Dysregulated FXR-FGF19 signaling and choline metabolism are associated with gut dysbiosis and hyperplasia in a novel pig model of pediatric NASH

Abstract

To investigate the role of bile acids (BAs) in the pathogenesis of diet-induced nonalcoholic steatohepatitis (NASH), we fed a “Western-style diet” [high fructose, high fat (HFF)] enriched with fructose, cholesterol, and saturated fat for 10 wk to juvenile Iberian pigs. We also supplemented probiotics with in vitro BA deconjugating activity to evaluate their potential therapeutic effect in NASH. Liver lipid and function, cytokines, and hormones were analyzed using commercially available kits. Metabolites, BAs, and fatty acids were measured by liquid chromatography-mass spectrometry. Histology and gene and protein expression analyses were performed using standard protocols. HFF-fed pigs developed NASH, cholestasis, and impaired enterohepatic Farnesoid-X receptor (FXR)-fibroblast growth factor 19 (FGF19) signaling in the absence of obesity and insulin resistance. Choline depletion in HFF livers was associated with decreased lipoprotein and cholesterol in serum and an increase of choline-containing phospholipids in colon contents and trimethylamine- N-oxide in the liver. Additionally, gut dysbiosis and hyperplasia increased with the severity of NASH, and were correlated with increased colonic levels of choline metabolites and secondary BAs. Supplementation of probiotics in the HFF diet enhanced NASH, inhibited hepatic autophagy, increased excretion of taurine and choline, and decreased gut microbial diversity. In conclusion, dysregulation of BA homeostasis was associated with injury and choline depletion in the liver, as well as increased biliary secretion, gut metabolism and excretion of choline-based phospholipids. Choline depletion limited lipoprotein synthesis, resulting in hepatic steatosis, whereas secondary BAs and choline-containing phospholipids in colon may have promoted dysbiosis, hyperplasia, and trimethylamine synthesis, causing further damage to the liver. NEW & NOTEWORTHY Impaired Farnesoid-X receptor (FXR)-fibroblast growth factor 19 (FGF19) signaling and cholestasis has been described in nonalcoholic fatty liver disease (NAFLD) patients. However, therapeutic interventions with FXR agonists have produced contradictory results. In a swine model of pediatric nonalcoholic steatohepatitis (NASH), we show that the uncoupling of intestinal FXR-FGF19 signaling and a decrease in FGF19 levels are associated with a choline-deficient phenotype of NASH and increased choline excretion in the gut, with the subsequent dysbiosis, colonic hyperplasia, and accumulation of trimethylamine- N-oxide in the liver.

Gut Microbiota Metabolites in NAFLD Pathogenesis and Therapeutic Implications

“…Choline can be converted by the gut microbiota into trimethylamine (TMA) directly or via betaine, and TMA is oxidized into trimethylamine N-oxide (TMAO). In NAFLD, the conversion of choline into TMA/TMAO are increased, leading to deficiency of choline and accumulation of TMA/TMAO [ 64 , 65 ]. Choline can promote very low-density lipoprotein transportation out of the liver and deficiency of choline results in accumulation of lipids in the liver.…”

“…Arao et al (2020) used a methionine/choline-deficient diet to establish a NASH model and found that mitochondrial DNA content was decreased [ 64 ]. Hernandez et al (2020) reported that high fructose-, fat- and cholesterol feeding in pigs caused hepatic steatosis and inflammation, with impaired FXR-FGF19 signaling [ 65 ]. The severity of NAFLD in this model was associated with increased choline metabolites.…”

The Neonatal and Juvenile Pig in Pediatric Drug Discovery and Development

“…Recent reports have pointed to the microbiota [ 233 ] and dietary fat sources [ 234 ] to play an important role. More recent studies have described in more detail the biochemical and signaling pathways that go astray and lead to NAFLD in the juvenile pig [ 237 ] and provide cues on possible therapeutic interventions.…”

%polynova_2way: A SAS macro for implementation of mixed models for metabolomics data

“…An experimental dataset from a previously published study in our lab [7] was used to illustrate the utilization of the %polynova_2way SAS macro. Data is provided in S1 Data file.…”

“…Before a model fitting to one-metabolite-at-a-time can be implemented with our %polynov-a_2way macro, data must be analyzed with multivariate statistics using software such as R prcomp() function [8], MetaboAnalyst or Primer-E (v.7; Primer-E Ltd., Plymouth, UK), as previously described [7]. Briefly, liver data was imported into Primer-E, log transformed into a normal distribution approximation and analyzed with a Euclidean distance matrix.…”