Mitochondrial metabolic modulation to minimize ischemic damage in donor heart

终末期心力衰竭（HF）患者需要机械循环支持，如果符合条件， 移植（HT）以挽救生命。多达20%的患者在接受心脏移植时死亡。的 由于目前供体心脏的唯一来源是病人， 不可逆的严重脑损伤（脑死亡后捐赠，DBD）。因此，迫切需要 扩大心脏捐献者的范围这种供体心脏的一个潜在来源是DCD（循环后捐赠）。 死亡）捐赠者。DCD供体增加了包括肝、肺和 肾不幸的是，DCD方案诱导了持续的热缺血时间，从而损害了心肌 排除了其用于临床移植。因此，缺血（ISC）形成DCD方案， 储存和再灌注（REP）相关损伤引起的心肌损伤联合收割机共同构成了额外的风险 加剧DCD心脏的损伤，从而妨碍其在临床移植中的使用。 虽然DCD心脏热缺血是不可避免的，但通过适当的治疗可以减轻REP损伤， 我们建议制定新的战略，以防止REP的发病， 损伤将减少对DCD心脏的损害。线粒体是心脏损伤的重要靶点和介质 我们以前的研究发现，在呼吸暂停时， ISC后心脏中REP的时间减少了心脏损伤。保护作用包括减少ROS 线粒体渗透性转换孔的产生和打开的抑制。我们建议 在早期REP之前和期间立即使用快速起效和可逆的呼吸抑制剂将 减少DCD心脏的心脏损伤。离体DCD人类心脏代表了一个独特的机会， 将来自强大的临床前数据的强大心脏保护作用转化为人类疾病。 在ISC-REP过程中受损的肌细胞激活了基于细胞的内部炎症机制，包括 NOD样受体蛋白3（NLRP 3）炎性小体。来自受损线粒体的ROS激活 NLRP 3炎性体并进一步维持组织损伤。该项目采用独特的方法， 联合收割机辅助干预，以钝化急性尿道驱动损伤并减轻长期REP 抑制炎性体信号传导的损伤。损伤的潜在机理关系 在DCD心脏模型中研究了线粒体和炎性体信号传导的活化。 我们假设最初的线粒体保护作用随后是线粒体的衰减- 激活的炎症信号将保护DCD心脏功能，使其能够用于心脏 移植我们的新策略是使用异戊巴比妥治疗快速，可逆和短暂 调节线粒体呼吸以减少急性ISC-REP损伤，随后抑制炎症 介导的损伤。本项目的目标是在复苏期间保护DCD心脏，维持 保存功能性线粒体，以达到类似于DBD的可移植心脏的质量。 使用大鼠DCD心脏模型，Aim 1将评价对电子传递可逆抑制的保护作用。 使用离体灌注心脏在REP开始时的转运。随后将是目标2， 这些有丝分裂保护DCD心脏的同基因异位HT将受到保护， 炎性小体介导的延迟再灌注损伤。然后将对异位移植的心脏进行评估 用于移植物存活、功能和病理学评估，包括炎性浸润的存在， MRI、超声心动图和组织学检查显示纤维化。我们计划的实验和预期的 这些结果将为DCD心脏的保护策略提供初步的见解。使用集成 治疗方法，我们的目标是走向使用DCD心脏的临床移植。

Patients with end stage heart failure (HF) require mechanical circulatory support, and if eligible, heart transplantation (HT) to save their life. Up to 20% of patients die while listed for heart transplantation. The supply of donor hearts has reached a plateau since the only current source of donor hearts consists of patients with irreversible severe brain damage (donation after brain death, DBD). Thus, there is an urgent need to expand the heart donor pool. A potential source of such donor hearts is from DCD (donation after circulatory death) donors. DCD donors have increased the transplantation rates of solid organs including liver, lungs and kidney. Unfortunately, DCD protocol induces a sustained warm ischemic time that damages myocardium precluding its use for clinical transplantation. Thus, the ischemia (ISC) form DCD protocol, the potential myocardial injury from storage, and reperfusion (REP) associated injury combine to represent additional risks to exacerbate injury in the DCD heart precluding their use in clinical transplantation. Although the warm ischemia is inevitable in DCD hearts, REP injury can be decreased through proper interventions applied at the onset of REP. We propose that development of new strategies to prevent REP injury will reduce damage to the DCD heart. Mitochondria are critical targets and mediators of cardiac injury during REP. Our previous studies found that temporary and reversible inhibition of mitochondrial respiration at the time of REP in hearts following ISC decreases cardiac injury. Protection involved the reduction of the ROS generation and inhibition of the opening of the mitochondrial permeability transition pores. We propose that the use of rapid onset and reversible inhibitors of respiration immediately before and during early REP will decrease cardiac injury in DCD hearts. The ex vivo DCD human heart represents a unique opportunity to translate this robust cardiac protection derived from strong pre-clinical data to human disease. Myocytes injured during ISC-REP activate internal cell-based mechanisms of inflammation including the NOD like receptor protein 3 (NLRP3) inflammasome. ROS derived from damaged mitochondria activates the NLRP3 inflammasome and perpetuates tissue injury further. This project takes the unique approach to combine complimentary interventions to blunt acute mitochondrial-driven injury and attenuate longer-term REP damage from inhibition of inflammasome signaling. The potential mechanistic relationship of damaged mitochondria and the activation of inflammasome signaling are studied in the DCD heart model. We hypothesize that the initial protection of mitochondria followed by attenuation of mitochondria- activated inflammatory signaling will protect the DCD heart function, enabling it to be used for heart transplantation. Our novel strategy is to use amobarbital treatment to rapidly, reversibly and transiently modulate mitochondrial respiration to decrease acute ISC-REP injury followed by inhibition of inflammation mediated injury. The goal of this project is to protect the DCD heart during reanimation, sustain the preservation of functional mitochondria in order to reach the quality of a transplantable heart similar to DBD. Using a rat model of DCD heart, Aim 1 will evaluate the protection from reversible inhibition of electron transport at the onset of REP using the perfused heart ex vivo. This will be followed by Aim 2 where the syngeneic heterotopic HT of these mito-protected DCD hearts will be subjected to protection from inflammasome mediated delayed reperfusion injury. The heterotopic transplanted hearts will then be evaluated for graft survival, function and pathological assessment including the presence of inflammatory infiltrates and fibrosis on MRI, ECHO cardiogram and histological examination. Our planned experiments and the anticipated results will provide the initial insight into the protective strategies for DCD hearts. Using an integrated treatment approach as proposed we aim to move toward use of DCD hearts for clinical transplantation.