Mehta Veela B, Zhou Yu, Radulescu Andrei, Besner Gail E
Department of Pediatric Surgery, The Research Institute at Nationwide Children's Hospital, The Ohio State University College of Medicine, Columbus, OH 43205, USA.
Growth Factors. 2008 Dec;26(6):301-15. doi: 10.1080/08977190802393596.
Heparin-binding EGF-like growth factor (HB-EGF) is a member of the epidermal growth factor (EGF) family of ligands that is expressed by many cell types including endothelial cells. We have previously shown that HB-EGF stimulates angiogenesis in vitro in human umbilical vein endothelial cells (HUVEC). Nitric oxide (NO) derived from endothelial nitric oxide synthase (eNOS) is an important regulator of angiogenesis. However, the role of HB-EGF in regulation of eNOS has not yet been investigated. Whether HB-EGF-induced endothelial cell migration and vascular network formation are mediated via production of NO from eNOS is also unknown. To address these questions, we stimulated HUVEC with HB-EGF and evaluated the expression of eNOS at the mRNA and protein levels. HB-EGF significantly upregulated expression of eNOS mRNA, stimulated eNOS protein production, and increased NO release from HUVEC. HB-EGF phosphorylated eNOS in a phosphatidylinositol 3-kinase (PI3K) dependent fashion, and stimulated in vitro angiogenesis. eNOS siRNA inhibited HB-EGF-stimulated HUVEC migration in a scratch assay. NG-nitro-L-arginine-methyl-ester (L-NAME) and L-N5-(1-lminoethyl)ornithine,dihydochloride (L-NIO) (specific inhibitors of eNOS) also abolished HB-EGF-induced HUVEC migration and angiogenesis. More importantly, we found that HB-EGF also promotes angiogenesis in vivo in the Marigel plug assay. Lastly, inhibition of the p38 MAPK pathway enhanced HB-EGF-induced EC migration and angiogenesis. We conclude that HB-EGF, through its interaction with EGF receptors (EGFR), stimulates eNOS activation and NO production via a PI3K-dependent pathway. Thus, activation of eNOS appears to be one of the key signaling pathways necessary for HB-EGF mediated angiogenesis. These novel findings highlight an important role for HB-EGF as a regulator of endothelial cell function.
肝素结合表皮生长因子样生长因子（HB-EGF）是表皮生长因子（EGF）配体家族的成员，由包括内皮细胞在内的多种细胞类型表达。我们之前已经表明，HB-EGF在体外可刺激人脐静脉内皮细胞（HUVEC）的血管生成。内皮型一氧化氮合酶（eNOS）产生的一氧化氮（NO）是血管生成的重要调节因子。然而，HB-EGF在eNOS调节中的作用尚未得到研究。HB-EGF诱导的内皮细胞迁移和血管网络形成是否通过eNOS产生NO介导也尚不清楚。为了解决这些问题，我们用HB-EGF刺激HUVEC，并在mRNA和蛋白质水平评估eNOS的表达。HB-EGF显著上调eNOS mRNA的表达，刺激eNOS蛋白的产生，并增加HUVEC中NO的释放。HB-EGF以磷脂酰肌醇3-激酶（PI3K）依赖的方式使eNOS磷酸化，并刺激体外血管生成。在划痕试验中，eNOS siRNA抑制HB-EGF刺激的HUVEC迁移。NG-硝基-L-精氨酸甲酯（L-NAME）和L-N5-（1-亚氨基乙基）鸟氨酸二盐酸盐（L-NIO）（eNOS的特异性抑制剂）也消除了HB-EGF诱导的HUVEC迁移和血管生成。更重要的是，我们发现在基质胶植入试验中，HB-EGF在体内也促进血管生成。最后，抑制p38丝裂原活化蛋白激酶（MAPK）途径增强了HB-EGF诱导的内皮细胞迁移和血管生成。我们得出结论，HB-EGF通过与表皮生长因子受体（EGFR）相互作用，经由PI3K依赖的途径刺激eNOS活化和NO产生。因此，eNOS的活化似乎是HB-EGF介导血管生成所必需的关键信号通路之一。这些新发现突出了HB-EGF作为内皮细胞功能调节因子的重要作用。
