Role of Vascular Smooth Muscle Bcl11b in Arterial Stiffness

动脉硬化（AS）是弹性动脉顺应性的逐渐丧失， 发展心血管疾病（CVD）。然而，AS的细胞和分子机制是 不太了解。确定这些机制可能会导致创新的战略，可以减缓或 逆转AS，从而降低发生CVD的风险。14号染色体上的基因沙漠位点， 基因Bcl 11b的下游，最近已被证明具有单核苷酸多态性 （SNPs）与AS高度相关。我们是第一个展示敲除Bcl 11b的人， 小鼠（BSMKO），无论是在全球范围内，特别是在血管平滑肌（VSM），引起升高的AS， 并增加血管紧张素II诱导的主动脉瘤的发病率。此外，我们发现Bcl 11b 在两种AS模型（高脂肪、高糖（HFHS）饮食的小鼠）的睾丸中表达下调 和老年小鼠），Bcl 11b转录调节收缩蛋白的表达，包括平滑 肌球蛋白（MYH 11）和平滑肌α-肌动蛋白（α-SMA）。综合考虑，我们假设 3 '-Bcl 11b基因沙漠区的SNP变体调节和抑制Bcl 11b的表达，发挥着重要作用。 在AS发病机制中的致病作用。 我们假设僵硬性垂体腺瘤Bcl 11b表达减少，刺激VSM的改变， 收缩表型和/或细胞外基质（ECM）重塑，或这些因子的组合， 从而损害主动脉的结构和功能完整性。在目标1中，我们将使用双轴机械 对野生型和BSMKO细胞的测试以及微观结构驱动的本构模型 分析基线时平滑肌细胞、弹性蛋白和胶原蛋白对主动脉壁硬度的贡献 以及在收缩激动剂刺激后。然后，我们将把双轴试验的结果与分子 VSM收缩、肌动蛋白聚合和粘着斑蛋白的表达， 到VSM的音调和僵硬。在目标2中，我们将使用染色质免疫沉淀（ChIP）测序， VSM匀浆以鉴定Bcl 11b VSM特异性DNA结合位点。我们还将检验假设 Bcl 11b通过募集组蛋白来表观遗传地调节MYH 11和α-SMA基因表达， 脱乙酰酶sirtuin-1在其基因启动子中的G/C基序。在目标3中，我们将使用 转基因小鼠，或者对于更多的翻译方法，通过施用AAV 2/5载体，以确定 如果增加Bcl 11b挽救受损的VSM特异性VSM收缩分子机制和/或 VSM细胞-细胞外基质相互作用（例如，收缩蛋白，粘着斑复合物，肌动蛋白 聚合）。VSM刚度和脉搏波速度连续降低， AS体内指数将确定靶向Bcl 11b是改善AS的可行策略 从而防止CVD。

Arterial stiffening (AS), the progressive loss of compliance in elastic arteries, increases the risk of developing cardiovascular diseases (CVDs). However, cellular and molecular mechanisms of AS are poorly understood. Determining these mechanisms may lead to innovative strategies that can slow or reverse AS and thus decrease the risk of developing CVD. A gene desert locus on chromosome 14, downstream of the gene Bcl11b, has recently been shown to harbor single nucleotide polymorphisms (SNPs) with a highly significant association with AS. We were the first to show knocking out Bcl11b in mice (BSMKO), both globally and specifically in vascular smooth muscle (VSM), caused elevated AS, and increased the incidence of angiotensin II-induced aortic aneurysms. In addition, we showed Bcl11b expression is downregulated in aortas of two models of AS (mice fed high fat, high sucrose (HFHS) diet and aged mice), and Bcl11b transcriptionally regulates contractile protein expression, including smooth muscle myosin (MYH11) and smooth muscle α-actin (α-SMA). Taken together, we hypothesize that SNP variants in the 3'-Bcl11b gene desert region regulate and suppress Bcl11b expression, playing a causative role in the pathogenesis of AS. We hypothesize that stiff aortas have decreased Bcl11b expression, stimulating alterations in VSM contractile phenotype and/or extracellular matrix (ECM) remodeling, or a combination of these factors, thereby impairing structural and functional integrity of the aorta. In Aim 1, we will use biaxial mechanical testing on wild type and BSMKO aortas together with a microstructurally-motivated constitutive model to dissect the contribution of smooth muscle cells, elastin and collagen to aortic wall stiffness, at baseline and after contractile agonist stimulation. We will then correlate the results of biaxial tests to molecular expressions of VSM contractile, actin polymerization, and focal adhesion proteins known to contribute to VSM tone and stiffness. In Aim 2, we will use chromatin immunoprecipitation (ChIP)-sequencing on VSM homogenates to identify Bcl11b VSM-specific DNA binding sites. We will also test the hypothesis that Bcl11b epigenetically regulates MYH11 and α-SMA gene expression by recruiting the histone deacetylase sirtuin-1 at G/C motifs in their gene promoters. In Aim 3, we will overexpress Bcl11b using transgenic mice or, for a more translational approach, by administering a AAV2/5 vector, to determine if increasing Bcl11b rescues impaired VSM-specific molecular mechanisms of VSM contraction and/or VSM cell-extracellular matrix interaction (e.g., contractile proteins, focal adhesion complexes, actin polymerization) in obese and aged mice. Definitive decreases in VSM stiffness and pulse wave velocity, the in vivo index of AS, would establish that targeting Bcl11b is a viable strategy for ameliorating AS and thus preventing CVD.