G-Protein-Signaling Modulator 2 Expression And Role In A CD133~+ Pancreatic Cancer Stem Cell Subset

Objective:To investigate the differential expression of GPSM2 in CD133~+PANC-1 cell subset and CD133~-PANC-1 cell subset,and to investigate the effect on proliferation,clone formation and migration of CD133~+PANC-1 cells by knocking down GPSM2.Method:In this study,the PANC-1 cell line with higher expression of GPSM2 protein was selected as the experimental cell line by western bloting(WB)pre-experiment.CD133~+and CD133~-subsets were selected from PANC-1 cell line by flow cytometry(FCM)using pancreatic cancer stem cells(PCSCs)surface molecular marker CD133antigen as a marker.The tumorigenic potential of the CD133~+PANC-1 cell subsets was assessed by cell cycle assay and subcutaneous tumor formation experiments in nude mice.Differential expression of GPSM2 was examined by real-time quantitative RT-PCR(RT-qPCR)and WB.To silence GPSM2 expression,a shRNA lentiviral vector targeting GPSM2 was constructed and stably transfected into CD133~+PANC-1 cells.The inhibitory efficiency of the GPSM2 gene was verified by RT-qPCR and WB.The proliferation,colony formation,and migration abilities of the transfected CD133~+PANC-1 cells were assessed by MTT,soft agar colony formation,and Transwell assays.Result:CD133~+and CD133~-cell subsets were successfully isolated from PANC-1 cells.The CD133 positive subset subcutaneously formed tumors in nude mice that were significantly bigger(343.05±57.59 mm~3 vs 176.86±32.58 mm~3,P<0.01)and denser(4.13±0.37 g vs 1.07±0.21 g,P<0.01)than those of the CD133~-cell subset.Cell cycle assay confirmed that the percentage of cells in G0/G1 phase in CD133~+PANC-1 cell subset was significantly higher than that in CD133~-PANC-1 cell subset[(65.34±3.25)%vs(56.56±4.87)%,P<0.01],which was consistent with the characteristics of stem cell cycle distribution.The GPSM2 mRNA and protein expression was significantly higher in CD133~+PANC-1 cell subset than in CD133~-subset.Stable downregulation of GPSM2 expression reduced the proliferation,colony formation,and migration abilities of CD133~+PANC-1 cells(P<0.05).Conclusions:The CD133~+PANC-1 cells have obvious stem cell characteristics and increased GPSM2 expression.Downregulation of GPSM2 significantly reduces the proliferation and migration ability of the cells.Therefore,GPSM2 may provide an important target for regulating PCSCs.