ATG7 Gene Silencing Enhances The Sensitivity Of Ovarian Cancer SKOV3 Cells To Cisplatin Chemotherapy

Objective:To investigate the effect of Autophagy-Related Gene 7（ATG7）silencing induced by small interfering RNA（siRNA）on the sensitivity of ovarian cancer SKOV3 cells to cisplatin（DDP）.Method:1.MTT method was used to select the optimal concentration and time of DDP for ovarian cancer SKOV3 cell line.2.The reactive oxygen species（ROS）detection kit was used to detect the levels of reactive oxygen species produced by ovarian cancer SKOV3 cell line when different concentrations of DDP were used.3.Western blot was used to detect the expression levels of autophagy-related proteins Beclin-1,P62,LC3-Ⅰ,LC3-Ⅱand ATG7 in ovarian cancer SKOV3 cell line with different concentrations of DDP.4.The IL-17RA levels of SKOV3 cell were detected by Western blot.5.The effect of DDP before and after siRNA-ATG7interference on the survival rate of ovarian cancer SKOV3 cell line was detected by MTT method.6.Annexin V-FITC/PI staining method was used to detect the effect of DDP on ovarian cancer SKOV3 cell line apoptosis rate before and after siRNA-ATG7 interference.7.Mitochondrial Membrane Potential Detection Kit（JC-1）detects the effects of DDP before and after siRNA-ATG7 interference on the early apoptosis of ovarian cancer SKOV3 cell line.8.The morphology of ovarian cancer SKOV3 cell line induced by DDP before and after interference with siRNA-ATG7 was observed with an inverted microscope.Results:1.MTT results showed that different concentrations of DDP(2,4,7,9,10,20,40,60μg·mL^-1)had a concentration-dependent and time-dependent effect on the proliferation inhibition of ovarian cancer SKOV3 cell line.2.The results of reactive oxygen species（ROS）test showed that the ovarian cancer SKOV3 cell line produced more reactive oxygen species when different concentrations of DDP(0,2,4,8μg·mL^-1)were applied.3.The results of Western blot showed that the expression levels of autophagy-related proteins Beclin-1 and ATG7 in ovarian cancer SKOV3 cell line increased,and the expression level of P62 decreased when DDP(0,2,4,8μg·mL^-1)was used at different concentrations.The conversion of LC3Ⅰto LC3Ⅱalso gradually increased.4.The results of Western blot showed that the interference effect of siRNA-ATG7 on ATG7 gene in ovarian cancer SKOV3 cell line was obvious.5.The results of MTT assay showed that the survival rate of ovarian cancer SKOV3 cell line induced by DDP was significantly lower than that before siRNA-ATG7 interference.6.The results of Annexin V-FITC/PI staining showed that the apoptosis rate of ovarian cancer SKOV3 cell line induced by DDP was significantly increased after interference compared with that before siRNA-ATG7 interference.7.The results of mitochondrial membrane potential detection kit（JC-1）showed that the early apoptotic rate of ovarian cancer SKOV3 cell line induced by DDP was significantly increased after interference compared with that before siRNA-ATG7 interference.8.The results of an inverted microscope observation showed that in the DDP/siRNA-ATG7combination group,cellular density was significantly reduced,cell-cell gap was enlarged,the number of suspended cells was significantly increased volume of cell cytoplasm was reduced,and cells gradually shrunk,rounded and lost their normal morphology with obscure boundaries.Conclusion:1.Autophagy may be involved in DDP resistance in ovarian cancer SKOV3 cells.2.ATG7 gene silencing enhances the sensitivity of ovarian cancer SKOV3 cells to cisplatin chemotherapy.