The bone marrow extracellular matrix: scaffold of hematopoiesis

骨髓细胞外基质：造血支架

• 批准号: 10338115
• 负责人: Shinobu Matsuura
• 金额: $ 10.51万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-02-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10338115
• 关键词: 3-Dimensional; Ablation; Address; Adhesives; Affect; Allogenic; Animals; Antibodies; Automobile Driving; Award; Blocking Antibodies; Blood; Blood Cells; Blood Vessels; Bone Marrow; Bone Marrow Diseases; CRISPR/Cas technology; Cell Proliferation; Cell-Matrix Junction; Cells; Complementary therapies; Complex; Computer Analysis; Dependence; Development; Disease; Disease Outcome; Dysmyelopoietic Syndromes; Endothelial Cells; Environment; Enzymes; Exons; Extracellular Matrix; Failure; Fiber; Foundations; Gene Mutation; Genes; Genetic; Genomics; Goals; Hematology; Hematopoiesis; Hematopoietic; Human; Integrins; JAK2 gene; K-Series Research Career Programs; Life; Ligands; Lysine; Malignant - descriptor; Mass Spectrum Analysis; Mediating; Mentorship; Modeling; Morphology; Mus; Mutation; Mutation Analysis; Myelofibrosis; Outcome; Oxides; Pathogenesis; Pathology; Patients; Pericytes; Play; Post Translational Modification Analysis; Post-Translational Protein Processing; Primary Myelofibrosis; Procedures; Production; Prognosis; Proliferating; Protein-Lysine 6-Oxidase; Proteomics; Publishing; Reporting; Research; Research Personnel; Sampling; Signal Transduction; Spatial Distribution; Stem cell transplant; Stromal Cells; Survival Rate; Techniques; Testing; Therapeutic; Tissues; Training; Transgenes; Transgenic Mice; base; career; confocal imaging; curative treatments; defined contribution; effective therapy; experience; high risk; human disease; human model; improved; in vivo; inhibitor; leukemia; mortality; mouse model; mutant; novel; programs; promoter; scaffold; targeted treatment; therapy development

ABSTRACT This proposal addresses the urgent need to understand disease mechanisms in Primary Myelofibrosis (PMF). PMF is a disease with a dismal prognosis. The only potentially curative therapy, allogeneic stem cell transplantation, is a high-risk procedure, with a mortality rate of at least 50%. Myelofibrosis is characterized by excessive production of extracellular matrix (ECM), the fibers in bone marrow, which leads to a progressive failure in blood cell production. A number of gene mutations, including JAK2V617F, have been reported in PMF. While it is thought that hematopoietic cells carrying gene mutations abnormally proliferate and secrete factors that stimulate stromal cells to produce ECM excessively, inhibition of JAK2 only partially improves disease outcomes, and complementary therapies that specifically target ECM production are not currently available. Dependence on the prevalent hypothesis, with its focus on gene mutations, is a critical barrier to progress in the field. This project proposes instead to address the problem from the ECM angle. Building on our published and preliminary studies with mouse and human samples, the central hypothesis of this proposal is that abnormal activation of integrins (adhesive molecules that mediate cell attachment to ECM) facilitates proliferation of PMF cells. As PMF proliferation arises from interaction between ECM and integrins, the question is whether proliferation is triggered by an abnormality in ECM formation or by direct integrin activation. Thus, I will explore two potential mechanisms behind this abnormal integrin activation, asking whether: (1) abnormally high ECM production occurs in the vascular niche, where PMF cells develop, and (2) Lysyl oxidase (LOX), an ECM enzyme upregulated in PMF, activates integrins in PMF through post-translational modification. These studies will be performed on the most representative mouse model of human PMF, mice transgenic for the JAK2V617F mutation. Effects of integrins on cell proliferation in PMF will be determined by blocking integrin function with antibodies and by deleting integrin genes using CRISPR/Cas9. I will use 3D confocal imaging and advanced computational analysis to determine the spatial distribution of cells, ECM, and vasculature in myelofibrotic bone marrow, clarifying the mechanisms of ECM production by detailed morphological analysis of the vascular niche. Proteomic analysis of post-translational modifications will be performed on integrins to determine specific lysine residues oxidized by LOX, which has implications on the development of therapies targeting integrins or LOX. This is a Career Development Award, and the PI of this application will have the mentorship of investigators with expertise in the field for the execution of the project. The project builds on previous experience of the PI in malignant hematology. The training and the expertise acquired during this award will provide a foundation for a successful independent career for the PI of this project.

摘要 该提案解决了了解原发性骨髓纤维化（PMF）疾病机制的迫切需要。 PMF是一种预后不良的疾病。唯一可能治愈的疗法，异基因干细胞 移植是一种高风险手术，死亡率至少为50%。骨髓纤维化的特征是 细胞外基质（ECM）的过度产生，骨髓中的纤维，导致进行性 血细胞生产的失败。许多基因突变，包括JAK 2 V617 F，已经在 PMF。虽然认为携带基因突变的造血细胞异常增殖和分泌 刺激基质细胞过度产生ECM的因子，JAK 2的抑制仅部分改善 疾病的结果，以及专门针对ECM产生的补充疗法目前还没有 available.对流行假说的依赖，其重点是基因突变，是一个关键的障碍， 外地的进展。本项目建议从企业内容管理的角度解决这一问题。基础上 我们发表的和初步的研究与小鼠和人类样本，这一建议的中心假设， 整合素（介导细胞附着于ECM的粘附分子）的异常激活促进了 PMF细胞增殖。由于PMF增殖是由ECM和整合素之间的相互作用引起的， 问题是增殖是由ECM形成的异常还是由直接的整合素活化触发的。 因此，我将探讨这种异常整合素激活背后的两种潜在机制，询问是否：（1） 异常高的ECM产生发生在PMF细胞发育的血管龛中，以及（2）赖氨酰氧化酶 (LOX)在PMF中上调的ECM酶通过翻译后修饰激活PMF中的整合素。 这些研究将在最具代表性的人类PMF小鼠模型（转基因小鼠）上进行 JAK 2 V617 F突变。将通过阻断PMF中整合素对细胞增殖的影响来确定整合素对PMF中细胞增殖的影响。 通过使用抗体和使用CRISPR/Cas9缺失整联蛋白基因来实现整联蛋白功能。我将使用3D共聚焦 成像和先进的计算分析，以确定细胞的空间分布，ECM， 骨髓纤维化骨髓血管，阐明ECM生产的机制，详细 维管生态位的形态学分析。翻译后修饰的蛋白质组学分析将是 对整联蛋白进行，以确定由LOX氧化的特定赖氨酸残基，这对 开发靶向整联蛋白或LOX的疗法。这是一个职业发展奖，这个奖的PI 申请人将得到具有该领域专门知识的调查员的指导，以执行该项目。 该项目建立在PI先前在恶性血液学方面的经验基础上。培训和专业知识 在此期间获得的奖项将提供一个成功的独立职业生涯的PI的基础， 项目