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.

先前的研究表明，肾脏在动脉压的长期控制中发挥着主导作用，并且在迄今为止研究的每种高血压遗传和实验模型中，压力与尿钠排泄的关系都转向更高的压力。然而，改变肾功能的因素以及所涉及的基因和途径仍有待确定。该项目之前的工作表明，压力尿钠与肾髓质血流量和间质静水压 (RIHP) 升高以及近端肾小管 Na+ 转运抑制有关。在上一个资助期间，我们发现 RIHP 升高会刺激肾脏形成 20-HETE，并且 20-HETE 通过抑制近端肾小管中的钠转运来促进压力尿钠反应。我们进一步证明，阻断 20-HETE 的形成会促进盐敏感性高血压和 获得令人兴奋的新证据表明，将Lewis大鼠的含有CYP4504A等位基因的5号染色体5 cM区域转移到Dahl盐敏感（SS）遗传背景上会增加CYP4A蛋白的肾脏表达并减弱高血压的发展。我们现在已经掌握了传输的初步数据 来自布朗诺瓦里 (BN) 大鼠的 5 号染色体转移到 SS 遗传背景上也反对 SS.5BN 康体品系中高血压的发展。这些发现支持我们的工作假设，即肾脏形成 20-HETE 的缺陷导致 SS 大鼠患高血压。该项目的目标是确定是否存在降低活性或 SS 大鼠肾脏中 CYP4A 基因之一的表达，并在高血压的发生中发挥因果作用，或者该途径是否可能次要参与，因为 CYP4A 亚型的表达受到 5 号染色体上的某些其他基因的调节。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脏保护作用是否依赖于肾脏表达的增加CYP4A蛋白和20-HETE的生产； 2) 确定四个 CYP4A 基因之一是否存在序列变异，从而降低 SS 大鼠肾脏中这些酶的表达或活性； 3) 使用转基因技术测试 CYP4A 基因中鉴定的任何序列变异是否会导致 SS 大鼠高血压和肾病的发生。这些研究的新颖之处在于，他们将采用我们在过去 5 年中开发的独特的 5 号染色体同系和同体品系 SS 大鼠，这是一种新的 用于测量 20-HETE 的 LC/MS/MS 测定、20-HETE 合成和作用的新型抑制剂、用于测量 CYP4A 亚型表达的实时 PCR 测定以及用于创建大鼠转基因品系的新慢病毒策略确定 20-HETE 肾脏形成中的遗传异常是否通过重置压力-尿钠排泄关系而导致 SS 大鼠高血压发生的研究是特别独特的，并且相关的，因为 CYP4A11 基因中减少 20-HETE 形成的多态性最近已与三个独立人群中的血压升高相关。(77,183,184) 这些研究将提供一种新的同源动物模型系统，以探索机制来解释如何肾脏生成 20-HETE 的缺陷可能导致人类患上盐敏感型高血压。所获得的结果也可能会刺激 开发上调该途径的药物来治疗高血压和肾脏疾病。