FIGURE 1. IL-21 inhibits human Treg differentiation and suppressor activity.

(A) Naïve CD4⁺ T cells were isolated from peripheral blood of SLE and matched healthy control (HC) subjects and cultured for 3 days in the presence of anti-CD3/CD28, TGF-β (5 ng/ml), and IL-2 (50 IU/ml) with or without IL-21 (10 ng/ml). Representative flow cytometry dot plots showing CD4⁺CD25⁺FOXP3⁺ Tregs (left). Numbers below the dot plots represent the frequency of CD4⁺CD25⁺FOXP3⁺ Tregs. Cumulative data from 7 matched HC and SLE subjects (right). Data were analyzed by a paired two-tailed t-test (**, p<0.01; ****, p<0.0001). (B) Magnetically sorted CD4⁺CD25⁻ responder T cells (Tresp cells) from HC and SLE donors were stained with CFSE and cultured for 5 days in the presence of plate-bound anti-CD3 and irradiated PBMCs with or without IL-21 (100 ng/ml) in the presence or absence of equal number of autologous CD4⁺CD25⁺ Tregs. Proliferation of Tresp cells was determined by calculating the division index of CFSE⁺ cells using FlowJo software. Representative flow cytometry histograms showing CFSE dilution by Tresp cells in the presence (solid line) or absence (dashed line) of Tregs (left). Numbers below the histograms represent % suppression of division index in the presence of Tregs. Cumulative data represent mean ± SE of Treg suppressive function from 8 pairs of matched HC and SLE subjects (right). Statistical analysis was performed by a paired two-tailed t-test (*, p<0.05).