Lipid Modulators of Pulmonary Vascular Tone

描述（由申请人提供）：我们的研究主要集中在Cyp4及其脂质产物20-HETE在肺血管功能和生物学中的作用。在过去的4年里，我们发现了Cyp4/20-HETE在调节肺动脉内皮细胞（PAEC） eNOS，介导vegf诱导的小肺动脉舒张中的独特作用。除了影响血管张力外，Cyp4还促进全身血管床的血管生成。我们的初步数据表明，在paec中，20-HETE具有一种新的抗凋亡活性，可以防止饥饿引起的caspase 3活性的增加。除了众所周知的造成氧化损伤的能力，活性氧（ROS）现在被理解为在包括细胞生长和血管生成在内的重要生理过程的信号传导中发挥关键作用。因此，我们探索了Cyp4/20-HETE影响paec中ROS生成和内皮细胞生长的潜力。我们的数据首次证明了20-HETE/ cyp4诱导分离paec和完整肺中ROS的增加。在我们的实验中，NADPH氧化酶和线粒体呼吸都是20-HETE处理的肺内皮中重要的ROS来源。抑制ROS可消除20-HETE对caspase 3活性的保护作用。这些观察结果表明，Cyp4/20- HETE的至少一个作用可能是通过刺激ROS的产生来保护肺动脉内皮细胞免受凋亡。通过这些作用，20-HETE/Cyp4可能有助于维持肺血管床的完整性。我们将验证20-HETE/Cyp4增加paec中的ROS并以ROS依赖的方式保护细胞凋亡的总体假设。在第一个目标中，我们将研究Cyp4/20-HETE在体外和体内增加paec ROS产生的机制，重点关注NADPH氧化酶和线粒体效应。在第二个目标中，我们将研究20-HETE刺激ROS产生的功能意义，探讨ROS在体内和体外对内皮细胞凋亡的保护作用。我们将与自由基和血管生物学中心的合作伙伴和专业知识一起，结合最先进的技术来研究Cyp4/20-HETE在肺内皮中的这些令人兴奋的新功能。

DESCRIPTION (provided by applicant): Our studies have focused on the role of Cyp4 and its lipid product, 20-HETE in pulmonary vascular function and biology. Over the last 4 years, we identified a unique role for Cyp4/20-HETE in regulating pulmonary arterial endothelial cell (PAEC) eNOS, mediating VEGF-induced relaxation of small pulmonary arteries. In addition to effects on vascular tone, Cyp4 promotes angiogenesis in systemic vascular beds. Our preliminary data demonstrate a novel, anti-apoptotic activity of 20-HETE in PAECs, with 20-HETE protecting against starvation-evoked increases in caspase 3 activity. Beyond their well recognized capacity to inflict oxidative injury, reactive oxygen species (ROS) are now understood to play key roles in signaling vital physiologic processes including cell growth and angiogenesis. Therefore we explored the potential of Cyp4/20-HETE to impact ROS production and endothelial cell growth in PAECs. Our data demonstrate for the first time 20-HETE/Cyp4-induced increases in ROS in isolated PAECs as well as in intact lungs. Both NADPH oxidase and mitochondrial respiration appear to be important ROS sources in pulmonary endothelium treated with 20-HETE in our experiments. Inhibition of ROS eliminates the protective effect of 20-HETE on caspase 3 activity. These observations raise the suggestion that at least one role of Cyp4/20- HETE may be to protect pulmonary arterial endothelial cells against apoptosis via stimulation of ROS production. By such actions, 20-HETE/Cyp4 may contribute to sustaining the integrity of the pulmonary vascular bed. We will test the overall hypothesis that 20-HETE/Cyp4 increases ROS in PAECs and protects against apoptosis in a ROS-dependent manner. In the first aim, we will investigate the mechanisms through which Cyp4/20-HETE increases ROS production of PAECs, both in vitro and in vivo, with a focus on NADPH oxidase and mitochondrial effects. In the second aim, we will study the functional implications of 20-HETE stimulated ROS production, probing the role of ROS in protection against apoptosis of endothelium in vivo and in vitro. Together with our collaborators and expertise available in the Free Radical and Vascular Biology Centers, we will use a combination of state of the art techniques to study these exciting and novel functions of Cyp4/20-HETE in lung endothelium.