A Novel Strategy To Study The Invasive Capability Of Adherent-Invasive Escherichia Coli By Using Human Primary Organoid-Derived Epithelial Monolayers

Over the last decades, Adherent-Invasive Escherichia coli (AIEC) has been linked to the pathogenesis of Crohn's Disease. AIEC's characteristics, as well as its interaction with the gut immune system and its role in intestinal epithelial barrier dysfunction, have been extensively studied. Nevertheless, the currently available techniques to investigate the cross-talk between this pathogen and intestinal epithelial cells (IECs) are based on the infection of immortalized cell lines. Despite their many advantages, cell lines cannot reproduce the conditions in tissues, nor do they reflect interindividual variability or gut location-specific traits. In that sense, the use of human primary cultures, either healthy or diseased, offers a system that can overcome all of these limitations. Here, we developed a new infection model by using freshly isolated human IECs. For the first time, we generated and infected monolayer cultures derived from human colonic organoids to study the mechanisms and effects of AIEC adherence and invasion on primary human epithelial cells. To establish the optimal conditions for AIEC invasion studies in human primary organoid-derived epithelial monolayers, we designed an infection-kinetics study to assess the infection dynamics at different time points, as well as with two multiplicities of infection (MOI). Overall, this method provides a model for the study of host response to AIEC infections, as well as for the understanding of the molecular mechanisms involved in adhesion, invasion and intracellular replication. Therefore, it represents a promising tool for elucidating the cross-talk between AIEC and the intestinal epithelium in healthy and diseased tissues.