Fig. 4

p38 inhibits T helper type 9 (T_H9) cell generation by Fas signaling. a Flow cytometric analysis of IL-9⁺ (interleukin-9⁺) cell frequency among CD4⁺ T cells after naive CD4⁺ T cells were stimulated with 10 μg ml⁻¹ ISO or Jo2 and with or without 0.4 μM p38 inhibitor SB203580 under T_H9-skewing conditions for 4 days. b Western blotting analysis of p-p38 proteins in naive CD4⁺ T cells stimulated with 10 μg ml⁻¹ ISO or Jo2 for 15 min and with or without 0.5 μM enzastaurin under T_H9-skewing conditions (up), and statistical analysis of the relative protein levels (down). c Flow cytometric analysis of IL-9⁺ cell frequency among CD4⁺ T cells after naive CD4⁺ T cells were stimulated with 10 μg ml⁻¹ ISO or Jo2 and with or without NFAT inhibitor (5 pM FK506 or 50 nM INCA-6) under T_H9-skewing conditions for 4 days. d, e Western blotting analysis of p-NFAT1 proteins in WT or Fas^lpr CD4⁺ T cells (up) (d) or WT CD4⁺ T cells stimulated with 10 μg ml⁻¹ ISO or Jo2 (e) under T_H9-skewing conditions for 15 min (up), and statistical analysis of the relative protein levels (down) (d, e). f Western blotting analysis of p-NFAT1 proteins in WT CD4⁺ T cells stimulated with 10 μg ml⁻¹ ISO or Jo2 and with or without 0.4 μM SB203580 under T_H9-skewing conditions for 15 min (up) and statistical analysis of the relative protein levels (down). g Flow cytometric analysis of IL-9⁺ cell frequency among CD4⁺ T cells after naive CD4⁺ T cells were stimulated with 10 μg ml⁻¹ Jo2 in the presence of 0.4 μM SB203580 or 50 nM INCA-6 plus 0.4 μM SB203580 under T_H9-skewing conditions for 4 days. AU, arbitrary units; NS, not significant; *P < 0.05, **P < 0.01, and ***P < 0.001 (unpaired Student’s t test (a, c, g)). Representative results from three independent experiments are shown (mean and s.d.). Relative protein levels = the indicated protein gray values/p38 (b) or NFAT1 (d–f) gray values