T Cell Response Toward Tissue-and Epidermal-Transglutaminases in Coeliac Disease Patients Developing Dermatitis Herpetiformis

Abstract

The reason why only few coeliac patients develop the cutaneous manifestation of the disease, named dermatitis herpetiformis (DH), is still unknown. Epidermal transglutaminase (TG3) has been described as the main autoantigen of humoral immunity in DH but the mechanisms leading to this autoimmune response remain obscure. Here we characterized T cells from skin, gut and peripheral blood of DH and coeliac disease (CD) patients, evaluated the impact of the gluten-free diet on circulating T lymphocytes' phenotype and investigated antigen specific T cell response toward epidermal and tissue transglutaminase (TG2). DH patients showed an increased frequency of skin-derived T cells producing TNFα when compared to CD patients. Moreover, circulating T cells producing TNFα and IL-17A positively correlated with clinical score of skin disease activity and decreased after gluten-free diet. Finally, TG2 and TG3-specific T cells resulted more reactive to antigens stimulation in DH patients and showed cross reactivity toward the two autoantigens in both the group of patients. Our data suggest a role of TNFα and IL-17A producing cells in the development of DH and, for the first time, show the existence of a crossed T cell response toward the two transglutaminases isoforms, thus suggesting new insights on T cells role in skin damage.

Keywords: T lymphocytes (CD4+); celiac disease; cross reactivity; epidermal transglutaminase; tissue transglutaminase (TG2).

Figure 1

TNFα-producing cells are enriched in skin samples from DH patients. (A) Intracellular cytokines production in in vitro expanded CD4+ T cells from PB (black columns), skin (grey columns) and gut (white columns): frequencies of cytokines producing of CD4+CD3+-gated cells were assessed in 10 DH patients, 6 CD patients and 4 healthy donors. Columns represent means ( ± SE). Bars * indicate p ≤ 0.05; bars ** indicate p ≤ 0.01. (B) Frequencies of intracellular cytokine producing CD4+ T cells in fresh samples from PB (black columns), skin (grey columns) and gut (white columns) of 5 DH patients. Columns represent means ( ± SE). Bars * indicate p ≤ 0.05. (C) Representative flow cytometric analysis obtained in one out of five subjects with DH and in only one healthy specimen. Percentages of CD4+ T cells are shown. PB,peripheral blood.

Figure 2

PB Cytokine-producing T cells frequencies correlation with disease activity score and evaluation after gluten free diet in DH patients. (A) Correlation between disease activity score PDAI and frequencies of circulating CD4+ T cells producing TNFα (left panel) and L-17A (right panel) was calculated in 15 DH patients. (B) Frequencies of intracellular cytokines producing CD4+ T cells in fresh samples from PB were evaluated in 8 DH patients before and after 1 year of gluten free diet (GFD). Columns represent means ( ± SE) of the indicated cytokine. *p ≤ 0.05. (C) Frequencies of circulating CD4+ T cells producing TNFα (left panel) and IL-17A (right panel) in fresh samples from PB were evaluated before (To) and after 1 and 2 year of gluten free diet (GFD) in 5 DH patients. ( ± SE). *p ≤ 0.05 compared to the To evaluation.

Figure 3

Transglutaminases-specific T cells proliferation assays. (A) Proliferation assay was performed on PB derived TCLs, induced in the presence of TG2 and TG3, from 15 DH patients (left panel) and 12 CD patients (right panel). Each TCLs was stimulated with/without both TG2 and TG3 antigens. Dots with the same shape and colour represent stimulation index (SI) of TCLs derived from the same patients. (B) Proliferation assay was performed on T cell clones (TCCs), derived from antigen specific TCLs from 1 DH patients. Each TCC was stimulated with/without both TG2 and TG3 antigens as well as streptokinase (SK) as negative unrelated control. Dots with the same colour represent SI of the same TCC.