Gene Expression Profiling of Apoptosis-Related Genes in Human Atherosclerosis

Abstract

Objective— Apoptosis substantially affects the cellularity and integrity of atherosclerotic plaques. It remains, however, unclear which key regulatory genes are involved. In this study, cDNA expression arrays were used to analyze transcript levels of 205 apoptosis-related genes in human carotid endarterectomy specimens versus nonatherosclerotic mammary arteries. Methods and Results— Seventeen genes with a 2- to 5-fold relative expression difference were identified. One of the most apparent changes in human plaques was the overexpression of death-associated protein (DAP) kinase (≈5-fold), a positive mediator of apoptotic cell death. Differential expression of DAP kinase mRNA in human plaques relative to mammary arteries was confirmed by quantitative reverse-transcription polymerase chain reaction. Western blotting and immunohistochemistry demonstrated enhanced levels of DAP kinase protein in the plaque with negligible expression in non-atherosclerotic vessels. DAP kinase was located predominantly in foam cells of smooth muscle cell (SMC) origin. Uptake of aggregated LDL by cultured aortic SMCs as well as exposure of SMCs to the short-chain acyl ceramide derivative N -hexanoyl- d -sphingosine (C 6 -ceramide) upregulated DAP kinase both at the mRNA and protein level. Conclusions— Our data demonstrate that cDNA array technology can identify novel genes that might participate in cell death pathways underlying atherogenesis.