Cryopreservation of Engineered Tissue Equivalents

描述（由申请人提供）：为了减少动物毒性试验的使用，公司已经开发出体外皮肤模型或等同物来取代Draize皮肤刺激试验。从2009年开始，欧盟禁止使用动物收集化妆品成分的毒理学数据，这进一步推动了这一发展。这些工程组织的皮肤等效物的供应是一个重要的问题，可以通过开发有效的冷冻保存方法来解决。目前，这些结构是定制的，需要几个星期的交货时间才能使用。冷冻保存的产品将在需要时提供，从而消除生产产品所需的前置时间。此外，可以对准备发货的库存进行质量控制检查，并且可以批量生产，从而为供应商节省成本。低温保存的皮肤模型还将为最终用户提供更大的灵活性来安排实验，而不必担心结构的可用性，并且可以从同一批次的产品中订购，以便更好地控制研究中的一致性。在第一阶段，我们的替代策略，玻璃化被用于成功低温保存皮肤构建表皮，存活率为91%。第二阶段提出了三个具体目标。玻璃化方案将针对插入井中的上皮组织结构进行优化，不仅包括表皮，还包括与毒性测试相关的其他上皮组织结构。此外，实验计划扩大玻璃化过程，以获得更多的样品，并确定最佳的储存和运输条件。这种优化的过程也适用于其他组织工程组织结构，如心血管组织等同物。

DESCRIPTION (provided by applicant): In an effort to reduce the use of animals for toxicity testing, companies have developed in vitro skin models or equivalents to replace the Draize skin irritation test. This development has been further pushed by the European Union regulation that has prohibited the use of animals for collecting toxicological data on cosmetic ingredients that began 2009. Supply of these engineered tissues skin equivalents is a significant issue that can be solved by development of effective cryopreservation methods. Presently, these constructs are made to order and require a lead time of several weeks before they can be used. A cryopreserved product would be available when needed eliminating lead time required to manufacture the product. Additionally, performance of quality control checks for stock that is ready to be shipped could be performed and large batches could be manufactured providing a cost savings to suppliers. A cryopreserved skin model would also provide the end- user greater flexibility for scheduling of experiments without concern regarding availability of the constructs and it would be possible to order from the same batch of product in order to have better control of uniformity within research studies. In Phase I, our alternate strategy, vitrification was used t successfully cryopreserve the skin construct Epiderm with a viability of 91%. Three specific aims are proposed for Phase II. The vitrification protocol will be optimized for epithelial tissue constructs in well inserts to include not only Epiderm but other epithelial tissue constructs relevant to toxicity testing. In addition, experiments are planned to scale up the vitrification process for larger numbers of samples and determine optimum storage and shipping conditions. This optimized process would also be applicable to other tissue engineered tissue constructs, such as cardiovascular tissue equivalents.