Journal
CLINICA CHIMICA ACTA
Volume 341, Issue 1-2, Pages 83-91Publisher
ELSEVIER
DOI: 10.1016/j.cccn.2003.11.009
Keywords
apoptosis; nitric oxide; vascular smooth muscle cells; mechanism
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Background: In contrast to the anti-apoptotic action of nitric oxide (NO) on endothelial cells, NO exerts a proapoptotic effect on vascular smooth muscle cells (VSMCs). This study was designed to elucidate the mechanism underlying NO-induced apoptosis in rat VSMCs. Methods and results: (1) Using the terminal deoxynucleotidyl transferase-mediated digoxigenin-11-dUTP nick end labeling (TUNEL) assay and fluorescence activated cell sorter (FACS) analyses, apoptosis of rat VSMCs were confirmed after exposure to sodium nitroprusside (SNP) (0.5 to 4 mmol/l), an exogenous NO donor. The effects of SNP were blocked by hemoglobin. (2) A universal caspase inhibitor, z-VAD-fmk, dose-dependently inhibited NO-induced apoptosis. VSMCs degraded Ac-DEVD-pNA rather than Ac-WHED-pNA after exposure to SNP. which suggested that the activation of caspase 3 rather than caspase I was involved in the process. Immunoblot analysis confirmed the activation of caspase-3. (3) Exposure to SNP induced the release of cytochrome c from the mitochondria to the cytosol, which was detected by immunoblot analysis of mitochondrial and cytosol fractions. (4) SNP exposure increased the ratio of Bax/Bcl-2 protein expression twofold by immunoblot analysis. Conclusions: The mechanism of NO-induced apoptosis in rat VSMCs involves an increase in the ratio of Bax/Bcl-2 gene expression, which leads to the release of cytochrome c from the mitochondria to the cytosol, finally activating caspase-3 and resultant apoptosis. (C) 2004 Elsevier B.V. All rights reserved.
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