title: Trypsin impaired epithelial barrier function and induced IL-8 secretion through basolateral PAR-2: a lesson from a stratified squamous epithelial model.
abstract: Immune-mediated injury by the protease-activated receptor-2-interleukin-8 (PAR-2-IL8) pathway may underlie the development of gastroesophageal reflux disease (GERD). However, the localization of PAR-2 and the mechanism of PAR-2 activation remain unclear. This study aimed to address these questions on an esophageal stratified squamous epithelial model and in the human esophageal mucosa of GERD patients. Normal human esophageal epithelial cells were cultured with the air-liquid interface system to establish the model. SLIGKV-NH2 (PAR-2 synthetic agonist), trypsin (PAR-2 natural activator), and weak acid (pH 4, 5, and 6) were added to either the apical or basolateral compartment to evaluate their effects on transepithelial electrical resistance (TEER) and IL-8 production. PAR-2 localization was examined both in the cell model and biopsies from GERD patients by immunohistochemistry. Apical trypsin stimulation induced IL-8 accompanied by decreased TEER in vitro, whereas the effective concentration from the basolateral side was 10 times lower. SLIGKV-NH2 from basolateral but not apical stimulation induced IL-8 production. Apical weak acid stimulation did not influence TEER or IL-8 production. Immunohistochemistry showed intense reactivity of PAR-2 in the basal and suprabasal layers after stimulation with trypsin. A similar PAR-2 reactivity that was mainly located at the basal and suprabasal layers was detected in GERD patients. In conclusion, the activation of the PAR-2-IL-8 pathway probably occurred at the basal and suprabasal layers, while the esophageal epithelial barrier may influence the activation of PAR-2. Under proton pump inhibitor therapy, refluxed trypsin may remain active and be a potential agent in the pathogenesis of refractory GERD.
authors.affiliation: Division of Upper Gastroenterology, Dept. of Internal Medicine, Hyogo College of Medicine, Nishinomiya, Japan.
authors.forename: Jing
authors.initials: J
authors.lastname: Shan
authors.forename: Tadayuki
authors.initials: T
authors.lastname: Oshima
authors.forename: Xin
authors.initials: X
authors.lastname: Chen
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Fukui
authors.forename: Jiro
authors.initials: J
authors.lastname: Watari
authors.forename: Hiroto
authors.initials: H
authors.lastname: Miwa
mesh_terms: D017097:Electric Impedance; D004847:Epithelial Cells / Q000378:metabolism; D004947:Esophagus / Q000378:metabolism; D005764:Gastroesophageal Reflux / Q000378:metabolism; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology; D044464:Receptor, PAR-2 / Q000378:metabolism*; D014357:Trypsin / Q000494:pharmacology*
chemical_list: D016209:Interleukin-8; D009842:Oligopeptides; D044464:Receptor, PAR-2; C515807:seryl-leucyl-isoleucyl-glycyl-lysyl-valinamide; D014357:Trypsin