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.

抽象的 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 是一种预后不佳的疾病。唯一有可能治愈的疗法，同种异体干细胞 移植是一种高风险手术，死亡率至少50%。骨髓纤维化的特点是 细胞外基质（ECM）（骨髓中的纤维）产生过多，导致进行性 血细胞生成失败。包括 JAK2V617F 在内的许多基因突变已在 PMF。虽然人们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 的因素，抑制 JAK2 只能部分改善 目前还没有专门针对 ECM 产生的疾病结果和补充疗法 可用的。对流行假设的依赖，其重点是基因突变，是一个关键障碍 该领域的进展。该项目建议从 ECM 角度解决该问题。建立在 我们已发表的针对小鼠和人类样本的初步研究，是该提案的中心假设 整合素（介导细胞附着至 ECM 的粘附分子）的异常激活促进 PMF 细胞的增殖。由于 PMF 增殖是由 ECM 和整合素之间的相互作用引起的， 问题是增殖是否是由 ECM 形成异常引发的，还是由直接整合素激活引发的。 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 产量发生在 PMF 细胞发育的血管生态位中，以及 (2) 赖氨酰氧化酶 (LOX) 是一种在 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 小鼠模型上进行，即转基因小鼠 JAK2V617F 突变。整合素对 PMF 中细胞增殖的影响将通过封闭来确定 整合素与抗体发挥作用，并使用 CRISPR/Cas9 删除整合素基因。我将使用 3D 共焦 成像和高级计算分析，以确定细胞、ECM 和细胞的空间分布 骨髓纤维化骨髓中的脉管系统，通过详细阐明 ECM 产生机制 血管生态位的形态学分析。翻译后修饰的蛋白质组学分析将 对整合素进行以确定被 LOX 氧化的特定赖氨酸残基，这对 开发针对整合素或 LOX 的疗法。这是一个职业发展奖，其 PI 应用程序将得到具有该领域专业知识的研究人员的指导来执行项目。 该项目建立在 PI 先前在恶性血液学方面的经验的基础上。培训和专业知识 在此奖项期间获得的奖项将为该 PI 的成功独立职业生涯奠定基础 项目。