钙激活中性蛋白酶在肝癌细胞迁移能力中的作用

摘要

Objective: To evaluate the effect of calcium-activated neutral protease ( calpain) on metastasis of hepatocarcinoma cell. Methods: Western blotting assay was used to detect the expression and zymohydrolysis of calpain-1, calpain-2 and Src in hepatoma cell or hepatocyte cell lines with different metastasis ability, while wound healing assay was used to detect the effect of calpain-2 inhibitor on the migration ability of MHCC97-H (hepatocarcinoma cell with high metastasis ability). Results: As Western blot result showing, the expression and the active low molecular weight (LMW) forms of calpain-1, calpain-2 and Src from zymohydrolysis were not consistent in different hepatic cell lines, in which the alteration model of calpain-2 and Src were related with the metastasis ability except calpain-1. In MHCC97-H cell line, the expression level and active LMW forms of calpain-2 and Src were reduced by calpain-2 inhibitor. Consistently, with the inhibitor of calpain-2, the migration of MHCC97-H was reduced, and the higher the inhibitor concentration was, the stronger the inhibition of cell migration ability was. Conclusions: In calpain family, the up-regulation of calpain-2 expression and in-crease of its activity might increase the migration ability of hepatoma cells by activate Src.%目的:观察不同转移能力肝癌细胞中钙激活中性蛋白酶(calpain)的表达情况,利用抑制剂抑制calpain活性,评价calpain对肝癌细胞迁移能力的影响.方法:用蛋白质印迹(Western blotting)检测不同转移能力肝癌细胞或肝细胞中calpain-1、calpain-2和Src的表达和酶解情况；用划痕法观察calpain-2抑制剂对高转移性肝癌细胞(MHCC97-H)迁移能力的影响.结果:Western blot结果显示,不同肝细胞系中calpain-1、calpain-2及Src的表达水平和剪切成为低分子量(LMW)活性形式的程度不一样,其中calpain-2和Src的表达与肝癌细胞的迁移能力有相关性,calpain-2抑制剂明显下调MHCC97-H细胞中calpain-2和Src的表达,减少其被剪切的程度；划痕实验结果显示,calpain-2抑制剂能明显减少MHCC97-H细胞向划痕区域迁移,而且抑制剂浓度越大,细胞迁移能力被抑制的程度就越重.结论:calpain-2的表达上调和活性增强可通过激活Src增加肝癌细胞的迁移能力.