CR3 and Dectin-1 Collaborate in Macrophage Cytokine Response through Association on Lipid Rafts and Activation of Syk-JNK-AP-1 Pathway
Fig 5
AP-1, but not NF-κB, mediates the collaborative cytokine response upon CR3 and Dectin-1 ligation.
FLDMs from Syk+/+, Syk-/- and Syk+/- embryos (A) and macrophages from WT, Itgam-/-, Clec7a-/- and Itgam-/-Clec7a-/- mice (B and C) were stimulated with or without (0 min) HK H. capsulatum for 30 and 60 min. Cell lysates were analyzed by Western blotting. The blot shown in (A) is the same one shown in Fig 4B. Macrophages from WT mice (D and E), and Itgam-/- and Clec7a-/- mice (F) were stimulated with or without iC3b-LB, dZ, LB, or the combination of iC3b-LB and dZ for 30 min (E) or 60 min (D and F). Cell lysates were analyzed by Western blotting. Data are representative of at least 3 independent experiments (A-F). (G and H) Macrophages from WT mice were transfected with small interfering RNA (siRNA) against c-Fos or c-Jun, followed by stimulation with HK H. capsulatum 48 h later. Cell lysates and culture supernatants were collected at 6 h after stimulation. Silencing of c-Fos and c-Jun was confirmed by Western blotting (G). TNF and IL-6 levels in culture supernatants were quantified by ELISA and are presented as the mean ± SD of relative levels of TNF and IL-6 (n = 3 from 3 independent experiments) (H). ** p ≦ 0.01, *** p ≦ 0.001 [2-tailed t-test].