Dual effect of oxidized LDL on cell cycle in human endothelial cells through oxidative stress

Background. Oxidized low-density lipoprotein (OxLDL) exerts proliferation a nd apoptosis in vascular cells, depending on its concentration and the: exp osure time. Various steps in the cell cycle and in the apoptotic signaling cascade are modulated by O-2(-), and OxLDL stimulates vascular O-2(-) forma tion. Here we studied the role of NADPH oxidase, a potential source for O-2 (-) formation after OxLDL stimulation, in cell proliferation, and we invest igated whether OxLDL influences anti-apoptotic genes in cultured human umbi lical vein endothelial cells (HUVEC). Methods and Results. OxLDL dose-dependently (10 to 300 mug/mL) stimulated O -2(-) formation in HUVEC (detected by cytochrome c assay and by chemilumine scence of lucigenin). Low OxLDL concentrations (5 to 10 mug/mL) induced pro liferation (detected by H-3-thymidine incorporation), while higher concentr ations (50 to 300 mug/mL) induced apoptotic cell death (detected by Annexin assay and DNA fragmentation). Proliferation was blocked by the antioxidant s SOD and catalase and by diphenyleneiodonium (10 mu mol/L), an inhibitor o f the O-2(-) generating NADPH oxidase. In addition, cells transfected with antisense oligonucleotides for NADPH oxidase showed a significantly reduced O-2(-) formation after stimulation with OxLDL. The OxLDL effect on apoptos is was also blocked by antioxidants. Since endothelial cells are protected against apoptosis through anti-apoptotic genes, we investigated whether OxL DL overcomes protection against apoptosis through suppression of the anti-a poptotic gene A20, a zinc-finger protein. OxLDL suppressed the expression o f A20 in a dose-dependent manner. Conclusion. These data indicate that OxLDL has a dual effect on cell cycle in HUVEC, inducing proliferation at low and apoptosis at higher concentrati ons. Both effects are mediated by O-2(-) formation, with NADPH oxidase bein g a major source for O-2(-). Thus, OxLDL contributes importantly to vascula r cellular turnover through the induction of oxidative stress.