Zwint高表达对肝癌细胞增殖和肝癌肝移植预后的影响

摘要

目的 探讨Zw10结合因子(Zwint)在原发性肝癌(肝癌)中的表达情况及其对肝癌肝移植预后的影响.方法 收集50例肝癌肝移植受体的肝癌组织、癌旁组织及临床资料.采用实时荧光定量聚合酶链反应(PCR)、蛋白免疫印迹(Western Blot)法和免疫组织化学(免疫组化)法,分别比较20例肝癌肝移植受体的20对肝癌组织及癌旁组织中Zwint信使核糖核酸(mRNA)表达情况和Zwint蛋白的表达水平;选取两个成功干扰Zwint表达的肝癌细胞系HepG-2(si-Zwint-1组和si-Zwint-2组),以空白对照作为si-NC组,采用细胞计数试剂盒(CCK)-8实验、平板克隆实验和细胞周期实验,比较各组细胞的增殖能力和细胞周期;采用Western Blot法和免疫组化法,分析肝癌组织和肝癌细胞中Zwint与细胞周期蛋白D1(cyclin D1)表达的一致性;以Zwint蛋白表达水平的中位数为分割点,将入组病例分高表达组(22例)和低表达组(28例),分析Zwint蛋白表达水平与肝癌肝移植受体临床特征、总体存活率及无复发存活率的关系.结果 实时荧光定量PCR结果提示肝癌组织中Zwint mRNA表达水平高于癌旁组织(P=0.03).Western Blot及免疫组化结果均提示肝癌组织中Zwint蛋白的表达水平高于癌旁组织(均为P<0.05).干扰肝癌细胞系HepG-2的Zwint基因后,CCK-8及平板克隆实验显示细胞增殖能力明显减弱(均为P<0.01),细胞周期阻滞于G1期(均为P<0.05).Zwint蛋白表达水平与肿瘤直径和肿瘤、淋巴、转移(TNM)分期密切相关(均为P<0.05).Zwint高表达组肝癌肝移植受体的总体存活率低于低表达组(P=0.02).结论 Zwint高表达于肝癌组织,其通过调节细胞周期促进肝癌细胞增殖,且Zwint表达水平与肝癌肝移植后预后呈负相关.%Objective To investigate the expression of zeste white 10 interactor (Zwint) in primary hepatocellular carcinoma (HCC) and its effect on the prognosis of liver transplantation for HCC. Methods HCC tissues, paracancerous tissues and clinical data of 50 liver transplant recipients for HCC were collected. The expression levels of Zwint messenger RNA (mRNA) and Zwint protein in 20 pairs of HCC tissues and paracancerous tissues of 20 liver transplant recipients for HCC were compared using real-time fluorescence quantitative polymerase chain reaction (PCR), Western Blot and immunohistochemistry (IHC). Two HCC cell lines HepG-2 which interfered with the expression of Zwint successfully were selected as si-Zwint-1 group and si-Zwint-2 group, and the blank control was taken as si-NC group. The cell proliferation and cell cycle of various groups were compared using cell counting kit (CCK) -8 experiment, flat-cloning assay and cell cycle experiment. The consistency of the expression of Zwint and cyclin D1 in HCC tissues and cells was analyzed using Western Blot and IHC. The enrolled patients were divided into high expression group (22 cases) and low expression group (28 cases) based on the median of Zwint protein expression level, and the relationship of the expression level of Zwint protein and clinical characteristics, overall survival rate and disease free survival rate of liver transplant recipients for HCC was analyzed. Results The results of real-time fluorescence quantitative PCR showed that the expression level of Zwint mRNA in HCC tissues was higher than that of paracancerous tissues (P=0.03). The results of Western Blot and IHC showed that the expression level of Zwint protein in HCC tissues was higher than that of paracancerous tissues(both P<0.05).After the Zwint gene of HCC cell line HepG-2 was interfered,CCK-8 and flat-cloning assay showed that the cell proliferation potential was significantly weakened (all P<0.01), and the cell cycle arrested at stage G1(all P<0.05). The expression level of Zwint protein was closely related to tumor diameter and tumor, node, metastasis (TNM) staging (all P<0.05). The overall survival rate of liver transplant recipients for HCC in the high Zwint expression group was lower than that of the low expression group (P=0.02). Conclusions Zwint is highly expressed in HCC tissues, and it can promote the proliferation of HCC cells through regulating cell cycle. The expression level of Zwint is negatively correlated with the prognosis of liver transplantation for HCC.