| Objective and background:Renal cell carcinoma (RCC) comprises approximately 3% of all malignancies,with a median age at diagnosis of 65 years. The rate of RCC has increased by 2% per year for the past years. The reason for thisincrease is unknown. Approximately 90% of renal tumors are RCC, and 80% of these are clear cell tumors Surgical resection remains the only effective therapy for clinically localized RCC; with options including radical nephrectomy and nephron-sparing surgery.Tumor growth and metastasis dependent on angiogenesis,what is more,VEGF could enhance specially the penetration of blood vessel ,which plays an important role in the physiological and pathological process,including the embryonic development,inflammation and tumor.Therefore,VEGF was the most potent target against tumor angiogenesis.RNAi is not only a technology in study of genome function,but also an effective method to treat disease,especially,in gene therapy of the tumor. In this study,to investigate the effects and mechanism of proliferation and invadation by siRNA silencing VEGFgene expression in RCC,acquire more information about the effect of VEGF on tumor growth in vitro and find the base for its future using in clinic and test the feasibility of make VEGF gene as target gene to treat RCC.Methods:ACHN cells were transfected with small interfering RNAs (siRNAs) by liposome transfection targeting the human VEGF by chemosynthesis, the expression of VEGF in the levels of protein was detected by Western blot,the cell growth curves was determined by typan blue staining, the inhibition of proliferation of ACHN cell was studied by methyl thiazolyl tetrazolium (MTT) assay,cell apoptosis was detected by TUNEL assay, the abilities of cell mobility and adhesion were detected by wound healing assay and adhesion test,the abilities of cell invasion was evaluated using Transwell chambers.Result: The cells of siRNA1~2 group transected with a specific VEGF-siRNA showed suppression of VEGF protein expression compared with those of the blank control group and negative controI group.The cell growth curves showed the growth of ACHN cells of blank control and negative controI were suppressed significantly(p﹤0.05).The inhibition rate(IR) of siRNA1 group(18.9%,32.7%,40.3%) and siRNA2 group(27.6%,50.6%,67.8%) were significantly higher than those in the blank control group and negative controI group respectively at 24h,48h,72h(p﹤0.05).The apoptosis index of siRNA1 group(10.7%,15.9%,20.3%) and siRNA2 group(17.3%,26.2%,37.4%) were obviously higher than those in the blank control group and negative controI group respectively at 24h,48h,72h(p﹤0.05).Moreover,the effects of siRNA2 on inhibition rate,apoptosis index and down-regulating expression of VEGF protein were stronger than the group of siRNA1. After siRNA interference, the abilities of adhesion of siRNA1~2 group,especially in siRNA2 group, compared with the blank control group,was decreased(81.5±3.1% vs 40.5±2.6%, p﹤0.05;81.5±3.1% vs 22.5±2.4% ,p﹤0.05);the abilities of migration of siRNA1~2 group,compared with the blank control group,was decreased(162±9 vs 81±5,p﹤0.05;162±9 vs 38±4,p﹤0.05),especially in siRNA2 group;the abilities of invasion of siRNA1~2 group were suppressed(p﹤0.05),compared with the blank control group,especially in siRNA2 group(p﹤0.05).Conclusion: VEGF overexpresses in renal cell carcinoma ACHN lines,plays an important role in the carcinogenesis and development of renal cell carcinoma , in cell proliferation,enhance cell apoptosis,adhesion and migration.siRNAs targeting against human VEGF by chemosynthesis may specially suppress the expression of VEGF protein in renal cell carcinoma cells, inhibit cell proliferation and enhance cell apoptosis,effectively inhibit adhesion,migration,and invasion of human renal cell carcinoma cells.The RNAi targeting VEGF may become as a novel gene therapeutic strategy for VEGF overexpressed renal cell carcinoma.
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