| MicroRNA is a kind of non-coding small RNA molecule,which regulate post-transcriptional of genes and plays a key role in the evolution of many tumors.Among the small RNAs associated with human tumors,microRNA-17-92 family is an earlier discovered microRNA family.MicroRNA-17-5p is an important member in the family of microRNA-17-92,which expresses in various malignant tumors abnormally and participates in the progress of regulating human multiple system tumors.Colon cancer is one of the most common malignant tumors in humans,and its occurrence is a complex process of multiple carcinogenic factors and multiple oncogenes interactions.However,as a carcinogenic factor,the mechanism of microRNA-17-5p in the pathogenesis of colon cancer remains unclear currently.Objective:This research was conducted to observe the effect of microRNA-17-5p on the biological behavior of colon cancer cell line HCT116,and investigated the mechanism of microRNA-17-5p on the occurrence and development of colon cancer.The purpose is to forecast its target and corroborate its function in colon cancer.Methods:1.Human colon cancer cell line HCT116 was selected for cell culture as the cell model in vitro.MicroRNA-17-5p mimics and negative control(NC)were transfected into HCT116 cells to construct the cell model of over-expression of microRNA-17-5p.The group of transfected with microRNA-17-5p mimics was used as experimental group,and the group of transfected with mimics control was used as negative control group.2.Real-time quantitative reverse transcription-PCR(q RT-PCR)was used to investigate the relative expression of microRNA-17-5p and transforming growth factor-β receptor II(TGFBR2)at m RNA level after48 hours of transfection in HCT116 cells.3.CCK-8 assay was used to detect Optical Density(OD)at 24 h,48h,72 h and 96 h after transfection.Cell growth curves were drawn to compare the changes of cell proliferation between the experimental group and the negative control group.4.Flow cytometry was used to observe the amount of early apoptotic HCT116 cells through Annexin-V-FITC kit after overexpression of microRNA-17-5p.5.Transwell was used to analyze the changes in HCT116 cell on the ability of invasion and migration after overexpression of microRNA-17-5p.6.Cell Scratch Healing Experiments was used to analyze the changes in the healing ability of HCT116 cell.7.Target Scan predicts the possible target gene of microRNA-17-5p.Western blot was used to detect the expression of transforming growth factor-beta receptor II(TGFBR2)at protein level after 48 hours of transfection in HCT116 cells.Results:1.The results of q RT-PCR showed that the expression of microRNA-17-5p in the experimental group was significantly higher than that in the negative control group(P=0.0001).2.The results of CCK-8 test showed that the OD value of cells in the experimental group was significantly higher than that in the negative control group after transfection of microRNA-17-5p mimics or mimics control(P=0.01,0.02,0.01).3.The results of flow cytometry showed that the amount of early apoptotic cells in the experimental group was significantly lower than that in the negative control group after transfection of microRNA-17-5p mimics or mimics control(P=0.002).4.The results of Transwell test showed that the number of invasive and migrating cells in the experimental group was significantly higher than that in the negative control group after transfection of microRNA-17-5p mimics or mimics control(P=0.04,0.004).5.The results of scratch healing test showed that the scratch healing rate of the experimental group was significantly faster than that of the negative control group after transfection of microRNA-17-5p mimics or mimics control(P=0.04,0.03,0.04).6.Bioinformatics predicts the target gene TGFBR2.The results of western blot and q RT-PCR showed that the expression of TGFBR2 in the experimental group was significantly lower than that in the negative control group after transfection of microRNA-17-5p mimics or mimics control at m RNA and protein level(P=0.03,0.01).Conclusions:1.In colon cancer HCT116 cells,the up-regulation of microRNA-17-5p can effectively promote proliferation,invasion and migration ability of cells and inhibit the early apoptotic of cells`.These results suggest that microRNA-17-5p can promote the development and metastasis of colon cancer cells,providing an evaluating evidence for the pathogenesis of colon cancer.2.In colon cancer HCT116 cells,the up-regulation of microRNA-17-5p expression can significantly reduce the expression of TGFBR2.These results suggest that microRNA-17-5p may play biological role by reducing TGFBR2 in colon cancer cells lines,providing a new potential target for the treatment of colon cancer. |
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