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目的:探讨Tat-SmacN7融合肽对人非小细胞肺癌H460细胞辐射增敏的作用及其机制。方法:WST-1法检测Tat-SamcN7对H460细胞的毒性作用,细胞克隆实验观察Tat-SamcN7对H460细胞的辐射增敏作用,流式细胞术检测H460细胞的凋亡率,ELISA法检测H460细胞的Caspase-3、Caspase-8和Caspase-9的活性。结果:0.01~100μmol/L的Tat-SmacN7融合肽与H460细胞共培养,细胞存活率均>80%;20μmol/L的Tat-SmacN7融合肽与H460细胞共培养,对细胞的辐射增敏比为1.63;Tat-SmacN7(20μmol/L)对细胞凋亡〔(10.87±0.45)%〕有主效应,F=54.389,P<0.001;4Gy照射对细胞凋亡〔(19.83±0.75)%〕有主效应,F=641.516,P<0.001;二者有交互效应,F=65.756,P<0.001。Tat-SmacN7对细胞Caspase-3活性(1.418±0.064)有主效应,F=67.029,P<0.001;照射对细胞Caspase-3活性(1.188±0.102)有主效应,F=29.509,P=0.001;两者有交互作用,F=11.328,P=0.01。Tat-SmacN7对细胞Caspase-8活性(1.867±0.208)有主效应,F=149.705,P<0.001;照射对细胞Caspase-8活性(1.227±0.123)有主效应,F=30.418,P=0.001;两者有交互作用,F=10.663,P=0.011。Tat-SmacN7对细胞Caspase-9活性(1.672±0.075)有主效应,F=127.027,P<0.001;照射对细胞Caspase-9活性(1.279±0.141)有主效应,F=44.281,P<0.001;两者有交互作用,F=9.782,P=0.014。结论:Tat-SmacN7单独作用对非小细胞肺癌H460细胞没有明显毒性作用,但能增强H460细胞对辐射的敏感性,具有辐射增敏作用,与其能增强细胞内Caspase-3、Caspase-8和Caspase-9的活性相关。
Objective: To investigate the effect and mechanism of Tat-SmacN7 fusion peptide on radiation-sensitization of human non-small cell lung cancer H460 cells. Methods: Toxic effects of Tat-SamcN7 on H460 cells were detected by WST-1 method. Radiation sensitization effect of Tat-SamcN7 on H460 cells was observed by cell clone assay. Flow cytometry was used to detect the apoptosis rate of H460 cells. H460 cells Caspase-3, Caspase-8 and Caspase-9 activity. Results: The Tat-SmacN7 fusion peptide of 0.01 ~ 100μmol / L was co-cultured with H460 cells. The cell viability was> 80%. The Tat-SmacN7 fusion protein of 20μmol / L was co-cultured with H460 cells. (19.83 ± 0.75)%], the main effect of Tat-SmacN7 (20μmol / L) on apoptosis was (10.87 ± 0.45)%, F = 54.389, Effect, F = 641.516, P <0.001; both had an interaction effect, F = 65.756, P <0.001. Tat-SmacN7 had the main effect on the activity of Caspase-3 (1.418 ± 0.064), F = 67.029, P <0.001. The main effect of Tat-SmacN7 on the activity of Caspase-3 was 1.188 ± 0.102, F = 29.509, P = Interaction between the two, F = 11.328, P = 0.01. Tat-SmacN7 had the main effect on the activity of Caspase-8 (1.867 ± 0.208), F = 149.705, P <0.001. The main effect of Tat-SmacN7 on the activity of Caspase-8 was 1.227 ± 0.123, F = 30.418, Interaction between the two, F = 10.663, P = 0.011. Tat-SmacN7 had the main effect on the activity of Caspase-9 (1.672 ± 0.075), F = 127.027, P <0.001. The main effect of Tat-SmacN7 on the activity of Caspase-9 was 1.279 ± 0.141, F = 44.281, Interaction between the two, F = 9.782, P = 0.014. CONCLUSION: Tat-SmacN7 alone has no obvious toxic effect on H460 cells, but enhances the sensitivity of H460 cells to radiation and enhances the radiosensitivity of H460 cells. It can enhance the expression of Caspase-3, Caspase-8 and Caspase -9 activity.