Anti-Coagulant and Cytoprotective activity in CCM pathogenesis

CCM 发病机制中的抗凝血和细胞保护活性

• 批准号: 10621253
• 负责人: Mark HOWARD Ginsberg
• 金额: $ 31.38万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10621253
• 关键词: Acute; Adult; Anticoagulants; Blocking Antibodies; Blood Vessels; Blood coagulation; Brain; CCM1 gene; Chronic; Cytoprotection; Deposition; Development; Disease; Disease Progression; Endothelial Cells; Endothelium; F2R gene; Factor V Leiden mutation; Hemophilia A; Hemorrhage; Hemosiderin; Human; Hypoxia; Hypoxia Inducible Factor; Impairment; Intercellular Junctions; Intervention; Lesion; MSH2 gene; Modeling; Monoclonal Antibodies; Morbidity - disease rate; Mus; Pathogenesis; Pathway interactions; Perinatal; Preclinical Testing; Protein C; Research; Resistance; Role; Sepsis; Signal Transduction; Source; Stroke; Symptoms; Testing; Thrombomodulin; activated Protein C; activated protein C receptor; cerebral cavernous malformations; factor V Leiden; gene function; insight; loss of function; mortality; mutant; neonatal mice; neurovascular; new therapeutic target; pharmacologic; prevent; therapeutic target

ABSTRACT Cerebral Cavernous Malformations (CCM) are subject to acute and chronic bleeding that is a major source of morbidity and mortality in this disease. We have found that endothelial cells within murine and human CCM express markedly increased levels of thrombomodulin (TM) and endothelial protein C receptor (EPCR), which lead to activation of endogenous anti-coagulant protein C. We hypothesize that CCM form an anti-coagulant vascular domain and that activated Protein C (APC) contributes to bleeding in CCM. APC can also exert a cytoprotective effect on endothelium by signaling via PAR1 resulting in, among other effects, stabilization of endothelial cell-cell junctions. Indeed, this cytoprotective effect of APC has been exploited by creation of APC loss of function mutants that selectively maintain cytoprotective activity. We thus hypothesize that APC cytoprotective activity may limit morbidity from CCM as it does in experimental stroke. To genetically test this hypothesis, we will examine the bleeding in the acute murine CCM models in Factor V Leiden (F5R504Q/wt) mice, which are resistant to the anti-coagulant effect of APC. As a second approach we will test the effect of MAPC1591, a monoclonal antibody that blocks the anti-coagulant activity of APC in acute models of CCM. To test the effects of cytoprotective activity, we will examine acute CCM lesion development and bleeding in F2rR46Q/R46Q mice bearing PAR1 that is selectively resistant to APC cleavage. Conversely, we will examine the effect of 3K3A-APC, a loss of function APC mutant that is selectively impaired in anti-coagulant function in acute and chronic CCM models. We will exploit our observation that a brief period of hypoxia or pharmacological stabilization of Hypoxia-inducible factor 1A markedly exacerbates acute CCM formation in perinatal mice and enables a robust subacute model that manifests in adult mice. This subacute model which manifests both bleeding and hemosiderin deposition, will be used to test the interventions described in aims 1 and 2. Completion of these Aims will provide mechanistic insight into the role of the marked increase of TM and EPCR that is observed in CCM and provide important preclinical tests of the idea that hemorrhage or disease progression can be influenced by manipulating pathways that are already being therapeutically targeted in diseases such as stroke, sepsis, and hemophilia.

摘要 脑海绵状血管畸形（CCM）易发生急性和慢性出血， 是该病发病率和死亡率的主要来源。我们发现内皮细胞 在鼠和人CCM中表达显著增加的血栓调节蛋白（TM）水平， 内皮蛋白C受体（EPCR），其导致内源性抗凝剂的激活 蛋白C我们假设CCM形成了一个抗凝血管域， 蛋白C（APC）有助于CCM的出血。APC也可以对细胞产生保护作用， 通过经由PAR1的信号传导，导致内皮细胞的稳定，以及其他作用， 细胞间连接事实上，APC的这种细胞保护作用已经通过产生 选择性维持细胞保护活性的APC功能丧失突变体。我们因此 假设APC细胞保护活性可能会限制CCM的发病率，就像在 实验性中风为了从遗传学上验证这一假设，我们将检查急性出血性休克患者的出血情况。 在对抗凝剂耐药的因子V Leiden（F5R504Q/wt）小鼠中的鼠CCM模型 APC的影响。作为第二种方法，我们将测试MAPC 1591（一种单克隆抗体）的作用。 在CCM急性模型中阻断APC的抗凝活性。为了测试 细胞保护活性，我们将检查急性CCM病变的发展和出血， 携带PAR 1的F2rR46Q/R46Q小鼠对APC切割具有选择性抗性。相反，我们将 检测3K3A-APC的作用，3K3A-APC是一种功能丧失的APC突变体， 急性和慢性CCM模型中的抗凝功能。我们将利用我们的观察， 短暂缺氧或缺氧诱导因子1A的药理学稳定显著 加剧了围产期小鼠的急性CCM形成，并能够建立一个稳健的亚急性模型， 在成年小鼠中表现出来。这种亚急性模型表现出出血和含铁血黄素 将利用沉积物来检验目标1和目标2所述的干预措施。完成这些 目的将提供对TM和EPCR显著增加的作用的机制性见解， 在CCM中观察到，并提供了重要的临床前试验，证明出血或疾病 可以通过操纵已经在治疗中的途径来影响进展， 针对中风、败血症和血友病等疾病。