Department of Pathology, Xiangya Medical College, Ministry of Health of China, Xiangya Hospital, Central South University, Changsha 410078, China.
Dis Esophagus. 2010 Feb;23(2):175-84. doi: 10.1111/j.1442-2050.2009.01001.x. Epub 2009 Jul 31.
beta-catenin has emerged as a key regulator of Wnt signaling pathway, which plays an important role in the development and progression of various cancers. Its accumulation in nucleus of the esophagus squamous epithelium might be the crucial step for the carcinogenesis of esophageal squamous cell carcinoma (ESCC). To detect the proteins correlated with beta-catenin function, we used the established cell lines of pGen-3-con (Eca109 cells transfected by control vector) and pGen-3-CTNNB1 (Eca109 cells transfected by beta-catenin siRNA) as cell models for further analysis. Two-dimensional gel electrophoresis technology was performed to separate the proteins of pGen-3-con and pGen-3-CTNNB1 cell lines, respectively. The differential protein spots were analyzed by software analysis, subjected to in-gel digestion, and identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Consequently, 13 differentially expressed proteins between the two cell lines were identified, of which 14-3-3sigma, prohibitin, and nm23-H1 were further verified by western blotting and quantitative real-time reverse transcriptase-polymerase chain reaction. Then, the tissue microarray and immunohistochemical analysis were employed to research their relationship in ESCC and their corresponding normal mucosa tissues. The upregulation of prohibitin or the downregulation of 14-3-3sigma and nm23-H1 proteins was significantly associated with the proliferation, invasion depth, and lymph node metastasis of ESCC. There were statistically significant correlations between the expression of beta-catenin and the three proteins. The results presented here might provide potential protein markers to elucidate the mechanism of beta-catenin-mediated biologic characteristics for ESCC.
β-连环蛋白已成为 Wnt 信号通路的关键调节因子，该通路在各种癌症的发生和发展中起着重要作用。其在食管鳞状上皮细胞核中的积累可能是食管鳞状细胞癌（ESCC）发生癌变的关键步骤。为了检测与β-连环蛋白功能相关的蛋白质，我们使用已建立的 pGen-3-con（用对照载体转染的 Eca109 细胞）和 pGen-3-CTNNB1（用β-连环蛋白 siRNA 转染的 Eca109 细胞）细胞系作为细胞模型进行进一步分析。分别使用二维凝胶电泳技术分离 pGen-3-con 和 pGen-3-CTNNB1 细胞系的蛋白质。通过软件分析对差异蛋白斑点进行分析，进行胶内消化，并通过基质辅助激光解吸/电离飞行时间质谱进行鉴定。结果，在两种细胞系之间鉴定出 13 种差异表达的蛋白质，其中 14-3-3sigma、 prohibitin 和 nm23-H1 进一步通过 Western blot 和实时定量逆转录聚合酶链反应进行验证。然后，使用组织微阵列和免疫组织化学分析研究它们在 ESCC 及其相应正常黏膜组织中的关系。 prohibitin 的上调或 14-3-3sigma 和 nm23-H1 蛋白的下调与 ESCC 的增殖、浸润深度和淋巴结转移显著相关。β-连环蛋白的表达与这三种蛋白质之间存在统计学显著相关性。这里呈现的结果可能为阐明β-连环蛋白介导的 ESCC 生物学特性的机制提供潜在的蛋白质标志物。
