The development of human enteroids as a model to study the human intestine offers tremendous opportunities to study the pathogenesis of infectious enteric disease. We propose to combine the substantial expertise from three institutions with long and impressive histories in the investigation of enteric diseases to exploit this powerful model to study the pathogenesis of four major etiologic agents of diarrheal disease: three pathotypes of diarrheagenic E.coli (enteroaggregative, enterohemorrhagic and enterotoxigenic E. coli) and Shigella. Investigators from the University of Maryland, Johns Hopkins University and the University of Virginia will collaborate in this Program Project Grant (PPG). The leadership team of this PPG comprises three internationally renowned experts in enteric diseases, with complementary training in microbiology, gastroenterology, molecular physiology, and pediatric infectious diseases. They will direct a multidisciplinary team of co-investigators with expertise in cell biology, molecular pathogenesis, and mucosal immunology. The overall goal is to increase understanding of the pathophysiology and potential treatments of these four important pathogens. The proposed studies will use normal human mini-intestines, called enteroids or colonoids, grown on monolayers to develop models that mimic human disease. Examination of pathophysiologic aspects common to the diseases studied will serve to integrate the projects. These aspects include the role of mucins, bacterial proteases called SPATES, enterotoxins and secreted cytokines and are in addition to pathophysiologic aspects specific to each infection. In addition, the contribution of cells involved in innate immunity will be examined by co-culture of the enteroids/colonoids with human macrophages, neutrophils and dendritic cells. These studies using human mini-intestines offer the possibility of revealing insights in disease pathophysiology that are specific to normal human intestine rather than the animal models and cancer cell line models used until now. Each of the four projects focuses on a specific pathogen: enteroaggregative E. coli, Shigella, enterohemorrhagic E. coli, entertoxigenic E. coli. Besides an Administrative Core, there is an Enteroid Core that provides human enteroids/colonoids and growth media, instructs all projects on how to produce enteroid/colonoid monolayers and an Immunology Core that measures cytokines and chemokines, and isolates human macrophages, neutrophils and dendritic cells. The Enteroid and Immunology Cores work together to develop co-culture systems of innate immune cells and enteroids/colonoids that will be used by the projects. The investigators will regularly interact by monthly joint laboratory meetings. The proposed project will yield many significant new insights into enteric disease caused by these important pathogens.
Overall Program Project Narrative This project will advance understanding of diarrheal diseases caused by four important enteric pathogens, including three types of diarrheagenic E.coli and Shigella. A recently developed model of the human intestine called enteroids allows the formation of human mini-intestines that can mimic important aspects of disease due to these organisms. Three universities with long histories of contributions in studying diarrheal diseases will collaborate in utilizing this novel model to study basic mechanisms of disease, characterize attenuated vaccine strains, and attempt to identify new drug targets that could lead to development of novel therapeutic agents.
