PROJECT SUMMARY/ABSTRACT This proposal comprises a three-year research and career development program for Rachel Niec, MD, PhD to achieve independence as an investigator at the intersection of immunology and intestinal epithelial biology. Dr. Niec completed her doctoral training in immunology at Sloan Kettering Institute and Internal Medicine Residency and Gastroenterology Fellowship at Weill Cornell Medical College. The research and career development activities will occur at Rockefeller University. Dr. Niec will engage in career development activities including didactics, workshops in grant writing and lab management, acquisition of technical skills and scientific expertise, conference presentations, and a comprehensive mentorship program through her dedicated mentoring team. Dr. Niec has a strong background in cell biology and, over the course of this K08 award at the Rockefeller University, aims to expand her skills in advanced microscopy, transcriptomics and stem cell (SC) biology to study how features of the intestinal SC (ISC) niche interpret diverse cues to direct SC activity in health and in disease. Inflammatory bowel diseases (IBD) are inflammatory conditions focused within the intestinal epithelium and driven by genetic, environmental, and microbial factors. Intestinal epithelial maintenance is dependent on ISC which in turn rely on their niche for local signals to direct their activity. While many niche factors emanate from local sources, intestinal epithelial tissues is subject to systemic changes suggesting vascular features may regulate ISC activity. While lymphatics abnormalities have long been appreciated as a feature of IBD, the vascular features of the intestinal niche that control ISC behavior are unknown. The central hypothesis is vasculature is a key regulator of ISC function, through direct signaling between vasculature, ISC and other niche cells. Two specific aims are proposed: (1) Define the contribution of lymphatic capillaries to the cellular network comprising the intestinal stem cell niche; (2) Determine the impact of lymphatic derived Reln and lymphatic:stem cell interactions on ISC maintenance and function. These aims will be addressed using tissue clearing and advanced 3D imaging, established and newly developed single cell and spatial resolution transcriptomics, and novel in vitro coculture and in vivo molecular genetic approaches. The significance of this proposal lies in its relevance to fundamental mechanism of epithelial maintenance and regeneration and to pathogenesis of IBD. The proposal is innovative in the combination of advanced methods in imaging and transcriptomics and their application to SC niche biology and intestinal inflammation. Long-term, Dr. Niec aims to apply the expertise gained to identify environmental and inflammatory signals that maintain and regulate the intestinal SC niche in homeostasis and disease to improve prevention and treatment of IBD.