Sugars as Novel Cryoprotectants for Primate Oocytes

糖作为灵长类卵母细胞的新型冷冻保护剂

• 批准号: 7243324
• 负责人: ALI EROGLU
• 金额: $ 31.1万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7243324
• 关键词: Area; Artemia; Basic Science; Biological Preservation; Birth; Calculi; Cell membrane; Cells; Chemicals; Condition; Coupled; Cryopreservation; Cryoprotective Agents; Culture Media; Cytosol; Derivation procedure; Desiccation; Development; Developmental Biology; Dimethyl Sulfoxide; Embryo; Embryonic Development; Ensure; Ethical Issues; Ethylene Glycols; Excision; Female infertility; Fertilization; Freezing; Glass; Goals; Human; Ice; Infertility; Injury; Intracellular Transport; Legal; Malignant Neoplasms; Mammalian Cell; Medical Technology; Methods; Modeling; Molecular Weight; Mus; Nature; Oocytes; Organism; Outcome; Pathway interactions; Permeability; Preservation Technique; Primates; Procedures; Propylene Glycols; Protocols documentation; Rana; Rate; Reproduction; Research; Research Personnel; Salts; Scheme; Science; Sodium Chloride; Solutions; Somatic Cell; Specific qualifier value; Stress; Testing; Thermodynamics; Toxic effect; Transition Temperature; Trehalose; Water; base; cell type; coping; cryobiology; embryonic stem cell; ethylene glycol; extracellular; improved; molecular size; nonhuman primate; novel; novel strategies; nuclear reprogramming; programs; research study; sugar

DESCRIPTION (provided by applicant): The overall goal of this proposal is to develop a novel preservation technique for primate oocytes. Oocytes are unique totipotent cells that offer exceptional prospects for studying fundamental developmental biology, as successfully explored in many areas including nuclear reprogramming and derivation of embryonic stem cells. Therefore, successful banking of oocytes from different model species would enormously contribute to the advancement of science, medical technologies, and conservation of endangered species. A cryopreservation method developed and thoroughly tested using non-human primate oocytes would not only facilitate basic research and conservation of endangered primates but would also be a major stepping-stone towards a reliable preservation protocol for human oocytes for infertility treatment. To this end, we propose a mechanistic novel approach based on survival schemes in nature where a variety of organisms such as frogs, tardigrades, and brine shrimp can survive freezing or almost complete drying. Accumulation of sugars is extremely critical for their survival strategy. By overcoming the permeability barrier of mammalian cell membranes to the sugars, recent studies have clearly demonstrated improved cryosurvival of somatic cells. Therefore, we hypothesize that when present both inside and outside cells, sugars protect oocytes against freezing-induced injury by their excellent glass formation abilities and direct interactions with subcellular components. We also hypothesize that combinations of sugars with conventional penetrating cryoprotectants and development of a novel sugar-based freezing medium will further improve cryosurvival. We will test these hypotheses systematically using intra- and extracellular sugars with different glass transition temperatures and molecular sizes, and then combining the most effective sugars with conventional cryoprotectants. Next, using a coupled mechanistic model of water transport and intracellular ice formation, we will establish an optimal thermodynamic pathway for oocyte freezing in the presence of sugar/cryoprotectant. Finally, we will test developmental capacity of cryopreserved oocytes to term. Our studies will contribute to understanding fundamental aspects of oocyte cryobiology as well as cryoprotective mechanisms of sugars. We predict that the proposed research will result in a reliable preservation technique for primate oocytes by circumventing many of the difficulties associated with conventional protocols.

描述（由申请人提供）：本提案的总体目标是开发一种新的灵长类动物卵母细胞保存技术。卵母细胞是一种独特的全能细胞，为研究基础发育生物学提供了特殊的前景，在许多领域，包括核重编程和胚胎干细胞的衍生都有成功的探索。因此，不同模式物种卵母细胞的成功建库将极大地促进科学、医疗技术和濒危物种保护的进步。使用非人类灵长类动物卵母细胞开发和彻底测试的冷冻保存方法不仅有利于濒危灵长类动物的基础研究和保护，而且也将是迈向用于不孕症治疗的人类卵母细胞的可靠保存协议的主要垫脚石。为此，我们提出了一种基于自然界生存方案的机械新方法，其中各种生物如青蛙，缓步动物和卤虫可以在冷冻或几乎完全干燥后存活。糖的积累对于它们的生存策略至关重要。通过克服哺乳动物细胞膜对糖的渗透性屏障，最近的研究已经清楚地证明了体细胞的冷冻存活改善。因此，我们假设，当存在于细胞内外时，糖通过其优异的玻璃形成能力和与亚细胞组分的直接相互作用来保护卵母细胞免受冷冻诱导的损伤。我们还假设，糖与传统的渗透性冷冻保护剂的组合和一种新的糖基冷冻介质的开发将进一步提高冷冻存活率。我们将使用具有不同玻璃化转变温度和分子大小的细胞内和细胞外糖系统地测试这些假设，然后将最有效的糖与传统的冷冻保护剂相结合。接下来，使用耦合的水运输和细胞内冰形成的机制模型，我们将建立一个最佳的热力学途径，在糖/冷冻保护剂的存在下，卵母细胞冷冻。最后，我们将测试冷冻保存的卵母细胞的发育能力。我们的研究将有助于了解卵母细胞冷冻生物学的基本方面以及糖的冷冻保护机制。我们预测，拟议中的研究将导致一个可靠的保存技术灵长类动物卵母细胞绕过许多与传统协议的困难。