Enterohemorrhagic Escherichia coli Reduces Mucus and Intermicrovillar Bridges in Human Stem Cell-Derived Colonoids

Abstract

BACKGROUND AND AIMS Enterohemorrhagic E. coli (EHEC) causes over 70,000 episodes of foodborne diarrhea annually in the USA. The early sequence of events which precede life-threatening hemorrhagic colitis and hemolytic uremic syndrome are not fully understood due to the initial asymptomatic phase of the disease and the lack of a suitable animal model. The aim of this study was to determine the initial molecular events in the interaction between EHEC and human colonic epithelium. METHODS Human colonoids derived from adult proximal colonic stem cells were developed into monolayers to study EHEC-epithelial interactions. Monolayer confluency and differentiation were monitored by transepithelial electrical resistance (TER) measurements. The monolayers were apically infected with EHEC and the progression of epithelial damage over time was assessed using biochemical and imaging approaches. RESULTS Human colonoid cultures recapitulate the differential protein expression patterns characteristic of the crypt and surface colonocytes. Mucus-producing differentiated colonoid monolayers are preferentially colonized by EHEC. Upon colonization, EHEC forms characteristic attaching and effacing lesions on the apical surface of colonoid monolayers. Mucin 2, a main component of colonic mucus, and protocadherin 24 (PCDH24), a microvillar resident protein, are targeted by EHEC at early stages of infection. The EHEC secreted serine protease, EspP, initiates brush border damage through PCDH24 reduction. CONCLUSIONS Human colonoid monolayers are a relevant pathophysiological model which allows the study of early molecular events during enteric infections. Colonoid monolayers provide access to both apical and basolateral surfaces, thus providing an advantage over 3D cultures to study host-pathogen interactions in a controllable and tractable manner. EHEC reduces colonic mucus and affects the brush border cytoskeleton in the absence of commensal bacteria.

Human Intestinal Enteroids: a New Model To Study Human Rotavirus Infection, Host Restriction, and Pathophysiology

“…The development of nontransformed human intestinal epithelial cell cultures that model intestinal biology (2) has stimulated interest in new studies of host-pathogen interactions, with most recently reported studies focusing on bacterial infections (1,7,(49)(50)(51)64). Here, we demonstrate the utility of such novel ex vivo multicellular, physiologically active cultures to study a human enteric virus (HRV) to gain new insight into interactions with human small intestinal cells in cultures established from different patients.…”

The mucosal barrier at a glance

“…Although this discussion has focused on inflammatory bowel disease and graft-versus-host disease, which has been studied extensively, other examples of disease-associated gut barrier loss abound, including those that occur within the contexts of intestinal infection (Halliez et al, 2016;In et al, 2016;Zolotarevsky et al, 2002), irritable bowel syndrome (Bertiaux-Vandaële et al, 2011;Martínez et al, 2013;Wu et al, 2016), celiac disease (Schumann et al, 2012;Setty et al, 2015;Szakál et al, 2010) and environmental enteric dysfunction (Kelly et al, 2016;Vinetz et al, 2016;Yu et al, 2016). Although studied to a lesser extent, barrier defects also occur in pulmonary, renal and dermatologic diseases.…”

A paradox of transcriptional and functional innate interferon responses of human intestinal enteroids to enteric virus infection

“…Is Reproducible Across HIEs from Different People. HIEs are novel, physiologically active, nontransformed cultures that are increasingly used to study human pathogen-host interactions (5,8,9,39,40). We first evaluated the variability in the transcriptional response to HRV infection in HIEs by analyzing transcriptomes from one jejunal HIE culture by microarray analysis.…”