EAPSI: Ultra-Rapid Freezing to Maintain Stem Cell Developmental Potential in Rainbow Trout

EAPSI：超快速冷冻可维持虹鳟鱼的干细胞发育潜力

• 批准号: 1515297
• 负责人: Tawny Scanlan
• 金额: $ 0.51万
• 依托单位: Scanlan Tawny N
• 依托单位国家: 美国
• 项目类别: Fellowship Award
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1515297&HistoricalAwards=false
• 关键词: EAPSI; Ultra; Rapid; Freezing; Maintain

The establishment of novel technologies for the long-term storage of fish genetic resources has become critical as aquatic species around the world are experiencing rapid declines due to overexploitation and global climate change. This project will examine an innovative area of reproductive stem cell biology to investigate optimal conditions for vitrification of spermatogonial stem cells from Rainbow trout (Oncorhynchus mykiss). Vitrification involves ultra-rapid cooling that results in instant solidification of the cell without crystallization. Development of a vitrification protocol is desirable since vitrification is fast and the process avoids trauma to cells by eliminating intracellular ice formation and instead creates a glass-like (vitreous) state, resulting in higher cell survival. This study will be conducted at the Tokyo University of Marine Sciences and Technology in Tokyo, Japan in collaboration with and under the supervision of Dr. Goro Yoshizaki, a renowned expert in fish reproductive stem cell biology. The proposed study will provide insight into fundamental aspects of stem cell cryobiology and could lead to real-world application of an inexpensive, economical method for preserving fish genetic resources for the conservation of endangered fish and aquaculture purposes.The proposed experiments for this project will use Gfp-vasa+ transgenic rainbow trout to 1) determine the effects of the vitrification process on spermatogonial stem cell survival and 2) investigate the functionality of vitrified spermatogonial stem cells. Stem cell survival will be quantified by using flow cytometry, which will separate cell populations into viable or non-viable based on fluorescence patterns. A germ cell transplantation assay will be utilized to determine if the viable vitrified stem cells retain their functionality potential to colonize and proliferate in recipients. This research will improve understanding of the complex effects of cryopreservation and vitrification on stem cell function while providing the information necessary to successfully implement stem cell research as a novel artificial reproductive technology method to maximize genetic diversity for the conservation of threatened aquatic species. This NSF EAPSI award is funded in collaboration with the Japan Society for the Promotion of Science (JSPS).

由于过度开发和全球气候变化，世界各地的水生物种正在迅速减少，因此建立长期储存鱼类遗传资源的新技术变得至关重要。本项目将研究生殖干细胞生物学的一个创新领域，研究虹鳟鱼精原干细胞玻璃化冷冻的最佳条件。玻璃化涉及超快速冷却，导致细胞瞬间固化而不结晶。玻璃化方案的开发是期望的，因为玻璃化是快速的，并且该过程通过消除细胞内冰形成而避免了对细胞的创伤，而是产生玻璃样（玻璃状）状态，从而导致更高的细胞存活。这项研究将在日本东京的东京海洋科技大学进行，并在鱼类生殖干细胞生物学著名专家Goro Yoshizaki博士的合作和监督下进行。这项拟议中的研究将为干细胞低温生物学的基本方面提供深入了解，并可能导致廉价，为保护濒危鱼类和水产养殖目的而保存鱼类遗传资源的经济方法。本项目的拟议实验将使用Gfp-vasa+转基因虹鳟鱼1）确定玻璃化过程对精原干细胞存活的影响，2）研究玻璃化冷冻精原干细胞的功能。将使用流式细胞术定量干细胞存活率，流式细胞术将基于荧光模式将细胞群分离为存活或非存活。将使用生殖细胞移植试验来确定活的玻璃化冷冻干细胞是否保留其在受体中定殖和增殖的功能潜力。这项研究将提高对冷冻保存和玻璃化对干细胞功能的复杂影响的理解，同时提供成功实施干细胞研究所需的信息，作为一种新的人工生殖技术方法，以最大限度地提高遗传多样性，保护受威胁的水生物种。 NSF EAPSI奖由日本科学促进会（JSPS）资助。