Effect of silencing of poly(ADP-ribose) polymerase-1 on cell apoptosis induced by hydroquinone in rat bone marrow mesenchymal stem cells / 中国职业医学

ABSTRACT

OBJECTIVE: To explore the effect of silencing of poly( ADP-ribose) polymerase-1( PARP-1) on cell apoptosis induced by hydroquinone( HQ) in rat bone marrow-derived mesenchymal stem cells( BMMSCs). METHODS: i) The RNA expression vectors for PARP-1 gene were transfected into BMMSCs. Neomycin was used to select the transfected cells that stably expressed PARP-1-shRNA. Western blotting was used to examine the gene silencing efficiency. ii) BMMSCs with PARP-1 silencing were sorted as the treated group,while BMMSCs with empty vector were considered as the control group.HQ dissolved in phosphate buffer solution at the concentrations of 0. 0,2. 5,5. 0,10. 0,20. 0,40. 0,80. 0,160. 0 and320. 0 μmol / L were given to both groups for 24 hours. The cell viability was detected by methyl thiazolyl tetrazolium assay,and the concentration of HQ was chosen for the following study based on cell viability. iii) Both groups were treated by HQ at concentrations of 0. 0-20. 0 μmol / L for 24 hours,then the apoptosis of BMMSCs was detected by flow cytometry. The PARP-1 mRNA expression was determined by real-time fluorescent quantitative polymerase chain reaction assay. RESULTS: i) PARP-1 silencing cells and empty vector control cells were successfully screened with a mass concentration of 400 mg / L neomycin,and confirmed by level of protein expression. The interference efficiency of PARP-1 gene and inhibition efficiency was 85. 00%. ii) Based on the result of cell viability,HQ at concentrations of 0. 0-20. 0 μmol / L was chosen for the following study. iii) Compared with the group treated by HQ at concentration of 0. 0 μmol / L,the rate of early apoptosis of control group increased significantly with HQ at concentration of 10. 0 μmol / L while that of treated group was increased significantly at concentration of 5. 0 μmol / L( P < 0. 05). In addition,at concentrations of 0. 0-10. 0 μmol / L,the rates of early apoptosis in both groups increased in a dose-dependent manner( P < 0. 01). Compared with the group treated by HQ at concentrations of 0. 0 μmol / L,the expression of PARP-1 mRNA of both groups increased significantly at the concentration of 5. 0 μmol / L( P < 0. 05). The expression of PARP-1 mRNA of treated group was less than that of control group with HQ at every concentration( P < 0. 05). At concentrations of 0. 0-20. 0 μmol / L,the expression of PARP-1 mRNA of both groups increased in a dose-dependent manner( P < 0. 01). CONCLUSION: Silencing PARP-1 in BMMSCs caused cell apoptosis. PARP-1 may participate in cell apoptosis induced by HQ.