摘要
AIM: To investigate the role of DNA-PKcs subunits inradiosensitization by hyperthermia on hepatocellularcarcinoma HepG2 cell lines.METHODS: Hep G2 cells were exposed to hyperthermiaand irradiation. Hyperthermia was given at 45.5 ℃Cellsurvival was determined by an in vitro clonogenic assay forthe cells treated with or without hyperthermia at varioustime points. DNA DSB rejoining was measured usingasymmetric field inversion gel electrophoresis (AFIGE). TheDNA-PKcs activities were measured using DNA-PKcs enzymeassay system.RESULTS: Hyperthermia can significantly enhanceirradiation-killing cells. Thermal enhancement ratio ascalculated at 10 % survival was 2.02. The difference inradiosensitivity between two treatment modes manifestedas a difference in the α components and the almost sameβ components, which α value was considerably higher inthe cells of combined radiation and hyperthermia ascompared with irradiating cells (1.07 Gy-1 versus 0.44 Gy1). Survival fraction showed 1 logarithm increase after an8-hour interval between heat and irradiation, whereas DNA-PKcs activity did not show any recovery. The cells wereexposed to heat 5 minutes only, DNA-PKcs activity wasinhibited at the nadir, even though the exposure time waslengthened. Whereas the ability of DNA DSB rejoining wasinhibited with the increase of the length of hyperthermictime. The repair kinetics of DNA DSB rejoining aftertreatment with Wortmannin is different from thehyperthermic group due to the striking high slow rejoiningcomponent.CONCLUSION: Determination with the cell extracts andthe peptide phosphorylation assay, DNA-PKcs activity wasinactivated by heat treatment at 45.5 C, and could notrestore. Cell survival is not associated with the DNA-PKcsinactivity after heat. DNA-PKcs is not a unique factor affectingthe DNA DSB repair. This suggests that DNA-PKcs do notplay a crucial role in the enhancement of cellularradiosensitivity by hyperthermia.
AIM:To investigate the role of DNA-PKcs subunits in radiosensitization by hyperthermia on hepatocellular carcinoma HepG_2 cell lines. METHODS:Hep G2 cells were exposed to hyperthermia and irradiation.Hyperthermia was given at 45.5℃.Cell survival was determined by an in vitro clonogenic assay for the cells treated with or without hyperthermia at various time points.DNA DSB rejoining was measured using asymmetric field inversion gel electrophoresis(AFIGE).The DNA-PKcs activities were measured using DNA-PKcs enzyme assay system. RESULTS:Hyperthermia can significantly enhance irradiation-killing cells.Thermal enhancement ratio as calculated at 10 % survival was 2.02.The difference in radiosensitivity between two treatment modes manifested as a difference in the α components and the almost same β components,which α value was considerably higher in the cells of combined radiation and hyperthermia as compared with irradiating cells(1.07 Gy^(-1)versus 0.44 Gy^(- 1)).Survival fraction showed 1 logarithm increase after an 8-hour interval between heat and irradiation,whereas DNA- PKcs activity did not show any recovery.The cells were exposed to heat 5 minutes only,DNA-PKcs activity was inhibited at the nadir,even though the exposure time was lengthened.Whereas the ability of DNA DSB rejoining was inhibited with the increase of the length of hyperthermic time.The repair kinetics of DNA DSB rejoining after treatment with Wortmannin is different from the hyperthermic group due to the striking high slow rejoining component.CONCLUSION: Determination with the cell extracts and the peptide phosphorylation assay, DNA-PKcs activity was inactivated by heat treatment at 45.5 °C, and could not restore. Cell survival is not associated with the DNA-PKcs inactivity after heat. DNA-PKcs is not a unique factor affecting the DNA DSB repair. This suggests that DNA-PKcs do not play a crucial role in the enhancement of cellular radiosensitivity by hyperthermia.
