MiR-92b-3p Promotes Malignant Biological Properties Of Colorectal Carcinoma Via Targeting FBXW7

Objective:Colorectal carcinoma is one of the most common cancers with high morbidity and mortality.The incidence of colorectal carcinoma increased year by year,threateing life and health of more people all over the world.MiRNA is a kind of non-coding RNA which is involved in cell proliferation,differentiation and apoptosis.The treatment of colorectal carcinoma is traditional surgery and chemotherapy and mi RNA is considered as potential diagnosis and treatment targets in recent years.A number of studies have explored the molecular mechanisms of miRNA in colorectal cancer.It is known that miR-92b-3p has a relatively high level of abnormal expression in colorectal cancer cells and tissues than in adjacent normal tissues.However,it is rarely known about the relationship between miR-92b-3p and colorectal cancer and its mechanism.Methods: Part 1:1.CCK8 assay was used to detect the proliferation and colony formation of HCT116 and HT29 cells treated with miR-92b-3p inhibitor;2.Flow cytometry was used to detect the changes of cell cycle and apoptosis after treated with miR-92b-3p inhibitor;3.Transwell assaywas used to detect the changes of cell invasion and migration after treated with miR-92b-3p inhibitor;Part 2: 1.Bioinformatics technology and public database were used to predict the target genes of miR-92b-3p;2.The dual luciferase reporter assay was used to verify the interaction between mi R-92b-3p and its target genes;3.We established FBXW7 over-expression cell lines in HCT116 and HT29,examined the effects of FBXW7 over-expression on proliferation and colony forming ability withMTS assay;4.The changes of cell cycle ratio and apoptosis level were detected by flow cytometry in FBXW7 overexpression cell lines;5.In FBXW7 overexpression HCT116 and HT29 cells,the migration and invasion ability of cells were arrayed by transwell.Part 3: The cells were divided into blank group,the inhibitor group,inhibitor+siFBXW7 group and NC group.MTS assay was used to detect the proliferation and colony formation ability of HCT116 and HT29 cells in four groups;2 Change of cell cycle and apoptosis in four groups were detected by flow cytometric;3.Change of cell migration and invasion ability in four groupse were detected by transwell assay;Part 4:We established a transplanted tumor mice model with four cell groups,of which the tumor size was recorded and the growth curve was drawn.The tumor tissue obtained from transplanted tumor mice were sliced and the structure was detected by HE staining.Results: Part one: 1 miR-92b-3p has significantly higher levels of expression in colorectal cancer cell lines HCT116 and HT29,and inhibition of mi R-92b-3p significantly inhibited the growth and colony formation of colorectal cancer cells.More colorectal cancer cells were arrested in G0/G1 phase,and the percentage of cells in S phase decreased in miR-92b-3p suppressed cells.The apoptosis of colorectal cancer cells were promoted by inhibiton of miR-92b-3p(p<0.05);3.Inhibition of miR-92b-3p suppresses the invasion and migration of colorectal cancer cells(p<0.05).Part two: 1.According to informatics prediction,FBXW7 is one of the direct acting target genes of mi R-92b-3p,which is verified by double luciferase reporter gene experiments.2.We established FBXW7over-expression cell lines in HCT116 and HT29,and found high level of FBXW7 inhibited the proliferation and colony formation ability of colorectal cancer cells(p<0.05);3.More HCT116 and HT29 cells with over-expression FBXW7 were blocked in the G0/G1 phase,meanwhile the proportion of S phase cells decreased(p<0.05).The corresponding apoptosis level of cancer cells has increased significantly(p<0.05).4.Over expression of FBXW7 can inhibit the migration and invasion of colorectal cancer cells were inhibited by over-expression of FBXW7Part three: 1.Proliferation and colony forming ability has been significantly inhibited in mi R-92b-3p inhibitor group,and the proliferationand colony formation of inhibitor+siFBXW7 group has been partially restored(p<0.05);2.More cells of inhibitor group were arrested in G0/G1 phase and more cells were induced into apoptosis,but the proportion of each cell phase and apoptosis level were partly restored in inhibitor+siFBXW7 group(p<0.05).3.Invasion and migration have been significantly inhibited in mi R-92b-3p inhibitor group,and the invasion and migration of inhibitor+siFBXW7 group has been partially restored(p<0.05);Part four: 1.It was found that the tumor size of mice in inhibitor group was significantly smaller compared with the control group,while the inhibitor+siFBXW7 group counteracted the previous effects in mice(p<0.05).2.Compared with the control group,the tumor tissue structure of the inhibitor group was significantly inhibited,while the inhibitor+siFBXW7 group of cancer tissue has been largely restored(p<0.05).Conclusion: In colorectal cancer,the high level of miR-92b-3p inhibits the FBXW7 expression level,resulting in higher proliferation and anti-apoptotic ability of cancer cells.This is the first report on the interaction of miR-92b-3p and FBXW7 and their effects on cell biology function in colorectal cancer,which has also been proved in mice model.It reveals the molecular mechanism of oncogene function with miR-92b-3p in colorectal cancer.