UNASSIGNED: The expression of ANCR in BCC tissues and cells was detected by qRT-PCR. Proliferation, invasion, migration and apoptosis of ANCR overexpressed or knock down TE354.T and A431 cells were examined by CCK8, transwell assay, wound healing assay and flow cytometry analysis, respectively. Western blot was performed to measure the expression of apoptosis-related proteins (BAX, BCL2 and Cleaved-caspase3), epithelial-mesenchymal transformation-related proteins (E-cadherin, N-cadherin, vimentin and β-catenin), and Hedgehog-pathway-related proteins (PTCH, GLI1 and SMO). RNA pull-down assay was used to analyze the relationship between ANCR and PTCH. The effect of ANCR on BCC growth in vivo was analyzed using xenograft model. TUNEL assay was used to determine the cell apoptosis.
UNASSIGNED: ANCR and Hedgehog pathway were more highly expressed in BCC tissues than in adjacent normal tissues. ANCR overexpression substantially promoted BCC cell proliferation, invasion, and migration, inhibited apoptosis, and up-regulated BCL2 and decreased the expression of BAX and Cleaved-caspase3 proteins. Additionally, the upregulation of N-cadherin, vimentin, β-catenin, PTCH, GLI1, and SMO expression, and downregulation of E-cadherin expression were observed after ANCR overexpression. Moreover, ANCR knockdown had the opposite effects. An RNA pull-down assay further revealed that ANCR is specifically bound to PTCH. In vivo experiments also showed that ANCR overexpression significantly increased tumor growth and decreased apoptosis, which was reversed by cyclopamine, a specific inhibitor of the Hedgehog signaling pathway.
UNASSIGNED: ANCR activates the Hedgehog signaling pathway by binding to PTCH, thereby promoting BCC progression; accordingly, ANCR could be a candidate therapeutic target in BCC.
■通过qRT-PCR检测ANCR在BCC组织和细胞中的表达。扩散,入侵,过表达或敲低TE354的ANCR的迁移和凋亡。T和A431细胞通过CCK8,transwell试验,伤口愈合试验和流式细胞术分析,分别。进行Westernblot以测量凋亡相关蛋白的表达(BAX,BCL2和Cleaved-caspase3),上皮-间质转化相关蛋白(E-cadherin,N-钙黏着蛋白,波形蛋白和β-连环蛋白),和Hedgehog通路相关蛋白(PTCH,GLI1和SMO)。采用RNA下拉法分析ANCR与PTCH的关系。使用异种移植模型分析ANCR对体内BCC生长的影响。TUNEL测定用于确定细胞凋亡。
■ANCR和Hedgehog通路在BCC组织中的表达高于邻近正常组织。ANCR过表达显著促进BCC细胞增殖,入侵,和移民,抑制细胞凋亡,并上调BCL2并降低BAX和Cleaved-caspase3蛋白的表达。此外,N-钙粘蛋白的上调,波形蛋白,β-连环蛋白,PTCH,GLI1和SMO表达式,ANCR过表达后观察到E-cadherin表达下调。此外,ANCR击倒具有相反的效果。RNA下拉测定进一步揭示ANCR与PTCH特异性结合。体内实验还表明,ANCR过表达显着增加了肿瘤的生长,减少了细胞凋亡,被环巴明逆转了,Hedgehog信号通路的特异性抑制剂。
■ANCR通过与PTCH结合激活Hedgehog信号通路,从而促进BCC进展;因此,ANCR可能是BCC的候选治疗靶标。