Whole heart suspended animation: leveraging the zebrafish to solve the organ shortage

目前，美国有超过123，000名患者在器官移植等待名单上，这个数字远远超过了 可用器官的供应，并继续每年增长约5%。在加拿大，每十分钟 有人被添加到等待名单中，每天约有22人在等待移植时死亡。一个主要原因是， 我们目前的器官保存和处理实践的局限性，这严重限制了保存 持续时间和不允许移植轻微受伤的器官。该项目旨在制定一项协议， 将心脏储存数周，以实现全球器官匹配并克服缺血耐受性的局限性， 来增加心脏捐献者的数量 我建议解决目前心脏移植障碍的策略，包括离体保存和 边缘器官的恢复，应该向大自然寻求灵感。更确切地说，是下降到 “假死”状态，其特征是生命过程减慢和耐应激性提高， 有希望我将系统地利用假死的两种方法; 1）冷冻耐受，2） 高温冬眠利用自然界中耐寒的树蛙的灵感，我试图开发一种 整个心脏保存的方案，该方案将在存在以下物质的情况下实现高的零度以下储存温度： 细胞外的冰，以及数周至数月的储存时间。此外，我将确定药理学试剂， 将战略性地降低代谢需求，以匹配氧气/营养供应，并增加热缺血 从自然界中冬眠的灵长类动物中获得灵感， 为了实现这一目标，我建议引入斑马鱼作为低温生物学的一种新的模型系统， 器官移植虽然斑马鱼已成为研究人类疾病的首选研究动物， 从未被用作开发器官处理和保存的创新方法的工具， 了解器官移植的基础生物学。典型的模式生物如斑马鱼 提供了许多优点，包括强大的遗传/生物化学工具，易于实时成像， 高通量研究，并且便宜且易于维护。通过扩大规模的实验， 通过在包括大鼠在内的哺乳动物模型系统中进行验证，本提案将弥补 研究领域。

There are currently over 123,000 patients on the organ transplant waiting list in the US, a number that far exceeds the supply of available organs, and that continues to grow ~5% each year. In Canada, every ten minutes someone is added to the waitlist and ~22 people die each day while waiting for a transplant. A major reason is the limitations of our current organ preservation and handling practices, which severely restricts preservation duration and disallows the transplantation of marginally injured organs. This project aims to develop a protocol to bank hearts for several weeks to enable global organ matching and overcome limitations of ischemia tolerance to increase the donor pool of hearts. I propose that strategies to solve current barriers in heart transplantation, including ex vivo preservation and recovery of marginal organs, should look to nature for inspiration. More specifically, the ability to descend into a state of “suspended animation”, characterized by the slowing of life processes and improved stress tolerance, holds promise. I will systematically leverage two methods of suspended animation; 1) freeze-tolerance, and 2) high-temperature hibernation. Using inspiration from freeze-tolerant wood frogs in nature, I seek to develop a protocol for whole heart preservation which will achieve high subzero storage temperatures in the presence of extracellular ice, and storage durations of weeks to months. Further, I will identify pharmacological agents which will strategically lower metabolic demands to match oxygen/nutrient availability, and increase warm ischemia tolerance from minutes to hours, using inspiration from hibernating primates in nature. To accomplish this goal, I propose to introduce the zebrafish as a novel model system for cryobiology and organ transplantation. While zebrafish has become a favored research animal for studying human disease, it has never been used as a tool to develop innovative approaches for organ handling and preservation or to understand the underlying biology of organ transplantation. Classical model organisms such as the zebrafish offer many advantages including robust genetic/biochemical tools, ease of real-time imaging, compatibility with high-throughput studies, and are cheap and easy to maintain. Complimented by scale-up experiments and validation in mammalian model systems, including rats, the present proposal will mend a critical gap in the research field.