BAG-1基因沉默对非小细胞肺癌放射敏感性的影响及机制

目的探讨BAG-1基因与非小细胞肺癌放射敏感性的关系,为非小细胞肺癌个体化治疗提供理论依据。方法对A549肺腺癌细胞、H460大细胞癌及NCL-H520鳞癌细胞三株非小细胞肺癌细胞株及人胚肾细胞株HEK293行细胞放射敏感性试验,采用Western blot法检测各细胞株中BAG-1蛋白表达情况。选择其中对放射线相对较抗拒、BAG-1蛋白表达最高的A549细胞株,构建干扰载体pGCsi-BAG-1并筛选BAG-1基因沉默细胞株,用shRNA-2转染,酶标仪检测细胞生长情况,流式细胞仪检测细胞凋亡率,放射敏感性试验观察细胞对放射线的敏感性。结果 shRNA-2转染的细胞株生长受到抑制,且随时间延长抑制作用明显,而阴性对照质粒转染的A549细胞株生长无明显变化;6 Gy射线照射后24 h,shRNA-2转染细胞凋亡率为(49.6±4.23)%,未转染的A549细胞为(15.3±2.11)%,阴性对照质粒转染细胞为(14.8±3.55)%,shRNA-2转染细胞凋亡率较未转染细胞增加了3.2倍(P<0.01),而阴性对照转染细胞与未转染细胞间相比差异无统计学意义;亲本A549细胞和阴性对照转染细胞之间的放射敏感性无差异(P>0.05)。而shRNA-2转染细胞的放射敏感性明显增加,与前二者比较差异有显著性(P<0.05)。结论 BAG-1基因沉默可增强非小细胞肺癌放射敏感性,可能机制为BAG-1基因沉默后抑制细胞生长,抗凋亡作用减弱,细胞内BAG-1分布发生改变,细胞对射线更敏感。

Effect and mechanism of BAG-1 gene silencing on radio-sensitivity in non-small cell lung cancer

Objective To explore the relationship between BAG-1 and radiation sensitivity of non-small cell lung cancer,so to provide for theoretical basis for individual therapy of non-small cell lung cancer.Methods Radiation sensitivity experiment was used in A549 cells,H460 cells,NCL-H520 cells and HEK293 cells,expression of BAG-1 protein was observed in different cell lines through Western blot,then the A549 cells which relatively resistant to radiation and expressed BAG-1 protein highest were chosen,and RNA interference vector pGCsi-BAG-1 was constructed,silencing BAG-1 gene cell lines shRNA-2 were selected,growth of cells were observed using microplate reader,apoptosis was detected through flow cytometry,radiosensitivity was studied using radio sensitivity test.Results Silencing BAG-1 gene shRNA-2 inhibited cell growth,which was closely related to growth time,shRNA-SC cells had no significant effect on cell growth;24 h after the radiation of 6 Gy X-ray,apoptosis rate of shRNA-2 transfected cells was(49.6±4.23)%,the control cells was(15.3±2.11)%,the shRNA-SC cells was(14.8±3.55)%,apoptosis rate of shRNA-2 transfected cells increased 3.2 fold compared with control cells(P<0.01),while which of shRNA-SC cells had no significant difference compared with control cells;no difference of radiosensitivity was found between the parent A549 cells and shRNA-SC cells,but the radiosensitivity of shRNA-2 transfected cells was significantly higher than that of the former cells(P>0.05).Conclusions BAG-1 gene silencing can enhance radiosensitivity of non-small cell lung cancer,its mechanism maybe that BAG-1 gene silencing can inhibit cell growth,increase apoptosis,and change intracellular distribution of BAG-1,make the cells more sensitive to radiation.