Nuclear FAK-mediated VSMC differentiation via epigenetic reprograming invascular diseases

VSMCs在血管损伤时去分化为增殖状态或转分化为巨噬细胞样 动脉粥样硬化进展过程中的细胞(MLCs)。VSMC表型切换由多个 转录和表观遗传变化导致收缩基因减少导致增殖增加 表达和增加基质产生，这对动脉粥样硬化病变不利，直接 针对这一过程的干预性研究一直缺乏。增加的基质和增长因素改变 整合素信号转导并导致粘着斑激酶(FAK)异常激活，从而可能促进VSMC 表型转换。我们证明了FAK是不活跃的，主要定位于VSMC的胞核。 健康的动脉。然而，血管损伤促进了FAK的激活和胞浆重定位，从而 细胞周期蛋白D1转录和细胞周期增加。而我们发现抑制VSMC中FAK的活性 诱导FAK的核定位和促进收缩基因转录，其潜在机制是通过 目前尚不清楚是哪种FAK调节了收缩基因。我们已经鉴定出DNA甲基转移酶3A (DNMT3A)和核小体重塑和脱乙酰酶(NuRD)复合体，两个关键的表观遗传抑制 机械，作为VSMCs中与核FAK互动的合作伙伴。FAK抑制降低DNMT3A和NuRD 组分表达，这与DNA甲基化减少和组蛋白活性增加有关 收缩基因启动子内的标记。使用遗传FAK细胞质(Cyto)限制的VSMCs，我们发现 核FAK是降低DNMT3A/NuRD和增加收缩基因表达所必需的。 此外，与WT小鼠相比，ApoE-/-；FAK-Cyto小鼠表现出动脉粥样硬化的增加，这表明 活跃的胞浆FAK加重动脉粥样硬化。此外，FAK显示胞浆定位增加。 与健康标本相比，人类动脉粥样硬化病变内的活性。重要的是，FAK抑制剂 ApoE-/-小鼠晚期动脉粥样硬化病变减少，这与DNMT3A和DNMT3A减少有关 随着纤维帽内ACTA2+细胞的增多，NuRD组分表达增强。我们的假设是FAK 催化抑制增强FAK核定位并通过减少表达促进VSMC分化 表观遗传调控因子DNMT3A和NuRD复合体。目标1将阐明其分子机制。 核FAK通过DNA甲基化、染色质表观遗传调控介导的VSMC表型转换 重塑和组蛋白修饰。目标2将研究DNMT3A和NuRD复合体在 使用DNMT3A和NuRD遗传模型研究血管损伤时的VSMC去分化。目标3将 评价FAK抑制剂阻断VSMC转分化及促进斑块稳定性的作用 早期和晚期动脉粥样硬化。这项研究将为VSMC表型转换提供新的见解 FAK通过DNMT3A和NuRD复合体的稳定性介导表观遗传调控。药物的治疗潜力 FAK抑制剂在减轻血管损伤和动脉粥样硬化中的内膜增厚方面也将得到评估。

VSMCs dedifferentiate into a proliferative state upon vessel injury or transdifferentiate into macrophage-like cells (MLCs) during atherosclerosis progression. VSMC phenotypic switching are driven by multiple transcriptional and epigenetic changes that lead to increased proliferation with reduced contractile gene expression and increased matrix production, which is detrimental to atherosclerotic lesions, direct interventional studies that target this process have been lacking. Increased matrix and growth factors alter integrin signaling and leads to aberrant focal adhesion kinase (FAK) activation, which may promote VSMC phenotypic switching. We demonstrated that FAK is inactive and primarily localized in the nuclei of VSMCs of healthy arteries. However, vessel injury promoted FAK activation and cytoplasmic relocalization, which increased cyclin D1 transcription and cell cycling. While we found that inhibition of FAK activity in VSMCs induced nuclear localization of FAK and increased contractile gene transcription, the underlying mechanism by which FAK regulates the contractile genes is not known. We have identified DNA methyltransferase 3A (DNMT3A) and the nucleosome remodeling and deacetylase (NuRD) complex, two key epigenetic repression machineries, as nuclear FAK-interacting partners in VSMCs. FAK inhibition decreased DNMT3A and NuRD component expression, which was associated with decreased DNA methylation and increased active histone marks within contractile gene promoters. Using genetic FAK cytoplasmic (Cyto) restricted VSMCs, we found that nuclear FAK is required for reducing DNMT3A/NuRD and for increasing contractile gene expression. Additionally, ApoE-/-;FAK-Cyto mice showed increased atherosclerosis compared to WT mice, suggesting that active cytoplasmic FAK exacerbates atherosclerosis. Further, FAK showed increased cytoplasmic localization and activity within human atherosclerotic lesions compared to healthy specimens. Importantly, FAK inhibitor reduced advanced atherosclerotic lesions in ApoE-/- mice, which was associated with reduced DNMT3A and NuRD component expression with increased ACTA2+ cells in the fibrous cap. Our hypothesis is that FAK catalytic inhibition forces FAK nuclear localization and promotes VSMC differentiation via reduced expression of epigenetic regulators DNMT3A and NuRD complex. Aim 1 will elucidate the molecular mechanism of nuclear FAK-mediated VSMC phenotypic switching via epigenetic modulation of DNA methylation, chromatin remodeling, and histone modification. Aim 2 will investigate the role of DNMT3A and the NuRD complex in VSMC dedifferentiation upon vascular injury using both DNMT3A and NuRD genetic models. Aim 3 will evaluate the effect of FAK inhibition on blocking VSMC transdifferentiation and promoting plaque stability in early and advanced atherosclerosis. This study will provide new insights into VSMC phenotype switching via FAK-mediated epigenetic control through DNMT3A and NuRD complex stability. The therapeutic potential of FAK inhibitors in alleviating intimal thickening in vascular injury and atherosclerosis will also be assessed.