Effects of combined treatment with 40°C hyperthermia and bleomycin on the accumulation of heat shock protein in murine L cells

Our research group has reported the enhanced cytotoxicity of combined treatment with bleomycin (BLM) and low hyperthermia at 40°C, using murine L cells, and suggested that post-heating could inhibit BLM-induced sublethal damage repair. For further understanding of the involved mechanisms, we subsequently investigated the kinetics of the cellular accumulations of inducible 72-kDa heat shock protein (hsp72) after 40°C hyperthermia and/or BLM treatment using the same cell line. Western blot analysis showed significantly enhanced accumulation of hsp72 after a low hyperthermia at 40°C for 40, 105 or 180 min, and no significant enhancement of it after exposure to 10 μg/ml BLM at 37°C for either 40 or 105 min. When the cells were heated in the presence of BLM, the accumulations of hsp72 were markedly suppressed, with the maxima of hsp72 accumulation decreasing to 38% and 63% of those induced by hyperthermia alone for 40 or 105 min, respectively. On the other hand, sequential treatment with hyperthermia either before or after BLM treatment did not show significant alteration of the heat-induced accumulations of hsp72. It was demonstrated that BLM was necessary during heating to effectively suppress the heat-induced accumulation of hsp72. This study indicates that the suppression of heat-induced accumulation of hsp72 by BLM may partially contribute to enhance cytotoxicity of the simultaneous treatment of 40°C hyperthermia and BLM.