Characterization of A Novel 65kDa E-selectin Ligand on G-CSF Mobilized Leukocytes

G-CSF 动员白细胞上新型 65kDa E-选择素配体的表征

• 批准号: 7460691
• 负责人: ROBERT SACKSTEIN
• 金额: $ 21.44万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7460691
• 关键词: Acute; Acute Lung Injury; Adhesions; Adhesives; Affinity; Antigens; Arthritis; Binding; Biological Assay; Biology; Blood Vessels; Bone Marrow; Bone Marrow Cells; Carbohydrates; Cardiac; Cell Adhesion Molecules; Cell surface; Cells; Chimera organism; Clinical; Condition; Dose; E-Selectin; Electrophoresis; Epitopes; Exhibits; Flow Cytometry; Gel; Glycoproteins; Granulocyte Colony-Stimulating Factor; Harvest; Hematopoietic; Hematopoietic Stem Cell Mobilization; Hematopoietic Stem Cell Transplantation; Heterogeneity; Human; In Vitro; Inflammatory; Inflammatory Response; Ischemia; L-Selectin; Lead; Leukocytes; Ligands; Liquid substance; Mass Spectrum Analysis; Mediating; Membrane; Membrane Proteins; Methods; Modification; Molecular; Myeloid Cells; Newborn Respiratory Distress Syndrome; P-selectin ligand protein; Pathologic; Peripheral Blood Stem Cell; Phase; Physiological; Preparation; Proteins; Range; Relative (related person); Series; Serum; Source; Time; Vascular Endothelium; Western Blotting; base; clinically relevant; comparative; glycosylation; glycosyltransferase; in vivo; insight; interest; leukocyte mediator; novel; novel strategies; peripheral blood; prevent; progenitor; protein E; research study

DESCRIPTION (provided by applicant): Granulocyte-colony stimulating factor (G-CSF)-mobilized peripheral blood (MPB) has replaced traditional bone marrow harvest as the preferred source of cells for hematopoietic stem cell transplantation. Though generally considered to be a safer alternative to harvest, there are increasing observations that G-CSF administration causes vascular and inflammatory complications due to enhanced adhesion of leukocytes to vascular endothelium. A major mediator of leukocyte-endothelial adhesive interactions is E-selectin, an inducible endothelial molecule that binds sialofucosylated carbohydrate antigens recognized by the mAb HECA-452. We reasoned that vascular and inflammatory complications may result from G-CSF-induced expression of E-selectin ligands on circulating leukocytes. We have found that P-selectin glycoprotein ligand-1 (PSGL-1) and Hematopoietic Cell E-and L-selectin Ligand (HCELL) are E-selectin ligands on G- CSF MPB leukocytes. Importantly, our studies also reveal that a novel HECA-452-reactive ~65kDa high affinity E-selectin ligand (hereinafter called "~65kDa protein") is expressed on G-CSF MPB leukocytes but not on native leukocytes and not on normal human bone marrow cells. This ~65kDa protein exhibits potent Ca2+-dependent E-selectin ligand activity, as evidenced by (i) binding to fluid phase E-selectin and (ii) avid binding of E-selectin- transfected cells under physiological shear flow conditions. Treatment of normal human bone marrow cells with G-CSF at a pharmacokinetically-relevant concentration in vitro induces the expression of this ~65kDa protein directly on myeloid cells. We hypothesize that this ~65kDa E-selectin ligand contributes to vascular and inflammatory complications following G-CSF administration. The specific aims of this proposal are: (1) To identify the ~65kDa protein serving as an E-selectin ligand on G-CSF MPB leukocytes; and (2) To identify the subset(s) of human leukocyte-series cells that express this glycoprotein following G-CSF administration in vivo and in vitro. The identification and the characterization of expression of this novel E-selectin ligand should provide insights into the biology of E-selectin ligands and greatly increase our understanding of G-CSF MPB leukocyte-endothelial interactions. It is anticipated that the results obtained will lead to new approaches for preventing or alleviating G-CSF-induced vascular and inflammatory complications

描述（由申请人提供）：粒细胞集落刺激因子（G-CSF）动员外周血（MPB）已经取代传统的骨髓收获作为造血干细胞移植的首选细胞来源。虽然通常认为G-CSF是一种更安全的替代方法，但越来越多的观察表明，由于白细胞对血管内皮的粘附增强，G-CSF会导致血管和炎症并发症。白细胞-内皮粘附相互作用的主要介质是e-选择素，e-选择素是一种可诱导的内皮分子，可结合被单克隆抗体hea -452识别的sialo focusylated碳水化合物抗原。我们推断，血管和炎症并发症可能是由g - csf诱导的循环白细胞上e -选择素配体的表达引起的。我们发现p -选择素糖蛋白配体-1 （PSGL-1）和造血细胞e -和l -选择素配体（HCELL）是G- CSF MPB白细胞上的e -选择素配体。重要的是，我们的研究还揭示了一种新的heca -452反应性~65kDa高亲和力e -选择素配体（以下简称“~65kDa蛋白”）在G-CSF MPB白细胞上表达，而不是在天然白细胞和正常人骨髓细胞上表达。这种~65kDa的蛋白表现出强大的Ca2+依赖性e -选择素配体活性，这可以通过(i)与液相e -选择素的结合和（ii）在生理剪切流动条件下与e -选择素转染的细胞的强烈结合来证明。体外药代动力学相关浓度的G-CSF处理正常人骨髓细胞可诱导该~65kDa蛋白直接在髓细胞上表达。我们假设这种~65kDa的e -选择素配体有助于G-CSF给药后的血管和炎症并发症。本提案的具体目的是：(1)鉴定作为G-CSF MPB白细胞上e -选择素配体的~65kDa蛋白；(2)鉴定在体内和体外给药G-CSF后表达该糖蛋白的人白细胞系列细胞亚群。这种新型e-选择素配体的鉴定和表达特征应该为e-选择素配体的生物学提供见解，并极大地增加我们对G-CSF MPB白细胞-内皮相互作用的理解。预计获得的结果将导致预防或减轻g - csf诱导的血管和炎症并发症的新方法