Mechanistic studies on the alleviation of ANIT-induced cholestatic liver injury by Polygala fallax Hemsl. polysaccharides.

ETHNOPHARMACOLOGICAL RELEVANCE: Polygala fallax Hemsl. is a traditional folk medicine commonly used by ethnic minorities in the Guangxi Zhuang Autonomous Region, and has a traditional application in the treatment of liver disease. Polygala fallax Hemsl. polysaccharides (PFPs) are of interest for their potential health benefits. AIM OF THIS STUDY: This study explored the impact of PFPs on a mouse model of cholestatic liver injury (CLI) induced by alpha-naphthyl isothiocyanate (ANIT), as well as the potential mechanisms. MATERIALS AND METHODS: A mouse CLI model was constructed using ANIT (80 mg/kg) and intervened with different doses of PFPs or ursodeoxycholic acid. Their serum biochemical indices, hepatic oxidative stress indices, and hepatic pathological characteristics were investigated. Then RNA sequencing was performed on liver tissues to identify differentially expressed genes and signaling pathways and to elucidate the mechanism of liver protection by PFPs. Finally, Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were used to verify the differentially expressed genes. RESULTS: Data analyses showed that PFPs reduced the levels of liver function-related biochemical indices, such as ALT, AST, AKP, TBA, DBIL, and TBIL. PFPs up-regulated the activities of SOD and GSH, down-regulated the contents of MDA, inhibited the release of IL-1ß, IL-6, and TNF-α, or promoted IL-10. Pathologic characterization of the liver revealed that PFPs reduced hepatocyte apoptosis or necrosis. The RNA sequencing indicated that the genes with differential expression were primarily enriched for the biosynthesis of primary bile acids, secretion or transportation of bile, the reactive oxygen species in chemical carcinogenesis, and the NF-kappa B signaling pathway. In addition, the results of qRT-PCR and Western blotting analysis were consistent with those of RNA sequencing analysis. CONCLUSIONS: In summary, this study showed that PFPs improved intrahepatic cholestasis and alleviated liver damage through the modulation of primary bile acid production, Control of protein expression related to bile secretion or transportation, decrease in inflammatory reactions, and inhibition of oxidative pressure. As a result, PFPs might offer a hopeful ethnic dietary approach for managing intrahepatic cholestasis.

Gancao decoction attenuates hepatic necroptosis via activating caspase 8 in cholestatic liver injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Gancao Decoction (GCD) is widely used to treat cholestatic liver injury. However, it is unclear whether is related to prevent hepatocellular necroptosis. AIM OF THE STUDY: The purpose of this study is to clarify the therapeutic effects of GCD against hepatocellular necroptosis induced by cholestasis and its active components. MATERIALS AND METHODS: We induced cholestasis model in wild type mice by ligating the bile ducts or in Nlrp3-/- mice by intragastrical administering Alpha-naphthylisothiocyanate (ANIT). Serum biochemical indices, liver pathological changes and hepatic bile acids (BAs) were measured to evaluate GCD's hepatoprotective effects. Necroptosis was assessed by expression of hallmarkers in mice liver. Moreover, the potential anti-necroptotic effect of components from GCD were investigated and confirmed in ANIT-induced cholestasis mice and in primary hepatocytes from WT mouse stimulated with Tumor Necrosis Factor alpha (TNF-α) and cycloheximide (CHX). RESULTS: GCD dose-dependently alleviated hepatic necrosis, reduced serum aminotranferase activity in both BDL and ANIT-induced cholestasis models. More importantly, the expression of hallmarkers of necroptosis, including MLKL, RIPK1 and RIPK3 phosphorylation (p- MLKL, p-RIPK1, p-RIPK3) were reduced upon GCD treatment. Glycyrrhetinic acid (GA), the main bioactive metabolite of GCD, effectively protected against ANIT-induced cholestasis, with decreased expression of p-MLKL, p-RIPK1 and p-RIPK3. Meanwhile, the expression of Fas-associated death domain protein (FADD), long isoform of cellular FLICE-like inhibitory protein (cFLIPL) and cleaved caspase 8 were upregulated upon GA treatment. Moreover, GA significantly increased the expression of active caspase 8, and reduced that of p-MLKL in TNF-α/CHX induced hepatocytes necroptosis. CONCLUSIONS: GCD substantially inhibits necroptosis in cholestatic liver injury. GA is the main bioactive component responsible for the anti-necroptotic effects, which correlates with upregulation of c-FLIPL and active caspase 8.

Management of Tacrolimus-Induced Toxicity With Normal Serum Levels After Liver Transplant.

Drug-induced liver injury after liver transplant occurs in 1.7% of patients. Tacrolimus is an effective immunosuppressant that is used to treat acute rejection. Although rare, it can cause toxicity, which is demonstrated by cholestatic liver injury. Here, we present a case of a young male patient who was diagnosed with Wilson disease, had penicillaminechelating therapy, and underwent living related liver transplant. Within 1 month posttransplant, he developed deranged, predominantly cholestatic pattern liver function tests. Laboratory parameters showed total bilirubin of 1.12 mg/ dL, alanine aminotransferase of 553 IU/L, gammaglutamyltransferase of 624 IU/L, and tacrolimus level of 10.2 ng/mL. After thorough evaluation, a liver biopsy was performed. Liver biopsy showed hepatocellular necrosis with centrilobular cholestasis without any evidence of graft rejection. However, with normal level of tacrolimus, the biopsy was suggestive of drug-induced liver injury. Thus, tacrolimus dose was reduced, resulting in improved liver function tests and patient discharge from the hospital. Tacrolimus is an effective immunosuppressant after liver transplant and has the ability to treat early acute rejection. The patient's liver biopsy showed hepatocellular necrosis with centrilobular cholestasis without any evidence of graft rejection. Cholestatic liver injury after tacrolimus usually resolves after dose reduction or by switching to another agent. With demonstrated tacrolimus-induced toxicity in liver transplant recipients, despite normal serum levels, transplant physicians should keep high index of suspicion regarding toxicity in the posttransplant setting.

Ursodeoxycholic acid and 18ß-glycyrrhetinic acid alleviate ethinylestradiol-induced cholestasis via downregulating RORγt and CXCR3 signaling pathway in iNKT cells.

Estrogen-induced intrahepatic cholestasis (IHC) is a mild but potentially serious risk and urges for new therapeutic targets and effective treatment. Our previous study demonstrated that RORγt and CXCR3 signaling pathway of invariant natural killer T (iNKT) 17 cells play pathogenic roles in 17α-ethinylestradiol (EE)-induced IHC. Ursodeoxycholic acid (UDCA) and 18ß-glycyrrhetinic acid (GA) present a protective effect on IHC partially due to their immunomodulatory properties. Hence in present study, we aim to investigate the effectiveness of UDCA and 18ß-GA in vitro and verify the accessibility of the above targets. Biochemical index measurement indicated that UDCA and 18ß-GA presented efficacy to alleviate EE-induced cholestatic cytotoxicity. Both UDCA and 18ß-GA exhibited suppression on the CXCL9/10-CXCR3 axis, and significantly restrained the expression of RORγt in vitro. In conclusion, our observations provide new therapeutic targets of UDCA and 18ß-GA, and 18ß-GA as an alternative treatment for EE-induced cholestasis.

Allocholic acid protects against α-naphthylisothiocyanate-induced cholestasis in mice by ameliorating disordered bile acid homeostasis.

Cholestasis is a pathological condition characterized by disruptions in bile flow, leading to the accumulation of bile acids (BAs) in hepatocytes. Allocholic acid (ACA), a unique fetal BA known for its potent choleretic effects, reappears during liver regeneration and carcinogenesis. In this research, we investigated the protective effects and underlying mechanisms of ACA against mice with cholestasis brought on by α-naphthylisothiocyanate (ANIT). To achieve this, we combined network pharmacology, targeted BA metabolomics, and molecular biology approaches. The results demonstrated that ACA treatment effectively reduced levels of serum AST, ALP, and DBIL, and ameliorated the pathological injury caused by cholestasis. Network pharmacology analysis suggested that ACA primarily regulated BA and salt transport, along with the signaling pathway associated with bile secretion, to improve cholestasis. Subsequently, we examined changes in BA metabolism using UPLC-MS/MS. The findings indicated that ACA pretreatment induced alterations in the size, distribution, and composition of the liver BA pool. Specifically, it reduced the excessive accumulation of BAs, especially cholic acid (CA), taurocholic acid (TCA), and ß-muricholic acid (ß-MCA), facilitating the restoration of BA homeostasis. Furthermore, ACA pretreatment significantly downregulated the expression of hepatic BA synthase Cyp8b1, while enhancing the expression of hepatic efflux transporter Mrp4, as well as the renal efflux transporters Mdr1 and Mrp2. These changes collectively contributed to improved BA efflux from the liver and enhanced renal elimination of BAs. In conclusion, ACA demonstrated its potential to ameliorate ANIT-induced liver damage by inhibiting BA synthesis and promoting both BA efflux and renal elimination pathways, thus, restoring BA homeostasis.

Exploring the effect and mechanism of cucurbitacin B on cholestatic liver injury based on network pharmacology and experimental verification.

ETHNOPHARMACOLOGICAL RELEVANCE: Cholestatic liver injury (CLI) is a pathologic process with the impairment of liver and bile secretion and excretion, resulting in an excessive accumulation of bile acids within the liver, which leads to damage to both bile ducts and hepatocytes. This process is often accompanied by inflammation. Cucumis melo L is a folk traditional herb for the treatment of cholestasis. Cucurbitacin B (CuB), an important active ingredient in Cucumis melo L, has significant anti-inflamamatory effects and plays an important role in diseases such as neuroinflammation, skin inflammation, and chronic hepatitis. Though numerous studies have confirmed the significant therapeutic effect of CuB on liver diseases, the impact of CuB on CLI remains uncertain. Consequently, the objective of this investigation is to elucidate the therapeutic properties and potential molecular mechanisms underlying the effects of CuB on CLI. AIM OF THE STUDY: The aim of this paper was to investigate the potential protective mechanism of CuB against CLI. METHODS: First, the corresponding targets of CuB were obtained through the SwissTargetPrediction and SuperPre online platforms. Second, the DisGeNET database, GeneCards database, and OMIM database were utilized to screen therapeutic targets for CLI. Then, protein-protein interaction (PPI) was determined using the STRING 11.5 data platform. Next, the OmicShare platform was employed for the purpose of visualizing the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. The molecular docking technique was then utilized to evaluate the binding affinity existing between potential targets and CuB. Subsequently, the impacts of CuB on the LO2 cell injury model induced by Lithocholic acid (LCA) and the CLI model induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) were determined by evaluating inflammation in both in vivo and in vitro settings. The potential molecular mechanism was explored by real-time quantitative polymerase chain reaction (RT-qPCR) and western blot (WB) techniques. RESULTS: A total of 122 CuB targets were collected and high affinity targets were identified through the PPI network, namely TLR4, STAT3, HIF1A, and NFKB1. GO and KEGG analyses indicated that the treatment of CLI with CuB chiefly involved the inflammatory pathway. In vitro study results showed that CuB alleviated LCA-induced LO2 cell damage. Meanwhile, CuB reduced elevated AST and ALT levels and the release of inflammatory factors in LO2 cells induced by LCA. In vivo study results showed that CuB could alleviate DDC-induced pathological changes in mouse liver, inhibit the activity of serum transaminase, and suppress the liver and systemic inflammatory reaction of mice. Mechanically, CuB downregulated the IL-6, STAT3, and HIF-1α expression and inhibited STAT3 phosphorylation. CONCLUSION: By combining network pharmacology with in vivo and in vitro experiments, the results of this study suggested that CuB prevented the inflammatory response by inhibiting the IL-6/STAT3/HIF-1α signaling pathway, thereby demonstrating potential protective and therapeutic effects on CLI. These results establish a scientific foundation for the exploration and utilization of natural medicines for CLI.

Effects of Yinzhihuang on Alleviating Cyclosporine A-Induced Cholestatic Liver Injury via Farnesoid X Receptor-Mediated Regulation of Transporters and Enzymes in Vitro and in Vivo.

Yinzhihuang (YZH), a traditional Chinese medicine prescription, was widely used to treat cholestasis. Cholestatic liver injury limited the use of the immunosuppressive drug cyclosporine A (CsA) in preventing organ rejection after solid organ transplantation. Clinical evidences suggested that YZH could enhance bile acids and bilirubin clearance, providing a potential therapeutic strategy against CsA-induced cholestasis. Nevertheless, it remains unclear whether YZH can effectively alleviate CsA-induced cholestatic liver injury, as well as the molecular mechanisms responsible for its hepatoprotective effects. The purpose of the present study was to investigate the hepatoprotective effects of YZH on CsA-induced cholestatic liver injury and explore its molecular mechanisms in vivo and vitro. The results demonstrated that YZH significantly improved the CsA-induced cholestatic liver injury and reduced the level of liver function markers in serum of Sprague-Dawley (SD) rats. Targeted protein and gene analysis indicated that YZH increased bile acids and bilirubin efflux into bile through the regulation of multidrug resistance-associated protein 2 (Mrp2), bile salt export pump (Bsep), sodium taurocholate cotransporting polypeptide (Ntcp) and organic anion transporting polypeptide 2 (Oatp2) transport systems, as well as upstream nuclear receptors farnesoid X receptor (Fxr). Moreover, YZH modulated enzymes involved in bile acids synthesis and bilirubin metabolism including Cyp family 7 subfamily A member 1 (Cyp7a1) and uridine 5'-diphosphate (UDP) glucuronosyltransferase family 1 member A1 (Ugt1a1). Furthermore, the active components geniposidic acid, baicalin and chlorogenic acid exerted regulated metabolic enzymes and transporters in LO2 cells. In conclusion, YZH may prevent CsA-induced cholestasis by regulating the transport systems, metabolic enzymes, and upstream nuclear receptors Fxr to restore bile acid and bilirubin homeostasis. These findings highlight the potential of YZH as a therapeutic intervention for CsA-induced cholestasis and open avenues for further research into its clinical applications.

Hepatic interleukin 32 attenuates liver injury through repression of necroptosis in cholestasis.

OBJECTIVE: We aimed to evaluate the association between interleukin (IL)-32 and necroptosis in cholestatic liver injury. METHODS: Levels of necroptosis-related markers in cholestatic and control patients, including the receptor-interacting serine-threonine kinase 3 (RIPK3), receptor-interacting serine-threonine kinase 1 (RIPK1), and mixed lineage kinase domain-like (MLKL) were measured. Animal experiments in C57BL/6J and transgenic mice with IL32ß/γ overexpression were also conducted to confirm the effect of IL-32 on necroptosis in cholestasis, which was induced by α-naphthylisothiocyanate (ANIT) and 1% lithocholic acid (LCA). PLC/PRF/5-ASBT and primary mouse hepatocytes were utilized for the investigation of the regulation and mechanism of IL-32 in cholestasis. RESULTS: In the liver tissues of cholestatic patients, the mRNA and protein expressions of RIPK1, RIPK3, and MLKL were increased and associated with IL-32 expression. In addition, expressions of these indicators in the liver of 1% LCA- and ANIT-induced mouse models were significantly increased, while they were markedly decreased in hIL32ßLTg and hIL32γLTg mice. After bile acid stimulation, IL-32 and phosphorylated Akt (p-Akt) expressions significantly elevated in a dose-dependent manner. After treated with tumor necrosis factor (TNF)-α, IL-32 inhibited MLKL expression in primary mouse hepatocytes. CONCLUSION: IL-32 is negatively associated with necroptosis in cholestatic patients. Moreover, IL-32 is induced by p-Akt and effectively attenuates necroptosis in ANIT- or 1% LCA-induced cholestasis.

A Case of Progressive Cholestatic Drug-Induced Liver Injury Due to Terbinafine.

INTRODUCTION: Terbinafine is commonly prescribed for onychomycosis. It rarely leads to severe, prolonged cholestatic drug-induced liver injury. Clinicians should remain vigilant for this complication. CASE PRESENTATION: A 62-year-old woman was started on terbinafine and developed mixed hepatocellular and cholestatic drug-induced liver injury, confirmed on liver biopsy. The injury became predominantly cholestatic. Unfortunately, she developed coagulopathy with elevated international normalized ratio and progressive drug-induced liver injury with severely elevated alkaline phosphatase and total bilirubin, requiring repeat liver biopsy. Fortunately, she did not develop acute liver failure. DISCUSSION: Prior case reports and series have documented severe cholestatic drug-induced liver injury (although with lesser degree of bilirubin elevation) due to terbinafine, which has very rarely been associated with acute liver failure, need for liver transplantation, and/or death. CONCLUSIONS: Non-acetaminophen drug-induced liver injury is idiosyncratic. Complications including acute liver failure and vanishing bile duct syndrome can be slow to develop, so monitoring for them is important over longitudinal follow-up.

The role of biliary events in treatment and survival of patients with advanced pancreatic ductal adenocarcinoma.

BACKGROUND: Systemic chemotherapy (CT) is the treatment of choice for advanced pancreatic ductal adenocarcinoma (PDAC). Biliary obstruction is common in this setting and may interfere with CT administration due to jaundice or cholangitis related to biliary stent malfunction. AIMS: To evaluate the impact of biliary events on CT administration and survival in patients with stage III-IV PDAC. METHODS: Patients enrolled in a randomized trial of nab-paclitaxel plus gemcitabine with/without capecitabine and cisplatin in advanced PDAC were included. Data on management of jaundice, biliary stents/complications and CT were prospectively collected and retrospectively analyzed. Modified overall (mOS) and progression-free (mPFS) survival were evaluated. RESULTS: Eighty-eight patients met the inclusion criteria (50% females; median age 65years). Seven of eight (87.5%) patients who placed plastic stents developed biliary complications versus 14/30 (46.7%) with metallic stents (p = 0.071). Patients without biliary complications completed planned CT in 64.2% versus 47.6% of cases (p = 0.207). CT completion was related to longer mOS (17 vs 12 months, p = 0.005) and mPFS (9 vs 6 months, p = 0.011). mOS was shorter when biliary complications occurred (12 vs 17 months, p = 0.937), as was mPFS (6 vs 8 months, p = 0.438). CONCLUSION: Complications related to biliary obstruction influence chemotherapy completion and survival in patients with advanced PDAC.

AMP-activated protein kinase-farnesoid X receptor pathway contributes to oleanolic acid-induced liver injury.

Natural pentacyclic triterpenoid oleanolic acid (OA) is used as an over-the-counter drug for acute and chronic hepatitis. However, clinical use of OA-containing herbal medicines has been reported to cause cholestasis, and the specific mechanism is unknown. The purpose of this study was to explore how OA causes cholestatic liver injury via the AMP-activated protein kinase (AMPK)-farnesoid X receptor (FXR) pathway. In animal experiments, it was found that OA treatment activated AMPK and decreased FXR and bile acid efflux transport proteins expression. When intervened with the specific inhibitor Compound C (CC), it was observed that AMPK activation was inhibited, the reduction of FXR and bile acid efflux transport protein expression was effectively alleviated, serum biochemical indicators were significantly reduced, and liver pathological damage brought about by OA was effectively ameliorated. In addition, OA was found to downregulate the expression of FXR and bile acid efflux transport proteins by activating the ERK1/2-LKB1-AMPK pathway in cellular experiments. The ERK1/2 inhibitor U0126 was used to pretreat primary hepatocytes, and this drastically reduced the phosphorylation levels of LKB1 and AMPK. The inhibition effects of OA on FXR and bile acid efflux transport proteins were also effectively alleviated after pretreatment with CC. In addition, OA-induced downregulation of FXR gene and protein expression levels was significantly prevented after silencing AMPKα1 expression in AML12 cells. Our study demonstrated that OA inhibited FXR and bile acid efflux transporters through the activation of AMPK, thus leading to cholestatic liver injury.

iNKT17 cells play a pathogenic role in ethinylestradiol-induced cholestatic hepatotoxicity.

IL-17 is closely associated with inflammation in intrahepatic cholestasis (IHC). Targeting IL-17 ameliorates IHC in mice. Invariant natural killer T (iNKT) cells are predominantly enriched in the liver and they mediate drug-induced liver injury through their secreted cytokines. However, whether iNKT17 cells are involved in ethinylestradiol (EE)-induced IHC remains unclear. In the present study, the administration of EE (10 mg/kg in vivo and 6.25 µM in vitro) promoted the activation and expansion of iNKT17 cells, which contributed to a novel hepatic iNKT17/Treg imbalance. iNKT cell-deficient Jα18-/- mice and the RORγt inhibitor digoxin (20 µg) alleviated EE-induced cholestatic hepatotoxicity and downregulated the IL-17 signalling pathway. In contrast, the co-administration of EE with recombinant IL-17 (1 µg) to Jα18-/- mice induced cholestatic hepatotoxicity and increased the infiltration of hepatic neutrophils and monocytes. Importantly, the administration of IL-17-/- iNKT cells (3.5 × 105) to Jα18-/- mice resulted in the attenuation of hepatotoxicity and the recruitment of fewer hepatic neutrophils and monocytes than the adoptive transfer of wild-type iNKT cells. These results indicated that iNKT17 cells could exert pathogenic effects. The recruitment and activation of iNKT17 cells could be attributed to the high level of CXCR3 expression on their surface. CXCL10 deficiency ameliorated EE-induced cholestatic liver damage, reduced hepatic CXCR3+ iNKT cells and inhibited RORγt expression. These findings suggest that iNKT17 cells play a key role in EE-induced cholestatic liver injury via CXCR3-mediated recruitment and activation. Our study provides new insights and therapeutic targets for cholestatic diseases.

Calculus Bovis Sativus alleviates estrogen cholestasis-induced gut and liver injury in rats by regulating inflammation, oxidative stress, apoptosis, and bile acid profiles.

ETHNOPHARMACOLOGICAL RELEVANCE: Natural Calculus Bovis (NCB) is a traditional Chinese medicine used for anti-inflammation, treating fever, pain, sedation, and recovering hepatobiliary function. Calculus Bovis Sativus (CBS), produced from in vitro artificial cultivation by bioengineering techniques, acts as an ideal substitute for NCB when treating various diseases. AIM OF THE STUDY: Gut-liver injury is an important pathological feature of several cholestatic liver diseases, including estrogen-induced cholestasis (EIC). The strong link between cholestatic liver injury and intestinal damage emphasizes the need of considering gut-liver integrity during treatment. The purpose of this study is to look into the pharmacological activities of CBS on EIC-induced gut and liver damage. MATERIALS AND METHODS: EIC-induced cholestatic rats were given oral gavage daily for five days with or without CBS (150 mg/kg). The liver/body weight, serum biochemistry, and tissue histopathology were then evaluated. Quantitative real-time PCR, Western blot analyses, and immunofluorescence were used to determine the gene expression associated with pathological alterations of the liver and intestine in EIC-induced cholestatic rats. Bile acid profiles within enterohepatic circulation were detected by liquid chromatography-mass spectrometry. RESULTS: CBS significantly reduced relative liver weight, restored serum biochemistry levels, and improved the hepatic and intestinal pathological damage in EIC model rats. CBS reduced EIC-induced hepatic inflammation by inactivation of the NF-κB signaling and inhibition of TNFα, IL-1ß, and IL-6 expression. CBS alleviated EIC-induced hepatic and intestinal oxidative stress by regulating Nrf2-GCLM/GCLC and Nrf2-HO-1 pathways, respectively. CBS treatment upregulated Bcl-2 and downregulated Bax and cleaved caspase3 to improve EIC-induced hepatic and intestinal cell apoptosis. Additionally, CBS reversed the disorders of bile acid profiles in the enterohepatic circulation by reducing bile acid accumulation in the liver and plasma and increasing bile excretion and intestinal reabsorption of bile acids. CONCLUSION: CBS alleviates EIC-induced hepatic and intestinal injury through regulating inflammation, oxidative stress, apoptosis, and bile acid profiles. These results suggest that CBS or drugs targeting the gut-liver axis may be effective therapeutic agents for cholestasis.

Severe azathioprine-induced liver injury 22 months after initiation of treatment.

Drug-induced liver injury (DILI) is the leading cause of acute liver failure in high-income countries. Acute cholestasis is one of the most common forms of hepatotoxicity induced by azathioprine. It usually begins during the first year of treatment, with most cases reported during the first month. We describe an uncommon case of DILI that occurred after 22 months of drug administration. A woman in her 50s was hospitalised because of jaundice and asthenia. She had been treated with azathioprine for myasthenia gravis during the last 2 years. Acute cholestatic injury was diagnosed. After ruling out most common causes of cholestasis, azathioprine was withdrawn and subsequent histological findings in liver biopsy were consistent with drug-induced cholestatic liver damage. After discontinuation of azathioprine, biochemical parameters progressively normalised and remarkable clinical improvement was achieved. With this report, we suggest that azathioprine should be considered among the causes of liver injury, despite long treatment duration.

Silymarin protects the liver from α-naphthylisothiocyanate-induced cholestasis by modulating the expression of genes involved in bile acid homeostasis.

Cholestasis is characterized by impaired bile flow which results in inflammation, cirrhosis, and ultimately liver failure. The current study is aimed to evaluate the anti-cholestatic effect of silymarin against α-naphthylisothiocyanate (ANIT) induced cholestasis. Mice were gavaged with various doses of silymarin or ursodeoxycholic acid (UDCA) for 19 days. Then they were challenged with α-naphthylisothiocyanate (ANIT) and after 48 hours the animals were sacrificed to obtain blood and liver sections. Serum levels of bilirubin, aspartate transaminase (AST), alanine transaminase (ALP), and liver histology were analyzed. mRNA expression of selected transporters (Bile salt export pump (BSEP) and sodium taurocholate cotransporting polypeptide (NTCP)) and proteins (farnesoid x receptor (FXR) and Cytochrome P450 Family 7 Subfamily A Member 1 (Cyp7a1)) involved in bile acids biosynthesis, excretion and uptake were also evaluated by quantitative PCR. The results indicated that the serum levels of bilirubin, AST, and ALP were significantly higher in a cholestatic model group as compared to an untreated control group. However, in silymarin groups, the serum level of these parameters is significantly lower than in a cholestatic model group. Liver histology also showed that silymarin prevents ANIT-induced hepatic injury. mRNA expression of FXR, BSEP, and NTCP was downregulated and expression of Cyp7a1 was upregulated in a cholestatic model group as compared to an untreated control group. However, in silymarin treatment groups, the expression of FXR, BSEP and NTCP was upregulated and the expression of Cyp7a1 was downregulated as compared to the cholestatic model group. In conclusion, silymarin could alleviate hepatic injury by modulating the expression of genes involved in bile acid homeostasis.

L-carbocisteine can cause cholestasis with vanishing bile duct syndrome in children: A case report.

RATIONALE: Vanishing bile duct syndrome (VBDS) is the acquired progressive destruction and disappearance of intrahepatic interlobular bile ducts in the absence of underlying liver or biliary tract disease, causing chronic cholestasis. Infections, drugs, toxins, malignant diseases, and certain immunological processes are associated with the development of this syndrome. There have been no reports of children developing VBDS as a consequence of the administration of L-carbocisteine. PATIENT CONCERNS: A 9-year-old Japanese girl presented with fever, jaundice, and skin rash. Laboratory investigations revealed elevated levels of serum transaminases, γ-glutamyltransferase, and bilirubin. Histopathological features were consistent with a diagnosis of VBDS. Drug-induced lymphocyte stimulation tests (DLST) were positive for L-carbocisteine. DIAGNOSIS: VBDS caused by L-carbocisteine. INTERVENTIONS: Ursodeoxycholic acid and discontinuation of L-carbocisteine. OUTCOMES: The patient responded to treatment based upon discontinuation of L-carbocisteine and administration of ursodeoxycholic acid. Her transaminase and bilirubin levels were normalized gradually. LESSONS: Physicians should be aware of the fact that L-carbocisteine can cause cholestasis with VBDS in children.

Protective Effect of Rhus chinensis Mill. Fruits on 3,5-Diethoxycarbonyl-1,4-Dihydrocollidine-Induced Cholestasis in Mice via Ameliorating Oxidative Stress and Inflammation.

This study focused on the preventive effects of the extracts of Rhus chinensis Mill. (RCM) fruits on cholestasis induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) in mice. The results showed that RCM extracts could significantly ameliorate DDC-induced cholestasis via multiple mechanisms, including (1) alleviating liver damage via enhancing antioxidant capacity, such as increasing the contents of glutathione, superoxide dismutase, and catalase and inhibiting the levels of malondialdehyde; (2) preventing liver inflammation by suppressing NF-κB pathway and reducing proinflammatory cytokines secretion (e.g., tumor necrosis factor-α, interleukin-1ß, and interleukin-6); (3) inhibiting liver fibrosis and collagen deposition by regulating the expression of transforming growth factor-ß and α-smooth muscle actin; (4) modulating abnormal bile acid metabolism through increasing the expression of bile salt export pump and multidrug resistance-associated protein 2. This study was the first to elucidate the potential preventive effect of RCM extracts on DDC-induced cholestasis in mice from multiple pathways, which suggested that RCM fruits could be considered as a potential dietary supplement to prevent cholestasis.

Fluorofenidone ameliorates cholestasis and fibrosis by inhibiting hepatic Erk/-Egr-1 signaling and Tgfß1/Smad pathway in mice.

Cholestasis is characterized by intrahepatic accumulation of bile acids (BAs), resulting in liver injury, fibrosis, and liver failure. To date, only ursodeoxycholic acid and obeticholic acid have been approved for the treatment of cholestasis. As fluorofenidone (AKF-PD) was previously reported to play significant anti-fibrotic and anti-inflammatory roles in various diseases, we investigated whether AKF-PD ameliorates cholestasis. A mouse model of cholestasis was constructed by administering a 0.1 % 3,5-diethoxycarbonyl-1,4-dihydroxychollidine (DDC) diet for 14 days. Male C57BL/6 J mice were treated with either AKF-PD or pirfenidone (PD) orally in addition to the DDC diet. Serum and liver tissues were subsequently collected and analyzed. We found that AKF-PD significantly reduced the levels of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and total bile salts (TBA), as well as hepatic bile acids (BAs) levels. Hepatic histological analyses demonstrated that AKF-PD markedly attenuated hepatic inflammation and fibrosis. Further mechanistic analyses revealed that AKF-PD markedly inhibited expression of Cyp7a1, an enzyme key to BAs synthesis, by increasing Fxr nuclear translocation, and decreased hepatic inflammation by attenuating Erk/-Egr-1-mediated expression of inflammatory cytokines and chemokines Tnfα, Il-1ß, Il-6, Ccl2, Ccl5 and Cxcl10. Moreover, AKF-PD was found to substantially reduce liver fibrosis via inhibition of Tgfß1/Smad pathway in our mouse model. Here, we found that AKF-PD effectively attenuates cholestasis and hepatic fibrosis in the mouse model of DDC-induced cholestasis. As such, AKF-PD warrants further investigation as a candidate drug for treatment of cholestasis.

Cloxacillin-induced acute vanishing bile duct syndrome: A case study and literature review.

Ductopenia is often regarded as a chronic process where ≥50% of portal tracts lack bile ducts, which is also known as vanishing bile duct syndrome (VBDS). One aetiology is drug-induced liver injury. Cloxacillin, an antistaphylococcal penicillin, typically causes "bland" cholestasis. We present the first case of cloxacillin-induced acute ductopenia or VBDS and a review of published cloxacillin-induced liver injuries. A 66-year-old woman with no prior liver disease, but known penicillin allergy, was treated for postcarotid angioplasty staphylococcal infection with 6 weeks of cloxacillin. She presented with a 2-week history of weakness and jaundice. Laboratory work-up showed elevated liver enzymes with a cholestatic pattern, hyperbilirubinemia and eosinophilia. She required ICU transfer for hypotension and was started empirically on prednisone. Liver biopsy revealed severe centrilobular cholestasis, mild necroinflammation and ductopenia with epithelial injury, but no ductular reaction. Two months later she was discharged on hydrocortisone and ursodiol with persistently elevated alkaline phosphatase and bilirubin. She was considered for liver transplantation but died of liver failure 4 months later. Four additional articles were found with histopathologic descriptions of cloxacillin-related liver injury. These included portal inflammation, cholestasis and mild necroinflammation. Clinical features were reported in two cases; both had mild symptoms with cholestatic liver enzymes and hyperbilirubinemia. Both patients recovered completely within 10-60 days. Cloxacillin-induced cholestasis can be secondary to acute ductopenia, which can result in worse clinical outcomes than previously described "bland" cholestasis. Liver biopsy is recommended to identify cases with acute VBDS.