title: Ex-vivo whole blood secretion of interferon (IFN)-γ and IFN-γ-inducible protein-10 measured by enzyme-linked immunosorbent assay are as sensitive as IFN-γ enzyme-linked immunospot for the detection of gluten-reactive T cells in human leucocyte antigen (HLA)-DQ2·5(+) -associated coeliac disease.
abstract: T cell cytokine release assays are used to diagnose infectious diseases, but not autoimmune or allergic disease. Coeliac disease (CD) is a common T cell-mediated disease diagnosed by the presence of gluten-dependent intestinal inflammation and serology. Many patients cannot be diagnosed with CD because they reduce dietary gluten before medical workup. Oral gluten challenge in CD patients treated with gluten-free diet (GFD) mobilizes gluten-reactive T cells measurable by interferon (IFN)-γ enzyme-linked immunospot (ELISPOT) or major histocompatibility complex (MHC) class II tetramers. Immunodominant peptides are quite consistent in the 90% of patients who possess HLA-DQ2·5. We aimed to develop whole blood assays to detect gluten-specific T cells. Blood was collected before and after gluten challenge from GFD donors confirmed to have CD (n = 27, all HLA-DQ2·5(+) ), GFD donors confirmed not to have CD (n = 6 HLA-DQ2·5(+) , 11 HLA-DQ2·5(-) ) and donors with CD not following GFD (n = 4, all HLA-DQ2·5(+) ). Plasma IFN-γ and IFN-γ inducible protein-10 (IP-10) were measured by enzyme-linked immunosorbent assay (ELISA) after whole blood incubation with peptides or gliadin, and correlated with IFN-γ ELISPOT. No T cell assay could distinguish between CD patients and controls prior to gluten challenge, but after gluten challenge the whole blood IFN-γ ELISA and the ELISPOT were both 85% sensitive and 100% specific for HLA-DQ2·5(+) CD patients; the whole blood IP-10 ELISA was 94% sensitive and 100% specific. We conclude that whole blood cytokine release assays are sensitive and specific for detection of gluten-reactive T cells in CD; further clinical studies addressing the utility of these tests in patients with an uncertain diagnosis of CD is warranted.
authors.affiliation: The Walter and Eliza Hall Institute of Medical Research, Parkville, Vic., Australia; Department of Medical Biology, The University of Melbourne, Parkville, Vic., Australia.
authors.forename: N
authors.initials: N
authors.lastname: Ontiveros
authors.forename: J A
authors.initials: JA
authors.lastname: Tye-Din
authors.forename: M Y
authors.initials: MY
authors.lastname: Hardy
authors.forename: R P
authors.initials: RP
authors.lastname: Anderson
mesh_terms: D000328:Adult; D000368:Aged; D002446:Celiac Disease / Q000097:blood / Q000175:diagnosis*; D054357:Chemokine CXCL10 / Q000097:blood* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D058501:Enzyme-Linked Immunospot Assay; D005260:Female; D005903:Gliadin / Q000276:immunology; D005983:Glutens / Q000276:immunology; D006683:HLA-DQ Antigens / Q000032:analysis / Q000276:immunology; D006801:Humans; D007371:Interferon-gamma / Q000097:blood* / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D013601:T-Lymphocytes / Q000276:immunology*; D055815:Young Adult
chemical_list: D054357:Chemokine CXCL10; D006683:HLA-DQ Antigens; C053186:HLA-DQ2 antigen; D005983:Glutens; D007371:Interferon-gamma; D005903:Gliadin
keywords: coeliac disease; diagnostics; gluten challenge