632436.fig.003a
(a)
632436.fig.003b
(b)
632436.fig.003c
(c)
632436.fig.003d
(d)
Figure 3: rSSP4 immunization induced CD4+CD25+ T cells with suppressive function in vitro. (a) A total of CD4+CD25 T cells (from naïve mice) and APCs (from nonimmunized mice) were cultured alone or in combination with titer quantities of rSSP4-primed CD4+CD25+ T cells in the presence of anti-CD3 and anti-CD28 antibodies plus antigen. Proliferation was quantified by 3H-Thymidine incorporation during the last 16 h of culture. (b) An aliquot of rSSP4-primed CD4+CD25+ sorted T cells was tested for FOXP3 expression. Results shown are representative of three independent experiments (c) and (d). Anti-TGF-β antibodies partially inhibit suppressor activity of CD4+CD25+ T cells. A total of 5 × 104 CD4+ T cells (from immunized mice) and 1 × 105 APCs (from nonimmunized mice) were cultured alone or in combination with titer quantities of rSSP4-primed CD4+CD25+ T cells in the presence of anti-CD3 and anti-CD28 antibodies plus antigen in the presence of 5 μg/mL of neutralizing anti-IL-10 (c) or anti-TGF-β (d) antibodies. Proliferation was quantified by 3H-Thymidine incorporation during the last 16 h of culture. Results are the average of three independent experiments done in duplicates.