Human carcinoma HSC-2 cells were more sensitive to a 24 h exposure to gallic acid than were normal human gingival fibroblasts. Acting as a prooxidant, gallic acid generated hydrogen peroxide in cell culture medium. The potency of gallic acid to HSC-2 cells was significantly lessened in the presence of scavengers of hydrogen peroxide, including catalase, pyruvate, and divalent cobalt cations, and was potentiated in the presence of the intracellular glutathione depleters, 1-chloro-2,4-dinitrobenzene, bis(2-chloroethyl)-N-nitrosourea, and DL-buthionine-[S,R]-sulfoximine. Exposure of HSC-2 cells to gallic acid decreased the level of intracellular reduced glutathione, caused lipid peroxidation, and increased the level of intracellular reactive oxygen species. Flow cytometric analyses of gallic acid-treated HSC-2 cells indicated a concentration-dependent response for the induction of apoptosis, which was reversed in the presence of divalent cobalt. Immunoblot analyses of gallic acid-treated cells showed proteolytic inactivation of poly(ADP-ribose) polymerase, an indication of apoptosis, which was also reversed in the presence of divalent cobalt cations. These studies demonstrated that the cytotoxic activities of gallic acid to HSC-2 cells were mediated through autooxidation of the polyphenol, leading to the induction of oxidative stress and thereby apoptotic cell death.
