AMPK抑制TGF-β信号通路减轻EMT诱导膀胱癌转移机制研究

Mechanism research of AMPK in inhibiting TGF-β signaling pathway and alleviating the metastasis of EMT-induced bladder cancer

目的探讨AMPK抑制TGF-β信号通路减轻EMT诱导膀胱癌转移机制。方法采用Lipofectamine2000介导AMPKα1转染至人膀胱移行上皮癌BIU-87,噻唑蓝(MTT)比色法和AnnexinV-FITC/PI双染法检测shRNA对细胞增殖和凋亡的作用。细胞划痕实验、Transwell小室侵袭实验检测体外细胞迁移和侵袭能力。Western blot检测E-cadherin、N-cadherin和Vimentin蛋白的表达情况。结果转染后的C4-2 DN细胞生长速度明显慢于未转染的BIU-87细胞及转染空载体的C4-2 E细胞,且在24 h内的细胞凋亡率均明显高于转染空载体的C4-2 E,组间差异具有统计学意义(P<0.05)。AMPK抑制TGF-β信号通路能够抑制人膀胱移行上皮癌BIU-87细胞的增殖和侵袭迁移(P<0.05)。同时,AMPK抑制TGF-β信号通路下调N-cadherin 和 Vimentin 蛋白水平,上调E-cadherin 蛋白表达。结论 AMPK抑制TGF-β信号通路下调N-cadherin 和 Vimentin 蛋白水平,上调E-cadherin 蛋白表达,减轻EMT诱导膀胱癌转移。

Objective To investigate the role of AMPK in inhibiting TGF-β signaling pathway and attenuating EMT-induced bladder cancer metastasis. Methods Lipofectamine2000 was used to mediate AMPKα1 transfection into human bladder transitional cell carcinoma BIU-87. The effect of shRNA on cell proliferation and apoptosis was detected by MTT colorimetric assay and AnnexinV-FITC/PI double staining. Cell scratch assay and Transwell chamber invasion assay were used to detect cell migration and invasion in vitro. The expression of E-cadherin, N-cadherin and Vimentin protein was detected by Western blot. Results The growth rate of C4-2 DN cells after transfection was significantly slower than that of untransfected BIU-87 cells and C4-2 E cells transfected with empty vector, and the apoptosis rate of C4-2 DN cells after transfection was significantly higher than that of C4-2 E cells transfected with empty vector transfection within 24 h, and the difference between groups was statistically significant (P<0.05). AMPK inhibits proliferation and invasion and migration of human bladder transitional epithelial carcinoma BIU-87 cells by inhibiting TGF-β signaling pathway (P<0.05). At the same time, AMPK inhibits TGF-β signaling pathway and down-regulates N-cadherin and Vimentin protein levels, and up-regulates E-cadherin protein expression. Conclusion AMPK inhibits TGF-β signaling pathway, down-regulates N-cadherin and Vimentin protein levels, up-regulates E-cadherin protein expression, and reduces EMT-induced bladder cancer metastasis.