Lipid macropinocytosis: a novel target in atherosclerotic cardiovascular disease

脂质巨胞饮作用：动脉粥样硬化性心血管疾病的新靶点

• 批准号: 10401923
• 负责人: Gabor Csanyi
• 金额: $ 38.5万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10401923
• 关键词: Actin-Binding Protein; Affinity; Antibodies; Antigen Presentation; Aorta; Apolipoprotein E; Area; Arterial Fatty Streak; Arteries; Atherosclerosis; Attenuated; Binding; Blocking Antibodies; CD36 gene; CD47 gene; Cardiovascular system; Cause of Death; Cell Fractionation; Cell membrane; Cells; Cholesterol; Chronic; Confocal Microscopy; Data; Development; Dissociation; Exhibits; Extracellular Matrix Proteins; Flow Cytometry; Fluorescence; Foam Cells; Genetic; High Pressure Liquid Chromatography; Human; In Vitro; Inflammation; Inflammatory; Knock-out; Knockout Mice; Knowledge; Link; Lipid-Laden Macrophage; Lipids; Lipoproteins; Low Density Lipoprotein Receptor; Low Density Lipoprotein oxidation; Low-Density Lipoproteins; Mediating; Membrane; Microscopy; Modification; Mus; NHE1; Other Genetics; Pathogenesis; Pathway interactions; Pharmacology; Phenotype; Phospholipase C; Phosphoric Monoester Hydrolases; Physiological; Play; Process; Protein Dephosphorylation; Protein Isoforms; Proteins; Receptor Signaling; Reporting; Resolution; Role; Signal Transduction; Small Interfering RNA; Smooth Muscle Myocytes; Subendothelial Layer; Techniques; Testing; Thrombospondin 1; Time; United States; Vascular Smooth Muscle; Western Blotting; Woman; cofilin; cost; health care delivery; in vivo; inhibitor; innovation; macrophage; men; mouse genetics; mutant; nanoparticle; novel; overexpression; platelet-derived growth factor BB; protein aminoacid sequence; receptor; scavenger receptor; therapeutic target; tool; uptake; vascular inflammation

Project Summary Atherosclerosis, the main underlying cause of death worldwide, is characterized by chronic inflammation and accumulation of lipids in the arterial wall. Excessive lipid accumulation by macrophages (MΦs) and vascular smooth muscle cells (SMCs) plays a key role in the initiation and progression of atherosclerosis. It is generally accepted that atherosclerosis arise from LDL modification in the arterial wall and its subsequent internalization by MΦs and SMCs through a variety of scavenger receptors. The role of scavenger receptor-independent lipid uptake in atherosclerosis, the physiological factors stimulating this process, and the signaling mechanisms involved remain poorly characterized. We reported that matrix protein thrombospondin-1 (TSP1) stimulates direct, scavenger receptor-independent uptake of unmodified, native LDL (nLDL) in MΦs. TSP1 via its cognate receptor CD47 activates actin-binding protein cofilin, leading to macropinocytosis of nLDL, and excessive cholesterol accumulation. The signaling mechanisms downstream of CD47 that stimulate macropinocytosis are unknown. Although phospholipase C (PLC) and slingshot phosphatase 1 (SSH1) have been shown to activate cofilin, their roles in TSP1-induced macropinocytosis have not yet been investigated. Moreover, the ability of TSP1-CD47 signaling to stimulate MΦ macropinocytosis in atherosclerotic vessels in vivo and the significance of lipid macropinocytosis in the pathogenesis of atherosclerosis remain to be determined. Our novel preliminary data show that TSP1 and CD47 knockout mice are protected from atherosclerosis and the macropinocytosis inhibitor EIPA decreases atherosclerotic lesion formation in hypercholesterolemic mice. We hypothesize that TSP1 via CD47 promotes lipid macropinocytosis in the arterial wall, contributing to lipid accumulation and the pathogenesis of atherosclerosis. The hypothesis will be tested via the following aims: (1) examining for the first time whether CD47 receptor signaling in MΦs stimulates macropinocytosis via PLC- and SSH1-mediated cofilin activation and contributes to atherosclerosis development; (2) exploring whether MΦs internalize lipoproteins via macropinocytosis in atherosclerotic arteries in vivo and that deletion of NHE1 (major target of EIPA) specifically in MΦs attenuates atherosclerosis, and (3) investigating whether TSP1 binding to CD47 stimulates macropinocytosis in MΦ-like SMCs via Nox1-mediated cofilin activation. The proposal will employ global and cell-specific knockout mice, and other genetic tools to test the hypothesis. Specific targeting of CD47 via multiple approaches (antibody blockade, siRNA/morpholino silencing, and CD47 activating peptide sequences) will provide confirmation of results obtained in genetic mutants. Multiple complementary techniques will be used to study macropinocytosis in vitro (pharmacological, genetic, fluorescence/high-resolution microscopy) and in vivo (cofilin mutants, AngioSPARK 680, MΦ-specific NHE1 knockouts). This innovative proposal has the potential to reveal important new mechanisms of lipid internalization and provide a paradigm shift in our knowledge about how atherosclerosis develops.

项目概要 动脉粥样硬化是全世界死亡的主要根本原因，其特点是慢性炎症和 动脉壁脂质积聚 巨噬细胞 (MΦs) 和血管脂质积聚过多。 平滑肌细胞（SMC）通常在动脉粥样硬化的发生和进展中发挥着关键作用。 公认动脉粥样硬化是由动脉壁中的低密度脂蛋白修饰及其随后的内化引起的 由MΦs和SMCs通过多种清道夫受体产生非清道夫受体依赖性脂质的作用。 动脉粥样硬化的摄取、刺激该过程的生理因素以及信号传导机制 我们报道了基质蛋白血小板反应蛋白-1 (TSP1) 的刺激作用。 MΦs 中的未修饰天然 LDL (nLDL) 通过其同源直接、不依赖清道夫受体的摄取。 受体 CD47 激活肌动蛋白结合蛋白 cofilin，导致 nLDL 巨胞饮作用，并过量 CD47 下游刺激巨胞饮作用的信号机制是胆固醇积累。 未知，尽管磷脂酶 C (PLC) 和弹弓磷酸酶 1 (SSH1) 已被证明可以激活。 cofilin 在 TSP1 诱导的巨胞饮作用中的作用尚未得到研究。 TSP1-CD47信号在体内刺激动脉粥样硬化血管MΦ巨胞饮作用及其意义 脂质巨胞饮作用在动脉粥样硬化发病机制中的作用仍有待我们的初步研究。 数据显示 TSP1 和 CD47 敲除小鼠免受动脉粥样硬化和巨胞饮作用 抑制剂 EIPA 可减少高胆固醇血症小鼠的动脉粥样硬化病变形成。 TSP1 通过 CD47 促进动脉壁中的脂质巨胞饮作用，有助于脂质积累和 动脉粥样硬化的发病机制将通过以下目标进行检验：（1）检查第一个目标。 MΦs 中的 CD47 受体信号传导是否通过 PLC 和 SSH1 介导的丝切蛋白刺激巨胞饮作用的时间 (2)探讨MΦs是否通过以下途径内化脂蛋白： 体内动脉粥样硬化动脉中的巨胞饮作用以及NHE1（EIPA的主要靶标）的特异性缺失 MΦs 中的 TSP1 可以减轻动脉粥样硬化，并且 (3) 研究 TSP1 与 CD47 的结合是否会刺激 通过 Nox1 介导的丝切蛋白激活 MΦ 样 SMC 中的巨胞饮作用 该提案将采用全局和 细胞特异性基因敲除小鼠和其他遗传工具通过多重特异性靶向 CD47 来检验这一假设。 方法（抗体阻断、siRNA/吗啉沉默和 CD47 激活肽序列）将 将使用多种互补技术来确认在基因突变体中获得的结果。 研究体外（药理学、遗传学、荧光/高分辨率显微镜）和体内巨胞饮作用 （cofilin 突变体、AngioSPARK 680、MΦ 特异性 NHE1 敲除）这一创新提案有潜力。 揭示脂质内化的重要新机制，并为我们的知识提供范式转变 动脉粥样硬化如何发展。

项目成果

PKCδ stimulates macropinocytosis via activation of SSH1-cofilin pathway.

• DOI: 10.1016/j.cellsig.2018.09.018
• 发表时间: 2019-01
• 期刊: Cellular signalling
• 影响因子: 4.8
• 作者: Singla B;Lin HP;Ghoshal P;Cherian-Shaw M;Csányi G
• 通讯作者: Csányi G

Nf1 heterozygous mice recapitulate the anthropometric and metabolic features of human neurofibromatosis type 1.

• DOI: 10.1016/j.trsl.2020.08.001
• 发表时间: 2021-03
• 期刊: Translational research : the journal of laboratory and clinical medicine
• 作者: Tritz R;Benson T;Harris V;Hudson FZ;Mintz J;Zhang H;Kennard S;Chen W;Stepp DW;Csanyi G;Belin de Chantemèle EJ;Weintraub NL;Stansfield BK
• 通讯作者: Stansfield BK

Sildenafil improves vascular endothelial function in patients with cystic fibrosis.

• DOI: 10.1152/ajpheart.00301.2018
• 发表时间: 2018-11
• 期刊: American journal of physiology. Heart and circulatory physiology
• 作者: Paula Rodriguez-Miguelez;N. Lee;M. Tucker;G. Csányi;K. McKie;C. Forseen;R. Harris