Recovery of Bacteroides thetaiotaomicron ameliorates hepatic steatosis in experimental alcohol-related liver disease - PubMed (original) (raw)

. 2022 Jan-Dec;14(1):2089006.

doi: 10.1080/19490976.2022.2089006.

Christoph Grander 1, Felix Grabherr 1, Lisa Mayr 1, Barbara Enrich 1, Julian Schwärzler 1, Marcello Dallio 1 [ 3](#full-view-affiliation-3 "Department of Precision Medicine, University of Campania "Luigi Vanvitelli", Naples, Italy."), Vidyasagar Naik Bukke 2, Archana Moola 2, Antonio Moschetta 4, Timon E Adolph 1, Carlo Sabbà 4, Gaetano Serviddio 2, Herbert Tilg 1

Affiliations

Moris Sangineto et al. Gut Microbes. 2022 Jan-Dec.

Abstract

Alcohol-related liver disease (ALD) is a major cause of liver disease and represents a global burden, as treatment options are scarce. Whereas 90% of ethanol abusers develop alcoholic fatty liver disease (AFLD), only a minority evolves to steatohepatitis and cirrhosis. Alcohol increases lipogenesis and suppresses lipid-oxidation implying steatosis, although the key role of intestinal barrier integrity and microbiota in ALD has recently emerged. Bacteroides thetaiotaomicron (Bt) is a prominent member of human and murine intestinal microbiota, and plays important functions in metabolism, gut immunity, and mucosal barrier. We aimed to investigate the role of Bt in the genesis of ethanol-induced liver steatosis. Bt DNA was measured in feces of wild-type mice receiving a Lieber-DeCarli diet supplemented with an increase in alcohol concentration. In a second step, ethanol-fed mice were orally treated with living Bt, followed by analysis of intestinal homeostasis and histological and biochemical alterations in the liver. Alcohol feeding reduced Bt abundance, which was preserved by Bt oral supplementation. _Bt_-treated mice displayed lower hepatic steatosis and triglyceride content. Bt restored mucosal barrier and reduced LPS translocation by enhancing mucus thickness and production of Mucin2. Furthermore, Bt up-regulated Glucagon-like peptide-1 (GLP-1) expression and restored ethanol-induced Fibroblast growth factor 15 (FGF15) down-regulation. Lipid metabolism was consequently affected as Bt administration reduced fatty acid synthesis (FA) and improved FA oxidation and lipid exportation. Moreover, treatment with Bt preserved the mitochondrial fitness and redox state in alcohol-fed mice. In conclusion, recovery of ethanol-induced Bt depletion by oral supplementation was associated with restored intestinal homeostasis and ameliorated experimental ALD. Bt could serve as a novel probiotic to treat ALD in the future.

Keywords: Alcohol-related liver disease; Bacteroides thetaiotaomicron; intestinal barrier; microbiota; mitochondria; steatosis.

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Conflict of interest statement

No potential conflict of interest was reported by the author(s).

Figures

Figure 1.

Figure 1.

Ethanol depletes Bt. (a) Design of Lieber-DeCarli model. (b) Serum ALT levels (Pair fed = n6; EtOH = n10). (c, d) Histological determination of hepatic steatosis with representative pictures of H&E staining (n = 5 per group). (e) Liver triglyceride content (Pair fed = n5; EtOH = n6). (f) Quantification of Bt DNA in feces measured by qPCR (n = 5 per group). (g) Quantification of Bt number (expressed in percentage) in LYBHI medium supplemented with different ethanol concentrations. Data are expressed in mean ± SEM; *p < .05, **p < .01, ***p < .001, ****p < .0001 according to two-tails student’s t-test.

Figure 2.

Figure 2.

Bt supplementation ameliorates alcoholic fatty liver disease. (a) Experimental design. (b) Quantification of Bt DNA in feces measured by qPCR (Pair fed groups = n4-5; EtOH groups = n5). (c, d) Histological determination of hepatic steatosis with representative pictures of H&E staining (Pair fed groups = n6; EtOH groups = n9). (e) Liver Triglyceride content in EtOH fed mice (n = 9–10). Data are expressed in mean ± SEM; *p < .05; **p < .01; ***p < .001, ****p < .0001 according to one-Way ANOVA followed by post hoc analysis (Bonferroni test) or two-tails student’s t-test.

Figure 3.

Figure 3.

Bt recovery restores mucus production in the colon. (a, b) Quantification of mucus thickness in colon sections stained with periodic acid–Schiff reaction (Pair fed = n4; EtOH groups = n9-10). (c) Number of goblet cells per crypt and (d, e) goblet cells diameter identified by periodic acid–Schiff reaction. (f) Serum LPS concentration (Pair fed groups = n6; EtOH groups = n6-8). Data are expressed in mean ± SEM; *p < .05; **p < .01; ***p < .001, ****p < .0001 according to one-Way ANOVA followed by post hoc analysis (Bonferroni test) or two-tails student’s t-test.

Figure 4.

Figure 4.

Bt promotes mucin2 production via downregulation of claudin1-Notch axis (a) Colonic expression of muc1 and muc2 fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n6; EtOH groups = n10). (b) Muc2 immunoreactivity (red) with representative confocal microscope image of murine colon in EtOH-fed mice. DAPI, blue. (c-d) Colonic expression of Claudin1 and MMP9 fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n5-6; EtOH groups = n9-10). (e) Representative pictures of protein levels of actin, ERK and pERK determined by western blot analysis (EtOH groups = n5). (f) Colonic expression of Hes1 fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n5-6; EtOH groups = n9-10). Data are expressed in mean ± SEM; *p < .05; **p < .01; ***p < .001, ****p < .0001 according to one-Way ANOVA followed by post hoc analysis (Bonferroni test).

Figure 5.

Figure 5.

Bt modulated GLP1 and FGF15 expression. (a) Serum levels of FGF15 and ileal expression of FGF15, IBABP, Ost-α and Ost-β fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n4-6; EtOH groups = n6-10). (b) Serum levels of GLP-1 and colonic expression of GLP-1, TGR5, GPR43 and PYY fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n4-6; EtOH groups = n6-10). Data are expressed in mean ± SEM; *p < .05; **p < .01; ***p < .001, according to one-Way ANOVA followed by post hoc analysis (Bonferroni test).

Figure 6.

Figure 6.

Bt reduced hepatic FA synthesis (a) Liver expression of FGFR4 fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n5-6; EtOH groups = n9-10). (b) Representative pictures of protein levels of actin, AMPK, pAMPK, SREBP-1c, and nuclear SREBP-1c, determined by western blot analysis and expression of SREBP-1c determined by qPCRin the liver (for western blot: Pair fed groups = n3; EtOH groups = n5; for qPCR: Pair fed groups = n5-6; EtOH groups = n9-10). (c, d) Liver expression of FASN and SCD-1 fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n4; EtOH groups = n8-10). Data are expressed in mean ± SEM; *p < .05, according to one-Way ANOVA followed by post hoc analysis (Bonferroni test).

Figure 7.

Figure 7.

Bt improved hepatic mitochondrial fitness and function (a) Liver expression of PPAR-α, TFAM, PGC-1α, PGC-1β fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n4-5; EtOH groups = n6-7). (b) Representative pictures of protein levels of actin, respiratory chain complexes I, II and V determined by western blot analysis (Pair fed groups = n3; EtOH groups = n5), and and hepatic ATP content (Pair fed groups = n4; EtOH groups = n5). (c) Liver expression of CPT-1a fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n4; EtOH groups = n5); and representative pictures of protein levels of actin and VDAC1determined by western blot analysis (Pair fed groups = n3; EtOH groups = n5). (d) Serum levels of Triglycerides (Pair fed groups = n5; EtOH groups = n9), and liver expression of DGAT1 fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n4-5; EtOH groups = n9-10). (e) Hepatic SOD enzymatic activity (Pair fed groups = n4; EtOH groups = n4) and expression of PRDX-3 and −5 fold over Pair fed vehicle group and determined by qPCR (Pair fed groups = n5-6; EtOH groups = n9); and serum MDA- and HNE- protein adducts (Pair fed groups = n4; EtOH groups = n4). Data are expressed in mean ± SEM; *p < .05, **p < .01; ***p < .001, ****p < .0001 according to one-Way ANOVA followed by post hoc analysis (Bonferroni test).

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