摘要
目的 观察山仙颗粒含药血清对Hepa1-6 肝癌细胞迁移、侵袭能力的影响,并基于上皮-间充质转化(EMT)和Wnt/β-catenin信号通路探讨其可能的分子机制.方法 采用SD大鼠制备山仙颗粒低、中、高剂量含药血清和空白血清.体外培养Hepa1-6 肝癌细胞,设空白血清组、KYA1797K组及山仙颗粒低、中、高剂量含药血清组.取各组干预12 h、24 h、36 h后的Hepa1-6 肝癌细胞,通过划痕实验检测细胞迁移能力,Transwell实验检测细胞侵袭能力,Western blot法检测细胞中E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)、波形蛋白(Vimentin)、β-cate-nin和磷酸化糖原合成酶激酶3β(p-GSK3β)蛋白表达情况.结果 与空白血清组比较,山仙颗粒中、高含药血清组及KYA1797K组干预12h后的细胞迁移距离和山仙颗粒各组及KYA1797K组干预24 h、36 h后的细胞迁移距离均明显缩短(P均<0.05);山仙颗粒各组及KYA1797K组干预12 h、24 h、36 h后的穿膜细胞数量均明显减少(P均<0.05),细胞中E-cadherin蛋白相对表达量均明显升高(P均<0.05),细胞中N-cadherin、Vimentin、β-catenin、p-GSK3β蛋白相对表达量均明显降低(P均<0.05).山仙颗粒低、中、高剂量含药血清组各时间点的细胞迁移距离呈剂量依赖性缩短,穿膜细胞数量呈剂量依赖性减少,细胞中E-cadherin、N-cadherin和Vimentin蛋白相对表达量呈剂量依赖性升高或降低,组间两两比较差异均有统计学意义(P均<0.05).结论 山仙颗粒可以剂量依赖性地抑制Hepa1-6 肝癌细胞的迁移和侵袭,其机制可能与调控Wnt/β-catenin通路,上调E-cadherin和下调N-cadherin、Vimentin的表达,从而抑制细胞EMT进程有关.
Abstract
Objective It is to observe the effects of serum containing Shanxian granules on migration and invasiveness of hepatocellular carcinoma Hepa1-6 cell line,and explore its possible molecular mechanisms based on epithelial-mesen-chymal transition(EMT)and Wnt/β-catenin signaling pathways.Methods The SD rats were used to prepare Shanxian granule low,medium and high dose drug-containing serum and blank serum.Hepa1-6 hepatocellular carcinoma cells were cultured in vitro,and divided into blank serum group,KYA1797K group and Shanxian granules low,medium and high dose drug-containing serum groups.Hepa1-6 hepatocellular carcinoma cells were taken from each group after 12 h,24 h and 36 h of intervention,and their migration ability was detected by scratch assay,invasion ability by Transwell assay,and the protein expressions of E-cadherin,N-cadherin,vimentin,β-cadherin and phosphorylated glycogen synthase kinase 3β(p-GSK3β)in the cells were detected by Western blotting.Results Compared with the blank serum group,the cell migra-tion distances after 12 hours of intervention in the medium and high-dose medicated serum groups of Shanxian granule and KYA1797K group,and the cell migration distance after 24 and 36 hours of intervention in every Shanxian granule group and KYA1797K group was also significantly reduced(all P<0.05).After12 h,24 h,and36 h of intervention,the numbers of invading cells in each group of Shanxian granule and KYA1797K group were significantly reduced(all P<0.05),while the relative expression of E-cadherin protein in the cells was significantly increased(all P<0.05),additionally,the relative expressions of N-cadherin,Vimentin,β-catenin,and p-GSK3β proteins in the cells were significantly decreased(all P<0.05).The cell migration distance at each time point in the low,medium,and high-dose medicated serum groups of Shanxian granule showed a dose-dependent reduction,the number of invading cells displayed a dose-dependent decrease,the relative expressions of E-cadherin,N-cadherin,and Vimentin proteins in the cells exhibited a dose-dependent increase or decrease,the differences between the groups were all statistically significant(all P<0.05).Conclusion Shanxian gran-ule exhibited a dose-dependent inhibition on the migration and invasion of Hepa1-6 hepatocellular carcinoma cells.This effect may be attributed to its ability to upregulate E-cadherin expression and downregulate the expression of N-cadherin and Vimentin through the Wnt/β-catenin pathway,ultimately inhibiting the epithelial-mesenchymal transition(EMT)process in Hepa1-6 hepatocellular carcinoma cells.
基金项目
陕西省重点研发计划(2024SF-YBXM-147)
陕西省重点研发计划(2019SF-303)
陕西省中医药局科研项目(SZY-KJCYC-2023-031)
陕西中医药大学科研项目(2023GP13)
维普
万方数据