Roles of reactive oxygen species on signal transductlon in apoptosis induced by hyperthermia

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Hyperthermia-induced apoptosis and its enhancement in the presence of a temperature-dependent free radical initiator, 2,2'-azobis(2-aminopropane) dihydrochloride(AAPH) were examined in human uterine cervical cancer cell lines, CaSki and HeLa. When both cell lines were treated with hyperthermia at 44℃ for 60min, minimal apoptosis was observed. When combined with nontoxic AAPH(50mM), significant enhancement of apoptosis was observed, where the initial rate of free radical formation was about twice as high than that at 37℃. Augmentation of the growth delay, lipid peroxidation(LPO), and increase in [Ca^<2+>]i were also observed after the combined treatment. A water-soluble vitamin E, Trolox, blocked the increase in [Ca^<2+>]i and an intracellular Ca^<2+> chelator, BAPTA-AM, prevented the DNA fragmentation induced by the combination. However, no significant change in mitochondrial membrane potential and expression of Bax and Bcl-2 was observed. A slight increase in Fas expression was observed only in CaSki cells after the combined treatment. These results indicate that hyperthermia and AAPH induce enhanced apoptosis and subsequent cell killing via an increase in LPO and the increase in [Ca^<2+>]i, but not the activation of a mitochondria-mediated apoptosis pathway in these cell lines. Since it is known that cancer cells are generally resistant to physical and chemical stress-induced apoptosis, free radical generators like AAPH appear to be a useful thermosensitizer for hyperthermic cancer therapy.In addition, enhancement of caspase-dependent apoptosis induced by hyperthermia in the presence a lipophilic free radical initiator, 2,2'-azobis(2,4-dimethylvaleronitrile)(AMVN), enhancement of hyperthermia-induced apoptosis by local anesthetics, and enhancement of hyperthermia-induced apoptosis by nonthermal effects of ultrasound in human lymphoma U937 cells were studied.