Pathogenesis Of Primary Sclerosing Cholangitis Research Articles

Prevotella copri ameliorates cholestasis and liver fibrosis in primary sclerosing cholangitis by enhancing the FXR signalling pathway

Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease characterized by bile duct inflammation, fibrosis, bile acid (BA) metabolism disorders and gut microbiota dysbiosis. At present, the aetiology and pathogenesis of PSC are not clear, and there is no specific or effective treatment available. Therefore, new research perspectives are needed to explore effective methods to treat PSC and improve symptoms. The intestinal microbiota of patients with PSC is known to be significantly different from that of healthy people. By comparing differentially abundant bacterial genera in PSC patients, it was found that the abundance of Prevotella copri (P. copri) was significantly decreased, suggesting that this species may have a protective effect against PSC disease. Therefore, comprehensively exploring the role and possible function of P. copri in the disease process is worthwhile. In this study, a PSC mouse model was established by feeding mice a customized diet supplemented with 0.1% (w/w) 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) for one week, and the abundance of P. copri was confirmed to be decreased in this model. Previous studies in patients and animal models have demonstrated that gut microbiota intervention is an acceptable treatment for some diseases. We found that intervention with P. copri could significantly improve cholestasis and liver fibrosis by enhancing the FXR-related signalling pathway in PSC mice. Together, through the overall effect of P. copri on intestinal microbiota structure and its association with BAs, we speculate that P. copri intervention might be as potential biological treatment of PSC.

Indole-3-Carboxaldehyde Restores Gut Mucosal Integrity and Protects from Liver Fibrosis in Murine Sclerosing Cholangitis.

Primary sclerosing cholangitis (PSC) is a long-term liver disease characterized by a progressive course of cholestasis with liver inflammation and fibrosis. Intestinal barrier dysfunction has been implicated in the pathogenesis of PSC. According to the “leaky gut” hypothesis, gut inflammation alters the permeability of the intestinal mucosa, with the translocation of gut-derived products that enter the enterohepatic circulation and cause hepatic inflammation. Thus, the administration of molecules that preserve epithelial barrier integrity would represent a promising therapeutic strategy. Indole-3-carboxaldehyde (3-IAld) is a microbial-derived product working at the interface between the host and the microbiota and is able to promote mucosal immune homeostasis in a variety of preclinical settings. Herein, by resorting to a murine model of PSC, we found that 3-IAld formulated for localized delivery in the gut alleviates hepatic inflammation and fibrosis by modulating the intestinal microbiota and activating the aryl hydrocarbon receptor-IL-22 axis to restore mucosal integrity. This study points to the therapeutic potential of 3-IAld in liver pathology.

The Management of Cholestatic Liver Diseases: Current Therapies and Emerging New Possibilities.

Primary biliary cholangitis (PBC) and primary sclerosing cholangitis (PSC) are two chronic cholestatic liver diseases affecting bile ducts that may progress to biliary cirrhosis. In the past few years, the increasing knowledge in the pathogenesis of both diseases led to a growing number of clinical trials and possible new targets for therapy. In this review, we provide an update on the treatments in clinical use and summarize the new drugs in trials for PBC and PSC patients. Farnesoid X Receptor (FXR) agonists and Pan-Peroxisome Proliferator-Activated Receptor (PPAR) agonists are the most promising agents and have shown promising results in both PBC and PSC. Fibroblast Growth Factor 19 (FGF19) analogues also showed good results, especially in PBC, while, although PBC and PSC are autoimmune diseases, immunosuppressive drugs had disappointing effects. Since the gut microbiome could have a potential role in the pathogenesis of PSC, recent research focused on molecules that could change the microbiome, with good results. The near future of the medical management of these diseases may include new treatments or a combination of multiple drugs targeting different signaling pathways at different stages of the diseases.

Downregulation of TGR5 (GPBAR1) in biliary epithelial cells contributes to the pathogenesis of sclerosing cholangitis

Primary sclerosing cholangitis (PSC) is characterized by chronic inflammation and progressive fibrosis of the biliary tree. The bile acid receptor TGR5 (GPBAR1) is found on biliary epithelial cells (BECs), where it promotes secretion, proliferation and tight junction integrity. Thus, we speculated that changes in TGR5-expression in BECs may contribute to PSC pathogenesis. TGR5-expression and -localization were analyzed in PSC livers and liver tissue, isolated bile ducts and BECs from Abcb4-/-, Abcb4-/-/Tgr5Tg and ursodeoxycholic acid (UDCA)- or 24-norursodeoxycholic acid (norUDCA)-fed Abcb4-/- mice. The effects of IL8/IL8 homologues on TGR5 mRNA and protein levels were studied. BEC gene expression was analyzed by single-cell transcriptomics (scRNA-seq) from distinct mouse models. TGR5 mRNA expression and immunofluorescence staining intensity were reduced in BECs of PSC and Abcb4-/- livers, in Abcb4-/- extrahepatic bile ducts, but not in intrahepatic macrophages. No changes in TGR5 BEC fluorescence intensity were detected in liver tissue of other liver diseases, including primary biliary cholangitis. Incubation of BECs with IL8/IL8 homologues, but not with other cytokines, reduced TGR5 mRNA and protein levels. BECs from Abcb4-/- mice had lower levels of phosphorylated Erk and higher expression levels of Icam1, Vcam1 and Tgfβ2. Overexpression of Tgr5 abolished the activated inflammatory phenotype characteristic of Abcb4-/- BECs. NorUDCA-feeding restored TGR5-expression levels in BECs in Abcb4-/- livers. Reduced TGR5 levels in BECs from patients with PSC and Abcb4-/- mice promote development of a reactive BEC phenotype, aggravate biliary injury and thus contribute to the pathogenesis of sclerosing cholangitis. Restoration of biliary TGR5-expression levels represents a previously unknown mechanism of action of norUDCA. Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease-associated with progressive inflammation of the bile duct, leading to fibrosis and end-stage liver disease. Bile acid (BA) toxicity may contribute to the development and disease progression of PSC. TGR5 is a membrane-bound receptor for BAs, which is found on bile ducts and protects bile ducts from BA toxicity. In this study, we show that TGR5 levels were reduced in bile ducts from PSC livers and in bile ducts from a genetic mouse model of PSC. Our investigations indicate that lower levels of TGR5 in bile ducts may contribute to PSC development and progression. Furthermore, treatment with norUDCA, a drug currently being tested in a phase III trial for PSC, restored TGR5 levels in biliary epithelial cells.

Single-cell atlas of hepatic T cells reveals expansion of liver-resident naive-like CD4+ T cells in primary sclerosing cholangitis

Background & AimsLittle is known about the composition of intrahepatic immune cells and their contribution to the pathogenesis of primary sclerosing cholangitis (PSC). Herein, we aimed to create an atlas of intrahepatic T cells and thereby perform an in-depth characterization of T cells in inflamed human liver.MethodsDifferent single-cell RNA sequencing methods were combined with in silico analyses on intrahepatic and peripheral T cells from patients with PSC (n = 11) and healthy donors (HDs, n = 4). Multi-parameter flow cytometry and functional in vitro experiments were conducted on samples from patients with PSC (n = 24), controls with other liver diseases and HDs.ResultsWe identified a population of intrahepatic naive-like CD4+ T cells, which was present in all liver diseases tested, but particularly expanded in PSC. This population had a transcriptome and T cell receptor repertoire similar to circulating naive T cells but expressed a set of genes associated with tissue residency. Their periductal location supported the concept of tissue-resident naive-like T cells in livers of patients with PSC. Trajectory inference suggested that these cells had the developmental propensity to acquire a T helper 17 (TH17) polarization state. Functional and chromatin accessibility experiments revealed that circulating naive T cells in patients with PSC were predisposed to polarize towards TH17 cells.ConclusionWe report the first atlas of intrahepatic T cells in PSC, which led to the identification of a previously unrecognized population of tissue-resident naive-like T cells in the inflamed human liver and to the finding that naive CD4+ T cells in PSC harbour the propensity to develop into TH17 cells.Lay summaryThe composition of intrahepatic immune cells in primary sclerosing cholangitis (PSC) and their contribution to disease pathogenesis is widely unknown. We analysed intrahepatic T cells and identified a previously uncharacterized population of liver-resident CD4+ T cells which are expanded in the livers of patients with PSC compared to healthy liver tissue and other liver diseases. These cells are likely to contribute to the pathogenesis of PSC and could be targeted in novel therapeutic approaches.

Senescent cholangiocytes release extracellular vesicles that alter target cell phenotype via the epidermal growth factor receptor.

Primary sclerosing cholangitis (PSC) is a chronic liver disease characterized by peribiliary inflammation and fibrosis. Cholangiocyte senescence is a prominent feature of PSC. Here, we hypothesize that extracellular vesicles (EVs) from senescent cholangiocytes influence the phenotype of target cells. EVs were isolated from normal human cholangiocytes (NHCs), cholangiocytes from PSC patients and NHCs experimentally induced to senescence. NHCs, malignant human cholangiocytes (MHCs) and monocytes were exposed to 108 EVs from each donor cell population and assessed for proliferation, MAPK activation and migration. Additionally, we isolated EVs from plasma of wild-type and Mdr2-/- mice (a murine model of PSC), and assessed mouse monocyte activation. EVs exhibited the size and protein markers of exosomes. The number of EVs released from senescent human cholangiocytes was increased; similarly, the EVs in plasma from Mdr2-/- mice were increased. Additionally, EVs from senescent cholangiocytes were enriched in multiple growth factors, including EGF. NHCs exposed to EVs from senescent cholangiocytes showed increased NRAS and ERK1/2 activation. Moreover, EVs from senescent cholangiocytes promoted proliferation of NHCs and MHCs, findings that were blocked by erlotinib, an EGF receptor inhibitor. Furthermore, EVs from senescent cholangiocytes induced EGF-dependent Interleukin 1-beta and Tumour necrosis factor expression and migration of human monocytes; similarly, Mdr2-/- mouse plasma EVs induced activation of mouse monocytes. The data continue to support the importance of cholangiocyte senescence in PSC pathogenesis, directly implicate EVs in cholangiocyte proliferation, malignant progression and immune cell activation and migration, and identify novel therapeutic approaches for PSC.

Gut microbiome in primary sclerosing cholangitis: A review.

Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease characterized by biliary inflammation and stricturing. Exploration of the pathogenesis of PSC in light of its association with inflammatory bowel disease (IBD) and the “gut-liver” axis is an emerging area of interest. A growing number of studies have begun to elucidate the role of the gut microbiota, its metabolites and its influence on host immune responses in the development of PSC and PSC-IBD. Studies of the fecal microbiota have highlighted enriched levels of certain species, including Veillonella, Streptococcus and Enterococcus, among others. A heightened immune response to enteric dysbiosis and bacterial translocation have also been implicated. For example, Klebsiella pneumoniae strains derived from gnotobiotic mice transplanted with PSC-IBD microbiota were found to induce pore formation in human intestinal epithelial cells and enhanced Th17 responses. Gut microbes have additionally been hypothesized to be implicated in PSC pathogenesis through their role in the synthesis of various metabolites, including bile acids (BAs), which function as signaling molecules with important gut and hepatic effects. An expanded knowledge of the gut microbiome as it relates to PSC offers critical insight into the development of microbe-altering therapeutic interventions, such as antibiotics, nutritional interventions and fecal microbial transplantation. Some of these have already shown some preliminary evidence of benefit. Despite exciting progress in the field, much work remains to be done; areas that are particularly lacking include functional characterization of the microbiome and examination of pediatric populations. In this review, we summarize studies that have investigated the microbiome in PSC and PSC-IBD as well as putative mechanisms, including the potential role of metabolites, such as BAs. We then briefly review the evidence for interventions with microbe-altering properties for treating PSC.

The transcription factor ETS1 promotes apoptosis resistance of senescent cholangiocytes by epigenetically up-regulating the apoptosis suppressor BCL2L1

Primary sclerosing cholangitis (PSC) is an idiopathic, progressive cholangiopathy. Cholangiocyte senescence is important in PSC pathogenesis, and we have previously reported that senescence is regulated by the transcription factor ETS proto-oncogene 1 (ETS1) and associated with overexpression of BCL2 like 1 (BCL2L1 or BCL-xL), an anti-apoptotic BCL2-family member. Here, we further explored the mechanisms regulating BCL-xL-mediated, apoptosis resistance in senescent cholangiocytes and uncovered that ETS1 and the histone acetyltransferase E1A-binding protein P300 (EP300 or p300) both promote BCL-xL transcription. Using immunofluorescence, we found that BCL-xL protein expression is increased both in cholangiocytes of livers from individuals with PSC and a mouse model of PSC. Using an in vitro model of lipopolysaccharide-induced senescence in normal human cholangiocytes (NHCs), we found increased BCL-xL mRNA and protein levels, and ChIP-PCRs indicated increased occupancy of ETS1, p300, and histone 3 Lys-27 acetylation (H3K27Ac) at the BCL-xL promoter. Using co-immunoprecipitation and proximity ligation assays, we further demonstrate that ETS1 and p300 physically interact in senescent but not control NHCs. Additionally, mutagenesis of predicted ETS1-binding sites within the BCL-xL promoter blocked luciferase reporter activity, and CRISPR/Cas9-mediated genetic deletion of ETS1 reduced senescence-associated BCL-xL expression. In senescent NHCs, TRAIL-mediated apoptosis was reduced ∼70%, and ETS1 deletion or RNAi-mediated BCL-xL suppression increased apoptosis. Overall, our results suggest that ETS1 and p300 promote senescent cholangiocyte resistance to apoptosis by modifying chromatin and inducing BCL-xL expression. These findings reveal ETS1 as a central regulator of both cholangiocyte senescence and the associated apoptosis-resistant phenotype.

Imaging Features of Primary Sclerosing Cholangitis: From Diagnosis to Liver Transplant Follow-up.

Primary sclerosing cholangitis (PSC) is a chronic progressive inflammatory disease of the bile ducts that leads to multifocal bile duct fibrosis, strictures, cholestasis, liver parenchymal changes, and ultimately cirrhosis. It more commonly occurs in young adults, with a variety of clinical and imaging manifestations. The cause of the disease is not known, but it has a strong association with inflammatory bowel disease and can overlap with other autoimmune diseases, including autoimmune hepatitis and immunoglobulin G4-related disease. Patients are predisposed to various hepatic and extrahepatic deteriorating complications, such as bile duct and gallbladder calculi, acute bacterial cholangitis, liver abscess, and portal hypertension, as well as malignancies including cholangiocarcinoma (CCA), gallbladder cancer, and colorectal carcinoma. Imaging has an essential role in diagnosis, surveillance, and detection of complications. MR cholangiopancreatography and endoscopic retrograde cholangiopancreatography have high specificity and sensitivity for detection of primary disease and assessment of disease progression. However, many patients with PSC are still diagnosed incidentally at US or CT. Novel imaging techniques such as transient elastography and MR elastography are used to survey the grade of liver fibrosis. Annual cancer surveillance is necessary in all PSC patients to screen for CCA and gallbladder cancer. Familiarity with PSC pathogenesis and imaging features across various classic imaging modalities and novel imaging techniques can aid in correct imaging diagnosis and guide appropriate management. The imaging features of the biliary system and liver parenchyma in PSC across various imaging modalities are reviewed. Imaging characteristics of the differential diagnosis of PSC, clinical associations, and complications are described. Finally, the role of imaging in evaluation of PSC progression, pre-liver transplant assessment, and post-liver transplant disease recurrence are discussed.©RSNA, 2019.

In vivo 1 H MRS of human gallbladder bile in understanding the pathophysiology of primary sclerosing cholangitis (PSC): Immune-mediated disease versus bile acid-induced injury.

Primary sclerosing cholangitis (PSC) has been considered to be either an "autoimmune disease" or a "bile acid-induced injury." In vitro MRS studies on PSC patients have limitations due to the contamination of bile with contrast agent (commonly administered during endoscopic retrograde cholangiopancreatography) and/or the use of patient cohorts with other diseases as controls. The objective of this study was to quantify biliary metabolites using in vivo 1 H MRS and gain insight into the pathogenesis of PSC. Biliary metabolites in 10 PSC patients and 14 healthy controls were quantified in vivo using 1 H MRS on a 3T MR scanner. The concentrations of total bile acids plus cholesterol, glycine-conjugated bile acids, taurine-conjugated bile acids, and choline-containing phospholipids (chol-PLs) were compared between the two groups. There were statistically significant decreases in the levels of the above mentioned biliary metabolites in the PSC patients compared with controls. The reduction in bile acid secretion in bile of PSC patients indicates accumulation of bile acids in hepatocytes. Moreover, reduction in the levels of chol-PLs in bile may increase the toxic effects of bile acids. Our findings suggest that the bile duct injury in PSC patients is most likely due to "bile acid-induced injury."

Gut pathobionts underlie intestinal barrier dysfunction and liver T helper 17 cell immune response in primary sclerosing cholangitis.

Primary sclerosing cholangitis (PSC) is a chronic inflammatory liver disease and its frequent complication with ulcerative colitis highlights the pathogenic role of epithelial barrier dysfunction. Intestinal barrier dysfunction has been implicated in the pathogenesis of PSC, yet its underlying mechanism remains unknown. Here, we identify Klebsiella pneumonia in the microbiota of patients with PSC and demonstrate that K. pneumoniae disrupts the epithelial barrier to initiate bacterial translocation and liver inflammatory responses. Gnotobiotic mice inoculated with PSC-derived microbiota exhibited T helper 17 (TH17) cell responses in the liver and increased susceptibility to hepatobiliary injuries. Bacterial culture of mesenteric lymph nodes in these mice isolated K. pneumoniae, Proteus mirabilis and Enterococcus gallinarum, which were prevalently detected in patients with PSC. A bacterial-organoid co-culture system visualized the epithelial-damaging effect of PSC-derived K. pneumoniae that was associated with bacterial translocation and susceptibility to TH17-mediated hepatobiliary injuries. We also show that antibiotic treatment ameliorated the TH17 immune response induced by PSC-derived microbiota. These results highlight the role of pathobionts in intestinal barrier dysfunction and liver inflammation, providing insights into therapeutic strategies for PSC.

M1-Polarized Macrophages Promote Self-Renewing Phenotype of Hepatic Progenitor Cells with Jagged1-Notch Signalling Involved: Relevance in Primary Sclerosing Cholangitis.

The immunologic interaction between parenchyma cells and encircling inflammatory cells is thought to be the most important mechanism of biliary damage and repair in primary sclerosing cholangitis (PSC). Monocytes/macrophages as master regulators of hepatic inflammation have been demonstrated to contribute to PSC pathogenesis. Macrophages coordinate with liver regeneration, and multiple phenotypes have been identified with diverse expressions of surface proteins and cytokine productions. We analyzed the expression of Notch ligand Jagged1 in polarized macrophages and investigated the relevance of Notch signalling activation in liver regeneration. M1 or M2 macrophages were generated from mouse bone marrow-derived macrophages (BMDMs) by classical or alternative activation, respectively. Then, the expression levels of Jagged1 (Jag1) of each phenotype were measured. The effects of polarized BMDMs on the expression of hepatic progenitor cell- (HPC-) specific markers and hairy and enhancer of split-1 (HES1) in HPCs in coculture were also analyzed. Monocyte-macrophage and Notch signalling-associated gene signatures were evaluated in the GEO database (access ID: GSE61260) by gene set enrichment analysis (GSEA). M1 macrophages were found associated with elevated Jag1 expression, which increased the fraction of HPC with self-renewing phenotypes (CD326+CD44+ or CD324+CD44+) and HES1 expression level in cocultured HPC. Blocking Jagged1 by siRNA or antibody in the coculture system attenuates HPC self-renewing phenotypes as well as HES1 expression in HPC. GSEA data show that macrophage activation and Notch signalling-associated gene signatures are enriched in PSC patients. These findings suggest that M1 macrophages promote an HPC self-renewing phenotype which is associated with Notch signalling activation within HPC. In the liver of PSC patients, the prevalence of activated macrophages, with M1 polarized accounting for the main part, is associated with increment of Notch signalling and enhancement of HPC self-renewal.

Inflammation and the Gut-Liver Axis in the Pathophysiology of Cholangiopathies.

Cholangiocytes, the epithelial cells lining the bile ducts, represent the unique target of a group of progressive diseases known as cholangiopathies whose pathogenesis remain largely unknown. In normal conditions, cholangiocytes are quiescent and participate to the final bile volume and composition. Following exogenous or endogenous stimuli, cholangiocytes undergo extensive modifications of their phenotype. Reactive cholangiocytes actively proliferate and release a set of proinflammatory molecules, which act in autocrine/paracrine manner mediating the cross-talk with other liver cell types and innate and adaptive immune cells. Cholangiocytes themselves activate innate immune responses against gut-derived microorganisms or bacterial products that reach the liver via enterohepatic circulation. Gut microbiota has been implicated in the development and progression of the two most common cholangiopathies, i.e., primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC), which have distinctive microbiota composition compared to healthy individuals. The impairment of intestinal barrier functions or gut dysbiosis expose cholangiocytes to an increasing amount of microorganisms and may exacerbate inflammatory responses thus leading to fibrotic remodeling of the organ. The present review focuses on the complex interactions between the activation of innate immune responses in reactive cholangiocytes, dysbiosis, and gut permeability to bacterial products in the pathogenesis of PSC and PBC.

Follicular T Helper Cell Signatures in Primary Biliary Cholangitis and Primary Sclerosing Cholangitis.

Primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC) are the most common cholestatic liver diseases. While PBC is generally accepted to be an autoimmune disorder characterized by pathognomonic autoantibodies against mitochondrial antigens, the pathogenesis of PSC is less precisely defined; however, some degree of altered immunity toward autoantigens has been suggested. Follicular T helper (Tfh) cells, a distinct clusters of differentiation (CD)4 T‐cell subset specialized in facilitating antibody responses, have been shown to contribute to humoral autoimmunity in various disorders; yet, there is only limited information on possible alterations of Tfh cells in the context of cholestatic liver diseases. Thus, we addressed this important question by analyzing the frequency, activation status, and function of Tfh cells and frequencies of regulatory follicular T helper (Tfr) cells in well‐defined cohorts of patients with PBC and patients with PSC. Interestingly, we observed a significant increase in circulating chemokine (C‐X‐C motif) receptor 5 (CXCR5)+programmed death 1 (PD‐1) +CD4+ Tfh cells in patients with PBC but not in those with PSC. Although the frequency of potentially pathogenic chemokine (C‐C motif) receptor 7 (CCR7)lowCXCR5+PD‐1+CD4+ Tfh cells was increased in both disorders compared to healthy donors, the increase was significantly more pronounced in PBC. Furthermore, in patients with PBC, Tfh cells displayed stronger expression of the activation markers OX40 and inducible costimulator of T cells, correlated with anti‐anti‐mitochondrial antibody M2 and immunoglobulin M titers, and were most significantly increased in patients with cirrhosis. Tfr cell numbers were similarly increased; however, Tfh/Tfr ratios were unaltered in PSC and PBC. These alterations did not correlate with increased secretion of the Tfh signature cytokine interleukin‐21 in sorted CD4 T cells. Conclusion: Significant alterations occur in the Tfh cell compartment in cholestatic liver diseases, suggesting that Tfh cells influence the pathogenesis of PBC and to a lesser extend PSC.

Differences in the Colon Microbiome between Patients with and without Recurrence of Primary Sclerosing Cholangitis

Background After liver transplantation (LT) for Primary Sclerosing Cholangitis (PSC), patients have a risk of up to 27% at recurrence of their liver disease (rPSC) with a negative impact on survival. The pathogenesis of both PSC and rPSC are largely unknown, but the idea of a multifactorial concept is broadly accepted. The gut microbiome is suggested to be one of these factors, by either altering the immune system and/or affecting the bile acid-gut microbiome axis. Coexisting Inflammatory Bowel Disease (IBD) in numerous patients with PSC emphasizes this theory. We have investigated whether the gut microbiome differs at time of transplant between patients with and without rPSC. Materials and Methods All patients who receive a LT at our center undergo a screening colonoscopy with biopsies. These biopsies, stored in FFPE, were used to extract bacterial DNA using the RTP Bacteria DNA Mini Kit (Stratec®). This DNA was used for 16S rRNA gene sequencing, revealing the microbiome of each patient. Using shrinkage-based variable selection we identified bacteria associated with rPSC. Recurrence PSC was defined according to the Mayo criteria, excluding secondary causes of post-LT biliary disease. Results and Discussion A total of 100 FFPE samples were available from patients who received a LT between 1987 and 2015 for PSC. Median time between colon sampling and LT was 7.2 months (range 1.7-32.8). The recurrence rate in this cohort was 14% (n=14) after a median of 5.0 years (range 0.5-10.1). We identified two bacteria (genus level) from different phyla that were significantly more abundant in the rPSC group: Microbacterium (phylum Actinobacteria) and Thermicanus (phylum Firmicutes). Log odds ratios were 174 and 55, respectively. Conclusion Despite similar disease pre-LT, we found that two genera, the Microbacterium and Thermicanus, were significantly more abundant in the pre-LT colon of patients who developed rPSC post-LT as compared to those who did not. This difference in the colon microbiome may contribute to the pathogenesis of rPSC, and might help to understand the link between IBD and rPSC.

Macrophages contribute to the pathogenesis of sclerosing cholangitis in mice

Macrophages contribute to liver disease, but their role in cholestatic liver injury, including primary sclerosing cholangitis (PSC), is unclear. We tested the hypothesis that macrophages contribute to the pathogenesis of, and are therapeutic targets for, PSC. Immune cell profile, hepatic macrophage number, localization and polarization, fibrosis, and serum markers of liver injury and cholestasis were measured in an acute (intrabiliary injection of the inhibitor of apoptosis antagonist BV6) and chronic (Mdr2-/- mice) mouse model of sclerosing cholangitis (SC). Selected observations were confirmed in liver specimens from patients with PSC. Because of the known role of the CCR2/CCL2 axis in monocyte/macrophage chemotaxis, therapeutic effects of the CCR2/5 antagonist cenicriviroc (CVC), or genetic deletion of CCR2 (Ccr2-/- mice) were determined in BV6-injected mice. We found increased peribiliary pro-inflammatory (M1-like) and alternatively-activated (M2-like) monocyte-derived macrophages in PSC compared to normal livers. In both SC models, genetic profiling of liver immune cells identified a predominance of monocytes/macrophages; immunohistochemistry confirmed peribiliary monocyte-derived macrophage recruitment (M1>M2-polarized), which paralleled injury onset and was reversed upon resolution in acute SC mice. PSC, senescent and BV6-treated human cholangiocytes released monocyte chemoattractants (CCL2, IL-8) and macrophage-activating factors in vitro. Pharmacological inhibition of monocyte recruitment by CVC treatment or CCR2 genetic deletion attenuated macrophage accumulation, liver injury and fibrosis in acute SC. Peribiliary recruited macrophages are a feature of both PSC and acute and chronic murine SC models. Pharmacologic and genetic inhibition of peribiliary macrophage recruitment decreases liver injury and fibrosis in mouse SC. These observations suggest monocyte-derived macrophages contribute to the development of SC in mice and in PSC pathogenesis, and support their potential as a therapeutic target. Primary sclerosing cholangitis (PSC) is an inflammatory liver disease which often progresses to liver failure. The cause of the disease is unclear and therapeutic options are limited. Therefore, we explored the role of white blood cells termed macrophages in PSC given their frequent contribution to other human inflammatory diseases. Our results implicate macrophages in PSC and PSC-like diseases in mice. More importantly, we found that pharmacologic inhibition of macrophage recruitment to the liver reduces PSC-like liver injury in the mouse. These exciting observations highlight potential new strategies to treat PSC.

The Microbiome in Primary Sclerosing Cholangitis: Current Evidence and Potential Concepts.

The close relationship between primary sclerosing cholangitis (PSC) and inflammatory bowel disease has inspired hypothetical models in which gut bacteria or bacterial products are key players in PSC pathogenesis. Several studies using high-throughput sequencing technology to characterize the gut microbiota in PSC have been published over the past years. They all report reduced diversity and significant shifts in the overall composition of the gut microbiota. However, it remains unclear as to whether the observed changes are primary or secondary to PSC development and further studies are needed to assess the biological implications of the findings. In the present article, we review the published data in perspective of similar studies in other diseases. We discuss aspects of methodology and study design that are relevant to interpretation of the data. Furthermore, we propose that interpretation and further assessments of findings are structured into conceptual compartments, and elaborate three such possible concepts relating to immune function (the "immunobiome"), host metabolism (the "endobiome"), and dietary and xenobiotic factors (the "xenobiome") in PSC.

Development and characterization of cholangioids from normal and diseased human cholangiocytes as an in vitro model to study primary sclerosing cholangitis

Primary sclerosing cholangitis (PSC) is an incurable, fibroinflammatory biliary disease for which there is no effective pharmacotherapy. We recently reported cholangiocyte senescence as an important phenotype in PSC while others showed that portal macrophages accumulate in PSC. Unfortunately, our ability to explore cholangiocyte senescence and macrophage accumulation has been hampered by limited in vitro models. Thus, our aim was to develop and characterize a three-dimensional (3D) model of normal and diseased bile ducts (cholangioids) starting with normal human cholangiocytes (NHC), senescent NHC (NHC-sen), and cholangiocytes from PSC patients. In 3D culture, NHCs formed spheroids of ~5000 cells with a central lumen of ~150 μm. By confocal microscopy and western blot, cholangioids retained expression of cholangiocyte proteins (cytokeratin 7/19) and markers of epithelial polarity (secretin receptor and GM130). Cholangioids are functionally active, and upon secretin stimulation, luminal size increased by ~80%. Cholangioids exposed to hydrogen peroxide exhibited cellular senescence and the senescence-associated secretory phenotype (SASP; increased IL-6, p21, SA‐β-Gal, yH2A.x and p16 expression). Furthermore, cholangioids derived from NHC-sen or PSC patients were smaller and had slower growth than the controls. When co-cultured with THP-1 macrophages, the number of macrophages associated with NHC-sen or PSC cholangioids was five- to seven-fold greater compared to co-culture with non-senescent NHC. We observed that NHC-sen and PSC cholangioids release greater number of extracellular vesicles (EVs) compared to controls. Moreover, conditioned media from NHC-sen cholangioids resulted in an ~2-fold increase in macrophage migration. In summary, we developed a method to generate normal and diseased cholangioids, characterized them morphologically and functionally, showed that they can be induced to senescence and SASP, and demonstrated both EV release and macrophage attraction. This novel model mimics several features of PSC, and thus will be useful for studying the pathogenesis of PSC and potentially identifying new therapeutic targets.

A Unique Triad of Ulcerative Colitis With Autoimmune Hemolytic Anemia and Primary Sclerosing Cholangitis

Inflammatory Bowel Diseases (IBD) lead to chronic intestinal inflammation along with extra-intestinal manifestations which could present prior to, subsequent with or after the diagnosis of IBD. We present a unique case of Ulcerative Colitis (UC), Autoimmune Hemolytic Anemia (AIHA) and Primary Sclerosing Cholangitis (PSC) diagnosed simultaneously in a patient. The unique triad of these diseases gives important clues to the immunological factors involved in the pathogenesis of these diseases. An 18-year-old male presented with 1 week of bloody diarrhea, abdominal cramps and nausea. Vitals were stable. Initial labs showed Hb 9.5 g/dL, MCV 102 fL, WBC 22.4 x 103/μL, total bilirubin 2.4 mg/dL, direct bilirubin 0.7 mg/dL, alkaline phosphatase 436 units/L, ALT 133 units/L and AST 83 units/L. Hepatitis panel and Clostridium difficile toxin assay were negative. Further work-up revealed reticulocyte count > 17%, LDH 529 units/L, haptoglobin < 10 mg/dl and positive direct Coomb's test. Peripheral smear showed hemolysis by a warm antibody. Hydrocortisone was started for AIHA. CT abdomen showed mild colonic wall thickening. Colonoscopy showed moderate pancolitis and biopsies showed findings consistent with UC. Mesalamine was started. MRCP showed irregular beaded appearance of intra and extrahepatic bile ducts. ERCP was performed and biopsies taken from the ducts showed chronic inflammation but no evidence of malignancy. ASMA and ANCA were positive. Liver biopsy ruled out overlap syndrome with autoimmune hepatitis. The patient clinically improved and was discharged to follow-up with a gastroenterologist. In patients with UC, extra-intestinal manifestations involving cutaneous, hepatobiliary and ocular systems are common. The concomitant presence of PSC is 2-7% and AIHA is 0.2 -1.7% in patients with UC. The extra-intestinal manifestations of UC could be due to associated autoimmune diseases or they could be immune-mediated due to damaged mucosal wall. The exact pathogenesis of PSC and AIHA is unknown. The absorption of toxins and antigenic substances from the mucosa in the circulation mediates inflammation in the biliary system leading to PSC. This also leads to warm anti-E antibody production which cross-react with intestinal microbiome and the E antigen at the surface of RBCs. In UC patients, a vigorous workup for early detection of other autoimmune diseases is important to improve the quality of life and to prevent mortality associated with them.Figure: Peripheral smear showing polychromatic RBCs and microspherocytes.Figure: Axial CT demonstrating mild wall thickening and featureless appearance of the rectosigmoid colon compatible with chronic ulcerative colitis (left). Descending colon at 4X magnification showing crypt distortion, cryptitis and crypt abscess (right).Figure: Left: Coronal thin section T2 weighted image showing a beaded appearance of the left hepatic and intrahepatic bile ducts with alternating strictures (arrows) and dilatation (arrowheads). Right: A) Fluoroscopic image from an ERCP depicting beading of the bilateral intrahepatic bile ducts due to segmental strictures (thin arrows). Guidewire in the left intrahepatic bile duct (thick arrow). B) ERCP image after biliary stent placement (arrowhead) with decompression of the biliary tree.

Small Duct Primary Sclerosing Cholangitis

Primary sclerosing cholangitis (PSC) is a well documented disease of the biliary tract associated with inflammatory bowel diseases (IBD), especially ulcerative colitis.1 Small-duct PSC, a variant of PSC, also has an association with inflammatory bowel disease.1 The exact pathogenesis of PSC is not well understood, but thought to be due to a combination of a genetic predisposition and gut structure.1 A 34 year-old male with a history significant for rheumatoid arthritis, portal vein thrombosis and ulcerative colitis presented to the hepatology clinic for evaluation of abnormal liver enzymes. He was diagnosed with ulcerative colitis at age 24 and over the years treated with Mesalamine, steroids, Azathioprine, Cyclosporine, and Infliximab. However, he was unresponsive to therapy and underwent total colectomy 2 years after diagnosis. The liver enzymes of concern had a mixed cholestatic and hepatocellular pattern with ALT 203, AST 104, Tbili 1.7, Dbili 0.6, and ALP 339. Hence, the differential was initially broad including possible autoimmune causes, vascular with the portal vein thromboses history, drug induced or primary sclerosing cholangitis (PSC) or primary biliary cirrhosis. A viral hepatitis screen was negative. Autoimmune testing revealed a positive anti-nuclear antibody titer. Magnetic resonance cholangiopancreatography (MRCP) revealed hypoattenuations correlating with his history of portal vein thromboses. Liver biopsy revealed small duct PSC. He is scheuled for imaging every 6 months to observe for cholangitis. A retrospective analysis of patients with PSC from 1980 to 2010 at 37 centers in Europe, North America, and Australia was completed by the International PSC Study Group.2 Of the 7121 patients, 65.5% were men, 89.8% had classical or large-duct disease, and 70.0% developed IBD at some point.2 Small-duct PSC was associated with a lower risk of liver transplant, death or malignancy compared with classic PSC.2 Small-duct PSC characterized a low-risk phenotype in both sexes.2 This suggests that small duct PSC is usually seen in younger individuals, in early stage disease, and suggests better outcomes.