背景:肝细胞癌(HCC)是全球最常见的恶性肿瘤之一,因为其进展迅速,转移或复发的发生率很高。越来越多的证据表明,表达CD58的肿瘤细胞与各种癌症的发展有关。本研究旨在揭示CD58在HCC进展中的功能意义和潜在机制。
方法:免疫组织化学染色(IHC),免疫印迹法检测CD58在肝癌组织和细胞中的表达。通过ELISA评估细胞上清液和血清中sCD58(CD58的可溶形式)的水平。CCK-8,集落形成,和异种移植试验用于检测CD58对体外和体内增殖的功能。进行Transwell测定和球体形成测定以评估CD58和sCD58对HCC细胞转移和自我更新能力的影响。西方印迹,免疫荧光(IF),TOP/FOPFlash报告基因测定,通过亚细胞分级分离实验研究肝癌细胞中CD58/sCD58与AKT/GSK-3β/β-catenin轴的分子调控。
结果:CD58在HCC组织中显著上调。CD58表达的升高与更多的卫星灶和血管浸润相关,肝癌患者的无瘤生存率和总生存率较差。与健康个体相比,肝癌患者血清中的sCD58水平较高。功能上,CD58在体外和体内促进HCC细胞的增殖。同时,CD58和sCD58诱导转移,体外肝癌细胞的自我更新和多能性。机械上,CD58通过增加AKT或GSK3β信号的磷酸化激活AKT/GSK-3β/β-catenin信号通路,促进Wnt/β-catenin靶蛋白的表达和TCF/LEF介导的转录活性。此外,AKT激活剂SC-79或抑制剂LY294002消除了CD58沉默对增殖的抑制作用,转移,和肝癌细胞的干性。
结论:综合来看,CD58通过激活AKT/GSK-3β/β-catenin通路促进HCC进展和转移,提示CD58是HCC的一个新的预后生物标志物和治疗靶点。
Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies worldwide because of rapid progression and high incidence of metastasis or recurrence. Accumulating evidence shows that CD58-expressing tumor cell is implicated in development of various cancers. The present study aimed to reveal the functional significance of CD58 in HCC progression and the underlying mechanisms.
Immunohistochemical staining (IHC), and western blotting were used to detect the expression of CD58 in HCC tissues and cells. The levels of sCD58 (a soluble form of CD58) in the cell supernatants and serum were assessed by ELISA. CCK-8, colony formation, and xenograft assays were used to detect the function of CD58 on proliferation in vitro and in vivo. Transwell assay and sphere formation assay were performed to evaluate the effect of CD58 and sCD58 on metastasis and self-renewal ability of HCC cells. Western blotting, immunofluorescence (IF), TOP/FOP Flash reporter assay, and subcellular fractionation assay were conducted to investigate the molecular regulation between CD58/sCD58 and AKT/GSK-3β/β-catenin axis in HCC cells.
CD58 was significantly upregulated in HCC tissues. Elevation of CD58 expression correlated with more satellite foci and vascular invasion, and poorer tumor-free and overall survival in HCC patients. Higher sCD58 levels were in HCC patients\' serum compared to healthy individuals. Functionally, CD58 promotes the proliferation of HCC cells in vitro and in vivo. Meanwhile, CD58 and sCD58 induce metastasis, self-renewal and pluripotency in HCC cells in vitro. Mechanistically, CD58 activates the AKT/GSK-3β/β-catenin signaling pathway by increasing phosphorylation of AKT or GSK3β signaling, promoting expression of Wnt/β-catenin target proteins and TCF/LEF-mediated transcriptional activity. Furthermore, AKT activator SC-79 or inhibitor LY294002 abolished the inhibitory effect of CD58 silencing on the proliferation, metastasis, and stemness of HCC cells.
Taken together, CD58 promotes HCC progression and metastasis via activating the AKT/GSK-3β/β-catenin pathway, suggesting that CD58 is a novel prognostic biomarker and therapeutic target for HCC.