Abstract 2005: Aberrant hedgehog signaling is responsible for the highly invasive behavior of a subpopulation of hepatoma cells
Abstract 2005: Aberrant hedgehog signaling is responsible for the highly invasive behavior of a subpopulation of hepatoma cells
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DOI：
10.1158/1538-7445.am2014-2005
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发表时间：
2014-10
期刊：
影响因子：
11.2
通讯作者：
Yahan Fan;Jia Ding;Jian Wu
中科院分区：
文献类型：
--
作者：
Yahan Fan;Jia Ding;Jian Wu
会议记录：2014 年 AACR 年会； 2014年4月5日至9日；加利福尼亚州圣地亚哥 背景：肝癌由细胞表面标志物、致瘤性、侵袭和转移能力方面的异质亚群组成。在我们之前的研究中，我们表明 CD133-/EpCAM- 肝癌亚群比其对应的亚群更具转移性；然而，其控制机制尚未探索（J Hepatol 2011；55：838-845）。本研究旨在阐明异常的hedgehog信号在转移发展中的重要性。方法：Huh-7 细胞经 FACS 富集为 CD133+/EpCAM+（双阳性，DP）和 CD133-/EpCAM-（双阴性，DN）亚群。双阴性细胞进一步进行转移细胞的Transwell选择（Transwell-select，TS）。在这些亚群中测定了转移率、基质金属蛋白酶 (MMP) 基因表达、上皮间质转化 (EMT) 标记、hedgehog 信号活性以及截短的 Gli1。结果：与 DN 和 DP 群体相比，TS 细胞表现出更高的转移活性，表现为 Transwell 侵袭率增加（105 vs. 68.7 和 58.0%，p<0.01），MMP1、2 和 9 基因的表达极度升高（36-3000 倍，p<0.05-0.001）。与 DN 和 DP 亚群相比，TS 细胞中 TGF-β1 基因表达增加了近 2 倍。免疫细胞化学显示，与 DP 细胞相比，TS 细胞失去了 E-钙粘蛋白，并且全部呈波形蛋白阳性。从 DP、DN 到 TS 亚群，Gli1 基因表达水平出现过渡性增加，这与通过 Western blot 分析和定量 RT-PCR 检测到的核组分中 Gli-2 和 Twist1 水平升高一致。此外，在高侵袭性肝癌细胞亚群中检测到了截短的 Gli1，它表明介导参与转移的分子的表达，并且在 TS 亚群中最高。然而，它没有出现在原代人肝细胞中。流式细胞术分析证实这些 TS 细胞在其子代中保持阴性细胞表面标记谱。结论：独特的 Transwell 选择细胞的高转移能力与 CD133 和 EpCAM 表面分子的阴性表达、显着的 EMT、增强的 hedgehog 活性和截短的 Gli1 的异常表达相关。截短的 Gli1 的出现可能是 Transwell 选择的双阴性亚群的高度侵袭性行为的原因。进一步探索异常hedgehog信号传导调节转移的分子机制将为肝癌转移的治疗干预建立分子靶点。引用格式：范雅涵，丁家，吴健。异常的刺猬信号传导导致肝癌细胞亚群的高度侵袭性行为。 [抽象的]。见：美国癌症研究协会第 105 届年会记录； 2014年4月5-9日；加利福尼亚州圣地亚哥。费城（宾夕法尼亚州）：AACR；癌症研究 2014;74(19 Suppl):摘要 nr 2005. doi:10.1158/1538-7445.AM2014-2005
Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Background: Hepatoma consists of heterogeneous subpopulations in terms of their cell surface markers, tumorigenicity, invasion and metastatic capability. In our previous study, we indicated that the CD133-/EpCAM- hepatoma subpopulation was more metastatic than its counterpart; however the controlling mechanisms are unexplored (J Hepatol 2011;55:838-845). The present study aims to delineate the significance of aberrant hedgehog signaling in the development of metastases. Methods: Huh-7 cells were FACS-enriched into CD133+/EpCAM+ (double positive, DP) and CD133-/EpCAM- (double negative, DN) subpopulations. The double negative cells further underwent Transwell-selection for metastatic cells (Transwell-selected, TS). The metastatic rate, matrix metalloproteinase (MMP) gene expression, epithelial mesenchymal transition (EMT) markers, and hedgehog signaling activities, as well as truncated Gli1 were determined in these subpopulations. Results: TS cells displayed much greater metastatic activity as evidenced by an increased Transwell invasion rate (105 vs. 68.7 and 58.0%, p<0.01), extremely elevated expression of MMP1, 2 and 9 genes (36-3000-fold, p<0.05-0.001) compared to DN and DP populations. There was a nearly 2-fold increase in TGF-β1 gene expression in TS cells compared to DN and DP subpopulations. TS cells lost E-cadherin and were all vimentin-positive as shown by immunocytochemistry in contrast to DP cells. There was a transitional increase in Gli1 gene expression levels from DP, DN to TS subpopulations, which is consistent with elevated Gli-2 and Twist1 levels in the nuclear fraction as detected by Western blot analysis and quantitative RT-PCR. Furthermore, truncated Gli1, which has been indicative of mediating expression of the molecules involved in metastasis, was detected in highly invasive hepatoma cell subpopulations and was the highest in the TS subpopulation; whereas, it did not appear in primary human hepatocytes. Flow cytometric analysis verified that these TS cells maintained a negative cell surface marker profile in their sub-passages. Conclusions: The highly metastatic capability of unique Transwell-selected cells is associated with negative expression of CD133 and EpCAM surface molecules, significant EMT, enhanced hedgehog activity and aberrant expression of truncated Gli1. The appearance of truncated Gli1 is probably responsible for highly invasive behavior of the transwell-selected double-negative subpopulation. Further exploration of molecular mechanisms of abnormal hedgehog signaling in modulating metastases will establish molecular targets for the therapeutic intervention of hepatoma metastases. Citation Format: Yahan Fan, Jia Ding, Jian Wu. Aberrant hedgehog signaling is responsible for the highly invasive behavior of a subpopulation of hepatoma cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2005. doi:10.1158/1538-7445.AM2014-2005