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Transforming growth factor-beta (TGF-beta) has been shown to induce apoptosis in normal hepatocytes and hepatoma cells both in vivo and in vitro. However, the mechanism by which TGF-beta induces apoptosis is not clear. The antiapoptotic activity of antioxidants including N-acetyl-L-cysteine (Ac-Cys), ascorbic acid and a novel free radical scavenger, carboxyfullerene (C60) on TGF-beta-treated human hepatoma Hep3B cells was examined. Only the water-soluble hexacarboxylic acid derivative of C60 was found to prevent TGF-beta-induced apoptosis. Antiapoptotic activity of C60 correlated its ability to eliminate TGF-beta-generated reactive oxygen species (ROSs). However, C60 did not interfere with TGF-beta-activated PAI-1 promoter activity in the Hep3B cells. These results indicate that the signaling pathway of TGF-beta-induced apoptosis may be related to the generation of ROSs and may be uncoupled from the TGF-beta-activated gene promoter activity. Furthermore, the regioisomer of C60 with a C3 symmetry was more potent in protecting cells from apoptosis than that with a D3 symmetry, and the C3 isomer had stronger interactions with lipid bilayers than the D3 isomer. The spectroscopic analysis revealed that the C3 isomer had stronger interactions with artificial lipid bilayers than the D3 isomer. Therefore, our study indicates that C60 may interact with membrane to eliminate TGF-beta-induced ROSs and to prevent apoptosis occur in human hepatoma cells.
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