苦参碱调控Wnt/β-catenin途径抑制宫颈癌细胞系Caski细胞侵袭迁移

Effects of matrine on invasion and migration of cervical cancer cell line Caski by Wnt/β-catenin pathway

目的:基于Wnt/β-catenin途径探索苦参碱抑制人宫颈癌细胞系Caski细胞侵袭迁移的分子机制。方法:Caski细胞用不同浓度苦参碱处理后,MTT法检测细胞增殖抑制情况;Transwell小室侵袭迁移实验检测细胞侵袭迁移情况;ELISA法检测细胞基质金属蛋白酶9(matrix metalloproteinase 9,MMP-9)和血管内皮生长因子(vascular endothelial growth factor,VEGF)的分泌变化;实时荧光定量PCR检测细胞糖原合成酶激酶3β(glycogen synthase kinase 3β,GSK-3β)、Wnt2B蛋白的表达情况;免疫蛋白印迹检测细胞β连环蛋白(β-catenin)、磷酸化β连环蛋白(p-β-catenin)含量。结果:苦参碱对Caski细胞的增殖具有抑制作用(P<0.05),且呈现时间、浓度依赖性;苦参碱抑制Caski细胞的侵袭迁移率(P<0.05),对Caski细胞分泌MMP-9、VEGF的能力具有显著抑制作用(P<0.05);苦参碱可显著降低Caski细胞Wnt2B mRNA的表达及β-catenin的蛋白含量,增加Caski细胞中GSK-3βmRNA的表达和p-β-catenin的蛋白含量。结论:苦参碱通过促进Caski细胞Wnt/β-catenin通路中GSK-3βmRNA表达,提高p-β-catenin蛋白含量,降低Wnt2B mRNA表达、β-catenin蛋白含量以及Caski细胞内MMP-9、VEGF的分泌,进而抑制Caski细胞的侵袭迁移。

Objective:This study aims to explore the molecular mechanism of matrine in inhibiting the invasion and migration of human cervical cancer cell line Caski cells based on the Wnt/β-catenin pathway. Methods:Caski cells were treated with different concentration of matrine. Methyl thiazolyl tetrazolium(MTT)assay was used to detect cell proliferation inhibition;Transwell chamber invasion and migration assay was used to detect cell invasion and migration;ELISA assay was used to detect cell matrix metalloprotein9(MMP-9)and vascular endothelial growth factor(VEGF)secretion;Quantitative real-time PCR(qRT-PCR)was used to detect cell glycogen synthase kinase3β(GSK-3β)and wingless/integrated 2 B(Wnt2 B)expression;Western blot was used to detect cell β-catenin,p-β-catenin protein content. Results:Matrine had inhibitory effect on the proliferation of cervical cancer cells Caski cells(P < 0.05)and showed a time and concentration dependence. Matrine inhibited the invasion and migration of Caski cells(P < 0.05),and it had a significant effect on Caski cells. The ability to secrete MMP-9 and VEGF has a significant inhibitory effect(P < 0.05);matrine can significantly reduce Wnt2 B mRNA expression and β-catenin protein content in Caski cells,and increase GSK-3β mRNA expression in Caski cells and p-β-catenin protein content. Conclusion:Matrine promotes the expression of GSK-3βmRNA in Wnt/β-catenin pathway of Caski cells,increases the content of p-β-catenin protein,reduces the expression of Wnt2 B mRNA,the content of β-catenin protein and the secretion of MMP-9 and VEGF in Caski cells,thereby inhibiting the invasion and migration of Caski cells.