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. 2019 Nov 8;10(3):462–474. doi: 10.1016/j.apsb.2019.11.004

Figure 4.

Figure 4

BA inhibition of IL-1β expression via down-regulation of IKK, ERK and P38. (A) RAW264.7 cells were transfected for 24 h with 0.45 μg of pNF-κB-Luc (NF-κB reporter plasmid) or pAP-1-Luc (AP-1 reporter plasmid) and 0.15 μg of pRL-SV40 (internal control). The cells were treated with RANKL for 24 h in the absence or presence of 10 μmol/L BA. The luciferase activity of each cell lysate was measured using a dual-luciferase assay system. The activity of firefly luciferase was normalized to that of the Renilla enzyme and expressed as fold increase relative to the activity of RANKL-untreated cells. (B–H) BMMs were cultured in the presence of M-CSF and were treated with RANKL for 24 h (B and C), the indicated times (D, E and G), 15 min (F) and 48 h (H) in presence of 10 μmol/L BA or 10 μmol/L inhibitors of ERK (U0126), P38 (SB202190), JNK (SP600125) and IKK (SC-514) (G). Cell lysates were subjected to immunoblotting analysis (C–E and G). The cells were stained with p65 antibody and DAPI, and then photographed under a fluorescence microscope. Scale bar, 10 μm (F). The transcription of individual genes was quantified by real-time PCR and presented as fold induction (B and H). ***P < 0.001 between the indicated groups (A, B and H) or vehicle vs. inhibitors in the presence of RANKL (H). ns, not significant.