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（E3），E3对心血管和肾功能具有强有力的作用。通过可溶性环氧化物水解酶（sEH）将雌二醇代谢成相应的二醇（DHE 2）。目前的研究涉及：（1）在生物化学和分子水平上表征CYP 2 J和CYP 2C亚家族P450;（2）评价CYP 2 J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（缺血性心脏病、高血压、动脉粥样硬化）动物模型中检查该途径。我们已经发现了许多哺乳动物CYP 2 J，尽管我们的大部分努力都集中在人类CYP 2 J2和小鼠CYP 2 J5上。人CYP 2 J2是在心脏和血管系统中表达的主要人P450，在那里它定位于心肌细胞和内皮细胞，并且在AA至雌二醇的代谢中具有活性。CYP 2 J2-衍生的内皮素是血管扩张剂，抑制精氨酸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，影响心脏电生理学，并保护心脏免受缺血性损伤。开发CYP 2 J2转基因小鼠（α-肌球蛋白重链启动子驱动的心脏特异性表达），以研究体内增加的雌二醇对心脏功能的影响。这些小鼠具有正常的基础心脏解剖结构和功能，改善缺血后左心室功能，缩短心脏动作电位，改变心脏电生理学，增强β-肾上腺素能受体反应性。类似地，表现出减少的EET水解的sEH缺失小鼠具有改善的缺血后功能恢复。我们还开发了转基因小鼠，其中CYP 2 J2仅在内皮细胞中表达（Tie 2启动子驱动），以检查CYP 2 J2产物对血管功能的作用。目前正在评估这些小鼠的表型。人CYP 2 J2基因已被克隆、测序和表征。我们已经确定了几种功能相关的CYP 2 J2多态性变体，其中一种与CYP 2 J2表达降低相关，并且在德国大型队列中，心血管疾病患者与正常患者相比存在更高的频率。我们还确定了sEH基因的功能相关多态性，并在美国的一个大型多中心队列中表明它们与心血管风险增加相关。CYP 2 J5是一种主要的鼠P450花生四烯酸环氧合酶，在肾脏中表达并定位于近端小管。为了评估P450及其类花生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Cyp 2 j5基因。CYP 2 J5敲除小鼠具有自发性高血压，其在高盐和低盐饮食下都持续存在。高血压在女性中更为严重，与循环雌二醇水平降低有关，并对雌激素补充剂有反应。与野生型小鼠相比，CYP 2 J5敲除雌性小鼠的近端肾小管转运速率增加，血管对血管紧张素II和内皮素的反应性增强，生育力降低。这种新的动物模型将导致更好地了解肾脏P450，性激素，肾脏类花生酸和血压调节之间复杂的相互关系。与这些发现一致，我们已经表明，在田纳西州的一个队列中，CYP 2 J2多态性变异与高血压之间存在关联。