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目的:研究发现泛素连接酶SIAH2在多种肿瘤中高表达,为了研究SIAH2和肝癌发病之间的相关性,利用RNA干扰技术观察泛素连接酶SIAH2基因表达抑制对人肝癌细胞株Hep G2细胞周期和凋亡的影响。方法:采用免疫组化检测50例HCC和相应癌旁组织中SIAH2蛋白的表达。构建特异性靶向SIAH2的重组干扰质粒转染人肝癌细胞Hep G2,采用RT-PCR和Western blotting技术检测重组质粒对SIAH2 mRNA和蛋白表达的影响,MTS比色法测定重组质粒转染对细胞体外增殖能力的影响,流式细胞术检测重组质粒转染后细胞周期和凋亡的变化。Transwell实验检测细胞体外侵袭能力变化。结果:SIAH2在肝癌组织中表达明显升高。靶向SIAH2的干扰RNA能显著抑制SIAH2 mRNA和蛋白的表达水平;与Hep G2-neo组、Hep G2-NC组和未转染Hep G2组相比,SIAH2干扰组(Hep G2-S3)细胞增殖速度明显减慢,细胞明显阻滞于G1期,细胞凋亡率明显升高,体外侵袭能力减弱。结论:SIAH2通过促进肝癌细胞增殖和侵袭在肝癌发病中发挥癌基因作用,上述研究为以SIAH2为靶点的肝癌基因治疗提供实验依据。
To study the correlation between SIAH2 and the pathogenesis of hepatocellular carcinoma (HCC). To study the effect of SIAH2 gene expression on the proliferation of human hepatocellular carcinoma cell line Hep G2 And the impact of apoptosis. Methods: Immunohistochemistry was used to detect the expression of SIAH2 protein in 50 cases of HCC and corresponding paracancerous tissues. The recombinant plasmid targeting SIAH2 was constructed and transfected into Hep G2 cells. The expression of SIAH2 mRNA and protein was detected by RT-PCR and Western blotting. The expression of SIAH2 mRNA and protein was detected by MTT assay. Proliferating ability, the changes of cell cycle and apoptosis after transfected with recombinant plasmid were detected by flow cytometry. Transwell assay was used to detect the in vitro invasiveness of cells. Results: The expression of SIAH2 in hepatocellular carcinoma was significantly increased. Compared with Hep G2-neo group, Hep G2-NC group and untransfected Hep G2 group, SIAH2 interference group (Hep G2-S3) cell proliferation was significantly inhibited by interfering RNA targeting SIAH2 The speed slowed down significantly, the cells were significantly arrested in G1 phase, the apoptosis rate was significantly increased, in vitro invasive ability weakened. Conclusion: SIAH2 can play an oncogene role in the pathogenesis of hepatocellular carcinoma by promoting the proliferation and invasion of hepatocellular carcinoma cells. These studies provide experimental evidence for gene therapy of hepatocellular carcinoma targeting SIAH2.