No abstract

Pemphigus vulgaris (PV) is the most severe autoimmune bullous skin disorder and is primarily associated with circulating autoantibodies (autoAb) against desmoglein 3 (Dsg3). In light of recent evidence that autoreactive T cells are critical for the induction and regulation of Ab production, the goal of this study was to characterize and quantitate autoreactive T cells in patients with PV and healthy controls. Peripheral Dsg3-reactive Th cells from 28 patients with acute-onset, chronic active, and remittent PV were quantitated by MACS secretion assay. Dsg3-reactive Th2 cells were detected at similar frequencies in all studied PV patients, while the number of autoreactive Th1 cells exceeded those of Th2 cells in chronic active PV. In contrast, healthy carriers of the PV-associated HLA class II alleles, DRB1*0402 and DQB1*0503, exhibited exclusively Dsg3-reactive Th1 cell responses, while healthy carriers of other HLA class II alleles did not. Moreover, the presence of IgG1 and IgG4 against Dsg3 was directly related to the ratio of Dsg3-reactive Th1/Th2 cells. T cell recognition of Dsg3 was restricted by HLA-DRB1*0402 and DQB1*0503 in PV patients and Dsg3-responsive healthy donors. These observations strongly suggest 1) that the appearance of Dsg3-reactive Th2 cells is restricted to patients with PV; 2) that specific HLA class II alleles that are prevalent in PV are critical for T cell recognition of Dsg3 in PV patients and Dsg3-responsive healthy donors; and 3) that autoAb production is associated with both Th1 and Th2 cells.

Pemphigus vulgaris (PV) is a severe autoimmune bullous skin disorder and is primarily associated with circulating autoantibodies against desmoglein 3 (Dsg3) that are presumably regulated by Th cells. The aim of this study was to identify Dsg3-specific T regulatory (Tr) cells that may help to maintain and restore natural tolerance against Dsg3. Dsg3-responsive IL-10-secreting Tr1 cells were isolated by MACS cytokine secretion assay from healthy carriers of the PV-associated HLA class II alleles, DRB1*0402 and DQB1*0503, but were only rarely detected in PV patients. The Dsg3-specific Tr1 cells secreted IL-10, TGF-β, and IL-5 upon Ag stimulation, proliferated in response to IL-2 but not to Dsg3 or mitogenic stimuli, and inhibited the proliferative response of Dsg3- and tetanus toxoid-responsive Th clones in an Ag-specific (Dsg3) and cell number-dependent manner. Moreover, their inhibitory effect was blocked by Ab against IL-10, TGF-β, and by paraformaldehyde fixation. These observations strongly suggest that 1) Dsg3-responsive Tr1 cells predominate in healthy individuals, 2) their growth requires the presence of IL-2, and 3) they exert their Dsg3-dependent inhibitory function by the secretion of IL-10 and TGF-β. Because autoaggressive T cells responsive to identical epitopes of Dsg3 were recently found both in PV patients and healthy individuals, the identified Tr1 cells may be critically involved in the maintenance and restoration of tolerance against Dsg3.

Pemphigus vulgaris is a severe autoimmune disease caused by autoantibodies against the cutaneous adhesion molecule, desmoglein 3 (Dsg3). The aim of this study was to characterize the specificity of autoreactive Th cells, which presumably regulate Dsg3-specific autoantibody production. Ninety-seven Th1 and Th2 clones isolated from 16 pemphigus patients and 12 HLA-matched healthy donors recognized the Dsg3 peptides, DG3(78-94), DG3(96-112), DG3(189-205), DG3(205-221), and DG3(250-266). Peptide DG3(96-112), and to a lesser extent DG3(250-266), was recognized by the majority of T cells from patients and healthy donors in association with HLA-DRB1*0402 and DQB1*0503 which were prevalent in the pemphigus patients and Dsg3-responsive healthy donors. Analyzing the Vβ-chain of the TCR of the DG3(96-112)-specific T cells showed no restricted TCR usage. Peptides DG3(342-358) and DG3(376-392) were exclusively recognized by T cell clones (n = 13) from patients while DG3(483-499) was only recognized by T cell clones (n = 3) from a healthy donor. All Dsg3 peptides contained conserved amino acids at relative positions 1, 4, and 6; amino acids with a positive charge at position 4 presumably represent anchor motifs for DRB1*0402. These findings demonstrate that T cell recognition of distinct Dsg3 peptides is restricted by distinct HLA class II molecules and is independent from the development of pemphigus vulgaris.

Pemphigus vulgaris (PV) is a severe autoimmune bullous skin disorder and is associated with autoantibodies against desmoglein (Dsg)3 that are regulated by Th2 cells. Recently, Dsg3-specific type 1 regulatory T cells (Tr1) were identified that are presumably critical for the maintenance of tolerance against Dsg3 because there is a much lower Dsg3-specific Tr1:Th2 ratio in the PV patients than in healthy individuals. The aim of this study was to down-regulate the transcription factor Foxp3 in Dsg3-specific Tr1 using antisense oligonucleotides because Foxp3 is constitutively expressed by the Dsg3-specific Tr1. Antisense-treated Dsg3-specific Tr1 clones lost expression of Foxp3, glucocorticoid-induced TNFR family-related receptor, and CTLA-4, and started to secrete IL-2, whereas the secretion of IL-5, TGF-β, and IL-10 remained unchanged. Moreover, antisense treatment induced a proliferative response to Dsg3 of the formerly anergic Tr1 and abrogated their suppressor activity on Dsg3-specific Th2 cell clones. Thus, inhibition of Foxp3 mRNA expression in the Tr1 induced a Th2-like phenotype. In conclusion, Foxp3 expression is inherent to Tr1 function, and modulation of Foxp3 expression in autoaggressive Th2 cells may provide a novel therapeutic approach aimed at restoring tolerance against Dsg3 in PV.