Effect of GTPBP4 silencing on radiosensitivity of EC9706 cells
Zhang Cuihong1, Lyu Xin2, Fan Cai1, Ma Bojing1, Zhang Yi1, Zhang Jianjun1
1Department of Radiation Oncology, 980th Hospital of PLA Joint Logistics Support Force, Shijiazhuang 050082, China; 2Department of Medical Services, The Central Hospital of Baixiang County, Xingtai 055450, China
Abstract：Objective To investigate the effect of GTPBP4 silencing by RNA interference on the radiosensitivity of esphageal cancer EC9706 cells line. Methods The expression data of GTPBP4 in esophageal cancer tissues was obtained from public Gene Expression Omnibus (GEO) database. Recombinant plasmid-mediated RNA interference (RNAi) was employed to transfect the esophageal cancer EC9706 cell to evaluate the influence of GTPBP4 silencing on the proliferation, apoptosis and radiosensitivity of esphageal cancer EC9706 cells. The expression levels of GTPBP4 mRNA and protein and apoptosis-associated proteins of Bax,cleaved caspase-9,cleaved caspase-3 and Bcl-2 were determined by qRT-PCR and Western blot. The cell proliferation was determined by MTT assay. The changes in cell apoptosis were detected AnnexinⅤ-FITC/PI double staining flow cytometry. The variations in radiosensitivity after radiation exposure were assessed by clone formation assay. Results The expression level of GTPBP4 in the esophageal cancer tissues was significantly higher than that in the normal adjacent esophageal tissues (P<0.001). qRT-PCR and Western blot demonstrated that the expression levels of GTPBP4 mRNA and protein in the GTPBP4-siRNA group were significantly lower than those in the blank and negative control groups (both P<0.001), suggesting that the plasmid was successfully transfected into the EC9706 cells. MTT assay indicated that the EC9706 cell proliferation rate was significantly inhibited (P<0.001). Flow cytometry found that the apoptosis rate was significantly increased in the GTPBP4-siRNA group (P<0.001). After GTPBP4 gene interference combined with radiotherapy, the cell sensitivity enhancement ratio was 1.716. The apoptosis rate of EC9706 cells was significantly increased in the GTPBP4-siRNA group (P<0.001). The expression levels of apoptosis-associated proteins including cleaved caspase-9, cleaved caspase-3 and Bax were significantly up-regulated, whereas that of Bcl-2 was significantly down-regulated in the EC9706 cells in the GTPBP4-siRNA group (P<0.001, P=0.001, P=0.001 and P=0.005). Conclusions GTPBP4 gene is highly expressed in human esophageal cancer tissues. RNAi technology can effectively inhibit the expression of GTPBP4 gene in the EC9706 cells, thereby suppressing cell proliferation, inducing cell apoptosis and enhancing the radiosensitivity of cells.
Zhang Cuihong,Lyu Xin,Fan Cai et al. Effect of GTPBP4 silencing on radiosensitivity of EC9706 cells[J]. Chinese Journal of Radiation Oncology, 2021, 30(5): 509-513.
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