Innate-Adaptive Immunoregulation in Liver Transplant Ischemia/Reperfusion Injury

项目3 -摘要/摘要 缺血和再灌注损伤（IRI）是移植过程中不可避免的结果，是移植过程中的一个关键限制。 人类肝脏移植IRI引起的细胞损伤可导致原发性无功能， 再次移植肝脏IRI也使受体易于发生急性和慢性排斥反应以及移植物丢失， 同时也减少了可移植器官的数量。小鼠模型中的关键观察结果表明，肝脏IRI 是由先天免疫系统和适应性免疫系统介导的。很少有人知道类似的机制是否 在人类肝脏移植中发挥作用。我们的中心假设是，识别DAMP及其相关的 PRR在人类OLT-IRI期间驱动骨髓细胞可塑性，这将使我们能够开发治疗干预措施， 改善奥尔特结果。目的1、确定TLR 9和TLR 7/NOD 2配体介导的分化 炎症性巨噬细胞对调节性巨噬细胞的极化以及与T细胞的串扰。我们会描述细胞的特征- 术前/术后游离DNA/TLR 9配体线粒体DNA、中性粒细胞胞外陷阱和HMGB 1/HMGB 2 再灌注血液，以确定它们对人OLT-IRI的贡献，巨噬细胞增殖和发育的能力， 同种异体反应性T细胞我们还将确定TLR 7和NOD 2配体对其调节的致耐受性作用。 减轻TLR 4和/或TLR 9极化和抑制同种免疫的潜力。表征 获得的受体血液和供体同种异体移植物活检的骨髓和T细胞表型的纵向变化 再灌注前和再灌注后将确定沿着内型特异性途径的新的靶向相互作用 TLR 4/TLR 9和TLR 7/NOD 2原位。目的2将确定PRR预处理的治疗潜力， 减轻骨髓活化和OLT-IRI。我们将评估TLR 4/9拮抗剂的治疗潜力， 用血液减轻体外IR介导的损伤、巨噬细胞炎症极化和T细胞同种异体反应性 从人奥尔特接受者和在IRI的啮齿动物模型中的体内。空间免疫特征的原位评估 对照与耐受器官中供体与受体骨髓细胞的比较将探索IRI介导的免疫细胞， 实质变化导致移植物中的调节与炎症状态。目标3将决定 DAMP/PRR内型对同种免疫产生和移植结果的影响我们将评估 DAMP/PRR内型，其指导同种异体反应性T细胞和供体特异性T细胞的产生和广度。 抗体的数据将与临床元数据、血液和移植物免疫表型、IRI和移植物 结果，以确定导致IRI易感性和更差结果的DAMP/PRR内型。

PROJECT 3 - SUMMARY/ABSTRACT Ischemia and reperfusion injury (IRI), the inevitable consequence of the transplant process, is a key limitation to human liver transplantation. The cellular damage incurred by IRI can lead to primary non-function necessitating re-transplantation. Liver IRI also predisposes the recipient to both acute and chronic rejection and graft loss, as well as, decreasing the pool of transplantable organs. Key observations in murine models indicate that liver IRI is mediated by both the innate and adaptive immune systems. Little is known whether similar mechanisms are at play in human liver transplantation. Our central hypothesis is that identifying the DAMPs and their associated PRRs driving myeloid cell plasticity during human OLT-IRI will permit us to develop therapeutic interventions to improve OLT outcomes. Aim 1, we will determine the TLR9 and TLR7/NOD2 ligands mediating differential polarization of inflammatory vs. regulatory macrophages and crosstalk with T cells. We will characterize the cell- free DNA/TLR9 ligands mitochondrial DNA, neutrophil extracellular traps, and HMGB1/RAGE in pre/post reperfusion blood to establish their contributions to human OLT-IRI, ability to polarize macrophages and develop alloreactive T cells. We will also determine the tolerogenic effect of TLR7 and NOD2 ligands for their regulatory potential to mitigate TLR4 and/or TLR9 polarization and suppression of alloimmunity. Characterization of longitudinal changes in myeloid and T cell phenotypes of recipient blood and donor allograft biopsies obtained pre- and post-reperfusion will identify new targetable interactions along the endotype-specific pathways TLR4/TLR9 and TLR7/NOD2 in situ. Aim 2 will determine the therapeutic potential of PRR preconditioning to mitigate myeloid activation and OLT-IRI. We will assess the therapeutic potential of TLR4/9 antagonists to mitigate IR mediated-injury, macrophage inflammatory polarization and T cell alloreactivity in vitro using blood from human OLT recipients, and in vivo in rodent models of IRI. In situ assessment of the spatial immune profile of donor vs. recipient myeloid cells in control vs. tolerized organs will explore the IRI-mediated immune cell and parenchymal changes contributing to a regulatory vs inflammatory state in the graft. Aim 3 will determine the impact of DAMP/PRR endotypes on generation of alloimmunity and graft outcome. We will assess the DAMP/PRR endotypes that direct the generation and breadth of alloreactive T cells and donor specific antibodies. Data will be integrated with clinical metadata, blood and graft immunophenotypes, IRI and transplant outcomes to determine DAMP/PRR endotypes responsible for susceptibility to IRI and worse outcomes.