Journal: The Febs Journal

Article Title: Butyrate suppresses mucosal inflammation in inflammatory bowel disease primarily through HDAC3 inhibition in monocytes and macrophages

doi: 10.1111/febs.70289

Figure Lengend Snippet: Butyrate induces MCT1 and IL‐18 mRNA expression in human colonic organoids without affecting inflammatory mediators and tight junction gene expression. (A) mRNA levels of colonic organoid cell markers cultured in expansion medium (EM) or differentiation medium (DM) ( n = 3). (B) Bright‐field images of colonic organoids cultured in DM and treated with 2 m m butyrate in the presence or absence of an inflammatory cytokine mix (CM) (5 ng·mL −1 of TNF‐α, IL‐1β, and IFN‐γ), for 6 h (objective 10×, scale bar: 250 μm). (C) Lactate dehydrogenase (LDH) release from organoids was measured in conditioned medium following treatments ( n = 5 or n = 6). (D) mRNA levels of SLC16A1 ( n = 6) and (E) representative image and quantification of MCT1 protein expression ( n = 3, objective 40×, scale bar: 100 μm, quantification of image resolution: 0.30 μm per pixel) were determined by TaqMan RT‐qPCR and immunofluorescence in PFA‐fixed/paraffin‐embedded organoids, respectively. (F) mRNA levels of NOS2 , TNFA , IL1B , CXCL8 , IRF1 , TFF1 , DUOX2 , LCN2 , IL18 , OCLN , CLDN3 , and TJP1 ( n = 4 or n = 6) were determined by RT‐qPCR. Statistical analysis: Paired t ‐test (A, D), F (except NOS2 ) and Wilcoxon matched‐pairs signed‐rank (F, NOS2 ). * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: Gene expression was analyzed using TaqMan Gene Expression Assays for SLC16A1 (Hs01560299_m1), HCAR2 (Hs02341584_s1), HDAC3 (Hs00187320_m1), IRF1 (Hs00971965_m1), TFF1 (Hs00907239_m1), DUOX2 (Hs00204187_m1), and LCN2 (Hs01008571_m1), MARCKS (Hs00158993_m1), and CD48 (Hs00381156_m1) (Applied Biosystems, Waltham, MA, USA) or TaqMan primers and probes from Eurogentec (Maastricht, The Netherlands; see Table ).

Techniques: Expressing, Gene Expression, Cell Culture, Quantitative RT-PCR, Immunofluorescence