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Hyperammonemia refers to elevated levels of ammonia in the blood, which is an important pathological feature of liver cirrhosis and hepatic failure. Preclinical studies suggest tropifexor (TXR), a novel non-bile acid agonist of Farnesoid X Receptor (FXR), has shown promising effects on reducing hepatic steatosis, inflammation, and fibrosis. This study evaluates the impact of TXR on hyperammonemia in a piglet model of cholestasis. We here observed blood ammonia significantly elevated in patients with biliary atresia (BA) and was positively correlated with liver injury. Targeted metabolomics and immunblotting showed glutamine metabolism and urea cycles were impaired in BA patients. Next, we observed that TXR potently suppresses bile duct ligation (BDL)-induced injuries in liver and brain with improving the glutamine metabolism and urea cycles. Within the liver, TXR enhances glutamine metabolism and urea cycles by up-regulation of key regulatory enzymes, including glutamine synthetase (GS), carbamoyl-phosphate synthetase 1 (CPS1), argininosuccinate synthetase (ASS1), argininosuccinate lyase (ASL), and arginase 1 (ARG1). In primary mice hepatocytes, TXR detoxified ammonia via increasing ureagenesis. Mechanically, TXR activating FXR to increase express enzymes that regulating ureagenesis and glutamine synthesis through a transcriptional approach. Together, these results suggest that TXR may have therapeutic implications for hyperammonemic conditions in cholestatic livers., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to disclose., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Biliary atresia (BA) is a cholestatic liver disease in neonates with devastating obstructive intrahepatic and extrahepatic biliary ducts. Owing to the lack of an early diagnostic marker and limited understanding of its pathogenesis, BA often leads to death within 2 years. Therefore, this study aimed to develop early diagnostic methods and investigate the underlying pathogenesis of liver injury in BA using metabolomics. Metabolomics and organoid combined energy metabolism analysis was used to obtain new insights into BA diagnosis and pathobiology using patient samples, mice liver organoids, and a zebrafish model. Metabolomics revealed that D-2-hydroxyglutarate (D-2-HG) levels were significantly elevated in the plasma and liver of patients with BA and closely correlated with liver injuries and impaired liver regeneration. D-2-HG suppressed the growth and expansion of liver organoids derived from the intrahepatic biliary ducts. The energy metabolism analysis demonstrated that D-2-HG inhibited mitochondrial respiration and ATP synthase; however, it increased aerobic glycolysis in organoids. In addition, D-2-HG exposure caused liver degeneration in zebrafish larvae. Mechanistically, D-2-HG inhibited the activation of protein kinase B and the mammalian target of rapamycin signaling. These findings reveal that D-2-HG may represent a novel noninvasive diagnostic biomarker and a potential therapeutic target for infants with BA., (© 2022. The Author(s).)

Biliary atresia (BA) is a severe cholestatic liver disease in children featuring cholestasis and liver fibrosis. The early diagnosis of BA is still challenging. This study aimed to evaluate the diagnostic values of matrix metalloprotease-7 (MMP-7), interleukin-8 (IL-8), and gamma-glutamyl transferase (GGT) in BA. Infants diagnosed with BA and non-BA between 2013 and 2018 were retrospectively analyzed. Plasma levels of MMP-7, IL-8, and GGT were measured in these infants. The receiver operating characteristic (ROC) curves and area under the ROC curve (AUC) were used to assess the diagnostic values of MMP-7, IL-8, and GGT. The expression of MMP-7 and IL-8 in the livers was detected by immunofluorescence staining. A total of 229 infants were enrolled in this study: 156 BA infants and 73 non-BA infants including 16 ones with infantile hepatitis syndrome. The plasma levels of MMP-7, IL-8, and GGT in BA infants had a median of 11.8 ng/mL (interquartile range, IQR: 5.3-57.5), 1.5 ng/mL (IQR: 1.0-2.8), and 381.0 U/L (IQR: 197.0-749.0), respectively, which were higher than non-BA subjects [MMP-7, 4.4 ng/mL (IQR: 3.3-6.1); IL-8, 0.7 ng/mL (IQR: 0.5-1.0); GGT, 59.0 U/L (IQR: 26.0-124.0)]. The AUC values of MMP-7, IL-8, and GGT for the diagnosis of BA were 0.8035, 0.8083, and 0.9126, respectively. The AUC values of MMP-7 + IL-8, MMP-7 + GGT, IL-8 + GGT, and MMP-7 + IL-8 + GGT for the diagnosis of BA were 0.8248, 0.9382, 0.9168, and 0.9392, respectively. The AUC values of MMP-7, IL-8, and GGT for differentiating BA infants with cholic stool from non-BA infants with cholic stool were 0.8006, 0.8258, and 0.9141, respectively. The expression of MMP-7 and IL-8 was increased in the cholangiocytes in BA livers.   Conclusion: Plasma MMP-7, IL-8, and GGT alone or a combination of them has good accuracy to differentiate BA from non-BA and may be reliable biomarkers for BA. What is Known: • Biliary atresia (BA) is a severe cholestatic liver disease in children featuring cholestasis and progressive liver fibrosis. • Although early diagnosis of BA is crucial for good outcomes, it remains a clinical challenge. What is New: • Plasma MMP-7, IL-8, and GGT alone or a combination of them has good accuracy to differentiate BA from non-BA. • Plasma MMP-7, IL-8, and GGT have good accuracy for differentiating BA infants with cholic stool from non-BA infants with cholic stool., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Early diagnosis and timely surgical Kasai portoenterostomy greatly improve the survival of patients with biliary atresia (BA), a neonatal cholestatic disease, which has encouraged investigators to develop newborn screening for BA. In this study, we used ultraperformance liquid chromatography-triple quadrupole mass spectrometry-based targeted metabolomics profiling to identify potential BA biomarkers in dried blood spots (DBS) collected from BA patients ( n = 21) and healthy controls ( n = 100). A distinctive metabolic profile comprising eight significantly differentially expressed metabolites, taurohyocholic acid (THCA), glutamic acid, 2-hydroxyglutaric acid, ketoleucine, indoleacetic acid, alpha-ketoisovaleric acid, glycocholic acid, and taurocholic acid (TCA), clearly distinguished BA infants from control neonates. Three metabolites, THCA, 2-hydroxyglutaric acid, and indoleacetic acid, were selected using linear regression and receiver operating characteristic (ROC) curve analysis and model construction. The area under the ROC curve for this model to discriminate between BA and comparison infants was 0.938 (95% confidence interval, CI: 0.874-1.000). A cutoff value of -0.336 produced a sensitivity of 90.48% (95% CI: 69.62% - 98.83%) and specificity of 92% (95% CI: 84.84% - 96.48%). In conclusion, the results suggest that metabolic markers in DBS obtained from newborns have a great potential for BA screening.

Parenteral nutrition-associated liver disease (PNALD) refers to a spectrum of conditions that can develop cholestasis, steatosis, fibrosis, and cirrhosis in the setting of parenteral nutrition (PN) use. Patient risk factors include short bowel syndrome, bacterial overgrowth and translocation, disturbance of hepatobiliary circulation, and lack of enteral feeding. A growing body of evidence suggests an intricate linkage between the gut microbiota and the pathogenesis of PNALD. In this review, we highlight current knowledge on the taxonomic and functional changes in the gut microbiota that might serve as noninvasive biomarkers. We also discuss the function of microbial metabolites and associated signaling pathways in the pathogenesis of PNALD. By providing the perspectives of microbiota-host interactions in PNALD for basic and translational research and summarizing current limitations of microbiota-based approaches, this review paves the path for developing novel and precise microbiota-based therapies in PNALD., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Biliary atresia (BA) is a neonatal liver disease featuring cholestasis and severe liver fibrosis (LF). Despite advances in the development of surgical treatment, lacking an early diagnostic marker and intervention of LF invariably leads to death from end-stage liver disease in the early years of life. We previously reported that knockout of sphingosine 1-phosphate receptor 2 (S1PR2) protected mice from bile duct ligation (BDL)-induced cholangiocyte proliferation and LF. Our recent studies further showed that both hepatic and serum exosomal long noncoding RNA H19 (lncRNAH19) levels are correlated with cholestatic injury in multidrug resistance 2 knockout (Mdr2 -/- ) mice. However, the role of lncRNAH19 in BA progression remains unclear. Here, we show that both hepatic and serum exosomal H19 levels are positively correlated with severity of fibrotic liver injuries in BA patients. H19 deficiency protects mice from BDL-induced cholangiocyte proliferation and LF by inhibiting bile-acid-induced expression and activation of S1PR2 and sphingosine kinase 2 (SphK2). Furthermore, H19 acts as a molecular sponge for members of the microRNA let-7 family, which results in up-regulation of high-mobility group AT-hook 2 (HMGA2), a known target of let-7 and enhancement of biliary proliferation. Conclusion: These results indicate that H19 plays a critical role in cholangiocyte proliferation and cholestatic liver injury in BA by regulating the S1PR2/SphK2 and let-7/HMGA2 axis. Serum exosomal H19 may represent a noninvasive diagnostic biomarker and potential therapeutic target for BA., (© 2019 by the American Association for the Study of Liver Diseases.)

Mitophagy is a major pathway for clearance of injured mitochondria. However, whether mitophagy is involved in the cholestasis-induced damages of hepatic mitochondria remains unknown. We here aimed to investigate the molecular links between cholestasis and hepatic mitophagy. We show that mitophagy is increased significantly in livers of biliary atresia (BA) that is cholestatic disease in infants. The mitochondrial-toxicity bile acids treatment increases the activities of mitophagy in hepatocytes. Mechanistically, we find that the prohibitin 2 (PHB2) is crucial for cholestasis-mediated mitophagy in vitro. On the one hand, PHB2 binds the autophagosomal membrane-associated protein LC3 upon injured mitochondria via an LC3-interaction region domain. On the other hand, PHB2 forms a ternary protein complex with sequestosome 1 (SQSTM1) and LC3, leading to loading of LC3 onto the damaged mitochondria. Altogether, our study suggests that PHB2 is required for cholestasis-induced mitophagy via LC3 onto the injured mitochondria.

Biliary atresia (BA) is a severe chronic cholestasis disorder of infants that leads to death if not treated on time. Neonatal hepatitis syndrome (NHS) is another leading cause of neonatal cholestasis confounding the diagnosis of BA. Recent studies indicate that altered bile acid metabolism is closely associated with liver injury and cholestasis. In this study, we systematically measured the bile acid metabolome in plasma of BA, NHS, and healthy controls. Liver bile acids were also measured using biopsy samples from 48 BA and 16 NHS infants undergoing operative cholangiography as well as 5 normal adjacent nontumor liver tissues taken from hepatoblastoma patients as controls. Both BA and NHS samples had significantly elevated bile acid levels in plasma compared to normal controls. BA patients showed a distinct bile acid profile characterized by the higher taurochenodeoxycholic acid (TCDCA) level and lower chenodeoxycholic acid (CDCA) level than those in NHS patients. The ratio of TCDCA to CDCA in plasma was significantly higher in BA compared to healthy infants (p < 0.001) or NHS (p < 0.001). The area under receiver operating characteristic curve for TCDCA/CDCA to differentiate BA from NHS was 0.923 (95% CI: 0.862-0.984). These findings were supported by significantly altered expression levels of bile acid transporters and nuclear receptors in liver including farnesoid X receptor (FXR), small heterodimer partner (SHP), bile salt export pump (BSEP), and multidrug resistant protein 3 (MDR3) in BA compared to NHS. Taken together, the plasma bile acid profiles are distinct in BA, NHS, and normal infants, as characterized by the ratio of TCDCA/CDCA differentially distributed among the three groups of infants.

Background & Aims: Cholestatic liver disease is associated with dysregulated expression of microRNAs (miRNAs). However, it remains unknown whether miRNAs are involved in the cholestasis-induced proliferation of cholangiocytes. In this study, we tested the hypothesis that miRNAs modulate cholangiocyte proliferation through effects on the IL-6 pathway, a known regulator of cholangiocyte proliferation., Methods: Expression of IL-6, Foxa2, and phosphorylated signal transducer activator of transcription 3 (STAT3) was investigated in patients with biliary atresia (BA) and in rats subjected to bile duct ligation (BDL). miRNA expression was determined in BA patients and BDL rats, with miRNA array and quantitative real-time PCR. Biological functions of miRNAs were studied using immunoblot, immunohistochemical and proliferation assays. Luciferase reporter assays and Western blots were performed to identify miRNA targets., Results: Hepatic interleukin-6 (IL-6) expression was significantly elevated in BA patients and BDL rats, while the expression of miR-124 was dramatically decreased in comparison to controls. Moreover, mRNA levels of STAT3 and IL-6 receptor (IL-6R) were inversely correlated with those of miR-124. Ectopic expression of miR-124 inhibited IL-6-mediated cholangiocyte proliferation in vitro and cholangiocyte hyperplasia in vivo, through a mechanism involving direct targeting of the 3'-untranslated region of STAT3 and IL-6R. We further demonstrated that miR-200 family members were significantly upregulated in cholestasis and inhibited FOXA2 expression in cholangiocytes, which further enhanced the expression of IL-6., Conclusions: Our findings suggest that downregulation of miR-124 and upregulation of miR-200 collaboratively promote bile duct proliferation through the IL-6/STAT3 pathway., (Copyright © 2014 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)