Jiang Rui, Niu Xiaoshuang, Huang Yuxiang, Wang Xiaosheng
Department of Radiation Oncology, Fudan University Shanghai Cancer Center, Shanghai 2000031, China.
Department of Radiation Oncology, Fudan University Shanghai Cancer Center, Shanghai 2000031, China
Acta Biochim Biophys Sin (Shanghai). 2016 Mar;48(3):229-37. doi: 10.1093/abbs/gmv134. Epub 2016 Feb 4.
Nasopharyngeal carcinoma (NPC) is one of the most common malignant tumors with poor prognosis and recurrence in South China. The hard eradication of NPC in clinic is predominantly due to cancer stem cells (CSCs). Increasing evidence revealed that the aberrant activation of Wnt/β-catenin was positively correlated with the produce of CSCs. To further investigate the effect of β-catenin on CSCs and tumorigenesis in NPC, a CNE2 cell line (pLKO.1-sh-β-catenin-CNE2) with stably suppressed expression of β-catenin was used in this study. The expressions of biomarkers in CSCs including c-myc, Nanog, Oct3/4, Sox2, EpCAM as well as adhesion-related proteins like E-cadherin and vimentin were analyzed by western blot analysis and immunofluorescent staining. The proliferation and migration abilities were investigated by MTT assay and Transwell assay, respectively. Cell cycle was analyzed by flow cytometry. Finally, xenograft was performed to determine the effect of β-catenin on oncogenesis in vivo. Results showed that the expressions of c-myc, Nanog, Oct3/4, Sox2, and EpCAM were all decreased in pLKO.1-sh-β-catenin-CNE2 cells. It was also found that vimentin was downregulated, while E-cadherin was upregulated. Results of MTT and Transwell assays suggested that the proliferation and migration abilities were impaired by silencing of β-catenin, and more cells were arrested in G1 phase when compared with the control. In vivo study indicated that the tumor growth was markedly suppressed in experimental group. Based on current findings, β-catenin may function as an essential protein for the maintenance of migration and proliferation abilities of NPC cells, and a complicated network consisting of c-myc, Nanog, Oct3/4, Sox2, EpCAM, E-cadherin, vimentin, and β-catenin may be involved in the inherent regulation mechanisms.
鼻咽癌(NPC)是中国南方最常见的恶性肿瘤之一,预后较差且易复发。临床上鼻咽癌难以根治主要归因于癌症干细胞(CSCs)。越来越多的证据表明,Wnt/β-连环蛋白的异常激活与癌症干细胞的产生呈正相关。为了进一步研究β-连环蛋白对鼻咽癌中癌症干细胞及肿瘤发生的影响,本研究使用了β-连环蛋白表达被稳定抑制的CNE2细胞系(pLKO.1-sh-β-catenin-CNE2)。通过蛋白质免疫印迹分析和免疫荧光染色分析了癌症干细胞中生物标志物c-myc、Nanog、Oct3/4、Sox2、EpCAM的表达以及E-钙黏蛋白和波形蛋白等黏附相关蛋白的表达。分别通过MTT法和Transwell法研究增殖和迁移能力。通过流式细胞术分析细胞周期。最后,进行异种移植以确定β-连环蛋白对体内肿瘤发生的影响。结果显示,pLKO.1-sh-β-catenin-CNE2细胞中c-myc、Nanog、Oct3/4、Sox2和EpCAM的表达均降低。还发现波形蛋白下调,而E-钙黏蛋白上调。MTT和Transwell试验结果表明,β-连环蛋白的沉默损害了增殖和迁移能力,与对照组相比,更多细胞停滞在G1期。体内研究表明,实验组的肿瘤生长明显受到抑制。基于目前的研究结果,β-连环蛋白可能是维持鼻咽癌细胞迁移和增殖能力的必需蛋白,由c-myc、Nanog、Oct3/4、Sox2、EpCAM、E-钙黏蛋白、波形蛋白和β-连环蛋白组成的复杂网络可能参与了内在调控机制。