Role of Cytochrome P450 Eichosanoids in Pressure-Natriuresis

细胞色素 P450 类二十烷酸在压力尿钠中的作用

• 批准号: 8230995
• 负责人: Richard J. Roman
• 金额: $ 25.56万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2013-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8230995
• 关键词: Alkane 1-monooxygenase; Alleles; Animal Model; Antihypertensive Agents; Attenuated; Biological Assay; Biological Models; Blood Pressure; Blood flow; CYP4A11 gene; Chromosomes, Human, Pair 5; Congenic Strain; Consomic Strain; Cytochrome P450; Data; Development; Experimental Models; Funding; Genes; Genetic; Genetic Models; Genetic Polymorphism; Goals; Human; Hydrostatic Pressure; Hydroxyeicosatetraenoic Acids; Hypertension; Kidney; Kidney Diseases; Measurement; Measures; Molecular Abnormality; Natriuresis; Pathway interactions; Play; Population; Production; Protein Isoforms; Proteins; Rat Strains; Rattus; Renal function; Role; Sodium; Techniques; Tern; Testing; Time; Transgenic Organisms; Variant; Work; drug development; enzyme activity; hypertension treatment; inhibitor/antagonist; interstitial; liquid chromatography mass spectrometry; man; novel; pressure; programs; response; salt sensitive; saluretic

Previous studies have indicated the kidney plays a dominant role in the long-tern control of arterial pressure and that the pressure-natriuresis relationship is shifted to higher pressures in every genetic and experimental model of hypertension that has been studied to date. However, the factors that alter renal function and the genes and pathways involved remain to be determined. Previous work done in this program revealed that pressure-natriuresis is associated with elevations in renal medullary blood flow and interstitial hydrostatic pressure (RIHP) and inhibition of Na+ transport in the proximal tubule. During the last funding period, we found that elevations in RIHP stimulate the renal formation of 20-HETE and that 20-HETE contributes to the pressure-natriuretic response by inhibiting sodium transport in the proximal tubule. We further demonstrated that blockade of the formation of 20-HETE promotes the development of salt-sensitive hypertension and obtained exciting new evidence that transfer of a 5 cM region of chromosome 5 containing the CYP4504A alleles from Lewis rats onto the Dahl salt-sensitive (SS) genetic background increases the renal expression of CYP4A protein and attenuates the development of hypertension. We now have preliminary data that transfer of chromosome 5 from the Brown Norwary (BN) rat onto the SS genetic background also opposes the development of hypertension in a SS.5BN consomic strain. These findings support our working hypothesis that a deficiency in the renal formation of 20-HETE contributes to the development of hypertension in SS rats. The goal of this project is to determine whether there is a sequence variant that reduces the activity or expression of one of the CYP4A genes in the kidney of SS rats and plays a causal role in the development of hypertension or if this pathway maybe secondarily involved because the expression of CYP4A isoforms are regulated by some other gene on chromosome 5. The Specific Aims are: 1) to determine whether the antihypertensive and renoprotective effects of transfer of chromosome 5 from the BN rat into the SS genetic background is dependent on an increase in the renal expression of CYP4A protein and the production of 20-HETE; 2) to determine if there is a sequence variant in one of the four CYP4A genes that reduces the expression or activity of these enzymes in the kidney of SS rats; and 3) to test if any of the sequence variants identified in the CYP4A genes contribute to the development of hypertension and renal disease in SS rats using transgenic techniques. The novel aspects of these studies are that they will employ unique chromosome 5 congenic and consomic strains of SS rats that we developed over the last 5 years, a new LC/MS/MS assay for measurement of 20-HETE, novel inhibitors of the synthesis and actions of 20-HETE, real time-PCR assays for measuring the expression of CYP4A isoforms and a new lentiviral strategy for creating transgenic strains of rats The proposed studies to determine if a genetic abnormality in the renal formation of 20-HETE contributes to the development of hypertension in SS rats by resetting the pressure-natriuretic relationship are especially unique and relevant, since a polymorphism in the CYP4A11 gene that reduces the formation of 20-HETE has recently been associated with elevated blood pressure in three independent human populations.(77,183,184) These studies will provide a new homologous animal model system to explore mechanisms to explain how a deficiency in the renal formation of 20-HETE could contribute to the development of salt-sensitive forms of hypertension in man. The results obtained may also spur the development of drugs that upregulate this pathway for the treatment of hypertension and renal disease.

以前的研究表明，肾脏在动脉血压的长期控制中起着主导作用，到目前为止，在所有已研究的高血压遗传和实验模型中，压力-钠尿关系都向更高的压力转移。然而，改变肾功能的因素以及涉及的基因和途径仍有待确定。该项目以前的工作表明，压力-钠尿与肾髓质血流量和间质静水压力(RIHP)的升高以及近端小管Na+转运的抑制有关。在上一次资助期间，我们发现RIHP的升高刺激了20-HETE的肾脏形成，并且20-HETE通过抑制近端小管的钠转运而促进了血压-利钠反应。我们进一步证明，阻断20-HETE的形成促进盐敏感型高血压的发展和 获得了令人振奋的新证据，即将5号染色体上包含CYP4504A等位基因的5 cM区域从Lewis大鼠转移到Dahl盐敏感(SS)遗传背景上，增加了肾脏CyP4A蛋白的表达，并减缓了高血压的发展。我们现在有初步数据表明 从棕色Norwary(BN)大鼠到SS遗传背景的5号染色体也反对SS.5BN单体品系中高血压的发展。这些发现支持了我们的工作假设，即肾脏形成20-HETE的缺陷有助于SS大鼠高血压的发展。这个项目的目标是确定是否存在降低活性的序列变体或 目的：1)研究5号染色体转导到SS大鼠肾脏中的降压和肾脏保护作用是否依赖于肾脏中CYP4A蛋白表达的增加和20-HETE的产生；2)确定四种CYP4A基因中的一种是否存在序列变异，从而降低SS大鼠肾脏中这些酶的表达或活性；以及3)利用转基因技术检测在CYP4A基因中发现的序列变异是否与SS大鼠的高血压和肾脏疾病的发生有关。这些研究的新颖之处在于，他们将使用我们在过去5年中培育的SS大鼠独特的5号染色体同源和共体品系，一种新的 用于测定20-HETE的LC/MS/MS分析方法、用于测定20-HETE的合成和作用的新型抑制剂、用于测定细胞色素P4A亚型表达的实时定量聚合酶链式反应分析方法以及用于创造转基因大鼠品系的新的慢病毒策略。本研究旨在确定20-HETE的肾脏结构的遗传异常是否通过重置压力-钠尿关系而导致SS大鼠高血压的发生具有特别独特和相关的意义。由于最近在三个独立的人类群体中发现了减少20-HETE形成的CYP4A11基因的多态与高血压有关。(77,183,184)这些研究将提供一个新的同源动物模型系统，以探索20-HETE的肾脏形成缺陷如何促进人类盐敏感型高血压的发展的机制。所获得的结果也可能刺激 为治疗高血压和肾脏疾病而开发上调这一途径的药物。