Anti-Atherogenic Mechanisms of Drebrin

我们发现，肌动蛋白结合蛋白Dreplastin在平滑肌细胞（SMC）中大量表达 并且在小鼠中响应动脉损伤和在人类中响应动脉粥样硬化而上调。比较WT 在Dbn-/+小鼠中，我们发现Dreplatin在体外和体内均抑制SMC迁移和增殖， 通过稳定肌动蛋白丝。在SMC特异性Dbn-/-（SMC-Dbn-/-）小鼠的研究中，我们发现， 平滑肌细胞舒张期限制血管紧张素II诱导的升主动脉重塑， 下调NADPH氧化酶1（NOX 1），减少SMC活性氧（ROS）的产生， 减少血管炎症。因此，Dreplatin不仅限制了SMC的迁移/增殖表型， 还包括血管损伤和血管紧张素II引起的促炎性SMC表型。一致地，我们 发现SMC-Dbn-/-/Ldlr-/-小鼠的动脉粥样硬化程度高于同类SMC-Dbn+/+/Ldlr-/-小鼠。因为 平滑肌细胞负调节动脉粥样硬化，我们的目标是确定其机制， DREBELINE调节SMC促炎信号传导，以及DREBELINE增强SMC表达是否能 抑制动脉粥样硬化形成。为此，我们对Dbn-/-和同源基因进行了SILAC/质谱研究。 WT SMC。我们发现，在Dbn-/-SMC中，ROS防御酶谷胱甘肽-S-转移酶μ1 （GSTM 1）下调，促炎细胞因子CX 3CL 1（fractalkine）上调：因此，我们将 研究这些蛋白在介导Dbn-/-SMC表型中的作用。因为内吞作用 NOX 1调节ROS的产生和促炎性NFκB信号的激活， 显示抑制内吞作用，我们将研究是否drepletin抑制NOX 1介导的ROS的产生， 抑制内吞作用。我们研究的一个主要焦点将是促动脉粥样硬化SMC衍生的泡沫细胞， SMC谱系追踪研究表明，动脉粥样硬化病变中约40%为泡沫细胞。我们 初步研究表明，与同源Dbnflox/flox SMCs相比，Dbn-/- SMCs被诱导为 胆固醇负荷的转分化表现出巨噬细胞标志物，CD 68， 和Kruppel样因子4（KLF 4），一种SMC向泡沫细胞转分化所需的转录因子。通过 将Dbnflox/flox和SMC-Dbn-/-小鼠颈总动脉移植到同种Apoe-/-小鼠颈总动脉中， 在小鼠中，我们还表明Drexylase缺陷增强了SMC向体内CD 68+细胞的转分化。我们 将检验Dreplasty通过限制SMC转分化为泡沫细胞来抑制动脉粥样硬化形成的假设。 为了做到这一点，我们将确定Dredrone是否抑制SMC向泡沫细胞的转分化和相关的促分化因子。 炎症信号，在体外和体内;确定是否dreplastin抑制SMC转分化，通过 ROS依赖性机制;并定义GSTM 1和CX 3CL 1在介导Dreplasma抑制性细胞凋亡中的作用。 对SMC转分化和促炎信号传导的影响。

We discovered that the actin-binding protein, Drebrin, is abundantly expressed in smooth muscle cells (SMCs) and is up-regulated in response to arterial injury in mice and to atherosclerosis in humans. Comparing WT with Dbn-/+ mice, we found that Drebrin inhibits SMC migration and proliferation, both in vitro and in vivo, through stabilization of actin filaments. In studies with SMC-specific Dbn-/- (SMC-Dbn-/-) mice, we found that SMC Drebrin limits angiotensin II-induced remodeling of the ascending aorta, in a manner that correlates with down-regulation of NADPH oxidase 1 (NOX1), decreased SMC reactive oxygen species (ROS) production and reduced vascular inflammation. Thus, Drebrin constrains not only the migratory/proliferative SMC phenotype but also the pro-inflammatory SMC phenotype evoked by vascular injury and angiotensin II. Congruently, we found that atherosclerosis is greater in SMC-Dbn-/-/Ldlr-/- than in congenic SMC-Dbn+/+/Ldlr-/- mice. Because SMC Drebrin negatively regulates atherosclerosis, our goals are to determine the mechanisms by which Drebrin regulates SMC pro-inflammatory signaling and whether enhanced SMC expression of Drebrin can inhibit atherogenesis. To this end, we performed SILAC/mass spectrometry studies on Dbn-/- and congenic WT SMCs. We found that whereas in Dbn-/- SMCs the ROS-defensive enzyme Glutathione-S-transferase µ1 (GSTM1) is down-regulated, the pro-inflammatory cytokine CX3CL1 (fractalkine) is up-regulated: thus we will investigate the role of these proteins in mediating the phenotype of Dbn-/- SMCs. Because endocytosis of NOX1 regulates ROS generation and activation of pro-inflammatory NFκB signaling and Drebrin has been shown to inhibit endocytosis, we will investigate whether Drebrin inhibits NOX1-mediated ROS generation by inhibiting endocytosis. A major focus of our studies will be pro-atherogenic SMC-derived foam cells, which SMC lineage tracing studies have shown, comprise ~40% of foam cells in atherosclerotic lesions. Our Preliminary Studies show that, compared with cognate Dbnflox/flox SMCs, Dbn-/- SMCs induced to transdifferentiate with cholesterol loading exhibited increased expression of the macrophage marker, CD68, and Kruppel-like factor 4 (KLF4), a transcription factor required for SMC-to-foam cell transdifferentiation. By grafting common carotid arteries from Dbnflox/flox and SMC-Dbn-/- mice into carotid arteries of congenic Apoe-/- mice, we also show that Drebrin deficiency augments transdifferentiation of SMCs to CD68+ cells in vivo. We will test the hypothesis that Drebrin inhibits atherogenesis by limiting SMC transdifferentiation into foam cells. To do so, we will establish whether Drebrin inhibits SMC-to-foam cell transdifferentiation and associated pro- inflammatory signaling, both in vitro and in vivo; determine if Drebrin inhibits SMC transdifferentiation through ROS-dependent mechanisms; and define the roles of GSTM1 and CX3CL1 in mediating Drebrin’s inhibitory effects on SMC transdifferentiation and pro-inflammatory signaling.