草苁蓉环烯醚萜苷(IGBR)对TGF-β1诱导的HepG2细胞上皮间质转化模型的影响

Effect of iridoid glycosides from Boschniakia rossica on epithelial-mesenchymal transition of HepG2 cells induced by transforming growth factor-beta 1

目的研究草苁蓉环烯醚萜苷(IGBR)对TGF-β1诱导肝癌HepG2细胞上皮间质转化(EMT)的影响作用。方法用10μg/L TGF-β1诱导HepG2肝癌细胞株构建肝癌细胞EMT模型。实验分为对照组、模型组与IGBR组3组,对照组用无血清DMEM处理,模型组用10μg/L TGF-β1处理,IGBR组用10μg/L TGF-β1和500 mg/L IGBR联合处理,培养48 h。利用细胞黏附实验、划痕愈合实验和Transwell小室实验观察细胞迁移和侵袭能力。RT-PCR法和Western Blot法检测细胞中E-钙黏蛋白、N-钙黏蛋白、波形蛋白的mRNA和蛋白表达,Western Blot法检测Slug、Twist1、ZEB1、p-STAT3、STAT3的蛋白表达。计量资料多组间比较采用单因素方差分析,进一步两两比较采用LSD-t检验;两组间比较采用成组t检验。结果TGF-β1诱导后,模型组HepG2细胞呈现长梭形改变;与模型组比较,IGBR组细胞黏附率降低,抑制细胞迁移、侵袭能力(P值均<0.05),E-钙黏蛋白的mRNA表达和蛋白表达均增高(P值均<0.05),N-钙黏蛋白和波形蛋白的mRNA表达和蛋白表达均降低(P值均<0.05),Slug、Twist1、ZEB1蛋白表达和p-STAT3蛋白表达均降低(P值均<0.05)。结论IGBR可抑制TGF-β1诱导的HepG2细胞EMT过程,从而减弱HepG2细胞黏附力和细胞迁移、侵袭能力,上调E-钙黏蛋白,下调N-钙黏蛋白和波形蛋白,上调Slug、Twist1、ZEB1、STAT3的蛋白表达,其作用可能通过抑制STAT3通路下调Slug、Twist1、ZEB1等EMT转录因子来实现。

Objective To investigate the effect of iridoid glycosides from Boschniakia rossica(IGBR)on epithelial-mesenchymal transition(EMT)of HepG2 hepatoma cells induced by transforming growth factor-beta 1(TGF-β1).Methods HepG2 hepatoma cells were induced by 10μg/L TGF-β1 to construct an EMT model of hepatoma cells.The cells were divided into control group(treated with serum-free DMEM),model group(treated with 10μg/L TGF-β1),and IGBR group(treated with 10μg/L TGF-β1 and 500 mg/L IGBR),and all cells were cultured for 48 hours.Cell adhesion assay,wound healing assay,and Transwell chamber assay were used to observe the migration and invasion abilities of cells.RT-PCR and Western blot were used to measure the mRNA and protein expression levels of E-cadherin,N-cadherin,and vimentin in cells,and Western blot was used to measure the protein expression levels of Slug,Twist1,ZEB1,p-STAT3,and STAT3.A one-way analysis of variance was used for comparison of continuous data between multiple groups,and the least significant difference t-test was used for further comparison between two groups;the independent-samples t test was used for comparison between two groups.Results After TGF-β1 induction,HepG2 cells in the model group showed long spindle-shape changes,while those in the control group showed polygonal epithelia-like changes.Compared with the model group,the IGBR group had a significant reduction in cell adhesion rate and significant inhibition of cell migration and invasion abilities(all P<0.05),as well as significant increases in the mRNA and protein expression levels of E-cadherin(P<0.05),significant reductions in the mRNA and protein expression levels of N-cadherin and vimentin(all P<0.05),and significant reductions in the protein expression levels of Slug,Twist1,ZEB1,and p-STAT3(all P<0.05).Conclusion IGBR can inhibit TGF-β1-induced EMT process in HepG2 cells,thereby attenuating cell adhesion,migration,and invasion abilities,and it can also upregulate E-cadherin,downregulate N-cadherin and vimentin,and upregulate the protein expression of Slug,Twist1,ZEB1,and STAT3,possibly by inhibiting the STAT3 pathway to downregulate the EMT transcription factors such as Slug,Twist1,and ZEB1.