Fig. 4: UDPG regulates inflammatory macrophages via P2Y₁₄ receptor.

a–c Pgm1, Ugp2 or Gys1 siRNA transfected BMDMs were stimulated with IFN-γ/LPS for 24 h, Nos2, Tnf, Il6 and Il1b expression was determined by real-time PCR. d Gys1, Pgm1 or Ugp2 siRNA transfected BMDMs were stimulated with IFN-γ/LPS for 24 h, UDPG in supernatants was determined by ELISA. e IFN-γ/LPS-stimulated BMDMs were treated with UDPG (0, 100 or 200 μM) for 24 or 36 h, Nos2, Tnf, Il6, and Il1b expression was determined by real-time PCR (left), iNOS, TNF and IL-6 expression was determined by western blot (middle) and ELISA (right). f, g Ugp2 or G6pdx siRNA transfected BMDMs were stimulated with IFN-γ/LPS ± UDPG (200 μM) for 24 or 36 h, Nos2, Tnf, Il6, and Il1b expression was determined by real-time PCR (left), iNOS, TNF and IL-6 expression was determined by western blot (middle) and ELISA (right). h Pygl or G6pdx siRNA transfected BMDMs were stimulated with IFN-γ/LPS for 24 h, UDPG in supernatants was determined by ELISA. i BMDMs were pretransfected with siRNA (Pygl or G6pdx) for 24 h prior to stimulation with IFN-γ/LPS for 6 h and switched to ¹³C-glucose for 6 h, lysed cells were analyzed by LC-MS/MS to determine m + 6-labeled UDPG. j, k P2Y₁₄ receptor expression in untreated, IFN-γ/LPS- or IL-4-treated BMDMs was determined by real-time PCR (j) and western blot (k). l, m IFN-γ/LPS-stimulated BMDMs were treated with UDPG (0, 100 or 200 μM) for 24 or 36 h, P2Y₁₄ expression was determined by real-time PCR (l) and western blot (m). n–p P2ry₁₄ siRNA transfected BMDMs were stimulated with or without IFN-γ/LPS for 24 or 36 h, Nos2, Tnf, Il6, and Il1b expression was determined by real-time PCR (n), iNOS, TNF, and IL-6 expression was determined by western blot (o) and ELISA (p). Unless otherwise specified, n = 3 biologically independent experiments were performed. Data are presented as mean ± SEM. P values were calculated using one-way ANOVA. **p < 0.01, ****p < 0.0001.