Nan Zhou1, Zeng-Peng Chi1, Wen-Jian Li2, Kai Zhao3, Shao-Ru Wang2, Qi-Min Wang4, Lei Tong4, Zong-Xuan He5, Hong-Yu Han4, Ying Wang6, Zheng-Gang Chen4. 1. College of Stomatology, Weifang Medical University, Weifang 261021, China. 2. School of Stomatology, Dalian Medical University, Dalian 116044, China. 3. School of Stomatology, Qingdao University, Qingdao 266003, China. 4. Dept. of Stomatology, Qingdao Municipal Hospital, Qingdao University, Qingdao 266071, China. 5. Dept. of Oral and Maxillafacial Surgery, The Affiliated Hospital of Qingdao University, Qingdao 266005, China. 6. Dept. of Stomatology, Fourth People's Hospital of Jinan, Jinan 250031, China.
Abstract
OBJECTIVES: The effect of isoprenylcysteine carboxymethyltransferase (ICMT) silencing on the migration and invasion of tongue squamous cell carcinoma was investigated by constructing the small interfering RNA (siRNA) of ICMT. METHODS: Through liposomal transfection, siRNA was transfected into human tongue squamous cell carcinoma CAL-27 and SCC-4 cells (ICMT-siRNA group) with a negative control group (transfected with NC-siRNA) and a blank control group (transfected with a transfection reagent but not with siRNA). Quantitative real-time polymerase chain reaction was performed to analyze the mRNA expression of ICMT and RhoA in each group of cells after transfection and to measure the silencing efficiency. Western blot was applied to examine the expression levels of ICMT, total RhoA, membrane RhoA, ROCK1, matrix metalloproteinase (MMP)-2, and MMP-9 proteins in each group. The migration and invasion abilities were evaluated via wound healing and Transwell motility assays. RESULTS: After CAL-27 and SCC-4 cells were transfected with ICMT-siRNA, the expression levels of ICMT genes and proteins decreased significantly in the experimental group compared with those in the negative and blank control groups (P<0.05). The mRNA and total protein expression levels of RhoA in the two groups were not significantly different (P>0.05). The expression levels of RhoA membrane protein, ROCK1, MMP-2, and MMP-9 decreased (P<0.05). The migration and invasion abilities were inhibited (P<0.05). CONCLUSIONS: The migration and invasion abilities of CAL-27 and SCC-4 cells were reduced significantly after the transfection of ICMT-siRNA, and the involved mechanism might be related to the RhoA-ROCK signaling pathway.
OBJECTIVES: The effect of isoprenylcysteine carboxymethyltransferase (ICMT) silencing on the migration and invasion of tongue squamous cell carcinoma was investigated by constructing the small interfering RNA (siRNA) of ICMT. METHODS: Through liposomal transfection, siRNA was transfected into humantongue squamous cell carcinoma CAL-27 and SCC-4 cells (ICMT-siRNA group) with a negative control group (transfected with NC-siRNA) and a blank control group (transfected with a transfection reagent but not with siRNA). Quantitative real-time polymerase chain reaction was performed to analyze the mRNA expression of ICMT and RhoA in each group of cells after transfection and to measure the silencing efficiency. Western blot was applied to examine the expression levels of ICMT, total RhoA, membrane RhoA, ROCK1, matrix metalloproteinase (MMP)-2, and MMP-9 proteins in each group. The migration and invasion abilities were evaluated via wound healing and Transwell motility assays. RESULTS: After CAL-27 and SCC-4 cells were transfected with ICMT-siRNA, the expression levels of ICMT genes and proteins decreased significantly in the experimental group compared with those in the negative and blank control groups (P<0.05). The mRNA and total protein expression levels of RhoA in the two groups were not significantly different (P>0.05). The expression levels of RhoA membrane protein, ROCK1, MMP-2, and MMP-9 decreased (P<0.05). The migration and invasion abilities were inhibited (P<0.05). CONCLUSIONS: The migration and invasion abilities of CAL-27 and SCC-4 cells were reduced significantly after the transfection of ICMT-siRNA, and the involved mechanism might be related to the RhoA-ROCK signaling pathway.