Kindlin-3 Signaling in Blood Cells

血细胞中的 Kindlin-3 信号转导

• 批准号: 9193133
• 负责人: Yan-Qing Ma
• 金额: $ 41.75万
• 依托单位: VERSITI WISCONSIN, INC.
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-01 至 2020-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9193133
• 关键词: Affect; Animals; Applications Grants; Binding; Binding Proteins; Biochemical; Biological Models; Blood Cells; Blood Platelets; Blood coagulation; Bone Marrow; Cells; Coagulation Process; Complement; Complex; Cytoplasmic Tail; Data; Deep Vein Thrombosis; Defect; Deposition; Development; Etiology; Evaluation; Event; Functional disorder; Generations; Goals; Growth; HL60; Hemorrhage; Hereditary Disease; Immune system; Infection; Inferior vena cava structure; Infiltration; Inflammation; Inflammatory Response; Integrin beta Chains; Integrins; Kinetics; Knock-in Mouse; Lead; Leukocyte Adhesion Deficiency; Leukocytes; Ligature; Maps; Mediating; Mediator of activation protein; Modeling; Molecular; Morbidity - disease rate; Mus; Mutation; Neutrophil Activation; Neutrophil Infiltration; Patients; Play; Prevention strategy; Production; Pulmonary Embolism; Reactive Oxygen Species; Recruitment Activity; Recurrence; Reporter; Research Project Grants; Resolution; Risk Factors; Role; Signal Pathway; Signal Transduction; Site; Testing; Therapeutic; Thromboembolism; Thromboplastin; Thrombosis; Thrombus; Venous; Venous Thrombosis; base; deep vein; design; extracellular; in vivo; insight; intravital microscopy; mortality; mutant; neutrophil; new therapeutic target; novel; prevent; treatment strategy

Deep vein thrombosis (DVT) is one of the major causes of morbidity and mortality worldwide. DVT is associated with multiple risk factors and occurs when a blood clot forms in the deep veins. Significantly, recent studies have revealed that neutrophils are essentially involved in DVT formation, indicating that DVT is a complex, coordinated dysfunction of both the coagulation cascade and the innate immune system. Studies in animal DVT models have demonstrated that initial neutrophil recruitment and subsequent release of neutrophil extracellular traps (NETs) promote thrombus growth; in addition, later neutrophil infiltration facilitates thrombus resolution, suggesting that neutrophils may play dual functional roles in DVT. Kindlin-3 is a recently identified integrin β cytoplasmic tail binding protein in blood cells and required for supporting integrin-mediated leukocyte recruitment and infiltration. Patients with kindlin-3 deficiency suffer from Type-III leukocyte adhesion deficiency, a rare genetic disorder characterized by recurrent infections and severe bleeding due to integrin dysfunction in both leukocytes and platelets. Importantly, our preliminary observations have suggested that kindlin-3 also carries out a number of integrin-independent functions in neutrophils, including suppressing NET release. Therefore, we hypothesize that kindlin-3 plays multiple roles in regulating the kinetics of DVT. The overall goal of this competitive application is to examine both integrin-dependent and integrin-independent functions of kindlin-3 in neutrophils in DVT formation, and to mechanistically interpret the novel function of kindlin-3 that regulates NET release. Two Specific Aims are proposed: (1) Specific Aim 1 will determine the importance of kindlin-3/integrin interactions in neutrophils in DVT formation. We will examine the kinetics of DVT formation in a standard inferior vena cava (IVC) ligature model in mice that carry an integrin-interaction disrupting mutation in kindlin-3 or a kindlin-3 deletion specifically in neutrophils. The kinetics and extent of neutrophil recruitment, infiltration in the IVC, NET release, thrombus growth and resolution will be quantified in these mice using a combination of both intravital microscopy and histological analysis. We expect that kindlin-3 plays important roles in both initial neutrophil recruitment to inflamed IVC where venous thromboemboli develop and later neutrophil infiltration into the formed thrombi that promotes DVT resolution. (2) Specific Aim 2 will determine the molecular mechanism by which neutrophil kindlin-3 regulates NET release in DVT. The key subdomain(s) and residue(s) in kindlin-3 responsible for regulating NET release will be mapped in informative model systems. The signaling intermediates between kindlin-3 and NET release will be identified by established biochemical approaches. Further, the contribution of kindlin-3/NETs signaling pathway to DVT formation will be evaluated in mice. Taken together, these studies will redefine our understanding of the complex mechanisms by which neutrophil kindlin-3 regulates the kinetics of DVT, thus providing opportunities for designing novel and targeted therapeutic strategies for prevention and treatment of DVT.

深静脉血栓形成（DVT）是世界范围内发病率和死亡率的主要原因之一。DVT是 与多种风险因素相关，并在深静脉中形成血块时发生。值得注意的是，最近 研究表明，中性粒细胞主要参与DVT的形成，这表明DVT是一种恶性肿瘤。 凝血级联和先天免疫系统的复杂协调功能障碍。研究 动物DVT模型已经证明，初始中性粒细胞募集和随后的中性粒细胞释放 细胞外陷阱（NET）促进血栓生长;此外，中性粒细胞浸润促进血栓形成 解决，这表明中性粒细胞可能发挥双重功能的作用，DVT。Kindlin-3是最近发现的一种 血细胞中的整合素β胞质尾结合蛋白，其是支持整合素介导的白细胞所必需的 招募和渗透。kindlin-3缺乏的患者患有III型白细胞粘附缺陷， 一种罕见的遗传性疾病，特征是由于整合素功能障碍引起的复发性感染和严重出血， 包括白细胞和血小板。重要的是，我们的初步观察表明，kindlin-3也 在中性粒细胞中执行许多整合素非依赖性功能，包括抑制NET释放。 因此，我们假设Kindlin-3在调节DVT动力学中起着多种作用。总目标 这项竞争性应用的目的是检查整合素依赖性和整合素非依赖性的功能， kindlin-3在DVT形成中的中性粒细胞中的表达，并从机制上解释kindlin-3的新功能， 控制NET的释放。提出了两个具体目标：（1）具体目标1将确定以下方面的重要性： 在DVT形成中中性粒细胞中的Kindlin-3/整合素相互作用。我们将研究DVT形成的动力学， 携带整合素相互作用破坏突变的小鼠的标准下腔静脉（IVC）结扎模型 在kindlin-3或kindlin-3缺失中，特别是在中性粒细胞中。中性粒细胞募集的动力学和程度， 将在这些小鼠中使用免疫组织化学方法定量IVC中的浸润、NET释放、血栓生长和消退。 活体显微镜检查和组织学分析的组合。我们希望kindlin-3能发挥重要作用 在初始中性粒细胞募集到静脉血栓栓塞发展的炎性IVC和随后 中性粒细胞浸润到形成的血栓中，促进深静脉血栓的消退。(2)具体目标2将确定 中性粒细胞kindlin-3调节DVT中NET释放的分子机制。密钥子域 和kindlin-3中负责调节NET释放的残基将被映射到信息模型系统中。 Kindlin-3和NET释放之间的信号传导中间体将通过已建立的生物化学方法鉴定。 接近。此外，Kindlin-3/NETs信号通路对DVT形成的贡献将在下文中评估。 小鼠总之，这些研究将重新定义我们对复杂机制的理解， 中性粒细胞Kindlin-3调节DVT的动力学，从而为设计新的靶向药物提供了机会。 预防和治疗DVT的治疗策略。