Fig. 8: Propionate and butyrate inhibit MAPK signaling pathway in macrophages.

a–d RAW 264.7 cells were pre-treated with or without sodium propionate (SP) at concentrations of 1.6 mM, 3.2 mM, and 6.4 mM, and sodium butyrate (SB) at concentrations of 0.2 mM, 0.4 mM, and 0.8 mM for 24 hours. Subsequently, the corresponding metabolites were combined with 1 μg/mL LPS and used to treat the cells for an additional 6 hours. The RAW 264.7 cells were then collected for western blotting and immunofluorescent staining, and the supernatants were collected for enzyme-linked immunosorbent assay (ELISA). Representative images showing immunofluorescence staining of CD86⁺ cells and CD206⁺ cells in RAW 264.7 cells (a, c), with bar plots displaying their quantitative analysis (b, d) in different groups (n = 3 independent experiments). e, f Bar plots showing the levels of IL-6, TNF-α, and IL-8 in the supernatant of RAW 264.7 medium determined by ELISA in different groups (n = 3 independent experiments). g, h Representative images showing the relative levels of p-JNK, p-ERK1/2, p-p38, and p-p65 in RAW 264.7 cells determined by western blotting in different groups (n = 3 independent experiments). i–l RAW 264.7 cells were pre-treated with SP (6.4 mM), SB (0.8 mM), and U46619 (10 μM, a p38 MAPK agonist) for 24 hours, and then were combined with 1 μg/mL LPS and used to treat the cells for another 6 hours. Representative images show immunofluorescence staining of CD86⁺ cells and CD206⁺ cells in RAW 264.7 cells (i, k), with bar plots displaying their quantitative analysis (j, l) in different groups (n = 3 independent experiments). Data are presented as mean ± SD. Statistical comparisons were performed using one-way ANOVA with Dunnett’s test for statistical significance. Scale bar, 50 μm. Source data are provided as a Source Data file. LPS lipopolysaccharide, CD86 cluster of differentiation 86, DAPI 4’,6-diamidino-2-phenylindole, IL-6 Interleukin-6, TNF-α tumor necrosis factor-α, p- phospho-, GAPDH glyceraldehyde-3-phosphate dehydrogenase.