Fig. 4: Role of OGDH in gut homeostasis.
From: Metabolic adaptations direct cell fate during tissue regeneration
a,b, ABP and immunofluorescence for lysozyme, GFP and cleaved caspase 3 (CC3) in intestinal tissue from TRE-shRenCag-rtTA3 and TRE-shOgdhCag-rtTA3 mice (a) and vehicle-treated and DM-αKG-treated mice (b) at the indicated time points. c, Heat map depicting time-course quantification (D indicates day) of OGDH (fluorescence intensity), BrdU (positive cells), Spdef (mRNA levels), lysozyme (Lyz) (positive cells) and CC3 (positive cells) in intestinal crypts from doxycycline-treated TRE-shRenCag-rtTA3 and TRE-shOgdhCag-rtTA3 mice or DM-αKG-treated C57Bl/6 mice (n = 5 per group). d,e, ABP and immunofluorescence for OGDH, lysozyme (Paneth cells), HNF4α (enterocytes) and ACE2 (mature enterocytes) (d), and CC3 (cell death) (e) in intestinal sections from TRE-shOgdhCag-rtTA3 mice concomitantly treated with or without DM-succinate and doxycycline for 6–7 days. Data are representative of shRen n = 3, shRen + succinate n = 3, shOgdh n = 8 and shOgdh + succinate n = 4 mice. f, Quantification from e. Each dot represents one crypt or villus for lysozyme and CC3 and one mouse for HNF4α. a.u., arbitrary units. g, Immunofluorescence for 5hmC and HNF4α in crypts from TRE-shRenCag-rtTA3, TRE-shOgdhCag-rtTA3 and DM-αKG-treated mice. Dashed lines outline crypts (top) and 5hmC+HNF4α− cells (bottom). h, Top, enzyme-linked immunosorbent assay (ELISA) of crypt lysates to measure intestinal 5hmC abundance in TRE-shRenCag-rtTA3, TRE-shOgdhCag-rtTA3, DM-αKG-treated and vehicle-treated mice. Each dot represents one mouse (shRen n = 4, shOgdh n = 9, vehicle n = 4, αKG n = 6). Bottom, relative 5hmC levels in the Spdef promoter within isolated crypts from shRenCag-rtTA3, TRE-shOgdhCag-rtTA3 and DM-αKG-treated mice, measured by qPCR at the indicated time points. Each dot represents one mouse (n ≥ 3 mice per group). i, Venn diagrams of upregulated (red) and downregulated (blue) genes in TRE-shOgdhCag-rtTA3 and αKG-treated mice versus controls (TRE-shRenCag-rtTA3 and vehicle-treated), with gene ontology (GO) analysis of the overlapping genes. Abs(score), absolute value of enrichment score. This figure is adapted from our published patent (WO2024229094A1)50. Data are mean ± s.e.m. Statistical significance was determined using one-way ANOVA followed by Tukey’s HSD test in f and two-tailed t-test in h.
