Nitro-nitrate fatty acid derivatives as novel cGMP-dependent and cGMP-independent signaling mediators

项目总结 有机硝酸酯是长期以来被推崇的心血管疾病(CVD)的治疗方法，是导致 世界范围内的死亡。值得注意的是，有机硝酸盐从未在生物学上被检测到，而且它们的血管扩张作用 由于硝酸盐耐受和内皮功能障碍的发展而减弱，突出了迫切需要 寻求另类疗法。在这种情况下，饮食干预可以降低心血管疾病的发病率和死亡率。虽然福利 在饮食干预中，临床上有很好的记录，关于其贡献的新数据不断涌现 个体饮食成分在调节有益的下游反应中的作用。食用水果， 蔬菜、肉类和乳制品富含共轭亚油酸(CLA)和无机盐。 阴离子亚硝酸盐(NO2−)和硝酸盐(NO3−)。强有力的证据表明，NO_2−和NO_3−很容易转化 通过代谢和炎症条件促进共轭亚油酸的硝化生成亲电的硝基烯烃 衍生物(NO2-FA)，可诱导亲核半胱氨酸(Cys)残基的可逆烷基化，从而 许多功能重要的调节蛋白的翻译后修饰(PTM)。临床前和临床 资料证实，NO2-FA可诱导多向适应性和抗炎基因表达反应。我们 最近报道，酸性的胃环境和共轭双结合导致了一种新的 NO2−介导的不饱和脂肪酸硝化机制是产生含有机硝酸盐的关键 脂肪酸衍生物，称为硝基-硝酸(NO2-ONO2-FA)。初步数据表明，NO2-ONO2- FA：A)不仅在体内通过共轭亚油酸+亚硝酸盐−的消化反应内源性产生，而且在体外也产生 心肌I/R损伤，b)存活于肠道吸收，被结合到乳糜米隆甘油三酯和REACH 体循环，c)是非亲电性，然后衰变为亲电性的NO2-FA产物 释放次级氮氧化物和d)可诱导协同有益的PTM效应和鸟苷环化酶 激活。根据这一认识，现在假设NO2-ONO2-FA衍生物促进血管扩张和 通过cGMP依赖和非cGMP抑制心血管炎症反应 信号动作。追求这个项目的基本原理是定义生化形成和独特的PK 和NO2-ONO2-FA的信号作用，即当前的数据支持是一个小分子介体，它显示 有机硝酸盐和半胱氨酸反应性亲电体的药理学。我们还更好地定义了 内源性产物来源于健康饮食和炎症的代谢反应。目标 该提案的内容是：1)定义NO2-ONO2-FA的代谢和炎症生成及PK；2)定义 量化体外和体内对NO2-ONO2-FA的心血管反应 血管保护性cGMP依赖和cGMP非依赖性信号转导。圆满完成拟议中的 研究计划可以为针对炎症相关CVD的安全药理策略奠定基础。

PROJECT SUMMARY Organic nitrate esters are long-appreciated treatments for cardiovascular disease (CVD), leading causes of death worldwide. Of note, organic nitrates have never been detected biologically, and their vasodilatory effects are attenuated by the development of nitrate tolerance and endothelial dysfunction, highlighting an urgent need for alternative therapies. In this context, dietary interventions reduce CVD incidence and mortality. While benefits of the dietary interventions are clinically well documented, new data continues to emerge as to the contributions of individual dietary constituents in mediating beneficial downstream responses. Consumption of fruits, vegetables, meat, and dairy products provides a rich source of conjugated linoleic acid (CLA) and the inorganic anions nitrite (NO2−), and nitrate (NO3−). Strong evidence reveals that NO2− and NO3− are readily transformed by metabolic and inflammatory conditions promoting nitration of CLA and yielding electrophilc nitroalkene derivatives (NO2-FA), which induce reversible alkylation of nucleophilic cysteine (Cys) residues with consequent post-translational modification (PTM) of many functionally-significant regulatory proteins. Preclinical and clinical data affirm that NO2-FA induce pleiotropic adaptive and anti-inflammatory gene expression responses. We recently reported that the acidic gastric environment and the conjugated double bound of CLA results in a novel NO2−-mediated unsaturated fatty acid nitration mechanism that is critical for generating organic nitrate-containing fatty acid derivatives, termed nitro-nitrate fatty acids (NO2-ONO2-FA). Preliminary data indicates that NO2-ONO2- FA: a) are not only endogenously-generated in vivo by digestive reactions of CLA + NO2− but also upon ex vivo myocardial I/R injury, b) survive intestinal absorption, are incorporated into chylomicron triglycerides and reach the systemic circulation, c) are non-electrophilic that then decay to an electrophilic NO2-FA product after releasing secondary nitrogen oxides and d) can induce concerted salutary PTM effects and guanylate cyclase activation. From this insight, it is now hypothesized that NO2-ONO2-FA derivatives promote vasodilation and inhibit cardiovascular inflammatory responses via both cGMP-dependent and cGMP-independent signaling actions. The rationale for pursuing this project is to define the biochemical formation and unique PK and signaling actions of NO2-ONO2-FA, that current data support is a small molecule mediator that displays the pharmacology of both organic nitrates and Cys-reactive electrophiles. We also better define the scope of endogenous products stemming from the metabolic reactions of both a healthy diet and inflammation. The goals of the proposal are: 1) to define the metabolic and inflammatory generation and PK of NO2-ONO2-FA and 2) to quantify the in vitro and in vivo cardiovascular responses to NO2-ONO2-FA in the context of both vasodilatory and vasoprotective cGMP-dependent and cGMP-independent signaling. Successful completion of the proposed research plan can lay the foundation for a safe pharmacological strategy targeting inflammatory-related CVD.