RESEARCH STRATEGY: The primary research objective of this proposal is to address how subtypes of enteroendocrine cells change their activity (secretion) or state of differentiation in response to injury signals. A better understanding of intestinal injury repair is critical to VA health priorities; the prevalence of inflammatory bowel disease (IBD) has more than doubled among VA patients since 1998 and at least half of patients won?t experience full mucosal healing with existing therapies. Hormones produced in intestinal enteroendocrine cells (EEC) are altered in patients with IBD and can promote epithelial healing, but there is little understanding of how EEC secretion is regulated during injury. The study of EECs has been limited because they are rare, heterogenous, and not amenable to long-term primary culture with existing methods. To address this gap, we have developed a new experimental system to culture EECs from primary colon samples in long-term monolayers that supports all known EEC subtypes. We have discovered that subtypes of EEC are dynamically altered with a simple experimental injury in vitro. The altered EECs increase the expression of factors that promote intestinal healing. The goal of this proposal is to determine the cellular and transcriptional basis of how subtypes of EEC respond to intestinal injury through altered activity or differentiation.
Our Aims are to (1) test the hypothesis that subtypes of EECs are adaptively reprogrammed by injury induced endoplasmic reticulum (ER) stress, (2) test the hypothesis that microenvironmental injury signals reprogram EECs to increase hormone secretion, and (3) define the transcriptional mechanisms of EEC reprogramming during injury at single-cell resolution. The expected outcome of this work is to establish the new paradigm that EECs undergo adaptive reprogramming during injury which will serve as the foundation for future independent grant proposals. Additionally, we expect to discover novel secreted factors or altered states of differentiation in EEC that would lead to new potential treatments for veterans suffering from intestinal or metabolic disorders. CANDIDATE/ENVIRONMENT: Dr. Brian Muegge is a senior fellow in the Division of Endocrinology at Washington University in St. Louis and the St. Louis VA Medical Center. He completed his M.D., Ph.D. training at Washington University where he performed graduate studies in Dr. Jeffrey Gordon?s laboratory. Dr. Muegge completed his residency in internal medicine at UCSF. He is conducting his fellowship research in the lab of his primary mentor Dr. Thaddeus Stappenbeck and co-mentor Dr. Carlos Bernal-Mizrachi at Washington University. Dr. Muegge has gained experience in intestinal stem cell culture and computational analysis of transcriptional data. He has developed a novel culture system that allows him to model EEC development and injury using primary cells. He now seeks to expand his expertise in new areas including lineage tracing, cell- cell signaling, and single cell RNA sequencing prior to starting his independent research career. CAREER DEVELOPMENT: This award will ensure that Dr. Muegge is able to establish a career as an independent physician scientist in the VA health care system. He has developed a formal 5-year training plan that will expand his scientific skills in intestinal development, in vitro models, and single cell transcriptomics, as well as academic skills in scientific publishing, grant writing, and laboratory management. Dr. Muegge?s training will be accomplished with experimental research, didactics, seminars, and national conferences. He has assembled a scientific advisory committee with the requisite experience and expertise to monitor and support his progress towards achieving his career goals. At the completion of the CDA-2, Dr. Muegge will be equipped with skills in cutting edge technology (single cell transcriptomics, in vitro stem cell culture systems) working in a unique area that sets him apart from others in the field (intestinal endocrine function during bowel injury). He will be uniquely prepared to address fundamental questions that require knowledge of intestinal biology, endocrinology, and computational biology.
Inflammatory Bowel Disease is a growing problem among veterans and new approaches to repair injured epithelium are needed. We have demonstrated that enteroendocrine cells, a rare and overlooked lineage in the intestinal epithelium, are dynamically altered during injury to promote the secretion of wound repairing hormones. We have developed a novel long-term culture system of primary intestinal enteroendocrine cells that recreates key features of normal development and in vivo injury. We propose to use this system, along with animal models and cutting-edge techniques of developmental biology and computational genomics, to dissect the mechanisms by which enteroendocrine cells are dynamically regulated during injury and identify secreted factors that promote wound healing. Successful completion of the proposed aims and training plan will prepare the candidate to become an independent physician scientist identifying translational therapies derived from enteroendocrine cells for veterans suffering from intestinal and metabolic disease.