Subzero preservation of vascular composite allografts

抽象的。VCA移植的技术可行性早在20年前就得到了论证，并在临床上初步应用 结果令人振奋。然而，终生免疫抑制的伦理考量确实经常排除VCA。 移植。为了避免长期免疫抑制的副作用，最有希望的方法是诱导 通过骨髓联合移植的混合嵌合体耐受。这是唯一一种在 实体器官移植的大型动物和临床研究。然而，目前临床试验中的耐受性方案需要 从最初的供者移植物恢复到受者，在3至7天内对受者进行预调节 移植。避免需要预适应来诱导混合嵌合体似乎并不容易实现和 在临床VCA移植案例中，当一个团队尝试时失败了。与肾脏相反，在VCA中不存在 活体捐赠，使得受者预先调理在临床上不现实。因此，一种延长嫁接的技术 保存以缩短受体条件作用的时间仍然是实现耐受性诱导的关键必要 在VCA移植中。 我们的总体目标是开发一种高低温保存方案，以延长血管组织的存活率 在移植前保存期间，从而创建实施耐受诱导方案的时间窗口。 这项研究的首要假设是减轻低温保护剂(CPA)加载期间的渗透休克， 减少CPA装卸过程中对内皮细胞的剪切损伤和毒性，并绕过失控 通过纳米温控从零度以下的温度复温期间的缺血，将使更好的和更长时间的保存 同种异体肢体移植。这一假设是基于我们之前的数据提出的，这些数据证明了 人体肝脏显示出可伸缩性的证据；然而，最初尝试直接对大鼠和猪的肢体采用相同的方案 导致过度水肿和对缺血损伤和血管阻力的极端敏感性 肝脏。为这项工作建立的多机构伙伴关系汇集了两个机构的独特专长： 1)麻省理工学院医学与外科工程中心的器官存储为零度以下，2)明尼苏达大学 医学工程研究所的独特纳米武器技术。 我们的预期贡献是开发了一种新的大鼠血管复合组织保存方案 这将总储存期从几小时延长到几天，并为接受者提供了时间来准备诱导耐受性。这 贡献是重大的，因为这里开发的方法可能是对临床方案的概念验证 对于VCA移植的耐受性诱导是实用的，因此可以使它们得到广泛的使用，而这是不可能的 目前。

ABSTRACT. The technical feasibility of VCA transplantation was demonstrated 20 years ago and the initial clinical outcomes have been encouraging. However, the ethical calculus of lifelong immunosuppression does often preclude VCA transplantation. To avoid the side effects of long-term immunosuppression, the most promising approach is the induction of tolerance by mixed chimerism via bone-marrow co-transplantation. This is the only strategy that has shown efficacy in large animal and clinical studies of solid organ transplantation. However, current tolerance protocols in clinical trials require somewhere between 3 to 7 days to pre-condition the recipient, measured from initial donor graft recovery to recipient transplantation. Sidestepping the need for preconditioning to induce mixed chimerism does not appear easy to achieve and failed in a clinical VCA transplant case when attempted by one team. Contrary to kidneys, in VCA there is no possibility of living donation, making advance recipient conditioning clinically unrealistic. Therefore, a technology to extend graft preservation to bridge the time for recipient conditioning remains a critical necessity that would enable tolerance induction in VCA transplants. Our overall goal is to develop a high subzero preservation protocol that will prolong the viability of vascular tissues during preservation prior to transplantation, thereby creating a time window to implement tolerance induction protocols. The overarching hypothesis of this study is that alleviating osmotic shocks during cryoprotective agent (CPA) loading, reducing shear injury and toxicity to endothelial cells during loading and unloading CPAs, and bypassing uncontrolled ischemia during rewarming from subzero temperatures by nanowarming, will enable superior and extended preservation of limb allografts. This hypothesis has been formulated based on our prior data demonstrating success in supercooling of human livers showing proof of scalability; however initial attempts to directly adopt the same protocol for rat and pig limbs led to excessive edema and identification of extreme sensitivity to ischemic injury and vascular resistance compared to livers. The multi-institutional partnership assembled for this work brings together the unique expertise of two institutions: 1) subzero organ storage at the MGH Center for Engineering in Medicine & Surgery, and 2) the University of Minnesota Institute for Engineering in Medicine’s unique technology of nanowarming. Our expected contribution here is the development of a new preservation protocol for rat vascularized composite tissues that extends total storage duration from hours to days, and provides time to prepare the recipient to induce tolerance. This contribution is significant because the methods developed here could be the proof-of-concept for a protocol that is clinically practical for tolerance induction in VCA transplants, and could therefore enable their wide-spread use which is not possible currently.