Enterohemorrhagic Escherichia coli reduce mucus and intermicrovillar bridges in human stem cell-derived colonoids

Julie In; Jennifer Foulke-Abel; Nicholas C Zachos; Anne-Marie Hansen; James B Kaper; Harris D Bernstein; Marc Halushka; Sarah Blutt; Mary K Estes; Mark Donowitz; Olga Kovbasnjuk

doi:10.1016/j.jcmgh.2015.10.001

Enterohemorrhagic Escherichia coli reduce mucus and intermicrovillar bridges in human stem cell-derived colonoids

Cell Mol Gastroenterol Hepatol. 2016 Jan 1;2(1):48-62.e3. doi: 10.1016/j.jcmgh.2015.10.001.

Authors

Affiliations

¹ Department of Medicine, Division of Gastroenterology and Hepatology, Johns Hopkins University School of Medicine, Baltimore, MD.
² Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD.
³ Genetics and Biochemistry Branch, National Institutes of Diabetes and Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD.
⁴ Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD.
⁵ Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, TX.

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.

Keywords: human colonoid monolayers; intestinal organoids; microvillar effacement; serine protease EspP.

Abstract

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