Enhanced Production of Both Th1- and Th2-Polarizing Cytokines by SOCS1-Silenced DCs and Activated CD4⁺ Th

<div><p>(A) Enhanced production of both Th1- and Th2-polarizing cytokines by SOCS1-silenced DCs. BM-DCs transfected with SOCS1 siRNA or control [<a href="http://www.plosmedicine.org/article/info:doi/10.1371/journal.pmed.0030011#pmed-0030011-b024" target="_blank">24</a>] were stimulated with LPS (100 ng/ml). Concentrations of various cytokines in the culture media were analyzed by ELISA 24 h after stimulation. NS, no stimulation. *<i>P</i> < 0.01, LV-SOCS1-siRNA-DCs versus LV-GFP-siRNA-DCs.</p> <p>(B) IL-12 production by transduced DCs. BM cells were cultured with mGM-CSF (20 μg/ml only or mGM-CSF and mIL-4 (20 μg/ml) [<a href="http://www.plosmedicine.org/article/info:doi/10.1371/journal.pmed.0030011#pmed-0030011-b024" target="_blank">24</a>] for 6 d and then transduced with LV-SOCS1-siRNA or LV-GFP-siRNA. The transduced DCs (5 × 10⁵/ml) were then stimulated with LPS (100 ng/ml) and plate-coated anti-CD40 mAb (5 μg/ml, BD Bioscience) in the presence or absence of IL-4. Concentrations of IL-12 (p70) in the culture media were analyzed by ELISA 24 h after stimulation and are presented from one of three independent experiments. *<i>P</i> < 0.01, LV-SOCS1-siRNA (GM-CSF + IL-4) versus LV-GFP-siRNA (GM-CSF + IL-4).</p> <p>(C–E) CD4⁺ Th responses induced by LV-SOCS1-siRNA-DCs. CD4⁺ T cells were isolated from pooled splenocytes of different groups of mice and subjected to the following assays. Numbers of IFN-γ-producing CD4⁺ T cell precursors were determined with the ELISPOT assay (C). ³H-thymidine incorporation rates of the isolated CD4⁺ T cells were determined on the fourth day of restimulation with gp120-pulsed DCs (D). Cytokine levels in the culture medium of isolated CD4⁺ cells stimulated with gp120-pulsed DCs for 48 h were determined by ELISA (E). The mean results (+ standard error) from one of three experiments are presented. *<i>P</i> < 0.01, LV-SOCS1-siRNA-DC versus LV-GFP-siRNA-DC mice.</p>