Tat-SmacN7诱导肿瘤细胞辐射敏感性的机理研究

Mechanisms of radiosensitization induced by Tat-SmacN7 in tumor cells

探讨Tat-SmacN7诱导食管癌细胞株EC109和非小细胞肺癌细胞株NCL-H460辐射敏感性的机制。采用体外培养EC109细胞和NCL-H460细胞,实验分为对照组、单纯给药组、单纯照射组和给药联合照射组,用RT-PCR检测CAPS8、CASP9和CASP3的转录水平,Fluorogenic caspase assay试剂盒检测Caspase-3、-8、和-9的活性,ELISA方法检测Caspase活性。结果显示,两种肿瘤细胞在使用Tat-SmacN7联合Caspase的抑制剂z-VAD-fmk后,存活曲线较不使用抑制剂组均有明显上移;Caspase-3、-8、和-9的活性在Tat-SmacN7组明显提高,提示Tat-SmacN7可通过细胞凋亡线粒体途径发挥辐射增敏作用,有望成为一种新的辐射增敏剂。

To explore the mechanisms of Tat-SmacN7 in radiosensitization by the activation of caspases and induction of apoptosis pathway in EC109 and NCL-H460 cancer cells. The cells were divided into four groups： the control group, the Tat-SmacN7 group, the radiation group, the combined Tat-SmacN7 with radiation group. The transcriptional level of caspase-3, -8 and -9 were fitted by RNA extraction, cDNA synthesis and quantitative real-time PCR. The activity of caspase-3, -8 and-9, was analyzed using a fluorogenic caspase assay with Ac-DEVD-AFC, Ac-IETD-AFC, Ac-LEHD-AFC as substrates, respectively, z-VAD-fmk treatment completely blocked apoptotic combination of Tat-SmacN7 and radiation in both EC109 and H460 cells. The activity and mRNA expression of caspase-3, -8 and -9 were found increased significantly in Tat-SmacN7 group. Tat-SmacN7 increases tumor radiosensitivity through mitochondria pathway of apoptosis, which is expected to be a new radiosensitization agent in the future.