title: A novel cell-based assay for inhibitory anti-muscarinic type 3 receptor antibodies in primary Sjögren's syndrome.
abstract: Inhibitory autoantibodies acting at the muscarinic acetylcholine receptor type 3 (M3R) are postulated to mediate autonomic dysfunction, including decreased salivary and lacrimal gland output and extra-glandular manifestations, in patients with primary Sjögren's syndrome. However, the contention that anti-M3R antibodies are pathogenic in patients remains untested, due to a lack of assays both sophisticated enough to detect inhibitory anti-M3R antibodies yet suitable for screening large patient cohorts. In the current study, we have established a cell-based bioassay of M3R activity, based on dual transfection of the M3R and a luciferase reporter gene. The bioassay is capable of capturing real-time agonist-mediated signalling of the M3R, which is inhibited specifically by patient IgG that have previously been demonstrated to have anti-M3R activity. The assay can be run in multi-well culture plates, and analysed using simple luminescence readers. As such, the new bioassay incorporating M3R-mediated luciferase transduction is the first assay adaptable to common diagnostic platforms that is capable of determining the presence in patient serum of functionally active anti-M3R autoantibodies. The new bioassay should prove useful for large cohort screening studies aiming to correlate the presence in patients of inhibitory anti-M3R antibodies with symptoms of both glandular and extra-glandular autonomic dysfunction.
authors.affiliation: Department of Immunology, Allergy & Arthritis, Flinders University, Bedford Park 5042, South Australia, Australia.
authors.forename: Isabell
authors.initials: I
authors.lastname: Bastian
authors.affiliation: Department of Immunology, Allergy & Arthritis, Flinders University, Bedford Park 5042, South Australia, Australia.
authors.forename: Tom P
authors.initials: TP
authors.lastname: Gordon
authors.affiliation: Department of Immunology, Allergy & Arthritis, Flinders University, Bedford Park 5042, South Australia, Australia. Electronic address: michael.jackson@flinders.edu.au.
authors.forename: Michael W
authors.initials: MW
authors.lastname: Jackson
mesh_terms: D001323:Autoantibodies / Q000097:blood*; D001324:Autoantigens / Q000276:immunology; D057809:HEK293 Cells; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D043587:Receptor, Muscarinic M3 / Q000276:immunology*; D012859:Sjogren's Syndrome / Q000276:immunology*; D014162:Transfection
chemical_list: D001323:Autoantibodies; D001324:Autoantigens; D043587:Receptor, Muscarinic M3
keywords: Anti-M3R antibodies; Autoantibodies; Functional antibodies; Muscarinic receptors; Sjögren’s syndrome