Ox-LDL induces apoptosis in human coronary artery endothelial cells: role of PKC, PTK, bcl-2, and Fas.
Oxidized low-density lipoprotein (ox-LDL) plays a critical role in the development of atherosclerosis. Recent studies show that ox-LDL may induce apoptosis of cultured rabbit smooth muscle cells and human macrophages. This study was designed to determine the modulation by ox-LDL of apoptosis in cultured human coronary arterial endothelial cells (HCAEC) during hypoxia-reoxygenation and to determine underlying mechanisms. When HCAEC were approximately 85% confluent, the cells were exposed to hypoxia (24 h)-reoxygenation (3 h), native LDL, or ox-LDL. Fragmented DNA end-labeling, DNA laddering, and light and electron microscopy were used to determine changes characteristic of apoptosis. Ox-LDL (20 microg/ml) increased apoptosis during hypoxia-reoxygenation compared with hypoxia-reoxygenation alone (P < 0.05). Low concentrations of ox-LDL (5 microg/ml) and native LDL (20 microg/ml) under identical conditions had no effect on the degree of apoptosis. Ox-LDL markedly decreased endogenous superoxide dismutase activity and increased lipid peroxidation in HCAEC. The presence of ox-LDL, but not native LDL, in cultured HCAEC resulted in the activation of protein kinase C (PKC) and protein tyrosine kinase (PTK). The specific PKC and PTK inhibitors significantly reduced ox-LDL-mediated apoptosis of HCAEC (P < 0.05). Hypoxia-reoxygenation significantly increased Fas expression and decreased bcl-2 expression in HCAEC lysate as determined by Western analysis. Ox-LDL further increased Fas expression and decreased bcl-2 expression. These data indicate that ox-LDL enhances hypoxia-reoxygenation-mediated apoptosis in HCAEC. Ox-LDL-mediated apoptosis of HCAEC appears to involve activation of PKC and PTK. In addition, ox-LDL modulates Fas and bcl-2 protein expression in HCAEC. This study also suggests that ox-LDL is more important than native LDL in hypoxia-reoxygenation-induced apoptosis.
Department of Medicine, University of Florida, and Veterans Affairs Medical Center, Gainesville, Florida 32610, USA.
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Yang
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Mehta
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D015687:Cell Hypoxia / Q000502:physiology; D002478:Cells, Cultured; D003331:Coronary Vessels; D003470:Culture Media; D053938:DNA Fragmentation; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D004789:Enzyme Activation; D006801:Humans; D015227:Lipid Peroxidation / Q000187:drug effects; D008077:Lipoproteins, LDL / Q000494:pharmacology* / Q000502:physiology; D010084:Oxidation-Reduction; D011493:Protein Kinase C / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D011817:Rabbits; D015398:Signal Transduction; D013482:Superoxide Dismutase / Q000378:metabolism; D019014:fas Receptor / Q000378:metabolism*
D003470:Culture Media; D008077:Lipoproteins, LDL; D019253:Proto-Oncogene Proteins c-bcl-2; D019014:fas Receptor; C099252:oxidized low density lipoprotein; D013482:Superoxide Dismutase; D011505:Protein-Tyrosine Kinases; D011493:Protein Kinase C