ABSTRACT
<p><b>OBJECTIVE</b>To study the effects of RhoA down-regulation by RNA interference on the invasion of tongue carcinoma Tca8113 and SCC-4.</p><p><b>METHODS</b>Determination of the human RhoA sequence as well as the design and constructionof a short specific small interfering RNAs (siRNA) were performed. The siRNA of RhoA gene was transfected into humantongue squamous cell carcinoma Tca8113 and SCC-4 cells line by Lipofectamine 2000. Quantitative real-time polymerasechain reaction was used to examine the mRNA expressionlevels of RhoA. Protein expressions of mRNA, galectin-3,and matrix metalloproteinase (MMP)-9 were evaluated byWestern blot. Transwell invasion assay was performed toassess the invasion ability of tongue carcinoma.</p><p><b>RESULTS</b>RhoA expressions in Tca8113 and SCC-4 cells were reducedsignificantly after transfection of RhoA-siRNA. Protein levels f galectin-3 and MVP-9 were also down-regulated significantly. Invasion ability was inhibited as well.</p><p><b>CONCLUSION</b>RhoA-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.</p>
Subject(s)
Humans , Carcinoma, Squamous Cell , Genetics , Metabolism , Pathology , Cell Line, Tumor , Down-Regulation , Galectin 3 , Metabolism , Gene Silencing , Matrix Metalloproteinase 9 , Metabolism , RNA Interference , RNA, Messenger , Metabolism , RNA, Small Interfering , Genetics , Tongue Neoplasms , Genetics , Metabolism , Pathology , TransfectionABSTRACT
<p><b>OBJECTIVE</b>This study investigated the effect of RhoA silencing through RNA interference on proliferation and growth of tongue cancer cells, as well as explored the possible mechanisms of this effect.</p><p><b>METHODS</b>SSC-4 tongue cancer cells were cultured in vitro and then transfected with small interfering RNA to knock down RhoA expression. The tested cells were divided into three groups: experimental group (experimental group 1: transfected with RhoA-siRNA-1; experi-mental group 2: transfected with RhoA-siRNA-2), negative control group (transfected by random sequence NC-siRNA), and blank control group (transfected with Lipofectamine). The expression levels of RhoA mRNA were respectively measured by quantitative real-time polymerase chain reaction and western blot assay. Moreover, the expression levels of cyclin D1, p21, and p27 and RhoA protein were evaluated by Western blot assay. Proliferation and growth potentiality were analyzed through evaluation of doubling times and methyl thiazolyl tetrazolium assessment.</p><p><b>RESULTS</b>The expression levels of RhoA gene and protein of experimental groups significantly decreased following siRNA transfection compared with those in the negative and blank control groups. The expression of cyclin D1 decreased significantly and that of p21 and p27 increased significantly. The doubling time was extended and the growth potentiality decreased.</p><p><b>CONCLUSIONS</b>The results indicated that RhoA silencing can inhibit proliferation of tongue cancer cells, whereas RhoA affects cell proliferation by regulating the cell cycle pathway. Thus, RhoA is a potential target in gene therapy for tongue cancer.</p>
Subject(s)
Humans , Cell Line, Tumor , Cell Proliferation , Cyclin D1 , Gene Silencing , Neoplasms, Squamous Cell , RNA, Messenger , RNA, Small Interfering , Real-Time Polymerase Chain Reaction , Tongue Neoplasms , Transfection , rhoA GTP-Binding ProteinABSTRACT
0.05); there was a significant difference between the groups, in which the thickness of tumor was less than 5 mm, and the other one, in which the thickness was greater than 5 mm (P