Characterization And Functional Significance Of P450 Ara

P450 Ara 的表征及功能意义

• 批准号: 7168262
• 负责人: Darryl C Zeldin
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7168262
• 关键词: androgens; angiotensin II; arachidonate; blood pressure; cardiovascular disorder risk; cardiovascular function; cytochrome P450; eicosanoid metabolism; enzyme activity; enzyme mechanism; epoxide hydrolase; estrogens; gel mobility shift assay; gene expression profiling; gene therapy; genetically modified animals; hormone regulation /control mechanism; human subject; ion transport; kallikreins; kidney function; laboratory mouse; pharmacogenetics; polymerase chain reaction; single nucleotide polymorphism; telemetry

Cytochromes P450 metabolize arachidonic acid (AA) to epoxyeicosatrienoic acids (EETs) which have potent effects on cardiovascular and renal function. EETs are metabolized to corresponding diols (DHETs) by soluble epoxide hydrolase (sEH). Current research involves: (1) characterization of CYP2J and CYP2C subfamily P450s at the biochemical and molecular levels; (2) evaluation of the functional roles of CYP2J products in cardiovascular and renal physiology; and (3) examination of this pathway in selected animal models of human disease (ischemic heart disease, hypertension, atherosclerosis). We have discovered a number of mammalian CYP2Js, although we have focused most of our efforts on human CYP2J2 and mouse CYP2J5. Human CYP2J2 is the major human P450 expressed in heart and vasculature, where it is localized to cardiac myocytes and endothelial cells, and is active in the metabolism of AA to EETs. CYP2J2-derived EETs are vasodilators, inhibit cytokine-induced endothelial cell adhesion molecule expression, induce tissue plasminogen activator gene expression, inhibit vascular smooth muscle cell migration, protect endothelial cells against hypoxia-reoxygenation injury, upregulate endothelial nitric oxide biosynthesis, affect cardiac electrophysiology, and protect the heart from ischemic injury. CYP2J2 transgenic mice (alpha-myosin heavy chain promoter driven cardiac-specific expression) were developed to study the effects of increased EETs on cardiac function in vivo. These mice have normal basal heart anatomy and function, improved post-ischemic left ventricular function, shortened cardiac action potential, altered cardiac electrophysiology, and enhanced beta-adrenergic receptor responsiveness. Similarly, sEH null mice which exhibit reduced EET hydrolysis have improved postischemic functional recovery. We have also developed transgenic mice in which CYP2J2 is expressed exclusively in endothelial cells (Tie2 promoter driven) to examine the role of CYP2J2 products on vascular function. The phenotype of these mice is currently being evaluated. The human CYP2J2 gene has been cloned, sequenced and characterized. We have identified several functionally relevant CYP2J2 polymorphic variants, one of which is associated with reduced CYP2J2 expression and is present at higher frequency in patients with cardiovascular disease vs. normals in a large German cohort. We have also identified functionally relevant polymorphisms in the sEH gene and have shown that they are associated with increased cardiovascular risk in a large multicenter cohort in the U.S. CYP2J5 is a major murine P450 arachidonic acid epoxygenase expressed in the kidney and localized to proximal tubules. To evaluate the role of this P450 and its eicosanoid products in renal function and blood pressure regulation, we disrupted the Cyp2j5 gene by homologous recombination. CYP2J5 null mice have spontaneous hypertension that persists on both high and low salt diets. The hypertension is much more severe in females, is associated with reduced circulating estradiol levels, and is responsive to estrogen supplementation. CYP2J5 null female mice also have increased proximal tubular transport rates, enhanced vascular responsiveness to angiotensin II and endothelin, and reduced fertility compared to wild type counterparts. This new animal model will lead to a better understanding of the complex interrelationship between renal P450s, sex hormones, renal eicosanoids and blood pressure regulation. Consistent with these findings, we have shown that there is an association between CYP2J2 polymorphic variants and hypertension in a cohort from Tennessee.

细胞色素 P450 将花生四烯酸 (AA) 代谢为环氧二十碳三烯酸 (EET)，对心血管和肾功能具有有效影响。 EET 通过可溶性环氧化物水解酶 (sEH) 代谢为相应的二醇 (DHET)。目前的研究包括：（1）CYP2J和CYP2C亚家族P450在生化和分子水平上的表征； (2)评价CYP2J产物在心血管和肾脏生理学中的功能作用； (3) 在选定的人类疾病动物模型（缺血性心脏病、高血压、动脉粥样硬化）中检查该通路。尽管我们将大部分精力集中在人类 CYP2J2 和小鼠 CYP2J5 上，但我们已经发现了许多哺乳动物 CYP2J。人CYP2J2是在心脏和脉管系统中表达的主要人P450，其定位于心肌细胞和内皮细胞，并且在AA到EET的代谢中活跃。 CYP2J2衍生的EET是血管扩张剂，抑制细胞因子诱导的内皮细胞粘附分子表达，诱导组织纤溶酶原激活剂基因表达，抑制血管平滑肌细胞迁移，保护内皮细胞免受缺氧-复氧损伤，上调内皮一氧化氮生物合成，影响心脏电生理，保护心脏免受 缺血性损伤。 CYP2J2 转基因小鼠（α-肌球蛋白重链启动子驱动心脏特异性表达）被开发用于研究 EET 增加对体内心脏功能的影响。这些小鼠具有正常的基础心脏解剖结构和功能，改善了缺血后左心室功能，缩短了心脏动作电位，改变了心脏电生理学，并增强了β-肾上腺素能受体反应性。类似地，EET 水解减少的 sEH 缺失小鼠改善了缺血后的功能恢复。我们还开发了 CYP2J2 只在内皮细胞中表达（Tie2 启动子驱动）的转基因小鼠，以检查 CYP2J2 产品对血管功能的作用。目前正在评估这些小鼠的表型。人类 CYP2J2 基因已被克隆、测序和表征。我们已经鉴定了几种功能相关的 CYP2J2 多态性变异，其中一种与 CYP2J2 表达降低相关，并且在一个大型德国队列中，与正常人相比，心血管疾病患者的出现频率更高。我们还鉴定了 sEH 基因中功能相关的多态性，并在美国的一个大型多中心队列中证明它们与心血管风险增加相关。CYP2J5 是一种主要的小鼠 P450 花生四烯酸环氧化酶，在肾脏中表达并定位于近端肾小管。为了评估 P450 及其类二十烷酸产物在肾功能和血压调节中的作用，我们通过同源重组破坏了 Cyp2j5 基因。 CYP2J5缺失小鼠患有自发性高血压，在高盐和低盐饮食下均持续存在。女性的高血压更为严重，与循环雌二醇水平降低有关，并且对补充雌激素有反应。与野生型小鼠相比，CYP2J5缺失的雌性小鼠还具有增加的近端肾小管转运率、增强的血管对血管紧张素II和内皮素的反应性以及降低的生育力。这种新的动物模型将有助于更好地理解肾 P450、性激素、肾类二十烷酸和血压调节之间复杂的相互关系。与这些发现一致的是，我们在田纳西州的一个队列中证明了 CYP2J2 多态性变异与高血压之间存在关联。