Role of Epoxygenated Fatty Acids in Modulating Pain

描述（由申请人提供）：慢性疼痛状态对新型疼痛疗法存在巨大需求。花生四烯酸（ARA）级联反应是一个重要的炎症性疼痛通路，许多研究涉及该级联反应的环氧合酶（考克斯）和脂氧合酶（lox）分支。然而，第三个分支，细胞色素P450途径的重要性，提供了新的替代疼痛调制。与炎性考克斯和LOX代谢物相反，细胞色素P450产生的ARA代谢物环氧二十碳三烯酸（Ehrs）是有效的生理抗炎/抗痛觉过敏分子。这些生物活性脂质具有非常短的半衰期，并且抑制它们由可溶性环氧化物水解酶（sEH，EC 3.3.2.3）介导的降解导致抗伤害感受。sEH抑制剂在炎症性和神经性疼痛模型中有效，但目前尚不清楚sEH抑制如何减轻疼痛，阻碍了对这种新途径的进一步研究。sEH在外周和中枢神经系统、神经元和神经胶质细胞中均有表达，并具有空间选择性 大脑区域之间的表达。sEH抑制剂（sEHI）阻断伤害性行为，同时保持正常的伤害性和运动功能。因此，在这项研究中，我们将测试的假设，环氧脂肪酸和sEHI在疼痛病理生理学的调节作用。为了解决这一假设，将监测两种不同疼痛模型中皮肤和腰脊髓中EpFA及其降解产物的水平。然后通过不同的方法在两种不同的疼痛模型中调节EpFA的水平; a）通过外源性EpFA，B）通过抑制sEH，c）通过抑制磷酸二酯酶和d）通过对sEH介导的降解有抗性的EpFA的新合成类似物。我们将利用角叉菜胶诱导的炎性疼痛模型来鉴定抗痛觉过敏的EpFA。抗伤害性EpFA的生物活性将在临床相关模型（切口疼痛模型）中得到验证。我们将使用行为分析，生物化学和代谢组学分析的组合，以了解与伤害性通路的EpFAs的相互作用。我们将产生一个特征/诊断LC/MS/MS生成的代谢组学配置文件的足底角叉菜胶和足底切口疼痛大鼠模型。特别是切口疼痛的生物活性脂质代谢物的变化方面的研究较少。目标 代谢物（80+生物活性脂质）来自AA级联的所有三个分支，包括主要的考克斯、lox、细胞色素P450和sEH产物，包括EPA和DHA的脂质代谢物，这允许我们研究生物级联之间的代谢物通量和串扰。拟议的研究将导致进一步了解的抗伤害性机制的行动EpFA和sEH抑制剂，并提供新的见解的潜在机制的lipidomic和生物化学的改变，在疼痛。这些实验产生的知识可能会导致开发新的药物类别，这些药物的靶向途径以前没有被认识到。

DESCRIPTION (provided by applicant): Great demand for novel pain therapies exists for chronic pain states. The arachidonic acid (ARA) cascade is a pivotal inflammatory pain pathway, and many studies addressed the cyclooxygenase (cox) and lipoxygenase (lox) branches of this cascade. However, the emerging importance of a third branch, the cytochrome P450 pathway, offers novel alternatives for pain modulation. In contrast to the inflammatory cox and lox metabolites, the cytochrome P450-generated ARA metabolites, epoxyeicosatrienoic acids (EETs), are potent physiological anti-inflammatory/antihyperalgesic molecules. These bioactive lipids have very short half lives and inhibiting their degradation mediated by soluble epoxide hydrolase (sEH, EC 3.3.2.3) leads to antinociception. Inhibitors of sEH are effective in models of inflammatory and neuropathic pain, but it is unclear how sEH inhibition attenuates pain, warranting further investigation of this novel pathway. The sEH is well expressed in the peripheral and central nervous system, in the neurons and the glial cells, with spatial selectivity of expression among brain regions. Inhibitors of sEH (sEHIs) block nocifensive behavior, while leaving intact normal nociceptive and motor function. Thus in this study we will test the hypothesis that epoxygenated fatty acids and sEHI have a modulatory role in pain pathophysiology. To address this hypothesis, the levels of the EpFAs and their degradation products in the skin and the lumbar spinal cord in two distinct models of pain will be monitored. Levels of EpFAs will then be modulated in two distinct models of pain by different approaches; a) by exogenous EpFAs, b) by inhibiting the sEH, c)by inhibiting phosphodiesterases and d) by novel synthetic analogues of EpFAs that are resistant to sEH mediated degradation. We will utilize the carrageenan induced inflammatory pain model to identify anti- hyperalgesic EpFAs. The bioactivity of anti-nociceptive EpFAs will be validated in a clinically relevant model, the incisional pain model. We will use a combination of behavioral assays, biochemical and metabolomic analysis to understand the interactions of EpFAs with nociceptive pathways. We will produce a characteristic/diagnostic LC/MS/MS generated metabolomic profile of the intraplantar carrageenan and the plantar incisional pain models in rats. The incisional pain in particular is less well studied in regard to alterations in bioactive lipid metabolites. The target metabolites (80+ bioactive lipids) are from all three branches of the AA cascade including major cox, lox, cytochrome P450 and the sEH products including lipid metabolites of EPA and DHA, allowing us to investigate metabolite flux and crosstalk between biological cascades. The proposed studies will result in an advanced understanding of the antinociceptive mechanism of action of EpFAs and sEH inhibitors, and provide novel insights to the underlying mechanisms of lipidomic and biochemical alterations in response to pain. The knowledge generated by these experiments is likely to lead to the development of novel classes of agents that target pathways that have not been recognized previously.