Identification, frequency, activation and function of CD4+ CD25highFoxP3+ regulatory T cells in children with juvenile idiopathic arthritis

The aim of the study was to test the frequency of CD4+ CD25highFoxP3 regulatory T cells in JIA patients and to assess their activation status and functional activity. The study involved 12 children with JIA and 35 healthy control subjects. PBMC were stained with monoclonal antibodies (anti-CD25, anti-CD4, anti-CD127, anti-CD69, anti-CD71, and anti-FoxP3). The samples were evaluated using flow cytometer. CD4+ CD25− and CD4+ CD25+ cells were isolated by negative and positive selection with magnetic microbeads. CD4+ CD25+ and CD4+ CD25− cells were cultured separately and co-cultured (1:1) with or without PHA. The percentage of Tregs in JIA patients was significantly decreased in comparison with controls (median, 3.2 vs. 4.6; P = 0.042). Relative fluorescence intensities of FoxP3 were higher in JIA patients than in controls (median, 9.1 vs. 6.8). The percentage of activated Tregs (CD71+) was significantly higher in JIA patients in comparison with controls (median, 6.5 vs. 2.8; P = 0.00043). CD4+ CD25+ cells derived from JIA patients and controls were anergic upon PHA stimulation, while CD4+ CD25− cells showed intensive proliferative response. The proliferation rate of CD4+ CD25− cells stimulated by PHA was decreased in co-cultures. In JIA patients, the inhibition of proliferation of CD4+ CD25− cells by CD4+ CD25+ cells was 37.9%, whereas in controls it was significantly lower (55.7%, P = 0.046). JIA patients had statistically lower percentage of Tregs in peripheral blood compared to controls. CD4+ CD25+ cells sorted from peripheral blood of JIA patients had statistically lower ability to suppress CD4+ CD25− cell proliferation in comparison with cells obtained from controls

Dermatophagoides pteronyssinus immunotherapy changes the T-regulatory cell activity

Abstract Subcutaneous specific immunotherapy (SCIT) has been shown to modify the Dermatophagoides pteronissinus (DP) allergic response, characterized by generation of Treg cells. However, studies have reported no changes in the proportion of Treg cells after immunotherapy, indicating that the effects may be due to modifications in their regulatory activities. We aimed to determine whether Tregs generated by DP-SCIT can switch the allergic response to tolerant and study the involvement of suppressive cytokines on it. Twenty-four DP-allergic rhinitis patients were recruited, 16 treated with DP-SCIT and 8 untreated. Treg and T effector cells were isolated before and after DP-SCIT, and cocultured in different combinations with α-IL-10, α-TGF-β blocking antibodies and nDer p 1. Treg cells after DP-SCIT increased Th1 and decreased Th2 and Th9 proliferation. Similarly, they increased IL-10 and decreased IL-4 and IL-9-producing cells. α-IL-10 affected the activity of Treg cells obtained after DP-SCIT only. Finally, DP-specific IgG4 levels, Treg percentage and IL-10 production were correlated after DP-SCIT. These results demonstrate that DP-SCIT induces Treg cells with different suppressive activities. These changes could be mediated by IL-10 production and appear to play an important role in the induction of the tolerance response leading to a clinical improvement of symptoms