Induction of the cytoprotective enzyme heme oxygenase-1 by statins is enhanced in vascular endothelium exposed to laminar shear stress and impaired by disturbed flow.

In addition to cholesterol-lowering properties, statins exhibit lipid-independent immunomodulatory, anti-inflammatory actions. However, high concentrations are typically required to induce these effects in vitro, raising questions concerning therapeutic relevance. We present evidence that endothelial cell sensitivity to statins depends upon shear stress.

KLF2-dependent, shear stress-induced expression of CD59: a novel cytoprotective mechanism against complement-mediated injury in the vasculature.

Complement activation may predispose to vascular injury and atherogenesis. The atheroprotective actions of unidirectional laminar shear stress led us to explore its influence on endothelial cell expression of complement inhibitory proteins CD59 and decay-accelerating factor. Human umbilical vein and aortic endothelial cells were exposed to laminar shear stress (12 dynes/cm(2)) or disturbed flow (+/- 5 dynes/cm(2) at 1Hz) in a parallel plate flow chamber.

Isolation and analysis of large and small vessel endothelial cells.

The in vitro isolation, propagation and study of endothelial cells (EC) is an invaluable means by which the function of the vascular endothelium in physiology and patho-physiology can be explored. In recent years heterogeneity between large and small vessel EC, between arteries and veins, and between microvascular EC derived from different organs has become increasingly apparent. This has led to the development of protocols for the isolation of these different EC.

A protein kinase Cepsilon-anti-apoptotic kinase signaling complex protects human vascular endothelial cells against apoptosis through induction of Bcl-2.

Endothelial cell apoptosis is associated with vascular injury and predisposes to atherogenesis. Endothelial cells express anti-apoptotic genes including Bcl-2, Bcl-XL and survivin, which also contribute to angiogenesis and vascular remodeling. We report a central role for protein kinase Cepsilon (PKCepsilon) in the regulation of Bcl-2 expression and cytoprotection of human vascular endothelium against apoptosis.

Aprotinin inhibits proinflammatory activation of endothelial cells by thrombin through the protease-activated receptor 1.

Objective: Thrombin is generated in significant quantities during cardiopulmonary bypass and mediates adverse events, such as platelet aggregation and proinflammatory responses, through activation of the high-affinity thrombin receptor protease-activated receptor 1, which is expressed on platelets and endothelium. Thus antagonism of protease-activated receptor 1 might have broad therapeutic significance. Aprotinin, used clinically to reduce transfusion requirements and the inflammatory response to bypass, has been shown to inhibit protease-activated receptor 1 on platelets in vitro and in vivo.

A role for proteinase-activated receptor 2 and PKC-epsilon in thrombin-mediated induction of decay-accelerating factor on human endothelial cells.

Thrombin, an important mediator of thrombosis and inflammation, may also enhance vascular cytoprotection. Thus thrombin induces expression of the complement-inhibitory protein decay-accelerating factor (DAF) in human umbilical vein endothelial cells (HUVECs), thus increasing protection against complement-mediated injury. Using PKC isozyme-specific peptide antagonists and adenoviral constructs, we have shown in the present study that PKC-epsilon is the primary isozyme involved in DAF induction by thrombin.

Cytokine stimulated vascular cell adhesion molecule-1 (VCAM-1) ectodomain release is regulated by TIMP-3.

Objectives: Vascular cell adhesion molecule-1 (VCAM-1) is a cell surface adhesion molecule involved in the recruitment of leukocytes to endothelial cells on arterial walls during the pathogenesis of atherosclerosis. The soluble ectodomain of VCAM-1 (sVCAM-1) is proteolytically released from the cell surface into the circulation, a process which is up-regulated in patients with cardiovascular or inflammatory disease. Here we investigate mechanisms involved in sVCAM-1 generation in response to cytokine stimulation.

Decay-accelerating factor induction on vascular endothelium by vascular endothelial growth factor (VEGF) is mediated via a VEGF receptor-2 (VEGF-R2)- and protein kinase C-alpha/epsilon (PKCalpha/epsilon)-dependent cytoprotective signaling pathway and is inhibited by cyclosporin A.

Decay-accelerating factor (DAF), a membrane-bound complement regulatory protein, is up-regulated on endothelial cells (ECs) following treatment with vascular endothelial growth factor (VEGF), providing enhanced protection from complement-mediated injury. We explored the signaling pathways involved in this response. Incubation of human umbilical vein ECs with VEGF induced a 3-fold increase in DAF expression.

Decay-accelerating factor induction by tumour necrosis factor-alpha, through a phosphatidylinositol-3 kinase and protein kinase C-dependent pathway, protects murine vascular endothelial cells against complement deposition.

We have shown that human endothelial cells (EC) are protected against complement-mediated injury by the inducible expression of decay-accelerating factor (DAF). To understand further the importance of DAF regulation, we characterized EC DAF expression on murine EC in vitro and in vivo using a model of glomerulonephritis. Flow cytometry using the monoclonal antibody (mAb) Riko-3 [binds transmembrane- and glycosylphosphatidylinositol (GPI)-anchored DAF], mAb Riko-4 (binds GPI-anchored DAF) and reverse transcription-polymerase chain reaction (RT-PCR), demonstrated that murine EC DAF is GPI-anchored.

Statin-induced expression of decay-accelerating factor protects vascular endothelium against complement-mediated injury.

Complement-mediated vascular injury is important in the pathophysiology of atherosclerosis and myocardial infarction. Because recent evidence shows that statins have beneficial effects on endothelial cell (EC) function independent of lipid lowering, we explored the hypothesis that statins modulate vascular EC resistance to complement through the upregulation of complement-inhibitory proteins. Human umbilical vein and aortic ECs were treated with atorvastatin or simvastatin, and decay-accelerating factor (DAF), membrane cofactor protein, and CD59 expression was measured by flow cytometry.

bFGF and VEGF synergistically enhance endothelial cytoprotection via decay-accelerating factor induction.

The complement-regulatory protein decay-accelerating factor (DAF) can be upregulated on endothelial cells (EC) by protein kinase C (PKC)-dependent and -independent pathways. We hypothesized that basic fibroblast growth factor (bFGF) might induce EC DAF expression, providing a cytoprotective mechanism for angiogenic neovessels against complement-mediated injury. Incubation of umbilical vein, aortic, and dermal EC with bFGF or vascular endothelial growth factor (VEGF) significantly increased DAF expression.

Conditional immortalization of growth factor-responsive cardiac endothelial cells from H-2K(b)-tsA58 mice.

Although mouse endothelial cells (EC) may advance our understanding of endothelial function, primary EC remain difficult to isolate. We have established a murine cardiac endothelial cell line (MCEC-1) from transgenic mice harboring a temperature-sensitive simian virus 40 large TAg gene (tsA58 TAg) under H-2K(b) class I promoter control. MCEC-1 cells were characterized by their ability to form tubes, Griffonia simplicifolia isolectin B4 binding, and CD31, intercellular adhesion molecule (ICAM)-2, and endoglin expression.