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. 2018 Mar 5;15(8):782–793. doi: 10.1038/cmi.2017.163

Figure 4.

Figure 4

The specific phenotype of IL-33-treated neutrophils is mediated by JNK and NF-κB signaling pathways. (a) Neutrophils freshly isolated from bone marrow were treated with IL-33 for 15, 30 and 120 min, respectively. The activities of JNK, p38, STAT1, Erk and p65 in cytoplasm were detected by western blotting. (b) The c-jun and p65 protein levels in nucleus of neutrophils after IL-33 treatment for 1 and 2 h, respectively, were determined by western blotting. (c and d) Neutrophils were pretreated with the indicated concentrations of JNK inhibitor (SP600125) and NF-κB inhibitor for 30 min and then stimulated with IL-33 for 24 h. The levels of IL-9, CCL2, CCL7, CCL12, IL-1R2 and IL-13Ra1 expression were detected by real-time PCR. (e and f) The concentrations of IL-9, IL-4, IL-5, IL-13, CCL2 and CCL7 in the supernatant of neutrophils pretreated with either SP600125 or NF-κB inhibitor and then cultured with IL-33 for 24 h were determined by ELISA assay kits. Assays were performed more than three times. Data are shown as mean±s.d. (n=3). *P<0.05, **P<0.01 and ***P<0.001 compared with control. CCL, C-C motif chemokine ligand; ELISA, enzyme-linked immunosorbent assay; ERK, extracellular signal–regulated kinase; IL, interleukin; JNK, c-Jun N-terminal kinase; NF, nuclear factor; STAT, signal transducer and activator of transcription factor.