Grape seed extract (GSE), rich in the
bioflavonoids commonly known as
procyanidins, is one of the most commonly consumed dietary supplements in the United States because of its several health benefits. Epidemiological studies show that many
prostate cancer (PCA) patients use herbal extracts as dietary supplements in addition to their
prescription drugs. Accordingly, in recent years, we have focused our attention on assessing the efficacy of GSE against PCA. Our studies showed that GSE inhibits growth and induces apoptotic death of human PCA cells in culture and in nude mice. Here, we performed detailed studies to define the molecular mechanism of GSE-induced apoptosis in advanced human PCA DU145 cells. GSE treatment of cells at various doses (50-200 micro g/ml) for 12-72 h resulted in a moderate to strong apoptotic death in a dose- and time-dependent manner. In the studies assessing the apoptotic-signaling pathway induced by GSE, we observed an increase in cleaved fragments of
caspases 3, 7 and 9 as well as PARP in GSE-treated cells after 48 and 72 h of treatment. Pre-treatment of cells with general
caspases inhibitor,
z-Val-Ala-Asp(OMe)-FMK or
caspase 3-like
proteases inhibitor [
z-Asp(OMe)-Glu(OMe)-Val-Asp(OMe)-FMK], almost completely (approximately 90%) inhibited the GSE-induced apoptotic cell death. In a later case, GSE-induced
caspase-3 activity was completely inhibited. Selective
caspase 9 inhibitor [z-Leu-Glu(OMe)-His-Asp(OMe)-FMK] showed only partial inhibition of GSE-induced apoptosis whereas GSE-induced
protease activity of
caspase 9 was completely inhibited. Upstream of
caspase cascade, GSE showed disappearance of mitochondrial membrane potential and an increase in
cytochrome c release in cytosol. Together, these results suggest that GSE possibly causes mitochondrial damage leading to
cytochrome c release in cytosol and activation of
caspases resulting in PARP cleavage and execution of apoptotic death of human PCA DU145 cells. Furthermore, GSE-caused
caspase 3-mediated apoptosis also involves other pathway(s) including
caspase 9 activation.