Murine models of vascular remodeling

血管重塑的小鼠模型

• 批准号: 8746649
• 负责人: Manfred Boehm
• 金额: $ 69.01万
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8746649
• 关键词: Acute; Arterial Injury; Attenuated; B-Lymphocytes; Binding; Binding Sites; Blood Vessels; Bone Marrow Transplantation; CD4 Positive T Lymphocytes; Cell Proliferation; Cells; Cellularity; Clinical; Complex; Data; Dependence; Development; Disease; Endothelial Cells; Environment; Event; Immune; Immune response; Infiltration; Inflammation; Inflammatory; Inflammatory Response; Injury; Interleukin-6; Knock-out; Knockout Mice; Laboratories; Lesion; Lung; Maps; Mechanics; Medial; Mediating; Mesenchymal; Modeling; Monocrotaline; Mus; Natural regeneration; Pathogenesis; Pathology; Phase; Phenotype; Physiologic intraventricular pressure; Play; Process; Production; Pulmonary Hypertension; Pulmonary artery structure; RANTES; Regulation; Right Ventricular Hypertrophy; Role; STAT3 gene; Signal Pathway; Signal Transduction; Small Interfering RNA; Smooth Muscle Myocytes; Stem cells; Stenosis; Structure of jugular vein; T cell response; T-Lymphocyte; TNF gene; Time; Up-Regulation; Vascular Endothelial Cell; Vascular remodeling; Veins; Wound Healing; arteriole; cell injury; femoral artery; human disease; in vivo; lung injury; macrophage; mouse model; neointima formation; neutralizing antibody; new therapeutic target; novel; p65; prevent; programs; promoter; pulmonary arterial hypertension; research study; response to injury; small hairpin RNA

Vascular wound repair is controlled by a complex interaction between local vascular cells and circulating immune and non-immune cells. Recently, the classic dogma of a predominantly local response to injury has been challenged by the identification of circulating vascular progenitor cells. The role of these vascular progenitor cells in vascular regeneration and their interaction with local vascular cells and infiltrating immune cells is poorly understood. My laboratory has undertaken a number of approaches to understand how these cells contribute to the overall vascular injury response and how they might ultimately be therapeutically manipulated. We studied events immediately following arterial injury and identified an acute STAT3- and NF-B (p65 subunit)-dependent upregulation of RANTES production by medial VSMCs, leading to early T cell and macrophage recruitment - processes also under the higher-order regulation of p21Cip1. Unique to VSMCs, RANTES production was initiated by TNF but not IL-6/gp130, and was dependent on binding of a p65-STAT3 complex to NF-B binding sites within the RANTES promoter, with shRNA knockdown of either STAT3 or p65 markedly attenuating RANTES production. In vivo, acute NF-B and STAT3 activation in medial VSMCs was identified, with acute RANTES production after injury significantly reduced in TNF-/- mice compared to controls. Finally, we generated mice with smooth muscle cell-specific conditional STAT3 knockout (STAT3fl/fl;SM22-Cre) and confirmed the STAT3-dependence of acute RANTES production by VSMCs. Together, these observations unify inflammatory events after vascular injury, demonstrating that VSMCs orchestrate the arterial inflammatory response program via acute RANTES production and subsequent inflammatory cell recruitment. Veins grafted into an arterial environment undergo vascular remodeling, a complex process of major clinical importance. Using lineage tracing experiments, we provide unprecedented evidence that cells of endothelial origin from the vein graft contribute to mesenchymal cellularity; during neointimal formation. In murine jugular veins grafted to femoral arteries, we found that endothelial cells lose their nascent markers and over time gain smooth muscle cell markers, indicative of endothelial to mesenchymal transition (EndoMT). This process is dependent on TGF- signaling, with early Smad activation. Antagonism of TGF- signaling by TGF- neutralizing antibody, siRNA-mediated Smad3 knockdown, or Smad3 haploinsufficiency resulted in decreased EndoMT and favorable vascular remodeling. Hence, we have identified EndoMT as a novel and pivotal mechanism underlying the stenosis-producing neointimal overgrowth of vein grafts, implying a potential a new therapeutic target to prevent vein graft pathology. Pulmonary hypertension is a vascular proliferative disease characterized by pulmonary artery remodeling due to dysregulated endothelial and smooth muscle cell proliferation. While the role inflammation plays in the development of the disease is not well defined, plexogenic lesions in human disease have been characterized by perivascular inflammation composed in part by T cells. We sought to explore the role of T cell infiltration on pulmonary vascular remodeling following endothelial cell damage. We induced endothelial cell damage using monocrotaline and isolated the role of T cells by utilizing Rag1tm1Mom mice and performing adoptive T cell transfer. We found that monocrotaline causes a pulmonary vascular endothelial cell injury which is followed by a perivascular inflammatory response. The infiltration of inflammatory cells is made up primarily of CD4+ T cells and leads to a progressive muscularization of small (<30m) arterioles. Pulmonary vascular proliferative changes were accompanied by progressive and persistent elevations in right ventricular pressure and right ventricular hypertrophy. Supporting the central role of CD4+ T cells in the inflammatory response, Rag1tm1Mom (Rag1-/-) mice which are devoid of T and B cells were protected from the development ofvascular injury when exposed to monocrotaline. Introduction of T cells from control mice into Rag1-/- mice reproduced the vascular injury phenotype. These data indicate that following endothelial cell damage CD4+ T cell infiltration participates in pulmonary vascular remodeling. This suggests that a CD4+ T cell immune response may contribute to the pathogenesis of inflammatory vascular lesions seen in some forms of pulmonary hypertension.

血管创伤修复受局部血管细胞与循环免疫和非免疫细胞之间的复杂相互作用控制。最近，对损伤的局部反应占主导地位的经典教条受到了循环血管祖细胞鉴定的挑战。这些血管祖细胞在血管再生中的作用及其与局部血管细胞和浸润免疫细胞的相互作用知之甚少。我的实验室已经采取了许多方法来了解这些细胞如何促进整体血管损伤反应，以及它们最终如何被治疗操纵。 我们研究了动脉损伤后立即发生的事件，并确定了中膜VSMC产生的RANTES的急性STAT 3和NF-B（p65亚基）依赖性上调，导致早期T细胞和巨噬细胞募集-也是在p21 Cip 1的高阶调节下进行的过程。VSMC特有的是，RANTES的产生是由TNF而不是IL-6/gp 130启动的，并且依赖于p65-STAT 3复合物与RANTES启动子内的NF-B结合位点的结合，而STAT 3或p65的shRNA敲低显著减弱RANTES的产生。在体内，中膜VSMC中的急性NF-B和STAT 3活化被鉴定，与对照组相比，TNF-/-小鼠损伤后的急性RANTES产生显著减少。最后，我们产生了平滑肌细胞特异性条件性STAT 3敲除小鼠（STAT 3fl/fl; SM 22-Cre），并证实了VSMC急性RANTES产生的STAT 3依赖性。总之，这些观察结果统一了血管损伤后的炎症事件，表明VSMC通过急性RANTES产生和随后的炎症细胞募集协调动脉炎症反应程序。 移植到动脉环境中的静脉经历血管重塑，这是一个具有重要临床意义的复杂过程。使用谱系追踪实验，我们提供了前所未有的证据表明，来自静脉移植物的内皮细胞有助于间充质细胞;在新生内膜形成过程中。在移植到股动脉的小鼠颈静脉中，我们发现内皮细胞失去了它们的新生标志物，并随着时间的推移获得平滑肌细胞标志物，这表明内皮细胞向间质细胞转化（EndoMT）。该过程依赖于TGF-β信号传导，具有早期Smad激活。TGF-中和抗体、siRNA介导的Smad 3敲低或Smad 3单倍不足对TGF-信号传导的拮抗作用导致EndoMT减少和有利的血管重塑。因此，我们已经将EndoMT确定为静脉移植物狭窄产生新生内膜过度生长的一种新的关键机制，这意味着潜在的预防静脉移植物病理学的新治疗靶点。 肺动脉高压是一种血管增生性疾病，其特征是由于内皮细胞和平滑肌细胞增殖失调导致的肺动脉重塑。虽然炎症在疾病发展中的作用尚未明确，但人类疾病中的丛源性病变的特征在于部分由T细胞组成的血管周围炎症。本研究旨在探讨T细胞浸润在内皮细胞损伤后肺血管重构中的作用。我们使用野百合碱诱导内皮细胞损伤，并通过利用Rag 1 tm 1 Mom小鼠和进行过继性T细胞转移来分离T细胞的作用。我们发现野百合碱导致肺血管内皮细胞损伤，随后是血管周围炎症反应。炎性细胞的浸润主要由CD 4 + T细胞组成，并导致小（<30 m）小动脉的进行性肌化。肺血管增生性改变伴随右心室压力和右心室肥大的进行性和持续性升高。支持CD 4 + T细胞在炎症反应中的中心作用，缺乏T和B细胞的Rag 1 tm 1 Mom（Rag 1-/-）小鼠在暴露于野百合碱时受到保护，免于血管损伤的发展。将来自对照小鼠的T细胞引入Rag 1-/-小鼠中再现了血管损伤表型。这些数据表明，内皮细胞损伤后，CD 4 + T细胞浸润参与肺血管重建。这表明，CD 4 + T细胞免疫应答可能有助于某些形式的肺动脉高压中所见的炎性血管病变的发病机制。