Background and aims: The a2-gliadin-33mer was previously shown to be important in the pathogenesis of celiac disease. We aimed to study mechanisms of its epithelial translocation and processing in respect to transcytotic and paracellular pathways.
Methods: Transepithelial passage of a fluorescence-labeled a2-gliadin-33mer was studied in Caco-2 with RP-HPLC, mass spectrometry, confocal laser scanning microscopy and FACS analysis. Endocytosis mechanisms were characterized with rab-GFP constructs transiently transfected into Caco-2 cells and in human duodenal biopsy specimens.
Results: The a2-gliadin-33mer dose-dependently crossed the epithelial barrier in apical-to-basal direction. Degradation analysis revealed translocation of the 33mer-polypeptide in uncleaved as well as in degraded form. Transcellular passage was identified by confocal laser scanning microscopy, inhibitor experiments and FACS analysis. Rab5 but not rab4 or rab7 vesicles were shown to be part of the transcytotic pathway. After interferon-g preincubation, translocation of the 33mer was increased by 40%. In mucosal biopsies of the duodenum, epithelial 33mer uptake was significantly higher in untreated CD patients than in healthy controls or CD patients on a gluten-free diet.
Conclusion: Epithelial translocation of the a2-gliadin-33mer occurs by transcytosis after partial degradation through a rab5 endocytosis compartment and is regulated by interferon-g. 33mer uptake is higher in acute CD than in controls and CD on gluten-free diet.