Ca²⁺ signalling is critical for autoantibody-induced blistering of human epidermis in pemphigus.

Abstract

Pemphigus is a severe bullous autoimmune skin disease. Pemphigus foliaceus (PF) is characterized by antidesmoglein (Dsg) 1 IgG causing epidermal blistering; mucosal pemphigus vulgaris (mPV) by anti-Dsg3 IgG inducing erosions in the mucosa; and mucocutaneous pemphigus vulgaris (PV) by affecting both, with autoantibodies targeting Dsg1 and Dsg3. To characterize the Ca²⁺ flux pathway and delineate its importance in pemphigus pathogenesis and clinical phenotypes caused by different antibody profiles. Immunoprecipitation, Ca²⁺ flux analysis, Western blotting, immunofluorescence staining, dissociation assays and a human skin ex vivo model were used. PV IgG and PF IgG, but neither Dsg3-specific monoclonal antibody (AK23) nor mPV IgG, caused Ca²⁺ influx in primary human keratinocytes. Phosphatidylinositol 4-kinase α interacts with Dsg1 but not with Dsg3. Its downstream target - phospholipase-C-γ1 (PLC) - was activated by PV IgG and PF IgG but not AK23 or mPV IgG. PLC releases inositol 1,4,5-trisphosphate (IP3) causing IP3 receptor (IP3R) activation and Ca²⁺ flux from the endoplasmic reticulum into the cytosol, which stimulates Ca²⁺ release-activated channels (CRAC)-mediated Ca²⁺ influx. Inhibitors against PLC, IP3R and CRAC effectively blocked PV IgG and PF IgG-induced Ca²⁺ influx; ameliorated alterations of Dsg1 and Dsg3 localization, and reorganization of keratin and actin filaments; and inhibited loss of cell adhesion in vitro. Finally, inhibiting PLC or IP3R was protective against PV IgG-induced blister formation and redistribution of Dsg1 and Dsg3 in human skin ex vivo. Ca²⁺ -mediated signalling is important for epidermal blistering and dependent on the autoantibody profile, which indicates different roles for signalling complexes organized by Dsg1 and Dsg3. Interfering with PLC and Ca²⁺ signalling may be a promising approach to treat epidermal manifestations of pemphigus.