小檗碱通过TGF-β/Smad通路抑制TGF-β1诱导的人肝癌HepG2细胞上皮间质转化的研究

Berberine inhibits TGF-β1-induced epithelial-mesenchymal transition in human liver cancer HepG2 cells via TGF-β/Smad pathway

目的探讨小檗碱(berberine)对转化生长因子-β1(transforming growth factor-β1,TGF-β1)诱导的人肝癌HepG2细胞上皮间质转化(epithelial-mesenchymal transition,EMT)的作用及其机制。方法MTT法检测小檗碱对HepG2细胞增殖活性;采用10 ng·L-1 TGF-β1诱导HepG2细胞EMT模型形成,并加入小檗碱处理;克隆形成实验、细胞划痕和Transwell小室实验分别检测HepG2细胞的克隆形成能力、迁移和侵袭能力;免疫荧光法检测EMT间质标志物Vimentin的表达;Western blot法检测EMT标志物蛋白(E-cadherin、N-cadherin、Snail)、基质金属蛋白酶(MMP-2)、TGF-β/Smad通路蛋白(Smad2、p-Smad2、Smad3、p-Smad3)的表达。结果小檗碱呈浓度时间依赖性抑制HepG2细胞活性;与TGF-β1组比较,小檗碱可以明显抑制HepG2细胞的克隆形成能力、迁移和侵袭能力;并且小檗碱可以抑制TGF-β1上调的E-cadherin蛋白表达,和TGF-β1下调的N-cadherin、Vimentin、Snail、MMP-2、p-Smad2、p-Smad3蛋白表达。结论小檗碱可能通过抑制TGF-β/Smad信号通路,干预TGF-β1诱导HepG2细胞的EMT进程,抑制HepG2细胞的迁移和侵袭能力。

Aim To investigate the effects of berberine on epithelial-mesenchymal transition(EMT)of human liver cancer HepG2 cells induced by transforming growth factor-β1(TGF-β1)and its mechanism.Methods MTT assay was used to detect the proliferation activity of berberine on HepG2 cells.After 10 ng·L-1 TGF-β1 was used to induce EMT model process of HepG2 cells,berberine was added to treat HepG2 cells.Colony formation,cell scratch and Transwell assays were used to detect the clonogenic,migratory and invasive capabilities of HepG2 cells.Immunofluorescence assay was used to detect the expression of EMT mesenchymal marker Vimentin.Western blot assay was used to detect the proteins expression of EMT marker(E-cadherin,N-cadherin,Snail),matrix metalloproteinase(MMP-2),TGF-β/Smad pathway(Smad2,p-Smad2,Smad3,p-Smad3)in HepG2 cells.Results Berberine inhibited the proliferation of HepG2 cells in a concentration-time dependent manner.Compared with TGF-β1 group,berberine could significantly inhibit the abilities of colony formation,migration and invasion of HepG2 cells.Berberine could significantly inhibit the expression of E-cadherin protein up-regulated by TGF-β1,and N-cadherin,Vimentin,Snail,MMP-2,p-Smad2,p-Smad2 proteins expression down-regulated by TGF-β1.Conclusions Berberine may interfere with the EMT process of HepG2 cells induced by TGF-β1 by inhibiting the TGF-β/Smad signaling pathway to inhibit the HepG2 cell migration and invasion.