Experiment Study On Silencing The Expression Of MDR1 Gene In Multidrug Resistent Gastric Cancer Cell 7901/VCR By ShRNA

Objective: To Inhibit transcription and expression of MDR1 gene in multidrug resistent gastric cancer cell 7901/VCR by RNA interference. Exploring the efficiency of inhibiting the expression of gastric cancer cell 7901/VCR in nude mice by pshRNA-MDR1. Investigating the inhibitory effect of multidrug resistent gastric cancer cell 7901/VCR with pshRNA-MDR1 combined with TRAIL. Setting up foundation for the tumor gene therapy .Methods: (1) Recombinant vectors of shRNA aiming at two MDR1 gene sequences were constructed and used to transfect the cell line 7901/VCR by Lipofectamine TM2000. The change of the drug resistent index was examined by MTT method. The change of the cell cycle was detected by flow cytometry. The mRNA concentration of MDR1 gene was determined by RT-PCR and the protein expression of MDR1 gene was detected by Western blot. (2) Multidrug resistent gastric cancer cell 7901/VCR was inoculated into subcutaneous tissue of nude mice and 7901/VCR transplanted tumor model was constructed. pshRNA-MDR1 was injected into the tumor, and the volume and weight of the tumor were measured. MDR1 expression level was detected by immunohistochemical method. (3) pshRNA-MDR1 and plasmid of TRAIL were transfected into the multidrug resistent gastric cancer cell 7901/VCR simultaneously to investigate the inhibitory effect of the tumor.Results: (1) The RNAi vectors were successfully constructed. After transfection, cell IC50 and drug resistent index was greatly reduced, G1 phase cells and apoptosis cell were increased . The expression of MDR1 mRNA and P-gp were all reduced. (2) Compared with control group, tumor sizes in group of injecting of pshRNA-MDR1 were decreased significantly. The expression level of MDR1 was also reduced obviously. (3) The pshRNA-MDR1 could significantly induce the apoptosis of the tumor with combination of plasmid TRAIL.Conclusion: In vitro experiments showed that pshRNA-MDR1 could effectively inhibit the expression of MDR1 of multidrug resistent gastric cancer cell 7901/VCR, This finding could be a foundation for tumor gene theraphy. In animal experiment, pshRNA-MDR1 inhibited the growth of tumor in nude mice and reduced the expression of MDR1 in transplanted tumor. The apoptosis of tumor cells were greatly increased after treatment of pshRNA-MDR1 combined with plasmid of TRAIL.