title: Enterohemorrhagic E. coli alters murine intestinal epithelial tight junction protein expression and barrier function in a Shiga toxin independent manner.
abstract: Shiga toxin (Stx) is implicated in the development of hemorrhagic colitis and hemolytic-uremic syndrome, but early symptoms of enterohemorrhagic Escherichia coli (EHEC) infection such as nonbloody diarrhea may be Stx independent. In this study, we defined the effects of EHEC, in the absence of Stx, on the intestinal epithelium using a murine model. EHEC colonization of intestines from two groups of antibiotic-free and streptomycin-treated C57Bl/6J mice were characterized and compared. EHEC colonized the cecum and colon more efficiently than the ileum in both groups; however, greater amounts of tissue-associated EHEC were detected in streptomycin-pretreated mice. Imaging of intestinal tissues of mice infected with bioluminescent EHEC further confirmed tight association of the bacteria with the cecum and colon. Greater numbers of EHEC were also cultured from stool samples obtained from streptomycin-pretreated mice, as compared with those that received no antibiotics. Transmission electron microscopy shows that EHEC infection leads to microvillous effacement of mouse colonocytes. Hematoxylin and eosin staining of the colonic tissues of infected mice revealed a slight increase in the number of lamina propria polymorphonuclear leukocytes. Transmucosal electrical resistance, a measure of epithelial barrier function, was reduced in the colonic tissues of infected animals. Increased mucosal permeability to 4- kDa FITC-dextran was also observed in the colonic tissues of infected mice. Immunofluorescence microscopy showed that EHEC infection resulted in redistribution of the tight junction (TJ) proteins occludin and claudin-3 and increased the expression of claudin-2, whereas ZO-1 localization remained unaltered. Quantitative real-time PCR showed that EHEC altered mRNA transcription of OCLN, CLDN2, and CLDN3. Most notably, claudin-2 expression was significantly increased and correlated with increased intestinal permeability. Our data indicate that C57Bl/6J mice serve as an in vivo model to study the physiological effects of EHEC infection on the intestinal epithelium and suggest that altered transcription of TJ proteins has a role in the increase in intestinal permeability.
authors.affiliation: Department of Medicine, Section of Digestive Diseases and Nutrition, University of Illinois at Chicago, Chicago, IL 60612-7323, USA.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Roxas
authors.forename: Athanasia
authors.initials: A
authors.lastname: Koutsouris
authors.forename: Amy
authors.initials: A
authors.lastname: Bellmeyer
authors.forename: Samuel
authors.initials: S
authors.lastname: Tesfay
authors.forename: Sandhya
authors.initials: S
authors.lastname: Royan
authors.forename: Kanakeshwari
authors.initials: K
authors.lastname: Falzari
authors.forename: Antoneicka
authors.initials: A
authors.lastname: Harris
authors.forename: Hao
authors.initials: H
authors.lastname: Cheng
authors.forename: Ki Jong
authors.initials: KJ
authors.lastname: Rhee
authors.forename: Gail
authors.initials: G
authors.lastname: Hecht
mesh_terms: D000818:Animals; D062465:Claudin-3; D003106:Colon / Q000378:metabolism / Q000382:microbiology; D003911:Dextrans; D003967:Diarrhea; D054324:Enterohemorrhagic Escherichia coli; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D004927:Escherichia coli Infections / Q000378:metabolism* / Q000382:microbiology*; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives; D006463:Hemolytic-Uremic Syndrome; D007082:Ileum / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism / Q000382:microbiology*; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D062793:Occludin; D010539:Permeability; D022621:Shiga Toxin / Q000378:metabolism; D019108:Tight Junctions / Q000378:metabolism / Q000382:microbiology / Q000502:physiology
chemical_list: D062465:Claudin-3; C568123:Cldn3 protein, mouse; D003911:Dextrans; D008565:Membrane Proteins; D062793:Occludin; C568986:Ocln protein, mouse; C015219:fluorescein isothiocyanate dextran; D022621:Shiga Toxin; D016650:Fluorescein-5-isothiocyanate