TFCP2/YAP转录复合体调控蛋白CPE对肝细胞肝癌的诊断价值

摘要

目的 探讨转录因子CP2 (TFCP2) /Yes相关蛋白 (YAP) 转录复合体调控蛋白——羧肽酶E (CPE) 在肝细胞肝癌 (HCC) 中的诊断价值.方法 采用酶联免疫吸附试验测定147例HCC患者、42例乙型肝炎患者、29例胃癌患者及100名健康志愿者 (正常对照组) 血清CPE水平.采用实时荧光定量聚合酶链反应 (PCR) 检测CPE mRNA水平, 免疫印迹法检测CPE、TFCP2及YAP蛋白表达, 双荧光报告实验检测CPE启动子基因的活性.CPE与甲胎蛋白 (AFP) 、丙氨酸氨基转移酶 (ALT) 、天门冬氨酸氨基转移酶 (AST) 的相关性采用Spearman等级相关分析.采用受试者工作特征 (ROC) 曲线评价CPE和AFP诊断HCC的价值.结果过表达YAP或TFCP2可促进CPE mRNA及蛋白的表达, 而YAP和TFCP2共同过表达可明显促进CPE mRNA和蛋白的表达.YAP或TFCP2的敲减可抑制CPE mRNA和蛋白表达, 而YAP及TFCP2共同敲减可明显抑制CPE mRNA和蛋白的表达.CPE启动子区域存在TFCP2结合基序, 突变该基序后, TFCP2和YAP对CPE启动子的调控作用丧失.HCC组血清CPE水平明显高于乙型肝炎组、胃癌组和正常对照组 (P<0.01) , 而乙型肝炎组、胃癌组和正常对照组之间差异无统计学意义 (P>0.05) .CPE与AFP、ALT、AST均呈正相关 (r值分别为0.644、0.454、0.351, P值分别为0.000、0.000、0.001) .ROC曲线分析显示, CPE诊断HCC的曲线下面积 (AUC) 为0.937, 明显高于AFP的AUC (0.679) .结论 CPE在HCC的诊断中具有较高的价值, 可作为HCC的血清学诊断标志物.%Objective To investigate the role of transcription factor CP2 (TFCP2) /Yes-associated protein (YAP) transcriptional complex co-regulated protein carboxypeptidase E (CPE) for the diagnosis of hepatocellular carcinoma (HCC) . Methods The levels of serum CPE in 147 HCC patients, 42 hepatitis B patients, 29 gastric cancer patients and 100 healthy subjects (healthy control group) were determined by enzyme-linked immunosorbent assay. Real-time ?uorescence quantitation polymerase chain reaction (PCR) was used to determine the expression of CPE mRNA. Western blotting was used to determine the expression of CPE, TFCP2 and YAP. The dual-luciferase reporter test was used to determine the activity of CPE promoter. The correlations between CPE and other indicators such as alpha-fetoprotein (AFP) , alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were analyzed using Spearman correlation analysis. Receiver operating characteristic (ROC) curve was used to compare the ef?ciency of CPE and AFP for the diagnosis of HCC. Results The overexpression of YAP or TFCP2 stimulated the expression of CPE mRNA and protein, and the overexpression of YAP and TFCP2 stimulated the expression of CPE mRNA and protein. The knockdown of YAP or TFCP2 inhibited the expression of CPE mRNA and protein, and the knockdown of YAP and TFCP2 inhibited the expression of CPE mRNA and protein. A TFCP2-binding-motif was existed in CPE promoter region. After mutation, TFCP2 and YAP lost the regulation function on CPE promoter.The level of serum CPE was higher in HCC group than those in healthy control, hepatitis B and gastric cancer groups (P<0.01) , and there was no statistical signi?cance among healthy control, hepatitis B and gastric cancer groups (P>0.05) . The level of serum CPE was positively correlated with the levels of AFP, ALT and AST (r=0.644, 0.454 and 0.351, P=0.000, 0.000 and 0.001, respectively) . The area under ROC curve (AUC) of CPE for the diagnosis of HCC was 0.937, which is better than that of AFP (0.679) . Conclusions CPE plays a role in the diagnosis of HCC, and CPE may be a diagnostic biomarker for HCC.