Ku80 表达抑制联合热疗对前列腺癌细胞放射敏感性影响的研究

Effects of inhibition of the expression of Ku80on radio sensitivity of prostate cancer cells

目的探讨抑制PC3细胞中Ku80表达水平并结合热疗对前列腺癌细胞的辐射敏感性的影响。方法构建调节Ku80表达的siRNA载体,筛选转染后的PC3-Ku80(Ku80组)、PC3-β-actin(Vector组)、PC3-Ku80-siRNA(RNAi组)、PC3-β-actin-siRNA(Scramble组)等4种稳定细胞株。在不同的条件下分别用γ射线照射后行Annexin V-FIC双染色及DAPI、TUNNEL染色检测细胞周期与凋亡情况,另取一组PC3-Ku80-siRNA细胞经42℃水浴30min,转37℃细胞培养12h再行射线照射,通过细胞周期与凋亡检测等了解不同组间放射效果的差异。结果 Western blot和RT-PCR分析表明转染后可明显调节PC3细胞中Ku80的表达,稳定转染的对数生长期细胞经γ射线照射,细胞周期与凋亡检测结果显示随着Ku80表达的降低,细胞对放射治疗更趋敏感,5组细胞随γ射线照射剂量的增加,细胞存活率下降,其中RNAi组的细胞存活率均低于对照组和Ku80组,5组比较差异有统计学意义(F=0.953,P<0.001)。PC3-Ku80-siRNA细胞经热休克处理后再行射线照射,其存活率明显低于未经热休克处理的细胞株。结论抑制前列腺癌细胞中Ku80蛋白表达水平,结合热休克,可明显提高肿瘤组织对放射治疗的敏感性,这可能是一个较为理想的综合治疗前列腺癌的方法。

Objective To investigate the effect of the expression level of Ku80 be adjusted by siR- NA Vector and heat shock condition on the radio sensitivity in prostate cancer cell. Methods To regulate endogenous Ku80 protein expression in prostate cancer cells by siRNA technology. Screening four kinds of stable cell lines, they are PC3-Ku80 （Ku80 group）, PC3-13 actin （Vector group）, PC3- Ku80-siRNA （RNAi group）, PC3-13-actin-siRNA （Scramble group） after transfection. Western blot and RT-PCR method were taken to detect the Ku80 protein and mRNA expression levels. The 4 groups cells in logarithmic growth phase were performed by Annexin V-FIC double staining and DA- PI, TUNNEL staining to assess the situation of the cell cycle and apoptosis. A group of PC3-Ku80- siRNA cells were treated at 42 ~C for 30 min and cultured at 37 ~C for 12 h before irradiation. Re- sults Western blot and RT-PCR analysis showed that the expression of Ku80 in PC3 cells were sig- nificantly regulated by the siRNA Vectors. Five stable cell lines were observed after y rays detec- tion. More sensitive to radiotherapy accompany with Ku80 expression decreasing. Under condition of heat s^ock, the significant cumulative magnifying effect were rendered in cells. Conclusions By inhibiting the Ku80 protein expression level in prostate cancer cells, combined with the heat shock, sensitivity of tumor tissue to radiation therapy ean he significantly improved, and this may he an ide- al method for the comprehensive treatment of prostate cancer.