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目的:研究mi R-718和早期生长反应蛋白3(EGR3)在肝癌细胞系Hep G2及SMMC-7721中对细胞恶性行为的作用。创新点:首次在肝癌细胞系Hep G2和SMMC-7721中发现miR-718作为抑癌基因及EGR3作为促癌基因起到调控肿瘤恶性行为的作用。方法:采用聚合酶链反应(PCR)方法构建mi R-718和EGR3的过表达及敲降质粒,并采用实时定量PCR(q RT-PCR)方法验证质粒有效性;采用MTT法和克隆形成实验检测肝癌细胞系HepG 2和SMMC-7721的生长增殖能力;采用Transwell迁移侵袭实验检测肝癌细胞系Hep G2和SMMC-7721的迁移和侵袭能力;采用增强型绿色荧光蛋白(EGFP)荧光报告载体实验验证miR-718的靶基因;采用qR T-PCR和蛋白质印迹(Western blot)检测相关基因表达水平的变化。结论:mi R-718在肝癌细胞系HepG 2和SMMC-7721中起到抑癌基因的作用;EGR3在肝癌细胞系Hep G2和SMMC-7721中起到促癌基因的作用;miR-718是通过靶定EGR3 m RNA 3’UTR下调EGR3的表达起到抑癌基因的作用。
AIM: To investigate the effect of mi R-718 and early growth responsive protein 3 (EGR3) on the malignant behavior of human hepatocellular carcinoma cell lines Hep G2 and SMMC-7721. Innovative point: For the first time in the liver cancer cell lines Hep G2 and SMMC-7721 found miR-718 as a tumor suppressor gene and EGR3 as a tumor-promoting gene play a regulatory role in tumor malignancy. Methods: The overexpression and knockdown plasmids of mi R-718 and EGR3 were constructed by polymerase chain reaction (PCR), and the plasmid validation was verified by real-time quantitative PCR (q RT-PCR). MTT assay and clonogenic assay The growth and proliferation of HepG 2 and SMMC-7721 cells were detected by flow cytometry. The migration and invasion ability of Hep G2 and SMMC-7721 cells were detected by Transwell invasion and invasion assay. The enhanced green fluorescent protein (EGFP) The target gene of miR-718 was detected by qRT-PCR and Western blot. Conclusions: mi R-718 plays a role of tumor suppressor gene in HepG 2 and SMMC-7721 cell lines. EGR3 plays a role of oncogene in hepatoma cell lines Hep G2 and SMMC-7721. Targeting EGR3 m RNA 3’UTR down-regulates the expression of EGR3 as a tumor suppressor gene.