Study On Transduction By Adenovirus-mediated RNA Interfering The Expression Of ERCC1 Gene Reversing Drug-resistance Of Ovarian Carcinoma Cells In Vitro

ObjectiveBased on transduction by adenovirus-mediated RNA interfering the expression of ERCC1 gene to observe whether the recombinate adenovirus (Ad-hTERT-ERCC1-siRNA)could enhance DDP's chemical sensitivity on ovary carcinoma cell line SKOV3 which is sensitive to DDP,and whether it could reverse the drug-resistance on ovary carcinoma cell line SKOV3/DDP which is resistant to DDP in some how,and if these effects have any relations with it's dosages,and to give the experimental basis for reversing the DDP-resistance on ovarian carcinoma.The recombinate adenovirus(Ad-hTERr-ERCC1-siRNA)and DDP were used only as comparison in vitro.Therefore,an effective therapeutic strategy for overcoming chemotherapy-resistance of ovarian cancer will be initiated.MethodsPart 1'SKOV3 cells were infected by direct infection method and with recombinant adenovirus infection,Taifan blue dye,MTT,Hoechst staining and flow cytometer were used to study recombinant adenovirus' inhibitory effects and chemical sensitivity-enhancing effects on ovary carcinoma cell line SKOV3 which is sensitive to DDP.Part 2:SKOV3/DDP cells were infected by direct infection method and with recombinant adenovirus infection,Taifan blue dye,MTT,Hoechst staining and flow cytometer were used to study recombinant adenovirus' DDP-resistance reversing effects on ovary carcinoma cell line SKOV3/DDP which is resistant to DDP.ResultsPart 11.DDP's IC₅₀is 7.76μg/ml on SKOV3,Ad-hTERT-ERCC1-siRNA has somewhat inhibitory effect on SKOV3;2.Drug sensitivity test showed that:apoptosis of SKOV3 cells increased as the infectious viral titer and the concentration of DDP enhanced,,cell survival rate significantly decreased,dose dependence existed;3.By Hoechst staining,as the infectious viral titer and the concentration of DDP enhanced,the cell apoptosis ascend clearly on SKOV3;4.Infection of adenovirus with ERCC1-siRNA and therapy of cisplatin could increase S stage,cell apoptosis,and G₁ stage predominantly decreased.Part 21.DDP's IC₅₀is 13.92μg/ml on SKOV3/DDP,drug fast is 1.79, Ad-hTERT-ERCC1-siRNA has somewhat inhibitory effect on SKOV3/DDP;2.Drug sensitivity test showed that:apoptosis of SKOV3/DDP cells increased as the infectious viral titer and the concentration of DDP enhanced,cell survival rate significantly decreased,dose dependence existed;3.By Hoechst staining,as the infectious viral titer and the concentration of DDP enhanced,the cell apoptosis ascend notably on SKOV3/DDP;4.Infection of adenovirus with ERCC1-siRNA and therapy of cisplatin could increase S stage,cell apoptosis,and G₀/G₁ stage predominantly increased.Conclusions1.Single infection of Ad-hTERT-ERCC1-siRNA has inhibitory effect to SKOV3 and SKOV3/DDP;2.Apoptosis of SKOV3 and SKOV3/DDP cells increased as the infectious viral titer and the concentration of DDP enhanced,cell survival rate significantly decreased, dose dependence existed;3.Ad-hTERT-ERCC1-siRNA could effectively make target gene expression in silence,change cell's cycle and increase the apoptosis-inducing rate of DDP,so that the chemical sensitivity of DDP to SKOV3 is enhanced,and the resistance of DDP to SKOV3/DDP is reversed.