Ferrochelatase & guanylate cyclase regulation in vascular dysfunction

铁螯合酶

• 批准号: 9127540
• 负责人: DONG SUN
• 金额: $ 41万
• 依托单位: NEW YORK MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2020-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9127540
• 关键词: Amino Acids; Aminolevulinic Acid; Angiotensin II; Antioxidants; Arteries; Biological; Blood Pressure Monitors; Blood Vessels; Cattle; Coronary; Coronary artery; Cyclic GMP; Dependence; Development; Disease; Disease model; Echocardiography; Enzymes; Evaluation; Functional disorder; Generations; Guanylate Cyclase; Heme; Human; Hypertension; Implant; Infusion procedures; Investigation; Iron; Literature; Mediating; Mesentery; Metabolic Diseases; Microcirculation; Mitochondria; Mitochondrial Matrix; Mus; Nitric Oxide; Organoids; Oxidases; Pharmaceutical Preparations; Process; Production; Proteolysis; Pump; Reactive Oxygen Species; Receptor, Angiotensin, Type 1; Regulation; Role; SOD2 gene; Site; Skeletal Muscle; Small Interfering RNA; Soluble Guanylate Cyclase; Source; Sulfur; Superoxides; System; Therapeutic; Tissues; Vascular Diseases; Vascular Smooth Muscle; Vasodilation; Vasodilator Agents; Work; base; biological systems; cofactor; cremaster muscle; drinking water; ferrochelatase; heme biosynthesis; in vivo; intravital imaging; mouse model; novel; novel therapeutic intervention; osmotic minipump; oxidation; prevent; protoporphyrin IX; public health relevance; receptor; response; targeted treatment

 DESCRIPTION (provided by applicant): The overall hypothesis for this project is that increased mitochondrial generation of superoxide causes a disruption of vascular smooth muscle heme biosynthesis by ferrochelatase (FECH), resulting in a loss of the beneficial vascular regulatory effects of soluble guanylate cyclase (sGC). It is known that oxidation of the sGC heme promotes loss of regulation by nitric oxide (NO) and sGC degradation by proteolysis. While the mitochondrial heme generating enzyme FECH has an iron-sulfur cluster essential for its stability, which is a potential target for disruption by superoxide, the literature appears to ack evidence for the consequences of FECH dysfunction on the heme-dependence of sGC regulation in any biological system. Due to the potential importance of angiotensin II (AngII) in promoting human vascular dysfunction in multiple diseases and previous evidence for mechanisms associated with increased superoxide generation in subcellular sites including mitochondria, and disruption of NO and its associated heme-dependent regulation of sGC/cGMP-mediated vasodilation, we hypothesized and found evidence for both FECH activity being disrupted by AngII and for FECH controlling the expression and NO-mediated activation of sGC. Studies in Aim 1 focus on showing how AngII regulation of increases in mitochondrial superoxide cause a disruption in heme biosynthesis by FECH that impairs vascular regulation by sGC in isolated mouse and bovine coronary arteries treated with AngII and siRNA or mechanistic probes modulating subcellular sources of superoxide and FECH. Arteries from mice deficient in AngII type-1 receptor (AT1R), Cu, Zn-SOD (SOD1), mitochondrial matrix SOD2, FECH, Nox1 and Nox2 oxidase will be used in these studies to define how AngII regulation of cytosolic and mitochondrial superoxide influences regulation by the sGC and FECH systems. Aim 2 investigates new preliminary observations suggesting how promoting availability of protoporphyrin IX (PpIX, an activator of sGC) and heme from δ-aminolevulinic acid (ALA) appears to protect ferrochelatase and sGC regulation potentially through a cGMP-mediated inhibition of mitochondrial superoxide that may involve cGMP preventing the depletion of SOD2. Studies in Aim 3 focus in defining the in vivo importance of the disruption of ferrochelatase and heme- associated sGC regulation of vascular function and protection provided by ALA, using a mouse model of osmotic minipump delivery of AngII. Radiotelemetry monitoring of blood pressure, echocardiography, and evaluation of changes in isolated arteries and in vivo arteriolar function in the skeletal muscle microcirculation will be evaluated. The role of subcellular sources of superoxide in the disruption of ferrochelatase will also be assessed in AngII-infused mice deficient in AT1R, SOD1, SOD2, Nox1 and Nox2, together with a therapy specifically targeting mitochondrial superoxide. These studies are expected to document that mitochondrial superoxide disruption of ferrochelatase has an important role in sGC-associated vascular dysfunction, which can be targeted with a beneficial ALA therapy regulating the activity, expression and NO-stimulation of sGC.

 描述（由申请人提供）：该项目的总体假设是，线粒体产生的超氧化物增加导致亚铁螯合酶（FECH）破坏血管平滑肌血红素生物合成，导致可溶性鸟苷酸环化酶（sGC）的有益血管调节作用丧失。已知sGC血红素的氧化促进一氧化氮（NO）调节的丧失和通过蛋白水解的sGC降解。虽然线粒体血红素生成酶FECH具有对其稳定性至关重要的铁-硫簇，其是超氧化物破坏的潜在靶标，但文献似乎缺乏FECH功能障碍对任何生物系统中sGC调节的血红素依赖性的后果的证据。由于血管紧张素II（AngII）在促进多种疾病中的人类血管功能障碍中的潜在重要性以及先前关于与包括线粒体在内的亚细胞位点中超氧化物生成增加以及NO的破坏及其相关的sGC/cGMP介导的血管舒张的血红素依赖性调节相关的机制的证据，我们假设并发现了FECH活性被AngII破坏以及FECH控制sGC表达和NO介导的活化的证据。目的1中的研究集中于显示AngII调节线粒体超氧化物的增加如何通过FECH引起血红素生物合成的破坏，这损害了用AngII和siRNA或调节超氧化物和FECH的亚细胞来源的机械探针处理的分离的小鼠和牛冠状动脉中sGC的血管调节。来自AngII 1型受体（AT 1 R）、Cu，Zn-SOD（SOD 1）、线粒体基质SOD 2、FECH、Nox 1和Nox 2氧化酶缺陷小鼠的动脉将用于这些研究，以确定AngII对胞质和线粒体超氧化物的调节如何影响sGC和FECH系统的调节。目的2研究了新的初步观察结果，表明促进原卟啉IX（PpIX，sGC的激活剂）和来自δ-氨基乙酰丙酸（ALA）的血红素的可用性似乎如何通过cGMP介导的线粒体超氧化物的抑制来保护亚铁螯合酶和sGC调节，这可能涉及cGMP防止SOD 2的耗尽。目的3中的研究集中于使用AngII的渗透性微泵递送的小鼠模型来定义由ALA提供的血管功能和保护的亚铁螯合酶和血红素相关sGC调节的破坏的体内重要性。将评价血压的无线遥测监测、超声心动图以及骨骼肌微循环中离体动脉和体内小动脉功能变化的评价。亚细胞来源的作用 还将在AT 1 R、SOD 1、SOD 2、Nox 1和Nox 2缺陷的AngII输注小鼠中评估超氧化物在亚铁螯合酶破坏中的作用，以及特异性靶向线粒体超氧化物的治疗。这些研究预计将证明亚铁螯合酶的线粒体超氧化物破坏在sGC相关的血管功能障碍中具有重要作用，这可以通过调节sGC的活性、表达和NO刺激的有益ALA疗法来靶向。