National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
Int J Biochem Cell Biol. 2009 Nov;41(11):2163-72. doi: 10.1016/j.biocel.2009.03.014. Epub 2009 Apr 5.
Tumor angiogenesis, induced by tumor-secreted pro-angiogenic factors, is an essential process for cancer development and metastasis. CD146 is identified as an endothelial cell adhesion molecule and implicated in blood vessel formation, however, its exact role in angiogenesis, particularly tumor angiogenesis, and its potential function of mediating downstream signaling are still unclear. In present study, we evidenced that silencing endogenous endothelial CD146 by RNAi significantly impaired hepatocarcinoma cell secretions-promoted tubular morphogenesis and -enhanced motility of endothelial cells. Biochemical studies revealed that CD146 was required for the activation of p38/IKK/NF kappaB signaling cascade and up-regulation of NF kappaB downstream pro-angiogenic genes, notably IL-8, ICAM-1 and MMP9, in response to tumor secretions. Interestingly, specific anti-CD146 mAb AA98, which bound a conformational epitope depending on C452-C499 disulfide bond, could abrogate NF kappaB activation and tumor angiogenesis, whereas another anti-CD146 mAb AA1 recognizing a linear epitope containing aa50-54 did not have such effects. Further structure-function analysis identified that C452-C499 disulfide bond within the fifth extracellular Ig domain was indispensible for CD146-mediated signaling and tube formation. Moreover, dimerization of CD146, which was enhanced by tumor secretions and suppressed by AA98 but not AA1, also relied on C452 and C499. Together, this study for the first time uncovered the pro-angiogenic role of CD146 and also pinpointed the key structural basis responsible for its signaling function and dimerization. These findings also suggested that CD146 might serve as not just a cell adhesion molecule but also a membrane signal receptor in tumor-induced angiogenesis.
肿瘤血管生成是肿瘤分泌的促血管生成因子诱导的，是癌症发展和转移的必要过程。CD146 被鉴定为内皮细胞黏附分子，参与血管形成，但它在血管生成，特别是肿瘤血管生成中的确切作用及其介导下游信号的潜在功能仍不清楚。在本研究中，我们通过 RNAi 沉默内源性内皮细胞 CD146，明显抑制肝癌细胞分泌促进管状形态发生和增强内皮细胞迁移的作用。生化研究表明，CD146 是肿瘤分泌物激活 p38/IKK/NF-κB 信号级联反应和上调 NF-κB 下游促血管生成基因所必需的，特别是白细胞介素 8、细胞间黏附分子 1 和基质金属蛋白酶 9。有趣的是，特异性抗 CD146 mAb AA98 与依赖于 C452-C499 二硫键的构象表位结合，可阻断 NF-κB 激活和肿瘤血管生成，而另一种识别包含 aa50-54 的线性表位的抗 CD146 mAb AA1 则没有这种作用。进一步的结构功能分析表明，第五个细胞外 Ig 结构域内的 C452-C499 二硫键对于 CD146 介导的信号转导和管状形成是必不可少的。此外，肿瘤分泌物增强而 AA98 但不是 AA1 抑制的 CD146 二聚化也依赖于 C452 和 C499。总之，这项研究首次揭示了 CD146 的促血管生成作用，并确定了负责其信号功能和二聚化的关键结构基础。这些发现还表明，CD146 不仅可以作为细胞黏附分子，还可以作为肿瘤诱导血管生成中的膜信号受体。
