Project 1: Megakaryocytes as Organizers of the Hematopoietic Environment

项目 1：巨核细胞作为造血环境的组织者

• 批准号: 10321580
• 负责人: Karin Maria Hoffmeister
• 金额: $ 62.02万
• 依托单位: VERSITI WISCONSIN, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10321580
• 关键词: Abnormal megakaryocyte; Affect; Alloimmunization; Blood Platelets; Bone Marrow; Bone Marrow Cells; Bone Marrow Transplantation; Cell surface; Cells; Clinical; Cluster Analysis; Data; Data Analyses; Defect; Development; Environment; Fluorouracil; GAG Gene; Galactosyltransferases; Gene Expression Profile; Genomics; Glycobiology; Glycosaminoglycan Degradation Pathway; Glycosaminoglycans; Goals; Hematopoietic; Hematopoietic stem cells; Hemorrhage; Heparan Sulfate Proteoglycan; Heparin Lyase; Heterogeneity; Human; Hyperactivity; Impairment; In Vitro; Injury; Integrins; Investigation; Joints; Knowledge; Libraries; Maintenance; Mediating; Megakaryocytes; Megakaryocytopoieses; Messenger RNA; Methods; Molecular; Molecular Target; Mus; Myelogenous; Patients; Phenotype; Platelet Transfusion; Polysaccharides; Population; Production; Proteins; Proteome; Reaction; Recovery; Risk; Role; Sialic Acids; Source; Stem cell transplant; Stress; Structure; Structure-Activity Relationship; Sulfate; Surface; Syndrome; Testing; Therapeutic; Thrombocytopenia; Thrombopoiesis; Transfusion; Virus Diseases; Work; base; cellular targeting; chemoradiation; chemotherapy; extracellular; genetic signature; glycosylation; heparanase; heparin proteoglycan; improved; in vivo; mRNA Expression; mimetics; mouse model; novel; prevent; programs; sialylation; single-cell RNA sequencing; transcriptome

Insufficient megakaryopoiesis and thrombopoiesis remain the main cause of persistent thrombocytopenia after hematopoietic stem and progenitor cell (HSPC) transplantation and after radiochemotherapy. Platelet transfusions are required to support patients with consequent increased risk of transfusion reactions, platelet alloimmunization, and transfusion-associated viral infections. Significant efforts are focused on identifying the most suitable cellular and molecular targets to enhance platelet production after bone marrow (BM) transplantation, chemotherapy, and to maximize the efficacy of in vitro human platelet production as an alternative method to increase the platelet supply for transfusion. Our ability to therapeutically optimize platelet recovery and in vitro platelet production is hindered by our paucity of knowledge of the molecular determinants that govern thrombopoiesis. Megakaryocytes (MKs) reside in the BM and maintain the continuous production of billions of circulating platelets in order to prevent bleeding. Recent work shows that MKs also maintain a functionally specific BM niche to support MK-biased HSPCs. Preliminary data suggest that: 1) MK-biased HSPCs express unusually high levels of α2,6-sialic acid (α2,6-Sia) which is unparalleled by cell intrinsic St6gal1 mRNA expression; 2) systemic ST6GAL1 deficiency promotes a myeloid skewed hematopoietic development profile, suggesting that α2,6-sialylation is a checkpoint of a particular HSPC population (Project 2); 3) B4galt1 deletion in MKs renders β1 integrin hyperactive and unexpectedly also regulates heparin sulfate proteoglycan (HS PG) and heparinase expression in MKs thereby severely impairing thrombopoiesis at steady state and following myeloablative injury; 4) Heparinase and glycosaminoglycan (GAG) mimetics improved thrombopoiesis in wild type and B4galt1-/- MKs, suggesting a role for GAGs in thrombopoiesis. The overarching hypothesis of this program is that “distinct cell-intrinsic and extrinsic glycan-mediated mechanisms regulate maintenance, differentiation, and function of hematopoietic cells”. Project 1 will test the specific hypothesis that glycosylation regulates MK-biased HSCs and thrombopoiesis in three aims: In Aim 1, a functionally defined MK-biased hematopoietic stem cell will be investigated together with Project 2, especially with respect to the heavily α2,6- sialylated cell surface despite the absence of St6gal1 expression necessary to generate this structure. We will establish the role of α2,6-Sia in MK-biased HSPCs. Based on the known requirement of the galactosyltransferase β4GalT1 in thrombopoiesis, Project 1 will also investigate the roles of β4GalT1, glycosaminoglycans (GAGs)/HS PG in thrombopoiesis at steady-state (Aim 2) and following myeloablative stress (Aim 3) using the novel combined shared “omics” together with Project 3 and standard approaches. A previously unknown role of β4GalT1 to regulate MK expression of HS PG will also be investigated. This Project will uncover new information to increase platelet production and help understand clinical conditions characterized by MK abnormalities.

巨毛毛菌和血小板不足仍然是持续性血小板细胞减少症的主要原因 造血干和祖细胞（HSPC）移植以及放射化学疗法后。血小板 需要输血来支持患者，从而增加输血反应的风险增加，血小板 同种免疫和输血相关的病毒感染。重大努力的重点是确定 最合适的细胞和分子靶标可增强骨髓后血小板的产生（BM） 移植，化学疗法，并最大程度地提高体外人血小板的效率 增加血小板供应进行翻译的替代方法。我们热优化血小板的能力 恢复和体外血小板的产生受到我们对分子确定剂知识的匮乏的阻碍 控制血小板。巨核细胞（MK）居住在BM中，并保持不断的生产 数十亿个流通的血小板以防止出血。最近的工作表明，MK还保持了 功能特定的BM利基市场支持MK偏置的HSPC。初步数据表明：1）MK偏向HSPCS 表达异常高的α2,6-硅酸（α2,6-sia），这是由细胞内在的ST6GAL1 mRNA无与伦比的 表达; 2）系统性ST6GAL1缺乏促进髓样偏斜的造血发育概况， 表明α2,6-元素是特定HSPC种群的检查点（项目2）； 3）b4galt1删除 在MKS中，β1整合素过度活跃和出乎意料地调节硫酸肝素蛋白聚糖（HS PG） 在MK中的肝素酶表达，因此在稳定状态下严重损害了血小板 骨髓损伤； 4）肝素酶和糖胺聚糖（GAG）MIMETICS改善了野外血小板 类型和B4GALT1 - / - MK，这表明GAG在血小板中的作用。总体假设 程序是“独特的细胞中性和外部聚糖介导的机制调节维护， 造血细胞的分化和功能”。项目1将检验糖基化的特定假设 在三个目的中调节MK偏向HSC和血小板的偏向：在AIM 1中，功能定义的MK偏置 造血干细胞将与项目2一起研究，尤其是关于严重的α2,6-- 尽管没有ST6GAL1表达来产生这种结构，但硫化细胞表面尽管没有ST6GAL1表达。我们将 确定α2,6-SIA在MK偏置的HSPC中的作用。根据已知要求 半乳糖基转移酶β4GALT1在血小板中，项目1还将研究β4GALT1的作用， 在稳态的血栓形成（AIM 2）和骨髓性的糖胺聚糖（GAGS）/HS PG（hs pg） 压力（AIM 3）使用新颖的共享“ OMICS”以及项目3和标准方法。一个 还将研究β4GALT1调节HS PG的MK表达的未知作用。这个项目 将发现新信息以增加血小板的产生并帮助了解临床状况的特征 由MK异常。