体外RNA干扰RhoA基因对舌癌细胞侵袭的影响

目的 应用RNA干扰技术抑制人舌癌细胞株Tca8113和SCC-4中RhoA基因的表达，探讨RhoA基因表达下调对舌癌细胞侵袭的影响。方法登录Genbank数据库确定人RhoA基因序列，针对RhoA的基因序列设计短链RNA，利用Lipofectamine2000介导法将RhoA-siRNA转染至舌癌Tca8113和SCC-4细胞。采用实时荧光定量聚合酶链反应检测转染后的舌癌细胞中RhoA mRNA的表达，蛋白质印迹法检测RhoA、半乳糖凝集素-3（galectin-3）和基质金属蛋白酶-9（MMP-9）蛋白的表达，Transwell小室检测细胞的侵袭能力。结果舌癌细胞转染RhoA-siRNA后，RhoA的基因和蛋白表达下降，galectin-3和MMP-9蛋白表达下降，细胞体外侵袭能力降低。结论RhoA-siRNA可以有效地抑制舌癌Tca8113和SCC-4细胞中RhoA的表达，通过降低galectin-3和MMP-9的表达从而降低细胞的侵袭能力。RhoA在舌癌的侵袭和转移中可能发挥重要的作用。

Effects of RhoA gene silencing by RNA interference on invasion of tongue carcinoma

ObjectiveTo study the effects of RhoA down-regulation by RNA interference on the invasion of tongue carcinoma Tca8113 and SCC-4. MethodsDetermination of the human RhoA sequence as well as the design and construction of a short specific small interfering RNAs (siRNA) were performed. The siRNA of RhoA gene was transfected into human tongue squamous cell carcinoma Tca8113 and SCC-4 cells line by Lipofectamine 2000. Quantitative real-time polymerase chain reaction was used to examine the mRNA expression levels of RhoA. Protein expressions of mRNA, galectin-3, and matrix metalloproteinase (MMP)-9 were evaluated by Western blot. Transwell invasion assay was performed to assess the invasion ability of tongue carcinoma. ResultsRhoA expressions in Tca8113 and SCC-4 cells were reduced significantly after transfection of RhoA-siRNA. Protein levels of galectin-3 and MMP-9 were also down-regulated significantly. Invasion ability was inhibited as well. ConclusionRhoA-siRNA can effectively inhibit RhoA expression in Tca8113 and SCC-4 cells. The invasion ability of tongue carcinoma cells decreased with down-regulation of the protein expressions of galectin-3 and MMP-9, indicating that RhoA-siRNA can inhibit invasion of tongue carcinoma. Results show that RhoA may play an important role in the processes of invasion and metastasis of tongue carcinoma.