Effect Of Bcl-2 And Bcl-xl SiRNA On Drug Sensitivity Of HepG2 Cells

AIM To investigate the changes of the drug sensitivity and mechanism in Bcl-2 and Bcl-xl siRNA transfected HepG2 cells.METHODS Bcl-2,Bcl-xl siRNA and negative siRNA expression vector were constructed and stably transfected into HepG2 cells. RT-PCR and Immunofluorescence were used to detect the target gene expression. Western blot was used to detect Bcl-2,Bcl-xl,Bax and caspase-3 protein expressiom.Drug sensitivity of the cells to 5-fluorouracil (5-FU) and lO-hydroxycamptothecin(HCPT) were analyzed with MTT and flow cytometry. RESULTS The mRNA and protein expression level of Bcl-2 and Bcl-xl in Bcl-2 siRNA,Bcl-xl siRNA and Bcl-2 siRNA/Bcl-xl siRNA stable transfected were reduced compared with negative siRNA transfected or untreated cells. The protein expression of Bax had no changed and caspase-3 was up-regulated when Bcl-2 and Bcl-xl protein were reduced. MTT results showed that Bcl-2 and Bcl-xl transfected had higher cell inhibitory after treated with 5-FU or HCPT. Added 5-FU (1300mg/L) and HCPT(0.72mg/L),FCM results demonstrated that sub G1 population increased in Bcl-2 siRNA/Bcl-xl siRNA co-transfected, Bcl-xl siRNA and Bcl-2 siRNA respectively transfected cells compared with negative siRNA or untreated cell(5-FU:59.1±1.19%, 53.2±3.22%, 46.2±1.62%vs 24.4±2.48%,or 22.7±2.61 P<0.05; HCPT: 83.4±3.09%, 57.1±1.52% , 51.5±1.89%vs 24.2±2.70% or 26.3±0.91%, P<0.05).