title: Yersinia pseudotuberculosis induces transcytosis  of nanoparticles across human intestinal villus  epithelium via invasin-dependent macropinocytosis.
abstract: Crohn's disease is characterized by a defect in intestinal barrier function, where bacteria are considered the most important inflammation-driving factor. Enteric bacteria, including E. coli and Yersinia spp, affect tight junctions in enterocytes, but little is known about bacterial effects on the transcellular pathway. Our objective was to study the short-term effects of Y. pseudotuberculosis on uptake of nanoparticles across human villus epithelium. Monolayers of human colon epithelium-derived Caco-2 cells and biopsies of normal human ileum were studied after 2 h exposure to Y. pseudotuberculosis expressing (inv+) or lacking (inv-) the bacterial adhesion molecule, invasin. Transepithelial transport of fluorescent nanoparticles (markers of transcytosis) was quantified by flow cytometry, and mechanisms explored by using inhibitors of endocytosis. Epithelial expressions of beta1-integrin and particle uptake pathways were studied by confocal microscopy. The paracellular pathway was assessed by electrical resistance (TER), mannitol flux, and expression of tight junction proteins occludin and caludin-4 by confocal microscopy. Inv+ Y. pseudotuberculosis adhered to the apical surface of epithelial cells and induced transcytosis of exogenous nanoparticles across Caco-2 monolayers (30-fold increase, P<0.01) and ileal mucosa (268+/-47% of control; P<0.01), whereas inv bacteria had no effect on transcytosis. The transcytosis was concentration-, particle size- and temperature-dependent, and possibly mediated via macropinocytosis. Y. pseudotuberculosis also induced apical expression of beta1-integrin on epithelial cells. A slight drop in TER was seen after exposure to inv+ Y. pseudotuberculosis, whereas mannitol flux and tight junction protein expression was unchanged. In summary, Y. pseudotuberculosis induced apical expression of beta1-integrin and stimulated uptake of nanoparticles via invasin-dependent transcytosis in human intestinal epithelium. Our findings suggest that bacterial factors may initiate transcytosis of luminal exogenous particles across human ileal mucosa, thus presenting a novel mechanism of intestinal barrier dysfunction.
authors.affiliation: Department of Pharmacy, Uppsala University, Uppsala, Sweden.
authors.forename: Eva G E
authors.initials: EG
authors.lastname: Ragnarsson
authors.forename: Ida
authors.initials: I
authors.lastname: Schoultz
authors.forename: Elisabet
authors.initials: E
authors.lastname: Gullberg
authors.forename: Anders H
authors.initials: AH
authors.lastname: Carlsson
authors.forename: Farideh
authors.initials: F
authors.lastname: Tafazoli
authors.forename: Maria
authors.initials: M
authors.lastname: Lerm
authors.forename: Karl-Eric
authors.initials: KE
authors.lastname: Magnusson
authors.forename: Johan D
authors.initials: JD
authors.lastname: Söderholm
authors.forename: Per
authors.initials: P
authors.lastname: Artursson
mesh_terms: D018829:Adhesins, Bacterial / Q000009:adverse effects; D000368:Aged; D018938:Caco-2 Cells; D020895:Enterocytes / Q000382:microbiology* / Q000502:physiology; D005260:Female; D006801:Humans; D007082:Ileum / Q000166:cytology / Q000503:physiopathology*; D019012:Integrin beta1 / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000382:microbiology / Q000503:physiopathology*; D008297:Male; D018613:Microscopy, Confocal; D008875:Middle Aged; D053758:Nanoparticles; D010873:Pinocytosis / Q000502:physiology*; D019108:Tight Junctions / Q000382:microbiology / Q000502:physiology; D015011:Yersinia pseudotuberculosis / Q000472:pathogenicity*
chemical_list: D018829:Adhesins, Bacterial; D019012:Integrin beta1; C053476:invasin, Yersinia