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目的探讨膜-细胞骨架联接蛋白ezrin在肝细胞肝癌生长和转移过程中的作用。方法分别应用免疫荧光、逆转录聚合酶链反应(RT-PCR)和Western blot检测ezrin和骨架蛋白在不同转移潜能肝癌细胞系中的表达。选取高转移潜能SF7721(SMMC-7721经转基因后稳定表达肝细胞生长因子．从而获得高转移潜能的细胞系)和 MHCC97-H细胞系为研究对象,通过RNA干扰技术下调SF7721和MHCC97-H细胞系中ezrin蛋白的表达,观察其运动和侵袭能力的变化:通过四甲基偶氮唑盐检测细胞增殖能力变化;电子显微镜观察细胞伪足,Transwell检测细胞的运动侵袭能力。结果免疫荧光显示ezrin和骨架蛋白表达于细胞质,且双色荧光证实两者存在共表达;高转移潜能细胞系SF7721,MHCC-I、MHCC97-H ezrin和骨架蛋白的表达明显高于低转移潜能细胞系SMMC-7721、 Hep3B、HepG2细胞(x2=13．277,P=0．010; x2=21．815,P<0．01)。β-肌动蛋白在高低转移潜能细胞系的表达差异无统计学意义。通过RNA干扰技术抑制ezrin蛋白表达后,SF7721和MHHC97-H的细胞的增殖和侵袭能力均显著下降。结论 ezrin和骨架蛋白的过表达与肝癌的转移潜能相关,通过下调ezrin的表达可明显抑制肝癌细胞系SMMC-7721和MHCC97-H细胞的增殖和运动侵袭能力。
Objective To investigate the role of membrane-associated cytoskeleton ezrin in the growth and metastasis of hepatocellular carcinoma. Methods Immunofluorescence, reverse transcription polymerase chain reaction (RT-PCR) and Western blot were used to detect the expression of ezrin and scaffold protein in different metastatic potential hepatocellular carcinoma cell lines. The high-metastatic potential SF7721 (SMMC-7721 transgenic cells stably expressing hepatocyte growth factor to obtain a high metastatic potential cell line) and MHCC97-H cell line were selected as experimental subjects, and the down-regulation of SF7721 and MHCC97-H cell lines by RNA interference The ezrin protein expression was observed and the changes of motility and invasion ability were observed. The changes of cell proliferation were detected by MTT assay. Cell pseudopodia were observed by electron microscope. Results Immunofluorescence showed that ezrin and scaffold proteins were expressed in the cytoplasm, and the two proteins were co-expressed by two-color fluorescence. The expression of SF7721, MHCC-I, MHCC97-H ezrin and scaffold proteins were significantly higher in the metastatic potential cell line than in the low metastatic potential cell line SMMC-7721, Hep3B, HepG2 cells (x2 = 13.277, P = 0.010; x2 = 21.815, P <0.01). There was no significant difference in the expression of β-actin between the cell lines of high and low metastatic potential. Inhibition of ezrin protein expression by RNA interference decreased the proliferation and invasion ability of SF7721 and MHHC97-H cells. Conclusion The overexpression of ezrin and scaffold proteins is related to the metastatic potential of HCC. The down-regulation of ezrin can significantly inhibit the proliferation and motility of hepatocellular carcinoma cell lines SMMC-7721 and MHCC97-H.