小檗碱通过调控ELK-3抑制人肝癌细胞的上皮-间质转化

Berberine inhibits the epithelial-mesenchymal transition of human liver cancer cells by regulating ELK-3

关键词:  小檗碱|肝癌细胞|上皮-间质转化|ELK-3

Key words:  berberine|liver cancer cells|epithelial-mesenchymal transition|ELK-3

摘要:

[目的] 研究小檗碱与ELK-3和人肝癌细胞的上皮-间质转化的关系及其作用机制。[方法] 将HuH7细胞分为空转组、pcDNA3.1-ELK-3组、小檗碱组和小檗碱+pcDNA3.1-ELK-3组。荧光倒置显微镜观察慢病毒转染后的细胞荧光状态；蛋白免疫印迹法（Western Blot）和实时荧光定量逆转录聚合酶链反应（RT-qPCR）检测ELK-3和ELK-3靶基因重组人早期生长反应蛋白-1（EGR-1）蛋白和基因表达量，上皮-间质转化相关分子钙黏附素E和波形蛋白的基因和蛋白表达量，以及乙型转化生长因子-1（TGF-β1）/丝裂原活化蛋白激酶（MAPK）信号通路中TGFβ1、细胞外调节蛋白激酶（ERK1/2）、磷酸化细胞外调节蛋白激酶（p-ERK1/2）、p38、磷酸化p38（p-p38）蛋白表达量；细胞划痕实验和体外Transwell小室实验观察HuH7细胞迁移和侵袭能力。[结果] 与空转组比较，小檗碱组的ELK-3、EGR-1和波形蛋白的基因和蛋白表达量显著降低（P<0.05），钙黏附素E基因和蛋白表达量显著增加（P<0.05），TGF-β1蛋白和p-ERK1/2/ERK1/2和p-p38/p38蛋白比值显著降低（P<0.05），细胞迁移和侵袭能力显著降低（P<0.05），而pcDNA3.1-ELK-3组的结果与之相反。与pcDNA3.1-ELK-3组比较，小檗碱+pcDNA3.1-ELK-3组的ELK-3、EGR-1和波形蛋白的基因和蛋白表达量显著降低（P<0.05），钙黏附素E蛋白和基因表达量显著增加（P<0.05），TGF-β1蛋白和p-ERK1/2/ERK1/2和p-p38/p38蛋白比值显著降低（P<0.05），并且细胞迁移和侵袭能力显著降低（P<0.05）。[结论] 小檗碱可通过减少ELK-3的表达量，从而抑制人肝癌细胞的上皮-间质转化。

Abstract:

[Objective] To study the relationship between berberine,ELK-3 and the epithelial-mesenchymal transition of human liver cancer cells and its mechanism.[Methods] HuH7 cells were divided into idler group,pcDNA3.1-ELK-3 group,berberine group and berberine + pcDNA3.1-ELK-3 group. We used fluorescence inverted microscope to observe the cell fluorescence status after lentivirus transfection. Western Blot and RT-qPCR detected the protein and gene expression of ELK-3 and ELK-3 target gene EGR-1 and epithelial-mesenchymal transition related molecule cadherin E,as well as the protein expression of TGFβ1,ERK1/2,p-ERK1/2,p38,and p-p38 in the TGF-β1/MAPK signaling pathway. Cell scratch test and in vitro Transwell chamber test observed the migration and invasion ability of HuH7 cells.[Results] Compared with the idling group,the gene and protein expression of ELK-3,EGR-1 and vimentin in the berberine group were significantly reduced (P<0.05),and the expression of cadherin E gene and protein were significantly increased (P<0.05),TGF-β1 protein,p-ERK1/2/ERK1/2 and p-p38/p38 protein ratios were significantly reduced(P<0.05),cell migration and invasion capabilities were significantly reduced (P<0.05),while the results of the pcDNA3.1-ELK-3 group were the opposite. Compared with the pcDNA3.1-ELK-3 group,the gene and protein expression of ELK-3,EGR-1 and vimentin in the berberine + pcDNA3.1-ELK-3 group were significantly reduced (P<0.05),the expression of cadherin E protein and gene were increased significantly (P<0.05),the ratio of TGF-β1 protein and p-ERK1/2/ERK1/2 and p-p38/p38 protein were decreased significantly (P<0.05),and cell migration and the invasion ability were significantly reduced (P<0.05).[Conclusion] Berberine could inhibit the epithelial-mesenchymal transition of human liver cancer cells by reducing the expression of ELK-3.