Role of Soluble Epoxide Hydrolase in Alcohol-Associated Liver Disease

酒精相关性肝病（ALD）是一系列肝脏疾病，范围从肝脂肪变性到肝硬化。 伴有不同程度纤维化和肝硬化的脂肪性肝炎。ALD是发病率、死亡率和并发症的主要原因。 美国和全世界的医疗费用。然而，没有FDA批准的治疗ALD的任何阶段。 对疾病进展的机制和介质也没有完全理解， 严重性。可溶性环氧化物水解酶（s-EH）是调节内源性代谢的主要酶， 生物活性脂质（例如，环氧脂肪酸（Ep-FAs）最近被认为是一种新兴的治疗靶点 多种疾病。该项目的总体目标是测试在以下情况下抑制s-EH的治疗效果： 酒精性肝脏疾病严重程度的不同阶段，并为使用s-EH抑制作为新的治疗方法提供机制基础 治疗酒精引起的肝脏病理。目标1.检测s-EH抑制剂的治疗效果 作为ALD的新治疗策略。我们将：i）测试是否通过药理学方法调节s-EH活性， 抑制或s-EH（Ephx 2）基因消融可有效减轻或预防EtOH诱导的肝损伤， ii）评估s-EH抑制是否导致血液和肝脏Ep-FA的稳定;和iii） 将Ep-FA水平的变化与肝损伤的标志物相关联。全身和靶向肝脏特异性递送s- EH抑制剂（t-TUCB和TPPU）以及WT和Ephx 2-/-小鼠（整体和肝脏特异性）将用于多个实验中。 ALD的动物模型，其产生疾病严重程度的不同阶段，并概括了不同的 人类ALD的特征。这允许在不同阶段严格评价s-EH抑制的效果 严重的ALD。将采用治疗和预防模式。目标2.探讨机制 这是实验性ALD中s-EH抑制的有益作用的基础。我们将确定Ep-FA是否 由s-EH抑制介导的稳定化在减轻EtOH诱导的肝损伤中起关键作用。我们将 测试n3与n6 Ep-FA是否通过增强M2巨噬细胞极化发挥更大的有益作用， 增加促恢复/促消退巨噬细胞表型，并通过Ep-FAs-PPARγ-CXCL 1介导 中性粒细胞浸润减少。s-EH抑制剂，WT，转基因fat-1小鼠（内源性转化n6 PUFA至n3 PUFA）和肝细胞特异性Pparγ-/-小鼠。将进行体内和体外研究， 执行。目标3：评价乙醇诱导的人ALD中s-EH和Ep-FA的变化。利用de- i）评价血浆Ep-FA的改变， 建立酒精中毒患者肝损伤生物标志物和全身炎症之间的关系， ii）确定n3-PUFA膳食补充剂对急性肝炎（AH）患者血浆Ep-FA的影响; 重度饮酒个体;和iii）测试s-EH抑制和Ep-FA（包括n3和n6 Ep-FA两者） 可有效降低全血中基础和LPS刺激的促炎细胞因子产生， 从AH患者获得的外周血单核细胞。

Alcohol-associated liver disease (ALD) is a spectrum of liver disorders ranging from hepatic steatosis to steatohepatitis with varying degrees of fibrosis and cirrhosis. ALD is a major cause of morbidity, mortality, and health care costs in the US and worldwide. However, there is no FDA-approved therapy for any stage of ALD. There is also an incomplete understanding of the mechanisms and mediators of disease progression and severity. Soluble epoxide hydrolase (s-EH), a master enzyme that regulates the metabolism of endogenous bioactive lipids (e.g., epoxy-fatty acids, Ep-FAs), has recently been recognized as an emerging therapeutic target in multiple diseases. The overall goal of this project is to test the therapeutic efficacy of s-EH inhibition at different stages of ALD severity, and to provide a mechanistic foundation for using s-EH inhibition as a novel therapy for alcohol-induced liver pathology. Aim 1. To test the therapeutic effectiveness of s-EH inhibition as a novel therapeutic strategy for ALD. We will: i) test whether modulation of s-EH activity by pharmacological inhibition or s-EH (Ephx2) genetic ablation can effectively attenuate or prevent EtOH-induced liver injury in experimental ALD; ii) evaluate whether s-EH inhibition leads to stabilization of blood and liver Ep-FAs; and iii) correlate changes in Ep-FA levels with markers of liver injury. Systemic and targeted liver-specific delivery of s- EH inhibitors (t-TUCB and TPPU), and WT and Ephx2 -/- mice (global and liver-specific) will be used in multiple animal models of ALD, which produce different stages of disease severity and which recapitulate different features of human ALD. This allows for a rigorous evaluation of the effects of s-EH inhibition at different stages of ALD severity. Treatment and prevention paradigms will be applied. Aim 2. To explore mechanism(s) underlying the beneficial effects of s-EH inhibition in experimental ALD. We will determine whether Ep-FA stabilization mediated by s-EH inhibition plays a critical role in attenuation of EtOH-induced liver injury. We will test whether n3 vs n6 Ep-FAs exert a greater beneficial effect by enhancing M2 macrophage polarization, increasing a pro-restorative/pro-resolving macrophage phenotype, and by Ep-FAs-PPARγ-CXCL1-mediated reduction of neutrophil infiltration. s-EH inhibitors, WT, transgenic fat-1 mice (which endogenously convert n6 PUFAs to n3 PUFAs), and hepatocyte-specific Pparγ-/- mice will be used. In vivo and in vitro studies will be performed. Aim 3. To evaluate EtOH-induced alterations in s-EH and Ep-FAs in human ALD. Utilizing de- identified human plasma and whole blood samples, we will: i) evaluate alterations in plasma Ep-FAs and establish relationships between biomarkers of liver injury and systemic inflammation in patients with alcohol- associated hepatitis (AH); ii) determine the effects of n3-PUFA dietary supplementation on plasma Ep-FAs in heavy drinking individuals; and iii) test whether s-EH inhibition and Ep-FAs (including both n3 and n6 Ep-FAs) can effectively decrease basal and LPS-stimulated pro-inflammatory cytokine production in whole blood and peripheral blood monocytes obtained from AH patients.