New Cell Desiccation Strategy Using Non-Isothermal Drying and Biophysical Models

使用非等温干燥和生物物理模型的新细胞干燥策略

基本信息

批准号：
8989096
负责人：
ALI EROGLU
金额：
$ 19.3万
依托单位：
AUGUSTA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-02-01 至 2019-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8989096
关键词：
Address Affect Age Algorithms Area Biological Preservation Biological Sciences Biophysics Cancer Patient Cancer Survivor Cell Membrane Permeability Cell Therapy Cells Chemicals Computer Simulation Containment Cryopreservation Cryoprotective Agents Dehydration Delayed Childbearing Dependence Desiccation Development Diagnosis Diffusion Disease Embryo Emotional Engineering Ethical Issues Exposure to Female Fertility Fertility Agents Fertilization in Vitro Freezing Future Germ Cells Glass Goals Health Human Injury Institutes Legal Life Liquid substance Long-Term Survivors Malignant Neoplasms Mammalian Cell Measurement Measures Medical Methods Mission Modeling Mus National Institute of Biomedical Imaging and Bioengineering Nature Nitrogen Oocytes Organism Outcome Ovarian Hyperstimulation Syndrome Polycystic Ovary Syndrome Polymers Premature Ovarian Failure Preparation Procedures Process Production Property Protocols documentation Quality Control Recording of previous events Recovery Regenerative Medicine Rehydrations Research Research Personnel Residual state Risk Safety Sampling Stem cells Sterility Strategic Planning Stress Techniques Technology Temperature Testing Theoretical model Thermodynamics Time Tissue Engineering Transition Temperature Transplantation Transportation Trehalose Water Woman Work base biobank biophysical model cancer statistics cancer therapy cell injury cell type clinical application cost cytotoxicity design extracellular gonad function improved infertility treatment innovation mathematical model meetings multidisciplinary novel novel strategies programs public health research simulation success sugar young woman

项目摘要

DESCRIPTION: Effective preservation technology for living cells is necessary in biomedical applications ranging from in vitro fertilization to tissue engineering and cell therapies, because the short shelf-life of (non-stabilized) cells imposes severe limitations on biobanking, long-distance transportation, and even safety/quality testing. Current preservation strategies are inadequate, especially for sensitive cell types such as oocytes. For example, cryopreservation requires exposure to high concentrations of penetrating chemicals that are used as cryoprotectants (CPAs) but have inherent cytotoxicity, and requires expensive containment cooled by liquid nitrogen (LN2) for storage and transportation. Moreover, slow-cooling approaches to cryopreservation yield unsatisfactory recovery of healthy oocytes, whereas recent oocyte vitrification approaches require unsafe direct contact with LN2, unreliable sample preparation methods, and much higher CPA concentrations. An alternative preservation strategy, anhydrobiosis (via desiccation), has the potential to stabilize cells at more convenient storage temperatures (e.g., room temperature), but current approaches to desiccation are damaging, because cells are exposed to prolonged osmotic stresses due to diffusion barriers that develop during drying; desiccation of mammalian oocytes is not currently possible. The development of desiccation procedures for human oocytes would allow preservation of fertility for cancer patients, and offer infertility treatment options for other disease states including premature ovarian failure, ovarian hyperstimulation syndrome, and polycystic ovary. The long-term goal of the proposed research program is to meet the critical need of effective, low-cost cell preservation technology by developing a novel, mathematically optimized approach to desiccation. The objective of this developmental (R21) application is to demonstrate feasibility of the proposed method in mouse and human oocytes. This goal will be achieved using a simulation- guided approach, and by taking advantage of unique protective properties of intra- and extracellular sugars. The investigators have previously used this strategy to improve cryopreservation outcome. Informed by preliminary studies indicating that oocytes can be dried to as little as 5% residual moisture content without significant loss of functionality, the central hypothesis is that oocytes can safely be brought into a glassy state at temperatures of -20�C (for storage in a conventional freezer) or 4�C (refrigerator storage) by using a non-isothermal desiccation approach in the presence of intra- and extracellular sugars, sugar polymers, and small amounts (d0.5 M) of a penetrating CPA. The proposed non-isothermal desiccation approach is innovative, because it circumvents the diffusion barrier problem that is inherent to isothermal desiccation, thus allowing faster, less damaging dehydration; other innovations include the development of novel desiccation solutions that use only minimal amounts of CPA, and the use of biophysics-based mathematical models and computer simulations to identify the optimal protocols for non-isothermal desiccation. The proposed research is significant, because its success will (i) shift current paradigms of cell preservation, and (ii) pave the way for meetin biopreservation needs in cell therapy and tissue engineering.

描述：在从体外受精到组织工程和细胞治疗的生物医学应用中，有效的活细胞保存技术是必要的，因为（非稳定的）细胞的保质期短对生物库、长途运输和运输造成了严重限制。甚至安全/质量测试也是不够的，特别是对于卵母细胞等敏感细胞类型，例如，冷冻保存需要暴露于用作冷冻保护剂（CPA）的高浓度渗透性化学物质。但具有固有的细胞毒性，并且需要通过液氮（LN2）冷却来进行昂贵的密封以进行储存和运输。此外，缓慢冷却的冷冻保存方法对健康卵母细胞的恢复效果并不令人满意，而最近的卵母细胞玻璃化冷冻方法需要不安全的直接接触液氮（LN2），不可靠的样品。制备方法和更高的 CPA 浓度另一种保存策略，脱水（通过干燥），有可能在更方便的储存温度下稳定细胞。（例如，室温），但目前的干燥方法具有破坏性，因为由于干燥过程中形成的扩散障碍，细胞会长期暴露在渗透压下；目前不可能开发出用于人类卵母细胞的干燥方法。允许癌症患者保留生育能力，并为其他疾病状态（包括卵巢早衰、卵巢过度刺激综合征和多囊卵巢）提供不孕治疗选择。研究计划是通过开发一种新颖的、经过数学优化的干燥方法来满足有效、低成本的细胞保存技术的关键需求。这一开发（R21）应用的目的是证明其可行性。该目标将通过模拟引导的方法并利用细胞内和细胞外糖的独特保护特性来实现。初步研究表明，卵母细胞可以干燥至残留水分含量低至 5%，而不会显着丧失功能，这是核心假设是，在存在内部和外部环境的情况下，通过使用非等温干燥方法，可以在 -20°C（用于传统冰箱中储存）或 4°C（冰箱储存）的温度下安全地将卵母细胞带入玻璃态。所提出的非等温干燥方法是创新的，因为它规避了等温固有的扩散屏障问题。干燥，从而实现更快、更小的脱水；其他创新包括开发仅使用最少量 CPA 的新型干燥解决方案，以及使用基于生物物理学的数学模型和计算机模拟来确定非等温干燥的最佳方案。拟议的研究意义重大，因为它的成功将（i）改变当前的细胞保存范式，并且（ii）为满足细胞治疗和组织工程中的生物保存需求铺平道路。