Novel Role of Thrombospondin-1 in Protection against Rupture of Abdominal Aortic Aneurysm

Thrombospondin-1 在预防腹主动脉瘤破裂中的新作用

• 批准号: 10383732
• 负责人: Bo Liu
• 金额: $ 45.99万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10383732
• 关键词: Abdominal Aortic Aneurysm; Actins; Address; Adoptive Transfer; Affect; Aneurysm; Angiotensin II; Animals; Aorta; Aortic Aneurysm; Aortic Rupture; Apoptotic; Area; Atherosclerosis; Attenuated; Blood Vessels; Bone Marrow; CD47 gene; Cells; Clinical; Complication; Conflict (Psychology); Coupled; Data; Deoxyribonuclease I; Detection; Development; Dilatation - action; Elastases; Female; Gene Deletion; Growth; Human; Imipramine; Immunohistochemistry; Impairment; In Situ Hybridization; Incidence; Infiltration; Inflammation; Infusion procedures; Knock-out; Knockout Mice; Knowledge; Life Style; Low-Density Lipoproteins; Mediating; Modeling; Molecular; Molecular Profiling; Monitor; Mouse Strains; Mus; Operative Surgical Procedures; Patients; Phagocytosis; Phenotype; Play; Preventive; Proteins; Publications; Reporting; Research; Role; Rupture; Ruptured Abdominal Aortic Aneurysm; Ruptured Aneurysm; Small Interfering RNA; Source; System; THBS1 gene; Telemetry; Testing; Thrombospondin 1; Time; Tissues; United States; Vascular Diseases; atherosclerotic plaque rupture; beta Aminopropionitrile; conditional knockout; extracellular; human disease; hypercholesterolemia; induced pluripotent stem cell; inhibitor; knock-down; macrophage; male; migration; mortality; mouse model; neutrophil; novel; polymerization; receptor; response; single-cell RNA sequencing; spatial relationship; therapeutic development; ultrasound; uptake

Abdominal aortic aneurysm (AAA) is the progressive weakening and dilation of the aorta. A substantial knowledge gap exists in the understanding of molecular mechanisms responsible for aneurysm rupture, the major cause of mortality among AAA patients. Following our prior report of elevated thrombospondin-1 (TSP1) in human and mouse aneurysmal tissues, we conducted single-cell RNA sequencing (scRNA-seq) analysis and identified macrophages (Mɸs) being the primary source of elevated TSP1 in mouse aneurysmal aorta. We subsequently generated Mɸ-specific Thbs1 knockout mice (Thbs1∆Mɸ) by crossing Lyz2-Cre with our newly constructed Thbs1flox/flox mice. When subjected to aneurysm induction by angiotensin II (Ang II) coupled with hypercholesterolemia, over 60% of Thbs1∆Mɸ died due to AAA rupture, an incidence that was 2.6 times higher than Thbs1wt. Intriguingly, Thbs1∆Mɸ mice that survived to the end of 28-day Ang II infusion showed less aneurysm dilation than Thbs1wt. Smaller aneurysmal expansion was also found when Thbs1∆Mɸ mice were challenged with perivascular application of CaCl2, an AAA model that does not produce rupture. We propose two specific aims to delineate the mechanisms through which Mɸ-specific Thbs1 gene deletion differentially affects aortic dilation and rupture with an emphasis on AAA rupture. Specific Aim 1 devotes to establishing the rupture- preventive function of Mɸ TSP1. Specifically, we will determine the aortic responses proceeding lethal rupture in male and female Thbs1∆Mɸ mice in the Ang II model followed by identifying rupture-associated molecular signatures through scRNA-seq, in situ hybridization and immunostaining. Furthermore, we will examine the effects of Mɸ-specific Thbs1 knockout using a different murine model that produces rupture in advanced stages of aneurysm, which is more relevant to human AAA than the early rupture produced by the Ang II model. Specific Aim 2 focuses on investigating molecular mechanisms of aneurysm rupture. Preliminary studies showed that compared to wildtype, Thbs1-/- Mɸs had significantly reduced ability to migrate or to engulf apoptotic cells as well as neutrophil extracellular traps (NETs). We will test whether Mɸ TSP1 promotes NET clearance through CD47- mediated actin polymerization. Next, we will establish the causal effect of impaired Mɸ migration and phagocytosis on aneurysm rupture of Thbs1∆Mɸ mice. We will first determine whether NET burden is increased in Thbs1∆Mɸ died from rupture, and the spatial relationship between NETs and Mɸs. Second, we will test whether restoring Mɸ migration in Thbs1∆Mɸ reduces NET accumulation via adoptive transfer strategies. Furthermore, we will examine whether enhancing or attenuating NET clearance affect aneurysm rupture in Thbs1∆Mɸ. Lastly, we will analyze TSP1 expression and its association with Mɸ and NET accumulation in ruptured and non-ruptured human AAA tissues. By dissecting the multifaceted functions of Mɸs through TSP1 manipulations, this project will produce significant impact on the understanding of aneurysm rupture.

腹主动脉瘤（AAA）是主动脉的逐渐减弱和扩张。实质性 知识差距存在于对负责动脉瘤破裂的分子机制的理解中， AAA患者死亡的主要原因。遵循我们先前关于升高血小板素-1（TSP1）的报告 在人类和小鼠动脉瘤组织中，我们进行了单细胞RNA测序（SCRNA-SEQ）分析和 鉴定出巨噬细胞（MɸS）是小鼠动脉瘤主动脉中TSP1升高的主要来源。我们 随后通过将LYZ2-CRE与我们的新近跨越LYZ2-CR 构建的Thbs1flox/Flox小鼠。当血管紧张素II（ANG II）诱导动脉瘤时， 高胆固醇血症，超过60％的THBS1ΔMɸ因AAA破裂而死亡，这一事件高2.6倍 比thbs1wt。有趣的是，生存到28天ANG II输注结束的THBS1ΔMɸ小鼠显示出较少的动脉瘤 扩张比THBS1WT。当THBS1ΔMɸ小鼠挑战时，还发现了较小的动脉瘤膨胀 CACL2的血管周期应用，这是一种不会产生破裂的AAA模型。我们提出了两个具体目标 描绘Mɸ特异性THBS1基因缺失差异影响主动脉的机制 扩张和破裂，重点是AAA破裂。特定的目标1致力于建立破裂 - mɸtsp1的预防功能。特别是，我们将确定进行致命破裂的主动脉反应 ANG II模型中的雄性和雌性THBS1ΔMɸ小鼠，然后鉴定与破裂相关的分子 通过SCRNA-SEQ的签名，原位杂交和免疫染色。此外，我们将研究 使用不同的鼠模型在高级阶段产生破裂的Mɸ特异性THBS1敲除效果 动脉瘤，与人类AAA相比，它比Ang II模型产生的早期破裂更相关。具体的 AIM 2专注于研究动脉瘤破裂的分子机制。初步研究表明 与WildType相比，THBS1 - /-MɸS的迁移能力显着降低或吞噬凋亡细胞的能力也显着降低 作为中性粒细胞外陷阱（网）。我们将测试MɸTSP1是否通过CD47-促进净清除率 介导的肌动蛋白聚合。接下来，我们将确定Mɸ迁移受损的因果关系 吞噬作用在THBS1ΔMɸ小鼠的动脉瘤破裂上。我们将首先确定净燃烧是否增加 在THBS1ΔMɸ中死于破裂以及网与Mɸ之间的空间关系。第二，我们将测试是否 通过自适应转移策略来恢复THBS1ΔMɸ中的Mɸ迁移可降低净积累。此外，我们 将检查增强或衰减净清除率是否影响THBS1ΔMɸ中的动脉瘤破裂。最后，我们 将分析TSP1的表达及其与Mɸ和净积累的关联 人AAA组织。通过通过TSP1操作解剖MɸS的多面功能，该项目 将对对动脉瘤破裂的理解产生重大影响。