BACKGROUND: Ischemia of the bile duct is a common feature in liver disease and transplantation, which represents a major cause of morbidity and mortality, especially after liver transplantation. Detailed knowledge of its pathogenesis remains incomplete due to the lack of appropriate in vitro models.
METHODS: To recapitulate biliary damage induced by ischemia and reperfusion in vitro, human intrahepatic cholangiocyte organoids (ICOs) were grown at low oxygen levels of 1% up to 72 h, followed by re-oxygenation at normal levels.
RESULTS: ICOs stressed by ischemia and subsequent re-oxygenation represented the dynamic change in biliary cell proliferation, upregulation of epithelial-mesenchymal transition (EMT)-associated markers, and the evocation of phase-dependent cell death programs similar to what is described in patients. Clinical-grade alpha-1 antitrypsin was identified as a potent inhibitor of both ischemia-induced apoptosis and necroptosis.
CONCLUSIONS: These findings demonstrate that ICOs recapitulate ischemic cholangiopathy in vitro and enable drug assessment studies for the discovery of new therapeutics for ischemic cholangiopathies.
BACKGROUND: Dutch Digestive FoundationMLDS D16-26; TKI-LSH (Topconsortium Kennis en Innovatie-Life Sciences & Health) grant RELOAD, EMC-LSH19002; Medical Delta program \"Regenerative Medicine 4D\"; China Scholarship Council No. 201706230252.

Clonorchiasis caused by Clonorchis sinensis is a mainly foodborne parasitic disease. It can lead to hepatobiliary duct inflammation, fibrosis, obstructive jaundice, liver cirrhosis, and even cholangiocarcinoma. Interleukin (IL)-10 is an immune-regulatory cytokine which plays an immunosuppressive role during infection. Our previous study found that IL-10 was increased in mice with C. sinensis infection. However, the role and mechanism of IL-10 playing in hepatobiliary injury induced by C. sinensis infection remain unknown. Herein, Il10+/+ mice and Il10+/- C57BL/6J mice were infected with C. sinensis. It was found that IL-10 deficiency aggravated biliary hyperplasia and exacerbated periductal fibrosis induced by C. sinensis infection. Moreover, IL-10 deficiency increased CD4+T cells and CD8+T cells but not macrophages in the liver of mice with infection. There were no apparent differences in Th1 and Treg cells between Il10+/+ and Il10+/- mice infected with C. sinensis. However, the proportion of Th17 cells in CD4+T cells in Il10+/- infected mice was significantly higher than that in Il10+/+ infected mice. IL-10 deficiency also enhanced the increase of Th17 cells induced by ESPs stimulation in vitro. Taken together, our results suggest that IL-10 plays a protective role in hepatobiliary injury in C57BL/6J mice induced by C. sinensis infection via inhibiting Th17 cells, which could deepen our understanding of the immunopathology of clonorchiasis.

BACKGROUND: Clonorchiasis caused by Clonorchis sinensis is a zoonotic parasitic disease characterized by cholangitis, biliary proliferation, biliary fibrosis, and even cholangiocarcinoma. Our previous study showed that the expression of interleukin (IL)-33 is increased in both humans and mice infected by C. sinensis, suggesting that IL-33 is potentially involved in the pathogenesis of clonorchiasis. However, the roles and potential mechanism of IL-33 underlying remain unknown.
METHODS: Wild-type (WT) and IL-33 knockout (KO) mice (BALB/c female mice) were orally infected with 45 metacercariae of C. sinensis for 8 weeks. Biliary injuries and fibrosis were extensively evaluated. Hepatic type II cytokines (IL-4, IL-13, and IL-10) were detected by ELISA.
RESULTS: For wild-type mice, we found that the mice infected with C. sinensis showed severe biliary injuries and fibrosis compared with the normal mice that were free from worm infection. In addition, the levels of type II cytokines such as IL-4, IL-13, and IL-10 in infected wild-type mice were significantly higher than in the control mice without infection (P < 0.05). However, IL-33 deficiency (IL-33 KO) prevents the augmentation of biliary injuries and fibrosis caused by C. sinensis infection. Furthermore, the increased levels of these type II cytokines induced by worm infection were also reversed in IL-33 KO mice.
CONCLUSIONS: Our present study demonstrates that IL-33 contributes to the pathogenesis of C. sinensis-induced biliary injuries and repair, which can potentially orchestrate type 2 responses. These findings highlight the pathophysiological role of IL-33 in the progression of clonorchiasis.

Liver and bile duct diseases often are associated with extensive cell death of cholangiocytes. Necroptosis represents a common mode of programmed cell death in cholangiopathy, however, detailed mechanistic knowledge is limited owing to the lack of appropriate in vitro models. To address this void, we investigated whether human intrahepatic cholangiocyte organoids (ICOs) can recapitulate cholangiopathy-associated necroptosis and whether this model can be used for drug screening.
We evaluated the clinical relevance of necroptosis in end-stage liver diseases and liver transplantation by immunohistochemistry. Cholangiopathy-associated programmed cell death was evoked in ICOs derived from healthy donors or patients with primary sclerosing cholangitis or alcoholic liver diseases by the various stimuli.
The expression of key necroptosis mediators, receptor-interacting protein 3 and phosphorylated mixed lineage kinase domain-like, in cholangiocytes during end-stage liver diseases was confirmed. The phosphorylated mixed lineage kinase domain-like expression was etiology-dependent. Gene expression analysis confirmed that primary cholangiocytes are more prone to necroptosis compared with primary hepatocytes. Both apoptosis and necroptosis could be specifically evoked using tumor necrosis factor α and second mitochondrial-derived activator of caspases mimetic, with or without caspase inhibition in healthy and patient-derived ICOs. Necroptosis also was induced by ethanol metabolites or human bile in ICOs from donors and patients. The organoid cultures further uncovered interdonor variable and species-specific drug responses. Dabrafenib was identified as a potent necroptosis inhibitor and showed a protective effect against ethanol metabolite toxicity.
Human ICOs recapitulate cholangiopathy-associated necroptosis and represent a useful in vitro platform for the study of biliary cytotoxicity and preclinical drug evaluation.

OBJECTIVE: To identify the species and abundance of microbes in the bile of patients with biliary injury after liver transplantation. To explore the potential microbial involvement, we evaluated the differences in biliary microbiota in patients with and without biliary injury after liver transplantation.
METHODS: Bile was collected by percutaneous transhepatic cholangiography drainage from 5 liver transplant recipients with biliary injury (case group) and from the T-tube in 4 liver transplant recipients without biliary injury (control group). 16S rRNA sequencing was performed on the bile samples.
RESULTS: The biliary bacterial phyla in patients after liver transplantation were Firmicutes, Proteobacteria, Actinobacteria, Bacteroidetes, and Acidobacteria in order of abundance from highest to lowest. There were differences in genus Prevotella (P = 0.03), Porphyromonas (P = 0.02), and Fusobacterium (P = 0.02) between the two groups, which were significantly abundant in the case group. There was no significant difference in the diversity analysis between the two groups (P > 0.05). The terpenoid backbone biosynthesis pathway and the histidine degradation pathway were significantly enriched in the bile samples in the case group. Compared with the control group, the pyruvate ferredoxin oxidoreductase pathway was depleted in the samples from the case group.
CONCLUSIONS: This is the first bile microbiota report using high-throughput 16S rRNA gene sequencing of bile material in liver transplant recipients. There were significant differences in bacterial abundance between patients with and without biliary tract injury after liver transplantation but no significant differences in the diversity. KEGG analysis showed that there were differences in metabolic pathways between the two groups.

The purpose of this study is to introduce and evaluate a new technique of repairing bile ducts by the tubular gastric wall with a vascularized pedicle. Both the end-to-end bile duct repair and Roux-en-Y hepatoenterostomy have limitations in the treatment of benign bile duct strictures after cholecystectomy. There are no other good choices to manage these cases, especially the bile duct transection injuries or partly missing common bile duct or hepatic duct. Eleven patients with partly missing common bile ducts in the Chinese People\'s Liberation Army General Hospital between January 2007 and December 2012 were retrospectively analyzed. The study comprised 8 females and 3 males, whose age ranged from 29 to 56 years. All patients underwent successful bile duct repair. The time of operations ranged from 210 minutes to 240 minutes. The maximal blood loss was less than 220 ml. There was no perioperative mortality and no case of gastric fistula. Postoperative complications occurred in 3 patients, including wound infection, bile leakage, and erosive gastritis. All complications were cured by conservative treatment. The mean follow-up time was 42 months. One patient was classified as Terblanche\'s grade II and 10 patients were classified as Terblanche\'s grade I. The observations indicate that this technique is a feasible and effective choice to manage low level biliary stricture after cholecystectomy, especially suitable to repair bile duct transection injuries or partly missing common bile duct or hepatic duct.

Nonanastomotic strictures (NAS) are common biliary complications after liver transplantation (LT). Delayed rearterialization induces biliary injury in several hours. However, whether this injury can be prolonged remains unknown. The correlation of this injury with NAS occurrence remains obscure. Different delayed rearterialization times were compared using a porcine LT model. Morphological and functional changes in bile canaliculus were evaluated by transmission electron microscopy and real-time PCR. Immunohistochemistry and TUNEL were performed to validate intrahepatic bile duct injury. Three months after LT was performed, biliary duct stricture was determined by cholangiography; the tissue of common bile duct was detected by real-time PCR. Bile canaliculi were impaired in early postoperative stage and then exacerbated as delayed rearterialization time was prolonged. Nevertheless, damaged bile canaliculi could fully recover in subsequent months. TNF-α and TGF-β expressions and apoptosis cell ratio increased in the intrahepatic bile duct only during early postoperative period in a time-dependent manner. No abnormality was observed by cholangiography and common bile duct examination after 3 months. Delayed rearterialization caused temporary injury to bile canaliculi and intrahepatic bile duct in a time-dependent manner. Injury could be fully treated in succeeding months. Solo delayed rearterialization cannot induce NAS after LT.

OBJECTIVE: Endoscopic placement of covered self-expandable metal stent (SEMS) has gained popularity in the management of benign biliary strictures (BBS). The existing SEMS has been designed primarily to palliate malignant biliary obstruction and has a high frequency of stent migration, difficulty in retrieval and stricture recurrence after stent removal. This study aimed to design a novel retrievable SEMS dedicated to the treatment of extrahepatic BBS and evaluate its clinical efficacy and safety.
METHODS: A short fully covered SEMS (FCSEMS) with a retrieval lasso was designed for the specific treatment of BBS. A total of 45 patients with segmental extrahepatic BBS were included in this study. The stent was placed entirely inside the bile duct with only the retrieval lasso extending from the papilla. The stents were recommended to be in situ for 6 to 12 months before removal.
RESULTS: The FCSEMS was successfully placed in all 45 patients. In all, 33 patients had their FCSEMS successfully removed after a mean period of 8.6 ± 3.7 (range 2-15.5) months. Stent migration occurred in 9.1% of the patients. During a mean follow-up of 18.9 months after stent removal, recurrent stricture was found in 2 (6.1%) patients and was successfully treated with a second FCSEMS. Overall, the strictures resolved in 30/33 (90.9%) patients.
CONCLUSIONS: Intraductal placement of a short FCSEMS is suitable for the treatment of segmental extrahepatic BBS. This new removable design offered prolonged stenting and drainage for BBS for up to one year with minimal complications.