Abstract 66: The Role of the Viral Nef Protein as a Mediator of HIV-1--Induced Endothelial Dysfunction

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Ting Wang
Keyword(s):  
Cardiovascular Disease ◽  
T Cells ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Endothelial Dysfunction ◽  
Binding Site ◽  
Cell Activation ◽  
Endothelial Activation ◽  
Endothelial Cell Activation ◽  
Hiv 1

With the prevalence of antiviral therapy in the developed world, many HIV-1-infected people die of diseases other than AIDS. One of the emerging major causes is cardiovascular disease, leading to the prediction that the majority of HIV-1 patients are expected to develop cardiovascular complications. Endothelial dysfunction is thought to be a key event in the development of cardiovascular diseases, particularly atherosclerosis. Assays testing the effect of HIV-1 on endothelial activation shows that direct contact with HIV-1 infected T cells enhance endothelial cell activation to a greater extent than HIV-1 alone, suggesting an intracellular HIV-1 protein is responsible for endothelial activation. The HIV-1 viral protein Nef, which is responsible for T cell activation and maintenance of high viral loads in vivo , has been shown to mediate its own transfer to bystander cells. We demonstrate here for the first time that Nef induces nanotube-like conduits connecting T cells and endothelial cells. We also show that Nef is transferred from T cells to endothelial cells via these nanotubes, and is necessary and sufficient for endothelial cell activation. Moreover, we show that SIV-infected macaques exhibit endothelial Nef expression in coronary arteries. Nef expression in endothelial cells causes endothelial apoptosis, ROS and MCP-1 production. Interestingly, a Nef SH3 binding site mutant abolishes Nef-induced apoptosis and ROS formation and reduces MCP-1 production in endothelial cells, suggesting that the Nef SH3 binding site is critical for Nef effects on endothelial cells. Nef induces apoptosis of endothelial cells through an NADPH oxidase- and ROS-dependent mechanism, while Nef-induced MCP-1 production is NF-kB dependent. Taken together, these data suggest that Nef can mediate its transfer from T cells to endothelial cells through nanotubes to enhance endothelial dysfunction.Thus, Nef is a promising new therapeutic target for reducing the risk for cardiovascular disease in the HIV-1 positive population.

scholarly journals Brucella abortus-Stimulated Platelets Activate Brain Microvascular Endothelial Cells Increasing Cell Transmigration through the Erk1/2 Pathway

Pathogens ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 708
Author(s):  
Ana María Rodríguez ◽  
Aldana Trotta ◽  
Agustina P. Melnyczajko ◽  
M. Cruz Miraglia ◽  
Kwang Sik Kim ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Brucella Abortus ◽  
Cell Activation ◽  
Transendothelial Migration ◽  
Endothelial Activation ◽  
Microvascular Endothelial Cell ◽  
Inflammatory Disorder ◽  
Endothelial Cell Activation ◽  
Microvascular Endothelial

Central nervous system invasion by bacteria of the genus Brucella results in an inflammatory disorder called neurobrucellosis. A common feature associated with this pathology is blood–brain barrier (BBB) activation. However, the underlying mechanisms involved with such BBB activation remain unknown. The aim of this work was to investigate the role of Brucella abortus-stimulated platelets on human brain microvascular endothelial cell (HBMEC) activation. Platelets enhanced HBMEC activation in response to B. abortus infection. Furthermore, supernatants from B. abortus-stimulated platelets also activated brain endothelial cells, inducing increased secretion of IL-6, IL-8, CCL-2 as well as ICAM-1 and CD40 upregulation on HBMEC compared with supernatants from unstimulated platelets. Outer membrane protein 19, a B. abortus lipoprotein, recapitulated B. abortus-mediated activation of HBMECs by platelets. In addition, supernatants from B. abortus-activated platelets promoted transendothelial migration of neutrophils and monocytes. Finally, using a pharmacological inhibitor, we demonstrated that the Erk1/2 pathway is involved in the endothelial activation induced by B. abortus-stimulated platelets and also in transendothelial migration of neutrophils. These results describe a mechanism whereby B. abortus-stimulated platelets induce endothelial cell activation, promoting neutrophils and monocytes to traverse the BBB probably contributing to the inflammatory pathology of neurobrucellosis.

scholarly journals A novel flow cytometry-based quantitative monocyte adhesion assay to estimate endothelial cell activation in vitro

BioTechniques ◽  
2020 ◽  
Vol 68 (6) ◽  
pp. 325-333
Author(s):  
Vinnyfred Vincent ◽  
Himani Thakkar ◽  
Anjali Verma ◽  
Atanu Sen ◽  
Nikhil Chandran ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Activation ◽  
Endothelial Activation ◽  
Adhesion Assay ◽  
Monocyte Adhesion ◽  
Endothelial Cell Activation ◽  
Functional Level

One of the earliest events in the development of atherosclerosis is endothelial activation, which is estimated in vitro at the functional level by quantifying monocyte adhesion. This involves the incubation of fluorescently labeled monocytes on top of cultured endothelial cells and quantifying the number of adhered monocytes. Currently, the quantification of adhered monocytes is done using microscopy or by lysing the cells and estimating the fluorescence. Here we present a novel flow cytometry-based method for the quantification of monocyte adhesion. This method could quantify the average number of monocytes adhered to a single endothelial cell after monocyte adhesion assay, and was also sensitive to the level of activation of endothelial cells. Flow cytometry-based quantification requires less time and effort compared with microscopy-based quantification.

Endothelial dysfunction as a possible link between C-reactive protein levels and cardiovascular disease

2000 ◽  
Vol 98 (5) ◽  
pp. 531-535 ◽  
Author(s):  
Stephen J. CLELAND ◽  
Naveed SATTAR ◽  
John R. PETRIE ◽  
Nita G. FOROUHI ◽  
Henry L. ELLIOTT ◽  
...  
Keyword(s):  
Cardiovascular Disease ◽  
Endothelial Cell ◽  
Endothelial Dysfunction ◽  
Cell Activation ◽  
Density Lipoprotein ◽  
Low Grade ◽  
C Reactive Protein ◽  
Endothelial Cell Activation ◽  
Reactive Protein ◽  
The Relationship

Low-grade chronic inflammation, characterized by elevated plasma concentrations of C-reactive protein (CRP), is associated with an increased risk of atherosclerotic cardiovascular disease. Endothelial cell activation is an early event in atherogenesis, and previous studies have reported correlations between indirect markers of endothelial cell activation and CRP concentration. Therefore, in the present study, we measured CRP concentration (and leptin concentration as an index of fat mass) in nine healthy subjects (mean age 53±8.1 years; body mass index 27±3.2 kg/m2; mean arterial blood pressure 101±9.0 mmHg) undergoing measurement of basal endothelial nitric oxide (NO) synthesis using intra-brachial infusions of NG-monomethyl-l-arginine (l-NMMA; a substrate inhibitor of endothelial NO synthase) and noradrenaline (a non-specific control vasoconstrictor). In univariate analysis, CRP concentration was correlated with (i) the percentage decrease in forearm blood flow (FBF) during l-NMMA infusion (r = 0.85, P = 0.004); and (ii) the serum leptin concentration (r = 0.65, P = 0.05). In multivariate analysis, the relationship between CRP concentration and the FBF response to l-NMMA remained significant when age and leptin (t = 2.65, P = 0.045), age and BMI (t = 3.69, P = 0.014), or age and low-density-lipoprotein-cholesterol plus high-density-lipoprotein-cholesterol (t = 3.37, P = 0.044), were included in regression models. In contrast, the response of FBF to noradrenaline was not significantly related to CRP concentration. These data demonstrate for the first time a relationship between low-grade chronic inflammation and basal endothelial NO synthesis (measured using an invasive method), and support the notion that endothelial dysfunction is a critical intermediate phenotype in the relationship between inflammation and cardiovascular disease.

106. IL-6 INCREASES THE SHEDDING OF NECROTIC TROPHOBLASTS FROM PLACENTAL EXPLANTS

2009 ◽  
Vol 21 (9) ◽  
pp. 25
Author(s):  
Q. Chen ◽  
L. Chen ◽  
B. Liu ◽  
H. Zhao ◽  
P. R. Stone ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Activation ◽  
Elevated Serum ◽  
Serum Levels ◽  
Endothelial Activation ◽  
Maternal Blood ◽  
Endothelial Cell Activation ◽  
Cell Monolayers ◽  
Cell Dysfunction

Preeclampsia (PE) is characterised by elevated maternal blood pressure, preceded by endothelial cell dysfunction. Dead trophoblasts, shed from the placenta may be one of the factors that trigger PE. Women with PE frequently have elevated serum levels of inflammatory markers such as, IL-6 and TNF a but their functional significance is unclear. In this study we investigated whether these or other cytokines can alter trophoblast shedding from placental explants. Placental explants were treated with 9 different cytokines for 72 hours. Shed trophoblasts then were harvested using our published method1. The numbers of trophoblasts shed were quantified by automated cell counter. Expression of active of caspases 3&7 by the shed trophoblasts was determined using a FLICA kit. The trophoblasts shed from cytokine-treated or control explants were exposed to endothelial cell monolayers and endothelial activation determined by ELISA for cell surface ICAM-1. Treatment of explants with IL-6 caused a 50% increase (p=0.001), while TNF a and TGF b 1, caused smaller significant increases in the numbers of trophoblasts shed. Trophoblasts shed from explants treated with IL-6, TGF b 1, or TGF b 3 expressed significantly less active caspases 3&7 than controls or trophoblasts shed from explants treated with other cytokines. Exposing trophoblasts shed from IL-6- or TGF b 1-treated explants to endothelial cells caused a significant (P<0.001) increase in endothelial activation. Normally trophoblasts shed from the placenta die by an apoptosis-like process and their phagocytosis by endothelial cells is silent but a shift to shedding of necrotic trophoblasts can lead to endothelial cell activation 2. However, it remains unclear what might trigger a shift from apoptotic to necrotic trophoblast death. This study suggests that IL-6 and possibly other cytokines can alter both the number and the nature of shed trophoblasts such that the trophoblast are more necrotic and their phagocytosis by maternal endothelial cells could contribute to the pathogenesis of preeclampsia.

scholarly journals Functional interactions of T cells with endothelial cells: the role of CD40L-CD40-mediated signals.

1995 ◽  
Vol 182 (6) ◽  
pp. 1857-1864 ◽  
Author(s):  
M J Yellin ◽  
J Brett ◽  
D Baum ◽  
A Matsushima ◽  
M Szabolcs ◽  
...  
Keyword(s):  
T Cells ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Surface ◽  
Cell Activation ◽  
Endothelial Cell Activation ◽  
Cd40 Expression

CD40 is expressed on a variety of cells, including B cells, monocytes, dendritic cells, and fibroblasts. CD40 interacts with CD40L, a 30-33-kD activation-induced CD4+ T cell surface molecule. CD40L-CD40 interactions are known to play key roles in B cell activation and differentiation in vitro and in vivo. We now report that normal human endothelial cells also express CD40 in situ, and CD40L-CD40 interactions induce endothelial cell activation in vitro. Frozen sections from normal spleen, thyroid, skin, muscle, kidney, lung, or umbilical cord were studied for CD40 expression by immunohistochemistry. Endothelial cells from all tissues studied express CD40 in situ. Moreover, human umbilical vein endothelial cells (HUVEC) express CD40 in vitro, and recombinant interferon gamma induces HUVEC CD40 upregulation. CD40 expression on HUVEC is functionally significant because CD40L+ Jurkat T cells or CD40L+ 293 kidney cell transfectants, but not control cells, upregulate HUVEC CD54 (intercellular adhesion molecule-1), CD62E (E-selectin), and CD106 (vascular cell adhesion molecule-1) expression in vitro. Moreover, the kinetics of CD40L-, interleukin 1-, or tumor necrosis factor alpha-induced CD54, CD62E, and CD106 upregulation on HUVEC are similar. Finally, CD40L-CD40 interactions do not induce CD80, CD86, or major histocompatibility complex class II expression on HUVEC in vitro. These results demonstrate that CD40L-CD40 interactions induce endothelial cell activation in vitro. Moreover, they suggest a mechanism by which activated CD4+ T cells may augment inflammatory responses in vivo by upregulating the expression of endothelial cell surface adhesion molecules.

scholarly journals Endothelial Activation in Orientia tsutsugamushi Infection Is Mediated by Cytokine Secretion From Infected Monocytes

2021 ◽  
Vol 11 ◽  
Author(s):  
Wiwit Tantibhedhyangkul ◽  
Sutthicha Matamnan ◽  
Asma Longkunan ◽  
Chawikan Boonwong ◽  
Ladawan Khowawisetsut
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Activation ◽  
Endothelial Activation ◽  
Cytokine Secretion ◽  
Cell Surface Expression ◽  
Orientia Tsutsugamushi ◽  
Endothelial Cell Activation ◽  
Chemokine Production ◽  
Delay In Diagnosis

Scrub typhus, caused by Orientia tsutsugamushi, is a common systemic infection in Asia. Delay in diagnosis and treatment can lead to vasculitis in the visceral organs and other complications. The mechanisms that drive endothelial activation and the inflammatory response in O. tsutsugamushi infection remain unknown. In addition, the interaction between monocytes and endothelial cells is still unclear. Here we demonstrate that O. tsutsugamushi-infected human dermal microvascular endothelial cells produced moderate levels of chemokines and low levels of IL-6 and IFN-β, but not TNF or IL-1β. Recombinant TNF and cytokine-rich supernatants from infected monocytes markedly enhanced chemokine production in infected endothelial cells. We also show that TNF and monocyte supernatants, but not O. tsutsugamushi infection of endothelial cells per se, upregulated the endothelial cell surface expression of ICAM-1, E-selectin, and tissue factor. This finding was consistent with the inability of O. tsutsugamushi to induce cytokine secretion from endothelial cells. The upregulation of surface molecules after stimulation with monocyte supernatants was significantly reduced by neutralizing anti-TNF antibodies. These results suggest that endothelial cell activation and response are mainly mediated by inflammatory cytokines secreted from monocytes.

Annexin A2 mediates endothelial cell activation by antiphospholipid/anti-β2 glycoprotein I antibodies

Blood ◽  
2005 ◽  
Vol 105 (5) ◽  
pp. 1964-1969 ◽  
Author(s):  
Jianwei Zhang ◽  
Keith R. McCrae
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Activation ◽  
Endothelial Activation ◽  
Annexin A2 ◽  
Dependent Manner ◽  
Endothelial Cell Activation ◽  
Glycoprotein I ◽  
Endothelial Surface ◽  
Increased Risk

AbstractPatients with antiphospholipid antibodies (APLAs) are at increased risk for arterial and venous thrombosis. Many APLAs associated with these events react with β2 glycoprotein I (β2GPI), and endothelial cell reactive antibodies that activate endothelial cells in a β2GPI-dependent manner occur commonly in these patients. We previously reported that β2GPI binds with high affinity to annexin A2 on the endothelial surface, though the relevance of this interaction to APLA/anti-β2GPI antibody–induced endothelial activation has not been determined. In this report, we confirm that anti-β2GPI antibodies activate endothelial cells in the presence of β2GPI, and demonstrate that anti–annexin A2 antibodies directly cause endothelial cell activation of a similar magnitude and with a similar time course. Moreover, bivalent anti–annexin A2 F(ab′)2 fragments also caused endothelial cell activation, whereas monomeric Fab fragments not only did not cause activation, but blocked activation induced by anti–annexin A2 antibodies and F(ab′)2 fragments, as well as that caused by anti-β2GPI antibodies in the presence of β2GPI. These observations suggest a novel pathway for endothelial activation induced by APLA/anti-β2GPI antibodies that is initiated by cross-linking or clustering of annexin A2 on the endothelial surface.

Abstract 48: Deletion of the Neuronal Guidance Molecule Epha2 Reduces Inflammation in Experimental Atherosclerosis

2013 ◽  
Vol 33 (suppl_1) ◽  
Author(s):  
Steven D Funk ◽  
Arif Yurdagul ◽  
Jonette Green ◽  
Patrick Albert ◽  
Marshall McInnis ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Activation ◽  
Experimental Atherosclerosis ◽  
Marker Genes ◽  
Endothelial Cell Activation ◽  
Enhanced Expression ◽  
Guidance Molecule ◽  
Neuronal Guidance ◽  
Deficient Mice

Neuronal guidance molecules are increasingly implicated in inflammatory responses. Recently, our group demonstrated enhanced expression of the neuronal guidance molecule EphA2 and its ephrinA1 ligand in mouse and human atherosclerotic plaques, and elucidated a novel proinflammatory function for EphA2 perpetuating proinflammatory gene expression during endothelial cell activation. However, a direct role for Eph/ephrins in atherosclerosis has never been demonstrated. We now show that knocking out the EphA2 gene in Western diet-fed ApoE mice blunts atherosclerotic plaque location at multiple sites. This reduction in atherosclerosis is associated with decreased monocyte infiltration and diminished expression of proinflammatory genes. EphA2 reduction may affect monocyte homing through multiple mechanisms, since reducing EphA2 expression in cytokine-activated endothelial cells does not affect endothelial adhesion molecule expression or monocyte rolling but significantly decreases firm adhesion in primary human monocytes. Like endothelial cells, plaque macrophages also express EphA2, and macrophages derived from EphA2 deficient mice show diminished expression of M1 marker genes and enhanced expression of M2 marker genes compared to their ApoE counterparts. Surprisingly, EphA2 deficient mice show significantly elevated plasma cholesterol. However, this elevation does not involve increased LDL levels but instead occurs due to elevations in plasma HDL levels. Taken together, the current data suggest EphA2 inhibition results in a multifaceted protective effect on experimental atherosclerosis characterized by reduced endothelial cell activation, monocyte recruitment, and M1/M2 polarization and enhanced circulating HDL levels.

Abstract 327: Mitochondrial Functional Variation Contributes to Endothelial Cell Activation

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
David M Krzywanski ◽  
Bing Cheng ◽  
Xinggui Shen ◽  
Christopher Kevil
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Ethnic Groups ◽  
Mitochondrial Function ◽  
Cell Activation ◽  
Oxidant Stress ◽  
Oxygen Utilization ◽  
Endothelial Cell Activation ◽  
Inflammatory Conditions ◽  
Oxidant Production

Vascular oxidant stress contributes to endothelial dysfunction and plays a critical role in early stage cardiovascular disease (CVD) development. Changes in endothelial function due to oxidant stress may contribute to CVD initiation and progression through the development of a pro-inflammatory environment. Differences in mitochondrial function may contribute to this process and provide insight into why age of onset and clinical outcomes differ amongst individuals form distinct ethnic groups; but no reports demonstrate distinct mitochondrial functional parameters between normal cells. Consequently, we hypothesized that significant variations in normal mitochondrial function and oxidant production exist between endothelial cells from donors representing different ethnic groups. Aspects of mitochondrial oxygen utilization and oxidant production were assessed under basal and inflammatory conditions in human aortic endothelial cells (HAECs) isolated from African Americans (AA) and Caucasians (CA). Bioenergetic analysis indicates that compared to CA, AA HAEC utilized significantly less oxygen for ATP production, possess a lower maximal respiratory capacity, and have reduced electron leak. Significant differences in mitochondrial membrane potential, decreased expression of endothelial nitric oxide synthase, and increased levels of superoxide were also observed and AA HAEC supporting a pro-inflammatory phenotype. As a marker of endothelial cell activation, AA HAEC expressed increased levels of intercellular cell adhesion molecule-1 under both basal and inflammatory conditions that could be partially mitigated but treatment with the mitochondrially targeted antioxidant MitoTEMPO. These data demonstrate that fundamental differences exist in mitochondrial oxygen utilization and oxidant production between CA and AA HAEC and that these changes may affect endothelial cell activation. These findings are consistent with the hypothesis that differences in “normal” mitochondrial function amongst ethnic groups could influence individual susceptibility by contributing to vascular inflammation, providing important insights into the mechanisms that contribute human CVD development.

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