| Hepatocellular carcinoma (HCC) is the most common primary liver malignancy worldwide and a leading cause of death. The aetiology of HCC seems to be multifactorial, and several events seem to be necessary for malignant transformation to occur. HCC has a poor prognosis with the number of deaths about 600,000 annually,and turns into a major global health problem. Surgical resection is a potentially curative treatment and is still the method of first choice in China and many other countries. With the evolution of liver transplantation techniques, transplantation has been successful for the treatment of early-stage HCC. Recent progress in diagnostic and treatment technologies has improved the long-term survival of patients with HCC, but the prognosis remains unfavourable. The invasiveness and metastasis of HCC seems to be the most important reason for its poor prognosis. This emphasizes the need to investigate the contribution of mechanism to tumor invasion and metastasis in HCC. Rho GTPases contribute to multiple cellular processes that could affect cancer progression, including cytoskeletal dynamics, cell cycle progression, cell migration, transcriptional regulation, and cell survival. As one of the important members of Rho family, RhoA has been implicated in virtually all stages of cancer progression. In this article, we would study the malignant behavior of HepG2 cells which are transfected by the RhoA-siRNA expression vector.Objective:The current studies were designed to construct the RhoA-siRNA expression vector, investigate the effects of RhoA-siRNA on RhoA expression in HepG2 cells and influence of RhoA-siRNA on proliferation and migration of HepG2 cells transfected with RhoA-siRNA.Methods:The HepG2 cells were cultured as usual. RhoA-siRNA DNA fragment was synthesized and cloned into the expression vector pGenesil-1. The constructed RhoA-siRNA plasmid was stably transfected into HepG2 cells by lipofectamine. The inhibitory effect of RhoA-siRNA on RhoA protein expression was detected by Western blot. The proliferation, migration, growth potentiality and cell cycle of transfected HepG2 cells and controls were evaluated by MTT, wounded healing,plate cloning formation test and FCM respectively. The data was analyzed by One-way ANOVA and Chi-Square test.Results:RhoA-siRNA can obviously down-regulate the RhoA protein expression in HepG2 cells. RhoA gene silencing significantly inhibited the proliferation of HepG2 cells. The result of flow cytometry showed that cells transfected with RhoA-siRNA were more in G0/G1 phase and lesser in S phase(P<0.05). The scratched cells of control groups were healed within 48h, but not happened in RNAi group. The clone formation rate of RhoA-RNAi group, HepG2 group and negative control group were 38.5%±3.0%, 67.0%±4.5% and 69.5%±6.0% respectively, revealing significantly different among the 3 groups (P<0.05).Conclusions:RhoA protein expression of HepG2 cells is down-regulate by RhoA-siRNA expression vector. RhoA-siRNA can effectively suppress the potentiality of proliferation and migration in HepG2 cells, which may provide a novel gene therapy on hepatocellular carcinoma.
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