Fig. 3.

Inhibitory effects of MPQP on LPS-induced NF-κB and MAPK pathways. RAW 264.7 macrophages were pre-incubated with MPQP (1, 2, 5, and 10 μM) for 2 h and stimulated with LPS (1 μg/ml) for 5 min (for detection of IκBα, IKKα/β, and IRAK1) or 15 min (for detection MAPKs, MKK3/6, and MKK4). Total cell lysates were prepared and analyzed by immunoblot analysis. The expression levels of (A) p-IκBα, IκBα, (B) p-JNK, JNK, p-ERK, ERK, p-p38, p38, (C) p-MKK3/6, MKK3/6, p-MKK4, MKK4, (D) IRAK1, p-IKKα/β, and IKKα/β were detected using specific antibodies. The relative expression levels of p-IκBα, IκBα, and IRAK1 were normalized to the α-tubulin levels. The phosphorylation levels of MAPKs, MKK3/6, MKK4, and IKKα/β were normalized to the corresponding MAPKs, MKK3/6, MKK4, and IKKα/β levels. Quantitative analyses of phosphorylation and protein levels are shown as bar graphs after normalization. All bar graphs are represented as the mean ± SEM and analyzed using one-way ANOVA from three independent experiments. ^#P < 0.0001 vs. LPS-untreated control groups. ^aP < 0.01 and ^bP < 0.001 vs LPS-treated groups.