In Vitro Inhibition Of Targeting The FLIP Gene By SiRNA Interference On PC3Prostate Cancer Cell

Background and objectives:Fas-associated death domain-like interleukin-1β-converting enzymeinhibitory protein (cellular-FLICE inhibitory protein, c-FLIP), referred to as FLIP,found its overexpression in various types of tumor cells, inhibition of Fas throughcompetitive, TNFR-1, DR4and TRAILR apoptotic pathway mediated by deathreceptor-like in the caspase-8(cysteine aspartate proteases8) apoptotic proteins canprevent apoptosis process, to promote excessive growth of tumor cells. FLIPoverexpression in PCa, inhibiting apoptosis in PCa cells, maintaining excessivegrowth of PCa cells, may be an effective target for gene therapy for PCa. This studywas designed siRNA targeting FLIP silencing gene expression in PC3cells and toexplore its feasibility as a target for gene therapy of PCa.Methods:The cultured PC3cells were divided into control group (plus media only),negative control group (NC-siRNA, liposomes) in the experimental group(FLIPL-siRNA, liposomes). QPCR detection of PC3cells FLIPL mRNA, caspase-8mRNA expression levels; western blot detection of PC3cells FLIP protein levels;MTT assay PC3cell growth inhibition rate, flow cytometry checking PC3cellapoptosis rate and Transwell model checking PC3cell invasiveness.Results:1)QPCR detection FLIPLmRNA expression levels after FLIPL-siRNAtransfected PC3cells48h. Relative to a blank PC3group and negative control PC3group, FLIPLmRNA expression levels in the experimental group PC3cells wassignificantly decreased (P <0.001); siRNA-1, siRNA-2, siRNA-3, and the inhibitionrate siRNA-4, respectively(69.9士2.2)%、(65.7士2.3)%、(75.4士1.6)%and (70.3士3.2)%,where siRNA-3strongest inhibition (P<0.05); while the blank PC3groupand negative control PC3group no difference between FLIPLmRNA expressionlevels of statistical significance (P>0.05). siRNA-3PC3group of caspase-8mRNAexpression levels were significantly higher than the blank PC3group and negative control PC3group (P <0.001), mRNA levels of the difference between the blank PC3group and negative control PC3group was not statistically significant (P>0.05).1)FLIPL-siRNA transfected PC3cells48h later,Western blot assay FLIPLproteinlevels. Relative to a blank PC3group, siRNA-3PC3group FLIPLprotein levelsdecreased significantly (P <0.01).3) MTT, flow cytometry, Transwell model showed siRNA-3to PC3cell growthinhibition rate, apoptosis rate, invasive, which are statistically significant (P <0.01).Conclusions:1) It was found that FLIP-siRNA can inhibit FLIPLgene expression and promoteexpression levels of caspase8gene.2) The experimental results show FLIP-siRNA targeting gene silencing inhibitthe growth of PC3cells and promote apoptosis and reduce its invasiveness.3) Experimental results suggest FLIPLgene may be an effective target for genetherapy of PCa(including AIPC).