Abstract
BACKGROUND: Pemphigus foliaceus (PF) is an autoimmune blistering disease caused by autoantibodies (Abs) against desmoglein 1 (Dsg1). PF sera contain polyclonal Abs which are heterogeneous mixture of both pathogenic and non-pathogenic Abs, as shown by isolation of monoclonal Abs (mAbs). OBJECTIVE: To investigate how pathogenic and non-pathogenic anti-Dsg1 Abs contribute to blister formation in PF. METHODS: Using organ-cultured human skin, we compared the effect of a single pathogenic anti-Dsg1 IgG mAb, a single non-pathogenic anti-Dsg1 IgG mAb, and their mixture on blister formation as analyzed by histology, subcellular localization of IgG deposits and desmosomal proteins by confocal microscopy, and desmosomal structure by electron microscopy. In addition, we measured keratinocyte adhesion by an in vitro dissociation assay. RESULTS: 24h after injection, a single pathogenic anti-Dsg1 IgG caused a subcorneal blister with IgG and Dsg1 localized linearly on the cell surface of keratinocytes. A single non-pathogenic anti-Dsg1 IgG bound linearly on the keratinocytes but did not induce blisters. A pathogenic and a non-pathogenic IgG mAb injected together caused an aberrant granular pattern of IgG and Dsg1 in the lower epidermis with blister formation in the superficial epidermis. Electron microscopy demonstrated that the mixture of mAbs shortened desmosomal lengths more than a single mAb in the basal and spinous layers. Furthermore, although Dsg1 clustering required both cross-linking of Dsg1 molecules by the non-pathogenic IgG plus a pathogenic antibody, the latter could be in the form of a monovalent single chain variable fragment, suggesting that loss of trans-interaction of Dsg1 is required for clustering. Finally, a p38MAPK inhibitor blocked Dsg1 clustering. When pathogenic strength was measured by the dissociation assay, a mixture of pathogenic and non-pathogenic IgG mAbs disrupted keratinocyte adhesion more than a single pathogenic mAb. This pathogenic effect was only partially suppressed by the p38MAPK inhibitor. CONCLUSION: These findings indicate that a polyclonal mixture of anti-Dsg1 IgG antibodies enhances pathogenic activity for blister formation associated with p38MAPK-dependent Dsg1 clustering and that not only pathogenic antibodies but also non-pathogenic antibodies coordinately contribute to blister formation in PF.