Effect Of Oxidized High Density Lipoprotein On Endothelial Progenitor Cells Function And Its Mechanism

Background:High density lipoprotein (HDL) levels inversely correlate with cardiovascular events due to the protective effects on the vascular wall. Endothelial progenitor cells (EPCs) play an important role in neovascularization in chronic ischemic disease. EPCs number and functions are influenced by a lot of factors and EPCs number and functions are important predictors for cardiovascular events. Recent studies demonstrated that HDL could directly stimulate EPCs differentiation and enhance ischemia-induced angiogenesis. However, a number of reports have indicated that HDL is susceptible to oxidation and structural modifications in case of atherosclerosis and diabetes. Unlike HDL, oxidized HDL (oxHDL) lose the important protective functions and impacts neovascularization negatively. However, the effect of oxHDL on EPCs is still unclear.Objective:The objective of this study is to investigate the effect of oxHDL on EPCs and the underlying mechanism.Method:EPCs were isolated from human peripheral venous blood. Then the whole study were divided into three parts:(1) The human oligonucleotide microarray was applied to isolate and classify the differentially expressed genes between untreated EPCs and ox-HDL treated EPCs. Quantitative real-time PCR and Western blot were used to confirm the results from the microarray.(2) Cells were cultured and stimulated with different concentrations of oxHDL (0-50μg/mL), HDL (50μg/mL) in the absence and presence anti-CD36 neutralizing antibody. EPCs apoptosis and migration, angiogenesis and intracellular reactive oxygen species (ROS) levels were assayed. And the expression of thrombospondin-1(TSP-1) and vascular endothelial growth factor (VEGF) were measured by real-time PCR and ELISA. (3)The expression of mitogen-activated protein kinase (MAPK) family proteins (p38, ERK and JNK) and NF-κB were measured by by Western blot and EMSA.Results:(1) The results of human oligonucleotide microarray showed oxHDL promoted the gene expression of CD36. Quantitative real-time PCR and Western blot results confirmed the results from the microarray. (2) oxHDL increased apoptosis and intracellular ROS levels and reduced the ability of migration and angiogenesis of EPCs in a dose dependent manner (all P<0.05). oxHDL also upregulated the expression of TSP-1 without affecting VEGF expression of EPCs. All oxHDL-mediated effects on EPCs could be significantly attenuated by pretreatment with anti-CD36 neutralizing antibody. (3) P38 MAPK and NF-κB were activated post oxHDL stimulation. All oxHDL-mediated effects on EPCs could be significantly attenuated by pretreatment with anti-CD36 neutralizing antibody.Conclution:These findings suggest that oxHDL promote apoptosis and inhibit migration and angiogenesis function of EPCs. These effects were mainly mediated via scavenger receptor CD36 and P38/MAPK, NF-κB signaling pathways. The ability of oxHDL to negatively influence EPCs biology may represent a novel pathological mechanism for the development and progression of cardiovascular disease.