Collaborative Research: Role of Endothelial Cell Activation in Hypoxia Tolerance of an Elite Diver, the Weddell Seal

该奖项全部或部分由《2021年美国救援计划法案》（公法117-2）资助。第一部分：非技术描述：威德尔海豹是一种标志性的南极物种，也是一名出色的潜水员，可以游到2000英尺深的水下，在水下停留45分钟。然而，对于任何哺乳动物来说，潜水时血液中的低氧浓度和回到水面后的恢复都是需要克服的挑战，这使得它们的潜水能力变得独特，这让科学家们着迷了几十年。该研究项目将评估威德尔海豹的生理机能，以保护该物种免受潜水的影响。这项工作将结合实验室实验，在实验室中，血管中的细胞将暴露在低氧条件下，类似于将在南极洲潜水的海豹中测量的条件。研究人员将测试一个新想法，即在长时间潜水之前进行几次短期潜水，让海豹适应自己。对潜水过程中释放到血液中的化合物的测量，结合对调节它们的基因的实验，将为帮助海豹忍受这些极端条件的生化途径提供线索。该项目允许记录单个海豹潜水，并将这些信息提供给更广泛的科学界，以寻求研究这些海豹，教育研究生和本科生以及博士后研究员，并为中学生制作科学宣传漫画书，以说明项目的科学活动，目标和成果。第二部分：技术描述：威德尔海豹是一种潜水冠军，对低血氧浓度（低氧血症）和血液供应不足（缺血）具有很高的天然耐受性。这个物种特有的过程保护它们的组织免受炎症和氧化应激的影响，在其他哺乳动物组织中也观察到这种生理条件。该项目旨在利用分子、细胞和代谢工具，了解保护海豹免受炎症和氧化应激的过程特征。假设在长时间潜水之前重复的短时间潜水可以预先密封组织并激活保护过程。这项工作的新方面是对内皮细胞的研究，内皮细胞感知氧气和血流的变化，提供了屏气和细胞功能之间的联系。该方法是一项实验室实验，结合了在南极洲麦克默多站外一个冰营2年的实地工作。本研究有三个主要目标：1)使用多种生理、生化和基因组工具，包括CRSPR/Cas9敲除和敲低方法，对暴露于氧气和流量变化的动脉内皮细胞进行实验室实验，以确定负责缺氧和缺血耐受性的分子途径。2)海豹长时间潜水时产生的血液代谢物的代谢组学分析。3)短潜期间海豹生理代谢组学和基因组学测定假设条件前耐受反应。在野外，海豹在一个孤立的冰洞中潜水后，将采集血液样本并记录其潜水表现。预计血液中会含有与实验室实验中确定的分子途径相关的代谢物。专家合作伙伴将提供现场支持，包括冰营、海豹潜水洞，以及与海豹潜水相关的遥测技术。该项目建立在以前nsf资助的海豹基因组和细胞资源生产项目的基础上。本科研究人员将从具有吸引少数族裔和少数族裔服务机构的记录的机构项目中招募。为了进一步增加极地扫盲培训和教育影响，实地小组将包括一个博客，在那里分享实地经验，并与一名为K-12学生和公众宣传设计的艺术家一起编写漫画书。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117-2). Part I: Non-technical description: The Weddell seal is an iconic Antarctic species and a superb diver, swimming down to 2,000 feet and staying underwater for up to 45 minutes. However, as for any mammal, the low oxygen concentrations in the blood during diving and the recovery once back at the surface are challenges that need to be overcome making their diving ability something unique that has fascinated scientists for decades. This research project will evaluate the underlying processes in Weddell seal’s physiology that protects this species from the consequences of diving. The work will combine laboratory experiments where cells that line the blood vessels will be exposed to conditions of low oxygen, similar to those that will be measured in diving seals in Antarctica. The investigarors will test a new idea that several short-term dives, performed before a long dive, allows seals to condition themselves. Measurements on the chemical compounds released to the blood during dives, combined with experiments on the genes that regulate them will provide clues on the biochemical pathways that help the seals tolerate these extreme conditions. The project allows for documentation of individual seal dives and provisioning of such information to the broader science community that seeks to study these seals, educating graduate and undergraduate students and a post-doctoral researcher and producing a science-outreach comic book for middle-school students to illustrate the project's science activities, goals and outcomes. Part II: Technical description: The Weddell seal is a champion diver with high natural tolerance for low blood oxygen concentration (hypoxemia) and inadequate blood supply (ischemia). The processes unique to this species protects their tissues from inflammation and oxidative stress observed in other mammalian tissues exposed to such physiological conditions. This project aims to understand the signatures of the processes that protect seals from inflammation and oxidant stress, using molecular, cellular and metabolic tools. Repetitive short dives before long ones are hypothesized to precondition seal tissues and activate the protective processes. The new aspect of this work is the study of endothelial cells, which sense changes in oxygen and blood flow, providing a link between breath-holding and cellular function. The approach is one of laboratory experiments combined with 2-years of field work in an ice camp off McMurdo Station in Antarctica. The study is structured by three main objectives: 1) laboratory experiments with arterial endothelial cells exposed to changes in oxygen and flow to identify molecular pathways responsible for tolerance of hypoxia and ischemia using several physiological, biochemical and genomic tools including CRSPR/Cas9 knochout and knockdown approaches. 2) Metabolomic analyses of blood metabolites produced by seals during long dives. And 3) Metabolomic and genomic determinations of seal physiology during short dives hypothesized to pre-condition tolerance responses. In the field, blood samples will be taken after seals dive in an isolated ice hole and its diving performance recorded. It is expected that the blood will contain metabolites that can be related to molecular pathways identified in lab experiments. Expert collaborators will provide field support, with the ice camp, dive hole for the seals, and telemetry associated with the seals’ dives. The project builds upon previous NSF-funded projects where the seal genome and cellular resources were produced. Undergraduate researchers will be recruited from institutional programs with a track record of attracting underrepresented minorities and a minority-serving institution. To further increase polar literacy training and educational impacts, the field team will include a blog where field experiences are shared and comic book preparation with an artist designed for K-12 students and public outreach.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.