Interleukin-6 decreases senescence and increases telomerase activity in malignant human cholangiocytes

BACKGROUND/AIMS: Cellular senescence results in irreversible growth arrest. In malignant cells, senescence is prevented by maintenance of chromosomal length by telomerase activity. Telomerase activity is increased in malignant, but not in normal cholangiocytes. Interleukin-6 (IL-6) is an autocrine promoter of cholangiocarcinoma growth. Our aims were to assess the relationship between IL-6 activated p38 mitogen-activated protein kinase (MAPK) pathways and senescence in malignant cholangiocytes. METHODS: Cell senescence and telomerase activity was assessed in Mz-ChA-1 malignant human cholangiocytes. The effect of inhibitors of p38 MAPK and telomerase activity on cell proliferation was assessed, and the interaction between these inhibitors was quantitated by median effects analysis. RESULTS: Mz-ChA-1 cells rapidly underwent senescence during repeated passaging. IL-6 increased telomerase activity and decreased cellular senescence during repeated passaging. However, basal telomerase activity was increased by inhibition of p38 MAPK. Inhibition of telomerase activity decreased IL-6 induced proliferation and had a synergistic effect with p38 MAPK inhibitors. Thus, IL-6 increases telomerase activity independent of p38 MAPK signaling and maintenance of telomerase activity promotes cholangiocarcinoma growth. CONCLUSION: Enhanced telomerase activity in response to IL-6 stimulation can prevent cellular senescence and thereby contribute to cholangiocarcinoma growth. Inhibition of telomerase activity may therefore be therapeutically useful in biliary tract malignancies.     

PyMEGABASE: Predicting Cell-Type-Specific Structural Annotations of Chromosomes Using the Epigenome

The folding patterns of interphase genomes in higher eukaryotes, as obtained from DNA-proximity-ligation or Hi-C experiments, are used to classify loci into structural classes called compartments and subcompartments. These structurally annotated (sub) compartments are known to exhibit specific epigenomic characteristics and cell-type-specific variations. To explore the relationship between genome structure and the epigenome, we present PyMEGABASE (PYMB), a maximum-entropy-based neural network model that predicts (sub) compartment annotations of a locus based solely on the local epigenome, such as ChIP-Seq of histone post-translational modifications. PYMB builds upon our previous model while improving robustness, capability to handle diverse inputs and user-friendly implementation. We employed PYMB to predict subcompartments for over a hundred human cell types available in ENCODE, shedding light on the links between subcompartments, cell identity, and epigenomic signals. The fact that PYMB, trained on data for human cells, can accurately predict compartments in mice suggests that the model is learning underlying physicochemical principles transferable across cell types and species. Reliable at higher resolutions (up to 5 kbp), PYMB is used to investigate compartment-specific gene expression. Not only can PYMB generate (sub) compartment information without Hi-C experiments, but its predictions are also interpretable. Analyzing PYMB’s trained parameters, we explore the importance of various epigenomic marks in each subcompartment prediction. Furthermore, the predictions of the model can be used as input for OpenMiChroM software, which has been calibrated to generate three-dimensional structures of the genome. Detailed documentation of PYMB is available at https://pymegabase.readthedocs.io, including an installation guide using pip or conda, and Jupyter/Colab notebook tutorials.     

Cadherin-6 is a putative tumor suppressor and target of epigenetically dysregulated miR-429 in cholangiocarcinoma

Cholangiocarcinoma (CC) is a rare malignancy of the extrahepatic or intrahepatic biliary tract with an outstanding poor prognosis. Non-surgical therapeutic regimens result in minimally improved survival of CC patients. Global genomic analyses identified a few recurrently mutated genes, some of them in genes involved in epigenetic patterning. In a previous study, we demonstrated global DNA methylation changes in CC, indicating major contribution of epigenetic alterations to cholangiocarcinogenesis. Here, we aimed at the identification and characterization of CC-related, differentially methylated regions (DMRs) in potential microRNA promoters and of genes targeted by identified microRNAs. Twenty-seven hypermethylated and 13 hypomethylated potential promoter regions of microRNAs, known to be associated with cancer-related pathways like Wnt, ErbB, and PI3K-Akt signaling, were identified. Selected DMRs were confirmed in 2 independent patient cohorts. Inverse correlation between promoter methylation and expression suggested miR-129-2 and members of the miR-200 family (miR-200a, miR-200b, and miR-429) as novel tumor suppressors and oncomiRs, respectively, in CC. Tumor suppressor genes deleted in liver cancer 1 (DLC1), F-box/WD-repeat-containing protein 7 (FBXW7), and cadherin-6 (CDH6) were identified as presumed targets in CC. Tissue microarrays of a representative and well-characterized cohort of biliary tract cancers (n=212) displayed stepwise downregulation of CDH6 and association with poor patient outcome. Ectopic expression of CDH6 on the other hand, delayed growth in the CC cell lines EGI-1 and TFK-1, together suggesting a tumor suppressive function of CDH6. Our work represents a valuable repository for the study of epigenetically altered miRNAs in cholangiocarcinogenesis and novel putative, CC-related tumor suppressive miRNAs and oncomiRs.     

Multiomics metabolic and epigenetics regulatory network in cancer: A systems biology perspective

Genetic, epigenetic, and metabolic alterations are all hallmarks of cancer. However, the epigenome and metabolome are both highly complex and dynamic biological networks in vivo. The interplay between the epigenome and metabolome contributes to a biological system that is responsive to the tumor microenvironment and possesses a wealth of unknown biomarkers and targets of cancer therapy. From this perspective, we first review the state of high-throughput biological data acquisition(i.e. multiomics data)and analysis(i.e. computational tools) and then propose a conceptual in silico metabolic and epigenetic regulatory network(MER-Net) that is based on these current high-throughput methods. The conceptual MER-Net is aimed at linking metabolomic and epigenomic networks through observation of biological processes, omics data acquisition, analysis of network information, and integration with validated database knowledge. Thus, MER-Net could be used to reveal new potential biomarkers and therapeutic targets using deep learning models to integrate and analyze large multiomics networks. We propose that MER-Net can serve as a tool to guide integrated metabolomics and epigenomics research or can be modified to answer other complex biological and clinical questions using multiomics data.     

Shotgun proteomics of human bile in hilar cholangiocarcinoma

The need to find biomarkers for hepatobiliary diseases including cholangiocarcinoma (CCA) has led to an interest in using bile as a proximal fluid in biomarker discovery experiments, although there are inherent challenges both in its acquisition and analysis. The study described here greatly extends previous studies that have started to characterise the bile proteome. Bile from four patients with hilar CCA was depleted of albumin and immunoglobulin G and analysed by GeLC-MS/MS. The number of proteins identified per bile sample was between 378 and 741. Overall, the products of 813 unique genes were identified, considerably extending current knowledge of the malignant bile proteome. Of these, 268 were present in at least 3 out of 4 patients. This data set represents the largest catalogue of bile proteins to date and together with other studies in the literature constitutes an important prelude to the potential promise of expression proteomics and subsequent validation studies in CCA biomarker discovery.     

肠道菌群的研究进展及与胆管癌的关系分析

人体肠道内生存着种类繁多的微生物,它们在维持人体健康及疾病的发生发展中发挥着重要的作用,被称为人体的“第二基因库”及“隐藏的器官”。近年来,随着人类微生物组计划（HMP）及人类肠道宏基因组计划（MetaHIT）的实施,关于肠道微生态的研究逐渐走向高潮,其中肠道菌群在胆管癌的发生发展中的作用逐渐显露出来。胆管癌作为一种预后极差且发病率逐渐升高的消化系肿瘤,其与肠道菌群的关系,特别是细菌逆行性感染及肠道微生态失衡在胆管癌病情进展中的作用应引起大家的重视。本文就肠道菌群的分布情况及其在胆管癌发生发展中的影响、检测方法及研究展望作一综述。     

Targeting chromatin dysregulation in organ fibrosis

Fibrosis leads to destruction of organ architecture accompanied by chronic inflammation and loss of function. Fibrosis affects nearly every organ in the body and accounts for ∼45% of total deaths worldwide. Over the past decade, tremendous progress has been made in understanding the basic mechanisms leading to organ fibrosis. However, we are limited with therapeutic options and there is a significant need to develop highly effective anti-fibrotic therapies. Recent advances in sequencing technologies have advanced the burgeoning field of epigenetics towards molecular understanding at a higher resolution. Here we provide a comprehensive review of the recent advances in chromatin regulatory processes, specifically DNA methylation, post-translational modification of histones, and chromatin remodeling complexes in kidney, liver and lung fibrosis. Although this research field is young, we discuss new strategies for potential therapeutic interventions for treating organ fibrosis.     

Targeting cholangiocarcinoma

Cholangiocarcinoma (CCA) represents a diverse group of epithelial cancers associated with the biliary tract, and can best be stratified anatomically into intrahepatic (iCCA), perihilar (pCCA) and distal (dCCA) subsets. Molecular profiling has identified genetic aberrations associated with these anatomic subsets. For example, IDH catalytic site mutations and constitutively active FGFR2 fusion genes are predominantly identified in iCCA, whereas KRAS mutations and PRKACB fusions genes are identified in pCCA and dCCA. Clinical trials targeting these specific driver mutations are in progress. However, The Tumor Genome Atlas (TCGA) marker analysis of CCA also highlights the tremendous molecular heterogeneity of this cancer rendering comprehensive employment of targeted therapies challenging. CCA also display a rich tumor microenvironment which may be easier to target. For example, targeting cancer associated fibroblasts for apoptosis with BH3-mimetics and/or and reversing T-cell exhaustion with immune check point inhibitors may help aid in the treatment of this otherwise devastating malignancy. Combinatorial therapy attacking the tumor microenvironment plus targeted therapy may help advance treatment for CCA. This article is part of a Special Issue entitled: Cholangiocytes in Health and Disease edited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.     

肝吸虫与胆管癌

肝吸虫病是一种重要的人兽共患寄生虫病,也是我国重点防治主要食源性寄生虫病之一。肝吸虫感染被国际癌症研究中心（IARC）确定为Ⅰ类致癌因素,其所致胆管癌是肝吸虫感染的严重后果之一。本文就目前国内外肝吸虫感染的流行病学及其诱发胆管癌的发病机制研究进展和成果做一综述,为今后学者进一步深入研究肝吸虫病提供参考资料。     

Epigenetic regulation in the carcinogenesis of cholangiocarcinoma

Cholangiocarcinoma (CCA) is a malignancy arising from the epithelial cells lining the biliary tract. Despite the existence of variation in incidence and etiology worldwide, its incidence is increasing globally in the past few decades. Surgery is the only curative treatment option for a minority of patients presented with early disease; while moderate effective chemotherapy remains the standard care for patients with locally advanced or metastatic diseases. In this article, we briefly review the molecular alterations that have been described in CCAs focusing on the role of epigenetic modification, including promoter methylation inactivation, histone modification and microRNA, in the carcinogenesis and progression of CCAs. This article is part of a Directed Issue entitled: Epigenetics dynamics in development and disease.     

Proteomic analysis of cholangiocarcinoma cell line

Cholangiocarcinoma (CCA), a malignant tumor derived from bile duct epithelium, occurs with a higher incidence in tropical countries, such as Thailand. Distinguishing CCA from hepatocellular carcinoma (HCC) of the liver often requires the use of histochemistry, so molecular markers for diagnosis and prognosis are still required. In this study, the two-dimensional (2-D) protein map of a Thai human bile duct epithelial carcinoma cell line (HuCCA-1) has been compared to human hepatocellular carcinoma cell lines (HepG2 and HCC-S102) and a human breast epithelial cancer cell line (MCF-7). Our results show that HuCCA-1 expressed a unique pattern of proteins. Forty-three major proteins were identified by matching to the map of MCF-7, and by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) and electrospray ionization-tandem MS (ESI-MS/MS). Cytokeratins CK8 and CK18 were overexpressed in both HuCCA-1 and HCC, while CK7 and CK19 were only expressed in HuCCA-1. Four specific proteins with MW/pI 57.2/5.21 (U1, vimentin), 42.2/6.20 (U2), 43.2/6.20 (U3, EF-TU), and 42.2/6.40 (U4, unidentified) were absent from HepG2. U2 showed high expression in HuCCA-1, while U1 and U4 showed high expression in HCC-S102. U2 could be separated in 2 proteins, U2/1 (alpha-enolase) and U2/2 (not identified) by using IPG pH 4-7. Galectin-3 showed high expression level in HuCCA-1 by 1-DE immunodetection, and gave only one spot with MW 32.9 kDa and pI 8.29 on 2-DE immunoblotting, Thus, certain proteins, namely CK7, CK19, U2/2 and galectin-3, may be good markers useful for differential diagnosis of cholangiocarcinoma compared to hepatocellular carcinoma.     

Autophagy in intra-hepatic cholangiocarcinoma

Intra-hepatic cholangiocarcinoma (IHCC) is a primary cancer of the liver that shares histological features with the hepatic bile ducts from which it is thought to arise. The incidence of this disease is increasing, possibly related to increased inflammatory liver diseases such as viral hepatitis and steatohepatitis (commonly called “fatty liver”), induced by obesity, diabetes, and other metabolic derangements. Its prognosis is generally poor with early metastasis and presently there are limited effective treatments. A basic understanding of the disease has long been hampered by limited tumor cell lines and good model systems. Similarly, such limitations have obstructed new therapeutic inroads.     

Autophagy in intra-hepatic cholangiocarcinoma

Intra-hepatic cholangiocarcinoma (IHCC) is a primary cancer of the liver that shares histological features with the hepatic bile ducts from which it is thought to arise. The incidence of this disease is increasing, possibly related to increased inflammatory liver diseases such as viral hepatitis and steatohepatitis (commonly called “fatty liver”), induced by obesity, diabetes, and other metabolic derangements. Its prognosis is generally poor with early metastasis and presently there are limited effective treatments. A basic understanding of the disease has long been hampered by limited tumor cell lines and good model systems. Similarly, such limitations have obstructed new therapeutic inroads.     

Cholangiocytes in the pathogenesis of primary sclerosing cholangitis and development of cholangiocarcinoma

Primary sclerosing cholangitis (PSC) is an idiopathic cholangiopathy strongly associated with inflammatory bowel disease (IBD) and characterized by cholestasis, chronic immune infiltration and progressive fibrosis of the intrahepatic and extrahepatic bile ducts. PSC confers a high risk of cholangiocarcinoma (CCA) with PSC-CCA representing the leading cause of PSC-associated mortality. PSC-CCA is derived from cholangiocytes and associated progenitor cells – a heterogeneous group of dynamic epithelial cells lining the biliary tree that modulate the composition and volume of bile production by the liver. Infection, inflammation and cholestasis can trigger cholangiocyte activation leading to an increased expression of adhesion and antigen-presenting molecules as well as the release of various inflammatory and fibrogenic mediators. As a result, activated cholangiocytes engage in a myriad of cellular processes, including hepatocellular proliferation, apoptosis, angiogenesis and fibrosis. Cholangiocytes can also regulate the recruitment of immune cells, mesenchymal cells, and endothelial cells that participate in tissue repair and destruction in settings of persistent inflammation. In PSC, the role of cholangiocytes and the mechanisms governing their transformation to PSC-CCA are unclear however localization of disease suggests that cholangiocytes are a key target and potential regulator of hepatobiliary immunity, fibrogenesis and tumorigenesis. Herein, we summarize mechanisms of cholangiocyte activation in PSC and highlight new insights into disease pathways that may contribute to the development of PSC-CCA. This article is part of a Special Issue entitled: Cholangiocytes in Health and Disease edited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.     

胆管癌化疗患者小肠细菌过度生长情况

目的 探讨胆管癌化疗患者化疗前后小肠细菌过度生长（SIBO）情况及应用酪酸梭菌治疗SIBO后能否改善肿瘤患者的临床症状。方法 选择62例于我院接受化疗的胆管癌患者作为观察组,选择同时段内于我院体检的健康志愿者50例作为对照组。观察组患者化疗前后及对照组均接受葡萄糖氢呼气试验（GHBT）检测SIBO阳性率。将观察组化疗后SIBO阳性患者分为益生菌治疗组和安慰剂组,比较其治疗前后SIBO阳性率及临床症状评分的变化。结果 观察组患者化疗前SIBO阳性率为45.16%（28/62）,化疗后SIBO阳性率为67.74%（42/62）。对照组SIBO阳性率为16.00％（8/50）。观察组患者化疗前后SIBO阳性率显著高于对照组,差异均有统计学意（均P＜0.05）。观察组患者化疗后SIBO阳性率高于化疗前,差异有统计学意义（χ2=6.43,P＜0.05）。益生菌治疗组患者经酪酸梭菌治疗后SIBO转阴率为52.38%（11/21）,安慰剂组为14.29%（3/21）,且两组患者给药后的临床症状积分对比差异有统计学意义（均P＜0.05）。结论 胆管癌患者SIBO阳性率更高,化疗可进一步加重SIBO的发生。酪酸梭菌治疗胆管癌患者化疗后SIBO效果较好,可提高SIBO转阴率,改善胆管癌患者化疗后腹痛、腹胀等不适症状。     

Synthesis of 14-deoxy-11,12-didehydroandrographolide analogues as potential cytotoxic agents for cholangiocarcinoma

A series of 14-deoxy-11,12-didehydroandrographolide analogues were synthesized from naturally occurring andrographolide and their cytotoxicity evaluated against nine cancer cell lines including cholangiocarcinoma. Analogues 5a and 5b exhibited the most potent cytotoxicity with ED₅₀s of 3.37 and 3.08?μM on KKU-M213 cell lines and 2.93 and 3.27?μM on KKU-100 cell lines, respectively. Selective cytotoxicity on cholangiocarcinoma cell lines identified in this study highlight the importance of structural modification in the development of drugs for this cancer.     

胆道微生态的研究进展

人体微生态是微生物与宿主在长期相互作用过程中形成的特殊生态系统,在人体的健康与疾病中起着重要作用。目前对消化系统微生态的研究主要集中在肠道,而胆道系统可能存在独特的微生态体系,它们与胆道系统的健康和疾病之间的关系尚不明确,研究人员探讨了胆道的微生物群与胆囊结石、胆总管结石、Oddi括约肌受损、胆总管结石复发、原发性硬化性胆管炎和胆道肿瘤等肝胆疾病的发生发展、诊断和治疗的关系。本文从健康人群、肝移植供肝者的胆道及以上疾病着手,对胆道微生态的相关研究和进展作一综述。