Bhagwat Shripad V, Petrovic Nenad, Okamoto Yasuhiro, Shapiro Linda H
Departments of Pathology, and Hematology/Oncology, St Jude Children's Research Hospital, Memphis, TN, USA.
Blood. 2003 Mar 1;101(5):1818-26. doi: 10.1182/blood-2002-05-1422. Epub 2002 Oct 24.
Angiogenesis, the formation of new blood vessels, is a critical step for tumor growth and metastasis and an integral component of the pathologic inflammatory response in arthritis and the proliferative retinopathies. The CD13/aminopeptidase N (CD13/APN) metalloprotease is an important regulator of angiogenesis where its expression on activated blood vessels is induced by angiogenic signals. Here, we show that cytokine induction of CD13/APN in endothelial cells is regulated by distinct Ras effector pathways involving Ras/mitogen-activated protein kinase (MAPK) or PI-3K. Signals transduced by activated Ras, Raf, and mitogen-induced extracellular kinase (MEK) stimulate transcription from the CD13/APN proximal promoter. Inhibition of these pathways and extracellular signal-regulated serine/threonine kinase (ERK-2) and PI-3K by expression of dominant-negative proteins or chemical inhibitors prevented induction of CD13/APN transcription in response to basic fibroblast growth factor (bFGF). We show that Ras-induced signal transduction is required for growth factor-induced angiogenesis, because inhibition of downstream mediators of Ras signaling (MEK or PI-3K) abrogated endothelial cell migration, invasion, and morphogenesis in vitro. Reintroduction of CD13/APN, a shared downstream target of these pathways, overrode the suppressive effect of these inhibitors and restored the function of endothelial cells in migration/invasion and capillary morphogenesis assays. Similarly, inhibition of MEK abrogated cell invasion and the formation of endothelial-lined capillaries in vivo, which was effectively rescued by addition of exogenous CD13/APN protein. These studies provide strong evidence that CD13/APN is an important target of Ras signaling in angiogenesis and is a limiting factor in angiogenic progression.
血管生成，即新血管的形成，是肿瘤生长和转移的关键步骤，也是关节炎和增殖性视网膜病变病理炎症反应的一个重要组成部分。CD13/氨肽酶N（CD13/APN）金属蛋白酶是血管生成的重要调节因子，其在活化血管上的表达由血管生成信号诱导。在此，我们表明内皮细胞中CD13/APN的细胞因子诱导受涉及Ras/丝裂原活化蛋白激酶（MAPK）或PI-3K的不同Ras效应途径调控。活化的Ras、Raf和丝裂原诱导的细胞外激酶（MEK）转导的信号刺激CD13/APN近端启动子的转录。通过表达显性负性蛋白或化学抑制剂抑制这些途径以及细胞外信号调节丝氨酸/苏氨酸激酶（ERK-2）和PI-3K，可阻止对碱性成纤维细胞生长因子（bFGF）的反应中CD13/APN转录的诱导。我们表明Ras诱导的信号转导是生长因子诱导的血管生成所必需的，因为抑制Ras信号的下游介质（MEK或PI-3K）可消除体外内皮细胞的迁移、侵袭和形态发生。重新引入这些途径的共同下游靶点CD13/APN，可克服这些抑制剂的抑制作用，并在迁移/侵袭和毛细血管形态发生试验中恢复内皮细胞的功能。同样，抑制MEK可消除体内细胞侵袭和内皮衬里毛细血管的形成，添加外源性CD13/APN蛋白可有效挽救这种情况。这些研究提供了强有力的证据，表明CD13/APN是血管生成中Ras信号的重要靶点，并且是血管生成进展中的一个限制因素。
