OOCYTE CRYOPRESERVATION IN RARE BIOMEDICAL MODELS

描述（由申请人提供）：候选人将接受培训，同时产生有关猫科动物卵母细胞冷冻生物学的学术知识。由于许多罕见的猫基因型繁殖不良，配子冷冻保存结合体外胚胎生产和受体雌性移植可以帮助繁殖和管理这些种群。目标是：1）研究卵母细胞的膜生物物理特性; 2）表征和减轻由两步冷冻引起的冷冻损伤;以及3）将新开发的卵母细胞冷冻保存方案应用于稀有猫科动物生物医学模型以辅助群体管理。由于这项技术在从垂死动物身上“拯救”配子方面的潜力，重点将放在未成熟的卵母细胞上。将表征冷冻保护剂暴露、直接冷却和保持在零度以下温度，然后直接浸入液氮中的影响。评价目标包括生物物理膜特性，细胞内冰的形成，细胞骨架的完整性，细胞质膜和与卵丘细胞的相互作用，并最终在体外和体内的发育能力。研究结果将应用于猫科动物模型，同时利用独特的访问选定的野生猫科动物物种，以更快地了解机制。这种基本和比较的方法提供了一个多学科的培训机会，使候选人能够作为一名生理学家，在生物物理学，冷冻生物学和卵母细胞体外培养方面具有专业知识。研究的好处将是对未成熟猫卵母细胞的生物物理特性以及调节其在冷却-冷冻-解冻中存活并发育成可行胚胎和后代的能力的因素的深入了解。实际的好处将是另一个工具，解决始终困难的任务，管理小种群的繁殖受损的猫科动物。与公共卫生的相关性：动物模型对于研究与通过生物医学研究理解和改善人类状况有关的机制至关重要。这种多学科的方法将提供新的卵母细胞冷冻保存协议有用的猫殖民地作为人类疾病的生物医学模型的管理。

DESCRIPTION (provided by applicant): The Candidate will receive training while producing scholarly knowledge about felid oocyte cryobiology. Since many rare cat genotypes reproduce poorly, gamete cryopreservation combined with in vitro embryo production and transfer in recipient females can help to reproduce and manage these populations. Objectives are to: 1) investigate membrane biophysical properties of the oocyte; 2) characterize and mitigate cryoinjuries caused by 2-step freezing; and 3) apply a newly developed oocyte cryopreservation protocol to rare felid biomedical models to assist in population management. Because of the potential of this technology for 'rescuing' gametes from dying animals, the focus will be on the immature oocyte. Impact of cryoprotectant exposure, direct cooling and holding at a subzero temperature followed by direct plunging into liquid nitrogen will be characterized. Evaluation targets include biophysical membrane properties, intracellular ice formation, integrity of the cytoskeleton, cytoplasmic membrane and interactions with cumulus cells, and eventually developmental competence in vitro and in vivo. Findings will be applied to felid models while taking advantage of unique access to selected wild felid species to more rapidly understand mechanisms. This fundamental and comparative approach provides a multidisciplinary training opportunity that will allow the Candidate to advance as a physiologist with expertise in biophysics, cryobiology and in vitro culture of oocytes. The research benefit will be a thorough understanding of the biophysical properties of the immature cat oocyte and the factors regulating its ability to survive cooling-freezing-thawing and to develop into a viable embryo and offspring. The practical benefit will be another tool for addressing the always-difficult task of managing small populations of reproductively compromised felids. Relevance to public health: Animal models are essential to studying mechanisms pertinent to understanding and improving the human condition through biomedical research. This multidisciplinary approach will provide new oocyte cryopreservation protocols useful to the management of cat colonies used as biomedical models for human diseases.