AIM: To clarify the mechanism underlying the anti-mutagenic and anti-cancer activities of Scorpio water extract (SWE).METHODS: Human hepatoma HepG2 cells were incubated with various concentrations of SWE. After 24-h i...AIM: To clarify the mechanism underlying the anti-mutagenic and anti-cancer activities of Scorpio water extract (SWE).METHODS: Human hepatoma HepG2 cells were incubated with various concentrations of SWE. After 24-h incubation,cytotoxicity and apoptosis evaluations were determined by MTT and DNA fragmentation assay, respectively. After treatment with SWE, mitochondrial membrane potential(MMP) was determined by measuring the retention of the dye 3,3'-dihexyloxacarbocyanine (DiOC6(3)) and the protein expression including cytochrome C and poly-(ADPribose) polymerase (PARP)were measured by Western blotting. Caspase-3 and -9 enzyme activities were measured using specific fluorescence dyes such as Ac-DEVD-AFC and Ac-LEHD-AFC.RESULTS: We found that treatment with SWE induced apoptosis as confirmed by discontinuous DNA fragmentation in cultured human hepatoma HepG2 cells. Our investigation also showed that SWE-induced apoptosis of HepG2 cells were associated with intracellular events including disruption of MMP, increased translocation of cytochrome C from mitochondria to cytosol, activation of caspase-3,and PARP. Pre-treatment of N-acetyl-Asp-Glu-Val-Asp-CHO(Ac-DEVD-CHO), a caspase-3 specific inhibitor, or cydosporin A (CsA), an inhibitor of MMP disruption, completely abolished SWE-induced DNA fragmentation.CONCLUSION: These results suggest that SWE possibly causes mitochondrial damage, leading to cytochrome C release into cytosol and activation of caspases resulting in PARP cleavage and execution of apoptotic cell death in HepG2 cells. These results further suggest that Scorpio may be a valuable agent of therapeutic intervention of human hepatomas.展开更多
文摘AIM: To clarify the mechanism underlying the anti-mutagenic and anti-cancer activities of Scorpio water extract (SWE).METHODS: Human hepatoma HepG2 cells were incubated with various concentrations of SWE. After 24-h incubation,cytotoxicity and apoptosis evaluations were determined by MTT and DNA fragmentation assay, respectively. After treatment with SWE, mitochondrial membrane potential(MMP) was determined by measuring the retention of the dye 3,3'-dihexyloxacarbocyanine (DiOC6(3)) and the protein expression including cytochrome C and poly-(ADPribose) polymerase (PARP)were measured by Western blotting. Caspase-3 and -9 enzyme activities were measured using specific fluorescence dyes such as Ac-DEVD-AFC and Ac-LEHD-AFC.RESULTS: We found that treatment with SWE induced apoptosis as confirmed by discontinuous DNA fragmentation in cultured human hepatoma HepG2 cells. Our investigation also showed that SWE-induced apoptosis of HepG2 cells were associated with intracellular events including disruption of MMP, increased translocation of cytochrome C from mitochondria to cytosol, activation of caspase-3,and PARP. Pre-treatment of N-acetyl-Asp-Glu-Val-Asp-CHO(Ac-DEVD-CHO), a caspase-3 specific inhibitor, or cydosporin A (CsA), an inhibitor of MMP disruption, completely abolished SWE-induced DNA fragmentation.CONCLUSION: These results suggest that SWE possibly causes mitochondrial damage, leading to cytochrome C release into cytosol and activation of caspases resulting in PARP cleavage and execution of apoptotic cell death in HepG2 cells. These results further suggest that Scorpio may be a valuable agent of therapeutic intervention of human hepatomas.
