Regulation of eNOS by S-Nitrosylation in Diet Associated Erectile Dysfunction

S-亚硝基化对饮食相关勃起功能障碍中 eNOS 的调节

基本信息

批准号：
8717081
负责人：
Justin David LaFavor
金额：
$ 5.15万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-01-01 至 2017-09-23
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8717081
关键词：
Active Sites Address Animal Sources Binding Biological Assay Consumption Coupling Cysteine Data Dependence Development Diamide Diet Dietary Intervention Dithiothreitol Dose Dyslipidemias Endothelial Cells Environment Erectile dysfunction Etiology Event Fatty Acids Fatty acid glycerol esters Fellowship Fibrosis Genetic Glutathione Disulfide Glutathione Reductase Health Immunoprecipitation Impairment Individual Insulin Resistance Intake Intraperitoneal Injections Knockout Mice Link Maintenance Mass Spectrum Analysis Measurement Measures Mediating Mediator of activation protein Metabolic syndrome Methods Modeling Modification Molecular Morphology Mus Nerve Nitric Oxide Nitric Oxide Synthase Nutrient Obesity Operative Surgical Procedures Oxidants Oxidation-Reduction Oxidative Stress Pathway interactions Pattern Penile Erection Physiological Post-Translational Protein Processing Prevention Protein S Proteins Proteome Proteomics Quality of life Rattus Reactive Oxygen Species Reducing Agents Regulation Research Rodent Role SKIL gene Signal Pathway Signal Transduction Signaling Molecule Site Staining method Stains Stimulus Sucrose Sulfhydryl Compounds Superoxides System Techniques Thioredoxin Tissues Training United States Vegetable Oils Western Blotting base dimer disulfide bond experience feeding genetic manipulation human NOS3 protein in vivo inhibitor/antagonist light microscopy mTOR protein monomer mouse model novel strategies nutrition penis protein structure function public health relevance response sugar vector

项目摘要

PROJECT SUMMARY A diet abundant in sugar and fats derived from animal sources and cheap vegetable oils has been termed the "Western diet". Consumption of the Western diet is now prevalent in the United States and has been spreading throughout the world over the past 15 years. Consumption of a Western style diet is implicated in the etiology of erectile dysfunction (ED), a condition strongly associated with reduced quality of life. Nitric oxide is a primary mediator of penile erection, which is produced by nitric oxide synthase. Under conditions of excessive oxidative stress, endothelial nitric oxide synthase (eNOS) may become uncoupled, whereby superoxide is produced rather than nitric oxide, which propagates further oxidative stress. It is thought that eNOS coupling status is regulated by adequate presence of co-factors and substrates. However, recent data indicate that eNOS uncoupling and dysfunctionality may be promoted by S-nitrosylation, which may be dependent on an oxidative shift in cellular redox state. Additionally, the mammalian target of rapamycin (mTOR) is a cellular signaling molecule that may induce oxidative stress in response to excessive fat and sugar consumption. The primary hypothesis of this proposal is that an oxidative shift in the penile redox environment, mediated by mTOR- dependent nutrient signaling in response to the Western diet promotes erectile dysfunction due to eNOS dysregulation via excessive S-nitrosylation induced by impairment of the denitrosylation system. Specific aims for the proposal are: 1) to determine if Western diet-induced ED is mediated by an oxidative shift in redox state dependent upon mTOR signaling; 2) to determine if the mTOR dependent shift in redox state exerts alteration of S-nitrosylation; 3) to determine if regulation of S-nitrosylation controls erectile function in a diet-dependent manner. The proposed research plan includes the use of a Western style rodent diet in combination with pharmacologic, molecular, proteomic, and physiological studies. These studies utilize the Western diet in combination with pharmacologic shifts in redox state, inhibition of mTOR, and genetic manipulation of proteins that regulate denitrosylation. Molecular studies proposed include assessment of protein structure, function, and post-translational modifications influencing eNOS, the mTOR signaling pathway, and the proteins that regulate cellular S-nitrosylation. Proteomic studies proposed include assessment of S-nitrosylation status of the penile proteome, as well as S-nitrosylation of individual thiol groups of penile eNOS. Molecular and proteomic studies will be enriched by physiologic erectile function measurements in response to these perturbations. The proposed studies include the use of a genetic mouse model where the regulation of S-nitrosylation is manipulated, and a rat model where the regulation of S-nitrosylation is manipulated specifically in the penis. It is anticipated that the findings of this proposal will provide greater understanding of molecular events leading to Western diet associated ED, and provide a basis for enhanced treatment.

项目概要富含来自动物来源的糖和脂肪以及廉价植物油的饮食被称为“ “西方饮食”。西方饮食的消费现在在美国很盛行并且正在蔓延过去15年遍布世界各地。西式饮食的消费与病因有关勃起功能障碍（ED），一种与生活质量下降密切相关的疾病。一氧化氮是主要的阴茎勃起的介质，由一氧化氮合酶产生。在过度氧化的条件下应激时，内皮一氧化氮合酶 (eNOS) 可能会解偶联，从而产生超氧化物而不是一氧化氮，它会进一步传播氧化应激。 eNOS 耦合状态被认为是通过充足的辅因子和底物的存在来调节。然而，最近的数据表明，eNOS S-亚硝基化可能会促进解偶联和功能障碍，这可能依赖于氧化细胞氧化还原状态的转变。此外，哺乳动物雷帕霉素靶标 (mTOR) 是一种细胞信号传导可能因过度脂肪和糖消耗而诱发氧化应激的分子。初级该提议的假设是，由 mTOR-介导的阴茎氧化还原环境中的氧化转变响应西方饮食的依赖营养信号会促进 eNOS 导致的勃起功能障碍去亚硝基化系统受损引起的过度 S-亚硝基化导致调节失调。具体目标该提案的目的是：1) 确定西方饮食引起的 ED 是否是由氧化还原状态的氧化转变介导的依赖于 mTOR 信号传导； 2) 确定 mTOR 依赖的氧化还原态转变是否会产生改变 S-亚硝基化； 3) 确定 S-亚硝基化的调节是否控制饮食依赖型患者的勃起功能方式。拟议的研究计划包括使用西式啮齿动物饮食并结合药理学、分子、蛋白质组学和生理学研究。这些研究利用西方饮食与氧化还原状态的药理学转变、mTOR 抑制和蛋白质基因操作相结合调节去亚硝基化。提议的分子研究包括评估蛋白质结构、功能和影响 eNOS、mTOR 信号通路和调节蛋白的翻译后修饰细胞S-亚硝基化。提议的蛋白质组学研究包括评估阴茎的 S-亚硝基化状态蛋白质组，以及阴茎 eNOS 各个硫醇基团的 S-亚硝基化。分子和蛋白质组学研究将通过响应这些扰动的生理勃起功能测量来丰富。这拟议的研究包括使用遗传小鼠模型，其中 S-亚硝基化的调节是以及在阴茎中专门操纵 S-亚硝基化调节的大鼠模型。它预计该提案的结果将有助于人们更好地了解导致西方饮食与ED相关，并为加强治疗提供依据。