siRNA沉默Skp2基因对肝癌HepG_2细胞生物学行为影响及机制

Effects of RNAi-silenced Skp2 Gene on the Biological Behavior of Hepatocellular Carcinoma HepG_2 Cells and Its Mechanism

目的观察Skp2在肝癌组织及细胞中表达,及其对HepG_2细胞增殖、凋亡及侵袭的影响。方法利用siRNA沉默HepG_2中Skp2的表达。采用qRT-PCR检测肝癌组织及细胞中Skp2 mRNA的表达,CCK-8、流式细胞术和Transwell小室测定干扰Skp2表达后对细胞的增殖、凋亡和侵袭力的影响,ELISA法测定MMP2和MMP9水平,Western blot法检测caspase-3、P27的表达。结果肝癌组织中Skp2的表达水平高于癌旁组织(P <0. 05)。与NC-Skp2组比较,siRNA-Skp2组Skp2 mRNA水平显著降低,细胞的增殖能力受抑制,凋亡能力增强,细胞侵袭力下调(P <0. 05)。此外,与NC-Skp2组比较,siRNA-Skp2组HepG_2细胞分泌MMP2和MMP9能力明显降低,caspase-3、P27蛋白水平明显增加(P <0. 05)。结论 Skp2的表达与细胞HepG_2的增殖、凋亡和侵袭密切有关,siRNA可以降低HepG_2细胞的增殖和侵袭,诱导细胞凋亡。

Objective To observe Skp2 expression levels in hepatocellular carcinoma patients,and its effect on cell proliferation,apoptosis and invasiveness of HepG2 cells.Methods Skp2 expression was silenced by small interfering RNA(siRNA)in HepG2 cells.The relative expression of Skp2 mRNA in hepatocarcinoma tissues and HepG2 cells were detected by real-time PCR.Cell proliferation was determined by CCK-8 method.Cell apoptosis was tested by flow cytometry.Invasiveness was tested by transwell assay.MMP2 and MMP9 levels were measured by ELISA,and the protein expression of caspase-3 and P27 was detected by Western blot analysis.Results Compared with the adjacent tumor tissue,Skp2 mRNA levels were significantly higher in hepatocellular carcinoma(P<0.05).Compared with the NC-Skp2 group,Skp2 mRNA levels was significantly decreased in the siRNA-Skp2 group,the proliferation ability and invasiveness of the cells was inhibited,and the apoptosis was enhanced(P<0.05).In addition,compared with NC-Skp2 group,the levels of MMP2 and MMP9 were significantly decreased in siRNA-Skp2 group,and levels of caspase-3 and P27 protein were significantly increased(P<0.05).Conclusion This study suggested that The Skp2 expression is closely related to cell proliferation,apoptosis and invasiveness of HepG2 cells.siRNA can decrease the proliferation and invasion of HepG2 cells and induce apoptosis.