Stanford Cooperative Research Center for Novel, Alternative Model Systems for Enteric Diseases

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 DESCRIPTION OF THE OVERALL U19 APPLICATION (provided by applicant): The human gastrointestinal tract exhibits diverse and essential roles in processes as widespread as digestion, absorption, secretion and immunity. However, these same crucial functions represent a significant vulnerability by which bacterial and viral gastrointestinal pathogens induce substantial morbidity and mortality in humans worldwide. While animal models are invaluable to the study of basic mechanisms of infectious disease, the interactions between some of the most prominent gastrointestinal pathogens with host tissues are species specific and host restricted, particularly for infections that have co-evolved with humans. Thus, the study of many pathogens has heretofore required animal-adapted strains that do not accurately recapitulate the human disease pathophysiology and host range restriction in mice, or alternatively oncogenically transformed human cell lines that do not model normal tissue characteristics. Recent advances in culture of primary tissues as 3-dimensional "organoids" that reproduce multilineage differentiation and organ architecture represent a promising technology to modeling the interaction of human enteric pathogens with human gastrointestinal epithelium in vitro. The overall goal of this application is to create alternative model systems for a variety of human enteric diseases and pathogen-associated specific immune responses using optimized 3D human gastrointestinal organoid cultures via the co-culture of (1) primary gastric and intestinal organoids with (2) bacterial/viral pathogens and/or (3) immune effector cells. We have thus assembled a multidisciplinary, collaborative, and synergistic team at Stanford University including Calvin Kuo (gastrointestinal organoid culture), Manuel Amieva (Helicobacter pylori), Harry Greenberg (Rotavirus), Sarah Heilshorn (microenvironment bioengineering), Sean Bendall (CyTOF), Elizabeth Mellins (immunology), and Denise Monack (Salmonella Typhi). Together, these PIs form the Stanford Novel, Alternative Models for Enteric Diseases Cooperative Research Center (Stanford NAMSED CRC) Program. The Stanford NAMSED CRC is composed of Cores A-C (Administration, Organoid Production, and Advanced Co-Culture Engineering and Single Cell Statistics of Gut Immunology "ACCESS-GI"), and Projects 1-2, which study bacterial and viral pathogen interactions with human GI organoids, respectively. Towards these goals, we employ human organoid technologies incorporating epithelial-only or epithelial/mesenchymal components to create tissue-specific models of bacterial (Helicobacter pylori, Salmonella Typhi, Salmonella Typhimurium) or viral (rotavirus) infections, with or without immune effector cells or stimuli. We deploy innovative supporting technologies including CyTOF mass cytometry, bioengineered extracellular matrices, microfluidics and CRISPR-based gene editing. Overall, the Stanford NAMSED represents a synergistic group devoted to the integrated modeling of enteric pathogens as an epithelial-immune organoid unit.