Isochoric Pressure Based Preservation of Cells, Tissues and Organs

 描述（由申请人提供）：生物医学领域的大量工作都需要保存活生物材料，包括医学研究和药物发现、器官和组织移植、基于细胞的治疗、生育和再生医学、应急准备和创伤护理。细胞保存在历史上依赖于通用的冷冻保存方法，其通常导致显著的细胞死亡、细胞凋亡和批次间差异。我们建议开发的技术，结合现有的保存方法，使用人类血管内皮细胞的方法基于自然。最终，这项技术可以扩大到组织，甚至整个器官。我们的方法是使用等容（恒定体积）压力系统，以实现生物系统在高的零下温度下的热稳定的非冷冻保存，该系统基于自然界中过冷和冬眠动物所采用的策略，用溶液制剂进行增强。该资助的目的是确定在-5至-20 ° C的温度范围内采用等容压力（<95 MPa）的冷冻保存溶液配方和方案，这些配方和方案与这些条件下的细胞存活相容。这项工作分为3个具体目标：在目标1中，我们在加州大学伯克利分校的子项目团队将对一系列候选低温恒温鸡尾酒和协议进行热力学分析，这些协议将压力限制在<95 MPa，并允许冷却到低至-20 ° C的温度而不完全冻结。在目标2中，将使用组合的高通量方法用人血管内皮细胞评价确保潜在非冷冻状态的电极导线条件。最后，在目标3中，新定义的低温恒温鸡尾酒和早期目标的最佳冷却/升温速率将用于测试等容储存后的细胞恢复，作为温度、压力和储存持续时间的函数。在证明使用这种创新方法进行冷冻保存的可行性之后，我们将计划进行第二阶段SBIR研究，其中进行进一步优化，并使用我们的技术评估各种细胞，组织和器官的冷冻保存研究和医学利益。

 DESCRIPTION (provided by applicant): Preservation of living biological materials is needed for a huge range of endeavors in biomedicine, spanning medical research and drug discovery, organ and tissue transplantation, cell-based therapies, fertility and regenerative medicine, emergency preparedness, and trauma care. Cell preservation has historically relied on one-size-fits-all cryopreservation approaches that often lead to significant cell death, apoptosis and batch-to- batch variation. We propose development of technology using an approach based on nature in combination with existing preservation methods using human vascular endothelial cells. Ultimately this technology can be scaled up for tissues and possibly whole organs. Our approach is to use an isochoric (constant volume) pressure system to achieve thermodynamically stable non-frozen preservation of biological systems at high sub-zero temperatures that is augmented with solution formulations based upon strategies employed by supercooling and hibernating animals in nature. The objective of this grant is to identify cryopreservation solution formulations and protocols employing isochoric pressure (<95MPa) at temperatures ranging from -5 to -20°C that are compatible with cell survival under these conditions. The work is divided into 3 specific aims: In aim 1 our subaward team at University of California, Berkeley, will perform thermodynamic profiling of a series of candidate cryostasis cocktails and protocols that will limit pressure to <95MPa and allow cooling to temperatures as low as -20°C without complete freezing. In aim 2 the lead conditions that ensures a potential non-frozen state will be evaluated with human vascular endothelial cells using a combinatorial, high throughput approach. Finally, in aim 3 the newly defined cryostasis cocktails and optimal cooling/warming rates from the earlier aims will be used to test cell recovery after isochoric storage as a function of temperature, pressure and storage duration. Following demonstration of feasibility using this innovative approach to cryopreservation we will plan Phase II SBIR studies in which further optimization is performed and cryopreservation of various cells, tissues and organs of research and medical interest are evaluated using our technology.