Fig. 4

GRIM19 regulates Treg/Th17 ratio via inhibiting STAT3 pathway. a Recipients (B/c mice) were intravenously injected with 5 × 10⁶ donor (WT or GRIM19 Tg) bone marrow cells and 1 × 10⁷ WT or GRIM19 Tg splenocytes after lethal irradiation. Intracellular immunostaining was performed for FOXP3⁺ Treg (CD4⁺CD25⁺) cells in spleen and LN. Data in the left panel are representatives of three independent experiments. Bars show the mean ± SD of at least three independent experiments. b Fourteen days after BMT, isolated spleens were analyzed by confocal microscopy for the expression of GITR and PD-1 among CD4⁺CD25⁺Foxp3⁺ regulatory T cells. Bars represent the SD of six mice per group. c The expression of signal transducer phosphorylated STAT5 in the spleens was analyzed by confocal microscopy on day 14 after BMT. Data are representatives of three independent experiments. d Splenic CD4⁺ T cells were sorted from B6 mice and transfected with mock or GRIM19 vector. Mock or GRIM19-transfected T cells were activated by stimulation with anti-CD3/CD28 for 3 days. Grim19 and Foxp3 mRNA levels were determined by real-time PCR. Data are expressed as mean ± SD of three independent experiments. *p < 0.05, **p < 0.01 vs. mock. e Splenic CD4⁺ T cell isolated from WT or GRIM19 Tg mice were activated by stimulation with anti-CD3/CD28 for 3 days. A STAT5 inhibitor was added at priming. Foxp3 mRNA level was determined by real-time PCR. Data are expressed as mean ± SD of three independent experiments