| Objective: To explore the role of LncRNA H19 as ceRNA in promoting the proliferation and metastasis of bladder cancer cells by antagonizing the expression of Lin28 b mRNA,a target gene regulated by let-7d.The reliable evidence of LncRNA H19 as a target gene and related signal transduction pathway of let-7d was obtained to elucidate the function of LncRNA H19 and to reveal the new molecular mechanism of proliferation,invasion and metastasis of bladder cancer.To provide further scientific basis for searching target genes and drugs for the diagnosis and treatment of bladder cancer.Methods: Through the chip data analysis in GEO database,we select the target gene that we want to study.From July 2015 to July 2016,35 patients with bladder cancer were collected from urology department of the first affiliated Hospital of China Medical University and received surgical treatment.The clinical specimens of 35 patients with bladder cancer and their adjacent normal tissues were collected.Certified by pathological tissue,accompanied by detailed clinical data.The qRT-PCR(Quantitative real-time polymerase chain reactionation was performed on 35 samples,and H19 and Lin28 b were selected as the research objects,and the correlation between H19 and Lin28 b expression in human bladder cancer was deduced by further qRT-PCR experiment and statistical calculation.Then we selected 4 bladder cancer cell lines and 1 normal bladder cell lines to measure the expression of H19 in each cell line.Then transfection of H19 Si RNA by transfection of liposome Lipo3000 to T24 of bladder cancer cells proved the best one of 3 different Si RNA.The synthetic H19 si RNAs were transfected into bladder cancer cell T24 by liposome Lipo3000,and Negative control NCwas used as control.The expression of H19,Lin28 b and Let-7d in the two experimental groups was compared by qRT-PCR experiment,and the relationship between H19 and Lin28 b was verified by Let-7d.The effect on bladder cancer cell proliferation was verified by CCK8 assay and plate cloning assay,and the effect on bladder cancer cell invasion was verified by scratch test and invasion test.The binding sites of H19 and Lin28 b to let-7d were predicted by the bioinformatics website Target Scan.and further verified by the published literature.In vitro,qRT-PCR and cell function experiments were used to verify the relationship between H19OLin28 b and let-7d and its effect on bladder cancer cells: transfection of synthetic H19 Si RNA and Let-7d inhibitor by liposome Lipo3000 was carried out into bladder cancer cell line 5637.Using 5637 bladder cancer cells transfected only with H19 Si RNA as control group,the expression of H19 Si RNA Lin28 b and Let-7d in two experimental groups were compared by qRT-PCR experiment to verify the relationship between H19 and Lin28 b via Let-7d.The effect on bladder cancer cell proliferation was verified by CCK8 assay and plate cloning assay,and the effect on bladder cancer cell invasion was verified by scratch test and invasion test.Finally,Western Blot was used to determine the expression of Lin28 b protein and its pathway c-Myc / Lin28 b / HMGA2 in vitro.Results: By qRT-PCR test,we found that the expression of H19 and Lin28 b in bladder cancer was significantly higher than that in paracancerous tissues or normal bladder tissues at the mRNA level.In addition,the enhanced expression of H19 and Lin28 b was also observed in bladder cancer cells,which confirmed the correctness of the high expression of H19 and Lin28 b in bladder cancer.Then we found a positive correlation between H 19 and Lin28 b expression in bladder cancer by statistical analysis.Then we screened out the most efficient si RNAs by transfecting H19 Si RNA into the cells in vitro,and confirmed that H19 could positively regulate the expression of Lin28 b from the mRNA level through qRT-PCR experiments in vitro.In addition,H19 Si RNA was used to interfere the expression of H19 in bladder cancer cells compared with that in NC group.CCK8 assay was used to demonstrate that decreasing the expression of H19 in bladder cancer cells could reduce the proliferation of bladder cancer cells.The results of scratch and transwell showed that decreasing the expression of H19 in bladder cancer cells could reduce the migration of bladder cells.The exact binding site between H19 / Lin28 b and Let-7d was obtained by bioinformatics prediction.Then we demonstrated the regulatory relationship between qRT-PCR and cell function experiments in bladder cancer cells: interfering with the expression of LncRNA H19 decreased the expression of target gene Lin28 bmRNA,and when the inhibitor was added with Let-7d,The co-expression of H19 and Lin28 b could be attenuated.The results of cell function test showed that when we transfected Let-7d inhibitor and H19 Si RNA in 5637 and transferred into H19 Si RNA only as control,the proliferation of CCK8 cells and the plate clone assay showed that the proliferation rate of bladder cancer cells increased significantly;and.Scratch test showed that the invasive ability of bladder cancer cells was significantly increased.Finally,we further found that H19 inhibited the expression of Lin28 b protein and reduced the expression of c-Myc / Linb / HMGA2 protein by Western blot analysis.These effects may lead to abnormal proliferation and metastasis of bladder cancer cells.Conclusion: 1)LncRNA H19 and Lin28 b were highly expressed in bladder cancer tissues and the expression of H19 and Lin28 b were positively correlated.2)LncRNA H19 acts as ceRNA to form H19 / Let-7d / Lin28 b network structure,which regulates the expression of Lin28b-b through the competitive binding of let-7d with mRNA of Lin28b-b,thus promoting the proliferation and metastasis of bladder cancer cells.3)In bladder cancer,H19 participates in the regulation of proliferation and metastasis of bladder cancer cells through c-Myc / Lin28 b / HMGA2 signaling pathway,and Lin28 b is the target of long chain noncoding RNA H19. 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