Organ banking for transplant--kidney cryopreservation by vitrification and novel nanowarming technology

摘要 慢性肾脏疾病是一个重大的医疗保健问题，影响超过15%的美国人口，耗资数十亿美元。 每年的医疗费用。移植是大多数疾病进展患者的最佳选择， 与透析相比，导致预期寿命的显著增加和生活质量的改善。的 据估计，美国潜在的已故捐赠者器官供应量将超过目前的移植数量 4至5倍，主要限制是可接受的移植器官数量， 在恢复和植入之间持续的缺血性损伤。一种冷冻保存或“银行”肾脏的方法， 器官移植将有效地消除器官分配供应链中时间的影响。这 这将为移植提供一个新的范例，可以改善供体/受体的匹配， 患者准备，促进耐受诱导方案，并增加器官利用率，同时改善移植物 患者生存率。一种有希望克服传统策略局限性的方法是 玻璃化--即器官冷却得太快，以至于它们不能经历从液体到固体的相变 冰在冷冻保护剂（CPA）的帮助下，器官进入稳定的玻璃样状态，其中活的 存储在理论上是不确定的。然而，关键的挑战是在没有冰形成或破裂的情况下重新升温： 如果复温太慢，就会形成冰晶，如果复温不均匀，热应力就会导致开裂。期间 在我们最初的R 01资金支持下，我们开发了一种称为“重新武装”的新方法，实现了这两个目标。 氧化铁纳米颗粒与CPA溶液一起沿着灌注到整个器官的脉管系统中。的 然后通过冷却将器官玻璃化，并通过将其置于射频线圈中按需重新加热， 在纳米颗粒中加热，因此从器官内加热。我们发现冷冻可以使 玻璃化的器官，包括肾脏，在动物模型中。我们最近第一次证明， 器官在移植后的体外和体内功能。此外，我们成功地进行了玻璃化冷冻， 人类大小（猪）肾脏的移植。这些新数据支持我们的方法的可行性， 冷冻保存和移植整个人体器官。然而，许多问题 包括与控制器官相比，受损的肾脏功能如何，发生了什么损伤（如果有的话） 以及如何扩大到人类大小的器官。在本次更新R 01中，我们建议：（1） 定量评估大鼠模型中冷冻保存和纳米臂肾移植功能，包括长期肾移植 长期保存，长期功能，损伤模式和宿主免疫反应的改变，（2）工程师 和优化放大，用于玻璃化冷冻人体大小的器官，和（3）玻璃化和冷冻人体大小的器官 肾脏，同时测量活力，结构完整性和器官功能。

ABSTRACT Chronic kidney disease is a significant healthcare issue affecting >15% of the U.S. population and costing billions in healthcare dollars annually. Transplantation is the best option for most patients with progressive disease, resulting in a significant increase in life expectancy and improved quality of life compared to dialysis. The potential U.S. deceased donor organ supply is estimated to exceed the current number of organs transplanted by a factor of 4- to 5-fold, with a major limitation to the number of acceptable organs for transplant being the ischemic injury sustained between recovery and implantation. A method to cryopreserve or “bank” kidneys prior to transplant would effectively remove the influence of time from the supply chain of organ distribution. This would allow a new paradigm for transplantation that would improve donor/recipient matching, allow for better patient preparation, facilitate tolerance induction protocols, and increase organ utilization while improving graft and patient survival. One promising approach that overcomes the limitations of conventional strategies is vitrification—that is, cooling organs so quickly that they cannot undergo the phase transition from liquid to solid ice. With the help of cryoprotective agents (CPAs), the organ enters a stable glass-like state wherein viable storage is theoretically indefinite. The critical challenge, however, is rewarming without ice formation or cracking: if rewarming is too slow, ice crystals form, and if rewarming is not uniform, thermal stress causes cracking. During our initial R01 funding, we developed a novel approach termed “nanowarming” that achieved both objectives. Iron oxide nanoparticles were perfused throughout the vasculature of the organ along with CPA solutions. The organ was then vitrified by cooling and rewarmed on-demand by placing it in a radiofrequency coil that induces heating in the nanoparticles and, therefore, from within the organ. We found that nanowarming could rewarm vitrified organs, including kidneys, in animal models. We have recently shown, for the first time, that nanowarmed organs function in vitro and in vivo following transplantation. Further, we showed successful vitrification and nanowarming of human-sized (porcine) kidneys. These new data support the feasibility of our approach to cryopreserve and nanowarm whole human organs for transplantation. Nevertheless, many questions remain, including how nanowarmed kidneys function compared to control organs, what, if any, injury occurs during nanowarming, and how to scale up to human-sized organs. In this renewal R01, we propose to: (1) Quantitatively assess cryopreserved and nanowarmed kidney transplant function in a rat model, including long- term preservation, long-term function, modes of injury, and alterations of the host immune response, (2) Engineer and optimize scale-up for nanowarming vitrified human-sized organs, and (3) Vitrify and nanowarm human-sized kidneys while measuring viability, structural integrity, and organ function.