The role of HLA and its coreceptors in endothelial cell activation and leukocyte recruitment in antibody-mediated transplant rejection

摘要 供体特异性HLAI和II类抗体的产生与慢性粒细胞白血病的发生有关 抗体介导的排斥反应(CAMR)和移植血管病变(TV)。CAMR的两个主要特征是 内皮细胞(EC)功能障碍和免疫细胞，特别是巨噬细胞的渗透。建议数 研究将探索新的范式，即人类白细胞抗原I类抗体与ITGB4和TLR4以及 在同种异体血管内皮细胞诱导中，人类白细胞抗原II和ITGB1协调不同的细胞内信号转导程序 促进单核细胞向移植物募集和巨噬细胞极化的功能。我们建议渗透到 单核细胞会遇到局部EC衍生因子和Ig G-FcγR信号，从而为其分化做好准备。在 移植后，促血管生成/促纤维化的巨噬细胞驱动新生内膜增厚。拮抗EC激活 和/或单核细胞募集可能是阻止CAMR和保护移植物免受感染的可行治疗靶点 电视。我们将采用原代人类EC培养模型，TV和人类的同种异体小鼠心脏移植模型。 心脏移植组织单核细胞HLAI、II信号通路及机制的电视解剖 招募和两极分化。目标1：确定复合体形成和功能的结构要求 人类白细胞抗原I类及其辅助受体ITGB4和TLR4在介导内皮细胞激活中的串扰 心脏TV中单核细胞募集和巨噬细胞极化。我们将：(1A)确定人类白细胞抗原I抗体如何- 活化的EC平衡渗入单核细胞，极化为具有不同表型和功能的巨噬细胞 体外；(1b)单核细胞募集、TV病变形成和浸润性巨噬细胞表型的特征 在MHC-I抗体介导的同种异体移植物损伤和发育过程中；(1c)确定结构要求 ITGB4、TLR4和HLAI之间的分子串扰对单核细胞募集和功能的影响 单抗激活的EC在体外对巨噬细胞的极化；以及(1D)表征对MHC I共刺激的需求 TLR4和ITGB4受体在单核细胞募集、巨噬细胞极化和TV病变形成中的作用 小鼠心脏移植。目标2.定义复合体形成和功能的结构要求 人类白细胞抗原II类及其辅受体ITGB1在介导单核细胞活化中的串扰 心脏TV中的募集和巨噬细胞极化。我们将：(2)确定人类白细胞抗原II抗体如何激活EC 平衡渗透单核细胞，在体外极化为表型和功能不同的巨噬细胞；(2b) 单核细胞募集、血管病变形成和浸润性巨噬细胞表型的特征 MHC II CAMR和TV的小鼠心脏移植模型；(2c)确定结构要求和 ITGB1和HLAII分子串扰对单核细胞募集和巨噬细胞功能的影响 抗体激活的血管内皮细胞的极化；(2d)表征MHC II辅助受体ITGB1在 小鼠心脏移植中单核细胞募集、巨噬细胞极化和TV损伤形成；以及 (2E)用TV确定人心脏移植移植物内巨噬细胞的表型。

ABSTRACT The production of donor specific HLA class I and class II antibodies is associated with development of chronic antibody-mediated rejection (cAMR) and transplant vasculopathy (TV). Two key features of cAMR are endothelial cell (EC) dysfunction and infiltration of immune cells, particularly macrophages. The proposed research will explore the novel paradigm that antibody crosslinking of HLA class I with ITGB4 and TLR4 and HLA class II with ITGB1 orchestrate divergent intracellular signaling programs in allograft endothelium inducing functions that promote monocyte recruitment to the graft and macrophage polarization. We propose infiltrating monocytes encounter local EC-derived factors and IgG-FcγR signals that prime them for differentiation. In the graft, the proangiogenic/profibrotic macrophages drive neointimal thickening. Antagonizing EC activation and/or monocyte recruitment may represent viable therapeutic targets to arrest cAMR and protect grafts from TV. We will employ a primary human EC culture model, a mouse cardiac allograft model of TV and human cardiac explants with TV to dissect the HLA I and II signaling pathways and mechanisms of monocyte recruitment and polarization. Aim 1: Define the structural requirements for complex formation and functional crosstalk between HLA Class I and its co-receptors ITGB4 and TLR4 in mediating endothelial cell activation, monocyte recruitment and macrophage polarization in cardiac TV. We will: (1a) determine how HLA I Ab- activated EC poise infiltrating monocytes to polarize to macrophages with distinct phenotypes and functions in vitro; (1b) characterize monocyte recruitment, TV lesion formation and phenotypes of infiltrating macrophages during MHC I antibody-mediated allograft injury and development of TV; (1c) define the structural requirements and functional effects of molecular crosstalk between ITGB4, TLR4 and HLA I on monocyte recruitment and macrophage polarization by Ab-activated EC in vitro; and (1d) characterize the requirement for MHC I co- receptors TLR4 and ITGB4 in monocyte recruitment, macrophage polarization and TV lesion formation in murine cardiac transplants. Aim 2. Define the structural requirements for complex formation and functional crosstalk between HLA class II and its co-receptor ITGB1 in mediating endothelial cell activation, monocyte recruitment and macrophage polarization in cardiac TV. We will: (2a) determine how HLA II Ab-activated EC poise infiltrating monocytes to polarize to macrophages with divergent phenotypes and functions in vitro; (2b) characterize monocyte recruitment, vascular lesion formation and phenotypes of infiltrating macrophages in a murine cardiac transplant model of MHC II cAMR and TV; (2c) define the structural requirements and functional effects of molecular crosstalk between ITGB1 and HLA II on monocyte recruitment and macrophage polarization by Ab-activated EC in vitro; (2d) characterize the requirement for MHC II coreceptor ITGB1 in monocyte recruitment, macrophage polarization and TV lesion formation in murine cardiac transplants; and (2e) define the phenotype profile of intragraft macrophages in human cardiac transplants with TV.