Modeling Shigella Interaction with Innate Cells in Enteroid Co-Cultures to Inform Vaccine Development

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Project 2: Modeling Shigella Interaction with Innate Cells in Enteroid Co-Cultures to Inform Vaccine Development Shigella is responsible for a significant burden of disease in multiple populations within the US and worldwide causing an estimated 163 million cases and >74,000 deaths per year. The greatest impact is in children under 5 years of age in developing countries where Shigella was identified as the most important pathogen causing diarrhea in 12-59 month old children. The widespread isolation of multiple drug resistant isolates limiting therapeutic interventions and the continued high levels of endemic disease underscore the significance of Shigella as a global health priority and reinforce the need for preventative interventions and vaccine development. Infection with Shigella results in diarrhea and dysentery following transcytosis of the epithelial barrier, invasion of gastrointestinal cells, intracellular replication and induction of a severe inflammatory response. Interaction with host innate immune cells including macrophages (MΦ), dendritic cells (DCs) and intraepithelial lymphocytes (IEL) are critical events in progression of disease symptoms as well as in initiation of a protective immune response. The human enteroid model provides a highly human relevant multicellular system that recapitulates important aspects of gastrointestinal physiology. Advances during the first P01 funding cycle included the addition of M cells and immune cell co-culture in enteroids. This complex model provides a system in which critical features of Shigella uptake, transcytosis, and engagement with immune cells can be identified which may serve as targets for interventional strategies. Using a series of isogenic Shigella strains with mutations in key virulence factors, we aim to define bacterial requirements for each stage of host engagement. Furthermore, the evaluation of a series of live attenuated vaccine strains that have been studied clinically will serve as tools to further understand bacterial requirements for the pathogenic process and allow determination of the utility of this model to distinguish promising vaccine candidates. Building on our current capability of M cell incorporation, we will use the M cell enteroid model to characterize sequential stages of Shigella-host interactions including the process of uptake and transcytosis, and subsequent interaction with critical innate immune cells. These studies will reveal novel aspects of host-pathogen interactions and ensuing innate immune responses to this pathogen that better reflect what occurs in humans; new insights will be provided that will guide intervention strategies.