Wohlwend High Pressure Freezer for the BIDMC Electron Microscopy Core

适用于 BIDMC 电子显微镜核心的 Wohlwend 高压冷冻机

• 批准号: 8825681
• 负责人: Susan J Hagen
• 金额: $ 19.94万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8825681
• 关键词: Advisory Committees; Biomedical Research; Brown Fat; Budgets; Cell Nucleus; Cells; Conflict (Psychology); Cytoplasm; Data; Diabetes Mellitus; Digestive System Disorders; Electron Microscopy; Electron Microscopy Facility; Employee Strikes; Energy Metabolism; Ensure; Exercise; Freeze Substitution; Freezing; Functional disorder; Funding; Goals; Grant; Health; Individual; Inflammation; Inflammatory Bowel Diseases; Intestines; Israel; Liver; Medical center; Metabolic; Methods; Microtomy; Minor; Mitochondria; Mitochondrial Matrix; Mitochondrial Swelling; Modeling; Morphologic artifacts; Morphology; Muscle Mitochondria; Obesity; Paneth Cells; Pathogenesis; Performance; Plastics; Postdoctoral Fellow; Process; Proteins; Recruitment Activity; Request for Applications; Research; Research Personnel; Role; Services; Skeletal Muscle; Stomach; Stress; Techniques; Thick; Tight Junctions; Time; Tissues; Training; United States National Institutes of Health; flexibility; improved; instrument; instrumentation; malignant stomach neoplasm; pressure; ranpirnase; repaired; response; success; tissue processing

 DESCRIPTION (provided by applicant): The Major and Minor Users for this application request funds to purchase a High Pressure Freezing (HPF) machine to carry-out ultrastructural studies for NIH-funded projects. The broad, long-term objective of this shared instrument is to improve the quality of tissue processing for electron microscopy (EM), in particular to improve the ultrastructure of mitochondria, ER, and other cellular components in brown adipose tissue (BAT), skeletal muscle, intestine, liver, and stomach. The requested instrument is the Wohlwend HPF, which was chosen for its flexibility, ease of use, excellent repair and service record, and outstanding performance. The machine will be placed into an existing EM facility, which is partially supported by the Beth Israel Deaconess Medical Center and partially supported by an active NIH P30 Center Grant for digestive diseases. The Major User group consists of 5 investigators with NIH-funded grants, one Minor User, who has an active biomedical research pro- gram, and dedicated time for outside users. The individual research goal for each user is different, but each requires improved tissue ultrastructure. Dr. Arany studies changes in skeletal muscle mitochondria in different models of metabolic pathophysiology. Drs. Cohen, Goodyear, and Tseng study the pathological significance of mitochondrial changes in BAT and/or liver, which relate to energy metabolism in obesity, exercise, or diabetes. Dr. Blumberg studies the role of intestinal Paneth cells in the pathogenesis of Inflammatory Bowel Disease, in particular the inflammation-induced activation of ER stress and the unfolded protein response. Lastly, Dr. Ha- gen studies the pathogenic role of tight junction abnormalities in stomach cancer pathogenesis. Preliminary data, collected with the assistance of an expert in the field who was also recruited as a consultant for this application, clearly demonstrate that HPF followed by super-quick freeze substitution methods resulted in significantly improved tissue morphology compared to conventional processing techniques. The overall benefit to the health-related goals of each project will be that HPF/SQFS limited cell shrinkage, mitochondrial swelling and disruption, and the extraction of important proteins in the mitochondrial matrix, lumen of the ER, nuclei, and cytoplasm. With the capability of the requested HPF machine to freeze thicker and larger tissue pieces, the user groups were able to prepare numerous thick plastic sections for analysis followed by numerous thin sections for EM analysis that were in the proper orientation and free of freezing damage. The differences in tissue ultrastructure using HPF/SQFS compared to conventional processing were so striking that the User group is reluctant to continue their studies with significant artifacts that cannot be fixed without the benefit of this machine. Administratively, the requested HPF will be managed by the EM Core Director and Research Associate, who will do the training, upkeep, and oversight of the instrument. There will be a local advisory committee to handle conflicts and new user applications. Lastly, there is strong institutional support for this instrumentation, with an appropriate budget, space, and financial commitment to ensure success.

 描述（由申请人提供）： 本申请的主要和次要用户请求资金购买高压冷冻 (HPF) 机器，以便为 NIH 资助的项目进行超微结构研究 该共享仪器的广泛、长期目标。旨在提高电子显微镜（EM）的组织处理质量，特别是改善棕色脂肪组织（BAT）、骨骼肌、所需的仪器是 Wohlwend HPF，因其灵活性、易用性、出色的维修和服务记录以及出色的性能而被选中。该机器将被放置在现有的 EM 设施中，该设施部分已建成。得到 Beth Israel Deaconess 医疗中心的支持，部分得到 NIH P30 消化系统疾病中心拨款的支持。主要用户组由 5 名获得 NIH 资助的研究人员组成，一名次要用户拥有积极的生物医学研究项目，每个用户的个人研究目标不同，但每个人都需要改善组织超微结构。Arany 博士、Cohen 博士和 Tseng 博士研究了不同的代谢病理生理学模型。 Blumberg 博士研究了 BAT 和/或肝脏中线粒体变化的病理意义，这些变化与肥胖、运动或糖尿病中的能量代谢有关，特别是肠道潘氏细胞在炎症性肠病发病机制中的作用。最后，哈根博士研究了紧密连接异常在胃癌发病机制中的致病作用，这些数据是在该领域专家的帮助下收集的。该应用的顾问清楚地证明，与传统处理技术相比，HPF 后采用超快速冷冻替代方法可显着改善组织形态，每个项目对健康相关目标的总体好处将是 HPF/SQFS 限制了细胞。收缩、线粒体肿胀和破坏，以及提取线粒体基质、内质网腔、细胞核和细胞质中的重要蛋白质。凭借所需的 HPF 机器冷冻更厚和更大的组织块的能力，用户组能够准备大量厚的塑料切片进行分析，然后准备大量薄切片进行电镜分析，这些切片的方向正确且无冷冻损伤。与传统处理相比，使用 HPF/SQFS 的组织超微结构差异非常显着，以至于用户组不愿意这样做。继续学习在管理上，如果没有这台机器就无法修复的重要工件，所请求的 HPF 将由 EM 核心主任和研究助理管理，他们将负责仪器的培训、维护和监督。最后，该工具有强有力的机构支持，并有适当的预算、空间和财务承诺，以确保成功。