ABCE1基因通过RNase L途径抑制人大细胞肺癌H460细胞凋亡

ABCE1 gene inhibit apoptosis of human large cell lung cancer H460 cells through RNase L pathway

目的研究ABCE1基因表达对人大细胞肺癌H460细胞增殖、凋亡的影响并初步探讨其机制。方法体外培养H460细胞,分为A组(转染ABCE1-siRNA-1组)、B组(转染ABCE1-siRNA-2组)、C组(转染空质粒组)以及D组(空白对照组)。将构建好的ABCE1的siRNA(小干扰RNA)载体转染入培养的细胞中,荧光显微镜下观察转染效率,Western Blot法检测ABCE1蛋白以及RNase L蛋白的表达,RT-PCR法检测ABCE1 mRNA的表达,MTT法分析细胞增殖能力,流式细胞术检测细胞凋亡率。结果将构建好的载体成功转入了H460细胞,转染后A、B组的细胞活性降低,ABCE1的表达受到阻断(P<0.01),RNase L蛋白含量明显增加(P<0.01),细胞的增殖能力明显降低并逐渐凋亡(P<0.01)。结论 ABCE1基因可促使人大细胞肺癌H460细胞增殖并抑制细胞凋亡,其机制是阻断了2-5A/RNase L细胞通路。

Objective To study the effect of ABCE1 gene on proliferation and apoptosis of human large cell lung cancer H460 cells and its mechanism. Methods In vitro cultured H460 cells were divided into group A, group B, group C and group D （normal control group）. The cells in groups A, B and C were transfected with ABCEI-siRNA-1, ABCEI-siRNA-2 and ABCEI-siRNA-N, respectively. The siRNA vector of constructed ABCE1 was transfected into H460 cells. The efficiency of transfection was observed under a fluorescence microscope. Expressions of ABCE1 protein, RNase L protein and ABCE1 mRNA were detected by Western blot and RT-PCR, respectively. The proliferation and apoptosis of H460 cells were detected by MTT and flow cytometry, respectively. Results The siRNA vector of constructed ABCE1 was successfully transfected into the H460 cells. The viability of H460 cells in groups A and B and the expression level of ABCE1 were significantly lower whereas the expression level of RNase L protein was significantly higher after transfection than before transfection （P 〈 0.01）. The proliferation rate of H460 cells was significantly lower whereas the apoptosis rate of H460 cells was significantly higher after transfecfion than before transfection （P 〈 0.01 ）. Conclusion ABCE1 gene promotes the proliferation and inhibit the apoptosis of H460 cells by blocking the 2-5A/RNase L pathway.