TRAIL Signalling Is Pro-Inflammatory in Eosinophilic Esophagitis

S A T U R D A Y 247 Functional Analysis of Calpain-14 in Eosinophilic Esophagitis Benjamin P. Davis, MD, Leah Claire Kottyan, PhD, Emily Stucke, BA, Joseph D. Sherrill, PhD, Marc E. Rothenberg, MD, PhD; Division of Allergy and Immunology, Department of Pediatrics, Cincinnati Children’s Hospital Medical Center, University of Cincinnati, Cincinnati, Ohio, USA, Center for Autoimmune Genomics and Etiology, Department of Pediatrics, Cincinnati Children’s Hospital Medical Center, University of Cincinnati, Cincinnati, Ohio, USA, Cincinnati, OH, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH, Children’s Hospital Medical Center, Cincinnati, OH. RATIONALE: We recently identified a genome-wide association susceptibility locus for eosinophilic esophagitis (EoE) at 2q23 wherein CAPN14 (p52.5 3 10210) is located. CAPN14 encodes for a member of the calpain protease family. We have shown that CAPN14 was specifically expressed in the esophagus, dynamically regulated as a function of disease activity and genetic haplotype and after exposure of epithelial cells to IL-13. Herein, we aimed to determine the function of calpain-14. METHODS: We overexpressed calpain-14 in an esophageal progenitor cell line (EPC2) grown in an air-liquid interface (ALI) culture and analyzed barrier effects by transepithial electrical resistance (TEER), fluorescein isothiocyanate (FITC)-dextran flux, and hematoxylin and eosin stain. We also identified potential calpain-14 targets via western blot and immunofluorescence (IF) of the ALI culture. RESULTS: Calpain-14 over-expression induced a 2-3 fold (p50.0034) decrease in TEER and a 2-3 fold (P<0.0001) increase in FITC-dextran flux, consistent with calpain-14 induced impaired barrier function (IBF). Histological analysis revealed that calpain-14 overexpression induced acantholysis, intraepithelial clefting, and epidermolysis. Western blot following calpain-14 overexpression revealed degradation of the desmosome protein, desmoglein-1 (DSG-1), as a low molecular weight band was increased 30-fold (p50.0009) compared to control. IF for DSG-1 demonstrated reduced expression and less defined desmosome-like structures following calpain-14 overexpression versus control. CONCLUSIONS: The genetic pathoetiology of EoE is mediated by genetic variation of CAPN14 and associated with an IL-13–inducible esophageal response involving calpain-14 induced IBF mediated by DSG-1 cleavage. The study was supported by the Consortium of Food Allergy Researchers (CoFAR) by US NIH grant U19 AI066738 from NIAID and NIDDK.