Antioxidant regulation of intestinal homeostasis and disease

项目摘要 这项建议详细说明了一项为期5年的培训计划，以帮助Sarah Short博士继续发展成为一名 独立的胃肠道研究人员。这项研究计划将重点阐明谷胱甘肽过氧化物酶1的作用。 在炎症性肠病(IBD)中普遍表达的硒酶和有效的抗氧化剂(GPX1) 和结肠炎相关性发育不良(CAD)。使用GPX1-/-MICE的令人信服的初步数据表明，与 许多抗氧化剂的缺失会加剧小鼠结肠炎，GPX1的缺失可以显著保护小鼠免受葡聚糖的伤害。 硫酸钠(DSS)诱导的结肠炎。GPX1缺乏也增加了3D有机化合物的存活率和茎 并改变组织培养细胞的新陈代谢，这可能额外促进再生和伤口愈合。 GPX1的表达也改变了免疫细胞的功能，就像GPX1-/-骨髓来源的巨噬细胞 对“M2”刺激反应增强，迁移能力下降。综上所述，这些结果表明GPX1 通过改变上皮细胞和免疫细胞功能来增加炎性损伤。 基于这些发现，这项建议的假设是GPX1在炎症性肠病中是有害的 通过改变干细胞功能、氧化还原动态平衡和免疫反应而导致的疾病。此外，抑制GPX1 活动可能是一种有效的治疗策略。这一假设将在两个具体目标上进行检验，以确定 GPX1如何促进肠上皮细胞动态平衡、氧化应激、结肠炎和结肠炎相关性 发育不良。第一个目标是研究上皮功能，利用肖特博士10多年的经验 在上皮细胞生物学和屏障功能方面。第二个目标将通过以下方式补充基于上皮的研究 确定GPX1缺失如何改变免疫细胞的招募、分化和功能，并确定它们是如何 变化会改变肠道损伤反应。除了是合乎逻辑的“下一步”实验 GPX1功能，这些实验为Short博士在各个方面的进一步发展提供了完美的框架 对导致肠道疾病的粘膜免疫学进行研究，并将包括新的流式细胞术培训， 趋化因子分析、骨髓移植和T细胞转移性结肠炎模型。肖特博士的职业生涯 通过与主要导师Christopher Williams博士的定期讨论，将进一步促进发展，以及 她的指导委员会由基思·威尔逊博士、杰里米·戈特尔博士和肖恩·戴维斯组成。所有研究和 培训将在Vanderbilt以及该机构、医学系和胃肠病科进行 都高度支持肖特博士的学术生涯，并全力支持她的申请。 肖特博士的最终目标是成为一名独立的学术研究员，专注于 调节IBD和结肠炎相关癌症的发展和严重程度，可以改善治疗 这些患者的选择。有趣的是，本申请中提出的两个具体目标都将评估GPX1作为 使用FDA批准且耐受性良好的硫普罗宁作为治疗靶点。总而言之，这些培训 经验将确保肖特博士准备好领导一项全面的IBD独立研究计划。

Project Summary This proposal details a 5-year training plan to aid the continued develop of Dr. Sarah Short, Ph.D. into an independent GI researcher. This research plan will focus on elucidating the role of glutathione peroxidase 1 (GPx1), a ubiquitously expressed selenoenzyme and potent antioxidant, in inflammatory bowel disease (IBD) and colitis-associated dysplasia (CAD). Compelling preliminary data using Gpx1-/- mice indicates that unlike many antioxidants whose loss exacerbates murine colitis, loss of GPx1 confers striking protection from dextran- sodium sulfate (DSS)-induced colitis. GPx1 deficiency also increases survival and stemness in 3D organoids and alters metabolism in tissue culture cells, which may additionally promote regeneration and wound healing. GPx1 expression also modifies immune cell function, as Gpx1-/- bone marrow-derived macrophages have heightened response to “M2” stimuli and decreased migratory ability. Together, these results suggest that GPx1 augments inflammatory injury through alterations in both epithelial and immune cell function. Based on these finding, the hypothesis of this proposal is that GPx1 is detrimental in inflammatory bowel disease by altering stem cell function, redox homeostasis, and immune responses. Further, inhibiting GPx1 activity may be an effective therapeutic strategy. This hypothesis will be tested in two specific aims to determine how GPx1 contributes to intestinal epithelial cell homeostasis, oxidative stress, colitis, and colitis-associated dysplasia. The first aim will investigate epithelial function, capitalizing on Dr. Short’s over 10 years of experience in epithelial cell biology and barrier function. The second aim will complement epithelial-based studies by determining how GPx1 loss alters immune cell recruitment, differentiation, and function, and identify how these changes modify intestinal injury responses. In addition to being the logical “next step” experiments in defining GPx1 function, these experiments provide the perfect framework to further Dr. Short’s development in aspects of mucosal immunology which contribute to intestinal diseases, and will include new training in flow cytometry, chemokine analysis, bone marrow transplantation, and the T-cell transfer colitis model. Dr. Short’s career development will be further enhanced by regular discussions with primary mentor, Dr. Christopher Williams, and her mentoring committee consisting of Drs. Keith Wilson, Jeremy Goettel, and Sean Davies. All studies and training will take place at Vanderbilt, and the institution, Department of Medicine, and Gastroenterology Division are highly supportive of Dr. Short’s academic career and fully support her application. Dr. Short’s ultimate goal is to become an independent academic researcher focusing on mechanisms which regulate development and severity of IBD and colitis-associated cancer that can lead to improved therapeutic options for these patients. Interestingly, both specific aims proposed in this application will evaluate GPx1 as a therapeutic target using tiopronin, which is FDA-approved and well-tolerated. Together, these training experiences will ensure Dr. Short is poised to direct a well-rounded independent research program in IBD.