Ezrin调控肝癌细胞侵袭性伪足的形成促进肝细胞癌侵袭转移

Ezrin promotes invasion and metastasis of hepatocellular carcinoma by regulating invadopodia formation

目的探讨Ezrin在肝癌细胞侵袭性伪足形成过程中的作用。方法利用GEPIA数据库分析肝癌组织中Ezrin mRNA与伪足标志物Cortactin mRNA的表达相关性。将收集的105例肝细胞癌患者的组织标本制作成组织芯片,采用免疫组化检测Ezrin蛋白和伪足标志物Cortactin蛋白在肝癌组织中的表达,并分析二者的表达相关性。分析Ezrin的表达水平与肝癌患者临床病理特征及预后的关系。敲低和上调肝癌细胞中Ezrin的表达,采用Transwell实验观察Ezrin蛋白的表达变化对肝癌细胞侵袭迁移能力的影响,采用Western blot和细胞免疫荧光技术检测Ezrin蛋白表达变化对肝癌细胞侵袭性伪足形成能力的影响。结果生物信息学分析结果提示肝癌组织中Ezrin mRNA与伪足标志物Cortactin mRNA的表达呈正相关(r=0.420,P<0.01)。肝癌组织芯片免疫组化结果显示Ezrin蛋白与伪足标志物Cortactin蛋白的表达水平呈正相关(r=0.491,P<0.001)。患者临床资料结果显示,Ezrin蛋白的高表达与肝癌患者的血管侵犯率(P=0.016)及肿瘤相关死亡率(P=0.018)相关,且Ezrin蛋白高表达患者的术后5年生存率及术后5年无瘤生存率均较低(P<0.05)。细胞实验结果显示,上调Ezrin蛋白的表达可促进肝癌细胞的侵袭和迁移(P<0.05),且其高表达可促进肝癌细胞侵袭性伪足的形成(P<0.05);敲低Ezrin蛋白表达可抑制肝癌细胞侵袭性伪足的形成,并降低肝癌细胞的侵袭和迁移能力。结论Ezrin可通过调控肝癌细胞侵袭性伪足的形成增强肝癌细胞的侵袭、迁移能力,进而促进肝癌进展。

Objective To explore the role of ezrin in the formation of invadopodia in hepatocellular carcinoma(HCC) cells. Methods GEPIA database was used to analyze the expression correlation between ezrin and the invadopodia marker cortactin in HCC tissues at mRNA level. Tissue specimens obtained from 105 HCC patients were made into tissue microarrays, which were subsequently detected by immunohistochemistry to analyze the expression correlation of ezrin with cortactin at protein level. Moreover, the relationship between ezrin level and the clinicopathological features as well as the prognosis of HCC patients were analyzed. Finally, ezrin expression was knocked down or up-regulated in HCC cells, respectively, and Transwell assay was performed to observe the effects of ezrin expression changes on the invasion and migration of HCC cells;Western blotting and immunofluorescence assay were also adopted to detect the role of ezrin in the formation of invadopodia in HCC cells. Results Bioinformatics analysis revealed that the mRNA level of ezrin was positively correlated with that of cortactin in HCC tissues(r=0.420, P<0.01), and immunohistochemistry assay also confirmed the positive correlation between ezrin and cortactin expression at protein level(r= 0.491, P<0.001). Analysis of clinical data showed that high expression of ezrin protein was correlated with vascular invasion rate(P=0.016) and tumor-related mortality rate(P=0.018) in HCC patients, and patients with higher ezrin level had both lower 5-year postoperative survival rate and 5-year tumor-free survival rate(P<0.05). In cell experiments, up-regulation of ezrin promoted the invasion and metastasis(P<0.05), and facilitated the invadopodia formation of HCC cells(P<0.05). In contrast, knockdown of ezrin suppressed the invadopodia formation and inhibited the invasion and metastasis of HCC cells. Conclusion Ezrin enhances the invasive and migratory abilities of HCC cells by regulating invadopodia formation and thus promotes the progression of HCC.