-01-01

    Diabetes is a pandemic disease with a higher occurrence in minority populations. The molecular mechanism to initiate diabetes-associated retinal angiogenesis remains largely unknown. We propose an inflammatory pathway of diabetic retinopathy in which macrophages in the diabetic eye provide TGFβ to retinal endothelial cells (REC) in the retinal microvasculature. In response to TGFβ, REC synthesize and secrete a pro-apoptotic BIGH3 (TGFβ-Induced Gene Human Clone 3) protein, which acts in an autocrine loop to induce REC apoptosis. Rhesus monkey retinal endothelial cells (RhREC) were treated with dMCM (cell media of macrophages treated with high glucose and LDL) and assayed for apoptosis (TUNEL), BIGH3 mRNA (qPCR), and protein (Western blots) expressions. Cells were also treated with ΤGFβ1 and 2 for BIGH3 mRNA and protein expression. Inhibition assays were carried out using antibodies for TGFβ1 and for BIGH3 to block apoptosis and mRNA expression. BIGH3 in cultured RhREC cells were identified by immunohistochemistry (IHC). Distribution of BIGH3 and macrophages in the diabetic mouse retina was examined with IHC. RhRECs treated with dMCM or TGFβ showed a significant increase in apoptosis and BIGH3 protein expression. Recombinant BIGH3 added to RhREC culture medium led to a dose-dependent increase in apoptosis. Antibodies (Ab) directed against BIGH3 and TGFβ, as well as TGFβ receptor blocker resulted in a significant reduction in apoptosis induced by either dMCM, TGFβ or BIGH3. IHC showed that cultured RhREC constitutively expressed BIGH3. Macrophage and BIGH3 protein were co-localized to the inner retina of the diabetic mouse eye. Our results support a novel inflammatory pathway for diabetic retinopathy. This pathway is initiated by TGFβ released from macrophages, which promotes synthesis and release of BIGH3 protein by REC and REC apoptosis.

  14. Transition of mesenchymal stem/stromal cells to endothelial cells.

    Science.gov (United States)

    Crisan, Mihaela

    2013-08-14

    Mesenchymal stem/stromal cells (MSCs) are heterogeneous. A fraction of these cells constitute multipotent cells that can self-renew and mainly give rise to mesodermal lineage cells such as adipocytes, osteocytes and chondrocytes. The ability of MSCs to differentiate into endothelial cells remains controversial. Isolation and in vitro manipulation of MSCs before clinical application are important steps. High numbers of MSCs are needed, requiring the in vitro expansion of these clinically important cells. To this end, a well-controlled procedure for MSC isolation and maintenance in culture is necessary.

  15. Cell Structure Controls Endothelial Cell Migration under Fluid Shear Stress.

    Science.gov (United States)

    Lin, Xiefan; Helmke, Brian P

    2009-06-01

    Cobblestone-shaped endothelial cells in confluent monolayers undergo triphasic mechanotaxis in response to steady unidirectional shear stress, but cells that are elongated and aligned on micropatterned substrates do not change their migration behavior in response to either perpendicular or parallel flow. Whether mechanotaxis of micropatterned endothelial cell layers is suppressed by elongated cytoskeletal structure or limited availability of adhesion area remains unknown. In this study, cells were examined on wide (100-200 μm) micropatterned lines after onset of shear stress. Cells in center regions of the lines exhibited cobblestone morphology and triphasic mechanotaxis behavior similar to that in unpatterned monolayers, whereas cells along the edges migrated parallel to the line axis regardless of the flow direction. When scratch wounds were created perpendicular to the micropatterned lines, the cells became less elongated before migrating into the denuded area. In sparsely populated lines oriented perpendicular to the flow direction, elongated cells along the upstream edge migrated parallel to the edge for 7 h before migrating parallel to the shear stress direction, even though adhesion area existed in the downstream direction. Thus, cytoskeletal structure and not available adhesion area serves as the dominant factor in determining whether endothelial mechanotaxis occurs in response to shear stress.

  16. Ionizing radiation activates vascular endothelial growth factor-A transcription in human umbilical vein endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Hyounji; Kim, Kwang Seok; Jeong, Jae Hoon; Lim, Young Bin [Radiation Cancer Biology Team, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2016-12-15

    Vascular endothelial growth factor (VEGF) is an essential paracrine factor for developmental and pathological angiogenesis. VEGF also exerts its effects in an autocrine manner in VEGF-producing cells. For instance, autocrine VEGF signaling occurs in tumor cells and contributes to key aspects of tumorigenesis, such as in the function of cancer stem cells and tumor initiation, which are independent of angiogenesis. In addition to tumors cells, non-transformed cells also express VEGF. For example, a VEGF dependent intracellular autocrine mechanism is crucial for the survival of hematopoietic stem cells and hematopoiesis. Stereotactic body radiation therapy (SBRT) is a novel treatment modality for early primary cancer and oligometastatic disease. SBRT delivers high-dose hypofractionated radiation, such as 20-60 Gy, to tumors in a single fraction or 2-5 fractions. As VEGF is a critical regulator of functional integrity and viability of vascular endothelial cells, we examined whether high-dose irradiation alters VEGF signaling by measuring the expression levels of VEGFA transcript. It is generally believed that endothelial cells do not produce VEGF in response to radiation. In present study, however, we provide the first demonstration of transcriptional regulation of VEGFA in human vascular endothelial cells by IR treatment. Irradiation with doses higher than 10 Gy in a single exposure triggers up-regulation of VEGFA transcription within 2 hours in HUVECs, whereas irradiation with 10 Gy does not alter VEGFA levels. Our data have shown that high-dose irradiation triggers immediate transactivation of VEGFA in human vascular endothelial cells.

  17. Dual role of lipoproteins in endothelial cell dysfunction in atherosclerosis.

    Science.gov (United States)

    Stancu, Camelia S; Toma, Laura; Sima, Anca V

    2012-08-01

    The endothelium is a key constituent of the vascular wall, being actively involved in maintaining the structural integrity and proper functioning of blood vessels. Hyperlipidemia, diabetes, hypertension, smoking and aging are important risk factors for the dysfunction of endothelial cells (EC). Circulating lipoproteins (Lp) synthesized and secreted from the intestine or liver have an important role in supplying peripheral tissues with fatty acids from triglyceride rich lipoproteins (TGRLp) for energy production or storage, and cholesterol from low density lipoproteins (LDL) or high density lipoproteins (HDL) for the synthesis of cellular membranes and steroid hormones. Under pathological conditions, Lp may suffer alterations in concentration and composition and become aggressors for EC. Modified LDL, remnant Lp, TGRLp lipolysis products, dysfunctional HDL are involved in the changes induced in EC morphology (reduced glycocalyx, overdeveloped endoplasmic reticulum, Golgi apparatus and basement membrane), loose intercellular junctions, increased oxidative and inflammatory stress, nitric oxide/redox imbalance, excess Lp transport and storage, as well as loss of anti-thrombotic properties, all of these being characteristics of endothelial dysfunction. Normal HDL are able to counteract the harmful effects of atherogenic Lp in EC but under persistent pathological conditions they lose the protective properties and become pro-atherogenic. This review summarises recent advances in understanding the role of Lp in the induction of endothelial dysfunction and the initiation and progression of atherosclerotic lesions. Its main focus is the antagonistic role of atherogenic Lp (LDL, VLDL, dysfunctional HDL) versus anti-atherogenic Lp (HDL), also pointing out the potential targets for arresting or reversing this process.

  18. Viability and proliferation of endothelial cells upon exposure to GaN nanoparticles

    Directory of Open Access Journals (Sweden)

    Tudor Braniste

    2016-09-01

    Full Text Available Nanotechnology is a rapidly growing and promising field of interest in medicine; however, nanoparticle–cell interactions are not yet fully understood. The goal of this work was to examine the interaction between endothelial cells and gallium nitride (GaN semiconductor nanoparticles. Cellular viability, adhesion, proliferation, and uptake of nanoparticles by endothelial cells were investigated. The effect of free GaN nanoparticles versus the effect of growing endothelial cells on GaN functionalized surfaces was examined. To functionalize surfaces with GaN, GaN nanoparticles were synthesized on a sacrificial layer of zinc oxide (ZnO nanoparticles using hydride vapor phase epitaxy. The uptake of GaN nanoparticles by porcine endothelial cells was strongly dependent upon whether they were fixed to the substrate surface or free floating in the medium. The endothelial cells grown on surfaces functionalized with GaN nanoparticles demonstrated excellent adhesion and proliferation, suggesting good biocompatibility of the nanostructured GaN.

  19. Human Endothelial Cell Models in Biomaterial Research.

    Science.gov (United States)

    Hauser, Sandra; Jung, Friedrich; Pietzsch, Jens

    2017-03-01

    Endothelial cell (EC) models have evolved as important tools in biomaterial research due to ubiquitously occurring interactions between implanted materials and the endothelium. However, screening the available literature has revealed a gap between material scientists and physiologists in terms of their understanding of these biomaterial-endothelium interactions and their relative importance. Consequently, EC models are often applied in nonphysiological experimental setups, or too extensive conclusions are drawn from their results. The question arises whether this might be one reason why, among the many potential biomaterials, only a few have found their way into the clinic. In this review, we provide an overview of established EC models and possible selection criteria to enable researchers to determine the most reliable and relevant EC model to use. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Tumor-Endothelium Cross Talk Blocks Recruitment of Neutrophils to Endothelial Cells: A Novel Mechanism of Endothelial Cell Anergy1

    OpenAIRE

    Blaheta, Roman A; Powerski, Maciej; Hudak, Lukasz; Juengel, Eva; Jonas, Dietger; von Knethen, Andreas; Doerr, Hans Willhelm; Cinatl, Jindrich

    2009-01-01

    Tumor cells have evolved effective strategies to escape the host immune response. The objective of this study was to determine whether tumor cells can condition endothelial cells in a specific manner to prevent subsequent adhesion of polymorphonuclear neutrophils (PMNs) and/or peripheral blood lymphocytes (PBLs). Human umbilical vein endothelial cells (HUVECs) and UKF-NB-4 neuroblastoma tumor cells were established in coculture on opposite sides of porous transwell filters. After 24 hours wit...

  1. Tumor-Endothelium Cross Talk Blocks Recruitment of Neutrophils to Endothelial Cells: A Novel Mechanism of Endothelial Cell Anergy

    OpenAIRE

    Blaheta, Roman A.; Powerski, Maciej; Hudak, Lukasz; Juengel, Eva; Jonas, Dietger; von Knethen, Andreas; Doerr, Hans Willhelm; Cinatl Jr, Jindrich

    2009-01-01

    Tumor cells have evolved effective strategies to escape the host immune response. The objective of this study was to determine whether tumor cells can condition endothelial cells in a specific manner to prevent subsequent adhesion of polymorphonuclear neutrophils (PMNs) and/or peripheral blood lymphocytes (PBLs). Human umbilical vein endothelial cells (HUVECs) and UKF-NB-4 neuroblastoma tumor cells were established in coculture on opposite sides of porous transwell filters. After 24 hours wit...

  2. Radiation-induced apoptosis in microvascular endothelial cells.

    OpenAIRE

    Langley, R. E.; Bump, E A; Quartuccio, S. G.; Medeiros, D; Braunhut, S. J.

    1997-01-01

    The response of the microvasculature to ionizing radiation is thought to be an important factor in the overall response of both normal tissues and tumours. It has recently been reported that basic fibroblast growth factor (bFGF), a potent mitogen for endothelial cells, protects large vessel endothelial cells from radiation-induced apoptosis in vitro. Microvessel cells are phenotypically distinct from large vessel cells. We studied the apoptotic response of confluent monolayers of capillary en...

  3. Autocrine VEGF isoforms differentially regulate endothelial cell behavior

    Directory of Open Access Journals (Sweden)

    Hideki Yamamoto

    2016-09-01

    Full Text Available Vascular endothelial growth factor A (VEGF is involved in all the essential biology of endothelial cells, from proliferation to vessel function, by mediating intercellular interactions and monolayer integrity. It is expressed as three major alternative spliced variants. In mice, these are VEGF120, VEGF164, and VEGF188, each with different affinities for extracellular matrices and cell surfaces, depending on the inclusion of heparin-binding sites, encoded by exons 6 and 7. To determine the role of each VEGF isoform in endothelial homeostasis, we compared phenotypes of primary endothelial cells isolated from lungs of mice expressing single VEGF isoforms in normoxic and hypoxic conditions. The differential expression and distribution of VEGF isoforms affect endothelial cell functions, such as proliferation, adhesion, migration and integrity, which are dependent on the stability of and affinity to VEGF receptor 2 (VEGFR2. We found a correlation between autocrine VEGF164 and VEGFR2 stability, which is also associated with increased expression of proteins involved in cell adhesion. Endothelial cells expressing only VEGF188, which localizes to extracellular matrices or cell surfaces, presented a mesenchymal morphology and weakened monolayer integrity. Cells expressing only VEGF120 lacked stable VEGFR2 and dysfunctional downstream processes, rendering the cells unviable. Endothelial cells expressing these different isoforms in isolation also had differing rates of apoptosis, proliferation, and signaling via nitric oxide (NO synthesis. These data indicate that autocrine signaling of each VEGF isoform has unique functions on endothelial homeostasis and response to hypoxia, due to both distinct VEGF distribution and VEGFR2 stability, which appears to be, at least partly, affected by differential NO production. This study demonstrates that each autocrine VEGF isoform has a distinct effect on downstream functions, namely VEGFR2-regulated endothelial cell

  4. Radioprotection of mouse CNS endothelial cells in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Lyubimova, N.; Coultas, P.; Martin, R. [Peter MacCallum Cancer institute, Melbourne, Victoria (Australia)

    1997-03-01

    After treatments with monoamine oxidase inhibitors and L-DOPA, the blood brain barrier causes a build-up of dopamine in brain capillary endothelial cells. Conversion of the dopamine to a fluorophore provides a marker which can be used to measure endothelial cell density by fluorescence microscopy. Earlier studies used this technique to monitor post-irradiation changes in endothelial cell density in rodent brain and showed loss within 24 hours of a sub-population of about 15 % of the endothelial cells. As a first step, rather than use the later endpoints of radionecrosis it was decided to examine directly whether Hoechst 33342 could protect against this rapid initial endothelial cell loss. Ten minutes after intravenous injection of Joechst 33342, in mouse brain the ligand was confined to endothelial cells and, for irradiation at this time, there was protection against endothelial cell loss over the first 24 hours after after exposure. Ablation of the sensitive subpopulation in unprotected mice took place over a dose range of 1 to 3 Gy {gamma}-rays but doses between 12 to 20 Gy were required in the presence of the ligand. This protection equated to a high dose modification factor of approximately 7 and may reflect suppression of apoptosis in this sensitive endothelial subpopulation. The extent to which there is enhanced survival in the endothelial population as a whole, and how the observed protection affects late CNS necrosis development, has yet to be determined. However these results suggest a potential use of DNA-binding radioprotectors with limited penetration in investigations of the relative significance of endothelial and parenchymal damage in normal tissue responses to ionising radiation. (authors)

  5. SOCS1 prevents graft arteriosclerosis by preserving endothelial cell function.

    Science.gov (United States)

    Qin, Lingfeng; Huang, Qunhua; Zhang, Haifeng; Liu, Renjing; Tellides, George; Min, Wang; Yu, Luyang

    The aim of this study was to determine the role of suppressor of cytokine signaling 1 (SOCS1) in graft arteriosclerosis (GA). GA, the major cause of late cardiac allograft failure, is initiated by immune-mediated endothelial activation resulting in vascular inflammation and consequent neointima formation. SOCS1, a negative regulator of cytokine signaling, is highly expressed in endothelial cells (ECs) and may prevent endothelial inflammatory responses and phenotypic activation. Clinical specimens of coronary arteries with GA, with atherosclerosis, or without disease were collected for histological analysis. SOCS1 knockout or vascular endothelial SOCS1 (VESOCS1) transgenic mice were used in an aorta transplant model of GA. Mouse aortic ECs were isolated for in vitro assays. Dramatic but specific reduction of endothelial SOCS1 was observed in human GA and atherosclerosis specimens, which suggested the importance of SOCS1 in maintaining normal endothelial function. SOCS1 deletion in mice resulted in basal EC dysfunction. After transplantation, SOCS1-deficient aortic grafts augmented leukocyte recruitment and neointima formation, whereas endothelial overexpression of SOCS1 diminished arterial rejection. Induction of endothelial adhesion molecules in early stages of GA was suppressed by the VESOCS1 transgene, and this effect was confirmed in cultured aortic ECs. Moreover, VESOCS1 maintained better vascular function during GA progression. Mechanistically, endothelial SOCS1, by modulating both basal and cytokine-induced expression of the adhesion molecules platelet/endothelial cell adhesion molecule-1, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1, restrained leukocyte adhesion and transendothelial migration during inflammatory cell infiltration. SOCS1 prevents GA progression by preserving endothelial function and attenuating cytokine-induced adhesion molecule expression in vascular endothelium. Copyright © 2014 American College of Cardiology

  6. Selective expression of erg isoforms in human endothelial cells.

    Science.gov (United States)

    Hewett, P W; Nishi, K; Daft, E L; Clifford Murray, J

    2001-04-01

    Erg and Fli-1 are closely related members of the ets family of transcription factors. There are at least five human Erg isoforms (Erg-1, Erg-2, Erg-3/p55(Erg), p49(Erg) and p38(Erg)) produced through differential mRNA splicing and alternative use of translational start codons. However, relatively little is known about the expression or function of these isoforms in vitro or their distribution in vivo. We used RT-PCR to screen a panel of primary and established human cell lines for erg and fli-1 consensus sequences. Whilst fli-1 was expressed in several human cell types, erg was detected mainly in endothelial cells. To identify which erg isoforms are expressed in endothelial cells we used RT-PCR, Northern blotting and 5'-RACE. Erg-3/p55(Erg) and p38(Erg)/p38(Erg)-like transcripts were detected in both microvascular and large vessel endothelial cells affinity-purified from different vascular beds. Moreover, these erg isoforms were present in both freshly isolated, and confluent endothelial cells following several passages in culture, indicating that endothelial erg expression in vitro may be broadly representative of that in vivo. The selective expression of the Erg-3/p55(Erg) and p38(Erg)/p38(Erg)-like isoforms in endothelial cells indicates their involvement in the regulation of endothelial-restricted genes.

  7. Using cultured endothelial cells to study endothelial barrier dysfunction: Challenges and opportunities

    OpenAIRE

    Aman, Jurjan; Weijers, Ester M.; Geerten P van Nieuw Amerongen; Malik, Asrar B.; van Hinsbergh, Victor W.M.

    2016-01-01

    Despite considerable progress in the understanding of endothelial barrier regulation and the identification of approaches that have the potential to improve endothelial barrier function, no drug- or stem cell-based therapy is presently available to reverse the widespread vascular leak that is observed in acute respiratory distress syndrome (ARDS) and sepsis. The translational gap suggests a need to develop experimental approaches and tools that better mimic the complex environment of the micr...

  8. Animal study on transplantation of human umbilical vein endothelial cells for corneal endothelial decompensation

    Directory of Open Access Journals (Sweden)

    Li Cui

    2014-06-01

    Full Text Available AIM: To explore the feasibility of culturing human umbilical vein endothelial cells(HUVECon acellular corneal stroma and performing the posterior lamellar endothelial keratoplasty(PLEKtreating corneal endothelial decompensation.METHODS: Thirty New-Zealand rabbits were divided into three groups randomly, 10 rabbits for experimental group, 10 for stroma group and 10 for control group. Corneal endothelial cells were removed to establish animal model of corneal endothelial failure. PLEK was performed on the rabbits of experimental group and stroma group, and nothing was transplantated onto the rabbits of control group with the deep layer excised only. Postoperative observation was taken for 3mo. The degree of corneal edema and central corneal thickness were recorded for statistical analysis.RESULTS: Corneas in experimental group were relieved in edema obviously compared with that in stroma group and the control group, and showed increased transparency 7d after the operation. The average density of endothelial cells was 2 026.4±129.3cells/mm2, and average central corneal thickness was 505.2±25.4μm in experimental group, while 1 535.6±114.5μm in stroma group and 1 493.5±70.2μm in control group 3mo after operation.CONCLUSION:We achieved preliminary success in our study that culturing HUVEC on acellular corneal stroma and performing PLEK for corneal endothelial decompensation. HUVEC transplanted could survive in vivo, and have normal biological function of keeping cornea transparent. This study provides a new idea and a new way clinically for the treatment of corneal endothelial diseases.

  9. Mechanical cell-substrate feedback explains pairwise and collective endothelial cell behavior in vitro

    NARCIS (Netherlands)

    R.F.M. van Oers (Rene); R.M.H. Merks (Roeland)

    2013-01-01

    htmlabstractIn vitro cultures of endothelial cells are a widely used model system of the collective behavior of endothelial cells during vasculogenesis and angiogenesis. When seeded in a extracellular matrix, endothelial cells can form blood vessel-like structures, including vascular networks and

  10. Chemosensitizing AML cells by targeting bone marrow endothelial cells.

    Science.gov (United States)

    Bosse, Raphael C; Wasserstrom, Briana; Meacham, Amy; Wise, Elizabeth; Drusbosky, Leylah; Walter, Glenn A; Chaplin, David J; Siemann, Dietmar W; Purich, Daniel L; Cogle, Christopher R

    2016-05-01

    Refractory disease is the greatest challenge in treating patients with acute myeloid leukemia (AML). Blood vessels may serve as sanctuary sites for AML. When AML cells were co-cultured with bone marrow endothelial cells (BMECs), a greater proportion of leukemia cells were in G0/G1. This led us to a strategy of targeting BMECs with tubulin-binding combretastatins, causing BMECs to lose their flat phenotype, degrade their cytoskeleton, cease growth, and impair migration despite unchanged BMEC viability and metabolism. Combretastatins also caused downregulation of BMEC adhesion molecules known to tether AML cells, including vascular cell adhesion molecule (VCAM)-1 and vascular endothelial (VE)-cadherin. When AML-BMEC co-cultures were treated with combretastatins, a significantly greater proportion of AML cells dislodged from BMECs and entered the G2/M cell cycle, suggesting enhanced susceptibility to cell cycle agents. Indeed, the combination of combretastatins and cytotoxic chemotherapy enhanced additive AML cell death. In vivo mice xenograft studies confirmed this finding by revealing complete AML regression after treatment with combretastatins and cytotoxic chemotherapy. Beyond highlighting the pathologic role of BMECs in the leukemia microenvironment as a protective reservoir of disease, these results support a new strategy for using vascular-targeting combretastatins in combination with cytotoxic chemotherapy to treat AML. Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

  11. Biophysical Cueing and Vascular Endothelial Cell Behavior

    Directory of Open Access Journals (Sweden)

    Joshua A. Wood

    2010-03-01

    Full Text Available Human vascular endothelial cells (VEC line the vessels of the body and are critical for the maintenance of vessel integrity and trafficking of biochemical cues. They are fundamental structural elements and are central to the signaling environment. Alterations in the normal functioning of the VEC population are associated with a number of vascular disorders among which are some of the leading causes of death in both the United States and abroad. VECs attach to their underlying stromal elements through a specialization of the extracellular matrix, the basement membrane. The basement membrane provides signaling cues to the VEC through its chemical constituents, by serving as a reservoir for cytoactive factors and through its intrinsic biophysical properties. This specialized matrix is composed of a topographically rich 3D felt-like network of fibers and pores on the nano (1–100 nm and submicron (100–1,000 nm size scale. The basement membrane provides biophysical cues to the overlying VECs through its intrinsic topography as well as through its local compliance (relative stiffness. These biophysical cues modulate VEC adhesion, migration, proliferation, differentiation, and the cytoskeletal signaling network of the individual cells. This review focuses on the impact of biophysical cues on VEC behaviors and demonstrates the need for their consideration in future vascular studies and the design of improved prosthetics.

  12. Activated Brain Endothelial Cells Cross-Present Malaria Antigen.

    Science.gov (United States)

    Howland, Shanshan W; Poh, Chek Meng; Rénia, Laurent

    2015-06-01

    In the murine model of cerebral malaria caused by P. berghei ANKA (PbA), parasite-specific CD8+ T cells directly induce pathology and have long been hypothesized to kill brain endothelial cells that have internalized PbA antigen. We previously reported that brain microvessel fragments from infected mice cross-present PbA epitopes, using reporter cells transduced with epitope-specific T cell receptors. Here, we confirm that endothelial cells are the population responsible for cross-presentation in vivo, not pericytes or microglia. PbA antigen cross-presentation by primary brain endothelial cells in vitro confers susceptibility to killing by CD8+ T cells from infected mice. IFNγ stimulation is required for brain endothelial cross-presentation in vivo and in vitro, which occurs by a proteasome- and TAP-dependent mechanism. Parasite strains that do not induce cerebral malaria were phagocytosed and cross-presented less efficiently than PbA in vitro. The main source of antigen appears to be free merozoites, which were avidly phagocytosed. A human brain endothelial cell line also phagocytosed P. falciparum merozoites. Besides being the first demonstration of cross-presentation by brain endothelial cells, our results suggest that interfering with merozoite phagocytosis or antigen processing may be effective strategies for cerebral malaria intervention.

  13. Activated Brain Endothelial Cells Cross-Present Malaria Antigen.

    Directory of Open Access Journals (Sweden)

    Shanshan W Howland

    2015-06-01

    Full Text Available In the murine model of cerebral malaria caused by P. berghei ANKA (PbA, parasite-specific CD8+ T cells directly induce pathology and have long been hypothesized to kill brain endothelial cells that have internalized PbA antigen. We previously reported that brain microvessel fragments from infected mice cross-present PbA epitopes, using reporter cells transduced with epitope-specific T cell receptors. Here, we confirm that endothelial cells are the population responsible for cross-presentation in vivo, not pericytes or microglia. PbA antigen cross-presentation by primary brain endothelial cells in vitro confers susceptibility to killing by CD8+ T cells from infected mice. IFNγ stimulation is required for brain endothelial cross-presentation in vivo and in vitro, which occurs by a proteasome- and TAP-dependent mechanism. Parasite strains that do not induce cerebral malaria were phagocytosed and cross-presented less efficiently than PbA in vitro. The main source of antigen appears to be free merozoites, which were avidly phagocytosed. A human brain endothelial cell line also phagocytosed P. falciparum merozoites. Besides being the first demonstration of cross-presentation by brain endothelial cells, our results suggest that interfering with merozoite phagocytosis or antigen processing may be effective strategies for cerebral malaria intervention.

  14. Activated Brain Endothelial Cells Cross-Present Malaria Antigen

    Science.gov (United States)

    Howland, Shanshan W.; Poh, Chek Meng; Rénia, Laurent

    2015-01-01

    In the murine model of cerebral malaria caused by P. berghei ANKA (PbA), parasite-specific CD8+ T cells directly induce pathology and have long been hypothesized to kill brain endothelial cells that have internalized PbA antigen. We previously reported that brain microvessel fragments from infected mice cross-present PbA epitopes, using reporter cells transduced with epitope-specific T cell receptors. Here, we confirm that endothelial cells are the population responsible for cross-presentation in vivo, not pericytes or microglia. PbA antigen cross-presentation by primary brain endothelial cells in vitro confers susceptibility to killing by CD8+ T cells from infected mice. IFNγ stimulation is required for brain endothelial cross-presentation in vivo and in vitro, which occurs by a proteasome- and TAP-dependent mechanism. Parasite strains that do not induce cerebral malaria were phagocytosed and cross-presented less efficiently than PbA in vitro. The main source of antigen appears to be free merozoites, which were avidly phagocytosed. A human brain endothelial cell line also phagocytosed P. falciparum merozoites. Besides being the first demonstration of cross-presentation by brain endothelial cells, our results suggest that interfering with merozoite phagocytosis or antigen processing may be effective strategies for cerebral malaria intervention. PMID:26046849

  15. Endothelial cells are intrinsically defective in xenophagy of Streptococcus pyogenes.

    Directory of Open Access Journals (Sweden)

    Shiou-Ling Lu

    2017-07-01

    Full Text Available Group A Streptococcus (GAS is deleterious pathogenic bacteria whose interaction with blood vessels leads to life-threatening bacteremia. Although xenophagy, a special form of autophagy, eliminates invading GAS in epithelial cells, we found that GAS could survive and multiply in endothelial cells. Endothelial cells were competent in starvation-induced autophagy, but failed to form double-membrane structures surrounding GAS, an essential step in xenophagy. This deficiency stemmed from reduced recruitment of ubiquitin and several core autophagy proteins in endothelial cells, as demonstrated by the fact that it could be rescued by exogenous coating of GAS with ubiquitin. The defect was associated with reduced NO-mediated ubiquitin signaling. Therefore, we propose that the lack of efficient clearance of GAS in endothelial cells is caused by their intrinsic inability to target GAS with ubiquitin to promote autophagosome biogenesis for xenophagy.

  16. Identification of laminin α5 short arm peptides active for endothelial cell attachment and tube formation.

    Science.gov (United States)

    Kikkawa, Yamato; Sugawara, Yumika; Harashima, Nozomi; Fujii, Shogo; Ikari, Kazuki; Kumai, Jun; Katagiri, Fumihiko; Hozumi, Kentaro; Nomizu, Motoyoshi

    2017-07-01

    Laminin-511, a major component of endothelial basement membrane, consists of α5, β1, and γ1 chains. The short arm region of the α5 chain is a structural feature of endothelial laminins. In this study, we identified active sequences for human umbilical vein endothelial cells (HUVECs) using recombinant proteins and synthetic peptides. The short arm of the α5 chain contains three globular domains [laminin N-terminal globular domain, laminin 4 domain a, and laminin 4 domain b (LN, L4a, and L4b)] and three rod-like elements [laminin epidermal growth factor-like domain a, b, and c (LEa, LEb, and LEc)]. The cell attachment assay using recombinant proteins showed that RGD-independent cell attachment sites were localized in the α5LN-LEa domain. Further, we synthesized 70 peptides covering the amino acid sequences of the α5LN-LEa domain. Of the 70 peptides, A5-16 (mouse laminin α5 230-243: LENGEIVVSLVNGR) potently exhibited endothelial cell attachment activity. An active sequence analysis using N-terminally and C-terminally truncated A5-16 peptides showed that the nine-amino acid sequence IVVSLVNGR was critical for the endothelial cell attachment activity. Cell adhesion to the peptides was dependent on both cations and heparan sulfate. Further, the A5-16 peptide inhibited the capillary-like tube formation of HUVECs with the cells forming small clumps with short tubes. The eight-amino acid sequence EIVVSLVN in the A5-16 peptide was critical to inhibit HUVEC tube formation. This amino acid sequence could be useful for grafts and thus modulate endothelial cell behavior for vascular surgery. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.

  17. Variety of corneal endothelial cell in glaucoma by confocal microscope

    Directory of Open Access Journals (Sweden)

    Hong-Liang Gao

    2014-10-01

    Full Text Available ATM: To define the causes of corneal endothelial cell damage, to investigate the preventive methods, and to observe the variety of corneal endothelial cell in glaucoma using confocal microscope.METHODS: Totally, 143 eyes of 97 patients with different types of glaucoma, and matched normal people were 20 cases, all 40 eyes. The cell density, cell area and cell variable coefficient were measured used confocal microscope. These indicatives of every kind of glaucoma were compared.RESULTS: The corneal endothelial cell density of normal group was 2 893.88±255.026/mm2, the group of acute angle-closure glaucoma(AACGwas 1 674.11±683.95/mm2, and the group of open angle glaucoma(OAGwas 2 687.22±391.87/mm2, the group of chronic angle-closure glaucoma(CACGwas 2 706.97±351.27/mm2. In all index the average cell density of corneal endothelial and the average area have statistical significance(F=62.950, 8.795; P=0.000, especially the group of AACG. CONCLUSION: The index of corneal endothelial cell in AACG is lower than that of normal. All index in OAG and CACG is difference with that of normal, but the difference has no statistical significance. And the dominant factor of damaged corneal endothelial is the time of intraocular hypertension.

  18. Activation of Endothelial Nitric Oxide (eNOS Occurs through Different Membrane Domains in Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Jason Tran

    Full Text Available Endothelial cells respond to a large range of stimuli including circulating lipoproteins, growth factors and changes in haemodynamic mechanical forces to regulate the activity of endothelial nitric oxide synthase (eNOS and maintain blood pressure. While many signalling pathways have been mapped, the identities of membrane domains through which these signals are transmitted are less well characterized. Here, we manipulated bovine aortic endothelial cells (BAEC with cholesterol and the oxysterol 7-ketocholesterol (7KC. Using a range of microscopy techniques including confocal, 2-photon, super-resolution and electron microscopy, we found that sterol enrichment had differential effects on eNOS and caveolin-1 (Cav1 colocalisation, membrane order of the plasma membrane, caveolae numbers and Cav1 clustering. We found a correlation between cholesterol-induced condensation of the plasma membrane and enhanced high density lipoprotein (HDL-induced eNOS activity and phosphorylation suggesting that cholesterol domains, but not individual caveolae, mediate HDL stimulation of eNOS. Vascular endothelial growth factor (VEGF-induced and shear stress-induced eNOS activity was relatively independent of membrane order and may be predominantly controlled by the number of caveolae on the cell surface. Taken together, our data suggest that signals that activate and phosphorylate eNOS are transmitted through distinct membrane domains in endothelial cells.

  19. Activation of Endothelial Nitric Oxide (eNOS) Occurs through Different Membrane Domains in Endothelial Cells.

    Science.gov (United States)

    Tran, Jason; Magenau, Astrid; Rodriguez, Macarena; Rentero, Carles; Royo, Teresa; Enrich, Carlos; Thomas, Shane R; Grewal, Thomas; Gaus, Katharina

    2016-01-01

    Endothelial cells respond to a large range of stimuli including circulating lipoproteins, growth factors and changes in haemodynamic mechanical forces to regulate the activity of endothelial nitric oxide synthase (eNOS) and maintain blood pressure. While many signalling pathways have been mapped, the identities of membrane domains through which these signals are transmitted are less well characterized. Here, we manipulated bovine aortic endothelial cells (BAEC) with cholesterol and the oxysterol 7-ketocholesterol (7KC). Using a range of microscopy techniques including confocal, 2-photon, super-resolution and electron microscopy, we found that sterol enrichment had differential effects on eNOS and caveolin-1 (Cav1) colocalisation, membrane order of the plasma membrane, caveolae numbers and Cav1 clustering. We found a correlation between cholesterol-induced condensation of the plasma membrane and enhanced high density lipoprotein (HDL)-induced eNOS activity and phosphorylation suggesting that cholesterol domains, but not individual caveolae, mediate HDL stimulation of eNOS. Vascular endothelial growth factor (VEGF)-induced and shear stress-induced eNOS activity was relatively independent of membrane order and may be predominantly controlled by the number of caveolae on the cell surface. Taken together, our data suggest that signals that activate and phosphorylate eNOS are transmitted through distinct membrane domains in endothelial cells.

  20. Magnetizable stent-grafts enable endothelial cell capture

    Energy Technology Data Exchange (ETDEWEB)

    Tefft, Brandon J. [Department of Cardiovascular Diseases, Mayo Clinic, Rochester, MN (United States); Uthamaraj, Susheil [Division of Engineering, Mayo Clinic, Rochester, MN (United States); Harburn, J. Jonathan [School of Medicine, Pharmacy and Health, Durham University, Stockton-on-Tees (United Kingdom); Hlinomaz, Ota [Department of Cardioangiology, St. Anne' s University Hospital, Brno (Czech Republic); Lerman, Amir [Department of Cardiovascular Diseases, Mayo Clinic, Rochester, MN (United States); Dragomir-Daescu, Dan [Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, MN (United States); Sandhu, Gurpreet S., E-mail: sandhu.gurpreet@mayo.edu [Department of Cardiovascular Diseases, Mayo Clinic, Rochester, MN (United States)

    2017-04-01

    Emerging nanotechnologies have enabled the use of magnetic forces to guide the movement of magnetically-labeled cells, drugs, and other therapeutic agents. Endothelial cells labeled with superparamagnetic iron oxide nanoparticles (SPION) have previously been captured on the surface of magnetizable 2205 duplex stainless steel stents in a porcine coronary implantation model. Recently, we have coated these stents with electrospun polyurethane nanofibers to fabricate prototype stent-grafts. Facilitated endothelialization may help improve the healing of arteries treated with stent-grafts, reduce the risk of thrombosis and restenosis, and enable small-caliber applications. When placed in a SPION-labeled endothelial cell suspension in the presence of an external magnetic field, magnetized stent-grafts successfully captured cells to the surface regions adjacent to the stent struts. Implantation within the coronary circulation of pigs (n=13) followed immediately by SPION-labeled autologous endothelial cell delivery resulted in widely patent devices with a thin, uniform neointima and no signs of thrombosis or inflammation at 7 days. Furthermore, the magnetized stent-grafts successfully captured and retained SPION-labeled endothelial cells to select regions adjacent to stent struts and between stent struts, whereas the non-magnetized control stent-grafts did not. Early results with these prototype devices are encouraging and further refinements will be necessary in order to achieve more uniform cell capture and complete endothelialization. Once optimized, this approach may lead to more rapid and complete healing of vascular stent-grafts with a concomitant improvement in long-term device performance. - Highlights: • Magnetic stent-grafts were made from 2205 steel stents and polyurethane nanofibers. • Stent-grafts remained patent and formed a thin and uniform neointima when implanted. • Stent-grafts captured endothelial cells labeled with magnetic nanoparticles.

  1. Vascular Mechanobiology Endothelial Cell Responses to Fluid Shear Stress

    National Research Council Canada - National Science Library

    Ando, Joji; Yamamoto, Kimiko

    2009-01-01

    Endothelial cells (ECs) lining blood vessel walls respond to shear stress, a fluid mechanical force generated by flowing blood, and the EC responses play an important role in the homeostasis of the circulatory system...

  2. Endothelial cell proliferation in swine experimental aneurysm after coil embolization.

    Directory of Open Access Journals (Sweden)

    Yumiko Mitome-Mishima

    Full Text Available After coil embolization, recanalization in cerebral aneurysms adversely influences long-term prognosis. Proliferation of endothelial cells on the coil surface may reduce the incidence of recanalization and further improve outcomes after coil embolization. We aimed to map the expression of proliferating tissue over the aneurysmal orifice and define the temporal profile of tissue growth in a swine experimental aneurysm model. We compared the outcomes after spontaneous thrombosis with those of coil embolization using histological and morphological techniques. In aneurysms that we not coiled, spontaneous thrombosis was observed, and weak, easily detachable proliferating tissue was evident in the aneurysmal neck. In contrast, in the coil embolization group, histological analysis showed endothelial-like cells lining the aneurysmal opening. Moreover, immunohistochemical and morphological analysis suggested that these cells were immature endothelial cells. Our results indicated the existence of endothelial cell proliferation 1 week after coil embolization and showed immature endothelial cells in septal tissue between the systemic circulation and the aneurysm. These findings suggest that endothelial cells are lead to and proliferate in the former aneurysmal orifice. This is the first examination to evaluate the temporal change of proliferating tissue in a swine experimental aneurysm model.

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

    KAUST Repository

    Cima, Igor

    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.

  4. Lonidamine Causes Inhibition of Angiogenesis-Related Endothelial Cell Functions

    Directory of Open Access Journals (Sweden)

    Donatella Del Bufalo

    2004-09-01

    Full Text Available The aim of this study was to assess whether lonidamine (LND interferes with some steps in angiogenesis progression. We report here, for the first time, that LND inhibited angiogenic-related endothelial cell functions in a dose-dependent manner (1-50 μg/ml. In particular, LND decreased proliferation, migration, invasion, and morphogenesis on matrigel of different endothelial cell lines. Zymographic and Western blot analysis assays showed that LND treatment produced a reduction in the secretion of matrix metalloproteinase-2 and metalloproteinase-9 by endothelial cells. Vessel formation in a matrigel plug was also reduced by LND. The viability, migration, invasion, and matrix metalloproteinase production of different tumor cell lines were not affected by low doses of LND (1-10 μg/ml, whereas 50 μg/ml LND, which corresponds to the dose used in clinical management of tumors, triggered apoptosis both in endothelial and tumor cells. Together, these data demonstrate that LND is a compound that interferes with endothelial cell functions, both at low and high doses. Thus, the effect of LND on endothelial cell functions, previously undescribed, may be a significant contributor to the antitumor effect of LND observed for clinical management of solid tumors.

  5. Paradoxic effects of metformin on endothelial cells and angiogenesis

    Science.gov (United States)

    Bruno, Antonino; Cantelmo, Anna R.; Albini, Adriana

    2014-01-01

    The biguanide metformin is used in type 2 diabetes management and has gained significant attention as a potential cancer preventive agent. Angioprevention represents a mechanism of chemoprevention, yet conflicting data concerning the antiangiogenic action of metformin have emerged. Here, we clarify some of the contradictory effects of metformin on endothelial cells and angiogenesis, using in vitro and in vivo assays combined with transcriptomic and protein array approaches. Metformin inhibits formation of capillary-like networks by endothelial cells; this effect is partially dependent on the energy sensor adenosine-monophosphate-activated protein kinase (AMPK) as shown by small interfering RNA knockdown. Gene expression profiling of human umbilical vein endothelial cells revealed a paradoxical modulation of several angiogenesis-associated genes and proteins by metformin, with short-term induction of vascular endothelial growth factor (VEGF), cyclooxygenase 2 and CXC chemokine receptor 4 at the messenger RNA level and downregulation of ADAMTS1. Antibody array analysis shows an essentially opposite regulation of numerous angiogenesis-associated proteins in endothelial and breast cancer cells including interleukin-8, angiogenin and TIMP-1, as well as selective regulation of angiopioetin-1, -2, endoglin and others. Endothelial cell production of the cytochrome P450 member CYP1B1 is upregulated by tumor cell supernatants in an AMPK-dependent manner, metformin blocks this effect. Metformin inhibits VEGF-dependent activation of extracellular signal-regulated kinase 1/2, and the inhibition of AMPK activity abrogates this event. Metformin hinders angiogenesis in matrigel pellets in vivo, prevents the microvessel density increase observed in obese mice on a high-fat diet, downregulating the number of white adipose tissue endothelial precursor cells. Our data show that metformin has an antiangiogenic activity in vitro and in vivo associated with a contradictory short

  6. Paradoxic effects of metformin on endothelial cells and angiogenesis.

    Science.gov (United States)

    Dallaglio, Katiuscia; Bruno, Antonino; Cantelmo, Anna R; Esposito, Alessia I; Ruggiero, Luca; Orecchioni, Stefania; Calleri, Angelica; Bertolini, Francesco; Pfeffer, Ulrich; Noonan, Douglas M; Albini, Adriana

    2014-05-01

    The biguanide metformin is used in type 2 diabetes management and has gained significant attention as a potential cancer preventive agent. Angioprevention represents a mechanism of chemoprevention, yet conflicting data concerning the antiangiogenic action of metformin have emerged. Here, we clarify some of the contradictory effects of metformin on endothelial cells and angiogenesis, using in vitro and in vivo assays combined with transcriptomic and protein array approaches. Metformin inhibits formation of capillary-like networks by endothelial cells; this effect is partially dependent on the energy sensor adenosine-monophosphate-activated protein kinase (AMPK) as shown by small interfering RNA knockdown. Gene expression profiling of human umbilical vein endothelial cells revealed a paradoxical modulation of several angiogenesis-associated genes and proteins by metformin, with short-term induction of vascular endothelial growth factor (VEGF), cyclooxygenase 2 and CXC chemokine receptor 4 at the messenger RNA level and downregulation of ADAMTS1. Antibody array analysis shows an essentially opposite regulation of numerous angiogenesis-associated proteins in endothelial and breast cancer cells including interleukin-8, angiogenin and TIMP-1, as well as selective regulation of angiopioetin-1, -2, endoglin and others. Endothelial cell production of the cytochrome P450 member CYP1B1 is upregulated by tumor cell supernatants in an AMPK-dependent manner, metformin blocks this effect. Metformin inhibits VEGF-dependent activation of extracellular signal-regulated kinase 1/2, and the inhibition of AMPK activity abrogates this event. Metformin hinders angiogenesis in matrigel pellets in vivo, prevents the microvessel density increase observed in obese mice on a high-fat diet, downregulating the number of white adipose tissue endothelial precursor cells. Our data show that metformin has an antiangiogenic activity in vitro and in vivo associated with a contradictory short

  7. Gap-junctional coupling between neutrophils and endothelial cells

    OpenAIRE

    Zahler, Stefan; Hoffmann, Anke; Gloe, Torsten; Pohl, Ulrich

    2003-01-01

    Communication between leukocytes and endothelial cells is crucial for inflammatory reactions. Paracrine cross-talk and outside-in signaling (via adhesion molecules) have been characterized as communication pathways to date. As leukocytes and endothelial cells express connexins, we considered intercellular communication via gap junctions an intriguing additional concept. We found that gap-junctional coupling between neutrophils and endothelium occurred in a time-dependent, bidirectional manner...

  8. Cellular and molecular biology of aging endothelial cells.

    Science.gov (United States)

    Donato, Anthony J; Morgan, R Garrett; Walker, Ashley E; Lesniewski, Lisa A

    2015-12-01

    Cardiovascular disease (CVD) is the leading cause of death in the United States and aging is a major risk factor for CVD development. One of the major age-related arterial phenotypes thought to be responsible for the development of CVD in older adults is endothelial dysfunction. Endothelial function is modulated by traditional CVD risk factors in young adults, but advancing age is independently associated with the development of vascular endothelial dysfunction. This endothelial dysfunction results from a reduction in nitric oxide bioavailability downstream of endothelial oxidative stress and inflammation that can be further modulated by traditional CVD risk factors in older adults. Greater endothelial oxidative stress with aging is a result of augmented production from the intracellular enzymes NADPH oxidase and uncoupled eNOS, as well as from mitochondrial respiration in the absence of appropriate increases in antioxidant defenses as regulated by relevant transcription factors, such as FOXO. Interestingly, it appears that NFkB, a critical inflammatory transcription factor, is sensitive to this age-related endothelial redox change and its activation induces transcription of pro-inflammatory cytokines that can further suppress endothelial function, thus creating a vicious feed-forward cycle. This review will discuss the two macro-mechanistic processes, oxidative stress and inflammation, that contribute to endothelial dysfunction with advancing age as well as the cellular and molecular events that lead to the vicious cycle of inflammation and oxidative stress in the aged endothelium. Other potential mediators of this pro-inflammatory endothelial phenotype are increases in immune or senescent cells in the vasculature. Of note, genomic instability, telomere dysfunction or DNA damage has been shown to trigger cell senescence via the p53/p21 pathway and result in increased inflammatory signaling in arteries from older adults. This review will discuss the current state

  9. Expansion and cryopreservation of porcine and human corneal endothelial cells.

    Science.gov (United States)

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

    2017-08-01

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

  10. Cytotoxicity of activated monocytes on endothelial cells.

    Science.gov (United States)

    Peri, G; Chiaffarino, F; Bernasconi, S; Padura, I M; Mantovani, A

    1990-02-15

    Unstimulated human monocytes did not express appreciable levels of cytotoxicity on normal human umbilical vein endothelial cells (EC) in a 24-48 hr TdR release assay. On activation with IFN-gamma and LPS, monocytes had appreciable cytotoxicity on EC. Monocyte cytotoxicity on EC was not dependent on the presence of contaminating lymphoid cells. Recombinant TNF, IL-1, and IL-6 as well as monocyte supernatants did not exert a cytotoxic effect on EC. Moreover, anti-TNF, anti-IL-1, and anti-IL-6 antibodies, as well as scavengers of reactive oxygen intermediates, did not affect the cytotoxicity of activated monocytes on EC. Antibodies against the beta-chain (CD18) of leukocyte integrins inhibited the adhesion and cytotoxicity of activated monocytes on EC. Pretreatment of EC with IL-1 augmented the adhesion of monocytes on EC. Normal monocytes were not cytotoxic on IL-1-pretreated EC and IL-1 treatment did not increase the susceptibility of EC to activated monocytes. Thus adhesion is necessary but not sufficient for monocyte killing of EC. Anti-alpha L (LFA-1) antibodies markedly reduced monocyte cytotoxicity on EC, although anti-alpha X (p150) antibodies had only a modest effect. Anti-alpha M (Mac-1/CR3) antibodies were intermediate inhibitors of EC killing by activated monocytes. Thus, alpha L, beta 2 (LFA-1), and, to a lesser extent, alpha M, beta 2 (Mac-1/CR3) and alpha X, beta 2 (p 150, 95) integrins are the main adhesive structures involved in the cytotoxic interaction of activated monocytes with EC. Monocyte-mediated damage of EC could play a role as a mechanism of tissue injury under conditions of local or systemic activation of mononuclear phagocytes.

  11. Circulating endothelial progenitor cells in obese children and adolescents.

    Science.gov (United States)

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

    2015-01-01

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

  12. Endothelial cell adhesion and proliferation to PEGylated polymers with covalently linked RGD peptides.

    Science.gov (United States)

    Wang, Xin; Heath, Daniel E; Cooper, Stuart L

    2012-03-01

    A nonfouling peptide grafted polymer was synthesized that can promote endothelial cell (EC) binding. The polymer was composed of hexyl methacrylate, methyl methacrylate, poly(ethylene glycol) methacrylate, and CGRGDS peptide. The peptide was incorporated into the polymer system either by a chain transfer reaction or by coupling to an acrylate-PEG-N-hydroxysuccinimide (NHS) comonomer. The introduction of PEG chains minimizes protein adsorption. Human umbilical vein ECs and endothelial colony forming cells were cultured on these surfaces in short term and long-term studies. A difference in number and morphology of ECs was observed depending on the method of peptide incorporation. Both cell types adhered better to polymer films containing NHS coupled RGD peptide after 2 h even in the presence of albumin but significant cell detachment occurred after 4 days. Polymer solutions were electrospun into fibrous scaffolds. Both nonfouling and peptide binding characteristics were retained after processing. Copyright © 2012 Wiley Periodicals, Inc.

  13. Conserved signaling through vascular endothelial growth (VEGF) receptor family members in murine lymphatic endothelial cells.

    Science.gov (United States)

    Coso, Sanja; Zeng, Yiping; Sooraj, Dhanya; Williams, Elizabeth D

    2011-10-15

    Lymphatic vessels guide interstitial fluid, modulate immune responses by regulating leukocyte and antigen trafficking to lymph nodes, and in a cancer setting enable tumor cells to track to regional lymph nodes. The aim of the study was to determine whether primary murine lymphatic endothelial cells (mLECs) show conserved vascular endothelial growth factor (VEGF) signaling pathways with human LECs (hLECs). LECs were successfully isolated from murine dermis and prostate. Similar to hLECs, vascular endothelial growth factor (VEGF) family ligands activated MAPK and pAkt intracellular signaling pathways in mLECs. We describe a robust protocol for isolation of mLECs which, by harnessing the power of transgenic and knockout mouse models, will be a useful tool to study how LEC phenotype contributes to alterations in lymphatic vessel formation and function. Crown Copyright © 2011. Published by Elsevier Inc. All rights reserved.

  14. Single-cell imaging of mechanotransduction in endothelial cells.

    Science.gov (United States)

    Lu, Shaoying; Wang, Yingxiao

    2014-01-01

    Endothelial cells (ECs) are constantly exposed to chemical and mechanical microenvironment in vivo. In mechanotransduction, cells can sense and translate the extracellular mechanical cues into intracellular biochemical signals, to regulate cellular processes. This regulation is crucial for many physiological functions, such as cell adhesion, migration, proliferation, and survival, as well as the progression of disease such as atherosclerosis. Here, we overview the current molecular understanding of mechanotransduction in ECs associated with atherosclerosis, especially those in response to physiological shear stress. The enabling technology of live-cell imaging has allowed the study of spatiotemporal molecular events and unprecedented understanding of intracellular signaling responses in mechanotransduction. Hence, we also introduce recent studies on mechanotransduction using single-cell imaging technologies. © 2014 Elsevier Inc. All rights reserved.

  15. CD34 marks angiogenic tip cells in human vascular endothelial cell cultures

    NARCIS (Netherlands)

    Siemerink, Martin J.; Klaassen, Ingeborg; Vogels, Ilse M. C.; Griffioen, Arjan W.; van Noorden, Cornelis J. F.; Schlingemann, Reinier O.

    2012-01-01

    The functional shift of quiescent endothelial cells into tip cells that migrate and stalk cells that proliferate is a key event during sprouting angiogenesis. We previously showed that the sialomucin CD34 is expressed in a small subset of cultured endothelial cells and that these cells extend

  16. Endothelial cells present antigens in vivo

    Directory of Open Access Journals (Sweden)

    Tellides George

    2004-03-01

    Full Text Available Abstract Background Immune recognition of vascular endothelial cells (EC has been implicated in allograft rejection, protection against pathogens, and lymphocyte recruitment. However, EC pervade nearly all tissues and predominate in none, complicating any direct test of immune recognition. Here, we examined antigen presentation by EC in vivo by testing immune responses against E. coli β-galactosidase (β-gal in two lines of transgenic mice that express β-gal exclusively in their EC. TIE2-lacZ mice express β-gal in all EC and VWF-lacZ mice express β-gal in heart and brain microvascular EC. Results Transgenic and congenic wild type FVB mice immunized with β-gal expression vector DNA or β-gal protein generated high titer, high affinity antisera containing comparable levels of antigen-specific IgG1 and IgG2a isotypes, suggesting equivalent activation of T helper cell subsets. The immunized transgenic mice remained healthy, their EC continued to express β-gal, and their blood vessels showed no histological abnormalities. In response to β-gal in vitro, CD4+ and CD8+ T cells from immunized transgenic and FVB mice proliferated, expressed CD25, and secreted IFN-γ. Infection with recombinant vaccinia virus encoding β-gal raised equivalent responses in transgenic and FVB mice. Hearts transplanted from transgenic mice into FVB mice continued to beat and the graft EC continued to express β-gal. These results suggested immunological ignorance of the transgene encoded EC protein. However, skin transplanted from TIE2-lacZ onto FVB mice lost β-gal+ EC and the hosts developed β-gal-specific antisera, demonstrating activation of host immune effector mechanisms. In contrast, skin grafted from TIE2-lacZ onto VWF-lacZ mice retained β-gal+ EC and no antisera developed, suggesting a tolerant host immune system. Conclusion Resting, β-gal+ EC in transgenic mice tolerize specific lymphocytes that would otherwise respond against β-gal expressed by EC within

  17. Lymphocyte traffic through sinusoidal endothelial cells is regulated by hepatocytes.

    Science.gov (United States)

    Edwards, Sarah; Lalor, Patricia F; Nash, Gerard B; Rainger, G Ed; Adams, David H

    2005-03-01

    Crosstalk between hepatic sinusoidal ECs and closely juxtaposed hepatocytes via vascular endothelial growth factor is essential for the maintenance of sinusoidal endothelial growth and differentiation. We propose that paracrine interactions between endothelial cells and hepatocytes also may be responsible for the unique complement of adhesion receptors expressed on sinusoidal endothelium that regulate the recruitment of lymphocytes into the liver. To address this hypothesis, we developed an in vitro model of the hepatic sinusoid in which flowing lymphocytes could interact with hepatic endothelium conditioned by the presence of hepatocytes. Human hepatic sinusoidal endothelial cells cocultured with hepatocytes were activated so that they supported the adhesion of lymphocytes at levels equivalent to those seen on endothelium stimulated with the inflammatory cytokine tumour necrosis factor-beta. Lymphocyte adhesion was supported by intracellular adhesion molecule 1, vascular cell adhesion molecule 1, and E-selectin, with an additional contribution from the novel adhesion receptor VAP-1. In conclusion, we show that interactions between hepatocytes and endothelial cells amplify leukocyte recruitment through the sinusoids by regulating the expression and function of endothelial adhesion molecules. These paracrine interactions may be responsible for the induction of the adhesion molecules that support constitutive lymphocyte recruitment to the liver as well as contributing significantly to the patterns of leukocyte adhesion seen during episodes of hepatic inflammation.

  18. The influence of biomaterials on endothelial cell thrombogenicity

    Science.gov (United States)

    McGuigan, Alison P.; Sefton, Michael V.

    2007-01-01

    Driven by tissue engineering and regenerative medicine, endothelial cells are being used in combination with biomaterials in a number of applications for the purpose of improving blood compatibility and host integration. Endothelialized vascular grafts are beginning to be used clinically with some success in some centers, while endothelial seeding is being explored as a means of creating a vasculature within engineered tissues. The underlying assumption of this strategy is that when cultured on artificial biomaterials, a confluent layer of endothelial cells maintain their non-thrombogenic phenotype. In this review the existing knowledge base of endothelial cell thrombogenicity cultured on a number of different biomaterials is summarized. The importance of selecting appropriate endpoint measures that are most reflective of overall surface thrombogenicity is the focus of this review. Endothelial cells inhibit thrombosis through three interconnected regulatory systems (1) the coagulation cascade (2) the cellular components of the blood such as leukocytes and platelets and (3) the complement cascade, and also through effects on fibrinolysis and vascular tone, the latter which influences blood flow. Thus, in order to demonstrate the thromobgenic benefit of seeding a biomaterial with EC, the conditions under which EC surfaces are more likely to exhibit lower thrombogenicity than unseeded biomaterial surfaces need to be consistent with the experimental context. The endpoints selected should be appropriate for the dominant thrombotic process that occurs under the given experimental conditions. PMID:17316788

  19. Cyclosporine Induces Endothelial Cell Release of Complement-Activating Microparticles

    Science.gov (United States)

    Renner, Brandon; Klawitter, Jelena; Goldberg, Ryan; McCullough, James W.; Ferreira, Viviana P.; Cooper, James E.; Christians, Uwe

    2013-01-01

    Defective control of the alternative pathway of complement is an important risk factor for several renal diseases, including atypical hemolytic uremic syndrome. Infections, drugs, pregnancy, and hemodynamic insults can trigger episodes of atypical hemolytic uremic syndrome in susceptible patients. Although the mechanisms linking these clinical events with disease flares are unknown, recent work has revealed that each of these clinical conditions causes cells to release microparticles. We hypothesized that microparticles released from injured endothelial cells promote intrarenal complement activation. Calcineurin inhibitors cause vascular and renal injury and can trigger hemolytic uremic syndrome. Here, we show that endothelial cells exposed to cyclosporine in vitro and in vivo release microparticles that activate the alternative pathway of complement. Cyclosporine-induced microparticles caused injury to bystander endothelial cells and are associated with complement-mediated injury of the kidneys and vasculature in cyclosporine-treated mice. Cyclosporine-induced microparticles did not bind factor H, an alternative pathway regulatory protein present in plasma, explaining their complement-activating phenotype. Finally, we found that in renal transplant patients, the number of endothelial microparticles in plasma increases 2 weeks after starting tacrolimus, and treatment with tacrolimus associated with increased C3 deposition on endothelial microparticles in the plasma of some patients. These results suggest that injury-associated release of endothelial microparticles is an important mechanism by which systemic insults trigger intravascular complement activation and complement-dependent renal diseases. PMID:24092930

  20. MicroRNAs in Hyperglycemia Induced Endothelial Cell Dysfunction

    Directory of Open Access Journals (Sweden)

    Maskomani Silambarasan

    2016-04-01

    Full Text Available Hyperglycemia is closely associated with prediabetes and Type 2 Diabetes Mellitus. Hyperglycemia increases the risk of vascular complications such as diabetic retinopathy, diabetic nephropathy, peripheral vascular disease and cerebro/cardiovascular diseases. Under hyperglycemic conditions, the endothelial cells become dysfunctional. In this study, we investigated the miRNA expression changes in human umbilical vein endothelial cells exposed to different glucose concentrations (5, 10, 25 and 40 mM glucose and at various time intervals (6, 12, 24 and 48 h. miRNA microarray analyses showed that there is a correlation between hyperglycemia induced endothelial dysfunction and miRNA expression. In silico pathways analyses on the altered miRNA expression showed that the majority of the affected biological pathways appeared to be associated to endothelial cell dysfunction and apoptosis. We found the expression of ten miRNAs (miR-26a-5p, -26b-5p, 29b-3p, -29c-3p, -125b-1-3p, -130b-3p, -140-5p, -192-5p, -221-3p and -320a to increase gradually with increasing concentration of glucose. These miRNAs were also found to be involved in endothelial dysfunction. At least seven of them, miR-29b-3p, -29c-3p, -125b-1-3p, -130b-3p, -221-3p, -320a and -192-5p, can be correlated to endothelial cell apoptosis.

  1. Thrombospondin-1 signaling through CD47 inhibits cell cycle progression and induces senescence in endothelial cells.

    Science.gov (United States)

    Gao, Qi; Chen, Kexin; Gao, Lu; Zheng, Yang; Yang, Yong-Guang

    2016-09-08

    CD47 signaling in endothelial cells has been shown to suppress angiogenesis, but little is known about the link between CD47 and endothelial senescence. Herein, we demonstrate that the thrombospondin-1 (TSP1)-CD47 signaling pathway is a major mechanism for driving endothelial cell senescence. CD47 deficiency in endothelial cells significantly improved their angiogenic function and attenuated their replicative senescence. Lack of CD47 also suppresses activation of cell cycle inhibitors and upregulates the expression of cell cycle promoters, leading to increased cell cycle progression. Furthermore, TSP1 significantly accelerates replicative senescence and associated cell cycle arrest in a CD47-dependent manner. These findings demonstrate that TSP1-CD47 signaling is an important mechanism driving endothelial cell senescence. Thus, TSP1 and CD47 provide attractive molecular targets for treatment of aging-associated cardiovascular dysfunction and diseases involving endothelial dysregulation.

  2. Does corneal hysteresis correlate with endothelial cell density?

    Science.gov (United States)

    Akova-Budak, Berna; Kıvanç, Sertaç Argun

    2015-05-21

    Our aim was to determine if there is a correlation between corneal biomechanical properties, endothelial cell count, and corneal pachymetry in healthy corneas. Ninety-two eyes of all subjects underwent complete ocular examination, including intraocular pressure measurement by Goldmann applanation tonometer, objective refraction, and slit-lamp biomicroscopy. Topographic measurements and corneal pachymetry were performed using a Scheimpflug-based (Pentacam, Oculus, Germany) corneal topographer. Corneal hysteresis (CH) and corneal resistance factor (CRF) were measured with an Ocular Response Analyzer (ORA, Reichert Ophthalmic Instruments, Buffalo, NY). Endothelial cell count measurement was done using a specular microscope (CellChek, Konan, USA). Right eye values of the subjects were taken for the study. The mean CH was 11.5±1.7 mmHg and the mean CRF was 11.2±1.4 mmHg. Mean intraocular pressure was 15.3±2.3 mmHg. The mean endothelial cell count was 2754±205 cells/mm2. No correlation was found between biomechanical properties of cornea and endothelial cell count. There was a significant positive correlation between CH, CRF, and corneal thickness (pcorneal biomechanical properties significantly correlated with corneal thickness. We found no correlation between CH and CRF with the endothelial cell density in normal subjects.

  3. Human iPSC-Derived Endothelial Cell Sprouting Assay in ...

    Science.gov (United States)

    Activation of vascular endothelial cells (ECs) by growth factors initiates a cascade of events in vivo consisting of EC tip cell selection, sprout formation, EC stalk cell proliferation, and ultimately vascular stabilization by support cells. Although EC functional assays can recapitulate one or more aspects of angiogenesis in vitro, they are often limited by a lack of definition to the substratum and lack of dependence on key angiogenic signaling axes. Here, we designed and characterized a chemically-defined model of endothelial sprouting behavior in vitro using human induced pluripotent stem cell-derived endothelial cells (iPSC-ECs). Thiol-ene photopolymerization was used to rapidly encapsulate iPSC-ECs at high density in poly(ethylene glycol) (PEG) hydrogel spheres and subsequently to rapidly encapsulate iPSC-EC-containing hydrogel spheres in a cell-free over-layer. The hydrogel sprouting array here maintained pro-angiogenic phenotype of iPSC-ECs and supported growth factor-dependent proliferation and sprouting behavior. The sprouting model responded appropriately to several reference pharmacological angiogenesis inhibitors, which suggests the functional role of vascular endothelial growth factor, NF-κB, matrix metalloproteinase-2/9, protein kinase activity, and β-tubulin in endothelial sprouting. A blinded screen of 38 putative vascular disrupting compounds (pVDCs) from the US Environmental Protection Agency’s ToxCast library identified five compounds th

  4. Endothelial glycocalyx on brain endothelial cells is lost in experimental cerebral malaria

    DEFF Research Database (Denmark)

    Hempel, Casper; Hyttel, Poul; Kurtzhals, Jørgen Al

    2014-01-01

    electron microscopy and measured circulating glycosaminoglycans by dot blot and ELISA. The glycocalyx was degraded in brain vasculature in cerebral and to a lesser degree uncomplicated malaria. It was affected on both intact and apoptotic endothelial cells. Circulating glycosaminoglycan levels suggested...

  5. Endothelial protein C receptor in renal tubular epithelial cells and ...

    African Journals Online (AJOL)

    Jane

    2011-07-20

    Jul 20, 2011 ... However, troglitazone had protective effects of EPCR on injured cells. Key words: Endothelial protein C receptor, renal tubular epithelial cell, troglitazone, tumor necrosis factor-α, interleukin-1β; high glucose. ..... induce apoptosis of vascular smooth muscle cells through an extracellular signal- regulated ...

  6. Hyperphosphatemia, Phosphoprotein Phosphatases, and Microparticle Release in Vascular Endothelial Cells

    OpenAIRE

    Abbasian, Nima; Burton, James O.; Herbert, Karl E.; Tregunna, Barbara-Emily; Brown, Jeremy R.; Ghaderi-Najafabadi, Maryam; Brunskill, Nigel J.; Goodall, Alison H.; Bevington, Alan

    2015-01-01

    Hyperphosphatemia in patients with advanced CKD is thought to be an important contributor to cardiovascular risk, in part because of endothelial cell (EC) dysfunction induced by inorganic phosphate (Pi). Such patients also have an elevated circulating concentration of procoagulant endothelial microparticles (MPs), leading to a prothrombotic state, which may contribute to acute occlusive events. We hypothesized that hyperphosphatemia leads to MP formation from ECs through an elevation of intra...

  7. Regulation of tissue morphogenesis by endothelial cell-derived signals

    Science.gov (United States)

    Ramasamy, Saravana K.; Kusumbe, Anjali P.; Adams, Ralf H.

    2016-01-01

    Summary Endothelial cells form an extensive network of blood vessels that has numerous essential functions in the vertebrate body. In addition to their well-established role as a versatile transport network, blood vessels can induce organ formation or direct growth and differentiation processes by providing signals in a paracrine (angiocrine) fashion. Tissue repair also requires the local restoration of vasculature. Endothelial cells are emerging as important signaling centers that coordinate regeneration and help to prevent deregulated, disease-promoting processes. Vascular cells are also part of stem cell niches and play key roles in hematopoiesis, bone formation and neurogenesis. Here, we will review these newly identified roles of endothelial cells in the regulation of organ morphogenesis, maintenance and regeneration. PMID:25529933

  8. Metformin improves endothelial function in aortic tissue and microvascular endothelial cells subjected to diabetic hyperglycaemic conditions.

    Science.gov (United States)

    Ghosh, Suparna; Lakshmanan, Arun P; Hwang, Mu Ji; Kubba, Haidar; Mushannen, Ahmed; Triggle, Chris R; Ding, Hong

    2015-12-01

    The cellular mechanisms whereby metformin, the first line drug for type 2 diabetes (T2DM), mediates its antidiabetic effects remain elusive, particularly as to whether metformin has a direct protective action on the vasculature. This study was designed to determine if a brief 3-h exposure to metformin protects endothelial function against the effects of hyperglycaemia. We investigated the protective effects of metformin on endothelial-dependent vasodilatation (EDV) in thoracic aortae from T2DM db/db mice and on high glucose (HG, 40 mM) induced changes in endothelial nitric oxide synthase (eNOS) signaling in mouse microvascular endothelial cells (MMECs) in culture. Exposure of aortae from db+/? non-diabetic control mice to high glucose (HG, 40 mM) containing Krebs for 3-h significantly (Pmetformin; metformin also improved ACh-induced EDV in aortae from diabetic db/db mice. Immunoblot analysis of MMECs cultured in HG versus NG revealed a significant reduction of the ratio of phosphorylated (p-eNOS)/eNOS and p-Akt/Akt, but not the expression of total eNOS or Akt. The 3-h exposure of MMECs to metformin significantly (Pmetformin can reverse/reduce the impact of HG on endothelial function, via mechanisms linked to increased phosphorylation of eNOS and Akt. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Development of Cell Analysis Software for Cultivated Corneal Endothelial Cells.

    Science.gov (United States)

    Okumura, Naoki; Ishida, Naoya; Kakutani, Kazuya; Hongo, Akane; Hiwa, Satoru; Hiroyasu, Tomoyuki; Koizumi, Noriko

    2017-11-01

    To develop analysis software for cultured human corneal endothelial cells (HCECs). Software was designed to recognize cell borders and to provide parameters such as cell density, coefficient of variation, and polygonality of cultured HCECs based on phase contrast images. Cultured HCECs with high or low cell density were incubated with Ca-free and Mg-free phosphate-buffered saline for 10 minutes to reveal the cell borders and were then analyzed with software (n = 50). Phase contrast images showed that cell borders were not distinctly outlined, but these borders became more distinctly outlined after phosphate-buffered saline treatment and were recognized by cell analysis software. The cell density value provided by software was similar to that obtained using manual cell counting by an experienced researcher. Morphometric parameters, such as the coefficient of variation and polygonality, were also produced by software, and these values were significantly correlated with cell density (Pearson correlation coefficients -0.62 and 0.63, respectively). The software described here provides morphometric information from phase contrast images, and it enables subjective and noninvasive quality assessment for tissue engineering therapy of the corneal endothelium.

  10. Using cultured endothelial cells to study endothelial barrier dysfunction: Challenges and opportunities.

    Science.gov (United States)

    Aman, Jurjan; Weijers, Ester M; van Nieuw Amerongen, Geerten P; Malik, Asrar B; van Hinsbergh, Victor W M

    2016-08-01

    Despite considerable progress in the understanding of endothelial barrier regulation and the identification of approaches that have the potential to improve endothelial barrier function, no drug- or stem cell-based therapy is presently available to reverse the widespread vascular leak that is observed in acute respiratory distress syndrome (ARDS) and sepsis. The translational gap suggests a need to develop experimental approaches and tools that better mimic the complex environment of the microcirculation in which the vascular leak develops. Recent studies have identified several elements of this microenvironment. Among these are composition and stiffness of the extracellular matrix, fluid shear stress, interaction of endothelial cells (ECs) with pericytes, oxygen tension, and the combination of toxic and mechanic injurious stimuli. Development of novel cell culture techniques that integrate these elements would allow in-depth analysis of EC biology that closely approaches the (patho)physiological conditions in situ. In parallel, techniques to isolate organ-specific ECs, to define EC heterogeneity in its full complexity, and to culture patient-derived ECs from inducible pluripotent stem cells or endothelial progenitor cells are likely to advance the understanding of ARDS and lead to development of therapeutics. This review 1) summarizes the advantages and pitfalls of EC cultures to study vascular leak in ARDS, 2) provides an overview of elements of the microvascular environment that can directly affect endothelial barrier function, and 3) discusses alternative methods to bridge the gap between basic research and clinical application with the intent of improving the translational value of present EC culture approaches. Copyright © 2016 the American Physiological Society.

  11. Characterization and comparison of embryonic stem cell-derived KDR+ cells with endothelial cells.

    Science.gov (United States)

    Sun, Xuan; Cheng, Lamei; Duan, Huaxin; Lin, Ge; Lu, Guangxiu

    2012-09-01

    Growing interest in utilizing endothelial cells (ECs) for therapeutic purposes has led to the exploration of human embryonic stem cells (hESCs) as a potential source for endothelial progenitors. In this study, ECs were induced from hESC lines and their biological characteristics were analyzed and compared with both cord blood endothelial progenitor cells (CBEPCs) and human umbilical vein endothelial cells (HUVECs) in vitro. The results showed that isolated embryonic KDR+ cells (EC-KDR+) display characteristics that were similar to CBEPCs and HUVECs. EC-KDR+, CBEPCs and HUVECs all expressed CD31 and CD144, incorporated DiI-Ac-LDL, bound UEA1 lectin, and were able to form tube-like structures on Matrigel. Compared with CBEPCs and HUVECs, the expression level of endothelial progenitor cell markers such as CD133 and KDR in EC-KDR+ was significantly higher, while the mature endothelial marker vWF was lowly expressed in EC-KDR+. In summary, the study showed that EC-KDR+ are primitive endothelial-like progenitors and might be a potential source for therapeutic vascular regeneration and tissue engineering. Copyright © 2012 Elsevier Inc. All rights reserved.

  12. Endothelial progenitor cells in vascular health: focus on lifestyle.

    Science.gov (United States)

    Van Craenenbroeck, Emeline M; Conraads, Viviane M

    2010-05-01

    Endothelial dysfunction, which is considered the functional equivalent of a disrupted balance between endothelial injury and repair, precedes overt atherosclerosis by many years. Although this phenomenon is part of the normal aging process, prevention of early and progressive endothelial dysfunction has become an important therapeutic target. Evidence has accumulated to show that endothelial progenitor cells (EPC), contribute substantially to preservation of a structurally and functionally intact endothelium. There has been considerable progress in our understanding of the various cell types that were in the past all covered by the term "EPC." EPC home to sites of endothelial injury and ischemia, where they proliferate, differentiate and integrate into the endothelial layer or exert a paracrine function by producing vascular growth factors. Although more emphasis has been put on the pharmacological approach of endothelial dysfunction, the effect of a healthy lifestyle, via mobilization and functional improvement of EPC, is increasingly recognized. This review will focus on successful lifestyle interventions that aim to maintain vascular health through beneficial actions on cell populations with vasculogenic potential ("EPC"). The role of physical activity and dietary recommendations, which are considered essential elements of a healthy lifestyle, will be particularly emphasized. A thorough understanding of the physiology of endothelial benefits, derived from such interventions, may help to implement these measures on top of classical drug therapy, but also provides a solid basis for primary prevention. The effects of additional elements of a comprehensive lifestyle advice, such as smoking cessation, weight and stress reduction, also comprise a modulation of EPC function and circulating numbers and are therefore included in this review as well. Copyright 2009 Elsevier Inc. All rights reserved.

  13. Circulating endothelial cell number and markers of endothelial dysfunction in previously preeclamptic women.

    Science.gov (United States)

    Tuzcu, Zeyneb Baspehlivan; Asicioglu, Ebru; Sunbul, Murat; Ozben, Beste; Arikan, Hakki; Koc, Mehmet

    2015-10-01

    Patients with preeclampsia (PE) have endothelial dysfunction and an increased future risk of cardiovascular (CV) mortality. The number of circulating endothelial cells (CECs) is markedly increased in conditions associated with a high degree of endothelial cell activation/injury including PE. We hypothesized that the number of CECs continues to be increased in women with a history of PE, reflecting ongoing endothelial cell activation/injury. CECs, flow-mediated vasodilation, levels of adhesion molecules and soluble vascular endothelial growth factor receptor-1 (sVEGFR1), and urine albumin/creatinine ratio were determined in 21 healthy women with ongoing normal pregnancy, 24 healthy currently nonpregnant women with a history of normal pregnancy, a total of 17 women with currently active mild (n = 11) or severe (n = 6) PE without hemolysis, elevated liver enzymes, and low platelet count (HELLP) syndrome, and 16 currently nonpregnant women with a history of mild (n = 10) or severe (n = 6) PE. Blood samples from women with active preeclampsia had higher CECs (9.9 ± 7.9 cells/mL) than healthy pregnant women (3.0 ± 4.1 cells/mL; P < .001), healthy nonpregnant women with a history of normal pregnancy (3.4 ± 4.0 cells/mL; P < .001), or women with a history of preeclampsia (2.4 ± 2.0 cells/mL; P < .001). The number of CECs were similar between women with a history of preeclampsia and healthy nonpregnant women with a history of normal pregnancy. Patients with active preeclampsia had significantly higher soluble vascular cell adhesion molecule-1, soluble E-selectin, sVEGFR1, and urinary albumin/creatinine ratio than healthy pregnant women. However, soluble vascular cell adhesion molecule-1, soluble E-selectin, urinary albumin/creatinine ratio were similar in women with a history of preeclampsia and healthy nonpregnant women with a history of normal pregnancy. However, women with a history of preeclampsia had higher sVEGFR1 levels than women with a history of normal

  14. Characterization of vascular endothelial progenitor cells from chicken bone marrow

    Directory of Open Access Journals (Sweden)

    Bai Chunyu

    2012-05-01

    Full Text Available Abstract Background Endothelial progenitor cells (EPC are a type of stem cell used in the treatment of atherosclerosis, vascular injury and regeneration. At present, most of the EPCs studied are from human and mouse, whereas the study of poultry-derived EPCs has rarely been reported. In the present study, chicken bone marrow-derived EPCs were isolated and studied at the cellular level using immunofluorescence and RT-PCR. Results We found that the majority of chicken EPCs were spindle shaped. The growth-curves of chicken EPCs at passages (P 1, -5 and -9 were typically “S”-shaped. The viability of chicken EPCs, before and after cryopreservation was 92.2% and 81.1%, respectively. Thus, cryopreservation had no obvious effects on the viability of chicken EPCs. Dil-ac-LDL and FITC-UAE-1 uptake assays and immunofluorescent detection of the cell surface markers CD34, CD133, VEGFR-2 confirmed that the cells obtained in vitro were EPCs. Observation of endothelial-specific Weibel-Palade bodies using transmission electron microscopy further confirmed that the cells were of endothelial lineage. In addition, chicken EPCs differentiated into endothelial cells and smooth muscle cells upon induction with VEGF and PDGF-BB, respectively, suggesting that the chicken EPCs retained multipotency in vitro. Conclusions These results suggest that chicken EPCs not only have strong self-renewal capacity, but also the potential to differentiate into endothelial and smooth muscle cells. This research provides theoretical basis and experimental evidence for potential therapeutic application of endothelial progenitor cells in the treatment of atherosclerosis, vascular injury and diabetic complications.

  15. Characterization of angiogenin receptors on bovine brain capillary endothelial cells.

    Science.gov (United States)

    Chamoux, M; Dehouck, M P; Fruchart, J C; Spik, G; Montreuil, J; Cecchelli, R

    1991-04-30

    The mitogenic effect of bovine milk angiogenin was studied on bovine brain capillary and aortic endothelial cells, smooth muscle cells and fibroblasts. The proliferation of only bovine brain capillary endothelial cells was detected at concentrations ranging from 10 to 1,000 ng/ml, with a maximum effect at 100 ng/ml. This mitogenic activity may be correlated with a specific binding of angiogenin which was demonstrated only to bovine brain capillary endothelial cells. [125I]-labeled angiogenin binding was time and concentration dependent and saturable. Scatchard analyses of binding data showed evidence of a single class of binding sites with an apparent dissociation constant of 5.10(-10)M. The molecular mass of the angiogenin receptor (49 kDa) was determined by ligand blotting.

  16. Ponatinib reduces viability, migration, and functionality of human endothelial cells.

    Science.gov (United States)

    Gover-Proaktor, Ayala; Granot, Galit; Shapira, Saar; Raz, Oshrat; Pasvolsky, Oren; Nagler, Arnon; Lev, Dorit L; Inbal, Aida; Lubin, Ido; Raanani, Pia; Leader, Avi

    2017-06-01

    Tyrosine kinase inhibitors (TKIs) have revolutionized the prognosis of chronic myeloid leukemia. With the advent of highly efficacious therapy, the focus has shifted toward managing TKI adverse effects, such as vascular adverse events (VAEs). We used an in vitro angiogenesis model to investigate the TKI-associated VAEs. Our data show that imatinib, nilotinib, and ponatinib reduce human umbilical vein endothelial cells (HUVECs) viability. Pharmacological concentrations of ponatinib induced apoptosis, reduced migration, inhibited tube formation of HUVECs, and had a negative effect on endothelial progenitor cell (EPC) function. Furthermore, in HUVECs transfected with VEGF receptor 2 (VEGFR2), the effect of ponatinib on tube formation and on all parameters representing normal endothelial cell function was less prominent than in control cells. This is the first report regarding the pathogenesis of ponatinib-associated VAEs. The antiangiogenic effect of ponatinib, possibly mediated by VEGFR2 inhibition, as shown in our study, is another piece in the intricate puzzle of TKI-associated VAEs.

  17. Focus on biological identity of endothelial progenitors cells.

    Science.gov (United States)

    Zaccone, V; Flore, R; Santoro, L; De Matteis, G; Giupponi, B; Li Puma, D D; Santoliquido, A

    2015-11-01

    Circulating Endothelial Progenitor Cells (EPCs) were discovered by Asahara et al in 1997 and defined as bone marrow CD34+/KDR+ cells endowed with angiogenic potentialities in vitro and in vivo. The most likely assumption is that EPCs consist of several cell subpopulations with functions targeted at accomplishing the post-natal neovascularization process in a synergic and complementary fashion. Indeed, the subsequent identification of numerous and differentiated hematic populations, characterized by the capacity to develop an endothelial phenotype, has posed a number of questions as to the real identity of EPCs. This concept does not represent a sterile speculation but rather it suggests important implications for the future practice of stem cell therapy. The aim of this report was to explore through a critical analysis the two main experimental methodologies, in vitro culture and flow cytometry, applied to EPCs, followed by a brief revaluation of the endothelial progenitors employing a globally functional approach.

  18. Dynamic Endothelial Cell Rearrangements Drive Developmental Vessel Regression

    Science.gov (United States)

    Franco, Claudio A.; Jones, Martin L.; Bernabeu, Miguel O.; Geudens, Ilse; Mathivet, Thomas; Rosa, Andre; Lopes, Felicia M.; Lima, Aida P.; Ragab, Anan; Collins, Russell T.; Phng, Li-Kun; Coveney, Peter V.; Gerhardt, Holger

    2015-01-01

    Patterning of functional blood vessel networks is achieved by pruning of superfluous connections. The cellular and molecular principles of vessel regression are poorly understood. Here we show that regression is mediated by dynamic and polarized migration of endothelial cells, representing anastomosis in reverse. Establishing and analyzing the first axial polarity map of all endothelial cells in a remodeling vascular network, we propose that balanced movement of cells maintains the primitive plexus under low shear conditions in a metastable dynamic state. We predict that flow-induced polarized migration of endothelial cells breaks symmetry and leads to stabilization of high flow/shear segments and regression of adjacent low flow/shear segments. PMID:25884288

  19. Rapid flow-induced responses in endothelial cells

    Science.gov (United States)

    Stamatas, G. N.; McIntire, L. V.

    2001-01-01

    Endothelial cells alter their morphology, growth rate, and metabolism in response to fluid shear stress. To study rapid flow-induced responses in the 3D endothelial cell morphology and calcium distribution, coupled fluorescence microscopy with optical sectioning, digital imaging, and numerical deconvolution techniques have been utilized. Results demonstrate that within the first minutes of flow application nuclear calcium is increasing. In the same time frame whole cell height and nuclear height are reduced by about 1 microm. Whole cell height changes may facilitate reduction of shear stress gradients on the luminal surface, whereas nuclear structural changes may be important for modulating endothelial growth rate and metabolism. To study the role of the cytoskeleton in these responses, endothelial cells have been treated with specific disrupters (acrylamide, cytochalasin D, and colchicine) of each of the cytoskeleton elements (intermediate filaments, microfilaments, and microtubules, respectively). None of these compounds had any effect on the shear-induced calcium response. Cytochalasin D and acrylamide did not affect the shear-induced nuclear morphology changes. Colchicine, however, completely abrogated the response, indicating that microtubules may be implicated in force transmission from the plasma membrane to the nucleus. A pedagogical model based on tensegrity theory principles is presented that is consistent with the results on the 3D endothelial morphology.

  20. Nanofiber density determines endothelial cell behavior on hydrogel matrix

    Energy Technology Data Exchange (ETDEWEB)

    Berti, Fernanda V., E-mail: fernanda@intelab.ufsc.br [Department of Chemical and Food Engineering, Federal University of Santa Catarina, 88040-900 Florianópolis, SC (Brazil); Rambo, Carlos R. [Department of Electrical Engineering, Federal University of Santa Catarina, 88040-900 Florianópolis, SC (Brazil); Dias, Paulo F. [Department of Cell Biology, Embryology and Genetics, Federal University of Santa Catarina, 88040-900 Florianópolis, SC (Brazil); Porto, Luismar M. [Department of Chemical and Food Engineering, Federal University of Santa Catarina, 88040-900 Florianópolis, SC (Brazil)

    2013-12-01

    When cultured under static conditions, bacterial cellulose pellicles, by the nature of the polymer synthesis that involves molecular oxygen, are characterized by two distinct surface sides. The upper surface is denser in fibers (entangled) than the lower surface that shows greater surface porosity. Human umbilical vein endothelial cells (HUVECs) were used to exploit how the microarchitecture (i.e., surface porosity, fiber network structure, surface topology, and fiber density) of bacterial cellulose pellicle surfaces influence cell–biomaterial interaction and therefore cell behavior. Adhesion, cell ingrowth, proliferation, viability and cell death mechanisms were evaluated on the two pellicle surface sides. Cell behavior, including secondary necrosis, is influenced only by the microarchitecture of the surface, since the biomaterial is extremely pure (constituted of cellulose and water only). Cell–cellulose fiber interaction is the determinant signal in the cell–biomaterial responses, isolated from other frequently present interferences such as protein and other chemical traces usually present in cell culture matrices. Our results suggest that microarchitecture of hydrogel materials might determine the performance of biomedical products, such as bacterial cellulose tissue engineering constructs (BCTECs). - Highlights: • Topography of BC pellicle is relevant to determine endothelial cells' fate. • Cell–biomaterial response is affected by the topography of BC-pellicle surface. • Endothelial cells exhibit different behavior depending on the BC topography. • Apoptosis and necrosis of endothelial cells were affected by the BC topography.

  1. Detection of histidine decarboxylase mRNA in human vascular smooth muscle and endothelial cells.

    Science.gov (United States)

    Tippens, A S; Gruetter, C A

    2004-06-01

    The objective of this study was to investigate histamine synthesis capability of human vascular smooth muscle and endothelial cells by detecting histidine decarboxylase (HDC) mRNA. HDC catalyzes exclusively the formation of histamine in mammalian cells. Experiments utilizing nested reverse transcription-polymerase chain reaction (nRT-PCR) were conducted to detect the presence of HDC mRNA. Human aortic smooth muscle cells (HAoSMC) and human aortic endothelial cells (HAEC) were cultured and RNA was extracted and amplified using two sets of HDC-specific primers. Rat liver and kidney RNA were isolated and amplified to serve as positive and negative controls, respectively. Gel electrophoresis of HAoSMC, HAEC and liver mRNA revealed bands coinciding with an expected product size of 440 base pairs. Sequence analysis revealed that the observed bands were the appropriate HDC amplicons. These findings are the first to indicate the presence of HDC mRNA in vascular smooth muscle cells and confirm the presence of HDC mRNA in endothelial cells which is consistent with an ability of these cell types to synthesize histamine in the vascular wall.

  2. LGN Directs Interphase Endothelial Cell Behavior via the Microtubule Network.

    Directory of Open Access Journals (Sweden)

    Catherine E Wright

    Full Text Available Angiogenic sprouts require coordination of endothelial cell (EC behaviors as they extend and branch. Microtubules influence behaviors such as cell migration and cell-cell interactions via regulated growth and shrinkage. Here we investigated the role of the mitotic polarity protein LGN in EC behaviors and sprouting angiogenesis. Surprisingly, reduced levels of LGN did not affect oriented division of EC within a sprout, but knockdown perturbed overall sprouting. At the cell level, LGN knockdown compromised cell-cell adhesion and migration. EC with reduced LGN levels also showed enhanced growth and stabilization of microtubules that correlated with perturbed migration. These results fit a model whereby LGN influences interphase microtubule dynamics in endothelial cells to regulate migration, cell adhesion, and sprout extension, and reveal a novel non-mitotic role for LGN in sprouting angiogenesis.

  3. Magnetizable stent-grafts enable endothelial cell capture

    Science.gov (United States)

    Tefft, Brandon J.; Uthamaraj, Susheil; Harburn, J. Jonathan; Hlinomaz, Ota; Lerman, Amir; Dragomir-Daescu, Dan; Sandhu, Gurpreet S.

    2017-04-01

    Emerging nanotechnologies have enabled the use of magnetic forces to guide the movement of magnetically-labeled cells, drugs, and other therapeutic agents. Endothelial cells labeled with superparamagnetic iron oxide nanoparticles (SPION) have previously been captured on the surface of magnetizable 2205 duplex stainless steel stents in a porcine coronary implantation model. Recently, we have coated these stents with electrospun polyurethane nanofibers to fabricate prototype stent-grafts. Facilitated endothelialization may help improve the healing of arteries treated with stent-grafts, reduce the risk of thrombosis and restenosis, and enable small-caliber applications. When placed in a SPION-labeled endothelial cell suspension in the presence of an external magnetic field, magnetized stent-grafts successfully captured cells to the surface regions adjacent to the stent struts. Implantation within the coronary circulation of pigs (n=13) followed immediately by SPION-labeled autologous endothelial cell delivery resulted in widely patent devices with a thin, uniform neointima and no signs of thrombosis or inflammation at 7 days. Furthermore, the magnetized stent-grafts successfully captured and retained SPION-labeled endothelial cells to select regions adjacent to stent struts and between stent struts, whereas the non-magnetized control stent-grafts did not. Early results with these prototype devices are encouraging and further refinements will be necessary in order to achieve more uniform cell capture and complete endothelialization. Once optimized, this approach may lead to more rapid and complete healing of vascular stent-grafts with a concomitant improvement in long-term device performance.

  4. Endothelial cells are essential for ovarian stromal tissue restructuring after xenotransplantation of isolated ovarian stromal cells

    National Research Council Canada - National Science Library

    Dath, C; Dethy, A; Van Langendonckt, A; Van Eyck, A S; Amorim, C A; Luyckx, V; Donnez, J; Dolmans, M M

    2011-01-01

    .... To help improve the grafting technique, we investigated whether short-term xenografting of a suspension containing ovarian stromal and endothelial cells without follicles could enhance graft survival...

  5. Endothelial Cells Enhance Tumor Cell Invasion through a Crosstalk Mediated by CXC Chemokine Signaling

    Directory of Open Access Journals (Sweden)

    Kristy A. Warner

    2008-02-01

    Full Text Available Field cancerization involves the lateral spread of premalignant or malignant disease and contributes to the recurrence of head and neck tumors. The overall hypothesis underlying this work is that endothelial cells actively participate in tumor cell invasion by secreting chemokines and creating a chemotactic gradient for tumor cells. Here we demonstrate that conditioned medium from head and neck tumor cells enhance Bcl-2 expression in neovascular endothelial cells. Oral squamous cell carcinoma-3 (OSCC3 and Kaposi's sarcoma (SLK show enhanced invasiveness when cocultured with pools of human dermal microvascular endothelial cells stably expressing Bcl-2 (HDMEC-Bcl-2, compared to cocultures with empty vector controls (HDMEC-LXSN. Xenografted OSCC3 tumors vascularized with HDMEC-Bcl-2 presented higher local invasion than OSCC3 tumors vascularized with control HDMEC-LXSN. CXCL1 and CXCL8 were upregulated in primary endothelial cells exposed to vascular endothelial growth factor (VEGF, as well as in HDMEC-Bcl-2. Notably, blockade of CXCR2 signaling, but not CXCR1, inhibited OSCC3 and SLK invasion toward endothelial cells. These data demonstrate that CXC chemokines secreted by endothelial cells induce tumor cell invasion and suggest that the process of lateral spread of tumor cells observed in field cancerization is guided by chemotactic signals that originated from endothelial cells.

  6. Treponema pallidum Invades Intercellular Junctions of Endothelial Cell Monolayers

    Science.gov (United States)

    Thomas, D. Denee; Navab, Mahamad; Haake, David A.; Fogelman, Alan M.; Miller, James N.; Lovett, Michael A.

    1988-05-01

    The pathogenesis of syphilis reflects invasive properties of Treponema pallidum, but the actual mode of tissue invasion is unknown. We have found two in vitro parallels of treponemal invasiveness. We tested whether motile T. pallidum could invade host cells by determining the fate of radiolabeled motile organisms added to a HeLa cell monolayer; 26% of treponemes associated with the monolayer in a trypsin-resistant niche, presumably between the monolayer and the surface to which it adhered, but did not attain intracellularity. Attachment of T. pallidum to cultured human and rabbit aortic and human umbilical vein endothelial cells was 2-fold greater than to HeLa cells. We added T. pallidum to aortic endothelial cells grown on membrane filters under conditions in which tight intercellular junctions had formed. T. pallidum was able to pass through the endothelial cell monolayers without altering tight junctions, as measured by electrical resistance. In contrast, heat-killed T. pallidum and the nonpathogen Treponema phagedenis biotype Reiter failed to penetrate the monolayer. Transmission electron micrographs of sections of the monolayer showed T. pallidum in intercellular junctions. Our in vitro observations suggest that these highly motile spirochetes may leave the circulation by invading the junctions between endothelial cells.

  7. Arecoline inhibits endothelial cell growth and migration and the attachment to mononuclear cells

    Directory of Open Access Journals (Sweden)

    Shuei-Kuen Tseng

    2014-09-01

    Conclusion: Arecoline impaired vascular endothelial cells by inhibiting their growth and migration and their adhesion to U937 mononuclear cells. These results reveal that arecoline may contribute to the pathogenesis of oral submucous fibrosis and cardiovascular diseases by affecting endothelial cell function in BQ chewers.

  8. 3D map of the human corneal endothelial cell

    OpenAIRE

    Zhiguo He; Fabien Forest; Philippe Gain; Damien Rageade; Aurélien Bernard; Sophie Acquart; Michel Peoc’h; Dennis M. Defoe; Gilles Thuret

    2016-01-01

    Corneal endothelial cells (CECs) are terminally differentiated cells, specialized in regulating corneal hydration and transparency. They are highly polarized flat cells that separate the cornea from the aqueous humor. Their apical surface, in contact with aqueous humor is hexagonal, whereas their basal surface is irregular. We characterized the structure of human CECs in 3D using confocal microscopy of immunostained whole corneas in which cells and their interrelationships remain intact. Hexa...

  9. Mechanotransduction in Endothelial Cells Studied with Fluorescence Imaging

    Energy Technology Data Exchange (ETDEWEB)

    Chien Shu [Departments of Bioengineering and Medicine and Institute of Engineering in Medicine, University of California, San Diego, La Jolla, California 92093-0427 (United States)

    2011-01-01

    Mechanotransduction involves the conversion of mechanical stimuli to intracellular signaling to modulate gene and protein expressions and hence cellular functions in endothelial cells, thus playing importance roles in the regulation of homeostasis in health and disease. The aim of this paper is to investigate the dynamics of mechanotransduction in endothelial cells by the use of fluorescent resonance energy transfer (FRET) to study the temporal and spatial activation of Src kinase and focal adhesion kinase, both of which play critical roles in many cellular processes. The results have contributed to the elucidation of the roles of these two important signaling molecules and their interactions in mediating mechanotransduction.

  10. High glucose induced endothelial to mesenchymal transition in human umbilical vein endothelial cell.

    Science.gov (United States)

    Yu, Chun-Hong; Suriguga; Gong, Meng; Liu, Wen-Juan; Cui, Ning-Xuan; Wang, Ying; Du, Xin; Yi, Zong-Chun

    2017-06-01

    Studies have shown that endothelial-to-mesenchymal transition (EndMT) could contribute to the progression of diabetic nephropathy, diabetic renal fibrosis, and cardiac fibrosis. The aim of this study was to investigate the influence of high glucose and related mechanism of MAPK inhibitor or specific antioxidant on the EndMT. In vitro human umbilical vein endothelial cells (HUVEC) were cultured with 11mM, 30mM, 60mM and 120mM glucose for 0, 24, 48, 72 and 168h. Endothelial cell morphology was observed with microscope, and RT-PCR was used to detect mRNA expression of endothelial markers VE-cadherin and CD31, mesenchymal markers α-SMA and collagen I, and transforming growth factor TGF-β1. Immunofluorescence staining was performed to detect the expression of CD31 and α-SMA. The concentration of TGF-β1 in the supernatant was detected by ELISA. ERK1/2 phosphorylation level was detected by Western blot analysis. High glucose induced EndMT and increased the TGF-β1 level in HUVEC cells. Cells in high glucose for 7 days showed a significant decrease in mRNA expression of CD31 and VE-cadherin, and a significant increase in that of α-SMA and collagen I, while lost CD31 staining and acquired α-SMA staining. ERK signaling pathway blocker PD98059 significantly attenuated the high glucose-induced increase in the ERK1/2 phosphorylation level. PD98059 and NAC both inhibited high glucose-induced TGF-β1 expression and attenuated EndMT marker protein synthesis. High glucose could induce HUVEC cells to undergo EndMT. NAC and ERK signaling pathway may play important role in the regulation of the TGF-β1 biosynthesis during high glucose-induced EndMT. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Glyoxalase 1-knockdown in human aortic endothelial cells - effect on the proteome and endothelial function estimates.

    Science.gov (United States)

    Stratmann, Bernd; Engelbrecht, Britta; Espelage, Britta C; Klusmeier, Nadine; Tiemann, Janina; Gawlowski, Thomas; Mattern, Yvonne; Eisenacher, Martin; Meyer, Helmut E; Rabbani, Naila; Thornalley, Paul J; Tschoepe, Diethelm; Poschmann, Gereon; Stühler, Kai

    2016-11-29

    Methylglyoxal (MG), an arginine-directed glycating agent, is implicated in diabetic late complications. MG is detoxified by glyoxalase 1 (GLO1) of the cytosolic glyoxalase system. The aim was to investigate the effects of MG accumulation by GLO1-knockdown under hyperglycaemic conditions in human aortic endothelial cells (HAECs) hypothesizing that the accumulation of MG accounts for the deleterious effects on vascular function. SiRNA-mediated knockdown of GLO1 was performed and MG concentrations were determined. The impact of MG on the cell proteome and targets of MG glycation was analysed, and confirmed by Western blotting. Markers of endothelial function and apoptosis were assessed. Collagen content was assayed in cell culture supernatant. GLO1-knockdown increased MG concentration in cells and culture medium. This was associated with a differential abundance of cytoskeleton stabilisation proteins, intermediate filaments and proteins involved in posttranslational modification of collagen. An increase in fibrillar collagens 1 and 5 was detected. The extracellular concentration of endothelin-1 was increased following GLO1-knockdown, whereas the phosphorylation and amount of eNOS was not influenced by GLO1-knockdown. The expression of ICAM-1, VCAM-1 and of MCP-1 was elevated and apoptosis was increased. MG accumulation by GLO1-knockdown provoked collagen expression, endothelial inflammation and dysfunction and apoptosis which might contribute to vascular damage.

  12. Human Endothelial Cells: Use of Heparin in Cloning and Long-Term Serial Cultivation

    Science.gov (United States)

    Thornton, Susan C.; Mueller, Stephen N.; Levine, Elliot M.

    1983-11-01

    Endothelial cells from human blood vessels were cultured in vitro, with doubling times of 17 to 21 hours for 42 to 79 population doublings. Cloned human endothelial cell strains were established for the first time and had similar proliferative capacities. This vigorous cell growth was achieved by addition of heparin to culture medium containing reduced concentrations of endothelial cell growth factor. The routine cloning and long-term culture of human endothelial cells will facilitate studying the human endothelium in vitro.

  13. Effects of irradiated biodegradable polymer in endothelial cell monolayer formation

    Energy Technology Data Exchange (ETDEWEB)

    Arbeitman, Claudia R.; Grosso, Mariela F. del [CONICET – Consejo Nacional de Investigaciones Científicas y Técnicas (Argentina); Gerencia de Investigación y Aplicaciones, TANDAR-CNEA (Argentina); Behar, Moni [Instituto de Física, UFRGS, Porto Alegre, RS (Brazil); García Bermúdez, Gerardo, E-mail: ggb@tandar.cnea.gov.ar [CONICET – Consejo Nacional de Investigaciones Científicas y Técnicas (Argentina); Gerencia de Investigación y Aplicaciones, TANDAR-CNEA (Argentina); Escuela de Ciencia y Tecnología, UNSAM (Argentina)

    2013-11-01

    In this work we study cell adhesion, proliferation and cell morphology of endothelial cell cultured on poly-L-lactide acid (PLLA) modified by heavy ion irradiation. Thin films of PLLA samples were irradiated with sulfur (S) at energies of 75 MeV and gold (Au) at 18 MeV ion-beams. Ion beams were provided by the Tandar (Buenos Aires, Argentina) and Tandetron (Porto Alegre, Brazil) accelerators, respectively. The growth of a monolayer of bovine aortic endothelial cells (BAEC) onto unirradiated and irradiated surfaces has been studied by in vitro techniques in static culture. Cell viability and proliferation increased on modified substrates. But the results on unirradiated samples, indicate cell death (necrosis/apoptosis) with the consequent decrease in proliferation. We analyzed the correlation between irradiation parameters and cell metabolism and morphology.

  14. Serglycin in Quiescent and Proliferating Primary Endothelial Cells

    Science.gov (United States)

    Reine, Trine M.; Vuong, Tram T.; Rutkovskiy, Arkady; Meen, Astri J.; Vaage, Jarle; Jenssen, Trond G.; Kolset, Svein O.

    2015-01-01

    Proteoglycans are fundamental components of the endothelial barrier, but the functions of the proteoglycan serglycin in endothelium are less described. Our aim was to describe the roles of serglycin in processes relevant for endothelial dysfunction. Primary human umbilical vein endothelial cells (HUVEC) were cultured in vitro and the expression of proteoglycans was investigated. Dense cell cultures representing the quiescent endothelium coating the vasculature was compared to sparse activated cell cultures, relevant for diabetes, cancer and cardiovascular disease. Secretion of 35S- proteoglycans increased in sparse cultures, and we showed that serglycin is a major component of the cell-density sensitive proteoglycan population. In contrast to the other proteoglycans, serglycin expression and secretion was higher in proliferating compared to quiescent HUVEC. RNAi silencing of serglycin inhibited proliferation and wound healing, and serglycin expression and secretion was augmented by hypoxia, mechanical strain and IL-1β induced inflammation. Notably, the secretion of the angiogenic chemokine CCL2 resulting from IL-1β activation, was increased in serglycin knockdown cells, while angiopoietin was not affected. Both serglycin and CCL2 were secreted predominantly to the apical side of polarized HUVEC, and serglycin and CCL2 co-localized both in perinuclear areas and in vesicles. These results suggest functions for serglycin in endothelial cells trough interactions with partner molecules, in biological processes with relevance for diabetic complications, cardiovascular disease and cancer development. PMID:26694746

  15. Tumor-endothelium cross talk blocks recruitment of neutrophils to endothelial cells: a novel mechanism of endothelial cell anergy.

    Science.gov (United States)

    Blaheta, Roman A; Powerski, Maciej; Hudak, Lukasz; Juengel, Eva; Jonas, Dietger; von Knethen, Andreas; Doerr, Hans Willhelm; Cinatl, Jindrich

    2009-10-01

    Tumor cells have evolved effective strategies to escape the host immune response. The objective of this study was to determine whether tumor cells can condition endothelial cells in a specific manner to prevent subsequent adhesion of polymorphonuclear neutrophils (PMNs) and/or peripheral blood lymphocytes (PBLs). Human umbilical vein endothelial cells (HUVECs) and UKF-NB-4 neuroblastoma tumor cells were established in coculture on opposite sides of porous transwell filters. After 24 hours with and without HUVEC conditioning, PMNs or PBLs were added to the HUVEC monolayer. Adhesion to conditioned HUVEC versus adhesion to nonconditioned HUVEC was compared. Effects on endothelial CD44v4, CD44v5, CD44v7, intercellular adhesion molecule 1 (ICAM-1), E-selectin, and vascular cell adhesion molecule 1 (VCAM-1) adhesion receptor expression were analyzed by flow cytometry, intracellular signaling proteins of the mitogen-activated protein kinase pathway and protein kinase C (PKC) subtypes quantified by Western blot analysis. Endothelial conditioning led to a distinct reduction in PMN but not in PBL adhesion to HUVEC. CD44 was significantly reduced, whereas ICAM-1, E-selectin, and VCAM-1 were not altered during HUVEC conditioning. Antibody blockade against CD44v4, CD44v5, and CD44v7 inhibited PMN but not PBL binding. The observed effects were caused by direct tumor cell-HUVEC contact because addition of isolated tumor cell membrane fragments but not of soluble cell culture supernatant to HUVEC induced the CD44 receptor loss. PKCalpha activity was strongly enhanced in conditioned HUVEC. Blocking PKC prevented the reduction in PMN binding, indicating that this protein is involved in PMN adhesion regulation. A novel tumor escape strategy is presented here. Cell contact-dependent adhesion of tumor cells to the vascular wall promotes down-regulation of endothelial CD44 receptor expression, impairing an effective neutrophil attack.

  16. Tumor-Endothelium Cross Talk Blocks Recruitment of Neutrophils to Endothelial Cells: A Novel Mechanism of Endothelial Cell Anergy

    Directory of Open Access Journals (Sweden)

    Roman A. Blaheta

    2009-10-01

    Full Text Available Tumor cells have evolved effective strategies to escape the host immune response. The objective of this study was to determine whether tumor cells can condition endothelial cells in a specific manner to prevent subsequent adhesion of polymorphonuclear neutrophils (PMNs and/or peripheral blood lymphocytes (PBLs. Human umbilical vein endothelial cells (HUVECs and UKF-NB-4 neuroblastoma tumor cells were established in coculture on opposite sides of porous transwell filters. After 24 hours with and without HUVEC conditioning, PMNs or PBLs were added to the HUVEC monolayer. Adhesion to conditioned HUVEC versus adhesion to nonconditioned HUVEC was compared. Effects on endothelial CD44v4, CD44v5, CD44v7, intercellular adhesion molecule 1 (ICAM-1, E-selectin, and vascular cell adhesion molecule 1 (VCAM-1 adhesion receptor expression were analyzed by flow cytometry, intracellular signaling proteins of the mitogen-activated protein kinase pathway and protein kinase C (PKC subtypes quantified by Western blot analysis. Endothelial conditioning led to a distinct reduction in PMN but not in PBL adhesion to HUVEC. CD44 was significantly reduced, whereas ICAM-1, E-selectin, and VCAM-1 were not altered during HUVEC conditioning. Antibody blockade against CD44v4, CD44v5, and CD44v7 inhibited PMN but not PBL binding. The observed effects were caused by direct tumor cell-HUVEC contact because addition of isolated tumor cell membrane fragments but not of soluble cell culture supernatant to HUVEC induced the CD44 receptor loss. PKCα activity was strongly enhanced in conditioned HUVEC. Blocking PKC prevented the reduction in PMN binding, indicating that this protein is involved in PMN adhesion regulation. A novel tumor escape strategy is presented here. Cell contact-dependent adhesion of tumor cells to the vascular wall promotes down-regulation of endothelial CD44 receptor expression, impairing an effective neutrophil attack.

  17. Synthetic heparan sulfate oligosaccharides inhibit endothelial cell functions essential for angiogenesis.

    Directory of Open Access Journals (Sweden)

    Claire L Cole

    2010-07-01

    Full Text Available Heparan sulfate (HS is an important regulator of the assembly and activity of various angiogenic signalling complexes. However, the significance of precisely defined HS structures in regulating cytokine-dependent angiogenic cellular functions and signalling through receptors regulating angiogenic responses remains unclear. Understanding such structure-activity relationships is important for the rational design of HS fragments that inhibit HS-dependent angiogenic signalling complexes.We synthesized a series of HS oligosaccharides ranging from 7 to 12 saccharide residues that contained a repeating disaccharide unit consisting of iduronate 2-O-sulfate linked to glucosamine with or without N-sulfate. The ability of oligosaccharides to compete with HS for FGF2 and VEGF165 binding significantly increased with oligosaccharide length and sulfation. Correspondingly, the inhibitory potential of oligosaccharides against FGF2- and VEGF165-induced endothelial cell responses was greater in longer oligosaccharide species that were comprised of disaccharides bearing both 2-O- and N-sulfation (2SNS. FGF2- and VEGF165-induced endothelial cell migration were inhibited by longer 2SNS oligosaccharide species with 2SNS dodecasaccharide activity being comparable to that of receptor tyrosine kinase inhibitors targeting FGFR or VEGFR-2. Moreover, the 2SNS dodecasaccharide ablated FGF2- or VEGF165-induced phosphorylation of FAK and assembly of F-actin in peripheral lamellipodia-like structures. In contrast, FGF2-induced endothelial cell proliferation was only moderately inhibited by longer 2SNS oligosaccharides. Inhibition of FGF2- and VEGF165-dependent endothelial tube formation strongly correlated with oligosaccharide length and sulfation with 10-mer and 12-mer 2SNS oligosaccharides being the most potent species. FGF2- and VEGF165-induced activation of MAPK pathway was inhibited by biologically active oligosaccharides correlating with the specific phosphorylation

  18. A role for activated endothelial cells in red blood cell clearance: implications for vasopathology.

    Science.gov (United States)

    Fens, Marcel H A M; van Wijk, Richard; Andringa, Grietje; van Rooijen, Karlijn L; Dijstelbloem, Hilde M; Rasmussen, Jan T; de Vooght, Karen M K; Schiffelers, Raymond M; Gaillard, Carlo A J M; van Solinge, Wouter W

    2012-04-01

    Phosphatidylserine exposure by red blood cells is acknowledged as a signal that initiates phagocytic removal of the cells from the circulation. Several disorders and conditions are known to induce phosphatidylserine exposure. Removal of phosphatidylserine-exposing red blood cells generally occurs by macrophages in the spleen and liver. Previously, however, we have shown that endothelial cells are also capable of erythrophagocytosis. Key players in the erythrophagocytosis by endothelial cells appeared to be lactadherin and α(v)-integrin. Phagocytosis via the phosphatidylserine-lactadherin-α(v)-integrin pathway is the acknowledged route for removal of apoptotic innate cells by phagocytes. Endothelial cell phagocytosis of red blood cells was further explored using a more (patho)physiological approach. Red blood cells were exposed to oxidative stress, induced by tert-butyl hydroperoxide. After opsonization with lactadherin, red blood cells were incubated with endothelial cells to study erythrophagocytosis and examine cytotoxicity. Red blood cells exposed to oxidative stress show alterations such as phosphatidylserine exposure and loss of deformability. When incubated with endothelial cells, marked erythrophagocytosis occurred in the presence of lactadherin under both static and flow conditions. As a consequence, intracellular organization was disturbed and endothelial cells were seen to change shape ('rounding up'). Increased expression of apoptotic markers indicated that marked erythrophagocytosis has cytotoxic effects. Activated endothelial cells show significant phagocytosis of phosphatidylserine-exposing and rigid red blood cells under both static and flow conditions. This results in a certain degree of cytotoxicity. We postulate that activated endothelial cells play a role in red blood cell clearance in vivo. Significant erythrophagocytosis can induce endothelial cell loss, which may contribute to vasopathological effects as seen, for instance, in sickle cell

  19. Allogeneic Mesenchymal Stem Cells Restore Endothelial Function in Heart Failure by Stimulating Endothelial Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Courtney Premer

    2015-05-01

    Interpretation: These findings reveal a novel mechanism whereby allogeneic, but not autologous, MSC administration results in the proliferation of functional EPCs and improvement in vascular reactivity, which in turn restores endothelial function towards normal in patients with HF. These findings have significant clinical and biological implications for the use of MSCs in HF and other disorders associated with endothelial dysfunction.

  20. Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture

    Directory of Open Access Journals (Sweden)

    Magnusson Magnus K

    2010-07-01

    Full Text Available Abstract Background Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D co-culture assay. Methods Breast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy. Results In co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors. Conclusion Breast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.

  1. Antiproliferative effect of elevated glucose in human microvascular endothelial cells

    Science.gov (United States)

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

    1998-01-01

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

  2. Ex Vivo Behaviour of Human Bone Tumor Endothelial Cells

    Energy Technology Data Exchange (ETDEWEB)

    Infante, Teresa [SDN-Foundation, Institute of Diagnostic and Nuclear Development, IRCCS, 80143 Naples (Italy); Cesario, Elena [Department of Biochemistry and Biophysics, Second University of Naples, 80138 Naples (Italy); Gallo, Michele; Fazioli, Flavio [Division of Skeletal Muscles Oncology Surgery, National Cancer Institute, Pascale Foundation, 80131 Naples (Italy); De Chiara, Annarosaria [Anatomic Pathology Unit, National Cancer Institute, Pascale Foundation, 80131 Naples (Italy); Tutucci, Cristina; Apice, Gaetano [Medical Oncology of Bone and Soft Sarcoma tissues Unit, National Cancer Institute, Pascale Foundation, 80131 Naples (Italy); Nigris, Filomena de, E-mail: filomena.denigris@unina2.it [Department of Biochemistry and Biophysics, Second University of Naples, 80138 Naples (Italy)

    2013-04-11

    Cooperation between endothelial cells and bone in bone remodelling is well established. In contrast, bone microvasculature supporting the growth of primary tumors and metastasis is poorly understood. Several antiangiogenic agents have recently been undergoing trials, although an extensive body of clinical data and experimental research have proved that angiogenic pathways differ in each tumor type and stage. Here, for the first time, we characterize at the molecular and functional level tumor endothelial cells from human bone sarcomas at different stages of disease and with different histotypes. We selected a CD31{sup +} subpopulation from biopsies that displayed the capability to grow as adherent cell lines without vascular endothelial growth factor (VEGF). Our findings show the existence in human primary bone sarcomas of highly proliferative endothelial cells expressing CD31, CD44, CD105, CD146 and CD90 markers. These cells are committed to develop capillary-like structures and colony formation units, and to produce nitric oxide. We believe that a better understanding of tumor vasculature could be a valid tool for the design of an efficacious antiangiogenic therapy as adjuvant treatment of sarcomas.

  3. Apical sorting of ADAMTS13 in vascular endothelial cells and Madin-Darby canine kidney cells depends on the CUB domains and their association with lipid rafts.

    Science.gov (United States)

    Shang, Dezhi; Zheng, X Wu; Niiya, Masami; Zheng, X Long

    2006-10-01

    ADAMTS13 biosynthesis appeared to occur mainly in hepatic stellate cells, but detection of ADAMTS13 mRNA in many other tissues suggests that vascular endothelium may also produce ADAMTS13. We showed that ADAMTS13 mRNA and protein were detectable in human umbilical vein endothelial cells, aortic endothelial cells, and endothelium-derived cell line (ECV304). ADAMTS13 in cell lysate or serum-free conditioned medium cleaved von Willebrand factor (VWF) specifically. ADAMTS13 and VWF were localized to the distinct compartments of endothelial cells. Moreover, ADAMTS13 was preferentially sorted into apical domain of ECV304 and Madin-Darby canine kidney (MDCK) cells. Apical sorting of ADAMTS13 depended on the CUB domains and their association with lipid rafts. A mutation in the second CUB domain of ADAMTS13 (4143-4144insA), naturally occurring in patients with inherited thrombotic thrombocytopenic purpura, resulted in a significant reduction of ADAMTS13 secretion and a reversal of its polarity in MDCK cells. These data demonstrated that ADAMTS13 is synthesized and secreted from endothelial cells; the apically secreted ADAMTS13 from endothelial cells may contribute significantly to plasma ADAMTS13 proteases. The data also suggest a critical role of the CUB domains and a novel cargo-selective mechanism for apical sorting of a soluble ADAMTS protease in polarized cells.

  4. Reelin expression in brain endothelial cells: an electron microscopy study.

    Science.gov (United States)

    Perez-Costas, Emma; Fenton, Erin Y; Caruncho, Hector J

    2015-03-24

    Reelin expression and function have been extensively studied in the brain, although its expression has been also reported in other tissues including blood. This raises the possibility that reelin might be able to cross the blood-brain barrier, which could be functionally relevant. Up-to-date no studies have been conducted to assess if reelin is present in the blood-brain barrier, which is mainly constituted by tightly packed endothelial cells. In this report we assessed the expression of reelin in brain capillaries using immunocytochemistry and electron microscopy. At the light microscope, reelin immunolabeling appeared in specific endothelial cells in brain areas that presented abundant diffuse labeling for this protein (e.g., layer I of the cortex, or the stratum lacunosum moleculare of the hippocampus), while it was mostly absent from capillaries in other brain areas (e.g., deeper cortical layers, or the CA1 layer of the hippocampus). As expected, at the electron microscope reelin labeling was observed in neurons of the cortex, where most of the labeling was associated with the rough endoplasmic reticulum. Importantly, reelin was also observed in some endothelial cells located in small capillaries, which confirmed the findings obtained at the light microscope. In these cells, reelin labeling was located primarily in caveolae (i.e., vesicles of transcytosis), and associated with the plasma membrane of the luminal side of endothelial cells. In addition, some scarce labeling was observed in the nuclear membrane. The presence of reelin immunolabeling in brain endothelial cells, and particularly in caveolar vesicles within these cells, suggests that reelin and/or reelin peptides may be able to cross the blood-brain barrier, which could have important physiological, pathological, and therapeutic implications.

  5. Immune-induced fever is mediated by IL-6 receptors on brain endothelial cells coupled to STAT3-dependent induction of brain endothelial prostaglandin synthesis.

    Science.gov (United States)

    Eskilsson, Anna; Mirrasekhian, Elahe; Dufour, Sylvie; Schwaninger, Markus; Engblom, David; Blomqvist, Anders

    2014-11-26

    The cytokine IL-6, which is released upon peripheral immune challenge, is critical for the febrile response, but the mechanism by which IL-6 is pyrogenic has remained obscure. Here we generated mice with deletion of the membrane bound IL-6 receptor α (IL-6Rα) on neural cells, on peripheral nerves, on fine sensory afferent fibers, and on brain endothelial cells, respectively, and examined its role for the febrile response to peripherally injected lipopolysaccharide. We show that IL-6Rα on neural cells, peripheral nerves, and fine sensory afferents are dispensable for the lipopolysaccharide-induced fever, whereas IL-6Rα in the brain endothelium plays an important role. Hence deletion of IL-6Rα on brain endothelial cells strongly attenuated the febrile response, and also led to reduced induction of the prostaglandin synthesizing enzyme Cox-2 in the hypothalamus, the temperature-regulating center in the brain, as well as reduced expression of SOCS3, suggesting involvement of the STAT signaling pathway. Furthermore, deletion of STAT3 in the brain endothelium also resulted in attenuated fever. These data show that IL-6, when endogenously released during systemic inflammation, is pyrogenic by binding to IL-6Rα on brain endothelial cells to induce prostaglandin synthesis in these cells, probably in concerted action with other peripherally released cytokines. Copyright © 2014 the authors 0270-6474/14/3415957-05$15.00/0.

  6. Oleic acid increases mitochondrial reactive oxygen species production and decreases endothelial nitric oxide synthase activity in cultured endothelial cells

    NARCIS (Netherlands)

    Gremmels, Hendrik; Bevers, Lonneke M.; Fledderus, Joost O.; Braam, Branko; Jan Van Zonneveld, Anton; Verhaar, Marianne C.; Joles, Jaap A.

    2015-01-01

    Elevated plasma levels of free fatty acids (FFA) are associated with increased cardiovascular risk. This may be related to FFA-induced elevation of oxidative stress in endothelial cells. We hypothesized that, in addition to mitochondrial production of reactive oxygen species, endothelial nitric

  7. Effect of propionyl-L-carnitine on human endothelial cells

    NARCIS (Netherlands)

    Hinsbergh, V.W.M. van; Scheffer, M.A.

    1991-01-01

    A possible protective effect of propionyl-L-carnitine on human endothelial cells was studied both under basal culture conditions and in the presence of agents capable of influencing oxidative damage, such as glucose/glucose oxidase and oxidized low-density lipoproteins. Propionyl-L-carnitine had no

  8. Dabigatran abrogates brain endothelial cell permeability in response to thrombin.

    Science.gov (United States)

    Hawkins, Brian Thomas; Gu, Yu-Huan; Izawa, Yoshikane; del Zoppo, Gregory John

    2015-06-01

    Atrial fibrillation (AF) increases the risk and severity of thromboembolic stroke. Generally, antithrombotic agents increase the hemorrhagic risk of thromboembolic stroke. However, significant reductions in thromboembolism and intracerebral hemorrhage have been shown with the antithrombin dabigatran compared with warfarin. As thrombin has been implicated in microvessel injury during cerebral ischemia, we hypothesized that dabigatran decreases the risk of intracerebral hemorrhage by direct inhibition of the thrombin-mediated increase in cerebral endothelial cell permeability. Primary murine brain endothelial cells (mBECs) were exposed to murine thrombin before measuring permeability to 4-kDa fluorescein isothiocyanate-dextran. Thrombin increased mBEC permeability in a concentration-dependent manner, without significant endothelial cell death. Pretreatment of mBECs with dabigatran completely abrogated the effect of thrombin on permeability. Neither the expressions of the endothelial cell β1-integrins nor the tight junction protein claudin-5 were affected by thrombin exposure. Oxygen-glucose deprivation (OGD) also increased permeability; this effect was abrogated by treatment with dabigatran, as was the additive effect of thrombin and OGD on permeability. Taken together, these results indicate that dabigatran could contribute to a lower risk of intracerebral hemorrhage during embolism-associated ischemia from AF by protection of the microvessel permeability barrier from local thrombin challenge.

  9. Store-operated calcium entry and increased endothelial cell permeability.

    Science.gov (United States)

    Norwood, N; Moore, T M; Dean, D A; Bhattacharjee, R; Li, M; Stevens, T

    2000-11-01

    We hypothesized that myosin light chain kinase (MLCK) links calcium release to activation of store-operated calcium entry, which is important for control of the endothelial cell barrier. Acute inhibition of MLCK caused calcium release from inositol trisphosphate-sensitive calcium stores and prevented subsequent activation of store-operated calcium entry by thapsigargin, suggesting that MLCK serves as an important mechanism linking store depletion to activation of membrane calcium channels. Moreover, in voltage-clamped single rat pulmonary artery endothelial cells, thapsigargin activated an inward calcium current that was abolished by MLCK inhibition. F-actin disruption activated a calcium current, and F-actin stabilization eliminated the thapsigargin-induced current. Thapsigargin increased endothelial cell permeability in the presence, but not in the absence, of extracellular calcium, indicating the importance of calcium entry in decreasing barrier function. Although MLCK inhibition prevented thapsigargin from stimulating calcium entry, it did not prevent thapsigargin from increasing permeability. Rather, inhibition of MLCK activity increased permeability that was especially prominent in low extracellular calcium. In conclusion, MLCK links store depletion to activation of a store-operated calcium entry channel. However, inhibition of calcium entry by MLCK is not sufficient to prevent thapsigargin from increasing endothelial cell permeability.

  10. Effects of hypergravity on the angiogenic potential of endothelial cells

    NARCIS (Netherlands)

    Costa-Almeida, R. (Raquel); Carvalho, D.T.O. (Daniel T.O.); Ferreira, M.J.S. (Miguel J.S.); Aresta, G. (Guilherme); Gomes, M.E. (Manuela E.); Van Loon, J.J.W.A. (Jack J.W.A.); K. van der Heiden (Kim); Granja, P.L. (Pedro L.)

    2016-01-01

    textabstractAngiogenesis, the formation of blood vessels from pre-existing ones, is a key event in pathology, including cancer progression, but also in homeostasis and regeneration. As the phenotype of endothelial cells (ECs) is continuously regulated by local biomechanical forces, studying

  11. Comparison of Endothelial Cell Loss by Specular Microscopy ...

    African Journals Online (AJOL)

    life restored.[1,2] Providing Early Childhood Care and. Education (ECCE) to 95% of those who need it (95% coverage level) would avert more than 3.5 million disability‑adjusted life years per year globally.[2]. Some degree of endothelial cell loss is inevitable after any type of cataract surgery.[3,4]. The number and.

  12. Differentiation of mesenchymal stem cells from human amniotic fluid to vascular endothelial cells.

    Science.gov (United States)

    Tancharoen, Waleephan; Aungsuchawan, Sirinda; Pothacharoen, Peraphan; Markmee, Runchana; Narakornsak, Suteera; Kieodee, Junjira; Boonma, Nonglak; Tasuya, Witoon

    2017-03-01

    Endothelial dysfunction is a principle feature of vascular-related disease. Endothelial cells have been acquired for the purposes of the restoration of damaged tissue in therapeutic angiogenesis. However, their use is limited by expansion capacity and the small amount of cells that are obtained. Human amniotic fluid mesenchymal stem cells (hAF-MSCs) are considered an important source for vascular tissue engineering. In this study, hAF-MSCs were characterized and then induced in order to differentiate into the endothelial-like cells. Human amniotic fluid cells (hAFCs) were obtained from amniocentesis at the second trimester of gestation. The cells were characterized as mesenchymal stem cells by flow cytometry. The results showed that the cells were positive for mesenchymal stem cell markers CD44, CD73, CD90 and HLA-ABC, and negative for CD31, Amniotic fluid stem cells marker: CD117, anti-human fibroblasts, HLA-DR and hematopoietic differentiation markers CD34 and CD45. The hAF-MSCs were differentiated into endothelial cells under the induction of vascular endothelial growth factor (VEGF) and analyzed for the expression of the endothelial-specific markers and function. The expression of the endothelial-specific markers was determined by reverse transcriptase-quantitative PCR (RT-qPCR), while immunofluorescent analysis demonstrated that the induced hAF-MSCs expressed von Willebrand factor (vWF), vascular endothelial growth factor receptor 2 (VEGFR2), CD31 and endothelial nitric oxide synthase (eNOS). The network formation assay showed that the induced hAF-MSCs formed partial networks. All results indicated that hAF-MSCs have the potential to be differentiated into endothelial-like cells, while human amniotic fluid might be a suitable source of MSCs for vascularized tissue engineering. Copyright © 2016 Elsevier GmbH. All rights reserved.

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  18. File list: DNS.CDV.05.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  19. File list: Oth.CDV.10.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  1. File list: ALL.CDV.05.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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  5. File list: ALL.CDV.20.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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  7. File list: Oth.CDV.50.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  9. File list: Pol.CDV.50.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  10. File list: Unc.CDV.05.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  11. File list: Pol.CDV.20.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  12. File list: His.CDV.50.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  13. File list: His.CDV.20.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  14. File list: DNS.CDV.20.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  15. DMPD: Lipopolysaccharide signaling in endothelial cells. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 16357866 Lipopolysaccharide signaling in endothelial cells. Dauphinee SM, Karsan A.... Lab Invest. 2006 Jan;86(1):9-22. (.png) (.svg) (.html) (.csml) Show Lipopolysaccharide signaling in endothe...lial cells. PubmedID 16357866 Title Lipopolysaccharide signaling in endothelial cells. Authors Dauphinee SM,

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  1. File list: His.CDV.50.AllAg.Primary_endothelial_cells [Chip-atlas[Archive

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  13. File list: Unc.CDV.50.AllAg.Primary_endothelial_cells [Chip-atlas[Archive

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  14. File list: DNS.CDV.20.AllAg.Primary_endothelial_cells [Chip-atlas[Archive

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  15. File list: DNS.CDV.50.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.CDV.50.AllAg.Brachiocephalic_endothelial_cells hg19 DNase-seq Cardiovascular Brachiocephalic endot...helial cells http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.CDV.50.AllAg.Brachiocephalic_endothelial_cells.bed ...

  16. Epigenetic regulation of tumor endothelial cell anergy : Silencing of intercellular adhesion molecule-1 by histone modifications

    NARCIS (Netherlands)

    Hellebrekers, Debby M. E. I.; Castermans, Karolien; Vire, Emmanuelle; Dings, Ruud P. M.; Hoebers, Nicole T. H.; Mayos, Kevin H.; Egbrink, Mirjam G. A. Oude; Molema, Grietje; Fuks, Francois; Griffloen, Arjan W.

    2006-01-01

    Tumors can escape from immunity by repressing leukocyte adhesion molecule expression on tumor endothelial cells and by rendering endothelial cells unresponsive to inflammatory activation. This endothelial cell anergy is induced by angiogenic growth factors and results in reduced leukocyte-vessel

  17. Role of endothelial cell metabolism in vessel sprouting.

    Science.gov (United States)

    De Bock, Katrien; Georgiadou, Maria; Carmeliet, Peter

    2013-11-05

    Endothelial cells (ECs) are quiescent for years but can plastically switch to angiogenesis. Vascular sprouting relies on the coordinated activity of migrating tip cells at the forefront and proliferating stalk cells that elongate the sprout. Past studies have identified genetic signals that control vascular branching. Prominent are VEGF, activating tip cells, and Notch, which stimulates stalk cells. After the branch is formed and perfused, ECs become quiescent phalanx cells. Now, emerging evidence has accumulated indicating that ECs not only adapt their metabolism when switching from quiescence to sprouting but also that metabolism regulates vascular sprouting in parallel to the control by genetic signals. Copyright © 2013 Elsevier Inc. All rights reserved.

  18. Endocytosis of Red Blood Cell Microparticles by Pulmonary Endothelial Cells is Mediated By Rab5.

    Science.gov (United States)

    Kim, Young; Abplanalp, William A; Jung, Andrew D; Schuster, Rebecca M; Lentsch, Alex B; Gulbins, Erich; Caldwell, Charles C; Pritts, Timothy A

    2018-03-01

    Microparticles are submicron vesicles shed from aging erythrocytes as a characteristic feature of the red blood cell (RBC) storage lesion. Exposure of pulmonary endothelial cells to RBC-derived microparticles promotes an inflammatory response, but the mechanisms underlying microparticle-induced endothelial cell activation are poorly understood. In the present study, cultured murine lung endothelial cells (MLECs) were treated with microparticles isolated from aged murine packed RBCs or vehicle. Microparticle-treated cells demonstrated increased expression of the adhesion molecules ICAM and E-selectin, as well as the cytokine, IL-6. To identify mechanisms that mediate these effects of microparticles on MLECs, cells were treated with microparticles covalently bound to carboxyfluorescein succinimidyl ester (CFSE) and cellular uptake of microparticles was quantified via flow cytometry. Compared with controls, there was a greater proportion of CFSE-positive MLECs from 15 min up to 24 h, suggesting endocytosis of the microparticles by endothelial cells. Colocalization of microparticles with lysosomes was observed via immunofluorescence, indicating endocytosis and endolysosomal trafficking. This process was inhibited by endocytosis inhibitors. SiRNA knockdown of Rab5 signaling protein in endothelial cells resulted in impaired microparticle uptake as compared with nonsense siRNA-treated cells, as well as an attenuation of the inflammatory response to microparticle treatment. Taken together, these data suggest that endocytosis of RBC-derived microparticles by lung endothelial cells results in endothelial cell activation. This response seems to be mediated, in part, by the Rab5 signaling protein.

  19. Endothelial cell-initiated extravasation of cancer cells visualized in zebrafish

    OpenAIRE

    Kanada, Masamitsu; Zhang, Jinyan; Libo YAN; Sakurai, Takashi; Terakawa, Susumu

    2014-01-01

    The extravasation of cancer cells, a key step for distant metastasis, is thought to be initiated by disruption of the endothelial barrier by malignant cancer cells. An endothelial covering-type extravasation of cancer cells in addition to conventional cancer cell invasion-type extravasation was dynamically visualized in a zebrafish hematogenous metastasis model. The inhibition of VEGF-signaling impaired the invasion-type extravasation via inhibition of cancer cell polarization and motility re...

  20. Changes in Zn homeostasis during long term culture of primary endothelial cells and effects of Zn on endothelial cell senescence.

    Science.gov (United States)

    Malavolta, Marco; Costarelli, Laura; Giacconi, Robertina; Basso, Andrea; Piacenza, Francesco; Pierpaoli, Elisa; Provinciali, Mauro; Ogo, Ogo A; Ford, Dianne

    2017-12-01

    Endothelial cell senescence and Zn nutritional status influence cardiovascular disease. The influence of Zn appears dichotomous, hence it is imperative to understand the relationship with cellular senescence to improve knowledge about the molecular and cellular basis of the disease. Here we aimed to determine: 1) the impact of chronic exposure to a moderately high dose of Zn on senescence of endothelial cells; 2) the changes in Zn homeostasis during the lifespan of primary cultured endothelial cells; and 3) the susceptibility of proliferating and senescent endothelial cells to cell death after short term exposure to increasing doses of Zn and of the Zn chelator TPEN. Chronic exposure to Zn accelerated senescence and untreated cells at later passages, where doubling time had increased, displayed relocation of labile Zn and altered expression of genes involved in the response to Zn toxicity, including SLC30A1, SLC39A6, SLC30A5, SLC30A10 and metallothioneins, indicating that senescent cells have altered zinc homeostasis. Most Zn-dependent genes that were expressed differently between early and late passages were correlated with changes in the expression of anti-apoptotic genes. Short-term treatment with a high dose of Zn leads to cell death, but only in the population of cells at both earlier and later passages that had already entered senescence. In contrast, Zn depletion led to death of cells at earlier but not later passages, which suggests that there are sub-populations of senescent cells that are resistant to Zn depletion. This resistant senescent cell population may accumulate under conditions of Zn deficiency and contribute to vascular pathology. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Corneal endothelial cell density and morphology in normal Iranian eyes

    Directory of Open Access Journals (Sweden)

    Fallah Mohammad

    2006-03-01

    Full Text Available Abstract Background We describe corneal endothelial cell density and morphology in normal Iranian eyes and compare endothelial cell characteristics in the Iranian population with data available in the literature for American and Indian populations. Methods Specular microscopy was performed in 525 eyes of normal Iranian people aged 20 to 85 years old. The studied parameters including mean endothelial cell density (MCD, mean cell area (MCA and coefficient of variation (CV in cell area were analyzed in all of the 525 eyes. Results MCD was 1961 ± 457 cell/mm2 and MCA was 537.0 ± 137.4 μm2. There was no statistically significant difference in MCD, MCA and CV between genders (Student t-test, P = 0.85, P = 0.97 and P = 0.15 respectively. There was a statistically significant decrease in MCD with age (P r = -0.64. The rate of cell loss was 0.6% per year. There was also a statistically significant increase in MCA (P r = 0.56 and CV (P r = 0.30 from 20 to 85 years of age. Conclusion The first normative data for the endothelium of Iranian eyes seems to confirm that there are no differences in MCD, MCA and CV between genders. Nevertheless, the values obtained in Iranian eyes seem to be different to those reported by the literature in Indian and American populations.

  2. Insulin resistance in vascular endothelial cells promotes intestinal tumour formation

    DEFF Research Database (Denmark)

    Wang, X; Häring, M-F; Rathjen, Thomas

    2017-01-01

    did not change intestinal tumour number or size distribution on either a low or high-fat diet. We therefore asked whether cells in the tumour stroma might explain the association between tumour formation and insulin resistance. To this end, we generated Apc(Min/+) mice with loss of insulin receptors......The risk of several cancers, including colorectal cancer, is increased in patients with obesity and type 2 diabetes, conditions characterised by hyperinsulinaemia and insulin resistance. Because hyperinsulinaemia itself is an independent risk factor for cancer development, we examined tissue...... in vascular endothelial cells. Strikingly, these mice had 42% more intestinal tumours than controls, no change in tumour angiogenesis, but increased expression of vascular cell adhesion molecule-1 (VCAM-1) in primary culture of tumour endothelial cells. Insulin decreased VCAM-1 expression and leukocyte...

  3. Effect of cytokines on tumour cell-endothelial interactions.

    Science.gov (United States)

    Cohen, M C; Bereta, M; Bereta, J

    1997-01-01

    The adherence of tumour cells to microvascular endothelium is believed to be a necessary step in their migration to sites of metastasis. It has been proposed that this process occurs when cell surface molecules on tumour cells bind to complementary sites on endothelial cells. The expression of these endothelial-derived cell adhesion molecules appears to be modulated by cytokines, a broad class of protein mediators which play important roles in immune and inflammatory reactions. It has been found by ourselves and others that exposure of endothelium to some cytokines augments the adhesion of inflammatory cells as well as tumour cells in in vitro assays. We used a murine model consisting of P815 mastocytoma cells and microvascular endothelium and found that pretreatment of endothelial monolayers with TNF-alpha, IL-1, LPS or PMA augmented the number of tumour cells that attach in a dose-dependent fashion. FACS analysis showed that the change in binding was due to an increase in the expression of VCAM-1 on the surface of the endothelial cell. Methylxanthines (caffeine and theophylline) as well as "classical" calcium-mobilizing agents (ionomycin and thapsigargin) inhibited the expression of VCAM-1 in MME. We also studied the possible mechanisms of TNF-alpha signal transduction in endothelial cells. We examined the involvement of protein kinases in the TNF-alpha effect. Although we found that inhibitors of PKC could inhibit the TNF-alpha effect, our studies suggest that the "classical" PKC pathway is not completely responsible for signaling since TNF-alpha did not cause translocation of PKC to the cell membrane and its effect could not be completely mimicked by PMA. We also studied the effect of TGF-beta on the binding of tumour cells to endothelium. Exposure of endothelium to TGF-beta led to the inhibition of both basal and TNF-alpha enhanced binding of P815 cells. Inhibitors of G-proteins do not abolish TGF-beta action, and PKC and PKA activators elicit an opposite

  4. ANTIBODIES DEFINING RAT ENDOTHELIAL-CELLS - RECA-1, A PAN-ENDOTHELIAL CELL-SPECIFIC MONOCLONAL-ANTIBODY

    NARCIS (Netherlands)

    DUIJVESTIJN, AM; VANGOOR, H; KLATTER, F; MAJOOR, GD; VANBUSSEL, E; VRIESMAN, PJCV

    We have been searching for antibodies reactive with rat endothelial cells. Two monoclonal antibodies (mAb), named RECA-1 and RECA-2 were produced and tested in immunoperoxidase staining on frozen sections of various rat tissues. Staining patterns were compared to those obtained with the mAbs OX-2,

  5. Nylon-3 polymers that enable selective culture of endothelial cells.

    Science.gov (United States)

    Liu, Runhui; Chen, Xinyu; Gellman, Samuel H; Masters, Kristyn S

    2013-11-06

    Substrates that selectively encourage the growth of specific cell types are valuable for the engineering of complex tissues. Some cell-selective peptides have been identified from extracellular matrix proteins; these peptides have proven useful for biomaterials-based approaches to tissue repair or regeneration. However, there are very few examples of synthetic materials that display selectivity in supporting cell growth. We describe nylon-3 polymers that support in vitro culture of endothelial cells but do not support the culture of smooth muscle cells or fibroblasts. These materials may be promising for vascular biomaterials applications.

  6. An Important Method in the Investigation of Vascular Pathologies: Endothelial Cell Culture

    Directory of Open Access Journals (Sweden)

    Yusufhan Yazır

    2012-12-01

    Full Text Available Endothelial cells line the interior surface of blood vessels and form an interface between circulating blood in the lumen and the rest of the vessel wall. Endothelial cells are involved in many aspects of vascular biology, including barrier function, vasoconstriction, coagulation and inflamation. The endothelial cells in different organs have different functions and surface phenotype. These cells express prostoglandin-I2, platelet activating factor, collagen, endothelin-1, laminin, fibronectin and growth factors including platelet derived growth factor, fibroblast growth factor. İn the cell culture, cells can be isolated, maintened and proliferate in the laboratory conditions. The techniques of the cell culture have allowed scientists to use the cells in vitro for experimental studies, such as the production of vaccine, antibody and enzime, drug research, cell-cell interactions. Human umbilical vein endothelial cell is a good source for endothelial cell, because it is cheaper, easy to find and has the basic features of the normal endothelial cells.

  7. Culture and Characterization of Circulating Endothelial Progenitor Cells in Patients with Renal Cell Carcinoma.

    Science.gov (United States)

    Gu, Wenyu; Sun, Wei; Guo, Changcheng; Yan, Yang; Liu, Min; Yao, Xudong; Yang, Bin; Zheng, Junhua

    2015-07-01

    Although emerging evidence demonstrates increased circulating endothelial progenitor cells in patients with solid tumors, to our knowledge it is still unknown whether such cells can be cultured from patients with highly angiogenic renal cell carcinoma. We cultured and characterized circulating endothelial progenitor cells from patients with renal cell carcinoma. The circulating endothelial progenitor cell level (percent of CD45(-)CD34(+) VEGF-R2(+) cells in total peripheral blood mononuclear cells) was quantified in 47 patients with renal cell carcinoma and 40 healthy controls. Peripheral blood mononuclear cells were then isolated from 33 patients with renal cell carcinoma and 30 healthy controls to culture and characterize circulating endothelial progenitor cells. The circulating endothelial progenitor cell level was significantly higher in patients with renal cell carcinoma than in healthy controls (0.276% vs 0.086%, p cells first emerged significantly earlier in patient than in control preparations (6.72 vs 14.67 days, p culture success rate (87.8% vs 40.0% of participants) and the number of colonies (10.06 vs 1.83) were significantly greater for patients than for controls (each p cell level correlated positively with the number of patient colonies (r = 0.762, p Cells cultured from patients and controls showed a similar growth pattern, immunophenotype, ability to uptake Ac-LDL and bind lectin, and form capillary tubes in vitro. However, significantly more VEGF-R2(+) circulating endothelial progenitor cells were found in preparations from patients with renal cell carcinoma than from healthy controls (21.1% vs 13.4%, p cell colonies, a higher cell culture success rate and more colonies were found for patients with renal cell carcinoma than for healthy controls. Results indicate the important significance of VEGF-R2(+) circulating endothelial progenitors in patients with renal cell carcinoma. Copyright © 2015 American Urological Association Education and Research

  8. Methylene blue modulates adhesion molecule expression on microvascular endothelial cells.

    Science.gov (United States)

    Werner, Isabella; Guo, Fengwei; Stock, Ulrich A; Lupinski, Michèle; Meybohm, Patrick; Moritz, Anton; Beiras-Fernandez, Andres

    2014-08-01

    As methylene blue (MB) has been recently proposed to preserve blood pressure in case of vasoplegic syndrome and shock, an entity directly related to systemic inflammation, we aimed to elucidate the effect of MB on the expression of adhesion-molecules in endothelial-cells. Human microvascular endothelial-cells (HuMEC-1) were treated with 10, 30 or 60 µM MB for 30 min and 2 h each. Additionally, the treated HuMEC-1 were co-cultured with either human peripheral blood mononuclear cells (PBMCs) or Jurkat cells (human T-lymphocytes) for 2 h. HuMEC-1 were analyzed after MB treatment and after co-culture experiments for expression of different adhesion-molecules (ICAM-1, VCAM-1, L-selectin, E-selectin) via FACS measurement and western blot analysis. The supernatants of the experiments were analyzed with regard to the soluble forms of the adhesion molecules. We found that MB is able to modulate the expression of adhesion-molecules on EC. Administration of MB increases the expression of E-selectin and VCAM-1 depending on the dosage and time of exposure. ICAM-1 measurements provide evidence that different circulating blood cells can differently alter the adhesion-molecule expression on EC after MB exposure. Our results provide evidence regarding the immunomodulatory effect of MB upon endothelial-cells after inflammation.

  9. Endothelial progenitor cells in sudden sensorineural hearing loss.

    Science.gov (United States)

    Quaranta, Nicola; Ramunni, Alfonso; De Luca, Concetta; Brescia, Paola; Dambra, Porzia; De Tullio, Giacomina; Vacca, Angelo; Quaranta, Antonio

    2011-04-01

    Endothelial progenitor cells (EPCs) are a unique subtype of circulating cells with properties similar to those of embryonal angioblasts. They have the potential to proliferate and to differentiate into mature endothelial cells. EPCs are reduced in patients with vascular risk factors due to a decreased mobilization, an increased consumption at the site of damage or a reduced half-life. The results of this study confirm the existence of an endothelial dysfunction in patients with sudden sensorineural hearing loss (SSHL) and support the vascular involvement in the pathogenesis of the disease. The aim of this study was to evaluate the concentration of EPCs in patients affected by SSHL. Twenty-one patients affected by SSHL were evaluated. The number of EPCs was analyzed by flow cytometry analysis of peripheral blood CD34+KDR+CD133+ cells. Circulating levels of EPCs were significantly lower in SSHL patients compared with controls. In particular, CD34+KDR+ cells and CD34+CD133+KDR+ cells were significantly reduced (p < 0.05).

  10. Erectile dysfunction precedes other systemic vascular diseases due to incompetent cavernous endothelial cell-cell junctions.

    Science.gov (United States)

    Ryu, Ji-Kan; Jin, Hai-Rong; Yin, Guo Nan; Kwon, Mi-Hye; Song, Kang-Moon; Choi, Min Ji; Park, Jin-Mi; Das, Nando Dulal; Kwon, Ki-Dong; Batbold, Dulguun; Lee, Tack; Gao, Zhen Li; Kim, Kyu-Won; Kim, Woo Jean; Suh, Jun-Kyu

    2013-08-01

    Erectile dysfunction is often a harbinger of cardiovascular disease. We sought to gain mechanistic insight at the cellular and molecular levels into why erectile dysfunction precedes the clinical consequences of cardiovascular disease. Diabetes was induced by intraperitoneal streptozotocin injection in 8-week-old C57BL/6J mice. At 8 weeks after diabetes induction, we determined the expression of endothelial cell-cell junction proteins and vascular endothelial permeability in the penis, heart and hind limb by systemic injection of various vascular space markers (350 Da to 2,000 kDa) or by immunohistochemical staining with antibody to oxidized low density lipoprotein. We also investigated the effect of recombinant Ang1 protein on cavernous endothelial permeability. Alterations in the integrity of the endothelial cell-cell junction, including a decrease in endothelial cell-cell junction proteins and an increase in vascular permeability to fluorescent tracers or oxidized low density lipoprotein, were prominent in the cavernous tissue of diabetic mice. In contrast, no significant changes in endothelial cell-cell junction proteins or vascular permeability were noted in heart or hind limb tissue according to the diabetic condition. Intracavernous injection of Ang1 protein, an anti-permeability factor, significantly decreased cavernous endothelial permeability to oxidized low density lipoprotein by restoring endothelial cell-cell junction proteins in diabetic mice. The incompetent cavernous endothelial cell-cell junction in the diabetic condition provides an important clue to why erectile dysfunction is highly prevalent and often precedes other systemic vascular diseases. Copyright © 2013 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  11. Young's modulus of elasticity of Schlemm's canal endothelial cells.

    Science.gov (United States)

    Zeng, Dehong; Juzkiw, Taras; Read, A Thomas; Chan, Darren W-H; Glucksberg, Matthew R; Ethier, C Ross; Johnson, Mark

    2010-02-01

    Schlemm's canal (SC) endothelial cells are likely important in the physiology and pathophysiology of the aqueous drainage system of the eye, particularly in glaucoma. The mechanical stiffness of these cells determines, in part, the extent to which they can support a pressure gradient and thus can be used to place limits on the flow resistance that this layer can generate in the eye. However, little is known about the biomechanical properties of SC endothelial cells. Our goal in this study was to estimate the effective Young's modulus of elasticity of normal SC cells. To do so, we combined magnetic pulling cytometry of isolated cultured human SC cells with finite element modeling of the mechanical response of the cell to traction forces applied by adherent beads. Preliminary work showed that the immersion angles of beads attached to the SC cells had a major influence on bead response; therefore, we also measured bead immersion angle by confocal microscopy, using an empirical technique to correct for axial distortion of the confocal images. Our results showed that the upper bound for the effective Young's modulus of elasticity of the cultured SC cells examined in this study, in central, non-nuclear regions, ranged between 1,007 and 3,053 Pa, which is similar to, although somewhat larger than values that have been measured for other endothelial cell types. We compared these values to estimates of the modulus of primate SC cells in vivo, based on images of these cells under pressure loading, and found good agreement at low intraocular pressure (8-15 mm Hg). However, increasing intraocular pressure (22-30 mm Hg) appeared to cause a significant increase in the modulus of these cells. These moduli can be used to estimate the extent to which SC cells deform in response to the pressure drop across the inner wall endothelium and thereby estimate the extent to which they can generate outflow resistance.

  12. Upcyte® Microvascular Endothelial Cells Repopulate Decellularized Scaffold

    Science.gov (United States)

    Dally, Iris; Hartmann, Nadja; Münst, Bernhard; Braspenning, Joris; Walles, Heike

    2013-01-01

    A general problem in tissue engineering is the poor and insufficient blood supply to guarantee tissue cell survival as well as physiological tissue function. To address this limitation, we have developed an in vitro vascularization model in which a decellularized porcine small bowl segment, representing a capillary network within a collagen matrix (biological vascularized scaffold [BioVaSc]), is reseeded with microvascular endothelial cells (mvECs). However, since the supply of mvECs is limited, in general, and as these cells rapidly dedifferentiate, we have applied a novel technology, which allows the generation of large batches of quasi-primary cells with the ability to proliferate, whilst maintaining their differentiated functionality. These so called upcyte mvECs grew for an additional 15 population doublings (PDs) compared to primary cells. Upcyte mvECs retained endothelial characteristics, such as von Willebrandt Factor (vWF), CD31 and endothelial nitric oxide synthase (eNOS) expression, as well as positive Ulex europaeus agglutinin I staining. Upcyte mvECs also retained biological functionality such as tube formation, cell migration, and low density lipoprotein (LDL) uptake, which were still evident after PD27. Initial experiments using MTT and Live/Dead staining indicate that upcyte mvECs repopulate the BioVaSc Scaffold. As with conventional cultures, these cells also express key endothelial molecules (vWF, CD31, and eNOS) in a custom-made bioreactor system even after a prolonged period of 14 days. The combination of upcyte mvECs and the BioVaSc represents a novel and promising approach toward vascularizing bioreactor models which can better reflect organs, such as the liver. PMID:22799502

  13. Endothelial progenitor cells physiology and metabolic plasticity in brain angiogenesis and blood-brain barrier modeling

    Directory of Open Access Journals (Sweden)

    Natalia Malinovskaya

    2016-12-01

    Full Text Available Currently, there is a considerable interest to the assessment of blood-brain barrier (BBB development as a part of cerebral angiogenesis developmental program. Embryonic and adult angiogenesis in the brain is governed by the coordinated activity of endothelial progenitor cells, brain microvascular endothelial cells, and non-endothelial cells contributing to the establishment of the BBB (pericytes, astrocytes, neurons. Metabolic and functional plasticity of endothelial progenitor cells controls their timely recruitment, precise homing to the brain microvessels, and efficient support of brain angiogenesis. Deciphering endothelial progenitor cells physiology would provide novel engineering approaches to establish adequate microfluidically-supported BBB models and brain microphysiological systems for translational studies.

  14. Hold Me, but Not Too Tight-Endothelial Cell-Cell Junctions in Angiogenesis.

    Science.gov (United States)

    Szymborska, Anna; Gerhardt, Holger

    2017-09-08

    Endothelial cell-cell junctions must perform seemingly incompatible tasks during vascular development-providing stable connections that prevent leakage, while allowing dynamic cellular rearrangements during sprouting, anastomosis, lumen formation, and functional remodeling of the vascular network. This review aims to highlight recent insights into the molecular mechanisms governing endothelial cell-cell adhesion in the context of vascular development. Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.

  15. Antiproliferative Effects of Drugs on Endothelial and Osteoblastic Cells and Altered Release of Angioregulatory Mediators by Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Hilde Kvestad

    2014-01-01

    Full Text Available The combined use of the histone deacetylase inhibitor valproic acid (VPA, the retinoic acid receptor-α agonist all-trans retinoic acid (ATRA, and the deoxyribonucleic acid polymerase-α inhibitor cytarabine (Ara-C is now considered for disease-stabilizing treatment of acute myeloid leukemia (AML. Leukemogenesis and leukemia cell chemoresistance seem to be supported by neighbouring stromal cells in the bone marrow, and we have therefore investigated the effects of these drugs on primary human endothelial cells and the osteoblastic Cal72 cell line. The results show that VPA and Ara-C have antiproliferative effects, and the antiproliferative/cytotoxic effect of Ara-C was seen at low concentrations corresponding to serum levels found during low-dose in vivo treatment. Furthermore, in functional assays of endothelial migration and tube formation VPA elicited an antiangiogenic effect, whereas ATRA elicited a proangiogenic effect. Finally, VPA and ATRA altered the endothelial cell release of angiogenic mediators; ATRA increased levels of CXCL8, PDGF-AA, and VEGF-D, while VPA decreased VEGF-D and PDGF-AA/BB levels and both drugs reduced MMP-2 levels. Several of these mediators can enhance AML cell proliferation and/or are involved in AML-induced bone marrow angiogenesis, and direct pharmacological effects on stromal cells may thus indirectly contribute to the overall antileukemic activity of this triple drug combination.

  16. Endothelial cell senescence with aging in healthy humans: prevention by habitual exercise and relation to vascular endothelial function.

    Science.gov (United States)

    Rossman, Matthew J; Kaplon, Rachelle E; Hill, Sierra D; McNamara, Molly N; Santos-Parker, Jessica R; Pierce, Gary L; Seals, Douglas R; Donato, Anthony J

    2017-11-01

    Cellular senescence is emerging as a key mechanism of age-related vascular endothelial dysfunction, but evidence in healthy humans is lacking. Moreover, the influence of lifestyle factors such as habitual exercise on endothelial cell (EC) senescence is unknown. We tested the hypothesis that EC senescence increases with sedentary, but not physically active, aging and is associated with vascular endothelial dysfunction. Protein expression (quantitative immunofluorescence) of p53, a transcription factor related to increased cellular senescence, and the cyclin-dependent kinase inhibitors p21 and p16 were 116%, 119%, and 128% greater (all P 0.05) in venous ECs from older exercising adults (57 ± 1 yr, n = 13). Furthermore, venous EC protein levels of p53 ( r  = -0.49, P = 0.003), p21 ( r  = -0.38, P = 0.03), and p16 ( r  = -0.58, P = 0.002) were inversely associated with vascular endothelial function (brachial artery flow-mediated dilation). Similarly, protein expression of p53 and p21 was 26% and 23% higher (both P 0.05) in older habitually exercising adults (59 ± 1 yr, n = 14). These data indicate that EC senescence is associated with sedentary aging and is linked to endothelial dysfunction. Moreover, these data suggest that prevention of EC senescence may be one mechanism by which aerobic exercise protects against endothelial dysfunction with age. NEW & NOTEWORTHY Our study provides novel evidence in humans of increased endothelial cell senescence with sedentary aging, which is associated with impaired vascular endothelial function. Furthermore, our data suggest an absence of age-related increases in endothelial cell senescence in older exercising adults, which is linked with preserved vascular endothelial function. Copyright © 2017 the American Physiological Society.

  17. Coniferyl Aldehyde Ameliorates Radiation Intestine Injury via Endothelial Cell Survival

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Ye Ji; Jung, Myung Gu; Lee, Yoonjin; Lee, Haejune [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Lee, Yunsil [Ewha Woman' s Univ., Seoul (Korea, Republic of); Ko, Younggyu [Korea Univ., Seoul (Korea, Republic of)

    2014-05-15

    Cancer treatments related gastrointestinal toxicity has also been recognized as a significant economic burden. Especially, extensive apoptosis of microvascular endothelial cell of the lamina propria is the primary lesion initiating intestinal radiation damage after abdominal radiation therapy. Coniferyl aldehyde (CA) is phenolic compounds isolated from cork stoppers, and one of the major pyrolysis products of lignin. Shi H. was support for the empirical use of CA as a medicinal food for cardiovascular diseases. CA has positive effect in broad way but there is no consequence in radiation induced intestine damage. Here, we investigate effect of CA on small intestine after abdominal IR to mice in this study. In this study, CA increased the survival rate in C3H mice against 13.5 Gy abdominal IR. We found CA protects small intestine via preventing endothelial cell apoptosis and enhancing their angiogenic activity. CA also showed protective effect on crypt cell survival. Endothelial cell survival may affect crypt cell protection against IR. From this data, we concluded that CA is effective for protection against abdominal radiation injury. CA could ameliorate side-effect of radiation therapy.

  18. Protection of Candida parapsilosis from neutrophil killing through internalization by human endothelial cells.

    Science.gov (United States)

    Glass, Kyle A; Longley, Sarah J; Bliss, Joseph M; Shaw, Sunil K

    2015-01-01

    Candida parapsilosis is a fungal pathogen that is associated with hematogenously disseminated disease in premature neonates, acutely ill or immunocompromised patients. In cell culture, C. parapsilosis cells are actively and avidly endocytosed by endothelial cells via actin polymerization mediated by N-WASP. Here we present evidence that C. parapsilosis that were internalized by endothelial cells remained alive, and avoided being acidified or otherwise damaged via the host cell. Internalized fungal cells reproduced intracellularly and eventually burst out of the host endothelial cell. When neutrophils were added to endothelium and C. parapsilosis, they patrolled the endothelial surface and efficiently killed most adherent fungal cells prior to endocytosis. But after endocytosis by endothelial cells, internalized fungal cells evaded neutrophil killing. Silencing endothelial N-WASP blocked endocytosis of C. parapsilosis and left fungal cells stranded on the cell surface, where they were susceptible to neutrophil killing. These observations suggest that for C. parapsilosis to escape from the bloodstream, fungi may adhere to and be internalized by endothelial cells before being confronted and phagocytosed by a patrolling leukocyte. Once internalized by endothelial cells, C. parapsilosis may safely replicate to cause further rounds of infection. Immunosurveillance of the intravascular lumen by leukocytes crawling on the endothelial surface and rapid killing of adherent yeast may play a major role in controlling C. parapsilosis dissemination and infected endothelial cells may be a significant reservoir for fungal persistence.

  19. Pharmacologically active microcarriers for endothelial progenitor cell support and survival.

    Science.gov (United States)

    Musilli, Claudia; Karam, Jean-Pierre; Paccosi, Sara; Muscari, Claudio; Mugelli, Alessandro; Montero-Menei, Claudia N; Parenti, Astrid

    2012-08-01

    The regenerative potential of endothelial progenitor cell (EPC)-based therapies is limited due to poor cell viability and minimal retention following application. Neovascularization can be improved by means of scaffolds supporting EPCs. The aim of the present study was to investigate whether human early EPCs (eEPCs) could be efficiently cultured on pharmacologically active microcarriers (PAMs), made with poly(d,l-lactic-coglycolic acid) and coated with adhesion/extracellular matrix molecules. They may serve as a support for stem cells and may be used as cell carriers providing a controlled delivery of active protein such as the angiogenic factor, vascular endothelial growth factor-A (VEGF-A). eEPC adhesion to fibronectin-coated PAMs (FN-PAMs) was assessed by means of microscopic evaluation and by means of Alamar blue assay. Phospho ERK(1/2) and PARP-1 expression was measured by means of Western blot to assess the survival effects of FN-PAMs releasing VEGF-A (FN-VEGF-PAMs). The Alamar blue assay or a modified Boyden chamber assay was employed to assess proliferative or migratory capacity, respectively. Our data indicate that eEPCs were able to adhere to empty FN-PAMs within a few hours. FN-VEGF-PAMs increased the ability of eEPCs to adhere to them and strongly supported endothelial-like phenotype and cell survival. Moreover, the release of VEGF-A by FN-PAMs stimulated in vitro HUVEC migration and proliferation. These data strongly support the use of PAMs for supporting eEPC growth and survival and for stimulating resident mature human endothelial cells. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. The secretome of endothelial progenitor cells promotes brain endothelial cell activity through PI3-kinase and MAP-kinase.

    Science.gov (United States)

    Di Santo, Stefano; Seiler, Stefanie; Fuchs, Anna-Lena; Staudigl, Jennifer; Widmer, Hans Rudolf

    2014-01-01

    Angiogenesis and vascular remodelling are crucial events in tissue repair mechanisms promoted by cell transplantation. Current evidence underscores the importance of the soluble factors secreted by stem cells in tissue regeneration. In the present study we investigated the effects of paracrine factors derived from cultured endothelial progenitor cells (EPC) on rat brain endothelial cell properties and addressed the signaling pathways involved. Endothelial cells derived from rat brain (rBCEC4) were incubated with EPC-derived conditioned medium (EPC-CM). The angiogenic response of rBCEC4 to EPC-CM was assessed as effect on cell number, migration and tubular network formation. In addition, we have compared the outcome of the in vitro experiments with the effects on capillary sprouting from rat aortic rings. The specific PI3K/AKT inhibitor LY294002 and the MEK/ERK inhibitor PD98059 were used to study the involvement of these two signaling pathways in the transduction of the angiogenic effects of EPC-CM. Viable cell number, migration and tubule network formation were significantly augmented upon incubation with EPC-CM. Similar findings were observed for aortic ring outgrowth with significantly longer sprouts. The EPC-CM-induced activities were significantly reduced by the blockage of the PI3K/AKT and MEK/ERK signaling pathways. Similarly to the outcome of the rBCEC4 experiments, inhibition of the PI3K/AKT and MEK/ERK pathways significantly interfered with capillary sprouting induced by EPC-CM. The present study demonstrates that EPC-derived paracrine factors substantially promote the angiogenic response of brain microvascular endothelial cells. In addition, our findings identified the PI3K/AKT and MEK/ERK pathways to play a central role in mediating these effects.

  1. Leptin-induced transphosphorylation of vascular endothelial growth factor receptor increases Notch and stimulates endothelial cell angiogenic transformation.

    Science.gov (United States)

    Lanier, Viola; Gillespie, Corey; Leffers, Merle; Daley-Brown, Danielle; Milner, Joy; Lipsey, Crystal; Webb, Nia; Anderson, Leonard M; Newman, Gale; Waltenberger, Johannes; Gonzalez-Perez, Ruben Rene

    2016-10-01

    Leptin increases vascular endothelial growth factor (VEGF), VEGF receptor-2 (VEGFR-2), and Notch expression in cancer cells, and transphosphorylates VEGFR-2 in endothelial cells. However, the mechanisms involved in leptin's actions in endothelial cells are not completely known. Here we investigated whether a leptin-VEGFR-Notch axis is involved in these leptin's actions. To this end, human umbilical vein and porcine aortic endothelial cells (wild type and genetically modified to overexpress VEGFR-1 or -2) were cultured in the absence of VEGF and treated with leptin and inhibitors of Notch (gamma-secretase inhibitors: DAPT and S2188, and silencing RNA), VEGFR (kinase inhibitor: SU5416, and silencing RNA) and leptin receptor, OB-R (pegylated leptin peptide receptor antagonist 2: PEG-LPrA2). Interestingly, in the absence of VEGF, leptin induced the expression of several components of Notch signaling pathway in endothelial cells. Inhibition of VEGFR and Notch signaling significantly decreased leptin-induced S-phase progression, proliferation, and tube formation in endothelial cells. Moreover, leptin/OB-R induced transphosphorylation of VEGFR-1 and VEGFR-2 was essential for leptin's effects. These results unveil for the first time a novel mechanism by which leptin could induce angiogenic features via upregulation/trans-activation of VEGFR and downstream expression/activation of Notch in endothelial cells. Thus, high levels of leptin found in overweight and obese patients might lead to increased angiogenesis by activating VEGFR-Notch signaling crosstalk in endothelial cells. These observations might be highly relevant for obese patients with cancer, where leptin/VEGFR/Notch crosstalk could play an important role in cancer growth, and could be a new target for the control of tumor angiogenesis. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Effect of Mitomycin-C augmented trabeculectomy on corneal endothelial cells

    Directory of Open Access Journals (Sweden)

    Reza Zarei

    2015-01-01

    Conclusion: MMC application in trabeculectomy seems to cause a small but significant corneal endothelial loss. Most of the damage occurs intraoperatively, or in the early postoperative period, however progressive endothelial cell loss is not a major concern.

  3. Stability of vision, topography, and endothelial cell density from 1 year to 2 years after deep lamellar endothelial keratoplasty surgery.

    Science.gov (United States)

    Ousley, Paula J; Terry, Mark A

    2005-01-01

    To evaluate whether the visual, topographic, and endothelial cell count results observed 1 year after deep lamellar endothelial keratoplasty (DLEK) surgery remain stable up to 2 years after surgery. Prospective, noncomparative, interventional case series. Twenty eyes of 20 patients with corneal edema from Fuchs' endothelial dystrophy. Deep lamellar endothelial keratoplasty endothelial replacement surgery, with a 9.0-mm or 9.5-mm scleral access incision and a specialized intrastromal trephine, was performed. Snellen visual acuities, corneal topography, and endothelial cell counts were prospectively measured preoperatively and 1 year and 2 years after DLEK. Uncorrected and best spectacle-corrected visual acuity (BSCVA), refractive and topographic astigmatism, mean corneal curvature, topographic regularity and symmetry, and endothelial cell density. At 1 year postoperatively, BSCVA averaged 20/50 (range, 20/25-20/200), spherical equivalents (SE) averaged -0.194+/-1.521 diopters (D), manifest refraction (MR) astigmatism averaged 2.04+/-1.05 D (range, 0.0-4.0 D), topographic astigmatism averaged 2.3+/-1.1 D, mean corneal curvature was 43.2+/-1.8 D, the surface regularity index (SRI) averaged 1.16+/-0.41, and the surface asymmetry index (SAI) averaged 1.05+/-1.09. At 2 years postoperatively, BSCVA averaged 20/48 (range, 20/25-20/200), SE averaged -0.369+/-1.267 D, MR astigmatism averaged 1.76+/-0.66 D (range, 0.75-3.0 D), topographic astigmatism averaged 2.4+/-1.1 D, mean corneal curvature was 43.6+/-1.8 D, the SRI averaged 1.13+/-0.44, and the SAI averaged 0.76+/-0.59. There was no significant change in visual or topographic parameters between 1 year and 2 years postoperatively (P>0.05). Endothelial cell counts averaged 2335+/-468 cells/mm(2) at 1 year and 2151+/-457 cells/mm(2) at 2 years postoperatively (P = 0.041). Deep lamellar endothelial keratoplasty provides stable refractions, corneal topography, and endothelial cell densities as long as 2 years after surgery

  4. Mechanical cell-matrix feedback explains pairwise and collective endothelial cell behavior in vitro.

    Directory of Open Access Journals (Sweden)

    René F M van Oers

    2014-08-01

    Full Text Available In vitro cultures of endothelial cells are a widely used model system of the collective behavior of endothelial cells during vasculogenesis and angiogenesis. When seeded in an extracellular matrix, endothelial cells can form blood vessel-like structures, including vascular networks and sprouts. Endothelial morphogenesis depends on a large number of chemical and mechanical factors, including the compliancy of the extracellular matrix, the available growth factors, the adhesion of cells to the extracellular matrix, cell-cell signaling, etc. Although various computational models have been proposed to explain the role of each of these biochemical and biomechanical effects, the understanding of the mechanisms underlying in vitro angiogenesis is still incomplete. Most explanations focus on predicting the whole vascular network or sprout from the underlying cell behavior, and do not check if the same model also correctly captures the intermediate scale: the pairwise cell-cell interactions or single cell responses to ECM mechanics. Here we show, using a hybrid cellular Potts and finite element computational model, that a single set of biologically plausible rules describing (a the contractile forces that endothelial cells exert on the ECM, (b the resulting strains in the extracellular matrix, and (c the cellular response to the strains, suffices for reproducing the behavior of individual endothelial cells and the interactions of endothelial cell pairs in compliant matrices. With the same set of rules, the model also reproduces network formation from scattered cells, and sprouting from endothelial spheroids. Combining the present mechanical model with aspects of previously proposed mechanical and chemical models may lead to a more complete understanding of in vitro angiogenesis.

  5. Adenosine formation in contracting primary rat skeletal muscle cells and endothelial cells in culture

    DEFF Research Database (Denmark)

    Hellsten, Ylva; Frandsen, Ulrik

    1997-01-01

    1. The present study examined the capacity for adenosine formation, uptake and metabolism in contracting primary rat muscle cells and in microvascular endothelial cells in culture. 2. Strong and moderate electrical simulation of skeletal muscle cells led to a significantly greater increase...... in the extracellular adenosine concentration (421 +/- 91 and 235 +/- 30 nmol (g protein)-1, respectively; P muscle cells (161 +/- 20 nmol (g protein)-1). The ATP concentration was lower (18%; P muscle cells....... 3. Addition of microvascular endothelial cells to the cultured skeletal muscle cells enhanced the contraction-induced accumulation of extracellular adenosine (P cells in culture alone did not cause extracellular accumulation of adenosine. 4. Skeletal muscle cells were...

  6. A role for activated endothelial cells in red blood cell clearance: implications for vasopathology

    DEFF Research Database (Denmark)

    Fens, Marcel H A M; van Wijk, Richard; Andringa, Grietje

    2012-01-01

    Background Phosphatidylserine exposure by red blood cells is acknowledged as a signal that initiates phagocytic removal of the cells from the circulation. Several disorders and conditions are known to induce phosphatidylserine exposure. Removal of phosphatidylserine-exposing red blood cells gener...... cells play a role in red blood cell clearance in vivo. Significant erythrophagocytosis can induce endothelial cell loss, which may contribute to vasopathological effects as seen, for instance, in sickle cell disease.......Background Phosphatidylserine exposure by red blood cells is acknowledged as a signal that initiates phagocytic removal of the cells from the circulation. Several disorders and conditions are known to induce phosphatidylserine exposure. Removal of phosphatidylserine-exposing red blood cells...... generally occurs by macrophages in the spleen and liver. Previously, however, we have shown that endothelial cells are also capable of erythrophagocytosis. Key players in the erythrophagocytosis by endothelial cells appeared to be lactadherin and αv-integrin. Phagocytosis via the phosphatidylserine...

  7. Acetylbritannilactone Modulates Vascular Endothelial Growth Factor Signaling and Regulates Angiogenesis in Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Jingshan Zhao

    Full Text Available The present study was conducted to determine the effects of 1-O-acetylbritannilactone (ABL, a compound extracted from Inula britannica L., on vascular endothelial growth factor (VEGF signaling and angiogenesis in endothelial cells (ECs. We showed that ABL promotes VEGF-induced cell proliferation, growth, migration, and tube formation in cultured human ECs. Furthermore, the modulatory effect of ABL on VEGF-induced Akt, MAPK p42/44, and p38 phosphorylation, as well as on upstream VEGFR-2 phosphorylation, were associated with VEGF-dependent Matrigel angiogenesis in vivo. In addition, animals treated with ABL (26 mg/kg/day recovered blood flow significantly earlier than control animals, suggesting that ABL affects ischemia-mediated angiogenesis and arteriogenesis in vivo. Finally, we demonstrated that ABL strongly reduced the levels of VEGFR-2 on the cell surface, enhanced VEGFR-2 endocytosis, which consistent with inhibited VE-cadherin, a negative regulator of VEGF signaling associated with VEGFR-2 complex formation, but did not alter VE-cadherin or VEGFR-2 expression in ECs. Our results suggest that ABL may serve as a novel therapeutic intervention for various cardiovascular diseases, including chronic ischemia, by regulating VEGF signaling and modulating angiogenesis.

  8. Acetylbritannilactone Modulates Vascular Endothelial Growth Factor Signaling and Regulates Angiogenesis in Endothelial Cells.

    Science.gov (United States)

    Zhao, Jingshan; Niu, Honglin; Li, Aiying; Nie, Lei

    2016-01-01

    The present study was conducted to determine the effects of 1-O-acetylbritannilactone (ABL), a compound extracted from Inula britannica L., on vascular endothelial growth factor (VEGF) signaling and angiogenesis in endothelial cells (ECs). We showed that ABL promotes VEGF-induced cell proliferation, growth, migration, and tube formation in cultured human ECs. Furthermore, the modulatory effect of ABL on VEGF-induced Akt, MAPK p42/44, and p38 phosphorylation, as well as on upstream VEGFR-2 phosphorylation, were associated with VEGF-dependent Matrigel angiogenesis in vivo. In addition, animals treated with ABL (26 mg/kg/day) recovered blood flow significantly earlier than control animals, suggesting that ABL affects ischemia-mediated angiogenesis and arteriogenesis in vivo. Finally, we demonstrated that ABL strongly reduced the levels of VEGFR-2 on the cell surface, enhanced VEGFR-2 endocytosis, which consistent with inhibited VE-cadherin, a negative regulator of VEGF signaling associated with VEGFR-2 complex formation, but did not alter VE-cadherin or VEGFR-2 expression in ECs. Our results suggest that ABL may serve as a novel therapeutic intervention for various cardiovascular diseases, including chronic ischemia, by regulating VEGF signaling and modulating angiogenesis.

  9. Stem cell-derived vascular endothelial cells and their potential application in regenerative medicine

    Science.gov (United States)

    Although a 'vascular stem cell' population has not been identified or generated, vascular endothelial and mural cells (smooth muscle cells and pericytes) can be derived from currently known pluripotent stem cell sources, including human embryonic stem cells and induced pluripotent stem cells. We rev...

  10. The Glycoprofile Patterns of Endothelial Cells in Usual Interstitial Pneumonia

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    A Barkhordari

    2014-09-01

    Full Text Available [THIS ARTICLE HAS BEEN RETRACTED FOR DUPLICATE PUBLICATION] Background: The pathological classification of cryptogenic fibrosing alveolitis has been a matter of debate and controversy for histopathologists. Objective: To identify and specify the glycotypes of capillary endothelial cells in usual interstitial pneumonia (UIP compared to those found in normal tissue. Methods: Sections of formalin-fixed, paraffin-embedded blocks from 16 cases of UIP were studied by lectin histochemistry with a panel of 27 biotinylated lectins and an avidin-peroxidase revealing system. Results: High expression of several classes of glycan was seen de novo in capillary endothelial cells from patients with UIP including small complex and bi/tri-antennary bisected complex N-linked sequences bolund by Concanavalin A and erythro-phytohemagglutinin, respectively, GalNAca1 residues bound by Helix pomatia and Maclura pomifera agglutinins, and L-fucosylated derivatives of type II glycan chains recognized by Ulex europaeus agglutinin-I. Glycans bound by agglutinins from Lycopersicon esculentum (β1,4GlcNAc and Wisteria floribunda (GalNAc as well as GlcNAc oligomers bound by Phytolacca americana and succinylated Wheat Germ agglutinin were also seen in the capillary endothelial cells of UIP. In contrast, L-fucosylated derivatives of type I glycan chains were absent in cells from cases of UIP when Anguilla anguilla agglutinin was applied, unlike the situation in normal tissue. Conclusion: These results may indicate existence of two distinct populations of endothelial cell in UIP with markedly different patterns of glycosylation, reflecting a pattern of differentiation and angiogenesis, which is not detectable morphologically.

  11. Cellular Cytotoxicity of Antiglaucoma Drugs in Cultured Corneal Endothelial Cells

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    Kwou-Yeung Wu

    2007-03-01

    Full Text Available In this study, the various antiglaucoma drugs including betaxolol, timolol, levobunolol, carteolol, brimonidine, dipivefrin, dorzolamide, brinzolamide, latanoprost, unoprostone, and pilocarpine were used to investigate the effects of cellular cytotoxicity in cultured bovine corneal endothelial cells. After exposure to the drugs in three dilutions, 1/100, 1/1,000, and 1/10,000, for 100 minutes, cells were estimated based on the release assay of lactate dehydrogenase (LDH enzyme. It was found that cellular LDH was significantly released in the medium only at 1/100th dilution of betaxolol, brimonidine, dorzolamide, dipivefrin, latanoprost and unoprostone to 130%, 123%, 145%, 157%, 128% and 237%, respectively, compared with controls upon exposure to drugs for 100 minutes. Moreover, benzalkonium chloride preservative at the concentrations ranging from 0.001 to 0.00001mg/mL did not affect cellular LDH release in bovine corneal endothelial cells. These results indicate that high concentrations of antiglaucoma drugs may induce cytotoxicity in corneal endothelial cells.

  12. Extended incubation times improve corneal endothelial cell transplantation success.

    Science.gov (United States)

    Insler, M S; Lopez, J G

    1991-05-01

    To investigate the ability of extended incubation times to improve the success of endothelial cell transplantation, eight human donor corneas were denuded of their native endothelium, seeded twice during a 1-hr interval with a suspension of cultured infant human corneal endothelial cells, and then incubated for 144 hr under standard conditions. Subsequently the corneas were transplanted into African green monkeys using routine penetrating keratoplasty techniques. Rotational autografts and corneas devoid of endothelial cells served as controls. The seeded corneas appeared hazy at the time of surgery (mean pachymetry 48 hr postoperatively, 0.794 mm). Six corneas (75%) subsequently cleared, yielding a mean corneal thickness of 0.541 +/- 0.040 and 0.554 +/- 0.040 at 6 and 12 postoperative months, respectively. All control eyes showed advanced edema (thickness, greater than 1.0 mm) and developed extensive neovascularization. Clinically, the extended postseeding incubation corneas were observed to clear more rapidly and stabilize their thickness earlier than corneas incubated for only 24-48 hr. Scanning electron microscopy of extended postseeding incubation corneas revealed an intact monolayer of contact-inhibited cells with the hexagonal mosaic typical of corneal endothelium in vivo and improved intercellular contact compared with corneas incubated for only 24-48 hr.

  13. Antiproliferative effect of elevated glucose in human microvascular endothelial cells

    Science.gov (United States)

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

    1998-01-01

    Diabetic microangiopathy has been implicated as a fundamental feature of the pathological complications of diabetes including retinopathy, neuropathy, and diabetic foot ulceration. However, previous studies devoted to examining the deleterious effects of elevated glucose on the endothelium have been performed largely in primary cultured cells of macrovessel origin. Difficulty in the harvesting and maintenance of microvascular endothelial cells in culture have hindered the study of this relevant population. Therefore, the objective of this study was to characterize the effect of elevated glucose on the proliferation and involved signaling pathways of an immortalized human dermal microvascular endothelial cell line (HMEC-1) that possess similar characteristics to their in vivo counterparts. Human dermal microvascular endothelial cells (HMEC-1) were grown in the presence of normal (5 mM) or high D-glucose (20 mM) for 14 days. The proliferative response of HMEC-1 was compared under these conditions as well as the cAMP and PKC pathways by in vitro assays. Elevated glucose significantly inhibited (P pathophysiology associated with diabetic microangiopathy.

  14. Endothelial progenitor cell subsets and preeclampsia: Findings and controversies

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    Armin Attar

    2017-10-01

    Full Text Available Vascular remodeling is an essential component of gestation. Endothelial progenitor cells (EPCs play an important role in the regulation of vascular homeostasis. The results of studies measuring the number of EPCs in normal pregnancies and in preeclampsia have been highly controversial or even contradictory because of some variations in technical issues and different methodologies enumerating three distinct subsets of EPCs: circulating angiogenic cells (CAC, colony forming unit endothelial cells (CFU-ECs, and endothelial colony-forming cells (ECFCs. In general, most studies have shown an increase in the number of CACs in the maternal circulation with a progression in the gestational age in normal pregnancies, while functional capacities measured by CFU-ECs and ECFCs remain intact. In the case of preeclampsia, mobilization of CACs and ECFCs occurs in the peripheral blood of pregnant women, but the functional capacities shown by culture of the derived colony-forming assays (CFU-EC and ECFC assays are altered. Furthermore, the number of all EPC subsets will be reduced in umbilical cord blood in the case of preeclampsia. As EPCs play an important role in the homeostasis of vascular networks, the difference in their frequency and functionality in normal pregnancies and those with preeclampsia can be expected. In this review, there was an attempt to provide a justification for these controversies.

  15. Human Brain Microvascular Endothelial Cells and Umbilical Vein Endothelial Cells Differentially Facilitate Leukocyte Recruitment and Utilize Chemokines for T Cell Migration

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    Shumei Man

    2008-01-01

    Full Text Available Endothelial cells that functionally express blood brain barrier (BBB properties are useful surrogates for studying leukocyte-endothelial cell interactions at the BBB. In this study, we compared two different endothelial cellular models: transfected human brain microvascular endothelial cells (THBMECs and human umbilical vein endothelial cells (HUVECs. With each grow under optimal conditions, confluent THBMEC cultures showed continuous occludin and ZO-1 immunoreactivity, while HUVEC cultures exhibited punctate ZO-1 expression at sites of cell-cell contact only. Confluent THBMEC cultures on 24-well collagen-coated transwell inserts had significantly higher transendothelial electrical resistance (TEER and lower solute permeability than HUVECs. Confluent THBMECs were more restrictive for mononuclear cell migration than HUVECs. Only THBMECs utilized abluminal CCL5 to facilitate T-lymphocyte migration in vitro although both THBMECs and HUVECs employed CCL3 to facilitate T cell migration. These data establish baseline conditions for using THBMECs to develop in vitro BBB models for studying leukocyte-endothelial interactions during neuroinflammation.

  16. Conversion of vascular endothelial cells into multipotent stem-like cells

    Science.gov (United States)

    Medici, Damian; Shore, Eileen M.; Lounev, Vitali Y.; Kaplan, Frederick S.; Kalluri, Raghu; Olsen, Bjorn R.

    2011-01-01

    Mesenchymal stem cells can give rise to several cell types, but variations depending on isolation method and tissue source have led to controversies about their usefulness in clinical medicine. Here we show that vascular endothelial cells can transform into multipotent stem-like cells by an ALK2 receptor-dependent mechanism. In lesions from patients with Fibrodysplasia Ossificans Progressiva, a disease where heterotopic ossification occurs as a result of activating ALK2 mutations, or from a mutant ALK2 transgenic mouse model, chondrocytes and osteoblasts express endothelial markers. Tie2-Cre lineage tracing also suggests an endothelial origin of these cells. Expressing mutant ALK2 in endothelial cells, or treatment with the ALK2 ligands TGF-β2 or BMP4, causes endothelial-mesenchymal transition and acquisition of a stem cell-like phenotype. In selective media, these cells differentiate into osteoblasts, chondrocytes, or adipocytes. The process is inhibited by ALK2-specific siRNA. Conversion of endothelial cells to stem-like cells may provide a novel approach to tissue engineering. PMID:21102460

  17. Human Induced Pluripotent Stem Cell-Derived Endothelial Cells for Three-Dimensional Microphysiological Systems.

    Science.gov (United States)

    Kurokawa, Yosuke K; Yin, Rose T; Shang, Michael R; Shirure, Venktesh S; Moya, Monica L; George, Steven C

    2017-08-01

    Microphysiological systems (MPS), or "organ-on-a-chip" platforms, aim to recapitulate in vivo physiology using small-scale in vitro tissue models of human physiology. While significant efforts have been made to create vascularized tissues, most reports utilize primary endothelial cells that hinder reproducibility. In this study, we report the use of human induced pluripotent stem cell-derived endothelial cells (iPS-ECs) in developing three-dimensional (3D) microvascular networks. We established a CDH5-mCherry reporter iPS cell line, which expresses the vascular endothelial (VE)-cadherin fused to mCherry. The iPS-ECs demonstrate physiological functions characteristic of primary endothelial cells in a series of in vitro assays, including permeability, response to shear stress, and the expression of endothelial markers (CD31, von Willibrand factor, and endothelial nitric oxide synthase). The iPS-ECs form stable, perfusable microvessels over the course of 14 days when cultured within 3D microfluidic devices. We also demonstrate that inhibition of TGF-β signaling improves vascular network formation by the iPS-ECs. We conclude that iPS-ECs can be a source of endothelial cells in MPS providing opportunities for human disease modeling and improving the reproducibility of 3D vascular networks.

  18. Uptake of gold nanoparticles in primary human endothelial cells

    DEFF Research Database (Denmark)

    Klingberg, Henrik; Oddershede, Lene B.; Löschner, Katrin

    2015-01-01

    Gold nanoparticles (AuNPs) are relevant in nanomedicine for drug delivery in the vascular system, where endothelial cells are the first point of contact. We investigated the uptake of 80 nm AuNPs in primary human umbilical vein endothelial cells (HUVECs) by flow cytometry, 3D confocal microscopy......, nano-scale 3D-imaging using focused ion beam/scanning electron microscopy (FIB/SEM), and single particle inductively coupled plasma-mass spectrometry (spICP-MS). HUVECs were cultured for 3 h or 24 h in medium with AuNPs in a concentration range of 1.25–10 μg ml−1. There was a concentration...

  19. Histone Deacetylase (HDAC Inhibitors Down-Regulate Endothelial Lineage Commitment of Umbilical Cord Blood Derived Endothelial Progenitor Cells

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    Horia Maniu

    2012-11-01

    Full Text Available To test the involvement of histone deacetylases (HDACs activity in endothelial lineage progression, we investigated the effects of HDAC inhibitors on endothelial progenitors cells (EPCs derived from umbilical cord blood (UCB. Adherent EPCs, that expressed the endothelial marker proteins (PCAM-1, CD105, CD133, and VEGFR2 revealed by flow cytometry were treated with three HDAC inhibitors: Butyrate (BuA, Trichostatin A (TSA, and Valproic acid (VPA. RT-PCR assay showed that HDAC inhibitors down-regulated the expression of endothelial genes such as VE-cadherin, CD133, CXCR4 and Tie-2. Furthermore, flow cytometry analysis illustrated that HDAC inhibitors selectively reduce the expression of VEGFR2, CD117, VE-cadherin, and ICAM-1, whereas the expression of CD34 and CD45 remained unchanged, demonstrating that HDAC is involved in endothelial differentiation of progenitor cells. Real-Time PCR demonstrated that TSA down-regulated telomerase activity probably via suppression of hTERT expression, suggesting that HDAC inhibitor decreased cell proliferation. Cell motility was also decreased after treatment with HDAC inhibitors as shown by wound-healing assay. The balance of acethylation/deacethylation kept in control by the activity of HAT (histone acetyltransferases/HDAC enzymes play an important role in differentiation of stem cells by regulating proliferation and endothelial lineage commitment.

  20. Corneal endothelial cell density and morphology in Phramongkutklao Hospital

    Directory of Open Access Journals (Sweden)

    Narumon Sopapornamorn

    2008-03-01

    Full Text Available Narumon Sopapornamorn1, Manapon Lekskul1, Suthee Panichkul21Department of Ophthalmology, Phramongkutklao Hospital, Bangkok, Thailand; 2Department of Obstetrics and Gynecology, Phramongkutklao College of Medicine, Bangkok, ThailandObjective: To describe the corneal endothelial density and morphology in patients of Phramongkutklao Hospital and the relationship between endothelial cell parameters and other factors.Methods: Four hundred and four eyes of 202 volunteers were included. Noncontact specular microscopy was performed after taking a history and testing the visual acuity, intraocular pressure measurement, Schirmer’s test and routine eye examination by slit lamp microscope. The studied parameters included mean endothelial cell density (MCD, coefficient of variation (CV, and percentage of hexagonality.Results: The mean age of volunteers was 45.73 years; the range being 20 to 80 years old. Their MCD (SD, mean percentage of CV (SD and mean (SD percentage of hexagonality were 2623.49(325 cell/mm2, 39.43(8.23% and 51.50(10.99%, respectively. Statistically, MCD decreased significantly with age (p < 0.01. There was a significant difference in the percentage of CV between genders. There was no statistical significance between parameters and other factors.Conclusion: The normative data of the corneal endothelium of Thai eyes indicated that, statistically, MCD decreased significantly with age. Previous studies have reported no difference in MCD, percentage of CV, and percentage of hexagonality between gender. Nevertheless, significantly different percentages of CV between genders were presented in this study.Keywords: Corneal endothelial cell, parameters, age, gender, smoking, Thailand

  1. An easily accessible sulfated saccharide mimetic inhibits in vitro human tumor cell adhesion and angiogenesis of vascular endothelial cells

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    Grazia Marano

    2012-05-01

    Full Text Available Oligosaccharides aberrantly expressed on tumor cells influence processes such as cell adhesion and modulation of the cell’s microenvironment resulting in an increased malignancy. Schmidt’s imidate strategy offers an effective method to synthesize libraries of various oligosaccharide mimetics. With the aim to perturb interactions of tumor cells with extracellular matrix proteins and host cells, molecules with 3,4-bis(hydroxymethylfuran as core structure were synthesized and screened in biological assays for their abilities to interfere in cell adhesion and other steps of the metastatic cascade, such as tumor-induced angiogenesis.The most active compound, (4-{[(β-D-galactopyranosyloxy]methyl}furan-3-ylmethyl hydrogen sulfate (GSF, inhibited the activation of matrix-metalloproteinase-2 (MMP-2 as well as migration of the human melanoma cells of the lines WM-115 and WM-266-4 in a two-dimensional migration assay. GSF inhibited completely the adhesion of WM-115 cells to the extracellular matrix (ECM proteins, fibrinogen and fibronectin.In an in vitro angiogenesis assay with human endothelial cells, GSF very effectively inhibited endothelial tubule formation and sprouting of blood vessels, as well as the adhesion of endothelial cells to ECM proteins. GSF was not cytotoxic at biologically active concentrations; neither were 3,4-bis{[(β-D-galactopyranosyloxy]methyl}furan (BGF nor methyl β-D-galactopyranoside nor 3,4-bis(hydroxymethylfuran, which were used as controls, eliciting comparable biological activity. In silico modeling experiments, in which binding of GSF to the extracellular domain of the integrin αvβ3 was determined, revealed specific docking of GSF to the same binding site as the natural peptidic ligands of this integrin. The sulfate in the molecule coordinated with one manganese ion in the binding site.These studies show that this chemically easily accessible molecule GSF, synthesized in three steps from 3,4-bis

  2. Toxicity of fatty acids on ECV-304 endothelial cells.

    Science.gov (United States)

    Masi, Laureane Nunes; Portioli-Sanches, Erica Paula; Lima-Salgado, Thaís Martins; Curi, Rui

    2011-12-01

    The effects of stearic (saturated) or oleic (monounsaturated) acids and their combination with ω-3 and ω-6 polyunsaturated fatty acids (PUFA) on death of endothelial cells (ECV-304 cell line) were investigated. We examined: loss of plasma membrane integrity, DNA fragmentation, accumulation of neutral lipids (NL) and release of reactive oxygen species (ROS). The fatty acids studied were: stearic (SA), oleic (OA), docosahexaenoic (DHA), eicosapentaenoic (EPA), linoleic (LA) and gamma-linolenic (γA) acids. SA at 150 μM induced cell death, did not lead to accumulation of NL and raised the release of ROS. ω-3 PUFA decreased ROS production, increased NL content but did not protect against ECV-304 cell death induced by SA. ω-6 PUFA inhibited SA-induced cell death, increased NL content and decreased ROS production. OA caused cell death but did not increase NL content and ROS production even at 300 μM. ω-3 and ω-6 FA associated with OA further increased cell death with no change in ROS production and NL content. Concluding, ω-6 PUFA had a greater protective effect than ω-3 PUFA on the deleterious effects caused by SA whereas OA had low cytotoxicity but, when associated with PUFA, presented marked toxic effects on ECV-304 endothelial cells. Copyright © 2011 Elsevier Ltd. All rights reserved.

  3. Size-dependent cytotoxicity of europium doped NaYF{sub 4} nanoparticles in endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Shizhu; Zhang, Cuimiao; Jia, Guang; Duan, Jianlei; Wang, Shuxiang, E-mail: wsx@hbu.edu.cn; Zhang, Jinchao, E-mail: jczhang6970@163.com

    2014-10-01

    Lanthanide-doped sodium yttrium fluoride (NaYF{sub 4}) nanoparticles exhibit novel optical properties which make them be widely used in various fields. The extensive applications increase the chance of human exposure to these nanoparticles and thus raise deep concerns regarding their riskiness. In the present study, we have synthesized europium doped NaYF{sub 4} (NaYF{sub 4}:Eu{sup 3+}) nanoparticles with three diameters and used endothelial cells (ECs) as a cell model to explore the potential toxic effect. The cell viability, cytomembrane integrity, cellular uptake, intracellular localization, intracellular reactive oxygen species (ROS), mitochondrial membrane potential (MMP), apoptosis detection, caspase-3 activity and expression of inflammatory gene were studied. The results indicated that these nanoparticles could be uptaken into ECs and decrease the cell viability, induce the intracellular lactate dehydrogenase (LDH) release, increase the ROS level, and decrease the cell MMP in a size-dependent manner. Besides that, the cells were suffered to apoptosis with the caspase-3 activation, and the inflammation specific gene expressions (ICAM1 and VCAM1) were also increased. Our results suggest that the damage pathway may be related to the ROS generation and mitochondrial damage. The results provide novel evidence to elucidate their toxicity mechanisms and may be helpful for more rational applications of these compounds in the future. - Highlights: • NaYF{sub 4}:Eu{sup 3+} nanoparticles with three diameters have been synthesized. • NaYF{sub 4}:Eu{sup 3+} nanoparticles could be uptaken by endothelial cells (ECs). • NaYF{sub 4}:Eu{sup 3+} nanoparticles show a significant cytotoxicity on ECs. • The size of NaYF{sub 4}:Eu{sup 3+} nanoparticles may be important to their toxicology effect.

  4. In vitro analysis of human periodontal microvascular endothelial cells.

    Science.gov (United States)

    Tsubokawa, Mizuki; Sato, Soh

    2014-08-01

    Endothelial cells (ECs) participate in key aspects of vascular biology, such as maintenance of capillary permeability, initiation of coagulation, and regulation of inflammation. According to previous reports, ECs have revealed highly specific characteristics depending on the organs and tissues. However, some reports have described the characteristics of the capillaries formed by human periodontal ECs. Therefore, the aim of the present study is to examine the functional characteristics of the periodontal microvascular ECs in vitro. Human periodontal ligament-endothelial cells (HPDL-ECs) and human gingiva-endothelial cells (HG-ECs) were isolated by immunoprecipitation with magnetic beads conjugated to a monoclonal anti-CD31 antibody. The isolated HPDL-ECs and HG-ECs were characterized to definitively demonstrate that these cell cultures represented pure ECs. Human umbilical-vein ECs and human dermal microvascular ECs were used for comparison. These cells were compared according to the proliferation potential, the formation of capillary-like tubes, the transendothelial electric resistance (TEER), and the expression of tight junction proteins. HPDL-ECs and HG-ECs with characteristic cobblestone monolayer morphology were obtained, as determined by light microscopy at confluence. Furthermore, the HPDL-ECs and HG-ECs expressed the EC markers platelet endothelial cell adhesion molecule-1 (also known as CD31), von Willebrand factor, and Ulex europaeus agglutinin 1, and the cells stained strongly positive for CD31 and CD309. In addition, the HPDL-ECs and HG-ECs were observed to form capillary-like tubes, and they demonstrated uptake of acetylated low-density lipoprotein. Functional analyses of the HPDL-ECs and HG-ECs showed that, compared to the control cells, tube formation persisted for only a brief period of time, and TEER was substantially reduced at confluence. Furthermore, the cells exhibited delocalization of zonula occludens-1 and occludin at cell-cell contact sites

  5. Label-free quantitative cell division monitoring of endothelial cells by digital holographic microscopy

    Science.gov (United States)

    Kemper, Björn; Bauwens, Andreas; Vollmer, Angelika; Ketelhut, Steffi; Langehanenberg, Patrik; Müthing, Johannes; Karch, Helge; von Bally, Gert

    2010-05-01

    Digital holographic microscopy (DHM) enables quantitative multifocus phase contrast imaging for nondestructive technical inspection and live cell analysis. Time-lapse investigations on human brain microvascular endothelial cells demonstrate the use of DHM for label-free dynamic quantitative monitoring of cell division of mother cells into daughter cells. Cytokinetic DHM analysis provides future applications in toxicology and cancer research.

  6. Endothelial cell counts after Descemet’s stripping automated endothelial keratoplasty versus penetrating keratoplasty in Asian eyes

    Science.gov (United States)

    Ang, Marcus; Mehta, Jodhbir S; Anshu, Arundhati; Wong, Hon Kiat; Htoon, Hla M; Tan, Donald

    2012-01-01

    Background The purpose of this study was to compare endothelial cell counts after Descemet’s stripping automated endothelial keratoplasty (DSAEK) and penetrating keratoplasty in Asian eyes. Methods This was a retrospective study of patients from our prospective Singapore Corneal Transplant Study cohort who received corneal transplantation in 2006–2008. We compared eyes that underwent DSAEK or penetrating keratoplasty for Fuchs’ endothelial dystrophy or pseudophakic and aphakic bullous keratopathy. Clinical data, and donor and recipient characteristics were recorded. Of 241 patients who met our inclusion criteria, 68 underwent DSAEK and 173 underwent penetrating keratoplasty. The main outcome measure was endothelial cell loss at 1 year. Secondary outcome measures were graft survival and visual outcomes at 1-year follow-up. Results There were no significant differences in baseline characteristics of patients between the treatment groups. Percent endothelial cell loss at 1-year follow-up was greater in penetrating keratoplasty eyes (40.9% ± 2.9%) compared with DSAEK eyes (22.4% ± 2.3%; P keratoplasty-treated eyes. Penetrating keratoplasty-treated eyes had worse astigmatism as compared with DSAEK-treated eyes (−3.0 ± 2.1 versus −1.7 ± 0.8; P keratoplasty-treated eyes had clear grafts (P = 0.479). Conclusion We report lower percent endothelial cell loss comparing DSAEK and penetrating keratoplasty at 1-year follow-up in Asian eyes, with comparable graft survival rates in both groups. PMID:22536049

  7. An aptamer ligand based liposomal nanocarrier system that targets tumor endothelial cells.

    Science.gov (United States)

    Ara, Mst Naznin; Matsuda, Takashi; Hyodo, Mamoru; Sakurai, Yu; Hatakeyama, Hiroto; Ohga, Noritaka; Hida, Kyoko; Harashima, Hideyoshi

    2014-08-01

    The objective of this study was to construct our recently developed aptamer-modified targeted liposome nano-carrier (Apt-PEG-LPs) system to target primary cultured mouse tumor endothelial cells (mTEC), both in vitro and in vivo. We first synthesized an aptamer-polyethylene glycol 2000-distearoyl phosphoethanolamine (Apt-PEG2000-DSPE). The conjugation of the Apt-PEG2000-DSPE was confirmed by MALDI-TOF mass spectroscopy. A lipid hydration method was used to prepare Apt-PEG-LPs, in which the outer surface of the PEG-spacer was decorated with the aptamer. Apt-PEG-LPs were significantly taken up by mTECs. Cellular uptake capacity was observed both quantitatively and qualitatively using spectrofluorometry, and confocal laser scanning microscopy (CLSM), respectively. In examining the extent of localization of aptamer-modified liposomes that entered the cells, approximately 39% of the Apt-PEG-LPs were not co-localized with lysotracker, indicating that they had escaped from endosomes. The uptake route involved a receptor mediated pathway, followed by clathrin mediated endocytosis. This Apt-PEG-LP was also applied for in vivo research whether this system could target tumor endothelial cells. Apt-PEG-LP and PEG5000-DSPE modified Apt-PEG-LP (Apt/PEG5000-LP) were investigated by human renal cell carcinoma (OS-RC-2 cells) inoculating mice using CLSM. Apt-PEG-LP and Apt/PEG5000-LP showed higher accumulation on tumor vasculature compared to PEG-LP and the co-localization efficacy of Apt-PEG-LP and Apt/PEG5000-LP on TEC were quantified 16% and 25% respectively, which was also better than PEG-LP (3%). The findings suggest that this system is considerable promise for targeting tumor endothelial cells to deliver drugs or genes in vitro and in vivo. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Fate of cerium dioxide nanoparticles in endothelial cells: exocytosis

    Science.gov (United States)

    Strobel, Claudia; Oehring, Hartmut; Herrmann, Rudolf; Förster, Martin; Reller, Armin; Hilger, Ingrid

    2015-05-01

    Although cytotoxicity and endocytosis of nanoparticles have been the subject of numerous studies, investigations regarding exocytosis as an important mechanism to reduce intracellular nanoparticle accumulation are rather rare and there is a distinct lack of knowledge. The current study investigated the behavior of human microvascular endothelial cells to exocytose cerium dioxide (CeO2) nanoparticles (18.8 nm) by utilization of specific inhibitors [brefeldin A; nocodazole; methyl-β-cyclodextrin (MβcD)] and different analytical methods (flow cytometry, transmission electron microscopy, inductively coupled plasma mass spectrometry). Overall, it was found that endothelial cells were able to release CeO2 nanoparticles via exocytosis after the migration of nanoparticle containing endosomes toward the plasma membrane. The exocytosis process occurred mainly by fusion of vesicular membranes with plasma membrane resulting in the discharge of vesicular content to extracellular environment. Nevertheless, it seems to be likely that nanoparticles present in the cytosol could leave the cells in a direct manner. MβcD treatment led to the strongest inhibition of the nanoparticle exocytosis indicating a significant role of the plasma membrane cholesterol content in the exocytosis process. Brefeldin A (inhibitor of Golgi-to-cell-surface-transport) caused a higher inhibitory effect on exocytosis than nocodazole (inhibitor of microtubules). Thus, the transfer from distal Golgi compartments to the cell surface influenced the exocytosis process of the CeO2 nanoparticles more than the microtubule-associated transport. In conclusion, endothelial cells, which came in contact with nanoparticles, e.g., after intravenously applied nano-based drugs, can regulate their intracellular nanoparticle amount, which is necessary to avoid adverse nanoparticle effects on cells.

  9. Fate of cerium dioxide nanoparticles in endothelial cells: exocytosis

    Energy Technology Data Exchange (ETDEWEB)

    Strobel, Claudia, E-mail: Claudia.Strobel@med.uni-jena.de [Jena University Hospital – Friedrich Schiller University Jena, Department of Experimental Radiology, Institute of Diagnostic and Interventional Radiology (Germany); Oehring, Hartmut [Jena University Hospital – Friedrich Schiller University Jena, Institute of Anatomy II (Germany); Herrmann, Rudolf [University of Augsburg, Department of Physics (Germany); Förster, Martin [Jena University Hospital – Friedrich Schiller University Jena, Department of Internal Medicine I, Division of Pulmonary Medicine and Allergy/Immunology (Germany); Reller, Armin [University of Augsburg, Department of Physics (Germany); Hilger, Ingrid, E-mail: ingrid.hilger@med.uni-jena.de [Jena University Hospital – Friedrich Schiller University Jena, Department of Experimental Radiology, Institute of Diagnostic and Interventional Radiology (Germany)

    2015-05-15

    Although cytotoxicity and endocytosis of nanoparticles have been the subject of numerous studies, investigations regarding exocytosis as an important mechanism to reduce intracellular nanoparticle accumulation are rather rare and there is a distinct lack of knowledge. The current study investigated the behavior of human microvascular endothelial cells to exocytose cerium dioxide (CeO{sub 2}) nanoparticles (18.8 nm) by utilization of specific inhibitors [brefeldin A; nocodazole; methyl-β-cyclodextrin (MβcD)] and different analytical methods (flow cytometry, transmission electron microscopy, inductively coupled plasma mass spectrometry). Overall, it was found that endothelial cells were able to release CeO{sub 2} nanoparticles via exocytosis after the migration of nanoparticle containing endosomes toward the plasma membrane. The exocytosis process occurred mainly by fusion of vesicular membranes with plasma membrane resulting in the discharge of vesicular content to extracellular environment. Nevertheless, it seems to be likely that nanoparticles present in the cytosol could leave the cells in a direct manner. MβcD treatment led to the strongest inhibition of the nanoparticle exocytosis indicating a significant role of the plasma membrane cholesterol content in the exocytosis process. Brefeldin A (inhibitor of Golgi-to-cell-surface-transport) caused a higher inhibitory effect on exocytosis than nocodazole (inhibitor of microtubules). Thus, the transfer from distal Golgi compartments to the cell surface influenced the exocytosis process of the CeO{sub 2} nanoparticles more than the microtubule-associated transport. In conclusion, endothelial cells, which came in contact with nanoparticles, e.g., after intravenously applied nano-based drugs, can regulate their intracellular nanoparticle amount, which is necessary to avoid adverse nanoparticle effects on cells.

  10. Outcome of rotational keratoplasty: comparison of endothelial cell loss in autografts vs allografts.

    Science.gov (United States)

    Bertelmann, Eckart; Hartmann, Christian; Scherer, Matthias; Rieck, Peter

    2004-10-01

    The nature of chronic endothelial cell loss in homologous corneal grafts is still unclear. Possible causes are cell migration to the recipient bed and chronic subclinical immune reaction. To compare endothelial cell loss after autologous rotational keratoplasty and homologous keratoplasty and present the clinical outcome of patients after rotational keratoplasty. In this open prospective study, we included 7 consecutive patients who underwent rotational keratoplasty between 1998 and 2000 in our hospital. Patients were examined clinically every 3 months, and visual acuity, astigmatism, and endothelial cell density were evaluated. Endothelial cell densities were compared with endothelial cell counts of 293 homologous keratoplasties. Mean follow-up for autologous grafts was 39 months. Mean increase in visual acuity was 3.5 lines. Mean astigmatism was 4.75 diopters in the autologous graft group. Mean preoperative endothelial cell density was 2058 (637 cells/mm(2)). Mean endothelial cell density after 1 year was 1865 (639 cells/mm(2)), which represents a mean +/- SD cell loss of 15% +/- 7.19%. At the end of follow-up, endothelial cell number after autologous grafting was 1630 +/- 622 cells/mm(2). Endothelial cell loss after 1 year in homologous grafts was 40% +/- 21.34%. There was 1 decompensation of autologous graft in the follow-up period. Main Outcome Measure Comparison of endothelial cell count at different postoperative time points using nonpaired t test. Endothelial cell loss in autologous grafts is significantly lower than in homologous grafts, which supports the hypothesis that chronic endothelial cell loss is due to chronic subclinical immune reactions in homologous grafts. Autologous keratoplasties can lead to good functional results and can be superior to homologous corneal grafting in suitable situations.

  11. Induced Pluripotent Stem Cell-Derived Endothelial Cells in Insulin Resistance and Metabolic Syndrome.

    Science.gov (United States)

    Carcamo-Orive, Ivan; Huang, Ngan F; Quertermous, Thomas; Knowles, Joshua W

    2017-11-01

    Insulin resistance leads to a number of metabolic and cellular abnormalities including endothelial dysfunction that increase the risk of vascular disease. Although it has been particularly challenging to study the genetic determinants that predispose to abnormal function of the endothelium in insulin-resistant states, the possibility of deriving endothelial cells from induced pluripotent stem cells generated from individuals with detailed clinical phenotyping, including accurate measurements of insulin resistance accompanied by multilevel omic data (eg, genetic and genomic characterization), has opened new avenues to study this relationship. Unfortunately, several technical barriers have hampered these efforts. In the present review, we summarize the current status of induced pluripotent stem cell-derived endothelial cells for modeling endothelial dysfunction associated with insulin resistance and discuss the challenges to overcoming these limitations. © 2017 American Heart Association, Inc.

  12. Antioxidant Effects of Sheep Whey Protein on Endothelial Cells

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    Efthalia Kerasioti

    2016-01-01

    Full Text Available Excessive production of reactive oxygen species (ROS may cause endothelial dysfunction and consequently vascular disease. In the present study, the possible protective effects of sheep whey protein (SWP from tert-butyl hydroperoxide- (tBHP- induced oxidative stress in endothelial cells (EA.hy926 were assessed using oxidative stress biomarkers. These oxidative stress biomarkers were glutathione (GSH and ROS levels determined by flow cytometry. Moreover, thiobarbituric acid-reactive substances (TBARS, protein carbonyls (CARB, and oxidized glutathione (GSSG were determined spectrophotometrically. The results showed that SWP at 0.78, 1.56, 3.12, and 6.24 mg of protein mL−1 increased GSH up to 141%, while it decreased GSSG to 46.7%, ROS to 58.5%, TBARS to 52.5%, and CARB to 49.0%. In conclusion, the present study demonstrated for the first time that SWP protected endothelial cells from oxidative stress. Thus, SWP may be used for developing food supplements or biofunctional foods to attenuate vascular disturbances associated with oxidative stress.

  13. Pulmonary endothelial cell DNA methylation signature in pulmonary arterial hypertension.

    Science.gov (United States)

    Hautefort, Aurélie; Chesné, Julie; Preussner, Jens; Pullamsetti, Soni S; Tost, Jorg; Looso, Mario; Antigny, Fabrice; Girerd, Barbara; Riou, Marianne; Eddahibi, Saadia; Deleuze, Jean-François; Seeger, Werner; Fadel, Elie; Simonneau, Gerald; Montani, David; Humbert, Marc; Perros, Frédéric

    2017-08-08

    Pulmonary arterial hypertension (PAH) is a severe and incurable pulmonary vascular disease. One of the primary origins of PAH is pulmonary endothelial dysfunction leading to vasoconstriction, aberrant angiogenesis and smooth muscle cell proliferation, endothelial-to-mesenchymal transition, thrombosis and inflammation. Our objective was to study the epigenetic variations in pulmonary endothelial cells (PEC) through a specific pattern of DNA methylation. DNA was extracted from cultured PEC from idiopathic PAH ( n = 11), heritable PAH ( n = 10) and controls ( n = 18). DNA methylation was assessed using the Illumina HumanMethylation450 Assay. After normalization, samples and probes were clustered according to their methylation profile. Differential clusters were functionally analyzed using bioinformatics tools. Unsupervised hierarchical clustering allowed the identification of two clusters of probes that discriminates controls and PAH patients. Among 147 differential methylated promoters, 46 promoters coding for proteins or miRNAs were related to lipid metabolism. Top 10 up and down-regulated genes were involved in lipid transport including ABCA1, ABCB4, ADIPOQ, miR-26A, BCL2L11. NextBio meta-analysis suggested a contribution of ABCA1 in PAH. We confirmed ABCA1 mRNA and protein downregulation specifically in PAH PEC by qPCR and immunohistochemistry and made the proof-of-concept in an experimental model of the disease that its targeting may offer novel therapeutic options. In conclusion, DNA methylation analysis identifies a set of genes mainly involved in lipid transport pathway which could be relevant to PAH pathophysiology.

  14. Endothelial progenitor cells as a therapeutic option in intracerebral hemorrhage

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    Juan Pías-Peleteiro

    2017-01-01

    Full Text Available Intracerebral hemorrhage (ICH is the most severe cerebrovascular disease, which represents a leading cause of death and disability in developed countries. However, therapeutic options are limited, so is mandatory to investigate repairing processes after stroke in order to develop new therapeutic strategies able to promote brain repair processes. Therapeutic angiogenesis and vasculogenesis hold promise to improve outcome of ICH patients. In this regard, circulating endothelial progenitor cells (EPCs have recently been suggested to be a marker of vascular risk and endothelial function. Moreover, EPC levels have been associated with good neurological and functional outcome as well as reduced residual hematoma volume in ICH patients. Finally, experimental and clinical studies indicate that EPC might mediate endothelial cell regeneration and neovascularization. Therefore, EPC-based therapy could be an excellent therapeutic option in ICH. In this mini-review, we discuss the present status of knowledge about the possible therapeutic role of EPCs in ICH, molecular mechanisms, and the future perspectives and strategies for their use in clinical practice.

  15. Endothelial progenitor cells give rise to pro-angiogenic smooth muscle-like progeny

    NARCIS (Netherlands)

    Moonen, Jan-Renier A. J.; Krenning, Guido; Brinker, Marja G. L.; Koerts, Jasper A.; van Luyn, Marja J. A.; Harmsen, Martin C.

    2010-01-01

    Reciprocal plasticity exists between endothelial and mesenchymal lineages. For instance, mature endothelial cells adopt a smooth muscle-like phenotype through transforming growth factor beta-1 (TGF beta 1)-driven endothelial-to-mesenchymal transdifferentiation (EndMT). Peripheral blood contains

  16. Changes of junctions of endothelial cells in coronary sclerosis: A review

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    Li-Zi Zhang

    2016-03-01

    Full Text Available Atherosclerosis, the major cause of cardiovascular diseases, has been a leading contributor to morbidity and mortality in the United States and it has been on the rise globally. Endothelial cell–cell junctions are critical for vascular integrity and maintenance of vascular function. Endothelial cell junctions dysfunction is the onset step of future coronary events and coronary artery disease. Keywords: Coronary atherosclerosis, Junctions, Endothelial cells

  17. Effects of pulsatile flow on cultured vascular endothelial cell morphology.

    Science.gov (United States)

    Helmlinger, G; Geiger, R V; Schreck, S; Nerem, R M

    1991-05-01

    Endothelial cells (EC) appear to adapt their morphology and function to the in vivo hemodynamic environment in which they reside. In vitro experiments indicate that similar alterations occur for cultured EC exposed to a laminar steady-state flow-induced shear stress. However, in vivo EC are exposed to a pulsatile flow environment; thus, in this investigation, the influence of pulsatile flow on cell shape and orientation and on actin microfilament localization in confluent bovine aortic endothelial cell (BAEC) monolayers was studied using a 1-Hz nonreversing sinusoidal shear stress of 40 +/- 20 dynes/cm2 (type I), 1-Hz reversing sinusoidal shear stresses of 20 +/- 40 and 10 +/- 15 dynes/cm2 (type II), and 1-Hz oscillatory shear stresses of 0 +/- 20 and 0 +/- 40 dynes/cm2 (type III). The results show that in a type I nonreversing flow, cell shape changed less rapidly, but cells took on a more elongated shape than their steady flow controls long-term. For low-amplitude type II reversing flow, BAECs changed less rapidly in shape and were always less elongated than their steady controls; however, for high amplitude reversal, BAECs did not stay attached for more than 24 hours. For type III oscillatory flows, BAEC cell shape remained polygonal as in static culture and did not exhibit actin stress fibers, such as occurred in all other flows. These results demonstrate that EC can discriminate between different types of pulsatile flow environments.(ABSTRACT TRUNCATED AT 250 WORDS)

  18. Effects of Surface Viscoelasticity on Cellular Responses of Endothelial Cells

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    Motahare-Sadat Hosseini

    2014-10-01

    Full Text Available Background: One area of nanoscience deals with nanoscopic interactions between nanostructured materials and biological systems. To elucidate the effects of the substrate surface morphology and viscoelasticity on cell proliferation, fractal analysis was performed on endothelial cells cultured on nanocomposite samples based on silicone rubber (SR and various concentrations of organomodified nanoclay (OC. Methods: The nanoclay/SR ratio was tailored to enhance cell behavior via changes in sample substrate surface roughness and viscoelasticity. Results: Surface roughness of the cured SR filled with negatively-charged nanosilicate layers had a greater effect than elasticity on cell growth. The surface roughness of SR nanocomposite samples increased with increasing the OC content, leading to enhanced cell growth and extracellular matrix (ECM remodeling. This was consistent with the decrease in SR segmental motions and damping factor as the primary viscoelastic parameters by the nanosilicate layers with increasing clay concentrations. Conclusions: The inclusion of clay nanolayers affected the growth and behavior of endothelial cells on microtextured SR.

  19. Mesenchymal Stromal Cells as Active Regulators of Lymphocyte Recruitment to Blood Vascular Endothelial Cells.

    Science.gov (United States)

    Mcgettrick, Helen M; Ward, Lewis S C; Rainger, George Edward; Nash, Gerard B

    2017-01-01

    Methods are described for analyzing adhesion and migration of isolated lymphocytes on endothelial cell monolayers which have been cocultured with different mesenchymal stromal cells, with or without additional cytokine treatment. The different cells types are grown on opposite sides of 3.0 or 0.4 μm pore filters, depending on whether migration through the whole construct is to be analyzed, or adhesion to the endothelial cells alone. Migration away from the sub-endothelial space and through the stromal layer can also be assessed by culturing mesenchymal stromal cells within a 3-D collagen gel overlaid with endothelial cells. Assays may be "static" or the filter-based constructs can be incorporated into flow chambers so that cell behavior can be directly observed under conditions simulating those in vivo. In general, by choice of method, one can evaluate efficiency of attachment, and ability of cells to migrate across the endothelial monolayer, through the filter and through the stromal cell layer in 2-D or 3-D. Fluorescence microscopic examination of fixed filters can be used, e.g., to ascertain whether lymphocytes are retained by stromal cells. In general, static assays have the higher throughput and greatest ease of use, while the flow-based assays are more physiologically relevant and allow detailed recording of cell behavior in real time.

  20. Endothelial cell contraction increases Candida adherence to exposed extracellular matrix.

    Science.gov (United States)

    Klotz, S A; Maca, R D

    1988-01-01

    Bovine vascular endothelial cells treated with EDTA, urea, or thrombin underwent a marked, reversible contraction resulting in exposure of the subendothelial extracellular matrix (ECM). Candida yeasts adhered more to contracted monolayers than to confluent monolayers (P less than 0.01) by preferentially adhering to the ECM. Two strains of Candida albicans and one strain of Candida tropicalis bound avidly to exposed ECM, but Pseudomonas aeruginosa did not. However, treatment of endothelium with forskolin, which induces cell shape changes without exposure of the ECM, did not cause an increase in adherence. Images PMID:3137171

  1. Talin Modulation by a Synthetic N-Acylurea Derivative Reduces Angiogenesis in Human Endothelial Cells

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    I-Rang Lim

    2017-01-01

    Full Text Available Talin is a focal adhesion protein that activates integrins and recruits other focal adhesion proteins. Talin regulates the interactions between integrins and the extracellular matrix, which are critical for endothelial cells during angiogenesis. In this study, we successfully synthesized a novel talin modulator, N-((2-(1H-indol-3-ylethylcarbamoyl-2-(benzo[d][1,3]dioxol-5-yloxyacetamide, referred to as KCH-1521. KCH-1521 was determined to bind talin and modulate downstream signaling molecules of talin. After 24 h of treatment, KCH-1521 changed the cell morphology of human umbilical vein endothelial cells (HUVECs and reduced focal adhesion protein expression including vinculin and paxillin. Talin downstream signaling is regulated via focal adhesion kinase (FAK, kinase B (AKT, and extracellular signal-regulated kinase (ERK pathways, however, treatment with KCH-1521 decreased phosphorylation of FAK, AKT, and ERK, leading to reduction of cell proliferation, survival, and angiogenesis. Interestingly, the expression of various angiogenic genes was significantly decreased after treatment with KCH-1521. Also, in vitro tube forming assay revealed that KCH-1521 reduced angiogenic networks in a time-dependent manner. To investigate the reversibility of its effects, KCH-1521 was removed after treatment. HUVECs recovered their morphology through rearrangement of the cytoskeleton and the expression of angiogenic genes was also recovered. By further optimization and in vivo studies of KCH-1521, a novel drug of talin modulation could be used to achieve therapeutic anti-angiogenesis for vascular diseases and cancers.

  2. Liver sinusoidal endothelial cells induce immunosuppressive IL-10-producing Th1 cells via the Notch pathway

    NARCIS (Netherlands)

    Neumann, Katrin; Rudolph, Christine; Neumann, Christian; Janke, Marko; Amsen, Derk; Scheffold, Alexander

    2015-01-01

    Under homeostasis, liver sinusoidal endothelial cells (LSECs) shift intrahepatic T-cell responses towards tolerance. However, the role of LSECs in the regulation of T-cell-induced liver inflammation is less clear. Here, we studied the capacity of LSECs to modulate pro-inflammatory Th1-cell

  3. Visfatin as a novel mediator released by inflamed human endothelial cells.

    Science.gov (United States)

    Romacho, Tania; Villalobos, Laura A; Cercas, Elena; Carraro, Raffaele; Sánchez-Ferrer, Carlos F; Peiró, Concepción

    2013-01-01

    Visfatin is a multifaceted adipokine whose circulating levels are enhanced in different metabolic diseases. Extracellular visfatin can exert various deleterious effects on vascular cells, including inflammation and proliferation. Limited evidence exists, however, on the capacity of human vascular cells to synthesize and release visfatin by themselves, under basal or pro-inflammatory conditions. Intracellular visfatin was detected by Western blot in non-stimulated human umbilical vein endothelial cells (HUVEC). However, exposing HUVEC for 18 h to a series of pro-inflammatory stimulus, such as interleukin (IL)-1β (1 to 10 ng/mL), tumor necrosis factor-α (1 to 10 ng/mL) or angiotensin II (10 pmol/L to 1 μmol/L) markedly enhanced intracellular visfatin content. Using IL-1β (10 ng/mL; 18 h), it was determined that the increase in intracellular visfatin, which was paralleled by enhanced visfatin mRNA levels, relied on a signalling mechanism involving both nuclear factor-κB and poly (ADP ribose) polymerase-1 activation. Moreover, IL-1β modified the sub-cellular localization of visfatin; while in non-stimulated HUVEC immunoreactive visfatin predominantly showed an intra-nuclear granular pattern, in IL-1β-inflamed cells an extra-nuclear filamentous staining, co-localising with F-actin fibers and suggesting a secretory pattern, was mainly found. Indeed, IL-1β promoted visfatin secretion, as determined by both ELISA and immunocytochemistry. Human endothelial cells synthesize and release visfatin, particularly in response to inflammation. We suggest that the inflamed endothelium can be a source of visfatin, which arises as a local inflammatory mediator and a potential therapeutic target to interfere with vascular inflammation.

  4. Visfatin as a novel mediator released by inflamed human endothelial cells.

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    Tania Romacho

    Full Text Available BACKGROUND: Visfatin is a multifaceted adipokine whose circulating levels are enhanced in different metabolic diseases. Extracellular visfatin can exert various deleterious effects on vascular cells, including inflammation and proliferation. Limited evidence exists, however, on the capacity of human vascular cells to synthesize and release visfatin by themselves, under basal or pro-inflammatory conditions. METHODS AND RESULTS: Intracellular visfatin was detected by Western blot in non-stimulated human umbilical vein endothelial cells (HUVEC. However, exposing HUVEC for 18 h to a series of pro-inflammatory stimulus, such as interleukin (IL-1β (1 to 10 ng/mL, tumor necrosis factor-α (1 to 10 ng/mL or angiotensin II (10 pmol/L to 1 μmol/L markedly enhanced intracellular visfatin content. Using IL-1β (10 ng/mL; 18 h, it was determined that the increase in intracellular visfatin, which was paralleled by enhanced visfatin mRNA levels, relied on a signalling mechanism involving both nuclear factor-κB and poly (ADP ribose polymerase-1 activation. Moreover, IL-1β modified the sub-cellular localization of visfatin; while in non-stimulated HUVEC immunoreactive visfatin predominantly showed an intra-nuclear granular pattern, in IL-1β-inflamed cells an extra-nuclear filamentous staining, co-localising with F-actin fibers and suggesting a secretory pattern, was mainly found. Indeed, IL-1β promoted visfatin secretion, as determined by both ELISA and immunocytochemistry. CONCLUSIONS: Human endothelial cells synthesize and release visfatin, particularly in response to inflammation. We suggest that the inflamed endothelium can be a source of visfatin, which arises as a local inflammatory mediator and a potential therapeutic target to interfere with vascular inflammation.

  5. Maslinic acid inhibits proliferation of renal cell carcinoma cell lines and suppresses angiogenesis of endothelial cells

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    Parth Thakor

    2017-03-01

    Full Text Available Despite the introduction of many novel therapeutics in clinical practice, metastatic renal cell carcinoma (RCC remains a treatment-re-sistant cancer. As red and processed meat are considered risk factors for RCC, and a vegetable-rich diet is thought to reduce this risk, research into plant-based therapeutics may provide valuable complementary or alternative therapeutics for the management of RCC. Herein, we present the antiproliferative and antiangiogenic effects of maslinic acid, which occurs naturally in edible plants, particularly in olive fruits, and also in a variety of medicinal plants. Human RCC cell lines (ACHN, Caki-1, and SN12K1, endothelial cells (human umbilical vein endothelial cell line [HUVEC], and primary cultures of kidney proximal tubular epithelial cells (PTEC were treated with maslinic acid. Maslinic acid was relatively less toxic to PTEC when compared with RCC under similar experimental conditions. In RCC cell lines, maslinic acid induced a significant reduction in proliferation, proliferating cell nuclear antigen, and colony formation. In HUVEC, maslinic acid induced a significant reduction in capillary tube formation in vitro and vascular endothelial growth factor. This study provides a rationale for incorporating a maslinic acid–rich diet either to reduce the risk of developing kidney cancer or as an adjunct to existing antiangiogenic therapy to improve efficacy.

  6. Recombinant Treponema pallidum protein Tp0965 activates endothelial cells and increases the permeability of endothelial cell monolayer.

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    Rui-Li Zhang

    Full Text Available The recombinant Treponema pallidum protein Tp0965 (rTp0965, one of the many proteins derived from the genome of T. pallidum subsp. pallidum, shows strong immunogenicity and immunoreactivity. In this study, we investigated the effects of rTp0965 on the endothelial barrier. Treatment of human umbilical vein endothelial cells (HUVECs with rTp0965 resulted in increased levels of ICAM-1, E-selectin, and MCP-1 mRNA and protein expression. These increases contributed to the adhesion and chemataxis of monocytes (THP-1 cells to HUVECs preincubated with rTp0965. In addition, rTp0965 induced reorganization of F-actin and decreased expression of claudin-1 in HUVECs. Interestingly, inhibition of the RhoA/ROCK signal pathway protected against rTp0965-induced higher endothelial permeability as well as transendothelial migration of monocytes. These data indicate that Tp0965 protein may play an important role in the immunopathogenesis of syphilis.

  7. Levamisole induced apoptosis in cultured vascular endothelial cells

    Science.gov (United States)

    Artwohl, Michaela; Hölzenbein, Thomas; Wagner, Ludwig; Freudenthaler, Angelika; Waldhäusl, Werner; Baumgartner-Parzer, Sabina M

    2000-01-01

    To better understand the anticancer activity of Levamisole (LMS), which serves as an adjuvant in colon cancer therapy in combination with 5-Fluorouracil, this study analyses LMS' ability to induce apoptosis and growth arrest in cultured human micro- and macrovascular endothelial cells (ECs) and fibroblasts. Cells exposed (24 h) to Levamisole (range: 0.5–2 mmol l−1) alone or in combination with antioxidants (10 mmol l−1 glutathione or 5 mmol l−1 N-Acetylcysteine or 0.1 mmol l−1 Tocopherol) were evaluated for apoptosis (3H-thymidine assays, in situ staining), mRNA/protein expression (Northern/Western blot), and proliferation (3H-thymidine incorporation). Levamisole dose-dependently increased apoptosis in ECs to 230% (HUVECs-human umbilical vein ECs), 525% (adult human venous ECs) and 600% (human uterine microvascular ECs) but not in fibroblasts compared to control cells (set as 100%). Levamisole increased in ECs integrin-dependent matrix adhesion, inhibited proliferation (−70%), reduced expression of survival factors such as clusterin (−30%), endothelin-1 (−43%), bcl-2 (−34%), endothelial NO-synthase (−32%) and pRb (Retinoblastoma protein: −89%), and increased that of growth arrest/death signals such as p21 (+73%) and bak (+50%). LMS (2 mmol l−1)-induced apoptosis was inhibited by glutathione (−50%) and N-Acetylcysteine (−36%), which also counteracted reduction by Levamisole of pRb expression, suggesting reactive oxygen species and pRb play a role in these processes. The ability of LMS to selectively induce apoptosis and growth arrest in endothelial cells potentially hints at vascular targeting to contribute to Levamisole's anticancer activity. PMID:11139434

  8. Interactions between endothelial progenitor cells (EPC) and titanium implant surfaces.

    Science.gov (United States)

    Ziebart, Thomas; Schnell, Anne; Walter, Christian; Kämmerer, Peer W; Pabst, Andreas; Lehmann, Karl M; Ziebart, Johanna; Klein, Marc O; Al-Nawas, Bilal

    2013-01-01

    Endothelial cells play an important role in peri-implant angiogenesis during early bone formation. Therefore, interactions between endothelial progenitor cells (EPCs) and titanium dental implant surfaces are of crucial interest. The aim of our in vitro study was to investigate the reactions of EPCs in contact with different commercially available implant surfaces. EPCs from buffy coats were isolated by Ficoll density gradient separation. After cell differentiation, EPC were cultured for a period of 7 days on different titanium surfaces. The test surfaces varied in roughness and hydrophilicity: acid-etched (A), sand-blasted-blasted and acid-etched (SLA), hydrophilic A (modA), and hydrophilic SLA (modSLA). Plastic and fibronectin-coated plastic surfaces served as controls. Cell numbers and morphology were analyzed by confocal laser scanning microscopy. Secretion of vascular endothelial growth factor (VEGF)-A was measured by enzyme-linked immunosorbent assay and expressions of iNOS and eNOS were investigated by real-time polymerase chain reaction. Cell numbers were higher in the control groups compared to the cells of titanium surfaces. Initially, hydrophilic titanium surfaces (modA and modSLA) showed lower cell numbers than hydrophobic surfaces (A and SLA). After 7 days smoother surfaces (A and modA) showed increased cell numbers compared to rougher surfaces (SLA and modSLA). Cell morphology of A, modA, and control surfaces was characterized by a multitude of pseudopodia and planar cell soma architecture. SLA and modSLA promoted small and plump cell soma with little quantity of pseudopodia. The lowest VEGF level was measured on A, the highest on modSLA. The highest eNOS and iNOS expressions were found on modA surfaces. The results of this study demonstrate that biological behaviors of EPCs can be influenced by different surfaces. The modSLA surface promotes an undifferentiated phenotype of EPCs that has the ability to secrete growth factors in great quantities. In

  9. Functional and gene expression analysis of hTERT overexpressed endothelial cells

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    Haruna Takano

    2008-09-01

    Full Text Available Haruna Takano1, Satoshi Murasawa1,2, Takayuki Asahara1,2,31Institute of Biomedical Research and Innovation, Kobe, Japan; 2RIKEN Center for Developmental Biology, Kobe 650-0047, Japan; 3Tokai University of School of Medicine, Tokai, JapanAbstract: Telomerase dysfunction contributes to cellular senescence. Recent advances indicate the importance of senescence in maintaining vascular cell function in vitro. Human telomerase reverse transcriptase (hTERT overexpression is thought to lead to resistance to apoptosis and oxidative stress. However, the mechanism in endothelial lineage cells is unclear. We tried to generate an immortal endothelial cell line from human umbilical vein endothelial cells using a no-virus system and examine the functional mechanisms of hTERT overexpressed endothelial cell senescence in vitro. High levels of hTERT genes and endothelial cell-specific markers were expressed during long-term culture. Also, angiogenic responses were observed in hTERT overexpressed endothelial cell. These cells showed a delay in senescence and appeared more resistant to stressed conditions. PI3K/Akt-related gene levels were enhanced in hTERT overexpressed endothelial cells. An up-regulated PI3K/Akt pathway caused by hTERT overexpression might contribute to anti-apoptosis and survival effects in endothelial lineage cells.Keywords: endothelial, telomerase, senescence, oxidative stress, anti-apoptosis, PI3K/Akt pathway

  10. A novel minimally-invasive method to sample human endothelial cells for molecular profiling.

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    Stephen W Waldo

    Full Text Available The endothelium is a key mediator of vascular homeostasis and cardiovascular health. Molecular research on the human endothelium may provide insight into the mechanisms underlying cardiovascular disease. Prior methodology used to isolate human endothelial cells has suffered from poor yields and contamination with other cell types. We thus sought to develop a minimally invasive technique to obtain endothelial cells derived from human subjects with higher yields and purity.Nine healthy volunteers underwent endothelial cell harvesting from antecubital veins using guidewires. Fluorescence-activated cell sorting (FACS was subsequently used to purify endothelial cells from contaminating cells using endothelial surface markers (CD34/CD105/CD146 with the concomitant absence of leukocyte and platelet specific markers (CD11b/CD45. Endothelial lineage in the purified cell population was confirmed by expression of endothelial specific genes and microRNA using quantitative polymerase chain reaction (PCR.A median of 4,212 (IQR: 2161-6583 endothelial cells were isolated from each subject. Quantitative PCR demonstrated higher expression of von Willebrand Factor (vWF, P<0.001, nitric oxide synthase 3 (NOS3, P<0.001 and vascular cell adhesion molecule 1 (VCAM-1, P<0.003 in the endothelial population compared to similarly isolated leukocytes. Similarly, the level of endothelial specific microRNA-126 was higher in the purified endothelial cells (P<0.001.This state-of-the-art technique isolates human endothelial cells for molecular analysis in higher purity and greater numbers than previously possible. This approach will expedite research on the molecular mechanisms of human cardiovascular disease, elucidating its pathophysiology and potential therapeutic targets.

  11. Effects of space mission factors on the morphology and function of endothelial cells.

    Science.gov (United States)

    Kapitonova, M Yu; Kuznetsov, S L; Froemming, G R A; Muid, S; Nor-Ashikin, M N K; Otman, S; Shahir, A R M; Nawawi, H

    2013-04-01

    The structure and functions of endothelial cells after space mission were studied by electron and laser confocal microscopy, image analysis, and MTT test. The endothelial cells changed significantly (proliferative activity, size, contours, shape, distribution of mitochondria and microtubules) in comparison with controls on the Earth. These changes indicated injuries in the cytoskeleton and impairment of the barrier function of the cells, which presumably contributed to the development of endothelial dysfunction.

  12. Barrier Functionality of Porcine and Bovine Brain Capillary Endothelial Cells

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    Ailar Nakhlband

    2011-09-01

    Full Text Available Introduction: To date, isolated cell based blood-brain barrier (BBB models have been widely used for brain drug delivery and targeting, due to their relatively proper bioelectrical and permeability properties. However, primary cultures of brain capillary endothelial cells (BCECs isolated from different species vary in terms of bioelectrical and permeability properties. Methods: To pursue this, in the current investigation, primary porcine and bovine BCECs (PBCECs and BBCECs, respectively were isolated and used as an in vitro BBB model. The bioelectrical and permeability properties were assessed in BCECs co-cultured with C6 cells with/without hydrocortisone (550 nM. The bioelectrical properties were further validated by means of the permeability coefficients of transcellular and paracellular markers. Results: The primary PBCECs displayed significantly higher trans-endothelial electrical resistance (~900 W.cm2 than BBCECs (~700 W.cm2 - both co-cultured with C6 cells in presence of hydrocortisone. Permeability coefficients of propranolol/diazepam and mannitol/sucrose in PBCECs were ~21 and ~2 (×10-6 cm.sec-1, where these values for BBCECs were ~25 and ~5 (×10-6 cm.sec-1. Conclusion: Upon our bioelectrical and permeability findings, both models display discriminative barrier functionality but porcine BCECs seem to provide a better platform than bovine BCECs for drug screening and brain targeting.

  13. Effects of hypergravity on the angiogenic potential of endothelial cells.

    Science.gov (United States)

    Costa-Almeida, Raquel; Carvalho, Daniel T O; Ferreira, Miguel J S; Aresta, Guilherme; Gomes, Manuela E; van Loon, Jack J W A; Van der Heiden, Kim; Granja, Pedro L

    2016-11-01

    Angiogenesis, the formation of blood vessels from pre-existing ones, is a key event in pathology, including cancer progression, but also in homeostasis and regeneration. As the phenotype of endothelial cells (ECs) is continuously regulated by local biomechanical forces, studying endothelial behaviour in altered gravity might contribute to new insights towards angiogenesis modulation. This study aimed at characterizing EC behaviour after hypergravity exposure (more than 1g), with special focus on cytoskeleton architecture and capillary-like structure formation. Herein, human umbilical vein ECs (HUVECs) were cultured under two-dimensional and three-dimensional conditions at 3g and 10g for 4 and 16 h inside the large diameter centrifuge at the European Space Research and Technology Centre (ESTEC) of the European Space Agency. Although no significant tendency regarding cytoskeleton organization was observed for cells exposed to high g's, a slight loss of the perinuclear localization of β-tubulin was observed for cells exposed to 3g with less pronounced peripheral bodies of actin when compared with 1g control cells. Additionally, hypergravity exposure decreased the assembly of HUVECs into capillary-like structures, with a 10g level significantly reducing their organization capacity. In conclusion, short-term hypergravity seems to affect EC phenotype and their angiogenic potential in a time and g-level-dependent manner. © 2016 The Author(s).

  14. Culture and detection of primary cilia in endothelial cell models.

    Science.gov (United States)

    Lim, Yi Chung; McGlashan, Sue R; Cooling, Michael T; Long, David S

    2015-01-01

    The primary cilium is a sensor of blood-induced forces in endothelial cells (ECs). Studies that have examined EC primary cilia have reported a wide range of cilia incidence (percentage of ciliated cells). We hypothesise that this variation is due to the diversity in culture conditions in which the cells are grown. We studied two EC types: human umbilical vein endothelial cells (HUVECs) and human microvascular endothelial cells (HMEC-1s). Both cell types were grown in media containing foetal bovine serum (FBS) at high (20 % FBS and 10 % FBS for HUVECs and HMEC-1s, respectively) or low (2 % FBS) concentrations. Cells were then either fixed at confluence, serum-starved or grown post-confluence for 5 days in corresponding expansion media (cobblestone treatment). For each culture condition, we quantified cilia incidence and length. HUVEC ciliogenesis is dependent on serum concentration during the growth phase; low serum (2 % FBS) HUVECs were not ciliated, whereas high serum (20 % FBS) confluent HUVECs have a cilia incidence of 2.1 ± 2.2 % (median ± interquartile range). We report, for the first time, the presence of cilia in the HMEC-1 cell type. HMEC-1s have between 2.2 and 3.5 times greater cilia incidence than HUVECs (p cilia compared to HUVECs (3.0 ± 1.0 μm versus 5.1 ± 2.4 μm, at confluence, p = 0.003). We demonstrate that FBS plays a role in determining the prevalence of cilia in HUVECs. In doing so, we highlight the importance of considering a commonly varied parameter (% FBS), in the experimental design. We recommend that future studies examining large blood vessel EC primary cilia use confluent HUVECs grown in high serum medium, as we found these cells to have a higher cilia incidence than low serum media HUVECs. For studies interested in microvasculature EC primary cilia, we recommend using cobblestone HMEC-1s grown in high serum medium, as these cells have a 19.5 ± 6.2 % cilia incidence.

  15. Exogenous endothelial cells as accelerators of hematopoietic reconstitution

    Directory of Open Access Journals (Sweden)

    Mizer J

    2012-11-01

    Full Text Available Abstract Despite the successes of recombinant hematopoietic-stimulatory factors at accelerating bone marrow reconstitution and shortening the neutropenic period post-transplantation, significant challenges remain such as cost, inability to reconstitute thrombocytic lineages, and lack of efficacy in conditions such as aplastic anemia. A possible means of accelerating hematopoietic reconstitution would be administration of cells capable of secreting hematopoietic growth factors. Advantages of this approach would include: a ability to regulate secretion of cytokines based on biological need; b long term, localized production of growth factors, alleviating need for systemic administration of factors that possess unintended adverse effects; and c potential to actively repair the hematopoietic stem cell niche. Here we overview the field of hematopoietic growth factors, discuss previous experiences with mesenchymal stem cells (MSC in accelerating hematopoiesis, and conclude by putting forth the rationale of utilizing exogenous endothelial cells as a novel cellular therapy for acceleration of hematopoietic recovery.

  16. Functional endothelial progenitor cells from cryopreserved umbilical cord blood

    Science.gov (United States)

    Lin, Ruei-Zeng; Dreyzin, Alexandra; Aamodt, Kristie; Dudley, Andrew C.; Melero-Martin, Juan M.

    2010-01-01

    Umbilical cord blood (UCB) is recognized as an enriched source of endothelial progenitor cells (EPCs) with potential therapeutic value. Because cryopreservation is the only reliable method for long-term storage of UCB cells, the clinical application of EPCs depends on our ability to acquire them from cryopreserved samples; however, the feasibility of doing so remains unclear. In this study we demonstrate that EPCs can be isolated from cryopreserved UCB-derived mononuclear cells (MNCs). The number of outgrowth EPC colonies that emerged in culture from cryopreserved samples was similar to that obtained from fresh UCB. Furthermore, EPCs obtained from cryopreserved MNCs were phenotypically and functionally indistinguishable from freshly isolated ones, including the ability to form blood vessels in vivo. Our results eliminate the necessity of performing cell isolation procedures ahead of future clinical needs and suggest that EPCs derived from cryopreserved UCB may be suitable for EPC-related therapies. PMID:20887663

  17. Effects of Parietaria judaica pollen extract on human microvascular endothelial cells.

    Science.gov (United States)

    Taverna, Simona; Flugy, Anna; Colomba, Paolo; Barranca, Marilisa; De Leo, Giacomo; Alessandro, Riccardo

    2008-08-08

    Pollinosis from Parietaria judaica is one of the main causes of allergy in the Mediterranean area. The present study is designed to assess if P. judaica pollens contain bioactive compounds able to elicit a functional response in endothelial cells. We have demonstrated that addition of pollen extract to human lung microvascular endothelial cells (HMVEC-L) induces a modification of cell morphology, actin cytoskeletal rearrangements and an increase in endothelial cell permeability. We further showed that the treatment of endothelial cells with pollen extract causes an increase of E-selectin and VCAM-1 protein levels as well as an increase of IL-8 production. The stimulation of cell-cell adhesion molecules was paralleled by a dose-dependent increase of adhesion of polymorphonuclear cells (PMNs) to HMVEC-L monolayer. Our results suggest for the first time that pollen affect directly endothelial cells (EC) modulating critical functions related to the inflammatory response.

  18. Dexamethasone Enhances ATP-Induced Inflammatory Responses in Endothelial Cells

    Science.gov (United States)

    Ding, Yi; Gao, Zhan-Guo; Jacobson, Kenneth A.

    2010-01-01

    The purinergic nucleotide ATP is released from stressed cells and is implicated in vascular inflammation. Glucocorticoids are essential to stress responses and are used therapeutically, yet little information is available that describes the effects of glucocorticoids on ATP-induced inflammation. In a human microvascular endothelial cell line, extracellular ATP-induced interleukin (IL)-6 secretion in a dose- and time-dependent manner. When cells were pretreated with dexamethasone, a prototypic glucocorticoid, ATP-induced IL-6 production was enhanced in a time- and dose-dependent manner. Mifepristone, a glucocorticoid receptor antagonist, blocked these effects. ATP-induced IL-6 release was significantly inhibited by a phospholipase C inhibitor [1-[6-[((17β)-3-methoxyestra-1,3,5[10]-trien-17-yl)amino]hexyl]-1H-pyrrole-2,5-dione (U73122)] (63.2 ± 3%, p dexamethasone induced mRNA expression of the purinergic P2Y2 receptor (P2Y2R) 1.8- ± 0.1-fold and, when stimulated with ATP, enhanced Ca2+ release and augmented IL-6 mRNA expression. Silencing of the P2Y2R by its small interfering RNA decreased ATP-induced IL-6 production by 81 ± 1% (p Dexamethasone enhanced the transcription rate of P2Y2R mRNA and induced a dose-related increase in the activity of the P2Y2R promoter. Furthermore, dexamethasone-enhanced ATP induction of adhesion molecule transcription and augmented the release of IL-8. Dexamethasone leads to an unanticipated enhancement of endothelial inflammatory mediator production by extracellular ATP via a P2Y2R-dependent mechanism. These data define a novel positive feedback loop of glucocorticoids and ATP-induced endothelial inflammation. PMID:20826566

  19. Endothelial cells control pancreatic cell fate at defined stages through EGFL7 signaling.

    Science.gov (United States)

    Kao, Der-I; Lacko, Lauretta A; Ding, Bi-Sen; Huang, Chen; Phung, Kathleen; Gu, Guoqiang; Rafii, Shahin; Stuhlmann, Heidi; Chen, Shuibing

    2015-02-10

    Although endothelial cells have been shown to affect mouse pancreatic development, their precise function in human development remains unclear. Using a coculture system containing human embryonic stem cell (hESC)-derived progenitors and endothelial cells, we found that endothelial cells play a stage-dependent role in pancreatic development, in which they maintain pancreatic progenitor (PP) self-renewal and impair further differentiation into hormone-expressing cells. The mechanistic studies suggest that the endothelial cells act through the secretion of EGFL7. Consistently, endothelial overexpression of EGFL7 in vivo using a transgenic mouse model resulted in an increase of PP proliferation rate and a decrease of differentiation toward endocrine cells. These studies not only identified the role of EGFL7 as the molecular handle involved in the crosstalk between endothelium and pancreatic epithelium, but also provide a paradigm for using hESC stepwise differentiation to dissect the stage-dependent roles of signals controlling organogenesis. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  20. Endothelial Cells Control Pancreatic Cell Fate at Defined Stages through EGFL7 Signaling

    Directory of Open Access Journals (Sweden)

    Der-I Kao

    2015-02-01

    Full Text Available Although endothelial cells have been shown to affect mouse pancreatic development, their precise function in human development remains unclear. Using a coculture system containing human embryonic stem cell (hESC-derived progenitors and endothelial cells, we found that endothelial cells play a stage-dependent role in pancreatic development, in which they maintain pancreatic progenitor (PP self-renewal and impair further differentiation into hormone-expressing cells. The mechanistic studies suggest that the endothelial cells act through the secretion of EGFL7. Consistently, endothelial overexpression of EGFL7 in vivo using a transgenic mouse model resulted in an increase of PP proliferation rate and a decrease of differentiation toward endocrine cells. These studies not only identified the role of EGFL7 as the molecular handle involved in the crosstalk between endothelium and pancreatic epithelium, but also provide a paradigm for using hESC stepwise differentiation to dissect the stage-dependent roles of signals controlling organogenesis.

  1. Glioblastoma cell-secreted interleukin-8 induces brain endothelial cell permeability via CXCR2.

    Directory of Open Access Journals (Sweden)

    Julie Dwyer

    Full Text Available Glioblastoma constitutes the most aggressive and deadly of brain tumors. As yet, both conventional and molecular-based therapies have met with limited success in treatment of this cancer. Among other explanations, the heterogeneity of glioblastoma and the associated microenvironment contribute to its development, as well as resistance and recurrence in response to treatments. Increased vascularity suggests that tumor angiogenesis plays an important role in glioblastoma progression. However, the molecular crosstalk between endothelial and glioblastoma cells requires further investigation. To examine the effects of glioblastoma-derived signals on endothelial homeostasis, glioblastoma cell secretions were collected and used to treat brain endothelial cells. Here, we present evidence that the glioblastoma secretome provides pro-angiogenic signals sufficient to disrupt VE-cadherin-mediated cell-cell junctions and promote endothelial permeability in brain microvascular endothelial cells. An unbiased angiogenesis-specific antibody array screen identified the chemokine, interleukin-8, which was further demonstrated to function as a key factor involved in glioblastoma-induced permeability, mediated through its receptor CXCR2 on brain endothelia. This underappreciated interface between glioblastoma cells and associated endothelium may inspire the development of novel therapeutic strategies to induce tumor regression by preventing vascular permeability and inhibiting angiogenesis.

  2. Circulating endothelial cells: a potential parameter of organ damage in sickle cell anemia?

    NARCIS (Netherlands)

    Strijbos, Michiel H.; Landburg, Precious P.; Nur, Erfan; Teerlink, Tom; Leebeek, Frank W. G.; Rijneveld, Anita W.; Biemond, Bart J.; Sleijfer, Stefan; Gratama, Jan W.; Duits, Ashley J.; Schnog, John-John B.

    2009-01-01

    Objective laboratory tools are needed to monitor developing organ damage in sickle cell disease (SCD). Circulating endothelial cells (CECs) are indicative of vascular injury. We determined whether elevated CEC can be detected in asymptomatic SCD with the CellSearch system and whether the CEC count

  3. Circulating endothelial cells : a potential parameter of organ damage in sickle cell anemia?

    NARCIS (Netherlands)

    Strijbos, Michiel H; Landburg, Precious P; Nur, Erfan; Teerlink, Tom; Leebeek, Frank W G; Rijneveld, Anita W; Biemond, Bart J; Sleijfer, Stefan; Gratama, Jan W; Duits, Ashley J; Schnog, John-John B

    2009-01-01

    Objective laboratory tools are needed to monitor developing organ damage in sickle cell disease (SCD). Circulating endothelial cells (CECs) are indicative of vascular injury. We determined whether elevated CEC can be detected in asymptomatic SCD with the CellSearch system and whether the CEC count

  4. Transcellular transport of CCL2 across brain microvascular endothelial cells.

    Science.gov (United States)

    Ge, Shujun; Song, Li; Serwanski, David R; Kuziel, William A; Pachter, Joel S

    2008-03-01

    The means by which the chemokine CCL2 produced in the brain parenchyma can recruit leukocytes lying behind the highly impervious endothelium of the blood-brain barrier (BBB) has remained a paradox. As other chemokines have been evidenced to stimulate their own synthesis and release by peripheral microvascular endothelial cells, and/or undergo transcytosis in the abluminal-to-luminal direction, we determined whether CCL2 experiences similar fates across brain microvascular endothelial cells (BMEC). Using cultured BMEC as a paradigm of the BBB, it was observed that exogenous unlabeled CCL2 actually depressed the release of endogenous CCL2, and further caused diminished CCL2 mRNA levels in these cells. On the other hand, exogenous (125)I-labeled CCL2 exhibited transport across BMEC in a manner that was sensitive to temperature, competition by excess unlabeled CCL2 but not unlabeled CCL3, knockdown of caveolin-1/caveolae, and elimination of the cognate CCL2 receptor CCR2. These results implied a facet of CCL2 transport by a transcellular mechanism partly involving binding of CCL2 to CCR2, and subsequent transfer to caveolae vesicles for transcytosis. This notion was supported by double-label immuno-electronmicroscopy, which revealed co-localization of caveolin-1 with exogenous CCL2, during this chemokine's transit across BMEC. Collectively, these findings provide a rationale by which CCL2, deposited on the abluminal side of the brain microvasculature during inflammatory episodes, can be relayed across the BBB to foster leukocyte recruitment.

  5. Behaviour of Endothelial Cells in a Tridimensional In Vitro Environment

    Directory of Open Access Journals (Sweden)

    Raif Eren Ayata

    2015-01-01

    Full Text Available Angiogenesis is a fundamental process in healing, tumor growth, and a variety of medical conditions. For this reason, in vitro angiogenesis is an area of interest for researchers. Additionally, in vitro angiogenesis is important for the survival of prevascularized tissue-engineering models. The aim of this study was to observe the self-tubular organization behaviour of endothelial cells in the self-assembly method. In this study, bilayered and dermal substitutes were prepared using the self-assembly method. Histological, immunostaining, and biochemical tests were performed. The behavioural dynamics of endothelial cells in this biological environment of supportive cells were observed, as were the steps of the in vitro angiogenic cascade with self-organizing capillary-like structures formation. The epidermal component of the substitutes was seen to promote network expansion and density. It also increased the quantity of angiogenic factors (VEGF and Ang-1 without increasing the proinflammatory factor (IL-8. In addition, the increased MMP activity contributed to matrix degradation, which facilitated capillary formation.

  6. Influence of stromal cells on lymphocyte adhesion and migration on endothelial cells

    OpenAIRE

    McGettrick, Helen M.; Buckley, Chris D.; Rainger, G.Ed; Nash, Gerard B

    2010-01-01

    Methods are described for analysing adhesion and migration of isolated lymphocytes on endothelial cell monolayers which have been co-cultured with different stromal cells, with or without additional cytokine treatment. The different cells types are grown on opposite sides of 3.0 or 0.4μm pore filters, depending on whether migration through the whole construct is to be analysed, or adhesion to the endothelial cells alone. Assays may be ‘static’ or filters can be incorporated into flow chambers...

  7. Sun Ginseng Protects Endothelial Progenitor Cells From Senescence Associated Apoptosis

    Science.gov (United States)

    Im, Wooseok; Chung, Jin-Young; Bhan, Jaejun; Lim, Jiyeon; Lee, Soon-Tae; Chu, Kon; Kim, Manho

    2012-01-01

    Endothelial progenitor cells (EPC) are a population of cells that circulate in the blood stream. They play a role in angiogenesis and, therefore, can be prognostic markers of vascular repair. Ginsenoside Rg3 prevents endothelial cell apoptosis through the inhibition of the mitochondrial caspase pathway. It also affects estrogen activity, which reduces EPC senescence. Sun ginseng (SG), which is heat-processed ginseng, has a high content of ginsenosides. The purpose of this study was to investigate the protective effects of SG on senescence-associated apoptosis in EPCs. In order to isolate EPCs, mononuclear cells of human blood buffy coats were cultured and characterized by their uptake of acetylated low-density lipoprotein (acLDL) and their binding of Ulex europaeus agglutinin I (ulex-lectin). Flow cytometry with annexin-V staining was performed in order to assess early and late apoptosis. Senescence was determined by β-galactosidase (β-gal) staining. Staining with 4′-6-Diamidino-2-phenylindole verified that most adherent cells (93±2.7%) were acLDL-positive and ulex-lectin-positive. The percentage of β-gal-positive EPCs was decreased from 93.8±2.0% to 62.5±3.6% by SG treatment. A fluorescence-activated cell sorter (FACS) analysis showed that 4.9% of EPCs were late apoptotic in controls. Sun ginseng decreased the apoptotic cell population by 39% in the late stage of apoptosis from control baseline levels. In conclusion, these results show antisenescent and antiapoptotic effects of SG in human-derived EPCs, indicating that SG can enhance EPC-mediated repair mechanisms. PMID:23717107

  8. [Circulating endothelial progenitor cell levels in treated hypertensive patients].

    Science.gov (United States)

    Maroun-Eid, C; Ortega-Hernández, A; Abad, M; García-Donaire, J A; Barbero, A; Reinares, L; Martell-Claros, N; Gómez-Garre, D

    2015-01-01

    Most optimally treated hypertensive patients still have an around 50% increased risk of any cardiovascular event, suggesting the possible existence of unidentified risk factors. In the last years there has been evidence of the essential role of circulating endothelial progenitor cells (EPCs) in the maintenance of endothelial integrity and function, increasing the interest in their involvement in cardiovascular disease. In this study, the circulating levels of EPCs and vascular endothelial growth factor (VEGF) are investigated in treated hypertensive patients with adequate control of blood pressure (BP). Blood samples were collected from treated hypertensive patients with controlled BP. Plasma levels of EPCs CD34+/KDR+ and CD34+/VE-cadherin+ were quantified by flow cytometry. Plasma concentration of VEGF was determined by ELISA. A group of healthy subjects without cardiovascular risk factors was included as controls. A total of 108 hypertensive patients were included (61±12 years, 47.2% men) of which 82.4% showed BP<140/90 mmHg, 91.7% and 81.5% controlled diabetes (HbA1c <7%) and cLDL (<130 or 100 mg/dL), respectively, and 85.2% were non-smokers. Around 45% of them were obese. Although patients had cardiovascular parameters within normal ranges, they showed significantly lower levels of CD34+/KDR+ and CD34+/VE-cadherin+ compared with healthy control group, although plasma VEGF concentration was higher in patients than in controls. Despite an optimal treatment, hypertensive patients show a decreased number of circulating EPCs that could be, at least in part, responsible for their residual cardiovascular risk, suggesting that these cells could be a therapeutic target. Copyright © 2015 SEHLELHA. Published by Elsevier España, S.L.U. All rights reserved.

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

    Science.gov (United States)

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

    2014-06-01

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

  10. 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

    Energy Technology Data Exchange (ETDEWEB)

    Vallon, Mario, E-mail: m.vallon@arcor.de [Nuklearmedizinische Klinik und Poliklinik, Technische Universitaet Muenchen, Ismaninger Strasse 22, 81675 Munich (Germany); Rohde, Franziska; Janssen, Klaus-Peter [Chirurgische Klinik und Poliklinik, Technische Universitaet Muenchen, Munich (Germany); Essler, Markus [Nuklearmedizinische Klinik und Poliklinik, Technische Universitaet Muenchen, Ismaninger Strasse 22, 81675 Munich (Germany)

    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.

  11. Endothelial-mesenchymal transition and its contribution to the emergence of stem cell phenotype

    Science.gov (United States)

    Medici, Damian; Kalluri, Raghu

    2012-01-01

    Vascular endothelial cells can demonstrate considerable plasticity to generate other cell types during embryonic development and disease progression. This process occurs through a cell differentiation mechanism known as endothelial-mesenchymal transition (EndMT). The generation of mesenchymal cells from endothelium is a crucial step in endothelial cell differentiation to several lineages including fibroblasts, myofibroblasts, mural cells, osteoblasts, chondrocytes, and adipocytes. Such differentiation patterns have been observed in systems of cardiac development, fibrosis, diabetic nephropathy, heterotopic ossification and cancer. Here we describe the EndMT program and discuss the current evidence of EndMT-mediated acquisition of stem cell characteristics and multipotent differentiation capabilities. PMID:22554794

  12. Endothelial progenitor cells and hypertension: current concepts and future implications.

    Science.gov (United States)

    Luo, Shengyuan; Xia, Wenhao; Chen, Cong; Robinson, Eric A; Tao, Jun

    2016-11-01

    The discovery of endothelial progenitor cells (EPCs), a group of cells that play important roles in angiogenesis and the maintenance of vascular endothelial integrity, has led to considerable improvements in our understanding of the circulatory system and the regulatory mechanisms of vascular homoeostasis. Despite lingering disputes over where EPCs actually originate and how they facilitate angiogenesis, extensive research in the past decade has brought about significant advancements in this field of research, establishing EPCs as an essential element in the pathogenesis of various diseases. EPC and hypertensive disorders, especially essential hypertension (EH, also known as primary hypertension), represent one of the most appealing branches in this area of research. Chronic hypertension remains a major threat to public health, and the exact pathologic mechanisms of EH have never been fully elucidated. Is there a relationship between EPC and hypertension? If so, what is the nature of such relationship-is it mediated by blood pressure alterations, or other factors that lie in between? How can our current knowledge about EPCs be utilized to advance the prevention and clinical management of hypertension? In this review, we set out to answer these questions by summarizing the current concepts about EPC pathophysiology in the context of hypertension, while attempting to point out directions for future research on this subject. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

  13. CD40-TRAF Signaling Upregulates CX3CL1 and TNF-α in Human Aortic Endothelial Cells but Not in Retinal Endothelial Cells

    Science.gov (United States)

    Greene, Jennifer A.; Portillo, Jose-Andres C.; Lopez Corcino, Yalitza; Subauste, Carlos S.

    2015-01-01

    CD40, CX3CL1 and TNF-α promote atheroma and neointima formation. CD40 and TNF-α are also central to the development of diabetic retinopathy while CX3CL1 may play a role in the pathogenesis of this retinopathy. The purpose of this study was to examine whether CD40 ligation increases CX3CL1 and TNF-α protein expression in human endothelial cells from the aorta and retina. CD154 (CD40 ligand) upregulated membrane-bound and soluble CX3CL1 in human aortic endothelial cells. CD154 triggered TNF-α production by human aortic endothelial cells. TNF Receptor Associated Factors (TRAF) are key mediators of CD40 signaling. Compared to human aortic endothelial cells that express wt CD40, cells that express CD40 with a mutation that prevents TRAF2,3 recruitment, or CD40 with a mutation that prevents TRAF6 recruitment exhibited a profound inhibition of CD154-driven upregulation of membrane bound and soluble CX3CL1 as well as of TNF-α secretion. While both CD154 and TNF-α upregulated CX3CL1 in human aortic endothelial cells, these stimuli could act independently of each other. In contrast to human aortic endothelial cells, human retinal endothelial cells did not increase membrane bound or soluble CX3CL1 expression or secrete TNF-α in response to CD154 even though CD40 ligation upregulated ICAM-1 and CCL2 in these cells. Moreover, TNF-α did not upregulate CX3CL1 in retinal endothelial cells. In conclusion, CD40 ligation increases CX3CL1 protein levels and induces TNF-α production in endothelial cells. However, endothelial cells are heterogeneous in regards to these responses. Human aortic but not retinal endothelial cells upregulated CX3CL1 and TNF-α in response to CD40 ligation, as well as upregulated CX3CL1 in response to TNF-α. These dissimilarities may contribute to differences in regulation of inflammation in large vessels versus the retina. PMID:26710229

  14. CD40-TRAF Signaling Upregulates CX3CL1 and TNF-α in Human Aortic Endothelial Cells but Not in Retinal Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Jennifer A Greene

    Full Text Available CD40, CX3CL1 and TNF-α promote atheroma and neointima formation. CD40 and TNF-α are also central to the development of diabetic retinopathy while CX3CL1 may play a role in the pathogenesis of this retinopathy. The purpose of this study was to examine whether CD40 ligation increases CX3CL1 and TNF-α protein expression in human endothelial cells from the aorta and retina. CD154 (CD40 ligand upregulated membrane-bound and soluble CX3CL1 in human aortic endothelial cells. CD154 triggered TNF-α production by human aortic endothelial cells. TNF Receptor Associated Factors (TRAF are key mediators of CD40 signaling. Compared to human aortic endothelial cells that express wt CD40, cells that express CD40 with a mutation that prevents TRAF2,3 recruitment, or CD40 with a mutation that prevents TRAF6 recruitment exhibited a profound inhibition of CD154-driven upregulation of membrane bound and soluble CX3CL1 as well as of TNF-α secretion. While both CD154 and TNF-α upregulated CX3CL1 in human aortic endothelial cells, these stimuli could act independently of each other. In contrast to human aortic endothelial cells, human retinal endothelial cells did not increase membrane bound or soluble CX3CL1 expression or secrete TNF-α in response to CD154 even though CD40 ligation upregulated ICAM-1 and CCL2 in these cells. Moreover, TNF-α did not upregulate CX3CL1 in retinal endothelial cells. In conclusion, CD40 ligation increases CX3CL1 protein levels and induces TNF-α production in endothelial cells. However, endothelial cells are heterogeneous in regards to these responses. Human aortic but not retinal endothelial cells upregulated CX3CL1 and TNF-α in response to CD40 ligation, as well as upregulated CX3CL1 in response to TNF-α. These dissimilarities may contribute to differences in regulation of inflammation in large vessels versus the retina.

  15. Dying endothelial cells stimulate proliferation of malignant glioma cells via a caspase 3-mediated pathway

    OpenAIRE

    Mao, Ping; Smith, Luke; XIE, WANFU; Wang, Maode

    2013-01-01

    Emerging evidence has indicated that apoptotic cells have a compensatory effect on the proliferation of neighboring cells. However, the potential role of dying vascular endothelial cells (ECs) in glioma tumor proliferation remains unclear. In the present study, three glioma cell lines were cocultured with dying ECs under various conditions to evaluate the effect of dying ECs on tumor proliferation using alamarBlue and trypan blue assays to assess cell proliferation and viability, respectively...

  16. Plastic compressed collagen as a novel carrier for expanded human corneal endothelial cells for transplantation.

    Directory of Open Access Journals (Sweden)

    Hannah J Levis

    Full Text Available Current treatments for reversible blindness caused by corneal endothelial cell failure involve replacing the failed endothelium with donor tissue using a one donor-one recipient strategy. Due to the increasing pressure of a worldwide donor cornea shortage there has been considerable interest in developing alternative strategies to treat endothelial disorders using expanded cell replacement therapy. Protocols have been developed which allow successful expansion of endothelial cells in vitro but this approach requires a supporting material that would allow easy transfer of cells to the recipient. We describe the first use of plastic compressed collagen as a highly effective, novel carrier for human corneal endothelial cells. A human corneal endothelial cell line and primary human corneal endothelial cells retained their characteristic cobblestone morphology and expression of tight junction protein ZO-1 and pump protein Na+/K+ ATPase α1 after culture on collagen constructs for up to 14 days. Additionally, ultrastructural analysis suggested a well-integrated endothelial layer with tightly opposed cells and apical microvilli. Plastic compressed collagen is a superior biomaterial in terms of its speed and ease of production and its ability to be manipulated in a clinically relevant manner without breakage. This method provides expanded endothelial cells with a substrate that could be suitable for transplantation allowing one donor cornea to potentially treat multiple patients.

  17. Plastic compressed collagen as a novel carrier for expanded human corneal endothelial cells for transplantation.

    Science.gov (United States)

    Levis, Hannah J; Peh, Gary S L; Toh, Kah-Peng; Poh, Rebekah; Shortt, Alex J; Drake, Rosemary A L; Mehta, Jodhbir S; Daniels, Julie T

    2012-01-01

    Current treatments for reversible blindness caused by corneal endothelial cell failure involve replacing the failed endothelium with donor tissue using a one donor-one recipient strategy. Due to the increasing pressure of a worldwide donor cornea shortage there has been considerable interest in developing alternative strategies to treat endothelial disorders using expanded cell replacement therapy. Protocols have been developed which allow successful expansion of endothelial cells in vitro but this approach requires a supporting material that would allow easy transfer of cells to the recipient. We describe the first use of plastic compressed collagen as a highly effective, novel carrier for human corneal endothelial cells. A human corneal endothelial cell line and primary human corneal endothelial cells retained their characteristic cobblestone morphology and expression of tight junction protein ZO-1 and pump protein Na+/K+ ATPase α1 after culture on collagen constructs for up to 14 days. Additionally, ultrastructural analysis suggested a well-integrated endothelial layer with tightly opposed cells and apical microvilli. Plastic compressed collagen is a superior biomaterial in terms of its speed and ease of production and its ability to be manipulated in a clinically relevant manner without breakage. This method provides expanded endothelial cells with a substrate that could be suitable for transplantation allowing one donor cornea to potentially treat multiple patients.

  18. Expression of an insulin-regulatable glucose carrier in muscle and fat endothelial cells

    Science.gov (United States)

    Vilaró, Senen; Palacín, Manuel; Pilch, Paul F.; Testar, Xavier; Zorzano, Antonio

    1989-12-01

    INSULIN rapidly stimulates glucose use in the major target tissues, muscle and fat, by modulating a tissue-specific glucose transporter isoform1-6. Access of glucose to the target tissue is restricted by endothelial cells which line the walls of nonfenestrated capillaries of fat and muscle7. Thus, we examined whether the capillary endothelial cells are actively involved in the modulation of glucose availability by these tissues. We report here the abundant expression of the muscle/fat glucose transporter isoform in endothelial cells, using an immunocytochemical analysis with a monoclonal antibody specific for this isoform1. This expression is restricted to endothelial cells from the major insulin target tissues, and it is not detected in brain and liver where insulin does not activate glucose transport. The expression of the muscle/fat transporter isoform in endothelial cells is significantly greater than in the neighbouring muscle and fat cells. Following administration of insulin to animals in vivo, there occurs a rapid increase in the number of muscle/fat transporters present in the lumenal plasma membrane of the capillary endothelial cells. These results document that insulin promotes the translocation of the muscle/fat glucose transporter in endothelial cells. It is therefore likely that endothelial cells play an important role in the regulation of glucose use by the major insulin target tissues in normal and diseased states.

  19. Circulating endothelial progenitor cells do not contribute to regeneration of endothelium after murine arterial injury

    DEFF Research Database (Denmark)

    Hagensen, Mette; Raarup, Merete Krog; Mortensen, Martin Bødtker

    2012-01-01

    AIMS: Endothelial regeneration after vascular injury, including percutaneous coronary intervention, is essential for vascular homeostasis and inhibition of neointima formation. Circulating endothelial progenitor cells (EPCs) have been implicated to contribute by homing and differentiating...... into endothelial cells (ECs). We tested this theory in a murine arterial injury model using carotid artery transplants and fluorescent reporter mice. METHODS AND RESULTS: Wire-injured carotid artery segments from wild-type mice were transplanted into TIE2-GFP transgenic mice expressing green fluorescent protein...... recipient mouse. Among 1186 ECs identified in re-endothelialized transplants (n= 5) by staining for von Willebrand Factor or vascular endothelial-cadherin, we did not find any blood-derived (GFP(+)) cells. CONCLUSION: Endothelial regeneration after vascular injury did not involve circulating EPCs...

  20. Effect of vitamin D on endothelial progenitor cells function.

    Science.gov (United States)

    Hammer, Yoav; Soudry, Alissa; Levi, Amos; Talmor-Barkan, Yeela; Leshem-Lev, Dorit; Singer, Joel; Kornowski, Ran; Lev, Eli I

    2017-01-01

    Endothelial progenitor cells (EPCs) are a population of bone marrow-derived cells, which have an important role in the process of endothelialization and vascular repair following injury. Impairment of EPCs, which occurs in patients with diabetes, was shown to be related to endothelial dysfunction, coronary artery disease (CAD) and adverse clinical outcomes. Recent evidence has shown that calcitriol, the active hormone of vitamin D, has a favorable impact on the endothelium and cardiovascular system. There is limited data on the effect of vitamin D on EPCs function. To examine the in vitro effects of Calcitriol on EPCs from healthy subjects and patients with diabetes. Fifty-one patients with type 2 diabetes (60±11 years, 40% women, HbA1C: 9.1±0.8%) and 23 healthy volunteers were recruited. EPCs were isolated and cultured with and without calcitriol. The capacity of the cells to form colony-forming units (CFUs), their viability (measured by MTT assay), KLF-10 levels and angiogenic markers were evaluated after 1 week of culture. In diabetic patients, EPC CFUs and cell viability were higher in EPCs exposed to calcitriol vs. EPCs not exposed to calcitriol [EPC CFUs: 1.25 (IQR 1.0-2.0) vs. 0.5 (IQR 0.5-1.9), p < 0.001; MTT:0.62 (IQR 0.44-0.93) vs. 0.52 (IQR 0.31-0.62), p = 0.001]. KLF-10 levels tended to be higher in EPCs exposed to vitamin D, with no differences in angiopoietic markers. In healthy subjects, calcitriol supplementation also resulted in higher cell viability [MTT: 0.23 (IQR 0.11-0.46) vs. 0.19 (0.09-0.39), p = 0.04], but without differences in CFU count or angiopoietic markers. In patients with diabetes mellitus, in vitro vitamin D supplementation improved EPCs capacity to form colonies and viability. Further studies regarding the mechanisms by which vitamin D exerts its effect are required.

  1. Effect of vitamin D on endothelial progenitor cells function.

    Directory of Open Access Journals (Sweden)

    Yoav Hammer

    Full Text Available Endothelial progenitor cells (EPCs are a population of bone marrow-derived cells, which have an important role in the process of endothelialization and vascular repair following injury. Impairment of EPCs, which occurs in patients with diabetes, was shown to be related to endothelial dysfunction, coronary artery disease (CAD and adverse clinical outcomes. Recent evidence has shown that calcitriol, the active hormone of vitamin D, has a favorable impact on the endothelium and cardiovascular system. There is limited data on the effect of vitamin D on EPCs function.To examine the in vitro effects of Calcitriol on EPCs from healthy subjects and patients with diabetes.Fifty-one patients with type 2 diabetes (60±11 years, 40% women, HbA1C: 9.1±0.8% and 23 healthy volunteers were recruited. EPCs were isolated and cultured with and without calcitriol. The capacity of the cells to form colony-forming units (CFUs, their viability (measured by MTT assay, KLF-10 levels and angiogenic markers were evaluated after 1 week of culture.In diabetic patients, EPC CFUs and cell viability were higher in EPCs exposed to calcitriol vs. EPCs not exposed to calcitriol [EPC CFUs: 1.25 (IQR 1.0-2.0 vs. 0.5 (IQR 0.5-1.9, p < 0.001; MTT:0.62 (IQR 0.44-0.93 vs. 0.52 (IQR 0.31-0.62, p = 0.001]. KLF-10 levels tended to be higher in EPCs exposed to vitamin D, with no differences in angiopoietic markers. In healthy subjects, calcitriol supplementation also resulted in higher cell viability [MTT: 0.23 (IQR 0.11-0.46 vs. 0.19 (0.09-0.39, p = 0.04], but without differences in CFU count or angiopoietic markers.In patients with diabetes mellitus, in vitro vitamin D supplementation improved EPCs capacity to form colonies and viability. Further studies regarding the mechanisms by which vitamin D exerts its effect are required.

  2. Endothelial cell counts after Descemet’s stripping automated endothelial keratoplasty versus penetrating keratoplasty in Asian eyes

    Directory of Open Access Journals (Sweden)

    Ang M

    2012-04-01

    Full Text Available Marcus Ang1,2, Jodhbir S Mehta1–4, Arundhati Anshu1,2, Hon Kiat Wong5, Hla M Htoon2, Donald Tan1–31Singapore National Eye Centre, 2Singapore Eye Research Institute, 3Department of Ophthalmology, National University Health Systems, 4Department of Clinical Sciences, Duke-NUS Graduate Medical School, 5Department of Ophthalmology, Tan Tock Seng Hospital, SingaporeBackground: The purpose of this study was to compare endothelial cell counts after Descemet’s stripping automated endothelial keratoplasty (DSAEK and penetrating keratoplasty in Asian eyes.Methods: This was a retrospective study of patients from our prospective Singapore Corneal Transplant Study cohort who received corneal transplantation in 2006–2008. We compared eyes that underwent DSAEK or penetrating keratoplasty for Fuchs’ endothelial dystrophy or pseudophakic and aphakic bullous keratopathy. Clinical data, and donor and recipient characteristics were recorded. Of 241 patients who met our inclusion criteria, 68 underwent DSAEK and 173 underwent penetrating keratoplasty. The main outcome measure was endothelial cell loss at 1 year. Secondary outcome measures were graft survival and visual outcomes at 1-year follow-up.Results: There were no significant differences in baseline characteristics of patients between the treatment groups. Percent endothelial cell loss at 1-year follow-up was greater in penetrating keratoplasty eyes (40.9% ± 2.9% compared with DSAEK eyes (22.4% ± 2.3%; P < 0.001. DSAEK-treated eyes had significantly superior uncorrected visual acuity (mean difference = 0.42 ± 0.0059; P < 0.001 and best spectacle-corrected visual acuity (mean difference = 0.14 ± 0.032; P < 0.001 as compared with penetrating keratoplasty-treated eyes. Penetrating keratoplasty-treated eyes had worse astigmatism as compared with DSAEK-treated eyes (-3.0 ± 2.1 versus -1.7 ± 0.8; P < 0.001. Graft survival at 1 year was comparable in both groups, ie, 66/68 (97.0% DSAEK-treated eyes

  3. Double suicide genes selectively kill human umbilical vein endothelial cells

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    Liu Lunxu

    2011-02-01

    Full Text Available Abstract Background To construct a recombinant adenovirus containing CDglyTK double suicide genes and evaluate the killing effect of the double suicide genes driven by kinase domain insert containing receptor (KDR promoter on human umbilical vein endothelial cells. Methods Human KDR promoter, Escherichia coli (E. coli cytosine deaminase (CD gene and the herpes simplex virus-thymidine kinase (TK gene were cloned using polymerase chain reaction (PCR. Plasmid pKDR-CDglyTK was constructed with the KDR promoter and CDglyTK genes. A recombinant adenoviral plasmid AdKDR-CDglyTK was then constructed and transfected into 293 packaging cells to grow and harvest adenoviruses. KDR-expressing human umbilical vein endothelial cells (ECV304 and KDR-negative liver cancer cell line (HepG2 were infected with the recombinant adenoviruses at different multiplicity of infection (MOI. The infection rate was measured by green fluorescent protein (GFP expression. The infected cells were cultured in culture media containing different concentrations of prodrugs ganciclovir (GCV and/or 5-fluorocytosine (5-FC. The killing effects were measured using two different methods, i.e. annexin V-FITC staining and terminal transferase-mediated dUTP nick end-labeling (TUNEL staining. Results Recombinant adenoviruses AdKDR-CDglyTK were successfully constructed and they infected ECV304 and HepG2 cells efficiently. The infection rate was dependent on MOI of recombinant adenoviruses. ECV304 cells infected with AdKDR-CDglyTK were highly sensitive to GCV and 5-FC. The cell survival rate was dependent on both the concentration of the prodrugs and the MOI of recombinant adenoviruses. In contrast, there were no killing effects in the HepG2 cells. The combination of two prodrugs was much more effective in killing ECV304 cells than GCV or 5-FC alone. The growth of transgenic ECV304 cells was suppressed in the presence of prodrugs. Conclusion AdKDR-CDglyTK/double prodrog system may be a useful

  4. Laminar Shear Stress Promotes Mitochondrial Homeostasis in Endothelial Cells.

    Science.gov (United States)

    Wu, Li-Hong; Chang, Hao-Chun; Ting, Pei-Ching; Wang, Danny Ling

    2017-12-08

    Vascular endothelial cells (ECs) are constantly subjected to flow-induced shear stress that is crucial for endothelial functions. Laminar shear stress (LSS) exerts atheroprotection to ECs. Mitochondrial homeostasis is essential for cellular survival. However, the effects of LSS on mitochondrial homeostasis in ECs remain unclear. Mitochondrial homeostasis in ECs exposed to LSS was examined. Cultured human umbilical vein ECs were subjected to LSS (12 dynes/cm2 ) generated by a parallel-plate flow chamber system. ECs subjected to LSS demonstrated an increment of mitochondria in tubular form coupled with the increase of fusion proteins (Mfn2, OPA1) and the decrease of fission protein (Fis1). An increase of both long- and short- OPA1 along with a higher protease YME1L level were observed. LSS triggered a rapid phosphorylation on S637 but a decrease on S616 of fission-controlled protein Drp1. Consistently, Drp1 translocation to mitochondria was decreased in sheared ECs, suggesting that LSS promotes mitochondrial fusion. Enhanced mitochondrial biogenesis in sheared ECs was shown by the increase of mitochondrial mass and its regulatory proeins (PGC1α, TFAM, Nrf1). LSS enhances the expression of mitochondrial antioxidant enzymes and improves mitochondrial functions indicated by the increase of mitochondrial membrane potential (ΔΨm) and ATP generation. TNF α treatment decreased mitochondrial tubular network and its functions in ECs. LSS mitigated TNFα-induced mitochondrial impairments in ECs. Our results clearly indicate that LSS promotes mitochondrial homeostasis and attenuates inflammation-induced mitochondrial impairments in ECs. Our results provide novel insights into the manner of mitochondrial dynamics and functions modulated by LSS that contribute to endothelial integrity. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

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

    Science.gov (United States)

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

    2017-02-01

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

  6. Metabolic Responses in Endothelial Cells Following Exposure to Ketone Bodies

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    Erika Meroni

    2018-02-01

    Full Text Available The ketogenic diet (KD is a high-fat, low-carbohydrate diet based on the induction of the synthesis of ketone bodies (KB. Despite its widespread use, the systemic impact of KD is not completely understood. The purpose of this study was to evaluate the effects of physiological levels of KB on HMEC-1 endothelial cells. To this aim, DNA oxidative damage and the activation of Nrf2, a known transcriptional factor involved in cell responses to oxidative stress, were assessed. The exposure of cells to KB exerted a moderate genotoxic effect, measured by a significant increase in DNA oxidative damage. However, cells pre-treated with KB for 48 h and subjected to a secondary oxidative insult (H2O2, significantly decreased DNA damage compared to control oxidized cells. This protection occurred by the activation of Nrf2 pathway. In KB-treated cells, we found increased levels of Nrf2 in nuclear extracts and higher gene expression of HO-1, a target gene of Nrf2, compared to control cells. These results suggest that KB, by inducing moderate oxidative stress, activate the transcription factor Nrf2, which induces the transcription of target genes involved in the cellular antioxidant defense system.

  7. Cell-cell interactions mediate cytoskeleton organization and collective endothelial cell chemotaxis.

    Science.gov (United States)

    Shamloo, Amir

    2014-09-01

    This study investigates the role of cell-cell and cell-ligand interactions in cytoskeleton organization of endothelial cells (ECs) and their directional migration within a microfluidic device. The migration of ECs in response to a biochemical factor was studied. Mathematical analysis of the cell migration pathways and cellular cytoskeleton revealed that directional migration, migration persistence length, migration speed, and cytoskeletal stress fiber alignment can be mediated by the level of cell contacts as well as the presence or absence of a biochemical polarizing factor. It was shown that in the presence of a biochemical polarizing factor, higher cell density and more frequent cell contacts has a reinforcing effect on collective cell chemotaxis. In contrast, in the absence of a polarizing factor, high cell density can decrease or suppress the ability of the cells to migrate. Also, the correlation of actin stress fiber organization and alignment with directional migration of ECs was investigated. It was shown that in the presence of a biochemical polarizing factor, stress fibers within the cytoskeleton of ECs can be significantly aligned parallel to the gradient direction when the cells have higher level of contacts. The results also show that the organization and alignment of actin stress fibers is mediated by cell adhesion junctions during collective cell migration and introduce cell-cell interactions as a key factor during collective cell chemotaxis. © 2014 Wiley Periodicals, Inc.

  8. CD34+ cells represent highly functional endothelial progenitor cells in murine bone marrow.

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    Junjie Yang

    Full Text Available 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.

  9. Increased adhesive and inflammatory properties in blood outgrowth endothelial cells from sickle cell anemia patients.

    Science.gov (United States)

    Sakamoto, Tatiana Mary; Lanaro, Carolina; Ozelo, Margareth Castro; Garrido, Vanessa Tonin; Olalla-Saad, Sara Teresinha; Conran, Nicola; Costa, Fernando Ferreira

    2013-11-01

    The endothelium plays an important role in sickle cell anemia (SCA) pathophysiology, interacting with red cells, leukocytes and platelets during the vaso-occlusive process and undergoing activation and dysfunction as a result of intravascular hemolysis and chronic inflammation. Blood outgrowth endothelial cells (BOECs) can be isolated from adult peripheral blood and have been used in diverse studies, since they have a high proliferative capacity and a stable phenotype during in vitro culture. This study aimed to establish BOEC cultures for use as an in vitro study model for endothelial function in sickle cell anemia. Once established, BOECs from steady-state SCA individuals (SCA BOECs) were characterized for their adhesive and inflammatory properties, in comparison to BOECs from healthy control individuals (CON BOECs). Cell adhesion assays demonstrated that control individual red cells adhered significantly more to SCA BOEC than to CON BOEC. Despite these increased adhesive properties, SCA BOECs did not demonstrate significant differences in their expression of major endothelial adhesion molecules, compared to CON BOECs. SCA BOECs were also found to be pro-inflammatory, producing a significantly higher quantity of the cytokine, IL-8, than CON BOECs. From the results obtained, we suggest that BOEC may be a good model for the in vitro study of SCA. Data indicate that endothelial cells of sickle cell anemia patients may have abnormal inflammatory and adhesive properties even outside of the chronic inflammatory and vaso-occlusive environment of patients. © 2013.

  10. Endothelial progenitor cells display clonal restriction in multiple myeloma

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    Dai Kezhi

    2006-06-01

    Full Text Available Abstract Background In multiple myeloma (MM, increased neoangiogenesis contributes to tumor growth and disease progression. Increased levels of endothelial progenitor cells (EPCs contribute to neoangiogenesis in MM, and, importantly, covary with disease activity and response to treatment. In order to understand the mechanisms responsible for increased EPC levels and neoangiogenic function in MM, we investigated whether these cells were clonal by determining X-chromosome inactivation (XCI patterns in female patients by a human androgen receptor assay (HUMARA. In addition, EPCs and bone marrow cells were studied for the presence of clonotypic immunoglobulin heavy-chain (IGH gene rearrangement, which indicates clonality in B cells; thus, its presence in EPCs would indicate a close genetic link between tumor cells in MM and endothelial cells that provide tumor neovascularization. Methods A total of twenty-three consecutive patients who had not received chemotherapy were studied. Screening in 18 patients found that 11 displayed allelic AR in peripheral blood mononuclear cells, and these patients were further studied for XCI patterns in EPCs and hair root cells by HUMARA. In 2 patients whose EPCs were clonal by HUMARA, and in an additional 5 new patients, EPCs were studied for IGH gene rearrangement using PCR with family-specific primers for IGH variable genes (VH. Results In 11 patients, analysis of EPCs by HUMARA revealed significant skewing (≥ 77% expression of a single allele in 64% (n = 7. In 4 of these patients, XCI skewing was extreme (≥ 90% expression of a single allele. In contrast, XCI in hair root cells was random. Furthermore, PCR amplification with VH primers resulted in amplification of the same product in EPCs and bone marrow cells in 71% (n = 5 of 7 patients, while no IGH rearrangement was found in EPCs from healthy controls. In addition, in patients with XCI skewing in EPCs, advanced age was associated with poorer clinical status

  11. Universal cancer vaccine: an update on the design of cancer vaccines generated from endothelial cells.

    Science.gov (United States)

    Lokhov, Petr G; Balashova, Elena E

    2013-07-01

    Among the potential cancer immunotherapies, vaccination against antigens expressed by endothelial cells lining the tumor vasculature represents one of the most attractive options because this approach may prevent the growth of any solid tumor. Therefore, endothelial cells can be used as a source of antigens for developing a so-called "universal" cancer vaccine. Unfortunately, efficient endothelial cell-based cancer vaccines have not yet been developed because previous approaches utilized direct endothelial cell immunizations which is not effective and can result in the elicitation of autoimmune responses associated with systemic autoimmune vasculitis. Recently, the heterogeneity of the endothelial cell surface was defined using an in vitro system as a means of developing antiangiogenic cancer vaccines. This analysis demonstrated that tumors induced specific changes to the microvascular of human endothelial cell (HMEC) surface thereby providing a basis for the design of endothelial cell-based vaccines that directly target the tumor endothelium. (1) This commentary further describes HMEC heterogeneity from the perspective of designing an endothelial cell-based universal (for the treatment of all solid tumors) cancer vaccine with high immunogenicity that does not pose the risk of eliciting autoimmunity.

  12. Time lapse phase contrast video microscopy of directed migration of human microvascular endothelial cells on matrigel

    NARCIS (Netherlands)

    Meade-Tollin, L. C.; van Noorden, C. J.

    2000-01-01

    Migration of microvascular endothelial cells is an early and critical step in angiogenesis. Formation of branching and polygonal cellular aggregates by endothelial cells on matrigel has often been considered to be an in vitro model for angiogenesis, although formation of lumens has not always been

  13. Pulsatile atheroprone shear stress affects the expression of transient receptor potential channels in human endothelial cells

    DEFF Research Database (Denmark)

    Thilo, Florian; Vorderwülbecke, Bernd J; Marki, Alex

    2012-01-01

    The goal of the study was to assess whether pulsatile atheroprone shear stress modulates the expression of transient receptor potential (TRP) channels, TRPC3, TRPC6, TRPM7, and TRPV1 mRNA, in human umbilical vascular endothelial cells. Exposure of cultured vascular endothelial cells to defined...

  14. Recent advances in understanding the roles of vascular endothelial cells in allergic inflammation.

    Science.gov (United States)

    Shoda, Tetsuo; Futamura, Kyoko; Orihara, Kanami; Emi-Sugie, Maiko; Saito, Hirohisa; Matsumoto, Kenji; Matsuda, Akio

    2016-01-01

    Allergic disorders commonly involve both chronic tissue inflammation and remodeling caused by immunological reactions to various antigens on tissue surfaces. Due to their anatomical location, vascular endothelial cells are the final responders to interact with various exogenous factors that come into contact with the epithelial surface, such as pathogen-associated molecular patterns (PAMPs) and antigens. Recent studies have shed light on the important roles of endothelial cells in the development and exacerbation of allergic disorders. For instance, endothelial cells have the greatest potential to produce several key molecules that are deeply involved in allergic inflammation, such as periostin and thymus and activation-regulated chemokine (TARC/CCL17). Additionally, endothelial cells were recently shown to be important functional targets for IL-33--an essential regulator of allergic inflammation. Notably, almost all endothelial cell responses and functions involved in allergic inflammation are not suppressed by corticosteroids. These corticosteroid-refractory endothelial cell responses and functions include TNF-α-associated angiogenesis, leukocyte adhesion, IL-33-mediated responses and periostin and TARC production. Therefore, these unique responses and functions of endothelial cells may be critically involved in the pathogenesis of various allergic disorders, especially their refractory processes. Here, we review recent studies, including ours, which have elucidated previously unknown pathophysiological roles of vascular endothelial cells in allergic inflammation and discuss the possibility of endothelium-targeted therapy for allergic disorders. Copyright © 2015 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

    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. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Corneal endothelial cell changes associated with cataract surgery in patients with type 2 diabetes mellitus

    DEFF Research Database (Denmark)

    Hugod, Mikkel; Storr-Paulsen, Allan; Norregaard, Jens Christian

    2011-01-01

    To investigate the corneal endothelial cell density and morphology in patients with and without diabetes after phacoemulsification with intraocular lens implantation.......To investigate the corneal endothelial cell density and morphology in patients with and without diabetes after phacoemulsification with intraocular lens implantation....

  17. Delta- and gamma-tocotrienol isomers are potent in inhibiting inflammation and endothelial activation in stimulated human endothelial cells

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    Suhaila Muid

    2016-07-01

    Full Text Available Background: Tocotrienols (TCTs are more potent antioxidants than α-tocopherol (TOC. However, the effectiveness and mechanism of the action of TCT isomers as anti-atherosclerotic agents in stimulated human endothelial cells under inflammatory conditions are not well established. Aims: 1 To compare the effects of different TCT isomers on inflammation, endothelial activation, and endothelial nitric oxide synthase (eNOS. 2 To identify the two most potent TCT isomers in stimulated human endothelial cells. 3 To investigate the effects of TCT isomers on NFκB activation, and protein and gene expression levels in stimulated human endothelial cells. Methods: Human umbilical vein endothelial cells were incubated with various concentrations of TCT isomers or α-TOC (0.3–10 µM, together with lipopolysaccharides for 16 h. Supernatant cells were collected and measured for protein and gene expression of cytokines (interleukin-6, or IL-6; tumor necrosis factor-alpha, or TNF-α, adhesion molecules (intercellular cell adhesion molecule-1, or ICAM-1; vascular cell adhesion molecule-1, or VCAM-1; and e-selectin, eNOS, and NFκB. Results: δ-TCT is the most potent TCT isomer in the inhibition of IL-6, ICAM-1, VCAM-1, and NFκB, and it is the second potent in inhibiting e-selectin and eNOS. γ-TCT isomer is the most potent isomer in inhibiting e-selectin and eNOS, and it is the second most potent in inhibiting is IL-6, VCAM-1, and NFκB. For ICAM-1 protein expression, the most potent is δ-TCT followed by α-TCT. α- and β-TCT inhibit IL-6 at the highest concentration (10 µM but enhance IL-6 at lower concentrations. γ-TCT markedly increases eNOS expression by 8–11-fold at higher concentrations (5–10 µM but exhibits neutral effects at lower concentrations. Conclusion: δ- and γ-TCT are the two most potent TCT isomers in terms of the inhibition of inflammation and endothelial activation whilst enhancing eNOS, possibly mediated via the NFκB pathway. Hence

  18. Featured Article: Differential regulation of endothelial nitric oxide synthase phosphorylation by protease-activated receptors in adult human endothelial cells.

    Science.gov (United States)

    Tillery, Lakeisha C; Epperson, Tenille A; Eguchi, Satoru; Motley, Evangeline D

    2016-03-01

    Protease-activated receptors have been shown to regulate endothelial nitric oxide synthase through the phosphorylation of specific sites on the enzyme. It has been established that PAR-2 activation phosphorylates eNOS-Ser-1177 and leads to the production of the potent vasodilator nitric oxide, while PAR-1 activation phosphorylates eNOS-Thr-495 and decreases nitric oxide production in human umbilical vein endothelial cells. In this study, we hypothesize a differential coupling of protease-activated receptors to the signaling pathways that regulates endothelial nitric oxide synthase and nitric oxide production in primary adult human coronary artery endothelial cells. Using Western Blot analysis, we showed that thrombin and the PAR-1 activating peptide, TFLLR, lead to the phosphorylation of eNOS-Ser-1177 in human coronary artery endothelial cells, which was blocked by SCH-79797 (SCH), a PAR-1 inhibitor. Using the nitrate/nitrite assay, we also demonstrated that the thrombin- and TFLLR-induced production of nitric oxide was inhibited by SCH and L-NAME, a NOS inhibitor. In addition, we observed that TFLLR, unlike thrombin, significantly phosphorylated eNOS-Thr-495, which may explain the observed delay in nitric oxide production in comparison to that of thrombin. Activation of PAR-2 by SLIGRL, a PAR-2 specific ligand, leads to dual phosphorylation of both catalytic sites but primarily regulated eNOS-Thr-495 phosphorylation with no change in nitric oxide production in human coronary artery endothelial cells. PAR-3, known as the non-signaling receptor, was activated by TFRGAP, a PAR-3 mimicking peptide, and significantly induced the phosphorylation of eNOS-Thr-495 with minimal phosphorylation of eNOS-Ser-1177 with no change in nitric oxide production. In addition, we confirmed that PAR-mediated eNOS-Ser-1177 phosphorylation was Ca(2+)-dependent using the Ca(2+) chelator, BAPTA, while eNOS-Thr-495 phosphorylation was mediated via Rho kinase using the ROCK inhibitor, Y-27632

  19. High syndecan-1 levels in acute myeloid leukemia are associated with bleeding, thrombocytopathy, endothelial cell damage, and leukocytosis

    DEFF Research Database (Denmark)

    Larsen, Anne Mette Vestskov; Leinøe, Eva Birgitte; Johansson, Pär I

    2013-01-01

    The risk of hemorrhage is influenced by multiple factors in acute myeloid leukemia (AML). We investigated whether hemorrhage in AML patients was associated with endothelial perturbation, potentially caused by thrombocytopenia, platelet dysfunction and leukocytosis. Biomarkers of endothelial......, higher age, endothelial cell activation and damage, and leukocytosis. We suggest that platelet dysfunction and leukocytosis in AML causes endothelial perturbation....

  20. Biomedical potential of silver nanoparticles synthesized from calli cells of Citrullus colocynthis (L.) Schrad

    National Research Council Canada - National Science Library

    Satyavani, K; Gurudeeban, S; Ramanathan, T; Balasubramanian, T

    2011-01-01

    .... In this present investigation, we report, biomedical potential of silver nanopaticles synthesized from calli extract of Citrullus colocynthis on Human epidermoid larynx carcinoma (HEp -2) cell line...

  1. Effects of Nebivolol on Endothelial Gene Expression during Oxidative Stress in Human Umbilical Vein Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Ulisse Garbin

    2008-01-01

    Full Text Available The endothelium plays a key role in the development of atherogenesis and its inflammatory and proliferative status influences the progression of atherosclerosis. The aim of this study is to compare the effects of two beta blockers such as nebivolol and atenolol on gene expression in human umbilical vein endothelial cells (HUVECs following an oxidant stimulus. HUVECs were incubated with nebivolol or atenolol (10 micromol/L for 24 hours and oxidative stress was induced by the addition of oxidized (ox-LDL. Ox-LDL upregulated adhesion molecules (ICAM-1, ICAM-2, ICAM-3, E-selectin, and P-selectin; proteins linked to inflammation (IL-6 and TNFalpha, thrombotic state (tissue factor, PAI-1 and uPA, hypertension such as endothelin-1 (ET-1, and vascular remodeling such as metalloproteinases (MMP-2, MMP-9 and protease inhibitor (TIMP-1. The exposure of HUVECs to nebivolol, but not to atenolol, reduced these genes upregulated by oxidative stress both in terms of protein and RNA expression. The known antioxidant properties of the third generation beta blocker nebivolol seem to account to the observed differences seen when compared to atenolol and support the specific potential protective role of this beta blocker on the expression of a number of genes involved in the initiation and progression of atherosclerosis.

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

    Directory of Open Access Journals (Sweden)

    Cai-Guo Yu

    2016-01-01

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

  3. Late Release of Circulating Endothelial Cells and Endothelial Progenitor Cells after Chemotherapy Predicts Response and Survival in Cancer Patients

    Directory of Open Access Journals (Sweden)

    Jeanine M. Roodhart

    2010-01-01

    Full Text Available We and others have previously demonstrated that the acute release of progenitor cells in response to chemotherapy actually reduces the efficacy of the chemotherapy. Here, we take these data further and investigate the clinical relevance of circulating endothelial (progenitor cells (CE(PCs and modulatory cytokines in patients after chemotherapy with relation to progression-free and overall survival (PFS/OS. Patients treated with various chemotherapeutics were included. Blood sampling was performed at baseline, 4 hours, and 7 and 21 days after chemotherapy. The mononuclear cell fraction was analyzed for CE(PC by FACS analysis. Plasma was analyzed for cytokines by ELISA or Luminex technique. CE(PCs were correlated with response and PFS/OS using Cox proportional hazard regression analysis. We measured CE(PCs and cytokines in 71 patients. Only patients treated with paclitaxel showed an immediate increase in endothelial progenitor cell 4 hours after start of treatment. These immediate changes did not correlate with response or survival. After 7 and 21 days of chemotherapy, a large and consistent increase in CE(PC was found (P < .01, independent of the type of chemotherapy. Changes in CE(PC levels at day 7 correlated with an increase in tumor volume after three cycles of chemotherapy and predicted PFS/OS, regardless of the tumor type or chemotherapy. These findings indicate that the late release of CE(PC is a common phenomenon after chemotherapeutic treatment. The correlation with a clinical response and survival provides further support for the biologic relevance of these cells in patients' prognosis and stresses their possible use as a therapeutic target.

  4. Altered mechanobiology of Schlemm's canal endothelial cells in glaucoma.

    Science.gov (United States)

    Overby, Darryl R; Zhou, Enhua H; Vargas-Pinto, Rocio; Pedrigi, Ryan M; Fuchshofer, Rudolf; Braakman, Sietse T; Gupta, Ritika; Perkumas, Kristin M; Sherwood, Joseph M; Vahabikashi, Amir; Dang, Quynh; Kim, Jae Hun; Ethier, C Ross; Stamer, W Daniel; Fredberg, Jeffrey J; Johnson, Mark

    2014-09-23

    Increased flow resistance is responsible for the elevated intraocular pressure characteristic of glaucoma, but the cause of this resistance increase is not known. We tested the hypothesis that altered biomechanical behavior of Schlemm's canal (SC) cells contributes to this dysfunction. We used atomic force microscopy, optical magnetic twisting cytometry, and a unique cell perfusion apparatus to examine cultured endothelial cells isolated from the inner wall of SC of healthy and glaucomatous human eyes. Here we establish the existence of a reduced tendency for pore formation in the glaucomatous SC cell--likely accounting for increased outflow resistance--that positively correlates with elevated subcortical cell stiffness, along with an enhanced sensitivity to the mechanical microenvironment including altered expression of several key genes, particularly connective tissue growth factor. Rather than being seen as a simple mechanical barrier to filtration, the endothelium of SC is seen instead as a dynamic material whose response to mechanical strain leads to pore formation and thereby modulates the resistance to aqueous humor outflow. In the glaucomatous eye, this process becomes impaired. Together, these observations support the idea of SC cell stiffness--and its biomechanical effects on pore formation--as a therapeutic target in glaucoma.

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    Lifescience Database Archive (English)

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  9. Yap/Taz transcriptional activity in endothelial cells promotes intramembranous ossification via the BMP pathway

    Science.gov (United States)

    Uemura, Mami; Nagasawa, Ayumi; Terai, Kenta

    2016-01-01

    Osteogenesis is categorized into two groups based on developmental histology, intramembranous and endochondral ossification. The role of blood vessels during endochondral ossification is well known, while their role in intramembranous ossification, especially the intertissue pathway, is poorly understood. Here, we demonstrate endothelial Yap/Taz is a novel regulator of intramembranous ossification in zebrafish. Appropriate blood flow is required for Yap/Taz transcriptional activation in endothelial cells and intramembranous ossification. Additionally, Yap/Taz transcriptional activity in endothelial cells specifically promotes intramembranous ossification. BMP expression by Yap/Taz transactivation in endothelial cells is also identified as a bridging factor between blood vessels and intramembranous ossification. Furthermore, the expression of Runx2 in pre-osteoblast cells is a downstream target of Yap/Taz transcriptional activity in endothelial cells. Our results provide novel insight into the relationship between blood flow and ossification by demonstrating intertissue regulation. PMID:27273480

  10. A small population of liver endothelial cells undergoes endothelial-to-mesenchymal transition in response to chronic liver injury.

    Science.gov (United States)

    Ribera, Jordi; Pauta, Montse; Melgar-Lesmes, Pedro; Córdoba, Bernat; Bosch, Anna; Calvo, Maria; Rodrigo-Torres, Daniel; Sancho-Bru, Pau; Mira, Aurea; Jiménez, Wladimiro; Morales-Ruiz, Manuel

    2017-11-01

    Rising evidence points to endothelial-to-mesenchymal transition (EndMT) as a significant source of the mesenchymal cell population in fibrotic diseases. In this context, we hypothesized that liver endothelial cells undergo EndMT during fibrosis progression. Cirrhosis in mice was induced by CCl 4 A transgenic mouse expressing a red fluorescent protein reporter under the control of Tie2 promoter (Tie2-tdTomato) was used to trace the acquisition of EndMT. Sinusoidal vascular connectivity was evaluated by intravital microscopy and high-resolution three-dimensional confocal microscopy. A modest but significant fraction of liver endothelial cells from both cirrhotic patients and CCl 4 -treated Tie2-tdTomato mice acquired an EndMT phenotype characterized by the coexpression of CD31 and α-smooth muscle actin, compared with noncirrhotic livers. Bone morphogenetic protein-7 (BMP-7) inhibited the acquisition of EndMT induced by transforming growth factor-β1 (TGF-β1) treatment in cultured primary mouse liver endothelial cells from control mice. EndMT was also reduced significantly in vivo in cirrhotic Tie2-tdTomato mice treated intraperitoneally with BMP-7 compared with untreated mice (1.9 ± 0.2 vs. 3.8 ± 0.3%, respectively; P livers correlated with a significant decrease in liver fibrosis ( P livers in both animal models and patients. BMP-7 treatment decreases the occurrence of the EndMT phenotype and has a positive impact on the severity of disease by reducing fibrosis and sinusoidal vascular disorganization. NEW & NOTEWORTHY A subpopulation of liver endothelial cells from cirrhotic patients and mice with liver fibrosis undergoes endothelial-to-mesenchymal transition. Liver endothelial cells from healthy mice could transition into a mesenchymal phenotype in culture in response to TGF-β1 treatment. Fibrotic livers treated chronically with BMP-7 showed lower EndMT acquisition, reduced fibrosis, and improved vascular organization. Copyright © 2017 the American

  11. Clock upregulates intercellular adhesion molecule-1 expression and promotes mononuclear cells adhesion to endothelial cells.

    Science.gov (United States)

    Gao, Yinghua; Meng, Dan; Sun, Ning; Zhu, Zhu; Zhao, Ran; Lu, Chao; Chen, Sifeng; Hua, Luchun; Qian, Ruizhe

    2014-01-10

    Clock is a basic helix-loop-helix (bHLH) transcription factor that plays important role in circadian rhythms of various physiological functions. Previous study showed that the expression of intercellular adhesion molecule-1 (ICAM-1) was reduced in the liver tissues of Clock mutant mice. However, how Clock regulates ICAM-1 expression and whether Clock affects cell adhesion function remain unknown. In the present study, we found that exogenous expression of Clock upregulated the gene expressions of ICAM-1 and other adhesion-related genes including VCAM1 and CCL-2, and increased the transcriptional activity of ICAM-1 in mouse brain microvascular endothelial cell lines. In contrast, loss of Clock decreased these gene expressions and ICAM-1 transcriptional activity. Chromatin immunoprecipitation (ChIP) assay revealed that Clock binds to the E-box-like enhancer of ICAM-1 gene. ICAM-1 gene showed rhythmic expression in endothelial cells after serum shock in vitro, suggesting ICAM-1 may be a Clock-controlled gene. Clock regulates the adhesion of mononuclear cells to endothelial cells via ICAM-1. Together, our findings show that Clock is a positive regulator of ICAM-1, and promotes the adhesion of mononuclear cells to endothelial cells. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. Tumor-Derived Factors and Reduced p53 Promote Endothelial Cell Centrosome Over-Duplication.

    Directory of Open Access Journals (Sweden)

    Zhixian Yu

    Full Text Available Approximately 30% of tumor endothelial cells have over-duplicated (>2 centrosomes, which may contribute to abnormal vessel function and drug resistance. Elevated levels of vascular endothelial growth factor A induce excess centrosomes in endothelial cells, but how other features of the tumor environment affect centrosome over-duplication is not known. To test this, we treated endothelial cells with tumor-derived factors, hypoxia, or reduced p53, and assessed centrosome numbers. We found that hypoxia and elevated levels of bone morphogenetic protein 2, 6 and 7 induced excess centrosomes in endothelial cells through BMPR1A and likely via SMAD signaling. In contrast, inflammatory mediators IL-8 and lipopolysaccharide did not induce excess centrosomes. Finally, down-regulation in endothelial cells of p53, a critical regulator of DNA damage and proliferation, caused centrosome over-duplication. Our findings suggest that some tumor-derived factors and genetic changes in endothelial cells contribute to excess centrosomes in tumor endothelial cells.

  13. Do Neural Cells Communicate with Endothelial Cells via Secretory Exosomes and Microvesicles?

    Directory of Open Access Journals (Sweden)

    Neil R. Smalheiser

    2009-01-01

    Full Text Available Neurons, glial, cells, and brain tumor cells tissues release small vesicles (secretory exosomes and microvesicles, which may represent a novel mechanism by which neuronal activity could influence angiogenesis within the embryonic and mature brain. If CNS-derived vesicles can enter the bloodstream as well, they may communicate with endothelial cells in the peripheral circulation and with cells concerned with immune surveillance.

  14. Hyperphosphatemia, Phosphoprotein Phosphatases, and Microparticle Release in Vascular Endothelial Cells.

    Science.gov (United States)

    Abbasian, Nima; Burton, James O; Herbert, Karl E; Tregunna, Barbara-Emily; Brown, Jeremy R; Ghaderi-Najafabadi, Maryam; Brunskill, Nigel J; Goodall, Alison H; Bevington, Alan

    2015-09-01

    Hyperphosphatemia in patients with advanced CKD is thought to be an important contributor to cardiovascular risk, in part because of endothelial cell (EC) dysfunction induced by inorganic phosphate (Pi). Such patients also have an elevated circulating concentration of procoagulant endothelial microparticles (MPs), leading to a prothrombotic state, which may contribute to acute occlusive events. We hypothesized that hyperphosphatemia leads to MP formation from ECs through an elevation of intracellular Pi concentration, which directly inhibits phosphoprotein phosphatases, triggering a global increase in phosphorylation and cytoskeletal changes. In cultured human ECs (EAhy926), incubation with elevated extracellular Pi (2.5 mM) led to a rise in intracellular Pi concentration within 90 minutes. This was mediated by PiT1/slc20a1 Pi transporters and led to global accumulation of tyrosine- and serine/threonine-phosphorylated proteins, a marked increase in cellular Tropomyosin-3, plasma membrane blebbing, and release of 0.1- to 1-μm-diameter MPs. The effect of Pi was independent of oxidative stress or apoptosis. Similarly, global inhibition of phosphoprotein phosphatases with orthovanadate or fluoride yielded a global protein phosphorylation response and rapid release of MPs. The Pi-induced MPs expressed VE-cadherin and superficial phosphatidylserine, and in a thrombin generation assay, they displayed significantly more procoagulant activity than particles derived from cells incubated in medium with a physiologic level of Pi (1 mM). These data show a mechanism of Pi-induced cellular stress and signaling, which may be widely applicable in mammalian cells, and in ECs, it provides a novel pathologic link between hyperphosphatemia, generation of MPs, and thrombotic risk. Copyright © 2015 by the American Society of Nephrology.

  15. Probing Leader Cells in Endothelial Collective Migration by Plasma Lithography Geometric Confinement

    Science.gov (United States)

    Yang, Yongliang; Jamilpour, Nima; Yao, Baoyin; Dean, Zachary S.; Riahi, Reza; Wong, Pak Kin

    2016-01-01

    When blood vessels are injured, leader cells emerge in the endothelium to heal the wound and restore the vasculature integrity. The characteristics of leader cells during endothelial collective migration under diverse physiological conditions, however, are poorly understood. Here we investigate the regulation and function of endothelial leader cells by plasma lithography geometric confinement generated. Endothelial leader cells display an aggressive phenotype, connect to follower cells via peripheral actin cables and discontinuous adherens junctions, and lead migrating clusters near the leading edge. Time-lapse microscopy, immunostaining, and particle image velocimetry reveal that the density of leader cells and the speed of migrating clusters are tightly regulated in a wide range of geometric patterns. By challenging the cells with converging, diverging and competing patterns, we show that the density of leader cells correlates with the size and coherence of the migrating clusters. Collectively, our data provide evidence that leader cells control endothelial collective migration by regualting the migrating clusters. PMID:26936382

  16. The effect of 193 nm excimer laser radiation on the human corneal endothelial cell density

    Energy Technology Data Exchange (ETDEWEB)

    Isager, P.; Hjortdal, J.Oe.; Ehlers, N. [Aarhus Univ. Hospital, Dept. of Ophthalmology, Aarhus (Denmark)

    1996-06-01

    The effect of 193 nm excimer laser radiation on human corneal endothelial cell density was examined. Fifty-five eyes from 35 patients underwent photorefractive keratectomy for myopia. Photomicrographs of the endothelium were taken a short time before the operation and on an average of 7 months postoperatively with a specular microscope. The average endothelial cell densities were preoperatively 3375 {+-} 266 cells/mm{sup 2} (means {+-} SD) and postoperatively 3348 {+-} 287 cells/mm{sup 2}, corresponding to a fall of 27 cells/mm{sup 2} (N = 55). This fall in endothelial cell density was not statistically significant. A significant correlation between the change in cell density and age of the patient was found, with older patients losing more cells (N = 35, 2p < 0.05). The magnification of the specular microscope was found to change with corneal thickness. The importance of correcting the endothelial cell densities for corneal thickness is discussed. (au) 14 refs.

  17. Probing Leader Cells in Endothelial Collective Migration by Plasma Lithography Geometric Confinement.

    Science.gov (United States)

    Yang, Yongliang; Jamilpour, Nima; Yao, Baoyin; Dean, Zachary S; Riahi, Reza; Wong, Pak Kin

    2016-03-03

    When blood vessels are injured, leader cells emerge in the endothelium to heal the wound and restore the vasculature integrity. The characteristics of leader cells during endothelial collective migration under diverse physiological conditions, however, are poorly understood. Here we investigate the regulation and function of endothelial leader cells by plasma lithography geometric confinement generated. Endothelial leader cells display an aggressive phenotype, connect to follower cells via peripheral actin cables and discontinuous adherens junctions, and lead migrating clusters near the leading edge. Time-lapse microscopy, immunostaining, and particle image velocimetry reveal that the density of leader cells and the speed of migrating clusters are tightly regulated in a wide range of geometric patterns. By challenging the cells with converging, diverging and competing patterns, we show that the density of leader cells correlates with the size and coherence of the migrating clusters. Collectively, our data provide evidence that leader cells control endothelial collective migration by regualting the migrating clusters.

  18. Gene expression analysis of embryonic stem cells expressing VE-cadherin (CD144 during endothelial differentiation

    Directory of Open Access Journals (Sweden)

    Libermann Towia

    2008-05-01

    Full Text Available Abstract Background Endothelial differentiation occurs during normal vascular development in the developing embryo. This process is recapitulated in the adult when endothelial progenitor cells are generated in the bone marrow and can contribute to vascular repair or angiogenesis at sites of vascular injury or ischemia. The molecular mechanisms of endothelial differentiation remain incompletely understood. Novel approaches are needed to identify the factors that regulate endothelial differentiation. Methods Mouse embryonic stem (ES cells were used to further define the molecular mechanisms of endothelial differentiation. By flow cytometry a population of VEGF-R2 positive cells was identified as early as 2.5 days after differentiation of ES cells, and a subset of VEGF-R2+ cells, that were CD41 positive at 3.5 days. A separate population of VEGF-R2+ stem cells expressing the endothelial-specific marker CD144 (VE-cadherin was also identified at this same time point. Channels lined by VE-cadherin positive cells developed within the embryoid bodies (EBs formed by differentiating ES cells. VE-cadherin and CD41 expressing cells differentiate in close proximity to each other within the EBs, supporting the concept of a common origin for cells of hematopoietic and endothelial lineages. Results Microarray analysis of >45,000 transcripts was performed on RNA obtained from cells expressing VEGF-R2+, CD41+, and CD144+ and VEGF-R2-, CD41-, and CD144-. All microarray experiments were performed in duplicate using RNA obtained from independent experiments, for each subset of cells. Expression profiling confirmed the role of several genes involved in hematopoiesis, and identified several putative genes involved in endothelial differentiation. Conclusion The isolation of CD144+ cells during ES cell differentiation from embryoid bodies provides an excellent model system and method for identifying genes that are expressed during endothelial differentiation and that

  19. Silicon microgrooves for contact guidance of human aortic endothelial cells

    Directory of Open Access Journals (Sweden)

    Sara Fernández-Castillejo

    2017-03-01

    Full Text Available Background: Micro- and nanoscale substrates have been fabricated in order to study the influence of the topography on the cellular response. The aim of this work was to prepare different collagen-coated silicon substrates displaying grooves and ridges to mimic the aligned and elongated endothelium found in linear vessels, and to use them as substrates to study cell growth and behaviour.Results: The influence of groove-shaped substrates on cell adhesion, morphology and proliferation were assessed, by comparing them to flat silicon substrates, used as control condition. Using human aortic endothelial cells, microscopy images demonstrate that the cellular response is different depending on the silicon surface, when it comes to cell adhesion, morphology (alignment, circularity and filopodia presence and proliferation. Moreover, these structures exerted no cytotoxic effect.Conclusion: The results suggest that topographical patterning influences cell response. Silicon groove substrates can be used in developing medical devices with microscale features to mimic the endothelium in lineal vessels.

  20. Cross talk with hematopoietic cells regulates the endothelial progenitor cell differentiation of CD34 positive cells.

    Science.gov (United States)

    Kwon, Sang-Mo; Lee, Jun-Hee; Lee, Sang-Hun; Jung, Seok-Yun; Kim, Da-Yeon; Kang, Song-Hwa; Yoo, So-Young; Hong, Jong-Kyu; Park, Ji-Hye; Kim, Jung-Hee; Kim, Sung-Wook; Kim, Yeon-Ju; Lee, Sun-Jin; Kim, Hwi-Gon; Asahara, Takayuki

    2014-01-01

    Despite the crucial role of endothelial progenitor cells (EPCs) in vascular regeneration, the specific interactions between EPCs and hematopoietic cells remain unclear. In EPC colony forming assays, we first demonstrated that the formation of EPC colonies was drastically increased in the coculture of CD34+ and CD34- cells, and determined the optimal concentrations of CD34+ cells and CD34- cells for spindle-shaped EPC differentiation. Functionally, the coculture of CD34+ and CD34- cells resulted in a significant enhancement of adhesion, tube formation, and migration capacity compared with culture of CD34+ cells alone. Furthermore, blood flow recovery and capillary formation were remarkably increased by the coculture of CD34+ and CD34- cells in a murine hind-limb ischemia model. To elucidate further the role of hematopoietic cells in EPC differentiation, we isolated different populations of hematopoietic cells. T lymphocytes (CD3+) markedly accelerated the early EPC status of CD34+ cells, while macrophages (CD11b+) or megakaryocytes (CD41+) specifically promoted large EPC colonies. Our results suggest that specific populations of hematopoietic cells play a role in the EPC differentiation of CD34+ cells, a finding that may aid in the development of a novel cell therapy strategy to overcome the quantitative and qualitative limitations of EPC therapy.

  1. RhoA GTPase regulates radiation-induced alterations in endothelial cell adhesion and migration

    Energy Technology Data Exchange (ETDEWEB)

    Rousseau, Matthieu; Gaugler, Marie-Helene; Rodallec, Audrey; Bonnaud, Stephanie; Paris, Francois [Inserm UMR U892, Centre de Recherche en Cancerologie Nantes-Angers CRCNA, Institut de Recherche Therapeutique IRT-UN, Universite de Nantes, 8 Quai Moncousu, BP 70721, F-44007 (France); Corre, Isabelle, E-mail: icorre@nantes.inserm.fr [Inserm UMR U892, Centre de Recherche en Cancerologie Nantes-Angers CRCNA, Institut de Recherche Therapeutique IRT-UN, Universite de Nantes, 8 Quai Moncousu, BP 70721, F-44007 (France)

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer We explore the role of RhoA in endothelial cell response to ionizing radiation. Black-Right-Pointing-Pointer RhoA is rapidly activated by single high-dose of radiation. Black-Right-Pointing-Pointer Radiation leads to RhoA/ROCK-dependent actin cytoskeleton remodeling. Black-Right-Pointing-Pointer Radiation-induced apoptosis does not require the RhoA/ROCK pathway. Black-Right-Pointing-Pointer Radiation-induced alteration of endothelial adhesion and migration requires RhoA/ROCK. -- Abstract: Endothelial cells of the microvasculature are major target of ionizing radiation, responsible of the radiation-induced vascular early dysfunctions. Molecular signaling pathways involved in endothelial responses to ionizing radiation, despite being increasingly investigated, still need precise characterization. Small GTPase RhoA and its effector ROCK are crucial signaling molecules involved in many endothelial cellular functions. Recent studies identified implication of RhoA/ROCK in radiation-induced increase in endothelial permeability but other endothelial functions altered by radiation might also require RhoA proteins. Human microvascular endothelial cells HMEC-1, either treated with Y-27632 (inhibitor of ROCK) or invalidated for RhoA by RNA interference were exposed to 15 Gy. We showed a rapid radiation-induced activation of RhoA, leading to a deep reorganisation of actin cytoskeleton with rapid formation of stress fibers. Endothelial early apoptosis induced by ionizing radiation was not affected by Y-27632 pre-treatment or RhoA depletion. Endothelial adhesion to fibronectin and formation of focal adhesions increased in response to radiation in a RhoA/ROCK-dependent manner. Consistent with its pro-adhesive role, ionizing radiation also decreased endothelial cells migration and RhoA was required for this inhibition. These results highlight the role of RhoA GTPase in ionizing radiation-induced deregulation of essential endothelial

  2. Shear Stress Inhibits Apoptosis of Ischemic Brain Microvascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Xiafeng Shen

    2013-01-01

    Full Text Available As a therapeutic strategy for ischemic stroke, to restore or increase cerebral blood flow (CBF is the most fundamental option. Laminar shear stress (LS, as an important force generated by CBF, mainly acts on brain microvascular endothelial cells (BMECs. In order to study whether LS was a protective factor in stroke, we investigated LS-intervented ischemic apoptosis of rat BMECs (rBMECs through PE Annexin V/7-AAD, JC-1 and Hoechst 33258 staining to observe the membranous, mitochondrial and nuclear dysfunction. Real-time PCR and western blot were also used to test the gene and protein expressions of Tie-2, Bcl-2 and Akt, which were respectively related to maintain membranous, mitochondrial and nuclear norm. The results showed that LS could be a helpful stimulus for ischemic rBMECs survival. Simultaneously, membranous, mitochondrial and nuclear regulation played an important role in this process.

  3. Kininogen-cytokeratin 1 interactions in endothelial cell biology.

    Science.gov (United States)

    Shariat-Madar, Z; Schmaier, A H

    1999-11-01

    This review brings available data together to put in perspective the binding properties of high molecular weight kininogen (HK) to cytokeratin 1 (CK1) to direct future investigations. We summarize our knowledge of the structure and function of HK and discuss the influence of HK on the biologic activities of the plasma kallikrein/kinin system. To better understand the role of HK in regulating the activation of the kallikrein/kinin system, we discuss the interaction of HK with endothelial cell CK1 and its influence on prekallikrein activation. Last, we address the question on how a cytoskeletal protein could be a membrane-binding site and its possible role in disease states.

  4. Endothelialization of a non-woven silk fibroin net for use in tissue engineering: growth and gene regulation of human endothelial cells.

    Science.gov (United States)

    Unger, R E; Peters, K; Wolf, M; Motta, A; Migliaresi, C; Kirkpatrick, C J

    2004-09-01

    We have previously shown that a biomaterial consisting of a non-woven fibroin net produced from silk (Bombyx mori) cocoons is an excellent scaffolding material for a wide variety of human cells of different tissue types. Endothelialization must take place for a biomaterial to be successful after implantation. Therefore, primary human endothelial cells and the human endothelial cell lines, HPMEC-ST1.6R and ISO-HAS-1, were examined for adherence and growth patterns on the fibroin nets by confocal laser scanning microscopy after vital staining of the cells and by electron microscopy. Endothelial cells adhered and spread along individual fibers of the nets and did not fill the gaps between individual fibers. Higher attachment and growth coverage was obtained if nets were first coated with gelatin, fibronectin or collagen type I. Proinflammatory markers of endothelial cells on the fibers exhibited a non-activated state and LPS-stimulated cells exhibited activation of these markers. Furthermore, a typical PECAM-1 localization at cell-cell contacts was observed. Scanning electron microscopic examination of fibroin nets after removal of cells did not demonstrate any changes to the fibroin structure. HUVEC and HDMEC on fibroin nets embedded in collagen type I gels formed microvessel-like structures. Thus, silk fibroin nets are a highly endothelial cell-compatible scaffolding material that support the growth, normal and inducible cell functions and angiogenesis potential of human endothelial cells in vitro similar to that observed in vivo.

  5. Hypertonic saline impedes tumor cell-endothelial cell interaction by reducing adhesion molecule and laminin expression.

    LENUS (Irish Health Repository)

    Shields, Conor J

    2012-02-03

    BACKGROUND: Hypertonic saline infusion dampens inflammatory responses and suppresses neutrophil-endothelial interaction by reducing adhesion molecule expression. This study tested the hypothesis that hypertonic saline attenuates tumor cell adhesion to the endothelium through a similar mechanism. METHODS: Human colon cancer cells (LS174T) were transfected with green fluorescent protein and exposed to lipopolysaccharide, tumor necrosis factor-alpha, and interleukin-6 under hypertonic and isotonic conditions for 1 and 4 hours. Confluent human umbilical vein endothelial cells were similarly exposed. Cellular apoptosis and expression of adhesion molecules and laminin were measured by flow cytometry. Tumor cell adhesion to endothelium and laminin was assessed with fluorescence microscopy. Data are represented as mean +\\/- standard error of mean, and an ANOVA test was performed to gauge statistical significance, with P <.05 considered significant. RESULTS: Hypertonic exposure significantly reduced tumor cell adhesion despite the presence of the perioperative cell stressors (42 +\\/- 2.9 vs 172.5 +\\/- 12.4, P <.05), attenuated tumor cell beta-1 integrin (14.43 vs 23.84, P <.05), and endothelial cell laminin expression (22.78 +\\/- 2.2 vs 33.74 +\\/- 2.4, P <.05), but did not significantly alter cell viability. CONCLUSION: Hypertonic saline significantly attenuates tumor cell adhesion to endothelium by inhibiting adhesion molecule and laminin expression. This may halt the metastatic behavior of tumor cells shed at surgery.

  6. Integrin-alpha IIb identifies murine lymph node lymphatic endothelial cells responsive to RANKL

    NARCIS (Netherlands)

    O.G. Cordeiro (Olga G.); M. Chypre (Mélanie); N. Brouard (Nathalie); S. Rauber (Simon); F. Alloush (Farouk); M. Romera-Hernandez (Monica); C. Bénézech (Cécile); Z. Li (Zhi); A. Eckly (Anita); M. Coles (Mark); A. Rot (Antal); H. Yagita (Hideo); C. Léon (Catherine); B. Ludewig (Burkhard); T. Cupedo (Tom); F. Lanza (François); C.G. Mueller (Christopher G.)

    2016-01-01

    textabstractMicroenvironment and activation signals likely imprint heterogeneity in the lymphatic endothelial cell (LEC) population. Particularly LECs of secondary lymphoid organs are exposed to different cell types and immune stimuli. However, our understanding of the nature of LEC activation

  7. MFG-E8 released by apoptotic endothelial cells triggers anti-inflammatory macrophage reprogramming.

    Directory of Open Access Journals (Sweden)

    Marie-Joëlle Brissette

    Full Text Available Apoptotic endothelial cells are an important component of the "response to injury" process. Several atherosclerosis risk factors such as hyperglycemia and oxidized low-density lipoproteins, and immune injuries, such as antibodies and complement, induce endothelial cell apoptosis. While endothelial cell apoptosis is known to affect neighboring vascular wall cell biology, its consequences on macrophage reprogramming are ill defined. In this study, we report that apoptosis of human and mouse endothelial cells triggers the release of milk fat globule-epidermal growth factor 8 (MFG-E8 and reprograms macrophages into an anti-inflammatory cells. We demonstrated that MFG-E8 is released by apoptotic endothelial cells in a caspase-3-dependent manner. When macrophages were exposed to conditioned media from serum-starved apoptotic endothelial cells, they adopt a high anti-inflammatory, low pro-inflammatory cytokine/chemokine secreting phenotype that is lost if MFG-E8 is absent from the media. Macrophage treatment with recombinant MFG-E8 recapitulates the effect of conditioned media. Finally, we showed that MFG-E8-mediated reprogramming of macrophages occurs through increased phosphorylation of signal transducer and activator of transcription-3 (STAT-3. Taken together, our study suggests a key role of MFG-E8 release from apoptotic endothelial cells in macrophage reprogramming and demonstrates the importance of the apoptotic microenvironment in anti-inflammatory macrophage responses.

  8. File list: NoD.CDV.10.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  9. File list: NoD.CDV.05.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  10. File list: InP.CDV.20.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  13. File list: InP.CDV.50.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  14. File list: InP.CDV.05.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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  19. File list: DNS.CDV.50.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  20. File list: DNS.CDV.05.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  1. File list: Pol.CDV.50.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  2. File list: InP.CDV.50.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  3. File list: DNS.CDV.20.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  4. File list: InP.CDV.05.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  5. File list: InP.CDV.10.AllAg.Primary_endothelial_cells [Chip-atlas[Archive

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  6. File list: NoD.CDV.10.AllAg.Primary_endothelial_cells [Chip-atlas[Archive

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  7. File list: InP.CDV.10.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  8. File list: NoD.CDV.50.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  12. File list: InP.CDV.20.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  13. File list: NoD.CDV.05.AllAg.Primary_umbilical_vein_endothelial_cells [Chip-atlas[Archive

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  14. File list: InP.CDV.05.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  15. File list: Oth.CDV.10.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  16. File list: His.CDV.10.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  17. File list: Pol.CDV.20.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  18. File list: InP.CDV.50.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  19. File list: DNS.CDV.10.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  1. File list: DNS.CDV.05.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  2. File list: Pol.CDV.10.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  3. File list: Unc.CDV.05.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  4. File list: DNS.CDV.50.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  5. File list: Oth.CDV.05.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  6. File list: Pol.CDV.05.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  7. File list: Unc.CDV.10.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  8. File list: Pol.CDV.50.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  9. File list: Oth.CDV.50.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  10. File list: His.CDV.20.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  11. File list: DNS.CDV.20.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  12. File list: His.CDV.50.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  13. File list: InP.CDV.10.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  14. File list: Unc.CDV.50.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  15. File list: Oth.CDV.20.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  16. File list: InP.CDV.20.AllAg.Coronary_artery_endothelial_cells [Chip-atlas[Archive

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  17. Neutrophil-mediated protection of cultured human vascular endothelial cells from damage by growing Candida albicans hyphae

    Energy Technology Data Exchange (ETDEWEB)

    Edwards, J.E. Jr.; Rotrosen, D.; Fontaine, J.W.; Haudenschild, C.C.; Diamond, R.D.

    1987-05-01

    Interactions were studied between human neutrophils and cultured human umbilical vein endothelial cells invaded by Candida albicans. In the absence of neutrophils, progressive Candida germination and hyphal growth extensively damaged endothelial cell monolayers over a period of 4 to 6 hours, as determined both by morphological changes and release of /sup 51/Cr from radiolabeled endothelial cells. Monolayers were completely destroyed and replaced by hyphae after 18 hours of incubation. In contrast, when added 2 hours after the monolayers had been infected with Candida, neutrophils selectively migrated toward and attached to hyphae at points of hyphal penetration into individual endothelial cells (observed by time-lapse video-microscopy). Attached neutrophils spread over hyphal surfaces both within and beneath the endothelial cells; neutrophil recruitment to initial sites of leukocyte-Candida-endothelial cell interactions continued throughout the first 60 minutes of observation. Neutrophil spreading and stasis were observed only along Candida hyphae and at sites of Candida-endothelial cell interactions. These events resulted in 58.0% killing of Candida at 2 hours and subsequent clearance of Candida from endothelial cell monolayers, as determined by microcolony counts and morphological observation. On introduction of additional neutrophils to yield higher ratios of neutrophils to endothelial cells (10 neutrophils:1 endothelial cell), neutrophil migration toward hyphal elements continued. Despite retraction or displacement of occasional endothelial cells by invading Candida and neutrophils, most endothelial cells remained intact, viable, and motile as verified both by morphological observations and measurement of /sup 51/Cr release from radiolabeled monolayers.

  18. File list: DNS.CDV.20.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

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  19. File list: InP.CDV.20.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

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  20. File list: Pol.CDV.05.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  1. File list: NoD.CDV.05.AllAg.Primary_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  2. File list: DNS.CDV.10.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  3. File list: NoD.CDV.05.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  4. File list: Pol.CDV.10.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  5. File list: ALL.CDV.05.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  6. File list: DNS.CDV.50.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  7. File list: Oth.CDV.05.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  8. File list: His.CDV.10.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  9. File list: Pol.CDV.50.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  10. File list: His.CDV.50.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  11. File list: His.CDV.05.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  12. File list: ALL.CDV.50.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  13. File list: ALL.CDV.20.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  14. File list: Oth.CDV.20.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.CDV.20.AllAg.Aortic_valve_endothelial_cells hg19 TFs and others Cardiovascular Aortic valve endo...thelial cells SRX285599 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.CDV.20.AllAg.Aortic_valve_endothelial_cells.bed ...

  15. File list: Pol.CDV.20.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  16. File list: InP.CDV.50.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  17. File list: DNS.CDV.05.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.CDV.05.AllAg.Aortic_valve_endothelial_cells hg19 DNase-seq Cardiovascular Aortic valve endo...thelial cells http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.CDV.05.AllAg.Aortic_valve_endothelial_cells.bed ...

  18. File list: ALL.CDV.10.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  19. File list: Oth.CDV.10.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  20. File list: Oth.CDV.50.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  1. File list: Unc.CDV.50.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.CDV.50.AllAg.Aortic_valve_endothelial_cells hg19 Unclassified Cardiovascular Aortic valve endo...thelial cells http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Unc.CDV.50.AllAg.Aortic_valve_endothelial_cells.bed ...

  2. File list: Unc.CDV.10.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  3. File list: NoD.CDV.20.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  4. File list: His.CDV.20.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  5. File list: NoD.CDV.10.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  6. File list: InP.CDV.10.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  7. File list: Unc.CDV.05.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  8. File list: InP.CDV.05.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  9. File list: Unc.CDV.20.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  10. File list: NoD.CDV.50.AllAg.Aortic_valve_endothelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  11. Effects of AMPK on high glucose stimulated apoptosis of endothelial cells via regulation of calcium influx

    Directory of Open Access Journals (Sweden)

    Ting LU

    2015-11-01

    Full Text Available Objective To investigate the inhibitory effect of adenosine monophosphate (AMP-dependent protein kinase (AMPK on high glucose-stimulated endothelial cell apoptosis and its mechanism. Methods MS-1 endothelial cells were cultured in vitro, and they were treated with AMPK agonist, AMPK inhibitor, 2-APB (a blocker of store operated Ca2+ channel (SOCC and (or high glucose, and a control group without any intervention were set up. TUNEL assay was performed to determine apoptotic cells. Laser scanning confocal microscopy was used to assess the Ca2+ influx into cells, and Western-blotting was performed to determine the expressions of Stim1 and Orai1 of the store operated Ca2+ channel (SOCC proteins. Results Apoptosis of endothelial cells was induced significantly, and the expressions of Stim1 and Orai1 were upregulated in high glucose group compared with that in control group (P<0.05. The rate of apoptosis of high glucose-induced endothelial cell was found to be increased in AMPK inhibitor group and decreased in AMPK agonist group, and the expressions of Stim1 and Orai1 were found to be down-regulated in AMPK agonist group as compared with that in high glucose group (P<0.05. Compared with the control group, high glucose stimulation significantly induced the Ca2+ influx to endothelial cells; compared with high glucose group, 2-APB significantly inhibited high glucose-induced Ca2+ influx to endothelial cells, and blocked the inducing effect of high-glucose on endothelial cell apoptosis. Compared with high glucose group, AMPK agonist significantly inhibited high glucose-induced cell Ca2+ influx. Conclusion By reducing the expressions of Stim1 and Orai1, AMPK may inhibit SOCC-mediated Ca2+ influx, and block the high glucose-stimulated endothelial cell apoptosis, thus play an important protective role in sustaining endothelial cell function. DOI: 10.11855/j.issn.0577-7402.2015.10.01

  12. Smooth muscle progenitor cells from peripheral blood promote the neovascularization of endothelial colony-forming cells

    Energy Technology Data Exchange (ETDEWEB)

    Joo, Hyung Joon; Seo, Ha-Rim [Department of Cardiology, Cardiovascular Center, College of Medicine, Korea University, Seoul (Korea, Republic of); Jeong, Hyo Eun [Department of Mechanical Engineering, Korea University, Seoul (Korea, Republic of); Choi, Seung-Cheol; Park, Jae Hyung; Yu, Cheol Woong; Hong, Soon Jun [Department of Cardiology, Cardiovascular Center, College of Medicine, Korea University, Seoul (Korea, Republic of); Chung, Seok [Department of Mechanical Engineering, Korea University, Seoul (Korea, Republic of); Lim, Do-Sun, E-mail: dslmd@kumc.or.kr [Department of Cardiology, Cardiovascular Center, College of Medicine, Korea University, Seoul (Korea, Republic of)

    2014-07-11

    Highlights: • Two distinct vascular progenitor cells are induced from adult peripheral blood. • ECFCs induce vascular structures in vitro and in vivo. • SMPCs augment the in vitro and in vivo angiogenic potential of ECFCs. • Both cell types have synergistic therapeutic potential in ischemic hindlimb model. - Abstract: Proangiogenic cell therapy using autologous progenitors is a promising strategy for treating ischemic disease. Considering that neovascularization is a harmonized cellular process that involves both endothelial cells and vascular smooth muscle cells, peripheral blood-originating endothelial colony-forming cells (ECFCs) and smooth muscle progenitor cells (SMPCs), which are similar to mature endothelial cells and vascular smooth muscle cells, could be attractive cellular candidates to achieve therapeutic neovascularization. We successfully induced populations of two different vascular progenitor cells (ECFCs and SMPCs) from adult peripheral blood. Both progenitor cell types expressed endothelial-specific or smooth muscle-specific genes and markers, respectively. In a protein array focused on angiogenic cytokines, SMPCs demonstrated significantly higher expression of bFGF, EGF, TIMP2, ENA78, and TIMP1 compared to ECFCs. Conditioned medium from SMPCs and co-culture with SMPCs revealed that SMPCs promoted cell proliferation, migration, and the in vitro angiogenesis of ECFCs. Finally, co-transplantation of ECFCs and SMPCs induced robust in vivo neovascularization, as well as improved blood perfusion and tissue repair, in a mouse ischemic hindlimb model. Taken together, we have provided the first evidence of a cell therapy strategy for therapeutic neovascularization using two different types of autologous progenitors (ECFCs and SMPCs) derived from adult peripheral blood.

  13. Bortezomib induces autophagic death in proliferating human endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Belloni, Daniela; Veschini, Lorenzo [Myeloma Unit, Department of Oncology, IRCCS H San Raffaele, Milan (Italy); Foglieni, Chiara [Department of Cardiology, IRCCS H San Raffaele, Milan (Italy); Dell' Antonio, Giacomo [Department of Pathology, IRCCS H San Raffaele, Milan (Italy); Caligaris-Cappio, Federico [Myeloma Unit, Department of Oncology, IRCCS H San Raffaele, Milan (Italy); Universita Vita-Salute IRCCS H San Raffaele, Milan (Italy); Ferrarini, Marina [Myeloma Unit, Department of Oncology, IRCCS H San Raffaele, Milan (Italy); Ferrero, Elisabetta, E-mail: elisabetta.ferrero@hsr.it [Myeloma Unit, Department of Oncology, IRCCS H San Raffaele, Milan (Italy)

    2010-04-01

    The proteasome inhibitor Bortezomib has been approved for the treatment of relapsed/refractory multiple myeloma (MM), thanks to its ability to induce MM cell apoptosis. Moreover, Bortezomib has antiangiogenic properties. We report that endothelial cells (EC) exposed to Bortezomib undergo death to an extent that depends strictly on their activation state. Indeed, while quiescent EC are resistant to Bortezomib, the drug results maximally toxic in EC switched toward angiogenesis with FGF, and exerts a moderate effect on subconfluent HUVEC. Moreover, EC activation state deeply influences the death pathway elicited by Bortezomib: after treatment, angiogenesis-triggered EC display typical features of apoptosis. Conversely, death of subconfluent EC is preceded by ROS generation and signs typical of autophagy, including intense cytoplasmic vacuolization with evidence of autophagosomes at electron microscopy, and conversion of the cytosolic MAP LC3 I form toward the autophagosome-associated LC3 II form. Treatment with the specific autophagy inhibitor 3-MA prevents both LC3 I/LC3 II conversion and HUVEC cell death. Finally, early removal of Bortezomib is accompanied by the recovery of cell shape and viability. These findings strongly suggest that Bortezomib induces either apoptosis or autophagy in EC; interfering with the autophagic response may potentiate the antiangiogenic effect of the drug.

  14. Salidroside inhibits endogenous hydrogen peroxide induced cytotoxicity of endothelial cells.

    Science.gov (United States)

    Zhao, Xingyu; Jin, Lianhai; Shen, Nan; Xu, Bin; Zhang, Wei; Zhu, Hongli; Luo, Zhengli

    2013-01-01

    Salidroside, a phenylpropanoid glycoside isolated from Rhodiola rosea L., shows potent antioxidant property. Herein, we investigated the protective effects of salidroside against hydrogen peroxide (H2O2)-induced oxidative damage in human endothelial cells (EVC-304). EVC-304 cells were incubated in the presence or absence of low steady states of H2O2 (3-4 µM) generated by glucose oxidase (GOX) with or without salidroside. 3(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione (GSH) assays were performed, together with Hoechst 33258 staining and flow cytometric analysis using Annexin-V and propidium iodide (PI) label. The results indicated that salidroside pretreatment attenuated endogenous H2O2 induced apoptotic cell death in EVC-304 cells in a dose-dependent pattern. Furthermore, Western blot data revealed that salidroside inhibited activation of caspase-3, 9 and cleavage of poly(ADP-ribose) polymerase (PARP) induced by endogenous H2O2. It also decreased the expression of Bax and rescued the balance of pro- and anti-apoptotic proteins. All these results demonstrated that salidroside may present a potential therapy for oxidative stress in cardiovascular and cerebrovascular diseases.

  15. Neuropilin-1 modulates interferon-γ-stimulated signaling in brain microvascular endothelial cells.

    Science.gov (United States)

    Wang, Ying; Cao, Ying; Mangalam, Ashutosh K; Guo, Yong; LaFrance-Corey, Reghann G; Gamez, Jeffrey D; Atanga, Pascal Aliihnui; Clarkson, Benjamin D; Zhang, Yuebo; Wang, Enfeng; Angom, Ramcharan Singh; Dutta, Kirthica; Ji, Baoan; Pirko, Istvan; Lucchinetti, Claudia F; Howe, Charles L; Mukhopadhyay, Debabrata

    2016-10-15

    Inflammatory response of blood-brain barrier (BBB) endothelial cells plays an important role in pathogenesis of many central nervous system inflammatory diseases, including multiple sclerosis; however, the molecular mechanism mediating BBB endothelial cell inflammatory response remains unclear. In this study, we first observed that knockdown of neuropilin-1 (NRP1), a co-receptor of several structurally diverse ligands, suppressed interferon-γ (IFNγ)-induced C-X-C motif chemokine 10 expression and activation of STAT1 in brain microvascular endothelial cells in a Rac1-dependent manner. Moreover, endothelial-specific NRP1-knockout mice, VECadherin-Cre-ERT2/NRP1flox/flox mice, showed attenuated disease progression during experimental autoimmune encephalomyelitis, a mouse neuroinflammatory disease model. Detailed analysis utilizing histological staining, quantitative PCR, flow cytometry and magnetic resonance imaging demonstrated that deletion of endothelial NRP1 suppressed neuron demyelination, altered lymphocyte infiltration, preserved BBB function and decreased activation of the STAT1-CXCL10 pathway. Furthermore, increased expression of NRP1 was observed in endothelial cells of acute multiple sclerosis lesions. Our data identify a new molecular mechanism of brain microvascular endothelial inflammatory response through NRP1-IFNγ crosstalk that could be a potential target for intervention of endothelial cell dysfunction in neuroinflammatory diseases. © 2016. Published by The Company of Biologists Ltd.

  16. An evolving new paradigm: endothelial cells--conditional innate immune cells.

    Science.gov (United States)

    Mai, Jietang; Virtue, Anthony; Shen, Jerry; Wang, Hong; Yang, Xiao-Feng

    2013-08-22

    Endothelial cells (ECs) are a heterogeneous population that fulfills many physiological processes. ECs also actively participate in both innate and adaptive immune responses. ECs are one of the first cell types to detect foreign pathogens and endogenous metabolite-related danger signals in the bloodstream, in which ECs function as danger signal sensors. Treatment with lipopolysaccharide activates ECs, causing the production of pro-inflammatory cytokines and chemokines, which amplify the immune response by recruiting immune cells. Thus, ECs function as immune/inflammation effectors and immune cell mobilizers. ECs also induce cytokine production by immune cells, in which ECs function as immune regulators either by activating or suppressing immune cell function. In addition, under certain conditions, ECs can serve as antigen presenting cells (antigen presenters) by expressing both MHC I and II molecules and presenting endothelial antigens to T cells. These facts along with the new concept of endothelial plasticity suggest that ECs are dynamic cells that respond to extracellular environmental changes and play a meaningful role in immune system function. Based on these novel EC functions, we propose a new paradigm that ECs are conditional innate immune cells. This paradigm provides a novel insight into the functions of ECs in inflammatory/immune pathologies.

  17. An evolving new paradigm: endothelial cells – conditional innate immune cells

    Science.gov (United States)

    2013-01-01

    Endothelial cells (ECs) are a heterogeneous population that fulfills many physiological processes. ECs also actively participate in both innate and adaptive immune responses. ECs are one of the first cell types to detect foreign pathogens and endogenous metabolite-related danger signals in the bloodstream, in which ECs function as danger signal sensors. Treatment with lipopolysaccharide activates ECs, causing the production of pro-inflammatory cytokines and chemokines, which amplify the immune response by recruiting immune cells. Thus, ECs function as immune/inflammation effectors and immune cell mobilizers. ECs also induce cytokine production by immune cells, in which ECs function as immune regulators either by activating or suppressing immune cell function. In addition, under certain conditions, ECs can serve as antigen presenting cells (antigen presenters) by expressing both MHC I and II molecules and presenting endothelial antigens to T cells. These facts along with the new concept of endothelial plasticity suggest that ECs are dynamic cells that respond to extracellular environmental changes and play a meaningful role in immune system function. Based on these novel EC functions, we propose a new paradigm that ECs are conditional innate immune cells. This paradigm provides a novel insight into the functions of ECs in inflammatory/immune pathologies. PMID:23965413

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

    Science.gov (United States)

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

    2017-07-01

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

  19. Mechanisms of xenogeneic baboon platelet aggregation and phagocytosis by porcine liver sinusoidal endothelial cells.

    Directory of Open Access Journals (Sweden)

    Qiang Peng

    Full Text Available Baboons receiving xenogeneic livers from wild type and transgenic pigs survive less than 10 days. One of the major issues is the early development of profound thrombocytopenia that results in fatal hemorrhage. Histological examination of xenotransplanted livers has shown baboon platelet activation, phagocytosis and sequestration within the sinusoids. In order to study the mechanisms of platelet consumption in liver xenotransplantation, we have developed an in vitro system to examine the interaction between pig endothelial cells with baboon platelets and to thereby identify molecular mechanisms and therapies.Fresh pig hepatocytes, liver sinusoidal and aortic endothelial cells were isolated by collagenase digestion of livers and processing of aortae from GTKO and Gal+ MGH-miniature swine. These primary cell cultures were then tested for the differential ability to induce baboon or pig platelet aggregation. Phagocytosis was evaluated by direct observation of CFSE labeled-platelets, which are incubated with endothelial cells under confocal light microscopy. Aurintricarboxylic acid (GpIb antagonist blocking interactions with von Willebrand factor/vWF, eptifibatide (Gp IIb/IIIa antagonist, and anti-Mac-1 Ab (anti-α(Mβ(2 integrin Ab were tested for the ability to inhibit phagocytosis.None of the pig cells induced aggregation or phagocytosis of porcine platelets. However, pig hepatocytes, liver sinusoidal and aortic endothelial cells (GTKO and Gal+ all induced moderate aggregation of baboon platelets. Importantly, pig liver sinusoidal endothelial cells efficiently phagocytosed baboon platelets, while pig aortic endothelial cells and hepatocytes had minimal effects on platelet numbers. Anti-MAC-1 Ab, aurintricarboxylic acid or eptifibatide, significantly decreased baboon platelet phagocytosis by pig liver endothelial cells (P<0.01.Although pig hepatocytes and aortic endothelial cells directly caused aggregation of baboon platelets, only pig liver

  20. Effect of Brugia malayi on the growth and proliferation of endothelial cells in vitro.

    Science.gov (United States)

    Rao, U R; Zometa, C S; Vickery, A C; Kwa, B H; Nayar, J K; Sutton, E T

    1996-08-01

    Athymic mice (C3H/HeN) parasitized by Brugia malayi develop massively dilated lymphatics. The lymphatic endothelial lining is perturbed, and numerous mononuclear and giant cells are closely apposed to the endothelium. The hyperplastic endothelial cells and low opening pressure of the lymphatics suggest abnormal multiplication of these cells may be important in the dilation. We studied the in vitro growth rate of human umbilical vein endothelial cells cultured with adult worms and microfilariae of B. malayi. The tetrazolium salt reduction assays were used to quantify possible direct mitogenic or inhibitory effects. The growth factor-induced proliferation of endothelial cells was significantly suppressed by 44-51% on day 1, 46-81% on day 3, and 45-79% on day 5 in cultures containing adult female worms, which had greater suppressor activity on endothelial cell proliferation than male worms, microfilariae, or soluble adult worm extract. Culture supernatant containing female worm excretory-secretory products significantly inhibited the growth and multiplication of cells, suggesting that adult female worms release antigens or proteins that have inhibitory activity on growth factors necessary for endothelial cell proliferation in vitro. Excess human recombinant epidermal growth factor and bovine brain extract partly reversed the inhibitory activity of worms in culture and restored the endothelial cell proliferation when incubated with worm culture supernatant. Indomethacin and BW 775Hcl failed to restore normal endothelial proliferation in the presence of female worms, suggesting that parasite-derived prostanoids and cyclooxygenase products did not cause the inhibition. Lymph from dilated lymphatics, but not serum from infected mice, increased the proliferation of cells in vitro. Together, these data demonstrate that excretory-secretory products of B. malayi parasites suppress vascular endothelial proliferation in vitro. Furthermore, increases in the number of these cells

  1. Towards a Biohybrid Lung: Endothelial Cells Promote Oxygen Transfer through Gas Permeable Membranes

    Directory of Open Access Journals (Sweden)

    Sarah Menzel

    2017-01-01

    Full Text Available In patients with respiratory failure, extracorporeal lung support can ensure the vital gas exchange via gas permeable membranes but its application is restricted by limited long-term stability and hemocompatibility of the gas permeable membranes, which are in contact with the blood. Endothelial cells lining these membranes promise physiological hemocompatibility and should enable prolonged application. However, the endothelial cells increase the diffusion barrier of the blood-gas interface and thus affect gas transfer. In this study, we evaluated how the endothelial cells affect the gas exchange to optimize performance while maintaining an integral cell layer. Human umbilical vein endothelial cells were seeded on gas permeable cell culture membranes and cultivated in a custom-made bioreactor. Oxygen transfer rates of blank and endothelialized membranes in endothelial culture medium were determined. Cell morphology was assessed by microscopy and immunohistochemistry. Both setups provided oxygenation of the test fluid featuring small standard deviations of the measurements. Throughout the measuring range, the endothelial cells seem to promote gas transfer to a certain extent exceeding the blank membranes gas transfer performance by up to 120%. Although the underlying principles hereof still need to be clarified, the results represent a significant step towards the development of a biohybrid lung.

  2. Cell adhesion and viability of human endothelial cells on electrospun polymer scaffolds

    OpenAIRE

    Matschegewski Claudia; Matthies Jörn-Bo; Grabow Niels; Schmitz Klaus-Peter

    2016-01-01

    The usage of electrospun polymer scaffolds is a promising approach for artificial heart valve design. This study aims at the evaluation of biological performance of nanofibrous polymer scaffolds poly(L-lactide) PLLA L210, PLLA L214 and polyamide-6 fabricated by electrospinning via analyzing viability, adhesion and morphology of human umbilical vein endothelial cells (EA.hy926). Nanofibrous surface topography was shown to influence cell phenotype and cell viability according to the observation...

  3. Endothelial cells are essential for ovarian stromal tissue restructuring after xenotransplantation of isolated ovarian stromal cells.

    Science.gov (United States)

    Dath, C; Dethy, A; Van Langendonckt, A; Van Eyck, A S; Amorim, C A; Luyckx, V; Donnez, J; Dolmans, M M

    2011-06-01

    Grafting of isolated follicles represents an approach to prevent the risk of reimplanting malignant cells with cryopreserved ovarian fragments. Optimal conditions and cell types required to sustain human follicular growth need to be identified. To help improve the grafting technique, we investigated whether short-term xenografting of a suspension containing ovarian stromal and endothelial cells without follicles could enhance graft survival and revascularization. In human ovary, CD34 selectively labels endothelial cells of blood vessels. A CD34-replete ovarian stromal cell group, including stromal and endothelial cells, was obtained after enzymatic digestion of fresh human ovarian cortex. Magnetic-activated cell sorting was used to establish a CD34-depleted ovarian stromal cell group. Proportions of CD34-positive cells were evaluated by flow cytometry and immunocytochemistry. Cell suspensions were embedded in human plasma clots and grafted (n = 10 for each group, 7 days) to the ovarian bursa of nude mice. Angiogenesis was quantified after human/mouse CD34 immunostaining. CD34-replete grafts had a well-organized and vascularized stromal structure, containing tubular components staining for human CD34 and corresponding to functional vessels, as evidenced by intraluminal red blood cells. CD34-depleted grafts tended to be smaller than CD34-replete grafts and poorly vascularized with central necrosis. Global microvessel density was higher in the CD34-replete than depleted group (337.9 versus 187.3 vessels/mm(2), P ovarian endothelial and stromal cells to ensure the formation of a well-vascularized and structured ovarian-like stroma after short-term xenografting, for future application in the transplantation of isolated follicles.

  4. Expansion of Endothelial Progenitor Cells in High Density Dot Culture of Rat Bone Marrow Cells

    Science.gov (United States)

    Wang, Ling; Kretlow, James D.; Zhou, Guangdong; Cao, Yilin; Liu, Wei; Zhang, Wen Jie

    2014-01-01

    In vitro expansion of endothelial progenitor cells (EPCs) remains a challenge in stem cell research and its application. We hypothesize that high density culture is able to expand EPCs from bone marrow by mimicking cell-cell interactions of the bone marrow niche. To test the hypothesis, rat bone marrow cells were either cultured in high density (2×105 cells/cm2) by seeding total 9×105 cells into six high density dots or cultured in regular density (1.6×104 cells/cm2) with the same total number of cells. Flow cytometric analyses of the cells cultured for 15 days showed that high density cells exhibited smaller cell size and higher levels of marker expression related to EPCs when compared to regular density cultured cells. Functionally, these cells exhibited strong angiogenic potentials with better tubal formation in vitro and potent rescue of mouse ischemic limbs in vivo with their integration into neo-capillary structure. Global gene chip and ELISA analyses revealed up-regulated gene expression of adhesion molecules and enhanced protein release of pro-angiogenic growth factors in high density cultured cells. In summary, high density cell culture promotes expansion of bone marrow contained EPCs that are able to enhance tissue angiogenesis via paracrine growth factors and direct differentiation into endothelial cells. PMID:25254487

  5. Influence of stromal cells on lymphocyte adhesion and migration on endothelial cells

    Science.gov (United States)

    McGettrick, Helen M.; Buckley, Chris D.; Rainger, G. Ed; Nash, Gerard B.

    2011-01-01

    Methods are described for analysing adhesion and migration of isolated lymphocytes on endothelial cell monolayers which have been co-cultured with different stromal cells, with or without additional cytokine treatment. The different cells types are grown on opposite sides of 3.0 or 0.4μm pore filters, depending on whether migration through the whole construct is to be analysed, or adhesion to the endothelial cells alone. Assays may be ‘static’ or filters can be incorporated into flow chambers so that cell behaviour can be directly observed under conditions simulating those in vivo. In general, by choice of method, one can evaluate efficiency of attachment, and ability of cells to migrate across the endothelial monolayer, through the filter and through the stromal cell layer. Fluorescence microscopic examination of fixed filters can be used e.g., to ascertain whether lymphocytes are retained by stromal cells. In general, static assays have the higher throughput and greatest ease of use, while the flow-based assays are more physiologically-relevant and allow detailed recording of cell behaviour in real time. PMID:20379868

  6. Fulvic Acid Attenuates Resistin-Induced Adhesion of HCT-116 Colorectal Cancer Cells to Endothelial Cells.

    Science.gov (United States)

    Huang, Wen-Shih; Yang, Jen-Tsung; Lu, Chien-Chang; Chang, Shun-Fu; Chen, Cheng-Nan; Su, Yu-Ping; Lee, Ko-Chao

    2015-12-09

    A high level of serum resistin has recently been found in patients with a number of cancers, including colorectal cancer (CRC). Hence, resistin may play a role in CRC development. Fulvic acid (FA), a class of humic substances, possesses pharmacological properties. However, the effect of FA on cancer pathophysiology remains unclear. The aim of this study was to investigate the effect of resistin on the endothelial adhesion of CRC and to determine whether FA elicits an antagonistic mechanism to neutralize this resistin effect. Human HCT-116 (p53-negative) and SW-48 (p53-positive) CRC cells and human umbilical vein endothelial cells (HUVECs) were used in the experiments. Treatment of both HCT-116 and SW-48 cells with resistin increases the adhesion of both cells to HUVECs. This result indicated that p53 may not regulate this resistin effect. A mechanistic study in HCT-116 cells further showed that this resistin effect occurs via the activation of NF-κB and the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Co-treating cells with both FA and resistin revealed that FA significantly attenuated the resistin-increased NF-κB activation and ICAM-1/VCAM-1 expression and the consequent adhesion of HCT-116 cells to HUVECs. These results demonstrate the role of resistin in promoting HCT-116 cell adhesion to HUVECs and indicate that FA might be a potential candidate for the inhibition of the endothelial adhesion of CRC in response to resistin.

  7. Resistance exercise increases endothelial progenitor cells and angiogenic factors.

    Science.gov (United States)

    Ross, Mark D; Wekesa, Antony L; Phelan, John P; Harrison, Michael

    2014-01-01

    Bone marrow-derived endothelial progenitor cells (EPC) are involved in vascular growth and repair. They increase in the circulation after a single bout of aerobic exercise, potentially related to muscle ischemia. Muscular endurance resistance exercise (MERE) bouts also have the potential to induce muscle ischemia if appropriately structured. The objective of this study is to determine the influence of a single bout of MERE on circulating EPC and related angiogenic factors. Thirteen trained men age 22.4 ± 0.5 yr (mean ± SEM) performed a bout of MERE consisting of three sets of six exercises at participants' 15-repetition maximum without resting between repetitions or exercises. The MERE bout duration was 12.1 ± 0.6 min. Blood lactate and HR were 11.9 ± 0.9 mmol·L and 142 ± 5 bpm, respectively, at the end of MERE. Blood was sampled preexercise and at 10 min, 2 h, and 24 h postexercise. Circulating EPC and serum concentrations of vascular endothelial growth factors (VEGF-A, VEGF-C, and VEGF-D), granulocyte colony stimulating factor, soluble Tie-2, soluble fms-like tyrosine kinase-1, and matrix metalloproteinases (MMP-1, MMP-2, MMP-3, MMP-9, and MMP-9) were higher (P < 0.05) in the postexercise period. Circulating EPC levels were unchanged at 10 min postexercise but higher at 2 h postexercise (P < 0.05). The concentration of most angiogenic factors and metalloproteinases were higher at 10 min postexercise (VEGF-A, +38%; VEGF-C, +40%; VEGF-D, +9%; soluble Tie-2, +15%; soluble fms-like tyrosine kinase-1, +24%; MMP-1, +62%; MMP-2, +3%; MMP-3, +54%; and MMP-9, +45%; all P < 0.05). Soluble E-selectin was lower (P < 0.05) at 2 and 24 h postexercise, with endothelial microparticles and thrombomodulin unchanged. Short intense bouts of MERE can trigger increases in circulating EPC and related angiogenic factors, potentially contributing to vascular adaptation and vasculoprotection.

  8. Hypotonic shock stimulates ascorbate release from coronary artery endothelial cells by a Ca2+ -independent pathway.

    Science.gov (United States)

    Davis, Kim A; Samson, Sue E; Wilson, John X; Grover, Ashok K

    2006-10-24

    In endothelial cells, anion channels open upon osmotic swelling during shear stress and hypotonic shock. Therefore, we examined the effects of hypotonic shock on release of the antioxidant anion ascorbate from pig coronary artery endothelial cells. Hypotonic shock potentiated ascorbate release from freshly isolated or cultured pig coronary artery endothelial cells; subsequently cultured endothelial cells were used. The hypotonic shock-induced increase in Asc release was rapid, depended on the degree of hypotonic shock, and not due to membrane leakiness. Stimulating P2Y2 like receptors in endothelial cells with ATP causes ascorbate release via a Ca2+ -mediated pathway. Hypotonic shock-induced release differed from the Ca2+-mediated Asc release because: (a) the increase in release with hypotonic shock was additive to that with ATP or A23187 (Ca2+ -ionophore), (b) apyrase, suramin or removing extracellular Ca2+ did not affect the hypotonic shock-stimulated release, (c) anion channel blockers inhibited the release by the two pathways differently, and (d) hypotonic shock increased the ascorbate release from endothelial cells and cultured smooth muscle cells whereas the Ca2+ -mediated ascorbate release occurred only in endothelial cells. Accumulation of ascorbate by endothelial cells was examined at extracellular ascorbate concentrations of 10 (Na+ -ascorbate symporter not saturated) and 5000 microM (Na+ -ascorbate symporter saturated). Hypotonic shock and A23187 decreased ascorbate accumulation at 10 microM ascorbate but increased it at 5000 microM. The effects of the two treatments were additive and also differed from each other with substitution of gluconate for extracellular chloride. Thus, ascorbate release from endothelial cells can be potentiated by two distinct pathways - hypotonic shock mediated and ATP/Ca2+ stimulated.

  9. Rapid synthesis of water-dispersible superparamagnetic iron oxide nanoparticles by a microwave-assisted route for safe labeling of endothelial progenitor cells.

    Science.gov (United States)

    Carenza, Elisa; Barceló, Verónica; Morancho, Anna; Montaner, Joan; Rosell, Anna; Roig, Anna

    2014-08-01

    We synthesize highly crystalline citrate-coated iron oxide superparamagnetic nanoparticles that are stable and readily dispersible in water by an extremely fast microwave-assisted route and investigate the uptake of magnetic nanoparticles by endothelial cells. Nanoparticles form large aggregates when added to complete endothelial cell medium. The size of the aggregates was controlled by adjusting the ionic strength of the medium. The internalization of nanoparticles into endothelial cells was then investigated by transmission electron microscopy, magnetometry and chemical analysis, together with cell viability assays. Interestingly, a sevenfold more efficient uptake was found for systems with larger nanoparticle aggregates, which also showed significantly higher magnetic resonance imaging effectiveness without compromising cell viability and functionality. We are thus presenting an example of a straightforward microwave synthesis of citrate-coated iron oxide nanoparticles for safe endothelial progenitor cell labeling and good magnetic resonance cell imaging with potential application for magnetic cell guidance and in vivo cell tracking. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  10. Endothelial cell death and intimal foam cell accumulation in the coronary artery of infected hypercholesterolemic minipigs

    DEFF Research Database (Denmark)

    Birck, Malene Muusfeldt; Saraste, Antti; Hyttel, Poul

    2013-01-01

    Apoptosis of endothelial cells (ECs) has been suggested to play a role in atherosclerosis. We studied the synergism of hypercholesterolemia with Chlamydia pneumoniae and influenza virus infections on EC morphology and intimal changes in a minipig model. The coronary artery was excised at euthanas...

  11. Soy diet inhibits expression of inflammation-induced vascular cell adhesion molecules in endothelial cells

    Science.gov (United States)

    Recently we reported that dietary soy attenuated atherogenesis in apolipoprotein E knockout (apoE-/-) mice. However, the molecular mechanisms contributing to the atheroprotective effect of soy-based diets is not clear. Since interactions between endothelial cells and monocytes play a pivotal role ...

  12. IL-20 activates human lymphatic endothelial cells causing cell signalling and tube formation

    DEFF Research Database (Denmark)

    Hammer, Troels; Tritsaris, Katerina; Hübschmann, Martin V

    2009-01-01

    IL-20 is an arteriogenic cytokine that remodels collateral networks in vivo, and plays a role in cellular organization. Here, we investigate its role in lymphangiogenesis using a lymphatic endothelial cell line, hTERT-HDLEC, which expresses the lymphatic markers LYVE-1 and podoplanin. Upon stimul...

  13. Extracellular Vesicles from Vascular Endothelial Cells Promote Survival, Proliferation and Motility of Oligodendrocyte Precursor Cells.

    Directory of Open Access Journals (Sweden)

    Masashi Kurachi

    Full Text Available We previously examined the effect of brain microvascular endothelial cell (MVEC transplantation on rat white matter infarction, and found that MVEC transplantation promoted remyelination of demyelinated axons in the infarct region and reduced apoptotic death of oligodendrocyte precursor cells (OPCs. We also found that the conditioned medium (CM from cultured MVECs inhibited apoptosis of cultured OPCs. In this study, we examined contribution of extracellular vesicles (EVs contained in the CM to its inhibitory effect on OPC apoptosis. Removal of EVs from the CM by ultracentrifugation reduced its inhibitory effect on OPC apoptosis. To confirm whether EVs derived from MVECs are taken up by cultured OPCs, we labeled EVs with PKH67, a fluorescent dye, and added them to OPC cultures. Many vesicular structures labeled with PKH67 were found within OPCs immediately after their addition. Next we examined the effect of MVEC-derived EVs on OPC behaviors. After 2 days in culture with EVs, there was significantly less pyknotic and more BrdU-positive OPCs when compared to control. We also examined the effect of EVs on motility of OPCs. OPCs migrated longer in the presence of EVs when compared to control. To examine whether these effects on cultured OPCs are shared by EVs from endothelial cells, we prepared EVs from conditioned media of several types of endothelial cells, and tested their effects on cultured OPCs. EVs from all types of endothelial cells we examined reduced apoptosis of OPCs and promoted their motility. Identification of the molecules contained in EVs from endothelial cells may prove helpful for establishment of effective therapies for demyelinating diseases.

  14. Adhesion of endothelial cells and adsorption of serum proteins on gas-plasma treated polytetrafluoroethylene

    NARCIS (Netherlands)

    Dekker, A.; Dekker, A.; Reitsma, K.; Beugeling, T.; Beugeling, T.; Bantjes, A.; Bantjes, A.; Feijen, Jan; van Aken, W.G.

    1991-01-01

    From in vitro experiments it is known that human endothelial cells show poor adhesion to hydrophobic polymers. The hydrophobicity of vascular prostheses manufactured from Teflon® or Dacron® may be the reason why endothelialization of these grafts does not occur after implantation in humans. We

  15. Experimental allogenic transplantation of cornea endothelial cells in cats.

    Science.gov (United States)

    Kiełbowicz, Z; Kuryszko, J; Strzadała, L

    2010-01-01

    The aim of the present study was assessing the possibility of experimental allogenic transplantation of cat cornea endothelial cells, multiplied in vitro, into the anterior chamber of the eyeball in recipient cats. The reason for undertaking the research is the need to develop a method that would help in the cornea treatment in animals with corneal opacification following cataract surgery, as well as lens dislocation, injuries and endothelium degeneration. Cats aged 10-12 months were used in the experiment. Cornea fragments consisting of the posterior limiting membrane and posterior epithelium were placed in Iscove's medium with addition of 10% foetal calf serum. Multiplied in vitro cells were injected into the anterior chamber of recipient cats. The cornea was subject to histological, histometric and SEM examination on the 3rd, 7th, 20th and 30th day after the surgery. Micromorphological examination of the cornea showed full restitution of its endothelium 30 days after transplantation. Complete regeneration of structures indispensable for normal functioning of the posterior epithelium occurred as a result of implantation. In this study the results show that implantation of the cells of posterior corneal epithelium of donor cats, multiplied into vitro and injected into the anterior chamber of recipient cats. The cornea regained its full function, the layer of the posterior epithelium was regenerated and the stroma stabilized, presenting the image of full and proper corneal translucency.

  16. Endothelial progenitor cells regenerate infracted myocardium with neovascularisation development

    Directory of Open Access Journals (Sweden)

    M.T. Abd El Aziz

    2015-03-01

    Full Text Available We achieved possibility of isolation, characterization human umbilical cord blood endothelial progenitor cells (EPCs, examination potency of EPCs to form new blood vessels and differentiation into cardiomyoctes in canines with acute myocardial infarction (AMI. EPCs were separated and cultured from umbilical cord blood. Their phenotypes were confirmed by uptake of double stains dioctadecyl tetramethylindocarbocyanine-labeled acetylated LDL and FITC-labeled Ulex europaeus agglutinin 1 (DILDL-UEA-1. EPCs of cord blood were counted. Human VEGFR-2 and eNOS from the cultured EPCs were assessed by qPCR. Human EPCs was transplanted intramyocardially in canines with AMI. ECG and cardiac enzymes (CK-MB and Troponin I were measured to assess severity of cellular damage. Histopathology was done to assess neovascularisation. Immunostaining was done to detect EPCs transdifferentiation into cardiomyocytes in peri-infarct cardiac tissue. qPCR for human genes (hVEGFR-2, and eNOS was done to assess homing and angiogenic function of transplanted EPCs. Cultured human cord blood exhibited an increased number of EPCs and significant high expression of hVEGFR-2 and eNOS genes in the culture cells. Histopathology showed increased neovascularization and immunostaining showed presence of EPCs newly differentiated into cardiomyocyte-like cells. Our findings suggested that hEPCs can mediate angiogenesis and differentiate into cardiomyoctes in canines with AMI.

  17. JNK2 promotes endothelial cell alignment under flow.

    Directory of Open Access Journals (Sweden)

    Cornelia Hahn

    Full Text Available Endothelial cells in straight, unbranched segments of arteries elongate and align in the direction of flow, a feature which is highly correlated with reduced atherosclerosis in these regions. The mitogen-activated protein kinase c-Jun N-terminal kinase (JNK is activated by flow and is linked to inflammatory gene expression and apoptosis. We previously showed that JNK activation by flow is mediated by integrins and is observed in cells plated on fibronectin but not on collagen or basement membrane proteins. We now show thatJNK2 activation in response to laminar shear stress is biphasic, with an early peak and a later peak. Activated JNK localizes to focal adhesions at the ends of actin stress fibers, correlates with integrin activation and requires integrin binding to the extracellular matrix. Reducing JNK2 activation by siRNA inhibits alignment in response to shear stress. Cells on collagen, where JNK activity is low, align slowly. These data show that an inflammatory pathway facilitates adaptation to laminar flow, thereby revealing an unexpected connection between adaptation and inflammatory pathways.

  18. Matrix stiffness regulates endothelial cell proliferation through septin 9.

    Directory of Open Access Journals (Sweden)

    Yi-Ting Yeh

    Full Text Available Endothelial proliferation, which is an important process in vascular homeostasis, can be regulated by the extracellular microenvironment. In this study we demonstrated that proliferation of endothelial cells (ECs was enhanced on hydrogels with high stiffness (HSG, 21.5 kPa in comparison to those with low stiffness (LSG, 1.72 kPa. ECs on HSG showed markedly prominent stress fibers and a higher RhoA activity than ECs on LSG. Blockade of RhoA attenuated stress fiber formation and proliferation of ECs on HSG, but had little effect on ECs on LSG; enhancement of RhoA had opposite effects. The phosphorylations of Src and Vav2, which are positive RhoA upstream effectors, were higher in ECs on HSG. The inhibition of Src/Vav2 attenuated the HSG-mediated RhoA activation and EC proliferation but exhibited nominal effects on ECs on LSG. Septin 9 (SEPT9, the negative upstream effector for RhoA, was significantly higher in ECs on LSG. The inhibition of SEPT9 increased RhoA activation, Src/Vav2 phosphorylations, and EC proliferation on LSG, but showed minor effects on ECs on HSG. We further demonstrated that the inactivation of integrin α(vβ(3 caused an increase of SEPT9 expression in ECs on HSG to attenuate Src/Vav2 phosphorylations and inhibit RhoA-dependent EC proliferation. These results demonstrate that the SEPT9/Src/Vav2/RhoA pathway constitutes an important molecular mechanism for the mechanical regulation of EC proliferation.

  19. Interleukin-19 decreases leukocyte-endothelial cell interactions by reduction in endothelial cell adhesion molecule mRNA stability

    Science.gov (United States)

    England, Ross N.; Preston, Kyle J.; Scalia, Rosario

    2013-01-01

    Vascular endothelial cell (EC) inflammation is a key event in the pathogenesis of multiple vascular diseases. We tested the hypothesis that interleukin-19 (IL-19), an anti-inflammatory Th2 interleukin, could have a direct anti-inflammatory effect on ECs to decrease inflammation. IL-19 can significantly decrease tumor necrosis factor (TNF)-α-driven intracellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 mRNA and protein abundance in cultured human coronary artery ECs (P adhesion to EC monolayers (P adhesion in wild-type mice as assayed by intravital microscopy (P cell adhesion and the first to propose reduction in HuR-mediated mRNA stability of ICAM-1 and VCAM-1 as a mechanism. Expression of IL-19 by ECs may represent a protective mechanism to promote resolution of the vascular response to inflammation. Function of IL-19 outside of the immune system is a novel concept, suggesting that resident vascular cells can adopt a Th2 phenotype, and has important ramifications for numerous inflammatory diseases. PMID:23596173

  20. CD13 is a novel mediator of monocytic/endothelial cell adhesion

    DEFF Research Database (Denmark)

    Mina-Osorio, Paola; Winnicka, Beata; O'Conor, Catherine

    2008-01-01

    During inflammation, cell surface adhesion molecules guide the adhesion and migration of circulating leukocytes across the endothelial cells lining the blood vessels to access the site of injury. The transmembrane molecule CD13 is expressed on monocytes and endothelial cells and has been shown...... to mediate homotypic cell adhesion, which may imply a role for CD13 in inflammatory monocyte trafficking. Here, we show that ligation and clustering of CD13 by mAb or viral ligands potently induce myeloid cell/endothelial adhesion in a signal transduction-dependent manner involving monocytic cytoskeletal...... rearrangement and filopodia formation. Treatment with soluble recombinant (r)CD13 blocks this CD13-dependent adhesion, and CD13 molecules from monocytic and endothelial cells are present in the same immunocomplex, suggesting a direct participation of CD13 in the adhesive interaction. This concept...

  1. Pathogen sensing pathways in human embryonic stem cell derived-endothelial cells: role of NOD1 receptors.

    Directory of Open Access Journals (Sweden)

    Daniel M Reed

    Full Text Available Human embryonic stem cell-derived endothelial cells (hESC-EC, as well as other stem cell derived endothelial cells, have a range of applications in cardiovascular research and disease treatment. Endothelial cells sense Gram-negative bacteria via the pattern recognition receptors (PRR Toll-like receptor (TLR-4 and nucleotide-binding oligomerisation domain-containing protein (NOD-1. These pathways are important in terms of sensing infection, but TLR4 is also associated with vascular inflammation and atherosclerosis. Here, we have compared TLR4 and NOD1 responses in hESC-EC with those of endothelial cells derived from other stem cells and with human umbilical vein endothelial cells (HUVEC. HUVEC, endothelial cells derived from blood progenitors (blood outgrowth endothelial cells; BOEC, and from induced pluripotent stem cells all displayed both a TLR4 and NOD1 response. However, hESC-EC had no TLR4 function, but did have functional NOD1 receptors. In vivo conditioning in nude rats did not confer TLR4 expression in hESC-EC. Despite having no TLR4 function, hESC-EC sensed Gram-negative bacteria, a response that was found to be mediated by NOD1 and the associated RIP2 signalling pathways. Thus, hESC-EC are TLR4 deficient but respond to bacteria via NOD1. This data suggests that hESC-EC may be protected from unwanted TLR4-mediated vascular inflammation, thus offering a potential therapeutic advantage.

  2. Micromanipulation of endothelial cells: Ultrasound-microbubble-cell interaction

    NARCIS (Netherlands)

    van Wamel, Annemieke; Bouakaz, Ayache; Versluis, Michel; de Jong, N.

    2004-01-01

    Ultrasound (US) in combination with contrast microbubbles has been shown to alter the permeability of cell membranes without affecting cell viability. This permeabilisation feature is used to design new drug delivery systems using US and contrast agents. The underlying mechanisms are still unknown.

  3. Ecscr regulates insulin sensitivity and predisposition to obesity by modulating endothelial cell functions.

    Science.gov (United States)

    Akakabe, Yoshiki; Koide, Masahiro; Kitamura, Youhei; Matsuo, Kiyonari; Ueyama, Tomomi; Matoba, Satoaki; Yamada, Hiroyuki; Miyata, Keishi; Oike, Yuichi; Ikeda, Koji

    2013-01-01

    Insulin resistance is closely associated with obesity and is one of the earliest symptoms of type-2 diabetes. Endothelial cells are involved in the pathogenesis of insulin resistance through their role in insulin delivery and adipose tissue angiogenesis. Here we show that Ecscr (endothelial cell surface expressed chemotaxis and apoptosis regulator; also known as ARIA), the transmembrane protein that regulates endothelial cell signalling, is highly expressed in white and brown adipose tissues, and regulates energy metabolism and glucose homeostasis by modulating endothelial cell functions. Ecscr-deficient mice fed a normal chow show improved glucose tolerance and enhanced insulin sensitivity. We demonstrate that Ecscr deletion enhances the insulin-mediated Akt/endothelial nitric oxide synthase activation in endothelial cells, which increases insulin delivery into the skeletal muscle. Ecscr deletion also protects mice on a high-fat diet from obesity and obesity-related metabolic disorders by enhancing adipose tissue angiogenesis. Conversely, targeted activation of Ecscr in endothelial cells impairs glucose tolerance and predisposes mice to diet-induced obesity. Our results suggest that the inactivation of Ecscr enhances insulin sensitivity and may represent a new therapeutic strategy for treating metabolic syndrome.

  4. Rapamycin Inhibits Proliferation of Hemangioma Endothelial Cells by Reducing HIF-1-Dependent Expression of VEGF

    Science.gov (United States)

    Medici, Damian; Olsen, Bjorn R.

    2012-01-01

    Hemangiomas are tumors formed by hyper-proliferation of vascular endothelial cells. This is caused by elevated vascular endothelial growth factor (VEGF) signaling through VEGF receptor 2 (VEGFR2). Here we show that elevated VEGF levels produced by hemangioma endothelial cells are reduced by the mTOR inhibitor rapamycin. mTOR activates p70S6K, which controls translation of mRNA to generate proteins such as hypoxia inducible factor-1 (HIF-1). VEGF is a known HIF-1 target gene, and our data show that VEGF levels in hemangioma endothelial cells are reduced by HIF-1α siRNA. Over-expression of HIF-1α increases VEGF levels and endothelial cell proliferation. Furthermore, both rapamycin and HIF-1α siRNA reduce proliferation of hemangioma endothelial cells. These data suggest that mTOR and HIF-1 contribute to hemangioma endothelial cell proliferation by stimulating an autocrine loop of VEGF signaling. Furthermore, mTOR and HIF-1 may be therapeutic targets for the treatment of hemangiomas. PMID:22900063

  5. Rapamycin inhibits proliferation of hemangioma endothelial cells by reducing HIF-1-dependent expression of VEGF.

    Directory of Open Access Journals (Sweden)

    Damian Medici

    Full Text Available Hemangiomas are tumors formed by hyper-proliferation of vascular endothelial cells. This is caused by elevated vascular endothelial growth factor (VEGF signaling through VEGF receptor 2 (VEGFR2. Here we show that elevated VEGF levels produced by hemangioma endothelial cells are reduced by the mTOR inhibitor rapamycin. mTOR activates p70S6K, which controls translation of mRNA to generate proteins such as hypoxia inducible factor-1 (HIF-1. VEGF is a known HIF-1 target gene, and our data show that VEGF levels in hemangioma endothelial cells are reduced by HIF-1α siRNA. Over-expression of HIF-1α increases VEGF levels and endothelial cell proliferation. Furthermore, both rapamycin and HIF-1α siRNA reduce proliferation of hemangioma endothelial cells. These data suggest that mTOR and HIF-1 contribute to hemangioma endothelial cell proliferation by stimulating an autocrine loop of VEGF signaling. Furthermore, mTOR and HIF-1 may be therapeutic targets for the treatment of hemangiomas.

  6. Interaction of human endothelial cells and nickel-titanium materials modified with silicon ions

    Energy Technology Data Exchange (ETDEWEB)

    Lotkov, Aleksandr I., E-mail: lotkov@ispms.tsc.ru; Kashin, Oleg A., E-mail: okashin@ispms.tsc.ru [Institute of Strength Physics and Materials Science SB RAS, Tomsk, 634055 (Russian Federation); Kudryavtseva, Yuliya A., E-mail: yulia-k1970@mail.ru; Antonova, Larisa V., E-mail: antonova.la@mail.ru; Matveeva, Vera G., E-mail: matveeva-vg@mail.ru; Sergeeva, Evgeniya A., E-mail: sergeewa.ew@yandex.ru [Research Institute for Complex Issues of Cardiovascular Diseases, Kemerovo, 650002 (Russian Federation); Kudryashov, Andrey N., E-mail: kudryashov@angioline.ru [Angioline Interventional Device Ltd, Novosibirsk, 630090 (Russian Federation)

    2015-10-27

    The paper studies the influence of chemical and phase compositions of NiTi surface layers modified with Si ions by plasma immersion implantation on their interaction with endothelial cells. It is shown that certain technological modes of Si ion implantation enhance the adhesion, proliferation, and viability of endothelial cells. It is found that the Si-modified NiTi surface is capable of stimulating the formation of capillary-like structures in the cell culture.

  7. Eimeria bovis infection modulates endothelial host cell cholesterol metabolism for successful replication

    OpenAIRE

    Hamid, Penny H.; Hirzmann, Joerg; Kerner, Katharina; Gimpl, Gerald; Lochnit, Guenter; Hermosilla, Carlos R.; Taubert, Anja

    2015-01-01

    During first merogony Eimeria bovis forms large macromeronts in endothelial host cells containing >120 000 merozoites I. During multiplication, large amounts of cholesterol are indispensable for the enormous offspring membrane production. Cholesterol auxotrophy was proven for other apicomplexan parasites. Consequently they scavenge cholesterol from their host cell apparently in a parasite-specific manner. We here analyzed the influence of E. bovis infection on endothelial host cell cholestero...

  8. Calcium handling in porcine coronary endothelial cells by gastrin-17.

    Science.gov (United States)

    Grossini, E; Molinari, C; Sigaudo, L; Biella, M; Mary, D A S G; Vacca, G

    2013-04-01

    In porcine coronary artery endothelial cells (PCAEC), gastrin-17 has recently been found to increase nitric oxide (NO) production by the endothelial NO synthase (eNOS) isoform through cholecystokinin 1/2 (CCK1/2) receptors and the involvement of protein kinase A (PKA), PKC and the β2-adrenoreceptor-related pathway. As eNOS is the Ca(2)(+)-dependent isoform of the enzyme, we aimed to examine the effects of gastrin-17 on Ca(2)(+) movements. Thus, experiments were performed in Fura-2-acetoxymethyl-ester-loaded PCAEC, where changes of cytosolic Ca(2)(+) ([Ca(2)(+)]c) caused by gastrin-17 were analysed and compared with those of CCK receptors and β2-adrenoreceptors agonists/antagonists. In addition, some experiments were performed by stimulating cells with gastrin-17 in the presence or absence of cAMP/PKA activator/inhibitor and of phospholipase C (PLC) and Ca(2)(+)-calmodulin dependent protein kinase II (CaMKII) blockers. The results have shown that gastrin-17 can promote a transient increase in [Ca(2)(+)]c mainly originating from an intracellular pool sensitive to thapsigargin and from the extracellular space. In addition, the response of cells to gastrin-17 was increased by the adenylyl cyclase activator and the β2-adrenoreceptor agonists and affected mainly by the CCK2 receptor agonists/antagonists. Moreover, the effects of gastrin-17 were prevented by β2-adrenoreceptors and CaMKII blockers and the adenylyl cyclase/PKA and PLC inhibitors. Finally, in PCAEC cultured in Na(+)-free medium or loaded with the plasma membrane Ca(2)(+) pump inhibitor, the gastrin-17-evoked Ca(2)(+) transient was long lasting. In conclusion, this study shows that gastrin-17 affected intracellular Ca(2)(+) homeostasis in PCAEC by both promoting a discharge of an intracellular pool and by interfering with the operation of store-dependent channels through mainly CCK2 receptors and PKA/PLC- and CaMKII-related signalling downstream of β2-adrenoreceptor stimulation.

  9. Responses of human endothelial cells to pathogenic and non-pathogenic Leptospira species.

    Directory of Open Access Journals (Sweden)

    Denise G Martinez-Lopez

    2010-12-01

    Full Text Available Leptospirosis is a widespread zoonotic infection that primarily affects residents of tropical regions, but causes infections in animals and humans in temperate regions as well. The agents of leptospirosis comprise several members of the genus Leptospira, which also includes non-pathogenic, saprophytic species. Leptospirosis can vary in severity from a mild, non-specific illness to severe disease that includes multi-organ failure and widespread endothelial damage and hemorrhage. To begin to investigate how pathogenic leptospires affect endothelial cells, we compared the responses of two endothelial cell lines to infection by pathogenic versus non-pathogenic leptospires. Microarray analyses suggested that pathogenic L. interrogans and non-pathogenic L. biflexa triggered changes in expression of genes whose products are involved in cellular architecture and interactions with the matrix, but that the changes were in opposite directions, with infection by L. biflexa primarily predicted to increase or maintain cell layer integrity, while L. interrogans lead primarily to changes predicted to disrupt cell layer integrity. Neither bacterial strain caused necrosis or apoptosis of the cells even after prolonged incubation. The pathogenic L. interrogans, however, did result in significant disruption of endothelial cell layers as assessed by microscopy and the ability of the bacteria to cross the cell layers. This disruption of endothelial layer integrity was abrogated by addition of the endothelial protective drug lisinopril at physiologically relevant concentrations. These results suggest that, through adhesion of L. interrogans to endothelial cells, the bacteria may disrupt endothelial barrier function, promoting dissemination of the bacteria and contributing to severe disease manifestations. In addition, supplementing antibiotic therapy with lisinopril or derivatives with endothelial protective activities may decrease the severity of leptospirosis.

  10. A role for the endothelial glycosaminoglycan hyaluronan in neutrophil recruitment by endothelial cells cultured for prolonged periods

    OpenAIRE

    Butler, Lynn M.; Rainger, G.Ed; Nash, Gerard B

    2009-01-01

    Glycosaminoglycans (GAGs) presented on the surface of endothelial cells (ECs) are believed to influence leukocyte recruitment during inflammation, but their roles remain uncertain. Here we report an in vitro model of prolonged culture of human EC in which the contributions of heparan sulphate (HS) and hyaluronan (HA) to the process of neutrophil recruitment could be studied. Previously, we reported that increasing EC culture duration (up to 20?days) enhanced neutrophil recruitment in response...

  11. An optimized method for mouse liver sinusoidal endothelial cell isolation

    Energy Technology Data Exchange (ETDEWEB)

    Meyer, Jeremy, E-mail: jeremy.meyer@hcuge.ch [Division of Digestive and Transplantation Surgery, University Hospitals of Geneva, Rue Gabrielle-Perret-Gentil 4, 1211 Genève 14 (Switzerland); Unit of Surgical Research, University of Geneva, Rue Michel-Servet 1, 1206 Genève (Switzerland); Lacotte, Stéphanie, E-mail: stephanie.lacotte@unige.ch [Unit of Surgical Research, University of Geneva, Rue Michel-Servet 1, 1206 Genève (Switzerland); Morel, Philippe, E-mail: philippe.morel@hcuge.ch [Division of Digestive and Transplantation Surgery, University Hospitals of Geneva, Rue Gabrielle-Perret-Gentil 4, 1211 Genève 14 (Switzerland); Unit of Surgical Research, University of Geneva, Rue Michel-Servet 1, 1206 Genève (Switzerland); Gonelle-Gispert, Carmen, E-mail: carmen.gonelle@unige.ch [Unit of Surgical Research, University of Geneva, Rue Michel-Servet 1, 1206 Genève (Switzerland); Bühler, Léo, E-mail: leo.buhler@hcuge.ch [Division of Digestive and Transplantation Surgery, University Hospitals of Geneva, Rue Gabrielle-Perret-Gentil 4, 1211 Genève 14 (Switzerland); Unit of Surgical Research, University of Geneva, Rue Michel-Servet 1, 1206 Genève (Switzerland)

    2016-12-10

    The objective of the present study was to develop an accurate and reproducible method for liver sinusoidal endothelial cell (LSEC) isolation in mice. Non-parenchymal cells were isolated using a modified two-step collagenase digestion combined with Optiprep density gradient centrifugation. LSEC were further purified using two prevalent methods, short-term selective adherence and CD146+ magnetic-activated cell sorting (MACS), and compared in terms of cell yield, viability and purity to our purification technique using CD11b cell depletion combined with long-term selective adherence. LSEC purification using our technique allowed to obtain 7.07±3.80 million LSEC per liver, while CD146+ MACS and short-term selective adherence yielded 2.94±1.28 and 0.99±0.66 million LSEC, respectively. Purity of the final cell preparation reached 95.10±2.58% when using our method. In contrast, CD146+ MACS and short-term selective adherence gave purities of 86.75±3.26% and 47.95±9.82%, respectively. Similarly, contamination by non-LSEC was the lowest when purification was performed using our technique, with a proportion of contaminating macrophages of only 1.87±0.77%. Further, isolated cells analysed by scanning electron microscopy presented typical LSEC fenestrations organized in sieve plates, demonstrating that the technique allowed to isolate bona fide LSEC. In conclusion, we described a reliable and reproducible technique for the isolation of high yields of pure LSEC in mice. This protocol provides an efficient method to prepare LSEC for studying their biological functions. - Highlights: • This protocol provides an efficient method to prepare primary mouse LSEC for studying their biological functions. • The liver cell dispersion step was improved by performing a retrograde cannulation of the liver. • The cell yield and the purity obtained were higher than comparative techniques in mice. • Contaminating macrophages were removed by introducing a CD11b- magnetic

  12. Fractalkine expression induces endothelial progenitor cell lysis by natural killer cells.

    Directory of Open Access Journals (Sweden)

    Dilyana Todorova

    Full Text Available BACKGROUND: Circulating CD34(+ cells, a population that includes endothelial progenitors, participate in the maintenance of endothelial integrity. Better understanding of the mechanisms that regulate their survival is crucial to improve their regenerative activity in cardiovascular and renal diseases. Chemokine-receptor cross talk is critical in regulating cell homeostasis. We hypothesized that cell surface expression of the chemokine fractalkine (FKN could target progenitor cell injury by Natural Killer (NK cells, thereby limiting their availability for vascular repair. METHODOLOGY/PRINCIPAL FINDINGS: We show that CD34(+-derived Endothelial Colony Forming Cells (ECFC can express FKN in response to TNF-α and IFN-γ inflammatory cytokines and that FKN expression by ECFC stimulates NK cell adhesion, NK cell-mediated ECFC lysis and microparticles release in vitro. The specific involvement of membrane FKN in these processes was demonstrated using FKN-transfected ECFC and anti-FKN blocking antibody. FKN expression was also evidenced on circulating CD34(+ progenitor cells and was detected at higher frequency in kidney transplant recipients, when compared to healthy controls. The proportion of CD34(+ cells expressing FKN was identified as an independent variable inversely correlated to CD34(+ progenitor cell count. We further showed that treatment of CD34(+ circulating cells isolated from adult blood donors with transplant serum or TNF-α/IFN-γ can induce FKN expression. CONCLUSIONS: Our data highlights a novel mechanism by which FKN expression on CD34(+ progenitor cells may target their NK cell mediated killing and participate to their immune depletion in transplant recipients. Considering the numerous diseased contexts shown to promote FKN expression, our data identify FKN as a hallmark of altered progenitor cell homeostasis with potential implications in better evaluation of vascular repair in patients.

  13. Lymphatic endothelial cells are a replicative niche for Mycobacterium tuberculosis

    Science.gov (United States)

    Lerner, Thomas R.; de Souza Carvalho-Wodarz, Cristiane; Repnik, Urska; Russell, Matthew R.G.; Borel, Sophie; Diedrich, Collin R.; Rohde, Manfred; Wainwright, Helen; Collinson, Lucy M.; Wilkinson, Robert J.; Griffiths, Gareth; Gutierrez, Maximiliano G.

    2016-01-01

    In extrapulmonary tuberculosis, the most common site of infection is within the lymphatic system, and there is growing recognition that lymphatic endothelial cells (LECs) are involved in immune function. Here, we identified LECs, which line the lymphatic vessels, as a niche for Mycobacterium tuberculosis in the lymph nodes of patients with tuberculosis. In cultured primary human LECs (hLECs), we determined that M. tuberculosis replicates both in the cytosol and within autophagosomes, but the bacteria failed to replicate when the virulence locus RD1 was deleted. Activation by IFN-γ induced a cell-autonomous response in hLECs via autophagy and NO production that restricted M. tuberculosis growth. Thus, depending on the activation status of LECs, autophagy can both promote and restrict replication. Together, these findings reveal a previously unrecognized role for hLECs and autophagy in tuberculosis pathogenesis and suggest that hLECs are a potential niche for M. tuberculosis that allows establishment of persistent infection in lymph nodes. PMID:26901813

  14. Influenza virus and endothelial cells: a species specific relationship

    Directory of Open Access Journals (Sweden)

    Kirsty Renfree Short

    2014-12-01

    Full Text Available Influenza A virus infection is an important cause of respiratory disease in humans. The original reservoirs of influenza A virus are wild waterfowl and shorebirds, where virus infection causes limited, if any, disease. Both in humans and in wild waterbirds, epithelial cells are the main target of infection. However, influenza virus can spread from wild bird species to terrestrial poultry. Here, the virus can evolve into highly pathogenic avian influenza (HPAI. Part of this evolution involves increased viral tropism for endothelial cells. HPAI virus infections not only cause severe disease in chickens and other terrestrial poultry species but can also spread to humans and back to wild bird populations. Here, we review the role of the endothelium in the pathogenesis of influenza virus infection in wild birds, terrestrial poultry and humans with a particular focus on HPAI viruses. We demonstrate that whilst the endothelium is an important target of virus infection in terrestrial poultry and some wild bird species, in humans the endothelium is more important in controlling the local inflammatory milieu. Thus, the endothelium plays an important, but species-specific, role in the pathogenesis of influenza virus infection.

  15. Influenza virus and endothelial cells: a species specific relationship.

    Science.gov (United States)

    Short, Kirsty R; Veldhuis Kroeze, Edwin J B; Reperant, Leslie A; Richard, Mathilde; Kuiken, Thijs

    2014-01-01

    Influenza A virus (IAV) infection is an important cause of respiratory disease in humans. The original reservoirs of IAV are wild waterfowl and shorebirds, where virus infection causes limited, if any, disease. Both in humans and in wild waterbirds, epithelial cells are the main target of infection. However, influenza virus can spread from wild bird species to terrestrial poultry. Here, the virus can evolve into highly pathogenic avian influenza (HPAI). Part of this evolution involves increased viral tropism for endothelial cells. HPAI virus infections not only cause severe disease in chickens and other terrestrial poultry species but can also spread to humans and back to wild bird populations. Here, we review the role of the endothelium in the pathogenesis of influenza virus infection in wild birds, terrestrial poultry and humans with a particular focus on HPAI viruses. We demonstrate that whilst the endothelium is an important target of virus infection in terrestrial poultry and some wild bird species, in humans the endothelium is more important in controlling the local inflammatory milieu. Thus, the endothelium plays an important, but species-specific, role in the pathogenesis of influenza virus infection.

  16. Involvement of leucocyte/endothelial cell interactions in anorexia nervosa.

    Science.gov (United States)

    Víctor, Víctor M; Rovira-Llopis, Susana; Saiz-Alarcón, Vanessa; Sangüesa, Maria C; Rojo-Bofill, Luis; Bañuls, Celia; de Pablo, Carmen; Álvarez, Ángeles; Rojo, Luis; Rocha, Milagros; Hernández-Mijares, Antonio

    2015-07-01

    Anorexia nervosa is a common psychiatric disorder in adolescence and is related to cardiovascular complications. Our aim was to study the effect of anorexia nervosa on metabolic parameters, leucocyte-endothelium interactions, adhesion molecules and proinflammatory cytokines. This multicentre, cross-sectional, case-control study employed a population of 24 anorexic female patients and 36 controls. We evaluated anthropometric and metabolic parameters, interactions between leucocytes polymorphonuclear neutrophils (PMN) and human umbilical vein endothelial cells (HUVEC), proinflammatory cytokines such as tumour necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) and soluble cellular adhesion molecules (CAMs) including E-selectin, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1). Anorexia nervosa was related to a decrease in weight, body mass index, waist circumference, systolic blood pressure, glucose, insulin and HOMA-IR, and an increase in HDL cholesterol. These effects disappeared after adjusting for BMI. Anorexia nervosa induced a decrease in PMN rolling velocity and an increase in PMN rolling flux and PMN adhesion. Increases in IL-6 and TNF-α and adhesion molecule VCAM-1 were also observed. This study supports the hypothesis of an association between anorexia nervosa, inflammation and the induction of leucocyte-endothelium interactions. These findings may explain, in part at least, the increased risk of vascular disease among patients with anorexia nervosa. © 2015 Stichting European Society for Clinical Investigation Journal Foundation.

  17. Endoglin is a novel endothelial cell specification gene.

    Science.gov (United States)

    Banerjee, Saswati; Dhara, Sujoy K; Bacanamwo, Methode

    2012-01-01

    Endothelial cells (EC) are important in vasculogenesis and organogenesis during development and in the pathogenesis of cancer and cardiovascular diseases. However, few EC specification factors are known and primary EC production remains inefficient. Based on recent studies implicating endoglin (Eng) in early vascular development and angiogenesis, we hypothesized that Eng may be an EC specification gene. Mouse embryonic stem cells (ESC) were treated with recombinant Eng or a plasmid expressing the Eng ORF, and differentiated in the presence or absence of bone morphogenic protein 4 (BMP4). Expression of the mesoderm and EC marker genes, the known mediators of EC specification and their downstream targets was monitored by quantitative PCR, western blot, immunocytochemistry, and flow cytometry. Functionality of the differentiated EC was assessed by in vitro angiogenesis assay and the induction of Icam1 expression in response to TNF-α treatment. Both recombinant Eng and forced Eng expression increased the number of functional EC expressing the EC marker genes VE-cadherin, vWF, and Tie2, and enhanced the effect of BMP4. The Eng-induced EC differentiation was independent of known mediators of EC specification such as Indian Hedgehog (IHH) and BMP4 or of BMP4/Smad1/5/8 signaling. These studies suggest that Eng is a novel EC specification gene. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. The Effects of kisspeptin-10 on Migration and Proliferation of Endothelial Cell

    Directory of Open Access Journals (Sweden)

    Fatemeh Golzar

    2015-01-01

    Full Text Available Background: Migration, expansion and survival of endothelial cells that are an important cellular component of blood vessels plays an important role in the induction of tumor growth. Kisspeptins (kp, peptides that bind to coupled-G protein receptor (GPR54, inhibit each step of metastatic cascade include invasion, migration and homing, angiogenesis, survival and proliferation. In this study we investigated effects of kisspeptin-10, the most potent member of kisspeptin family, on Migration and proliferation of endothelial cells that are necessary for angiogenesis and tumor metastasis. Materials and Methods: We compared migration of Human Umbilical Vein Endothelial Cells (HUVECs were treated with 10-100 or 500 nM kp-10 for 24 hours and no treated cells using an in vitro trans membrane migration assay and HUVEC proliferation of treated endothelial cells with 10-100 or 500 nM kp-10 for 48 hours and no treated cells was measured by MTT Cell Proliferation Assay Kit. Analysis of data was performed using the Kruskal-Wallis test followed by the Mann-Whitney test. Results: Migration and proliferation of endothelial cells were increased at lower concentration of kp-10 specially at 100 nM while higher concentration reduced both migration and proliferation. Conclusion: Our data showed that different concentrations of kp-10 have distinct effects on migration and proliferation of endothelial cells.

  19. A nanoengineered peptidic delivery system with specificity for human brain capillary endothelial cells

    DEFF Research Database (Denmark)

    Wu, Linping; Moghimi, Seyed Moein

    2016-01-01

    avidity of the majority of the so-called ‘brain-specific’ nanoparticles to the brain capillary endothelial cells has been poor, even during in vitro conditions. We have addressed this issue and designed a versatile peptidic nanoplatform with high binding avidity to the human cerebral capillary endothelial...... cells. This was achieved by selecting an appropriate phage-derived peptide with high specificity for human brain capillary endothelial cells, which following careful structural modifications spontaneously formed a nanoparticle-fiber network. The peptidic network was characterized fully and its uptake...... by the human brain capillary endothelial cell line hCMEC/D3 was confirmed by live-cell fluorescent microscopy and quantified by flow cytometry. Recognition and internalization was medicated by two receptors leading to endolysosomal accumulation. Furthermore, the network was capable of delivering functional si...

  20. Effect of onion peel extract on endothelial function and endothelial progenitor cells in overweight and obese individuals.

    Science.gov (United States)

    Choi, Eun-Yong; Lee, Hansongyi; Woo, Jong Shin; Jang, Hyun Hee; Hwang, Seung Joon; Kim, Hyun Soo; Kim, Woo-Sik; Kim, Young-Seol; Choue, Ryowon; Cha, Yong-Jun; Yim, Jung-Eun; Kim, Weon

    2015-09-01

    Acute or chronic intake of polyphenol-rich foods has been reported to improve endothelial function. Quercetin, found abundantly in onion, is a potent antioxidant flavonoid. The aim of this study was to investigate whether consumption of onion peel extract (OPE) improves endothelial function in healthy overweight and obese individuals. This was a randomized double-blind, placebo-controlled study. Seventy-two healthy overweight and obese participants were randomly assigned to receive a red, soft capsule of OPE (100 mg quercetin/d, 50 mg quercetin twice daily; n = 36 participants) or an identical placebo capsule (n = 36) for 12 wk. Endothelial function, defined by flow-mediated dilation (FMD), circulating endothelial progenitor cells (EPCs) by flow cytometry, and laboratory test were determined at baseline and after treatment. Baseline characteristics and laboratory findings did not significantly differ between the two groups. Compared with baseline values, the OPE group showed significantly improved FMD at 12 wk (from 12.5 ± 5.2 to 15.2 ± 6.1; P = 0.002), whereas the placebo group showed no difference. Nitroglycerin-mediated dilation did not change in either group. EPC counts (44.2 ± 25.6 versus 52.3 ± 18.6; P = 0.005) and the percentage of EPCs were significantly increased in the OPE group. When FMD was divided into quartiles, rate of patients with endothelial dysfunction defined as lowest quartile (cutoff value, 8.6%) of FMD improved from 26% to 9% by OPE. Medium-term administration of OPE an improvement in FMD and circulating EPCs. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Regulation of staphylococcal enterotoxin B-elicited nitric oxide production by endothelial cells.

    Science.gov (United States)

    LeClaire, R D; Kell, W M; Sadik, R A; Downs, M B; Parker, G W

    1995-01-01

    The effect of staphylococcal enterotoxin B (SEB)-elicited inducible nitric oxide synthase (iNOS) in mouse endothelial cells was investigated. Results showed that SEB stimulated the same level of NO production in gamma interferon (IFN-gamma)-primed cells as did trichloroacetic acid-extracted lipopolysaccharide. The kinetics of induced NO production and expression of mRNA for iNOS differed markedly in endothelial and macrophage cells. Induced endothelial nitrite production was transient and was 15 to 20% of that generated by macrophage cells; mRNA levels peaked by 2 h and then steadily declined, whereas macrophage message levels continually increased. The ability of endothelial cells to produce SEB-induced NO depended on priming with IFN-gamma, although detectable mRNA could be elicited by SEB alone. Induction of endothelial iNOS mRNA was inhibited by cycloheximide, which indicated a requirement for de novo protein synthesis. Niacinamide and interleukin-10 significantly reduced SEB-induced endothelial NO production. Both are reported to affect IFN-gamma-induced class II major histocompatibility complex (MHC) expression on antigen-presenting cells. Niacinamide reduced iNOS mRNA levels and markedly reduced IFN-gamma induction of endothelial class II MHC surface antigen. Interleukin-10 did not consistently reduce iNOS mRNA expression and had no effect on IFN-gamma induction of endothelial class II MHC surface antigen. These results suggest that SEB interacts with IFN-gamma-primed endothelial cells to elicit induced NO and that this induction can be effectively modulated at the receptor or transcriptional level. PMID:7529748

  2. Palmitoylation of caveolin-1 in endothelial cells is post-translational but irreversible

    Science.gov (United States)

    Parat, M. O.; Fox, P. L.

    2001-01-01

    Caveolin-1 is a palmitoylated protein involved in assembly of signaling molecules in plasma membrane subdomains termed caveolae and in intracellular cholesterol transport. Three cysteine residues in the C terminus of caveolin-1 are subject to palmitoylation, which is not necessary for caveolar targeting of caveolin-1. Protein palmitoylation is a post-translational and reversible modification that may be regulated and that in turn may regulate conformation, membrane association, protein-protein interactions, and intracellular localization of the target protein. We have undertaken a detailed analysis of [(3)H]palmitate incorporation into caveolin-1 in aortic endothelial cells. The linkage of palmitate to caveolin-1 was hydroxylamine-sensitive and thus presumably a thioester bond. However, contrary to expectations, palmitate incorporation was blocked completely by the protein synthesis inhibitors cycloheximide and puromycin. In parallel experiments to show specificity, palmitoylation of aortic endothelial cell-specific nitric-oxide synthase was unaffected by these reagents. Inhibitors of protein trafficking, brefeldin A and monensin, blocked caveolin-1 palmitoylation, indicating that the modification was not cotranslational but rather required caveolin-1 transport from the endoplasmic reticulum and Golgi to the plasma membrane. In addition, immunophilin chaperones that form complexes with caveolin-1, i.e. FK506-binding protein 52, cyclophilin A, and cyclophilin 40, were not necessary for caveolin-1 palmitoylation because agents that bind immunophilins did not inhibit palmitoylation. Pulse-chase experiments showed that caveolin-1 palmitoylation is essentially irreversible because the release of [(3)H]palmitate was not significant even after 24 h. These results show that [(3)H]palmitate incorporation is limited to newly synthesized caveolin-1, not because incorporation only occurs during synthesis but because the continuous presence of palmitate on caveolin-1 prevents

  3. Distribution of blood-group-related carbohydrate antigens on oral endothelial cells.

    Science.gov (United States)

    Bryne, M; Lilleholt, S; Thrane, P S; Koppang, H S; Dabelsteen, E

    1993-05-01

    Recent studies indicate that carbohydrate structures are involved in various endothelial functions such as inflammatory processes, adhesion of metastatic cells to endothelium and endothelial differentiation. In this paper we report the endothelial expression of various blood-group-related carbohydrate structures in normal oral mucosa using 15 different monoclonal antibodies reacting with type 1, 2 or 3 carbohydrate chains. Twenty biopsies, including normal oral mucosa, from secretor individuals comprised nine blood group O, nine A, one B and one AB. Endothelial staining was compared with epithelial staining in the same biopsies. Five blood-group-related carbohydrate antigens were detected on endothelial cells. The H type 2 antigen, which is the precursor of A, B and AB antigens, has previously been believed to be a universal marker for endothelial cells. All blood group O individuals (n = 9) showed strong H antigen staining in the endothelium of most vessels. However, of blood group A, B and AB individuals (n = 11), four showed heterogenous H antigen staining. In addition, we found that six out of ten blood group A or AB individuals, who expressed A or A and B antigens on spinous squamous cells and glandular epithelium, showed either heterogenous or no staining for these structures on their endothelial cells. It is concluded that there are differences between the biosynthesis of blood-group-related carbohydrate antigens in oral endothelium and epithelium.

  4. Comparison of Endothelial Differentiation Capacities of Human and Rat Adipose-Derived Stem Cells.

    Science.gov (United States)

    Orbay, Hakan; Devi, Kamaljit; Williams, Priscilla A; Dehghani, Tima; Silva, Eduardo A; Sahar, David E

    2016-12-01

    The authors compared the endothelial differentiation capacities of human and rat adipose-derived stem cells to determine whether human adipose-derived stem cells can be a source of endothelial cells clinically. Human and rat adipose-derived stem cells were harvested and characterized with flow cytometry and trilineage differentiation. Cells from passages III through V were fed with endothelial cell differentiation medium for up to 3 weeks. Cells were harvested after 1, 2, and 3 weeks, and endothelial differentiation was evaluated with quantitative reverse-transcriptase polymerase chain reaction, flow cytometry, and angiogenic sprouting assays. Both human and rat adipose-derived stem cells were CD90, CD44, and CD31 before differentiation. The cells were successfully differentiated into adipogenic, osteogenic, and chondrogenic lineages. Expression of endothelial cell-specific genes peaked at the second week of differentiation in both human and rat cells. The fold changes in expression of CD31, vascular endothelial growth factor receptor-1, nitric oxide synthase, and von Willebrand factor genes at week 2 were 0.4 ± 0.1, 34.7 ± 0.3, 2.03 ± 0.25, and 12.5 ± 0.3 respectively, in human adipose-derived stem cells; and 1.5 ± 1.01, 21.6 ± 1.7, 17.9 ± 0.6, and 11.2 ± 1.3, respectively, in rat cells. The percentages of CD31 cells were 0.2, 0.64, and 1.6 in human cell populations and 0.5, 5.91, and 11.5 in rat cell populations at weeks 1, 2, and 3, respectively. Rat adipose-derived stem cell-derived endothelial cells displayed enhanced sprouting capability compared with the human cells. Human adipose-derived stem cells responded less strongly to EGM-2MV endothelial differentiation medium than did the rat cells. Still, the human cells have the potential to become a clinical source of endothelial cells with modifications in the differentiation conditions.

  5. Induced Pluripotent Stem (iPS) Cell Culture Methods and Induction of Differentiation into Endothelial Cells.

    Science.gov (United States)

    Chatterjee, Ishita; Li, Fei; Kohler, Erin E; Rehman, Jalees; Malik, Asrar B; Wary, Kishore K

    2016-01-01

    The study of stem cell behavior and differentiation in a developmental context is complex, time-consuming, and expensive, and for this reason, cell culture remains a method of choice for developmental and regenerative biology and mechanistic studies. Similar to ES cells, iPS cells have the ability to differentiate into endothelial cells (ECs), and the route for differentiation appears to mimic the developmental process that occurs during the formation of an embryo. Traditional EC induction methods from embryonic stem (ES) cells rely mostly on the formation of embryoid body (EB), which employs feeder or feeder-free conditions in the presence or absence of supporting cells. Similar to ES cells, iPS cells can be cultured in feeder layer or feeder-free conditions. Here, we describe the iPS cell culture methods and induction differentiation of these cells into ECs. We use anti-mouse Flk1 and anti-mouse VE-cadherin to isolate and characterize mouse ECs, because these antibodies are commercially available and their use has been described in the literature, including by our group. The ECs produced by this method have been used by our laboratory, and we have demonstrated their in vivo potential. We also discuss how iPS cells differ in their ability to differentiate into endothelial cells in culture.

  6. Degradable poly(apigenin) polymer inhibits tumor cell adhesion to vascular endothelial cells.

    Science.gov (United States)

    Cochran, David B; Gray, Lindsay N; Anderson, Kimberly W; Dziubla, Thomas D

    2016-10-01

    Cancer and the inflammatory system share a complex intertwined relationship. For instance, in response to an injury or stress, vascular endothelial cells will express cell adhesion molecules as a means of recruiting leukocytes. However, circulating tumor cells (CTCs) have been shown to highjack this expression for the adhesion and invasion during the metastatic cascade. As such, the initiation of endothelial cell inflammation, either by surgical procedures (cancer resection) or chemotherapy can inadvertently increase the metastatic potential of CTCs. Yet, systemic delivery of anti-inflammatories, which weaken the entire immune system, may not be preferred in some treatment settings. In this work, we demonstrate that a long-term releasing flavone-based polymer and subsequent nanoparticle delivery system can inhibit tumor cell adhesion, through the suppression of endothelial cell adhesion molecule expression. The degradation of a this anti-inflammatory polymer provides longer term, localized release profile of active therapeutic drug in nanoparticle form as compared with that of the free drug, permitting more targeted anti-metastatic therapies. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 1438-1447, 2016. © 2015 Wiley Periodicals, Inc.

  7. Morphological heterogeneity with normal expression but altered function of G proteins in porcine cultured regenerated coronary endothelial cells

    Science.gov (United States)

    Borg-Capra, Catherine; Fournet-Bourguignon, Marie-Pierre; Janiak, Philip; Villeneuve, Nicole; Bidouard, Jean-Pierre; Vilaine, Jean-Paul; Vanhoutte, Paul M

    1997-01-01

    Experiments were designed to investigate whether the pertussis toxin-dependent endothelial dysfunction following balloon injury is due to a reduced expression or an insufficient function of G-proteins. Endothelium-dependent responses of porcine coronary arteries were examined in vitro by use of conventional organ chambers. Morphological analysis was performed by isolating and culturing the endothelial cells from these arteries. The expression of Gi-proteins in regenerated endothelial cells was measured by Western blots and immunolabelling. The function of G-proteins was assessed by measuring the GTPase activity of cultured endothelial cells. Eight days following denudation, endothelial regrowth was confirmed by histological examination and by demonstrating the presence of endothelium-dependent relaxations to bradykinin and 5-hydroxytryptamine (5-HT). In primary culture, the regenerated endothelial cells displayed a ‘cobblestone' pattern as seen with native endothelial cells. Twenty eight days after denudation, the endothelium-dependent relaxations induced by 5-HT were impaired, but those to bradykinin were maintained. However, the latter were reduced when endothelium-dependent hyperpolarization was prevented. Twenty eight days after denudation, multinucleated giant cells were present in the regenerated but not in the native cultured endothelial cell populations. These regenerated endothelial cells incorporated less tritiated thymidine than native endothelial cells. The intensities of the bands on the immunoblot of the regenerated endothelial cells, when several antibodies against Giα1/α2/α3 were used, were the same as those obtained in native endothelial cells. The immunolabelling with the same antibodies was similar between the giant cells and the regenerated endothelial cells of normal size. The hydrolysis of GTP was lower in regenerated than in native endothelial cell membranes. In conclusion, endothelium-dependent relaxations mediated by Gi-proteins are

  8. ERG is required for the differentiation of embryonic stem cells along the endothelial lineage

    Directory of Open Access Journals (Sweden)

    Le Bras Alexandra

    2009-12-01

    Full Text Available Abstract Background The molecular mechanisms that govern stem cell differentiation along the endothelial lineage remain largely unknown. Ets related gene (ERG has recently been shown to participate in the transcriptional regulation of a number of endothelial specific genes including VE-cadherin (CD144, endoglin, and von Willebrand's Factor (vWF. The specific role of the ETS factor ERG during endothelial differentiation has not been evaluated. Results ERG expression and function were evaluated during the differentiation of embryonic stem cells into embryoid bodies (EB. The results of our study demonstrate that ERG is first expressed in a subpopulation of vascular endothelial growth factor receptor 2 (VEGF-R2 expressing cells that also express VE-cadherin. During ES cell differentiation, ERG expression remains restricted to cells of the endothelial lineage that eventually coalesce into primitive vascular structures within embryoid bodies. ERG also exhibits an endothelial cell (EC-restricted pattern during embryogenesis. To further define the role of ERG during ES cell differentiation, we used a knockdown strategy to inhibit ERG expression. Delivery of three independent shRNA led to 70-85% reductions in ERG expression during ES cell differentiation compared to no change with control shRNA. ERG knockdown was associated with a marked reduction in the number of ECs, the expression of EC-restricted genes, and the formation of vascular structures. Conclusion The ETS factor ERG appears to be a critical regulator of EC differentiation.

  9. Reduced proliferation of endothelial colony-forming cells in unprovoked venous thromboembolic disease as a consequence of endothelial dysfunction

    Science.gov (United States)

    Hernandez-Lopez, Rubicel; Chavez-Gonzalez, Antonieta; Torres-Barrera, Patricia; Moreno-Lorenzana, Dafne; Lopez-DiazGuerrero, Norma; Santiago-German, David; Isordia-Salas, Irma; Smadja, David; C. Yoder, Mervin; Majluf-Cruz, Abraham

    2017-01-01

    Background Venous thromboembolic disease (VTD) is a public health problem. We recently reported that endothelial colony-forming cells (ECFCs) derived from endothelial cells (EC) (ECFC-ECs) from patients with VTD have a dysfunctional state. For this study, we proposed that a dysfunctional status of these cells generates a reduction of its proliferative ability, which is also associated with senescence and reactive oxygen species (ROS). Methods and results Human mononuclear cells (MNCs) were obtained from peripheral blood from 40 healthy human volunteers (controls) and 50 patients with VTD matched by age (20−50 years) and sex to obtain ECFCs. We assayed their proliferative ability with plasma of patients and controls and supernatants of cultures from ECFC-ECs, senescence-associated β-galactosidase (SA-β-gal), ROS, and expression of ephrin-B2/Eph-B4 receptor. Compared with cells from controls, cells from VTD patients showed an 8-fold increase of ECFCs that emerged 1 week earlier, reduced proliferation at long term (39%) and, in passages 4 and 10, a highly senescent rate (30±1.05% vs. 91.3±15.07%, respectively) with an increase of ROS and impaired expression of ephrin-B2/Eph-4 genes. Proliferation potential of cells from VTD patients was reduced in endothelial medium [1.4±0.22 doubling population (DP)], control plasma (1.18±0.31 DP), or plasma from VTD patients (1.65±0.27 DP). Conclusions As compared with controls, ECFC-ECs from individuals with VTD have higher oxidative stress, proliferation stress, cellular senescence, and low proliferative potential. These findings suggest that patients with a history of VTD are ECFC-ECs dysfunctional that could be associated to permanent risk for new thrombotic events. PMID:28910333

  10. Photodynamic response of an endothelial hybridoma cell line using zinc(II) tetrasubstituted phthalocyanines

    Science.gov (United States)

    Cruse-Sawyer, Janet E.; Dixon, B.; Roberts, David J.; Griffiths, John; Brown, Stanley B.

    1995-03-01

    The EAhy 926 cell is a hybridoma line derived from human endothelium and A549/8 cells. They display stable endothelial characteristics and may provide an indication of how endothelial cells respond to photodynamic therapy. Cells were grown as monolayers, seeded at a density of 104 cells/35 mm dish, and then incubated with zinc (II) tetrasubstituted phthalocyanines (carboxylated, sulphonated, pyridinium or diethanolamine sulphonamide). After 24 hours, the cells were illuminated with laser light at 680 nm or 692 nm as appropriate. The response to each photosensitizer was evaluated using cell proliferation, clonogenicity, and release of tissue factor.

  11. Chorein Sensitivity of Actin Polymerization, Cell Shape and Mechanical Stiffness of Vascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Ioana Alesutan

    2013-09-01

    Full Text Available Background/Aims: Endothelial cell stiffness plays a key role in endothelium-dependent control of vascular tone and arterial blood pressure. Actin polymerization and distribution of microfilaments is essential for mechanical cell stiffness. Chorein, a protein encoded by the VPS13A gene, defective in chorea-acanthocytosis (ChAc, is involved in neuronal cell survival as well as cortical actin polymerization of erythrocytes and blood platelets. Chorein is expressed in a wide variety of further cells, yet nothing is known about the impact of chorein on cells other than neurons, erythrocytes and platelets. The present study explored whether chorein is expressed in human umbilical vein endothelial cells (HUVECs and addressed the putative role of chorein in the regulation of cytoskeletal architecture, stiffness and survival of those cells. Methods: In HUVECs with or without silencing of the VPS13A gene, VPS13A mRNA expression was determined utilizing quantitative RT-PCR, cytoskeletal organization visualized by confocal microscopy, G/F actin ratio and phosphorylation status of focal adhesion kinase quantified by western blotting, cell death determined by flow cytometry, mechanical properties studied by atomic force microscopy (AFM and cell morphology analysed by scanning ion conductance microscopy (SICM. Results: VPS13A mRNA expression was detectable in HUVECs. Silencing of the VPS13A gene attenuated the filamentous actin network, decreased the ratio of soluble G-actin over filamentous F-actin, reduced cell stiffness and changed cell morphology as compared to HUVECs silenced with negative control siRNA. These effects were paralleled by a significant decrease in FAK phosphorylation following VPS13A silencing. Moreover, silencing of the VPS13A gene increased caspase 3 activity and induced necrosis in HUVECs. Conclusions: Chorein is a novel regulator of cytoskeletal architecture, cell shape, mechanical stiffness and survival of vascular endothelial cells.

  12. Impaired Endothelial Repair Capacity of Early Endothelial Progenitor Cells in Hypertensive Patients With Primary Hyperaldosteronemia: Role of 5,6,7,8-Tetrahydrobiopterin Oxidation and Endothelial Nitric Oxide Synthase Uncoupling.

    Science.gov (United States)

    Chen, Long; Ding, Mei-Lin; Wu, Fang; He, Wen; Li, Jin; Zhang, Xiao-Yu; Xie, Wen-Li; Duan, Sheng-Zhong; Xia, Wen-Hao; Tao, Jun

    2016-02-01

    Although hyperaldosteronemia exerts detrimental impacts on vascular endothelium in addition to elevating blood pressure, the effects and molecular mechanisms of hyperaldosteronemia on early endothelial progenitor cell (EPC)-mediated endothelial repair after arterial damage are yet to be determined. The aim of this study was to investigate the endothelial repair capacity of early EPCs from hypertensive patients with primary hyperaldosteronemia (PHA). In vivo endothelial repair capacity of early EPCs from PHAs (n=20), age- and blood pressure-matched essential hypertension patients (n=20), and age-matched healthy subjects (n=20) was evaluated by transplantation into a nude mouse carotid endothelial denudation model. Endothelial function was evaluated by flow-mediated dilation of brachial artery in human subjects. In vivo endothelial repair capacity of early EPCs and flow-mediated dilation were impaired both in PHAs and in essential hypertension patients when compared with age-matched healthy subjects; however, the early EPC in vivo endothelial repair capacity and flow-mediated dilation of PHAs were impaired more severely than essential hypertension patients. Oral spironolactone improved early EPC in vivo endothelial repair capacity and flow-mediated dilation of PHAs. Increased oxidative stress, oxidative 5,6,7,8-tetrahydrobiopterin degradation, endothelial nitric oxide synthase uncoupling and decreased nitric oxide production were found in early EPCs from PHAs. Nicotinamide adenine dinucleotide phosphate oxidase subunit p47(phox) knockdown or 5,6,7,8-tetrahydrobiopterin supplementation attenuated endothelial nitric oxide synthase uncoupling and enhanced in vivo endothelial repair capacity of early EPCs from PHAs. In conclusion, PHAs exhibited more impaired endothelial repair capacity of early EPCs than did essential hypertension patients independent of blood pressure, which was associated with mineralocorticoid receptor-dependent oxidative stress and subsequently 5

  13. Upregulation of the low density lipoprotein receptor at the blood-brain barrier: intercommunications between brain capillary endothelial cells and astrocytes

    OpenAIRE

    1994-01-01

    In contrast to the endothelial cells in large vessels where LDL receptors are downregulated, brain capillary endothelial cells in vivo express an LDL receptor. Using a cell culture model of the blood-brain barrier consisting of a coculture of brain capillary endothelial cells and astrocytes, we observed that the capacity of endothelial cells to bind LDL is enhanced threefold when cocultured with astrocytes. We next investigated the ability of astrocytes to modulate endothelial cell LDL recept...

  14. Fibroblast growth factor signaling pathway in endothelial cells is activated by BMPER to promote angiogenesis.

    Science.gov (United States)

    Esser, Jennifer S; Rahner, Susanne; Deckler, Meike; Bode, Christoph; Patterson, Cam; Moser, Martin

    2015-02-01

    Previously, we have identified bone morphogenetic protein endothelial cell precursor-derived regulator (BMPER) to increase the angiogenic activity of endothelial cells in a concentration-dependent manner. In this project, we now investigate how BMPER acts in concert with key molecules of angiogenesis to promote blood vessel formation. To assess the effect of BMPER on angiogenesis-related signaling pathways, we performed an angiogenesis antibody array with BMPER-stimulated endothelial cells. We detected increased basic fibroblast growth factor (bFGF/FGF-2) expression after BMPER stimulation and decreased expression of thrombospondin-1. Additionally, FGF receptor-1 expression, phosphorylation, FGF signaling pathway activity, and cell survival were increased. Consistently, silencing of BMPER by small interfering RNA decreased bFGF and FGF receptor-1 expression and increased thrombospondin-1 expression and cell apoptosis. Next, we investigated the interaction of BMPER and the FGF signaling pathway in endothelial cell function. BMPER stimulation increased endothelial cell angiogenic activity in migration, Matrigel, and spheroid assays. To block FGF signaling, an anti-bFGF antibody was used, which effectively inhibited the proangiogenic BMPER effect. Accordingly, BMPER-silenced endothelial cells under bFGF stimulation showed decreased angiogenic activity compared with bFGF control. We confirmed these findings in vivo by subcutaneous Matrigel injections with and without bFGF in C57BL/6_Bmper(+/-) mice. Aortic ring assays of C57BL/6_Bmper(+/-) mice confirmed a specific effect for bFGF but not for vascular endothelial growth factor. Taken together, the proangiogenic BMPER effect in endothelial cells is mediated by inhibition of antiangiogenic thrombospondin-1 and enhanced expression and activation of the FGF signaling pathway that is crucial in the promotion of angiogenesis. © 2014 American Heart Association, Inc.

  15. Chronic hypoxia attenuates VEGF signaling and angiogenic responses by downregulation of KDR in human endothelial cells.

    Science.gov (United States)

    Olszewska-Pazdrak, Barbara; Hein, Travis W; Olszewska, Paulina; Carney, Darrell H

    2009-05-01

    Coronary artery disease results in progressive vascular stenosis associated with chronic myocardial ischemia. Vascular endothelial growth factor (VEGF) stimulates endothelial cell angiogenic responses to revascularize ischemic tissues; however, the effect of chronic hypoxia on the responsiveness of endothelial cells to VEGF remains unclear. We, therefore, investigated whether hypoxia alters VEGF-stimulated signaling and angiogenic responses in primary human coronary artery endothelial (HCAE) cells. Exposure of HCAE cells to hypoxia (1% O(2)) for 24 h decreased VEGF-stimulated endothelial cell migration ( approximately 82%), proliferation ( approximately 30%), and tube formation. Hypoxia attenuated VEGF-stimulated activation of endothelial nitric oxide (NO) synthase (eNOS) ( approximately 72%) and reduced NO production in VEGF-stimulated cells from 237 +/- 38.8 to 61.3 +/- 28.4 nmol/l. Moreover, hypoxia also decreased the ratio of phosphorylated eNOS to total eNOS in VEGF-stimulated cells by approximately 50%. This effect was not observed in thrombin-stimulated cells, suggesting that hypoxia specifically inhibited VEGF signaling upstream of eNOS phosphorylation. VEGF-induced activation of Akt, ERK1/2, p38, p70S6 kinases, and S6 ribosomal protein was also attenuated in hypoxic cells. Moreover, VEGF-stimulated phosphorylation of VEGF receptor-2 (KDR) at Y996 and Y1175 was decreased by hypoxia. This decrease correlated with a 70 +/- 12% decrease in KDR protein expression. Analysis of mRNA from these cells showed that hypoxia reduced steady-state levels of KDR mRNA by 52 +/- 16% and decreased mRNA stability relative to normoxic cells. Our findings demonstrate that chronic hypoxia attenuates VEGF-stimulated signaling in HCAE cells by specific downregulation of KDR expression. These data provide a novel explanation for the impaired angiogenic responses to VEGF in endothelial cells exposed to chronic hypoxia.

  16. Effect of syncytiotrophoblast microvillous membrane treatment on gene expression in human umbilical vein endothelial cells

    DEFF Research Database (Denmark)

    Høgh, Mette; Tannetta, D; Sargent, I

    2006-01-01

    Objective Syncytiotrophoblast membrane fragments (STBM) exist in the peripheral circulation in pregnant women and it has been shown that the level of circulating STBM is significantly increased with pre-eclampsia compared with uncomplicated pregnancies. STBM could be one of the factors which...... directly causes the endothelial cell dysfunction of pre-eclampsia. This study investigates the effect of STBM on endothelial cell gene expression. Design Human umbilical vein endothelial cells were cultured in the presence and absence of STBM. At specified time points, total RNA was purified from...... results. Results Overall, the results do not show any great changes in gene expression in endothelial cells after STBM treatment (28 genes changed two-fold or more out of approximately 10 000 genes examined by microarray). In general, the changes observed are consistent with inhibition of proliferation...

  17. Effect of syncytiotrophoblast microvillous membrane treatment on gene expression in human umbilical vein endothelial cells

    DEFF Research Database (Denmark)

    Hoegh, A M; Tannetta, D; Sargent, I

    2006-01-01

    OBJECTIVE: Syncytiotrophoblast membrane fragments (STBM) exist in the peripheral circulation in pregnant women and it has been shown that the level of circulating STBM is significantly increased with pre-eclampsia compared with uncomplicated pregnancies. STBM could be one of the factors which...... directly causes the endothelial cell dysfunction of pre-eclampsia. This study investigates the effect of STBM on endothelial cell gene expression. DESIGN: Human umbilical vein endothelial cells were cultured in the presence and absence of STBM. At specified time points, total RNA was purified from...... results. RESULTS: Overall, the results do not show any great changes in gene expression in endothelial cells after STBM treatment (28 genes changed two-fold or more out of approximately 10,000 genes examined by microarray). In general, the changes observed are consistent with inhibition of proliferation...

  18. Protective properties of artichoke (Cynara scolymus) against oxidative stress induced in cultured endothelial cells and monocytes.

    Science.gov (United States)

    Zapolska-Downar, Danuta; Zapolski-Downar, Andrzej; Naruszewicz, Marek; Siennicka, Aldona; Krasnodebska, Barbara; Kołdziej, Blanka

    2002-11-01

    It is currently believed that oxidative stress and inflammation play a significant role in atherogenesis. Artichoke extract exhibits hypolipemic properties and contains numerous active substances with antioxidant properties in vitro. We have studied the influence of aqueous and ethanolic extracts from artichoke on intracellular oxidative stress stimulated by inflammatory mediators (TNFalpha and LPS) and ox-LDL in endothelial cells and monocytes. Oxidative stress which reflects the intracellular production of reactive oxygen species (ROS) was followed by measuring the oxidation of 2', 7'-dichlorofluorescin (DCFH) to 2', 7'-dichlorofluorescein (DCF). Agueous and ethanolic extracts from artichoke were found to inhibit basal and stimulated ROS production in endothelial cells and monocytes in dose dependent manner. In endothelial cells, the ethanolic extract (50 microg/ml) reduced ox-LDL-induced intracellular ROS production by 60% (partichoke extracts have marked protective properties against oxidative stress induced by inflammatory mediators and ox-LDL in cultured endothelial cells and monocytes.

  19. Fluid shear stress induces the phosphorylation of small heat shock proteins in vascular endothelial cells

    National Research Council Canada - National Science Library

    Li, S; Piotrowicz, R S; Levin, E G; Shyy, Y J; Chien, S

    1996-01-01

    .... In bovine aortic endothelial cells stably transfected with the wild-type human HSP27 gene, shear stress induced the phosphorylation of both the exogenous human HSP27 and the endogenous bovine HSP25...

  20. Low density lipoprotein inhibits accumulation of nitrites in murine brain endothelial cell cultures.

    Science.gov (United States)

    Bereta, M; Bereta, J; Cohen, S; Cohen, M C

    1992-07-15

    Endothelial cells produce nitric oxide which is considered to serve as a major source of endothelial derived relaxing factor activity. It has been demonstrated that activation of mouse brain endothelium by TNF-alpha and IFN-gamma led to accumulation of nitrite which is presumably formed by oxidation of nitric oxide. A number of studies suggest that reactive oxygen species produced by cytokine-activated cells are involved in the conversion of nitric oxide to nitrites and nitrates. We investigated whether low density lipoprotein (LDL), acting as a radical scavenger, is able to inhibit nitrite accumulation in mouse brain endothelial cell cultures and in a cell-free system in which sodium nitroprusside was used as a source of nitric oxide. A comparison of these two models indicates the active involvement of LDL in suppressing nitrite accumulation in murine endothelial cultures.