A Novel Strategy to Study the Invasive Capability of Adherent-Invasive Escherichia coli by Using Human Primary Organoid-Derived Epithelial Monolayers

Abstract

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.

Mimicking the Intestinal Host–Pathogen Interactions in a 3D In Vitro Model: The Role of the Mucus Layer

“…In vitro systems based on intestinal organoids offer complex and elegant models containing all the intestinal cell types that self-organize into crypt and villus domains [ 13 , 50 ]. Although most of these studies have used organoids with a closed lumen as models [ 51 ], in the last years organoid-derived monolayers have been developed easing the access to the apical compartment and have been employed to study, for example, the invasive kinetics of intestinal pathogens [ 52 , 53 , 54 ]. However, these models fail to recapitulate the 3D crypt-villus axis characteristic of the small intestine and the mucus layer is not well represented [ 55 ].…”

Adherent-Invasive and Non-Invasive Escherichia coli Isolates Differ in Their Effects on Caenorhabditis elegans’ Lifespan

“…Hence, integrated and/or more relevant models are required. The use of human intestinal cells derived from a patient’s colonic organoids has deepened our understanding of the physiopathology of IBD, inter-individual variability, and the lesions’ precise location in the gut [ 51 ]. Although the organoid model offers many advantages, it is still time-consuming and costly and requires the patient’s consent for access to biopsies.…”

The World of Organoids: Gastrointestinal Disease Modelling in the Age of 3R and One Health with Specific Relevance to Dogs and Cats

“…Aside from functional characteristics, ODMs have also been developed much further as co-culture models with bacteria. Mayorgas et al used human ODMs as a proxy for the infection with invasive E. coli [ 100 ]. A slightly more complex system has been introduced by Sasaki et al [ 101 ].…”