The Effect And Mechanism Of KSRP On Regulating The Proliferation, Cycle And Apoptosis Of Hepatocellular Carcinoma Cell Line HepG2

Objective: This study aims to investigate the effects of K-H type splicing regulatory protein（KSRP）on the proliferation,cell cycle and apoptosis of hepatocellular carcinoma（HCC）cell line HepG2.Methods: 1.The GEO database was used to examine the expression level of KSRP in tumor tissues and corresponding adjacent non-tumorous tissues of HCC patients.2.The lentivirus was used to knock-down or overexpress KSRP,puromycin was used to screen the stably expressed HepG2 cell lines which knock-down or overexpressed KSRP,respectively.And western blot was used to detect the expression levels of KSRP protein in HepG2 control group,overexpression group and knock-down group,respectively,to verify the effects of overexpression and knock-down of KSRP.3.The HepG2 cell proliferation rates in the KSRP overexpression or knock-down group and the control group at 24 h,48h,72 h and 96 h were detected by CCK-8 assay.4.The RNA in HepG2 cells was extracted by the Trizol method,and the differentially expressed cell proliferation-related genes in sh KSRP and the control group were detected by RNA-seq.5.The changes of HepG2 cell cycle distribution in the sh KSRP group and the control group were detected by flow cytometry.6.The double-staining method of Annexin V-FITC/PI was used to detect the apoptosis of HepG2 cells in the sh KSRP group and the control group.Results: 1.The expression of KSRP in the tumor tissues of HCC patients was higher than that in the adjacent tissues（P<0.001）.2.The results of CCK-8 showed that the proliferation of HepG2 cells with KSRP knock-down was significantly decreased（P<0.01）.3.RNA-seq showed that the knock-down of KSRP could inhibit the expression of MKI67 gene（P<0.05）.4.Flow cytometry confirmed that knocking down KSRP could block HepG2 cells in the S phase（cell cycle）,and promote apoptosis,and the difference in the apoptosis rate was statistically significant（P<0.05）.Conclusion: Knocking down KSRP can arrest HepG2 cells at S phase of the cell cycle,inhibit cell proliferation and increase the apoptosis rate.This may be closely related to the down-regulation of proliferation-related gene MKI67.