Serum Bile Acid Levels Research Articles

Duodenal-jejunal bypass increases intraduodenal bile acids and upregulates duodenal SIRT1 expression in high-fat diet and streptozotocin-induced diabetic rats.

BACKGROUNDThe mechanisms underlying diabetes remission after duodenal-jejunal bypass (DJB) remain elusive. In DJB surgery, the duodenum is excluded. However, the duodenum has emerged as an important regulator of glucose homeostasis, and elevated duodenal SIRT1 leads to improved hepatic insulin sensitivity. After DJB, bile acids (BAs) in the duodenum are not mixed and diluted by the ingested food. And activation of BA receptors promotes SIRT1 expression in many tissues. We hypothesized that BA-mediated upregulation of SIRT1 may contribute to diabetic control after DJB.AIMTo investigate the surgical effects of DJB on duodenal SIRT1 expression and uncover the potential crosslinks between BAs and SIRT1.METHODSTwenty diabetic rats were randomly allocated to the sham (n = 10) and DJB (n = 10) groups. Body weight, food intake, fasting blood glucose (FBG), serum and intraduodenal total BA (TBA) levels were measured accordingly. Oral glucose tolerance test (OGTT) and intraperitoneal pyruvate tolerance test (ipPTT) were performed to evaluate the effects of surgeries on systemic glucose disposal and hepatic gluconeogenesis. The key genes of BA signaling pathway in the duodenal mucosa, including farnesoid X receptor (FXR), small heterodimer partner (SHP), and Takeda G-protein-coupled receptor 5 (TGR5) were evaluated by real-time quantitative polymerase chain reaction 8 wk postoperatively. The duodenal SIRT1, AMPK, and phosphorylated AMPK (p-AMPK) levels were evaluated by western blotting. Rat small intestine epithelial IEC-6 cells were treated with GW4064 and INT-777 to verify the effects of BAs on SIRT1 expression in enterocytes.RESULTSThe DJB group exhibited body weight and food intake comparable to those of the sham group at all postoperative time points. The FBG level and area under the curve for the OGTT and ipPTT were significantly lower in the DJB group. The DJB group exhibited higher fasting and postprandial serum TBA levels than the sham group at both 2 and 8 wk postoperatively. At 8 wk after surgery, the DJB group showed higher intraluminal TBA concentration, upregulated mRNA expression of FXR and SHP, and elevated protein expression of SIRT1 and p-AMPK in the descending and horizontal segments of the duodenum. Activation of FXR and TGR5 receptors by GW4064 and INT-777 increased the mRNA and protein expression of SIRT1 and promoted the phosphorylation of AMPK in IEC-6 cells.CONCLUSIONDJB elevates intraduodenal BA levels and activates the duodenal BA signaling pathway, which may upregulate duodenal SIRT1 and further contribute to improved glucose homeostasis after DJB.

Biochemical and Bioinformatic Characterization of Patients with a Sodium Taurocholate Cotransporting Polypeptide Mutation

Background: SLC10A1 codes for the sodium taurocholate cotransporting polypeptide (NTCP). The SLC10A1S267F mutation is associated with loss of function of bile acid (BA) uptake and defined as a new type of hypercholanemia. This kind of hypercholanemia is characterized by high levels of serum BA. However, limited studies have been conducted on this topic. Objectives: This study aimed to describe the biochemical and bioinformatic characterization of patients with an SLC10A1S267F mutation, as well as to dissect pathogenesis in hypercholanemia. Methods: In this study, a total of 12 individuals (including 5 homozygous, 3 heterozygous, and 4 wild-type individuals) were recruited. Whole-genome sequencing (WGS) and Sanger sequencing were used to confirm the genotype. Tests of liver function, renal function, and serum lipid level, in addition to routine blood tests, were performed to evaluate the clinical consequences of patients with an SLC10A1S267F mutation. The ClinVar website and protein prediction tools were used to analyze other cholesterol and BAs related gene mutations in SLC10A1S267F patients, as well as to evaluate their possible effects on serum BA levels of patients. Results: All SLC10A1S267F homozygous patients displayed high levels of BAs. Liver and renal functions were generally normal. According to previous reports, homozygous patients are prone to vitamin D deficiency and deviated blood lipids. However, all homozygous individuals had normal levels of blood lipids, thyroid hormones, and vitamin D (25(OH)D). Moreover, except for the SLC10A1S267F mutation, according to the WGS results, multiple gene mutations were found in 5 homozygous and might affect the level of BAs, but the SLC10A1S267F mutation still is the most important reason resulting in a high level of BAs. Conclusions: This study provided a more detailed description of the SLC10A1S267F mutation-induced hypercholanemia, delivering a new idea that there might be some mutations in SLC10A1S267F homozygotes, probably influencing BA metabolism.

Trends in gestational age at delivery for intrahepatic cholestasis of pregnancy and adoption of society guidelines

Intrahepatic cholestasis of pregnancy is associated with a significant risk of stillbirth, which contributes to variation in clinical management. Recent Society for Maternal-Fetal Medicine guidance recommends delivery at 36 weeks of gestation for patients with serum bile acid levels of >100 μmol/L, consideration for delivery between 36 and 39 weeks of gestation stratified by bile acid level, and against preterm delivery for those with clinical features of cholestasis without bile acid elevation. This study aimed to investigate institutional practices before the publication of the new delivery timing recommendations to establish the maternal and neonatal effects of late preterm, early-term, and term deliveries in the setting of cholestasis. This study examined maternal and neonatal outcomes of 441 patients affected by cholestasis delivering 484 neonates in a 4-hospital system over a 30-month period. Logistic and linear regression analyses were performed to assess neonatal outcomes concerning peak serum bile acid levels at various gestational ages controlling for maternal comorbidities, multiple pregnancies, and neonatal birthweight. With the clinical flexibility afforded by the new guidelines, pregnancy prolongation to term may have been achieved in 91 patients (21%), and 286 patients (74%) with bile acid elevation could have delivered at a later gestational age. Preterm deliveries of patients with bile acid levels of >10 μmol/L were associated with higher rates of neonatal intensive care unit admission and adverse neonatal outcomes than early-term deliveries. Study data suggested an opportunity for education and practice change to reflect current Society for Maternal-Fetal Medicine guidelines in efforts to reduce potential neonatal morbidities associated with late preterm deliveries among pregnancies affected by cholestasis.

Fat Malabsorption and Ursodeoxycholic Acid Treatment in Children With Reduced Organic Solute Transporter-α (SLC51A) Expression.

Objectives:A bile acid homeostasis disorder was suspected in 2 siblings and their second cousin who presented in infancy with fat malabsorption, severe fat-soluble vitamin deficiency, rickets, and mild liver involvement. Our aims were to identify the genetic cause, describe the disease, and evaluate the response to ursodeoxycholic acid (UDCA) treatment.Methods:Whole exome sequencing, immunohistochemistry of duodenal biopsies and candidate variant testing in a cell-based model was performed. Fecal fat excretion, serum bile acids, 7α-hydroxy-4-cholesten-3-one (C4), and fibroblast growth factor 19 (FGF19) were quantified in both siblings on and off UDCA treatment.Results:A novel homozygous variant of SLC51A, which encodes the bile acid carrier organic solute transporter (OST)-α, was identified in all affected children. OSTα protein expression was readily detected by immunohistochemistry in duodenum of pediatric control subjects but not in the affected siblings. The siblings had low serum levels of bile acids and C4 and high serum levels of FGF19 consistent with repression of hepatic bile acid synthesis. On treatment with UDCA, fecal fat excretion was reduced and serum levels of C4, FGF19, and liver enzymes normalized.Conclusions:We report an apparent deficiency of OSTα associated with early onset fat malabsorption and mild liver involvement. The clinical presentation partially overlaps previous reports for 3 patients with OSTα or OSTβ deficiency and extends the clinical spectrum associated with loss of SLC51A expression. Our data suggest that repression of hepatic bile acid synthesis contributes to fat malabsorption in OSTα-OSTβ deficiency but can be partly reversed with UDCA treatment.

NTCP Deficiency Affects the Levels of Circulating Bile Acids and Induces Osteoporosis.

BackgroundThe p.Ser267Phe mutation in the SLC10A1 gene can cause NTCP deficiency. However, the full clinical presentation of p.Ser267Phe homozygous individuals and its long-term consequences remain unclear. Hence, in the present study, we characterized the phenotypic characteristics of NTCP deficiency and evaluated its long-term prognosis.MethodsTen NTCP p.Ser267Phe homozygous individuals were recruited and a comprehensive medical evaluation with a 5-year follow-up observation was performed. The phenotypic characteristics of NTCP deficiency were also demonstrated using an NTCP-global knockout mouse model.ResultsDuring the 5-year follow-up observation of 10 NTCP p.Ser267Phe homozygous adults, we found that the most common phenotypic features of NTCP deficiency in adults were hypercholanemia, vitamin D deficiency, bone loss, and gallbladder abnormalities. The profile of bile acids (BAs) in the serum was significantly altered in these individuals and marked by both elevated proportion and concentration of primary and conjugated BAs. Moreover, the NTCP deficiency led to increased levels of serum BAs, decreased levels of vitamin D, and aggravated the osteoporotic phenotype induced by estrogen withdrawal in mice.ConclusionsBoth mice and humans with NTCP deficiency presented hypercholanemia and were more prone to vitamin D deficiency and aggravated osteoporotic phenotype. Therefore, we recommend monitoring the levels of BAs and vitamin D, bone density, and abdominal ultrasounds in individuals with NTCP deficiency.

Risk Factor Analysis of Hepatic Encephalopathy and the Establishment of Diagnostic Model.

To identify laboratory diagnostic indicators of hepatic encephalopathy (HE), the present study established a HE diagnostic model to explore the diagnostic value of serum homocysteine, lactic acid, procalcitonin, and bile acid levels in HE identification. 371 patients with liver cirrhosis were selected as research objects, who were admitted to the Department of Hepatic Diseases, Affiliated Hospital of Northwest Minzu University from August 2019 to August 2020. The Spearman correlation results indicated that between lactic acid, procalcitonin, bile acid, serum homocysteine, and HE, the coefficients were -0.15, 0.41, 0.29, and -0.19, respectively. Univariate and multivariate analysis methods were adopted for inpatient analysis to identify the influencing factors of HE occurrence, and the diagnosis of the HE identification model was subsequently constructed. The univariate logistic regression showed that risk of developing HE increased as bile acid level (P = 0.00434) and serum homocysteine (P = 0.058) increased. Multivariate logistic regression diagnostic model of bile acid level and serum homocysteine revealed that the AUC value of the area under the ROC curve was 0.7201, indicating that the diagnostic model produced a satisfactory evaluation effect. The model formula referred logistic (P) = −2.4544 + 0.0117 bile acid levels + 0.0198 serum homocysteine. In this study, the HE diagnostic model was established using logistic regression analysis, which could benefit patients in early HE differential diagnosis. Particularly, combined detection of serum homocysteine and bile acid levels was considered to be more significant.

Editorial: higher levels of certain serum bile acids in non‐alcoholic fatty liver disease–new insights from Guatemala.Authors’ reply

LINKED CONTENTThis article is linked to Rivera–Andrade et al papers. To view these articles, visit https://doi.org/10.1111/apt.16948 and https://doi.org/10.1111/apt.16962

Histopathological liver steatosis linked with high parenteral glucose and amino acid supply in infants with short bowel syndrome.

Steatosis is a common feature of intestinal failure-associated liver disease (IFALD) in adult and older pediatric patients receiving long-term parenteral nutrition (PN). There are limited clinical data concerning steatosis in infants with short bowel syndrome (SBS). We investigated early histopathological steatosis and its association to PN. In this retrospective study, 31 patients with SBS had a diagnostic liver biopsy taken at the median age of 5 (IQR 3-8) months. Follow-up biopsy was available for 24 patients at the median age of 29 (IQR 14-52) months. We evaluated the biopsies for steatosis and other histopathological signs of IFALD and compared results with patient characteristics, PN composition, and liver biochemistry. Diagnostic biopsies revealed steatosis in 8 (26%) patients. At the age of 3 months, patients with steatosis had received higher amounts of parenteral glucose: median 15.1 (IQR 12.4-17.2) vs 12.3 (8.7-14.4) g/kg/d (P = 0.04), amino acids: 2.9 (2.5-3.4) vs 2.2 (1.6-2.7) g/kg/d (P = 0.03), and energy: 87 (80-98) vs 73 (54-79) kcal/kg/d (P = 0.01) than those without steatosis. We detected no significant differences in parenteral lipid intake between the groups. Steatosis also associated with increased serum bile acid (P = 0.02), alanine aminotransferase (P = 0.0002), and aspartate aminotransferase (P = 0.001) levels. In this cohort, high parenteral glucose, amino acid, and energy provision associated with liver steatosis in infants with SBS. We recommend monitoring of bile acid and transaminase levels while aiming for PN with balanced macronutrient supply according to current recommendations to protect the liver from steatosis.

The Role of Bile Acid in Improving Glucose Tolerance of Non-Obese Diabetic Rats After Proximal Small Bowel Bypass.

An increase in bile acid (BA) levels after metabolic surgery is an important mechanism for improving glucose metabolism. However, the mechanisms underlying elevated BA levels and the regulatory mechanism of glucose metabolism remain unclear. In this study, we used the Goto-Kakizaki rat model to investigate the mechanism of BA elevation by comparing side-to-side jejunoileal bypass plus proximal loop ligation (SSJIBL) and bile ileum diversion (BID) as well as to explore the mechanism of BA metabolism in regulating blood glucose. The results showed that the fed blood glucose of rats in both the SSJIBL and BID groups was significantly lower than that of the SHAM group on days 2 and 14 after the operation. The oral glucose tolerance test (OGTT) improved in the SSJIBL and BID groups at day 14 postoperatively. The expression of CYP27A1 in the livers of the SSJIBL and BID groups was significantly increased. In addition, total serum BA levels in the SSJIBL and BID groups were significantly increased. Moreover, serum levels of lithocholic acid (LCA) and deoxycholic acid (DCA) were significantly higher in the SSJIBL group than in the SHAM group and negatively correlated with the area under the glucose tolerance curve (AUC-OGTT). In conclusion, increased BA synthesis may be an important cause of elevated total serum BA levels, and LCA and DCA are closely associated with improved glucose metabolism.

Lower systemic inflammation is associated with gut firmicutes dominance and reduced liver injury in a novel ambulatory model of parenteral nutrition

Background Total Parenteral Nutrition (TPN) provides lifesaving nutritional support to patients unable to maintain regular enteral nutrition (EN). Unfortunately, cholestasis is a significant side effect affecting 20–40% of paediatric patients. While the aetiology of TPN-associated injury remains ill-defined, an altered enterohepatic circulation in the absence of gut luminal nutrient content during TPN results in major gut microbial clonal shifts, resulting in metabolic endotoxemia and systemic inflammation driving liver injury and cholestasis. Hypothesis To interrogate the role of gut microbiota, using our novel ambulatory TPN piglet model, we hypothesized that clonal reduction of bacteria in Firmicutes phylum (predominant in EN) and an increase in pathogenic Gram-negative bacteria during TPN correlates with an increase in serum lipopolysaccharide and systemic inflammatory cytokines, driving liver injury. Methods Upon institutional approval, 16 animals were allocated to receive either TPN (n = 7) or EN only (n = 9). The TPN group was subdivided into a low systemic inflammation (TPN-LSI) and high systemic inflammation (TPN-HSI) based on the level of serum lipopolysaccharide. Culture-independent identification of faecal bacterial populations was determined by 16S rRNA. Results Piglets on TPN, in the TPN-HSI group, noted a loss of enterocyte protective Firmicutes bacteria and clonal proliferation of potent inflammatory and lipopolysaccharide containing pathogens: Fusobacterium, Bacteroidetes and Campylobacter compared to EN animals. Within the TPN group, the proportion of Firmicutes phylum correlated with lower portal lipopolysaccharide levels (r = −0.89). The TPN-LSI had a significantly lower level of serum bile acids compared to the TPN-HSI group (7.3 vs. 60.4 mg/dL; p = .018), increased day 14 weight (5.67 vs. 5.07 kg; p = .017) as well as a 13.7-fold decrease in serum conjugated bilirubin. Conclusion We demonstrate a novel relationship between the gut microbiota and systemic inflammation in a TPN animal model. Pertinently, the degree of gut dysbiosis correlated with the severity of systemic inflammation. This study underscores the role of gut microbiota in driving liver injury mechanisms during TPN and supports a paradigm change in therapeutic targeting of the gut microbiota to mitigate TPN-related injury. KEY MESSAGES This study identified a differential link between gut microbiota and inflammation—the higher the dysbiosis, the worse the systemic inflammatory markers. Higher levels of Firmicutes species correlated with reduced inflammation.

Intrahepatic Cholestasis of Pregnancy Increases Inflammatory Susceptibility in Neonatal Offspring by Modulating Gut Microbiota.

Intrahepatic cholestasis of pregnancy (ICP) is a liver disease of pregnancy that is characterized by increased bile acid levels in maternal serum. Studies have shown that cholestatic pregnancy can result in long-term metabolic disturbances in the offspring. However, how ICP shapes the offspring’s immunity and predisposition to inflammatory disorders at an early stage is unknown. In this study, we investigated the effect of maternal cholestasis on neonatal offspring metabolism and immune function. We compared 71 neonates with ICP mothers and 63 neonates with healthy mothers and found that the incidence of jaundice and infection was significantly higher in ICP offspring. Maternal serum total bile acid level was associated with blood cell counts in full-term ICP offspring. In animal experiments, a compensatory activation of hepatic and ileal farnesoid X receptor (FXR) and altered gut microbiota in the first week were found in ICP offspring. We also investigated lipopolysaccharide (LPS)-induced inflammatory responses in neonatal rats and found that ICP offspring were more susceptible to inflammation. To understand the correlation between congenital abnormal FXR activation and tissue immunity dysregulation, we assessed the effects of the FXR agonist GW4064 and FXR antagonist E/Z-GS in ICP offspring after LPS exposure. The expression of several pro-inflammatory cytokines significantly decreased after treatment with E/Z-GS but increased after treatment with GW4064. Treatment with the probiotic Lactobacillus rhamnosus LRX01 that inhibits FXR expression in the ileum reduced susceptibility to LPS exposure in ICP offspring. The current study indicated that cholestatic pregnancy may increase the susceptibility of the offspring to inflammation by altering bile acid metabolism and gut microbiota at an early stage. We suggest that supplementation with Lactobacillus rhamnosus LRX01, which inhibits FXR expression in the ileum, may improve intestinal immunity in ICP offspring.

Screening of Biomarkers and Toxicity Mechanisms of Rifampicin-Induced Liver Injury Based on Targeted Bile Acid Metabolomics.

Rifampicin (RIF) is a critical first-line drug for tuberculosis. However, long-term or high-dose treatment with RIF can induce severe liver injury; the underlying mechanism of this effect has not yet been clarified. This study was performed to screen reliable and sensitive biomarkers in serum bile acids (BAs) using targeted BA metabolomics and evaluate the toxicity mechanisms underlying RIF-induced liver injury through the farnesoid x receptor (Fxr)-multidrug resistance-associated proteins (Mrps) signaling pathway. Thirty-two Institute of Cancer Research mice were randomly divided into four groups, and normal saline, isoniazid 75 mg/kg + RIF 177 mg/kg (RIF-L), RIF-L, or RIF 442.5 mg/kg (RIF-H) was orally administered by gavage for 21 days. After treatment, changes in serum biochemical parameters, hepatic pathological conditions, BA levels, Fxr expression, and BA transporter levels were measured. RIF caused notable liver injury and increased serum cholic acid (CA) levels. Decline in the serum secondary BAs (deoxycholic acid, lithocholic acid, taurodeoxycholic acid, and tauroursodeoxycholic acid) levels led to liver injury in mice. Serum BAs were subjected to metabolomic assessment using partial least squares discriminant and receiver operating characteristic curve analyses. CA, DCA, LCA, TDCA, and TUDCA are potential biomarkers for early detection of RIF-induced liver injury. Furthermore, RIF-H reduced hepatic BA levels and elevated serum BA levels by suppressing the expression of Fxr and Mrp2 messenger ribonucleic acid (mRNA) while inducing that of Mrp3 and Mrp4 mRNAs. These findings provide evidence for screening additional biomarkers based on targeted BA metabolomics and provide further insights into the pathogenesis of RIF-induced liver injury.

Survival Benefit of Statin with Anti-Angiogenesis Efficacy in Lung Cancer-Associated Pleural Fluid through FXR Modulation.

Simple SummaryOur previous works showed that pleural fluid from lung cancer significantly induced endothelial proliferation, migration, and angiogenesis. Since endothelial metabolism was a key step in angiogenesis, we investigated the role of bile acid signaling and FXR expression in pleural angiogenesis. Elevated bile acid levels in lung-cancer-associated pleural fluid (LCPF) were characterized with positive FXR staining in pleural microvessels. We then confirmed the inhibitory effect of an FXR antagonist on LCPF-induced endothelial migration and angiogenesis. Due to the elevated protein expression in the cholesterol metabolism caused by LCPF, lipid-lowering agents with the efficacy needed to counteract LCPF-regulated angiogenesis were evaluated. Statin showed the potent efficacy needed to suppress LCPF-induced endothelial proliferation, migration, and angiogenesis through FXR inhibition. Following that, Kaplan–Meier analysis showed the survival benefit of statin exposure in patients with lung adenocarcinoma with LCPF. Our results suggest that targeting endothelial FXR signaling with statin treatment could ameliorate the angiogenesis activity of LCPF.Lung cancer-related pleural fluid (LCPF) presents as a common complication with limited treatment. Beyond its function in lipid digestion, bile acid was identified as a potent carcinogen to stimulate tumor proliferation. Previous research indicated a correlation between serum bile acid levels and the risk of developing several gastrointestinal cancers. Our study identified elevated bile acid levels in LCPF and increased farnesoid X receptor (FXR) expression as bile acid nuclear receptors in pleural microvessels of lung adenocarcinoma. Additionally, LCPF stimulated the expression of proteins involved in bile acid synthesis and cholesterol metabolism in HUVECs including CYP7A1, StAR, HMGCR, and SREBP2. LCPF-induced endothelial motility and angiogenesis were counteracted by using β-muricholic acid as an FXR antagonist. Moreover, we investigated the efficacy of cholesterol-lowering medications, such as cholestyramine, fenofibrate, and atorvastatin, in regulating LCPF-regulated angiogenesis. Along with suppressing endothelial proliferation and angiogenesis, atorvastatin treatment reversed cholesterol accumulation and endothelial junction disruption caused by LCPF. Statin treatment inhibited LCPF-induced endothelial FXR expression as well as the downstream proteins RXR and SHP. Based on the positive findings of suppressing endothelial angiogenesis, our group further incorporated the effect of statin on clinical patients complicated with LCPF. A Kaplan–Meier analysis revealed the clinical benefit of statin exposure in patients with lung adenocarcinoma with LCPF. Conclusively, our study demonstrated the ability of statin to alleviate LCPF-induced angiogenesis in patients with LCPF via FXR modulation.

Therapeutic Effect of Acer tegmentosum Maxim Twig Extract in Bile Duct Ligation-Induced Acute Cholestasis in Mice.

Cholestatic liver disease, or cholestasis, is a condition characterized by liver inflammation and fibrosis following a bile duct obstruction and an intrahepatic accumulation of bile acids. Inhibiting inflammation is a promising therapeutic strategy for cholestatic liver diseases. Acer tegmentosum Maxim extract (ATE) is best known for its anti-inflammatory and antioxidative properties. In this study, we investigated the effects of ATE on liver injury and fibrosis in mice with bile duct ligation (BDL)-induced cholestasis through analysis of gene expression, cytokines, and histological examination. Oral administration of ATE (20 or 50 mg/kg) for 14 days significantly attenuated hepatocellular necrosis compared to vehicle-treated BDL mice, which was accompanied by the reduced level of serum bile acids and bilirubin. We determined that ATE treatment reduced liver inflammation, oxidative stress, and fibrosis. These beneficial effects of ATE were concurrent with the decreased expression of genes involved in the NF-κB pathway, suggesting that the anti-inflammatory effect of ATE could be a possible mechanism against cholestasis-associated liver injury. Our findings substantiate ATE's role as an alternative therapeutic agent for cholestasis-induced liver injury and fibrosis.

Antarctic krill oil ameliorates liver injury in rats exposed to alcohol by regulating bile acids metabolism and gut microbiota

Bile acids (BAs) metabolism plays an important role in alcohol liver disease through the gut microflora-bile acids-liver axis. Antarctic Krill Oil (AKO) has protective effects on the liver, while whether AKO can protect against liver injury caused by alcohol is unclear. This study investigated the effects of AKO on BAs metabolism and intestinal microbiota in a rat model of alcohol-induced liver disease. Sprague-Dawley rats were randomly divided into five groups: control group, model group, low-dose AKO-treatment group (100 mg/kg/d), high-dose AKO-treatment group (200 mg/kg/d), and AKO control group (200 mg/kg/d). Administration of alcohol (8 to 10 mL/kg/d) for 16 weeks induced liver injury in rats. We found that AKO supplementation significantly protected the liver against alcohol-induced injury, evidenced by allayed hepatic histopathological changes, and inhibited the alcohol-induced elevation of serum biochemical indices. Furthermore, AKO could regulate BAs metabolism by activating the intestinal-hepatic FXR-FGF15-FGFR4 signaling axis with subsequently decreased cholesterol 7α-hydroxylase (CYP7A1) and sterol 12α-hydroxylase (CYP8B1) levels, reduced hepatic BAs production, decreased serum BAs level and increased fecal excretion of BAs. Additionally, 16S rDNA sequencing revealed that the gut microbiome richness and composition were altered in alcohol-treated rats in comparison to the control and AKO-administrated rats. Spearman's correlation analysis showed that differential gut bacterial genera correlated with the levels of BAs profiles in the serum, liver, and feces. These findings suggested that AKO dietary supplementation may protect against alcohol-induced liver injury through modulating BAs metabolism and altering the gut microbiome.

Lipocalin‐2 Stimulates Proliferation and a Profibrotic Cholangiocyte Phenotype during Cholestasis

Lipocalin‐2 (LCN2), also known as neutrophil gelatinase‐associated lipocalin (NGAL), is a secreted glycoprotein that has been shown to regulate cell proliferation, innate immunity, metabolism, and tumor metastasis in several cell types. LCN2 expression is elevated in hepatic tissues in experimental liver injury models and patients with alcoholic hepatitis. LCN2 has been shown to stimulate proliferation and metastasis of cholangiocarcinoma. However, the role of cholangiocyte LCN2 and LCN2 receptors (NGALR/24p3‐R) during cholestasis and primary sclerosing cholangitis (PSC) has not been evaluated. Therefore, we aimed to assess the role of LCN2 and its receptor, 24p3‐R, in acute and chronic cholestatic models.MethodsLCN2 and 24p3‐R were evaluated in liver sections, total liver, and isolated cholangiocytes from wild‐type (WT; C57) normal and BDL (7 days) mice and WT (FVB/NJ) and Mdr2‐/‐ mice (model of primary sclerosing cholangitis, PSC) by immunohistochemistry and qPCR, respectively. LCN2 levels were evaluated by ELISA in cholangiocyte supernatants (6 hrs) collected from animal models and serum from BDL and Mdr2‐/‐ and WT controls and late‐stage PSC patients and healthy human controls. Serum bile acid (BA) levels were measured using a commercial BA assay kit in BDL, Mdr2‐/‐, and control mice. In vitro, both human (H69) and mouse pooled (MPC) cholangiocytes were treated with either 50 ng/ml LCN2 or 100 mM taurocholic acid (TCA) for 24 hrs after which the expression of proliferation, fibrosis, LCN2, and 24p3‐R genes was determined via qPCR.ResultsThere were significant increases in LCN2 and 24p3‐R immunostaining in BDL mice compared to controls. By qPCR significant increases were observed for both LCN2 and 24p3‐R in total liver and isolated cholangiocytes from BDL and Mdr2‐/‐ mice compared to WT. Increased levels of LCN2 were found in serum from PSC patients and serum and supernatants from isolated cholangiocytes from BDL and Mdr2‐/‐ mice compared to control. In vitro, MPC cells treated with LCN2 had increases in fibrotic marker Col1a1, proliferation marker PCNA, and inflammatory markers IL‐1b and IL‐6. Also, significant increases were seen in fibrotic markers (aSMA, FN‐1, and TGF‐b1) and LCN2 receptor 24p3‐R in H69 cells treated with LCN2 compared to untreated cells. As expected, increased serum BA levels were observed in both BDL and Mdr2‐/‐ mice compared to controls. H69 cells treated with TCA also had significant increases in FN‐1, TGF‐b1, and the LCN2 receptor 24p3‐R, indicating that BA may regulate 24p3‐R expression.ConclusionLCN2 and 24p3‐R expression is elevated in cholestatic models and stimulates a profibrotic cholangiocyte phenotype. These findings suggest that LCN2 may play a crucial role in the pathogenesis of cholestatic liver injury.

Bile acid‐mediated activation of brown fat protects from alcohol‐induced steatosis and liver injury

Background & AimsAlcohol associated liver disease (AALD) is one of the most common causes of liver injury and failure. Limited knowledge of the mechanisms underlying AALD impedes the development of efficacious therapies. Bile acid (BA) signaling was shown to participate in progression of AALD, the mechanisms of which remain poorly understood.Approach & ResultsMice were exposed to chronic‐plus‐one‐binge ethanol feeding to induce AALD. Interestingly, ethanol feeding increased expression of the thermogenesis genes downstream of TGR5 (Takeda G protein‐coupled BA receptor 5) in brown adipose tissue (BAT) of wild‐type (WT) but not TGR5 knockout (KO) mice. TGR5 deficiency significantly diminished BAT activity and energy expenditure in mice after ethanol feeding. Alcohol increased serum BA levels in mice and humans through altering BA transportation rather than increasing hepatic BA synthesis, which activated TGR5 signaling in BAT. TGR5 deficiency did not alter alcohol‐stimulated adipose lipolysis, de novo hepatic lipogenesis, or alcohol effects on hepatic β‐oxidation of free fatty acids (FFA). However, compared to ethanol‐fed WT mice, ethanol‐fed TGR5 KO mice showed less FFA β‐oxidation in BAT, leading to increased circulation and liver uptake of FFA. Importantly, in ethanol‐fed mice housed under thermoneutral conditions, TGR5‐mediated fat burning in BAT was unchanged. Finally, administration of a specific TGR5 agonist significantly activated TGR5 signaling in BAT to increase thermogenesis, reduce serum FFA level and ameliorate hepatic steatosis and injury in AALD mice.ConclusionsBA signaling plays a protective role in AALD by enhancing BAT thermogenesis and BAT crosstalk with the liver via TGR5 activation. Targeting TGR5 in BAT may be a promising approach for the treatment of AALD.

Angelica keiskei Root Extract Attenuates Bile Duct Ligation-Induced Liver Injury in Mice.

Although multiple studies have shown that Angelica keiskei of the Umbelliferae family has potent anti-inflammatory and antioxidative activities and that it reduces the serum bile acids in humans, whether A. keiskei has protective effects against cholestasis-induced liver injury remains unexplored until now. This study tests the hypothesis that Angelica keiskei root extract (AKE) alleviates liver injury, inflammation, and fibrosis in mouse models of acute cholestasis induced by bile duct ligation (BDL). Oral administration of AKE (200 or 500 mg/kg) attenuated hepatocellular necrosis and significantly reduced serum levels of bile acids and bilirubin in BDL mice. The critical enzyme of bile acid synthesis, CYP7A1, was repressed by AKE, suggesting that reduced bile acid production may contribute to liver protection. Moreover, we determined through gene expression and cytokine analysis and histological examination that AKE treatment decreased liver inflammation, oxidative stress, and fibrosis. AKE also suppressed the NF-κB pathway, suggesting this as a possible mediator of its anti-inflammatory effect. Our findings substantiate that AKE may be promising for treating cholestatic liver diseases in the future.

The impact of assisted reproductive technology in twin pregnancies complicated by intrahepatic cholestasis: a five-year retrospective study

BackgroundIntrahepatic cholestasis of pregnancy is one of the common complications during pregnancy, and ursodeoxycholic acid has been recommended as the first-line drug. However, if the assisted reproductive technology may increase adverse perinatal outcomes of twin pregnancies complicated by intrahepatic cholestasis is disputed, we aimed to investigate perinatal outcomes between twin pregnancies by assisted reproductive technology versus spontaneous conception, based on these women accepted ursodeoxycholic acid treatment.MethodsFrom January 2014 to January 2019, we retrospectively analysed the clinical data of twin pregnant women with intrahepatic cholestasis, excluding those who did not receive ursodeoxycholic acid treatment. In total, 864 women were included, among whom 500 conceived by assisted reproductive technology and 364 conceived by spontaneous conception. The primary assessment for perinatal outcomes included premature birth, meconium-stained amniotic fluid, low Apgar score, neonatal intensive care unit and still birth, and secondary indicators were serum bile acid and liver enzymes level during medication, so we also finished subgroup analysis based on different elevated bile acid level and drug usage. The statistical analysis was performed by SPSS 22.0.ResultsThe study demonstrated that compared to spontaneous conception, assisted reproductive technology conceived twin pregnancies diagnosed as intrahepatic cholestasis earlier (p = 0.003), and lower birth weight (p = 0.001), less incidence of preterm delivery (p = 0.000) and neonatal intensive care unit admission (p = 0.001), but the rate of meconium-stained amniotic fluid, low Apgar score and still birth have no statistic differences. Moreover, the subgroup analysis showed no significant difference in elevated bile acid levels and medication between assisted reproductive technology and spontaneous conception groups.ConclusionsThe assisted reproductive technology may increase the risk of early-onset intrahepatic cholestasis in twin pregnancies, but it does not seem to increase adverse effects on bile acid levels and perinatal outcomes. Regardless of ursodeoxycholic acid used alone or combination, the effect of bile acid reduction and improving perinatal outcomes in twin pregnancies is limited. Our conclusions still need more prospective randomized controlled studies to confirm.

The role of inflammatory markers derived from complete blood count results in the diagnosis of intrahepatic cholestasis of pregnancy

Objective: This study aimed to investigate complete blood count (CBC) markers, particularly those indicating systemic inflammation, in healthy subjects and those with mild and severe intrahepatic cholestasis of pregnancy (ICP).
 Material and Method: Seventy-nine subjects with ICP and 100 healthy age-matched women who were matched based on their scheduled singleton deliveries (on the same day) were included. ICP was defined as the increase in serum bile acid levels ≥10 μmol/L coupled with pruritus which could not be explained by other conditions. Patients with ICP were further categorized to mild and severe ICP according to bile acid levels, as follows: Mild ICP (