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Brazilin Suppresses Inflammation via the Down-regulation of IRAK4 in LPS-stimulated Raw264.7 Macrophage

Kye-Yoon Yoon, Kui-Jin Kim, Hyung-Sun Youn, Sei-Ryang Oh, Boo-Yong Lee

Journal of Food and Nutrition Research. 2015, 3(9), 575-580 doi:10.12691/jfnr-3-9-3
  • Figure 1. The effect of brazilin on NF-κB luciferase activity in LPS-stimulated Raw264.7 cells. (A) The chemical structure of brazilin. (B) NF-κB luciferase activity of brazilin. Cells were incubated with increasing concentration of the brazilein for 1h and then co-treated with 1 μg/mL LPS for 10 h. DMSO was used as a vehicle. Data are mean ± SD (n=3)
  • Figure 2. Brazilin down-regulated the expression of iNOS and COX2 protein in LPS stimulated Raw264.7. Raw 264.7 macrophage cells (5×105 /well) were pretreated with 10, 30, or 50 μM brazilin for 1 h and then induced with 1 μg/mL LPS for an 24 h. The expression of iNOS and COX2 protein were detected by western blotting using iNOS, COX2, and GAPDH antibodies. DMSO was used as a vehicle. Data are expressed as mean ± SD (n=3)
  • Figure 3. Brazilin decreased the nitric oxide production and phosphorylation of JNK in LPS-stimulated Raw264.7 macrophage cell. (A) The production of soluble nitrite oxide. Culture media was subsequently isolated and nitrite concentrations determined. (B) The expression of p-JNK, p-p38MAPK, pERK, and ERK in LPS-stimulated Raw264.7 macrophage cell. Cells were pretreated with 10, 30 or 50 μM brazilin for 1 h and then stimulated with 1 μg/mL LPS for 30 min or 24 h. DMSO was used as a vehicle. Equal amounts of protein in cell lysates were analyzed by Western blot. Data are mean ± SD (n=3)
  • Figure 4. LPS increased the expression of IRAK4 protein, whereas brazilin diminished the expression of IRAK4 in Raw264.7 macrophage cell. Cells were pretreated with 10, 30 or 50 μM brazilein for 1 h and then stimulated with 1 μg/mL LPS for 30 min or 24 h. DMSO was used as a vehicle. Equal amounts of protein in cell lysates were analyzed by Western blot
  • Figure 5. The transcription levels of pro-inflammatory cytokines were down-regulated by brazilin in LPS-induced Raw264.7. Cells were pretreated with 10, 30, or 50 μM brazilein for 1 h, stimulated with LPS (1 μg/mL), and incubated for a further 24 h. DMSO was used as a vehicle. Data are expressed as mean ± SD (n=3). (A) Total RNA was amplified by RT-PCR using the indicated primers for IL-1β, TNFα, and IL-6. (B-D) The relative mRNA expression of IL-1β, TNFα, and IL-6.were quantified using the ImageJ program. Data are mean ± SD (n=3)