Extended Storage of Tissues and Organs in Subzero Environments

描述(由申请人提供)： 美国目前有大约10万名患者在等待器官移植，这个数字远远超过了可用器官的供应，而且每年还在继续增长约5%。最有希望的解决方案，生物人工组织和器官构建以及捐赠器官再工程方法，最终都受到生物保存技术的限制，因为在实验室制备的任何组织工程产品都必须储存一段时间才能使用。目前整个器官保存的黄金标准是在冰上冷藏长达72小时，在此期间器官不断变质。一种更好的生物保存方法，可以延长组织保存时间，超出目前的限制，目前还没有开发出来。这种方法将为组织和器官保存、组织和器官运输以及组织和器官移植提供关键的使能技术。本研究的目的是延长肝组织在过冷保存介质中的低温非冷冻保存的存活时间。本研究的中心假设依赖于两个现象：1)3-O-甲基葡萄糖(3OMG)可降低SZNF的稳定温度，且无主要毒副作用；2)常温灌流复温可减轻再灌注损伤。我们的假设是基于我们的初步发现，确立了3OMG作为一种对肝细胞毒性最小的冷冻保护剂，并建立了常温灌流可以显著逆转缺血的损害效应。这项研究的基本原理是，如果能够在避免防冻剂毒性的同时实现过冷保存，那么器官新陈代谢可以进一步放缓，从而将缺氧/缺血损伤减少到最低水平。建立零度以下的不冻保存技术将是该领域的一项受欢迎的创新。本文描述的工作将有助于开发这种过冷储存的使能技术，并为评估保存后的肝脏和生物人工器官的活性建立定量标准。虽然我们将重点放在肝脏上，但我们预计这里建立的方案也将作为其他组织工程产品的非冷冻保存的基础，例如人工器官替代品和种子支架结构。 公共卫生相关说明(申请人提供)：目前有97,000名患者在等待移植名单上，这个数字每年增加~5%。向公众提供更多捐献器官以及组织工程替代品的一个关键瓶颈是有限的保存期限。这项研究的目的是延长器官和生物人工替代品的存活保存时间，使其能够在零度以下的温度下延长保存时间，而不会结冰。这项研究的结果有望通过增加捐赠器官的可用性和使更多的移植成为可能，直接改善公共卫生。

DESCRIPTION (provided by applicant): There are currently ~100,000 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. The most promising solutions, bioartificial tissue and organ construction and donor organ reengineering methodologies, are both ultimately limited by biopreservation technologies, as any tissue engineered products prepared in a laboratory will have to be stored for a period of time until utilization. The current gold standard for whole organ preservation is cold storage on ice for up to 72 hours, during which time the organ continuously deteriorates. A superior biopreservation method that extends the tissue storage time beyond current limitations is yet to be developed. Such a method would provide a crucial enabling technology for tissue and organ preservation, tissue and organ transport, and tissue and organ transplantation. The objective of this study is to extend the viable preservation time of hepatic tissues by sub-zero non- freezing (SZNF) storage in a supercooled preservation medium. The central hypothesis of this study relies on two phenomena: 1) that 3-O-methyl-glucose (3OMG) lowers achievable stable SZNF temperature without major toxic side effects, and that 2) rewarming by normothermic perfusion reduces reperfusion damage. Our hypothesis has been formulated based on our preliminary findings establishing 3OMG as a minimally toxic cryoprotectant for hepatocytes, and establishing that normothermic perfusion can significantly reverse the damaging effects of ischemia. The rationale of the study is that if supercooled preservation can be achieved while avoiding antifreeze toxicity, then organ metabolism can be further slowed thereby reducing anoxic/ischemic damage to minimal levels. Establishment of a sub-zero nonfreezing preservation technology will be a welcome innovation to the field. The work described herein will help develop this enabling technology of supercooled storage, and also establish quantitative standards for evaluating the liver and bioartificial organ viability following preservation. While we focus on the liver, we expect that the protocols established here will also serve as the basis for subzero nonfreezing preservation of other tissue engineered products, such as artificial organ substitutes and seeded scaffold constructs. Public Health Relevance Statement (provided by applicant): There are currently 97,000 patients on the transplant waiting list, and the number increases by ~5% every year. A critical bottleneck in making more donor organs as well tissue engineering alternatives available to the public is the limited preservation duration. The objective of this study is to extend the viable preservation time of organs and bioartificial alternatives by enabling extended storage at sub-freezing temperatures without ice formation. The results of this study are expected to directly improve public health by increasing donor organ availability and making more transplantations possible.