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. 2021 May 18;12:655212. doi: 10.3389/fimmu.2021.655212

Figure 3.

Figure 3

In vivo DMF treatment of EAE-affected mice exerts a pro-inflammatory effect on IEC that is dependent on the activation of HCAR2/ERK1/2 pathway. (A, B) In-vivo DMF treatment of EAE-affected mice results in increased expression of pro-inflammatory cytokines by IEC, which is HCAR2-dependent. Tnf and Il1b mRNA expression in IEC isolated from naïve and EAE-affected WT (A) or HCAR2-KO (B) mice, that were treated or not with DMF. Data are presented as fold induction of gene expression in cells isolated from EAE-affected mice (treated or not with DMF) over cells isolated from naïve mice. (C, D) In EAE, the pro-inflammatory effect of DMF on IEC involves the HCAR2/ERK1/2 pathway, rather than the HCAR2/COX-2 pathway. Western blotting for COX-2 (C) and ERK1/2 (D) in IEC from naïve and EAE-affected WT (C, D) and HCAR2-KO (D) mice that were treated or not with DMF. GAPDH was assessed as loading control. One representative blot is shown and quantification by densitometric analysis of bands is presented as gly- or ungly-COX-2 over GAPDH (C), or P-ERK1/2 over tot-ERK1/2, normalized to GAPDH (D). Average clinical scores of mice at the time of the analysis are: WT-EAE-affected mice: 3 ± 0.17 (SEM); WT-EAE-affected mice treated with DMF: 2.75 ± 0.11; HCAR2-KO-EAE-affected mice: 3.1 ± 0.24; HCAR2-KO-EAE-affected mice treated with DMF: 3.05 ± 0.27. Results are shown as mean ± SEM of at least N = 3 experiments with at least N = 3 mice per group per experiment. *P < 0.05 and ***P < 0.001.