Concentration-dependent modes of cell death in Chinese hamster V79 cells after treatments with H2O2

Deterioration in the clonogenic ability of Chinese hamster V79 cells was observed after treatments with low concentrations of H2O2 (0.06-0.2 mM) for 1 h and subsequent incubation for 6-8 days, whereas no loss of ability of the cells to exclude trypan blue after treatments with H2O2 for 1 h and subsequent incubation for 6 h was observed in this concentration range. The loss of the dye-exclusion ability became observable when the cells were treated with concentrations greater than 0.9 mM H2O2 for 1 h and subsequently incubated for 6 h. Agarose gel electrophoresis of DNA extracted from cells treated with 1-5 mM H2O2 showed specific regular fragmentation of DNA, suggesting that apoptotic cell death was induced. The induction of apoptotic cell death was further confirmed by observing the protective effects of several modifiers, a protein synthesis inhibitor (cycloheximide), a Ca2+-chelator (BAPTA-AM) and an antioxidative compound (PBN), against cell survival and DNA fragmentation. From these results, it was concluded that apoptotic cell death was induced by H2O2 in Chinese hamster cells at high concentrations, where their clonogenic ability completely disappeared.