Enhancer of zeste homolog 2-mediated epigenetic activation of acid sphingomyelinase in endothelial dysfunction during obesity

项目摘要 内皮功能障碍是肥胖的早期缺陷，这是增加心血管疾病的主要原因。 发病率和死亡率，如动脉硬化、动脉粥样硬化和高血压。最近的研究 证明了酸性鞘磷脂酶（ASM）-神经酰胺信号传导在Nlrp 3的激发中的关键作用 肥胖和糖尿病期间的炎性小体和内皮功能障碍。本建议旨在 探索一种新的机制，介导ASM基因在EC中表达的转录控制，并确定 ASM表达和活性失调如何促进肥胖患者血管损伤。zeste增强子同源物 2（Ezh 2）是一种组蛋白甲基转移酶，通常抑制甲基化基因，作为一种关键的 基因表达的表观遗传调控机制。在初步研究中，我们发现Ezh 2的缺失 功能增加内膜中ASM表达和神经酰胺水平，导致内膜中新生内膜病变。 高脂饮食（HFD）小鼠的颈动脉。这种Ezh 2介导的ASM基因表达的抑制 和神经酰胺信号也在培养的EC中得到证实。基于这些观察，我们提出了一个 内皮细胞Ezh 2功能丧失上调ASM基因表达并增加神经酰胺的假设 在高脂血症条件下产生，其触发Nlrp 3炎性小体活化并产生 内皮损伤导致颈动脉壁新生内膜损伤。为了验证这个假设， 提出以下具体目标。特异性目标1将决定内皮ASM激活 由于Ezh 2功能的丧失在肥胖的早期阶段导致内皮功能障碍或损伤， 内皮特异性Ezh 2敲除小鼠（Ezh 2 ecKO）和它们的野生型同窝仔。具体目标2次尝试 为了测试Ezh 2调节的ASM活化如何通过研究以下因素的作用导致内皮功能障碍或损伤： 神经酰胺和富含神经酰胺的膜筏、Nlrp 3炎性小体活化、焦亡、内皮细胞凋亡、 依赖性血管舒张、内皮间连接破坏和适应性内皮祖细胞着陆 或分化。在具体目标3中，我们将探讨Ezh 2功能丧失的分子机制。 激活ASM，主要关注组蛋白和DNA甲基化在来自Ezh 2 ecKO的培养EC中的作用 小鼠和它们的野生型同窝仔。这一发现将为内皮细胞损伤的发病机制提供新的见解。 并鉴定Ezh 2-ASM途径作为预防或治疗空泡病治疗靶点 与肥胖有关。

Project Summary Endothelial dysfunction is an early defect in obesity, which is a major contributor to increased cardiovascular morbidity and mortality such as arterial stiffness, atherosclerosis, and hypertension. Recent studies have demonstrated a critical role of acid sphingomyelinase (ASM)-ceramide signaling in instigation of Nlrp3 inflammasomes and endothelial dysfunction during obesity and diabetes. The present proposal seeks to explore a novel mechanism mediating transcriptional control of ASM gene expression in ECs and determine how dysregulated ASM expression and activity promote vascular injury in obesity. Enhancer of zeste homolog 2 (Ezh2) is a histone methyltransferase that normally suppresses methylated genes, serving as a crucial epigenetic regulatory mechanism in gene expression. In preliminary studies, we found that loss of Ezh2 function increased ASM expression and ceramide levels in the intima leading to neointimal lesions in the carotid arteries of mice fed high fat diet (HFD). Such Ezh2-mediated suppression of ASM gene expression and ceramide signaling were also confirmed in cultured ECs. Based on these observations, we propose a hypothesis that loss of endothelial Ezh2 function upregulates ASM gene expression and augments ceramide production under hyperlipidemic conditions, which trigger Nlrp3 inflammasome activation and produce endothelial injury resulting in subsequent neointimal lesions on the carotid arterial wall. To test this hypothesis, the following Specific Aims are proposed. Specific Aim 1 will determine whether endothelial ASM activation due to loss of Ezh2 function contributes to endothelial dysfunction or injury at the early stage of obesity using endothelium-specific Ezh2 knockout mice (Ezh2ecKO) and their wild type littermates. Specific Aim 2 attempts to test how Ezh2-regulated ASM activation leads to endothelial dysfunction or injury by studying the role of ceramide and ceramide-enriched membrane rafts, Nlrp3 inflammasome activation, pyroptosis, endothelium- dependent vasodilation, inter-endothelial junction disruption, and adaptive endothelial progenitor cell landing or differentiation. In Specific Aim 3, we will explore the molecular mechanisms by which loss of Ezh2 function activates ASM with a main focus on the roles of histone and DNA methylation in cultured ECs from Ezh2ecKO mice and their wild type littermates. The findings will provide new insights into the pathogenesis of endothelial dysfunction and identify Ezh2-ASM pathway as therapeutic target for prevention or treatment of vaculopathy associated with obesity.