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Sample records for human arterial endothelial

  1. Muscle contraction induced arterial shear stress increases endothelial nitric oxide synthase phosphorylation in humans.

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    Casey, Darren P; Ueda, Kenichi; Wegman-Points, Lauren; Pierce, Gary L

    2017-10-01

    We determined if local increases in brachial artery shear during repetitive muscle contractions induce changes in protein expression of endothelial nitric oxide synthase (eNOS) and/or phosphorylated (p-)eNOS at Ser1177, the primary activation site on eNOS, in endothelial cells (ECs) of humans. Seven young male subjects (25 ± 1 yr) performed 20 separate bouts (3 min each) of rhythmic forearm exercise at 20% of maximum over a 2-h period. Each bout of exercise was separated by 3 min of rest. An additional six male subjects (24 ± 1 yr) served as time controls (no exercise). ECs were freshly isolated from the brachial artery using sterile J-wires through an arterial catheter at baseline and again after the 2-h exercise or time control period. Expression of eNOS or p-eNOS Ser1177 in ECs was determined via immunofluorescence. Brachial artery mean shear rate was elevated compared with baseline and the time control group throughout the 2-h exercise protocol (P 0.05 for both). Our novel results suggest that elevations in brachial artery shear increase eNOS Ser1177 phosphorylation in the absence of changes in total eNOS in ECs of young healthy male subjects, suggesting that this model is sufficient to alter posttranslational modification of eNOS activity in vivo in humans.NEW & NOTEWORTHY Elevations in brachial artery shear in response to forearm exercise increased endothelial nitric oxide synthase Ser1177 phosphorylation in brachial artery endothelial cells of healthy humans. Our present study provides the first evidence in humans that muscle contraction-induced increases in conduit arterial shear lead to in vivo posttranslational modification of endothelial nitric oxide synthase activity in endothelial cells. Copyright © 2017 the American Physiological Society.

  2. Endothelial shear stress: a critical determinant of arterial remodeling and arterial stiffness in humans--a carotid 3.0-T MRI study.

    NARCIS (Netherlands)

    Duivenvoorden, R.; Bavel, E. van; Groot, E. de; Stroes, E.S.; Disselhorst, J.A.; Hutten, B.A.; Lameris, J.S.; Kastelein, J.J.; Nederveen, A.J.

    2010-01-01

    BACKGROUND: Low endothelial shear stress (ESS) elicits endothelial dysfunction. However, the relationship between ESS and arterial remodeling and arterial stiffness is unknown in humans. We developed a 3.0-T MRI protocol to evaluate the contribution of ESS to arterial remodeling and stiffness.

  3. Double-stranded RNA attenuates the barrier function of human pulmonary artery endothelial cells.

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    Zoltán Bálint

    Full Text Available Circulating RNA may result from excessive cell damage or acute viral infection and can interact with vascular endothelial cells. Despite the obvious clinical implications associated with the presence of circulating RNA, its pathological effects on endothelial cells and the governing molecular mechanisms are still not fully elucidated. We analyzed the effects of double stranded RNA on primary human pulmonary artery endothelial cells (hPAECs. The effect of natural and synthetic double-stranded RNA (dsRNA on hPAECs was investigated using trans-endothelial electric resistance, molecule trafficking, calcium (Ca(2+ homeostasis, gene expression and proliferation studies. Furthermore, the morphology and mechanical changes of the cells caused by synthetic dsRNA was followed by in-situ atomic force microscopy, by vascular-endothelial cadherin and F-actin staining. Our results indicated that exposure of hPAECs to synthetic dsRNA led to functional deficits. This was reflected by morphological and mechanical changes and an increase in the permeability of the endothelial monolayer. hPAECs treated with synthetic dsRNA accumulated in the G1 phase of the cell cycle. Additionally, the proliferation rate of the cells in the presence of synthetic dsRNA was significantly decreased. Furthermore, we found that natural and synthetic dsRNA modulated Ca(2+ signaling in hPAECs by inhibiting the sarco-endoplasmic Ca(2+-ATPase (SERCA which is involved in the regulation of the intracellular Ca(2+ homeostasis and thus cell growth. Even upon synthetic dsRNA stimulation silencing of SERCA3 preserved the endothelial monolayer integrity. Our data identify novel mechanisms by which dsRNA can disrupt endothelial barrier function and these may be relevant in inflammatory processes.

  4. Growth hormone-releasing peptide ghrelin inhibits homocysteine-induced endothelial dysfunction in porcine coronary arteries and human endothelial cells.

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    Hedayati, Nasim; Annambhotla, Suman; Jiang, Jun; Wang, Xinwen; Chai, Hong; Lin, Peter H; Yao, Qizhi; Chen, Changyi

    2009-01-01

    Ghrelin, a novel growth hormone-releasing peptide, is implicated to play a protective role in cardiovascular tissues. However, it is not clear whether ghrelin protects vascular tissues from injury secondary to risk factors such as homocysteine (Hcy). This study investigated the effect and potential mechanisms of ghrelin on Hcy-induced endothelial dysfunction. Porcine coronary artery rings were incubated for 24 hours with ghrelin (100 ng/mL), Hcy (50 microM), or ghrelin plus Hcy. Endothelial vasomotor function was evaluated using the myograph tension model. The response to the thromboxane A(2)analog U46619, bradykinin, and sodium nitroprusside was analyzed. Endothelial nitric oxide synthase (eNOS) expression was determined using real-time polymerase chain reaction and immunohistochemistry staining, and superoxide anion production was documented lucigenin-enhanced chemiluminescence analysis. Human coronary artery endothelial cells (HCAECs) were treated with different concentrations of Hcy, ghrelin, or antighrelin receptor antibody for 24 hours, and eNOS protein levels were determined by Western blot analysis. Maximal contraction with U46619 and endothelium-independent vasorelaxation with sodium nitroprusside were not different among the four groups. However, endothelium-dependent vasorelaxation with bradykinin (10(-6) M) was significantly reduced by 34% with Hcy compared with controls (P ghrelin to Hcy had a protective effect, with 61.6% relaxation, which was similar to controls (64.7%). Homocysteine significantly reduced eNOS expression, whereas ghrelin cotreatment effectively restored eNOS expression to the control levels. Superoxide anion levels, which were increased by 100% with Hcy, returned to control levels with ghrelin cotreatment. Ghrelin also effectively blocked the Hcy-induced decrease of eNOS protein levels in HCAECs in a concentration-dependent manner. Antighrelin receptor antibody effectively inhibited the effect of ghrelin. Ghrelin has a protective

  5. Transcriptional Remodeling of Ion Channel Subunits by Flow Adaptation in Human Coronary Artery Endothelial Cells

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    Kefaloyianni, Eirini; Coetzee, William A.

    2011-01-01

    Endothelial cells (ECs) are constantly exposed to blood flow-induced shear forces in the vessels and this is a major determinant of endothelial function. Ion channels have a major role in endothelial function and in the control of vascular tone. We hypothesized that shear force is a general regulator of ion channel expression, which will have profound effects on endothelial function. We examined this hypothesis using large-scale quantitative real-time RT-PCR. Human coronary artery ECs were exposed to two levels of flow-induced shear stress for 24 h, while control cells were grown under static conditions. The expression of ion channel subunits was compared between control and flow-adapted cells. We used primers against 55 ion channel and exchanger subunits and were able to detect 54 subunits. Five dyn/cm2 of shear induced downregulation of 1 (NCX1) and upregulation of 18 subunits, including KCa2.2, KCa2.3, CX37, Kv1.5 and HCN2. Fifteen dyn/cm2 of shear stress induced the expression of 30 ion channel subunits, including KCa2.3, KCa2.2, CX37, Kir2.3 and KCa3.1. Our data demonstrate that substantial remodeling of endothelial ion channel subunit expression occurs with flow adaptation and suggest that altered ion channel expression may significantly contribute to vascular pathology associated with flow-induced alterations. PMID:21389733

  6. GLP-1 inhibits VEGFA-mediated signaling in isolated human endothelial cells and VEGFA-induced dilation of rat mesenteric arteries

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    Rotbøl, Cecilie Egholm; Khammy, Makhala Michell; Dalsgaard, Thomas

    2016-01-01

    to PLCγ activation, Src, and endothelial NOS (eNOS) signaling, thereby controlling endothelial vessel tone. By using RT-PCR analysis, we found mRNA for the GLP-1 receptor (GLP-1R) in human dermal microvascular endothelial cells (HDMEC), human retinal microvascular endothelial cells, and rat arteries...

  7. Biological Atomic Force Microscopy for Imaging Gold-Labeled Liposomes on Human Coronary Artery Endothelial Cells

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    Ana-María Zaske

    2013-01-01

    Full Text Available Although atomic force microscopy (AFM has been used extensively to characterize cell membrane structure and cellular processes such as endocytosis and exocytosis, the corrugated surface of the cell membrane hinders the visualization of extracellular entities, such as liposomes, that may interact with the cell. To overcome this barrier, we used 90 nm nanogold particles to label FITC liposomes and monitor their endocytosis on human coronary artery endothelial cells (HCAECs in vitro. We were able to study the internalization process of gold-coupled liposomes on endothelial cells, by using AFM. We found that the gold-liposomes attached to the HCAEC cell membrane during the first 15–30 min of incubation, liposome cell internalization occurred from 30 to 60 min, and most of the gold-labeled liposomes had invaginated after 2 hr of incubation. Liposomal uptake took place most commonly at the periphery of the nuclear zone. Dynasore monohydrate, an inhibitor of endocytosis, obstructed the internalization of the gold-liposomes. This study showed the versatility of the AFM technique, combined with fluorescent microscopy, for investigating liposome uptake by endothelial cells. The 90 nm colloidal gold nanoparticles proved to be a noninvasive contrast agent that efficiently improves AFM imaging during the investigation of biological nanoprocesses.

  8. Recombinant human activated protein C improves endotoxemia-induced endothelial dysfunction: a blood-free model in isolated mouse arteries.

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    Sennoun, Nacira; Baron-Menguy, Celine; Burban, Mélanie; Lecompte, Thomas; Andriantsitohaina, Ramaroson; Henrion, Daniel; Mercat, Alain; Asfar, Pierre; Levy, Bruno; Meziani, Ferhat

    2009-07-01

    Recombinant human activated protein C (rhAPC) is one of the treatment panels for improving vascular dysfunction in septic patients. In a previous study, we reported that rhAPC treatment in rat endotoxemia improved vascular reactivity, although the mechanisms involved are still under debate. In the present study, we hypothesized that rhAPC may improve arterial dysfunction through its nonanticoagulant properties. Ten hours after injection of LPS in mice (50 mg/kg ip), aortic rings and mesenteric arteries were isolated and incubated with or without rhAPC for 12 h. Aortic rings were mounted in a myograph, after which arterial contractility and endothelium-dependent relaxation were measured in the presence or absence of nitric oxide synthase or cyclooxygenase inhibitors. Flow (shear stress)-mediated dilation with or without the above inhibitors was also measured in mesenteric resistance arteries. Protein expression was assessed by Western blotting. Lipopolysaccharide (LPS) reduced aortic contractility to KCl and phenylephrine as well as dilation to acetylcholine. LPS also reduced flow-mediated dilation in mesenteric arteries. In rhAPC-treated aorta and mesenteric arteries, contractility and endothelial responsiveness to vasodilator drug and shear stress were improved. rhAPC treatment also improved LPS-induced endothelial dysfunction; this effect was associated with an increase in the phosphorylated form of endothelial nitric oxide synthase and protein kinase B as well as cyclooxygenase vasodilatory pathways, thus suggesting that these pathways, together with the decrease in nuclear factor-kappaB activation and inducible nitric oxide synthase expression in the vascular wall, are implicated in the endothelial effect of rhAPC. In conclusion, ex vivo application of rhAPC improves arterial contractility and endothelial dysfunction resulting from endotoxemia in mice. This finding provides important insights into the mechanism underlying rhAPC-induced improvements on arterial

  9. Colocalization of serum amyloid a with microtubules in human coronary artery endothelial cells.

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    Lakota, Katja; Resnik, Nataša; Mrak-Poljšak, Katjuša; Sodin-Šemrl, Snežna; Veranič, Peter

    2011-01-01

    Serum amyloid A (SAA) acts as a major acute phase protein and represents a sensitive and accurate marker of inflammation. Besides its hepatic origin, as the main source of serum SAA, this protein is also produced extrahepatically. The mRNA levels of SAA become significantly elevated following proinflammatory stimuli, as well as, are induced through their own positive feedback in human primary coronary artery endothelial cells. However, the intracellular functions of SAA are so far unknown. Colocalization of SAA with cytoskeletal filaments has previously been proposed, so we analyzed the colocalization of SAA with all three cytoskeletal elements: actin filaments, vimentin filaments, and microtubules. Immunofluorescent double-labeling analyses confirmed by PLA method revealed a strict colocalization of SAA with microtubules and a very infrequent attachment to vimentin while the distribution of actin filaments appeared clearly separated from SAA staining. Also, no significant colocalization was found between SAA and endomembranes labeled with the fluorescent lipid stain DiO₆. However, SAA appears to be located also unbound in the cytosol, as well as inside the nucleus and within nanotubes extending from the cells or bridging neighboring cells. These different locations of SAA in endothelial cells strongly indicate multiple potential functions of this protein.

  10. Colocalization of Serum Amyloid A with Microtubules in Human Coronary Artery Endothelial Cells

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    Katja Lakota

    2011-01-01

    Full Text Available Serum amyloid A (SAA acts as a major acute phase protein and represents a sensitive and accurate marker of inflammation. Besides its hepatic origin, as the main source of serum SAA, this protein is also produced extrahepatically. The mRNA levels of SAA become significantly elevated following proinflammatory stimuli, as well as, are induced through their own positive feedback in human primary coronary artery endothelial cells. However, the intracellular functions of SAA are so far unknown. Colocalization of SAA with cytoskeletal filaments has previously been proposed, so we analyzed the colocalization of SAA with all three cytoskeletal elements: actin filaments, vimentin filaments, and microtubules. Immunofluorescent double-labeling analyses confirmed by PLA method revealed a strict colocalization of SAA with microtubules and a very infrequent attachment to vimentin while the distribution of actin filaments appeared clearly separated from SAA staining. Also, no significant colocalization was found between SAA and endomembranes labeled with the fluorescent lipid stain DiO6. However, SAA appears to be located also unbound in the cytosol, as well as inside the nucleus and within nanotubes extending from the cells or bridging neighboring cells. These different locations of SAA in endothelial cells strongly indicate multiple potential functions of this protein.

  11. Facilitative interaction between angiotensin II and oxidised LDL in cultured human coronary artery endothelial cells

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    Jawahar L Mehta

    2001-03-01

    Full Text Available Background Several studies have shown that angiotensin II (Ang II and oxidised low-density lipoprotein (ox-LDL are critical factors in atherosclerosis. In this study, we examined the molecular basis of mutually facilitative interactions between Ang II and ox-LDL in human coronary artery endothelial cells (HCAECs.Methods and results We observed that incubation of cultured HCAECs with Ang II (10-12 to 10-6 M for 24 hours caused a concentration-dependent increase in the expression of mRNA and protein of a specialised receptor for ox-LDL (LOX-1. These effects of Ang II were completely blocked by pretreatment of HCAECs with candesartan (10-6 M, a specific AT1-receptor blocker, but not by PD 123319 (10-6 M, a specific AT2-receptor blocker. On the other hand, incubation of HCAECs with ox-LDL (10 and 40 µg/ml for 24 hours progressively upregulated AT1-, but not AT 2-, receptor mRNA and protein. Pretreatment of cells with the anti-oxidant alpha-tocopherol (1—5 x 10-6 M inhibited the upregulation of AT1-receptor expression induced by ox-LDL (p<0.05. To determine the significance of expression of AT1-receptors and LOX-1, we measured cell injury in response to Ang II and ox-LDL. Incubation of cells with both ox-LDL and Ang II synergistically increased cell injury, measured as cell viability and LDH release, compared with either ox-LDL or Ang II alone (both p<0.05. Alpha-tocopherol, as well as candesartan, attenuated cell injury in response to Ang II and ox-LDL (both p<0.05.Conclusions These observations show that Ang II upregulates a novel endothelial receptor for ox-LDL (LOX-1 gene expression and ox-LDL in turn upregulates Ang II AT 1receptor gene expression. This interaction between Ang II and ox-LDL further augments cell injury in HCAECs. These findings provide basis for the use of AT1-receptor blockers and anti-oxidants in designing therapy for atherosclerosis and myocardial ischaemia.

  12. Sympathetic Innervation Promotes Arterial Fate by Enhancing Endothelial ERK Activity.

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    Pardanaud, Luc; Pibouin-Fragner, Laurence; Dubrac, Alexandre; Mathivet, Thomas; English, Isabel; Brunet, Isabelle; Simons, Michael; Eichmann, Anne

    2016-08-19

    Arterial endothelial cells are morphologically, functionally, and molecularly distinct from those found in veins and lymphatic vessels. How arterial fate is acquired during development and maintained in adult vessels is incompletely understood. We set out to identify factors that promote arterial endothelial cell fate in vivo. We developed a functional assay, allowing us to monitor and manipulate arterial fate in vivo, using arteries isolated from quails that are grafted into the coelom of chick embryos. Endothelial cells migrate out from the grafted artery, and their colonization of host arteries and veins is quantified. Here we show that sympathetic innervation promotes arterial endothelial cell fate in vivo. Removal of sympathetic nerves decreases arterial fate and leads to colonization of veins, whereas exposure to sympathetic nerves or norepinephrine imposes arterial fate. Mechanistically, sympathetic nerves increase endothelial ERK (extracellular signal-regulated kinase) activity via adrenergic α1 and α2 receptors. These findings show that sympathetic innervation promotes arterial endothelial fate and may lead to novel approaches to improve arterialization in human disease. © 2016 American Heart Association, Inc.

  13. Effects of some anti-diabetic and cardioprotective agents on proliferation and apoptosis of human coronary artery endothelial cells

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    Eriksson Linnéa

    2012-03-01

    Full Text Available Abstract Background The leading cause of death for patients suffering from diabetes is macrovascular disease. Endothelial dysfunction is often observed in type 2 diabetic patients and it is considered to be an important early event in the pathogenesis of atherogenesis and cardiovascular disease. Many drugs are clinically applied to treat diabetic patients. However, little is known whether these agents directly interfere with endothelial cell proliferation and apoptosis. This study therefore aimed to investigate how anti-diabetic and cardioprotective agents affect human coronary artery endothelial cells (HCAECs. Methods The effect of anti-diabetic and cardioprotective agents on HCAEC viability, proliferation and apoptosis was studied. Viability was assessed using Trypan blue exclusion; proliferation in 5 mM and 11 mM of glucose was analyzed using [3H]thymidine incorporation. Lipoapoptosis of the cells was investigated by determining caspase-3 activity and the subsequent DNA fragmentation after incubation with the free fatty acid palmitate, mimicking diabetic lipotoxicity. Results Our data show that insulin, metformin, BLX-1002, and rosuvastatin improved HCAEC viability and they could also significantly increase cell proliferation in low glucose. The proliferative effect of insulin and BLX-1002 was also evident at 11 mM of glucose. In addition, insulin, metformin, BLX-1002, pioglitazone, and candesartan significantly decreased the caspase-3 activity and the subsequent DNA fragmentation evoked by palmitate, suggesting a protective effect of the drugs against lipoapoptosis. Conclusion Our results suggest that the anti-diabetic and cardioprotective agents mentioned above have direct and beneficial effects on endothelial cell viability, regeneration and apoptosis. This may add yet another valuable property to their therapeutic effect, increasing their clinical utility in type 2 diabetic patients in whom endothelial dysfunction is a prominent feature

  14. Oxidation modifies the structure and function of the extracellular matrix generated by human coronary artery endothelial cells.

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    Chuang, Christine Y; Degendorfer, Georg; Hammer, Astrid; Whitelock, John M; Malle, Ernst; Davies, Michael J

    2014-04-15

    ECM (extracellular matrix) materials, such as laminin, perlecan, type IV collagen and fibronectin, play a key role in determining the structure of the arterial wall and the properties of cells that interact with the ECM. The aim of the present study was to investigate the effect of peroxynitrous acid, an oxidant generated by activated macrophages, on the structure and function of the ECM laid down by HCAECs (human coronary artery endothelial cells) in vitro and in vivo. We show that exposure of HCAEC-derived native matrix components to peroxynitrous acid (but not decomposed oxidant) at concentrations >1 μM results in a loss of antibody recognition of perlecan, collagen IV, and cell-binding sites on laminin and fibronectin. Loss of recognition was accompanied by decreased HCAEC adhesion. Real-time PCR showed up-regulation of inflammation-associated genes, including MMP7 (matrix metalloproteinase 7) and MMP13, as well as down-regulation of the laminin α2 chain, in HCAECs cultured on peroxynitrous acid-treated matrix compared with native matrix. Immunohistochemical studies provided evidence of co-localization of laminin with 3-nitrotyrosine, a biomarker of peroxynitrous acid damage, in type II-III/IV human atherosclerotic lesions, consistent with matrix damage occurring during disease development in vivo. The results of the present study suggest a mechanism through which peroxynitrous acid modifies endothelial cell-derived native ECM proteins of the arterial basement membrane in atherosclerotic lesions. These changes to ECM and particularly perlecan and laminin may be important in inducing cellular dysfunction and contribute to atherogenesis.

  15. [The stimulation of human pulmonary artery endothelial cells by cigarette smoke extract contributed to cell senescence and induced human pulmonary artery smooth cell migration].

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    Cai, L; Zhu, P C; Wang, Y E; Gao, Y T; Ao, Q L

    2017-06-12

    Objective: To observe the senescent effect of human pulmonary arterial endothelial cells (HPAEC) stimulated by cigarette smoke extract (CSE) and the effect of secretion of senescent cells on human pulmonary arterial smooth muscles cell (HPASMC) proliferation and migration. Methods: HPAEC was treated with different concentrations of CSE in vitro and cell proliferation was determined by CCK8, senescence cells analyzed by detecting the β-gal activity, and the senescent proteins of cells measured by Western blot. The concentration of senescence-associated secretory phenotype (SASP) was detected by ELISA and the expression of MCP-1 and TGF-β1 was measured by Real-time PCR. The number of the proliferated cells was measured by Transwell assay and immunoflurescence. Results: The HPAEC was aging with the stimulation concentration of CSE increasing and the stimulation time prolonging (Pcells increased as the exposure time prolonged. Compared with the control group, cell viability of 48 h group(1.8±0.1) and 72 h group (1.8±0.1) decreased significantly. The flow cytometry showed a significant difference between the CSE group(14.1±1.2) and the control group(28.5±1.8) in S phase(Pcell cycle arrest. The SASP was increasing as the CSE-exposure prolonged. Compared with the control group(177±39), the 48 h group(460±43) and the 72 h group(609±64) showed a marked increase in MCP-1(Pcells could secrete SASP which induced HPASMC proliferation. After different times of conditioned medium stimulation, HPASMC proliferated especially at 72 h(P<0.05) . The immnoflorescence and Transwell assay confirmed this finding. Conclusion: CSE could induce senescence of HPAEC and SASP production which improved HPASMC proliferation and migration.

  16. High glucose-induced apoptosis in human coronary artery endothelial cells involves up-regulation of death receptors

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    Yamamoto Seiji

    2011-08-01

    Full Text Available Abstract Background High glucose can induce apoptosis in vascular endothelial cells, which may contribute to the development of vascular complications in diabetes. We evaluated the role of the death receptor pathway of apoptotic signaling in high glucose-induced apoptosis in human coronary artery endothelial cells (HCAECs. Methods HCAECs were treated with media containing 5.6, 11.1, and 16.7 mM of glucose for 24 h in the presence or absence of tumor necrosis factor (TNF-α. For detection of apoptosis, DNA fragmentation assay was used. HCAEC expression of death receptors were analyzed by the PCR and flow cytometry methods. Also, using immunohistochemical techniques, coronary expression of death receptors was assessed in streptozotocin-nicotinamide-induced type 2 diabetic mice. Results Exposure of HCAECs to high glucose resulted in a significant increase in TNF-R1 and Fas expression, compared with normal glucose. High glucose increased TNF-α production by HCAECs and exogenous TNF-α up-regulated TNF-R1 and Fas expression in HCAECs. High glucose-induced up-regulation of TNF-R1 and Fas expression was undetectable in the presence of TNF-α. Treatment with TNF-R1 neutralizing peptides significantly inhibited high glucose-induced endothelial cell apoptosis. Type 2 diabetic mice displayed appreciable expression of TNF-R1 and Fas in coronary vessels. Conclusions In association with increased TNF-α levels, the death receptors, TNF-R1 and Fas, are up-regulated in HCAECs under high glucose conditions, which could in turn play a role in high glucose-induced endothelial cell apoptosis.

  17. Effects of incretin agonists on endothelial nitric oxide synthase expression and nitric oxide synthesis in human coronary artery endothelial cells exposed to TNFα and glycated albumin.

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    Garczorz, Wojciech; Francuz, Tomasz; Siemianowicz, Krzysztof; Kosowska, Agnieszka; Kłych, Agnieszka; Aghdam, Mohammad Reza F; Jagoda, Krystyna

    2015-02-01

    There have been a number of beneficial effects of incretin agonists on the cardiovascular system. Glycated albumin (GA) and tumor necrosis factor (TNFα) may lead to endothelial dysfunction. Due to reports of cardioprotective effects of incretin agonist, we wanted to determine if GLP-1 and exendin-4 can reverse diminished production of nitric oxide (NO) after treatment with TNFα and GA. The objective of our experiment was to study the interaction between incretin agonists and proinflammatory substances like TNFα and GA on production of NO in HCAEC. Human vascular endothelial cells from the coronary artery (HCAEC) were used. The mRNA expression and protein level of endothelial nitric oxide synthase (eNOS) and inducible (iNOS) were quantified. NO production was measured in cells using DAF-FM/DA and flow cytometry. TNFα (10 ng/mL) decreased eNOS: mRNA by 90% and protein level by 31%. TNFα also decreased NO by 33%. GA (500 μg/mL) neither affected eNOS expression nor the protein level, but inhibited nearly all formation of NO in endothelium. GLP-1 (100 nM) and exendin-4 (1 and 10nM) decreased the amount of NO compared to control. Incubation of HCAEC with TNFα and incretin agonists did not change or moderately reduce the amount of NO compared to TNFα alone. TNFα and GA decrease production of NO in HCAEC, presumably by inducing reactive oxygen species or eNOS uncoupling. Incretin agonists in tested concentrations in the presence of l-arginine were not able to reverse this effect and instead led to a further reduction in NO production. Copyright © 2014 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  18. The mechanism of TGF-β/miR-155/c-Ski regulates endothelial-mesenchymal transition in human coronary artery endothelial cells.

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    Wang, Juan; He, Wen; Xu, Xiao; Guo, Liping; Zhang, Yin; Han, Suxia; Shen, Difei

    2017-08-31

    Human coronary artery endothelial cells (HCAECs) have the potential to undergo fibrogenic endothelial-mesenchymal transition (EndMT), which results in matrix-producing fibroblasts and thereby contributes to the pathogenesis of cardiac fibrosis. Recently, the profibrotic cytokine transforming growth factor-β (TGF-β) is shown to be the crucial pathogenic driver which has been verified to induce EndMT. C-Ski is an important regulator of TGF-β signaling. However, the detailed role of c-Ski and the molecular mechanisms by which c-Ski affects TGF-β-induced EndMT in HCAECs are not largely elucidated. In the present study, we treated HCAECs with TGF-β of different concentrations to induce EndMT. We found that overexpression of c-Ski in HCAECs either blocked EndMT via hindering Vimentin, Snail, Slug, and Twist expression while enhancing CD31 expression, with or without TGF-β treatment. In contrast, suppression of c-Ski further enhanced EndMT. Currently, miRNA expression disorder has been frequently reported associating with cardiac fibrosis. By using online tools, we regarded miR-155 as a candidate miRNA that could target c-Ski, which was verified using luciferase assays. C-Ski expression was negatively regulated by miR-155. TGF-β-induced EndMT was inhibited by miR-155 silence; the effect of TGF-β on Vimentin, CD31, Snail, Slug, and Twist could be partially restored by miR-155. Altogether, these findings will shed light on the role and mechanism by which miR-155 regulates TGF-β-induced HCAECs EndMT via c-Ski to affect cardiac fibrosis, and miR-155/c-Ski may represent novel biomarkers and therapeutic targets in the treatment of cardiac fibrosis. © 2017 The Author(s).

  19. LPS, but not Angiotensin ll, lnduces Direct Pro-lnflammatory Effects in Cultured Mouse Arteries and Human Endothelial and Vascular Smooth Muscle Cells

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    Outzen, Emilie M; Zaki, Marina; Mehryar, Rahila

    2017-01-01

    resistance-sized arteries (MRA) supported by experiments in cultured human primary endothelial and vascular smooth muscle cells. Results showed that 24-hr organ culture of mouse MRA with 10 nM Ang II had, unlike 100 ng/mL LPS, no effects on IL-6 or MCP-1 secretion, VCAM1 mRNA expression or endothelial......]-Ang II had no concentration- or time-dependent effects on IL-6 and MCP-1 secretion in human umbilical vein endothelial cells (HUVEC) and human aortic smooth muscle cells (HASMC). AGTR1 or AGTR2 mRNA expression were undetectable in HUVEC, whereas HASMC expressed only AGTR1 mRNA. In summary, contrary...

  20. Cigarette smoke but not electronic cigarette aerosol activates a stress response in human coronary artery endothelial cells in culture.

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    Teasdale, Jack E; Newby, Andrew C; Timpson, Nicholas J; Munafò, Marcus R; White, Stephen J

    2016-06-01

    It is generally acknowledged that e-cigarettes are unlikely to be as harmful as conventional cigarettes, but there is little data that quantifies their relative harms. We investigated the biological response to e-cigarette aerosol exposure (versus conventional cigarette smoke exposure) at the cellular level, by exposing human coronary artery endothelial cells (HCAEC) to aqueous filtered extracts of e-cigarette aerosol or cigarette smoke and looking at gene expression changes consistent with a stress response. This included genes controlled by the oxidant-stress sensing transcription factor NFR2 (NFE2L2), and cytochrome P450 family members. Cigarette smoke extract (CSE) was created using mainstream smoke from a single cigarette drawn through 10ml of endothelial cell growth media MV2. Electronic cigarette aerosol extract (eCAE) was created using the same apparatus, using a constant power output of 10.8w (4.2V) and 18mg/ml nicotine solution. eCAE was generated using 5 cycles of 5s heat with at least 10s in between each puff to allow the coil to cool, air being drawn through the device at 70ml/minute. HCAEC responded to the noxious components in CSE, resulting in activation of NRF2 and upregulation of cytochrome p450. However, eCAE did not induce NRF2 nuclear localisation, upregulation of NRF2-activated genes, or the upregulation of cytochrome p450. The use of e-cigarettes as a substitute for conventional cigarettes is likely to reduce immediate tobacco-related harm, at least with respect to cardiovascular harms. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  1. Nuclear IL-33 regulates soluble ST2 receptor and IL-6 expression in primary human arterial endothelial cells and is decreased in idiopathic pulmonary arterial hypertension

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    Shao, Dongmin [Section of Vascular Biology, National Heart and Lung Institute, Imperial College London, London (United Kingdom); Perros, Frédéric [Faculté de Médecine, Université Paris-Sud, Paris, Clamart (France); Caramori, Gaetano [Dipartimento di Scienze Mediche, Sezione di Medicina Interna e Cardiorespiratoria, Centro Interdipartimentale per lo Studio delle Malattie Infiammatorie delle Vie Aeree e Patologie Fumo-Correlate, University of Ferrara, Ferrara (Italy); Meng, Chao [Section of Vascular Biology, National Heart and Lung Institute, Imperial College London, London (United Kingdom); Department of Geriatrics, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai (China); Dormuller, Peter [Faculté de Médecine, Université Paris-Sud, Paris, Clamart (France); Chou, Pai-Chien [Airways Disease, National Heart and Lung Institute (United Kingdom); Church, Colin [Scottish Pulmonary Vascular Unit, University of Glasgow (United Kingdom); Papi, Alberto; Casolari, Paolo [Dipartimento di Scienze Mediche, Sezione di Medicina Interna e Cardiorespiratoria, Centro Interdipartimentale per lo Studio delle Malattie Infiammatorie delle Vie Aeree e Patologie Fumo-Correlate, University of Ferrara, Ferrara (Italy); Welsh, David; Peacock, Andrew [Scottish Pulmonary Vascular Unit, University of Glasgow (United Kingdom); Humbert, Marc [Faculté de Médecine, Université Paris-Sud, Paris, Clamart (France); Adcock, Ian M. [Airways Disease, National Heart and Lung Institute (United Kingdom); Wort, Stephen J., E-mail: s.wort@imperial.ac.uk [Section of Vascular Biology, National Heart and Lung Institute, Imperial College London, London (United Kingdom)

    2014-08-15

    Highlights: • Nuclear IL-33 expression is reduced in vascular endothelial cells from PAH patients. • Knockdown of IL-33 leads to increased IL-6 and sST2 mRNA expression. • IL-33 binds homeobox motifs in target gene promoters and recruits repressor proteins. - Abstract: Idiopathic pulmonary arterial hypertension (IPAH) is an incurable condition leading to right ventricular failure and death and inflammation is postulated to be associated with vascular remodelling. Interleukin (IL)-33, a member of the “alarmin” family can either act on the membrane ST2 receptor or as a nuclear repressor, to regulate inflammation. We show, using immunohistochemistry, that IL-33 expression is nuclear in the vessels of healthy subjects whereas nuclear IL-33 is markedly diminished in the vessels of IPAH patients. This correlates with reduced IL-33 mRNA expression in their lung. In contrast, serum levels of IL-33 are unchanged in IPAH. However, the expression of the soluble form of ST2, sST2, is enhanced in the serum of IPAH patients. Knock-down of IL-33 in human endothelial cells (ECs) using siRNA is associated with selective modulation of inflammatory genes involved in vascular remodelling including IL-6. Additionally, IL-33 knock-down significantly increased sST2 release from ECs. Chromatin immunoprecipitation demonstrated that IL-33 bound multiple putative homeodomain protein binding motifs in the proximal and distal promoters of ST2 genes. IL-33 formed a complex with the histone methyltransferase SUV39H1, a transcriptional repressor. In conclusion, IL-33 regulates the expression of IL-6 and sST2, an endogenous IL-33 inhibitor, in primary human ECs and may play an important role in the pathogenesis of PAH through recruitment of transcriptional repressor proteins.

  2. Arterial endothelial function measurement method and apparatus

    Science.gov (United States)

    Maltz, Jonathan S; Budinger, Thomas F

    2014-03-04

    A "relaxoscope" (100) detects the degree of arterial endothelial function. Impairment of arterial endothelial function is an early event in atherosclerosis and correlates with the major risk factors for cardiovascular disease. An artery (115), such as the brachial artery (BA) is measured for diameter before and after several minutes of either vasoconstriction or vasorelaxation. The change in arterial diameter is a measure of flow-mediated vasomodification (FMVM). The relaxoscope induces an artificial pulse (128) at a superficial radial artery (115) via a linear actuator (120). An ultrasonic Doppler stethoscope (130) detects this pulse 10-20 cm proximal to the point of pulse induction (125). The delay between pulse application and detection provides the pulse transit time (PTT). By measuring PTT before (160) and after arterial diameter change (170), FMVM may be measured based on the changes in PTT caused by changes in vessel caliber, smooth muscle tone and wall thickness.

  3. Pregnancy Augments VEGF-Stimulated In Vitro Angiogenesis and Vasodilator (NO and H2S) Production in Human Uterine Artery Endothelial Cells.

    Science.gov (United States)

    Zhang, Hong-Hai; Chen, Jennifer C; Sheibani, Lili; Lechuga, Thomas J; Chen, Dong-Bao

    2017-07-01

    Augmented uterine artery (UA) production of vasodilators, including nitric oxide (NO) and hydrogen sulfide (H2S), has been implicated in pregnancy-associated and agonist-stimulated rise in uterine blood flow that is rate-limiting to pregnancy health. Developing a human UA endothelial cell (hUAEC) culture model from main UAs of nonpregnant (NP) and pregnant (P) women for testing a hypothesis that pregnancy augments endothelial NO and H2S production and endothelial reactivity to vascular endothelial growth factor (VEGF). Main UAs from NP and P women were used for developing hUAEC culture models. Comparisons were made between NP- and P-hUAECs in in vitro angiogenesis, activation of cell signaling, expression of endothelial NO synthase (eNOS) and H2S-producing enzymes cystathionine β-synthase (CBS) and cystathionine γ-lyase, and NO/H2S production upon VEGF stimulation. NP- and P-hUAECs displayed a typical cobblestone-like shape in culture and acetylated low-density lipoprotein uptake, stained positively for endothelial and negatively for smooth muscle markers, maintained key signaling proteins during passage, and had statistically significant greater eNOS and CBS proteins in P- vs NP-hUAECs. Treatment with VEGF stimulated in vitro angiogenesis and eNOS protein and NO production only in P-hUEACs and more robust cell signaling in P- vs NP-hUAECs. VEGF stimulated CBS protein expression, accounting for VEGF-stimulated H2S production in hUAECs. Comparisons between NP- and P-hUAECs reveal that pregnancy augments VEGF-stimulated in vitro angiogenesis and NO/H2S production in hUAECs, showing that the newly established hUAEC model provides a critical in vitro tool for understanding human uterine hemodynamics.

  4. Pollutant particles enhanced H2O2 production from NAD(P)H oxidase and mitochondria in human pulmonary artery endothelial cells.

    Science.gov (United States)

    Li, Zhuowei; Hyseni, Xhevahire; Carter, Jacqueline D; Soukup, Joleen M; Dailey, Lisa A; Huang, Yuh-Chin T

    2006-08-01

    Particulate matter (PM) induces oxidative stress and cardiovascular adverse health effects, but the mechanistic link between the two is unclear. We hypothesized that PM enhanced oxidative stress in vascular endothelial cells and investigated the enzymatic sources of reactive oxygen species and their effects on mitogen-activated protein kinase (MAPK) activation and vasoconstriction. We measured the production of extracellular H2O2, activation of extracellular signal-regulated kinases1/2 (ERK1/2) and p38 MAPKs in human pulmonary artery endothelial cells (HPAEC) treated with urban particles (UP; SRM1648), and assessed the effects of H2O2 on vasoconstriction in pulmonary artery ring and isolated perfused lung. Within minutes after UP treatment, HPAEC increased H2O2 production that could be inhibited by diphenyleneiodonium (DPI), apocynin (APO), and sodium azide (NaN3). The water-soluble fraction of UP as well as its two transition metal components, Cu and V, also stimulated H2O2 production. NaN3 inhibited H2O2 production stimulated by Cu and V, whereas DPI and APO inhibited only Cu-stimulated H2O2 production. Inhibitors of other H2O2-producing enzymes, including Nomega-methyl-L-argnine, indomethacin, allopurinol, cimetidine, rotenone, and antimycin, had no effects. DPI but not NaN3 attenuated UP-induced pulmonary vasoconstriction and phosphorylation of ERK1/2 and p38 MAPKs. Knockdown of p47phox gene expression by small interfering RNA attenuated UP-induced H2O2 production and phosphorylation of ERK1/2 and p38 MAPKs. Intravascular administration of H2O2 generated by glucose oxidase increased pulmonary artery pressure. We conclude that UP induce oxidative stress in vascular endothelial cells by activating NAD(P)H oxidase and the mitochondria. The endothelial oxidative stress may be an important mechanism for PM-induced acute cardiovascular health effects.

  5. Endothelial derived hyperpolarization in renal interlobar arteries

    DEFF Research Database (Denmark)

    Brasen, Jens Christian; Sørensen, Charlotte M.

    2015-01-01

    In small arteries, vascular smooth muscle cells (VSMCs) and endothelial cells (ECs) are connect by myoendothelial junctions (MEJ), usually extending from the EC. Ca2+ activated K+ channels (IKCa and SKCa) located in the MEJ are suggested to play a role in NO-independent endothelium derived...

  6. Targeting Pulmonary Endothelial Hemoglobin α Improves Nitric Oxide Signaling and Reverses Pulmonary Artery Endothelial Dysfunction.

    Science.gov (United States)

    Alvarez, Roger A; Miller, Megan P; Hahn, Scott A; Galley, Joseph C; Bauer, Eileen; Bachman, Timothy; Hu, Jian; Sembrat, John; Goncharov, Dmitry; Mora, Ana L; Rojas, Mauricio; Goncharova, Elena; Straub, Adam C

    2017-12-01

    Pulmonary hypertension is characterized by pulmonary endothelial dysfunction. Previous work showed that systemic artery endothelial cells (ECs) express hemoglobin (Hb) α to control nitric oxide (NO) diffusion, but the role of this system in pulmonary circulation has not been evaluated. We hypothesized that up-regulation of Hb α in pulmonary ECs contributes to NO depletion and pulmonary vascular dysfunction in pulmonary hypertension. Primary distal pulmonary arterial vascular smooth muscle cells, lung tissue sections from unused donor (control) and idiopathic pulmonary artery (PA) hypertension lungs, and rat and mouse models of SU5416/hypoxia-induced pulmonary hypertension (PH) were used. Immunohistochemical, immunocytochemical, and immunoblot analyses and transfection, infection, DNA synthesis, apoptosis, migration, cell count, and protein activity assays were performed in this study. Cocultures of human pulmonary microvascular ECs and distal pulmonary arterial vascular smooth muscle cells, lung tissue from control and pulmonary hypertensive lungs, and a mouse model of chronic hypoxia-induced PH were used. Immunohistochemical, immunoblot analyses, spectrophotometry, and blood vessel myography experiments were performed in this study. We find increased expression of Hb α in pulmonary endothelium from humans and mice with PH compared with controls. In addition, we show up-regulation of Hb α in human pulmonary ECs cocultured with PA smooth muscle cells in hypoxia. We treated pulmonary ECs with a Hb α mimetic peptide that disrupts the association of Hb α with endothelial NO synthase, and found that cells treated with the peptide exhibited increased NO signaling compared with a scrambled peptide. Myography experiments using pulmonary arteries from hypoxic mice show that the Hb α mimetic peptide enhanced vasodilation in response to acetylcholine. Our findings reveal that endothelial Hb α functions as an endogenous scavenger of NO in the pulmonary endothelium

  7. 1α,25-Dihydroxyvitamin D(3) inhibits vascular cellular adhesion molecule-1 expression and interleukin-8 production in human coronary arterial endothelial cells.

    Science.gov (United States)

    Kudo, Keiko; Hasegawa, Shunji; Suzuki, Yasuo; Hirano, Reiji; Wakiguchi, Hiroyuki; Kittaka, Setsuaki; Ichiyama, Takashi

    2012-11-01

    Kawasaki disease is an acute febrile vasculitis of childhood that is associated with elevated production of inflammatory cytokines, causing damage to the coronary arteries. The production of proinflammatory cytokines and expression of adhesion molecules in human coronary arterial endothelial cells (HCAECs) is regulated by nuclear transcription factor-κB (NF-κB) activation. We have previously reported that the active form of vitamin D, 1α,25-dihydroxyvitamin D(3) (1α,25-(OH)(2)D(3)), inhibits tumor necrosis factor-α (TNF-α)-induced NF-κB activation. In this study, we examined the anti-inflammatory effects of 1α,25-(OH)(2)D(3) on TNF-α-induced adhesion molecule expression (vascular cellular adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1)) and cytokine production (interleukin-6 (IL-6) and IL-8) in HCAECs. Pretreatment with 1α,25-(OH)(2)D(3) significantly inhibited TNF-α-induced VCAM-1 expression and IL-8 production in HCAECs. Our results suggest that adjunctive 1α,25-(OH)(2)D(3) therapy may modulate the inflammatory response during Kawasaki disease vasculitis. Copyright © 2012 Elsevier Ltd. All rights reserved.

  8. Northern contaminant mixtures induced morphological and functional changes in human coronary artery endothelial cells under culture conditions typifying high fat/sugar diet and ethanol exposure.

    Science.gov (United States)

    Florian, Maria; Yan, Jin; Ulhaq, Saad; Coughlan, Melanie; Laziyan, Mahemuti; Willmore, William; Jin, Xiaolei

    2013-11-16

    It has been reported that Northern populations are exposed to mixtures of various environmental contaminants unique to the Arctic (Northern contaminant mixtures - NCM) at a large range of concentrations, depending on their geological location, age, lifestyle and dietary habits. To determine if these contaminants may contribute to a cardiovascular health risk, especially when combined with a high fat and sugar diet and ethanol exposure, we treated human coronary artery endothelial cells (HCAEC) with two mixtures of 4 organic (NCM1) or 22 organic and inorganic (NCM2) chemicals detected in Northerners' blood during 2004-2005 in the presence or absence of low-density lipoprotein (1.5mg/ml), very-low-density lipoprotein (1.0mg/ml) and glucose (10mmol/L) (LVG), and in the absence or presence of 0.1% ethanol. After 24h of exposure, cell morphology and markers of cytotoxicity and endothelial function were examined. NCM1 treatment did not affect cell viability, but increased cell size, disrupted cell membrane integrity, and decreased cell density, uptake of small peptides, release of endothelin-1 (ET-1) and plasminogen activator inhibitor (PAI), while causing no changes in endothelial nitric oxide synthase (eNOS) protein expression and nitric oxide (NO) release. In contrast, NCM2 decreased cell viability, total protein yield, uptake of small peptides, eNOS protein expression, and NO release and caused membrane damage, but caused no changes in the secretion of ET-1, prostacyclin and PAI. The presence of LVG and/or alcohol did or did not influence the effects of NCM1 or NCM2 depending on the endpoint and the mixture examined. These results suggested that the effects of one or one group of contaminants may be altered by the presence of other contaminants, and that with or without the interaction of high fat and sugar diet and/or ethanol exposure, NCMs at the concentrations used caused endothelial dysfunction in vitro. It remains to be investigated if these effects of NCMs also

  9. Aspirin protects human coronary artery endothelial cells against atherogenic electronegative LDL via an epigenetic mechanism: a novel cytoprotective role of aspirin in acute myocardial infarction.

    Science.gov (United States)

    Chang, Po-Yuan; Chen, Yi-Jie; Chang, Fu-Hsiung; Lu, Jonathan; Huang, Wen-Huei; Yang, Tzu-Ching; Lee, Yuan-Teh; Chang, Shwu-Fen; Lu, Shao-Chun; Chen, Chu-Huang

    2013-07-01

    L5 is the most negatively charged subfraction of human low-density lipoprotein (LDL) and is the only subfraction of LDL capable of inducing apoptosis in cultured vascular endothelial cells (ECs) by inhibiting fibroblast growth factor-2 (FGF2) transcription. We examined whether plasma L5 levels are elevated in patients with ST-segment elevation myocardial infarction (STEMI) and whether aspirin provides epigenetic protection of human coronary artery ECs (HCAECs) exposed to L5. Plasma L5 levels were compared between patients with STEMI (n = 10) and control subjects with chest pain syndrome but a normal coronary arteriogram (n = 5). L5 was isolated from the plasma of STEMI patients and control subjects, and apoptosis, FGF2 expression, and FGF2 promoter methylation were examined in HCAECs treated with L5 and aspirin. Plasma L5 levels were significantly higher in STEMI patients than in control subjects (P aspirin (0.2 mM) attenuated the adverse effects of L5 on HCAEC survival, FGF2 expression, and FGF2 promoter methylation. In contrast, high concentrations of aspirin (≥1.0 mM) accentuated the effects of L5. Our results show that L5 levels are significantly increased in STEMI patients. Furthermore, L5 impairs HCAEC function through CpG methylation of the FGF2 promoter, which is suppressed in the presence of low-concentration aspirin. Our results provide evidence of a novel mechanism of aspirin in the prevention of MI.

  10. Activation of group IVC phospholipase A{sub 2} by polycyclic aromatic hydrocarbons induces apoptosis of human coronary artery endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Tithof, Patricia K. [University of Tennessee, Department of Pathobiology, College of Veterinary Medicine, Knoxville, TN (United States); University of Tennessee, Center for Environmental Biotechnology, Knoxville, TN (United States); Richards, Sean M. [University of Tennessee, Department of Biological and Environmental Sciences, Chattanooga, TN (United States); Elgayyar, Mona A. [University of Tennessee, Department of Pathobiology, College of Veterinary Medicine, Knoxville, TN (United States); Menn, Fu-Minn; Vulava, Vijay M.; McKay, Larry; Sanseverino, John; Sayler, Gary [University of Tennessee, Center for Environmental Biotechnology, Knoxville, TN (United States); Tucker, Dawn E.; Leslie, Christina C. [National Jewish Medical and Research Center, Department of Pediatrics, Denver, CO (United States); Lu, Kim P. [Texas A and M University, Department of Biology, College Station, TX (United States); Ramos, Kenneth S. [University of Louisville, Department of Biochemistry and Molecular Biology, Louisville, KY (United States)

    2011-06-15

    Exposure to environmental pollutants, such as polycyclic aromatic hydrocarbons (PAHs) found in coal tar mixtures and tobacco sources, is considered a significant risk factor for the development of heart disease in humans. The goal of this study was to determine the influence of PAHs present at a Superfund site on human coronary artery endothelial cell (HCAEC) phospholipase A{sub 2} (PLA{sub 2}) activity and apoptosis. Extremely high levels of 12 out of 15 EPA high-priority PAHs were present in both the streambed and floodplain sediments at a site where an urban creek and its adjacent floodplain were extensively contaminated by PAHs and other coal tar compounds. Nine of the 12 compounds and a coal tar mixture (SRM 1597A) activated group IVC PLA{sub 2} in HCAECs, and activation of this enzyme was associated with histone fragmentation and poly (ADP) ribose polymerase (PARP) cleavage. Genetic silencing of group IVC PLA{sub 2} inhibited both {sup 3}H-fatty acid release and histone fragmentation by PAHs and SRM 1597A, indicating that individual PAHs and a coal tar mixture induce apoptosis of HCAECs via a mechanism that involves group IVC PLA{sub 2}. Western blot analysis of aortas isolated from feral mice (Peromyscus leucopus) inhabiting the Superfund site showed increased PARP and caspase-3 cleavage when compared to reference mice. These data suggest that PAHs induce apoptosis of HCAECs via activation of group IVC PLA{sub 2}. (orig.)

  11. Bromodomain and extra-terminal protein mimic JQ1 decreases inflammation in human vascular endothelial cells: Implications for pulmonary arterial hypertension.

    Science.gov (United States)

    Mumby, Sharon; Gambaryan, Natalia; Meng, Chao; Perros, Frederic; Humbert, Marc; Wort, S John; Adcock, Ian M

    2017-01-01

    Nuclear factor kappa B (NF-kB)-mediated inflammatory gene expression and vascular endothelial cell proliferation/remodelling are implicated in the pathophysiology of the fatal disease, pulmonary arterial hypertension (PAH). Bromodomain and extra-terminal (BET) proteins are essential for the expression of a subset of NF-kB-induced inflammatory genes. BET mimics including JQ1+ prevent binding of BETs to acetylated histones and down-regulate the expression of selected genes. The effects of JQ1+ on the proliferation of primary human pulmonary microvascular endothelial cells (HPMECs) from healthy subjects were measured by bromodeoxyuridine (BrdU) incorporation. Cell cycle progression was assessed by flow cytometry; mRNA and protein levels of cyclin-dependent kinases (CDKs), inhibitors and cytokines were determined by reverse transcription-quantitative PCR (RT-qPCR), Western blotting or ELISA. Histone acetyltransferase (HAT) and deacetylase (HDAC) activities were determined in nuclear extracts from whole lung of PAH and control patients. JQ1+ significantly inhibited IL6 and IL8 (IL6 and CXCL8) mRNA and protein in HPMECs compared with its inactive enantiomer JQ1-. JQ1+ decreased NF-kB p65 recruitment to native IL6 and IL8 promoters. JQ1+ showed a concentration-dependent decrease in HPMEC proliferation compared with JQ1--treated cells. JQ1+ induced G1 cell cycle arrest by increasing the expression of the CDK inhibitors (CDKN) 1A (p21 cip ) and CDKN2D (p19 INK4D ) and decreasing that of CDK2, CDK4 and CDK6. JQ1+ also inhibited serum-stimulated migration of HPMECs. Finally, HAT activity was significantly increased in the lung of PAH patients. Inhibition of BETs in primary HPMECs decreases inflammation and remodelling. BET proteins could be a target for future therapies for PAH. © 2016 The Authors. Respirology published by John Wiley & Sons Australia, Ltd on behalf of Asian Pacific Society of Respirology.

  12. Heart-type fatty-acid-binding protein (FABP3 is a lysophosphatidic acid-binding protein in human coronary artery endothelial cells

    Directory of Open Access Journals (Sweden)

    Ryoko Tsukahara

    2014-01-01

    Full Text Available Fatty-acid-binding protein 3, muscle and heart (FABP3, also known as heart-type FABP, is a member of the family of intracellular lipid-binding proteins. It is a small cytoplasmic protein with a molecular mass of about 15 kDa. FABPs are known to be carrier proteins for transporting fatty acids and other lipophilic substances from the cytoplasm to the nucleus, where these lipids are released to a group of nuclear receptors such as peroxisome proliferator-activated receptors (PPARs. In this study, using lysophosphatidic acid (LPA-coated agarose beads, we have identified FABP3 as an LPA carrier protein in human coronary artery endothelial cells (HCAECs. Administration of LPA to HCAECs resulted in a dose-dependent increase in PPARγ activation. Furthermore, the LPA-induced PPARγ activation was abolished when the FABP3 expression was reduced using small interfering RNA (siRNA. We further show that the nuclear fraction of control HCAECs contained a significant amount of exogenously added LPA, whereas FABP3 siRNA-transfected HCAECs had a decreased level of LPA in the nucleus. Taken together, these results suggest that FABP3 governs the transcriptional activities of LPA by targeting them to cognate PPARγ in the nucleus.

  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 modulation of coronary vasomotor tone in humans. Effects of atherosclerosis and risk factors for coronary artery disease.

    Science.gov (United States)

    Zeiher, A M

    1994-03-01

    Tests in patients with atherosclerosis or running the risk to get coronary heart diseases have been reported, the objective of which was to detect the importance the physiologic and pathophysiologic regulatory mechanisms of coronary blood flow have in early diagnosis. The reaction of coronary vessels after the infusion of acetylcholine compared to this they showed after the infusion of papaverine was tested, this is to say the responsiveness of coronary vessels to pharmacologic influences caused by the endothelium and aiming directly at the striatal muscles. Both, quantitative coronary angiography and intracoronary sonography served as measuring instruments. The results revealed that progressive endothelial dysfunction in the coronary vessels of risk patients can be detected by the help of pharmacologically differentiated active substances. In patients with increased cholesterin levels, the function of the endothelium is impaired, although a constriction of the lumen has not been observed. Some gradually more distinct dysfunctions have been found in patients with structural symptoms of coronary atherosclerosis and above all in patients with angiographically provable lumen alterations.

  15. High trans but not saturated fat beverage causes an acute reduction in postprandial vascular endothelial function but not arterial stiffness in humans.

    Science.gov (United States)

    Lane-Cordova, Abbi D; Witmer, Jordan R; Dubishar, Kaitlyn; DuBose, Lyndsey E; Chenard, Catherine A; Siefers, Kyle J; Myers, Janie E; Points, Lauren J; Pierce, Gary L

    2016-10-01

    A diet high in trans-fatty acids (TFAs) is associated with a higher risk of cardiovascular disease (CVD) than a diet high in saturated fatty acids (SFAs), but the mechanisms remain unclear. We hypothesized that a beverage high in TFAs would cause a larger reduction in postprandial endothelial function and an increase in arterial stiffness, in part from greater reductions in insulin sensitivity, compared with a beverage high in SFAs. Eleven healthy adults (aged 47±5 years) ingested a warm test beverage (520 kcal, 56 g total fat, 5 g carbohydrate, 1 g protein) high in either TFAs or SFAs in a randomized cross-over study. Ingestion of the beverage high in TFAs (p<0.01) but not high in SFAs (p=0.49) decreased endothelial function (brachial artery flow-mediated dilation, mmΔ) at 3-4 hours (p<0.01 for time; p=0.034 for interaction), but did not alter aortic stiffness or carotid β-stiffness. The homeostasis model of insulin resistance (interaction p=0.062) tended to decrease after SFAs but not TFAs. A beverage high in TFAs but not SFAs results in a postprandial reduction in endothelial function and a trend for decreased insulin sensitivity, potentially explaining the higher risk of CVD with a diet high in TFAs. © The Author(s) 2016.

  16. The mechanism of TGF-β/miR-155/c-Ski regulates endothelial–mesenchymal transition in human coronary artery endothelial cells

    Science.gov (United States)

    Wang, Juan; He, Wen; Xu, Xiao; Guo, Liping; Zhang, Yin

    2017-01-01

    Human coronary artery endothelial cells (HCAECs) have the potential to undergo fibrogenic endothelial–mesenchymal transition (EndMT), which results in matrix-producing fibroblasts and thereby contributes to the pathogenesis of cardiac fibrosis. Recently, the profibrotic cytokine transforming growth factor-β (TGF-β) is shown to be the crucial pathogenic driver which has been verified to induce EndMT. C-Ski is an important regulator of TGF-β signaling. However, the detailed role of c-Ski and the molecular mechanisms by which c-Ski affects TGF-β-induced EndMT in HCAECs are not largely elucidated. In the present study, we treated HCAECs with TGF-β of different concentrations to induce EndMT. We found that overexpression of c-Ski in HCAECs either blocked EndMT via hindering Vimentin, Snail, Slug, and Twist expression while enhancing CD31 expression, with or without TGF-β treatment. In contrast, suppression of c-Ski further enhanced EndMT. Currently, miRNA expression disorder has been frequently reported associating with cardiac fibrosis. By using online tools, we regarded miR-155 as a candidate miRNA that could target c-Ski, which was verified using luciferase assays. C-Ski expression was negatively regulated by miR-155. TGF-β-induced EndMT was inhibited by miR-155 silence; the effect of TGF-β on Vimentin, CD31, Snail, Slug, and Twist could be partially restored by miR-155. Altogether, these findings will shed light on the role and mechanism by which miR-155 regulates TGF-β-induced HCAECs EndMT via c-Ski to affect cardiac fibrosis, and miR-155/c-Ski may represent novel biomarkers and therapeutic targets in the treatment of cardiac fibrosis. PMID:28607031

  17. Peripheral Endothelial Function After Arterial Switch Operation for D-looped Transposition of the Great Arteries.

    Science.gov (United States)

    Sun, Heather Y; Stauffer, Katie Jo; Nourse, Susan E; Vu, Chau; Selamet Tierney, Elif Seda

    2017-06-01

    Coronary artery re-implantation during arterial switch operation in patients with D-looped transposition of the great arteries (D-TGA) can alter coronary arterial flow and increase shear stress, leading to local endothelial dysfunction, although prior studies have conflicting results. Endothelial pulse amplitude testing can predict coronary endothelial dysfunction by peripheral arterial testing. This study tested if, compared to healthy controls, patients with D-TGA after arterial switch operation had peripheral endothelial dysfunction. Patient inclusion criteria were (1) D-TGA after neonatal arterial switch operation; (2) age 9-29 years; (3) absence of known cardiovascular risk factors such as hypertension, diabetes, hypercholesterolemia, vascular disease, recurrent vasovagal syncope, and coronary artery disease; and (4) ability to comply with overnight fasting. Exclusion criteria included (1) body mass index ≥85th percentile, (2) use of medications affecting vascular tone, or (3) acute illness. We assessed endothelial function by endothelial pulse amplitude testing and compared the results to our previously published data in healthy controls (n = 57). We tested 20 D-TGA patients (16.4 ± 4.8 years old) who have undergone arterial switch operation at a median age of 5 days (0-61 days). Endothelial pulse amplitude testing indices were similar between patients with D-TGA and controls (1.78 ± 0.61 vs. 1.73 ± 0.54, p = 0.73).In our study population of children and young adults, there was no evidence of peripheral endothelial dysfunction in patients with D-TGA who have undergone arterial switch operation. Our results support the theory that coronary arterial wall thickening and abnormal vasodilation reported in these patients is a localized phenomenon and not reflective of overall atherosclerotic burden.

  18. Vascular endothelial growth factor-induced nitric oxide- and PGI2-dependent relaxation in human internal mammary arteries: a comparative study with KDR and Flt-1 selective mutants.

    Science.gov (United States)

    Wei, Wei; Jin, Hongkui; Chen, Zhi-Wu; Zioncheck, Thomas F; Yim, Anthony P C; He, Guo-Wei

    2004-11-01

    The role of the vascular endothelial growth factors (VEGF) receptors (KDR and Flt-1) and their characteristics in VEGF-induced vasodilation in human vessels is unclear. This study investigated the in vitro vasorelaxant effects of KDR-selective (KDR-SM) and Flt-1-selective mutants (Flt-1-SM) in the human internal mammary artery (IMA). IMA segments (n = 183) taken from 48 patients were studied in organ baths. The cumulative concentration (-12 to -8 log10M)-relaxation curves were established for VEGF, KDR-SM, Flt-1-SM, and placenta growth factor (PlGF) in the absence or presence of indomethacin (INDO, 7 microM), N-nitro-L-arginine (L-NNA, 300 microM), L-NNA + oxyhemoglobin (HbO, 20 microM), or INDO + L-NNA + HbO. The VEGF-induced relaxation was abolished in endothelium-denuded IMA. In the endothelium-intact vessel rings, VEGF (63.2 +/- 3.9%) induced significantly more (P < 0.001) relaxation than Flt-1-SM (28.5 +/- 4.3%, 95% CI 18.1-51.3%), and PlGF (26.0 +/- 4.7%, 95% CI 17.6-56.8%). The maximal relaxation induced by KDR-SM (53.0 +/- 4.0%) was only slightly less than that by VEGF (P = 0.075) but significantly more than that by Flt-1-SM (P = 0.001, 95% CI 7.8-41.1%). Pretreatment of INDO or L-NNA + HbO significantly (P < 0.001) inhibited the relaxation by VEGF (21.2 +/- 3.9% or 23.3 +/- 4.3%) and KDR-SM (9.8 +/- 8.2% or 10.1 +/- 17.8%). INDO + L-NNA + HbO completely inhibited the relaxation by VEGF, KDR-SM, or Flt-1-SM. KDR may be the dominant receptor in mediating the VEGF-mediated relaxation, which is regulated by both PGI2 and nitric oxide but probably not by endothelium-derived hyperpolarizing factor, in the human IMA. This study gives insight into the characteristics of the VEGF-mediated vasodilation and provides a scientific basis for potential clinical application of VEGF/KDR-SM in ischemic heart disease.

  19. Ticagrelor, but not clopidogrel, reduces arterial thrombosis via endothelial tissue factor suppression

    OpenAIRE

    Reiner, Martin F.; Akhmedov, Alexander; Stivala, Simona; Keller, Stephan; Gaul, Daniel S.; Bonetti, Nicole R.; Savarese, Gianluigi; Glanzmann, Martina; Zhu, Cuicui; Ruf, Wolfram; Yang, Zhihong; Christian M Matter; Lüscher, Thomas F; Camici, Giovanni G.; Beer, Juerg H.

    2017-01-01

    The P2Y12 antagonist ticagrelor reduces mortality in patients with acute coronary syndrome (ACS), compared with clopidogrel, and the mechanisms underlying this effect are not clearly understood. Arterial thrombosis is the key event in ACS; however, direct vascular effects of either ticagrelor or clopidogrel with focus on arterial thrombosis and its key trigger tissue factor have not been previously investigated.Methods and results: Human aortic endothelial cells were treated with ticagr...

  20. Carbon black nanoparticles and vascular dysfunction in cultured endothelial cells and artery segments

    DEFF Research Database (Denmark)

    Vesterdal, Lise K; Mikkelsen, Lone; Folkmann, Janne K

    2012-01-01

    Exposure to small size particulates is regarded as a risk factor for cardiovascular disease. We investigated effects of exposure to nanosized carbon black (CB) in human umbilical vein endothelial cells (HUVECs) and segments of arteries from rodents. The CB exposure was associated with increased...

  1. Aberrant Chloride Intracellular Channel 4 Expression Contributes to Endothelial Dysfunction in Pulmonary Arterial Hypertension

    Science.gov (United States)

    Wojciak-Stothard, Beata; Abdul-Salam, Vahitha B.; Lao, Ka Hou; Tsang, Hilda; Irwin, David C.; Lisk, Christina; Loomis, Zoe; Stenmark, Kurt R.; Edwards, John C; Yuspa, Stuart H.; Howard, Luke S.; Edwards, Robert J.; Rhodes, Christopher J.; Gibbs, J Simon R.; Wharton, John; Zhao, Lan; Wilkins, Martin R.

    2014-01-01

    Background Chloride intracellular channel 4 (CLIC4) is highly expressed in the endothelium of remodelled pulmonary vessels and plexiform lesions of patients with pulmonary arterial hypertension (PAH). CLIC4 regulates vasculogenesis through endothelial tube formation. Aberrant CLIC4 expression may contribute to the vascular pathology of PAH. Methods and Results CLIC4 protein expression was increased in plasma and blood-derived endothelial cells from patients with idiopathic PAH (IPAH) and in the pulmonary vascular endothelium of 3 rat models of pulmonary hypertension. CLIC4 gene deletion markedly attenuated the development of chronic hypoxia-induced pulmonary hypertension in mice. Adenoviral overexpression of CLIC4 in cultured human pulmonary artery endothelial cells compromised pulmonary endothelial barrier function and enhanced their survival and angiogenic capacity, while CLIC4 shRNA had an inhibitory effect. Similarly, inhibition of CLIC4 expression in blood-derived endothelial cells from patients with IPAH attenuated the abnormal angiogenic behaviour that characterises these cells. The mechanism of CLIC4 effects involves p65-mediated activation of nuclear factor-κB, followed by stabilisation of hypoxia-inducible factor-1α and increased downstream production of vascular endothelial growth factor and endothelin-1. Conclusions Increased CLIC4 expression is an early manifestation and mediator of endothelial dysfunction in pulmonary hypertension. PMID:24503951

  2. Activation of endothelial and epithelial K(Ca) 2.3 calcium-activated potassium channels by NS309 relaxes human small pulmonary arteries and bronchioles

    DEFF Research Database (Denmark)

    Kroigaard, Christel; Dalsgaard, Thomas; Nielsen, Gorm

    2012-01-01

    BACKGROUND AND PURPOSE: Small (K(Ca) 2) and intermediate (K(Ca) 3.1) conductance calcium-activated potassium channels (K(Ca) ) may contribute to both epithelium- and endothelium-dependent relaxations, but this has not been established in human pulmonary arteries and bronchioles. Therefore, we inv...

  3. Blood flow and arterial endothelial dysfunction: Mechanisms and implications

    Science.gov (United States)

    Barakat, Abdul I.

    2013-06-01

    The arterial endothelium exquisitely regulates vascular function, and endothelial dysfunction plays a critical role in the development of atherosclerosis. Atherosclerotic lesions develop preferentially at arterial branches and bifurcations where the blood flow is disturbed. Understanding the basis for this observation requires elucidating the effects of blood flow on the endothelial cell (EC) function. The goal of this review is: (1) to describe our current understanding of the relationships between arterial blood flow and atherosclerosis, (2) to present the wide array of flow-induced biological responses in ECs, and (3) to discuss the mechanisms by which ECs sense, transmit, and transduce flow-derived mechanical forces. We conclude by presenting some future perspectives in the highly interdisciplinary field of EC mechanotransduction.

  4. Endothelial Nitric Oxide Mediates Caffeine Antagonism of Alcohol-Induced Cerebral Artery Constriction

    Science.gov (United States)

    Chang, Jennifer; Fedinec, Alexander L.; Kuntamallappanavar, Guruprasad; Leffler, Charles W.; Bukiya, Anna N.

    2016-01-01

    Despite preventive education, the combined consumption of alcohol and caffeine (particularly from “energy drinks”) continues to rise. Physiologic perturbations by separate intake of ethanol and caffeine have been widely documented. However, the biologic actions of the alcohol-caffeine combination and their underlying subcellular mechanisms have been scarcely studied. Using intravital microscopy on a closed-cranial window and isolated, pressurized vessels, we investigated the in vivo and in vitro action of ethanol-caffeine mixtures on cerebral arteries from rats and mice, widely recognized models to address cerebrovascular pathophysiology and pharmacology. Caffeine at concentrations found in human circulation after ingestion of one to two cups of coffee (10 µM) antagonized the endothelium-independent constriction of cerebral arteries evoked by ethanol concentrations found in blood during moderate-heavy alcohol intoxication (40–70 mM). Caffeine antagonism against alcohol was similar whether evaluated in vivo or in vitro, suggesting independence of systemic factors and drug metabolism, but required a functional endothelium. Moreover, caffeine protection against alcohol increased nitric oxide (NO•) levels over those found in the presence of ethanol alone, disappeared upon blocking NO• synthase, and could not be detected in pressurized cerebral arteries from endothelial nitric-oxide synthase knockout (eNOS−/−) mice. Finally, incubation of de-endothelialized cerebral arteries with the NO• donor sodium nitroprusside (10 µM) fully restored the protective effect of caffeine. This study demonstrates for the first time that caffeine antagonizes ethanol-induced cerebral artery constriction and identifies endothelial NO• as the critical caffeine effector on smooth muscle targets. Conceivably, situations that perturb endothelial function and/or NO• availability will critically alter caffeine antagonism of alcohol-induced cerebrovascular constriction without

  5. Endothelial Nitric Oxide Mediates Caffeine Antagonism of Alcohol-Induced Cerebral Artery Constriction.

    Science.gov (United States)

    Chang, Jennifer; Fedinec, Alexander L; Kuntamallappanavar, Guruprasad; Leffler, Charles W; Bukiya, Anna N; Dopico, Alex M

    2016-01-01

    Despite preventive education, the combined consumption of alcohol and caffeine (particularly from "energy drinks") continues to rise. Physiologic perturbations by separate intake of ethanol and caffeine have been widely documented. However, the biologic actions of the alcohol-caffeine combination and their underlying subcellular mechanisms have been scarcely studied. Using intravital microscopy on a closed-cranial window and isolated, pressurized vessels, we investigated the in vivo and in vitro action of ethanol-caffeine mixtures on cerebral arteries from rats and mice, widely recognized models to address cerebrovascular pathophysiology and pharmacology. Caffeine at concentrations found in human circulation after ingestion of one to two cups of coffee (10 µM) antagonized the endothelium-independent constriction of cerebral arteries evoked by ethanol concentrations found in blood during moderate-heavy alcohol intoxication (40-70 mM). Caffeine antagonism against alcohol was similar whether evaluated in vivo or in vitro, suggesting independence of systemic factors and drug metabolism, but required a functional endothelium. Moreover, caffeine protection against alcohol increased nitric oxide (NO•) levels over those found in the presence of ethanol alone, disappeared upon blocking NO• synthase, and could not be detected in pressurized cerebral arteries from endothelial nitric-oxide synthase knockout (eNOS(-/-)) mice. Finally, incubation of de-endothelialized cerebral arteries with the NO• donor sodium nitroprusside (10 µM) fully restored the protective effect of caffeine. This study demonstrates for the first time that caffeine antagonizes ethanol-induced cerebral artery constriction and identifies endothelial NO• as the critical caffeine effector on smooth muscle targets. Conceivably, situations that perturb endothelial function and/or NO• availability will critically alter caffeine antagonism of alcohol-induced cerebrovascular constriction without

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

    Directory of Open Access Journals (Sweden)

    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.

  7. Biomarkers of coagulation, fibrinolysis, endothelial function, and inflammation in arterialized venous blood

    DEFF Research Database (Denmark)

    Gram, Anne Sofie; Skov, Jane; Ploug, Thorkil

    2014-01-01

    Effects of venous blood arterialization on cardiovascular risk markers are still unknown. We evaluated biomarkers of inflammation, coagulation, fibrinolysis, and endothelial function in arterialized compared with regular venous blood. Cubital venipunctures were obtained from 10 healthy volunteers...

  8. Ticagrelor, but not clopidogrel, reduces arterial thrombosis via endothelial tissue factor suppression.

    Science.gov (United States)

    Reiner, Martin F; Akhmedov, Alexander; Stivala, Simona; Keller, Stephan; Gaul, Daniel S; Bonetti, Nicole R; Savarese, Gianluigi; Glanzmann, Martina; Zhu, Cuicui; Ruf, Wolfram; Yang, Zhihong; Matter, Christian M; Lüscher, Thomas F; Camici, Giovanni G; Beer, Juerg H

    2017-01-01

    The P2Y12 antagonist ticagrelor reduces mortality in patients with acute coronary syndrome (ACS), compared with clopidogrel, and the mechanisms underlying this effect are not clearly understood. Arterial thrombosis is the key event in ACS; however, direct vascular effects of either ticagrelor or clopidogrel with focus on arterial thrombosis and its key trigger tissue factor have not been previously investigated. Human aortic endothelial cells were treated with ticagrelor or clopidogrel active metabolite (CAM) and stimulated with tumour necrosis factor-alpha (TNF-α); effects on procoagulant tissue factor (TF) expression and activity, its counter-player TF pathway inhibitor (TFPI) and the underlying mechanisms were determined. Further, arterial thrombosis by photochemical injury of the common carotid artery, and TF expression in the murine endothelium were examined in C57BL/6 mice treated with ticagrelor or clopidogrel. Ticagrelor, but not CAM, reduced TNF-α-induced TF expression via proteasomal degradation and TF activity, independently of the P2Y12 receptor and the equilibrative nucleoside transporter 1 (ENT1), an additional target of ticagrelor. In C57BL/6 mice, ticagrelor prolonged time to arterial occlusion, compared with clopidogrel, despite comparable antiplatelet effects. In line with our in vitro results, ticagrelor, but not clopidogrel, reduced TF expression in the endothelium of murine arteries. Ticagrelor, unlike clopidogrel, exhibits endothelial-specific antithrombotic properties and blunts arterial thrombus formation. The additional antithrombotic properties displayed by ticagrelor may explain its greater efficacy in reducing thrombotic events in clinical trials. These findings may provide the basis for new indications for ticagrelor. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2016. For permissions, please email: journals.permissions@oup.com.

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

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  12. 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...

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

    Science.gov (United States)

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

    2001-09-01

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

  14. The role of tissue renin angiotensin aldosterone system in the development of endothelial dysfunction and arterial stiffness

    Directory of Open Access Journals (Sweden)

    Annayya R Aroor

    2013-10-01

    Full Text Available Epidemiological studies support the notion that arterial stiffness is an independent predictor of adverse cardiovascular events contributing significantly to systolic hypertension, impaired ventricular-arterial coupling and diastolic dysfunction, impairment in myocardial oxygen supply and demand, and progression of kidney disease. Although arterial stiffness is associated with aging, it is accelerated in the presence of obesity and diabetes. The prevalence of arterial stiffness parallels the increase of obesity that is occurring in epidemic proportions and is partly driven by a sedentary life style and consumption of a high fructose, high salt and high fat western diet. Although the underlying mechanisms and mediators of arterial stiffness are not well understood, accumulating evidence supports the role of insulin resistance and endothelial dysfunction. The local tissue renin angiotensin aldosterone system (RAAS in the vascular tissue and immune cells and perivascular adipose tissue is recognized as an important element involved in endothelial dysfunction which contributes significantly to arterial stiffness. Activation of vascular RAAS is seen in humans and animal models of obesity and diabetes, and associated with enhanced oxidative stress and inflammation in the vascular tissue. The cross talk between angiotensin and aldosterone underscores the importance of mineralocorticoid receptors in modulation of insulin resistance, decreased bioavailability of nitric oxide, endothelial dysfunction and arterial stiffness. In addition, both innate and adaptive immunity are involved in this local tissue activation of RAAS. In this review we will attempt to present a unifying mechanism of how environmental and immunological factors are involved in this local tissue RAAS activation, and the role of this process in the development of endothelial dysfunction and arterial stiffness and targeting tissue RAAS activation.

  15. 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.

  16. Shear stress mediates endothelial adaptations to exercise training in humans.

    Science.gov (United States)

    Tinken, Toni M; Thijssen, Dick H J; Hopkins, Nicola; Dawson, Ellen A; Cable, N Timothy; Green, Daniel J

    2010-02-01

    Although episodic changes in shear stress have been proposed as the mechanism responsible for the effects of exercise training on the vasculature, this hypothesis has not been directly addressed in humans. We examined brachial artery flow-mediated dilation, an index of NO-mediated endothelial function, in healthy men in response to an acute bout of handgrip exercise and across an 8-week period of bilateral handgrip training. Shear stress responses were attenuated in one arm by cuff inflation to 60 mm Hg. Similar increases were observed in grip strength and forearm volume and girth in both limbs. Acute bouts of handgrip exercise increased shear rate (P<0.005) and flow-mediated dilation percentage (P<0.05) in the uncuffed limb, whereas no changes were evident in the cuffed arm. Handgrip training increased flow-mediated dilation percentage in the noncuffed limb at weeks 2, 4, and 6 (P<0.001), whereas no changes were observed in the cuffed arm. Brachial artery peak reactive hyperemia, an index of resistance artery remodeling, progressively increased with training in the noncuffed limb (P<0.001 and 0.004); no changes were evident in the cuffed arm. Neither acute nor chronic shear manipulation during exercise influenced endothelium-independent glyceryl trinitrate responses. These results demonstrate that exercise-induced changes in shear provide the principal physiological stimulus to adaptation in flow-mediated endothelial function and vascular remodeling in response to exercise training in healthy humans.

  17. INSTRUMENTAL AND DIAGNOSTIC CRITERIA OF HEMODYNAMIC DISORDERS AND ENDOTHELIAL DYSFUNCTION CORRECTION IN PREGNANTS WITH ARTERIAL HYPERTENSION

    Directory of Open Access Journals (Sweden)

    S. M. Heryak

    2014-12-01

    Conclusions. It was found that the brachial artery ultrasound measuring and occlusive plethysmography procedure by Dietz is an early and safe method of endothelial dysfunction diagnostic in pregnants with hypertension. Doppler ultrasound of blood flow in uterine, umbilical arteries, and middle cerebral arteries of the fetus allows timely diagnosis of the side effect of antihypertensive drugs on the fetus. The therapy of choice for pregnants with Stage II Arterial Hypertension should be based on methyldopa and calcium channel antagonists or selective beta-blockers combination. Highly selective beta-blockers with vasodilative effect (nebivolol hydrochloride and L-arginine (Tivortin allow to prevent perinatal adverse effects of antihypertensive therapy, to correct hemodynamic disorders and endothelial dysfunction in pregnants with arterial hypertension. KEY WORDS: arterial hypertension, uterine-placental hemodynamics, endothelial dysfunction

  18. Endothelial function in myometrial resistance arteries of normal pregnant women perfused with syncytiotrophoblast microvillous membranes

    NARCIS (Netherlands)

    van Wijk, M. J.; Boer, K.; Nisell, H.; Smarason, A. K.; van Bavel, E.; Kublickiene, K. R.

    2001-01-01

    OBJECTIVE: To investigate the effects of syncytiotrophoblast microvillous membranes (STBM) in concentrations, found in vivo in women with pre-eclampsia, on endothelial function in isolated resistance arteries. SETTING: Department of Obstetrics and Gynaecology, Huddinge University Hospital,

  19. Microvascular coronary artery spasm presents distinctive clinical features with endothelial dysfunction as nonobstructive coronary artery disease.

    Science.gov (United States)

    Ohba, Keisuke; Sugiyama, Seigo; Sumida, Hitoshi; Nozaki, Toshimitsu; Matsubara, Junichi; Matsuzawa, Yasushi; Konishi, Masaaki; Akiyama, Eiichi; Kurokawa, Hirofumi; Maeda, Hirofumi; Sugamura, Koichi; Nagayoshi, Yasuhiro; Morihisa, Kenji; Sakamoto, Kenji; Tsujita, Kenichi; Yamamoto, Eiichiro; Yamamuro, Megumi; Kojima, Sunao; Kaikita, Koichi; Tayama, Shinji; Hokimoto, Seiji; Matsui, Kunihiko; Sakamoto, Tomohiro; Ogawa, Hisao

    2012-10-01

    distinctive clinical features and endothelial dysfunction that are important to recognize as nonobstructive coronary artery disease so that optimal care with calcium channel blockers can be provided. URL: www.umin.ac.jp/ctr. Unique identifier: UMIN000003839.

  20. Study of lipoproteins and arterial intima interaction based on arterial endothelial cells real geometrical structure

    Science.gov (United States)

    Glukhova, O. E.; Kirillova, I. V.; Maslyakova, G. N.; Kossovich, E. L.; Zayarsky, D. A.; Fadeev, A. A.

    2013-02-01

    An original methodology is developed for scanning of the arterial intima morphology using the atomic force microscopy. The probing nanolaboratory NTEGRASpectra (NT-MDT, Russia) was itilized. The pictures of the coronary artery intima topology were obtained with the resolution of 1 nm. The 3D model of the `endothelial cell surface - low density lipoprotein (LDL)' complex was constructed. Using the ANSYS software, the deformation of LDL particle was found as well as the stress distribution at the moment of the macromolecule and endothelial surface collision. The largest normal and tangential stresses are found in the area of LDL interaction with the surface. These stresses are 2.173 and 0.053 kPa, respectively. It was shown that the LDL structure is being highly strained, which leads to the molecule compression and crease. Therefore, one can conclude that at the moment of LDL entering the intercellular hiatus the macromolecule will be suffering the overall deformations and large modification of its structure.

  1. 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

  2. Evaluation of a static stretching intervention on vascular endothelial function and arterial stiffness.

    Science.gov (United States)

    Shinno, Hiromi; Kurose, Satoshi; Yamanaka, Yutaka; Higurashi, Kyoko; Fukushima, Yaeko; Tsutsumi, Hiromi; Kimura, Yutaka

    2017-06-01

    Maintenance and enhancement of vascular endothelial function contribute to the prevention of cardiovascular disease and prolong a healthy life expectancy. Given the reversible nature of vascular endothelial function, interventions to improve this function might prevent arteriosclerosis. Accordingly, we studied the effects of a 6-month static stretching intervention on vascular endothelial function (reactive hyperaemia peripheral arterial tonometry index: RH-PAT index) and arterial stiffness (brachial-ankle pulse wave velocity: baPWV) and investigated the reversibility of these effects after a 6-month detraining period following intervention completion. The study evaluated 22 healthy, non-smoking, premenopausal women aged ≥40 years. Subjects were randomly assigned to the full-intervention (n = 11; mean age: 48.6 ± 2.8 years) or a half-intervention that included a control period (n = 11; mean age: 46.9 ± 3.6 years). Body flexibility and vascular endothelial function improved significantly after 3 months of static stretching. In addition to these improvements, arterial stiffness improved significantly after a 6-month intervention. However, after a 6-month detraining period, vascular endothelial function, flexibility, and arterial stiffness all returned to preintervention conditions, demonstrating the reversibility of the obtained effects. A 3-month static stretching intervention was found to improve vascular endothelial function, and an additional 3-month intervention also improved arterial stiffness. However, these effects were reversed by detraining.

  3. Evaluation of endothelial function by peripheral arterial tonometry and relation with the nitric oxide pathway

    DEFF Research Database (Denmark)

    Hedetoft, Morten; Olsen, Niels Vidiendal

    2014-01-01

    by flow-mediated dilation in the brachial artery, but the two methods are not interchangeable. We have reviewed the recent literature in an effort to evaluate peripheral arterial tonometry as a method to assess the function of the endothelium and additionally suggest directions for future research......Endothelial dysfunction is an important component in the development of cardiovascular diseases. Endothelial function may be evaluated by peripheral arterial tonometry (PAT) which measures the vasodilator function in the microvasculature of the fingertip during reactive hyperaemia. The reactive...

  4. The development of 3-D, in vitro, endothelial culture models for the study of coronary artery disease

    Directory of Open Access Journals (Sweden)

    Fraser Richard

    2009-10-01

    Full Text Available Abstract The response of the vascular endothelium to wall shear stress plays a central role in the development and progression of atherosclerosis. Current studies have investigated endothelial response using idealized in vitro flow chambers. Such cell culture models are unable to accurately replicate the complex in vivo wall shear stress patterns arising from anatomical geometries. To better understand this implication, we have created both simplified/tubular and anatomically realistic in vitro endothelial flow models of the human right coronary artery. A post-mortem vascular cast of the human left ventricular outflow tract was used to create geometrically accurate silicone elastomer models. Straight, tubular models were created using a custom made mold. Following the culture of human abdominal aortic endothelial cells within the inner lumen, cells were exposed to steady flow (Re = 233 for varying time periods. The resulting cell morphology was analyzed in terms of shape index and angle of orientation relative to the flow direction. In both models a progressive elongation and alignment of the endothelium in the flow direction was observed following 8, 12, and 24 hours. This change, however, was significantly less pronounced in the anatomical model (as observed from morphological variations indicative of localized flow features. Differences were also observed between the inner and outer walls at the disease-prone proximal region. Since morphological adaptation is a visual indication of endothelial shear stress activation, the use of anatomical models in endothelial genetic and biochemical studies may offer better insight into the disease process.

  5. H2S inhibits pulmonary arterial endothelial cell inflammation in rats with monocrotaline-induced pulmonary hypertension.

    Science.gov (United States)

    Feng, Shasha; Chen, Siyao; Yu, Wen; Zhang, Da; Zhang, Chunyu; Tang, Chaoshu; Du, Junbao; Jin, Hongfang

    2017-03-01

    This study aimed to determine whether hydrogen sulfide (H 2 S) inhibits pulmonary arterial endothelial inflammation in rats with monocrotaline (MCT)-induced pulmonary hypertension and its possible mechanisms. Twenty-four male Wistar rats were divided randomly into control, MCT, and MCT+H 2 S treatment groups. Human pulmonary arterial endothelial cells (HPAEC) were cultured and divided into four groups: control, MCT, MCT+H 2 S, and H 2 S. Pulmonary artery pressure was determined using a right cardiac catheterization procedure 3 weeks after MCT administration. Pulmonary vascular morphological changes and inflammatory infiltration were measured. Endogenous H 2 S levels, cystathionine-γ-lyase (CSE) expression, and inflammatory cytokines were determined both in vivo and in vitro. In addition, phosphorylation of NF-κB p65 and IκBα was detected by western blotting, and NF-κB p65 nuclear translocation, as well as its DNA-binding activity, was determined. Pulmonary hypertension and vascular remolding developed 3 wks after MCT administration, with elevated lung tissue inflammatory infiltration and cytokine level associated with activation of the NF-κB pathway, both in vivo and in vitro. However, the endogenous H 2 S/CSE pathway was downregulated in MCT rats. By contrast, an H 2 S donor markedly reduced pulmonary artery pressure, pulmonary vascular structural remolding, and increased lung inflammatory infiltration and cytokine levels of MCT-treated rats. Meanwhile, H 2 S reversed the activation of the NF-κB pathway successfully. The downregulated pulmonary arterial endothelial H 2 S/CSE pathway is involved in the pulmonary inflammatory response in MCT-treated pulmonary hypertensive rats. H 2 S attenuated endothelial inflammation by inhibiting the NF-κB pathway.

  6. 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...

  7. 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.

  8. An anti-inflammatory effect of red microalga polysaccharides in coronary artery endothelial cells.

    Science.gov (United States)

    Levy-Ontman, Oshrat; Huleihel, Mahmoud; Hamias, Rachel; Wolak, Talya; Paran, Esther

    2017-09-01

    Polysaccharides (PSs) produced by the red microalga Porphyridium sp. were reported to exhibit anti-inflammatory bioactivities in the human skin. The primary goal of the present research was to assess whether PSs attenuate inflammatory processes by interfering with tumour necrosis factor-alpha (TNF-α)-induced inflammation, in human coronary artery endothelial cells (HCAECs). Functional and inflammatory markers were quantified in TNF-α-stimulated HCAECs, with and without pre-treatment with PSs. The expression/activation of these markers was assessed by Western immunoblotting and a luciferase reporter assay. NO levels were measured using the Griess method and intracellular reactive oxygen stress (ROS) was determined with the fluorescent probe 2',7'-dichlorodihydro-fluorescein diacetate (H2DCFDA). The TNF-α-induced up-regulation of inter-cellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) translocation, as well as IκB degradation were significantly attenuated in cells pre-treated with PSs. In addition, PSs were able to inhibit NF-κB activation as well as TNF-α-induced oxidative stress in HCAECs. Endothelial function was also improved, as measured by increased nitric oxide (NO) formation and decreased endothelin (ET-1) protein expression. This is the first report that demonstrates the anti-inflammatory effect and vaso-relaxing property of red microalgae PSs in a HCAEC-TNF-α induced system. This study lays the foundation for basic research concerning the PS mode of action in biochemical processes involving endothelial dysfunction, and it also holds potential for applied research, possibly promoting the use of PSs as a therapeutic agent or food additive to improve vascular health. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Experimental sleep restriction causes endothelial dysfunction in healthy humans.

    Science.gov (United States)

    Calvin, Andrew D; Covassin, Naima; Kremers, Walter K; Adachi, Taro; Macedo, Paula; Albuquerque, Felipe N; Bukartyk, Jan; Davison, Diane E; Levine, James A; Singh, Prachi; Wang, Shihan; Somers, Virend K

    2014-11-25

    Epidemiologic evidence suggests a link between short sleep duration and cardiovascular risk, although the nature of any relationship and mechanisms remain unclear. Short sleep duration has also been linked to an increase in cardiovascular events. Endothelial dysfunction has itself been implicated as a mediator of heightened cardiovascular risk. We sought to determine the effect of 8 days/8 nights of partial sleep restriction on endothelial function in healthy humans. Sixteen healthy volunteers underwent a randomized study of usual sleep versus sleep restriction of two-thirds normal sleep time for 8 days/8 nights in a hospital-based clinical research unit. The main outcome was endothelial function measured by flow-mediated brachial artery vasodilatation (FMD). Those randomized to sleep restriction slept 5.1 hours/night during the experimental period compared with 6.9 hours/night in the control group. Sleep restriction was associated with significant impairment in FMD (8.6±4.6% during the initial pre-randomization acclimation phase versus 5.2±3.4% during the randomized experimental phase, P=0.01) whereas no change was seen in the control group (5.0±3.0 during the acclimation phase versus 6.73±2.9% during the experimental phase, P=0.10) for a between-groups difference of -4.40% (95% CI -7.00 to -1.81%, P=0.003). No change was seen in non-flow mediated vasodilatation (NFMD) in either group. In healthy individuals, moderate sleep restriction causes endothelial dysfunction. ClinicalTrials.gov. Unique identifier: NCT01334788. © 2014 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

  10. Iptakalim attenuates hypoxia-induced pulmonary arterial hypertension in rats by endothelial function protection.

    Science.gov (United States)

    Zhu, Rong; Bi, Li-Qing; Wu, Su-Ling; Li, Lan; Kong, Hui; Xie, Wei-Ping; Wang, Hong; Meng, Zi-Li

    2015-08-01

    The present study aimed to investigate the protective effects of iptakalim, an adenosine triphosphate (ATP)-sensitive potassium channel opener, on the inflammation of the pulmonary artery and endothelial cell injury in a hypoxia-induced pulmonary arterial hypertension (PAH) rat model. Ninety-six Sprague-Dawley rats were placed into normobaric hypoxia chambers for four weeks and were treated with iptakalim (1.5 mg/kg/day) or saline for 28 days. The right ventricle systolic pressures (RVSP) were measured and small pulmonary arterial morphological alterations were analyzed with hematoxylin and eosin staining. Enzyme-linked immunosorbent assay (ELISA) was performed to analyze the content of interleukin (IL)-1β and IL-10. Immunohistochemical analysis for ED1(+) monocytes was performed to detect the inflammatory cells surrounding the pulmonary arterioles. Western blot analysis was performed to analyze the expression levels of platelet endothelial cell adhesion molecule-1 (PECAM-1) and endothelial nitric oxide synthase (eNOS) in the lung tissue. Alterations in small pulmonary arteriole morphology and the ultrastructure of pulmonary arterial endothelial cells were observed via light and transmission electron microscopy, respectively. Iptakalim significantly attenuated the increase in mean pulmonary artery pressure, RVSP, right ventricle to left ventricle plus septum ratio and small pulmonary artery wall remodeling in hypoxia-induced PAH rats. Iptakalim also prevented an increase in IL-1β and a decrease in IL-10 in the peripheral blood and lung tissue, and alleviated inflammatory cell infiltration in hypoxia-induced PAH rats. Furthermore, iptakalim enhanced PECAM-1 and eNOS expression and prevented the endothelial cell injury induced by hypoxic stimuli. Iptakalim suppressed the pulmonary arteriole and systemic inflammatory responses and protected against the endothelial damage associated with the upregulation of PECAM-1 and eNOS, suggesting that iptakalim may represent a

  11. Arterial Injury and Endothelial Repair: Rapid Recovery of Function after Mechanical Injury in Healthy Volunteers

    Directory of Open Access Journals (Sweden)

    Lindsey Tilling

    2014-01-01

    Full Text Available Objective. Previous studies suggest a protracted course of recovery after mechanical endothelial injury; confounders may include degree of injury and concomitant endothelial dysfunction. We sought to define the time course of endothelial function recovery using flow-mediated dilation (FMD, after ischaemia-reperfusion (IR and mechanical injury in patients and healthy volunteers. The contribution of circulating CD133+/CD34+/VEGFR2+ “endothelial progenitor” (EPC or repair cells to endothelial repair was also examined. Methods. 28 healthy volunteers aged 18–35 years underwent transient forearm ischaemia induced by cuff inflation around the proximal biceps and radial artery mechanical injury induced by inserting a wire through a cannula. A more severe mechanical injury was induced using an arterial sheath and catheter inserted into the radial artery of 18 patients undergoing angiography. Results. IR and mechanical injury produced immediate impairment of FMD (from 6.5 ± 1.2% to 2.9 ± 2.2% and from 7.4 ± 2.3% to 1.5 ± 1.6% for IR and injury, resp., each P<0.001 but recovered within 6 hours and 2 days, respectively. FMD took up to 4 months to recover in patients. Circulating EPC did not change significantly during the injury/recovery period in all subjects. Conclusions. Recovery of endothelial function after IR and mechanical injury is rapid and not associated with a change in circulating EPC.

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  20. Chronic hypoxia promotes pulmonary artery endothelial cell proliferation through H2O2-induced 5-lipoxygenase.

    Directory of Open Access Journals (Sweden)

    Kristi M Porter

    Full Text Available Pulmonary Hypertension (PH is a progressive disorder characterized by endothelial dysfunction and proliferation. Hypoxia induces PH by increasing vascular remodeling. A potential mediator in hypoxia-induced PH development is arachidonate 5-Lipoxygenase (ALOX5. While ALOX5 metabolites have been shown to promote pulmonary vasoconstriction and endothelial cell proliferation, the contribution of ALOX5 to hypoxia-induced proliferation remains unknown. We hypothesize that hypoxia exposure stimulates HPAEC proliferation by increasing ALOX5 expression and activity. To test this, human pulmonary artery endothelial cells (HPAEC were cultured under normoxic (21% O2 or hypoxic (1% O2 conditions for 24-, 48-, or 72 hours. In a subset of cells, the ALOX5 inhibitor, zileuton, or the 5-lipoxygenase activating protein inhibitor, MK-886, was administered during hypoxia exposure. ALOX5 expression was measured by qRT-PCR and western blot and HPAEC proliferation was assessed. Our results demonstrate that 24 and 48 hours of hypoxia exposure have no effect on HPAEC proliferation or ALOX5 expression. Seventy two hours of hypoxia significantly increases HPAEC ALOX5 expression, hydrogen peroxide (H2O2 release, and HPAEC proliferation. We also demonstrate that targeted ALOX5 gene silencing or inhibition of the ALOX5 pathway by pharmacological blockade attenuates hypoxia-induced HPAEC proliferation. Furthermore, our findings indicate that hypoxia-induced increases in cell proliferation and ALOX5 expression are dependent on H2O2 production, as administration of the antioxidant PEG-catalase blocks these effects and addition of H2O2 to HPAEC promotes proliferation. Overall, these studies indicate that hypoxia exposure induces HPAEC proliferation by activating the ALOX5 pathway via the generation of H2O2.

  1. Relationship between upper and lower limb conduit artery vasodilator function in humans.

    Science.gov (United States)

    Thijssen, Dick H J; Rowley, Nicola; Padilla, Jaume; Simmons, Grant H; Laughlin, M Harold; Whyte, Greg; Cable, N Timothy; Green, Daniel J

    2011-07-01

    Brachial artery flow-mediated dilation (FMD) is a strong predictor of future cardiovascular disease and is believed to represent a "barometer" of systemic endothelial health. Although a recent study [Padilla et al. Exp Biol Med (Maywood) 235: 1287-1291, 2010] in pigs confirmed a strong correlation between brachial and femoral artery endothelial function, it is unclear to what extent brachial artery FMD represents a systemic index of endothelial function in humans. We conducted a retrospective analysis of data from our laboratory to evaluate relationships between the upper (i.e., brachial artery) vs. lower limb (superficial femoral n = 75; popliteal artery n = 32) endothelium-dependent FMD and endothelium-independent glyceryl trinitrate (GTN)-mediated dilation in young, healthy individuals. We also examined the relationship between FMD assessed in both brachial arteries (n = 42). There was no correlation between brachial and superficial femoral artery FMD (r(2) = 0.008; P = 0.46) or between brachial and popliteal artery FMD (r(2) = 0.003; P = 0.78). However, a correlation was observed in FMD between both brachial arteries (r(2) = 0.34; P < 0.001). Brachial and superficial femoral artery GTN were modestly correlated (r(2) = 0.13; P = 0.007), but brachial and popliteal artery GTN responses were not (r(2) = 0.08; P = 0.11). Collectively, these data indicate that conduit artery vasodilator function in the upper limbs (of healthy humans) is not predictive of that in the lower limbs, whereas measurement of FMD in one arm appears to be predictive of FMD in the other. These data do not support the hypothesis that brachial artery FMD in healthy humans represents a systemic index of endothelial function.

  2. 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.

  3. Copper Dependence of Angioproliferation in Pulmonary Arterial Hypertension in Rats and Humans

    Science.gov (United States)

    Mizuno, Shiro; Guignabert, Christophe; Al Hussaini, Aysar A.; Farkas, Daniela; Ruiter, Gerrina; Kraskauskas, Donatas; Fadel, Elie; Allegood, Jeremy C.; Humbert, Marc; Noordegraaf, Anton Vonk; Spiegel, Sarah; Farkas, Laszlo; Voelkel, Norbert F.

    2012-01-01

    Obliteration of the vascular lumen by endothelial cell growth is a hallmark of many forms of severe pulmonary arterial hypertension. Copper plays a significant role in the control of endothelial cell proliferation in cancer and wound-healing. We sought to determine whether angioproliferation in rats with experimental pulmonary arterial hypertension and pulmonary microvascular endothelial cell proliferation in humans depend on the proangiogenic action of copper. A copper-depleted diet prevented, and copper chelation with tetrathiomolybdate reversed, the development of severe experimental pulmonary arterial hypertension. The copper chelation–induced reopening of obliterated vessels was caused by caspase-independent apoptosis, reduced vessel wall cell proliferation, and a normalization of vessel wall structure. No evidence was found for a role of super oxide–1 inhibition or lysyl–oxidase–1 inhibition in the reversal of angioproliferation. Tetrathiomolybdate inhibited the proliferation of human pulmonary microvascular endothelial cells, isolated from explanted lungs from control subjects and patients with pulmonary arterial hypertension. These data suggest that the inhibition of endothelial cell proliferation by a copper-restricting strategy could be explored as a new therapeutic approach in pulmonary arterial hypertension. It remains to be determined, however, whether potential toxicity to the right ventricle is offset by the beneficial pulmonary vascular effects of antiangiogenic treatment in patients with pulmonary arterial hypertension. PMID:22162909

  4. 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...

  5. Effects of cigarette smoke on endothelial function of pulmonary arteries in the guinea pig

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    Martínez Anna

    2009-08-01

    Full Text Available Abstract Background Cigarette smoking may contribute to pulmonary hypertension in chronic obstructive pulmonary disease by altering the structure and function of pulmonary vessels at early disease stages. The objectives of this study were to evaluate the effects of long-term exposure to cigarette smoke on endothelial function and smooth muscle-cell proliferation in pulmonary arteries of guinea pigs. Methods 19 male Hartley guinea pigs were exposed to the smoke of 7 cigarettes/day, 5 days/week, for 3 and 6 months. 17 control guinea pigs were sham-exposed for the same periods. Endothelial function was evaluated in rings of pulmonary artery and aorta as the relaxation induced by ADP. The proliferation of smooth muscle cells and their phenotype in small pulmonary vessels were evaluated by immunohistochemical expression of α-actin and desmin. Vessel wall thickness, arteriolar muscularization and emphysema were assessed morphometrically. The expression of endothelial nitric oxide synthase (eNOS was evaluated by Real Time-PCR. Results Exposure to cigarette smoke reduced endothelium-dependent vasodilatation in pulmonary arteries (ANOVA p Conclusion In the guinea pig, exposure to cigarette smoke induces selective endothelial dysfunction in pulmonary arteries, smooth muscle cell proliferation in small pulmonary vessels and reduced lung expression of eNOS. These changes appear after 3 months of exposure and precede the development of pulmonary emphysema.

  6. Endothelial function and gene expression in perivascular adipose tissue from internal mammary arteries of obese patients with coronary artery disease.

    Science.gov (United States)

    Cybularz, Maria; Langbein, Heike; Zatschler, Birgit; Brunssen, Coy; Deussen, Andreas; Matschke, Klaus; Morawietz, Henning

    2017-11-01

    Obesity is a risk factor for endothelial dysfunction and atherosclerosis. However, perivascular adipose tissue can release adipokines and other unknown adipose-derived relaxing factors. Therefore, we investigated the impact of obesity on vascular function and expression of genes in perivascular adipose tissue from internal mammary arteries of patients with coronary artery disease undergoing coronary artery bypass grafting. The vessel function was compared between groups of patients with a body-mass index (BMI) between 25 and 30 kg/m 2 . The groups did not differ in age, gender (males), and ejection fraction. Vascular segments of internal mammary arteries were examined in a Mulvany myograph. Following preconstriction with noradrenaline, dose-response curves were assessed for relaxation with acetylcholine and sodium nitroprusside. Maximum contraction in response to potassium and noradrenaline was increased in obese patients with a BMI >30 kg/m 2 . EC50 of endothelium-dependent relaxation was impaired in patients with a BMI above 25, but below 30 kg/m 2 . Sodium nitroprusside-mediated maximal relaxation was not different between study groups. Integrin alpha X chain (ITGAX/CD11c) and macrophage mannose receptor (MRC1/CD206) expression was reduced in perivascular adipose tissue of patients with a BMI above 30 kg/m 2 , while adiponectin (ADPQ) expression was increased in the same tissue. Our data suggest a partially reduced endothelial function in internal mammary arteries of adipose patients with a BMI between 25 and 30 kg/m 2 undergoing coronary artery bypass grafting surgery. Increased adiponectin expression in perivascular tissue might contribute to maintenance of endothelial function in obese patients with a BMI above 30 kg/m 2 . Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Eugenol dilates mesenteric arteries and reduces systemic BP by activating endothelial cell TRPV4 channels.

    Science.gov (United States)

    Peixoto-Neves, Dieniffer; Wang, Qian; Leal-Cardoso, Jose H; Rossoni, Luciana V; Jaggar, Jonathan H

    2015-07-01

    Eugenol, a vanilloid molecule found in some dietary plants, relaxes vasculature in part via an endothelium-dependent process; however, the mechanisms involved are unclear. Here, we investigated the endothelial cell-mediated mechanism by which eugenol modulates rat mesenteric artery contractility and systemic BP. The isometric tension of rat mesenteric arteries (size 200-300 μm) was measured using wire myography; non-selective cation currents (ICat ) were recorded in endothelial cells using patch clamp electrophysiology. Mean arterial pressure (MAP) and heart rate (HR) were determined in anaesthetized rats. Eugenol relaxed endothelium-intact arteries in a concentration-dependent manner and this effect was attenuated by endothelium denudation. L-NAME, a NOS inhibitor, a combination of TRAM-34 and apamin, selective blockers of intermediate and small conductance Ca(2+) -activated K(+) channels, respectively, and HC-067047, a TRPV4 channel inhibitor, but not indomethacin, a COX inhibitor, reduced eugenol-induced relaxation in endothelium-intact arteries. Eugenol activated HC-067047-sensitive ICat in mesenteric artery endothelial cells. Short interfering RNA (siRNA)-mediated TRPV4 knockdown abolished eugenol-induced ICat activation. An i.v. injection of eugenol caused an immediate, transient reduction in both MAP and HR, which was followed by prolonged, sustained hypotension in anaesthetized rats. This sustained hypotension was blocked by HC-067047. Eugenol activates TRPV4 channels in mesenteric artery endothelial cells, leading to vasorelaxation, and reduces systemic BP in vivo. Eugenol may be therapeutically useful as an antihypertensive agent and is a viable molecular candidate from which to develop second-generation TRPV4 channel activators that reduce BP. © 2015 The British Pharmacological Society.

  8. ROCK2 mediates the proliferation of pulmonary arterial endothelial cells induced by hypoxia in the development of pulmonary arterial hypertension

    OpenAIRE

    Qiao, Feng; ZOU, ZHITIAN; Liu, Chunhui; Zhu, Xiaofeng; Wang, Xiaoqiang; Yang, Chengpeng; JIANG, TENGJIAO; Chen, Ying

    2016-01-01

    It has been reported that RhoA activation and Rho-kinase (ROCK) expression are increased in chronic hypoxic lungs, and the long-term inhibition of ROCK markedly improves the survival of patients with pulmonary arterial hypertension (PAH). However, whether Rho-kinase α (ROCK2) participates in regulation of the growth of pulmonary arterial endothelial cells (PAECs) remains unknown. The aim of the present study was to investigate the effect of hypoxia on the proliferation of PAECs and the role o...

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

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    Daniel Kotschy

    2015-08-01

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

  10. Microvascular Coronary Artery Spasm Presents Distinctive Clinical Features With Endothelial Dysfunction as Nonobstructive Coronary Artery Disease

    OpenAIRE

    Ohba, Keisuke; Sugiyama, Seigo; Sumida, Hitoshi; Nozaki, Toshimitsu; Matsubara, Junichi; Matsuzawa, Yasushi; Konishi, Masaaki; Akiyama, Eiichi; Kurokawa, Hirofumi; Maeda, Hirofumi; Sugamura, Koichi; Nagayoshi, Yasuhiro; Morihisa, Kenji; Sakamoto, Kenji; Tsujita, Kenichi

    2012-01-01

    Background Angina without significant stenosis, or nonobstructive coronary artery disease, attracts clinical attention. Microvascular coronary artery spasm (microvascular CAS) can cause nonobstructive coronary artery disease. We investigated the clinical features of microvascular CAS and the therapeutic efficacy of calcium channel blockers. Methods and Results Three hundred seventy consecutive, stable patients with suspected angina presenting nonobstructive coronary arteries (

  11. Tumor endothelial inflammation predicts clinical outcome in diverse human cancers.

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    Sean P Pitroda

    Full Text Available Vascular endothelial cells contribute to the pathogenesis of numerous human diseases by actively regulating the stromal inflammatory response; however, little is known regarding the role of endothelial inflammation in the growth of human tumors and its influence on the prognosis of human cancers.Using an experimental model of tumor necrosis factor-alpha (TNF-α-mediated inflammation, we characterized inflammatory gene expression in immunopurified tumor-associated endothelial cells. These genes formed the basis of a multivariate molecular predictor of overall survival that was trained and validated in four types of human cancer.We report that expression of experimentally derived tumor endothelial genes distinguished pathologic tissue specimens from normal controls in several human diseases associated with chronic inflammation. We trained these genes in human cancer datasets and defined a six-gene inflammatory signature that predicted significantly reduced overall survival in breast cancer, colon cancer, lung cancer, and glioma. This endothelial-derived signature predicted outcome independently of, but cooperatively with, standard clinical and pathological prognostic factors. Consistent with these findings, conditioned culture media from human endothelial cells stimulated by pro-inflammatory cytokines accelerated the growth of human colon and breast tumors in immunodeficient mice as compared with conditioned media from untreated endothelial cells.This study provides the first prognostic cancer gene signature derived from an experimental model of tumor-associated endothelial inflammation. These findings support the notion that activation of inflammatory pathways in non-malignant tumor-infiltrating endothelial cells contributes to tumor growth and progression in multiple human cancers. Importantly, these results identify endothelial-derived factors that could serve as potential targets for therapy in diverse human cancers.

  12. Endothelial Protective Monocyte Patrolling in Large Arteries Intensified by Western Diet and Atherosclerosis

    Science.gov (United States)

    Quintar, Amado; McArdle, Sara; Wolf, Dennis; Marki, Alex; Ehinger, Erik; Vassallo, Melanie; Miller, Jacqueline; Mikulski, Zbigniew; Ley, Klaus

    2017-01-01

    Rationale: Nonclassical mouse monocyte (CX3CR1high, Ly-6Clow) patrolling along the vessels of the microcirculation is critical for endothelial homeostasis and inflammation. Because of technical challenges, it is currently not established how patrolling occurs in large arteries. Objective: This study was undertaken to elucidate the molecular, migratory, and functional phenotypes of patrolling monocytes in the high shear and pulsatile environment of large arteries in healthy, hyperlipidemic, and atherosclerotic conditions. Methods and Results: Applying a new method for stable, long-term 2-photon intravital microscopy of unrestrained large arteries in live CX3CR1-GFP (green fluorescent protein) mice, we show that nonclassical monocytes patrol inside healthy carotid arteries at a velocity of 36 μm/min, 3× faster than in microvessels. The tracks are less straight but lead preferentially downstream. The number of patrolling monocytes is increased 9-fold by feeding wild-type mice a Western diet or by applying topical TLR7/8 (Toll-like receptor) agonists. A similar increase is seen in CX3CR1+/GFP/apoE−/− mice on chow diet, with a further 2- to 3-fold increase on Western diet (22-fold over healthy). In plaque conditions, monocytes are readily captured onto the endothelium from free flow. Stable patrolling is unaffected in CX3CR1-deficient mice and involves the contribution of LFA-1 (lymphocyte-associated antigen 1) and α4 integrins. The endothelial damage in atherosclerotic carotid arteries was assessed by electron microscopy and correlates with the number of intraluminal patrollers. Abolishing patrolling monocytes in Nr4a1−/− apoE−/− mice leads to pronounced endothelial apoptosis. Conclusions: Arterial patrolling is a prominent new feature of nonclassical monocytes with unique molecular and kinetic properties. It is highly upregulated in hyperlipidemia and atherosclerosis in a CX3CR1-independent fashion and plays a potential role in endothelial protection

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

    Science.gov (United States)

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

    2014-04-14

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

  14. Clopidogrel improves microvascular endothelial function in subjects with stable coronary artery disease.

    Science.gov (United States)

    Willoughby, Scott R; Luu, Lee-Jen; Cameron, James D; Nelson, Adam J; Schultz, Carlee D; Worthley, Stephen G; Worthley, Matthew I

    2014-06-01

    Clopidogrel therapy has recently been shown to reduce cardiovascular events in patients with stable vascular disease. This benefit may be due to effects not exclusively related to platelet aggregation. The aim of this study was to evaluate the effect of clopidogrel therapy on microvascular endothelial function in subjects with stable coronary artery disease (CAD). Forty subjects with stable CAD were randomised to clopidogrel therapy (75mg/day) or control. Blood and endothelial function testing occurred at baseline, one week and three months following randomisation. Microvascular endothelial function was assessed via reactive hyperaemic index (RHI). Platelet function was assessed by adenosine diphosphate (ADP)-induced whole blood aggregation and the VerifyNow™ system. Plasma markers of endothelial function (asymmetric dimethylarginine, ADMA) and oxidative stress (myeloperoxidase, MPO) were also tested. The primary endpoint was endothelial function assessment (RHI) at three months. At one week RHI increased by 20±10% in the clopidogrel group; this effect was maintained at three months (21±9% increase from baseline; Pmicrovascular endothelial function in patients with stable CAD. This effect is independent of its effects on ADP-induced platelet reactivity. Copyright © 2014 Australian and New Zealand Society of Cardiac and Thoracic Surgeons (ANZSCTS) and the Cardiac Society of Australia and New Zealand (CSANZ). Published by Elsevier B.V. All rights reserved.

  15. Can fish oil supplementation improve endothelial function in asymptomatic offspring of patients with peripheral arterial disease?

    Directory of Open Access Journals (Sweden)

    Spark JI

    2013-07-01

    Full Text Available J Ian Spark,1 Christopher L Delaney,1 Richard B Allan,1 Melissa HL Ho,2 Michelle D Miller21Department of Vascular Surgery, Flinders Medical Centre and Flinders University, 2Department of Nutrition and Dietetics, Flinders University, Bedford Park, Adelaide, South Australia, AustraliaBackground: Peripheral arterial disease affects 10%–25% of adults aged .55 years, and while a multitude of risk factors exist, one key influence is genetics. Rather than awaiting the onset of debilitating symptoms, interventions that target high-risk individuals and prevent or delay the onset of symptoms would have widespread impact. The aim of this study is to implement a 12-week fish oil intervention (10 mL/day containing approximately 1.5 g of eicosapentaenoic acid and 1 g of docosahexaenoic acid, with the intention of improving endothelial function, inflammation, and lipid status in a high-risk population, ie, those with impaired endothelial function and a parent with symptomatic peripheral arterial disease.Methods: This is a parallel-group, double-blind, randomized controlled trial involving administration of fish oil containing either about 1.5 g of docosahexaenoic acid and 1 g of docosahexaenoic acid (intervention or about 0.15 g of eicosapentaenoic acid and about 0.1 g of docosahexaenoic acid for 12 consecutive weeks (control. The participants are 100 offspring of adults with diagnosed peripheral arterial disease who themselves have an ankle-brachial pressure index ≥0.9 but impaired endothelial function according to peripheral arterial tonometry. Measures performed at baseline and at 6 and 12 weeks include flow-mediated dilatation, C-reactive protein, absolute neutrophil and lymphocyte counts, tumor necrosis factor-α, interleukin-1ß, and interleukin-6 levels, thromboxane and prostacyclin, lipid status, and homocysteine, nitrite, and nitrate levels. Participants will be phoned fortnightly to monitor adherence and side effects, while participants will

  16. Microfluidic assay of circulating endothelial cells in coronary artery disease patients with angina pectoris.

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    Shuiyu Chen

    Full Text Available Circulating endothelial cells (CECs are widely reported as a promising biomarker of endothelial damage/dysfunction in coronary artery disease (CAD. The two popular methods of CEC quantification include the use of immunomagnetic beads separation (IB and flow cytometry analysis (FC; however, they suffer from two main shortcomings that affect their diagnostic and prognostic responses: non-specific bindings of magnetic beads to non-target cells and a high degree of variability in rare cell identification, respectively. We designed a microfluidic chip with spatially staggered micropillars for the efficient harvesting of CECs with intact cellular morphology in an attempt to revisit the diagnostic goal of CEC counts in CAD patients with angina pectoris.A label-free microfluidic assay that involved an in-situ enumeration and immunofluorescent identification (DAPI+/CD146+/VEGFR1+/CD45- of CECs was carried out to assess the CEC count in human peripheral blood samples. A total of 55 CAD patients with angina pectoris [16 with chronic stable angina (CSA and 39 with unstable angina (UA], together with 15 heathy controls (HCs were enrolled in the study.CEC counts are significantly higher in both CSA and UA groups compared to the HC group [respective medians of 6.9, 10.0 and 1.5 cells/ml (p < 0.01]. Further, a significant elevation of CEC count was observed in the three UA subgroups [low risk (5.3 vs. intermediate risk (10.8 vs. high risk (18.0 cells/ml, p < 0.001 classified in accordance to the TIMI NSTEMI/UA risk score system. From the receiver-operating characteristic curve analysis, the AUCs for distinguishing CSA and UA from HC were 0.867 and 0.938, respectively. The corresponding sensitivities were 87.5% and 84.6% and the specificities were 66.7% and 86.7%, respectively.Our microfluidic assay system is efficient and stable for CEC capture and enumeration. The results showed that the CEC count has the potential to be a promising clinical biomarker for the

  17. Nicorandil attenuates monocrotaline-induced vascular endothelial damage and pulmonary arterial hypertension.

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    Makoto Sahara

    Full Text Available BACKGROUND: An antianginal K(ATP channel opener nicorandil has various beneficial effects on cardiovascular systems; however, its effects on pulmonary vasculature under pulmonary arterial hypertension (PAH have not yet been elucidated. Therefore, we attempted to determine whether nicorandil can attenuate monocrotaline (MCT-induced PAH in rats. MATERIALS AND METHODS: Sprague-Dawley rats injected intraperitoneally with 60 mg/kg MCT were randomized to receive either vehicle; nicorandil (5.0 mg·kg(-1·day(-1 alone; or nicorandil as well as either a K(ATP channel blocker glibenclamide or a nitric oxide synthase (NOS inhibitor N(ω-nitro-L-arginine methyl ester (L-NAME, from immediately or 21 days after MCT injection. Four or five weeks later, right ventricular systolic pressure (RVSP was measured, and lung tissue was harvested. Also, we evaluated the nicorandil-induced anti-apoptotic effects and activation status of several molecules in cell survival signaling pathway in vitro using human umbilical vein endothelial cells (HUVECs. RESULTS: Four weeks after MCT injection, RVSP was significantly increased in the vehicle-treated group (51.0±4.7 mm Hg, whereas it was attenuated by nicorandil treatment (33.2±3.9 mm Hg; P<0.01. Nicorandil protected pulmonary endothelium from the MCT-induced thromboemboli formation and induction of apoptosis, accompanied with both upregulation of endothelial NOS (eNOS expression and downregulation of cleaved caspase-3 expression. Late treatment with nicorandil for the established PAH was also effective in suppressing the additional progression of PAH. These beneficial effects of nicorandil were blocked similarly by glibenclamide and l-NAME. Next, HUVECs were incubated in serum-free medium and then exhibited apoptotic morphology, while these changes were significantly attenuated by nicorandil administration. Nicorandil activated the phosphatidylinositol 3-kinase (PI3K/Akt and extracellular signal-regulated kinase (ERK

  18. Are retinal microvascular abnormalities associated with large artery endothelial dysfunction and intima-media thickness? The Hoorn Study

    NARCIS (Netherlands)

    van Hecke, M.V.; Dekker, J.M.; Nijpels, G.; Stolk, R.P.; Henry, R.M.A.; Heine, R.J.; Bouter, L.M.; Stehouwer, C.D.A.; Polak, B.C.P.

    It has been hypothesized that microvascular dysfunction affects endothelial dysfunction of the large arteries, which may explain the relationship of microvascular disease with macrovascular disease. The aim of the present study was to investigate the relationship of retinal microvascular disorders

  19. Assessment of Post-Occlusive Reactive Hyperaemia in the Evaluation of Endothelial Function in Patients with Lower Extremity Artery Disease

    Directory of Open Access Journals (Sweden)

    Carasca Cosmin

    2017-09-01

    Full Text Available Background: The aim was to assess endothelial function with photoplethysmography (PPG, by post-occlusive reactive hyperaemia (PORH combined with alprostadil challenge test in patients with peripheral artery disease (PAD.

  20. Erythropoietin Attenuates Pulmonary Vascular Remodeling in Experimental Pulmonary Arterial Hypertension through Interplay between Endothelial Progenitor Cells and Heme Oxygenase

    NARCIS (Netherlands)

    van Loon, Rosa Laura E; Bartelds, Beatrijs; Wagener, Frank A D T G; Affara, Nada; Mohaupt, Saffloer; Wijnberg, Hans; Pennings, Sebastiaan W C; Takens, Janny; Berger, Rolf M F

    2015-01-01

    BACKGROUND: Pulmonary arterial hypertension (PAH) is a pulmonary vascular disease with a high mortality, characterized by typical angio-proliferative lesions. Erythropoietin (EPO) attenuates pulmonary vascular remodeling in PAH. We postulated that EPO acts through mobilization of endothelial

  1. Endothelial Function in Adolescents with a History of Premature Coronary Artery Disease in One Parent

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    M Hashemi

    2006-01-01

    Full Text Available Background: In young adults, a family history of premature coronary artery disease (CAD, as well as genetic and environmental factors are independent risk factors for coronary artery disease. Methods: Endothelial function was studied in 30 children (21 boys and 9 girls with mean age of 14.9 +/- 2.3 years old of patients with documented CAD (men 45 and women 50 years old. Chidren did not have any history of diabetes mellitus, dyslipidemia, hypertension, and smoking (active/passive. Using vascular ultrasound, we measured resting Basal Brachial artery Diameter (BBD and Endothelium-Dependent Dilatation (EDD in response to increased flow and sublingual glyceryltrinitrate (GTN, an Endothelium-Independent Dilation (EID. These parameters were also measured in 30 control subjects with normal parents (18 boys and 12 girls with mean age of 14.2 +/- 2/5years old and results were compared with each other. Results: Adolescents in CAD group had abnormal Endothelial Dependent Dilatation or EDD/BBD (8.5 +/- 3.4% vs 11.8 +/- 4.5% in control subjects; P= 0.003.Endothelial Independent Dilatation (EID/BBD in the positive fimily history group was significantly more than control subjects (18.5 +/- 6.7% vs 11.9 +/- 5.2%; P <0.001. EDD/EID or the index of endothelial function was significantly lower in the positive family history group (0.92 +/- 0.05 vs 1+/- 0.03; P<0.001. There was no difference in EDD/EID index between those with history of premature CAD in mother (7 cases and those with history of premature CAD in father (23 cases (0.92 +/- 0.04 vs 0.91+/- 0.05. Conclusion: Normal adolescents without any cardiovascular risk factors but a history of premature coronary artery disease in one parent may have endothelial dysfunction, and there is no difference whether the CAD is in mother or father. Keywords: Endothelial dependent dilation, family history, CAD risk factors, premature coronary artery disease

  2. Quercetin Inhibits Pulmonary Arterial Endothelial Cell Transdifferentiation Possibly by Akt and Erk1/2 Pathways

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    Shian Huang

    2017-01-01

    Full Text Available This study aimed to investigate the effects and mechanisms of quercetin on pulmonary arterial endothelial cell (PAEC transdifferentiation into smooth muscle-like cells. TGF-β1-induced PAEC transdifferentiation models were applied to evaluate the pharmacological actions of quercetin. PAEC proliferation was detected with CCK8 method and BurdU immunocytochemistry. Meanwhile, the identification and transdifferentiation of PAECs were determined by FVIII immunofluorescence staining and α-SMA protein expression. The related mechanism was elucidated based on the levels of Akt and Erk1/2 signal pathways. As a result, quercetin effectively inhibited the TGF-β1-induced proliferation and transdifferentiation of the PAECs and activation of Akt/Erk1/2 cascade in the cells. In conclusion, quercetin is demonstrated to be effective for pulmonary arterial hypertension (PAH probably by inhibiting endothelial transdifferentiation possibly via modulating Akt and Erk1/2 expressions.

  3. [The altered endothelial function in patients with arterial hypertension and different forms of atrial fibrillation].

    Science.gov (United States)

    Podzolkov, V I; Tarzimanova, A I; Mokhammadi, L N

    2014-01-01

    The role of endothelial function in the development of cardiovascular diseases has recently attracted attention of many researchers due to increasingly more data suggesting the relationship between endothelial dysfunction (ED) and disturbed cardiac rhythms including atrial fibrillation (AF). ED is known to precede lesions in target organs related to arterial hypertension (AH) which makes the study of endothelial function as an early marker of vascular lesions in AH and AF a topical issue. To study changes of endothelial function in patients with AH and AF. Group 1 included 84 patients with AH (inclusion criteria: essential AH and confirmed paroxysm of AF), group 2 contained 20 patients with AH and permanent AF, control group was comprised of 30 AH patients without AF. The vasomotor function of endothelium was evaluated from reactive hyperemia determined by the ultrasonic method, blood samples for biochemical analysis and determination of Willebrand factor (WF) were taken during fasting. Patients of group 2 showed significant changes of endothelium-dependent vasodilation of the brachial artery. Its diameter within 60 sec after decompression increased by 5.8 +/- 0.9% and 12.3 +/- 1.2% in groups 1 and 3 respectively (p < 0.05). In group 2, collagen-binding activity of WF increased significantly to 1500 +/-140 U/100 ml compared with 1060 +/- 120 and 840 +/- 110 in groups 2 and 3 (p < 0.05). Patients with AH and persistent AF had altered endothelial function in the form of significant decrease of endothelium-dependent vasodilation of the brachial artery and increase of collagen-binding activity of WF.

  4. Endothelial cells derived from human embryonic stem cells

    Science.gov (United States)

    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.

  5. Arterial stiffness and endothelial dysfunction independently and synergistically predict cardiovascular and renal outcome in patients with type 1 diabetes

    DEFF Research Database (Denmark)

    Theilade, S; Lajer, Maria Stenkil; Jorsal, Anders

    2012-01-01

    To evaluate whether pulse pressure alone or with placental growth factor as estimates of arterial stiffness and endothelial dysfunction, predicts mortality, cardiovascular disease and progression to end-stage renal disease in patients with Type 1 diabetes.......To evaluate whether pulse pressure alone or with placental growth factor as estimates of arterial stiffness and endothelial dysfunction, predicts mortality, cardiovascular disease and progression to end-stage renal disease in patients with Type 1 diabetes....

  6. 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.

  7. Endothelial dysfunction, carotid artery plaque burden, and conventional exercise-induced myocardial ischemia as predictors of coronary artery disease prognosis

    Directory of Open Access Journals (Sweden)

    Ishihara Masayuki

    2008-12-01

    Full Text Available Abstract Background While both flow-mediated vasodilation (FMD in the brachial artery (BA, which measures endothelium-dependent vasodilatation, and intima-media thickness (IMT in the carotid artery are correlated with the prognosis of coronary artery disease (CAD, it is not clear which modality is a better predictor of CAD. Furthermore, it has not been fully determined whether either of these modalities is superior to conventional ST-segment depression on exercise stress electrocardiogram (ECG as a predictor. Thus, the goal of the present study was to compare the predictive value of FMD, IMT, and stress ECG for CAD prognosis. Methods and Results A total of 103 consecutive patients (62 ± 9 years old, 79 men with clinically suspected CAD had FMD and nitroglycerin-induced dilation (NTG-D in the BA, carotid artery IMT measurement using high-resolution ultrasound, and exercise treadmill testing. The 73 CAD patients and 30 normal coronary patients were followed for 50 ± 15 months. Fifteen patients had coronary events during this period (1 cardiac death, 2 non-fatal myocardial infarctions, 3 acute heart failures, and 9 unstable anginas. On Kaplan-Meier analysis, only FMD and stress ECG were significant predictors for cardiac events. Conclusion Brachial endothelial function as reflected by FMD and conventional exercise stress testing has comparable prognostic value, whereas carotid artery plaque burden appears to be less powerful for predicting future cardiac events.

  8. Regulation of endothelial barrier function during flow-induced conversion to an arterial phenotype.

    Science.gov (United States)

    Seebach, Jochen; Donnert, Gerald; Kronstein, Romy; Werth, Sebastian; Wojciak-Stothard, Beata; Falzarano, Darryl; Mrowietz, Christof; Hell, Stefan W; Schnittler, Hans-J

    2007-08-01

    Flow-induced conversion of endothelial cells into an elongated arterial phenotype requires a coordinated regulation of cell junctions. Here we investigated the effect of acute and chronic flow on junction regulation. Using an extended experimental setup that allows analyses of endothelial barrier function under flow conditions, we found a flow-induced upregulation of the transendothelial electrical resistance within minutes. This was accompanied by an increase in actin filaments along the junctions and vascular endothelial (VE)-cadherin clustering, which was identified at nanoscale resolution by stimulated emission depletion microscopy. In addition, a transient tyrosine phosphorylation of VE-cadherin and catenins occurred within minutes following the onset of flow. VE-cadherin and actin distribution were maintained under chronic flow over 24 h and associated with the upregulation of VE-cadherin and alpha-catenin expression, thus compensating for the cell elongation-mediated increase in cell border length. Importantly, all observed effects were rac1 dependent as verified by the inhibitory effect of dominant negative N17rac1. These results show that flow-induced conversion of endothelial cells into an arterial phenotype occurs while intercellular junctions remain intact. The data place rac1 in a central multimodal regulatory position that might be important in the development of vascular diseases, such as arteriosclerosis.

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

    Directory of Open Access Journals (Sweden)

    Lucas Felipe de Oliveira

    2015-01-01

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

  10. Retrograde flow and shear rate acutely impair endothelial function in humans.

    Science.gov (United States)

    Thijssen, Dick H J; Dawson, Ellen A; Tinken, Toni M; Cable, N Timothy; Green, Daniel J

    2009-06-01

    Changes in arterial shear stress induce functional and structural vasculature adaptations. Recent studies indicate that substantial retrograde flow and shear can occur through human conduit arteries. In animals, retrograde shear is associated with atherogenic effects. The aim of this study was to examine the impact of incremental levels of retrograde shear on endothelial function in vivo. On 3 separate days, we examined bilateral brachial artery flow-mediated dilation, an index of NO-mediated endothelial function, in healthy men (24+/-3 years) before and after a 30-minute intervention consisting of cuff inflation to 25, 50, or 75 mm Hg. Cuff inflations resulted in "dose"-dependent increases in retrograde shear rate, compared with the noncuffed arm, within subjects (P<0.001). Flow-mediated dilation in the cuffed arm did not change in response to the 25-mm Hg stimulus but decreased significantly after both the 50- and 75-mm Hg interventions (P<0.05). The decrease in flow-mediated dilation after the 75-mm Hg intervention was significantly larger than that observed after a 50-mm Hg intervention (P=0.03). In the noncuffed arm, no changes in shear rate or flow-mediated dilation were observed. These results demonstrate that an increase in retrograde shear rate induces a dose-dependent attenuation of endothelial function in humans. This finding contributes to our understanding regarding the possible detrimental effects of retrograde shear rate in vivo.

  11. Sphingosine-1-Phosphate Signaling Regulates Myogenic Responsiveness in Human Resistance Arteries.

    Directory of Open Access Journals (Sweden)

    Sonya Hui

    Full Text Available We recently identified sphingosine-1-phosphate (S1P signaling and the cystic fibrosis transmembrane conductance regulator (CFTR as prominent regulators of myogenic responsiveness in rodent resistance arteries. However, since rodent models frequently exhibit limitations with respect to human applicability, translation is necessary to validate the relevance of this signaling network for clinical application. We therefore investigated the significance of these regulatory elements in human mesenteric and skeletal muscle resistance arteries. Mesenteric and skeletal muscle resistance arteries were isolated from patient tissue specimens collected during colonic or cardiac bypass surgery. Pressure myography assessments confirmed endothelial integrity, as well as stable phenylephrine and myogenic responses. Both human mesenteric and skeletal muscle resistance arteries (i express critical S1P signaling elements, (ii constrict in response to S1P and (iii lose myogenic responsiveness following S1P receptor antagonism (JTE013. However, while human mesenteric arteries express CFTR, human skeletal muscle resistance arteries do not express detectable levels of CFTR protein. Consequently, modulating CFTR activity enhances myogenic responsiveness only in human mesenteric resistance arteries. We conclude that human mesenteric and skeletal muscle resistance arteries are a reliable and consistent model for translational studies. We demonstrate that the core elements of an S1P-dependent signaling network translate to human mesenteric resistance arteries. Clear species and vascular bed variations are evident, reinforcing the critical need for further translational study.

  12. 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.

  13. 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.

  14. Endothelial NLRP3 inflammasome activation and arterial neointima formation associated with acid sphingomyelinase during hypercholesterolemia

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    Saisudha Koka

    2017-10-01

    Full Text Available The NLRP3 inflammasome has been reported to be activated by atherogenic factors, whereby endothelial injury and consequent atherosclerotic lesions are triggered in the arterial wall. However, the mechanisms activating and regulating NLRP3 inflammasomes remain poorly understood. The present study tested whether acid sphingomyelinase (ASM and ceramide associated membrane raft (MR signaling platforms contribute to the activation of NLRP3 inflammasomes and atherosclerotic lesions during hypercholesterolemia. We found that 7-ketocholesterol (7-Keto or cholesterol crystal (ChC markedly increased the formation and activation of NLRP3 inflammasomes in mouse carotid arterial endothelial cells (CAECs, as shown by increased colocalization of NLRP3 with ASC or caspase-1, enhanced caspase-1 activity and elevated IL-1β levels, which were markedly attenuated by mouse Asm siRNA, ASM inhibitor- amitriptyline, and deletion of mouse Asm gene. In CAECs with NLRP3 inflammasome formation, membrane raft (MR clustering with NADPH oxidase subunits was found remarkably increased as shown by CTXB (MR marker and gp91phox aggregation indicating the formation of MR redox signaling platforms. This MR clustering was blocked by MR disruptor (MCD, ROS scavenger (Tempol and TXNIP inhibitor (verapamil, accompanied by attenuation of 7-Keto or ChC-induced increase in caspase-1 activity. In animal experiments, Western diet fed mice with partially ligated left carotid artery (PLCA were found to have significantly increased neointimal formation, which was associated with increased NLRP3 inflammasome formation and IL-1β production in the intima of Asm+/+ mice but not in Asm-/- mice. These results suggest that Asm gene and ceramide associated MR clustering are essential to endothelial inflammasome activation and dysfunction in the carotid arteries, ultimately determining the extent of atherosclerotic lesions.

  15. Endothelial dysfunction in obstructive sleep apnea measured by peripheral arterial tone response in the finger to reactive hyperemia.

    Science.gov (United States)

    Itzhaki, Sarah; Lavie, Lena; Pillar, Giora; Tal, Galit; Lavie, Peretz

    2005-05-01

    The aim of this study was to investigate endothelial functioning in sleep apnea patients using a novel plethysmographic device that monitors peripheral arterial tone response in the finger to reactive hyperemia induced by forearm ischemia. Forty-six sleep apnea patients, 74.0% men, mean age 46.8 +/- 9.3 years, and 17 control subjects without sleep apnea, 64.7% men, mean age 47.1 +/- 6.7 years. Eight-bed Technion Sleep Medicine Center in Haifa, Israel. Endothelial functioning assessed by the reactive hyperemia peripheral arterial tone index was measured twice, before sleep and after waking from sleep monitored by polysomnography in the laboratory. The reactive hyperemia peripheral arterial tone index was calculated as the average amplitude of the peripheral arterial tone signal after the cuff deflation divided by the average amplitude before the cuff inflation. Morning index of endothelial functioning was significantly lower in patients with moderate to severe sleep apnea (apnea-hypopnea index > or = 30) than in patients with mild sleep apnea (30 disease had significantly lower morning and evening indexes of endothelial functioning than patients without such a history. Multivariate analysis revealed that apnea-hypopnea index and sleep efficiency were significant predictors of the morning index. Measurements of the response of the peripheral arterial tone in the finger to reactive hyperemia can be used as a substitute for the brachial artery ultrasound technique to measure endothelial functioning in patients with sleep apnea.

  16. Twenty-four-hour exposure to altered blood flow modifies endothelial Ca2+-activated K+ channels in rat mesenteric arteries

    DEFF Research Database (Denmark)

    Hilgers, Rob H P; Janssen, Ger M J; Fazzi, Gregorio E

    2010-01-01

    remodeling. In rats, mesenteric arteries were exposed to increased [+90%, high flow (HF)] or reduced blood flow [-90%, low flow (LF)] and analyzed 24 h later. There were no detectable changes in arterial structure or in expression level of endothelial nitric-oxide synthase, SK3, or IK1. Arterial relaxing......We tested the hypothesis that changes in arterial blood flow modify the function of endothelial Ca2+-activated K+ channels [calcium-activated K+ channel (K(Ca)), small-conductance calcium-activated K+ channel (SK3), and intermediate calcium-activated K+ channel (IK1)] before arterial structural...... arteries, the balance between the NO/prostanoid versus EDHF response was unaltered. However, the contribution of IK1 to the EDHF response was enhanced, as indicated by a larger effect of TRAM-34 and a larger residual NS309-induced relaxation in the presence of UCL 1684. Reduction of blood flow selectively...

  17. Induced cytoskeletal changes in bovine pulmonary artery endothelial cells by resveratrol and the accompanying modified responses to arterial shear stress

    Directory of Open Access Journals (Sweden)

    Lerea Kenneth M

    2001-01-01

    Full Text Available Abstract Background Atherosclerosis and coronary heart disease (CHD are significant contributors to morbidity and mortality in developed countries. A noted exception is the low mortality of CHD in France, particularly the southwest region. This phenomenon, commonly referred to as the French paradox, may be associated with high consumption of red wine. We investigate whether the cardioprotective activity of red wine may involve the grape skin-derived polyphenol, resveratrol. We further test the possibility that resveratrol acts by modulating structural and functional changes in endothelial cells lining the blood vessel wall. Results Bovine pulmonary artery endothelial cells (BPAEC were incubated with resveratrol, with and without concurrent exposure to simulated arterial shear stress. Resveratrol significantly affected proliferation and shape of BPAEC; growth was suppressed and cells became elongated, based on morphologic analysis of rhodamine-conjugated phalloidin stained F-actin by confocal microscopy. Using selective signaling inhibitors, we showed that the resveratrol-induced cellular phenotype was dependent on intracellular calcium and tyrosine kinase activities, and assembly of actin microfilaments and microtubules, but was unrelated to PKC activity. Exposure to simulated arterial flow revealed that, whereas controls cells easily detached from the culture support in a time-dependent manner, resulting in total cell loss after a 5 min challenge with simulated arterial flow conditions, a significant percentage of the treated cells remained attached to the cultured plastic coverslips under identical experimental conditions, suggesting that they adhered more strongly to the surface. Western blot analysis shows that whereas cells treated with 25 μM and 100 μM resveratrol had no change in total ERK1/2, treatment did result in an increase in phosphorylated ERK1/2, which probably involved stabilization of the active enzyme. An increase in nitric

  18. Flow-mediated dilation and peripheral arterial tonometry are disturbed in preeclampsia and reflect different aspects of endothelial function

    OpenAIRE

    Mannaerts, Dominique; Faes, Ellen; Goovaerts, Inge; Stoop, Tibor; Cornette, Jerome; Gyselaers, Wilfried; Spaanderman, Marc; Craenenbroeck, van, Emeline M.; Jacquemyn, Yves

    2017-01-01

    Abstract: Endothelial function and arterial stiffness are known to be altered in preeclamptic pregnancies. Previous studies have shown conflicting results regarding the best technique for assessing vascular function in pregnancy. In this study, we made a comprehensive evaluation of in vivo vascular function [including flow-mediated dilatation (FMD), peripheral arterial tonometry (PAT), and arterial stiffness] in preeclamptic patients and compared them with normal pregnancies. In addition, we ...

  19. HABITUAL FLAVONOID INTAKE AND ENDOTHELIAL FUNCTION IN HEALTHY HUMANS

    Science.gov (United States)

    Fisher, Naomi DL; Hurwitz, Shelley; Hollenberg, Norman K

    2013-01-01

    Objective Endothelial function, as measured by non-invasive techniques, is known to vary widely within populations. Our study was designed to test the hypothesis that this variation is determined in large part by a person’s habitual dietary intake of flavonoids. Methods This was an analytical study examining the relationship between endothelial function and dietary flavonoids in 19 healthy older adults (mean age 72 years). The study took place in the inpatient Clinical Research Center of the Brigham and Women’s Hospital. Habitual flavonoid intake was assessed via a focused food frequency questionnaire. Endothelial function, measured as the reactive hyperemia response to one dose of flavonoid-rich cocoa, was recorded with a plethysmograpic device via peripheral arterial tonometry (PAT). Results Background flavonoid intake and the reactive hyperemia (RH)-PAT response were significantly correlated (r=0.7, p=0.001); subjects with higher habitual flavonoid intake showed a significantly greater RH-PAT response than did lower consumers. PAT response to cocoa was also significantly correlated with simultaneous flavanol concentration in the blood (r=0.5, p=0.03). Conclusion Individual variation in endothelial function among healthy older people, measured as PAT response to flavonoid-rich cocoa, is highly dependent upon usual daily flavonoid consumption. These data raise the possibility that the consumption of fruits and vegetables dictates basal endothelial function, likely related to their flavonoid content and influence on nitric oxide. PMID:23378455

  20. 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.

  1. Evaluation of endothelial function by VOP and inflammatory biomarkers in patients with arterial hypertension.

    Science.gov (United States)

    Junqueira, Camillo L C; Magalhães, Maria Eliane C; Brandão, Andréa Araújo; Ferreira, Esmeralci; Junqueira, Adriana S M; Neto, José Firmino N; Souza, Maria das Graças C; Bottino, Daniel Alexandre; Bouskela, Eliete

    2018-02-01

    Hypertension is associated with microcirculatory impairment. Our objectives were to evaluate endothelial function and inflammatory biomarkers in patients with resistant (RH) and mild to moderate (MMH) arterial hypertension in comparison to normotensives (control group-CG). Three groups, 25 patients each, have been investigated, by anamnesis, venous occlusion plethysmography (VOP) and serum determination of adhesion molecules (VCAM, ICAM), adiponectin, endothelin and C-reactive protein (CRP). Patients not using statins and with or without blood pressure control were also analyzed. RH group showed smaller percentage increase of maximum forearm blood flow (FBF) (endothelial-dependent vasodilatation) than controls (p < 0.05), but no significant difference could be detected between MMH and CG groups on maximum FBF and minimum vascular resistance post-ischemia. RH and MMH groups showed higher resistance averages compared to controls (p < 0.05). Uncontrolled BP in hypertensive patients showed worse results for blood flow and resistance. Endothelial-independent vasodilatation was not affected. Endothelin levels were higher in RH and MMH groups (p < 0.05) not using statins. CRP was significantly higher only in RH compared to CG (p < 0.05). In conclusion patients with severe hypertension and lack of blood pressure control showed greater impairment of endothelial function with higher CRP and endothelin levels.

  2. Release of a leukocyte activation inhibitor by staurosporine-treated pulmonary artery endothelial cells.

    Science.gov (United States)

    Chen, X; Catravas, J D

    1998-07-01

    Bovine pulmonary arterial endothelial cells (BPAE) treated with the protein kinase C (PKC) inhibitor staurosporine inhibited O-2. generation by neutrophils exposed to phorbol myristate acetate (PMA) but did not affect O-2. generated enzymatically by xanthine/xanthine oxidase (X/XO). Similar results were obtained with conditioned medium from staurosporine-pretreated BPAE. The inhibitory effects of staurosporine-treated BPAE on O-2. generation were not altered by the superoxide dismutase inhibitor diethylcarbamazine. This BPAE-derived inhibitor was continuously released from staurosporine-pretreated BPAE for at least 5 h. The exact nature of the inhibitor remains unknown, but it appears to be a positively charged molecule with molecular weight neutrophils with staurosporine or conditioned medium from staurosporine-treated BPAE prevented the neutrophil-mediated decrease in endothelium-bound angiotensin-converting enzyme activity and cytotoxicity in BPAE. In contrast, staurosporine potentiated the H2O2- and X/XO-mediated endothelial cytotoxicity. These data suggest that staurosporine-treated endothelial cells release a soluble factor that inhibits neutrophil activation and protects endothelial cells from neutrophil-mediated injury.

  3. Nilotinib Does Not Alter the Secretory Functions of Carotid Artery Endothelial Cells in a Prothrombotic or Antithrombotic Fashion.

    Science.gov (United States)

    Katgı, Abdullah; Sevindik, Ömür Gökmen; Gökbulut, Aysun Adan; Özsan, Güner Hayri; Yüksel, Faize; Solmaz, Şerife Medeni; Alacacıoğlu, İnci; Özcan, Mehmet Ali; Demirkan, Fatih; Baran, Yusuf; Pişkin, Özden

    2015-10-01

    There have been concerns about the possible prothrombotic effects of nilotinib, especially in patients having cardiovascular risk factors. The potential mechanism behind the increased risk of thromboembolic events is still not clear. In this study, we aimed to evaluate possible harmful effects of nilotinib on endothelial cells. To this aim, we examined proliferative capacity and secretory functions of healthy human carotid artery endothelial cells (HCtAECs) in response to nilotinib. 3-(4,5-Dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation method was used to determine antiproliferative effects of nilotinib on HCtAECs. The HCtAECs were incubated with 5, 10, and 100 nmol/L doses of nilotinib for 72 hours. Then, in order to assess the endothelial function, levels of nitric oxide (NO), von Willebrand factor (vWF), tissue plasminogen activator, plasminogen activator inhibitor 1 (PAI-1), and endothelin 1 (ET-1) were evaluated using enzyme-linked immunosorbent assay from tissue culture supernatants. There were slight but statistically significant decreases in cell proliferation in response to nilotinib. Nilotinib increased the secretion of t-PA, PAI-1, and vWF in a dose-dependent manner when compared with the untreated control group. The ET-1 secretion was lower in 5 nmol/L and higher in 10 and 100 nmol/L nilotinib-treated cells as compared to untreated cells. Regarding NO secretion, lower levels were observed in 5 and 10 nmol/L, and higher levels were detected in 100 nmol/L nilotinib-treated cells as compared to untreated control group cells. Considering the results obtained in our study, nilotinib does not affect the functions of endothelial cells either in a prothrombotic or an antithrombotic fashion, despite a dose-dependent decline in cell viability. © The Author(s) 2014.

  4. Intravascular Papillary Endothelial Hyperplasia (Masson’s Tumor) of the Radial Artery: A Case Report

    Energy Technology Data Exchange (ETDEWEB)

    Stark, Christopher, E-mail: Christopher.stark@uvmhealth.org [University of Vermont Medical Center, Department of Radiology (United States); Olsen, Daniel [Mayo Clinic, Department of Pathology (United States); Morris, Christopher [University of Vermont Medical Center, Department of Radiology (United States); Bertges, Daniel [University of Vermont Medical Center, Department of Surgery (United States); Najarian, Kenneth [University of Vermont Medical Center, Department of Radiology (United States)

    2016-11-15

    Intravascular papillary endothelial hyperplasia (IPEH), often referred to as Masson’s tumor, is a benign non-neoplastic vascular lesion of the skin and subcutaneous tissues. Although it is rare, knowledge of the existence of IPEH is important as it can mimic other benign and malignant tumors, most notably angiosarcoma. IPEH remains an incompletely understood entity; however, most consider it to be the result of reactive endothelial proliferation following thrombus formation within a vessel, vascular malformation, or adjacent to a vessel. In this article, we report a case of IPEH arising within an arteriovenous malformation of the radial artery and present accompanying multimodality imaging and pathology figures. We will also describe the clinical presentation, pathophysiology, histology, imaging features, and management of IPEH.

  5. Assessment of endothelial function by brachial artery flow mediated dilatation in microvascular disease

    Directory of Open Access Journals (Sweden)

    Naidu Otikunta

    2011-12-01

    Full Text Available Abstract Background Cardiac syndrome X is an important therapeutic and diagnostic challenge to physician. Study of Csx patients may help to understand the pathophysiology of coronary microcirculation and to gain an insight on the management of these group patients. Methods We measured the flow mediated dilation of the brachial artery both endothelium dependent and independent vasodilatation by high resolution ultrasound in 30 cardiac syndrome X patients and matched with 30 healthy control subjects. Results Significantly decreased flow mediated dilatation was observed in patients when compared to control (9.42 ± 7.20 vs 21.11 ± 9.16 p 11.11(p Conclusions The study suggests impairment of endothelial function in cardiac syndrome X patients. Increased Systolic blood pressure and body mass index may increase the risk of impairment of endothelial function in this group of patients.

  6. Endothelial Mineralocorticoid Receptor Mediates Parenchymal Arteriole and Posterior Cerebral Artery Remodeling During Angiotensin II-Induced Hypertension.

    Science.gov (United States)

    Diaz-Otero, Janice M; Fisher, Courtney; Downs, Kelsey; Moss, M Elizabeth; Jaffe, Iris Z; Jackson, William F; Dorrance, Anne M

    2017-12-01

    The brain is highly susceptible to injury caused by hypertension because the increased blood pressure causes artery remodeling that can limit cerebral perfusion. Mineralocorticoid receptor (MR) antagonism prevents hypertensive cerebral artery remodeling, but the vascular cell types involved have not been defined. In the periphery, the endothelial MR mediates hypertension-induced vascular injury, but cerebral and peripheral arteries are anatomically distinct; thus, these findings cannot be extrapolated to the brain. The parenchymal arterioles determine cerebrovascular resistance. Determining the effects of hypertension and MR signaling on these arterioles could lead to a better understanding of cerebral small vessel disease. We hypothesized that endothelial MR signaling mediates inward cerebral artery remodeling and reduced cerebral perfusion during angiotensin II (AngII) hypertension. The biomechanics of the parenchymal arterioles and posterior cerebral arteries were studied in male C57Bl/6 and endothelial cell-specific MR knockout mice and their appropriate controls using pressure myography. AngII increased plasma aldosterone and decreased cerebral perfusion in C57Bl/6 and MR-intact littermates. Endothelial cell MR deletion improved cerebral perfusion in AngII-treated mice. AngII hypertension resulted in inward hypotrophic remodeling; this was prevented by MR antagonism and endothelial MR deletion. Our studies suggest that endothelial cell MR mediates hypertensive remodeling in the cerebral microcirculation and large pial arteries. AngII-induced inward remodeling of cerebral arteries and arterioles was associated with a reduction in cerebral perfusion that could worsen the outcome of stroke or contribute to vascular dementia. © 2017 American Heart Association, Inc.

  7. Effects of glucagon-like peptide-1 on endothelial function in type 2 diabetes patients with stable coronary artery disease

    DEFF Research Database (Denmark)

    Nyström, Thomas; Gutniak, Mark K; Zhang, Qimin

    2004-01-01

    (I)) in two groups: 1) 12 type 2 diabetes patients with stable coronary artery disease and 2) 10 healthy subjects with normal endothelial function and S(I). Subjects underwent infusion of recombinant GLP-1 or saline in a random crossover study. Endothelial function was measured by postischemic FMD of brachial...... artery, using ultrasonography. S(I) [in (10(-4) dl.kg(-1).min(-1))/(muU/ml)] was measured by hyperinsulinemic isoglycemic clamp technique. In type 2 diabetic subjects, GLP-1 infusion significantly increased relative changes in brachial artery diameter from baseline FMD(%) (3.1 +/- 0.6 vs. 6.6 +/- 1.0%, P...... FMD(%) (11.9 +/- 0.9 vs. 10.3 +/- 1.0%, P = NS) nor S(I) (14.8 +/- 1.8 vs. 11.6 +/- 2.0, P = NS). We conclude that GLP-1 improves endothelial dysfunction...

  8. 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.

  9. Short- and long-term black tea consumption reverses endothelial dysfunction in patients with coronary artery disease.

    Science.gov (United States)

    Duffy, S J; Keaney, J F; Holbrook, M; Gokce, N; Swerdloff, P L; Frei, B; Vita, J A

    2001-07-10

    Epidemiological studies suggest that tea consumption decreases cardiovascular risk, but the mechanisms of benefit remain undefined. Endothelial dysfunction has been associated with coronary artery disease and increased oxidative stress. Some antioxidants have been shown to reverse endothelial dysfunction, and tea contains antioxidant flavonoids. Methods and Results-- To test the hypothesis that tea consumption will reverse endothelial dysfunction, we randomized 66 patients with proven coronary artery disease to consume black tea and water in a crossover design. Short-term effects were examined 2 hours after consumption of 450 mL tea or water. Long-term effects were examined after consumption of 900 mL tea or water daily for 4 weeks. Vasomotor function of the brachial artery was examined at baseline and after each intervention with vascular ultrasound. Fifty patients completed the protocol and had technically suitable ultrasound measurements. Both short- and long-term tea consumption improved endothelium- dependent flow-mediated dilation of the brachial artery, whereas consumption of water had no effect (Pcaffeine (200 mg) had no short-term effect on flow-mediated dilation. Plasma flavonoids increased after short- and long-term tea consumption. Short- and long-term black tea consumption reverses endothelial vasomotor dysfunction in patients with coronary artery disease. This finding may partly explain the association between tea intake and decreased cardiovascular disease events.

  10. Interleukin-33 induces urokinase in human endothelial cells--possible impact on angiogenesis.

    Science.gov (United States)

    Stojkovic, S; Kaun, C; Heinz, M; Krychtiuk, K A; Rauscher, S; Lemberger, C E; de Martin, R; Gröger, M; Petzelbauer, P; Huk, I; Huber, K; Wojta, J; Demyanets, S

    2014-06-01

    Urokinase-type plasminogen activator (u-PA) plays a pivotal role in extracellular proteolysis and is thought to be critically involved in the modulation of angiogenesis. Interleukin (IL)-33 is a member of the IL-1 cytokine family, which is thought to act as danger signal that is released from cells after injury. IL-33 is involved in the pathogenesis of various inflammatory diseases and previously was shown to induce angiogenesis and inflammatory activation of endothelial cells. We investigated the impact of IL-33 on u-PA in endothelial cells as a new possible function for IL-33. We could demonstrate that IL-33 upregulated u-PA mRNA expression and protein production in human coronary artery and human umbilical vein endothelial cells in a time- and concentration-dependent manner via interaction with its receptor ST2 and activation of the nuclear factor-κB pathway but independent of autocrine IL-1-induced effects. The hydroxymethylglutaryl-coenzyme A reductase inhibitor simvastatin abrogated the IL-33-induced increase in u-PA, thus providing further evidence for pleiotropic effects of statins. IL-33 induced u-PA-dependent capillary-like tube formation and vessel sprouting. In human carotid atherosclerotic plaques (n = 16), u-PA mRNA positively correlated with IL-33 mRNA expression (r = 0.780, P < 0.001). Furthermore, IL-33 and u-PA protein were detected in endothelial cells in these samples using fluorescence immunohistochemistry. We hypothesize that IL-33, representing a danger signal that is released after tissue damage, in addition to its role in the inflammatory activation of endothelial cells, is involved in u-PA-driven angiogenesis, a process that has been shown before to be linked to inflammation in various pathologies. © 2014 International Society on Thrombosis and Haemostasis.

  11. Release of soluble vascular endothelial growth factor receptor-1 (sFlt-1 during coronary artery bypass surgery

    Directory of Open Access Journals (Sweden)

    Orsel Isabelle

    2007-09-01

    Full Text Available Abstract Background This study was conducted to follow plasma concentrations of sFlt-1 and sKDR, two soluble forms of the vascular endothelial growth factor (VEGF receptor in patients undergoing coronary artery bypass graft (CABG surgery with extracorporeal circulation (ECC. Methods Plasma samples were obtained before, during and after surgery in 15 patients scheduled to undergo CABG. Levels of sFlt-1 and KDR levels were investigated using specific ELISA. Results A 75-fold increase of sFlt-1 was found during cardiac surgery, sFlt-1 levels returning to pre-operative values at the 6th post-operative hour. In contrast sKDR levels did not change during surgery. The ECC-derived sFlt-1 was functional as judge by its inhibitory effect on the VEGF mitogenic response in human umbilical vein endothelial cells (HUVECs. Kinetic experiments revealed sFlt-1 release immediately after the beginning of ECC suggesting a proteolysis of its membrane form (mFlt-1 rather than an elevated transcription/translation process. Flow cytometry analysis highlighted no effect of ECC on the shedding of mFlt-1 on platelets and leukocytes suggesting vascular endothelial cell as a putative cell source for the ECC-derived sFlt-1. Conclusion sFlt-1 is released during CABG with ECC. It might be suggested that sFlt-1 production, by neutralizing VEGF and/or by inactivating membrane-bound Flt-1 and KDR receptors, might play a role in the occurrence of post-CABG complication.

  12. 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.

  13. 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

  14. Endothelial and lipoprotein lipases in human and mouse placenta

    DEFF Research Database (Denmark)

    Lindegaard, Marie L S; Olivecrona, Gunilla; Christoffersen, Christina

    2005-01-01

    Placenta expresses various lipase activities. However, a detailed characterization of the involved genes and proteins is lacking. In this study, we compared the expression of endothelial lipase (EL) and LPL in human term placenta. When placental protein extracts were separated by heparin-Sepharos...

  15. PDGF-BB/KLF4/VEGF Signaling Axis in Pulmonary Artery Endothelial Cell Angiogenesis.

    Science.gov (United States)

    Liang, Songhe; Yu, Hao; Chen, Xinxin; Shen, Tingting; Cui, Zhongqi; Si, Genle; Zhang, JunTing; Cheng, Yue; Jia, Shiwei; Song, Shasha; Zhang, Xiang; Yu, Xiufeng

    2017-01-01

    Accumulating evidence suggests that platelet-derived growth factor-BB (PDGF-BB) and vascular endothelial growth factor(VEGF) play a role in the progression of pulmonary arterial hypertension (PAH).Since chronic hypoxia is responsible for intimal hyperplasia and disordered angiogenesis of pulmonary arteries, which are histological hallmarks of PAH, we explored the role of the PDGF-BB/KLF4/VEGF signaling axis in the angiogenesis of pulmonary artery endothelial cells (PAECs). Adult male Wistar rats were used to study hypoxia-induced or monocrotaline (MCT)-induced right ventricular (RV) remodeling as well as systolic function and hemodynamics using echocardiography and a pressure-volume admittance catheter. Morphometric analyses of lung vasculature and RV vessels were performed. The results revealed that both the PDGF receptor-tyrosine kinase inhibitor imatinib and the multi-targeted VEGF and PDGF receptor inhibit or sunitinib malate reversed hypoxia-induced increases in right ventricular systolic pressure (RVSP), right ventricular function and thickening of the medial walls. Mechanistically VEGF/VEGFR and PDGF/PDGFR formed a biological complex. We also showed that PDGF-BBincreasedKLF4 promoter activity transcriptionally activating VEGF expression, which regulates PAEC proliferation; migration; and the cell-cycle transition from G0/G1phase to S phase and G2/M-phase and eventually leads to PAEC angiogenesis Conclusion: Our study indicates that hypoxia-induced angiogenesis of PAECs is associated with increased levels of PDGF-BB/KLF4/VEGF, which contribute to pulmonary vascular remodeling. Overall, our study contributes to a better understanding of PAH pathogenesis. © 2017 The Author(s). Published by S. Karger AG, Basel.

  16. Engineered Microvasculature in PDMS Networks Using Endothelial Cells Derived from Human Induced Pluripotent Stem Cells

    Science.gov (United States)

    Sivarapatna, Amogh; Ghaedi, Mahboobe; Xiao, Yang; Han, Edward; Aryal, Binod; Zhou, Jing; Fernandez-Hernando, Carlos; Qyang, Yibing; Hirschi, Karen K.

    2017-01-01

    In this study, we used a polydimethylsiloxane (PDMS)-based platform for the generation of intact, perfusion-competent microvascular networks in vitro. COMSOL Multiphysics, a finite-element analysis and simulation software package, was used to obtain simulated velocity, pressure, and shear stress profiles. Transgene-free human induced pluripotent stem cells (hiPSCs) were differentiated into partially arterialized endothelial cells (hiPSC-ECs) in 5 d under completely chemically defined conditions, using the small molecule glycogen synthase kinase 3β inhibitor CHIR99021 and were thoroughly characterized for functionality and arterial-like marker expression. These cells, along with primary human umbilical vein endothelial cells (HUVECs), were seeded in the PDMS system to generate microvascular networks that were subjected to shear stress. Engineered microvessels had patent lumens and expressed VE-cadherin along their periphery. Shear stress caused by flowing medium increased the secretion of nitric oxide and caused endothelial cells s to align and to redistribute actin filaments parallel to the direction of the laminar flow. Shear stress also caused significant increases in gene expression for arterial markers Notch1 and EphrinB2 as well as antithrombotic markers Kruppel-like factor 2 (KLF-2)/4. These changes in response to shear stress in the microvascular platform were observed in hiPSC-EC microvessels but not in microvessels that were derived from HUVECs, which indicated that hiPSC-ECs may be more plastic in modulating their phenotype under flow than are HUVECs. Taken together, we demonstrate the feasibly of generating intact, engineered microvessels in vitro, which replicate some of the key biological features of native microvessels. PMID:28901188

  17. Flow-mediated dilation and peripheral arterial tonometry are disturbed in preeclampsia and reflect different aspects of endothelial function.

    Science.gov (United States)

    Mannaerts, Dominique; Faes, Ellen; Goovaerts, Inge; Stoop, Tibor; Cornette, Jerome; Gyselaers, Wilfried; Spaanderman, Marc; Van Craenenbroeck, Emeline M; Jacquemyn, Yves

    2017-11-01

    Endothelial function and arterial stiffness are known to be altered in preeclamptic pregnancies. Previous studies have shown conflicting results regarding the best technique for assessing vascular function in pregnancy. In this study, we made a comprehensive evaluation of in vivo vascular function [including flow-mediated dilatation (FMD), peripheral arterial tonometry (PAT), and arterial stiffness] in preeclamptic patients and compared them with normal pregnancies. In addition, we assessed the relation between vascular function and systemic inflammation. Fourteen patients with preeclampsia (PE) and 14 healthy pregnant controls were included. Endothelial function was determined by FMD and PAT and arterial stiffness by carotid-femoral pulse-wave velocity and augmentation index. Systemic inflammation was assessed using mean platelet volume (MPV) and neutrophil-lymphocyte ratio (NLR). The reactive hyperemia index, assessed using PAT, is decreased at the third trimester compared with the first trimester in a normal, uncomplicated pregnancy (P = 0.001). Arterial stiffness is significantly higher in PE versus normal pregnancy (P function, obtained by FMD, is deteriorated in PE versus normal pregnancy (P = 0.015), whereas endothelial function assessment by PAT is improved in PE versus normal pregnancy (P = 0.001). Systemic inflammation (MPV and NLR) increases during normal pregnancy. FMD and PAT are disturbed in PE. Endothelial function, assessed by FMD and PAT, shows distinct results. This may indicate that measurements with FMD and PAT reflect different aspects of endothelial function and that PAT should not be used as a substitute for FMD as a measure of endothelial function in pregnancy. Copyright © 2017 the American Physiological Society.

  18. Endothelial progenitor cells from human fetal aorta cure diabetic foot in a rat model.

    Science.gov (United States)

    Zhao, Wan-Ni; Xu, Shi-Qing; Liang, Jian-Feng; Peng, Liang; Liu, Hong-Lin; Wang, Zai; Fang, Qing; Wang, Meng; Yin, Wei-Qin; Zhang, Wen-Jian; Lou, Jin-Ning

    2016-12-01

    Recent evidence has suggested that circulating endothelial progenitor cells (EPCs) can repair the arterial endothelium during vascular injury. However, a reliable source of human EPCs is needed for therapeutic applications. In this study, we isolated human fetal aorta (HFA)-derived EPCs and analyzed the capacity of EPCs to differentiate into endothelial cells. In addition, because microvascular dysfunction is considered to be the major cause of diabetic foot (DF), we investigated whether transplantation of HFA-derived EPCs could treat DF in a rat model. EPCs were isolated from clinically aborted fetal aorta. RT-PCR, fluorescence-activated cell sorting, immunofluorescence, and an enzyme-linked immunosorbent assay were used to examine the expressions of CD133, CD34, CD31, Vascular Endothelial Growth Factor Receptor 2 (VEGFR2), von Willebrand Factor (vWF), and Endothelial Leukocyte Adhesion Molecule-1 (ELAM-1). Morphology and Dil-uptake were used to assess function of the EPCs. We then established a DF model by injecting microcarriers into the hind-limb arteries of Goto-Kakizaki rats and then transplanting the cultured EPCs into the ischemic hind limbs. Thermal infrared imaging, oxygen saturation apparatus, and laser Doppler perfusion imaging were used to monitor the progression of the disease. Immunohistochemistry was performed to examine the microvascular tissue formed by HFA-derived EPCs. We found that CD133, CD34, and VEGFR2 were expressed by HFA-derived EPCs. After VEGF induction, CD133 expression was significantly decreased, but expression levels of vWF and ELAM-1 were markedly increased. Furthermore, tube formation and Dil-uptake were improved after VEGF induction. These observations suggest that EPCs could differentiate into endothelial cells. In the DF model, temperature, blood flow, and oxygen saturation were reduced but recovered to a nearly normal level following injection of the EPCs in the hind limb. Ischemic symptoms also improved. Injected EPCs were

  19. ROLE OF NON INVASIVE STUDY OF BRACHIAL ARTERY FLOW MEDIATED VASODILATATION WITH CORRELATION TO ENDOTHELIAL DYSFUNCTION IN HYPERTENSIVE PATIENTS

    Directory of Open Access Journals (Sweden)

    Srikanth Bodepudi

    2016-09-01

    Full Text Available BACKGROUND The endothelium is an early target of cardiovascular diseases like hypertension and Diabetes mellitus (DM. In hypertension, endothelial dysfunction has been shown at the level of both resistance and conduit arteries. The fact that Forearm's Brachial artery endothelial dysfunction is a marker of future cardiovascular events in patients with hypertension stresses the importance of the clinical evaluation of endothelial function. MATERIALS AND METHODS This is a prospective study of hypertensive patients admitted in King George Hospital of Andhra Medical College, Visakhapatnam, India during May 2015 to August 2016 and diagnosed as per the criteria laid down by JNC 7 Classification. Patients of Diabetes Mellitus, Hyperthyroidism, Heart Failure, Coronary Artery Disease, Smokers and Peripheral Vascular Disease which are known to affect endothelial dysfunction were excluded from the study. RESULTS The study group comprised of 50 subjects and the control group had 30 subjects. There are 18 males (59.4% and 12 females (39.6% among Controls and 31 males (62% and 19 females (38% in the Cases group. In this study, the mean age among the controls is (55.6±13.63 and among the cases is (56.4±15.24. Mean FMD (Flow-mediated dilation among Cases is 8.15 and mean FMD among controls is 19.3. Mean Hyperemic Flow among Cases is 70.1 and among Controls is 121. 26% of the males have Endothelial Dysfunction and 9% of the females among cases have Endothelial Dysfunction. CONCLUSION FMD% is a diagnostic aid for evaluation of endothelial function. It is an experimental tool to measure endothelial dysfunction which is a fundamental basis for atherogenesis and CAD. It is a user friendly, non- invasive, cheap, reliable, reproducible technique for risk stratification of CAD.

  20. 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.

  1. 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.

  2. 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

  3. Resveratrol inhibits neointimal formation after arterial injury through an endothelial nitric oxide synthase-dependent mechanism.

    Science.gov (United States)

    Breen, Danna M; Dolinsky, Vernon W; Zhang, Hangjun; Ghanim, Husam; Guo, June; Mroziewicz, Margaret; Tsiani, Evangelia L; Bendeck, Michelle P; Dandona, Paresh; Dyck, Jason R B; Heximer, Scott P; Giacca, Adria

    2012-06-01

    Revascularization procedures used for treatment of atherosclerosis often result in restenosis. Resveratrol (RSV), an antioxidant with cardiovascular benefits, decreases neointimal formation after arterial injury by a mechanism that is still not fully clarified. Our main objective was to address the role of nitric oxide synthases (NOSes) and more specifically the endothelial-NOS (eNOS) isoform as a mediator of this effect. RSV (4 mg/kg/day, s.c.) alone or in combination with the NOS inhibitor N-nitro-L-arginine methyl ester (L-NAME) (2 mg/kg/day, s.c.) was given to Sprague-Dawley rats beginning at 3 days before arterial (carotid or aortic) injury. RSV reduced neointimal formation by 50% (P<0.01), decreased intimal cell proliferation by 37% (P<0.01) and reduced inflammatory markers such as PECAM and MMP-9 mRNA. These effects of RSV were all abolished by coadministration of l-NAME. Oral RSV (beginning at 5 days before arterial injury) reduced neointimal thickness after femoral wire injury in mice, however this effect was not observed in eNOS knockout mice. This is the first report of RSV decreasing neointimal cell proliferation and neointimal growth through an eNOS-dependent mechanism. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  4. Constitutive description of human femoropopliteal artery aging.

    Science.gov (United States)

    Kamenskiy, Alexey; Seas, Andreas; Deegan, Paul; Poulson, William; Anttila, Eric; Sim, Sylvie; Desyatova, Anastasia; MacTaggart, Jason

    2017-04-01

    Femoropopliteal artery (FPA) mechanics play a paramount role in pathophysiology and the artery's response to therapeutic interventions, but data on FPA mechanical properties are scarce. Our goal was to characterize human FPAs over a wide population to derive a constitutive description of FPA aging to be used for computational modeling. Fresh human FPA specimens ([Formula: see text]) were obtained from [Formula: see text] predominantly male (80 %) donors 54±15 years old (range 13-82 years). Morphometric characteristics including radius, wall thickness, opening angle, and longitudinal pre-stretch were recorded. Arteries were subjected to multi-ratio planar biaxial extension to determine constitutive parameters for an invariant-based model accounting for the passive contributions of ground substance, elastin, collagen, and smooth muscle. Nonparametric bootstrapping was used to determine unique sets of material parameters that were used to derive age-group-specific characteristics. Physiologic stress-stretch state was calculated to capture changes with aging. Morphometric and constitutive parameters were derived for seven age groups. Vessel radius, wall thickness, and circumferential opening angle increased with aging, while longitudinal pre-stretch decreased ([Formula: see text]). Age-group-specific constitutive parameters portrayed orthotropic FPA stiffening, especially in the longitudinal direction. Structural changes in artery wall elastin were associated with reduction of physiologic longitudinal and circumferential stretches and stresses with age. These data and the constitutive description of FPA aging shed new light on our understanding of peripheral arterial disease pathophysiology and arterial aging. Application of this knowledge might improve patient selection for specific treatment modalities in personalized, precision medicine algorithms and could assist in device development for treatment of peripheral artery disease.

  5. Lack of association between Chlamydia Pneumoniae serology and endothelial dysfunction of coronary arteries

    Directory of Open Access Journals (Sweden)

    Oehme Albrecht

    2005-04-01

    Full Text Available Abstract Background Recent publications brought up the hypothesis that an infection with Chlamydia Pneumoniae (CP might be a major cause of coronary artery disease (CAD. Therefore, we investigated whether endothelial dysfunction (ED as a precursor of atherosclerosis might be detectable in patients with previous infection with CP but without angiographic evidence of CAD. Methods We included 16 patients (6 male / 10 female of 52 consecutive patients with normal coronary angiography who had typical angina pectoris and pathologic findings in the stress test. Exclusion criteria were: active smoker, elevated cholesterol, hypertension, age > 65 years, diabetes mellitus, treatment with ACE-inhibitors, or known CAD. Blood sample analysis for serum titer against CP (aCP-IgG was performed after coronary angiography. We looked for endothelial dysfunction analyzing the diameter of the left anterior descending coronary artery (LAD before and after acetylcholine (ACh i. c. Quantitative analysis of luminal diameter (LD was performed in at least two planes during baseline conditions and after ACh for 2 minutes in dosages of 7.2 μg/min and 36 μg/min with an infusion speed of 2 ml/min. Using Doppler guide wire, the coronary flow velocity was measured continuously in the LAD. The coronary flow velocity reserve (CFVR was measured after 20 μg adenosine i. c. Results 10 patients had an elevated aCP-IgG (> 1:8. 6 patients with negative titers (aCP-IgG ≤ 1:8 served as control (CTRL. Both groups were comparable in age, gender, angina class, results of non-invasive stress-test and the baseline values of LD and flow. In the CP positive group 3 patients (30% did not show an increase of LD after ACh as evidence of ED. In the CTRL group 4 patients (67 % had ED. There was no association between aCP-IgG and changes of coronary blood flow after ACh. All patients showed normal CFVR (3.0 ± 0.27 irrespective of their aCP-IgG values. Conclusion In patients with typical

  6. Drastic shift from positive to negative estrogen effect on bone morphogenetic protein signaling in pulmonary arterial endothelial cells under hypoxia.

    Science.gov (United States)

    Ichimori, Hiroaki; Kogaki, Shigetoyo; Takahashi, Kunihiko; Ishida, Hidekazu; Narita, Jun; Nawa, Nobutoshi; Baden, Hiroki; Uchikawa, Toshiki; Okada, Yoko; Ozono, Keiichi

    2013-01-01

    To investigate the possible role of sex hormones in the pathogenesis of pulmonary arterial hypertension (PAH), the effect of β-estradiol (E2) on bone morphogenetic protein (BMP) signaling, a key signaling pathway involved in PAH, was studied in human pulmonary arterial endothelial cells (HPAEC). BMP signaling molecules, including BMP receptor, Smad1/5/8 and Id1, were studied in HPAEC under 1% O2 (hypoxia) and 21% O2 (normoxia) as well as the effect of hypoxia-inducible factor (HIF)-1α expression in the presence of E2 on BMP signaling. The effects of an estrogen receptor (ER) antagonist (ICI 182,780) and cycloheximide, and the interaction of ER with Smad or HIF-1α were also studied. In the presence of E2, BMP signaling was augmented under normoxia but suppressed under hypoxia. HIF-1α accumulation suppressed BMP signaling, whereas HIF-1α inhibition augmented signaling. These effects were cancelled by ICI 182,780. Moreover, binding between ER, HIF-1α and phosphorylated (p)-Smad1/5/8 proteins occurred only under hypoxia. On inhibition of de novo synthesis with cycloheximide, however, p-Smad1/5/8 expression was suppressed only under normoxia. The effects of E2 on BMP signaling in HPAEC altered depending on O2 concentration and different mechanisms may be involved. BMP and sex hormones may play an important role in PAH development.

  7. Endothelial dysfunction assessment by noninvasive peripheral arterial tonometry in patients with chronic obstructive pulmonary disease compared with healthy subjects.

    Science.gov (United States)

    Malerba, Mario; Radaeli, Alessandro; Nardin, Matteo; Clini, Enrico; Carpagnano, Giovanna Elisiana; Sciatti, Edoardo; Salghetti, Francesca; Bonadei, Ivano; Platto, Fabio; Vizzardi, Enrico

    2017-08-05

    Patients with chronic obstructive pulmonary disease (COPD) have an increased risk of cardiovascular disease. The endothelial dysfunction likely plays a central role in increasing cardiovascular risk. This cross-sectional, study investigated the prevalence and extent of endothelial dysfunction in patients with stable COPD. Peripheral arterial tonometry (PAT) was measured by post-ischemic reactive hyperemia index (RHI) in 16 COPD patients, 16 healthy controls and 16 subjects with treated systemic arterial hypertension (AH) and analysed with covariates condition (dyslipidemia, and medications). The prevalence of endothelial dysfunction was significantly higher in COPD group than in the other groups. Mean RHI was significantly lower in COPD patients compared with the other groups. At linear regression FEV 1 and RHI were directly correlated (Spearman index = 0.553; P = .026). COPD patients in groups C and D according to Global Initiative for Chronic Obstructive Lung Disease (GOLD) stages showed lower RHI compared with patients classified as A and B (P < .01). At multiple regression analysis the presence of dyslipidemia, COPD and AH were associated with the presence of endothelial dysfunction. Endothelial dysfunction in stable COPD patients is probably implicated in the high cardiovascular comorbidity. This study suggests the potential utility of endothelial dysfunction evaluation in patients with COPD to a timely assessment and treatment for cardiac complications. © 2017 John Wiley & Sons Ltd.

  8. Inhibition of vascular endothelial growth factor (VEGF)-induced endothelial proliferation, arterial relaxation, vascular permeability and angiogenesis by dobesilate.

    Science.gov (United States)

    Angulo, Javier; Peiró, Concepción; Romacho, Tania; Fernández, Argentina; Cuevas, Begoña; González-Corrochano, Rocío; Giménez-Gallego, Guillermo; de Tejada, Iñigo Sáenz; Sánchez-Ferrer, Carlos F; Cuevas, Pedro

    2011-09-30

    Vascular endothelial growth factor (VEGF) is a key factor in angiogenesis and vascular permeability which is associated with many pathological processes. 2,5-hydroxybenzene sulfonate (DHBS; dobesilate) is a small molecule with anti-angiogenic activity that has been described as an inhibitor of fibroblast growth factors (FGF). The aim of the present study was to evaluate the effects of DHBS on VEGF-induced actions. The effects of DHBS were evaluated on VEGF-induced proliferation in human umbilical vein endothelial cells (HUVEC) and rat aorta relaxation, as well as on in vivo VEGF-induced skin vascular permeability and neovascularization in rats. DHBS at 50 and 100 μM concentration significantly inhibited the proliferation of HUVEC induced by VEGF (10 ng/ml), without significantly affecting HUVEC proliferation in the absence of VEGF. Rapid VEGF-induced activation of Akt in HUVEC was also prevented by DHBS (100 μM). Additionally, DHBS (2 μM) specifically inhibited the relaxation of rat aorta induced by VEGF (0.1 to 30 ng/ml), but not endothelium-dependent relaxation to acetylcholine (1 nM to 10 μM). The in vivo enhancement of vascular permeability caused by VEGF injection (50 μl at 10 ng/ml) in rat skin was also inhibited by DHBS co-administration (200 μM) (74.8±3.8% inhibition of dye extravasation). Administration of DHBS (200 mg/kg/day; i.p.) also reduced VEGF-induced angiogenesis in vivo. DHBS inhibits main responses elicited in vitro and in vivo by VEGF. As a dual antagonist of VEGF and FGF activities, DHBS could be of therapeutic interest in the treatment of diseases related to VEGF/FGF overproduction and excessive angiogenesis. Copyright © 2011 Elsevier B.V. All rights reserved.

  9. Innervation of the human middle meningeal artery

    DEFF Research Database (Denmark)

    Edvinsson, L; Gulbenkian, S; Barroso, C P

    1998-01-01

    The majority of nerve fibers in the middle meningeal artery and branching arterioles are sympathetic, storing norepinephrine and neuropeptide Y (NPY). A sparse supply of fibers contain acetylcholinesterase activity and immunoreactivity toward vasoactive intestinal peptide (VIP), peptidine histidine...... methionine (PHM), and calcitonin gene-related peptide (CGRP). Only few substance P and neuropeptide K immunoreactive fibers are noted. Electronmicroscopy shows axons and terminals at the adventitial medial border of the human middle meningeal artery, with a fairly large distance to the smooth muscle cells...

  10. 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.

  11. 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.

  12. Effects of dark chocolate and cocoa consumption on endothelial function and arterial stiffness in overweight adults.

    Science.gov (United States)

    West, Sheila G; McIntyre, Molly D; Piotrowski, Matthew J; Poupin, Nathalie; Miller, Debra L; Preston, Amy G; Wagner, Paul; Groves, Lisa F; Skulas-Ray, Ann C

    2014-02-01

    The consumption of cocoa and dark chocolate is associated with a lower risk of CVD, and improvements in endothelial function may mediate this relationship. Less is known about the effects of cocoa/chocolate on the augmentation index (AI), a measure of vascular stiffness and vascular tone in the peripheral arterioles. We enrolled thirty middle-aged, overweight adults in a randomised, placebo-controlled, 4-week, cross-over study. During the active treatment (cocoa) period, the participants consumed 37 g/d of dark chocolate and a sugar-free cocoa beverage (total cocoa = 22 g/d, total flavanols (TF) = 814 mg/d). Colour-matched controls included a low-flavanol chocolate bar and a cocoa-free beverage with no added sugar (TF = 3 mg/d). Treatments were matched for total fat, saturated fat, carbohydrates and protein. The cocoa treatment significantly increased the basal diameter and peak diameter of the brachial artery by 6% (+2 mm) and basal blood flow volume by 22%. Substantial decreases in the AI, a measure of arterial stiffness, were observed in only women. Flow-mediated dilation and the reactive hyperaemia index remained unchanged. The consumption of cocoa had no effect on fasting blood measures, while the control treatment increased fasting insulin concentration and insulin resistance (P= 0·01). Fasting blood pressure (BP) remained unchanged, although the acute consumption of cocoa increased resting BP by 4 mmHg. In summary, the high-flavanol cocoa and dark chocolate treatment was associated with enhanced vasodilation in both conduit and resistance arteries and was accompanied by significant reductions in arterial stiffness in women.

  13. Mutant LRP6 Impairs Endothelial Cell Functions Associated with Familial Normolipidemic Coronary Artery Disease

    Directory of Open Access Journals (Sweden)

    Jian Guo

    2016-07-01

    Full Text Available Mutations in the genes low-density lipoprotein (LDL receptor-related protein-6 (LRP6 and myocyte enhancer factor 2A (MEF2A were reported in families with coronary artery disease (CAD. We intend to determine the mutational spectrum of these genes among hyperlipidemic and normolipidemic CAD families. Forty probands with early-onset CAD were recruited from 19 hyperlipidemic and 21 normolipidemic Chinese families. We sequenced all exons and intron-exon boundaries of LRP6 and MEF2A, and found a novel heterozygous variant in LRP6 from a proband with normolipidemic CAD. This variant led to a substitution of histidine to tyrosine (Y418H in an evolutionarily conserved domain YWTD in exon 6 and was not found in 1025 unrelated healthy individuals. Co-segregated with CAD in the affected family, LRP6Y418H significantly debilitated the Wnt3a-associated signaling pathway, suppressed endothelial cell proliferation and migration, and decreased anti-apoptotic ability. However, it exhibited no influences on low-density lipoprotein cholesterol uptake. Thus, mutation Y418H in LRP6 likely contributes to normolipidemic familial CAD via impairing endothelial cell functions and weakening the Wnt3a signaling pathway.

  14. Effects of cyclic intermittent hypoxia on ET-1 responsiveness and endothelial dysfunction of pulmonary arteries in rats.

    Directory of Open Access Journals (Sweden)

    Zhuo Wang

    Full Text Available Obstructive sleep apnoea (OSA is a risk factor for cardiovascular disorders and in some cases is complication of pulmonary hypertension. We simulated OSA by exposing rats to cyclic intermittent hypoxia (CIH to investigate its effect on pulmonary vascular endothelial dysfunction. Sprague-Dawley Rats were exposed to CIH (FiO2 9% for 1 min, repeated every 2 min for 8 h/day, 7 days/wk for 3 wk, and the pulmonary arteries of normoxia and CIH treated rats were analyzed for expression of endothelin-1 (ET-1 and ET receptors by histological, immunohistochemical, RT-PCR and Western Blot analyses, as well as for contractility in response to ET-1. In the pulmonary arteries, ET-1 expression was increased, and ET-1 more potently elicited constriction of the pulmonary artery in CIH rats than in normoxic rats. Exposure to CIH induced marked endothelial cell damage associated with a functional decrease of endothelium-dependent vasodilatation in the pulmonary artery. Compared with normoxic rats, ETA receptor expression was increased in smooth muscle cells of the CIH rats, while the expression of ETB receptors was decreased in endothelial cells. These results demonstrated endothelium-dependent vasodilation was impaired and the vasoconstrictor responsiveness increased by CIH. The increased responsiveness to ET-1 induced by intermittent hypoxia in pulmonary arteries of rats was due to increased expression of ETA receptors predominantly, meanwhile, decreased expression of ETB receptors in the endothelium may also participate in it.

  15. Comparative gene expression analysis between coronary arteries and internal mammary arteries identifies a role for the TES gene in endothelial cell functions relevant to coronary artery disease.

    Science.gov (United States)

    Archacki, Stephen R; Angheloiu, George; Moravec, Christine S; Liu, Hui; Topol, Eric J; Wang, Qing Kenneth

    2012-03-15

    Coronary artery disease (CAD) is the leading cause of death worldwide. It has been established that internal mammary arteries (IMA) are resistant to the development of atherosclerosis, whereas left anterior descending (LAD) coronary arteries are athero-prone. The contrasting properties of these two arteries provide an innovative strategy to identify the genes that play important roles in the development of atherosclerosis. We carried out microarray analysis to identify genes differentially expressed between IMA and LAD. Twenty-nine genes showed significant differences in their expression levels between IMA and LAD, which included the TES gene encoding Testin. The role of TES in the cardiovascular system is unknown. Here we show that TES is involved in endothelial cell (EC) functions relevant to atherosclerosis. Western blot analysis showed higher TES expression in IMA than in LAD. Reverse transcription polymerase chain reaction and western blot analyses showed that TES was consistently and markedly down-regulated by more than 6-fold at both mRNA and protein levels in patients with CAD compared with controls without CAD (P= 0.000049). The data suggest that reduced TES expression is associated with the development of CAD. Knockdown of TES expression by small-interfering RNA promoted oxidized-LDL-mediated monocyte adhesion to ECs, EC migration and the transendothelial migration of monocytes, while the over-expression of TES in ECs blunted these processes. These results demonstrate association between reduced TES expression and CAD, establish a novel role for TES in EC functions and raise the possibility that reduced TES expression increases susceptibility to the development of CAD.

  16. 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.

  17. The effects of plant stanol ester consumption on arterial stiffness and endothelial function in adults: a randomised controlled clinical trial.

    Science.gov (United States)

    Gylling, Helena; Halonen, Janne; Lindholm, Harri; Konttinen, Jussi; Simonen, Piia; Nissinen, Markku J; Savolainen, Aslak; Talvi, Airi; Hallikainen, Maarit

    2013-07-10

    The hypocholesterolemic effect of plant stanol ester consumption has been studied extensively, but its effect on cardiovascular health has been less frequently investigated. We studied the effects of plant stanol esters (staest) on arterial stiffness and endothelial function in adults without lipid medication. Ninety-two asymptomatic subjects, 35 men and 57 women, mean age of 50.8±1.0 years (SEM) were recruited from different commercial companies. It was randomized, controlled, double-blind, parallel trial and lasted 6 months. The staest group (n=46) consumed rapeseed oil-based spread enriched with staest (3.0 g of plant stanols/d), and controls (n=46) the same spread without staest. Arterial stiffness was assessed via the cardio-ankle vascular index (CAVI) in large and as an augmentation index (AI) in peripheral arteries, and endothelial function as reactive hyperemia index (RHI). Lipids and vascular endpoints were tested using analysis of variance for repeated measurements. At baseline, 28% of subjects had a normal LDL cholesterol level (≤3.0 mmol/l) and normal arterial stiffness (cholesterol concentrations declined by 6.6, 10.2, and 10.6% compared with controls (pcholesterol levels achieved by staest was related to the improvement in RHI (r=-0.452, p=0.006 and -0.436, p=0.008). Lowering LDL and non-HDL cholesterol by 10% with staest for 6 months reduced arterial stiffness in small arteries. In subgroup analyses, staest also had a beneficial effect on arterial stiffness in large arteries in men and on endothelial function. Further research will be needed to confirm these results in different populations. Clinical Trials Register # NCT01315964.

  18. 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.

  19. Impaired autonomic regulation of resistance arteries in mice with low vascular endothelial growth factor or upon vascular endothelial growth factor trap delivery

    DEFF Research Database (Denmark)

    Storkebaum, Erik; Ruiz de Almodovar, Carmen; Meens, Merlijn

    2010-01-01

    BACKGROUND: Control of peripheral resistance arteries by autonomic nerves is essential for the regulation of blood flow. The signals responsible for the maintenance of vascular neuroeffector mechanisms in the adult, however, remain largely unknown. METHODS AND RESULTS: Here, we report that VEGF( ...... for VEGF in the maintenance of arterial neuroeffector function and may help us better understand how VEGF inhibitors cause vascular regulation defects in cancer patients.......( partial differential/ partial differential) mice with low vascular endothelial growth factor (VEGF) levels suffer defects in the regulation of resistance arteries. These defects are due to dysfunction and structural remodeling of the neuroeffector junction, the equivalent of a synapse between autonomic...... nerve endings and vascular smooth muscle cells, and to an impaired contractile smooth muscle cell phenotype. Notably, short-term delivery of a VEGF inhibitor to healthy mice also resulted in functional and structural defects of neuroeffector junctions. CONCLUSIONS: These findings uncover a novel role...

  20. Kinetics of circulating endothelial progenitor cells in patients undergoing carotid artery surgery

    Directory of Open Access Journals (Sweden)

    Kalender G

    2016-12-01

    Full Text Available G Kalender,1 A Kornberger,2 M Lisy,1 Andres Beiras-Fernandez,2 UA Stock2 1Deparment of General, Thoracic and Vascular Surgery, Hoechst Hospital, 2Department of Thoracic and Cardiovascular Surgery, University Hospital Frankfurt, Frankfurt am Main, Germany Aim: Endothelial progenitor cells (EPCs are primitive cells found in the bone marrow and peripheral blood (PB. In particular, the potential of EPCs to differentiate into mature endothelial cells remains of high interest for clinical applications such as bio-functionalized patches for autologous seeding after implantation. The objective of this study was to determine EPCs’ kinetics in patients undergoing carotid artery thromboendarterectomy (CTEA and patch angioplasty. Methods: Twenty CTEA patients were included (15 male, mean age 76 years. PB samples were taken at 1 day preoperatively, and at 1, 3, and 5 days postoperatively. Flow cytometric analysis was performed for CD34, CD133, KDR, and CD45. Expression of KDR, SDF-1α, and G-CSF was analyzed by means of enzyme-linked immunosorbent assay. Results: Fluorescence-activated cell sorting analysis revealed 0.031%±0.016% (% of PB mononuclear cells KDR+ cells and 0.052%±0.022% CD45-/CD34+/CD133+ cells, preoperatively. A 33% decrease of CD45–/CD34+/CD133+ cells was observed at day 1 after surgery. However, a relative number (compared to initial preoperative values of CD45-/CD34+/CD133+ cells was found on day 3 (82% and on day 5 (94% postoperatively. More profound upregulated levels of CD45–CD34+/CD133+ cells were observed for diabetic (+47% compared to nondiabetic and male (+38% compared to female patients. No significant postoperative time-dependent differences were found in numbers of KDR+ cells and the concentrations of the cytokines KDR and G-CSF. However, the SDF-1α levels decreased significantly on day 1 postoperatively but returned to preoperative levels by day 3. Conclusion: CTEA results in short-term downregulation of circulating

  1. Endothelial and lipoprotein lipases in human and mouse placenta

    DEFF Research Database (Denmark)

    Lindegaard, Marie Louise Skakkebæk; Olivecrona, Gunilla; Christoffersen, Christina

    2005-01-01

    Placenta expresses various lipase activities. However, a detailed characterization of the involved genes and proteins is lacking. In this study, we compared the expression of endothelial lipase (EL) and LPL in human term placenta. When placental protein extracts were separated by heparin...... protein associated with both cell types. In mouse placentas, lack of LPL expression resulted in increased EL mRNA expression. These results suggest that the cellular expression of EL and LPL in human placenta is different. Nevertheless, the two lipases might have overlapping functions in the mouse...... placenta. Our data also suggest that the major portions of both proteins are stored in an inactive form in human term placenta....

  2. 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...

  3. The Influence of Endothelial Function and Myocardial Ischemia on Peak Oxygen Consumption in Patients with Coronary Artery Disease

    Directory of Open Access Journals (Sweden)

    Simon L. Bacon

    2012-01-01

    Full Text Available Impaired endothelial function has been shown to limit exercise in coronary artery disease (CAD patients and has been implicated in myocardial ischemia. However, the association of endothelial function and ischemia on peak exercise oxygen consumption (VO2 has not been previously reported. A total of 116 CAD patients underwent standard exercise stress testing, during which VO2 was measured. On a separate day, endothelial-dependent and -independent function were assessed by ultrasound using flow-mediated arterial vasodilation (FMD and sublingual glyceryl trinitrate administration (GTNMD of the brachial artery. Patients with exercise-induced myocardial ischemia had lower FMD than nonischemic patients (3.64±0.57 versus 4.98±0.36, P=.050, but there was no difference in GTNMD (14.11±0.99 versus 15.47±0.63, P=.249. Analyses revealed that both FMD (P=.006 and GTNMD (P=.019 were related to peak VO2. However, neither the presence of ischemia (P=.860 nor the interaction of ischemia with FMD (P=.382 and GTNMD (P=.151 was related to peak VO2. These data suggest that poor endothelial function, potentially via impaired NO production and smooth muscle dysfunction, may be an important determinant of exercise capacity in patients with CAD, independent of myocardial ischemia.

  4. Effects of percutaneous lower-extremity arterial interventions on endothelial function and inflammation response in patients with both type 2 diabetes and lower-extremity peripheral arterial disease.

    Science.gov (United States)

    Du, Yaping; Wang, Fujun; Qi, Huiqing; Ding, Haixia; Hou, Lin; Gao, Qian; Tan, Miao; Liu, Yueqin; Xing, Na; Sun, Jin

    2015-01-01

    The high incidence and damage of PAD in people with diabetes has aroused wide attention. We aimed to examine effects of percutaneous lower-extremity arterial interventions (PLEAIs) on endothelial function and inflammation response in type 2 diabetes (T2D) patients with lower-extremity peripheral arterial disease (PAD). 78 T2D inpatients with PAD were selected into the treatment group. Their venous levels of von Willebrand Factor (vWF) and high sensitivity C reactive protein (hsCRP) were measured. Blood samples were collected from the arterial sheath for vWF and hsCRP tests. Venous levels of vWF and hsCRP were monitored at 24 hours, 48 hours, 1 week, and 2 weeks post PLEAIs. Prior to PLEAIs, venous levels of vWF and hsCRP in the treatment group were significantly higher than the control group. The arterial levels of vWF and hsCRP were 117.9%±15.1% and 5.19±0.76 mg/L in the control group, while those levels in the treatment group before intervention were also significantly higher than in the control group. In the treatment group prior to inventions, vWF and hsCRP levels of arterial ischemic regions were significantly higher than the non-ischemic regions. The vWF level of arterial ischemic regions after treatment was significantly higher than that prior to treatment. PLEAIs applied to those patients may lead to worse endothelial dysfunction and activated inflammatory response during treatment and 1 week after treatment, which indicates an emerging necessary of early protection or care on endothelial function and inflammatory reaction during and post PLEAIs.

  5. Evaluation of the brachial artery endothelial function in chronic alcohol consumption among males by high-frequency ultrasonography.

    Science.gov (United States)

    Luo, Runlan; Shen, Jiaqi; Zhou, Qiao; Liu, Yue; Li, Guangsen

    2017-02-01

    There is evidence suggesting that different volumes of chronic alcohol consumption have different effects on the endothelium. Therefore, using high-frequency ultrasonography, we evaluate the effects of the different volume and duration of alcohol intake on brachial artery endothelial function in chronic drinkers. Ninety-two male chronic episodic alcoholics were grouped by alcohol intake amount and duration: mild (group B, n=30); ≤90 mg ethanol daily, 3-5 days/wk for 5-8 years; moderate (group C, n=30); 90-150 mg ethanol daily, 3-5 days/wk for 9-20 years; and severe (group D, n=32); ≥150 mg ethanol daily, 6-7 days/wk for more than 10 years. Thirty male nondrinkers were recruited as the control group A. High-frequency ultrasonography was used to measure brachial artery diameter during rest, during reactive hyperemia and following the administration of nitroglycerin. Endothelial-dependent brachial artery flow-mediated dilatation (FMD) and endothelial-independent brachial artery nitrate-mediated dilatation (NMD) were calculated. Flow-mediated dilatation values for group C and D were significantly lower than those for group A and B (VC =7.63±0.22, VD =5.85±0.23 vs VA =13.35±0.35, VB =12.81±0.36, Pconsumption caused endothelial dysfunction, even damaging vascular smooth muscle cells in cases of heavy alcohol consumption, while abstinence and chronic mild alcoholics caused no effect on endothelial function. © 2016, Wiley Periodicals, Inc.

  6. Effects of a 12-week alpine skiing intervention on endothelial progenitor cells, peripheral arterial tone and endothelial biomarkers in the elderly.

    Science.gov (United States)

    Niederseer, David; Steidle-Kloc, Eva; Mayr, Matthias; Müller, Edith E; Cadamuro, Janne; Patsch, Wolfgang; Dela, Flemming; Müller, Erich; Niebauer, Josef

    2016-07-01

    Endothelial dysfunction occurs early during atherogenesis and it can be normalized by exercise training. Unfortunately, patients' compliance with exercise prescription remains low, often because the given choices do not appeal to them. In Alpine regions, skiing is a popular mode of exercise, and therefore we set out to assess whether it can induce antiatherogenic effects. We randomized 42 subjects into a group of 12weeks of guided skiing (intervention group, IG, n=22; 12 males/10 females; age: 66.6±2.1years) or a control group (CG, n=20; 10 males/10 females; age: 67.3±4.4years). Early (CD3-CD34+CD45+) and late endothelial progenitor cells (EPCs; CD45dimCD34+KDR+ peripheral blood mononuclear cells, PBMCs), peripheral arterial tonometry and endothelial biomarkers were assessed at the beginning and end of the study. In the IG, participants completed 28.5±2.6 skiing days at an average heart rate of 72.7±8.5% of their maximum heart rate. Changes in early (IG: +0.001±0.001% PBMC; CG: -0.001±0.001% PBMC; IG vs. CG: pskiing induces several beneficial effects on markers of atherogenesis including EPCs, peripheral arterial tone and homocysteine. Our findings suggest that recreational alpine skiing may serve as a further mode of preventive exercise training, which might result in improved compliance with current recommendations. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  7. Genetic engineering with endothelial nitric oxide synthase improves functional properties of endothelial progenitor cells from patients with coronary artery disease: an in vitro study.

    Science.gov (United States)

    Kaur, Savneet; Kumar, T R Santhosh; Uruno, Akira; Sugawara, Akira; Jayakumar, Karunakaran; Kartha, Chandrasekharan Cheranellore

    2009-11-01

    Recent studies have reported a marked impairment in the number and functions of endothelial progenitor cells (EPCs) in patients with coronary artery disease (CAD). In view of an important role of eNOS in angiogenesis, in the present study, we evaluated the effects of eNOS gene transfer in ex vivo expanded EPCs isolated from patients with CAD. The expanded EPCs were transfected with mammalian expression vector pcDNA3.1-eNOS containing the full-length human eNOS gene using lipofectamine. About 35-40% of the eNOS-EPCs had higher expression of eNOS as compared to untransfected EPCs. EPCs transfected with pcDNA3.0-EGFP, the plasmid vector expressing green fluorescent protein (GFP) were used as control. The untransfected, GFP-transfected and eNOS-transfected EPCs were compared in terms of important functional attributes of angiogenesis such as proliferation, migration, differentiation and adhesion/integration into tube-like structures in vitro. Functional studies revealed that in the presence of defined growth conditions, compared to the untransfected and GFP-transfected cells, eNOS-EPCs from patients with CAD have a significant increase in [3H] thymidine-labeled DNA (P < 0.01), migration (14.6 +/- 1.8 and 16.5 +/- 1.9 vs. 23.5 +/- 3.4 cells/field, P < 0.01), ability to differentiate into endothelial-like spindle-shaped cells (46 +/- 4.5 and 56.5 +/- 2.1 vs. 93.2 +/- 6.6 cells/field, P < 0.001) and also incorporation into tube-like structures on the matrigel (GFP-EPCs: 21.25 +/- 2.9 vs. GFP-eNOS-EPCs: 34.5 +/- 5.5 cells/field, P < 0.05). We conclude that eNOS gene transfection is a valuable approach to augment angiogenic properties of ex vivo expanded EPCs and eNOS-modified EPCs may offer significant advantages than EPCs alone in terms of their clinical use in patients with myocardial ischemia.

  8. Paracrine effects of bone marrow-derived endothelial progenitor cells: cyclooxygenase-2/prostacyclin pathway in pulmonary arterial hypertension.

    Directory of Open Access Journals (Sweden)

    Dong-Mei Jiang

    Full Text Available BACKGROUND: Endothelial dysfunction is the pathophysiological characteristic of pulmonary arterial hypertension (PAH. Some paracrine factors secreted by bone marrow-derived endothelial progenitor cells (BMEPCs have the potential to strengthen endothelial integrity and function. This study investigated whether BMEPCs have the therapeutic potential to improve monocrotaline (MCT-induced PAH via producing vasoprotective substances in a paracrine fashion. METHODS AND RESULTS: Bone marrow-derived mononuclear cells were cultured for 7 days to yield BMEPCs. 24 hours or 3 weeks after exposure to BMEPCs in vitro or in vivo, the vascular reactivity, cyclooxygenase-2 (COX-2 expression, prostacyclin (PGI2 and cAMP release in isolated pulmonary arteries were examined respectively. Treatment with BMEPCs could improve the relaxation of pulmonary arteries in MCT-induced PAH and BMEPCs were grafted into the pulmonary bed. The COX-2/prostacyclin synthase (PGIS and its progenies PGI2/cAMP were found to be significantly increased in BMEPCs treated pulmonary arteries, and this action was reversed by a selective COX-2 inhibitor, NS398. Moreover, the same effect was also observed in conditioned medium obtained from BMEPCs culture. CONCLUSIONS: Implantation of BMEPCs effectively ameliorates MCT-induced PAH. Factors secreted in a paracrine fashion from BMEPCs promote vasoprotection by increasing the release of PGI2 and level of cAMP.

  9. AXL-Mediated Productive Infection of Human Endothelial Cells by Zika Virus.

    Science.gov (United States)

    Liu, Shufeng; DeLalio, Leon J; Isakson, Brant E; Wang, Tony T

    2016-11-11

    The mosquito-borne Zika virus (ZIKV) is now recognized as a blood-borne pathogen, raising an important question about how the virus gets into human bloodstream. The imminent threat of the ZIKV epidemic to the global blood supply also demands novel therapeutics to stop virus transmission though transfusion. We intend to characterize ZIKV tropism for human endothelial cells (ECs) and provide potential targets for intervention. We conducted immunostaining, plaque assay, and quantitative reverse transcription-polymerase chain reaction of ZIKV RNA to evaluate the possible infection of ECs by ZIKV. Both the African and the South American ZIKV strains readily infect human umbilical vein endothelial cells and human ECs derived from aortic and coronary artery, as well as the saphenous vein. Infected ECs released infectious progeny virus. Compared with the African strains, South American ZIKV isolates replicate faster in ECs and are partially cytopathic, suggesting enhanced virulence of these isolates. Flow cytometric analyses showed that the susceptibility of ECs positively correlated with the cell surface levels of tyrosine-protein kinase receptor UFO (AXL) receptor tyrosine kinase. Gain- and loss-of-function studies further revealed that AXL is required for ZIKV entry at a postbinding step. Finally, small-molecule inhibitors of the AXL kinase significantly reduced ZIKA infection of ECs. We identified EC as a key cell type for ZIKV infection. These data support the view of hematogenous dissemination of ZIKV and implicate AXL as a new target for antiviral therapy. © 2016 American Heart Association, Inc.

  10. Relaxation of human isolated mesenteric arteries by vasopressin and desmopressin.

    Science.gov (United States)

    Martínez, M C; Vila, J M; Aldasoro, M; Medina, P; Flor, B; Lluch, S

    1994-01-01

    1. The effects of vasopressin and deamino-8-D-arginine vasopressin (DDAVP, desmopressin) were studied in artery rings (0.8-1 mm in external diameter) obtained from portions of human omentum during the course of abdominal operations (27 patients). 2. In arterial rings under resting tension, vasopressin produced concentration-dependent, endothelium-independent contractions with an EC50 of 0.59 +/- 0.12 nM. The V1 antagonist d(CH2)5Tyr(Me)AVP (1 microM) and the mixed V1-V2 antagonist desGly-d(CH2)5D-Tyr(Et)ValAVP (0.01 microM) displaced the control curve to vasopressin to the right in a parallel manner without differences in the maximal responses. In the presence of indomethacin (1 microM) the contractile response to vasopressin was significantly increased (P < 0.01). 3. In precontracted arterial rings, previously treated with the V1 antagonist, d(CH2)5Tyr(Me)AVP (1 microM), vasopressin produced endothelium-dependent relaxation. This relaxation was reduced significantly (P < 0.05) by indomethacin (1 microM) and unaffected by the V1-V2 receptor antagonist desGly-d(CH2)5D-Tyr(Et)ValAVP (1 microM) or by NG-nitro-L-arginine methyl ester (L-NAME, 0.1 mM). 4. The selective V2 receptor agonist, DDAVP, caused endothelium-independent, concentration-dependent relaxations in precontracted arterial rings that were inhibited by the mixed V1-V2 receptor antagonist, but not by the V1 receptor antagonist or by pretreatment with indomethacin or L-NAME. 5. Results from this study suggest that vasopressin is primarily a constrictor of human mesenteric arteries by V1 receptor stimulation; vasopressin causes dilatation only during V1 receptor blockade. The relaxation appears to be mediated by the release of vasodilator prostaglandins from the endothelial cell layer and is independent of V2 receptor stimulation or release of nitric oxide.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7834191

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

    Science.gov (United States)

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

    2017-10-09

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

  12. 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.

  13. 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...

  14. Activation of calpain-1 in human carotid artery atherosclerotic lesions

    Directory of Open Access Journals (Sweden)

    Pedro Luis M

    2009-06-01

    Full Text Available Abstract Background In a previous study, we observed that oxidized low-density lipoprotein-induced death of endothelial cells was calpain-1-dependent. The purpose of the present paper was to study the possible activation of calpain in human carotid plaques, and to compare calpain activity in the plaques from symptomatic patients with those obtained from patients without symptoms. Methods Human atherosclerotic carotid plaques (n = 29, 12 associated with symptoms were removed by endarterectomy. Calpain activity and apoptosis were detected by performing immunohistochemical analysis and TUNEL assay on human carotid plaque sections. An antibody specific for calpain-proteolyzed α-fodrin was used on western blots. Results We found that calpain was activated in all the plaques and calpain activity colocalized with apoptotic cell death. Our observation of autoproteolytic cleavage of the 80 kDa subunit of calpain-1 provided further evidence for enzyme activity in the plaque samples. When calpain activity was quantified, we found that plaques from symptomatic patients displayed significantly lower calpain activity compared with asymptomatic plaques. Conclusion These novel results suggest that calpain-1 is commonly active in carotid artery atherosclerotic plaques, and that calpain activity is colocalized with cell death and inversely associated with symptoms.

  15. Positive Feedback Regulation of Agonist-Stimulated Endothelial Ca2+ Dynamics by KCa3.1 Channels in Mouse Mesenteric Arteries

    DEFF Research Database (Denmark)

    Qian, Xun; Francis, Michael; Köhler, Ralf

    2014-01-01

    Intermediate and small conductance KCa channels IK1 (KCa3.1) and SK3 (KCa2.3) are primary targets of endothelial Ca(2+) signals in the arterial vasculature, and their ablation results in increased arterial tone and hypertension. Activation of IK1 channels by local Ca(2+) transients from internal ...... stores or plasma membrane channels promotes arterial hyperpolarization and vasodilation. Here, we assess arteries from genetically altered IK1 knockout mice (IK1(-/-)) to determine whether IK1 channels exert a positive feedback influence on endothelial Ca(2+) dynamics....

  16. Overexpression of tissue-nonspecific alkaline phosphatase (TNAP) in endothelial cells accelerates coronary artery disease in a mouse model of familial hypercholesterolemia

    Science.gov (United States)

    Romanelli, Filippo; Corbo, AnthonyMarco; Salehi, Maryam; Yadav, Manisha C.; Salman, Soha; Petrosian, David; Rashidbaigi, Omid J.; Chait, Jesse; Kuruvilla, Jes; Plummer, Maria; Radichev, Ilian; Margulies, Kenneth B.; Gerdes, A. Martin; Pinkerton, Anthony B.; Millán, José Luis; Savinov, Alexei Y.

    2017-01-01

    Objective Overexpression of tissue-nonspecific alkaline phosphatase (TNAP) in endothelium leads to arterial calcification in mice. The purpose of this study was to examine the effect of elevated endothelial TNAP on coronary atherosclerosis. In addition, we aimed to examine endogenous TNAP activity in human myocardium. Approach and results A vascular pattern of TNAP activity was observed in human non-failing, ischemic, and idiopathic dilated hearts (5 per group); no differences were noted between groups in this study. Endothelial overexpression of TNAP was achieved in mice harboring a homozygous recessive mutation in the low density lipoprotein receptor (whc allele) utilizing a Tie2-cre recombinase (WHC-eTNAP mice). WHC-eTNAP developed significant coronary artery calcification at baseline compared WHC controls (4312 vs 0μm2 alizarin red area, pcoronary arteries of WHC-eTNAP was increased compared to WHC controls (121633 vs 9330μm2 oil red O area, pCoronary lesions in WHC-eTNAP mice exhibited intimal thickening, calcifications, foam cells, and necrotic cores. This was accompanied by the reduction in body weight and left ventricular ejection fraction (19.5 vs. 23.6g, pcoronary calcium (78838 vs.144622μm2) and lipids (30754 vs. 77317μm2); improved body weight (22.4 vs.18.8g) and ejection fraction (59 vs. 47%). The effects of SBI-425 were significant in the direct comparisons with placebo but disappeared after TNAP-negative placebo-treated group was included in the models as healthy controls. Conclusions Endogenous TNAP activity is present in human cardiac tissues. TNAP overexpression in vascular endothelium in mice leads to an unusual course of coronary atherosclerosis, in which calcification precedes lipid deposition. The prevalence and significance of this mechanism in human atherosclerosis requires further investigations. PMID:29023576

  17. Effects of a 12-week alpine skiing intervention on endothelial progenitor cells, peripheral arterial tone and endothelial biomarkers in the elderly

    DEFF Research Database (Denmark)

    Niederseer, David; Steidle-Kloc, Eva; Mayr, Matthias

    2016-01-01

    : +0.18±0.76) and CG (-0.39±0.85; p=0.045), as did homocysteine (IG: -1.3±1.3μmol/l; CG: -0.4±1.4μmol/l; p=0.037) while other endothelial biomarkers remained essentially unchanged. CONCLUSIONS: This study shows that skiing induces several beneficial effects on markers of atherogenesis including EPCs......, peripheral arterial tone and homocysteine. Our findings suggest that recreational alpine skiing may serve as a further mode of preventive exercise training, which might result in improved compliance with current recommendations....

  18. Differential effect of amylin on endothelial-dependent vasodilation in mesenteric arteries from control and insulin resistant rats.

    Directory of Open Access Journals (Sweden)

    Mariam El Assar

    Full Text Available Insulin resistance (IR is frequently associated with endothelial dysfunction and has been proposed to play a major role in cardiovascular disease (CVD. On the other hand, amylin has long been related to IR. However the role of amylin in the vascular dysfunction associated to IR is not well addressed. Therefore, the aim of the study was to assess the effect of acute treatment with amylin on endothelium-dependent vasodilation of isolated mesenteric arteries from control (CR and insulin resistant (IRR rats and to evaluate the possible mechanisms involved. Five week-old male Wistar rats received 20% D-fructose dissolved in drinking water for 8 weeks and were compared with age-matched CR. Plasmatic levels of glucose, insulin and amylin were measured. Mesenteric microvessels were dissected and mounted in wire myographs to evaluate endothelium-dependent vasodilation to acetylcholine. IRR displayed a significant increase in plasmatic levels of glucose, insulin and amylin and reduced endothelium-dependent relaxation when compared to CR. Acute treatment of mesenteric arteries with r-amylin (40 pM deteriorated endothelium-dependent responses in CR. Amylin-induced reduction of endothelial responses was unaffected by the H2O2 scavenger, catalase, but was prevented by the extracellular superoxide scavenger, superoxide dismutase (SOD or the NADPH oxidase inhibitor (VAS2870. By opposite, amylin failed to further inhibit the impaired relaxation in mesenteric arteries of IRR. SOD, or VAS2870, but not catalase, ameliorated the impairment of endothelium-dependent relaxation in IRR. At concentrations present in insulin resistance conditions, amylin impairs endothelium-dependent vasodilation in mircrovessels from rats with preserved vascular function and low levels of endogenous amylin. In IRR with established endothelial dysfunction and elevated levels of amylin, additional exposure to this peptide has no effect on endothelial vasodilation. Increased superoxide

  19. 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

  20. Contribution of Endothelial-to-Mesenchymal Transition to the Pathogenesis of Human Cerebral and Orbital Cavernous Malformations.

    Science.gov (United States)

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

    2017-07-01

    The analysis of gene-targeted mouse mutants has demonstrated that endothelial-to-mesenchymal transition (EndMT) is crucial to the onset and progression of cerebral cavernous malformations (CMs). It has also been shown that Notch and ephrin/Eph signaling are involved in EndMT. However, their roles in the pathogenesis of human intracranial CMs remain unclear. To elucidate the contribution of EndMT, the Notch pathway, and ephrin-B2/EphB4 signaling to the pathogenesis of human intracranial CMs. Eight human intracranial CMs (5 cerebral and 3 orbital CMs) were immunohistochemically investigated. CD31 (an endothelial marker) and EndMT markers, such as α-smooth muscle actin (a mesenchymal marker) and CD44 (a mesenchymal stem cell marker), were expressed in the endothelial layer of vascular sinusoids in all cases, suggesting that endothelial cells (ECs) have acquired mesenchymal and stem-cell-like characteristics and undergone EndMT in all cerebral and orbital CMs. EndMT was observed in about 70% and 35% of ECs in cerebral and orbital CMs, respectively. In all cases, Notch3 was expressed in the endothelial layer, indicating that ECs of vascular sinusoids have acquired mesenchymal features. In all cases, both ephrin-B2 and EphB4 were detected in the endothelial layer, suggesting that ECs of vascular sinusoids are immature or malformed cells and have both arterial and venous characteristics. EndMT plays a critical role in the pathogenesis of human cerebral and orbital CMs. Modulating EndMT is expected to be a new therapeutic strategy for cerebral and orbital CMs.

  1. Utilization of Pulsatile flow to Decellularize the Human Umbilical Arteries to Make Small-Caliber Blood Vessel Scaffolds.

    Science.gov (United States)

    Chen, Shengjie; Li, Jingxing; Dong, Peiqing

    2013-09-01

    To explore the effect of pulsatile flow in the decellularization process of small blood vessels. A total of 30 human umbilical cords were used in the current study. The umbilical cords were flushed with 0.25% trypsin/0.01% EDTA for 30 minutes, followed by treatment with 1% sodium dodecyl sulfate sodium dodecyl sulfate for 3 hours. The effectiveness of decellularization on the umbilical artery wall was analyzed by mechanical analysis. The scaffolds' biocompatibility was observed by co-culture with the human umbilical endothelial cells. The maximum stress of the arteries before and after denuding was 3.55 ± 0.42N and 3.50 ± 0.43N, respectively. Under the pressure of 300 mmHg, 28 pieces of umbilical cords remained intact before and after the flushing, while 2 pieces ruptured under 300 mmHg. There was no significant difference in mechanical properties between flushed and control arteries. Isolated human umbilical endothelial cells grow and spread well on the decellularized umbilical artery scaffolds. Decellularization by pulsatile flow in human umbilical artery is a convenient, rapid and efficient approach to increase the availability of small caliber blood vessel scaffolds. Decellularization; Human umbilical artery; Scaffolds; Small diameter blood vessel; Vascular tissue engineering.

  2. Aurantio-obtusin relaxes systemic arteries through endothelial PI3K/AKT/eNOS-dependent signaling pathway in rats

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    Shuzhen Li

    2015-07-01

    Full Text Available Aurantio-obtusin is a natural effective compound isolated from Semen Cassiae, which possesses hypotensive and hypolipidemic effects. Although its hypotensive effect have been clarified, mechanisms Aurantio-obtusin relaxes systemic arteries remain unclear. This study was to investigate effects and mechanisms of Aurantio-obtusin on isolated mesenteric arteries (MAs. We examined MAs relaxation induced by Aurantio-obtusin on rat isolated MAs, expression and activity of endothelial nitric oxide synthase (eNOS and protein kinase B (AKT, and nitric oxide (NO production in bovine artery endothelial cells (BAECs. Findings showed Aurantio-obtusin elicited dose-dependent vasorelaxation with phenylephrine (PE precontracted rat MA rings (diameter: 200–300 μm, which can be diminished by denudation of endothelium and inhibition of eNOS activity, while having no effect on rat isolated pulmonary artery (PA rings. Aurantio-obtusin increased NO production by promoting phosphorylations of eNOS at Ser-1177 and Thr-495 in endothelial cells. Aurantio-obtusin also promoted phosphorylations of Akt at Ser-473. PI3K inhibitor LY290042 could diminish vasorelaxation induced by Aurantio-obtusin. Moreover Aurantio-obtusin also elicited dose-dependent vasorelaxation effect with PE precontracted MA rings (diameter: 100–150 μm. Therefore, vasorelaxation induced by Aurantio-obtusin was dependent on endothelium integrity and NO production, which mediated by endothelial PI3K/Akt/eNOS pathway. Results suggest Aurantio-obtusin may offer therapeutic effects in hypertension, as a new potential vasodilator.

  3. Pulmonary artery endothelial cell phenotypic alterations in a large animal model of pulmonary arteriovenous malformations after the Glenn shunt.

    Science.gov (United States)

    Kavarana, Minoo N; Mukherjee, Rupak; Eckhouse, Shaina R; Rawls, William F; Logdon, Christina; Stroud, Robert E; Patel, Risha K; Nadeau, Elizabeth K; Spinale, Francis G; Graham, Eric M; Forbus, Geoffrey A; Bradley, Scott M; Ikonomidis, John S; Jones, Jeffrey A

    2013-10-01

    Longevity of the superior cavopulmonary connection (SCPC) is limited by the development of pulmonary arteriovenous malformations (PAVM). The goal of this study was to determine whether phenotypic changes in pulmonary artery endothelial cells (PAEC) that favor angiogenesis occur with PAVM formation. A superior vena cava to right pulmonary artery connection was constructed in 5 pigs. Pulmonary arteries were harvested at 6 to 8 weeks after surgery to establish cultures of PAEC and smooth muscle cells, to determine cell proliferation, gene expression, and tubule formation. Abundance of proteins related to angiogenesis was measured in lung tissue. Contrast echocardiography revealed right-to-left shunting, consistent with PAVM formation. While the proliferation of smooth muscle cells from the right pulmonary artery (shunted side) and left pulmonary artery (nonshunted side) were similar, right PAEC proliferation was significantly higher. Expression profiles of genes encoding cellular signaling proteins were higher in PAECs from the right pulmonary artery versus left pulmonary artery. Protein abundance of angiopoietin-1, and Tie-2 (angiopoietin receptor) were increased in the right lung (both p SCPC concomitantly with differential changes in PAEC proliferative ability and phenotype. Moreover, there was a significant increase in the angiopoietin/Tie-2 complex in the right lung, which may provide novel therapeutic targets to attenuate PAVM formation after a SCPC. Copyright © 2013 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

  4. Cytotoxicity of Voriconazole on Cultured Human Corneal Endothelial Cells▿

    OpenAIRE

    Han, Sang Beom; Shin, Young Joo; Hyon, Joon Young; Wee, Won Ryang

    2011-01-01

    The purpose of the present study was to evaluate the toxicity of voriconazole on cultured human corneal endothelial cells (HCECs). HCECs were cultured and exposed to various concentrations of voriconazole (5.0 to 1,000 μg/ml). Cell viability was measured using a Cell Counting Kit-8 (CCK-8) and live/dead viability/cytotoxicity assays. Cell damage was assessed using phase-contrast microscopy after 24 h of exposure to voriconazole. To analyze the effect of voriconazole on the intercellular barri...

  5. Lidocaine Prevents Oxidative Stress-Induced Endothelial Dysfunction of the Systemic Artery in Rats With Intermittent Periodontal Inflammation.

    Science.gov (United States)

    Saito, Takumi; Yamamoto, Yasuhiro; Feng, Guo-Gang; Kazaoka, Yoshiaki; Fujiwara, Yoshihiro; Kinoshita, Hiroyuki

    2017-06-01

    Periodontal inflammation causes endothelial dysfunction of the systemic artery. However, it is unknown whether the use of local anesthetics during painful dental procedures alleviates periodontal inflammation and systemic endothelial function. This study was designed to examine whether the gingival or systemic injection of lidocaine prevents oxidative stress-induced endothelial dysfunction of the systemic artery in rats with intermittent periodontal inflammation caused by lipopolysaccharides (LPS). Some rats received 1500 µg LPS injections to the gingiva during a week interval from the age of 8 to 11 weeks (LPS group). Lidocaine (3 mg/kg), LPS + lidocaine (3 mg/kg), LPS + lidocaine (1.5 mg/kg), and LPS + lidocaine (3 mg/kg, IP) groups simultaneously received gingival 1.5 or 3 mg/kg or IP 3 mg/kg injection of lidocaine on the same schedule as the gingival LPS. Isolated aortas or mandibles were subjected to the evaluation of histopathologic change, isometric force recording, reactive oxygen species, and Western immunoblotting. Mean blood pressure and heart rate did not differ among the control, LPS, LPS + lidocaine (3 mg/kg), and lidocaine (3 mg/kg) groups. LPS application reduced acetylcholine (ACh, 10 to 10 mol/L)-induced relaxation (29% difference at ACh 3 × 10 mol/L, P = .01), which was restored by catalase. Gingival lidocaine (1.5 and 3 mg/kg) dose dependently prevented the endothelial dysfunction caused by LPS application (24.5%-31.1% difference at ACh 3 × 10 mol/L, P = .006 or .001, respectively). Similar to the gingival application, the IP injection of lidocaine (3 mg/kg) restored the ACh-induced dilation of isolated aortas from rats with the LPS application (27.5% difference at ACh 3 × 10 mol/L, P lidocaine (3 mg/kg), or the combination. The LPS induced a 4-fold increase in the protein expression of tumor necrosis factor-α in the periodontal tissue (P lidocaine (3 mg/kg) coadministration partly reduced the levels. Lidocaine application also decreased

  6. Preeclampsia Is Associated with Increased Central Aortic Pressure, Elastic Arteries Stiffness and Wave Reflections, and Resting and Recruitable Endothelial Dysfunction

    Directory of Open Access Journals (Sweden)

    Juan Torrado

    2015-01-01

    Full Text Available Introduction. An altered endothelial function (EF could be associated with preeclampsia (PE. However, more specific and complementary analyses are required to confirm this topic. Flow-mediated dilation (FMD, low-flow-mediated constriction (L-FMC, and hyperemic-related changes in carotid-radial pulse wave velocity (PWVcr offer complementary information about “recruitability” of EF. Objectives. To evaluate, in healthy and hypertensive pregnant women (with and without PE, central arterial parameters in conjunction with “basal and recruitable” EF. Methods. Nonhypertensive (HP and hypertensive pregnant women (gestational hypertension, GH; preeclampsia, PE were included. Aortic blood pressure (BP, wave reflection parameters (AIx@75, aortic pulse wave velocity (PWVcf and PWVcr, and brachial and common carotid stiffness and intima-media thickness were measured. Brachial FMD and L-FMC and hyperemic-related change in PWVcr were measured. Results. Aortic BP and AIx@75 were elevated in PE. PE showed stiffer elastic but not muscular arteries. After cuff deflation, PWVcr decreased in HP, while GH showed a blunted PWVcr response and PE showed a tendency to increase. Maximal FMD and L-FMC were observed in HP followed by GH; PE did not reach significant arterial constriction. Conclusion. Aortic BP and wave reflections as well as elastic arteries stiffness are increased in PE. PE showed both “resting and recruitable” endothelial dysfunctions.

  7. Epidermal Growth Factor Rescues Endothelial Dysfunction in Primary Human Tissues In Vitro.

    Science.gov (United States)

    Hastie, Roxanne; Tong, Stephen; Hannan, Natalie J; Brownfoot, Fiona; Cannon, Ping; Kaitu'u-Lino, Tu'uhevaha J

    2017-09-01

    Preeclampsia is a hypertensive disorder of pregnancy, responsible for over 60 000 maternal deaths annually. Endothelial dysfunction is a central aspect to its pathophysiology, and currently, no medical therapeutic is available for its treatment. In this study, we aim to investigate the effect of epidermal growth factor (EGF) on endothelial dysfunction using primary human tissues. We performed a number of in vitro assays that mimic the vascular endothelial dysfunction that occurs in preeclampsia. Epidermal growth factor reduced the expression of vascular cell adhesion molecule-1, a marker of endothelial dysfunction, after insult with tumor necrosis factor α (TNF-α) or serum from women with preeclampsia. Additionally, after TNF-α insult, EGF reduced tube disruption and the adhesion of monocytes to primary human umbilical vein endothelial cells (HUVECs). Our findings suggest that EGF reduces endothelial dysfunction in primary HUVECs. Epidermal growth factor may have potential as a novel peptide treatment for preeclampsia and other diseases where there is endothelial dysfunction.

  8. CLINICAL IMPORTANCE OF ENDOTHELIAL DYSFUNCTION AND INSULIN RESISTANCE SYNDROME IN PATIENTS WITH GOUT ASSOCIATED WITH ARTERIAL HYPERTENSION

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    N. N. Kushnarenko

    2015-09-01

    Full Text Available Aim. To study the endothelium status and determine the correlation between endothelial dysfunction and glucose metabolism in men with gout associated with arterial hypertension (HT.Material and methods. Patients (n=175, all are males with gout were enrolled into the study. Ambulatory blood pressure monitoring (ABPM was performed in all patients. Endothelial function was studied in tests with reactive hyperemia (endothelium-dependent reaction and nitroglycerin (endothelium independent reaction in brachial artery by ultrasonic Doppler examination. The level of nitrite-nitrate and endothelin-1 in blood serum was determined by ELISA technique. Fasting blood glucose and oral glucose tolerance tests were performed as well as fasting insulin blood level was determined by immunoenzyme method. Insulin-resistance index (HOMA-IR was calculated. Patients with HOMA- IR>2.77 were considered as insulin-resistant.Results. Patients with gout demonstrated endothelial deterioration associated with activation of nitroxid producing function, elevation in endothelin-1 serum level (1.36 fmol/ml [0.91; 2.32 fmol/ml] vs 0.19 fmol/ml [0.16; 0.27 fmol/ml] in controls, p<0.05 and impairments of endothelium-dependent vasodilation (6.4% [3.3; 7.3%] vs 17.8% [12.7; 23.9%] in controls, p<0.05. The revealed changes were the most marked in patients with gout associated with HT. The correlation between some endothelial dysfunction in- dices and glucose metabolism was observed.Conclusion. ABPM, brachial artery endothelium-dependent vasodilation and glucose metabolism status should be studied in patients with gout. Complex treatment of cardiovascular diseases in patients with gout should include ω-3 polyunsaturated fatty acids, angiotensin receptor antagonists should be used for antihypertensive therapy.

  9. CLINICAL IMPORTANCE OF ENDOTHELIAL DYSFUNCTION AND INSULIN RESISTANCE SYNDROME IN PATIENTS WITH GOUT ASSOCIATED WITH ARTERIAL HYPERTENSION

    Directory of Open Access Journals (Sweden)

    N. N. Kushnarenko

    2013-01-01

    Full Text Available Aim. To study the endothelium status and determine the correlation between endothelial dysfunction and glucose metabolism in men with gout associated with arterial hypertension (HT.Material and methods. Patients (n=175, all are males with gout were enrolled into the study. Ambulatory blood pressure monitoring (ABPM was performed in all patients. Endothelial function was studied in tests with reactive hyperemia (endothelium-dependent reaction and nitroglycerin (endothelium independent reaction in brachial artery by ultrasonic Doppler examination. The level of nitrite-nitrate and endothelin-1 in blood serum was determined by ELISA technique. Fasting blood glucose and oral glucose tolerance tests were performed as well as fasting insulin blood level was determined by immunoenzyme method. Insulin-resistance index (HOMA-IR was calculated. Patients with HOMA- IR>2.77 were considered as insulin-resistant.Results. Patients with gout demonstrated endothelial deterioration associated with activation of nitroxid producing function, elevation in endothelin-1 serum level (1.36 fmol/ml [0.91; 2.32 fmol/ml] vs 0.19 fmol/ml [0.16; 0.27 fmol/ml] in controls, p<0.05 and impairments of endothelium-dependent vasodilation (6.4% [3.3; 7.3%] vs 17.8% [12.7; 23.9%] in controls, p<0.05. The revealed changes were the most marked in patients with gout associated with HT. The correlation between some endothelial dysfunction in- dices and glucose metabolism was observed.Conclusion. ABPM, brachial artery endothelium-dependent vasodilation and glucose metabolism status should be studied in patients with gout. Complex treatment of cardiovascular diseases in patients with gout should include ω-3 polyunsaturated fatty acids, angiotensin receptor antagonists should be used for antihypertensive therapy.

  10. Assessment of pulmonary endothelial function during invasive testing in children and adolescents with idiopathic pulmonary arterial hypertension.

    Science.gov (United States)

    Apitz, Christian; Zimmermann, Rainer; Kreuder, Joachim; Jux, Christian; Latus, Heiner; Pons-Kühnemann, Joern; Kock, Ines; Bride, Peter; Kreymborg, Karsten Grosse; Michel-Behnke, Ina; Schranz, Dietmar

    2012-07-10

    The purpose of our study was to assess pulmonary endothelial function by vasodilator response to acetylcholine (Ach) administered in segmental pulmonary arteries in children with idiopathic pulmonary arterial hypertension (IPAH). We hypothesized that there was a relationship among pulmonary endothelial response to Ach, severity of the disease, and clinical outcome. IPAH may be associated with pulmonary endothelial dysfunction; however, data regarding the impact of endothelial dysfunction on severity and prognosis of this disease are limited. Forty-three children and adolescents (mean age: 10.4 ± 5.5 years) with IPAH were included in the study. Changes in pulmonary blood flow in response to Ach were determined using intravascular Doppler flow measurements. Pulmonary flow reserve (PFR) was calculated as the ratio of pulmonary blood flow velocity in response to Ach relative to baseline values. Mean PFR of all patients was 1.58 ± 0.67. Mean follow-up after catheterization was 55.7 ± 41.9 months. Freedom from serious cardiovascular events (lung transplantation or death) was 83% after 2 years, 76% after 3 years, and 57% after 5 years. PFR was related significantly to World Health Organization functional class. Receiver-operating characteristic curves revealed a PFR of 1.4 as the best cutoff value. Kaplan-Meier analysis demonstrated that a PFR of pulmonary endothelial response to Ach and prognosis of children with IPAH. As an adjunct to the usual testing protocol, this method provides additional information for therapeutic guidance. Copyright © 2012 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.

  11. Fourier transform method to determine human corneal endothelial morphology

    Science.gov (United States)

    Masters, Barry R.; Lee, Yim-Kul; Rhodes, William T.

    1991-08-01

    The statistical evaluation of the size, shape, density and regularity of the cells in the human corneal endothelium is an important diagnostic technique. A method based on the Fourier transform of the cell boundaries was developed which can yield these statistical properties. The development of a hybrid optical/digital technique to obtain these statistical perimeters is our goal. The input images were tracings of human endothelial cell patterns. The optical Fourier transform of each image was obtained, and the radial projection and the angular correlation function were plotted versus distance and angle respectively. The average size of the cells was obtained from the first peak of the radial projection. The width of this peak is related to the coefficient of variation of the average cell size. The separation of the peaks in the normalized angular correlation plot is related to cell shape. This method is suitable for rapid analysis of large numbers of endothelial cell images. This technique may have potential for diagnostic ophthalmology.

  12. Endothelial and Neuronal Nitric Oxide Activate Distinct Pathways on Sympathetic Neurotransmission in Rat Tail and Mesenteric Arteries.

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    Joana Beatriz Sousa

    Full Text Available Nitric oxide (NO seems to contribute to vascular homeostasis regulating neurotransmission. This work aimed at assessing the influence of NO from different sources and respective intracellular pathways on sympathetic neurotransmission, in two vascular beds. Electrically-evoked [3H]-noradrenaline release was assessed in rat mesenteric and tail arteries in the presence of NO donors or endothelial/neuronal nitric oxide synthase (NOS inhibitors. The influence of NO on adenosine-mediated effects was also studied using selective antagonists for adenosine receptors subtypes. Location of neuronal NOS (nNOS was investigated by immunohistochemistry (with specific antibodies for nNOS and for Schwann cells and Confocal Microscopy. Results indicated that: 1 in mesenteric arteries, noradrenaline release was reduced by NO donors and it was increased by nNOS inhibitors; the effect of NO donors was only abolished by the adenosine A1 receptors antagonist; 2 in tail arteries, noradrenaline release was increased by NO donors and it was reduced by eNOS inhibitors; adenosine receptors antagonists were devoid of effect; 3 confocal microscopy showed nNOS staining in adventitial cells, some co-localized with Schwann cells. nNOS staining and its co-localization with Schwann cells were significantly lower in tail compared to mesenteric arteries. In conclusion, in mesenteric arteries, nNOS, mainly located in Schwann cells, seems to be the main source of NO influencing perivascular sympathetic neurotransmission with an inhibitory effect, mediated by adenosine A1 receptors activation. Instead, in tail arteries endothelial NO seems to play a more relevant role and has a facilitatory effect, independent of adenosine receptors activation.

  13. Markers of inflammation, endothelial activation, and arterial stiffness in hypertensive heart disease and the effects of treatment: results from the SILVHIA study.

    Science.gov (United States)

    Jekell, Andreas; Malmqvist, Karin; Wallén, N Håkan; Mörtsell, David; Kahan, Thomas

    2013-12-01

    We assessed the contribution of blood pressure (BP), inflammation, and endothelial activation to the development of structural vascular and cardiac changes in hypertension. Furthermore, the effects of antihypertensive therapy were studied. We studied 114 patients with hypertension and left ventricular hypertrophy and 38 matched hypertensive subjects without cardiac hypertrophy and 38 normotensive subjects. The group with hypertension and cardiac hypertrophy were randomized to treatment with an angiotensin receptor blocker (irbesartan) or a beta-adrenergic receptor blocker (atenolol) for 48 weeks. Markers of inflammation (high-sensitive C-reactive protein, interleukin-6, leukocyte counts), vascular function (ambulatory aortic stiffness index, arterial compliance, and pulse pressure), and endothelial activation (E-selectin, intracellular adhesion molecule-1, vascular adhesion molecule-1) were assessed. Markers of inflammation and arterial stiffness were lowest in the normotensive group and highest in patients with hypertensive heart disease; endothelial markers were similar between groups. Inflammation was independently related to BP. Markers of arterial stiffness were independently related to BP and to a lesser extent to left ventricular mass. Antihypertensive treatment improved arterial compliance; inflammatory and endothelial markers remained unchanged. In conclusion, markers of inflammation and arterial stiffness are independently related to BP. Antihypertensive therapy seems to improve arterial stiffness, but effects on markers of inflammation and endothelial activation are small.

  14. HIV-1/cocaine induced oxidative stress disrupts tight junction protein-1 in human pulmonary microvascular endothelial cells: role of Ras/ERK1/2 pathway.

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    Pranjali Dalvi

    Full Text Available Intravenous drug use (IVDU is the major risk factor in the development of HIV-related pulmonary arterial hypertension (HRPAH; however, the pathogenesis of HRPAH in association with IVDU has yet to be characterized. Endothelial injury is considered to be an initiating factor for pulmonary vascular remodeling in animal models of PAH. Our previous study shows that simultaneous exposure to HIV-Trans-activator of transcription (Tat and cocaine exacerbates both disruption of tight junction proteins and permeability of human pulmonary artery endothelial cells compared with either treatment alone. We here now demonstrate that this HIV-Tat and cocaine mediated endothelial dysfunction accompanies with increase in hydrogen peroxide and superoxide radicals generation and involves redox sensitive signaling pathway. Pretreatment with antioxidant cocktail attenuated the cocaine and Tat mediated disassembly of Zonula Occludens (ZO-1 and enhancement of endothelial monolayer permeability. Furthermore, inhibition of NADPH oxidase by apocynin or siRNA-mediated knockdown of gp-91(phox abolished the Tat/cocaine-induced reactive oxygen species (ROS production, suggesting the NADPH oxidase mediated generation of oxidative radicals. In addition, ROS dependent activation of Ras and ERK1/2 Kinase was observed to be mediating the TJP-1 disassembly, and endothelial dysfunction in response to cocaine and Tat exposure. In conclusion, our findings demonstrate that Tat/cocaine -mediated production of ROS activate Ras/Raf/ERK1/2 pathway that contributes to disruption of tight junction protein leading to pulmonary endothelial dysfunction associated with pulmonary vascular remodeling.

  15. The effect of insulin to decrease neointimal growth after arterial injury is endothelial nitric oxide synthase-dependent.

    Science.gov (United States)

    Guo, June; Breen, Danna M; Pereira, Troy J; Dalvi, Prasad S; Zhang, Hangjun; Mori, Yusaku; Ghanim, Husam; Tumiati, Laura; Fantus, I George; Bendeck, Michelle P; Dandona, Paresh; Rao, Vivek; Dolinsky, Vernon W; Heximer, Scott P; Giacca, Adria

    2015-07-01

    In vitro, insulin has mitogenic effects on vascular smooth muscle cells (VSMC) but also has protective effects on endothelial cells by stimulating nitric oxide (NO) production and endothelial nitric oxide synthase (eNOS) expression. Furthermore, NOS inhibition attenuates the effect of insulin to inhibit VSMC migration in vitro. Using an in vivo model, we have previously shown that insulin decreases neointimal growth and cell migration and increases re-endothelialization after arterial injury in normal rats. Since insulin can stimulate NOS, and NO can decrease neointimal growth, we hypothesized that NOS, and more specifically eNOS was required for the effects of insulin in vivo. Rats were given subcutaneous insulin implants (3 U/day) alone or with the NOS inhibitor l-NAME (2 mg kg(-1) day(-1)) 3 days before arterial (carotid or aortic) balloon catheter injury. Insulin decreased both neointimal area (P < 0.01) and cell migration (P < 0.01), and increased re-endothelialization (P < 0.05). All of these effects were prevented by the co-administration of l-NAME. Insulin was found to decrease inducible NOS expression (P < 0.05) but increase eNOS phosphorylation (P < 0.05). These changes were also translated at the functional level where insulin improved endothelial-dependent vasorelaxation. To further study the NOS isoform involved in insulin action, s.c. insulin (0.1 U/day) was given to wild-type and eNOS knockout mice. We found that insulin was effective at decreasing neointimal formation in wild-type mice after wire injury of the femoral artery, whereas this effect of insulin was absent in eNOS knockout mice. These results show that the vasculoprotective effect of insulin after arterial injury is mediated by an eNOS-dependent mechanism. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  16. Interaction of Tissue Engineering Substrates with Serum Proteins and Its Influence on Human Primary Endothelial Cells.

    Science.gov (United States)

    Mohan, Tamilselvan; Niegelhell, Katrin; Nagaraj, Chandran; Reishofer, David; Spirk, Stefan; Olschewski, Andrea; Stana Kleinschek, Karin; Kargl, Rupert

    2017-02-13

    Polymer-based biomaterials particularly polycaprolactone (PCL) are one of the most promising substrates for tissue engineering. The surface chemistry of these materials plays a major role since it governs protein adsorption, cell adhesion, viability, degradation, and biocompatibility in the first place. This study correlates the interaction of the most abundant serum proteins (albumin, immunoglobulins, fibrinogen) with the surface properties of PCL and its influence on the morphology and metabolic activity of primary human arterial endothelial cells that are seeded on the materials. Prior to that, thin films of PCL are manufactured by spin-coating and characterized in detail. A quartz crystal microbalance with dissipation (QCM-D), a multiparameter surface plasmon resonance spectroscopy instrument (MP-SPR), wettability data, and atomic force microscopy are combined to elucidate the pH-dependent protein adsorption on the PCL substrates. Primary endothelial cells are cultured on the protein modified polymer, and conclusions are drawn on the significant impact of type and form of proteins coatings on cell morphology and metabolic activity.

  17. Nitric oxide scavenging causes remodeling of the endoplasmic reticulum, Golgi apparatus and mitochondria in pulmonary arterial endothelial cells.

    Science.gov (United States)

    Lee, Jason E; Yuan, Huijuan; Liang, Feng-Xia; Sehgal, Pravin B

    2013-09-01

    The dependence of the structure and function of cytoplasmic organelles in endothelial cells on constitutively produced intracellular nitric oxide (NO) remains largely unexplored. We previously reported fragmentation of the Golgi apparatus in cells exposed to NO scavengers or after siRNA-mediated knockdown of eNOS. Others have reported increased mitochondrial fission in response to an NO donor. Functionally, we previously reported that bovine pulmonary arterial endothelial cells (PAECs) exposed to the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO) developed a prosecretory phenotype characterized by prolonged secretion of soluble proteins. In the present study, we investigated whether NO scavenging led to remodeling of the endoplasmic reticulum (ER). Live-cell DAF-2DA imaging confirmed the presence of intracellular NO in association with the BODIPY C5-ceramide-labeled Golgi apparatus. Untreated human PAECs displayed a pattern of peripheral tubulo-reticular ER with a juxtanuclear accumulation of ER sheets. Cells exposed to c-PTIO showed a dramatic increase in ER sheets as assayed using immunofluorescence for the ER structural protein reticulon-4b/Nogo-B and the ER-resident GTPase atlastin-3, live-cell fluorescence assays using RTN4-GFP and KDEL-mCherry, and electron microscopy methods. These ER changes were inhibited by the NO donor diethylamine NONOate, and also produced by L-NAME, but not D-NAME or 8-br-cGMP. This ER remodeling was accompanied by Golgi fragmentation and increased fibrillarity and function of mitochondria (uptake of tetramethyl-rhodamine, TMRE). Despite Golgi fragmentation the functional ER/Golgi trafficking unit was preserved as seen by the accumulation of Sec31A ER exit sites adjacent to the dispersed Golgi elements and a 1.8-fold increase in secretion of soluble cargo. Western blotting and immunopanning data showed that RTN4b was increasingly ubiquitinated following c-PTIO exposure, especially in the

  18. Cytotoxicity of Voriconazole on Cultured Human Corneal Endothelial Cells▿

    Science.gov (United States)

    Han, Sang Beom; Shin, Young Joo; Hyon, Joon Young; Wee, Won Ryang

    2011-01-01

    The purpose of the present study was to evaluate the toxicity of voriconazole on cultured human corneal endothelial cells (HCECs). HCECs were cultured and exposed to various concentrations of voriconazole (5.0 to 1,000 μg/ml). Cell viability was measured using a Cell Counting Kit-8 (CCK-8) and live/dead viability/cytotoxicity assays. Cell damage was assessed using phase-contrast microscopy after 24 h of exposure to voriconazole. To analyze the effect of voriconazole on the intercellular barrier, immunolocalization of zonula occludens 1 (ZO1) was performed. A flow cytometric assay was performed to evaluate the apoptotic and necrotic effects of voriconazole on HCECs. Cytotoxicity tests demonstrated the dose-dependent toxic effect of voriconazole on HCECs. Voriconazole concentrations of ≥100 μg/ml led to a significant reduction in cell viability. The morphological characteristics of HCECs also changed in a dose-dependent manner. Increasing concentrations of voriconazole resulted in fading staining for ZO1. Higher concentrations of voriconazole resulted in an increased number of propidium iodide (PI)-positive cells, indicating activation of the proapoptotic pathway. In conclusion, voriconazole may have a dose-dependent toxic effect on cultured HCECs. The results of this study suggest that although voriconazole concentrations of up to 50 μg/ml do not decrease cell viability, intracameral voriconazole concentrations of ≥100 μg/ml may increase the risk of corneal endothelial damage. PMID:21768517

  19. Determination of Early and Late Endothelial Progenitor Cells in Peripheral Circulation and Their Clinical Association with Coronary Artery Disease

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    Shotoku Tagawa

    2015-01-01

    Full Text Available The clinical implications of early and late endothelial progenitor cells (EPCs in coronary artery disease (CAD remain unclear. We investigated endothelial dysfunction in CAD by simultaneously examining early and late EPC colony formation and gene expression of specific surface markers in EPCs. EPCs were extracted from a total of 83 subjects with (n=47 and without (n=36 CAD. Early and late EPC colonies were formed from mononuclear cells extracted from peripheral blood. We found that fewer early EPC colonies were produced in the CAD group (7.2 ± 3.l/well than those in the control group (12.4 ± 1.4/well, p<0.05, and more late EPC colonies were produced in the CAD group (0.8 ± 0.2/well than those in the control group (0.25 ± 0.02/well, p<0.05. In the CAD group, the relative expression of CD31 and KDR of early and late EPCs was lower than in the control group. These results demonstrate that CAD patients could have increased late EPC density and that early and late EPCs in CAD patients exhibited immature endothelial characteristics. We suggest that changes in EPC colony count and gene expression of endothelial markers may have relation with development of CAD.

  20. Effects of the dual TP receptor antagonist and thromboxane synthase inhibitor EV-077 on human endothelial and vascular smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Petri, Marcelo H. [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden); Tellier, Céline; Michiels, Carine [NARILIS, URBC, University of Namur, Namur (Belgium); Ellertsen, Ingvill [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden); Dogné, Jean-Michel [Department of Pharmacy, Namur Thrombosis and Hemostasis Center, University of Namur, Namur (Belgium); Bäck, Magnus, E-mail: Magnus.Back@ki.se [Department of Medicine, Karolinska Institutet and Center for Molecular Medicine, Karolinska University Hospital, Stockholm (Sweden)

    2013-11-15

    Highlights: •EV-077 reduced TNF-α induced inflammation in endothelial cells. •The thromboxane mimetic U69915 enhanced vascular smooth muscle cell proliferation. •EV-077 inhibited smooth muscle cell proliferation. -- Abstract: The prothrombotic mediator thromboxane A{sub 2} is derived from arachidonic acid metabolism through the cyclooxygenase and thromboxane synthase pathways, and transduces its effect through the thromboxane prostanoid (TP) receptor. The aim of this study was to determine the effect of the TP receptor antagonist and thromboxane synthase inhibitor EV-077 on inflammatory markers in human umbilical vein endothelial cells and on human coronary artery smooth muscle cell proliferation. To this end, mRNA levels of different proinflammatory mediators were studied by real time quantitative PCR, supernatants were analyzed by enzyme immune assay, and cell proliferation was assessed using WST-1. EV-077 significantly decreased mRNA levels of ICAM-1 and PTX3 after TNFα incubation, whereas concentrations of 6-keto PGF1α in supernatants of endothelial cells incubated with TNFα were significantly increased after EV-077 treatment. Although U46619 did not alter coronary artery smooth muscle cell proliferation, this thromboxane mimetic enhanced the proliferation induced by serum, insulin and growth factors, which was significantly inhibited by EV-077. In conclusion, EV-077 inhibited TNFα-induced endothelial inflammation and reduced the enhancement of smooth muscle cell proliferation induced by a thromboxane mimetic, supporting that the thromboxane pathway may be associated with early atherosclerosis in terms of endothelial dysfunction and vascular hypertrophy.

  1. Pulmonary Artery Endothelial Cell Phenotypic Alterations in a Large Animal Model of Pulmonary Arteriovenous Malformations Following the Glenn Shunt

    Science.gov (United States)

    Kavarana, Minoo N.; Mukherjee, Rupak; Eckhouse, Shaina R.; Rawls, William F.; Logdon, Christina; Stroud, Robert E.; Patel, Risha K.; Nadeau, Elizabeth K.; Spinale, Francis G.; Graham, Eric M.; Forbus, Geoffrey A.; Bradley, Scott M.; Ikonomidis, John S.; Jones, Jeffrey A.

    2014-01-01

    Background: Longevity of the superior cavopulmonary connection (SCPC) is limited by the development of pulmonary arteriovenous malformations (PAVM). The goal of this study was to determine whether phenotypic changes in pulmonary artery endothelial cells (PAEC) that favor angiogenesis occur with PAVM formation. Methods: A superior vena cava to right pulmonary artery connection was constructed in 5 pigs. Pulmonary arteries were harvested at 6-8 weeks following surgery to establish cultures of PAEC and smooth muscle cells, to determine cell proliferation, gene expression, and tubule formation. Abundance of proteins related to angiogenesis was measured in lung tissue. Results: Contrast echocardiography revealed right-to-left shunting, consistent with PAVM formation. While the proliferation of smooth muscle cells from the right pulmonary artery (RPA) (shunted side) and left pulmonary artery (LPA) (non- shunted side) were similar, right PAEC proliferation was significantly higher. Expression profiles of genes encoding cellular signaling proteins were higher in PAECs from the RPA vs. LPA. Protein abundance of angiopoietin-1, and Tie-2 (angiopoietin receptor) were increased in the right lung (both pSCPC. This study found that PAVM development occurred concomitantly with differential changes in PAEC proliferative ability and phenotype. Moreover, there was a significant increase in the angiopoietin/Tie-2 complex in the right lung, which may provide novel therapeutic targets to attenuate PAVM formation following a SCPC. PMID:23968766

  2. 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.

  3. Successful transplantation of in vitro expanded human corneal endothelial precursors to corneal endothelial surface using a nanocomposite sheet

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    Parikumar P

    2011-01-01

    Full Text Available Background: Though the transplantation of in vitro expanded human corneal endothelial precursors in animal models of endothelial damage by injecting into the anterior chamber has been reported, the practical difficulties of accomplishing such procedure in human patients have been a hurdle to clinical translation. Here we report the successful transplantation of in vitro expanded human corneal precursor cells to an animal eye using a transparent Nano-composite sheet and their engraftment.Materials and Methods: Human Corneal endothelial cells (HCEC were isolated from human cadaver eyes with informed consent and expanded in the lab using a sphere forming assay in a novel Thermoreversible Gelation Polymer (TGP for 26 days. HCEC obtained by sphere forming assay were seeded in a novel Nano-composite sheet, which was made of PNIPA-NC gels by in-situ, free-radical polymerization of NIPA monomer in the presence of exfoliated clay (synthetic hectorite “Laponite XLG” uniformly dispersed in aqueous media. After a further seven days in vitro culture of HCEC in the Nano-composite sheet, cells were harvested and transplanted on cadaver-bovine eyes (n=3. The cells were injected between the corneal endothelial layer and the Nano-composite sheet that had been placed prior to the injection in close proximity to the endothelial layer. After three hours, the transplanted Nano-composite sheets were removed from the bovine eyes and subjected to microscopic examination. The corneas were subjected to Histo-pathological studies along with controls. Results: HCEC formed sphere like colonies in TGP which expressed relevant markers as confirmed by RT-PCR. Microscopic studies of the Nanosheets and histopathological studies of the cornea of the Bull’s eye revealed that the HCEC got engrafted to the corneal endothelial layer of the bovine eyes with no remnant cells in the Nanosheet. Conclusion: Transplantation of in vitro expanded donor human corneal endothelial cells

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

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    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.

  5. Components of the interleukin-6 transsignalling system are associated with the metabolic syndrome, endothelial dysfunction and arterial stiffness.

    Science.gov (United States)

    Weiss, Thomas W; Arnesen, Harald; Seljeflot, Ingebjorg

    2013-07-01

    The metabolic syndrome (MetS) is an increasing epidemiologic challenge and cardiovascular risk factor. Interleukin-6 (IL-6) is a cytokine that exerts its biological function via a complex orchestration of soluble and membrane bound receptors. We have investigated associations between IL-6 and its soluble receptors, soluble IL-6 receptor (sIL-6r) and soluble glycoprotein 130 (sGP130) and the metabolic syndrome. Furthermore, we have investigated possible associations with endothelial dysfunction and arterial stiffness. A total of 563 subjects were included in this study. The Adult Treatment Panel III criteria of the National Cholesterol Education Program were used for the definition of MetS. We used commercially available ELISA to analyse circulating levels of the markers. Pulse wave propagation time (PWP) was determined to assess arterial stiffness. The criteria for having MetS were filled by 221 subjects. sGP130, sIL-6r and IL-6 levels were elevated in subjects with MetS (p<0.05 for all markers), and are associated with increasing components of MetS. Particularly hypertriglyceridaemia, hypertension and fasting plasma glucose (FPG) seem to carry this association. sGP130 (p<0.01), IL-6 (p<0.05) and partially sIL-6r (p<0.05) correlated with markers of endothelial function (E-selectin, I-CAM-1, V-CAM-1) and inversely with PWP after adjustment for relevant covariates. sGP130, sIL-6r and IL-6 were significantly elevated in subjects with MetS. In addition, sGP130, IL-6 and partially sIL-6r were associated with markers of endothelial function and arterial stiffness. This finding sheds new light on the role of these inflammatory cytokines in subjects with MetS and the development and progression of clinically silent atherosclerosis. Copyright © 2013 Elsevier Inc. All rights reserved.

  6. The influence of propofol on P-selectin expression and nitric oxide production in re-oxygenated human umbilical vein endothelial cells.

    LENUS (Irish Health Repository)

    Corcoran, T B

    2012-02-03

    BACKGROUND: Reperfusion injury is characterized by free radical production and endothelial inflammation. Neutrophils mediate much of the end-organ injury that occurs, requiring P-selectin-mediated neutrophil-endothelial adhesion, and this is associated with decreased endothelial nitric oxide production. Propofol has antioxidant properties in vitro which might abrogate this inflammation. METHODS: Cultured human umbilical vein endothelial cells were exposed to 20 h of hypoxia and then returned to normoxic conditions. Cells were treated with saline, Diprivan 5 microg\\/l or propofol 5 microg\\/l for 4 h after re-oxygenation and were then examined for P-selectin expression and supernatant nitric oxide concentrations for 24 h. P-selectin was determined by flow cytometry, and culture supernatant nitric oxide was measured as nitrite. RESULTS: In saline-treated cells, a biphasic increase in P-selectin expression was demonstrated at 30 min (P = 0.01) and 4 h (P = 0.023) after re-oxygenation. Propofol and Diprivan prevented these increases in P-selectin expression (P < 0.05). Four hours after re-oxygenation, propofol decreased endothelial nitric oxide production (P = 0.035). CONCLUSION: This is the first study to demonstrate an effect of propofol upon endothelial P-selectin expression. Such an effect may be important in situations of reperfusion injury such as cardiac transplantation and coronary artery bypass surgery. We conclude that propofol attenuates re-oxygenation-induced endothelial inflammation in vitro.

  7. 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

  8. 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

  9. Endothelial lipase is highly expressed in macrophages in advanced human atherosclerotic lesions

    DEFF Research Database (Denmark)

    Bartels, Emil D; Nielsen, John E; Lindegaard, Marie Louise Skakkebæk

    2007-01-01

    Endothelial lipase (EL) is expressed in endothelial cells, and affects plasma lipoprotein metabolism by hydrolyzing phospholipids in HDL. To determine the cellular expression of EL mRNA and protein in human atherosclerotic lesions, we performed in situ hybridization and immunohistochemical studies...

  10. 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...

  11. Association of impaired endothelial glycocalyx with arterial stiffness, coronary microcirculatory dysfunction, and abnormal myocardial deformation in untreated hypertensives.

    Science.gov (United States)

    Ikonomidis, Ignatios; Voumvourakis, Astrinos; Makavos, George; Triantafyllidi, Helen; Pavlidis, George; Katogiannis, Konstantinos; Benas, Dimitris; Vlastos, Dimitris; Trivilou, Paraskevi; Varoudi, Maria; Parissis, John; Iliodromitis, Efstathios; Lekakis, John

    2018-03-02

    We investigated the association of endothelial glycocalyx damage with arterial stiffness, impairment of coronary microcirculatory function, and LV myocardial deformation in 320 untreated hypertensives and 160 controls. We measured perfused boundary region (PBR) of the sublingual microvessels, a marker inversely related with glycocalyx thickness, coronary flow reserve (CFR), and Global Longitudinal strain (GLS) by echocardiography, pulse wave velocity (PWV), and central systolic blood pressure (cSBP). Hypertensives had higher PBR, PWV cSBP, and lower CFR and GLS than controls (P < .05). In hypertensives, increased PBR was associated with increased cSBP, PWV, and decreased CFR and GLS after adjustment for age, sex, BMI, smoking LV mass, heart rate, hyperlipidemia, and office SBP (P < .05). PBR had an additive value to PWV, CFR, and office SBP for the prediction of abnormal GLS (x 2  = 2.4-3.8, P for change = .03). Endothelial glycocalyx is impaired in untreated hypertensives and is related to arterial stiffness, coronary, and myocardial dysfunction. ©2018 Wiley Periodicals, Inc.

  12. Evaluation of statin therapy on endothelial function in hypercholesterolemic rabbits by automatic measurement of arterial wall movement using ultrasound images.

    Science.gov (United States)

    Rahmani-Cherati, Tavoos; Mokhtari-Dizaji, Manijhe; Vajhi, Alireza; Rostami, Abdorrazzagh

    2014-10-01

    The aim of this study was to evaluate arterial endothelial function, assessed as acetylcholine-mediated dilation (AMD), in a hypercholesterolemic atherosclerotic rabbit model to investigate the effects of atorvastatin in the atherosclerotic process, using a new computerized analysis model and ultrasound images. Twenty-seven rabbits were fed a high-cholesterol (2%) diet for 6 wk and then divided into three groups for an additional 9 wk: Group A received regular chow food, group B received a 2% cholesterol-rich diet plus atorvastatin drug, and group C received regular chow food plus atorvastatin. Ultrasound examinations of endothelial function of the rabbit abdominal aorta artery were performed immediately after the 6 weeks (0 wk) and then 3, 6 and 9 wk after that. For off-line analysis, a computerized analysis method for evaluating instantaneous changes in the wall of the rabbit abdominal aorta was used. As parameters of improvement resulting from treatment, endothelium-dependent acetylcholine-induced dilation and endothelium-independent nitroglycerin-induced dilation were evaluated in treated rabbits. Differences among groups were tested using analysis of variance. On histopathology, intima-media thickness decreased after treatment in all groups. There were no significant differences in arterial diameter and blood velocity changes among treated rabbits at 0, 3, 6 and 9 wk of treatment in all groups, except in end-diastolic velocity, radial strain percentage, pulse index and resistance index in group C. In group A, AMD did not significantly improve after 3, 6 and 9 wk, as compared with 0 wk. Atorvastatin treatment significantly increased AMD (18%) at 3 wk in group B, compared with week 0. AMD significantly increased after 3 (26%), 6 (124%) and 9 (182%) wk in group C, compared with 0 wk. It is concluded that the new automatic method enables accurate and repeated evaluation of endothelial function during the progression and regression of atherosclerosis. Also, the

  13. Cannabidiol causes endothelium-dependent vasorelaxation of human mesenteric arteries via CB1 activation.

    Science.gov (United States)

    Stanley, Christopher P; Hind, William H; Tufarelli, Cristina; O'Sullivan, Saoirse E

    2015-09-01

    The protective effects of cannabidiol (CBD) have been widely shown in preclinical models and have translated into medicines for the treatment of multiple sclerosis and epilepsy. However, the direct vascular effects of CBD in humans are unknown. Using wire myography, the vascular effects of CBD were assessed in human mesenteric arteries, and the mechanisms of action probed pharmacologically. CBD-induced intracellular signalling was characterized using human aortic endothelial cells (HAECs). CBD caused acute, non-recoverable vasorelaxation of human mesenteric arteries with an Rmax of ∼ 40%. This was inhibited by cannabinoid receptor 1 (CB1) receptor antagonists, desensitization of transient receptor potential channels using capsaicin, removal of the endothelium, and inhibition of potassium efflux. There was no role for cannabinoid receptor-2 (CB2) receptor, peroxisome proliferator activated receptor (PPAR)γ, the novel endothelial cannabinoid receptor (CBe), or cyclooxygenase. CBD-induced vasorelaxation was blunted in males, and in patients with type 2 diabetes or hypercholesterolemia. In HAECs, CBD significantly reduced phosphorylated JNK, NFκB, p70s6 K and STAT5, and significantly increased phosphorylated CREB, ERK1/2, and Akt levels. CBD also increased phosphorylated eNOS (ser1177), which was correlated with increased levels of ERK1/2 and Akt levels. CB1 receptor antagonism prevented the increase in eNOS phosphorylation. This study shows, for the first time, that CBD causes vasorelaxation of human mesenteric arteries via activation of CB1 and TRP channels, and is endothelium- and nitric oxide-dependent. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Cardiology.

  14. Harmonic skeleton guided evaluation of stenoses in human coronary arteries.

    Science.gov (United States)

    Yang, Yan; Zhu, Lei; Haker, Steven; Tannenbaum, Allen R; Giddens, Don P

    2005-01-01

    This paper presents a novel approach that three-dimensionally visualizes and evaluates stenoses in human coronary arteries by using harmonic skeletons. A harmonic skeleton is the center line of a multi-branched tubular surface extracted based on a harmonic function, which is the solution of the Laplace equation. This skeletonization method guarantees smoothness and connectivity and provides a fast and straightforward way to calculate local cross-sectional areas of the arteries, and thus provides the possibility to localize and evaluate coronary artery stenosis, which is a commonly seen pathology in coronary artery disease.

  15. Platelet Endothelial Cell Adhesion Molecule-1 Gene Polymorphisms are Associated with Coronary Artery Lesions in the Chronic Stage of Kawasaki Disease.

    Science.gov (United States)

    Lu, Wen-Hsien; Huang, Sin-Jhih; Yuh, Yeong-Seng; Hsieh, Kai-Sheng; Tang, Chia-Wan; Liou, Huei-Han; Ger, Luo-Ping

    2017-05-01

    Kawasaki disease is the most common cause of pediatric acquired heart disease. The role of platelet endothelial cell adhesion molecule-1 in the inflammatory process has been documented. To date, no report has investigated the relationship between coronary artery lesions of Kawasaki disease and platelet endothelial cell adhesion molecule-1 polymorphisms. A total of 114 Kawasaki disease children with coronary artery lesions and 185 Kawasaki disease children without coronary artery lesions were recruited in this study. The TaqMan assay was conducted to identify the genotype in this case-control study. In three single nucleotide polymorphisms (Leu125Val, Ser563Asn, and Arg670Gly) of platelet endothelial cell adhesion molecule-1, we found that the Leu-Ser-Arg haplotype was associated with a significantly increased risk for coronary artery lesions in the chronic stage (odds ratio 3.05, 95% confidence interval 1.06-8.80, p = 0.039), but not for coronary artery lesions in the acute stage. Analysis based on the diplotypes of platelet endothelial cell adhesion molecule-1 also showed that Kawasaki disease with one or two alleles of Leu-Ser-Arg had a significantly increased risk of chronic coronary artery lesions (odds ratio 3.38, 95% confidence interval 1.11-10.28, p = 0.032) and had increased platelet counts after Kawasaki disease was diagnosed, as compared to those with other diplotypes. The haplotype of platelet endothelial cell adhesion molecule-1 Leu-Ser-Arg might be associated with the increased platelet counts and the following risk of chronic coronary artery lesions in a dominant manner in Kawasaki disease.

  16. Body Mass Index Is Associated With Microvascular Endothelial Dysfunction in Patients With Treated Metabolic Risk Factors and Suspected Coronary Artery Disease.

    Science.gov (United States)

    van der Heijden, Dirk J; van Leeuwen, Maarten A H; Janssens, Gladys N; Lenzen, Mattie J; van de Ven, Peter M; Eringa, Etto C; van Royen, Niels

    2017-09-14

    Obesity is key feature of the metabolic syndrome and is associated with high cardiovascular morbidity and mortality. Obesity is associated with macrovascular endothelial dysfunction, a determinant of outcome in patients with coronary artery disease. Here, we compared the influence of obesity on microvascular endothelial function to that of established cardiovascular risk factors such as diabetes mellitus, hypertension, hypercholesterolemia, and smoking in patients with suspected coronary artery disease. Endothelial function was assessed during postocclusive reactive hyperemia of the brachial artery and downstream microvascular beds in 108 patients who were scheduled for coronary angiography. In all patients, microvascular vasodilation was assessed using peripheral arterial tonometry; laser Doppler flowmetry and digital thermal monitoring were performed. Body mass index was significantly associated with decreased endothelium-dependent vasodilatation measured with peripheral arterial tonometry (r=0.23, P=0.02), laser Doppler flowmetry (r=0.30, Pmicrovascular vasodilatation. Especially in diabetic patients, endothelial function was not significantly reduced (control versus diabetes mellitus, mean±SEM or median [interquartile range], peripheral arterial tonometry: 1.90±0.20 versus 1.67±0.20, P=0.19, laser Doppler flowmetry: 728% [interquartile range, 427-1110] versus 589% [interquartile range, 320-1067] P=0.28, and digital thermal monitoring: 6.6±1.0% versus 2.5±1.7%, P=0.08). In multivariate linear regression analysis, body mass index was the only risk factor that significantly attenuated endothelium-dependent vasodilatation using all 3 microvascular function tests. Higher body mass index is associated with reduced endothelial function in patients with suspected coronary artery disease, even after adjustment for treated diabetes mellitus, hypertension, hypercholesterolemia, and smoking. © 2017 The Authors. Published on behalf of the American Heart Association

  17. Progesterone amplifies oxidative stress signal and promotes NO production via H2O2 in mouse kidney arterial endothelial cells.

    Science.gov (United States)

    Yuan, Xiao-Hua; Fan, Yang-Yang; Yang, Chun-Rong; Gao, Xiao-Rui; Zhang, Li-Li; Hu, Ying; Wang, Ya-Qin; Jun, Hu

    2016-01-01

    The role of progesterone on the cardiovascular system is controversial. Our present research is to specify the effect of progesterone on arterial endothelial cells in response to oxidative stress. Our result showed that H2O2 (150 μM and 300 μM) induced cellular antioxidant response. Glutathione (GSH) production and the activity of Glutathione peroxidase (GPx) were increased in H2O2-treated group. The expression of glutamate cysteine ligase catalytic subunit (GCLC) and modifier subunit (GCLM) was induced in response to H2O2. However, progesterone absolutely abolished the antioxidant response through increasing ROS level, inhibiting the activity of Glutathione peroxidase (GPx), decreasing GSH level and reducing expression of GClC and GCLM. In our study, H2O2 induced nitrogen monoxide (NO) production and endothelial nitric oxide synthase (eNOS) expression, and progesterone promoted H2O2-induced NO production. Progesterone increased H2O2-induced expression of hypoxia inducible factor-α (HIFα) which in turn regulated eNOS expression and NO synthesis. Further study demonstrated that progesterone increased H2O2 concentration of culture medium which may contribute to NO synthesis. Exogenous GSH decreased the content of H2O2 of culture medium pretreated by progesterone combined with H2O2 or progesterone alone. GSH also inhibited expression of HIFα and eNOS, and abolished NO synthesis. Collectively, our study demonstrated for the first time that progesterone inhibited cellular antioxidant effect and increased oxidative stress, promoted NO production of arterial endothelial cells, which may be due to the increasing H2O2 concentration and amplified oxidative stress signal. Copyright © 2015. Published by Elsevier Ltd.

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

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    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.

  19. 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.

  20. Influence of endothelial cell seeding on platelet deposition and patency in small-diameter Dacron arterial grafts

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    Allen, B.T.; Long, J.A.; Clark, R.E.; Sicard, G.A.; Hopkins, K.T.; Welch, M.J.

    1984-01-01

    Serial platelet deposition, surface topography, and patency were evaluated in control (N . 28) and endothelial cell-seeded (N . 28) small-diameter (4 mm inner diameter) USCI Dacron grafts implanted in the carotid and femoral arteries of dogs. All dogs received aspirin (325 mg) daily for 2 weeks starting 24 hours prior to graft implantation. Endothelial cell seeding was performed by mixing suspensions of autologous endothelial cells that had been enzymatically harvested from segments of external jugular vein with blood that was used to preclot the prostheses. The platelet deposition on each graft was quantitated by means of indium 111-labeled platelets and technetium 99m-labeled red cells in a dual-isotope platelet-imaging technique. Platelet deposition on seeded grafts 24 hours after implantation was significantly higher than on the controls (p less than 0.05). Two weeks after implantation platelet deposition on seeded prostheses had decreased to a level significantly lower than that on the controls and continued to decline on serial studies up to 7 months. In contrast to seeded grafts, platelet accumulation on control grafts dramatically increased after the withdrawal of aspirin therapy and was associated with a sharp rise in control graft thromboses. Cumulative 7-month patency for seeded prostheses was significantly higher than for the controls (96% and 29%, respectively; p less than 0.001). We conclude that endothelial cell seeding in combination with short-term aspirin therapy is a simple, reliable diameter Dacron prostheses. Abrupt withdrawal of aspirin therapy may be contraindicated in nonseeded control grafts because it results in increased platelet deposition and thrombosis.

  1. Establishment and long-term culture of human cystic fibrosis endothelial cells.

    Science.gov (United States)

    Plebani, Roberto; Tripaldi, Romina; Lanuti, Paola; Recchiuti, Antonio; Patruno, Sara; Di Silvestre, Sara; Simeone, Pasquale; Anile, Marco; Venuta, Federico; Prioletta, Marco; Mucilli, Felice; Del Porto, Paola; Marchisio, Marco; Pandolfi, Assunta; Romano, Mario

    2017-11-01

    Endothelial cell (EC) dysfunction has been reported in cystic fibrosis (CF) patients. Thus, the availability of CF EC is paramount to uncover mechanisms of endothelial dysfunction in CF. Using collagenase digestion, we isolated cells from small fragments of pulmonary artery dissected from non-CF lobes or explanted CF lungs. These cells were a heterogeneous mixture, containing variable percentages of EC. To obtain virtually pure pulmonary artery endothelial cells (PAEC), we developed an easy, inexpensive, and reliable method, based on the differential adhesion time of pulmonary artery cells collected after collagenase digestion. With this method, we obtained up to 95% pure non-CF and CF-PAEC. Moreover, we also succeed at immortalizing both PAEC and CF-PAEC, which remained viable and with unchanged phenotype and proliferation rate over the 30th passage. These cells recapitulated cystic fibrosis transmembrane conductance regulator expression and functions of the parental cells. Thus, we isolated for the first time endothelial cells from CF patients, providing a valuable tool to define the emerging role of EC in CF lung and vascular disease.

  2. 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

  3. Serum can overcome contact inhibition in confluent human pulmonary artery smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Victor Solodushko

    Full Text Available Pulmonary artery endothelial cells (PAEC in an intact vessel are continually exposed to serum, but unless injured, do not proliferate, constrained by confluence. In contrast, pulmonary artery smooth muscle cells (PASMC attain, and maintain, confluence in the presence of minimal serum, protected from serum's stimulatory effects except when the endothelial barrier becomes more permeable. We hypothesized therefore, that confluent PASMC may be less constrained by contact inhibition in the presence of serum than PAEC and tested this idea by exposing confluent non-transformed human PAEC and PASMC to media containing increasing concentrations of fetal bovine serum (FBS and determining cell growth over 7 days. PAEC that had attained confluence in low serum did not proliferate even when exposed to 5% serum, the highest concentration tested. In contrast, PASMC that attained confluence in low serum did proliferate once serum levels were increased, an effect that was dose dependent. Consistent with this observation, PASMC had more BrdU incorporation and a greater percentage of cells in S phase in 5% compared to 0.2% FBS, whereas no such difference was seen in PAEC. These results suggest that confluent human PAEC are resistant to the stimulatory effects of serum, whereas confluent PASMC can proliferate when serum levels are increased, an effect mediated in part by differences in phosphoinositide 3-kinase activation. This observation may be relevant to understanding the PASMC hyperplasia observed in humans and animals with pulmonary hypertension in which changes in endothelial permeability due to hypoxia or injury expose the underlying smooth muscle to serum.

  4. 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.

  5. In vivo serial MR imaging of magnetically labeled endothelial progenitor cells homing to the endothelium injured artery in mice.

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    Jun Chen

    Full Text Available BACKGROUND: Emerging evidence of histopathological analyses suggests that endothelial progenitor cells (EPCs play an important role in vascular diseases. Neointimal hyperplasia can be reduced by intravenous transfusion of EPCs after vascular injury in mice. Therefore, it would be advantageous to develop an in vivo technique that can explore the temporal and spatial migration of EPCs homing to the damaged endothelium noninvasively. METHODOLOGY/PRINCIPAL FINDINGS: The left carotid common artery (LCCA was injured by removal of endothelium with a flexible wire in Kunming mice. EPCs were collected by in vitro culture of spleen-derived mouse mononuclear cells (MNCs. EPCs labeling was carried out in vitro using Fe₂O₃-poly-L-lysine (Fe₂O₃-PLL. In vivo serial MR imaging was performed to follow-up the injured artery at different time points after intravenous transfusion of EPCs. Vessel wall areas of injured artery were computed on T₂WI. Larger MR signal voids of vessel wall on T₂WI was revealed in all 6 mice of the labeled EPC transfusion group 15 days after LCCA injury, and it was found only in 1 mouse in the unlabeled EPC transfusion group (p = 0.015. Quantitative analyses of vessel wall areas on T₂WI showed that the vessel wall areas of labeled EPC transfusion group were less than those of unlabeled EPC transfusion group and control group fifteen days after artery injury (p<0.05. Histopathological analyses confirmed accumulation and distribution of transfused EPCs at the injury site of LCCA. CONCLUSIONS/SIGNIFICANCE: These data indicate that MR imaging might be used as an in vivo method for the tracking of EPCs homing to the endothelium injured artery.

  6. Effect of pistachio nut consumption on endothelial function and arterial stiffness.

    Science.gov (United States)

    Kasliwal, Ravi R; Bansal, Manish; Mehrotra, Rahul; Yeptho, Kajal Pandya; Trehan, Naresh

    2015-05-01

    Previous studies have demonstrated beneficial effects of regular consumption of pistachio nuts on glycemic, lipid, and oxidative stress parameters. The aim of this study was to determine its effect on vascular health, which has not been adequately studied so far. In this open label, randomized parallel-group study, 60 adults with mild dyslipidemia were randomized to lifestyle modification (LSM) alone or LSM with consumption of 80 g (in-shell) pistachios (equivalent to 40 g or 1.5 oz shelled pistachios) daily for 3 mo. Biochemical parameters, brachial artery flow-mediated vasodilation (BAFMD), and carotid-femoral and brachial-ankle pulse wave velocity (cfPWV and baPWV, respectively) were measured before and after the intervention. At 3 mo, there was no change in any of the clinical or biochemical parameters in the LSM group. However, the patients in the pistachio group had a significant increase in high-density lipoprotein cholesterol (HDL-C; 35.7 ± 8.8 mg/dL versus 37.8 ± 10.1 mg/dL; P = 0.04) and a reduction in low-density lipoprotein cholesterol (137.2 ± 32.6 mg/dL versus 127.6 ± 34.0 mg/dL; P = 0.02), total cholesterol (TC)-to-HDL-C ratio (5.8 ± 1.3 mg/dL versus 5.3 ± 1.1 mg/dL; P = 0.001), and fasting blood sugar (88.8 ± 7.1 mg/dL versus 86.6 ± 6.3 mg/dL; P = 0.05). Additionally, whereas LSM alone was associated with no improvement in BAFMD or PWV, individuals in the pistachio group had significant reduction in left baPWV (1261.7 ± 187.5 cm/sec versus 1192.4 ± 152.5 cm/sec; P = 0.02) and statistically nonsignificant improvement in most other parameters, including BAFMD. As a result, at 3 mo the patients in the pistachio group had lower cfPWV (770.9 ± 96.5 cm/sec versus 846.4 ± 162.0 cm/sec; P = 0.08), lower left baPWV (1192.4 ± 152.5 cm/sec versus 1326.3 ± 253.7 cm/sec; P = 0.05), and lower average baPWV (1208.2 ± 118.4 cm/sec versus 1295.8 ± 194.1 cm/sec; P = 0.08) compared

  7. Adhesion Molecule Expression in Human Endothelial Cells under Simulated Microgravity

    Science.gov (United States)

    Rudimov, E. G.; Andreeva, E. R.; Buravkova, L. B.

    2013-02-01

    High gravisensitivity of endothelium is now well recognized. Therefore, the microgravity can be one of the main factors affecting the endothelium in space flight. In this work we studied the effects of gravity vector randomization (3D-clinorotation in RPM) on the viability of endothelial cells from human umbilical vein (HUVEC) and the expression of adhesion molecules on its surface. After RPM exposure, HUVEC conditioning medium was collected for cytokines evaluation, a part of vials was used for immunocytochemistry and other one - for cytofluorimetric analysis of ICAM-I, VCAM-I, PECAM-I, E-selectin, Endoglin, VE-cadherin expression. The viability of HUVEC and constitutive expression of EC marker molecules PECAM-I and Endoglin were similar in all experimental groups both after 6 and 24 hrs of exposure. There were no differences in ICAM-I and E-selectin expression on HUVEC in 3 groups after 6 hrs of exposure. 24 hrs incubation has provoked decrease in ICAM-I and E-selectin expression. Thus, gravity vector randomization can lead to the disruption of ECs monolayer.

  8. 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.

  9. A hot water extract of Curcuma longa inhibits adhesion molecule protein expression and monocyte adhesion to TNF-α-stimulated human endothelial cells.

    Science.gov (United States)

    Kawasaki, Kengo; Muroyama, Koutarou; Yamamoto, Norio; Murosaki, Shinji

    2015-01-01

    The recruitment of arterial leukocytes to endothelial cells is an important step in the progression of various inflammatory diseases. Therefore, its modulation is thought to be a prospective target for the prevention or treatment of such diseases. Adhesion molecules on endothelial cells are induced by proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), and contribute to the recruitment of leukocytes. In the present study, we investigated the effect of hot water extract of Curcuma longa (WEC) on the protein expression of adhesion molecules, monocyte adhesion induced by TNF-α in human umbilical vascular endothelial cells (HUVECs). Treatment of HUVECs with WEC significantly suppressed both TNF-α-induced protein expression of adhesion molecules and monocyte adhesion. WEC also suppressed phosphorylation and degradation of nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (IκBα) induced by TNF-α in HUVECs, suggesting that WEC inhibits the NF-κB signaling pathway.

  10. Affective Measures of Anger and Hostility and Brachial Artery Endothelial Function during Mental Stress and Forearm Occlusion

    Science.gov (United States)

    2001-06-06

    cardiovascular diseases (Barefoot , Dahlstrom, & Williams, 1983; Shekelle, Gale, Ostfeld, & Paul, 1983). Funher, hostility predicts the severity of myocard ial... infarct ion and sudden cardiac death, in CAD patients (Powell & Thorenson. 1985), although results in this domain have been mixed. However, even in...POMS) Figure 4 · p =.08 REFERENCES Anderson, T.1. (1999). Assessment and treatment of endothelial dysfunction in humans. lournal of the American

  11. Body mass index is associated with microvascular endothelial dysfunction in patients with treated metabolic risk factors and suspected coronary artery disease

    NARCIS (Netherlands)

    D.J. Van Der Heijden (Dirk J.); M.A.H. van Leeuwen (Maarten); G.N. Janssens (Gladys N.); M.J. Lenzen (Mattie); P.M. van de Ven (Peter); E.C. Eringa (Etto ); N. van Royen (Niels)

    2017-01-01

    textabstractBackground--Obesity is key feature of the metabolic syndrome and is associated with high cardiovascular morbidity and mortality. Obesity is associated with macrovascular endothelial dysfunction, a determinant of outcome in patients with coronary artery disease. Here, we compared the

  12. Arterial stiffness, endothelial and cognitive function in subjects with type 2 diabetes in accordance with absence or presence of diabetic foot syndrome.

    Science.gov (United States)

    Tuttolomondo, Antonino; Casuccio, Alessandra; Guercio, Giovanni; Maida, Carlo; Del Cuore, Alessandro; Di Raimondo, Domenico; Simonetta, Irene; Di Bona, Danilo; Pecoraro, Rosaria; Della Corte, Vittoriano; Gulotta, Eliana; Gulotta, Gaspare; Pinto, Antonio

    2017-01-06

    Endothelial dysfunction is an early marker of cardiovascular disease so endothelial and arterial stiffness indexes are good indicators of vascular health. We aimed to assess whether the presence of diabetic foot is associated with arterial stiffness and endothelial function impairment. We studied 50 subjects with type 2 diabetes mellitus and diabetic foot syndrome (DFS) compared to 50 diabetic subjects without diabetic foot, and 53 patients without diabetes mellitus, by means of the mini mental state examination (MMSE) administered to evaluate cognitive performance. Carotid-femoral pulse wave velocity (PWV) and augmentation index (Aix) were also evaluated by Applanation tonometry (SphygmoCor version 7.1), and the RH-PAT data were digitally analyzed online by Endo-PAT2000 using reactive hyperemia index (RHI) values. In comparison to diabetic subjects without diabetic foot the subjects with diabetic foot had higher mean values of PWV, lower mean values of RHI, and lower mean MMSE. At multinomial logistic regression PWV and RHI were significantly associated with diabetic foot presence, whereas ROC curve analysis had good sensitivity and specificity in arterial PWV and RHI for diabetic foot presence. Pulse wave velocity and augmentation index, mean RHI values, and mean MMSE were effective indicators of diabetic foot. Future research could address these issues by means of longitudinal studies to evaluate cardiovascular event incidence in relation to arterial stiffness, endothelial and cognitive markers.

  13. 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.

  14. 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.

  15. Computational modeling of hypertensive growth in the human carotid artery

    Science.gov (United States)

    Sáez, Pablo; Peña, Estefania; Martínez, Miguel Angel; Kuhl, Ellen

    2014-06-01

    Arterial hypertension is a chronic medical condition associated with an elevated blood pressure. Chronic arterial hypertension initiates a series of events, which are known to collectively initiate arterial wall thickening. However, the correlation between macrostructural mechanical loading, microstructural cellular changes, and macrostructural adaptation remains unclear. Here, we present a microstructurally motivated computational model for chronic arterial hypertension through smooth muscle cell growth. To model growth, we adopt a classical concept based on the multiplicative decomposition of the deformation gradient into an elastic part and a growth part. Motivated by clinical observations, we assume that the driving force for growth is the stretch sensed by the smooth muscle cells. We embed our model into a finite element framework, where growth is stored locally as an internal variable. First, to demonstrate the features of our model, we investigate the effects of hypertensive growth in a real human carotid artery. Our results agree nicely with experimental data reported in the literature both qualitatively and quantitatively.

  16. 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...

  17. Peripheral Endothelial Function and Coronary Flow Velocity Reserve Are Not Associated in Women with Angina and No Obstructive Coronary Artery Disease

    DEFF Research Database (Denmark)

    Flintholm Raft, Kristoffer; Frestad, Daria; Michelsen, Marie Mide

    2017-01-01

    PURPOSE: We investigated whether impaired flow-mediated dilation (FMD) and plasma biomarkers reflecting endothelial dysfunction are associated with coronary microvascular dysfunction (CMD) in women with angina and no obstructive coronary artery disease (CAD). METHODS: Patients (n = 194) were...... randomly selected women with angina pectoris and no obstructive CAD (artery by high-resolution ultrasound. Coronary flow velocity reserve (CFVR) was assessed by transthoracic...... Doppler flow echocardiography (TTDE) of the left anterior descending artery during rest and high-dose dipyridamole infusion. CMD was defined as CFVR

  18. Endothelial Dysfunction After ST-segment Elevation Myocardial Infarction and Long-term Outcome: A Study With Reactive Hyperemia Peripheral Artery Tonometry.

    Science.gov (United States)

    Kandhai-Ragunath, Jasveen J; Doggen, Carine J M; Jørstad, Harald T; Doelman, Cees; de Wagenaar, Bjorn; IJzerman, Maarten J; Peters, Ron J G; von Birgelen, Clemens

    2016-07-01

    Long-term data on the relationship between endothelial dysfunction after ST-segment elevation myocardial infarction and future adverse clinical events are scarce. The aim of this study was to noninvasively assess whether endothelial dysfunction 4 weeks to 6 weeks after primary percutaneous coronary intervention for acute ST-segment elevation myocardial infarction predicts future clinical events. This prospective cohort study was performed in 70 patients of the RESPONSE randomized trial, who underwent noninvasive assessment of endothelial function 4 weeks to 6 weeks after primary percutaneous coronary intervention. Endothelial function was measured by the reactive hyperemia peripheral artery tonometry method; an index<1.67 identified endothelial dysfunction. The reactive hyperemia peripheral artery tonometry index measured on average 1.90±0.58. A total of 35 (50%) patients had endothelial dysfunction and 35 (50%) patients had normal endothelial function. Periprocedural "complications" (eg, cardiogenic shock, total atrioventricular block) were more common in patients with endothelial dysfunction than in those without (25.7% vs 2.9%; P<.01). During 4.0±1.7 years of follow-up, 20 (28.6%) patients had major adverse cardiovascular events: events occurred in 9 (25.7%) patients with endothelial dysfunction and in 11 (31.5%) patients with normal endothelial function (P=.52). There was an association between the prevalence of diabetes mellitus at baseline and the occurrence of major adverse cardiovascular events during follow-up (univariate analysis: hazard ratio=2.8; 95% confidence interval, 1.0-7.8; P<.05), and even in multivariate analyses the risk appeared to be increased, although not significantly (multivariate analysis: hazard ratio=2.5; 95% confidence interval, 0.8-7.5). In this series of patients who survived an ST-segment elevation myocardial infarction, endothelial dysfunction, as assessed by reactive hyperemia peripheral artery tonometry 4 weeks to 6 weeks

  19. Omega-3 fatty acids supplementation improves endothelial function and arterial stiffness in hypertensive patients with hypertriglyceridemia and high cardiovascular risk.

    Science.gov (United States)

    Casanova, Marcela A; Medeiros, Fernanda; Trindade, Michelle; Cohen, Célia; Oigman, Wille; Neves, Mario Fritsch

    2017-01-01

    Association between hypertriglyceridemia and cardiovascular (CV) disease is still controversial. The purpose of this study was to compare omega-3 and ciprofibrate effects on the vascular structure and function in low and high CV risk hypertensive patients with hypertriglyceridemia. Twenty-nine adults with triglycerides 150-499 mg/dL were divided into low (omega-3 fatty acids 1800 mg/d or ciprofibrate 100 mg/d for 12 weeks. Treatment was switched after 8-week washout. Clinical evaluation and vascular tests were assessed at baseline and after intervention. Peripheral (131 ± 3 to 125 ± 3 mm Hg, P omega-3. In conclusion, omega-3 improved arterial stiffness and endothelial function, pointing out the beneficial effect of this therapy on vascular aging, in high-risk patients. Copyright © 2016 American Society of Hypertension. Published by Elsevier Inc. All rights reserved.

  20. Bipolar disorder and related mood states are not associated with endothelial function of small arteries in adults without heart disease.

    Science.gov (United States)

    Tong, Brian; Abosi, Oluchi; Schmitz, Samantha; Myers, Janie; Pierce, Gary L; Fiedorowicz, Jess G

    2017-12-19

    Individuals with bipolar disorder are at increased risk for adverse cardiovascular disease (CVD) events. This study aimed to assess endothelial function and wave reflection, a risk factor for CVD, as measured by finger plethysmography in bipolar disorder to investigate whether CVD risk was higher in bipolar disorder and altered during acute mood episodes. We hypothesized that EndoPAT would detect a lower reactive hyperemia index (RHI) and higher augmentation index (AIX) in individuals with bipolar disorder compared with controls. Second, we predicted lower RHI and higher AIX during acute mood episodes. Reactive hyperemia index and augmentation index, measures of microvascular endothelial function and arterial pressure wave reflection respectively, were assessed using the EndoPAT 2000 device in a sample of 56 participants with a DSM-IV diagnosis of bipolar I disorder with 82 measures spanning different mood states (mania, depression, euthymia) and cross-sectionally in 26 healthy controls. RHI and AIX were not different between adults with and without bipolar disorder (mean age 40.3 vs. 41.2years; RHI: 2.04±0.67 vs. 2.05±0.51; AIX@75 (AIX adjusted for heart rate of 75): 1.4±19.7 vs. 0.8±22.4). When modeled in linear mixed models with a random intercept (to account for repeated observations of persons with bipolar disorder) and adjusting for age and sex, there were no significant differences between those with bipolar disorder and controls (p=0.89 for RHI; p=0.85 for AIX@75). Microvascular endothelial function and wave reflection estimated by finger plethysmography were unable to detect differences between adults with and without bipolar disorder or changes with mood states. Future research is necessary to identify more proximal and sensitive, yet relevant, biomarkers of abnormal mood-related influences on CVD risk or must target higher risk samples. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Endothelial nitric oxide gene polymorphisms, nitric oxide production and coronary artery disease risk in a South Indian population.

    Science.gov (United States)

    Angeline, T; Isabel, W; Tsongalis, Gregory J

    2010-12-01

    Nitric oxide (NO) synthesized by vascular endothelial cells, is a vasodilator agent produced from endothelial NO synthase (eNOS). It has been reported that decreased bioavailability of NO plays an important role in the development and progression of atherosclerosis. Electrocardiographically proven 100 patients with acute myocardial infarction and 100 age and sex matched healthy individuals with normal coronary arteries were included for the study. The genotypes of a 27-bp insertion/deletion in intron 4 (eNOS 4b/4a) and G894T polymorphism in exon 7, were determined by PCR analysis based on the banding pattern on gel electrophoresis. The genotype frequencies were calculated following the Hardy-Weinberg law. Serum NO level was also estimated by the Griess method. NO levels in AMI patients were higher than those of the healthy subjects (median [interquartile range], (14.36[12.42-15.78]) μM compared with 11.28[10.32-11.89]) μM; p<0.001; Mann-Whitney rank sum test, U=285. Mutant "T" allele frequency of the eNOS-G894T polymorphism was found to be comparatively higher (0.29) in AMI patients than among the controls (0.17). The calculated Odds ratio showed that the occurrence of mutant allele "T" was 1.6 fold as frequent in cases than controls [OR=1.6 (95%CI 0.898 to 2.833)]. To conclude, in the present study, (i) NO levels were found to be increased in patients than in controls, (ii) the homozygous mutant (TT) genotype confers genetic susceptibility to coronary artery disease (iii) both the eNOS 4a/b and G894T polymorphisms were not associated with serum NO levels in a South Indian Tamil population. Copyright © 2010 Elsevier Inc. All rights reserved.

  2. Evidence for a Role of Vascular Endothelium in the Control of Arterial Wall Viscosity in Humans.

    Science.gov (United States)

    Roca, Frederic; Iacob, Michele; Remy-Jouet, Isabelle; Bellien, Jeremy; Joannides, Robinson

    2018-01-01

    Arterial wall viscosity (AWV) is a major cause of energy dissipation along the arterial tree. Its determinants remain controversial but an active endothelial regulation has been suggested. Our objective was to assess in humans the physiological role of endothelium-derived nitric oxide (NO), epoxyeicosatrienoic acids and the effect of modulating smooth muscle tone in the regulation of AWV. We simultaneously measured radial artery diameter, wall thickness, and arterial pressure in healthy volunteers during the local infusion of inhibitors of NO-synthase ( N G -monomethyl-l-arginine), epoxyeicosatrienoic acids synthesis by cytochrome P450 (fluconazole), the epoxyeicosatrienoic acids cellular targets calcium-activated potassium channels (tetraethylammonium), alone and in combination. AWV was estimated from the relative viscosity expressed as the ratio of the area of the hysteresis loop of the pressure-diameter relationship to the area under the loading phase. Arterial tone was assessed by measuring change in wall stiffness and midwall stress. N G -monomethyl-l-arginine paradoxically reduced relative viscosity (34.9±8.9%-28.9±8.6%). Conversely, relative viscosity was not modified by fluconazole (33.5±15.5%-32.0±13.6%) but increased by tetraethylammonium (31.7±6.6%-35.7±8.0%). This increase was more marked with N G -monomethyl-l-arginine+fluconazole (31.1±10.7%-43.3±13.2%) and N G -monomethyl-l-arginine+tetraethylammonium (29.5±2.3%-41.5±11.1%) compared with inhibitors alone. Sodium nitroprusside decreased AWV (35.4±2.9%-28.7±2.0%). These effects were associated with parallel change in tone but of different magnitude for similar variations in viscosity, suggesting tone-dependent and independent mechanisms. In conclusion, this is the first demonstration that the endothelial factors, NO and epoxyeicosatrienoic acids, regulate AWV in humans and support the role of arterial tone in this regulation. URL: https://eudract.ema.europa.eu. Unique identifier: RCB2007-A

  3. Constitutively Expressed IFITM3 Protein in Human Endothelial Cells Poses an Early Infection Block to Human Influenza Viruses.

    Science.gov (United States)

    Sun, Xiangjie; Zeng, Hui; Kumar, Amrita; Belser, Jessica A; Maines, Taronna R; Tumpey, Terrence M

    2016-12-15

    A role for pulmonary endothelial cells in the orchestration of cytokine production and leukocyte recruitment during influenza virus infection, leading to severe lung damage, has been recently identified. As the mechanistic pathway for this ability is not fully known, we extended previous studies on influenza virus tropism in cultured human pulmonary endothelial cells. We found that a subset of avian influenza viruses, including potentially pandemic H5N1, H7N9, and H9N2 viruses, could infect human pulmonary endothelial cells (HULEC) with high efficiency compared to human H1N1 or H3N2 viruses. In HULEC, human influenza viruses were capable of binding to host cellular receptors, becoming internalized and initiating hemifusion but failing to uncoat the viral nucleocapsid and to replicate in host nuclei. Unlike numerous cell types, including epithelial cells, we found that pulmonary endothelial cells constitutively express a high level of the restriction protein IFITM3 in endosomal compartments. IFITM3 knockdown by small interfering RNA (siRNA) could partially rescue H1N1 virus infection in HULEC, suggesting IFITM3 proteins were involved in blocking human influenza virus infection in endothelial cells. In contrast, selected avian influenza viruses were able to escape IFITM3 restriction in endothelial cells, possibly by fusing in early endosomes at higher pH or by other, unknown mechanisms. Collectively, our study demonstrates that the human pulmonary endothelium possesses intrinsic immunity to human influenza viruses, in part due to the constitutive expression of IFITM3 proteins. Notably, certain avian influenza viruses have evolved to escape this restriction, possibly contributing to virus-induced pneumonia and severe lung disease in humans. Avian influenza viruses, including H5N1 and H7N9, have been associated with severe respiratory disease and fatal outcomes in humans. Although acute respiratory distress syndrome (ARDS) and progressive pulmonary endothelial damage

  4. Forkhead box O-1 modulation improves endothelial insulin resistance in human obesity.

    Science.gov (United States)

    Karki, Shakun; Farb, Melissa G; Ngo, Doan T M; Myers, Samantha; Puri, Vishwajeet; Hamburg, Naomi M; Carmine, Brian; Hess, Donald T; Gokce, Noyan

    2015-06-01

    Increased visceral adiposity has been closely linked to insulin resistance, endothelial dysfunction, and cardiometabolic disease in obesity, but pathophysiological mechanisms are poorly understood. We sought to investigate mechanisms of vascular insulin resistance by characterizing depot-specific insulin responses and gain evidence that altered functionality of transcription factor forkhead box O-1 (FOXO-1) may play an important role in obesity-related endothelial dysfunction. We intraoperatively collected paired subcutaneous and visceral adipose tissue samples from 56 severely obese (body mass index, 43 ± 7 kg/m(2)) and 14 nonobese subjects during planned surgical operations, and characterized depot-specific insulin-mediated responses using Western blot and quantitative immunofluorescence techniques. Insulin signaling via phosphorylation of FOXO-1 and consequent endothelial nitric oxide synthase stimulation was selectively impaired in the visceral compared with subcutaneous adipose tissue and endothelial cells of obese subjects. In contrast, tissue actions of insulin were preserved in nonobese individuals. Pharmacological antagonism with AS1842856 and biological silencing using small interfering RNA-mediated FOXO-1 knockdown reversed insulin resistance and restored endothelial nitric oxide synthase activation in the obese. We observed profound endothelial insulin resistance in the visceral adipose tissue of obese humans which improved with FOXO-1 inhibition. FOXO-1 modulation may represent a novel therapeutic target to diminish vascular insulin resistance. In addition, characterization of endothelial insulin resistance in the adipose microenvironment may provide clues to mechanisms of systemic disease in human obesity. © 2015 American Heart Association, Inc.

  5. Low concentrations of lithium and cyclooxygenase inhibitors enhance endothelin-1 (ET-1)-induced contractions in human temporal artery, but not in porcine ophthalmic artery.

    Science.gov (United States)

    Bakken, I J; Vincent, M B; White, L R; Cappelen, J; Skaanes, K O; Sjaastad, O

    1992-11-01

    Endothelins are a recently discovered group of potent vasoconstrictor peptides synthesized by endothelial cells and other tissues in various species, which seem to participate in the regulation of vascular tonus. Abnormalities in vasoactivity in the head may be an important event in headache pathophysiology, although the mechanisms responsible for such constrictions and/or dilations are not known. The endothelium and its constrictor peptide, endothelin, may play a key role in such mechanisms. Of the various drugs used in the treatment of headache, lithium is an accepted treatment for cluster headache, and indomethacin is the drug of choice for the associated condition chronic paroxysmal hemicrania. The mechanism of action of these drugs in these headaches is not known. Due to the possible involvement of endothelin in headache disorders, the objective of this study was to verify the effects of lithium and cyclooxygenase inhibitors (indomethacin, acetylsalicylic acid and naproxen) on endothelin-1 (ET-1)-induced contractions in isolated human temporal arteries and porcine ophthalmic arteries. It was found that all drugs increased the (ET-1)-induced contractions in human temporal arteries. Conversely, there were no significant changes induced by the drugs in porcine ophthalmic arteries. These results are consistent with the variation of activity often seen in different vascular beds and between species. The potential importance of such reactions for the understanding of vascular changes putatively involved in headache development and treatment is discussed.

  6. Substrates for Expansion of Corneal Endothelial Cells towards Bioengineering of Human Corneal Endothelium

    Directory of Open Access Journals (Sweden)

    Jesintha Navaratnam

    2015-09-01

    Full Text Available Corneal endothelium is a single layer of specialized cells that lines the posterior surface of cornea and maintains corneal hydration and corneal transparency essential for vision. Currently, transplantation is the only therapeutic option for diseases affecting the corneal endothelium. Transplantation of corneal endothelium, called endothelial keratoplasty, is widely used for corneal endothelial diseases. However, corneal transplantation is limited by global donor shortage. Therefore, there is a need to overcome the deficiency of sufficient donor corneal tissue. New approaches are being explored to engineer corneal tissues such that sufficient amount of corneal endothelium becomes available to offset the present shortage of functional cornea. Although human corneal endothelial cells have limited proliferative capacity in vivo, several laboratories have been successful in in vitro expansion of human corneal endothelial cells. Here we provide a comprehensive analysis of different substrates employed for in vitro cultivation of human corneal endothelial cells. Advances and emerging challenges with ex vivo cultured corneal endothelial layer for the ultimate goal of therapeutic replacement of dysfunctional corneal endothelium in humans with functional corneal endothelium are also presented.

  7. Human endothelial progenitor cells internalize high-density lipoprotein.

    Directory of Open Access Journals (Sweden)

    Kaemisa Srisen

    Full Text Available Endothelial progenitor cells (EPCs originate either directly from hematopoietic stem cells or from a subpopulation of monocytes. Controversial views about intracellular lipid traffic prompted us to analyze the uptake of human high density lipoprotein (HDL, and HDL-cholesterol in human monocytic EPCs. Fluorescence and electron microscopy were used to investigate distribution and intracellular trafficking of HDL and its associated cholesterol using fluorescent surrogates (bodipy-cholesterol and bodipy-cholesteryl oleate, cytochemical labels and fluorochromes including horseradish peroxidase and Alexa Fluor® 568. Uptake and intracellular transport of HDL were demonstrated after internalization periods from 0.5 to 4 hours. In case of HDL-Alexa Fluor® 568, bodipy-cholesterol and bodipy-cholesteryl oleate, a photooxidation method was carried out. HDL-specific reaction products were present in invaginations of the plasma membrane at each time of treatment within endocytic vesicles, in multivesicular bodies and at longer periods of uptake, also in lysosomes. Some HDL-positive endosomes were arranged in form of "strings of pearl"- like structures. HDL-positive multivesicular bodies exhibited intensive staining of limiting and vesicular membranes. Multivesicular bodies of HDL-Alexa Fluor® 568-treated EPCs showed multilamellar intra-vacuolar membranes. At all periods of treatment, labeled endocytic vesicles and organelles were apparent close to the cell surface and in perinuclear areas around the Golgi apparatus. No HDL-related particles could be demonstrated close to its cisterns. Electron tomographic reconstructions showed an accumulation of HDL-containing endosomes close to the trans-Golgi-network. HDL-derived bodipy-cholesterol was localized in endosomal vesicles, multivesicular bodies, lysosomes and in many of the stacked Golgi cisternae and the trans-Golgi-network Internalized HDL-derived bodipy-cholesteryl oleate was channeled into the lysosomal

  8. Endothelial Microparticles From Acute Coronary Syndrome Patients Induce Premature Coronary Artery Endothelial Cell Aging and Thrombogenicity: Role of the Ang II/AT1 Receptor/NADPH Oxidase-Mediated Activation of MAPKs and PI3-Kinase Pathways.

    Science.gov (United States)

    Abbas, Malak; Jesel, Laurence; Auger, Cyril; Amoura, Lamia; Messas, Nathan; Manin, Guillaume; Rumig, Cordula; León-González, Antonio J; Ribeiro, Thais P; Silva, Grazielle C; Abou-Merhi, Raghida; Hamade, Eva; Hecker, Markus; Georg, Yannick; Chakfe, Nabil; Ohlmann, Patrick; Schini-Kerth, Valérie B; Toti, Florence; Morel, Olivier

    2017-01-17

    Microparticles (MPs) have emerged as a surrogate marker of endothelial dysfunction and cardiovascular risk. This study examined the potential of MPs from senescent endothelial cells (ECs) or from patients with acute coronary syndrome (ACS) to promote premature EC aging and thrombogenicity. Primary porcine coronary ECs were isolated from the left circumflex coronary artery. MPs were prepared from ECs and venous blood from patients with ACS (n=30) and from healthy volunteers (n=4) by sequential centrifugation. The level of endothelial senescence was assessed as senescence-associated β-galactosidase activity using flow cytometry, oxidative stress using the redox-sensitive probe dihydroethidium, tissue factor activity using an enzymatic Tenase assay, the level of target protein expression by Western blot analysis, platelet aggregation using an aggregometer, and shear stress using a cone-and-plate viscometer. Senescence, as assessed by senescence-associated β-galactosidase activity, was induced by the passaging of porcine coronary artery ECs from passage P1 to P4, and was associated with a progressive shedding of procoagulant MPs. Exposure of P1 ECs to MPs shed from senescent P3 cells or circulating MPs from ACS patients induced increased senescence-associated β-galactosidase activity, oxidative stress, early phosphorylation of mitogen-activated protein kinases and Akt, and upregulation of p53, p21, and p16. Ex vivo, the prosenescent effect of circulating MPs from ACS patients was evidenced only under conditions of low shear stress. Depletion of endothelial-derived MPs from ACS patients reduced the induction of senescence. Prosenescent MPs promoted EC thrombogenicity through tissue factor upregulation, shedding of procoagulant MPs, endothelial nitric oxide synthase downregulation, and reduced nitric oxide-mediated inhibition of platelet aggregation. These MPs exhibited angiotensin-converting enzyme activity and upregulated AT1 receptors and angiotensin

  9. 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.

  10. Middle-Term Dietary Supplementation with Red Yeast Rice Plus Coenzyme Q10 Improves Lipid Pattern, Endothelial Reactivity and Arterial Stiffness in Moderately Hypercholesterolemic Subjects.

    Science.gov (United States)

    Cicero, Arrigo F G; Morbini, Martino; Rosticci, Martina; D''Addato, Sergio; Grandi, Elisa; Borghi, Claudio

    2016-01-01

    The aim of our study was to investigate whether treatment with red yeast rice added with Coenzyme Q10 is associated with changes in endothelial function and arterial stiffness. This double blind, placebo-controlled, randomized clinical trial was carried out on 40 non-smoker moderately hypercholesterolemic subjects (ClinicalTrial.gov ID NCT02492464). After 4 weeks of diet and physical activity, patients were allocated to treatment with placebo or with an active product containing 10 mg monacolins and 30 mg Coenzyme Q10, to be assumed for 6 months. Endothelial reactivity and arterial stiffness have been measured through the validated Vicorder® device. During monacolin treatment, patients experienced a more favorable percentage change in low density lipoprotein (LDL)-cholesterol (after monacolin treatment: -26.3%; after placebo treatment: +3.4%, p < 0.05). Endothelial reactivity (pulse volume displacement after monacolin treatment: +6.0%; after placebo treatment: -0.3%, p < 0.05), and arterial stiffness (pulse wave velocity (PWV) after monacolin treatment: -4.7%; after placebo: +1.1%, p < 0.05) also significantly improved only after monacolin treatment. The long-term assumption of the tested dietary supplement is associated with an improvement in LDL-cholesterolemia, endothelial reactivity and PWV in moderately hypercholesterolemic subjects. © 2016 S. Karger AG, Basel.

  11. Diabetes status differentiates endothelial function and plasma nitrite response to exercise stress in peripheral arterial disease following supervised training☆,☆☆

    Science.gov (United States)

    Allen, Jason D.; Stabler, Thomas; Kenjale, Aarti A.; Ham, Katherine L.; Robbins, Jennifer L.; Duscha, Brian D.; Kraus, William E.; Annex, Brian H

    2013-01-01

    Aims To determine if type 2 diabetes mellitus (T2D) differentiates endothelial function and plasma nitrite response (a marker of nitric oxide bioavailability) during exercise in peripheral arterial disease (PAD) subjects prior to and following 3 months supervised exercise training (SET). Methods In subjects with T2D + PAD (n = 13) and PAD-only (n = 14), endothelial function was measured using brachial artery flow-mediated dilation. On a separate day, venous blood draws were performed at rest and 10 min following a symptom-limited graded treadmill test (SL-GXT). Plasma samples were snap-frozen for analysis of nitrite by reductive chemiluminescence. All testing was repeated following 3 months of SET. Results Prior to training both groups demonstrated endothelial dysfunction, which was correlated with a net decrease in plasma nitrite following a SL-GXT (p ≤ 0.05). Following SET, the PAD-only group demonstrated an improvement in endothelial function (p ≤ 0.05) and COT (p ≤ 0.05), which was related to a net increase in plasma nitrite following the SL-GXT (both p ≤ 0.05). The T2D + PAD group had none of these increases. Conclusions T2D in the presence of PAD attenuated improvements in endothelial function, net plasma nitrite, and COT following SET. This suggests that T2D maybe associated with an inability to endogenously increase vascular NO bioavailability to SET. PMID:24355663

  12. Gas6 stimulates angiogenesis of human retinal endothelial cells and of zebrafish embryos via ERK1/2 signaling.

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    Young Sook Kim

    Full Text Available AIM: To determine if growth arrest-specific 6 (Gas6 plays an important role in the regulation of angiogenesis in human retinal microvascular endothelial cells (HRMECs and in vessel development of zebrafish. METHODS: Proliferation, wound-healing cell migration, and tube formation were measured in HRMECs treated with recombinant human Gas6 (rhGas6. Sprague-Dawley rat aortas in Matrigels were treated with rhGas6, and microvessel sprouting emanating from arterial rings was analyzed. Transgenic zebrafish embryos (flk:GFP were microinjected with rhGas6 at 50 hours post-fertilization (hpf, and ectopic sprouting of subintestinal vessels (SIVs was observed under a confocal microscope. Morpholino oligonucleotides (MOs were microinjected to knockdown gas6 in zebrafish embryos, and intersegmental vessel impairment was observed. The effect of the extracellular signal-regulated kinase (ERK1/2 inhibitor on the migration of HRMECs and on vessel development in zebrafish embryos was tested. RESULTS: rhGas6 stimulated proliferation, migration, and tube formation in HRMECs in a dose-dependent manner. In rat aortas, rhGas6 induced vessel outgrowth, and the sprouting length was longer than that of controls. The rhGas6-microinjected zebrafish embryos had significantly increased vessel outgrowth in the SIVs. Recombinant human vascular endothelial growth factor (rhVEGF served as a positive control. Knockdown of gas6 inhibited angiogenesis in the developing vessels of zebrafish. The ERK1/2 inhibitor inhibited HRMEC migration and intersegmental vessel formation in zebrafish embryos. CONCLUSIONS/INTERPRETATIONS: These data suggest that Gas6 plays a pivotal role in proliferation, migration, and sprouting of angiogenic endothelial cells in the retina and in zebrafish embryos. Furthermore, Gas6 induced angiogenic processes are induced via phosphorylation of ERK1/2.

  13. 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.

  14. Hemorheological abnormalities in human arterial hypertension

    Science.gov (United States)

    Lo Presti, Rosalia; Hopps, Eugenia; Caimi, Gregorio

    2014-05-01

    Blood rheology is impaired in hypertensive patients. The alteration involves blood and plasma viscosity, and the erythrocyte behaviour is often abnormal. The hemorheological pattern appears to be related to some pathophysiological mechanisms of hypertension and to organ damage, in particular left ventricular hypertrophy and myocardial ischemia. Abnormalities have been observed in erythrocyte membrane fluidity, explored by fluorescence spectroscopy and electron spin resonance. This may be relevant for red cell flow in microvessels and oxygen delivery to tissues. Although blood viscosity is not a direct target of antihypertensive therapy, the rheological properties of blood play a role in the pathophysiology of arterial hypertension and its vascular complications.

  15. Expression of vascular endothelial growth factor and its two receptors in normal human endometrium

    Institute of Scientific and Technical Information of China (English)

    王海燕; 陈贵安

    2003-01-01

    Objectives: We try to demonstrate the expression of vascular endothelial growthfactor (VEGF) and its receptors, flt-1 and KDR, in normal human emdometrium duringthe menstrual cycle.Methods: Immunohistochemical method was used to observe the expression ofVEGF and its two receptors in emdometrium throughout the normal menstrual cyclemeanwhile the isoforms of VEGF were also detected by Western blot analysis. The en-dothelial cells of micro-vessels were marked with Ⅷ factor antibody.Results: VEGF and its receptors existed in endometrial glandular, stromal and vas-cular endothelial cells of human endometrium. Their expressions were higher in the mid-secretory phase of menstrual cycle and highest at menstruation. VEGF121 and VEGF165were the predominant isoforms in normal human endometrium.Conclusion: The expression of VEGF and its two receptors showed cycle-dependentin human endometrium, probably involved in embryonic implantation and endometrialproliferation and differentiation.

  16. 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.

  17. Non-endothelial endothelin counteracts hypoxic vasodilation in porcine large coronary arteries

    Directory of Open Access Journals (Sweden)

    Fröbert Ole

    2011-05-01

    Full Text Available Abstract Background The systemic vascular response to hypoxia is vasodilation. However, reports suggest that the potent vasoconstrictor endothelin-1 (ET-1 is released from the vasculature during hypoxia. ET-1 is reported to augment superoxide anion generation and may counteract nitric oxide (NO vasodilation. Moreover, ET-1 was proposed to contribute to increased vascular resistance in heart failure by increasing the production of asymmetric dimethylarginine (ADMA. We investigated the role of ET-1, the NO pathway, the potassium channels and radical oxygen species in hypoxia-induced vasodilation of large coronary arteries. Results In prostaglandin F2α (PGF2α, 10 μM-contracted segments with endothelium, gradual lowering of oxygen tension from 95 to 1% O2 resulted in vasodilation. The vasodilation to O2 lowering was rightward shifted in segments without endothelium at all O2 concentrations except at 1% O2. The endothelin receptor antagonist SB217242 (10 μM markedly increased hypoxic dilation despite the free tissue ET-1 concentration in the arterial wall was unchanged in 1% O2 versus 95% O2. Exogenous ET-1 reversed hypoxic dilation in segments with and without endothelium, and the hypoxic arteries showed an increased sensitivity towards ET-1 compared to the normoxic controls. Without affecting basal NO, hypoxia increased NO concentration in PGF2α-contracted arteries, and an NO synthase inhibitor, L-NOARG,(300 μM, NG-nitro-L-Arginine reduced hypoxic vasodilation. NO-induced vasodilation was reduced in endothelin-contracted preparations. Arterial wall ADMA concentrations were unchanged by hypoxia. Blocking of potassium channels with TEA (tetraethylammounium chloride(10 μM inhibited vasodilation to O2 lowering as well as to NO. The superoxide scavenger tiron (10 μM and the putative NADPH oxidase inhibitor apocynin (10 μM leftward shifted concentration-response curves for O2 lowering without changing vasodilation to 1% O2. PEG (polyethylene

  18. Responses of human endothelial cells to pathogenic and non-pathogenic Leptospira species.

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    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.

  19. Arterial endothelial cytokines guide extravillous trophoblast invasion towards spiral arteries; an in-vitro study with the trophoblast cell line ACH-3P and female non-uterine endothelial cells.

    Science.gov (United States)

    Weiss, Gregor; Huppertz, Berthold; Siwetz, Monika; Lang, Ingrid; Moser, Gerit

    2016-02-01

    Invasion of extravillous trophoblasts (EVT) is tightly linked to appropriate cell to cell contact as well as paracrine guidance of EVT by maternal uterine cells, conducted by a variety of locally expressed cytokines. Here we investigated the interaction of the first trimester trophoblast cell line ACH-3P with adult iliac arterial (AEC) and venous endothelial cells (VEC). The impact of ACH-3P conditioned medium (Cdm), obtained at 2.5% and 21% oxygen, on endothelial cell viability (LDH-Assay) and network formation (Matrigel-Assay) was tested. We investigated cytokine expression of AEC- and VEC-Cdm and confirmed results with ELISA analysis, and investigated the influence of Cdm on ACH-3P proliferation and invasion. Additionally, direct co-culture experiments with ACH-3P and AEC on Matrigel were performed. A subset of experiments was verified with primary trophoblasts as well as with first trimester placenta in situ specimens. ACH-3P-Cdm significantly enhanced cell viability of AEC and VEC after 72 h. ACH-3P-Cdm at 2.5% oxygen stabilized endothelial network structures in Matrigel up to 24 h, similar to the effect of a direct co-culture of AEC and ACH-3P. AEC and VEC showed a similar pattern of secreted cytokines. However, elevated levels of cytokines secreted by AEC were found for GRO, IL-6, MMP-1 and uPAR. ELISA confirmed elevated concentrations of IL-6 and uPAR in AEC compared to VEC. ACH-3P and primary trophoblasts more likely invaded towards AEC-Cdm than towards VEC-Cdm. Addition of IL-6 to Cdm increased the invasion potential of both cell types. AEC- and VEC-Cdm reduced ACH-3P cell proliferation after 24 h of culture. IL-6 was highly expressed in uterine AEC compared to VEC as visualized by immunohistochemistry. The presented results clearly demonstrate that cytokines of both cell types, AEC and trophoblasts, differentially contribute to successful guidance and interaction in the process of trophoblast invasion. Copyright © 2015 Elsevier Ltd. All rights

  20. 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

  1. 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.

  2. 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...

  3. 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...

  4. 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.

  5. Vasculogenesis, angiogenesis and the molecular organisation of endothelial junctions in the early human placenta.

    Science.gov (United States)

    Leach, Lopa; Babawale, Michael O; Anderson, Mark; Lammiman, Michael

    2002-01-01

    Vasculogenesis and angiogenesis are regulated by the capacity of endothelial cells to adhere to each other and form new tubes. The presence and role of junctional adhesion molecules during physiological vasculogenesis is unknown. Using ultrastructural and immunocytochemical approaches, we compared the junctional phenotype of developing vessels of the first-trimester human placenta with vessels in the last trimester; the latter include newly formed terminal capillaries and the quiescent vascular bed. First-trimester placental vessels contained the adherens junctional molecules, vascular endothelial cadherin and alpha- and beta-catenin but lacked plakoglobin, the component of fully differentiated adherens junctions. Furthermore, these vessels did not contain the transmembrane tight junctional molecules occludin and claudin-1 and -2. This profile reflects the phenotype of terminal capillaries but differs from large vessels of the full-term placenta. Electron microscopic studies revealed that endothelial tight junctions are present in the first-trimester placenta. Thus, occludin and claudin-1 appear to play no part in the formation of endothelial tight junctions, but are a later requirement. In the early placenta, the predominant growth factor appears to be vascular endothelial growth factor (VEGF), whilst at term, angiopoietin-1 was present in large vessels, with intense angiopoietin-2 immunofluorescence (and VEGF) located in terminal villous capillaries. Thus, endothelial junctions in the human placenta possess two distinct molecular phenotypes, i.e. stable or dynamic, dependent on maturity and plasticity. These distinct phenotypes may be influenced by the angiopoietins/VEGF present in the placenta. Copyright 2002 S. Karger AG, Basel

  6. Role of Rutin on Nitric Oxide Synthesis in Human Umbilical Vein Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Azizah Ugusman

    2014-01-01

    Full Text Available Nitric oxide (NO, produced by endothelial nitric oxide synthase (eNOS, is a major antiatherogenic factor in the blood vessel. Oxidative stress plays an important role in the pathogenesis of various cardiovascular diseases, including atherosclerosis. Decreased availability of endothelial NO promotes the progression of endothelial dysfunction and atherosclerosis. Rutin is a flavonoid with multiple cardiovascular protective effects. This study aimed to investigate the effects of rutin on eNOS and NO production in cultured human umbilical vein endothelial cells (HUVEC. HUVEC were divided into four groups: control; oxidative stress induction with 180 μM H2O2; treatment with 300 μM rutin; and concomitant induction with rutin and H2O2 for 24 hours. HUVEC treated with rutin produced higher amount of NO compared to control (P<0.01. In the oxidative stress-induced HUVEC, rutin successfully induced cells’ NO production (P<0.01. Rutin promoted NO production in HUVEC by inducing eNOS gene expression (P<0.05, eNOS protein synthesis (P<0.01, and eNOS activity (P<0.05. Treatment with rutin also led to increased gene and protein expression of basic fibroblast growth factor (bFGF in HUVEC. Therefore, upregulation of eNOS expression by rutin may be mediated by bFGF. The results showed that rutin may improve endothelial function by augmenting NO production in human endothelial cells.

  7. Endothelial hyperpermeability in severe pulmonary arterial hypertension: role of store-operated calcium entry

    Science.gov (United States)

    Zhou, Chun; Townsley, Mary I.; Alexeyev, Mikhail; Voelkel, Norbert F.

    2016-01-01

    Here, we tested the hypothesis that animals with severe pulmonary arterial hypertension (PAH) display increased sensitivity to vascular permeability induced by activation of store-operated calcium entry. To test this hypothesis, wild-type and transient receptor potential channel 4 (TRPC4) knockout Fischer 344 rats were given a single injection of Semaxanib (SU5416; 20 mg/kg) followed by 3 wk of exposure to hypoxia (10% oxygen) and a return to normoxia (21% oxygen) for an additional 2–3 wk. This Semaxanib/hypoxia/normoxia (i.e., SU5416/hypoxia/normoxia) treatment caused PAH, as evidenced by development of right ventricular hypertrophy, pulmonary artery medial hypertrophy, and occlusive lesions within precapillary arterioles. Pulmonary artery pressure was increased fivefold in Semaxanib/hypoxia/normoxia-treated animals compared with untreated, Semaxanib-treated, and hypoxia-treated controls, determined by isolated perfused lung studies. Thapsigargin induced a dose-dependent increase in permeability that was dependent on TRPC4 in the normotensive perfused lung. This increase in permeability was accentuated in PAH lungs but not in Semaxanib- or hypoxia-treated lungs. Fluid accumulated in large perivascular cuffs, and although alveolar fluid accumulation was not seen in histological sections, Evans blue dye conjugated to albumin was present in bronchoalveolar lavage fluid of hypertensive but not normotensive lungs. Thus PAH is accompanied by a TRPC4-dependent increase in the sensitivity to edemagenic agents that activate store-operated calcium entry. PMID:27422996

  8. Sex differences in endothelial function in porcine coronary arteries: a role for H2O2 and gap junctions?

    Science.gov (United States)

    Wong, P S; Roberts, R E; Randall, M D

    2014-06-01

    Cardiovascular risk is higher in men and postmenopausal women compared with premenopausal women. This may be due to sex differences in endothelial function. Here, sex differences in endothelial function of porcine coronary arteries (PCAs) were investigated. Distal PCAs were studied under myographic conditions and after precontraction with U46619. Concentration-response curves to bradykinin were constructed in the presence of a range of inhibitors. In male and female PCAs, bradykinin produced comparable vasorelaxant responses. Inhibition of NO and prostanoid synthesis produced greater inhibition in males compared with females. Removing H2 O2 with PEG-catalase reduced the maximum relaxation in the absence, but not the presence of L-NAME and indomethacin in females, and had no effect in males. Blocking gap junctions with 100 µM carbenoxolone or 18α-glycyrrhetinic acid further inhibited the endothelium-derived hyperpolarization (EDH)-mediated response in females but not in males. In female PCAs, the maximum EDH-mediated response was reduced by inhibiting SKCa with apamin and by inhibiting IKCa with TRAM-34, or with both. In male PCAs, at maximum bradykinin concentration, the EDH-mediated response was reduced in the presence of apamin but not TRAM-34. Western blot did not detect any differences in connexins 40 or 43 or in IKCa expression between male and female PCAs. H2 O2 mediated some part of endothelium-dependent vasorelaxation in female PCAs and EDH was more important in females, with differences in the contribution of gap junctions and IKCa channels. These findings may contribute to understanding vascular protection in premenopausal women. © 2014 The British Pharmacological Society.

  9. Gene expression microarray data from human microvascular endothelial cells supplemented with a low concentration of niacin

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    Jennifer M. Hughes-Large

    2016-03-01

    Full Text Available The systemic lipid modifying drug, niacin, can directly improve human microvascular endothelial cell angiogenic function under lipotoxic conditions, possibly through activation of niacin receptors “Niacin receptor activation improves human microvascular endothelial cell angiogenic function during lipotoxicity” (Hughes-Large et al. 2014. Here we provide accompanying data collected using Affymetrix GeneChip microarrays to identify changes in gene expression in human microvascular endothelial cells treated with 10 μM niacin. Statistical analyses of robust multi-array average (RMA values revealed that only 16 genes exhibited greater than 1.3-fold differential expression. Of these 16, only 5 were identified protein coding genes, while 3 of the remaining 11 genes appeared to be small nuclear/nucleolar RNAs. Altered expression of EFCAB4B, NAP1L2, and OR13C8 was confirmed by real time quantitative PCR.

  10. Nicotine promotes vascular endothelial growth factor secretion by human trophoblast cells under hypoxic conditions and improves the proliferation and tube formation capacity of human umbilical endothelial cells.

    Science.gov (United States)

    Zhao, Hongbo; Wu, Lanxiang; Wang, Yahui; Zhou, Jiayi; Li, Ruixia; Zhou, Jiabing; Wang, Zehua; Xu, Congjian

    2017-04-01

    Pre-eclampsia, characterized as defective uteroplacental vascularization, remains the major cause of maternal and fetal mortality and morbidity. Previous epidemiological studies demonstrated that cigarette smoking reduced the risk of pre-eclampsia. However, the molecular mechanism remains elusive. In the present study, it is demonstrated that a low dose of nicotine decreased soluble vascular endothelial growth factor receptor 1 (sFlt1) secretion in human trophoblast cells under hypoxic conditions. Nicotine was then observed to promote vascular endothelial growth factor (VEGF) secretion by reducing sFlt1 secretion and increasing VEGF mRNA transcription. Further data showed that nicotine enhanced hypoxia-mediated hypoxia-inducible factor-1α (HIF-1α) expression and HIF-1α small interfering RNA abrogated nicotine-induced VEGF secretion, indicating that HIF-1α may be responsible for nicotine-mediated VEGF transcription under hypoxic conditions. Moreover, conditioned medium from human trophoblast cells treated with nicotine under hypoxic conditions promoted the proliferation and tube formation capacity of human umbilical endothelial cells (HUVEC) by promoting VEGF secretion. These findings indicate that nicotine may promote VEGF secretion in human trophoblast cells under hypoxic conditions by reducing sFlt1 secretion and up-regulating VEGF transcription and improve the proliferation and tube formation of HUVEC cells, which may contribute to elucidate the protective effect of cigarette smoking against pre-eclampsia. Copyright © 2017 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  11. Haemodynamic assessment of human coronary arteries is affected by degree of freedom of artery movement.

    Science.gov (United States)

    Javadzadegan, Ashkan; Yong, Andy S C; Chang, Michael; Ng, Martin K C; Behnia, Masud; Kritharides, Leonard

    2017-02-01

    Abnormal haemodynamic parameters are associated with atheroma plaque progression and instability in coronary arteries. Flow recirculation, shear stress and pressure gradient are understood to be important pathogenic mediators in coronary disease. The effect of freedom of coronary artery movement on these parameters is still unknown. Fluid-structure interaction (FSI) simulations were carried out in 25 coronary artery models derived from authentic human coronaries in order to investigate the effect of degree of freedom of movement of the coronary arteries on flow recirculation, wall shear stress (WSS) and wall pressure gradient (WPG). Each FSI model had distinctive supports placed upon it. The quantitative and qualitative differences in flow recirculation, maximum wall shear stress (MWSS), areas of low wall shear stress (ALWSS) and maximum wall pressure gradient (MWPG) for each model were determined. The results showed that greater freedom of movement was associated with lower MWSS, smaller ALWSS, smaller flow recirculation zones and lower MWPG. With increasing percentage diameter stenosis (%DS), the effect of degree of freedom on flow recirculation and WSS diminished. Freedom of movement is an important variable to be considered for computational modelling of human coronary arteries, especially in the setting of mild to moderate stenosis. 3D: Three-dimensional; 3DR: Three-dimensional Reconstruction; 3D-QCA: Three-dimensional quantitative coronary angiography; ALWSS: Areas of low wall shear stress; CAD: Coronary artery disease; CFD: Computational fluid dynamics; %DS: Diameter stenosis percentage; EPCS: End point of counter-rotating streamlines; FSI: Fluid-structure interaction; IVUS: Intravascular ultrasound; LAD: Left anterior descending; MWSS: Maximum wall shear stress; SST: Shear stress transport; TAWSS: Time-averaged wall shear stress; WSS: wall shear stress; WPG: Wall pressure gradient; MWPG: Maximum wall pressure gradient; FFR: Fractional flow reserve; i

  12. Correlation of mid-pregnancy uterine artery notch with preeclampsia incidence risk as well as serum endothelial injury and placental hypoperfusion

    Directory of Open Access Journals (Sweden)

    Xue-Jing Guo

    2016-10-01

    Full Text Available Objective: To study the correlation of mid-pregnancy uterine artery notch with preeclampsia incidence risk as well as serum endothelial injury and placental hypoperfusion. Methods: A total of 2 800 cases of singleton primiparas who received antenatal examination in Maternal and Child Health Hospital of Pudong New Area Shanghai between April 2014 and October 2015 were selected for prospective study and divided into notch group and non-notch group according to the uterine artery notch at 22-24 weeks of gestation, serum was collected at 24-30 weeks, 31-36 weeks and 37-41 weeks of gestation respectively to determine endothelial injury indexes VEGF, PLGF, sFlt-1 and sEng in serum, ultrasonography was conducted to determine uterine spiral arterial resistance index (RI and pulsatility index (PI, and the levels of apoptosis molecules in placenta tissue were determined after delivery. Results: Preeclampsia incidence rate of notch group was significantly higher than that of non-notch group; at 24-30 weeks, 31- 36 weeks and 37-41 weeks of gestation, serum VEGF and PLGF levels of notch group were significantly lower than those of non-notch group, sFlt-1 and sEng levels were significantly higher than those of non-notch group, and uterine spiral artery PI and RI were significantly higher than those of non-notch group; after delivery, p57kip2, Bax, Caspase-3, Fas and FasL levels in placenta tissue of notch group were significantly higher than those of non-notch group. Conclusion: The risk of preeclampsia is high in pregnant women with mid-pregnancy (22-24 weeks of gestation uterine artery notch and it will cause maternal endothelial injury, spiral artery remodeling dysfunction and placental hypoperfusion.

  13. Cultivation of Human Microvascular Endothelial Cells on Topographical Substrates to Mimic the Human Corneal Endothelium

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    Jie Shi Chua

    2013-03-01

    Full Text Available Human corneal endothelial cells have a limited ability to replicate in vivo and in vitro. Allograft transplantation becomes necessary when an accident or trauma results in excessive cell loss. The reconstruction of the cornea endothelium using autologous cell sources is a promising alternative option for therapeutic or in vitro drug testing applications. The native corneal endothelium rests on the Descemet’s membrane, which has nanotopographies of fibers and pores. The use of synthetic topographies mimics the native environment, and it is hypothesized that this can direct the behavior and growth of human microvascular endothelial cells (HMVECs to resemble the corneal endothelium. In this study, HMVECs are cultivated on substrates with micron and nano-scaled pillar and well topographies. Closely packed HMVEC monolayers with polygonal cells and well-developed tight junctions were formed on the topographical substrates. Sodium/potassium (Na+/K+ adenine triphosphatase (ATPase expression was enhanced on the microwells substrate, which also promotes microvilli formation, while more hexagonal-like cells are found on the micropillars samples. The data obtained suggests that the use of optimized surface patterning, in particular, the microtopographies, can induce HMVECs to adopt a more corneal endothelium-like morphology with similar barrier and pump functions. The mechanism involved in cell contact guidance by the specific topographical features will be of interest for future studies.

  14. Effects of Music Therapy on Endothelial Function in Patients With Coronary Artery Disease Participating in Aerobic Exercise Therapy.

    Science.gov (United States)

    Deljanin Ilic, Marina; Pavlovic, Radmila F; Kocic, Gordana; Simonovic, Dejan; Lazarevic, Gordana

    2017-05-01

    Context • Pleasant music that evokes a positive emotional response may activate brain pathways of the insular cortex, central nucleus of the amygdala, and lateral hypothalamus, which are involved in the integration of emotional and ambient sensory input, with corresponding autonomic responses. Exercise training can improve endothelium-dependent vasodilatation, both in epicardial coronary vessels and in resistance vessels, for patients with coronary heart disease. Objective • The aim of the present study was to evaluate the effects on endothelial function when patients with stable coronary artery disease (CAD) listened to their favorite music. Design • The study was a randomized controlled trial. Setting • The study occurred at the Institute of Cardiology, Niska Banja, Faculty of Medicine, University of Nis (Nis, Serbia). Participants • Participants were 74 patients with stable CAD. Intervention • Participants were randomly assigned to 1 of 3 groups: (1) exercise training only (T) group (n = 33), (2) listening to music and exercise training (MT) group (n = 31), and listening to music only (M) group (n = 10). Participants in the T and MT groups received usual medical care and underwent 3 wk of supervised aerobic exercise training. In addition to the exercise training, participants in the MT group listened to their favorite music for 1.5 h every day. Participants in the M group received the usual medical care and listened to their favorite music for 1.5 h every day. Outcome Measures • At baseline and postintervention, outcomes were assessed through measurement of the changes in circulating blood markers of endothelial function-the stable end product of nitric oxide (NOx), asymmetric dimethylarginine, symmetric dimethylarginine, and xanthine oxidase-and through the results of submaximal or symptom-limited exercise test. Results • After 3 wk, the NOx significantly increased in both in MT and T groups, with P favorite music in addition to participating in

  15. Retrograde flow and shear rate acutely impair endothelial function in humans.

    NARCIS (Netherlands)

    Thijssen, D.H.J.; Dawson, E.A.; Tinken, T.M.; Cable, N.T.; Green, D.J.

    2009-01-01

    Changes in arterial shear stress induce functional and structural vasculature adaptations. Recent studies indicate that substantial retrograde flow and shear can occur through human conduit arteries. In animals, retrograde shear is associated with atherogenic effects. The aim of this study was to

  16. Non-endothelial endothelin counteracts hypoxic vasodilation in porcine large coronary arteries

    DEFF Research Database (Denmark)

    Hedegaard, Elise Røge; Stankevicius, Edgaras; Simonsen, Ulf

    2011-01-01

    ) vasodilation. Moreover, ET-1 was proposed to contribute to increased vascular resistance in heart failure by increasing the production of asymmetric dimethylarginine (ADMA). We investigated the role of ET-1, the NO pathway, the potassium channels and radical oxygen species in hypoxia-induced vasodilation...... superoxide dismutase)(70 u/ml) affect vasodilation to O2 lowering. The mitochondrial inhibitors rotenone (1 μM) and antimycin A (1 μM) both inhibited hypoxic vasodilatation. CONCLUSION: The present results in porcine coronary arteries suggest NO contributes to hypoxic vasodilation, probably through K channel...

  17. Upregulation of transmembrane endothelial junction proteins in human cerebral cavernous malformations.

    Science.gov (United States)

    Burkhardt, Jan-Karl; Schmidt, Dörthe; Schoenauer, Roman; Brokopp, Chad; Agarkova, Irina; Bozinov, Oliver; Bertalanffy, Helmut; Hoerstrup, Simon P

    2010-09-01

    Cerebral cavernous malformations (CCMs) are among the most prevalent cerebrovascular malformations, and endothelial cells seem to play a major role in the disease. However, the underlying mechanisms, including endothelial intercellular communication, have not yet been fully elucidated. In this article, the authors focus on the endothelial junction proteins CD31, VE-cadherin, and occludin as important factors for functional cell-cell contacts known as vascular adhesion molecules and adherence and tight junctions. Thirteen human CCM specimens and 6 control tissue specimens were cryopreserved and examined for the presence of VE-cadherin, occludin, and CD31 by immunofluorescence staining. Protein quantification was performed by triplicate measurements using western blot analysis. Immunofluorescent analyses of the CCM sections revealed a discontinuous pattern of dilated microvessels and capillaries as well as increased expression of occludin, VE-cadherin, and CD31 in the intima and in the enclosed parenchymal tissue compared with controls. Protein quantification confirmed these findings by showing upregulation of the levels of these proteins up to 2-6 times. A protocol enabling the molecular and morphological examination of the intercellular contact proteins in human CCM was validated. The abnormal and discontinuous pattern in these endothelial cell-contact proteins compared with control tissue explains the loose intercellular junctions that are considered to be one of the causes of CCM-associated bleeding or transendothelial oozing of erythrocytes. Despite the small number of specimens, this study demonstrates for the first time a quantitative analysis of endothelial junction proteins in human CCM.

  18. Effect of Australian Propolis from Stingless Bees (Tetragonula carbonaria) on Pre-Contracted Human and Porcine Isolated Arteries

    Science.gov (United States)

    Massaro, Flavia C.; Brooks, Peter R.; Wallace, Helen M.; Nsengiyumva, Vianne; Narokai, Lorraine; Russell, Fraser D.

    2013-01-01

    Bee propolis is a mixture of plant resins and bee secretions. While bioactivity of honeybee propolis has been reported previously, information is limited on propolis from Australian stingless bees (Tetragonula carbonaria). The aim of this study was to investigate possible vasomodulatory effects of propolis in KCl-precontracted porcine coronary arteries using an ex vivo tissue bath assay. Polar extracts of propolis produced a dose-dependent relaxant response (EC50=44.7±7.0 μg/ml), which was unaffected by endothelial denudation, suggesting a direct effect on smooth muscle. Propolis markedly attenuated a contractile response to Ca2+ in vessels that were depolarised with 60 mM KCl, in Ca2+-free Krebs solution. Propolis (160 µg/ml) reduced vascular tone in KCl pre-contracted vessels to near-baseline levels over 90 min, and this effect was partially reversible with 6h washout. Some loss in membrane integrity, but no loss in mitochondrial function was detected after 90 min exposure of human cultured umbilical vein endothelial cells to 160 µg/ml propolis. We conclude that Australian stingless bee (T. carbonaria) propolis relaxes porcine coronary artery in an endothelial-independent manner that involves inhibition of voltage-gated Ca2+ channels. This effect is partially and slowly reversible upon washout. Further studies are required to determine the therapeutic potential of Australian stingless bee propolis for conditions in which vascular supply is compromised. PMID:24260567

  19. Human Bone Derived Collagen for the Development of an Artificial Corneal Endothelial Graft. In Vivo Results in a Rabbit Model.

    Directory of Open Access Journals (Sweden)

    Natalia Vázquez

    Full Text Available Corneal keratoplasty (penetrating or lamellar using cadaveric human tissue, is nowadays the main treatment for corneal endotelial dysfunctions. However, there is a worldwide shortage of donor corneas available for transplantation and about 53% of the world's population have no access to corneal transplantation. Generating a complete cornea by tissue engineering is still a tough goal, but an endothelial lamellar graft might be an easier task. In this study, we developed a tissue engineered corneal endothelium by culturing human corneal endothelial cells on a human purified type I collagen membrane. Human corneal endothelial cells were cultured from corneal rims after corneal penetrating keratoplasty and type I collagen was isolated from remnant cancellous bone chips. Isolated type I collagen was analyzed by western blot, liquid chromatography -mass spectrometry and quantified using the exponentially modified protein abundance index. Later on, collagen solution was casted at room temperature obtaining an optically transparent and mechanically manageable membrane that supports the growth of human and rabbit corneal endothelial cells which expressed characteristic markers of corneal endothelium: zonula ocluddens-1 and Na+/K+ ATPase. To evaluate the therapeutic efficiency of our artificial endothelial grafts, human purified type I collagen membranes cultured with rabbit corneal endothelial cells were transplanted in New Zealand white rabbits that were kept under a minimal immunosuppression regimen. Transplanted corneas maintained transparency for as long as 6 weeks without obvious edema or immune rejection and maintaining the same endothelial markers that in a healthy cornea. In conclusion, it is possible to develop an artificial human corneal endothelial graft using remnant tissues that are not employed in transplant procedures. This artificial endothelial graft can restore the integrality of corneal endothelium in an experimental model of

  20. Endothelial Differentiation of Human Adipose-Derived Stem Cells on Polyglycolic Acid/Polylactic Acid Mesh

    Science.gov (United States)

    Deng, Meng; Gu, Yunpeng; Liu, Zhenjun; Qi, Yue; Ma, Gui E.; Kang, Ning

    2015-01-01

    Adipose-derived stem cell (ADSC) is considered as a cell source potentially useful for angiogenesis in tissue engineering and regenerative medicine. This study investigated the growth and endothelial differentiation of human ADSCs on polyglycolic acid/polylactic acid (PGA/PLA) mesh compared to 2D plastic. Cell adhesion, viability, and distribution of hADSCs on PGA/PLA mesh were observed by CM-Dil labeling, live/dead staining, and SEM examination while endothelial differentiation was evaluated by flow cytometry, Ac-LDL/UEA-1 uptake assay, immunofluorescence stainings, and gene expression analysis of endothelial related markers. Results showed hADSCs gained a mature endothelial phenotype with a positive ratio of 21.4 ± 3.7% for CD31+/CD34− when induced in 3D mesh after 21 days, which was further verified by the expressions of a comprehensive range of endothelial related markers, whereas hADSCs in 2D induced and 2D/3D noninduced groups all failed to differentiate into endothelial cells. Moreover, compared to 2D groups, the expression for α-SMA was markedly suppressed in 3D cultured hADSCs. This study first demonstrated the endothelial differentiation of hADSCs on the PGA/PLA mesh and pointed out the synergistic effect of PGA/PLA 3D culture and growth factors on the acquisition of mature characteristic endothelial phenotype. We believed this study would be the initial step towards the generation of prevascularized tissue engineered constructs. PMID:26106426

  1. Endothelial Differentiation of Human Adipose-Derived Stem Cells on Polyglycolic Acid/Polylactic Acid Mesh

    Directory of Open Access Journals (Sweden)

    Meng Deng

    2015-01-01

    Full Text Available Adipose-derived stem cell (ADSC is considered as a cell source potentially useful for angiogenesis in tissue engineering and regenerative medicine. This study investigated the growth and endothelial differentiation of human ADSCs on polyglycolic acid/polylactic acid (PGA/PLA mesh compared to 2D plastic. Cell adhesion, viability, and distribution of hADSCs on PGA/PLA mesh were observed by CM-Dil labeling, live/dead staining, and SEM examination while endothelial differentiation was evaluated by flow cytometry, Ac-LDL/UEA-1 uptake assay, immunofluorescence stainings, and gene expression analysis of endothelial related markers. Results showed hADSCs gained a mature endothelial phenotype with a positive ratio of 21.4 ± 3.7% for CD31+/CD34− when induced in 3D mesh after 21 days, which was further verified by the expressions of a comprehensive range of endothelial related markers, whereas hADSCs in 2D induced and 2D/3D noninduced groups all failed to differentiate into endothelial cells. Moreover, compared to 2D groups, the expression for α-SMA was markedly suppressed in 3D cultured hADSCs. This study first demonstrated the endothelial differentiation of hADSCs on the PGA/PLA mesh and pointed out the synergistic effect of PGA/PLA 3D culture and growth factors on the acquisition of mature characteristic endothelial phenotype. We believed this study would be the initial step towards the generation of prevascularized tissue engineered constructs.

  2. Pdgfrα functions in endothelial-derived cells to regulate neural crest cells and the development of the great arteries.

    Science.gov (United States)

    Aghajanian, Haig; Cho, Young Kuk; Rizer, Nicholas W; Wang, Qiaohong; Li, Li; Degenhardt, Karl; Jain, Rajan

    2017-09-01

    Originating as a single vessel emerging from the embryonic heart, the truncus arteriosus must septate and remodel into the aorta and pulmonary artery to support postnatal life. Defective remodeling or septation leads to abnormalities collectively known as conotruncal defects, which are associated with significant mortality and morbidity. Multiple populations of cells must interact to coordinate outflow tract remodeling, and the cardiac neural crest has emerged as particularly important during this process. Abnormalities in the cardiac neural crest have been implicated in the pathogenesis of multiple conotruncal defects, including persistent truncus arteriosus, double outlet right ventricle and tetralogy of Fallot. However, the role of the neural crest in the pathogenesis of another conotruncal abnormality, transposition of the great arteries, is less well understood. In this report, we demonstrate an unexpected role of Pdgfra in endothelial cells and their derivatives during outflow tract development. Loss of Pdgfra in endothelium and endothelial-derived cells results in double outlet right ventricle and transposition of the great arteries. Our data suggest that loss of Pdgfra in endothelial-derived mesenchyme in the outflow tract endocardial cushions leads to a secondary defect in neural crest migration during development. © 2017. Published by The Company of Biologists Ltd.

  3. Effect of acute hypobaric hypoxia on the endothelial glycocalyx and digital reactive hyperemia in humans

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    Pär I Johansson

    2014-11-01

    Full Text Available Introduction: Hypoxia is associated with increased capillary permeability. This study tested whether acute hypobaric hypoxia involves degradation of the endothelial glycocalyx. Methods: We exposed 12 subjects to acute hypobaric hypoxia (equivalent to 4,500 m for 2-4 hours and measured venous blood concentrations of biomarkers reflecting endothelial and glycocalyx degradation (catecholamines, syndecan-1, soluble CD40 ligand, protein C, soluble thrombomodulin, tissue-type plasminogen activators, histone-complexed DNA fragments and nitrite/nitrate. Endothelial function was assessed by the hyperemic response to brachial artery occlusion by peripheral arterial tonometry. Results: Compared with normoxic baseline levels, hypoxia increased concentrations of syndecan-1 from 22 (95% confidence interval: 17-27 to 25 (19-30 ng/ml (p < 0.02 and protein C from 76 (70-83 % to 81 (74-88 % (p < 0.02. Nitrite/nitrate decreased from 23 (18-27 μM at baseline to 19 (14-24 μM and 18 (14-21 μM in hypoxia and recovery, respectively (p < 0.05. Other biomarkers remained unchanged. The post-occlusion/pre-occlusion ratio (reactive hyperemia index, RHI decreased from 1.80 (1.52–2.07 in normoxia to 1.62 (1.28–1.96 after 2 to 4 hours of hypobaric hypoxia and thereafter increased to 2.43 (1.99-2.86 during normoxic recovery (p < 0.01. Conclusions: The increase in syndecan-1 and protein C suggests that acute hypobaric hypoxia produces minor degree of glycocalyx degradation and overall cellular damage. After hypoxia RHI rebounded to higher than baseline levels suggesting improved endothelial functionality.

  4. Coagulopathy, catecholamines, and biomarkers of endothelial damage in experimental human endotoxemia and in patients with severe sepsis

    DEFF Research Database (Denmark)

    Ostrowski, Sisse R; Berg, Ronan M G; Windeløv, Nis A

    2013-01-01

    The aim of this study was to investigate associations between circulating catecholamines, endothelial damage, and coagulopathy in experimental human endotoxemia and septic patients.......The aim of this study was to investigate associations between circulating catecholamines, endothelial damage, and coagulopathy in experimental human endotoxemia and septic patients....

  5. Passive biaxial mechanical response of aged human iliac arteries.

    Science.gov (United States)

    Schulze-Bauer, Christian A J; Mörth, Christian; Holzapfel, Gerhard A

    2003-06-01

    Inflation and extension tests of arteries are essential for the understanding of arterial wall mechanics. Data for such tests of human arteries are rare. At autopsy we harvested 10 non-diseased external iliac arteries of aged subjects (52-87 yrs). Structural homogeneity was ensured by means of ultrasound imaging, and anamneses of patients were recorded. We measured the axial in situ stretches, load-free geometries and opening angles. Passive biaxial mechanical responses of preconditioned cylindrical specimens were studied in 37 degrees C calcium-free Tyrode solution under quasistatic loading conditions. Specimens were subjected to pressure cycles varying from 0 to 33.3 kPa (250 mmHg) at nine fixed axial loads, varying from 0 to 9.90N. For the description of the load-deformation behavior we employed five "two-dimensional" orthotropic strain-energy functions frequently used in arterial wall mechanics. The associated constitutive models were compared in regard to their ability of representing the experimental data. Histology showed that the arteries were of the muscular type. In contrast to animal arteries they exhibited intimal layers of considerable thickness. The average ratio of wall thickness to outer diameter was 7.7, which is much less than observed for common animal arteries. We found a clear correlation between age and the axial in situ stretch lambda is (r = -0.72, P = 0.03), and between age and distensibility of specimens, i.e. aged specimens are less distensible. Axial in situ stretches were clearly smaller (1.07 +/- 0.09, mean +/- SD) than in animal arteries. For one specimen lambda is was even smaller than 1.0, i.e. the vessel elongated axially upon excision. The nonlinear and anisotropic load-deformation behavior showed small hystereses. For the majority of specimens we observed axial stretches smaller than 1.3 and circumferential stretches smaller than 1.1 for the investigated loading range. Data from in situ inflation tests showed a significant

  6. You're Only as Old as Your Arteries: Translational Strategies for Preserving Vascular Endothelial Function with Aging

    Science.gov (United States)

    Kaplon, Rachelle E.; Gioscia-Ryan, Rachel A.; LaRocca, Thomas J.

    2014-01-01

    Endothelial dysfunction develops with age and increases the risk of age-associated vascular disorders. Nitric oxide insufficiency, oxidative stress, and chronic low-grade inflammation, induced by upregulation of adverse cellular signaling processes and imbalances in stress resistance pathways, mediate endothelial dysfunction with aging. Healthy lifestyle behaviors preserve endothelial function with aging by inhibiting these mechanisms, and novel nutraceutical compounds that favorably modulate these pathways hold promise as a complementary approach for preserving endothelial health. PMID:24985329

  7. Short- and long-term influence of diet and simvastatin on brachial artery endothelial function

    DEFF Research Database (Denmark)

    Jensen, Lisette Okkels; Thayssen, Per; Petersen, Knud Erik

    2006-01-01

    BACKGROUND: Endothelium-dependent dilation (EDD) has often been studied in patients with hypercholesterolemia without overt coronary atherosclerosis where an improvement after statin treatment has been documented within few weeks. The aim of the study was to assess the short-term effect of diet...... as well as short- and long-term effect of simvastatin on EDD in patients with hypercholesterolemia and ischemic heart disease. METHODS: In 43 male patients with hypercholesterolemia and ischemic heart disease, EDD of the brachial artery was measured at baseline, after 3 months on a lipid-lowering diet......, and after another 1, 3 and 12 months on simvastatin at 40 mg/day. RESULTS: Three months changes in EDD were neither influenced by diet nor short-term simvastatin therapy (4.9% vs. 4.9% vs. 4.8%, p=ns). Twelve months simvastatin treatment, however, significantly improved EDD by 32.7% (4.9% vs. 6.5%, p=0...

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

    Directory of Open Access Journals (Sweden)

    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.

  9. Endothelial dysfunction and brachial intima-media thickness: long term cardiovascular risk with claudication related to peripheral arterial disease: a prospective analysis.

    Directory of Open Access Journals (Sweden)

    Franz Hafner

    Full Text Available OBJECTIVE: Endothelial dysfunction plays a key role in the development, progression, and clinical manifestation of atherosclerosis, and in symptomatic peripheral arterial disease, endothelial dysfunction and enlarged intima-media thickness might be associated with increased cardiovascular risk. Flow-mediated dilatation and serologic parameters are used to evaluate individual endothelial function. Brachial intima-media thickness, a less recognized parameter of cardiovascular risk, is independently associated with coronary artery disease. The aim of this study was to evaluate the prognostic value of ultrasound and serologic parameters of endothelial function in relation to cardiovascular mortality in peripheral arterial disease. DESIGN: monocentric, prospective cohort study. METHODS: Flow mediated dilatation and brachial intima-media thickness were assessed in 184 (124 male patients with peripheral arterial disease (Rutherford stages 2-3. Serologic parameters of endothelial function included asymmetric dimethylarginine (ADMA, symmetric dimethylarginine (SDMA, and L-homoarginine. Cardiovascular events were recorded during a follow-up of 99.1±11.1 months. Subjects who died of noncardiovascular causes were excluded from further analysis. RESULTS: Eighty-two patients (44.6% died during follow-up after a mean duration of 49.7±28.3 months. There were 49 cardiovascular deaths (59.8% and 33 other deaths (40.2%. Flow mediated dilatation was associated with cardiovascular death [1.17% (0.0, 4.3 vs. 4.1% (1.2, 6.4, p<0.001]. Intima-media thickness was greater in patients who succumbed to cardiovascular disease [0.37 mm (0.30, 0.41] than in survivors [0.21 mm (0.15, 0.38, p<0.001]. Brachial intima-media thickness above 0.345 mm was most predictive of cardiovascular death, with sensitivity and specificity values of 0.714 and 0.657, respectively (p<0.001. Furthermore, ADMA levels above 0.745 µmol/l and SDMA levels above 0.825 µmol/l were significantly

  10. Early gene response of human brain endothelial cells to Listeria monocytogenes

    Science.gov (United States)

    The gene expression of human brain microvascular endothelial cells (HBMEC) to Listeria monocytogenes at 4 hour infection was analyzed. Four hours after infection, the expression of 456 genes of HBMEC had changed (p<0.05). We noted that many active genes were involved in the formyl-methionylleucylph...

  11. Subcellular localization and mechanism of secretion of vascular endothelial growth factor in human skeletal muscle

    DEFF Research Database (Denmark)

    Høier, Birgitte; Prats Gavalda, Clara; Qvortrup, Klaus

    2013-01-01

    The subcellular distribution and secretion of vascular endothelial growth factor (VEGF) was examined in skeletal muscle of healthy humans. Skeletal muscle biopsies were obtained from m.v. lateralis before and after a 2 h bout of cycling exercise. VEGF localization was conducted on preparations...

  12. UP-REGULATION OF ANTITHROMBOTIC ECTONUCLEOTIDASES BY ASPIRIN IN HUMAN ENDOTHELIAL-CELLS IN-VITRO

    NARCIS (Netherlands)

    CHEUNG, PK; VISSER, J; BAKKER, WW

    1994-01-01

    Ecto ATP-diphosphohydrolase (apyrase) activity of human endothelial cells following aspirin treatment has been studied in-vitro. It was shown by HPLC analysis of supernatant samples that pre-incubation of the cultures with aspirin resulted in a significantly increased turnover of supplemented ATP

  13. Effects of phthalates on the human corneal endothelial cell line B4G12

    DEFF Research Database (Denmark)

    Krüger, Tanja; Cao, Yi; Kjærgaard, Søren K.

    2012-01-01

    Phthalates are industrial chemicals used in many cosmetics. We evaluated an in vitro model for eye irritancy testing using the human corneal endothelial cell line B4G12. Cell proliferation and toxicity were assessed after exposing to di-n-butyl phthalate (DBP), benzyl butyl phthalate (BBP), di-2...

  14. Transfection of small numbers of human endothelial cells by electroporation and synthetic amphiphiles

    NARCIS (Netherlands)

    van Leeuwen, E B; van der Veen, A Y; Hoekstra, D; Engberts, J B; Halie, M R; van der Meer, J; Ruiters, M H

    OBJECTIVES: This study compared the efficiency of electroporation and synthetic amphiphiles. (SAINT-2pp/DOPE) in transfecting small numbers of human endothelial cells. METHODS AND RESULTS: Optimal transfection conditions were tested and appeared to be 400 V and 960 microF for electroporation and a

  15. Characterization of human endothelial cell urokinase-type plasminogen activator receptor protein and messenger RNA

    DEFF Research Database (Denmark)

    Barnathan, E S; Kuo, A; Karikó, K

    1990-01-01

    Human umbilical vein endothelial cells in culture (HUVEC) express receptors for urokinase-type plasminogen activators (u-PA). The immunochemical nature of this receptor and its relationship to u-PA receptors expressed by other cell types is unknown. Cross-linking active site-blocked u-PA to HUVEC...

  16. Potassium currents in cultured human pulmonary arterial smooth muscle cells.

    Science.gov (United States)

    Peng, W; Karwande, S V; Hoidal, J R; Farrukh, I S

    1996-04-01

    In this study, using whole cell and single-channel configurations of the patch-clamp technique, we characterized K+ currents (IK) in cultured human pulmonary arterial smooth muscle cells. The net whole cell outward membrane current (IKo) was activated at potentials positive to -60 mV. One component of IKo, IK(dr), was inhibited by 4-aminopyridine (4-AP) and high concentrations of tetraethylammonium (TEA) but was Ca2+ and charybdotoxin (CTX) insensitive. The other component of IKo, IK(Ca), was voltage and Ca2+ dependent and was inhibited by CTX and low concentrations of TEA. Activation of IKo in single-channel recordings was voltage dependent and demonstrated a high-conductance channel (245 +/- 2 pS) that was Ca2+ and CTX sensitive [IK(Ca)] and a low-conductance channel (109 +/- 2 pS) that was inhibited by 4-AP [IK(dr)] but was insensitive to low concentrations of TEA or to an increase in intracellular [Ca2+]. In isolated pulmonary arterial rings, TEA and 4-AP caused an additive increase in arterial tension. To our knowledge these data provide the first characterization of the IK in human pulmonary arterial smooth muscle cells and indicate that IK(Ca) and IK(dr) play an important role in maintaining pulmonary vascular tone. The data confirm previous observations in pulmonary smooth muscle cells of animal models.

  17. K ATP channels in pig and human intracranial arteries

    DEFF Research Database (Denmark)

    Ploug, Kenneth Beri; Sørensen, Mette Aaskov; Strøbech, Lotte

    2008-01-01

    Clinical trials suggest that synthetic ATP-sensitive K(+) (K(ATP)) channel openers may cause headache and migraine by dilating cerebral and meningeal arteries. We studied the mRNA expression profile of K(ATP) channel subunits in the pig and human middle meningeal artery (MMA) and in the pig middle...... cerebral artery (MCA). We determined the order of potency of four K(ATP) channel openers when applied to isolated pig MMA and MCA, and we examined the potential inhibitory effects of the Kir6.1 subunit specific K(ATP) channel blocker PNU-37883A on K(ATP) channel opener-induced relaxation of the isolated...... pig MMA and MCA. Using conventional RT-PCR, we detected the mRNA transcripts of the K(ATP) channel subunits Kir6.1 and SUR2B in all the examined pig and human intracranial arteries. Application of K(ATP) channel openers to isolated pig MMA and MCA in myographs caused a concentration...

  18. Oxygen radicals induce human endothelial cells to express GMP-140 and bind neutrophils.

    Science.gov (United States)

    Patel, K D; Zimmerman, G A; Prescott, S M; McEver, R P; McIntyre, T M

    1991-02-01

    The initial step in extravasation of neutrophils (polymorphonuclear leukocytes [PMNs]) to the extravascular space is adherence to the endothelium. We examined the effect of oxidants on this process by treating human endothelial cells with H2O2, t-butylhydroperoxide, or menadione. This resulted in a surface adhesive for PMN between 1 and 4 h after exposure. The oxidants needed to be present only for a brief period at the initiation of the assay. Adhesion was an endothelial cell-dependent process that did not require an active response from the PMN. The adhesive molecule was not platelet-activating factor, which mediates PMN adherence when endothelial cells are briefly exposed to higher concentrations of H2O2 (Lewis, M. S., R. E. Whatley, P. Cain, T. M. McIntyre, S. M. Prescott, and G. A. Zimmerman. 1988. J. Clin. Invest. 82:2045-2055), nor was it ELAM-1, an adhesive glycoprotein induced by cytokines. Oxidant-induced adhesion did not require protein synthesis, was inhibited by antioxidants, and, when peroxides were the oxidants, was inhibited by intracellular iron chelators. Granule membrane protein-140 (GMP-140) is a membrane-associated glycoprotein that can be translocated from its intracellular storage pool to the surface of endothelial cells where it acts as a ligand for PMN adhesion (Geng, J.-G., M. P. Bevilacqua, K. L. Moore, T. M. McIntyre, S. M. Prescott, J. M. Kim, G. A. Bliss, G. A. Zimmerman, and R. P. McEver. 1990. Nature (Lond). 343:757-760). We found that endothelial cells exposed to oxidants expressed GMP-140 on their surface, and that an mAb against GMP-140 or solubilized GMP-140 completely blocked PMN adherence to oxidant-treated endothelial cells. Thus, exposure of endothelial cells to oxygen radicals induces the prolonged expression of GMP-140 on the cell surface, which results in enhanced PMN adherence.

  19. Relationship between arterial vascular calcifications seen on screening mammograms and biochemical markers of endothelial injury

    Energy Technology Data Exchange (ETDEWEB)

    Pidal, Diego [Unidad de Investigacion del, Hospital de Jove, Gijon (Spain)], E-mail: dpidal@hotmail.com; Sanchez Vidal, M Teresa [Servicio de Medicina Interna, Hospital de Jove (Spain)], E-mail: medicinainterna@hospitaldejove.com; Rodriguez, Juan Carlos [Unidad de Investigacion del, Hospital de Jove, Gijon (Spain); Servicio de Cirugia General, Hospital de Jove (Spain); Instituto Universitario de Oncologia del Principado de Asturias, Oviedo (Spain)], E-mail: investigacion@hospitaldejove.com; Corte, M Daniela [Unidad de Investigacion del, Hospital de Jove, Gijon (Spain); Instituto Universitario de Oncologia del Principado de Asturias, Oviedo (Spain)], E-mail: mdanielac@hotmail.com; Pravia, Paz [Servicio de Radiodiagnostico, Hospital de Jove (Spain)], E-mail: radiologia@hospitaldejove.com; Guinea, Oscar [Servicio de Radiodiagnostico, Hospital de Jove (Spain)], E-mail: oscarfguinea@seram.org; Pidal, Ivan [Unidad de Investigacion del, Hospital de Jove, Gijon (Spain)], E-mail: ivanpida@hotmail.com; Bongera, Miguel [Unidad de Investigacion del, Hospital de Jove, Gijon (Spain)], E-mail: mbchoppy@hotmail.com; Escribano, Damaso [Servicio de Medicina Interna, Hospital de Jove (Spain)], E-mail: medicinainterna@hospitaldejove.com; Gonzalez, Luis O. [Unidad de Investigacion del, Hospital de Jove, Gijon (Spain)], E-mail: lovidiog@telefonica.net; Diez, M Cruz [Servicio de Cirugia General, Hospital de Jove (Spain)], E-mail: cirugiageneral@hospitaldejove.com; Venta, Rafael [Servicio de Analisis Clinicos, Hospital de San Agustin, Aviles (Spain); Departamento de Bioquimica y Biologia Molecular, Universidad de Oviedo (Spain)], E-mail: rafael.venta@sespa.princast.es; Vizoso, Francisco J. [Unidad de Investigacion del, Hospital de Jove, Gijon (Spain); Servicio de Cirugia General, Hospital de Jove (Spain); Instituto Universitario de Oncologia del Principado de Asturias, Oviedo (Spain)], E-mail: fjvizoso@telefonica.net

    2009-01-15

    To assess whether breast arterial calcifications (BAC) are associated with altered serum markers of cardiovascular risk, mammograms and records from 1759 women (age range: 45-65 years) screened for breast cancer were revised. One hundred and forty seven (8.36%) women showed BAC. A total of 136 women with BAC and controls (mean age: 57 and 55 years, respectively) accepted entering the study. There were no significant differences in serum levels of urea, glucose, uric acid, creatinine, total cholesterol, HDL-C, LDL-C, folic acid, vitamin B{sub 12}, TSH or cysteine, between both groups of patients. However, women with BAC showed higher serum levels of triglycerides (p = 0.006), homocysteine (p = 0.002) and hs-CRP (p = 0.003) than women without BAC. Likewise, we found a significantly higher percentage of cases with an elevated LDL-C/HDL-C ratio (coronary risk index >2) amongst women with BAC than in women without BAC (56.7 and 38.2%, respectively; p = 0.04). Our results indicate that the finding of BAC identify women showing altered serum markers of cardiovascular risk.

  20. [Liposome-mediated human CD40 gene transfection and human umbilical vein endothelial ECV-304 cells].

    Science.gov (United States)

    Wang, Wei-rong; Lin, Rong; Yang, Yu-cong; Gan, Wei-jie; Liu, Jun-tian; Lü, She-min

    2005-12-01

    To construct an eukaryotic expression vector containing human CD40 gene for its efficient, continuous and stable expression in human umbilical vein endothelial ECV-304 cells. The recombinant plasmid pUCD40 was digested with endonucleases to obtain human CD40 gene fragment, which was cloned into pCDNA3.1 vector to construct recombinant eukaryotic expression vector pCDNA3.1(+)/CD40. The recombinant vector was identified by enzyme digestion before introduced into ECV-304 cells via liposome, with the positive cell clones selected with G418. The stable transfection and expression of CD40 in ECV-304 cells were identified by reverse transcription (RT)-PCR, Western blotting and flow cytometry, respectively. Enzyme digestion analysis showed that target gene had been cloned into the recombinant vector. The transfected ECV-304 cells successfully expressed human CD40 as determined by RT-PCR and Western-blotting, and 95% of the cells were CD40-positive as shown by flow cytometry. The recombinant eukaryotic expression vector pCDNA3.1(+)/CD40 has been successfully constructed, which is capable of stable transfection and expression of CD40 in ECV-304 cells to facilitate further investigation of the roles of CD40 molecule in antiatherosclerotic drug development.

  1. Markers of thrombogenesis and fibrinolysis and their relation to inflammation and endothelial activation in patients with idiopathic pulmonary arterial hypertension.

    Directory of Open Access Journals (Sweden)

    Grzegorz Kopeć

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

  2. Isolation and characterization of human umbilical cord-derived endothelial colony-forming cells

    Science.gov (United States)

    Zhang, Hao; Tao, Yanling; Ren, Saisai; Liu, Haihui; Zhou, Hui; Hu, Jiangwei; Tang, Yongyong; Zhang, Bin; Chen, Hu

    2017-01-01

    Endothelial colony-forming cells (ECFCs) are a population of endothelial progenitor cells (EPCs) that display robust proliferative potential and vessel-forming capability. Previous studies have demonstrated that a limited number of ECFCs may be obtained from adult bone marrow, peripheral blood and umbilical cord (UC) blood. The present study describes an effective method for isolating ECFCs from human UC. The ECFCs derived from human UC displayed the full properties of EPCs. Analysis of the growth kinetics, cell cycle and colony-forming ability of the isolated human UC-ECFCs indicated that the cells demonstrated properties of stem cells, including relative stability and rapid proliferation in vitro. Gene expression of Fms related tyrosine kinase 1, kinase insert domain receptor, vascular endothelial cadherin, cluster of differentiation (CD)31, CD34, epidermal growth factor homology domains-2, von Willebrand factor and endothelial nitric oxide synthase was assessed by reverse transcription-polymerase chain reaction. The cells were positive for CD34, CD31, CD73, CD105 and vascular endothelial growth factor receptor-2, and negative for CD45, CD90 and human leukocyte antigen-antigen D related protein according to flow cytometry. 1,1′-dioctadecyl-3,3,3′,3′-tetra-methyl-indocarbocyanine perchlorate-labeled acetylated low-density lipoprotein and fluorescein isothiocyanate-Ulex europaeus-l were used to verify the identity of the UC-ECFCs. Matrigel was used to investigate tube formation capability. The results demonstrated that the reported technique is a valuable method for isolating human UC-ECFCs, which have potential for use in vascular regeneration. PMID:29067104

  3. Isolation and characterization of human umbilical cord-derived endothelial colony-forming cells.

    Science.gov (United States)

    Zhang, Hao; Tao, Yanling; Ren, Saisai; Liu, Haihui; Zhou, Hui; Hu, Jiangwei; Tang, Yongyong; Zhang, Bin; Chen, Hu

    2017-11-01

    Endothelial colony-forming cells (ECFCs) are a population of endothelial progenitor cells (EPCs) that display robust proliferative potential and vessel-forming capability. Previous studies have demonstrated that a limited number of ECFCs may be obtained from adult bone marrow, peripheral blood and umbilical cord (UC) blood. The present study describes an effective method for isolating ECFCs from human UC. The ECFCs derived from human UC displayed the full properties of EPCs. Analysis of the growth kinetics, cell cycle and colony-forming ability of the isolated human UC-ECFCs indicated that the cells demonstrated properties of stem cells, including relative stability and rapid proliferation in vitro. Gene expression of Fms related tyrosine kinase 1, kinase insert domain receptor, vascular endothelial cadherin, cluster of differentiation (CD)31, CD34, epidermal growth factor homology domains-2, von Willebrand factor and endothelial nitric oxide synthase was assessed by reverse transcription-polymerase chain reaction. The cells were positive for CD34, CD31, CD73, CD105 and vascular endothelial growth factor receptor-2, and negative for CD45, CD90 and human leukocyte antigen-antigen D related protein according to flow cytometry. 1,1'-dioctadecyl-3,3,3',3'-tetra-methyl-indocarbocyanine perchlorate-labeled acetylated low-density lipoprotein and fluorescein isothiocyanate-Ulex europaeus-l were used to verify the identity of the UC-ECFCs. Matrigel was used to investigate tube formation capability. The results demonstrated that the reported technique is a valuable method for isolating human UC-ECFCs, which have potential for use in vascular regeneration.

  4. Nitrobenzylthioinosine (NBT), a nucleoside transport inhibitor, protects against Shiga toxin cytotoxicity in human microvascular endothelial cells.

    Science.gov (United States)

    Ohmi, K; Kiyokawa, N; Sekino, T; Suzuki, T; Mimori, K; Taguchi, T; Nakajima, H; Katagiri, Y U; Fujimoto, J; Nakao, H; Takeda, T

    2001-01-01

    Infections with Shiga toxin (Stx)-producing Escherichia coli (STEC) cause microvascular endothelial cell damage, resulting in hemorrhagic colitis and hemolytic uremic syndrome. The prevention of endothelial cell damage is therefore a crucial step in overcoming this disorder. Here, we report that nitrobenzylthioinosine (NBT), a nucleoside transport inhibitor, has a protective effect against the cytotoxicity of Stxs in human microvascular endothelial cells (HMVECs). The relative viability of cells treated with 1.5-15 pM of Stx1 was reduced to 10-20% of that without Stx1. However, the viability of cells treated with NBT (10-100 microM) remained higher than 80%, even in the presence of Stx1. NBT also protected against Stx1 cytotoxicity in sodium butyrate-treated hypersensitive HMVECs. The protective effect of NBT against Stx cytotoxicity may be due to the depletion of ATP in the cells, thereby inhibiting the entry of Stx1.

  5. Human breast microvascular endothelial cells retain phenotypic traits in long-term finite life span culture

    DEFF Research Database (Denmark)

    Sigurdsson, Valgardur; Fridriksdottir, Agla J R; Kjartansson, Jens

    2007-01-01

    uptake of low-density lipoprotein, and had E-selectin induced upon treatment with tumor necrosis factor-alpha. The first signs of senescence in passage 14 were accompanied by gain of trisomy 11. At passage 18 cells showed chromosomal aberrations and growth arrest as revealed by beta......Attempts to study endothelial-epithelial interactions in the human breast have been hampered by lack of protocols for long-term cultivation of breast endothelial cells (BRENCs). The aim of this study was to establish long-term cultures of BRENCs and to compare their phenotypic traits......-galactosidase staining. We demonstrate here that breast microvasculature may serve as a large-scale source for expansion of BRENCs with molecular and functional traits preserved. These cells will form the basis for studies on the role of endothelial cells in breast morphogenesis....

  6. DPP-4 inhibition protects human umbilical vein endothelial cells from hypoxia-induced vascular barrier impairment

    Directory of Open Access Journals (Sweden)

    Naoko Hashimoto

    2017-09-01

    Full Text Available Dipeptidyl peptidase-4 (DPP-4 inhibitors are relatively new class of anti-diabetic drugs. Some protective effects of DPP-4 on cardiovascular disease have been described independently from glucose-lowering effect. However, the detailed mechanisms by which DPP-4 inhibitors exert on endothelial cells remain elusive. The purpose of this research was to determine the effects of DPP-4 inhibitor on endothelial barrier function. Human umbilical vein endothelial cells (HUVECs were cultured and exposed to hypoxia in the presence or absence of Diprotin A, a DPP-4 inhibitor. Immunocytochemistry of vascular endothelial (VE- cadherin showed that jagged VE-cadherin staining pattern induced by hypoxia was restored by treatment with Diprotin A. The increased level of cleaved β-catenin in response to hypoxia was significantly attenuated by Diprotin A, suggesting that DPP-4 inhibition protects endothelial adherens junctions from hypoxia. Subsequently, we found that Diprotin A inhibited hypoxia-induced translocation of NF-κB from cytoplasm to nucleus through decreasing TNF-α expression level. Furthermore, the tube formation assay showed that Diprotin A significantly restored hypoxia-induced decrease in number of tubes by HUVECs. These results suggest that DPP-4 inhibitior protects HUVECs from hypoxia-induced barrier impairment.

  7. Oxidative stress and apoptosis induced by iron oxide nanoparticles in cultured human umbilical endothelial cells.

    Science.gov (United States)

    Zhu, Mo-Tao; Wang, Yun; Feng, Wei-Yue; Wang, Bing; Wang, Meng; Ouyang, Hong; Chai, Zhi-Fang

    2010-12-01

    Recent epidemiologic researches indicate that exposure to ultrafine particles (nanoparticles) is an independent risk factor for several cardiovascular diseases. The induction of endothelial injuries is hypothesized to be an attractive mechanism involved in these cardiovascular diseases. To investigate this hypothesis, the widely used iron nanomaterials, ferric oxide (Fe2O3) and ferriferrous oxide (Fe3O4) nanoparticles were incubated with human umbilical endothelial cells (ECV304 cells) at different concentrations of 2, 20, 100 microg/mL. The cell viability, the rate of apoptosis, the apoptotic nuclear morphology and the mitochondria membrane potential were measured to estimate the cell necrosis and apoptosis caused by the nanoparticle exposure. The stimulation of superoxide anion (O2*-) and nitric oxide (NO) were examined to evaluate the stress responses of endothelial cells. Our results indicated that both the Fe2O3 and Fe3O4 nanoparticles could generate oxidative stress as well as the significant increase of nitric oxide in ECV304 cells. The loss of mitochondria membrane potential and the apoptotic chromatin condensation in the nucleus were observed as the early signs of apoptosis. It is inferred the stress response might be an important mechanism involving in endothelial cells apoptosis and death, and these injuries in endothelial cells might play a key role in downstream cardiovascular diseases such as atheroscelerosis, hypertension and myocardial infarction (MI).

  8. Recombinant human soluble thrombomodulin attenuates FK506-induced endothelial dysfunction through prevention of Akt inactivation.

    Science.gov (United States)

    Eguchi, Ryoji; Fujimori, Yoshihiro; Okada, Masaya; Tamaki, Hiroya; Wakabayashi, Ichiro; Ogawa, Hiroyasu

    2014-04-15

    Thrombomodulin (TM), a transmembrane glycoprotein on vascular endothelial cells, is a naturally occurring anticoagulant. Recombinant human soluble TM (rTM), composed of the extracellular domain of TM, also shows anti-coagulant and anti-inflammatory activity, but the effects of rTM on microangiopathy remain unclear. We reported that FK506 induced endothelial dysfunction through inactivation of Akt and extracellular-regulated kinase 1/2 using a three-dimensional culture blood vessel model. In the present study, we examined the effects of rTM on FK506-induced endothelial dysfunction. We found that rTM suppressed FK506-induced endothelial cell death, but not the breakdown of capillary-like tube structures. rTM prevented FK506-induced inactivation of Akt, but not of extracellular-regulated kinase 1/2. Akt inhibition by LY294002 abrogated the preventive effect of rTM on FK506-induced Akt inactivation and the suppressive effect of rTM on FK506-induced cell death. These results suggest that rTM attenuates FK506-induced endothelial dysfunction through prevention of Akt inactivation. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Concise Review: An Update on the Culture of Human Corneal Endothelial Cells for Transplantation.

    Science.gov (United States)

    Parekh, Mohit; Ferrari, Stefano; Sheridan, Carl; Kaye, Stephen; Ahmad, Sajjad

    2016-02-01

    The cornea forms the front window of the eye, enabling the transmission of light to the retina through a crystalline lens. Many disorders of the cornea lead to partial or total blindness, and therefore corneal transplantation becomes mandatory. Recently, selective corneal layer (as opposed to full thickness) transplantation has become popular because this leads to earlier rehabilitation and visual outcomes. Corneal endothelial disorders are a common cause of corneal disease and transplantation. Corneal endothelial transplantation is successful but limited worldwide because of lower donor corneal supply. Alternatives to corneal tissue for endothelial transplantation therefore require immediate attention. The field of human corneal endothelial culture for transplantation is rapidly emerging as a possible viable option. This manuscript provides an update regarding these developments. Significance: The cornea is the front clear window of the eye. It needs to be kept transparent for normal vision. It is formed of various layers of which the posterior layer (the endothelium) is responsible for the transparency of the cornea because it allows the transport of ions and solutes to and from the other layers of the cornea. Corneal blindness that results from the corneal endothelial dysfunction can be treated using healthy donor tissues. There is a huge demand for human donor corneas but limited supply, and therefore there is a need to identify alternatives that would reduce this demand. Research is underway to understand the isolation techniques for corneal endothelial cells, culturing these cells in the laboratory, and finding possible options to transplant these cells in the patients. This review article is an update on the recent developments in this field. ©AlphaMed Press.

  10. Arterial Disease in Patients With Human Immunodeficiency Virus Infection

    OpenAIRE

    Stein, James H.; Currier, Judith S.; HSUE, Priscilla Y

    2014-01-01

    With advances in antiretroviral therapy, individuals with human immunodeficiency virus (HIV) infection are living longer and increasingly die of non-HIV related diseases such as cardiovascular disease (CVD). Several observational studies suggest that HIV-infected patients on ART are at increased CVD risk; however, the precise mechanisms underlying the association between HIV infection and CVD risk are uncertain. Atherosclerosis and arterial disease in HIV-infected individuals is a multifactor...

  11. Effect of SR manipulation on conduit artery dilation in humans.

    Science.gov (United States)

    Carter, Howard H; Dawson, Ellen A; Birk, Gurpreet K; Spence, Angela L; Naylor, Louise H; Cable, N Timothy; Thijssen, Dick H J; Green, Daniel J

    2013-01-01

    The impact of manipulating shear stress on conduit artery vasodilation has not been comprehensively described in vivo. We hypothesized that manipulation of SR through the brachial and radial arteries would be associated with corresponding changes in diameter. We performed a series of studies involving the following: (1) leg cycle exercise at increasing intensities (≈70 and 85% maximum heart rate [HRmax]) with simultaneous bilateral measurement of SR in the radial arteries; (2) leg cycle exercise for 30 minutes at 80% HRmax with simultaneous bilateral measurement of velocity and diameter in the brachial arteries; and (3) bilateral forearm heating for 30 minutes with simultaneous bilateral measurement of brachial artery diameter and blood velocity. Cycling and forearm heating interventions were performed in the presence of unilateral cuff inflation throughout the experiment, or starting during the intervention (15 minutes), to manipulate SR responses. Cuff placement was associated with lower radial artery SR responses (cuffed versus uncuffed, 248±49 versus 349±105 L/s 85% HRmax; P<0.01), and diameter responses were similarly attenuated (2.45±0.30 versus 2.78±0.20 mm 85% HRmax; P<0.05). Exercise performed at 80% HRmax in the presence of unilateral cuff inflation also reduced brachial artery SR (cuffed versus uncuffed; 258±107 versus 454±157 L/s; P<0.01) and diameter (3.96±0.39 versus 4.20±0.45 mm). Finally, cuff inflation decreased the impact of forearm heating on brachial SR (cuffed versus uncuffed; 262±97 versus 440±106 L/s; P<0.01) and diameter (4.35±0.54 versus 4.87±0.47 mm; P<0.05). Similar significant differences between the cuffed and uncuffed limbs in SR and diameter were observed when cuff inflation occurred during exercise or heating. Our findings strongly implicate SR as an important stimulus to increase conduit artery diameter in humans.

  12. Effect of Moderate Aerobic Exercise Training on Endothelial Function and Arterial Stiffness in CKD Stages 3-4: A Randomized Controlled Trial.

    Science.gov (United States)

    Van Craenenbroeck, Amaryllis H; Van Craenenbroeck, Emeline M; Van Ackeren, Katrijn; Vrints, Christiaan J; Conraads, Viviane M; Verpooten, Gert A; Kouidi, Evangelia; Couttenye, Marie M

    2015-08-01

    Evidence of a beneficial effect of exercise training on mediators of vascular disease is accumulating in chronic kidney disease (CKD), but its effect on vascular function in vivo still has to be established. The present study was designed to investigate whether a formal aerobic exercise training program improves peripheral endothelial function in patients with CKD stages 3 to 4. Randomized controlled trial with a parallel-group design. 48 patients with CKD stages 3 to 4 without established cardiovascular disease were randomly assigned to either an exercise training program or usual care. 40 patients completed the study (exercise training, 19; usual care, 21). The 3-month home-based aerobic training program consisted of 4 daily cycling sessions of 10 minutes each at a target heart rate, calculated as 90% of the heart rate achieved at the anaerobic threshold. Patients in the usual-care group were given standard therapy. The primary outcome was peripheral endothelial function. Secondary outcomes were aerobic capacity, arterial stiffness, numbers of endothelial (EPCs) and osteogenic progenitor cells (OPCs), migratory function of circulatory angiogenic cells, and health-related quality of life. Endothelial function was assessed with flow-mediated dilation of the brachial artery, aerobic capacity by peak oxygen uptake (VO(2peak)), arterial stiffness by carotid-femoral pulse wave velocity, numbers of EPCs and OPCs by flow cytometry, circulatory angiogenic cell function by an in vitro migratory assay, and quality of life by the Kidney Disease Quality of Life-Short Form questionnaire. Exercise training significantly improved VO(2peak) and quality of life, but not in vivo vascular function (flow-mediated dilation and carotid-femoral pulse wave velocity) or cellular markers for vascular function (EPC and OPC count and circulatory angiogenic cell migratory function). Short duration and intermittent nature of the exercise intervention. In patients with CKD stages 3 to 4 without

  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. Association of endothelial nitric oxide synthase gene polymorphisms with coronary artery disease: an updated meta-analysis and systematic review.

    Directory of Open Access Journals (Sweden)

    Himanshu Rai

    Full Text Available Several association studies of endothelial nitric oxide synthase (NOS3 gene polymorphisms with respect to coronary artery disease (CAD have been published in the past two decades. However, their association with the disease, especially among different ethnic subgroups, still remains controversial. This prompted us to conduct a systematic review and an updated structured meta-analysis, which is the largest so far (89 articles, 132 separate studies, and a sample size of 69,235, examining association of three polymorphic forms of the NOS3 gene (i.e. Glu298Asp, T786-C and 27 bp VNTR b/a with CAD. In a subgroup analysis, we tested their association separately among published studies originating predominantly from European, Middle Eastern, Asian, Asian-Indian and African ancestries. The pooled analysis confirmed the association of all the three selected SNP with CAD in three different genetic models transcending all ancestries worldwide. The Glu298Asp polymorphism showed strongest association (OR range = 1.28-1.52, and P<0.00001 for all comparisons, followed by T786-C (OR range = 1.34-1.42, and P<0.00001 for all comparisons and 4b/a, (OR range = 1.19-1.41, and P ≤ 0.002 for all comparisons in our pooled analysis. Subgroup analysis revealed that Glu298Asp (OR range = 1.54-1.87, and P<0.004 for all comparisons and 4b/a (OR range = 1.71-3.02, and P<0.00001 for all comparisons have highest degree of association amongst the Middle Easterners. On the other hand, T786-C and its minor allele seem to carry a highest risk for CAD among subjects of Asian ancestry (OR range = 1.61-1.90, and P ≤ 0.01 for all comparisons.

  15. Erythropoietin attenuates pulmonary vascular remodeling in experimental pulmonary arterial hypertension through interplay between endothelial progenitor cells and heme-oxygenase

    Directory of Open Access Journals (Sweden)

    Rosa L.E. Loon

    2015-08-01

    Full Text Available BackgroundPulmonary arterial hypertension (PAH is a pulmonary vascular disease with a high mortality, characterized by typical angio-proliferative lesions. Erythropoietin (EPO attenuates pulmonary vascular remodeling in PAH. We postulated that EPO acts through mobilization of endothelial progenitor cells (EPCs and activation of the cytoprotective enzyme heme oxygenase-1 (HO1.MethodsRats with flow-associated PAH, resembling pediatric PAH, were treated with HO-1 inducer EPO in the presence or absence of the selective HO-activity-inhibitor tin-mesoporphyrin (SnMP. HO-activity, circulating EPCs and pulmonary vascular lesions were assessed after 3 weeks.ResultsIn PAH-rats, circulating EPCs were decreased and HO-activity was increased compared to control. EPO-treatment restored circulating EPCs and improved pulmonary vascular remodeling, as shown by a reduced wall thickness and occlusion rate of the intra-acinar vessels. Inhibition of HO-activity with SnMP aggravated PAH. Moreover, SnMP treatment abrogated EPO-induced amelioration of pulmonary vascular remodeling, while surprisingly further increasing circulating EPCs as compared with EPO alone.ConclusionsIn experimental PAH, EPO treatment restored the number of circulating EPC’s to control level, improved pulmonary vascular remodeling, and showed important interplay with HO-activity. Inhibition of increased HO-activity in PAH-rats exacerbated progression of pulmonary vascular remodeling, despite the presence of restored numbers of circulating EPC’s. We suggest that both EPO-induced HO1 and EPCs are promising targets to ameliorate the pulmonary vasculature in PAH.

  16. Relationship between upper and lower limb conduit artery vasodilator function in humans

    NARCIS (Netherlands)

    Thijssen, D.H.J.; Rowley, N.; Padilla, J.; Simmons, G.H.; Laughlin, M.H.; Whyte, G.; Cable, N.T.; Green, D.J.

    2011-01-01

    Brachial artery flow-mediated dilation (FMD) is a strong predictor of future cardiovascular disease and is believed to represent a "barometer" of systemic endothelial health. Although a recent study [Padilla et al. Exp Biol Med (Maywood) 235: 1287-1291, 2010] in pigs confirmed a strong correlation

  17. Tissue factor is induced by interleukin-33 in human endothelial cells: a new link between coagulation and inflammation.

    Science.gov (United States)

    Stojkovic, Stefan; Kaun, Christoph; Basilio, Jose; Rauscher, Sabine; Hell, Lena; Krychtiuk, Konstantin A; Bonstingl, Cornelia; de Martin, Rainer; Gröger, Marion; Ay, Cihan; Holnthoner, Wolfgang; Eppel, Wolfgang; Neumayer, Christoph; Huk, Ihor; Huber, Kurt; Demyanets, Svitlana; Wojta, Johann

    2016-05-04

    Tissue factor (TF) is the primary trigger of coagulation. Elevated levels of TF are found in atherosclerotic plaques, and TF leads to thrombus formation when released upon plaque rupture. Interleukin (IL)-33 was previously shown to induce angiogenesis and inflammatory activation of endothelial cells (ECs). Here, we investigated the impact of IL-33 on TF in human ECs, as a possible new link between inflammation and coagulation. IL-33 induced TF mRNA and protein in human umbilical vein ECs and coronary artery ECs. IL-33-induced TF expression was ST2- and NF-κB-dependent, but IL-1-independent. IL-33 also increased cell surface TF activity in ECs and TF activity in ECs-derived microparticles. IL-33-treated ECs reduced coagulation time of whole blood and plasma but not of factor VII-deficient plasma. In human carotid atherosclerotic plaques (n = 57), TF mRNA positively correlated with IL-33 mRNA expression (r = 0.691, p < 0.001). In this tissue, IL-33 and TF protein was detected in ECs and smooth muscle cells by immunofluorescence. Furthermore, IL-33 and TF protein co-localized at the site of clot formation within microvessels in plaques of patients with symptomatic carotid stenosis. Through induction of TF in ECs, IL-33 could enhance their thrombotic capacity and thereby might impact on thrombus formation in the setting of atherosclerosis.

  18. Inhibition of TGF-β signaling enables human corneal endothelial cell expansion in vitro for use in regenerative medicine.

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    Naoki Okumura

    Full Text Available Corneal endothelial dysfunctions occurring in patients with Fuchs' endothelial corneal dystrophy, pseudoexfoliation syndrome, corneal endotheliitis, and surgically induced corneal endothelial damage cause blindness due to the loss of endothelial function that maintains corneal transparency. Transplantation of cultivated corneal endothelial cells (CECs has been researched to repair endothelial dysfunction in animal models, though the in vitro expansion of human CECs (HCECs is a pivotal practical issue. In this study we established an optimum condition for the cultivation of HCECs. When exposed to culture conditions, both primate and human CECs showed two distinct phenotypes: contact-inhibited polygonal monolayer and fibroblastic phenotypes. The use of SB431542, a selective inhibitor of the transforming growth factor-beta (TGF-β receptor, counteracted the fibroblastic phenotypes to the normal contact-inhibited monolayer, and these polygonal cells maintained endothelial physiological functions. Expression of ZO-1 and Na(+/K(+-ATPase maintained their subcellular localization at the plasma membrane. Furthermore, expression of type I collagen and fibronectin was greatly reduced. This present study may prove to be the substantial protocol to provide the efficient in vitro expansion of HCECs with an inhibitor to the TGF-β receptor, and may ultimately provide clinicians with a new therapeutic modality in regenerative medicine for the treatment of corneal endothelial dysfunctions.

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

    Science.gov (United States)

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

    2015-05-01

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

  20. Comparative analysis of preservation method and intermittent perfusion volume on the expression of endothelial and inflammatory markers by coronal artery and myocardium in porcine donor hearts.

    Science.gov (United States)

    Hu, Zhibin; Yan, Zhikun; Wang, Haitao; Meng, Qun; Pan, Xiaohua

    2014-01-01

    Although continuous perfusion of donor hearts for preservation during transportation has been widely applied, intermittent perfusion has been suggested as an alternative. The aim of this study was to identify the optimal intermittent perfusion protocol by investigating the effects of perfusion volume on endothelial and inflammatory marker expression by the coronary artery. Donor porcine hearts were perfused with various volumes of Celsior solution supplemented with diazoxide (50, 100, 150, 200, and 250 ml) every 2 h for 30 min each for a 10 h period. The effects on cardiomyocytes and vascular endothelial cell morphology and marker expression were compared to the immersion control group. Whereas an incomplete endothelial cell layer with disorganized connective tissue was observed in the control and 50, 100, and 150 ml intermittent perfusion groups, transmission electron microscopic analysis revealed a complete endothelial cell layer in the intima with an organized subendothelium. A perfusion volume-dependent increase in eNOS expression that coincided with a decrease in ET-1, ICAM-1, vWF, and P-selectin expression was detected (all p porcine donor heart over a clinically relevant preservation period.

  1. Vasomotor control in mice overexpressing human endothelial nitric oxide synthase.

    Science.gov (United States)

    van Deel, Elza D; Merkus, Daphne; van Haperen, Rien; de Waard, Monique C; de Crom, Rini; Duncker, Dirk J

    2007-08-01

    Nitric oxide (NO) plays a key role in regulating vascular tone. Mice overexpressing endothelial NO synthase [eNOS-transgenic (Tg)] have a 20% lower systemic vascular resistance (SVR) than wild-type (WT) mice. However, because eNOS enzyme activity is 10 times higher in tissue homogenates from eNOS-Tg mice, this in vivo effect is relatively small. We hypothesized that the effect of eNOS overexpression is attenuated by alterations in NO signaling and/or altered contribution of other vasoregulatory pathways. In isoflurane-anesthetized open-chest mice, eNOS inhibition produced a significantly greater increase in SVR in eNOS-Tg mice compared with WT mice, consistent with increased NO synthesis. Vasodilation to sodium nitroprusside (SNP) was reduced, whereas the vasodilator responses to phosphodiesterase-5 blockade and 8-bromo-cGMP (8-Br-cGMP) were maintained in eNOS-Tg compared with WT mice, indicating blunted responsiveness of guanylyl cyclase to NO, which was supported by reduced guanylyl cyclase activity. There was no evidence of eNOS uncoupling, because scavenging of reactive oxygen species (ROS) produced even less vasodilation in eNOS-Tg mice, whereas after eNOS inhibition the vasodilator response to ROS scavenging was similar in WT and eNOS-Tg mice. Interestingly, inhibition of other modulators of vascular tone [including cyclooxygenase, cytochrome P-450 2C9, endothelin, adenosine, and Ca-activated K(+) channels] did not significantly affect SVR in either eNOS-Tg or WT mice, whereas the marked vasoconstrictor responses to ATP-sensitive K(+) and voltage-dependent K(+) channel blockade were similar in WT and eNOS-Tg mice. In conclusion, the vasodilator effects of eNOS overexpression are attenuated by a blunted NO responsiveness, likely at the level of guanylyl cyclase, without evidence of eNOS uncoupling or adaptations in other vasoregulatory pathways.

  2. Decellularized Matrix from Tumorigenic Human Mesenchymal Stem Cells Promotes Neovascularization with Galectin-1 Dependent Endothelial Interaction

    Science.gov (United States)

    Burns, Jorge S.; Kristiansen, Malthe; Kristensen, Lars P.; Larsen, Kenneth H.; Nielsen, Maria O.; Christiansen, Helle; Nehlin, Jan; Andersen, Jens S.; Kassem, Moustapha

    2011-01-01

    Background Acquisition of a blood supply is fundamental for extensive tumor growth. We recently described vascular heterogeneity in tumours derived from cell clones of a human mesenchymal stem cell (hMSC) strain (hMSC-TERT20) immortalized by retroviral vector mediated human telomerase (hTERT) gene expression. Histological analysis showed that cells of the most vascularized tumorigenic clone, -BD11 had a pericyte-like alpha smooth muscle actin (ASMA+) and CD146+ positive phenotype. Upon serum withdrawal in culture, -BD11 cells formed cord-like structures mimicking capillary morphogenesis. In contrast, cells of the poorly tumorigenic clone, -BC8 did not stain for ASMA, tumours were less vascularized and serum withdrawal in culture led to cell death. By exploring the heterogeneity in hMSC-TERT20 clones we aimed to understand molecular mechanisms by which mesenchymal stem cells may promote neovascularization. Methodology/Principal Findings Quantitative qRT-PCR analysis revealed similar mRNA levels for genes encoding the angiogenic cytokines VEGF and Angiopoietin-1 in both clones. However, clone-BD11 produced a denser extracellular matrix that supported stable ex vivo capillary morphogenesis of human endothelial cells and promoted in vivo neovascularization. Proteomic characterization of the -BD11 decellularized matrix identified 50 extracellular angiogenic proteins, including galectin-1. siRNA knock down of galectin-1 expression abrogated the ex vivo interaction between decellularized -BD11 matrix and endothelial cells. More stable shRNA knock down of galectin-1 expression did not prevent -BD11 tumorigenesis, but greatly reduced endothelial migration into -BD11 cell xenografts. Conclusions Decellularized hMSC matrix had significant angiogenic potential with at least 50 angiogenic cell surface and extracellular proteins, implicated in attracting endothelial cells, their adhesion and activation to form tubular structures. hMSC -BD11 surface galectin-1 expression was

  3. Anti-thrombotic and pro-fibrinolytic effects of levosimendan in human endothelial cells in vitro.

    Science.gov (United States)

    Krychtiuk, Konstantin A; Kaun, Christoph; Hohensinner, Philipp J; Stojkovic, Stefan; Seigner, Jacqueline; Kastl, Stefan P; Zuckermann, Andreas; Eppel, Wolfgang; Rauscher, Sabine; de Martin, Rainer; Maurer, Gerald; Huber, Kurt; Wojta, Johann; Speidl, Walter S

    2017-03-01

    Levosimendan is an inodilator for the treatment of acute decompensated heart failure (HF). Data from clinical studies suggest that levosimendan is particularly effective in HF due to myocardial infarction. After acute revascularization, no reflow-phenomenon is a common complication that may lead to pump failure and cardiogenic shock. Our aim was to examine whether levosimendan interferes with the pro-thrombotic phenotype of activated endothelial cells in vitro. Human heart microvascular endothelial cells (HHMEC) and human umbilical vein endothelial cells (HUVEC) were treated with interleukin-1β (IL-1β) (200U/mL) or thrombin (5U/mL) and co-treated with or without levosimendan (0.1-10μM) for 2-24h. In addition, flow experiments were performed. Effects on plasminogen activator inhibitor-1 (PAI-1) and tissue factor (TF) expression and activity were measured by rt-PCR, specific ELISA and flow cytometry. Treatment with IL-1β or thrombin significantly increased the expression of PAI-1 and TF in endothelial cells. Co-treatment with levosimendan strongly attenuated the effects of IL-1β and thrombin on PAI-1 and TF mRNA by up to 50% and 45%, in a dose- and time-dependent manner. Similar results were obtained under flow conditions. Furthermore, co-treatment with levosimendan dampened the antigen production of PAI-1 and the surface expression of TF by 35% and 45%, respectively. Additionally, levosimendan diminished both TF and PAI-1 activity. Levosimendan down-regulates the expression of the pro-thrombotic and anti-fibrinolytic biomolecules TF and PAI-1 in activated human endothelial cells. Our findings may, at least in part, explain some of the beneficial effects of levosimendan after myocardial reperfusion. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Effect of polyhexanide and gentamycin on human osteoblasts and endothelial cells.

    Science.gov (United States)

    Ince, Akif; Schütze, Norbert; Hendrich, Christian; Jakob, Franz; Eulert, Jochen; Löhr, Jochen F

    2007-03-10

    Infection of total joint replacements is painful, disabling and difficult to treat because of the increasing bacterial resistance against antibiotics. In view of this, antiseptics show limited bacterial tolerance and have a broad-spectrum antimicrobial activity. However, the application of antiseptics to bone is insufficiently studied in literature. Therefore, we investigated the biocompatibility of the antiseptic polyhexanide with bone related cells and asked whether supplementation to bone cement is appropriate in the management of total arthroplasty infections. We performed an in vitro study with immortalised human foetal osteoblast cells (hFOB 1.19) and human endothelial cells (EAhy 926). The cultured cells were exposed to media containing various concentrations of gentamicin (12.5-800 microg/ml) and polyhexanide (0.0006-0.01%) for six hours. We measured the phase-contrast microscopy images, the cell viability, cell number and the alkaline phosphatase activity as a parameter for osteogenic function. The exposure of hFOB and endothelial cells to polyhexanide showed a severe reduction of viability and cell number. Gentamicin did not have negative effects on hFOB and endothelial cell number and viability. The alkaline phosphatase activity of hFOB showed a significant decrease after exposure to polyhexanide and gentamicin. The viability and the cell number of endothelial cells seem more negatively affected by polyhexanide than the parameters of the hFOB-cells. The exposure of human osteoblasts and endothelial cells to polyhexanide at concentrations with questionable antibacterial activity resulted in severe cell damage whereas exposure to high dosed gentamicin did not. These results raise questions as to the feasibility of using antiseptics in bone cement for the treatment of total arthroplasty infections. Further in vivo studies are necessary to show the in vivo relevance of these in vitro findings.

  5. Action of shiga toxin type-2 and subtilase cytotoxin on human microvascular endothelial cells.

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    María M Amaral

    Full Text Available The hemolytic uremic syndrome (HUS associated with diarrhea is a complication of Shiga toxin (Stx-producing Escherichia coli (STEC infection. In Argentina, HUS is endemic and responsible for acute and chronic renal failure in children younger than 5 years old. The human kidney is the most affected organ due to the presence of very Stx-sensitive cells, such as microvascular endothelial cells. Recently, Subtilase cytotoxin (SubAB was proposed as a new toxin that may contribute to HUS pathogenesis, although its action on human glomerular endothelial cells (HGEC has not been described yet. In this study, we compared the effects of SubAB with those caused by Stx2 on primary cultures of HGEC isolated from fragments of human pediatric renal cortex. HGEC were characterized as endothelial since they expressed von Willebrand factor (VWF and platelet/endothelial cell adhesion molecule 1 (PECAM-1. HGEC also expressed the globotriaosylceramide (Gb3 receptor for Stx2. Both, Stx2 and SubAB induced swelling and detachment of HGEC and the consequent decrease in cell viability in a time-dependent manner. Preincubation of HGEC with C-9 -a competitive inhibitor of Gb3 synthesis-protected HGEC from Stx2 but not from SubAB cytotoxic effects. Stx2 increased apoptosis in a time-dependent manner while SubAB increased apoptosis at 4 and 6 h but decreased at 24 h. The apoptosis induced by SubAB relative to Stx2 was higher at 4 and 6 h, but lower at 24 h. Furthermore, necrosis caused by Stx2 was significantly higher than that induced by SubAB at all the time points evaluated. Our data provide evidence for the first time how SubAB could cooperate with the development of endothelial damage characteristic of HUS pathogenesis.

  6. Asymmetric dimethylarginine (ADMA) elevation and arginase up-regulation contribute to endothelial dysfunction related to insulin resistance in rats and morbidly obese humans.

    Science.gov (United States)

    El Assar, Mariam; Angulo, Javier; Santos-Ruiz, Marta; Ruiz de Adana, Juan Carlos; Pindado, María Luz; Sánchez-Ferrer, Alberto; Hernández, Alberto; Rodríguez-Mañas, Leocadio

    2016-06-01

    The presence of insulin resistance (IR) is determinant for endothelial dysfunction associated with obesity. Although recent studies have implicated the involvement of mitochondrial superoxide and inflammation in the defective nitric oxide (NO)-mediated responses and subsequent endothelial dysfunction in IR, other mechanisms could compromise this pathway. In the present study, we assessed the role of asymmetric dimethylarginine (ADMA) and arginase with respect to IR-induced impairment of endothelium-dependent vasodilatation in human morbid obesity and in a non-obese rat model of IR. We show that both increased ADMA and up-regulated arginase are determinant factors in the alteration of the l-arginine/NO pathway associated with IR in both models and also that acute treatment of arteries with arginase inhibitor or with l-arginine significantly alleviate endothelial dysfunction. These results help to expand our knowledge regarding the mechanisms of endothelial dysfunction that are related to obesity and IR and establish potential therapeutic targets for intervention. Insulin resistance (IR) is determinant for endothelial dysfunction in human obesity. Although we have previously reported the involvement of mitochondrial superoxide and inflammation, other mechanisms could compromise NO-mediated responses in IR. We evaluated the role of the endogenous NOS inhibitor asymmetric dimethylarginine (ADMA) and arginase with respect to IR-induced impairment of l-arginine/NO-mediated vasodilatation in human morbid obesity and in a non-obese rat model of IR. Bradykinin-induced vasodilatation was evaluated in microarteries derived from insulin-resistant morbidly obese (IR-MO) and non-insulin-resistant MO (NIR-MO) subjects. Defective endothelial vasodilatation in IR-MO was improved by l-arginine supplementation. Increased levels of ADMA were detected in serum and adipose tissue from IR-MO. Serum ADMA positively correlated with IR score and negatively with pD2 for bradykinin. Gene

  7. Direct Reprogramming of Human Dermal Fibroblasts Into Endothelial Cells Using ER71/ETV2.

    Science.gov (United States)

    Lee, Sangho; Park, Changwon; Han, Ji Woong; Kim, Ju Young; Cho, Kyuwon; Kim, Eun Jae; Kim, Sangsung; Lee, Shin-Jeong; Oh, Se Yeong; Tanaka, Yoshiaki; Park, In-Hyun; An, Hyo Jae; Shin, Claire Min; Sharma, Shraya; Yoon, Young-Sup

    2017-03-03

    Direct conversion or reprogramming of human postnatal cells into endothelial cells (ECs), bypassing stem or progenitor cell status, is crucial for regenerative medicine, cell therapy, and pathophysiological investigation but has remained largely unexplored. We sought to directly reprogram human postnatal dermal fibroblasts to ECs with vasculogenic and endothelial transcription factors and determine their vascularizing and therapeutic potential. We utilized various combinations of 7 EC transcription factors to transduce human postnatal dermal fibroblasts and found that ER71/ETV2 (ETS variant 2) alone best induced endothelial features. KDR+ (kinase insert domain receptor) cells sorted at day 7 from ER71/ETV2-transduced human postnatal dermal fibroblasts showed less mature but enriched endothelial characteristics and thus were referred to as early reprogrammed ECs (rECs), and did not undergo maturation by further culture. After a period of several weeks' transgene-free culture followed by transient reinduction of ER71/ETV2, early rECs matured during 3 months of culture and showed reduced ETV2 expression, reaching a mature phenotype similar to postnatal human ECs. These were termed late rECs. While early rECs exhibited an immature phenotype, their implantation into ischemic hindlimbs induced enhanced recovery from ischemia. These 2 rECs showed clear capacity for contributing to new vessel formation through direct vascular incorporation in vivo. Paracrine or proangiogenic effects of implanted early rECs played a significant role in repairing hindlimb ischemia. This study for the first time demonstrates that ER71/ETV2 alone can directly reprogram human postnatal cells to functional, mature ECs after an intervening transgene-free period. These rECs could be valuable for cell therapy, personalized disease investigation, and exploration of the reprogramming process. © 2016 American Heart Association, Inc.

  8. Large-scale simulation of the human arterial tree.

    Science.gov (United States)

    Grinberg, L; Anor, T; Madsen, J R; Yakhot, A; Karniadakis, G E

    2009-02-01

    1. Full-scale simulations of the virtual physiological human (VPH) will require significant advances in modelling, multiscale mathematics, scientific computing and further advances in medical imaging. Herein, we review some of the main issues that need to be resolved in order to make three-dimensional (3D) simulations of blood flow in the human arterial tree feasible in the near future. 2. A straightforward approach is computationally prohibitive even on the emerging petaflop supercomputers, so a three-level hierarchical approach based on vessel size is required, consisting of: (i) a macrovascular network (MaN); (ii) a mesovascular network (MeN); and (iii) a microvascular network (MiN). We present recent simulations of MaN obtained by solving the 3D Navier-Stokes equations on arterial networks with tens of arteries and bifurcations and accounting for the neglected dynamics through proper boundary conditions. 3. A multiscale simulation coupling MaN-MeN-MiN and running on hundreds of thousands of processors on petaflop computers will require no more than a few CPU hours per cardiac cycle within the next 5 years. The rapidly growing capacity of supercomputing centres opens up the possibility of simulation studies of cardiovascular diseases, drug delivery, perfusion in the brain and other pathologies.

  9. In vivo imaging of the effect of LPS on arterial endothelial cells: molecular imaging of heat shock protein 60 expression

    NARCIS (Netherlands)

    Wick, M.C.; Mayerl, Christina; Backovic, Aleksandar; Zee, R. van der; Jaschke, Werner; Dietrich, Hermann; Wick, G.

    2008-01-01

    Bacterial endotoxins are known as stress factors for endothelial cells. In 20 normocholesterolemic New Zealand White (NZW) rabbits, endothelial stress was induced by intravenous (i.v.) injection of lipopolysaccharide (LPS), while eight NZW rabbits were sham-treated or served as untreated

  10. IMPACT OF ANTICOAGULANT THERAPY ON THE INDICATORS OF ARTERIAL STIFFNESS AND ENDOTHELIAL DYSFUNCTION IN PATIENTS WITH ATRIAL FIBRILLATION AFTER CARDIOEMBOLIC STROKE

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    I. A. Zolotovskaya

    2016-01-01

    Full Text Available Objective: to investigate the impact of anticoagulant therapy on the indicators of arterial stiffness and endothelial dysfunction (ED in patients with atrial fibrillation (AF after cardioembolic stroke (CES.Patients and methods. The investigation enrolled 93 patients with AF after CES. The patients were divided into two groups: a study group in which all patients (n=48 received anticoagulants and a comparison group (n=45 in which the patients did not take anticoagulants, although the latter had been prescribed. The follow-up duration was 180.5±5.5 days. During this time the patients visited their doctors twice: the first visit was at baseline, the second one was after 24 weeks.All the patients underwent three-dimensional sphygmography. The levels of glucose, creatinine, total cholesterol, high-density lipoproteins, and triglycerides were determined. The markers of endothelial function, such as von Willebrand factor, plasminogen, antithrombin III,  and tissue plasminogen activator inhibitor-1, were estimated.Results. All the patients in the study and comparison groups had a higher comorbidity index of obvious somatic pathology. The patients from both groups showed changes in all cardiac morphometric and functional parameters. It was noted that after 6 months of a follow-up, there was a statistically significant positive trend in the indicators of arterial stiffness in the patients taking anticoagulants. The study group showed a significant correlation of blood pressure with the argumentation index. The 6-month follow-up revealed a strong correlation between the cardioankle vascular index and all the indicators of ED in the study group. After 24 weeks, in both groups there were 12 (12.9% deaths: 3 (6.25% and 8 (17.8% in the study and comparison groups, respectively.Conclusion. The findings suggest that in patients with AF after CES, the indicators of arterial stiffness and endothelial function tend to improve during anticoagulant therapy.

  11. Effect of high-glucose conditions on human periodontal ligament endothelial cells: in vitro analysis.

    Science.gov (United States)

    Maruyama, Kosuke; Sato, Soh

    2017-01-01

    Endothelial cells participate in key aspects of vascular biology, such as maintenance of capillary permeability and regulation of inflammation. According to previous reports, endothelial cells have revealed highly specific characteristics depending on the organs and tissues. In particular, periodontal endothelial cells have a higher permeability than vascular endothelial cells of other types of tissue. Periodontal disease is not only a chronic disease in oral, but also affect the entire body. Diabetes and periodontal disease are closely related, with periodontal disease even been referred to as the sixth complication of disease. However, no reports have investigated the pathophysiology of microvascular in periodontal tissue once diabetes has developed. Therefore, the aim of the present study is to investigate changes in the properties of human periodontal endothelial cells (HPDLECs) that were cultured under high-glucose conditions. We isolated HPDLECs from human periodontal ligament cells. HPDLECs were cultured under high-glucose (5.5, 11.0, 22.0 mM) and investigated proliferation, apoptosis, tube formation and the expression of cell adhesion molecules. A 5.5 mM (100 mg/dl) control was used in this study. HPDLECs stimulated with high glucose concentration exhibited suppression of cell proliferation and an increased percentage of apoptosis-positive cells. This results suggested that apoptosis was caused by TNF-α expression. The expression levels cell adhesion molecules increased. These results suggest that when HPDLECs are stimulated with a high glucose concentrations, PKC in the intracellular cell substrate is activated, increasing the expression of intercellular and vascular adhesion molecules. Thus, the results of this study demonstrate that diabetes exacerbates periodontal disease.

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

    Science.gov (United States)

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

    2016-10-01

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

  13. Heparanase mediates vascular endothelial growth factor gene transcription in high-glucose human retinal microvascular endothelial cells

    Science.gov (United States)

    Wang, Jingwei; Leng, Xuan; Hu, Yijun; Shen, Huangxuan; Song, Xin

    2017-01-01

    Purpose To observe the nuclear expression and interaction of heparanase and RNA polymerase II (RNA Pol II), an enzyme that catalyzes the transcription of DNA in eukaryotic cells) in human retinal microvascular endothelial cells (HRECs) under high glucose condition and to investigate the association of heparanase with the transcription activity of the vascular endothelial growth factor (VEGF) gene promoter. Methods Cultured HRECs were maintained for 3 days in media with high or normal glucose. The expressions of heparanase and RNA Pol II in each group were analyzed with immunofluorescence. Co-immunoprecipitation was applied to detect the interaction of heparanase and Pol II proteins. Cells in both groups were used for chromatin immunoprecipitation (ChIP) with anti-heparanase and anti-RNA Pol II antibodies to identify high-confidence heparanase-binding regions across the entire VEGF gene promoter. Moreover, real-time PCR was used to demonstrate the interaction between heparanase and the VEGF gene promoter region. Results The immunofluorescence studies showed that the nuclear expression of heparanase was intense in high-glucose HRECs but faint in the normal group; RNA Pol II in the nucleus was also intense in high glucose HRECs, and the distribution of heparanase was consistent with that of RNA Pol II. The co-immunoprecipitation data showed that heparanase combined with RNA Pol II in HRECs cells treated with high glucose, and the molecular size of HPA interacted with RNA Pol II was 50 kDa, while no combination of two proteins was evident in normal HRECs cells. Real-time PCR–based ChIP results showed that the high-confidence HPA-binding region was −1155 to −1018 (containing hypoxia response element) in the VEGF gene promoter, and the cells treated with high glucose showed increases in heparanase and RNA Pol II in the VEGF gene promoter region compared with the normal glucose treated cells (t = –3.244, p = 0.032; t = –6.096, p = 0.004, respectively

  14. Low-dose lithium stabilizes human endothelial barrier by decreasing MLC phosphorylation and universally augments cholinergic vasorelaxation capacity in a direct manner

    Directory of Open Access Journals (Sweden)

    Bert Bosche

    2016-12-01

    Full Text Available Lithium at plasma concentrations up to 1 mmol/L has been used in patients suffering from bipolar disorder for decades and has recently been shown to reduce the risk for ischemic stroke in these patients. The risk for stroke and thromboembolism depend not only on cerebral but also on general endothelial function and health; the entire endothelium as an organ is therefore pathophysiologically relevant. Regardless, the knowledge about the direct impact of lithium on endothelial function remains poor. We conducted an experimental study using lithium as pharmacologic pretreatment for murine, porcine and human vascular endothelium. We predominantly investigated endothelial vasorelaxation capacities in addition to human basal and dynamic (thrombin-/PAR-1 receptor agonist-impaired barrier functioning including myosin light chain phosphorylation (MLC-P. Low-dose therapeutic lithium concentrations (0.4 mmol/L significantly augment the cholinergic endothelium-dependent vasorelaxation capacities of cerebral and thoracic arteries, independently of central and autonomic nerve system influences. Similar concentrations of lithium (0.2-0.4 mmol/L significantly stabilized the dynamic thrombin-induced and PAR-1 receptor agonist-induced permeability of human endothelium, while even the basal permeability appeared to be stabilized. The lithium-attenuated dynamic permeability was mediated by a reduced endothelial MLC-P known to be followed by a lessening of endothelial cell contraction and paracellular gap formation. The well-known lithium-associated inhibition of inositol monophosphatase/glycogen synthase kinase-3-β signaling-pathways involving intracellular calcium concentrations in neurons seems to similarly occur in endothelial cells, too, but with different down-stream effects such as MLC-P reduction. This is the first study discovering low-dose lithium as a drug directly stabilizing human endothelium and ubiquitously augmenting cholinergic endothelium

  15. Changes in brachial artery endothelial function and resting diameter with moderate-intensity continuous but not sprint interval training in sedentary men.

    Science.gov (United States)

    Shenouda, Ninette; Gillen, Jenna B; Gibala, Martin J; MacDonald, Maureen J

    2017-10-01

    Moderate-intensity continuous training (MICT) improves peripheral artery function in healthy adults, a phenomenon that reverses as continued training induces structural remodeling. Sprint interval training (SIT) elicits physiological adaptations similar to MICT, despite a lower exercise volume and time commitment; however, its effect on peripheral artery function and structure is largely unexplored. We compared peripheral artery responses to 12 wk of MICT and SIT in sedentary, healthy men (age = 27 ± 8 yr). Participants performed MICT (45 min of cycling at 70% peak heart rate; n = 10) or SIT (3 × 20-s "all out" cycling sprints with 2 min of recovery; n = 9), and responses were compared with a nontraining control group (CTL, n = 6). Allometrically scaled brachial flow-mediated dilation (FMD) increased 2.2% after 6 wk of MICT and returned to baseline levels by 12 wk, but did not change in SIT or CTL (group × time interaction, P = 0.04). Brachial artery diameter increased after 6 and 12 wk (main effect, P = 0.03), with the largest increases observed in MICT. Neither training protocol affected popliteal relative FMD and diameter, or central and lower limb arterial stiffness (carotid distensibility, central and leg pulse wave velocity) (P > 0.05 for all). Whereas earlier and more frequent measurements are needed to establish the potential presence and time course of arterial responses to low-volume SIT, our findings suggest that MICT was superior to the intense, but brief and intermittent SIT stimulus at inducing brachial artery responses in healthy men.NEW & NOTEWORTHY We compared the effects of 12 wk of moderate-intensity continuous training (MICT) and sprint interval training (SIT) on peripheral artery endothelial function and diameter, and central and lower limb stiffness in sedentary, healthy men. Whereas neither training program affected the popliteal artery or stiffness indexes, we observed changes in brachial artery function and diameter with MICT but not SIT

  16. p27 Nuclear localization and growth arrest caused by perlecan knockdown in human endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Sakai, Katsuya; Oka, Kiyomasa [Kringle Pharma Joint Research Division for Regenerative Drug Discovery, Center for Advanced Science and Innovation, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871 (Japan); Matsumoto, Kunio [Division of Tumor Dynamics and Regulation, Cancer Research Institute, Kanazawa University, 13-1 Takaramachi, Kanazawa 920-0934 (Japan); Nakamura, Toshikazu, E-mail: nakamura@casi.osaka-u.ac.jp [Kringle Pharma Joint Research Division for Regenerative Drug Discovery, Center for Advanced Science and Innovation, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871 (Japan)

    2010-02-12

    Perlecan, a secreted heparan sulfate proteoglycan, is a major component of the vascular basement membrane and participates in angiogenesis. Here, we used small interference RNA-mediated knockdown of perlecan expression to investigate the regulatory function of perlecan in the growth of human vascular endothelial cells. Basic fibroblast growth factor (bFGF)-induced ERK phosphorylation and cyclin D1 expression were unchanged by perlecan deficiency in endothelial cells; however, perlecan deficiency inhibited the Rb protein phosphorylation and DNA synthesis induced by bFGF. By contrast to cytoplasmic localization of the cyclin-dependent kinase inhibitor p27 in control endothelial cells, p27 was localized in the nucleus and its expression increased in perlecan-deficient cells, which suggests that p27 mediates inhibition of Rb phosphorylation. In addition to the well-characterized function of perlecan as a co-receptor for heparin-binding growth factors such as bFGF, our results suggest that perlecan plays an indispensible role in endothelial cell proliferation and acts through a mechanism that involves subcellular localization of p27.

  17. Effects of depolarizing or non-depolarizing preservation solutions on human endothelial cells during cold hypoxia.

    Science.gov (United States)

    Hidalgo, M A; Mann, D J; Fuller, B J; Green, C J

    1996-02-01

    1. Hypothermic storage of whole organs flushed with a preservation solution is common practice in clinical transplantation. This procedure leaves vascular endothelial cells in direct contact with the preservation solution during the length of the cold ischaemic period. 2. Aiming to study the effects of organ preservation on vascular endothelium, we subjected cultures of human umbilical vein endothelial cells to hypoxic and hypothermic storage conditions in vitro for 3 or 16 h. Four preservation solutions with different levels of sodium and potassium were tested. Morphometric analysis and 51Cr leakage index were used to assess monolayer continuity, cell viability and membrane integrity. 3. Hypothermic storage resulted in severe changes in endothelial cell morphology with formation of intercellular gaps that destroyed monolayer continuity after only 3h. Cellular blebbing was a common feature in seriously damaged cells. 4. Morphometric analysis and 51Cr leakage results correlated well. No significant differences between the solutions tested were found after 3h of hypothermic hypoxic storage. After 16h, viability and monolayer continuity were significantly better preserved (Mann-Whitney, P storage, vascular endothelial cells appeared morphologically deformed and poorly attached in vitro. Lactobionate-based preservation solutions were more effective in preserving viability and continuity. Protection of vascular endothelium under cold hypoxic conditions could be a critical factor in successfully preserving organs for transplantation.

  18. eNOS-dependent antisenscence effect of a calcium channel blocker in human endothelial cells.

    Directory of Open Access Journals (Sweden)

    Toshio Hayashi

    Full Text Available Senescence of vascular endothelial cells is an important contributor to the pathogenesis of age-associated vascular disorders such as atherosclerosis. We investigated the effects of antihypertensive agents on high glucose-induced cellular senescence in human umbilical venous endothelial cells (HUVECs. Exposure of HUVECs to high glucose (22 mM for 3 days increased senescence-associated- β-galactosidase (SA-β-gal activity, a senescence marker, and decreased telomerase activity, a replicative senescence marker. The calcium channel blocker nifedipine, but not the β1-adrenergic blocking agent atenolol or the angiotensin-converting enzyme inhibitor perindopril, reduced SA-β-gal positive cells and prevented a decrease in telomerase activity in a high-glucose environment. This beneficial effect of nifedipine was associated with reduced reactive oxygen species (ROS and increased endothelial nitric oxide synthase (eNOS activity. Thus, nifedipine prevented high glucose-induced ROS generation and increased basal eNOS phosphorylation level at Ser-1177. Treatment with N (G-nitro-L-arginine (L-NAME and transfection of small interfering RNA (siRNA targeting eNOS eliminated the anti-senscence effect of nifedipine. These results demonstrate that nifedipine can prevent endothelial cell senescence in an eNOS-dependent manner. The anti-senescence action of nifedipine may represent a novel mechanism by which it protects against atherosclerosis.

  19. Urea immunoliposome inhibits human vascular endothelial cell proliferation for hemangioma treatment

    Science.gov (United States)

    2013-01-01

    Background Urea injection has been used in hemangioma treatment as sclerotherapy. It shrinks vascular endothelial cells and induces degeneration, necrosis, and fibrosis. However, this treatment still has disadvantages, such as lacking targeting and difficulty in controlling the urea dosage. Thus, we designed a urea immunoliposome to improve the efficiency of treatment. Methods The urea liposome was prepared by reverse phase evaporation. Furthermore, the urea immunoliposome was generated by coupling the urea liposome with a vascular endothelial growth factor receptor (VEGFR) monoclonal antibody using the glutaraldehyde cross-linking method. The influence of the urea immunoliposome on cultured human hemangioma vascular endothelial cells was observed preliminarily. Results Urea immunoliposomes showed typical liposome morphology under a transmission electron microscope, with an encapsulation percentage of 54.4% and a coupling rate of 36.84% for anti-VEGFR. Treatment with the urea immunoliposome significantly inhibited the proliferation of hemangioma vascular endothelial cells (HVECs) in a time- and dose-dependent manner. Conclusions The urea immunoliposome that we developed distinctly and persistently inhibited the proliferation of HVECs and is expected to be used in clinical hemangioma treatment. PMID:24266957

  20. Compartmentalization of vascular endothelial growth factor to the epithelial surface of the human lung.

    Science.gov (United States)

    Kaner, R J; Crystal, R G

    2001-04-01

    Based on assessment of mRNA expression, the lung is a major site of expression of the vascular endothelial growth factor (VEGF) gene, largely from type II alveolar epithelial cells. With the knowledge that VEGF can function to induce vascular leak, we hypothesized that to protect the lung from pulmonary edema, the VEGF produced in the lung must be compartmentalized from the pulmonary endothelium, and thus must be compartmentalized to the surface of the respiratory epithelium. To assess this hypothesis, we quantified the levels of VEGF in human respiratory epithelial lining fluid recovered by bronchoalveolar lavage from normal individuals. Strikingly, human respiratory epithelial lining fluid contains 11 +/- 5 ng/mL as quantified by ELISA, a 500-fold greater concentration than plasma (22 +/- 10 pg/mL, p Damocles sword" poised to induce lung endothelial permeability in conditions of acute lung injury when the integrity of the alveolar epithelial barrier is breached.

  1. Allopurinol protects human glomerular endothelial cells from high glucose-induced reactive oxygen species generation, p53 overexpression and endothelial dysfunction.

    Science.gov (United States)

    Eleftheriadis, Theodoros; Pissas, Georgios; Antoniadi, Georgia; Liakopoulos, Vassilios; Stefanidis, Ioannis

    2017-11-01

    Mitochondrial reactive oxygen species (ROS) overproduction in capillary endothelial cells is a prerequisite for the development of diabetic nephropathy. Inhibition of xanthine oxidase, another ROS generator, ameliorates experimental diabetic nephropathy. To test the hypothesis that the initial high glucose-induced ROS production by the mitochondria activates xanthine oxidase, which afterward remains as the major source of ROS, we cultured primary human glomerular endothelial cells (GEnC) under normal or high-glucose conditions, with or without the xanthine oxidase inhibitor allopurinol. ROS generation and nitric oxide synthase (NOS) activity were assessed by chemiluminescence or colorimetrically. Levels of intercellular adhesion molecule 1 (ICAM-1), p53 and phosphorylated p53 (p-p53) were assessed by western blotting. Allopurinol prevented high glucose-induced ROS generation indicating that xanthine oxidase is the major source of ROS. Allopurinol protected GEnC from endothelial dysfunction since it prevented the high glucose-induced decrease in NOS activity and increase in ICAM-1 expression. Allopurinol reduced p53 and p-p53 levels induced by high glucose suggesting an axis of xanthine oxidase-derived ROS, DNA damage, p53 stabilization and endothelial dysfunction that may contribute to the pathogenesis of diabetic nephropathy. Allopurinol protects GEnC from high glucose-induced ROS generation, p53 overexpression and endothelial dysfunction. These data provide a pathogenetic mechanism that supports the results of experimental and clinical studies about the beneficial effect of xanthine oxidase inhibitors on the development of diabetic nephropathy.

  2. Study of intracellular signaling pathways triggered by natural antioxidants in human endothelial cells

    OpenAIRE

    Cossu, Annalisa

    2012-01-01

    Cardiovascular (CV) benefits of Natural Antioxidant (NA) supplementation are contradictory. Endothelial Cells (EC) are pivotal player on CV diseases onset/progression and thus represent a good model to study the NA impact on vascular pathophysiology. We show that two NA, coumaric acid and resveratrol, affect intracellular ROS levels and cell physiology in human EC. While at lower doses both compounds were antioxidant, at mildly high doses they became pro-oxidant, eliciting cell death by apopt...

  3. Characterization of Influenza Virus-Induced Leukocyte Adherence to Human Umbilical Vein Endothelial Cell Monolayers

    Science.gov (United States)

    1993-07-01

    with other viruses. HL-60 cell adherence to endothelial cell virus type A, which did not infect human venous or bovine monolayers was modulated by...LEUCOCYTE ADHERENC:E TO [NDOTIIELIL (FS1% A. B reawsd on parainfluenza virus-infected airway epithelial Poiy-iiysine Codled IPLC) Wells PLC.Wells cells...an antibody against ICAN1- I has no significant effect PLC Wells Virus on parainfluenza -induced neutrophil adherence (58). In 25 *HSV-intected HUVEC

  4. 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...

  5. CXCL12 enhances angiogenesis through CXCR7 activation in human umbilical vein endothelial cells

    OpenAIRE

    ZHANG Min; Qiu, Lisha; Zhang, Yanyan; Xu, Dongsheng; Zheng, Jialin C.; Jiang, Li

    2017-01-01

    Angiogenesis is the process by which new vessels form from existing vascular networks. Human umbilical vein endothelial cells (HUVECs) may contribute to the study of vascular repair and angiogenesis. The chemokine CXCL12 regulates multiple cell functions, including angiogenesis, mainly through its receptor CXCR4. In contrast to CXCL12/CXCR4, few studies have described roles for CXCR7 in vascular biology, and the downstream mechanism of CXCR7 in angiogenesis remains unclear. The results of the...

  6. Experience With Intravascular Ultrasound Imaging Of Human Atherosclerotic Arteries

    Science.gov (United States)

    Mallery, John A.; Gessert, James M.; Maciel, Mario; Tobis, John M.; Griffith, James M.; Berns, Michael W.; Henry, Walter L.

    1989-08-01

    Normal human arteries have a well-defined structure on intravascular images. The intima appears very thin and is most likely represented by a bright reflection arising from the internal elastic lamina. The smooth muscle tunica media is echo-lucent on the ultrasound image and appears as a dark band separating the intima from the adventitia. The adventitia is a brightly reflective layer of variable thickness. The thickness of the intima, and therefore of the atherosclerotic plaque can be accurately measured from the ultrasound images and correlates well with histology. Calcification within the wall of arteries is seen as bright echo reflection with shadowing of the peripheral wall. Fibrotic regions are highly reflective but do not shadow. Necrotic liquid regions within advanced atherosclerotic plaques are seen on ultrasound images as large lucent zones surrounded by echogenic tissue. Imaging can be performed before and after interventional procedures, such as laser angioplasty, balloon angioplasty and atherectomy. Intravascular ultrasound appears to provide an imaging modality for identifying the histologic characteristics of diseased arteries and for quantifying plaque thickness. It might be possible to perform such quantification to evaluate the results of interventional procedures.

  7. Construction and analysis of SSH cDNA library of human vascular endothelial cells related to gastrocarcinoma

    OpenAIRE

    Liu, Yong-Bo; Wei, Zhao-Xia; Li, Li; Li, Hang-Sheng; Chen, Hui; Li, Xiao-Wen

    2003-01-01

    AIM: To construct subtracted cDNA libraries of human vascular endothelial cells (VECs) related to gastrocarcinoma using suppression substractive hybridization (SSH) and to analyze cDNA libraries of gastrocarcinoma and VECs in Cancer Gene Anatomy Project (CGAP) database.

  8. Characterization of ionizing radiation-induced unfolded protein response in human vascular endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Eun Ju; Lee, Yoon Jin; Kang, Seong Man [Laboratory of Radiation Effect, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2013-04-15

    Misfolded or unfolded proteins within the endoplasmic reticulum (ER stress), viral infection, or amino acid deprivation induce eukaryotic translation initiation factor 2α phosphorylation (eIF2α) in eukaryotic cells, repressing global protein synthesis coincident with preferential translation of activating transcription factor 4 (ATF4). ATF4 is a transcriptional activator of genes involved in amino acid metabolism, cellular redox homeostasis, and regulation of apoptosis. When the eIF2α/ATF4 pathway is initiated by ER stress, the pathway is referred toas the unfolded protein response (UPR). In addition to DNA, proteins may be initial and important targets of ionizing radiation (IR), and the damaged protein can trigger ER stress pathway. Recent investigations suggested that IR induces ER stress followed by UPR in various cell types including intestinal epithelial cells. We conducted this study to determine whether IR can activate UPR in human vascular endothelial cells. Our data have shown that IR increased PERK-dependent eIF2α phosphorylation accompanied by induction in ATF4 protein levels in human vascular endothelial cells without alterations in expressions of XBP-1s and GRP78. Based on these data, we suggest that IR selectively activates PERK branch of unfolded protein response in human vascular endothelial cells.

  9. Human Haemato-Endothelial Precursors: Cord Blood CD34+ Cells Produce Haemogenic Endothelium

    Science.gov (United States)

    Pelosi, Elvira; Castelli, Germana; Martin-Padura, Ines; Bordoni, Veronica; Santoro, Simona; Conigliaro, Alice; Cerio, Anna Maria; De Santis Puzzonia, Marco; Marighetti, Paola; Biffoni, Mauro; Alonzi, Tonino; Amicone, Laura; Alcalay, Myriam; Bertolini, Francesco; Testa, Ugo; Tripodi, Marco

    2012-01-01

    Embryologic and genetic evidence suggest a common origin of haematopoietic and endothelial lineages. In the murine embryo, recent studies indicate the presence of haemogenic endothelium and of a common haemato-endothelial precursor, the haemangioblast. Conversely, so far, little evidence supports the presence of haemogenic endothelium and haemangioblasts in later stages of development. Our studies indicate that human cord blood haematopoietic progenitors (CD34+45+144−), triggered by murine hepatocyte conditioned medium, differentiate into adherent proliferating endothelial precursors (CD144+CD105+CD146+CD31+CD45−) capable of functioning as haemogenic endothelium. These cells, proven to give rise to functional vasculature in vivo, if further instructed by haematopoietic growth factors, first switch to transitional CD144+45+ cells and then to haematopoietic cells. These results highlight the plasticity of haemato-endhothelial precursors in human post-natal life. Furthermore, these studies may provide highly enriched populations of human post-fetal haemogenic endothelium, paving the way for innovative projects at a basic and possibly clinical level. PMID:23226561

  10. Human haemato-endothelial precursors: cord blood CD34+ cells produce haemogenic endothelium.

    Directory of Open Access Journals (Sweden)

    Elvira Pelosi

    Full Text Available Embryologic and genetic evidence suggest a common origin of haematopoietic and endothelial lineages. In the murine embryo, recent studies indicate the presence of haemogenic endothelium and of a common haemato-endothelial precursor, the haemangioblast. Conversely, so far, little evidence supports the presence of haemogenic endothelium and haemangioblasts in later stages of development. Our studies indicate that human cord blood haematopoietic progenitors (CD34+45+144-, triggered by murine hepatocyte conditioned medium, differentiate into adherent proliferating endothelial precursors (CD144+CD105+CD146+CD31+CD45- capable of functioning as haemogenic endothelium. These cells, proven to give rise to functional vasculature in vivo, if further instructed by haematopoietic growth factors, first switch to transitional CD144+45+ cells and then to haematopoietic cells. These results highlight the plasticity of haemato-endhothelial precursors in human post-natal life. Furthermore, these studies may provide highly enriched populations of human post-fetal haemogenic endothelium, paving the way for innovative projects at a basic and possibly clinical level.

  11. 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.

  12. Bradykinin-evoked changes in cytosolic calcium and membrane currents in cultured bovine pulmonary artery endothelial cells.

    Science.gov (United States)

    Cannell, M B; Sage, S O

    1989-12-01

    1. Cultured bovine pulmonary artery endothelial cells were voltage clamped using a single microelectrode while cytosolic free calcium concentration ([Ca2+]i) was simultaneously measured using the fluorescent calcium indicator, Indo-1. 2. The resting current-voltage relationship was non-linear and exhibited marked inward rectification near the resting potential. In about 60% of cells examined, superfusion of saline resulted in a hyperpolarization and decrease in inward current. This result did not depend on the presence of agonist and is consistent with the presence of extracellular potassium accumulation in restricted spaces around the cell and the known dependence of the inward rectifier. In other cells there was no effect. 3. Resting [Ca2+]i was sensitive to membrane potential, decreasing continuously with membrane depolarization over the range -70 to +60 mV. This result is consistent with a simple pump-leak model and suggests that voltage-dependent calcium channels are not present in these cells. 4. Bradykinin (10 microM) increased [Ca2+]i after a delay of approximately 3 s. [Ca2+]i reached a peak after a further 3 s and declined over several minutes. 5. During the rise in [Ca2+]i evoked by application of bradykinin, there were no changes in the current-voltage relationship of the cell. These results question the role of a receptor-operated non-selective cation channel in mediating the increase in [Ca2+]i. This observation, coupled with the observed delay in the agonist-evoked response suggests that a second messenger system is involved in mediating the increase in [Ca2+]i. 6. Changes in the current-voltage relation started to occur about 30 s after the application of agonist. These changes could be explained by the activation of large-conductance potassium and non-selective cation channels with a reversal potential near 0 mV. The latter channels may mediate the plateau phase of the agonist-evoked response. 7. The results are discussed with respect to the

  13. Association of Endothelial Nitric Oxide Synthase Gene Polymorphisms with Coronary Artery Disease: An Updated Meta-Analysis and Systematic Review

    Science.gov (United States)

    Parveen, Farah; Kapoor, Aditya; Sinha, Nakul

    2014-01-01

    Several association studies of endothelial nitric oxide synthase (NOS3) gene polymorphisms with respect to coronary artery disease (CAD) have been published in the past two decades. However, their association with the disease, especially among different ethnic subgroups, still remains controversial. This prompted us to conduct a systematic review and an updated structured meta-analysis, which is the largest so far (89 articles, 132 separate studies, and a sample size of 69,235), examining association of three polymorphic forms of the NOS3 gene (i.e. Glu298Asp, T786-C and 27bp VNTR b/a) with CAD. In a subgroup analysis, we tested their association separately among published studies originating predominantly from European, Middle Eastern, Asian, Asian-Indian and African ancestries. The pooled analysis confirmed the association of all the three selected SNP with CAD in three different genetic models transcending all ancestries worldwide. The Glu298Asp polymorphism showed strongest association (OR range = 1.28–1.52, and P<0.00001 for all comparisons), followed by T786-C (OR range = 1.34–1.42, and P<0.00001 for all comparisons) and 4b/a, (OR range = 1.19–1.41, and P≤0.002 for all comparisons) in our pooled analysis. Subgroup analysis revealed that Glu298Asp (OR range = 1.54–1.87, and P<0.004 for all comparisons) and 4b/a (OR range = 1.71–3.02, and P<0.00001 for all comparisons) have highest degree of association amongst the Middle Easterners. On the other hand, T786-C and its minor allele seem to carry a highest risk for CAD among subjects of Asian ancestry (OR range = 1.61–1.90, and P≤0.01 for all comparisons). PMID:25409023

  14. Isolation of human lymphatic malformation endothelial cells, their in vitro characterization and in vivo survival in a mouse xenograft model.

    Science.gov (United States)

    Lokmic, Zerina; Mitchell, Geraldine M; Koh Wee Chong, Nicholas; Bastiaanse, Jacqueline; Gerrand, Yi-Wen; Zeng, Yiping; Williams, Elizabeth D; Penington, Anthony J

    2014-01-01

    Human lymphatic vascular malformations (LMs), also known as cystic hygromas or lymphangioma, consist of multiple lymphatic endothelial cell-lined lymph-containing cysts. No animal model of this disease exists. To develop a mouse xenograft model of human LM, CD34(Neg)CD31(Pos) LM lymphatic endothelial cells (LM-LEC) were isolated from surgical specimens and compared to foreskin CD34(Neg)CD31(Pos) lymphatic endothelial cells (LECs). Cells were implanted into a mouse tissue engineering model for 1, 2 and 4 weeks. In vitro LM-LECs showed increased proliferation and survival under starvation conditions (P lymphatic malformations.

  15. Isoflurane Protects Against Human Endothelial Cell Apoptosis by Inducing Sphingosine Kinase-1 via ERK MAPK

    Directory of Open Access Journals (Sweden)

    H. Thomas Lee

    2012-01-01

    Full Text Available Endothelial dysfunction is a major clinical problem affecting virtually every patient requiring critical care. Volatile anesthetics are frequently used during the perioperative period and protect the heart and kidney against ischemia and reperfusion injury. We aimed to determine whether isoflurane, the most commonly used volatile anesthetic in the USA, protects against endothelial apoptosis and necrosis and the mechanisms involved in this protection. Human endothelial EA.hy926 cells were pretreated with isoflurane or carrier gas (95% room air + 5% CO2 then subjected to apoptosis with tumor necrosis factor-α or to necrosis with hydrogen peroxide. DNA laddering and in situ Terminal Deoxynucleotidyl Transferase Biotin-dUTP Nick-End Labeling (TUNEL staining determined EA.hy926 cell apoptosis and percent LDH released determined necrosis. We also determined whether isoflurane modulates the expression and activity of sphingosine kinase-1 (SK1 and induces the phosphorylation of extracellular signal regulated kinase (ERK MAPK as both enzymes are known to protect against cell death. Isoflurane pretreatment significantly decreased apoptosis in EA.hy926 cells as evidenced by reduced TUNEL staining and DNA laddering without affecting necrosis. Mechanistically, isoflurane induces the phosphorylation of ERK MAPK and increased SK1 expression and activity in EA.hy926 cells. Finally, selective blockade of SK1 (with SKI-II or S1P1 receptor (with W146 abolished the anti-apoptotic effects of isoflurane. Taken together, we demonstrate that isoflurane, in addition to its potent analgesic and anesthetic properties, protects against endothelial apoptosis most likely via SK1 and ERK MAPK activation. Our findings have significant clinical implication for protection of endothelial cells during the perioperative period and patients requiring critical care.

  16. Impaired Endothelial Regeneration Through Human Parvovirus B19-Infected Circulating Angiogenic Cells in Patients With Cardiomyopathy.

    Science.gov (United States)

    Schmidt-Lucke, Caroline; Zobel, Thomas; Schrepfer, Sonja; Kuhl, Uwe; Wang, Dong; Klingel, Karin; Becher, Peter Moritz; Fechner, Henry; Pozzuto, Tanja; Van Linthout, Sophie; Lassner, Dirk; Spillmann, Frank; Escher, Felicitas; Holinski, Sebastian; Volk, Hans-Dieter; Schultheiss, Heinz-Peter; Tschope, Carsten

    2015-10-01

    Human parvovirus B19 (B19V) is a common pathogen in microvascular disease and cardiomyopathy, owing to infection of endothelial cells. B19V replication, however, is almost restricted to erythroid progenitor cells (ErPCs). Endothelial regeneration attributable to bone marrow-derived circulating angiogenic cells (CACs) is a prerequisite for organ function. Because of many similarities of ErPCs and CACs, we hypothesized that B19V is a perpetrator of impaired endogenous endothelial regeneration. B19V DNA and messenger RNA from endomyocardial biopsy specimens, bone marrow specimens, and circulating progenitor cells were quantified by polymerase chain reaction analysis. The highest B19V DNA concentrations were found in CD34(+)KDR(+) cells from 17 patients with chronic B19V-associated cardiomyopathy. B19V replication intermediates could be detected in nearly half of the patients. Furthermore, chronic B19V infection was associated with impaired endothelial regenerative capacity. B19V infection of CACs in vitro resulted in expression of transcripts encoding B19V proteins. The capsid protein VP1 was identified as a novel inducer of apoptosis, as were nonstructural proteins. Inhibition studies identified so-called death receptor signaling with activation of caspase-8 and caspase-10 to be responsible for apoptosis induction. B19V causally impaired endothelial regeneration with spreading of B19V in CACs in an animal model in vivo. We thus conclude that B19V infection and damage to CACs result in dysfunctional endogenous vascular repair, supporting the emergence of primary bone marrow disease with secondary end-organ damage. © The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  17. Niacin receptor activation improves human microvascular endothelial cell angiogenic function during lipotoxicity.

    Science.gov (United States)

    Hughes-Large, Jennifer M; Pang, Dominic K T; Robson, Debra L; Chan, Pak; Toma, Jelena; Borradaile, Nica M

    2014-12-01

    Niacin (nicotinic acid) as a monotherapy can reduce vascular disease risk, but its mechanism of action remains controversial, and may not be dependent on systemic lipid modifying effects. Niacin has recently been shown to improve endothelial function and vascular regeneration, independent of correcting dyslipidemia, in rodent models of vascular injury and metabolic disease. As a potential biosynthetic precursor for NAD(+), niacin could elicit these vascular benefits through NAD(+)-dependent, sirtuin (SIRT) mediated responses. Alternatively, niacin may act through its receptor, GPR109A, to promote endothelial function, though endothelial cells are not known to express this receptor. We hypothesized that niacin directly improves endothelial cell function during exposure to lipotoxic conditions and sought to determine the potential mechanism(s) involved. Angiogenic function in excess palmitate was assessed by tube formation following treatment of human microvascular endothelial cells (HMVEC) with either a relatively low concentration of niacin (10 μM), or nicotinamide mononucleotide (NMN) (1 μM), a direct NAD(+) precursor. Although both niacin and NMN improved HMVEC tube formation during palmitate overload, only NMN increased cellular NAD(+) and SIRT1 activity. We further observed that HMVEC express GRP109A. Activation of this receptor with either acifran or MK-1903 recapitulated niacin-induced improvements in HMVEC tube formation, while GPR109A siRNA diminished the effect of niacin. Niacin, at a low concentration, improves HMVEC angiogenic function under lipotoxic conditions, likely independent of NAD(+) biosynthesis and SIRT1 activation, but rather through niacin receptor activation. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  18. Shear resistance of human umbilical endothelial cells on different materials covered with or without extracellular matrix: controlled in-vitro study.

    Science.gov (United States)

    Hoepken, S; Fuhrmann, R; Jung, F; Franke, R P

    2009-01-01

    A variety of medical grade polymeric materials are used in tissue engineering and biomedical technology. Dense non-porous polymeric foils were used as substrates for endothelial cell layers. Half a the test samples (polymers and control materials) were seeded with bovine corneal endothelial cells (BCEC) which more or less covered the substrates with an extracellular matrix (ECM) in the consecutive culturing period. Afterwards the ECM covered as well as the uncovered materials were seeded with human umbilical venous endothelial cells (HUVEC). HUVEC seeded samples were cultured either under static or under dynamical conditions in a cone/plate rheometer with a mean low arterial shear stress of 8.2 dyn/cm2 to simulate the flow conditions in a coronary vein graft. With the exemption of polyvinyl chloride all other materials could be coated with ECM at least partially. Under static conditions the best results with respect to complete coverage with ECM and HUVEC were seen on polyester and polyurethane. Under shear load, however, the complete HUVEC layer together with the ECM detached from the polymer surface within a short time. ECM and HUVEC remained no longer than 43 minutes on anyone of the materials tested. The materials as supplied and tested were clearly not appropriate as implants in contact to the flowing blood.

  19. Stretch-induced human myometrial cytokines enhance immune cell recruitment via endothelial activation.

    Science.gov (United States)

    Lee, Yu-Hui; Shynlova, Oksana; Lye, Stephen J

    2015-03-01

    Spontaneous term labour is associated with amplified inflammatory events in the myometrium including cytokine production and leukocyte infiltration; however, potential mechanisms regulating such events are not fully understood. We hypothesized that mechanical stretch of the uterine wall by the growing fetus facilitates peripheral leukocyte extravasation into the term myometrium through the release of various cytokines by uterine myocytes. Human myometrial cells (hTERT-HM) were subjected to static mechanical stretch; stretch-conditioned media was collected and analysed using 48-plex Luminex assay and ELISA. Effect of stretch-conditioned media on cell adhesion molecule expression of human uterine microvascular endothelial cells (UtMVEC-Myo) was detected by quantitative polymerase chain reaction (qPCR) and flow cytometry; functional assays testing leukocyte-endothelial interactions: adhesion of leukocytes to endothelial cells and transendothelial migration of calcein-labelled primary human neutrophils as well as migration of THP-1 monocytic cells were assessed by fluorometry. The current in vitro study demonstrated that mechanical stretch (i) directly induces secretion of multiple cytokines and chemokines by hTERT-HM cells (IL-6, CXCL8, CXCL1, migration inhibitory factor (MIF), VEGF, G-CSF, IL-12p70, bFGF and platelet-derived growth factor subunit B (PDGF-bb), Pcytokines (ii) enhance leukocyte adhesion to the endothelium of the surrounding uterine microvasculature by (iii) inducing the expression of endothelial cell adhesion molecules and (iv) directing the transendothelial migration of peripheral leukocytes. (vi) Chemokine-neutralizing antibodies and broad-spectrum chemokine inhibitor block leukocyte migration. Our data provide a proof of mechanical regulation for leukocyte recruitment from the uterine blood vessels to the myometrium, suggesting a putative mechanism for the leukocyte infiltrate into the uterus during labour and postpartum involution.

  20. Stretch-induced human myometrial cytokines enhance immune cell recruitment via endothelial activation

    Science.gov (United States)

    Lee, Yu-Hui; Shynlova, Oksana; Lye, Stephen J

    2015-01-01

    Spontaneous term labour is associated with amplified inflammatory events in the myometrium including cytokine production and leukocyte infiltration; however, potential mechanisms regulating such events are not fully understood. We hypothesized that mechanical stretch of the uterine wall by the growing fetus facilitates peripheral leukocyte extravasation into the term myometrium through the release of various cytokines by uterine myocytes. Human myometrial cells (hTERT-HM) were subjected to static mechanical stretch; stretch-conditioned media was collected and analysed using 48-plex Luminex assay and ELISA. Effect of stretch-conditioned media on cell adhesion molecule expression of human uterine microvascular endothelial cells (UtMVEC-Myo) was detected by quantitative polymerase chain reaction (qPCR) and flow cytometry; functional assays testing leukocyte–endothelial interactions: adhesion of leukocytes to endothelial cells and transendothelial migration of calcein-labelled primary human neutrophils as well as migration of THP-1 monocytic cells were assessed by fluorometry. The current in vitro study demonstrated that mechanical stretch (i) directly induces secretion of multiple cytokines and chemokines by hTERT-HM cells (IL-6, CXCL8, CXCL1, migration inhibitory factor (MIF), VEGF, G-CSF, IL-12p70, bFGF and platelet-derived growth factor subunit B (PDGF-bb), Pstretch-induced cytokines (ii) enhance leukocyte adhesion to the endothelium of the surrounding uterine microvasculature by (iii) inducing the expression of endothelial cell adhesion molecules and (iv) directing the transendothelial migration of peripheral leukocytes. (vi) Chemokine-neutralizing antibodies and broad-spectrum chemokine inhibitor block leukocyte migration. Our data provide a proof of mechanical regulation for leukocyte recruitment from the uterine blood vessels to the myometrium, suggesting a putative mechanism for the leukocyte infiltrate into the uterus during labour and postpartum involution

  1. The impact of the ketogenic diet on arterial morphology and endothelial function in children and young adults with epilepsy: a case-control study.

    Science.gov (United States)

    Coppola, Giangennaro; Natale, Francesco; Torino, Annarita; Capasso, Rosanna; D'Aniello, Alfredo; Pironti, Erica; Santoro, Elena; Calabrò, Raffaele; Verrotti, Alberto

    2014-04-01

    The present study aimed to assess the impact of the ketogenic diet on arterial morphology and endothelial function of the big vessels of the neck and on cardiac diastolic function, in a cohort of epileptic children and young adults treated with the ketogenic diet. Patients were recruited based on the following inclusion criteria: (1) patients who were or had been on the ketogenic diet for a time period of at least six months. Each patient underwent measurement of carotid intima media thickness, carotid artery stiffness, echocardiography, and diastolic function assessment. Patients with drug resistant epilepsy, matched for number, age and sex and never treated with ketogenic diet, were recruited as controls. The population study was composed by 43 epilepsy patients (23 males), aged between 19 months and 31 years (mean 11 years). Twenty-three patients were or had been treated with ketogenic diet, and 20 had never been on it (control group). Subjects treated with the ketogenic diet had higher arterial stiffness parameters, including AIx and β-index and higher serum levels of cholesterol or triglycerides compared to those who had never been on the diet (control group) (pketogenic diet, before the increase of the intima media thickness. This supports that arterial stiffness is an early marker of vascular damage. Copyright © 2013 British Epilepsy Association. Published by Elsevier Ltd. All rights reserved.

  2. Endothelial dysfunction and smoking factor in patients with coronary heart disease while testing the radial artery prior to coronary bypass surgery

    Directory of Open Access Journals (Sweden)

    O. V. Kamenskaya

    2015-10-01

    Full Text Available Patients (111 males with coronary heart disease (CHD and angina, II-III functional class, accompanied by arterial hypertension (AH, I-III degree, in 77 % of cases (mean age was 62.00.76 years were examined prior to coronary bypass surgery. The patients were divided into two groups: group 1 consisted of 56 nonsmoking patients (112 upper extremities and group 2 - 55 smoking patients (110 upper extremities with the experience of smoking being 331.6 years on average. The thumb of both the right and left hand was tested for peripheral microcirculatory blood flow (MBF by using a laser-Doppler flowmetry method (LDF. This quantitative method was developed at our clinic to assess the adequacy of collateral circulation in the hand. The method compares MBF provided by the ulnar artery, with the radial artery occluded, with the background level. Some features of various clinical parameters in the group of smokers suffering from coronary artery disease were analyzed in comparison with those of non-smokers. Significant differences were found in the indicators of lipid profile: increased levels of total cholesterol and low-density lipoprotein (LDL against the background of low high-density lipoprotein (HDL in the group of smoking patients. We found out that the smokers had to apply for surgical treatment by 7 years (on average earlier; they had a significantly greater number of heart attacks and showed the lowest body mass index (BMI. The LDF test helped to establish that smoking in patients with CHD enhances the endothelial dysfunction, while significantly reducing the reactivity of the microvascular bed under the influence of hyperemic stimulus. Sampling the radial artery for coronary artery bypass grafting in smokers tended to give negative results.

  3. Effect of intravitreal anti-vascular endothelial growth factor therapy on the risk of arterial thromboembolic events: a meta-analysis.

    Directory of Open Access Journals (Sweden)

    Jin-Wei Cheng

    Full Text Available BACKGROUND: Intravitreal anti-vascular endothelial growth factor (VEGF monoclonal antibodies are used in ocular neovascular diseases. A consensus has emerged that intravenous anti-VEGF can increase the risk of arterial thromboembolic events. However, the role of intravitreal anti-VEGF in arterial thromboembolism is controversial. Therefore, we did a systematic review and meta-analysis to investigate the effects of intravitreal anti-VEGF on the risk of arterial thromboembolic events. METHODS: Electronic databases were searched to identify relevant randomized clinical trials comparing intravitreal anti-VEGF with controls. Criteria for inclusion in our meta-analysis included a study duration of no less than 12 months, the use of a randomized control group not receiving any intravitreal active agent, and the availability of outcome data for arterial thromboembolic events, myocardial infarction, cerebrovascular accidents, and vascular death. The risk ratios and 95% CIs were calculated using a fixed-effects or random-effects model, depending on the heterogeneity of the included studies. RESULTS: A total of 4942 patients with a variety of ocular neovascular diseases from 13 randomized controlled trials were identified and included for analysis. There was no significant difference between intravitreal anti-VEGF and control in the risk of all events, with risk ratios of 0.87 (95% CI, 0.64 to 1.19 for arterial thromboembolic events, 0.96 (95% CI, 0.55-1.68 for cerebrovascular accidents, 0.69 (95% CI 0.40-1.21 for myocardial infarctions, and 0.68 (95% CI, 0.37-1.27 for vascular death. CONCLUSIONS: The strength evidence suggests that the intravitreal use of anti-VEGF antibodies is not associated with an increased risk of arterial thromboembolic events.

  4. The use of adipose mesenchymal stem cells and human umbilical vascular endothelial cells on a fibrin matrix for endothelialized skin substitute.

    Science.gov (United States)

    Sánchez-Muñoz, Isabel; Granados, Rosario; Holguín Holgado, Purificación; García-Vela, José Antonio; Casares, Celia; Casares, Miguel

    2015-01-01

    In recent years, the reconstruction of human skin by tissue engineering represents a clinical challenge and has offered a therapeutic alternative. Avascular engineered skin equivalents have been available for several years and used to treat wounds due to burns, nonhealing ulcers, and surgical excisions. They are constituted by different types of cultured cells included in a three-dimensional structure that permits cellular proliferation to create tissue substitutes. The major drawback of these artificial skin substitutes is their lack of blood supply, since the endurance and cell proliferation of the substitute depend on an adequate oxygen and nutrient supply and on toxin removal. These functions are served by the vascular system. We have produced a new model of endothelialized skin substitute that promotes the formation of capillary-like structures by seeding human umbilical vein endothelial cells (HUVECs) with dermal fibroblasts and human adipose-derived mesenchymal stem cells (hADMSCs) in a fibrin matrix. Dermal fibroblasts and hADMSCs produce extracellular matrix that stimulates cellular growth and proliferation. hADMSCs secrete significant quantities of angiogenic and antiapoptotic factors (vascular endothelial growth factor and hepatocyte growth factor), which induce in vitro differentiation of these cells into endothelial cells promoting angiogenesis and participating in tissue repair and skin regeneration processes. We obtained the artificial skin substitute with similar structure to native skin, including dermis and epidermis. We demonstrated that endothelial cells (CD31 and von Willebrand factor positive) proliferated and organized themselves into capillary-like structures within the fibrin matrix. The epidermis showed a complete epithelization by squamous cells (AE1/AE3 cytokeratin positive) with intracytoplasmic keratohyalin granules, hyperkeratosis, and parakeratosis. We have established a novel artificial skin substitute that facilitates the formation

  5. Mapping the distinctive populations of lymphatic endothelial cells in different zones of human lymph nodes.

    Directory of Open Access Journals (Sweden)

    Saem Mul Park

    Full Text Available The lymphatic sinuses in human lymph nodes (LNs are crucial to LN function yet their structure remains poorly defined. Much of our current knowledge of lymphatic sinuses derives from rodent models, however human LNs differ substantially in their sinus structure, most notably due to the presence of trabeculae and trabecular lymphatic sinuses that rodent LNs lack. Lymphatic sinuses are bounded and traversed by lymphatic endothelial cells (LECs. A better understanding of LECs in human LNs is likely to improve our understanding of the regulation of cell trafficking within LNs, now an important therapeutic target, as well as disease processes that involve lymphatic sinuses. We therefore sought to map all the LECs within human LNs using multicolor immunofluorescence microscopy to visualize the distribution of a range of putative markers. PROX1 was the only marker that uniquely identified the LECs lining and traversing all the sinuses in human LNs. In contrast, LYVE1 and STAB2 were only expressed by LECs in the paracortical and medullary sinuses in the vast majority of LNs studied, whilst the subcapsular and trabecular sinuses lacked these molecules. These data highlight the existence of at least two distinctive populations of LECs within human LNs. Of the other LEC markers, we confirmed VEGFR3 was not specific for LECs, and CD144 and CD31 stained both LECs and blood vascular endothelial cells (BECs; in contrast, CD59 and CD105 stained BECs but not LECs. We also showed that antigen-presenting cells (APCs in the sinuses could be clearly distinguished from LECs by their expression of CD169, and their lack of expression of PROX1 and STAB2, or endothelial markers such as CD144. However, both LECs and sinus APCs were stained with DCN46, an antibody commonly used to detect CD209.

  6. [Daily blood pressure profiles and endothelial functions during long-term treatment of arterial hypertension with angiotensin-converting enzyme inhibitors].

    Science.gov (United States)

    Simonenko, V B; Fisun, A Ia; Mikhaĭlov, A A; Kuroedov, A V; Brizhan, M V; Butikov, V P; Tsokolov, A V

    2004-01-01

    Clinical practice has been recently applied new approaches to classifying patients with arterial hypertension (AH), which are based on the determination of the summary risk suggesting the prognosis of the disease. This approach is quite efficient due to the fact that it takes into account a combination of the comparable clinical symptoms of AH, which characterize its course. The paper presents data on the values of blood pressure (BP) in AH and on daily BP profiles (DBPP) in 494 patients with different categories of a risk and on their endothelial function. It also gives the results of achievement of target BP levels in patients with AH having different categories of a risk during long-term (36-month) treatment with enapril and perindopril, as well as data on its impact on endothelial vasomotor function. The findings suggest that the higher the risk is, the more severe the hypertensive syndrome is; that the redistribution of DBPP occurs towards prognostically poor types and endothelial dysfunction (EDF) progresses. Treatment with angiotensin-converting enzyme (ACE) inhibitors results not only to the achievement of target BP levels and to the normalization of DBPP, but also to the amelioration of EDF. Moreover, the co-administration of ACE inhibitors and isosorbide-5-mononitrates causes additional antiischemic effects.

  7. Upregulation of Human Endogenous Retrovirus-K Is Linked to Immunity and Inflammation in Pulmonary Arterial Hypertension.

    Science.gov (United States)

    Saito, Toshie; Miyagawa, Kazuya; Chen, Shih-Yu; Tamosiuniene, Rasa; Wang, Lingli; Sharpe, Orr; Samayoa, Erik; Harada, Daisuke; Moonen, Jan-Renier A J; Cao, Aiqin; Chen, Pin-I; Hennigs, Jan K; Gu, Mingxia; Li, Caiyun G; Leib, Ryan D; Li, Dan; Adams, Christopher M; Del Rosario, Patricia A; Bill, Matthew; Haddad, Francois; Montoya, Jose G; Robinson, William H; Fantl, Wendy J; Nolan, Garry P; Zamanian, Roham T; Nicolls, Mark R; Chiu, Charles Y; Ariza, Maria E; Rabinovitch, Marlene

    2017-11-14

    Immune dysregulation has been linked to occlusive vascular remodeling in pulmonary arterial hypertension (PAH) that is hereditary, idiopathic, or associated with other conditions. Circulating autoantibodies, lung perivascular lymphoid tissue, and elevated cytokines have been related to PAH pathogenesis but without a clear understanding of how these abnormalities are initiated, perpetuated, and connected in the progression of disease. We therefore set out to identify specific target antigens in PAH lung immune complexes as a starting point toward resolving these issues to better inform future application of immunomodulatory therapies. Lung immune complexes were isolated and PAH target antigens were identified by liquid chromatography tandem mass spectrometry, confirmed by enzyme-linked immunosorbent assay, and localized by confocal microscopy. One PAH antigen linked to immunity and inflammation was pursued and a link to PAH pathophysiology was investigated by next-generation sequencing, functional studies in cultured monocytes and endothelial cells, and hemodynamic and lung studies in a rat. SAM domain and HD domain-containing protein 1 (SAMHD1), an innate immune factor that suppresses HIV replication, was identified and confirmed as highly expressed in immune complexes from 16 hereditary and idiopathic PAH versus 12 control lungs. Elevated SAMHD1 was localized to endothelial cells, perivascular dendritic cells, and macrophages, and SAMHD1 antibodies were prevalent in tertiary lymphoid tissue. An unbiased screen using metagenomic sequencing related SAMHD1 to increased expression of human endogenous retrovirus K (HERV-K) in PAH versus control lungs (n=4). HERV-K envelope and deoxyuridine triphosphate nucleotidohydrolase mRNAs were elevated in PAH versus control lungs (n=10), and proteins were localized to macrophages. HERV-K deoxyuridine triphosphate nucleotidohydrolase induced SAMHD1 and proinflammatory cytokines (eg, interleukin 6, interleukin 1β, and tumor

  8. Constitutive modeling of human femoropopliteal artery biaxial stiffening due to aging and diabetes.

    Science.gov (United States)

    Desyatova, Anastasia; MacTaggart, Jason; Kamenskiy, Alexey

    2017-12-01

    Atherosclerotic obstructive disease of the femoropopliteal artery (Peripheral Arterial Disease, PAD) is notorious for high treatment failure rates. Older age and diabetes mellitus (DM) are among the major risk factors for PAD, and both are associated with increased arterial stiffness. Our goal was to develop a constitutive model describing multiaxial arterial stiffening, and use it to portray aging of normal and diabetic human femoropopliteal arteries (FPA). Fresh human FPAs (n=744) were obtained from 13-82-year-old donors. Arteries were tested using planar biaxial extension, and their behavior was modeled with a constitutive relation that included stiffening functions of age. FPA diameter, wall thickness, circumferential, and longitudinal opening angles increased with age, while longitudinal pre-stretch decreased. Diameter and circumferential opening angle did not change with age in subjects with DM. Younger FPAs were more compliant longitudinally but became more isotropic with age. Arteries with DM stiffened significantly faster in the circumferential direction than arteries without DM. Constitutive model accurately portrayed orthotropic stiffening with age of both normal and diabetic arteries. Constitutive description of FPA aging contributes to understanding of arterial pathophysiology and can help improve fidelity of computational models investigating device-artery interaction in PAD repair by providing more personalized arterial properties. We have analyzed n=744 human femoropopliteal artery (FPA) specimens using biaxial tensile testing to derive constitutive description of FPA aging in diabetic and non-diabetic subjects. The proposed model allows determination of FPA mechanical properties for subjects of any given age in the range of 13-82years. These results contribute to understanding of FPA pathophysiology and can help improve fidelity of computational models investigating device-artery interaction in peripheral arterial disease repair by providing more

  9. Ionizing radiation enhances IL-6 and IL-8 production by human endothelial cells

    Directory of Open Access Journals (Sweden)

    A. Van Der Meeren

    1997-01-01

    Full Text Available Irradiation exposure is known to induce an inflammatory reaction. Endothelial cells play a crucial role both in the inflammatory process and in radiation damage. Therefore, supernatants and cell lysates of 60Co-irradiated human umbilical vein endothelial cells (HUVEC have been assessed for the presence of pro-inflammatory cytokines. After gamma irradiation, interleukin (IL-1α, IL-1β and tumor necrosis factor (TNF-α remained undetectable in both cell supernatants and cell lysates. However, a dose-dependent increase in the production of IL-6 and IL-8 has been demonstrated up to 6 days after exposure. These data indicate that the pro-inflammatory cytokines IL-6 and IL-8 may be involved in the inflammatory response of vascular endothelium induced by exposure to ionizing radiation.

  10. The cytotoxicity evaluation of magnetic iron oxide nanoparticles on human aortic endothelial cells

    OpenAIRE

    Ge, Gaoyuan; WU, HENGFANG; Xiong,Fei; Zhang, Yu; Guo, Zhirui; Bian, Zhiping; Xu, Jindan; Gu, Chunrong; Gu, Ning; Chen, Xiangjian; Yang, Di

    2013-01-01

    One major obstacle for successful application of nanoparticles in medicine is its potential nanotoxicity on the environment and human health. In this study, we evaluated the cytotoxicity effect of dimercaptosuccinic acid-coated iron oxide (DMSA-Fe2O3) using cultured human aortic endothelial cells (HAECs). Our results showed that DMSA-Fe2O3 in the culture medium could be absorbed into HAECs, and dispersed in the cytoplasm. The cytotoxicity effect of DMSA-Fe2O3 on HAECs was dose-dependent, and ...

  11. Endothelial cell-borne platelet bridges selectively recruit monocytes in human and mouse models of vascular inflammation.

    Science.gov (United States)

    Kuckleburg, Christopher J; Yates, Clara M; Kalia, Neena; Zhao, Yan; Nash, Gerard B; Watson, Steve P; Rainger, George Ed

    2011-07-01

    Cells of the monocyte lineage are the most abundant inflammatory cells found in atherosclerotic lesions. Dominance of the inflammatory infiltrate by monocytes indicates that there is a disease-driven mechanism supporting their selective recruitment. Previous studies have demonstrated that interactions between endothelial cells (ECs) and platelets may promote monocyte recruitment. In this study, we sought to expand on this knowledge using a complex coculture model of the diseased vessel wall. Using primary human cells in an in vitro flow-based adhesion assay, we found that secretory arterial smooth muscle cells (SMCs), cocultured with ECs, promote preferential recruitment of monocytes from blood in a TGF-β1-dependent manner. Approximately 85% of leucocytes recruited to the endothelium were CD14(+). Formation of adhesive platelet bridges on ECs was essential for monocyte recruitment as platelet removal or inhibition of adhesion to the ECs abolished monocyte recruitment. Monocytes were recruited from flow by platelet P-selectin and activated by EC-derived CC chemokine ligand 2 (CCL2), although the presentation of CCL2 to adherent monocytes was dependent upon platelet activation and release of CXC chemokine ligand 4 (CXCL4). In an intravital model of TGF-β1-driven vascular inflammation in mice, platelets were also necessary for efficient leucocyte recruitment to vessels of the microcirculation in the cremaster muscle. In this study, we have demonstrated that stromal cells found within the diseased artery wall may promote the preferential recruitment of monocytes and this is achieved by establishing a cascade of interactions between SMCs, ECs, platelets, and monocytes.

  12. SOX17 Regulates Conversion of Human Fibroblasts Into Endothelial Cells and Erythroblasts by Dedifferentiation Into CD34+ Progenitor Cells.

    Science.gov (United States)

    Zhang, Lianghui; Jambusaria, Ankit; Hong, Zhigang; Marsboom, Glenn; Toth, Peter T; Herbert, Brittney-Shea; Malik, Asrar B; Rehman, Jalees

    2017-06-20

    The mechanisms underlying the dedifferentiation and lineage conversion of adult human fibroblasts into functional endothelial cells have not yet been fully defined. Furthermore, it is not known whether fibroblast dedifferentiation recapitulates the generation of multipotent progenitors during embryonic development, which give rise to endothelial and hematopoietic cell lineages. Here we established the role of the developmental transcription factor SOX17 in regulating the bilineage conversion of fibroblasts by the generation of intermediate progenitors. CD34+ progenitors were generated after the dedifferentiation of human adult dermal fibroblasts by overexpression of pluripotency transcription factors. Sorted CD34+ cells were transdifferentiated into induced endothelial cells and induced erythroblasts using lineage-specific growth factors. The therapeutic potential of the generated cells was assessed in an experimental model of myocardial infarction. Induced endothelial cells expressed specific endothelial cell surface markers and also exhibited the capacity for cell proliferation and neovascularization. Induced erythroblasts expressed erythroid surface markers and formed erythroid colonies. Endothelial lineage conversion was dependent on the upregulation of the developmental transcription factor SOX17, whereas suppression of SOX17 instead directed the cells toward an erythroid fate. Implantation of these human bipotential CD34+ progenitors into nonobese diabetic/severe combined immunodeficiency (NOD-SCID) mice resulted in the formation of microvessels derived from human fibroblasts perfused with mouse and human erythrocytes. Endothelial cells generated from human fibroblasts also showed upregulation of telomerase. Cell implantation markedly improved vascularity and cardiac function after myocardial infarction without any evidence of teratoma formation. Dedifferentiation of fibroblasts to intermediate CD34+ progenitors gives rise to endothelial cells and

  13. Thalidomide inhibits inflammatory and angiogenic activation of human intestinal microvascular endothelial cells (HIMEC)

    Science.gov (United States)

    Stein, Daniel J.; Nelson, Victoria M.; Otterson, Mary F.; Shaker, Reza; Binion, David G.

    2010-01-01

    The glutamic acid derivative thalidomide is a transcriptional inhibitor of TNF-α but is also known to affect human blood vessels, which may underlie its teratogenicity. Thalidomide has been used in the treatment of refractory Crohn's disease (CD), but the therapeutic mechanism is not defined. We examined the effect of thalidomide on primary cultures of human intestinal microvascular endothelial cells (HIMEC), the relevant endothelial cell population in inflammatory bowel disease (IBD), to determine its effect on endothelial activation, leukocyte interaction, and VEGF-induced angiogenesis. HIMEC cultures were pretreated with thalidomide before activation with either TNF-α/LPS or VEGF. A low-shear-stress flow adhesion assay with either U-937 or whole blood was used to assess HIMEC activation following TNF-α/LPS, and a Wright's stain identified adherent leukocytes. Expression of cell adhesion molecules (E-selectin, intercellular adhesion molecule-1, vascular cell adhesion molecule-1) was assessed using radioimmunoassay. Effects of thalidomide on NF-κB activation, cyclooxygenase (COX)-2, and inducible nitric oxide synthase (iNOS) expression in TNF-α/LPS-activated HIMEC were determined by RT-PCR and Western blotting. Thalidomide blocked adhesion of both U-937 and whole blood leukocytes by 50% in HIMEC, inhibiting binding of all classes of leukocytes. Thalidomide also blocked NF-κB and cell adhesion molecule expression in HIMEC. In marked contrast, thalidomide did not affect either iNOS or COX-2 expression, two key molecules that play a role in the downregulation of HIMEC activation. VEGF-induced HIMEC transmigration, growth, proliferation, tube formation, and Akt phosphorylation were significantly inhibited by thalidomide. In summary, thalidomide exerted a potent effect on HIMEC growth and activation, suggesting that it may also function via an endothelial mechanism in the treatment of CD. PMID:19926820

  14. Glucosamine exposure reduces proteoglycan synthesis in primary human endothelial cells in vitro

    Directory of Open Access Journals (Sweden)

    Trine M. Reine

    2016-09-01

    Full Text Available Purpose: Glucosamine (GlcN supplements are promoted for medical reasons, for example, for patients with arthritis and other joint-related diseases. Oral intake of GlcN is followed by uptake in the intestine, transport in the circulation and thereafter delivery to chondrocytes. Here, it is postulated to have an effect on synthesis and turnover of extracellular matrix constituents expressed by these cells. Following uptake in the intestine, serum levels are transiently increased, and the endothelium is exposed to increased levels of GlcN. We investigated the possible effects of GlcN on synthesis of proteoglycans (PGs, an important matrix component, in primary human endothelial cells. Methods: Primary human endothelial cells were cultured in vitro in medium with 5 mM glucose and 0–10 mM GlcN. PGs were recovered and analysed by western blotting, or by SDS-PAGE, gel chromatography or ion-exchange chromatography of 35S-PGs after 35S-sulphate labelling of the cells. Results: The synthesis and secretion of 35S-PGs from cultured endothelial cells were reduced in a dose- and time-dependent manner after exposure to GlcN. PGs are substituted with sulphated glycosaminoglycan (GAG chains, vital for PG function. The reduction in 35S-PGs was not related to an effect on GAG chain length, number or sulphation, but rather to the total expression of PGs. Conclusion: Exposure of endothelial cells to GlcN leads to a general decrease in 35S-PG synthesis. These results suggest that exposure to high levels of GlcN can lead to decreased matrix synthesis, contrary to what has been claimed by supporters of such supplements.

  15. Aging-related changes in oxidative stress response of human endothelial cells.

    Science.gov (United States)

    Conti, Valeria; Corbi, Graziamaria; Simeon, Vittorio; Russomanno, Giusy; Manzo, Valentina; Ferrara, Nicola; Filippelli, Amelia

    2015-08-01

    Oxidative stress is strongly associated with aging and age-related diseases and plays a crucial role in endothelial dysfunction development. To better understand the molecular mechanisms of aging and stress response in humans, we examined changes to young and older human endothelial cells over time (72, 96 and 120 h), before and after H2O2-induced stress. We measured the expression of the deacetylase Sirtuin 1 (Sirt1) and its transcriptional target Forkhead box O3a (Foxo3a); TBARS, a well-known marker of overall oxidative stress, and catalase activity as index of antioxidation. Moreover, we quantified levels of cellular senescence by senescence-associated β galactosidase (SA-βgal) assay. Under oxidative stress induction older cells showed a progressive decrease of Sirt1 and Foxo3a expression, persistently high TBARS levels with high, but ineffective Cat activity to counteract such levels. In addition cellular senescence drastically increased in older cells compared with Young cells both in presence and in the absence of oxidative stress. By following the cell behavior during the time course, we can hypothesize that while in young cells an oxidative stress induction stimulated an adequate response through activation of molecular factor crucial to counteract oxidative stress, the older cells are not able to adequately adapt themselves to external stress stimuli. During their life, endothelial cells impair the ability to defend themselves from oxidative stress stimuli. This dysfunction involves the pathway of Sirt1 a critical regulator of oxidative stress response and cellular lifespan, underlining its crucial role in endothelial homeostasis control during aging and age-associated diseases.

  16. Efficient nanoparticle mediated sustained RNA interference in human primary endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Mukerjee, Anindita; Shankardas, Jwalitha; Ranjan, Amalendu P; Vishwanatha, Jamboor K, E-mail: Jamboor.vishwanatha@unthsc.edu [Department of Molecular Biology and Immunology and Institute for Cancer Research, Graduate School of Biomedical Sciences, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States)

    2011-11-04

    Endothelium forms an important target for drug and/or gene therapy since endothelial cells play critical roles in angiogenesis and vascular functions and are associated with various pathophysiological conditions. RNA mediated gene silencing presents a new therapeutic approach to overcome many such diseases, but the major challenge of such an approach is to ensure minimal toxicity and effective transfection efficiency of short hairpin RNA (shRNA) to primary endothelial cells. In the present study, we formulated shAnnexin A2 loaded poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles which produced intracellular small interfering RNA (siRNA) against Annexin A2 and brought about the downregulation of Annexin A2. The per cent encapsulation of the plasmid within the nanoparticle was found to be 57.65%. We compared our nanoparticle based transfections with Lipofectamine mediated transfection, and our studies show that nanoparticle based transfection efficiency is very high ({approx}97%) and is more sustained compared to conventional Lipofectamine mediated transfections in primary retinal microvascular endothelial cells and human cancer cell lines. Our findings also show that the shAnnexin A2 loaded PLGA nanoparticles had minimal toxicity with almost 95% of cells being viable 24 h post-transfection while Lipofectamine based transfections resulted in only 30% viable cells. Therefore, PLGA nanoparticle based transfection may be used for efficient siRNA transfection to human primary endothelial and cancer cells. This may serve as a potential adjuvant treatment option for diseases such as diabetic retinopathy, retinopathy of prematurity and age related macular degeneration besides various cancers.

  17. Efficient nanoparticle mediated sustained RNA interference in human primary endothelial cells

    Science.gov (United States)

    Mukerjee, Anindita; Shankardas, Jwalitha; Ranjan, Amalendu P.; Vishwanatha, Jamboor K.

    2011-11-01

    Endothelium forms an important target for drug and/or gene therapy since endothelial cells play critical roles in angiogenesis and vascular functions and are associated with various pathophysiological conditions. RNA mediated gene silencing presents a new therapeutic approach to overcome many such diseases, but the major challenge of such an approach is to ensure minimal toxicity and effective transfection efficiency of short hairpin RNA (shRNA) to primary endothelial cells. In the present study, we formulated shAnnexin A2 loaded poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles which produced intracellular small interfering RNA (siRNA) against Annexin A2 and brought about the downregulation of Annexin A2. The per cent encapsulation of the plasmid within the nanoparticle was found to be 57.65%. We compared our nanoparticle based transfections with Lipofectamine mediated transfection, and our studies show that nanoparticle based transfection efficiency is very high (~97%) and is more sustained compared to conventional Lipofectamine mediated transfections in primary retinal microvascular endothelial cells and human cancer cell lines. Our findings also show that the shAnnexin A2 loaded PLGA nanoparticles had minimal toxicity with almost 95% of cells being viable 24 h post-transfection while Lipofectamine based transfections resulted in only 30% viable cells. Therefore, PLGA nanoparticle based transfection may be used for efficient siRNA transfection to human primary endothelial and cancer cells. This may serve as a potential adjuvant treatment option for diseases such as diabetic retinopathy, retinopathy of prematurity and age related macular degeneration besides various cancers.

  18. Effects of Pycnogenol on endothelial function in patients with stable coronary artery disease: a double-blind, randomized, placebo-controlled, cross-over study.

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    Enseleit, Frank; Sudano, Isabella; Périat, Daniel; Winnik, Stephan; Wolfrum, Mathias; Flammer, Andreas J; Fröhlich, Georg M; Kaiser, Priska; Hirt, Astrid; Haile, Sarah R; Krasniqi, Nazmi; Matter, Christian M; Uhlenhut, Klaus; Högger, Petra; Neidhart, Michel; Lüscher, Thomas F; Ruschitzka, Frank; Noll, Georg

    2012-07-01

    Extracts from pine tree bark containing a variety of flavonoids have been used in traditional medicine. Pycnogenol is a proprietary bark extract of the French maritime pine tree (Pinus pinaster ssp. atlantica) that exerts antioxidative, anti-inflammatory, and anti-platelet effects. However, the effects of Pycnogenol on endothelial dysfunction, a precursor of atherosclerosis and cardiovascular events, remain still elusive. Twenty-three patients with coronary artery disease (CAD) completed this randomized, double-blind, placebo-controlled cross-over study. Patients received Pycnogenol (200 mg/day) for 8 weeks followed by placebo or vice versa on top of standard cardiovascular therapy. Between the two treatment periods, a 2-week washout period was scheduled. At baseline and after each treatment period, endothelial function, non-invasively assessed by flow-mediated dilatation (FMD) of the brachial artery using high-resolution ultrasound, biomarkers of oxidative stress and inflammation, platelet adhesion, and 24 h blood pressure monitoring were evaluated. In CAD patients, Pycnogenol treatment was associated with an improvement of FMD from 5.3 ± 2.6 to 7.0 ± 3.1 (P Pycnogenol treatment, while no change was observed in the placebo group (mean difference 0.06 pg/mL with an associated 95% CI (0.01, 0.11), P = 0.012]. Inflammation markers, platelet adhesion, and blood pressure did not change after treatment with Pycnogenol or placebo. This study provides the first evidence that the antioxidant Pycnogenol improves endothelial function in patients with CAD by reducing oxidative stress.

  19. Regulation of human heme oxygenase in endothelial cells by using sense and antisense retroviral constructs.

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    Quan, S; Yang, L; Abraham, N G; Kappas, A

    2001-10-09

    Our objective was to determine whether overexpression and underexpression of human heme oxygenase (HHO)-1 could be controlled on a long-term basis by introduction of the HO-1 gene in sense (S) and antisense (AS) orientation with an appropriate vector into endothelial cells. Retroviral vector (LXSN) containing viral long terminal repeat promoter-driven human HO-1 S (LSN-HHO-1) and LXSN vectors containing HHO-1 promoter (HOP)-controlled HHO-1 S and AS (LSN-HOP-HHO-1 and LSN-HOP-HHO-1-AS) sequences were constructed and used to transfect rat lung microvessel endothelial cells (RLMV cells) and human dermal microvessel endothelial cells (HMEC-1 cells). RLMV cells transduced with HHO-1 S expressed human HO-1 mRNA and HO-1 protein associated with elevation in total HO activity compared with nontransduced cells. Vector-mediated expression of HHO-1 S or AS under control of HOP resulted in effective production of HO-1 or blocked induction of endogenous human HO-1 in HMEC-1 cells, respectively. Overexpression of HO-1 AS was associated with a long-term decrease (45%) of endogenous HO-1 protein and an increase (167%) in unmetabolized exogenous heme in HMEC-1 cells. Carbon monoxide (CO) production in HO-1 S- or AS-transduced HMEC-1 cells after heme treatment was increased (159%) or decreased (50%), respectively, compared with nontransduced cells. HO-2 protein levels did not change. These findings demonstrate that HHO-1 S and AS retroviral constructs are functional in enhancing and reducing HO activity, respectively, and thus can be used to regulate cellular heme levels, the activity of heme-dependent enzymes, and the rate of heme catabolism to CO and bilirubin.

  20. Heterogeneity in conduit artery function in humans: impact of arterial size.

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    Thijssen, Dick H J; Dawson, Ellen A; Black, Mark A; Hopman, Maria T E; Cable, N Timothy; Green, Daniel J

    2008-11-01

    To determine whether conduit artery size affects functional responses, we compared the magnitude, time course, and eliciting shear rate stimulus for flow-mediated dilation (FMD) in healthy men (n = 20; 31 +/- 7 yr). Upper limb (brachial and radial) and lower limb (common and superficial femoral) FMD responses were simultaneously assessed, whereas popliteal responses were measured in the same subjects during a separate visit. Glyceryl trinitrate (GTN)-mediated responses were similarly examined. Edge detection and wall tracking of high-resolution B-mode arterial ultrasound images, combined with synchronized Doppler waveform envelope analysis, were used to calculate conduit artery diameter, blood flow, and shear rate continuously across the cardiac cycle. Baseline artery size correlated inversely with the FMD response (r = -0.57, P < 0.001). Within-artery comparisons revealed a significant inverse correlation between artery size and FMD% for the radial (r = -0.66, P = 0.001), brachial (r = -0.55, P = 0.01), and popliteal artery (r = -0.48, P = 0.03), but not for the superficial and common femoral artery. Normalization of FMD responses for differences in eliciting shear rate did not abolish the between-artery relationship for artery function and size (r = -0.48, P < 0.001), suggesting that differences between artery function responses were not entirely due to size-related differences in shear rate. This was reinforced by a significant between-artery correlation for GTN responses and baseline artery size (r = -0.74, P < 0.001). In summary, systematic differences exist in vascular function responses of conduit arteries that differ in size. This raises the possibility that differences in artery size within or between individuals may influence functional responses.

  1. Association between human cartilage glycoprotein 39 (YKL-40 and arterial stiffness in essential hypertension

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    Ma Wei-hong

    2012-05-01

    Full Text Available Abstract Background YKL-40, a proposed marker of inflammation and endothelial dysfunction, is associated with atherosclerosis and an increased cardiovascular mortality in the general population. However, the relationship between YKL-40 and arterial stiffness in hypertensive patients has not been adequately assessed. Methods The relationship between serum levels of YKL-40 and arterial stiffness was evaluated in 93 essential hypertensive subjects and 80 normal subjects. Essential hypertensive subjects were divided into two groups based upon urinary albumin-to-creatinine ratio (ACR: nonmicroalbuminuric group, (ACR n = 50 and microalbuminuric group (ACR ≥30 mg/g, n = 43. Large artery wall stiffness was assessed by measuring femoral arterial stiffness and carotid-femoral pulse wave velocity (cf-PWV. Serum levels of YKL-40 were determined by enzyme-linked immunosorbent assay (ELISA. Results The study demonstrated that YKL-40,cf-PWV and femoral arterial stiffness were increased significantly (PPr = 0.44, P = 0.000 and femoral arterial stiffness ( r = 0.42, P =0.001. Multiple linear stepwise regression analysis showed that YKL-40 was the impact factor of arterial stiffness ( P Conclusion YKL-40 levels are elevated in essential hypertension subjects with an independent association between increasing YKL-40 levels and increasing arterial stiffness. The study suggests it played a positive role of YKL-40 in the progressing vascular complications in patients with essential hypertension.

  2. Pathogen sensing pathways in human embryonic stem cell derived-endothelial cells: role of NOD1 receptors.

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    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.

  3. Grape-derived polyphenols improve aging-related endothelial dysfunction in rat mesenteric artery: role of oxidative stress and the angiotensin system.

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    Noureddine Idris Khodja

    Full Text Available Aging is characterized by the development of an endothelial dysfunction, which affects both the nitric oxide (NO- and the endothelium-derived hyperpolarizing factor (EDHF-mediated relaxations, associated with vascular oxidative stress and the activation of the angiotensin system. This study investigated whether red wine polyphenols (RWPs, antioxidants and potent stimulators of NO- and EDHF-mediated relaxations improve aging-related endothelial dysfunction, and, if so, examined the underlying mechanism. Mesenteric artery reactivity was determined in organ chambers, vascular oxidative stress by dihydroethidine and MitoSOX staining, and expression of target proteins by immunohistochemical staining. Control young rats (16 weeks received solvent (ethanol, 3% v/v, and middle-aged rats (46 weeks either solvent or RWPs (100 mg/kg/day in the drinking water. The acetylcholine-induced endothelium-dependent NO component was slightly reduced whereas the EDHF component was markedly blunted in rings of middle-aged rats compared to young rats. The endothelial dysfunction was associated with oxidative stress, an upregulation of angiotensin II and AT1 receptors and a down-regulation of SK(Ca, IK(Ca, and angiotensin converting enzyme. Intake of RWPs for either one or two weeks improved the NO and the EDHF components of the relaxation, and normalized oxidative stress, the expression of SK(Ca, IK(Ca and the components of the angiotensin system. The protective effect of the 2-week RWPs treatment persisted for one and two weeks following stopping intake of RWPs. Thus, intake of RWPs caused a persistent improvement of the endothelial function, particularly the EDHF component, in middle-aged rats and this effect seems to involve the normalization of the expression of SK(Ca, IK(Ca and the angiotensin system.

  4. Grape-derived polyphenols improve aging-related endothelial dysfunction in rat mesenteric artery: role of oxidative stress and the angiotensin system.

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    Idris Khodja, Noureddine; Chataigneau, Thierry; Auger, Cyril; Schini-Kerth, Valérie B

    2012-01-01

    Aging is characterized by the development of an endothelial dysfunction, which affects both the nitric oxide (NO)- and the endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxations, associated with vascular oxidative stress and the activation of the angiotensin system. This study investigated whether red wine polyphenols (RWPs), antioxidants and potent stimulators of NO- and EDHF-mediated relaxations improve aging-related endothelial dysfunction, and, if so, examined the underlying mechanism. Mesenteric artery reactivity was determined in organ chambers, vascular oxidative stress by dihydroethidine and MitoSOX staining, and expression of target proteins by immunohistochemical staining. Control young rats (16 weeks) received solvent (ethanol, 3% v/v), and middle-aged rats (46 weeks) either solvent or RWPs (100 mg/kg/day) in the drinking water. The acetylcholine-induced endothelium-dependent NO component was slightly reduced whereas the EDHF component was markedly blunted in rings of middle-aged rats compared to young rats. The endothelial dysfunction was associated with oxidative stress, an upregulation of angiotensin II and AT1 receptors and a down-regulation of SK(Ca), IK(Ca), and angiotensin converting enzyme. Intake of RWPs for either one or two weeks improved the NO and the EDHF components of the relaxation, and normalized oxidative stress, the expression of SK(Ca), IK(Ca) and the components of the angiotensin system. The protective effect of the 2-week RWPs treatment persisted for one and two weeks following stopping intake of RWPs. Thus, intake of RWPs caused a persistent improvement of the endothelial function, particularly the EDHF component, in middle-aged rats and this effect seems to involve the normalization of the expression of SK(Ca), IK(Ca) and the angiotensin system.

  5. Decellularized matrix from tumorigenic human mesenchymal stem cells promotes neovascularization with galectin-1 dependent endothelial interaction.

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    Jorge S Burns

    Full Text Available BACKGROUND: Acquisition of a blood supply is fundamental for extensive tumor growth. We recently described vascular heterogeneity in tumours derived from cell clones of a human mesenchymal stem cell (hMSC strain (hMSC-TERT20 immortalized by retroviral vector mediated human telomerase (hTERT gene expression. Histological analysis showed that cells of the most vascularized tumorigenic clone, -BD11 had a pericyte-like alpha smooth muscle actin (ASMA+ and CD146+ positive phenotype. Upon serum withdrawal in culture, -BD11 cells formed cord-like structures mimicking capillary morphogenesis. In contrast, cells of the poorly tumorigenic clone, -BC8 did not stain for ASMA, tumours were less vascularized and serum withdrawal in culture led to cell death. By exploring the heterogeneity in hMSC-TERT20 clones we aimed to understand molecular mechanisms by which mesenchymal stem cells may promote neovascularization. METHODOLOGY/PRINCIPAL FINDINGS: Quantitative qRT-PCR analysis revealed similar mRNA levels for genes encoding the angiogenic cytokines VEGF and Angiopoietin-1 in both clones. However, clone-BD11 produced a denser extracellular matrix that supported stable ex vivo capillary morphogenesis of human endothelial cells and promoted in vivo neovascularization. Proteomic characterization of the -BD11 decellularized matrix identified 50 extracellular angiogenic proteins, including galectin-1. siRNA knock down of galectin-1 expression abrogated the ex vivo interaction between decellularized -BD11 matrix and endothelial cells. More stable shRNA knock down of galectin-1 expression did not prevent -BD11 tumorigenesis, but greatly reduced endothelial migration into -BD11 cell xenografts. CONCLUSIONS: Decellularized hMSC matrix had significant angiogenic potential with at least 50 angiogenic cell surface and extracellular proteins, implicated in attracting endothelial cells, their adhesion and activation to form tubular structures. hMSC -BD11 surface galectin-1

  6. Loss of CD34 expression in aging human choriocapillaris endothelial cells.

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    Elliott H Sohn

    Full Text Available Structural and gene expression changes in the microvasculature of the human choroid occur during normal aging and age-related macular degeneration (AMD. In this study, we sought to determine the impact of aging and AMD on expression of the endothelial cell glycoprotein CD34. Sections from 58 human donor eyes were categorized as either young (under age 40, age-matched controls (> age 60 without AMD, or AMD affected (>age 60 with early AMD, geographic atrophy, or choroidal neovascularization. Dual labeling of sections with Ulex europaeus agglutinin-I lectin (UEA-I and CD34 antibodies was performed, and the percentage of capillaries labeled with UEA-I but negative for anti-CD34 was determined. In addition, published databases of mouse and human retinal pigment epithelium-choroid were evaluated and CD34 expression compared between young and old eyes. Immunohistochemical studies revealed that while CD34 and UEA-I were colocalized in young eyes, there was variable loss of CD34 immunoreactivity in older donor eyes. While differences between normal aging and AMD were not significant, the percentage of CD34 negative capillaries in old eyes, compared to young eyes, was highly significant (p = 3.8×10(-6. Endothelial cells in neovascular membranes were invariably CD34 positive. Published databases show either a significant decrease in Cd34 (mouse or a trend toward decreased CD34 (human in aging. These findings suggest that UEA-I and endogenous alkaline phosphatase activity are more consistent markers of aging endothelial cells in the choroid, and suggest a possible mechanism for the increased inflammatory milieu in the aging choroid.

  7. Human vascular endothelial cells transport foreign exosomes from cow's milk by endocytosis.

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    Kusuma, Rio Jati; Manca, Sonia; Friemel, Taylor; Sukreet, Sonal; Nguyen, Christopher; Zempleni, Janos

    2016-05-15

    Encapsulation of microRNAs in exosomes confers protection against degradation and a vehicle for shuttling of microRNAs between cells and tissues, and cellular uptake by endocytosis. Exosomes can be found in foods including milk. Humans absorb cow's milk exosomes and deliver the microRNA cargo to peripheral tissues, consistent with gene regulation by dietary nucleic acids across species boundaries. Here, we tested the hypothesis that human vascular endothelial cells transport milk exosomes by endocytosis, constituting a step crucial for the delivery of dietary exosomes and their cargo to peripheral tissues. We tested this hypothesis by using human umbilical vein endothelial cells and fluorophore-labeled exosomes isolated from cow's milk. Exosome uptake followed Michaelis-Menten kinetics (Vmax = 0.057 ± 0.004 ng exosome protein × 40,000 cells/h; Km = 17.97 ± 3.84 μg exosomal protein/200 μl media) and decreased by 80% when the incubation temperature was lowered from 37°C to 4°C. When exosome surface proteins were removed by treatment with proteinase K, or transport was measured in the presence of the carbohydrate competitor d-galactose or measured in the presence of excess unlabeled exosomes, transport rates decreased by 45% to 80% compared with controls. Treatment with an inhibitor of endocytosis, cytochalasin D, caused a 50% decrease in transport. When fluorophore-labeled exosomes were administered retro-orbitally, exosomes accumulated in liver, spleen, and lungs in mice. We conclude that human vascular endothelial cells transport bovine exosomes by endocytosis and propose that this is an important step in the delivery of dietary exosomes and their cargo to peripheral tissues. Copyright © 2016 the American Physiological Society.

  8. Effects of drospirenone on adhesion molecule expression and monocyte adherence in human endothelial cells.

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    Ito, Fumitake; Mori, Taisuke; Takaoka, Osamu; Tanaka, Yukiko; Koshiba, Akemi; Tatsumi, Hiroshi; Iwasa, Koichi; Kitawaki, Jo

    2016-06-01

    A major concern in hormone replacement therapy is the associated increased risk of cardiovascular diseases. A progestogen without the unfavorable effects on cardiovascular disease should be explored. Monocyte adhesion to endothelial cells is an important initial event in atherosclerosis. In this study, the effects of the alternative progestogen drospirenone (DRSP) on monocyte adhesion in human umbilical venous endothelial cells (HUVECs) were examined. In HUVECs treated with estrogens and progestogens, including DRSP and medroxyprogesterone acetate (MPA), the expression of the adhesion molecules E-selectin, P-selectin, ICAM-1, and VCAM-1 were examined by real-time PCR and using an enzyme-linked immunosorbent assay. A flow chamber system was used to investigate the effects of DRSP on U937 monocytoid cell adherence to HUVEC monolayers. All experimental data were compared using one-way Analysis of Variance. Upregulation of adhesion molecule mRNA or protein was not seen in HUVECs treated with DRSP alone or with 17β-estradiol+DRSP. DRSP alone, 17β-estradiol+DRSP or ethinylestradiol+DRSP did not increase the number of adherent monocytoid cells to HUVECs in the flow chamber system. However, MPA significantly enhanced the monocytoid cell adherence (Padhesion molecules or monocytoid cell adherence to endothelial cells, indicating that DRSP could reduce the risk of atherogenesis caused by MPA. These results suggest that DRSP may be an alternative to MPA in hormone replacement therapy. Copyright © 2016. Published by Elsevier Ireland Ltd.

  9. Commercial single-walled carbon nanotubes effects in fibrinolysis of human umbilical vein endothelial cells.

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    Rodríguez-Yáñez, Yury; Bahena-Uribe, Daniel; Chávez-Munguía, Bibiana; López-Marure, Rebeca; González-Monroy, Stuart; Cisneros, Bulmaro; Albores, Arnulfo

    2015-08-01

    Recent studies have demonstrated that carbon nanotubes (CNTs) induce platelet aggregation, endothelial dysfunction and vascular thrombosis. However, there is little information on the effects of CNTs on fibrinolysis. We investigated the role of pristine-commercial single-walled carbon nanotubes (SWCNTs) with fibrinolysis and their contribution to the induction of pro-thrombotic processes in human vein endothelial cells (HUVEC). SWCNTs alone produced concentration-dependent oxidation, as measured by a dithiothreitol oxidation assay. Internalized SWCNTs were located in HUVEC treated with 25 μg/ml using transmission electron microscopy, whereas treatment with 50 μg/ml compromised cell viability, and oxidative stress increased significantly at 5 μg/ml. The study showed that in HUVEC treated with 25 μg SWCNT/ml, fibrinolysis-related gene expression and protein levels had increased by 3-12 h after treatment (serpine-1: 13-fold; PLAT: 11-fold and PLAU: 2-fold), but only the PAI-1 protein was increased (1.5-fold), whereas tissue and urokinase plasminogen activator proteins (tPA and uPA, respectively) tended to decrease. In summary, pristine SWCNTs treatment resulted in evident HUVEC damage caused by cell fiber contact, internalization, and oxidative stress due to contaminant metals. The generation of endothelial dysfunction, as shown by the altered expression of genes and proteins involved in fibrinolysis, suggest that SWCNTs display pro-thrombotic effects. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  10. Implanted neonatal human dermal fibroblasts influence the recruitment of endothelial cells in mice.

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    Guerreiro, Susana G; Brochhausen, Christoph; Negrão, Rita; Barbosa, Mário A; Unger, Ronald E; Kirkpatrick, C James; Soares, Raquel; Granja, Pedro L

    2012-01-01

    The vascularization of new tissue within a reasonable time is a crucial prerequisite for the success of different cell- and material-based strategies. Considering that angiogenesis is a multi-step process involving humoral and cellular regulatory components, only in vivo assays provide the adequate information about vessel formation and the recruitment of endothelial cells. The present study aimed to investigate if neonatal human dermal fibroblasts could influence in vivo neovascularization. Results obtained showed that fibroblasts were able to recruit endothelial cells to vascularize the implanted matrix, which was further colonized by murine functional blood vessels after one week. The vessels exhibited higher levels of hemoglobin, compared with the control matrix, implanted without fibroblasts, in which no vessel formation could be observed. No significant differences were detected in systemic inflammation. The presence of vessels originated from the host vasculature suggested that host vascular response was involved, which constitutes a fundamental aspect in the process of neovascularization. Fibroblasts implanted within matrigel increased the presence of endothelial cells with positive staining for CD31 and for CD34 and the production of collagen influencing the angiogenic process and promoting the formation of microvessels. New strategies in tissue engineering could be delineated with improved angiogenesis using neonatal fibroblasts.

  11. Reactive oxygen species modulate the barrier function of the human glomerular endothelial glycocalyx.

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    Anurag Singh

    Full Text Available Reactive oxygen species (ROS play a key role in the pathogenesis of proteinuria in glomerular diseases like diabetic nephropathy. Glomerular endothelial cell (GEnC glycocalyx covers the luminal aspect of the glomerular capillary wall and makes an important contribution to the glomerular barrier. ROS are known to depolymerise glycosaminoglycan (GAG chains of proteoglycans, which are crucial for the barrier function of GEnC glycocalyx. The aim of this study is to investigate the direct effects of ROS on the structure and function of GEnC glycocalyx using conditionally immortalised human GEnC. ROS were generated by exogenous hydrogen peroxide. Biosynthesis and cleavage of GAG chains was analyzed by radiolabelling (S(35 and (3H-glucosamine. GAG chains were quantified on GEnC surface and in the cell supernatant using liquid chromatography and immunofluorescence techniques. Barrier properties were estimated by measuring trans-endothelial passage of albumin. ROS caused a significant loss of WGA lectin and heparan sulphate staining from the surface of GEnC. This lead to an increase in trans-endothelial albumin passage. The latter could be inhibited by catalase and superoxide dismutase. The effect of ROS on GEnC was not mediated via the GAG biosynthetic pathway. Quantification of radiolabelled GAG fractions in the supernatant confirmed that ROS directly caused shedding of HS GAG. This finding is clinically relevant and suggests a mechanism by which ROS may cause proteinuria in clinical conditions associated with high oxidative stress.

  12. Mechanism of the toxicity induced by natural humic acid on human vascular endothelial cells.

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    Kihara, Yusuke; Yustiawati; Tanaka, Masato; Gumiri, Sulmin; Ardianor; Hosokawa, Toshiyuki; Tanaka, Shunitz; Saito, Takeshi; Kurasaki, Masaaki

    2014-08-01

    Humic acid (HA), a group of high-molecular weight organic compounds characterized by an ability to bind heavy metals, is normally found in natural water. Although the impairment of vascular endothelial cells in the presence of humic substances has been reported to be involved in some diseases, the mechanisms responsible for this involvement remain unclear. In this study, we examined the cytotoxicity of HA obtained from peatland in Central Kalimantan, Indonesia, to human vascular endothelial cells, as well as the mechanisms behind these effects. It was found that 50 mg/L HA showed cytotoxicity, which we considered to be mediated by apoptosis through the mitochondrial pathway because of an increase in the expression of caspases 6 and 9 in response to HA administration. In addition, this cytotoxicity was enhanced when cells in this experimental system were exposed to oxidative stress, while it was decreased by the addition of vitamin C. Thus, we conclude that the apoptosis induced by HA depends upon oxidative stress. Furthermore, an iron chelator, DFO, showed a tendency to decrease HA-induced cytotoxicity, suggesting that iron may potentially mediate HA-induced oxidative stress. In conclusion, long-term consumption of HA-rich water obtained from our study area may cause damage to endothelial cells and subsequent chronic health problems. Copyright © 2012 Wiley Periodicals, Inc., a Wiley company.

  13. A Fermented Whole Grain Prevents Lipopolysaccharides-Induced Dysfunction in Human Endothelial Progenitor Cells

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    Laura Giusti

    2017-01-01

    Full Text Available Endogenous and exogenous signals derived by the gut microbiota such as lipopolysaccharides (LPS orchestrate inflammatory responses contributing to development of the endothelial dysfunction associated with atherosclerosis in obesity, metabolic syndrome, and diabetes. Endothelial progenitor cells (EPCs, bone marrow derived stem cells, promote recovery of damaged endothelium playing a pivotal role in cardiovascular repair. Since healthy nutrition improves EPCs functions, we evaluated the effect of a fermented grain, Lisosan G (LG, on early EPCs exposed to LPS. The potential protective effect of LG against LPS-induced alterations was evaluated as cell viability, adhesiveness, ROS production, gene expression, and NF-kB signaling pathway activation. Our results showed that LPS treatment did not affect EPCs viability and adhesiveness but induced endothelial alterations via activation of NF-kB signaling. LG protects EPCs from inflammation as well as from LPS-induced oxidative and endoplasmic reticulum (ER stress reducing ROS levels, downregulating proinflammatory and proapoptotic factors, and strengthening antioxidant defense. Moreover, LG pretreatment prevented NF-kB translocation from the cytoplasm into the nucleus caused by LPS exposure. In human EPCs, LPS increases ROS and upregulates proinflammatory tone, proapoptotic factors, and antioxidants. LG protects EPCs exposed to LPS reducing ROS, downregulating proinflammatory and proapoptotic factors, and strengthening antioxidant defenses possibly by inhibiting NF-κB nuclear translocation.

  14. The influence of statins on the free intracellular calcium concentration in human umbilical vein endothelial cells

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    Figulla Hans R

    2004-05-01

    Full Text Available Abstract Background Statins are cholesterol-lowering drugs that are widely used to reduce the risk of cardiac infarction. Their beneficial clinical effects, however, are not restricted to their influence on cholesterol production. As several studies have shown that they have a potency of relaxing blood vessels. Methods We measured the effects of statins on the intracellular free calcium concentration ([Ca2+]i in human umbilical vein endothelial cells (HUVEC after acute application and 24-h-preincubation of statins. Results Incubation of the cells for 24 h with cerivastatin or fluvastatin significantly increased the resting [Ca2+]i. For cerivastatin this effect manifested at a concentration of 1 μM. Increase of resting [Ca2+]i in the presence of cerivastatin also occurred when the nitric oxide synthase was inhibited. Transient Ca2+ release induced by histamine was not affected. Conclusions The increase of resting [Ca2+]i after incubation with cerivastatin or fluvastatin may provide an explanation for the direct effects of statins on the endothelial-dependent vasodilatation and restoration of endothelial activity in vivo.

  15. Impact of introducer sheath coating on endothelial function in humans after transradial coronary procedures.

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    Dawson, Ellen A; Rathore, Sudhir; Cable, N Timothy; Wright, D Jay; Morris, John L; Green, Daniel J

    2010-04-01

    The aim of this study was to compare the impact of transradial catheterization with hydrophilic-coated catheter sheaths versus uncoated sheaths on NO-mediated endothelial-dependent and -independent vasodilator function. Thirty-five subjects undergoing transradial catheterization were recruited and assessed before and the day after catheterization. A subgroup was also assessed 3 to 4 months after catheterization. Subjects received hydrophilic-coated sheaths (n=15) or uncoated sheaths (n=20). Radial artery flow-mediated dilatation and endothelium- and NO-dependent arterial dilatation were assessed within the region of sheath placement. Glyceryl trinitrate endothelium-independent NO-mediated function was also assessed. The noncatheterized arm provided an internal control. Flow-mediated dilatation in the catheterized arm decreased from 10.3+/-3.8% to 5.3+/-3.3% and 8.1+/-2.4% to 5.2+/-3.7% in the coated and uncoated groups, respectively (P<0.01). These values returned toward baseline levels approximately 3 months later (coated, 6.4+/-1.4%; uncoated, 9.4+/-4.1%; P<0.05) versus postprocedure. Glyceryl trinitrate decreased from 14.8+/-7.2% to 9.5+/-4.1% (P<0.05) in the coated group and from 12.2+/-4.6% to 7.5+/-4.2% (P<0.01) in the uncoated group. Values returned to baseline at approximately 3 months (coated, 16.6+/-5.6%; uncoated, 12.1+/-3.9%; P<0.05). There was no difference in the magnitude of decrease in flow-mediated dilatation or glyceryl trinitrate between coated and uncoated groups. No changes in function occurred in the noncatheterized arm. Placement of a catheter sheath inside the radial artery disrupts vasodilator function, which recovers after 3 months. No differences were evident between hydrophilic-coated and uncoated sheaths.

  16. Knockout of CD8 delays reendothelialization and accelerates neointima formation in injured arteries of mouse via TNF-α inhibiting the endothelial cells migration.

    Directory of Open Access Journals (Sweden)

    Jun-Meng Zhang

    Full Text Available OBJECTIVE: Delayed or impaired reendothelialization is a major cause of stent thrombosis in the interventional treatment of coronary heart disease. T cells are involved in neointima formation of injured arteries. However, the regulated mechanism of reendothelialization and the role of CD8 T cell in reendothelialization are unclear. METHODS AND RESULTS: Immunofluorescence staining showed that CD8 positive cells were increased in wire injured femoral artery of mice. On day 21 after injury, elastin staining showed that knockout of CD8 (CD8(-/- significantly increased intimal thickness and a ratio of intima to media by 1.8 folds and 1.9 folds respectively in injured arteries. Evans blue staining showed that knockout of CD8 delayed the reendothelialization area on day 7 after injury (18.8±0.5% versus 42.1±5.6%, p<0.05. In vitro, a migration assay revealed that CD8(-/- T cells co-cultured with WT macrophages significantly inhibited the migration of the endothelial cells (ECs; compared to CD4(+ T cells, and CD8(+ T cells could promote the ECs migration. Furthermore, real-time PCR analysis showed that knockout of CD8 increased the level of tumor necrosis factor α (TNF-α in injured arteries and cytometric bead cytokine array showed that TNF-α was elevated in cultured CD8(-/- T cells. Finally, a wound-healing assay showed that recombinant TNF-α significantly inhibited the migration of ECs. CONCLUSION: Our study suggested that CD8(+ T cells could promote the reendothelialization and inhibit the neointima formation after the artery wire injury, and this effect is at least partly dependent on decreasing TNF-α production promoting ECs migration.

  17. Human neutrophil elastase induces endothelial cell apoptosis by activating the PERK-CHOP branch of the unfolded protein response.

    Science.gov (United States)

    Grechowa, Irina; Horke, Sven; Wallrath, Anja; Vahl, Christian-Friedrich; Dorweiler, Bernhard

    2017-09-01

    Human neutrophil elastase impacts on atherosclerotic plaque stability by inducing apoptosis in endothelial cells. Our aim was to investigate the proapoptotic mechanism of elastase on endothelial cells and to evaluate the presence of elastase in human plaque material. Human endothelial cells were treated with purified human neutrophil elastase. Apoptosis was assayed by capsase-3/7 activation, TUNEL, and sub-G 1 assay. Activation of unfolded protein response (UPR) effector molecules binding Ig protein, soluble X-binding protein-1, protein kinase RNA-like ER kinase (PERK), and C/EBP-homologous protein (CHOP) was analyzed by RT-PCR, immunocytochemistry, and Western blot. Genetic silencing of CHOP was achieved by small interfering RNA. Elastase induces autophagic-apoptotic forms of endothelial cell death in a time- and dose-dependent manner, in conjunction with a significant increase in phosphorylation/expression of the canonical UPR-activation markers PERK and CHOP. By using CHOP knockdown, we identified CHOP as a key mediator of elastase-induced endothelial cell death. Immunohistochemical analysis of human rupture-prone plaque specimens confirmed the presence of elastase and colocalization with apoptosis. We have demonstrated for the first time that the PERK-CHOP branch of the UPR is causally involved in elastase-induced apoptosis of endothelial cells. Ex vivo analysis of human rupture-prone plaques confirmed the presence of elastase and its colocalization with markers of apoptosis. This novel role of elastase underlines the potential of combined targeting of elastase and endoplasmic reticulum stress in the prevention of plaque progression and cardiovascular events.-Grechowa, I., Horke, S., Wallrath, A., Vahl, C.-F., Dorweiler, B. Human neutrophil elastase induces endothelial cell apoptosis by activating the PERK-CHOP branch of the unfolded protein response. © FASEB.

  18. Efficient gene disruption in cultured primary human endothelial cells by CRISPR/Cas9.

    Science.gov (United States)

    Abrahimi, Parwiz; Chang, William G; Kluger, Martin S; Qyang, Yibing; Tellides, George; Saltzman, W Mark; Pober, Jordan S

    2015-07-03

    The participation of endothelial cells (EC) in many physiological and pathological processes is widely modeled using human EC cultures, but genetic manipulation of these untransformed cells has been technically challenging. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 nuclease (Cas9) technology offers a promising new approach. However, mutagenized cultured cells require cloning to yield homogeneous populations, and the limited replicative lifespan of well-differentiated human EC presents a barrier for doing so. To create a simple but highly efficient method using CRISPR/Cas9 to generate biallelic gene disruption in untransformed human EC. To demonstrate proof-of-principle, we used CRISPR/Cas9 to disrupt the gene for the class II transactivator. We used endothelial colony forming cell-derived EC and lentiviral vectors to deliver CRISPR/Cas9 elements to ablate EC expression of class II major histocompatibility complex molecules and with it, the capacity to activate allogeneic CD4(+) T cells. We show the observed loss-of-function arises from biallelic gene disruption in class II transactivator that leaves other essential properties of the cells intact, including self-assembly into blood vessels in vivo, and that the altered phenotype can be rescued by reintroduction of class II transactivator expression. CRISPR/Cas9-modified human EC provides a powerful platform for vascular research and for regenerative medicine/tissue engineering. © 2015 American Heart Association, Inc.

  19. Suitability of endogenous reference genes for gene expression studies with human intraocular endothelial cells

    Directory of Open Access Journals (Sweden)

    Wei Ruoxin

    2013-02-01

    Full Text Available Abstract Background The use of quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR has become widely applied as a method to measure transcript abundance. In order to be reflective of biological processes during health and disease this method is dependent on normalisation of data against stable endogenous controls. However, these genes can vary in their stability in different cell types. The importance of reference gene validation for a particular cell type is now well recognised and is an important step in any gene expression study. Results Cultured primary human choroidal and retinal endothelial cells were treated with the immunostimulant polyinosinic: polycytidylic acid or untreated. qRT-PCR was used to quantify the expression levels of 10 commonly used endogenous control genes, TBP, HPRT1, GAPDH, GUSB, PPIA, RPLP0, B2M, 18S rRNA, PGK1 and ACTB. Three different mathematical algorithms, GeNorm, NormFinder, and BestKeeper were used to analyse gene stability to give the most representative validation. In choroidal endothelial cells the most stable genes were ranked as HPRT1 and GUSB by GeNorm and NormFinder and HPRT1 and PPIA by BestKeeper. In retinal endothelial cells the most stable genes ranked were TBP and PGK1 by GeNorm and NormFinder and HPRT1 by BestKeeper. The least stable gene for both cell types was 18S with all 3 algorithms. Conclusions We have identified the most stable endogenous control genes in intraocular endothelial cells. It is suggested future qRT-PCR studies using these cells would benefit from adopting the genes identified in this study as the most appropriate endogenous control genes.

  20. Oxidation of 2-cys peroxiredoxins in human endothelial cells by hydrogen peroxide, hypochlorous acid, and chloramines.

    Science.gov (United States)

    Stacey, Melissa M; Vissers, Margreet C; Winterbourn, Christine C

    2012-08-01

    Reactive oxygen species released from neutrophils during vascular inflammation could contribute to endothelial dysfunction seen in diseases such as atherosclerosis. Activated neutrophils generate hydrogen peroxide (H(2)O(2)) and hypochlorous acid (HOCl), as well as chloramines that are formed when HOCl reacts with amino compounds. These oxidants preferentially target thiol groups and thiol-containing proteins. The peroxiredoxins (Prxs) are thiol proteins that have high reactivity with H(2)O(2) and may also be sensitive to HOCl and chloramines. We have investigated human umbilical vein endothelial cells and shown that their cytoplasmic (Prx1 and Prx2) and mitochondrial (Prx3) Prxs are oxidized when they are exposed to H(2)O(2), HOCl, or cell-permeable chloramines. H(2)O(2) converted the Prxs to hyperoxidized, inactive forms, with little accumulation of disulfide-linked dimers. The oxidized Prxs were reduced over hours, presumably due to the action of endothelial sulfiredoxin. In contrast to the hyperoxidation seen with H(2)O(2), HOCl and the chloramine derivatives of glycine and ammonia converted the Prxs to disulfide-linked dimers and dimerization was reversed within 10-30 min of oxidant removal. HOCl treatment caused thioredoxin reductase (TrxR) inhibition with no reversal of dimerization. The cytotoxicity of ammonia chloramine was increased when cells were pretreated with H(2)O(2) to hyperoxidize the Prxs, or when the chloramine was added in the presence of the TrxR inhibitor, auranofin. We describe the novel observation that exposure of nucleated cells to inflammatory oxidants results in the accumulation of Prxs in the dimeric form. Endothelial cell Prxs are sensitive targets for neutrophil-derived oxidants and may protect against their damaging effects.

  1. Impact of calcium signaling during infection of Neisseria meningitidis to human brain microvascular endothelial cells.

    Science.gov (United States)

    Asmat, Tauseef M; Tenenbaum, Tobias; Jonsson, Ann-Beth; Schwerk, Christian; Schroten, Horst

    2014-01-01

    The pili and outer membrane proteins of Neisseria meningitidis (meningococci) facilitate bacterial adhesion and invasion into host cells. In this context expression of meningococcal PilC1 protein has been reported to play a crucial role. Intracellular calcium mobilization has been implicated as an important signaling event during internalization of several bacterial pathogens. Here we employed time lapse calcium-imaging and demonstrated that PilC1 of meningococci triggered a significant increase in cytoplasmic calcium in human brain microvascular endothelial cells, whereas PilC1-deficient meningococci could not initiate this signaling process. The increase in cytosolic calcium in response to PilC1-expressing meningococci was due to efflux of calcium from host intracellular stores as demonstrated by using 2-APB, which inhibits the release of calcium from the endoplasmic reticulum. Moreover, pre-treatment of host cells with U73122 (phospholipase C inhibitor) abolished the cytosolic calcium increase caused by PilC1-expressing meningococci demonstrating that active phospholipase C (PLC) is required to induce calcium transients in host cells. Furthermore, the role of cytosolic calcium on meningococcal adherence and internalization was documented by gentamicin protection assay and double immunofluorescence (DIF) staining. Results indicated that chelation of intracellular calcium by using BAPTA-AM significantly impaired PilC1-mediated meningococcal adherence to and invasion into host endothelial cells. However, buffering of extracellular calcium by BAPTA or EGTA demonstrated no significant effect on meningococcal adherence to and invasion into host cells. Taken together, these results indicate that meningococci induce calcium release from intracellular stores of host endothelial cells via PilC1 and cytoplasmic calcium concentrations play a critical role during PilC1 mediated meningococcal adherence to and subsequent invasion into host endothelial cells.

  2. Impact of calcium signaling during infection of Neisseria meningitidis to human brain microvascular endothelial cells.

    Directory of Open Access Journals (Sweden)

    Tauseef M Asmat

    Full Text Available The pili and outer membrane proteins of Neisseria meningitidis (meningococci facilitate bacterial adhesion and invasion into host cells. In this context expression of meningococcal PilC1 protein has been reported to play a crucial role. Intracellular calcium mobilization has been implicated as an important signaling event during internalization of several bacterial pathogens. Here we employed time lapse calcium-imaging and demonstrated that PilC1 of meningococci triggered a significant increase in cytoplasmic calcium in human brain microvascular endothelial cells, whereas PilC1-deficient meningococci could not initiate this signaling process. The increase in cytosolic calcium in response to PilC1-expressing meningococci was due to efflux of calcium from host intracellular stores as demonstrated by using 2-APB, which inhibits the release of calcium from the endoplasmic reticulum. Moreover, pre-treatment of host cells with U73122 (phospholipase C inhibitor abolished the cytosolic calcium increase caused by PilC1-expressing meningococci demonstrating that active phospholipase C (PLC is required to induce calcium transients in host cells. Furthermore, the role of cytosolic calcium on meningococcal adherence and internalization was documented by gentamicin protection assay and double immunofluorescence (DIF staining. Results indicated that chelation of intracellular calcium by using BAPTA-AM significantly impaired PilC1-mediated meningococcal adherence to and invasion into host endothelial cells. However, buffering of extracellular calcium by BAPTA or EGTA demonstrated no significant effect on meningococcal adherence to and invasion into host cells. Taken together, these results indicate that meningococci induce calcium release from intracellular stores of host endothelial cells via PilC1 and cytoplasmic calcium concentrations play a critical role during PilC1 mediated meningococcal adherence to and subsequent invasion into host endothelial cells.

  3. Particulate matter disrupts human lung endothelial barrier integrity via ROS- and p38 MAPK-dependent pathways.

    Science.gov (United States)

    Wang, Ting; Chiang, Eddie T; Moreno-Vinasco, Liliana; Lang, Gabriel D; Pendyala, Srikanth; Samet, Jonathan M; Geyh, Alison S; Breysse, Patrick N; Chillrud, Steven N; Natarajan, Viswanathan; Garcia, Joe G N

    2010-04-01

    Epidemiologic studies have linked exposure to airborne pollutant particulate matter (PM) with increased cardiopulmonary mortality and morbidity. The mechanisms of PM-mediated lung pathophysiology, however, remain unknown. We tested the hypothesis that PM, via enhanced oxidative stress, disrupts lung endothelial cell (EC) barrier integrity, thereby enhancing organ dysfunction. Using PM collected from Ft. McHenry Tunnel (Baltimore, MD), we assessed PM-mediated changes in transendothelial electrical resistance (TER) (a highly sensitive measure of barrier function), reactive oxygen species (ROS) generation, and p38 mitogen-activated protein kinase (MAPK) activation in human pulmonary artery EC. PM induced significant dose (10-100 microg/ml)- and time (0-10 h)-dependent EC barrier disruption reflected by reduced TER values. Exposure of human lung EC to PM resulted in significant ROS generation, which was directly involved in PM-mediated EC barrier dysfunction, as N-acetyl-cysteine (NAC, 5 mM) pretreatment abolished both ROS production and barrier disruption induced by PM. Furthermore, PM induced p38 MAPK activation and HSP27 phosphorylation, events that were both attenuated by NAC. In addition, PM-induced EC barrier disruption was partially prevented by the p38 MAP kinase inhibitor SB203580 (10 microM) as well as by reduced expression of either p38 MAPK beta or HSP27 (siRNA). These results demonstrate that PM induces ROS generation in human lung endothelium, resulting in oxidative stress-mediated EC barrier disruption via p38 MAPK- and HSP27-dependent pathways. These findings support a novel mechanism for PM-induced lung dysfunction and adverse cardiopulmonary outcomes.

  4. Particulate Matter Disrupts Human Lung Endothelial Barrier Integrity via ROS- and p38 MAPK–Dependent Pathways

    Science.gov (United States)

    Wang, Ting; Chiang, Eddie T.; Moreno-Vinasco, Liliana; Lang, Gabriel D.; Pendyala, Srikanth; Samet, Jonathan M.; Geyh, Alison S.; Breysse, Patrick N.; Chillrud, Steven N.; Natarajan, Viswanathan; Garcia, Joe G. N.

    2010-01-01

    Epidemiologic studies have linked exposure to airborne pollutant particulate matter (PM) with increased cardiopulmonary mortality and morbidity. The mechanisms of PM-mediated lung pathophysiology, however, remain unknown. We tested the hypothesis that PM, via enhanced oxidative stress, disrupts lung endothelial cell (EC) barrier integrity, thereby enhancing organ dysfunction. Using PM collected from Ft. McHenry Tunnel (Baltimore, MD), we assessed PM-mediated changes in transendothelial electrical resistance (TER) (a highly sensitive measure of barrier function), reactive oxygen species (ROS) generation, and p38 mitogen-activated protein kinase (MAPK) activation in human pulmonary artery EC. PM induced significant dose (10–100 μg/ml)- and time (0–10 h)-dependent EC barrier disruption reflected by reduced TER values. Exposure of human lung EC to PM resulted in significant ROS generation, which was directly involved in PM-mediated EC barrier dysfunction, as N-acetyl-cysteine (NAC, 5 mM) pretreatment abolished both ROS production and barrier disruption induced by PM. Furthermore, PM induced p38 MAPK activation and HSP27 phosphorylation, events that were both attenuated by NAC. In addition, PM-induced EC barrier disruption was partially prevented by the p38 MAP kinase inhibitor SB203580 (10 μM) as well as by reduced expression of either p38 MAPK β or HSP27 (siRNA). These results demonstrate that PM induces ROS generation in human lung endothelium, resulting in oxidative stress–mediated EC barrier disruption via p38 MAPK- and HSP27-dependent pathways. These findings support a novel mechanism for PM-induced lung dysfunction and adverse cardiopulmonary outcomes. PMID:19520919

  5. Human Galectin-3 Promotes Trypanosoma cruzi Adhesion to Human Coronary Artery Smooth Muscle Cells

    OpenAIRE

    Kleshchenko, Yuliya Y.; Moody, Tapria N.; Furtak, Vyacheslav A.; Ochieng, Josiah; Lima, Maria F.; Villalta, Fernando

    2004-01-01

    Human galectin-3 binds to the surface of Trypanosoma cruzi trypomastigotes and human coronary artery smooth muscle (CASM) cells. CASM cells express galectin-3 on their surface and secrete it. Exogenous galectin-3 increased the binding of T. cruzi to CASM cells. Trypanosome binding to CASM cells was enhanced when either T. cruzi or CASM cells were preincubated with galectin-3. Cells stably transfected with galectin-3 antisense show a dramatic decrease in galectin-3 expression and very little T...

  6. Human β-Defensin 3 Reduces TNF-α-Induced Inflammation and Monocyte Adhesion in Human Umbilical Vein Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Tianying Bian

    2017-01-01

    Full Text Available The aim of this study was to investigate the role of human β-defensin 3 (hBD3 in the initiation stage of atherosclerosis with human umbilical vein endothelial cells (HUVECs triggered by tumor necrosis factor- (TNF- α. The effects of hBD3 on TNF-α-induced endothelial injury and inflammatory response were evaluated. Our data revealed that first, hBD3 reduced the production of interleukin-6 (IL-6, IL-8, monocyte chemoattractant protein-1 (MCP-1, and macrophage migration inhibitory factor (MIF in HUVECs in a dose-dependent manner. In addition, hBD3 significantly prevented intracellular reactive oxygen species (ROS production by HUVECs. Second, western blot analysis demonstrated that hBD3 dose-dependently suppressed the protein levels of intracellular adhesion molecule-1 (ICAM-1 and vascular cell adhesion molecule-1 (VCAM-1 in TNF-α-induced HUVECs. As a result, hBD3 inhibited monocyte adhesion to TNF-α-treated endothelial cells. Additionally, hBD3 suppressed TNF-α-induced F-actin reorganization in HUVECs. Third, hBD3 markedly inhibited NF-κB activation by decreasing the phosphorylation of IKK-α/β, IκB, and p65 subunit within 30 min. Moreover, the phosphorylation of p38 and c-Jun N-terminal protein kinase (JNK in the mitogen-activated protein kinase (MAPK pathway were also inhibited by hBD3 in HUVECs. In conclusion, hBD3 exerts anti-inflammatory and antioxidative effects in endothelial cells in response to TNF-α by inhibiting NF-κB and MAPK signaling.

  7. Generation of novel monoclonal antibodies for the enrichment and characterization of human corneal endothelial cells (hCENC) necessary for the treatment of corneal endothelial blindness.

    Science.gov (United States)

    Ding, Vanessa; Chin, Angela; Peh, Gary; Mehta, Jodhbir S; Choo, Andre

    2014-01-01

    Corneal transplantation is the primary treatment option to restore vision for patients with corneal endothelial blindness. Although the success rate of treatment is high, limited availability of transplant grade corneas is a major obstacle. Tissue-engineered corneal endothelial grafts constructed using cultivated human corneal endothelial cells (hCENC) isolated from cadaveric corneas may serve as a potential graft source. Currently, tools for the characterization of cultured hCENC and enrichment of hCENC from potential contaminating cells such as stromal fibroblasts are lacking. In this study, we describe the generation and characterization of novel cell surface monoclonal antibodies (mAbs) specific for hCENC. These mAbs could be used for enrichment and characterization of hCENC. Out of a total of 389 hybridomas, TAG-1A3 and TAG-2A12 were found to be specific to the corneal endothelial monolayer by immunostaining of frozen tissue sections. Both mAbs were able to clearly identify hCENC with good 'cobblestone-like' morphology from multiple donors. The antigen targets for TAG-1A3 and TAG-2A12 were found to be CD166/ALCAM and Peroxiredoxin-6 (Prdx-6), respectively, both of which have not been previously described as markers of hCENC. Additionally, unlike other Prdx-6 mAbs, TAG-2A12 was found to specifically bind cell surface Prdx-6, which was only expressed on hCENC and not on other cell types screened such as human corneal stromal fibroblasts (hCSF) and human pluripotent stem cells (hPSC). From our studies, we conclude that TAG-1A3 and TAG-2A12 are promising tools to quantitatively assess hCENC quality. It is also noteworthy that the binding specificity of TAG-2A12 could be used for the enrichment of hCENC from cell mixtures of hCSF and hPSC.

  8. Effects of cognate, non-cognate and synthetic CXCR4 and ACKR3 ligands on human lung endothelial cell barrier function.

    Directory of Open Access Journals (Sweden)

    You-Hong Cheng

    Full Text Available Recent evidence suggests that chemokine CXCL12, the cognate agonist of chemokine receptors CXCR4 and ACKR3, reduces thrombin-mediated impairment of endothelial barrier function. A detailed characterization of the effects of CXCL12 on thrombin-mediated human lung endothelial hyperpermeability is lacking and structure-function correlations are not available. Furthermore, effects of other CXCR4/ACKR3 ligands on lung endothelial barrier function are unknown. Thus, we tested the effects of a panel of CXCR4/ACKR3 ligands (CXCL12, CXCL11, ubiquitin, AMD3100, TC14012 and compared the CXCR4/ACKR3 activities of CXCL12 variants (CXCL12α/β, CXCL12(3-68, CXCL121, CXCL122, CXCL12-S-S4V, CXCL12-R47E, CXCL12-K27A/R41A/R47A with their effects on human lung endothelial barrier function in permeability assays. CXCL12α enhanced human primary pulmonary artery endothelial cell (hPPAEC barrier function, whereas CXCL11, ubiquitin, AMD3100 and TC14012 were ineffective. Pre-treatment of hPPAEC with CXCL12α and ubiquitin reduced thrombin-mediated hyperpermeability. CXCL12α-treatment of hPPAEC after thrombin exposure reduced barrier function impairment by 70% (EC50 0.05-0.5nM, which could be antagonized with AMD3100; ubiquitin (0.03-3μM was ineffective. In a human lung microvascular endothelial cell line (HULEC5a, CXCL12α and ubiquitin post-treatment attenuated thrombin-induced hyperpermeability to a similar degree. CXCL12(3-68 was inefficient to activate CXCR4 in Presto-Tango β-arrestin2 recruitment assays; CXCL12-S-S4V, CXCL12-R47E and CXCL12-K27A/R41A/R47A showed significantly reduced potencies to activate CXCR4. While the potencies of all proteins in ACKR3 Presto-Tango assays were comparable, the efficacy of CXCL12(3-68 to activate ACKR3 was significantly reduced. The potencies to attenuate thrombin-mediated hPPAEC barrier function impairment were: CXCL12α/β, CXCL121, CXCL12-K27A/R41A/R47A > CXCL12-S-S4V, CXCL12-R47E > CXCL122 > CXCL12(3-68. Our findings

  9. Lipoprotein lipase and endothelial lipase in human testis and in germ cell neoplasms

    DEFF Research Database (Denmark)

    Nielsen, J E; Lindegaard, M L; Friis-Hansen, L

    2009-01-01

    The aim of this study was to investigate endothelial lipase (EL, LIPG) and lipoprotein lipase (LPL) mRNA and protein expression in normal human testis and testicular germ cell tumours (GCT). Both EL and LPL were expressed in normal seminiferous tubules and in the interstitial compartment. EL m......RNA and protein were found in all germ cells as well as in Sertoli and Leydig cells. EL mRNA was abundant in pre-invasive carcinoma in situ (CIS) cells and GCTs, and EL protein was present in the cytoplasm of these cells. LPL mRNA was also relatively abundant in germ cells, Sertoli cells, CIS cells and GCTs...

  10. Exercise-mediated changes in conduit artery wall thickness in humans: role of shear stress

    NARCIS (Netherlands)

    Thijssen, D.H.J.; Dawson, E.A.; Munckhof, I.C. van den; Tinken, T.M.; Drijver, E. den; Hopkins, N.; Cable, N.T.; Green, D.J.

    2011-01-01

    Episodic increases in shear stress have been proposed as a mechanism that induces training-induced adaptation in arterial wall remodeling in humans. To address this hypothesis in humans, we examined bilateral brachial artery wall thickness using high-resolution ultrasound in healthy men across an

  11. MAPK signaling pathway regulates cerebrovascular receptor expression in human cerebral arteries

    DEFF Research Database (Denmark)

    Ansar, Saema; Eftekhari, Sajedeh; Waldsee, Roya

    2013-01-01

    if the upregulation of contractile cerebrovascular receptors after 48 h of organ culture of human cerebral arteries involves MAPK pathways and if it can be prevented by a MEK1/2 inhibitor. Human cerebral arteries were obtained from patients undergoing intracranial tumor surgery. The vessels were divided into ring...

  12. Endothelial-derived tissue factor pathway inhibitor regulates arterial thrombosis but is not required for development or hemostasis.

    NARCIS (Netherlands)

    White, T.A.; Johnson, T.; Zarzhevsky, N.; Tom, C.; Delacroix, S.; Holroyd, E.W.; Maroney, S.A.; Singh, R.; Pan, S.; Fay, W.P.; Deursen, J.M.A. van; Mast, A.E.; Sandhu, G.S.; Simari, R.D.

    2010-01-01

    The antithrombotic surface of endothelium is regulated in a coordinated manner. Tissue factor pathway inhibitor (TFPI) localized at the endothelial cell surface regulates the production of FXa by inhibiting the TF/VIIa complex. Systemic homozygotic deletion of the first Kunitz (K1) domain of TFPI

  13. Endothelial SIRT1 prevents adverse arterial remodeling by facilitating HERC2-mediated degradation of acetylated LKB1

    DEFF Research Database (Denmark)

    Bai, Bo; Man, Andy W C; Yang, Kangmin

    2016-01-01

    Aims-SIRT1 exerts potent activity against cellular senescence and vascular ageing. By decreasing LKB1 protein levels, it promotes the survival and regeneration of endothelial cells. The present study aims to investigate the molecular mechanisms underlying SIRT1-mediated LKB1 degradation for the p...

  14. Uterine Vasculature Remodeling in Human Pregnancy Involves Functional Macrochimerism by Endothelial Colony Forming Cells of Fetal Origin

    Science.gov (United States)

    Sipos, Peter I; Rens, Willem; Schlecht, HÉlène; Fan, Xiaohu; Wareing, Mark; Hayward, Christina; Hubel, Carl A; Bourque, Stephane; Baker, Philip N; Davidge, Sandra T; Sibley, Colin P; Crocker, Ian P

    2013-01-01

    The potency of adult-derived circulating progenitor endothelial colony forming cells (ECFCs) is drastically surpassed by their fetal counterparts. Human pregnancy is associated with robust intensification of blood flow and vascular expansion in the uterus, crucial for placental perfusion and fetal supply. Here, we investigate whether fetal ECFCs transmigrate to maternal bloodstream and home to locations of maternal vasculogenesis, primarily the pregnant uterus. In the first instance, endothelial-like cells, originating from mouse fetuses expressing paternal eGFP, were identified within uterine endothelia. Subsequently, LacZ or enhanced green fluorescent protein (eGFP)-labeled human fetal ECFCs, transplanted into immunodeficient (NOD/SCID) fetuses on D15.5 pregnancy, showed similar integration into the mouse uterus by term. Mature endothelial controls (human umbilical vein endothelial cells), similarly introduced, were unequivocally absent. In humans, SRY was detected in 6 of 12 myometrial microvessels obtained from women delivering male babies. The copy number was calculated at 175 [IQR 149–471] fetal cells per millimeter square endothelium, constituting 12.5% of maternal vessel lumina. Cross-sections of similar human vessels, hybridized for Y-chromosome, positively identified endothelial-associated fetal cells. It appears that through ECFC donation, fetuses assist maternal uterine vascular expansion in pregnancy, potentiating placental perfusion and consequently their own fetal supply. In addition to fetal growth, this cellular mechanism holds implications for materno-fetal immune interactions and long-term maternal vascular health. PMID:23554274

  15. Human very Small Embryonic-like Cells Support Vascular Maturation and Therapeutic Revascularization Induced by Endothelial Progenitor Cells.

    Science.gov (United States)

    Guerin, Coralie L; Rossi, Elisa; Saubamea, Bruno; Cras, Audrey; Mignon, Virginie; Silvestre, Jean-Sébastien; Smadja, David M

    2017-08-01

    Very small embryonic-like stem cells (VSELs) are major pluripotent stem cells defined as cells of small size being Lineage- negative, CD133-positive, and CD45-negative. We previously described that human bone marrow VSELs were able to differentiate into endothelial cells and promoted post-ischemic revascularization in mice with surgically induced critical limb ischemia. In the present work, we isolated bone marrow VSELs from patients with critical limb ischemia and studied their ability to support endothelial progenitor cells therapeutic capacity and revascularization potential. Sorted bone marrow VSELs cultured in angiogenic media were co-injected with endothelial progenitor cells and have been show to trigger post-ischemic revascularization in immunodeficient mice, and support vessel formation in vivo in Matrigel implants better than human bone marrow mesenchymal stem cells. In conclusion, VSELs are a potential new source of therapeutic cells that may give rise to cells of the endothelial and perivascular lineage in humans. VSELs are the first real vasculogenic stem cells able to differentiate in endothelial and perivascular lineage in human adult described from now. Thus, because VSELs presence have been proposed in adult tissues, we think that VSELs are CD45 negative stem cells able to give rise to vascular regeneration in human tissues and vessels.

  16. The effect of candesartan on pentraxin-3 plasma levels as marker of endothelial dysfunction in patients with essential arterial hypertension.

    Science.gov (United States)

    Buda, V; Andor, M; Cristescu, C; Voicu, M; Cochera, F; Tuduce, P; Petrescu, L; Tomescu, M C

    2017-08-01

    In the last decades, the studies performed on the field of endothelial dysfunction confirmed the fact that the starting point of this pathology is the inflammation. Several inflammatory biomarkers had been discovered and studied, ones showing systemic inflammation, and others being more specific biomarkers and showing the local inflammation. Pentraxin-3 (PTX3) is a new inflammatory biomarker, from the same family as high-selectivity C-reactive protein (hs-CRP), but it is a more specific biomarker, due to its local production: the endothelial cells and not the liver like in the case of hs-CRP. Several antihypertensive classes of drugs seem to have a positive impact on reducing the local endothelial inflammation, beyond their effect of lowering the blood pressure, so this study aims to analyze the effect of candesartan on the two inflammatory biomarkers: PTX3 and CRP, compared with other antihypertensive drugs, in hypertensive patients with endothelial dysfunction. A total of 365 patients were included in the study: 127 hypertensive patients were under treatment with candesartan, 134 patients were under treatment with other hypotensive medication (beta blockers, calcium channel blockers, and diuretics), both groups with controlled values of blood pressure, and 104 were normotensive persons. Classical methods of assessing the endothelial function were correlated with these biochemical markers. The patients treated with candesartan had a significant lower value of PTX3 and hs-CRP, compared with those under treatment with other antihypertensive medication as follows: PTX3: 0.61 ± 0.49 vs 0.95 ± 1.04 ng/ml, P = 0.006 and hs-CRP: 0.19 ± 0.20 vs 0.20 ± 0.22 mg/dl, P = 0.54. Candesartan decreases PTX3 and hs-CRP plasma levels more powerful than other classes of antihypertensive drugs (beta blockers, calcium channel blockers, and diuretics), so we may assume that candesartan has a more potent action in reversing endothelial dysfunction and that it offers a

  17. Differentiation, Evaluation, and Application of Human Induced Pluripotent Stem Cell-Derived Endothelial Cells.

    Science.gov (United States)

    Lin, Yang; Gil, Chang-Hyun; Yoder, Mervin C

    2017-11-01

    The emergence of induced pluripotent stem cell (iPSC) technology paves the way to generate large numbers of patient-specific endothelial cells (ECs) that can be potentially delivered for regenerative medicine in patients with cardiovascular disease. In the last decade, numerous protocols that differentiate EC from iPSC have been developed by many groups. In this review, we will discuss several common strategies that have been optimized for human iPSC-EC differentiation and subsequent studies that have evaluated the potential of human iPSC-EC as a cell therapy or as a tool in disease modeling. In addition, we will emphasize the importance of using in vivo vessel-forming ability and in vitro clonogenic colony-forming potential as a gold standard with which to evaluate the quality of human iPSC-EC derived from various protocols. © 2017 American Heart Association, Inc.

  18. Scutellarin promotes in vitro angiogenesis in human umbilical vein endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Zhong-Xiu-Zi [Department of Anatomy, Basic Medical Science College, Harbin Medical University, Harbin (China); Huang, Da-Yong [Department of Oncology, The Second Clinical Hospital, Harbin Medical University, Harbin (China); Li, Hai-Xia; Zhang, Li-Na; Lv, Yan-Hong; Cui, Hai-Dong [Department of Anatomy, Basic Medical Science College, Harbin Medical University, Harbin (China); Zheng, Jin-Hua, E-mail: jhzhenghrbmu@yahoo.cn [Department of Anatomy, Basic Medical Science College, Harbin Medical University, Harbin (China)

    2010-09-10

    Research highlights: {yields} It has been shown that scutellarin exhibits a variety of pharmacological actions, including anti-oxidative, anti-inflammatory, vasodilator as well as cardiovascular and cerebrovascular ischemia protective effects, indicating beneficial vascular effects of scutellarin. Therefore, it is speculated that scutellarin may be able to stimulate angiogenesis, which could be beneficial in the treatment of ischemic disease, wound healing and tissue regeneration. {yields} The purpose of the present study was to elucidate the direct angiogenic actions of scutellarin on human umbilical vein endothelial cells (HUVECs) in vitro. {yields} Our results showed that scutellarin to directly induce in vitro angiogenesis, which is closely correlated with upregulated MMP-2 expression, suggesting a potential for increasing angiogenesis. -- Abstract: Angiogenesis is critical to a wide range of physiological and pathological processes. Scutellarin, a major flavonoid of a Chinese herbal medicine Erigeron breviscapus (Vant.) Hand. Mazz. has been shown to offer beneficial effects on cardiovascular and cerebrovascular functions. However, scutellarin's effects on angiogenesis and underlying mechanisms are not fully elucidated. Here, we studied angiogenic effects of scutellarin on human umbilical vein endothelial cells (HUVECs) in vitro. Scutellarin was found by MTT assay to induce proliferation of HUVECs. In scutellarin-treated HUVECs, a dramatic increase in migration was measured by wound healing assay; Transwell chamber assay found significantly more invading cells in scutellarin-treated groups. Scutellarin also promoted capillary-like tube formation in HUVECs on Matrigel, and significantly upregulated platelet endothelial cell adhesion molecule-1 at both mRNA and protein levels. Scutellarin's angiogenic mechanism was investigated in vitro by measuring expression of angiogenic factors associated with cell migration and invasion. Scutellarin strongly

  19. The cytotoxicity evaluation of magnetic iron oxide nanoparticles on human aortic endothelial cells.

    Science.gov (United States)

    Ge, Gaoyuan; Wu, Hengfang; Xiong, Fei; Zhang, Yu; Guo, Zhirui; Bian, Zhiping; Xu, Jindan; Gu, Chunrong; Gu, Ning; Chen, Xiangjian; Yang, Di

    2013-05-07

    One major obstacle for successful application of nanoparticles in medicine is its potential nanotoxicity on the environment and human health. In this study, we evaluated the cytotoxicity effect of dimercaptosuccinic acid-coated iron oxide (DMSA-Fe2O3) using cultured human aortic endothelial cells (HAECs). Our results showed that DMSA-Fe2O3 in the culture medium could be absorbed into HAECs, and dispersed in the cytoplasm. The cytotoxicity effect of DMSA-Fe2O3 on HAECs was dose-dependent, and the concentrations no more than 0.02 mg/ml had little toxic effect which were revealed by tetrazolium dye assay. Meanwhile, the cell injury biomarker, lactate dehydrogenase, was not significantly higher than that from control cells (without DMSA-Fe2O3). However, the endocrine function for endothelin-1 and prostacyclin I-2, as well as the urea transporter function, was altered even without obvious evidence of cell injury in this context. We also showed by real-time PCR analysis that DMSA-Fe2O3 exposure resulted in differential effects on the expressions of pro- and anti-apoptosis genes of HAECs. Meanwhile, it was noted that DMSA-Fe2O3 exposure could activate the expression of genes related to oxidative stress and adhesion molecules, which suggested that inflammatory response might be evoked. Moreover, we demonstrated by in vitro endothelial tube formation that even a small amount of DMSA-Fe2O3 (0.01 and 0.02 mg/ml) could inhibit angiogenesis by the HAECs. Altogether, these results indicate that DMSA-Fe2O3 have some cytotoxicity that may cause side effects on normal endothelial cells.

  20. Plant proteolytic enzyme papain abrogates angiogenic activation of human umbilical vein endothelial cells (HUVEC) in vitro

    Science.gov (United States)

    2013-01-01

    Background Vascular endothelial growth factor (VEGF) is a key regulator of physiologic and pathogenic angiogenesis in diseases such as cancer and diabetic retinopathy. It is known that cysteine proteases from plants, like bromelain and papain are capable to suppress inflammatory activation. Recent studies have demonstrated that they may interfere with angiogenesis related pathways as well. The aim of this study was to investigate the anti-angiogenic effects of papain on human umbilical vein endothelial cells (HUVEC) in vitro. Methods Cell viability after prolonged treatment with papain was investigated by life cell staining and lactate dehydrogenase release assay. Angiogenic activation was assessed by ELISA against phosphorylated proteins AKT, MEK1/2, ERK1/2, SAPK/JNK and p38-MAPK. Growth inhibition was determined by means of an MTT-assay and cell migration by means of a scratch assay. Capability to form a capillary network was investigated using a tube formation assay. Results Papain did not induce proteolysis or cell detachment of HUVEC in a concentration range between 0 and 25 μg/mL. Four hours treatment with 10 μg/mL papain resulted in a reduced susceptibility of endothelial cells to activation by VEGF as determined by phosphorylation levels of Akt, MEK1/2, SAPK/JNK. Papain exerted a distinct inhibitory effect on cell growth, cell migration and tube formation with inhibition of tube formation detectable at concentrations as low as 1 μg/mL. Bromelain and ficin displayed similar effects with regard to cell growth and tube formation. Conclusion Papain showed a strong anti-angiogenic effect in VEGF activated HUVEC. This effect may be due to interference with AKT, MEK1/2 and SAPK/JNK phosphorylation. Two other plant derived cysteine proteases displayed similar inhibition of HUVEC cell growth and tube formation. These findings indicate that plant proteolytic enzymes may have potential as preventive and therapeutic agents against angiogenesis related human diseases

  1. Human endothelial and platelet septin SEPT11: cloning of novel variants and characterisation of interaction partners.

    Science.gov (United States)

    Bartsch, Ingrid; Bläser, Susanne; Röseler, Sabrina; Sandrock, Kirstin; Busse, Anja; Huber, Michael; Rempp, Hansjörg; Lieber, Mareike; Horn, Julia; Brendle, Cornelia; Zieger, Barbara

    2010-12-01

    Septins are cytoskeletal GTPases forming heteropolymeric complexes involved in processes characterised by active membrane movement such as cytokinesis, vesicle trafficking, and exocytosis. Septins are expressed in non-mitotic cells such as neurons and platelets. SEPT11 belongs to the SEPT6 group and was identified as interaction partner of SEPT5. We cloned and characterised novel SEPT11 variants and investigated interaction partners of SEPT11 in platelets and human umbilical vein endothelial cells. An endothelial cell library was used for cloning novel SEPT11 variants. Using Northern analysis the different SEPT11 transcripts were illustrated. Interaction studies were performed using yeast two-hybrid system, precipitation, FRET, and immunofluorescence microscopy. We demonstrate that SEPT11 partners with SEPT2, SEPT4 and SEPT7 using yeast two-hybrid system and precipitation. The interaction of SEPT11 with SEPT7 is also demonstrated by FRET. In addition to the known SEPT11 transcript (SEPT11_v1) we identified a novel SEPT11 variant (SEPT11_v2) as interaction partner of SEPT4 and SEPT7. Library screening of an endothelial cell library also revealed the presence of this novel SEPT11_v2 transcript. In addition, a third SEPT11 variant (SEPT11_v3) was identified. Expression of SEPT11_v1 and of SEPT11_v2 and SEPT11_v3 in human brain regions was investigated by Northern analysis. Further interaction partners of SEPT11 are characterised using immunofluorescence. Co-localisation of SEPT2, SEPT4, SEPT7 and SEPT11 with tubulin and transferrin receptor (endocytotic marker) is demonstrated. In addition, co-localisation of SEPT4 and SEPT11 with the vesicle-associated protein synaptobrevin 1 (VAMP1), but not clearly with actin, was shown. Only SEPT2 and SEPT7 definitely co-localised with actin, but not clearly with VAMP1.