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Linking Sex Differences in Non-Alcoholic Fatty Liver Disease to Bile Acid Signaling, Gut Microbiota, and High Fat Diet

  • John Y.L. Chiang
    Correspondence
    Address correspondence to John Y.L. Chiang, Ph.D., Department of Integrative Medical Sciences, College of Medicine, Northeast Ohio Medical University, 4209 SR 44, Rootstown, OH 44272.
    Affiliations
    Department of Integrative Medical Sciences, College of Medicine, Northeast Ohio Medical University, Rootstown, Ohio
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      Sex differences in liver cancer and metabolic diseases have been recognized; males have higher prevalence and a higher death rate than females.
      • Siegel R.L.
      • Miller K.D.
      • Jemal A.
      Cancer statistics, 2016.
      Non-alcoholic fatty liver disease has emerged as a major health concern worldwide. Non-alcoholic fatty liver disease covers a spectrum of liver diseases from simple steatosis to non-alcoholic steatohepatitis, which is characterized by hepatic inflammation and fibrosis. Some non-alcoholic steatohepatitis patients develop to hepatocellular carcinoma and require liver transplant.
      Bile acids are steroid-derived amphipathic molecules that act as detergents for absorption of nutrients, steroids, and drugs in the intestine, and also as serve as signaling molecules that activate nuclear farnesoid X receptor (FXR) and the membrane G-protein bile acid receptor-1 (Gpbar-1, aka TGR5).
      • Chiang J.Y.
      Bile acids: regulation of synthesis.
      Bile acid signaling plays a critical role in regulation of metabolic homeostasis via the gut-to-liver axis. Bile acids activate FXR to regulate lipid, glucose, and energy metabolism. Deficiency of FXR impairs bile acid and lipoprotein metabolism and aggravates Western high fat diet (WD)–induced obesity and diabetes. In this issue of The American Journal of Pathology, Jena et al
      • Jena P.K.
      • Sheng L.
      • Liu H.-X.
      • Kalanetra K.M.
      • Mirsoian A.
      • Murphy W.J.
      • French S.W.
      • Krishnan V.V.
      • Mills D.A.
      • Wan Y.-J.Y.
      Western diet–induced dysbiosis in farnesoid X receptor knockout mice causes persistent hepatic inflammation after antibiotic treatment.
      reported a comprehensive study of WD-induced dysbiosis in both male and female Fxr−/− mice compared to wild-type mice.
      In the intestine, activation of FXR induces fibroblast growth factor 15 (FGF15, or FGF19 in humans) to inhibit bile acid synthesis in the liver. Intestinal bacteria bile salt hydrolases de-conjugate taurine-conjugated bile acids followed by bacteria 7α-dehydroxylase activity, which converts the primary bile acids, cholic acid and chenodeoxycholic acid, to secondary bile acids, deoxycholic acid and lithocholic acid, respectively. In humans, taurine- and glycine-conjugated cholic acid, chenodeoxycholic acid, and deoxycholic acid are the major bile acids in a highly hydrophobic bile acid pool, whereas in mice taurine chenodeoxycholic acid is converted to Tα-muricohlic acid and Tβ-muricholic acid, and taurine cholic acid (TCA) and Tα/β-MCAs are major bile acids in a highly hydrophilic bile acid pool.
      Bile acids control gut bacteria overgrowth to protect intestinal barrier function. Among all bile acids, deoxycholic acid has the highest bactericidal activity. Ablation of the Fxr gene in mice (Fxr−/−) impairs bile acid and lipoprotein metabolism and aggravates WD-induced hepatic steatosis.
      • Sinal C.J.
      • Tohkin M.
      • Miyata M.
      • Ward J.M.
      • Lambert G.
      • Gonzalez F.J.
      Targeted disruption of the nuclear receptor FXR/BAR impairs bile acid and lipid homeostasis.
      WD increases biliary secretion of bile acids and reshapes the gut microbiota in obesity by increasing Firmicutes and decreasing Bacteriodetes.
      • Clarke S.F.
      • Murphy E.F.
      • Nilaweera K.
      • Ross P.R.
      • Shanahan F.
      • O'Toole P.W.
      • Cotter P.D.
      The gut microbiota and its relationship to diet and obesity: new insights.
      Feeding a cholic acid–containing diet increases the gut Firmicutes to Bacteriodetes ratio, which is also increased in obese mice.
      • Islam K.B.
      • Fukiya S.
      • Hagio M.
      • Fujii N.
      • Ishizuka S.
      • Ooka T.
      • Ogura Y.
      • Hayashi T.
      • Yokota A.
      Bile acid is a host factor that regulates the composition of the cecal microbiota in rats.
      Dietary saturated fats increase TCA and promote bile-tolerant and sulfur-producing Bilophila wadsworthia to increase pro-inflammatory cytokines and colitis.
      • Devkota S.
      • Wang Y.
      • Musch M.W.
      • Leone V.
      • Fehlner-Peach H.
      • Nadimpalli A.
      • Antonopoulos D.A.
      • Jabri B.
      • Chang E.B.
      Dietary-fat-induced taurocholic acid promotes pathobiont expansion and colitis in Il10-/- mice.
      Consistently, an animal based–diet increases abundance of B. wadsworthia and Bacteroides, and decreases abundance of Firmicutes.
      • David L.A.
      • Maurice C.F.
      • Carmody R.N.
      • Gootenberg D.B.
      • Button J.E.
      • Wolfe B.E.
      • Ling A.V.
      • Devlin A.S.
      • Varma Y.
      • Fischbach M.A.
      • Biddinger S.B.
      • Dutton R.J.
      • Turnbaugh P.J.
      Diet rapidly and reproducibly alters the human gut microbiome.
      Obese and non-alcoholic fatty liver disease patients also have reduced Firmicutes and increased Proteobacteria,
      • Turnbaugh P.J.
      • Backhed F.
      • Fulton L.
      • Gordon J.I.
      Diet-induced obesity is linked to marked but reversible alterations in the mouse distal gut microbiome.
      which causes dysbiosis and contributes to liver inflammation.
      • De Minicis S.
      • Rychlicki C.
      • Agostinelli L.
      • Saccomanno S.
      • Candelaresi C.
      • Trozzi L.
      • Mingarelli E.
      • Facinelli B.
      • Magi G.
      • Palmieri C.
      • Marzioni M.
      • Benedetti A.
      • Svegliati-Baroni G.
      Dysbiosis contributes to fibrogenesis in the course of chronic liver injury in mice.
      These studies suggest a link between dietary fats, bile acids, and gut microbiota.
      Jena et al
      • Jena P.K.
      • Sheng L.
      • Liu H.-X.
      • Kalanetra K.M.
      • Mirsoian A.
      • Murphy W.J.
      • French S.W.
      • Krishnan V.V.
      • Mills D.A.
      • Wan Y.-J.Y.
      Western diet–induced dysbiosis in farnesoid X receptor knockout mice causes persistent hepatic inflammation after antibiotic treatment.
      reported a comprehensive study of WD-induced dysbiosis in both male and female Fxr−/− mice compared to wild-type mice. They found that male Fxr−/− mice had more severe hepatic inflammation and steatosis than female mice, and WD-feeding aggravates hepatic steatosis more in male than female Fxr−/− mice. To study the role of the gut microbiota in hepatic inflammation, they treated control diet– and WD-fed Fxr−/− mice with different antibiotics, Abx (ampicillin, neomycine, metronidazole, and vancomycin), vancomycin, and polymyxin B. Abx eliminated hepatic neutrophils and lymphocytes in control diet, but not in WD-fed Fxr−/− mice. Vancomycin and polymyxin B reduced hepatic lymphocytes. These antibiotics have differential effect on reducing inflammatory and fibrogenic gene mRNA expression in control diet– and WD-fed mice. Gut microbiome analysis showed that Fxr−/− mice had reduced Firmicutes and increased Proteobacteria, which could be reversed by Abx, but Proteobacteria and Bacteroides persisted in WD-fed Fxr−/− mice. Interestingly, they found that reducing hepatic inflammation by antibiotics was associated with decreased free and conjugated secondary bile acids, and also caused changes in gut microbiota. The same group has recently reported that the sex differences in WD-induced hepatic steatosis, insulin resistance, bile acids, and microbiota profiles are all FXR-dependent.
      • Sheng L.
      • Jena P.K.
      • Liu H.-X.
      • Kalanetra K.M.
      • Gonzalez F.J.
      • French S.W.
      • Krishnan V.V.
      • Mills D.A.
      • Wan Y.-J.Y.
      Gender differences in bile acids and microbiota in relationship with gener dissimilarity in steatosis induced by diet and FXR inactivation.
      Male WD-fed Fxr−/− mice had more severe insulin resistance that may contribute to higher severity of hepatic steatosis compared to female mice. Fxr deficiency increases TCA and T-βMCA, and the gut microbiota profiles in male WD-fed Fxr−/− mice are distinctly different from control diet–fed Fxr−/− mice. WD also shifts the gut microbiota in Fxr−/− mice by decreasing Firmicutes and increasing Proteobacteria.
      • Turnbaugh P.J.
      • Backhed F.
      • Fulton L.
      • Gordon J.I.
      Diet-induced obesity is linked to marked but reversible alterations in the mouse distal gut microbiome.
      Analysis of the gut microbiota revealed differential effects of WD on several metabolic pathways in male and female Fxr−/− mice. All these data provide convincing evidence that bile acid/FXR signaling plays a critical role in mediating sex difference in dysbiosis and non-alcoholic fatty liver disease. How the gut microbiota differentially alters metabolic pathways in male and female mice, and how high fat diet influences the gut microbiomes and metabolic pathways remains to be studied in detail.
      Growth hormone/STAT5 signaling has been shown to cause sexual dimorphic expression of the male dominant enzyme oxysterol 7α-hydroxylase (Cyp7b1) and regulate bile acid synthesis and lipid metabolism.
      • Donepudi A.C.
      • Boehme S.
      • Li F.
      • Chiang J.Y.
      G-protein-coupled bile acid receptor plays a key role in bile acid metabolism and fasting-induced hepatic steatosis in mice.
      A recent study reports that the incidence of streptozotocin (STZ)-high fat diet (HFD)-induced hepatocellular carcinoma is significantly higher in male mice than female mice.
      • Xie G.
      • Wang X.
      • Zhao A.
      • Yan J.
      • Chen W.
      • Jiang R.
      • Ji J.
      • Huang F.
      • Zhang Y.
      • Lei S.
      • Ge K.
      • Zheng X.
      • Rajani C.
      • Alegado R.A.
      • Liu J.
      • Liu P.
      • Nicholson J.
      • Jia W.
      Sex-dependent effects on gut microbiota regulate hepatic carcinogenic outcomes.
      Interestingly, metagenomic analysis showed differences in gut bacteria involved in bile acid metabolism between normal male and female mice. STZ-HFD treatment amplified the observed sex differences in gut microbiota. At the phylum level, female mice have a higher ratio of Firmicutes to Bacteroidetes than male mice. STZ-HFD treatment reduced Firmicutes and Bacteroidetes, and markedly increased Proteobacteria in female mice, whereas STZ-HFD treatment increased Firmicutes, decreased Bacteroidetes, and increased Proteobacteria much less in male mice. STZ-HFD treatment caused more intrahepatic retention of hydrophobic bile acids (TCA, taurine chenodeoxycholic acid, and taurine lithocholic acid) in male mice compared to female mice. Interestingly, STZ-HFD treatment strongly reduced Cyp7b1 mRNA expression, consistent with a much higher increase of TCA in male than female mice, and an altered gut microbiota between male and female mice. In diabetic patients, bile acid pool size and 12α-hydroxylated bile acids to non-12α-hydroxylated bile acids ratio in serum is increased. It is known that insulin and glucagon, and fasting and refeeding regulates bile acid synthesis.
      • Li T.
      • Chiang J.Y.
      Bile acid signaling in metabolic disease and drug therapy.
      These recent advances suggest that the sex disparity in diabetes, non-alcoholic steatohepatitis, and liver cancer is linked to sex differences in regulation of bile acid metabolism, which is influenced by insulin, glucocorticoid, and growth hormone levels in males and females. Future study is needed to show how hormones integrate diet, bile acid signaling, and gut microbiota in the gut-to-liver axis to determine sex disparities in metabolic diseases.

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