Dept. of Pharmacology and Toxicology, Medical College of Georgia, Georgia Health Sciences University, 1120 15th St., Augusta, GA 30912, USA.
Am J Physiol Cell Physiol. 2011 May;300(5):C979-88. doi: 10.1152/ajpcell.00368.2010. Epub 2011 Feb 9.
Nitric oxide (NO) mediates endothelial angiogenesis via inducing the expression of integrin α(v)β(3). During angiogenesis, endothelial cells adhere to and migrate into the extracellular matrix through integrins. Collagen IV binds to integrin α(v)β(3), leading to integrin activation, which affects a number of signaling processes in endothelial cells. In the present study, we evaluated the role of collagen IV in NO-induced angiogenesis. We found that NO donor 2,2'-(hydroxynitrosohydrazino)bis-ethanamine (NOC-18) causes increases in collagen IV mRNA and protein in lung endothelial cells and collagen IV release into the medium. Addition of collagen IV into the coating of endothelial culture increases endothelial monolayer wound repair, proliferation, and tube formation. Inhibition of collagen IV synthesis using gene silencing attenuates NOC-18-induced increases in monolayer wound repair, cell proliferation, and tube formation as well as in the phosphorylation of focal adhesion kinase (FAK). Integrin blocking antibody LM609 prevents NOC-18-induced increase in endothelial monolayer wound repair. Inhibition of protein kinase G (PKG) using the specific PKG inhibitor KT5823 or PKG small interfering RNA prevents NOC-18-induced increases in collagen IV protein and mRNA and endothelial angiogenesis. Together, these results indicate that NO promotes collagen IV synthesis via a PKG signaling pathway and that the increase in collagen IV synthesis contributes to NO-induced angiogenesis of lung endothelial cells through integrin-FAK signaling. Manipulation of collagen IV could be a novel approach for the prevention and treatment of diseases such as alveolar capillary dysplasia, severe pulmonary arterial hypertension, and tumor invasion.
一氧化氮（NO）通过诱导整合素α(v)β(3)的表达来介导内皮细胞血管生成。在血管生成过程中，内皮细胞通过整合素黏附并迁移到细胞外基质中。胶原 IV 与整合素 α(v)β(3)结合，导致整合素激活，从而影响内皮细胞中的许多信号转导过程。在本研究中，我们评估了胶原 IV 在 NO 诱导的血管生成中的作用。我们发现，NO 供体 2,2'-(羟亚硝基羟氨基)双乙胺（NOC-18）导致肺内皮细胞中胶原 IV mRNA 和蛋白增加，并将胶原 IV 释放到培养基中。将胶原 IV 添加到内皮细胞培养物的涂层中会增加内皮单层伤口修复、增殖和管状形成。使用基因沉默抑制胶原 IV 合成会减弱 NOC-18 诱导的单层伤口修复、细胞增殖和管状形成以及粘着斑激酶（FAK）的磷酸化增加。整合素阻断抗体 LM609 可阻止 NOC-18 诱导的内皮单层伤口修复增加。使用特异性蛋白激酶 G（PKG）抑制剂 KT5823 或 PKG 小干扰 RNA 抑制 PKG 可防止 NOC-18 诱导的胶原 IV 蛋白和 mRNA 增加以及内皮血管生成。综上所述，这些结果表明，NO 通过 PKG 信号通路促进胶原 IV 合成，而胶原 IV 合成的增加有助于 NO 诱导的肺内皮细胞血管生成通过整合素-FAK 信号通路。胶原 IV 的操纵可能是预防和治疗肺泡毛细血管发育不良、严重肺动脉高压和肿瘤侵袭等疾病的新方法。
