Cell Death Pathways and Heart Transplant Rejection

同种异体移植物和接受死亡供体器官的患者的长期存活率仍然很低。那里 有证据表明，冷缺血（CI）和随后的再灌注在移植过程中， 细胞损伤，导致诱导针对供体组织的T细胞同种免疫，并最终移植 排斥反应缺血再灌注（IR）引发细胞损伤的机制，损伤的方式， 以及这种损伤如何影响同种异体反应还知之甚少。为了更好地模仿人类已故捐赠者 在心脏移植方面，费尔柴尔德实验室开发了一种同种异体心脏移植模型， 在移植前进行延长的CI，然后施用CTLA 4 Ig共刺激因子， 封锁接受CI的同种异体心脏被排斥，而未接受CI的心脏存活。 使用该模型的初步研究表明，受体中的补体缺乏延长了 CI处理的同种异体移植物的存活率，与循环TNFα降低相关。由于TNFα是一种已知的 坏死性凋亡，一种炎性细胞死亡，另外的研究表明，来自CYLD缺陷供体的器官， （其在坏死性凋亡中有缺陷）比野生型器官存活更长时间，而来自SHARPIN- 缺陷供体（其具有加速的坏死性凋亡）被迅速排斥。根据这些研究，我们 提出并将测试移植期间CI/IR后补体激活的统一假设 导致TNFα的诱导，然后诱导供体组织中细胞的坏死性凋亡。这又 炎性细胞损伤激活T细胞同种免疫以引起排斥。这项合作研究带来了 以高度协同的方式，将三个实验室的独特优势结合起来， 假说. (1)在目标1中，我们将研究甘露糖结合凝集素（MBL）启动的补体的作用 在我们的同种异体移植模型中，使用MBL、TNF和 TNFR敲除/敲入。(2)在目标2中，将研究TNF细胞死亡途径在CI引发的排斥反应中的作用。 进一步使用来源于CYLD，RIPK 1，RIPK 3， Caspase-8和SHARPIN。(3)坏死性凋亡释放损伤相关分子模式（DAMP）， 在目标3中，我们将研究DAMP如何被宿主抗原呈递细胞感知，以及这如何导致 诱导T细胞同种异体反应。所有这三个目标都非常重要，因为它们将提供关于 在移植物排斥的（1）起始、（2）细胞损伤和（3）效应期， 移植中的治疗性阻断。该方法具有直接和积极影响的强大潜力 移植患者的结果。

The long-term survival of allografts and patients receiving deceased donor organs remain poor. There is evidence to suggest that cold ischemia (CI) and the subsequent reperfusion during transplants causes cellular injury, which leads to an induction of T cell alloimmunity against donor tissue and ultimately graft rejection. The mechanism by which ischemia reperfusion (IR) initiates cellular injury, the manner of the injury, and how this injury impacts alloresponse are poorly understood. To better mimic human deceased donor transplants, the Fairchild laboratory developed an allograft heart transplant model whereby the donor organ is subjected to extended CI prior to transplantation followed by administration of an CTLA4Ig costimulatory blockade. Allograft hearts subjected to CI were rejected whereas those not subjected to CI survived. Preliminary studies using this model indicated that complement deficiency in the recipients prolonged the survival of CI-treated allografts, correlating with reduced circulating TNFα. As TNFα is a known inducer of necroptosis, an inflammatory cell death, additional studies showed that organs from CYLD-deficient donors, (which are defective in necroptosis) survived longer than wild type organs whereas organs from SHARPIN- deficient donors (which have accelerated necroptosis) were rapidly rejected. Based on these studies, we have proposed and will test the unifying hypothesis that complement activation following CI/IR during transplants leads to the induction of TNFα, which then induces necroptosis of cells in donor tissues. In turn, this inflammatory cellular injury activates T cell alloimmunity to cause rejection. This collaborative study brings together in a highly synergistic manner the unique strengths of three laboratories to address the above hypothesis. (1) In Aim 1, we will examine the role of mannose-binding lectin (MBL)-initiated complement activation in triggering TNF-dependent necroptosis in our allograft transplant model using MBL, TNF and TNFRs knockouts/knockins. (2) In Aim 2, the role of the TNF cell death pathway in CI-initiated rejection will be further dissected using donor organs derived from various knockouts/knockins of CYLD, RIPK1, RIPK3, Caspase-8 and SHARPIN. (3) Necroptosis releases damage-associated molecular patterns (DAMPs) and thus in Aim 3, we will examine how DAMPs are sensed by host antigen-presenting cells and how this leads to induction of the T cell alloresponse. All three aims are highly significant, as they will provide insights into which molecules during (1) initiation, (2) cellular injury and (3) effector phases of graft rejection may be targeted for therapeutic blockade in transplants. The approach has strong potential to directly and positively impact outcomes of transplant patients.