Intracellular signal pathway of heat-induced apoptosis in human lung cancer cell lines

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This study was designed to detect heat-induced apoptosis in non-small human lung cancer cells with two different histological types and to elucidate its signaling. We analyzed the chronological changes in intracellular Ca^<2+> concentrations ([Ca^<2+>]_i) during heat-induced apoptosis in human lung cancer cell lines LK-2 (squamous cell carcinoma) and LU65A Gage cell carcinoma). In LK-2 cells, increased [Ca^<2+>]_i levels were maintained at levels between 250-350 nM 9 h after heat-shock. Treatment with BAPTA, an intracellular Ca^<2+> chelator, prior to heat-shock decreased the fequency of heat-induced apoptosis in LK-2, while thapsigargin, a selective endoplasmic reticulum Ca^-ATPase inhibitor did not change the number of apoptotic cells, regardless of the presence or absence of Ca^<2+>-supplemented medium. In LU65A cells, treatment with BAPTA or thapsigargin did not alter the apoptotic rates. Western blotting demonstrated that although expression of Bax and Bcl-2 were not changed by heat-shock, P53 expession was elevated in LK-2, but not LU65A cells. Immunohistochemistry showed that p53 was localized predominantly in the cytoplasmic of LK-2 cells, suggesting that p53 protein is not functional in LK-2. Heat-shock also elevated caspases 3, 8 and 9 activity in both cell lines. We conclude that a temporal increase in [Ca^<2+>]_i is the important initiating factor in hyperthermia-induced apoptosis in LK-2 cells and that, in these two lung cancer cell lines, apoptosis may occur through 'cross-talk' between P53-independent mitochondrial and death receptor pathways.