Identification, frequency, activation and function of CD4+ CD25(high)FoxP3+ regulatory T cells in children with juvenile idiopathic arthritis.

The aim of the study was to test the frequency of CD4+ CD25(high)FoxP3 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.