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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cal-pain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。
OBJECTIVE: To observe the expression of calpain in hepatocarcinoma cells with different metastatic potential, use the inhibitor to inhibit the activity of cal-pain and evaluate the effect of calpain on the migration ability of hepatoma cells. Methods: Western blotting was used to detect the expression of calpain-1, calpain-2 and Src in hepatoma cells or hepatocytes with different metastatic potential and the enzymatic hydrolysis. The calpain-2 inhibitor Effect of migration ability of cells (MHCC97-H). Results: Western blot results showed that the expression levels of calpain-1, calpain-2 and Src in different hepatocellular lines were different from those of low molecular weight (LMW) forms, of which the expressions of calpain-2 and Src were significantly different Calpain-2 inhibitor significantly down-regulated the expression of calpain-2 and Src in MHCC97-H cells and decreased the extent of their cleavage. Scratch assay showed that the calpain-2 inhibitor significantly decreased MHCC97-H cells migrated to the scratch area, and the greater the inhibitor concentration, the more severe the cell migration was inhibited. Conclusion: The up-regulation of calpain-2 and the enhancement of its activity can increase the migration ability of hepatoma cells by activating Src.