SMC macropinocytosis: a novel target in atherosclerotic vascular disease

SMC巨胞饮作用：动脉粥样硬化性血管疾病的新靶点

基本信息

批准号：
10735697
负责人：
Gabor Csanyi
金额：
$ 56.37万
依托单位：
AUGUSTA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-05-15 至 2028-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10735697
关键词：
3-Dimensional Actins Adopted Antibodies Aorta Area Arterial Fatty Streak Arteries Atherosclerosis Attenuated Blocking Antibodies Blood Brain CD36 gene CD47 gene Cell Lineage Cells Cessation of life Cre driver Data Development Disease Female Fluorescence Foam Cells Genetic Genomics Goals Human Image Immunoelectron Microscopy In Vitro Knockout Mice Leukocytes Ligands Lipids Low-Density Lipoproteins Mediating Membrane Microscopy Molecular Morbidity - disease rate Mus Myocardial Infarction Myocardium Other Genetics Oxygen PTPNS1 gene Phenotype Physiological Polymers Process Proteins Regulation Resolution Role Rupture Signal Transduction Smooth Muscle Myocytes Specificity Stroke Techniques Testing Therapeutic Intervention Thrombospondin 1 Vascular Smooth Muscle effective therapy genetic approach high resolution imaging human morbidity human mortality imaging approach in vivo indexing insight male mortality mouse model new therapeutic target novel pharmacologic polymerization prevent protein aminoacid sequence receptor reconstruction scavenger receptor single-cell RNA sequencing therapeutic target tool transdifferentiation uptake

项目摘要

Project Summary Atherosclerosis is a leading cause of morbidity and mortality globally. Recently, cell lineage tracing, single-cell RNA sequencing and human genomic studies have been integrated to demonstrate that a) majority of plaque foam cells are of vascular smooth muscle cell (SMC) origin and b) SMC can undergo a fate switch to transitional, multipotential cells that can adopt plaque altering phenotypes. Although these results identify SMC as potential therapeutic targets, most current treatments for atherosclerosis have little direct impact on SMC. The endocytic processes by which SMC take up lipids and become foam cells in the arterial wall are not clearly defined. In addition, the mechanisms underlying SMC phenotypic switching in the arterial wall remain largely unknown. Using a combination of high-resolution imaging, 3D cell reconstruction, and LDL immunolabeling, we have recently demonstrated that foam cells (lineage unknown) in human and murine atherosclerotic plaques internalize LDL via macropinocytosis. Novel preliminary data using SMC lineage tracing identifies SMC subsets performing macropinocytosis in the arterial wall in vivo. Stimulation of macropinocytosis of LDL in SMC promotes dedifferentiation and phenotype switching into plaque-promoting phenotypes in vitro. Further, pharmacological blockade of macropinocytosis using the Na+/H+ exchanger 1 (NHE1) blocker, EIPA, inhibits LDL uptake in atherosclerotic arteries and abrogates atherosclerosis development in multiple murine models of atherosclerosis. Preliminary data also show that the matricellular protein thrombospondin-1 (TSP1) stimulates macropinocytosis via CD47 in SMC, leading to foam cell formation and phenotypic switching, and global Cd47-/- mice are protected from atherosclerosis. Based on these observations, we hypothesize that SMC macropinocytosis drives atherosclerosis through foam cell formation and regulation of SMC phenotypic switching. The hypothesis will be tested via the following specific aims: Aim 1: SMC internalize LDL via macropinocytosis in atherosclerotic arteries. Aim 2: SMC-specific inhibition of macropinocytosis attenuates transdifferentiation of SMC into a plaque- promoting phenotype and inhibits atherosclerosis. Aim 3: TSP1 via CD47 stimulates SMC macropinocytosis, promotes SMC phenotypic switching and contributes to atherosclerosis development. The proposal will employ SMC-specific knockout mice (Nhe1-/-, Cd47-/- and Cd36-/-), primary human and murine aortic SMC, a vascular SMC-restricted Cre driver mouse model (Itga8-CreERT2+/-; mTmG+/-), and other genetic tools to test the hypothesis. Multiple complementary techniques will be used to study SMC macropinocytosis in vitro (pharmacological, genetic, fluorescence/SEM imaging) and in vivo (TEM/IEM microscopy, SMC-specific Nhe1 knockout mice). At their conclusion, the proposed studies will define key mechanism(s) promoting SMC foam cell formation and phenotypic switching and potentially identify new targets for therapeutic interventions of atherosclerosis.

项目概要动脉粥样硬化是全球发病和死亡的主要原因。最近，细胞谱系追踪、单细胞 RNA 测序和人类基因组研究相结合，证明：a) 大多数斑块泡沫细胞起源于血管平滑肌细胞 (SMC)，并且 b) SMC 可以经历命运转变为过渡性、可以采用斑块改变表型的多能细胞。尽管这些结果表明 SMC 具有潜力治疗目标，目前大多数动脉粥样硬化治疗对 SMC 几乎没有直接影响。内吞性 SMC 吸收脂质并成为动脉壁泡沫细胞的过程尚不清楚。在此外，动脉壁中 SMC 表型转换的机制仍然很大程度上未知。通过结合高分辨率成像、3D 细胞重建和 LDL 免疫标记，我们获得了最近证明，人类和小鼠动脉粥样硬化斑块中的泡沫细胞（谱系未知）通过巨胞饮作用内化 LDL。使用 SMC 谱系追踪的新颖初步数据识别 SMC 子集在体内的动脉壁中进行巨胞饮作用。刺激 SMC 中 LDL 的巨胞饮作用可促进体外去分化和表型转变为促进斑块的表型。此外，药理使用 Na+/H+ 交换器 1 (NHE1) 阻断剂 EIPA 阻断巨胞饮作用，抑制 LDL 摄取动脉粥样硬化并消除多种动脉粥样硬化小鼠模型中的动脉粥样硬化发展。初步数据还表明，基质细胞蛋白血小板反应蛋白-1 (TSP1) 可刺激巨胞饮作用通过SMC中的CD47，导致泡沫细胞形成和表型转换，全局Cd47-/-小鼠受到保护来自动脉粥样硬化。基于这些观察，我们假设 SMC 巨胞饮作用驱动通过泡沫细胞形成和调节 SMC 表型转换来调节动脉粥样硬化。假设将是通过以下具体目标进行测试：目标 1：SMC 通过动脉粥样硬化中的巨胞饮作用内化 LDL 动脉。目标 2：SMC 特异性抑制巨胞饮作用，减弱 SMC 向斑块的转分化促进表型并抑制动脉粥样硬化。目标 3：TSP1 通过 CD47 刺激 SMC 巨胞饮作用，促进 SMC 表型转换并有助于动脉粥样硬化的发展。该提案将采用 SMC 特异性敲除小鼠（Nhe1-/-、Cd47-/- 和 Cd36-/-）、原代人类和小鼠主动脉 SMC、血管 SMC 限制性 Cre 驱动小鼠模型 (Itga8-CreERT2+/-; mTmG+/-) 和其他遗传工具来测试假设。将使用多种互补技术来研究 SMC 巨胞饮作用的体外（药理学、遗传、荧光/SEM 成像）和体内（TEM/IEM 显微镜、SMC 特异性 Nhe1 基因敲除小鼠）。结论是，拟议的研究将确定促进 SMC 泡沫的关键机制细胞形成和表型转换，并可能确定治疗干预的新靶点动脉粥样硬化。