Eicosapentaenoic acid activation of free fatty acid receptor 4 prevents fibrosis, microvascular rarefaction, and diastolic dysfunction; implications for heart failure preserved ejection fraction

超过一半的心力衰竭(HF)诊断是心力衰竭伴保留射血分数(HFpEF)，这是一种复杂的 心力衰竭症状但左心室射血分数“正常”的综合征。HFpEF的特点是舒张期 功能障碍，充盈受损，肥大，钠尿肽升高。目前，还没有治疗方法 可提高HFpEF患者的存活率。HFpEF的病理重塑包括间质纤维化和 微血管疏松，导致舒张期僵硬、脑室充盈受损和运动 不宽容。纤维化是由于成纤维细胞激活导致细胞外基质产生增加所致， 成纤维细胞增殖，成纤维细胞向肌成纤维细胞转化。微血管稀疏与 炎症和内皮细胞死亡，导致氧气输送障碍，降低收缩和 舒张期储备，并加剧运动不耐受。我们发现ω-3-多不饱和脂肪酸(ω-3- 多不饱和脂肪酸)抑制心肌成纤维细胞转化，预防间质纤维化和舒张期功能障碍 部分类似于HFpEF(横动脉缩窄，TAC)的小鼠模型。此外，我们发现， 血中二十碳五烯酸(EPA)水平与预防间质纤维化有关，但EPA与此无关 在体内掺入成纤维细胞膜，这是一种传统的作用机制。相反，我们发现这是免费的 脂肪酸受体4(FFAR4)是一种G蛋白偶联的长链脂肪酸受体，足以和 防止心脏成纤维细胞转化所需的药物。其他组织报告称，ω3-PUFA 诱导后肢缺血血管生成，EPA激活FFAR4诱导血管内皮生长因子-A表达。因此， EPA在HFpEF中可能有预防纤维化和微血管疏松的双重好处。在人类中，ω3- 多不饱和脂肪酸可减少冠心病患者的猝死，但有效性问题仍然存在。然而，未能做到 达到足够的ω3-多不饱和脂肪酸水平，或一个治疗指数，可能解释先前的负面结果。因此，我们 假设饮食中摄取足够的EPA(治疗水平)和激活FFAR4可以预防纤维化， HFpEF中的微血管疏松和舒张期功能障碍。为了测试这一点，我们将：1)确定红细胞 EPA浓度(EPA指数)与预防纤维化、微血管疏松和舒张压相关 TAC后的功能障碍。我们还将通过cpdA将FFAR4作为潜在的靶点，cpdA是一种新的FFAR4配体 TAC后逆转重构的治疗方法。2)确定高水平的EPA是否可以防止 人类左心室肥厚和纤维化以及HFpEF的进展 动脉粥样硬化的民族研究试验(MESA)。3)使用成纤维细胞特异性FFAR4基因敲除小鼠来确定 FFAR4是EPA介导的预防纤维化、微血管疏松和舒张期功能障碍所必需的 跟在TAC后面。其影响将是：1)确定EPA的新的心脏保护功能，2)定义 EPA介导的心脏保护的治疗指标，3)为EPA的临床疗效提供证据 FFAR4，以防止HFpEF，以及4)描述了EPA的一种新的作用机制，即激活FFAR4。

Over half of all heart failure (HF) diagnoses are HF with preserved ejection fraction (HFpEF), a complex syndrome defined by symptoms of HF but a `normal' left ventricular EF. HFpEF is characterized by diastolic dysfunction, impaired filling, hypertrophy, and elevated natriuretic peptides. Presently, there are no treatments that improve survival in patients with HFpEF. Pathologic remodeling in HFpEF includes interstitial fibrosis and microvascular rarefaction, which contribute to diastolic stiffness, impaired ventricular filling, and exercise intolerance. Fibrosis results from activation of fibroblasts leading to increased extracellular matrix production, fibroblast proliferation, and fibroblast-to-myofibroblast transformation. Microvascular rarefaction is associated with inflammation and endothelial cell death, leading to impaired oxygen delivery, reducing systolic and diastolic reserve, and exacerbating exercise intolerance. We found that ω3-polyunsaturated fatty acids (ω3- PUFA) inhibited myofibroblast transformation and prevented interstitial fibrosis and diastolic dysfunction in a mouse model that partially resembles HFpEF (transverse aortic constriction, TAC). Further, we found that blood levels of eicosapentaenoic acid (EPA) correlated with prevention of interstitial fibrosis, but EPA was not incorporated into fibroblast membranes in vivo, a traditional mechanism of action. Rather, we found that free fatty acid receptor 4 (FFAR4), a G-protein coupled receptor for long-chain fatty acids, was sufficient and required to prevent myofibroblast transformation in cardiac fibroblasts. Other groups reported that ω3-PUFAs induce angiogenesis in hind-limb ischemia, and EPA activates FFAR4 to induce VEGF-A expression. Thus, EPA might have a dual benefit in HFpEF by preventing fibrosis and microvascular rarefaction. In humans, ω3- PUFAs reduce sudden death in coronary heart disease, but questions of efficacy remain. However, failure to achieve adequate ω3-PUFA levels, or a therapeutic index, likely explains prior negative results. Thus, we hypothesize that sufficient dietary uptake of EPA (therapeutic level) and activation of FFAR4 prevents fibrosis, microvascular rarefaction, and diastolic dysfunction in HFpEF. To test this we will: 1) Determine if erythrocyte EPA concentration (EPA-index) correlates with prevention of fibrosis, microvascular rarefaction, and diastolic dysfunction following TAC. We will also target FFAR4 with cpdA, a novel FFAR4 ligand, as a potential therapeutic approach to reverse remodeling following TAC. 2) Determine if high plasma levels of EPA prevent progression of left ventricular hypertrophy and fibrosis as well as HFpEF in humans using data from the Multi- Ethnic Study of Atherosclerosis trial (MESA). 3) Use fibroblast-specific FFAR4 knockout mice to determine if FFAR4 is required for EPA mediated prevention of fibrosis, microvascular rarefaction, and diastolic dysfunction following TAC. The impact will be to: 1) Identify novel cardioprotective functions of EPA, 2) Define a therapeutic index for EPA-mediated cardioprotection, 3) Provide evidence for the clinical efficacy of EPA/ FFAR4 to prevent HFpEF, and 4) Delineate a novel mechanism of action for EPA, activation of FFAR4.