The Expression And Significance Of Gli-2 In Bladder Cancer Cell Lines T24 And Its Influences On Vegf And Mmp-9 Expression

Objective:To investigate the relationship between Gli-2 expression and clinicopathological characteristic through detecting the expression of Gli-2, a key gene of Hedgehog signaling pathway, in bladder cancer tissues. To observe the role of Gli-2 on cell proliferation and its underlying mechanisms in bladder cancer cell lines T24, and its influence on the protein expressions of VEGF and MMP-9.Methods:(1)The expression levels of Gli-2 in 77 cases of bladder carcinoma tissue specimens were detected by immunohistochemistry(IHC), and their clinicopathological significance was analyzed.(2)Gli-2 expression was down-regulated in bladder cancer cell lines T24 by small interfering RNA(si RNA). The total RNA and proteins were extracted respectively. RT-PCR was used to detect Gli-2 m RNA expression, western blot to analyse expression levels of Gli-2, VEGF and MMP-9 protein, CCK-8 assay and flow cytometry to evaluate cell proliferation, apoptosis and cell cycle,respectively.Results:(1)Gli-2 expression was observed in 100% of bladder cancer tissues. Gli-2expression was intimately associated with pathological stage, and the expression levels were significantly higher in high stage bladder cancer compared with low stage(P<0.001). And the expression in muscle invasive bladder cancer was significantly higher than that in non-muscle invasive bladder cancer(P <0.001), however, only marginal significance was observed between Gli-2 expression and tumor grade(P = 0.052).(2)After down-regulation of Gli-2 expression in T24 cells by si RNA, the m RNA expression levels of Gli-2 gene in the experiment group were significantly decreased, and statistical significance was observed compared with the control group(t =-3.031, P <0.05). In addition, Gli-2-si RNA treatment in bladder cells resulted in a significant reduction of Gli-2 m RNA and protein expression, and the expression levels of VEGF and MMP-9 protein were significantly reduced. Gli-2 expression knockdown produced the decreased proliferation of bladder cancer cells through evaluated CCK-8 test results(P<0.001). Flow cytometry was used to detect the cell cycle and apoptosis in bladder carcinoma cell lines T24, by which(27.8 ± 2.1)% of the cells were found arrested in G2 / M phase in the experiment group, while only(6.61 ±1.1)% in the control group, and the difference was statistically significant(P<0.05). Number of apoptosis cells was greatly increased, and was about(11.63±1.09)%, which was significantly higher than that in control group,(3.43±1.05)%. The difference was statistically significant(t=4.015, P <0.05).Conclusion:Gli-2 positive expression was associated with clinical pathological stage and the expression was significantly higher in muscle invasive bladder cancer.Gli-2 may participate in bladder cancer caicinogenesis through regulating cell proliferation and apoptosis, may become a new potential therapeutic target for the treatment of bladder cancer..