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）是主动脉进行性减弱和扩张。 对导致动脉瘤破裂的分子机制的理解存在知识差距 根据我们之前的报告，血小板反应蛋白-1 (TSP1) 升高。 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) 分析并 确定巨噬细胞 (Mɸs) 是小鼠主动脉瘤 TSP1 升高的主要来源。 随后通过将 Lyz2-Cre 与我们新的杂交产生了 Mɸ 特异性 Thbs1 敲除小鼠 (Thbs1ΔMɸ) 构建了 Thbs1flox/flox 小鼠，当接受血管紧张素 II (Ang II) 联合诱导动脉瘤时。 高胆固醇血症，超过 60% 的 Thbs1ΔMɸ 因 AAA 破裂而死亡，发病率高出 2.6 倍 有趣的是，存活至 28 天 Ang II 输注结束的 Thbs1ΔMɸ 小鼠显示出较少的动脉瘤。 当 Thbs1ΔMɸ 小鼠受到挑战时，也发现了比 Thbs1wt 更小的动脉瘤扩张。 CaCl2 的血管周围应用，一种不会产生破裂的 AAA 模型，我们提出了两个具体目标。 描述 Mɸ 特异性 Thbs1 基因缺失对主动脉产生不同影响的机制 扩张和破裂，重点是 AAA 破裂。具体目标 1 致力于确定破裂。 Mɸ TSP1 的预防功能具体来说，我们将确定致命性破裂时的主动脉反应。 Ang II 模型中的雄性和雌性 Thbs1ΔMɸ 小鼠，然后识别破裂相关分子 此外，我们将通过 scRNA-seq、原位杂交和免疫染色来检查特征。 使用不同的小鼠模型进行 Mɸ 特异性 Thbs1 敲除的效果，该模型在晚期阶段产生破裂 动脉瘤，这比 Ang II 模型产生的早期破裂与人类 AAA 更相关。 目标 2 重点研究动脉瘤破裂的分子机制。初步研究表明。 与野生型相比，Thbs1-/- Mɸs 迁移或吞噬凋亡细胞的能力也显着降低 作为中性粒细胞胞外陷阱 (NET)，我们将测试 Mɸ TSP1 是否通过 CD47- 促进 NET 清除。 接下来，我们将确定 Mɸ 迁移受损的因果效应和 Thbs1ΔMɸ小鼠动脉瘤破裂的吞噬作用我们首先确定NET负荷是否增加。 在 Thbs1ΔMɸ 中，NETs 和 Mɸs 之间的空间关系是否因破裂而死亡。 恢复 Thbs1ΔMɸ 中的 Mɸ 迁移可通过采用转移策略减少 NET 积累。 将检查增强或减弱 NET 清除是否会影响 Thbs1ΔMɸ 的动脉瘤破裂。 将分析破裂和非破裂中 TSP1 表达及其与 Mɸ 和 NET 积累的关联 该项目通过 TSP1 操作剖析 Mɸs 的多方面功能。 将对动脉瘤破裂的认识产生重大影响。