Regulation of intestinal senescence during cholestatic liver disease modulates barrier function and liver disease progression.

Background & Aims: Senescence has been reported to have differential functions in cholangiocytes and hepatic stellate cells (HSCs) during human and murine cholestatic disease, being detrimental in biliary cells and anti-fibrotic in HSCs. Cholestatic liver disease is associated with loss of intestinal barrier function and changes in the microbiome, the mechanistic cause of which is undetermined. Methods: Intestinal samples were analysed from controls and patients with primary sclerosing cholangitis, as well as wild-type (WT) and p16-3MR transgenic mice. Cholestatic liver disease was induced by bile duct ligation (BDL) and DDC diet feeding. Fexaramine was used as an intestinal-restricted FXR agonist and antibiotics were given to eliminate the intestinal microbiome. Senescent cells were eliminated in p16-3MR mice with ganciclovir and in WT mice with the senolytic drug ABT-263. In vitro studies were done in intestinal CaCo-2 cells and organoids were generated from intestinal crypts isolated from mice. Results: Herein, we show increased senescence in intestinal epithelial cells (IECs) in patients with primary sclerosing cholangitis and in mice after BDL and DDC diet feeding. Intestinal senescence was increased in response to reduced exposure to bile acids and increased presence of lipopolysaccharide in vitro and in vivo during cholestatic liver disease. Senescence of IECs was associated with lower proliferation but increased intestinal stem cell activation, as supported by increased organoid growth from intestinal stem cells. Elimination of senescent cells with genetic and pharmacological approaches exacerbated liver injury and fibrosis during cholestatic liver disease, which was associated with increased IEC apoptosis and permeability. Conclusions: Senescence occurs in IECs during cholestatic disease and the elimination of senescent cells has a detrimental impact on the gut-liver axis. Our results point to cell-specific rather than systemic targeting of senescence as a therapeutic approach to treat cholestatic liver disease. Impact and implications: Cholestatic liver disease associates with the dysregulation of intestinal barrier function, while the mechanisms mediating the disruption of the gut-liver axis remain largely undefined. Here, we demonstrate that senescence, a cellular response to stress, is activated in intestinal cells during cholestatic liver disease in humans and mice. Mechanistically, we demonstrate that the reduction of bile acids and the increased presence of bacterial products mediate the activation of intestinal senescence during cholestatic liver disease. Importantly, the elimination of these senescent cells promotes further damage to the intestine that aggravates liver disease, with increased tissue damage and fibrosis. Our results provide evidence that therapeutic strategies to treat cholestatic liver disease by eliminating senescent cells may have unwanted effects in the intestine and support the need to develop cell/organ-specific approaches.

British Society of Gastroenterology guidelines for the diagnosis and management of cholangiocarcinoma.

These guidelines for the diagnosis and management of cholangiocarcinoma (CCA) were commissioned by the British Society of Gastroenterology liver section. The guideline writing committee included a multidisciplinary team of experts from various specialties involved in the management of CCA, as well as patient/public representatives from AMMF (the Cholangiocarcinoma Charity) and PSC Support. Quality of evidence is presented using the Appraisal of Guidelines for Research and Evaluation (AGREE II) format. The recommendations arising are to be used as guidance rather than as a strict protocol-based reference, as the management of patients with CCA is often complex and always requires individual patient-centred considerations.

Regulation of immune responses in primary biliary cholangitis: a transcriptomic analysis of peripheral immune cells.

BACKGROUND AIMS: In patients with primary biliary cholangitis (PBC), the serum liver biochemistry measured during treatment with ursodeoxycholic acid-the UDCA response-accurately predicts long-term outcome. Molecular characterization of patients stratified by UDCA response can improve biological understanding of the high-risk disease, thereby helping to identify alternative approaches to disease-modifying therapy. In this study, we sought to characterize the immunobiology of the UDCA response using transcriptional profiling of peripheral blood mononuclear cell subsets. METHODS: We performed bulk RNA-sequencing of monocytes and TH1, TH17, TREG, and B cells isolated from the peripheral blood of 15 PBC patients with adequate UDCA response ("responders"), 16 PBC patients with inadequate UDCA response ("nonresponders"), and 15 matched controls. We used the Weighted Gene Co-expression Network Analysis to identify networks of co-expressed genes ("modules") associated with response status and the most highly connected genes ("hub genes") within them. Finally, we performed a Multi-Omics Factor Analysis of the Weighted Gene Co-expression Network Analysis modules to identify the principal axes of biological variation ("latent factors") across all peripheral blood mononuclear cell subsets. RESULTS: Using the Weighted Gene Co-expression Network Analysis, we identified modules associated with response and/or disease status (q<0.05) in each peripheral blood mononuclear cell subset. Hub genes and functional annotations suggested that monocytes are proinflammatory in nonresponders, but antiinflammatory in responders; TH1 and TH17 cells are activated in all PBC cases but better regulated in responders; and TREG cells are activated-but also kept in check-in responders. Using the Multi-Omics Factor Analysis, we found that antiinflammatory activity in monocytes, regulation of TH1 cells, and activation of TREG cells are interrelated and more prominent in responders. CONCLUSIONS: We provide evidence that adaptive immune responses are better regulated in patients with PBC with adequate UDCA response.

Multitrait genome-wide analyses identify new susceptibility loci and candidate drugs to primary sclerosing cholangitis.

Primary sclerosing cholangitis (PSC) is a rare autoimmune bile duct disease that is strongly associated with immune-mediated disorders. In this study, we implemented multitrait joint analyses to genome-wide association summary statistics of PSC and numerous clinical and epidemiological traits to estimate the genetic contribution of each trait and genetic correlations between traits and to identify new lead PSC risk-associated loci. We identified seven new loci that have not been previously reported and one new independent lead variant in the previously reported locus. Functional annotation and fine-mapping nominated several potential susceptibility genes such as MANBA and IRF5. Network-based in silico drug efficacy screening provided candidate agents for further study of pharmacological effect in PSC.

The Emerging Role of Bile Acids in the Pathogenesis of Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD) is a chronic immune-mediated inflammatory disorder of the gastrointestinal tract that arises due to complex interactions between host genetic risk factors, environmental factors, and a dysbiotic gut microbiota. Although metagenomic approaches have attempted to characterise the dysbiosis occurring in IBD, the precise mechanistic pathways interlinking the gut microbiota and the intestinal mucosa are still yet to be unravelled. To deconvolute these complex interactions, a more reductionist approach involving microbial metabolites has been suggested. Bile acids have emerged as a key class of microbiota-associated metabolites that are perturbed in IBD patients. In recent years, metabolomics studies have revealed a consistent defect in bile acid metabolism with an increase in primary bile acids and a reduction in secondary bile acids in IBD patients. This review explores the evolving evidence that specific bile acid metabolites interact with intestinal epithelial and immune cells to contribute to the inflammatory milieu seen in IBD. Furthermore, we summarise evidence linking bile acids with intracellular pathways that are known to be relevant in IBD including autophagy, apoptosis, and the inflammasome pathway. Finally, we discuss how novel experimental and bioinformatics approaches could further advance our understanding of the role of bile acids and inform novel therapeutic strategies in IBD.

Intestinal Microbiome-Macrophage Crosstalk Contributes to Cholestatic Liver Disease by Promoting Intestinal Permeability in Mice.

BACKGROUND AND AIMS: Mounting evidence supports an association between cholestatic liver disease and changes in the composition of the microbiome. Still, the role of the microbiome in the pathogenesis of this condition remains largely undefined. APPROACH AND RESULTS: To address this, we have used two experimental models, administering alpha-naphtylisocyanate or feeding a 0.1% 3,5-diethoxycarbonyl-1,4-dihydrocollidine diet, to induce cholestatic liver disease in germ-free mice and germ-free mice conventionalized with the microbiome from wild-type, specific pathogen-free animals. Next, we have inhibited macrophage activation by depleting these cells using clodronate liposomes and inhibiting the inflammasome with a specific inhibitor of NOD-, LRR-, and pyrin domain-containing protein 3. Our results demonstrate that cholestasis, the accumulation of bile acids in the liver, fails to promote liver injury in the absence of the microbiome in vivo. Additional in vitro studies supported that endotoxin sensitizes hepatocytes to bile-acid-induced cell death. We also demonstrate that during cholestasis, macrophages contribute to promoting intestinal permeability and to altered microbiome composition through activation of the inflammasome, overall leading to increased endotoxin flux into the cholestatic liver. CONCLUSIONS: We demonstrate that the intestinal microbiome contributes to cholestasis-mediated cell death and inflammation through mechanisms involving activation of the inflammasome in macrophages.

British Society of Gastroenterology and UK-PSC guidelines for the diagnosis and management of primary sclerosing cholangitis.

These guidelines on the management of primary sclerosing cholangitis (PSC) were commissioned by the British Society of Gastroenterology liver section. The guideline writing committee included medical representatives from hepatology and gastroenterology groups as well as patient representatives from PSC Support. The guidelines aim to support general physicians, gastroenterologists and surgeons in managing adults with PSC or those presenting with similar cholangiopathies which may mimic PSC, such as IgG4 sclerosing cholangitis. It also acts as a reference for patients with PSC to help them understand their own management. Quality of evidence is presented using the AGREE II format. Guidance is meant to be used as a reference rather than for rigid protocol-based care as we understand that management of patients often requires individual patient-centred considerations.

Top research priorities in liver and gallbladder disorders in the UK.

OBJECTIVES: There is a mismatch between research questions considered important by patients, carers and healthcare professionals and the research performed in many fields of medicine. The non-alcohol-related liver and gallbladder disorders priority setting partnership was established to identify the top research priorities in the prevention, diagnostic and treatment of gallbladder disorders and liver disorders not covered by the James-Lind Alliance (JLA) alcohol-related liver disease priority setting partnership. DESIGN: The methods broadly followed the principles of the JLA guidebook. The one major deviation from the JLA methodology was the final step of identifying priorities: instead of prioritisation by group discussions at a consensus workshop involving stakeholders, the prioritisation was achieved by a modified Delphi consensus process. RESULTS: A total of 428 unique valid diagnostic or treatment research questions were identified. A literature review established that none of these questions were considered 'answered' that is, high-quality systematic reviews suggest that further research is not required on the topic. The Delphi panel achieved consensus (at least 80% Delphi panel members agreed) that a research question was a top research priority for six questions. Four additional research questions with highest proportion of Delphi panel members ranking the question as highly important were added to constitute the top 10 research priorities. CONCLUSIONS: A priority setting process involving patients, carers and healthcare professionals has been used to identify the top 10priority areas for research related to liver and gallbladder disorders. Basic, translational, clinical and public health research are required to address these uncertainties.

Fine-Tuning of Sirtuin 1 Expression Is Essential to Protect the Liver From Cholestatic Liver Disease.

Cholestasis comprises aetiologically heterogeneous conditions characterized by accumulation of bile acids in the liver that actively contribute to liver damage. Sirtuin 1 (SIRT1) regulates liver regeneration and bile acid metabolism by modulating farnesoid X receptor (FXR); we here investigate its role in cholestatic liver disease. We determined SIRT1 expression in livers from patients with cholestatic disease, in two experimental models of cholestasis, as well as in human and murine liver cells in response to bile acid loading. SIRT1-overexpressing (SIRToe ) and hepatocyte-specific SIRT1-KO (knockout) mice (SIRThep-/- ) were subjected to bile duct ligation (BDL) and were fed with a 0.1% DDC (3,5-diethoxycarbonyl-1,4-dihydrocollidine) diet to determine the biological relevance of SIRT1 during cholestasis. The effect of NorUDCA (24-norursodeoxycholic acid) was tested in BDL/SIRToe mice. We found that SIRT1 was highly expressed in livers from cholestatic patients, mice after BDL, and Mdr2 knockout mice (Mdr2-/- ) animals. The detrimental effects of SIRT1 during cholestasis were validated in vivo and in vitro. SIRToe mice showed exacerbated parenchymal injury whereas SIRThep-/- mice evidenced a moderate improvement after BDL and 0.1% DDC feeding. Likewise, hepatocytes isolated from SIRToe mice showed increased apoptosis in response to bile acids, whereas a significant reduction was observed in SIRThep-/- hepatocytes. Importantly, the decrease, but not complete inhibition, of SIRT1 exerted by norUDCA treatment correlated with pronounced improvement in liver parenchyma in BDL/SIRToe mice. Interestingly, both SIRT1 overexpression and hepatocyte-specific SIRT1 depletion correlated with inhibition of FXR, whereas modulation of SIRT1 by NorUDCA associated with restored FXR signaling. Conclusion: SIRT1 expression is increased during human and murine cholestasis. Fine-tuning expression of SIRT1 is essential to protect the liver from cholestatic liver damage.

Factors Associated With Outcomes of Patients With Primary Sclerosing Cholangitis and Development and Validation of a Risk Scoring System.

We sought to identify factors that are predictive of liver transplantation or death in patients with primary sclerosing cholangitis (PSC), and to develop and validate a contemporaneous risk score for use in a real-world clinical setting. Analyzing data from 1,001 patients recruited to the UK-PSC research cohort, we evaluated clinical variables for their association with 2-year and 10-year outcome through Cox-proportional hazards and C-statistic analyses. We generated risk scores for short-term and long-term outcome prediction, validating their use in two independent cohorts totaling 451 patients. Thirty-six percent of the derivation cohort were transplanted or died over a cumulative follow-up of 7,904 years. Serum alkaline phosphatase of at least 2.4 × upper limit of normal at 1 year after diagnosis was predictive of 10-year outcome (hazard ratio [HR] = 3.05; C = 0.63; median transplant-free survival 63 versus 108 months; P < 0.0001), as was the presence of extrahepatic biliary disease (HR = 1.45; P = 0.01). We developed two risk scoring systems based on age, values of bilirubin, alkaline phosphatase, albumin, platelets, presence of extrahepatic biliary disease, and variceal hemorrhage, which predicted 2-year and 10-year outcomes with good discrimination (C statistic = 0.81 and 0.80, respectively). Both UK-PSC risk scores were well-validated in our external cohort and outperformed the Mayo Clinic and aspartate aminotransferase-to-platelet ratio index (APRI) scores (C statistic = 0.75 and 0.63, respectively). Although heterozygosity for the previously validated human leukocyte antigen (HLA)-DR*03:01 risk allele predicted increased risk of adverse outcome (HR = 1.33; P = 0.001), its addition did not improve the predictive accuracy of the UK-PSC risk scores. Conclusion: Our analyses, based on a detailed clinical evaluation of a large representative cohort of participants with PSC, furthers our understanding of clinical risk markers and reports the development and validation of a real-world scoring system to identify those patients most likely to die or require liver transplantation.

A review of the medical treatment of primary sclerosing cholangitis in the 21st century.

Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease that progresses to end-stage liver disease and cirrhosis. Recurrent biliary inflammation is thought to lead to dysplasia, and as such PSC confers a high risk of cholangiocarcinoma. PSC accounts for 10% of all UK liver transplants, although transplantation does not guarantee a cure with 20% recurrence in the graft. At present there are no effective medical treatment options for PSC, and trials of novel therapeutic agents are limited by the time taken to reach clinically significant endpoints with no well defined early surrogate markers for disease outcome. Moreover, PSC appears to be a heterogeneous disease with regards to disease distribution, associated inflammatory bowel disease and subsequent disease outcome, further compounding the issue. Thus existing trials have taken place in heterogeneous groups, are likely to be underpowered to detect any individual subgroups effect. The current mainstay of medical treatment is still with ursodeoxycholic acid, although there is no evidence that it alters long-term outcome. Small pilot studies of immunosuppressive agents have taken place, but despite evidence that may support studies in larger groups, these have not been conducted. Recent advances in our understanding of the disease pathogenesis may therefore pave the way for trials of novel therapeutic agents in PSC, even given the limitations described. This review explores the controversial evidence underlying current treatment strategies and discounted treatments, and explores prospective agents that may bring new hope to the treatment of PSC in the 21st century.

Albumin reduces paracentesis-induced circulatory dysfunction and reduces death and renal impairment among patients with cirrhosis and infection: a systematic review and meta-analysis.

BACKGROUND: Studies have suggested that albumin has a value in cirrhotic patients undergoing paracentesis but its value in infection and sepsis is less clear. We planned to perform a meta-analysis of the risk of adverse outcomes in cirrhotic patients with and without albumin use. METHODS: We searched MEDLINE and EMBASE in January 2013 for randomized studies of cirrhotic patients that reported the risk of adverse events and mortality with albumin and no albumin exposure. We performed random effects meta-analysis and assessed heterogeneity using the I² statistic. RESULTS: Our review included 16 studies covering 1,518 patients. The use of albumin in paracentesis was associated with significantly reduced risk of paracentesis-induced circulatory dysfunction (OR 0.26 95%, CI 0.08-0.93) and there was a nonsignificant difference in death, encephalopathy, hyponatraemia, readmission, and renal impairment. Compared to the other volume expanders, albumin use showed no difference in clinical outcomes. In cirrhotic patients with any infection, there was a significant reduction in mortality (OR 0.46 95%, CI 0.25-0.86) and renal impairment (OR 0.34 95%, CI 0.15-0.75) when albumin was used. CONCLUSION: The use of albumin in cirrhotic patients is valuable in patients with any infection and it reduces the risk of circulatory dysfunction among patients undergoing paracentesis.

Therapeutic advances in the management of chronic hepatitis B infection.

Hepatitis B virus (HBV) is a small nonenveloped DNA virus that is a member of the Hepadnaviridae family. Chronic HBV infection is estimated to effect more than 350 million people worldwide with over 2 billion people being exposed to the virus. Risk factors for chronic infection include age of exposure to the virus, concurrent immunosuppression and HIV infection. Individuals chronically infected are 200 times more likely to develop hepatocellular carcinoma (HCC) than uninfected individuals and are at risk of developing cirrhosis and the risks of decompensated liver disease. This article focuses on the recent therapeutic advances that reduce the risk of developing these complications, those that prevent the spread of HBV and strategies for the prevention of post-liver-transplantation recurrence of HBV.

Dense genotyping of immune-related disease regions identifies nine new risk loci for primary sclerosing cholangitis.

Primary sclerosing cholangitis (PSC) is a severe liver disease of unknown etiology leading to fibrotic destruction of the bile ducts and ultimately to the need for liver transplantation. We compared 3,789 PSC cases of European ancestry to 25,079 population controls across 130,422 SNPs genotyped using the Immunochip. We identified 12 genome-wide significant associations outside the human leukocyte antigen (HLA) complex, 9 of which were new, increasing the number of known PSC risk loci to 16. Despite comorbidity with inflammatory bowel disease (IBD) in 72% of the cases, 6 of the 12 loci showed significantly stronger association with PSC than with IBD, suggesting overlapping yet distinct genetic architectures for these two diseases. We incorporated association statistics from 7 diseases clinically occurring with PSC in the analysis and found suggestive evidence for 33 additional pleiotropic PSC risk loci. Together with network analyses, these findings add to the genetic risk map of PSC and expand on the relationship between PSC and other immune-mediated diseases.

Fine mapping and replication of genetic risk loci in primary sclerosing cholangitis.

BACKGROUND AND AIMS: Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease characterized by progressive inflammation and fibrosis of the bile ducts eventually leading to biliary cirrhosis. Recent genetic studies in PSC have identified associations at 2q13, 2q35, 3p21, 4q27, 13q31 and suggestive association at 10p15. The aim of this study was to further characterize and refine the genetic architecture of PSC. METHODS: We analyzed previously reported associated SNPs at four of these non-HLA loci and 59 SNPs tagging the IL-2/IL-21 (4q27) and IL2RA (10p15) loci in 992 UK PSC cases and 5162 healthy UK controls. RESULTS: The most associated SNPs identified were rs3197999 (3p21 (MST1), p = 1.9 × 10⁻6, OR(A vs G) = 1.28, 95% CI (1.16-1.42)); rs4147359 (10p15 (IL2RA), p = 2.6 × 10⁻4, OR(A vs G) = 1.20, 95% CI (1.09-1.33)) and rs12511287 (4q27 (IL-2/IL-21), p = 3.0 × 10⁻4, OR(A vs T) = 1.21, 95% CI (1.09-1.35)). In addition, we performed a meta-analysis for selected SNPs using published summary statistics from recent studies. We observed genome-wide significance for rs3197999 (3p21 (MST1), P (combined) = 3.8 × 10⁻¹²) and rs4147359 (10p15 (IL2RA), P (combined) = 1.5 × 10⁻8). CONCLUSION: We have for the first time confirmed the association of PSC with genetic variants at 10p15 (IL2RA) locus at genome-wide significance and replicated the associations at MST1 and IL-2/IL-21 loci in a large homogeneous UK population. These results strongly implicate the role of IL-2/IL2RA pathway in PSC and provide further confirmation of MST1 association.

Endoscopic ultrasound: a review of current diagnostic and therapeutic applications.

Endoscopic ultrasound (EUS) has become important in a variety of clinical settings. Echoendoscopes may be categorised into radial and linear configurations. Radial devices are used for diagnostic imaging, whereas linear echoendoscopes also facilitate image guided tissue sampling and intervention. EUS is an established primary diagnostic tool for a number of conditions including choledocholithiasis and biliary microlithiasis. It is therefore well suited to the investigation of the aetiology of pancreatitis where simpler measures fail to identify the aetiology. It can also be used to identify chronic non-calcific pancreatitis. EUS is important in the secondary evaluation of abnormalities detected by other imaging modalities-for example, cystic pancreatic lesions. The high resolution of EUS allows more detailed image based analysis than other imaging modalities. The ability to sample cyst fluid significantly increases the accuracy of lesion characterisation. Most importantly, EUS has become indispensable in the staging of a variety of upper gastrointestinal tract tumours. If resection is being considered, the high resolution images obtained via EUS are invaluable for local tumour staging. EUS guided tissue sampling permits accurate nodal staging without relying on lymph node size as proxy for malignant infiltration. In patients with contraindications to magnetic resonance imaging, EUS is an alternative for the staging of rectal carcinoma. It is used in the staging of lung cancer, often in combination with endobronchial ultrasound. Finally, EUS is used therapeutically in image guided drainage (such as gastrocystostomy in pancreatic pseudocyst) and coeliac plexus neurolysis in patients with abdominal pain caused by pancreatic cancer or pancreatitis.