NR4A2 knockdown prevents IL-17 secretion but not RORγt upregulation.

<p>Naïve CD4⁺ T cells were transfected by electroporation with NR4A2-specific siRNA or scrambled control siRNA. Cells were then activated with 5 µg/ml plate-bound CD3-specific mAb and 0.5 µg/ml soluble CD28-specific mAb. <b>A:</b> IFN-γ production by cells activated in the presence or absence of 10 ng/ml IL-12 after 96 hours of culture. <b>B:</b> IL-17 production by cells activated in the presence of 20 ng/ml IL-6, 20 ng/ml IL-23, and TGF-β at a range of concentrations after 96 hours of culture. Significant differences between control and NR4A2 siRNA-treatments were tested with a student’s t-test, *p<0.05. <b>C:</b> IL-17 and IFN-γ intracellular cytokine staining for transfected T cells (control siRNA, left plots; NR4A2 siRNA, right plots) cultured for 96 hours in the presence of 10 ng/ml IL-12 (Th1 conditions, top row plots) or 20 ng/ml IL-6, 20 ng/ml IL-23, and 3 ng/ml TGF-β (Th17 conditions, bottom row plots). <b>D:</b> RORγt RNA expression as measured by real time PCR by activated T cells cultured under Th17 polarizing conditions at a range of timepoints. Data are representative of 5 independent experiments.