Alpha-Ketoglutarate Promotes Goblet Cell Differentiation and Alters Urea Cycle Metabolites in DSS-Induced Colitis Mice

Abstract

The metabolite, alpha-ketoglutarate (aKG), shows promise as an approach for ameliorating colitis, but much remains unknown about the full extent of its effects on the metabolome and mucosal barrier. To further elucidate this matter, C57BL/6 male mice received drinking water with or without 1% aKG for three weeks, then were subjected to 2.5% dextran sulfate sodium (DSS) induction for 7 days followed by 7 days of recovery. Cecal content and intestinal tissue samples were analyzed for changes in metabolite profile and signaling pathways. Gas chromatography-mass spectrometry (GC-MS) metabolomics revealed a separation between the metabolome of mice treated with or without aKG; putrescine and glycine were significantly increased; and ornithine and amide products, oleamide and urea were significantly decreased. Based on a pathway analysis, aKG treatment induced metabolite changes and enriched glutathione metabolism and the urea cycle. Additionally, signaling pathways committing epithelial cells to the secretory lineage were elevated in aKG-treated mice. Consistently, aKG supplementation increased goblet cells staining, mRNA expression of mucin 2, and, trefoil factor 3 and Krüppel-like factor 4, markers of goblet cell differentiation. These data suggest the ameliorating the effects of aKG against chemically induced colitis involves a reduction in harmful metabolites and the promotion of goblet cell differentiation, resulting in a more-fortified mucus layer.

Keywords: alpha-ketoglutarate; colitis; goblet cells; metabolome; mucin.

Figure 1

PCA of the cecal metabolome of dextran sulfate sodium (DSS)-induced colitis mice receiving water with or without alpha-ketoglutarate (aKG). CON: control mice without aKG supplementation in drinking water; aKG: mice receiving 1% aKG. n = 7–8 mice per group.

Figure 2

Cecal metabolite profiles in dextran sulfate sodium (DSS)-induced colitis mice receiving water with or without alpha-ketoglutarate. Fold changes related to CON, #: p < 0.10, *: p < 0.05, **: p < 0.01.

Figure 3

Pathway enrichment analysis of the differential metabolites in cecal samples of dextran sulfate sodium (DSS)-induced colitis mice receiving water with or without alpha-ketoglutarate.

Figure 4

Goblet cells in dextran sulfate sodium (DSS)-induced colitis mice supplemented with or without alpha-ketoglutarate (aKG). (A) Representative Alcian blue staining; (B) Goblet cell density; (C) mRNA expression of Muc2; (D) mRNA expression of goblet cell differentiation markers. CON: control mice without aKG; aKG: mice receiving 1% aKG; Mean ± SEM, n = 4–7 mice per group. *: p < 0.05.

Figure 5

Alpha-ketoglutarate (aKG) supplementation upregulates signaling transcription factors committing cells to the secretory cell lineage. (A) MATH1 immunoblotting; (B) mRNA expression of Notch1 and Hes1. CON: control mice without aKG; aKG: mice receiving 1% aKG in drinking water. Mean ± SEM, n = 7–8 mice per group. #: p < 0.10, *: p < 0.05.