Anti-Atherogenic Mechanisms of Drebrin

Drebrin 的抗动脉粥样硬化机制

• 批准号: 10532356
• 负责人: NEIL J. FREEDMAN
• 金额: $ 52.33万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-12-16 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10532356
• 关键词: Actin-Binding Protein; Angiotensin II; Antiatherogenic; Aorta; Apolipoprotein E; Arterial Fatty Streak; Arterial Injury; Arteries; Atherosclerosis; CX3CL1 gene; Carotid Arteries; Cd68; Cell Lineage; Cell Proliferation; Cells; Cholesterol; Common carotid artery; Congestive Heart Failure; Development; Disease; Down-Regulation; Dynamin; Endocytosis; Endocytosis Inhibition; Enzymes; Exhibits; Foam Cells; Fractalkine; GKLF protein; Generations; Genes; Glutathione Metabolism Pathway; Glutathione S-Transferase; Goals; Human; In Vitro; Inflammatory; Knock-in; Knowledge; LoxP-flanked allele; Macrophage; Mass Spectrum Analysis; Mediating; Medical; Memory; Microfilaments; Mission; Molecular; Morbidity - disease rate; Mus; Myocardial Infarction; NADPH Oxidase 1; Neurons; Nuclear; Nuclear Translocation; Phagocytes; Phenotype; Prevalence; Process; Production; Proteins; Public Health; Reactive Oxygen Species; Research; Role; Signal Transduction; Smooth Muscle Myocytes; Stroke; Testing; United States National Institutes of Health; Vascular Smooth Muscle; ascending aorta; atherogenesis; burden of illness; cell motility; congenic; cytokine; disability; drebrins; in vivo; mortality; novel therapeutic intervention; oxidized low density lipoprotein; response; transcription factor; transdifferentiation; vascular inflammation; vascular injury; western diet

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.

我们发现肌动蛋白结合蛋白Drebrin在平滑肌细胞(SMC)中大量表达。 在小鼠的动脉损伤和人类的动脉粥样硬化中表达上调。比较WT 在DBN-/+小鼠身上，我们发现Drebrin在体外和体内都抑制了SMC的迁移和增殖， 通过稳定肌动蛋白细丝。在对SMC特异性DBN-/-(SMC-DBN-/-)小鼠的研究中，我们发现 SMC Drebrin抑制血管紧张素II诱导的升主动脉重塑，其方式与 下调NADPH氧化酶1(NOX1)，减少SMC活性氧自由基(ROS)的产生，并 减少血管炎症。因此，Drebrin不仅抑制迁移/增殖的SMC表型 也包括由血管损伤和血管紧张素II引起的促炎性SMC表型。 发现SMC-DBN-/-/Ldlr-/-小鼠的动脉粥样硬化程度比同基因SMC-DBN+/+/Ldlr-/-小鼠更严重。因为 SMC Drebrin负性调节动脉粥样硬化，我们的目标是确定其机制 Drebrin调节SMC促炎信号及增强SMC表达是否可以 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/MS研究 WT SMCs。我们发现，在DBN-/-SMC中，ROS防御酶谷胱甘肽-S-转移酶µ1 (GSTM1)下调，促炎症细胞因子CX3CL1(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。因为细菌的内吞作用 NOX1调节ROS的产生和激活促炎症的NFκB信号和Drebrin已被 我们将研究Drebrin是否通过以下方式抑制NOX1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的SMC来源的泡沫细胞，它 SMC谱系追踪研究表明，在动脉粥样硬化病变中，SMC约占泡沫细胞的40%。我们的 初步研究表明，与同源Dbnflx/FLOX SMC相比，DBN-/-SMC诱导 随着胆固醇负荷的增加，巨噬细胞标志物CD68的表达增加， Kruppel样因子4(KLF4)，SMC向泡沫细胞转分化所需的转录因子。通过 Dbnflx/Flox和SMC-DBN-/-小鼠颈总动脉移植到同种APOE-/-的颈动脉 我们还发现，在体内，Drebrin缺乏会增强SMC向CD68+细胞的转分化。我们 将验证Drebrin通过限制SMC向泡沫细胞的转分化来抑制动脉粥样硬化形成的假设。 为此，我们将确定Drebrin是否抑制SMC到泡沫细胞的转分化和相关的促泡沫细胞分化。 体外和体内的炎症信号；确定Drebrin是否通过 ROS依赖的机制；并确定GSTM1和CX3CL1在介导Drebrin抑制中的作用 对SMC转分化和促炎信号的影响。

项目成果

Drebrin attenuates atherosclerosis by limiting smooth muscle cell transdifferentiation.

• DOI: 10.1093/cvr/cvab156
• 发表时间: 2021-04
• 期刊: Cardiovascular research
• 影响因子: 10.8
• 作者: Jiao‐Hui Wu;Lisheng Zhang;Igor Nepliouev;L. Brian;Taiqin Huang;Kamie P Snow;B. Schickling;E. Hauser;F. Miller;N. Freedman;Jonathan A Stiber