The role of HMGB-1 and innate immunity in promoting angiogenesis after ischemia

描述（由申请人提供）：此提案描述了Ulka Sachdev博士的职业目标和研究计划，她是匹兹堡大学血管外科外科助理教授。Sachdev博士在纽约西奈山医学院（Mount Sinai School of Medicine）完成了普通外科和血管外科培训，并在她的培训和早期教师生涯中积极参与基础科学研究。她的目标是成为一名独立资助的外科科学家，并最终进行转化研究以帮助患者。这一建议源于对缺血诱导血管生成的四年研究，以及HMGB-1在这一现象中的作用。外周动脉疾病是导致功能障碍和发病率的重要原因。随着腿部动脉闭塞性疾病的严重程度增加，高达40%的不可重建性疾病患者可发生严重肢体缺血，导致截肢。在肢体缺血情况下，促进血管生成的机制有待进一步研究，以优化不能进行手术血运重建的患者的医学治疗。炎症是血管生成的一个组成部分，最近的证据表明核蛋白HMGB-1在介导炎症的全身效应中起作用，也可能是促血管生成的。HMGB-1激活先天免疫的受体，包括toll样受体（TLR） 2和4，以及晚期糖基化终产物受体（RAGE）。本研究旨在探讨HMGB-1和TLRs在缺血时促进血管生成的作用。初步，Sachdev博士已经证明HMGB-1可以促进内皮细胞（EC）管的形成，这是一种血管生成表型，当与缺氧刺激结合时，可以增加EC的增殖，并可能通过自噬介导。她还表明，HMGB-1抗体阻断了内皮管的形成，并导致股动脉结扎的小鼠更大的肌肉坏死，这是一种公认的血管生成模型。抑制自噬也会破坏内皮管的形成。缺乏TLR4的小鼠也表现出比对照组更大的肌肉坏死区域，TLR2KO小鼠也是如此。最后，她发现缺血心肌细胞与正常缺血心肌细胞相比，细胞核中HMGB-1的丢失，这表明组织缺血可能导致HMGB-1释放到周围环境中，使其可介导血管生成事件。本研究有三个主要目的，旨在了解HMGB-1对体外血管细胞和体内缺血诱导血管生成的影响。第一个目的是确定EC血管生成行为中自噬与HMGB1释放之间的时间关系。第二个目的是确定HMGB-1和自噬在体内骨骼肌血管生成中的功能。第三个目的是确定tlr在介导HMGB1血管生成作用中的作用。

DESCRIPTION (provided by applicant): This proposal describes the career goals and research plan for Dr. Ulka Sachdev, who is an Assistant Professor of Surgery in the Division of Vascular Surgery at the University of Pittsburgh. Dr. Sachdev completed her General Surgery and Vascular Surgery training at Mount Sinai School of Medicine in New York, and has been active in basic science research throughout her training and early faculty career. Her goal is to become an independently funded surgeon-scientist, and eventually perform translational research to assist patients. This proposal culminates from four years of research on ischemia-induced angiogenesis, and the role of HMGB-1 within that phenomenon. Peripheral artery disease is a cause of significant functional disability and morbidity. With increasing severity of arterial occlusive disease in the leg, critical limb ischemia can occur resulting in amputation in up to 40% of patients with non-reconstructable disease. Further study is required to evaluate the mechanisms that promote angiogenesis in the setting of limb ischemia, in order to optimize medical treatments for patients who can not undergo surgical revascularization. Inflammation is a component of angiogenesis, and recent evidence suggests that the nuclear protein HMGB-1 which is known to play a role in mediating systemic effects of inflammation, may also be pro-angiogenic. HMGB-1 activates receptors of innate immunity, including Toll-like Receptors (TLR) 2 and 4, as well as the Receptor for Advanced Glycation End-products (RAGE). The purpose of this proposal is to evaluate the role of HMGB-1 and TLRs in promoting angiogenesis in response to ischemia. Preliminarily, Dr. Sachdev has shown that HMGB-1 enhances endothelial cell (EC) tube formation, which is an angiogenic phenotype, increases EC proliferation when coupled with hypoxic stimulus, and may be mediated by autophagy. She has also shown that antibody to HMGB-1 blocks endothelial tube formation, and results in greater muscle necrosis when given to mice that have undergone femoral artery ligation, which is a recognized model of angiogenesis. Inhibition of autophagy also disrupts endothelial tube formation. Mice deficient in TLR4 also demonstrate greater areas of muscle necrosis than controls, as do TLR2KO mice. Finally, she has shown that ischemic myocytes demonstrate loss of HMGB-1 from nuclei compared to normoxic myocytes, suggesting that tissue ischemia may lead to release of HMGB-1 into the surrounding milieu, rendering it available to mediate angiogenic events. There are three main aims of this proposal, directed at understanding the effect of HMGB-1 on both vascular cells in vitro, and ischemia-induced angiogenesis in vivo. The first aim is to determine the temporal relationship between autophagy and HMGB1 release during EC angiogenic behavior. The second aim is to determine the function of HMGB-1 and autophagy in skeletal muscle angiogenesis in vivo. The third aim is to identify a role for the TLRs in mediating the angiogenic actions of HMGB1. PUBLIC HEALTH RELEVANCE: The following proposal aims to elucidate if the danger signal HMGB-1 which is released by necrotic or stressed cells, as well as its receptors, helps promote the formation of new blood vessels through the process of angiogenesis in the setting of tissue ischemia. This project will help understand the mechanisms that govern neovascularization, and may help patients with peripheral vascular disease who can not undergo revascularization by bypass or endovascular therapies.