LACTOSAMINYL GLYCANS IN MEGAKARYOCYTES AND THROMBOPOIESIS

延迟血小板恢复和伴随的出血并发症是骨髓抑制/骨髓清除放化疗后发病率和死亡率的主要决定因素。造血关键依赖于时间表达的粘附分子的相互作用，这些粘附分子创造了相关的生长环境 在骨髓中。不同粘附分子的阶段依赖性展示和功能受到造血细胞因子和趋化因子的严格控制，其促进或抑制造血干细胞（HSC）的增殖、分化、成熟和迁移。这些作用部分是通过调节细胞表面乳糖胺聚糖介导的，特别是那些含有末端唾液酸和岩藻糖修饰的乳糖胺聚糖。这些结构在造血骨髓细胞内的表达发生在特定的阶段， 和特定血统的时尚。在迄今为止使用（β，4）-半乳糖基转移酶P4GalTl缺陷的小鼠的研究中，我们已经获得了乳糖胺聚糖是血小板生成的关键调节剂的直接证据。 在这个项目中，我们试图定义小鼠和人巨核细胞祖细胞（MKP）和巨核细胞（MK）上表达的不同乳糖胺聚糖，以及呈现这些聚糖对血小板生成至关重要的相关蛋白质支架和脂质。我们将定义表面的变化 本文研究了由血栓形成趋化因子和细胞因子或骨髓内皮细胞（BMEC）诱导的乳糖胺聚糖，并研究了这些乳糖胺结构如何介导与体外和体内造血关键的BMEC的粘附相互作用。为了获得可转化为实际治疗策略的知识，我们将把我们的研究扩展到人类细胞，以具体确定p4GalT1缺陷如何影响人类细胞。 血小板生成和乳糖胺聚糖表面表达。我们将确定相关的蛋白质支架和脂质，目前的乳糖胺聚糖的关键血小板生成在人类MK和分析的影响，趋化因子和细胞因子或BMEC乳糖胺聚糖的表达。我们将研究人MK表面乳糖胺聚糖的变化如何影响体外和体内BMEC的粘附功能。 这些研究将解决有关血小板生成的糖生物学的基本问题。我们还将研究血小板作为外源性糖基化介质的能力。生成的信息将用于制定策略，以调节关键末端乳糖胺聚糖的表达，从而在骨髓抑制/骨髓清除放化疗后以及在其他血小板减少症疾病（包括骨髓衰竭状态）中增强血小板生成和血小板生成。

Delayed platelet recovery, and attendant bleeding complications, is a major determinant of morbidity and mortality following myelosuppressive/myeloablatlve chemoradiotherapy. Hematopoiesis is critically dependent on the interplay of temporally expressed adhesion molecules that create relevant growth niches within the bone marrow. The stage-dependent display and function of distinct adhesion molecules is tightly controlled by hematopoietic cytokines and chemokines, which promote or inhibit hematopoietic stem cell (HSC) proliferation, differentiation, maturation and migration. These effects are mediated in part by modulation of cell surface lactosaminyl glycans, especially those containing terminal sialic acid and fucose modifications. Expression of these structures within hematopoietic marrow cells occurs in a stage-specific and lineage-specific fashion. In studies to date using mice deficient in the (pi,4)-galactosyltransferase, P4GalTl, we have obtained direct evidence that lactosaminyl glycans are key regulators of thrombopoiesis. In this project, we seek to define the distinct lactosaminyl glycans expressed on mouse and human megakaryocyte progenitors (MKPs) and megakaryocytes (MKs), and the pertinent protein scaffolds and lipids that present these glycans critical for thrombopoiesis. We will define the changes in surface lactosaminyl glycans induced by thrombopoletic chemokines and cytokines or bone marrow endothelial cells (BMECs), and examine how these lactosaminyl structures mediate adhesive interactions with BMECs critical to hematopoiesis in vitro and in vivo. To obtain knowledge translatable to practical therapeutic strategies, we will extend our studies to human cells to specifically determine how deficiency in p4GalT1 affects human thrombopoiesis and lactosaminoglycan surface expression. We will determine the pertinent protein scaffolds and lipids that present lactosaminoglycans critical for thrombopoiesis in human MKs and analyze the effects of chemokines and cytokines or BMECs on lactosaminoglycan expression. We will examine how changes in surface lactosaminoglycans on human MKs affect adhesive functions with BMECs in vitro and in vivo. These studies will address fundamental questions regarding the glycobiology of thrombopoiesis. We will also investigate the capacity of platelets to serve as mediators of extrinsic glycosylation. The generated information will serve to develop strategies to modulate expression of key terminal lactosaminyl glycans to enhance thrombopoiesis and platelet production following myelosuppressive/myeloablatlve chemoradiotherapy and in other conditions of thrombocytopenia, including bone marrow failure states.