RESUMO
PURPOSE: To examine whether a synthetic bile acid derivatives (HS-1200) sensitizes the radiation-induced apoptosis in human breast cancer cells (MCF-7) and to investigate the underlying mechanism. MATERIALS AND MEHTODS: Human breast cancer cells (MCF-7) in exponential growth phase were treated with HS-1200 for 24 hours at 37degrees C with 5% CO2 in air atmosphere. After removal of HS-1200, cells were irradiated with 2~8 Gy X-ray, and then cultured in drug-free media for 24-96 hours. The effect of radiation on the clonogenicity of MCF-7 cells was determined with clonogenic cell survival assay with 16muM of HS-1200. The induction of apoptosis was determined using agarose gel electrophoresis and Hoechst staining. The expression level of apoptosis-related molecules, such as PARP, Bax, Bcl-2, Bak and AIF, were assayed by Western blotting analysis with 40muM of HS-1200 combined with 8 Gy irradiation. To examine the cellular location of cytochrome c, bax and AIF immunofluorescent stainings were undertaken RESULTS: Treatment of MCF-7 cells with 40muM of HS-1200 combined with 8 Gy irradiation showed several changes associated with enhanced apoptosis by agarose gel electrophoresis and Hoechst staining. HS-1200 combined with 8 Gy irradiation treatment also enhanced production of PARP cleavage products and increased Bax/Bcl-2 ratio by Western blotting. Loss of mitochondrial membrane potential (delta psi m) and increased cytochrome c staining indicated that cytochrome c had been released from the mitochondria in HS-1200 treated cells. CONCLUSION: We demonstrated that combination treatment with a synthetic chenodeoxycholic acid derivative HS-1200 and irradiation enhanced radiation-induced apoptosis of human breast cancer cells (MCF-7). We suggest that the increased Bax/Bcl-2 ratio in HS-1200 co-treatment group underlies the increased radiosensitivity of MCF-7 cells. Further futures studies are remained elusive.