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

Authors: HERNANDEZ, GABRIELLA - California Polytechnic State University; SMITH, VICTORIA - California Polytechnic State University; MELYNK, MEGAN - California Polytechnic State University; BURD, MATTHEW - California Polytechnic State University; SPRAYBERRY, KIMBERLY - California Polytechnic State University; EDWARDS, MARK - California Polytechnic State University; PETERSON, DANIEL - California Polytechnic State University; BENNET, DARIN - California Polytechnic State University; FANTER, ROB - California Polytechnic State University; COLUMBUS, DANIEL - Prairie Swine Centre; STEIBEL, JUAN - Michigan State University; GLANZ, HUNTER - California Polytechnic State University; IMMOOS, CHAD - California Polytechnic State University; RICE, MARGARET - California Polytechnic State University; SANTIAGO-RODRIGUEZ, TASHA - California Polytechnic State University; BLANK, JASON - California Polytechnic State University; VANDERKELEN, JENNIFER - California Polytechnic State University; KITTS, CHRISTOPHER - California Polytechnic State University; PICCOLO, BRIAN - Arkansas Children'S Nutrition Research Center (ACNC); LA FRANO, MICHAEL - California Polytechnic State University; Burrin, Douglas - Doug; MAJ, MAGDALENA - California Polytechnic State University; MANJARIN, RODRIGO - California Polytechnic State University

Submitted to: American Journal of Physiology - Gastrointestinal and Liver Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/25/2020
Publication Date: 3/1/2020
Citation: Hernandez, G.V., Smith, V.A., Melynk, M., Burd, M.A., Sprayberry, K.A., Edwards, M.S., Peterson, D.G., Bennet, D.C., Fanter, R.K., Columbus, D.A., Steibel, J., Glanz, H., Immoos, C., Rice, M.S., Santiago-Rodriguez, T.M., Blank, J.M., Vanderkelen, J.J., Kitts, C.L., Piccolo, B.D., La Frano, M.R., Burrin, D.G., Maj, M., Manjarin, R. 2020. Dysregulated FXR-FGF19 signaling and choline metabolism is associated with gut dysbiosis and hyperplasia in a novel pig model of pediatric NASH. American Journal of Physiology - Gastrointestinal and Liver Physiology. 318(3):G582-G609. https://doi.org/10.1152/ajpgi.00344.2019.
DOI: https://doi.org/10.1152/ajpgi.00344.2019

Interpretive Summary: Non-alcoholic fatty liver disease (NAFLD) represents the major cause of pediatric chronic liver pathology in the United States. The cause of NAFLD in children has been linked to excessive consumption of a nutrient rich diet, but it is not clear how specific nutrients lead to liver disease. Most animal studies of NAFLD have used adult mice that do not model the effects of how early nutrition may cause liver disease in children. Feeding diets high in fat and sugar have been shown to trigger increased bile secretion from the liver for the digestion of fat. The impact of increased bile secretion on gut function and whether this contributes to liver disease in young children is poorly understood because there are not good animal models to test this question. The aim of this study was to develop a new model of pediatric NAFLD using a novel strain of Iberian pigs that have genetic mutation that increases their propensity to overeat and become obese. We used the novel Iberian pig model to test whether feeding a diet high in fat and sugar during infancy would lead to signs of NAFLD after three months. Our results confirmed that feeding a high fat diet to juvenile pigs resulted in increased blood markers of disease and accumulation of fat and tissue injury in the liver similar to that found in pediatric NAFLD. We also found that juvenile pigs fed a high fat diet caused depletion of key nutrients, choline and phospholipids, in the liver that are secreted in bile and important for fat metabolism. This study establishes an important new animal model using the Iberian pig that can be used for future studies to establish how nutrition during childhood leads to pediatric fatty liver disease.

Technical Abstract: To investigate the role of bile acids (BA) in the pathogenesis of diet-induced non-alcoholic steatohepatitis (NASH), we fed a "Western-style diet" (HFF) enriched with fructose, cholesterol and saturated fat for 10 weeks 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, while secondary BAs and choline-containing phospholipids in colon may have promoted dysbiosis, hyperplasia and trimethylamine synthesis, causing further damage to the liver.