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.

以往的研究表明，肾脏在动脉压的长期控制中起着主导作用，并且迄今为止研究的每一种高血压遗传和实验模型中，压力-尿钠排泄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。本项目中以前的工作表明，压力性尿钠排泄与肾髓质血流量和间质流体静压（RIHP）升高以及近端小管Na+转运抑制相关。在上一个资助期间，我们发现RIHP升高刺激肾脏形成20-HETE，20-HETE通过抑制近端小管中的钠转运促进压力-利钠反应。我们进一步证明，阻断20-HETE的形成可促进盐敏感性高血压的发展， 获得了令人兴奋的新证据，将含有来自刘易斯大鼠的CYP 4504 A等位基因的5号染色体的5 cM区域转移到Dahl盐敏感（SS）遗传背景上，增加了CYP 4A蛋白的肾脏表达，并减弱了高血压的发展。我们现在有初步的数据， 将来自Brown Norwary（BN）大鼠的5号染色体转移到SS遗传背景上也对抗SS.5BN同源品系中高血压的发展。这些发现支持了我们的工作假设，即肾形成的20-HETE的不足有助于SS大鼠高血压的发展。这个项目的目标是确定是否有一个序列变异，减少活动或 SS大鼠肾脏中CYP 4A基因之一的表达，并在高血压的发生中起因果作用，或者该途径可能次要参与，因为CYP 4A亚型的表达受5号染色体上的其他基因调节。具体目标是：1）确定将5号染色体从BN大鼠转移到SS遗传背景中的抗高血压和肾保护作用是否依赖于CYP 4A蛋白质的肾表达和20-HETE的产生的增加; 2）确定在四种CYP 4A基因之一中是否存在降低SS大鼠肾中这些酶的表达或活性的序列变体;和3）使用转基因技术测试在CYP 4A基因中鉴定的任何序列变体是否有助于SS大鼠中高血压和肾病的发展。这些研究的新颖之处在于，它们将采用我们在过去5年中开发的独特的5号染色体同源和同源的SS大鼠品系，这是一种新的 用于测量20-HETE（20-HETE合成和作用的新型抑制剂）的LC/MS/MS分析，测定CYP 4A亚型表达的真实的时间PCR测定法和建立大鼠转基因品系的新慢病毒策略拟定的研究旨在确定20-HETE肾形成中的遗传异常是否通过重置压力-利钠关系是特别独特和相关的，因为CYP 4A 11基因中减少20-HETE形成的多态性最近在三个独立的人群中与血压升高相关。(77 183，184）这些研究将提供一种新的同源动物模型系统，以探索解释肾脏形成20-HETE的缺陷如何有助于人类盐敏感型高血压的发展的机制。 开发上调这一途径的药物，用于治疗高血压和肾脏疾病。