Center for Engineering in Medicine and Surgery, Harvard Medical School and Massachusetts General Hospital, Boston, Massachusetts, USA.
Shriners Hospitals for Children - Boston, Boston, Massachusetts, USA.
FASEB J. 2023 Oct;37(10):e23187. doi: 10.1096/fj.202300076R.
Despite decades of effort, the preservation of complex organs for transplantation remains a significant barrier that exacerbates the organ shortage crisis. Progress in organ preservation research is significantly hindered by suboptimal research tools that force investigators to sacrifice translatability over throughput. For instance, simple model systems, such as single cell monolayers or co-cultures, lack native tissue structure and functional assessment, while mammalian whole organs are complex systems with confounding variables not compatible with high-throughput experimentation. In response, diverse fields and industries have bridged this experimental gap through the development of rich and robust resources for the use of zebrafish as a model organism. Through this study, we aim to demonstrate the value zebrafish pose for the fields of solid organ preservation and transplantation, especially with respect to experimental transplantation efforts. A wide array of methods were customized and validated for preservation-specific experimentation utilizing zebrafish, including the development of assays at multiple developmental stages (larvae and adult), methods for loading and unloading preservation agents, and the development of viability scores to quantify functional outcomes. Using this platform, the largest and most comprehensive screen of cryoprotectant agents (CPAs) was performed to determine their toxicity and efficiency at preserving complex organ systems using a high subzero approach called partial freezing (i.e., storage in the frozen state at -10°C). As a result, adult zebrafish cardiac function was successfully preserved after 5 days of partial freezing storage. In combination, the methods and techniques developed have the potential to drive and accelerate research in the fields of solid organ preservation and transplantation.
尽管经过了几十年的努力，复杂器官的保存仍然是移植的一个重大障碍，加剧了器官短缺危机。器官保存研究的进展受到不理想的研究工具的严重阻碍，这些工具迫使研究人员在通量和可转移性之间做出牺牲。例如，简单的模型系统，如单细胞单层或共培养物，缺乏天然组织结构和功能评估，而哺乳动物整个器官是具有混杂变量的复杂系统，不适合高通量实验。为了应对这一挑战，不同的领域和行业通过开发丰富而强大的资源，将斑马鱼作为一种模式生物，来弥补这一实验空白。通过这项研究，我们旨在展示斑马鱼在实体器官保存和移植领域的价值，特别是在实验移植方面。我们专门针对斑马鱼开发了多种方法并对其进行了验证，包括在多个发育阶段（幼虫和成年）进行保存特定实验的方法、加载和卸载保存剂的方法，以及开发用于量化功能结果的活力评分的方法。利用这一平台，我们对大量的冷冻保护剂（CPAs）进行了最大和最全面的筛选，以确定它们在使用高亚低温方法（即部分冷冻，即在-10°C 的冷冻状态下储存）保存复杂器官系统时的毒性和效率。结果，成年斑马鱼的心脏功能在经过 5 天的部分冷冻储存后成功保存。总之，开发的方法和技术有可能推动和加速实体器官保存和移植领域的研究。
