Preservation of human organs at subzero temperatures has been an elusive goal for decades. The major complication hindering successful subzero preservation is the formation of ice at temperatures below freezing. Supercooling, or subzero non-freezing, preservation completely avoids ice formation at subzero temperatures. We previously showed that rat livers can be viably preserved three times longer by supercooling as compared to hypothermic preservation at +4 °C. Scalability of supercooling preservation to human organs was intrinsically limited because of volume-dependent stochastic ice formation at subzero temperatures. However, we recently adapted the rat preservation approach so it could be applied to larger organs. Here, we describe a supercooling protocol that averts freezing of human livers by minimizing air-liquid interfaces as favorable sites of ice nucleation and uses preconditioning with cryoprotective agents to depress the freezing point of the liver tissue. Human livers are homogeneously preconditioned during multiple machine perfusion stages at different temperatures. Including preparation, the protocol takes 31 h to complete. Using this protocol, human livers can be stored free of ice at -4 °C, which substantially extends the ex vivo life of the organ. To our knowledge, this is the first detailed protocol describing how to perform subzero preservation of human organs.

中文翻译：

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过冷状态下人体肝脏的零度以下非冷冻保存。

几十年来，在零下温度下保存人体器官一直是一个难以实现的目标。阻碍成功零下保存的主要并发症是在低于冰点的温度下形成冰。过冷或零度以下非冷冻保存完全避免了在零度以下温度下结冰。我们之前表明，与 +4 °C 下的低温保存相比，过冷可以有效地保存大鼠肝脏的三倍。由于在零度以下温度下与体积相关的随机冰形成，过冷保存对人体器官的可扩展性本质上受到限制。然而，我们最近调整了大鼠保存方法，以便将其应用于更大的器官。这里，我们描述了一种过冷方案，该方案通过最大限度地减少空气-液体界面作为冰核形成的有利部位，并使用冷冻保护剂进行预处理来降低肝组织的冰点，从而避免人类肝脏冻结。人类肝脏在不同温度下的多个机器灌注阶段进行均匀预处理。包括准备在内，该协议需要 31 小时才能完成。使用该协议，人类肝脏可以在 -4°C 下无冰储存，这大大延长了器官的离体寿命。据我们所知，这是第一个描述如何对人体器官进行零下保存的详细协议。人类肝脏在不同温度下的多个机器灌注阶段进行均匀预处理。包括准备在内，该协议需要 31 小时才能完成。使用该协议，人类肝脏可以在 -4°C 下无冰储存，这大大延长了器官的离体寿命。据我们所知，这是第一个描述如何对人体器官进行零下保存的详细协议。人类肝脏在不同温度下的多个机器灌注阶段进行均匀预处理。包括准备在内，该协议需要 31 小时才能完成。使用该协议，人类肝脏可以在 -4°C 下无冰储存，这大大延长了器官的离体寿命。据我们所知，这是第一个描述如何对人体器官进行零下保存的详细协议。