Proliferation Inbitory Effect Of Shrna-Mediated Silence Of Plce Gene Expression On Renal Carinoma Cells

PART ONEObjective: To investigate the proliferation inhibitory effect of short hairpin RNA (shRNA)-mediated silence of PLCεgene expression on renal carcinoma 786-0 cells.Methods: Liposome was employed to mediate the transfection of both the recombinant plasmid (pGenesil-PLCε) and the control plasmid(pGenesil-NP)into the 786-0 cells. RT-PCR was used to detect the PLCεmRNA expression level after being transfected ; MTT assay was conducted to detect the proliferation inhibitory rate after being transfected with the recombinant plasmid; Flow cytometry(FCM) was performed to analyze the change of cell cycle;Results: The expression of PLCεmRNA was significantly inhibited by recombinant plasmid transfection, and the inhibitory rate reached 69.7%; The proliferation inhibitory rates were respectively 21.2%,31.6% and 32.7% after being transfected for 24h,48h and 72h; The distribution of its cell cycle changed as showed by the results from FCM, in which the number of cells in the G0/G1 phase increased, and that in both the S phase and G2/M phase decreased, cell cycle was blocked at the G0/G1 phase, and the subdiploid"apoptotic peak"appeared at the same time.Conclusions: The proliferation of the renal carcinoma 786-0 cell line was remarkably inhibited by RNA interference-mediated blockage of PLCεgene expression, the distribution of cell cycle changed and the cell proliferation decreased ,which implied the PLCεgene as the molecular target of gene therapy for renal carcinoma and established theory foundation of biological therapy to tumor aiming at PLCεgene .PART TWOObjective: To explore the mechanism of the proliferation inhibitory effect of shRNA-mediated silence of PLCεgene expression on renal carcinoma 786-0 cells.Methods: RT-PCR and immunocytochemistry were employed to analyze the expression status of both P27 and Ki67.Results: The expression of P27 was up-regulated and that of Ki67 was down-regulated as indicated by the results from both RT-PCR and immunocytochemistry. Conclusions: The proliferation of the renal carcinoma 786-0 cell line was inhibited by RNA interference-mediated blockage of PLCεgene expression, and the mechanism of which may partially be ascribed to its induction on P27 up-regulation which cell cycle was blocked at G1 phase and Ki67 down-regulation which the cell proliferation decreased. The results of our research enrich the cognition of pathogenesis to renal cell carcinoma and provide potent theory base to curing tumor by interfering molecular target of renal cell carcinoma.