Figure 5.

OP enhances Foxp3⁺ cells and Foxp3⁺ RORγ⁺ cells in culture with CD4⁺ T cells from EW-sensitised mice. (A,B) Spleen CD4⁺ T cells from naïve mice were cultured without stimuli (RPMI), or with OP, RA, OP+IL-4, or RA+IL-4 for 2 days, in the absence or presence of TGF-β, followed by stimulation with anti-CD3 and anti-CD28 for 2 additional days.The percentage of Foxp3⁺, RORγ⁺, Foxp3⁺ RORγ⁺, IL-10⁺, and GATA3⁺ cellswithin the total CD4⁺ T cell population was assayed by flow cytometry (A), and gene expression of Aldh1a1, Aldh1a2, Tgfb1, Il6, Tlr2, Tlr4, and Tlr5 was assayed by qPCR, normalised to the reference gene Actb, and expressed relative to CD4⁺ T cells cultured in RPMI (B). (C,D) Spleen CD4⁺ T cells from EW-sensitised mice were cultured without stimuli (RPMI) or with OP, OVA, or LPS for 2 days, followed by stimulation with anti-CD3 and anti-CD28 for 2 additional days.The percentage of Foxp3⁺, RORγ⁺, and Foxp3⁺ RORγ⁺ cellswithin the total CD4⁺ T cell population was assayed by flow cytometry(C), and gene expression of Aldh1a1, Aldh1a2, Tgfb1, Il6, Tlr2, Tlr4, and Tlr5 was assayed by qPCR, normalised to the reference gene Actb, and expressed relative to CD4⁺ T cells cultured in RPMI (D). Results of an experiment representative of 3 separate experiments performed in triplicate are shown. Data are means ± SEM. Different letters indicate statistically significant differences (p < 0.05) calculated using one-way ANOVA, followed by Tukey’s post-hoc test (A,C) or the Mann–Whitney U test (B,D).