A Macrophage Cation Channel in Prevention and Recovery from Inflammatory Injury

巨噬细胞阳离子通道在炎症损伤预防和恢复中的作用

• 批准号: 9259894
• 负责人: MICHAEL D CARRITHERS
• 金额: --
• 依托单位: JESSE BROWN VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-10-01 至 2018-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9259894
• 关键词: ATF2 gene; Address; Adoptive Cell Transfers; Adoptive Transfer; Alternative Splicing; Animal Model; Anti-Inflammatory Agents; Anti-inflammatory; Autoimmune Diseases; Autoimmune Process; Biochemical; Bone Marrow; Brain; Calcium; Calcium Oscillations; Cardiac; Caring; Cations; Cell Therapy; Cells; Chronic; Chronic Disease; Clinical; Couples; Cyclic AMP; Cytology; Data; Disease; Endosomes; Experimental Autoimmune Encephalomyelitis; Frequencies; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Genomics; Goals; Home environment; Human; Image; Immune; Immunophenotyping; In Vitro; Inflammation; Inflammatory; Injury; Knock-in; Laboratories; Lesion; Lymphocyte Function; Mediating; Modeling; Molecular; Multiple Sclerosis; Multiple Sclerosis Lesions; Mus; Neurologic; Nodule; Pathway interactions; Phagocytes; Phagocytosis; Pharmaceutical Preparations; Pharmacology; Phenotype; Prevention; Property; Proteins; Proteomics; RNA Splicing; Recovery; Regulation; Research; Resolution; Role; Services; Signal Pathway; Signal Transduction; Site; Sodium Channel; Spinal Cord; Spinal Cord Lesions; Symptoms; T-Lymphocyte; Techniques; Testing; Therapeutic; Therapeutic Effect; Tissues; Transgenes; Transgenic Mice; Transgenic Organisms; Treatment Protocols; Variant; Veterans; Wild Type Mouse; Work; base; cell type; central nervous system injury; cost; disability; experimental study; in vivo; innovation; intercellular communication; macrophage; mouse model; multiple sclerosis patient; multiple sclerosis treatment; novel; novel strategies; organelle movement; polarized cell; potassium ion; prevent; protein expression; public health relevance; release of sequestered calcium ion into cytoplasm; repaired; selective expression; sodium ion; targeted treatment; transcription factor; voltage; young adult

DESCRIPTION (provided by applicant): Existing therapies for autoimmune and inflammatory diseases have minimal effects on resolution and recovery from chronic injury. There is also a fundamental gap in understanding how macrophages regulate cellular phenotype to mediate either injury or repair. The long-term goal is to develop treatments that prevent and reverse chronic inflammatory injury. Using a combination of cellular, genomic, proteomic and in vivo approaches, a novel splice variant of the human sodium channel gene, SCN5A, has been identified as a central regulator of anti-inflammatory macrophage signaling. This splice variant encodes a novel, endosomal cation channel that couples intracellular calcium flux to downstream signaling and gene transcription. Because this splice variant is not expressed in mice, a transgenic knock-in model, the C57BL6cfms-hSCN5A mouse, was developed to study its in vivo function in macrophages. Transfer of bone marrow derived macrophages from these mice mediates clinical recovery in the experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). The objective of this study is to characterize the SCN5A signaling network in macrophages and determine how it regulates clinical recovery in the EAE model of chronic inflammatory disease. The central hypothesis is that persistent expression of SCN5A in macrophages initiates and maintains a cellular phenotype that enhances recovery from inflammatory injury. The rationale for this study is that understanding this mechanism will lead to innovative approaches to prevent and treat chronic inflammatory disease. The hypothesis will be tested in two specific aims: 1. Determine how the SCN5A variant cation channel regulates macrophage phenotype. 2. Analyze how SCN5A+ macrophages mediate recovery during EAE. Based on preliminary data, the working model is that endosomal channel activity mediates short-term biochemical signaling and long-term gene expression patterns that determine and maintain cellular phenotype. In Aim 1, channel-dependent regulation of cytosolic calcium, cyclic AMP, and activating transcription factor 2 (ATF2) pathways will be examined. The experimental approaches will include a combination of existing biochemical, imaging and molecular techniques in mouse and human primary macrophages. In Aim 2, the EAE model will be utilized to assess this pathway in vivo and determine how SCN5A+ macrophages mediate recovery from inflammatory injury. In vivo gene and protein expression will be analyzed and correlated with in vitro assessment of SCN5A regulation of macrophage vesicular secretion and intercellular communication. The proposed work is innovative because it represents a new approach to regulate macrophage function and treat chronic inflammatory disease. It is significant because promotion of recovery in MS and other chronic inflammatory diseases represents an unmet therapeutic need.

描述(由申请人提供)： 现有的治疗自身免疫性和炎症性疾病的方法对慢性损伤的缓解和恢复影响很小。在理解巨噬细胞如何调节细胞表型以调节损伤或修复方面也存在着根本的差距。长期目标是开发预防和逆转慢性炎症性损伤的治疗方法。结合细胞、基因组、蛋白质组和体内研究方法，人类钠通道基因的一种新的剪接变异体SCN5A已被确定为抗炎巨噬细胞信号转导的中心调节因子。这种剪接变异体编码一种新的内体阳离子通道，它将细胞内的钙流量耦合到下游信号和基因转录。由于这种剪接变体在小鼠中不表达，因此建立了转基因敲入模型C57BL6cfms-hSCN5A小鼠，以研究其在巨噬细胞中的体内功能。在多发性硬化症(MS)的实验性自身免疫性脑脊髓炎(EAE)模型中，从这些小鼠身上移植骨髓来源的巨噬细胞可以促进临床恢复。本研究的目的是确定巨噬细胞中SCN5A信号网络的特征，并确定它如何调节慢性炎症性疾病EAE模型的临床恢复。中心假设是SCN5A在巨噬细胞中的持续表达启动并维持了促进炎性损伤恢复的细胞表型。这项研究的基本原理是，了解这一机制将导致预防和治疗慢性炎症性疾病的创新方法。这一假设将在两个特定的目标下得到验证：1.确定SCN5A变异阳离子通道如何调节巨噬细胞的表型。2.分析SCN5A+巨噬细胞在EAE中的恢复作用。根据初步数据，工作模型是内体通道活动调节决定和维持细胞表型的短期生化信号和长期基因表达模式。在目标1中，将研究胞浆钙、环磷酸腺苷和激活转录因子2(ATF2)通路的通道依赖性调节。实验方法将包括现有的生化、成像和分子技术的组合，用于小鼠和人类的原代巨噬细胞。在目标2中，将利用EAE模型在体内评估这一途径，并确定SCN5A+巨噬细胞如何介导炎性损伤的恢复。在体内，基因和蛋白的表达将被分析，并与体外评估SCN5A对巨噬细胞囊泡分泌和细胞间通讯的调节有关。这项拟议的工作具有创新性，因为它代表了一种调节巨噬细胞功能和治疗慢性炎症性疾病的新方法。这一点意义重大，因为促进多发性硬化症和其他慢性炎症性疾病的康复是一种尚未得到满足的治疗需求。