Inhibition of glutamine synthetase in monocytes from patients with acute-on-chronic liver failure resuscitates their antibacterial and inflammatory capacity.

OBJECTIVE: Acute-on-chronic liver failure (ACLF) is associated with dysfunctional circulating monocytes whereby patients become highly susceptible to bacterial infections. Here, we identify the pathways underlying monocyte dysfunction in ACLF and we investigate whether metabolic rewiring reinstates their phagocytic and inflammatory capacity. DESIGN: Following phenotypic characterisation, we performed RNA sequencing on CD14+CD16- monocytes from patients with ACLF and decompensated alcoholic cirrhosis. Additionally, an in vitro model mimicking ACLF patient-derived features was implemented to investigate the efficacy of metabolic regulators on monocyte function. RESULTS: Monocytes from patients with ACLF featured elevated frequencies of interleukin (IL)-10-producing cells, reduced human leucocyte antigen DR isotype (HLA-DR) expression and impaired phagocytic and oxidative burst capacity. Transcriptional profiling of isolated CD14+CD16- monocytes in ACLF revealed upregulation of an array of immunosuppressive parameters and compromised antibacterial and antigen presentation machinery. In contrast, monocytes in decompensated cirrhosis showed intact capacity to respond to inflammatory triggers. Culturing healthy monocytes in ACLF plasma mimicked the immunosuppressive characteristics observed in patients, inducing a blunted phagocytic response and metabolic program associated with a tolerant state. Metabolic rewiring of the cells using a pharmacological inhibitor of glutamine synthetase, partially restored the phagocytic and inflammatory capacity of in vitro generated- as well as ACLF patient-derived monocytes. Highlighting its biological relevance, the glutamine synthetase/glutaminase ratio of ACLF patient-derived monocytes positively correlated with disease severity scores. CONCLUSION: In ACLF, monocytes feature a distinct transcriptional profile, polarised towards an immunotolerant state and altered metabolism. We demonstrated that metabolic rewiring of ACLF monocytes partially revives their function, opening up new options for therapeutic targeting in these patients.

The economic impact of using single-operator cholangioscopy for the treatment of difficult bile duct stones and diagnosis of indeterminate bile duct strictures.

BACKGROUND AND STUDY AIM: Conventional endoscopic retrograde cholangiopancreatography (ERCP) combines endoscopy and radiography to diagnose and treat pathological conditions of the bile duct. The aim of the present analysis was to evaluate the clinical and economic impact of the use of single-operator intraductal cholangioscopy (IDC), which allows for direct visualization of the bile duct, as an alternative to ERCP for the treatment of difficult bile duct stones and the diagnosis of bile duct strictures. PATIENTS AND METHODS: The clinical and economic consequences of single-operator IDC use were evaluated using two decision-tree models, one for management of difficult-to-remove stones and one for stricture diagnosis. A hospital perspective was adopted. Data to populate the models were derived from two Belgian hospitals that specialize in endoscopic procedures of the bile duct. Overall, the examined population consisted of 62 patients with difficult stones and 49 patients with indeterminate strictures. RESULTS: In the model for difficult stone management, the use of IDC determined a decrease in the number of procedures (- 27 % relative reduction) and costs (- €73 000; - 11 % relative reduction) when compared with ERCP. In the model for stricture diagnosis, the use of IDC determined a decrease in the number of procedures (- 31 % relative reduction) and costs (- €13 000; - 5 % relative variation) when compared with ERCP. CONCLUSIONS: The single-operator IDC system performed better than ERCP for the treatment of difficult bile duct stones and the diagnosis of bile duct strictures, and reduced the overall expenditure in hospitals in Belgium.

Janus-kinase-2 relates directly to portal hypertension and to complications in rodent and human cirrhosis.

OBJECTIVE: Angiotensin II (AngII) activates via angiotensin-II-type-I receptor (AT1R) Janus-kinase-2 (JAK2)/Arhgef1 pathway and subsequently RHOA/Rho-kinase (ROCK), which induces experimental and probably human liver fibrosis. This study investigated the relationship of JAK2 to experimental and human portal hypertension. DESIGN: The mRNA and protein levels of JAK2/ARHGEF1 signalling components were analysed in 49 human liver samples and correlated with clinical parameters of portal hypertension in these patients. Correspondingly, liver fibrosis (bile duct ligation (BDL), carbon tetrachloride (CCl4)) was induced in floxed-Jak2 knock-out mice with SM22-promotor (SM22Cre+-Jak2f/f). Transcription and contraction of primary myofibroblasts from healthy and fibrotic mice and rats were analysed. In two different cirrhosis models (BDL, CCl4) in rats, the acute haemodynamic effect of the JAK2 inhibitor AG490 was assessed using microsphere technique and isolated liver perfusion experiments. RESULTS: Hepatic transcription of JAK2/ARHGEF1 pathway components was upregulated in liver cirrhosis dependent on aetiology, severity and complications of human liver cirrhosis (Model for End-stage Liver disease (MELD) score, Child score as well as ascites, high-risk varices, spontaneous bacterial peritonitis). SM22Cre+- Jak2f/f mice lacking Jak2 developed less fibrosis and lower portal pressure (PP) than SM22Cre--Jak2f/f upon fibrosis induction. Myofibroblasts from SM22Cre+-Jak2f/f mice expressed less collagen and profibrotic markers upon activation. AG490 relaxed activated hepatic stellate cells in vitro. In cirrhotic rats, AG490 decreased hepatic vascular resistance and consequently the PP in vivo and in situ. CONCLUSIONS: Hepatic JAK2/ARHGEF1/ROCK expression is associated with portal hypertension and decompensation in human cirrhosis. The deletion of Jak2 in myofibroblasts attenuated experimental fibrosis and acute inhibition of JAK2 decreased PP. Thus, JAK2 inhibitors, already in clinical use for other indications, might be a new approach to treat cirrhosis with portal hypertension.

A new intraductal radiofrequency ablation device for inoperable biliopancreatic tumors complicated by obstructive jaundice: the IGNITE-1 study.

Background and study aims Biliary stenting of unresectable malignant bile duct obstruction is generally accepted as the standard of care but it can be hampered by tumor ingrowth and stent dysfunction. We aimed to test the feasibility, safety, and biliary patency rate of a new endoscopically applied intraductal radiofrequency ablation (RFA) device. Patients and methods Eighteen patients with inoperable malignant biliary obstruction underwent endoscopic retrograde cholangiopancreatography (ERCP)-directed RFA and stenting. Results Between December 2014 and November 2015, 18 patients underwent RFA to the intended region, with no complications within 3 months of the procedure. Bilirubin levels post-RFA and stenting decreased significantly (7.8 ±â€Š1 mg/dL to 1.7 ±â€Š0.4 mg/dL; P < 0.001). At 90 and 180 days post-intervention, biliary patency was maintained in 80 % and 69 % of patients still alive at that time, respectively. The median overall stent patency was 110 days (range 16 - 374), with a median patient survival of 227 days (range 16 - 374). Conclusion Intraductal RFA using a new device in patients with inoperable biliopancreatic cancer complicated by jaundice appeared feasible and safe with acceptable biliary patency. Randomized trials with prolonged follow-up are warranted.ClinTrials.gov: NCT02468076.

The FXR agonist obeticholic acid prevents gut barrier dysfunction and bacterial translocation in cholestatic rats.

Bacterial translocation (BTL) drives pathogenesis and complications of cirrhosis. Farnesoid X-activated receptor (FXR) is a key transcription regulator in hepatic and intestinal bile metabolism. We studied potential intestinal FXR dysfunction in a rat model of cholestatic liver injury and evaluated effects of obeticholic acid (INT-747), an FXR agonist, on gut permeability, inflammation, and BTL. Rats were gavaged with INT-747 or vehicle during 10 days after bile-duct ligation and then were assessed for changes in gut permeability, BTL, and tight-junction protein expression, immune cell recruitment, and cytokine expression in ileum, mesenteric lymph nodes, and spleen. Auxiliary in vitro BTL-mimicking experiments were performed with Transwell supports. Vehicle-treated bile duct-ligated rats exhibited decreased FXR pathway expression in both jejunum and ileum, in association with increased gut permeability through increased claudin-2 expression and related to local and systemic recruitment of natural killer cells resulting in increased interferon-γ expression and BTL. After INT-747 treatment, natural killer cells and interferon-γ expression markedly decreased, in association with normalized permeability selectively in ileum (up-regulated claudin-1 and occludin) and a significant reduction in BTL. In vitro, interferon-γ induced increased Escherichia coli translocation, which remained unaffected by INT-747. In experimental cholestasis, FXR agonism improved ileal barrier function by attenuating intestinal inflammation, leading to reduced BTL and thus demonstrating a crucial protective role for FXR in the gut-liver axis.

The expanding role of the bile acid receptor farnesoid X in the intestine and its potential clinical implications.

Knowledge about the role of farnesoid X receptor (FXR) in the intestine is rapidly expanding. In pre-clinical animal models of inflammatory bowel disease and bile duct ligation, FXR activation has proven to directly target the three pillars of intestinal homeostasis: intestinal permeability, inflammation and bacterial translocation. The protective role of FXR-ligands on this homeostasis has implications for many intestinal pathologies like inflammatory bowel disease, ischemia reperfusion injury, the metabolic syndrome, colon cancer and even diarrhea. In this review, we summarize the mechanisms by which FXR-activation exerts these protective effects and we discuss its potential clinical applications.

Obeticholic acid, a farnesoid X receptor agonist, improves portal hypertension by two distinct pathways in cirrhotic rats.

UNLABELLED: The farnesoid X receptor (FXR) is a nuclear bile acid receptor involved in bile acid homeostasis, hepatic and intestinal inflammation, liver fibrosis, and cardiovascular disease. We studied the effect of short-term treatment with obeticholic acid (INT-747), a potent selective FXR agonist, on intrahepatic hemodynamic dysfunction and signaling pathways in different rat models of cirrhotic portal hypertension (PHT). For this, thioacetamide (TAA)-intoxicated and bile-duct-ligated (BDL) rats were used as models. After gavage of two doses of 30 mg/kg of INT-747 or vehicle within 24 hours, in vivo hemodynamics were assessed. Additionally, we evaluated the direct effect of INT-747 on total intrahepatic vascular resistance (IHVR) and intrahepatic vascular tone (endothelial dysfunction and hyperresponsiveness to methoxamine) by means of an in situ liver perfusion system and on hepatic stellate cell contraction in vitro. FXR expression and involved intrahepatic vasoactive pathways (e.g., endothelial nitric oxide synthase [eNOS], Rho-kinase, and dimethylarginine dimethylaminohydrolase [DDAH]) were analyzed by immunohistochemistry, reverse-transcriptase polymerase chain reaction, or western blotting. In both cirrhotic models, FXR expression was decreased. Treatment with INT-747 in TAA and BDL reactivated the FXR downstream signaling pathway and decreased portal pressure by lowering total IHVR without deleterious systemic hypotension. In the perfused TAA and BDL cirrhotic liver, INT-747 improved endothelial vasorelaxation capacity, but not hyperresponsiveness. In both groups, this was associated with an increased eNOS activity, which, in TAA, related to down-regulation of Rho-kinase and in BDL to up-regulation of DDAH-2. CONCLUSION: FXR agonist INT-747 improves PHT in two different rat models of cirrhosis by decreasing IHVR. This hemodynamic effect relates to increased intrahepatic eNOS activity by pathways that differ depending on the etiology of cirrhosis.

A first case of Chimaeribacter species catheter-related bloodstream infection.

We present a case of a 80-year-old patient with a catheter-related bloodstream infection caused by Chimaeribacter species. The Chimaeribacter genus represents a novel genus within the Yersiniaceae family. To the best of our knowledge, as of today, nothing is known about the pathogenicity of Chimaeribacter species, nor about the appropriate antimicrobial management. In the present case, we demonstrate, for the first time, a potential clinical relevance of the Chimaeribacter species. Antimicrobial susceptibility data are presented.

Increased intrahepatic resistance in severe steatosis: endothelial dysfunction, vasoconstrictor overproduction and altered microvascular architecture.

Non-alcoholic fatty liver disease can progress to steatohepatitis and fibrosis, and is also associated with impaired liver regeneration. The pathophysiology remains elusive. We recently showed that severe steatosis is associated with an increase in portal pressure, suggesting liver flow impairment. The objective of this study is to directly assess total intrahepatic resistance and its potential functional and structural determinants in an in situ perfusion model. Male Wistar rats fed a control (n = 30) or a methionine-choline-deficient (MCD) diet (n = 30) for 4 weeks were compared. Liver tissue and serum analysis, in vivo haemodynamic measurements, in situ perfusion experiments and vascular corrosion casts were performed. The MCD group showed severe steatosis without inflammation or fibrosis on histology. Serum levels and liver tissue gene expression of interleukin (IL)-6, tumour necrosis factor-α, IL-1ß and interferon-γ, liver tissue myeloperoxidase activity and liver immunohistochemistry with anti-CD68 and anti-α smooth muscle actin were comparable between groups, excluding significant inflammation. Flow-pressure curves were significantly different between groups for all flows (slope values: 0.1636 ± 0.0605 mm Hg/ml/min in controls vs 0.7270 ± 0.0408 mm Hg/ml/min in MCD-fed rats, P < 0.001), indicating an increased intrahepatic resistance, which was haemodynamically significant (portocaval pressure gradient 2.2 ± 1.1 vs 8.2 ± 1.3 mm Hg in controls vs MCD, P<0.001). Dose-response curves to acetylcholine were significantly reduced in MCD-fed rats (P < 0.001) as was the responsiveness to methoxamine (P<0.001). Vascular corrosion casts showed a replacement of the regular sinusoidal anatomy by a disorganized pattern with multiple interconnections and vascular extensions. Liver phosphorylated endothelial NO synthase (eNOS)/eNOS and serum nitrite/nitrate were not increased in severe steatosis, whereas liver thromboxane synthase expression, liver endothelin-1 (ET-1) expression and serum andothelin-1 concentration were significantly increased. Severe steatosis induces a haemodynamically significant increase in intrahepatic resistance, which precedes inflammation and fibrogenesis. Both functional (endothelial dysfunction and increased thromboxane and ET-1 synthesis) and structural factors are involved. This phenomenon might significantly contribute to steatosis-related disease.

Bench-to-beside review: acute-on-chronic liver failure - linking the gut, liver and systemic circulation.

The concept of acute-on-chronic liver failure (ACLF) was introduced recently to describe a subset of patients with chronic liver disease presenting with profound deterioration of liver function and rapidly evolving multi-organ failure. ACLF is frequently accompanied by the development of severe inflammatory response syndrome and has a high mortality. To date, treatment options are limited and exclusively supportive. Over the last few years, some insights have been generated in the pathophysiology of ACLF. A key role for the interaction of innate immune dysfunction, enhanced bacterial translocation from the gut, and circulatory dysfunction has been proposed. In this respect, therapeutic strategies have been examined, with variable success, in experimental studies in animals and humans. This review focuses on potentially relevant pathophysiological elements in the development of ACLF and points out promising treatment modalities in ACLF.

Age Matching of Elderly Liver Grafts With Elderly Recipients Does Not Have a Synergistic Effect on Long-term Outcomes When Both Are Carefully Selected.

BACKGROUND: Older donors and recipients are increasingly considered for liver transplantation. Both donor and recipient age have a negative impact on outcomes. Large registry analyses show that older donors are frequently matched to older recipients. Whether age-related risks accumulate in a synergic negative effect on outcomes because of donor-recipient age matching is poorly understood. METHODS: We investigated the impact of donor-recipient age interaction on patient and death-censored graft survival in multivariate Cox regressions in 849 transplants (January 2000 to December 2015). RESULTS: Donors 70 years or older did not affect long-term patient or graft survival. Recipient age independently increased the risk of death (hazard ratio [HR], 1.03; 95% confidence interval [CI], 1.02-1.05, P < 0.0001), but donor-recipient age interaction was noninfluential. The negative impact of recipient age on patient survival was significant as early as 6 months after transplantation (HR, 1.06; 95% CI, 1.03-1.09; P = 0.00008). The adjusted risk of death was significant for patients aged 60 to 69 years (HR, 1.995; 95% CI, 1.40-2.85; P < 0.0001) and 70 years or older (HR, 2.001; 95% CI, 1.10-2.66; P = 0.04). In contrast, the risk of graft loss was not influenced by recipient age (HR, 1.02; 95% CI, 0.996-1.04; P = 0.11) or age interaction. CONCLUSIONS: Older livers can be safely used in older recipients without jeopardizing graft and patient survival if other risk factors are minimized.

Thrombosis of a portal vein aneurysm: a case report with literature review.

OBJECTIVES: Portal vein aneurysm is an unusual vascular dilatation of the portal vein. The etiology, diagnosis and management are ill-defined. METHODS: A case of a portal vein aneurysm complicated with complete thrombosis is presented with a literature review providing an overview of the etiology, clinical presentation and management. RESULTS: Portal venous aneurysms represent approximately 3% of all venous aneurysms with a reported prevalence of 0.06%. The reported incidence is on the rise with increasing use of modern imaging techniques in clinical practice. Usually, portal vein aneurysms are incidental findings and patients are asymptomatic. They can be congenital or acquired and portal hypertension represents the most frequent cause of the acquired version. Various complications such as biliary tract compression, portal vein thrombosis, and rupture can occur. Treatment options are conservative management or surgery. Surgical treatment is currently reserved for symptomatic patients with severe abdominal pain, symptoms of pressure effect or with expanding aneurysms, and/or complications such as thrombosis or rupture. CONCLUSION: Conservative management seems the best option in the majority of patients. A multidisciplinary approach discussing the best option on a case-by-case base in light of their individual underlying risk and symptoms is advised.

Therapeutic intranodal lymphangiography for refractory chylous ascites complicating acute necrotic pancreatitis.

Chylous ascites is a rare complication of acute pancreatitis. However, the incidence of intraperitoneal chyle leakage related to severe pancreatitis may be much higher. This is probably the result of direct damage to the cisterna chyli or its tributaries by pancreatic enzymes. In this case, conservative treatment failed to resolve the chyle leak. For the first time, to our knowledge, ultrasound guided therapeutic intranodal lymphangiography was shown to be a successful, minimally invasive treatment option in chylous ascites complicating acute necrotic pancreatitis.

Farnesoid X Receptor Activation Attenuates Intestinal Ischemia Reperfusion Injury in Rats.

INTRODUCTION: The farnesoid X receptor (FXR) is abundantly expressed in the ileum, where it exerts an enteroprotective role as a key regulator of intestinal innate immunity and homeostasis, as shown in pre-clinical models of inflammatory bowel disease. Since intestinal ischemia reperfusion injury (IRI) is characterized by hyperpermeability, bacterial translocation and inflammation, we aimed to investigate, for the first time, if the FXR-agonist obeticholic acid (OCA) could attenuate intestinal ischemia reperfusion injury. MATERIAL AND METHODS: In a validated rat model of intestinal IRI (laparotomy + temporary mesenteric artery clamping), 3 conditions were tested (n = 16/group): laparotomy only (sham group); ischemia 60min+ reperfusion 60min + vehicle pretreatment (IR group); ischemia 60min + reperfusion 60min + OCA pretreatment (IR+OCA group). Vehicle or OCA (INT-747, 2*30mg/kg) was administered by gavage 24h and 4h prior to IRI. The following end-points were analyzed: 7-day survival; biomarkers of enterocyte viability (L-lactate, I-FABP); histology (morphologic injury to villi/crypts and villus length); intestinal permeability (Ussing chamber); endotoxin translocation (Lipopolysaccharide assay); cytokines (IL-6, IL-1-ß, TNFα, IFN-γ IL-10, IL-13); apoptosis (cleaved caspase-3); and autophagy (LC3, p62). RESULTS: It was found that intestinal IRI was associated with high mortality (90%); loss of intestinal integrity (structurally and functionally); increased endotoxin translocation and pro-inflammatory cytokine production; and inhibition of autophagy. Conversely, OCA-pretreatment improved 7-day survival up to 50% which was associated with prevention of epithelial injury, preserved intestinal architecture and permeability. Additionally, FXR-agonism led to decreased pro-inflammatory cytokine release and alleviated autophagy inhibition. CONCLUSION: Pretreatment with OCA, an FXR-agonist, improves survival in a rodent model of intestinal IRI, preserves the gut barrier function and suppresses inflammation. These results turn FXR into a promising target for various conditions associated with intestinal ischemia.

FXR agonist obeticholic acid reduces hepatic inflammation and fibrosis in a rat model of toxic cirrhosis.

Hepatic inflammation drives hepatic stellate cells (HSC), resulting in liver fibrosis. The Farnesoid-X receptor (FXR) antagonizes inflammation through NF-κB inhibition. We investigated preventive and therapeutic effects of FXR agonist obeticholic acid (OCA) on hepatic inflammation and fibrosis in toxic cirrhotic rats. Cirrhosis was induced by thioacetamide (TAA) intoxication. OCA was given during or after intoxication with vehicle-treated rats as controls. At sacrifice, fibrosis, hemodynamic and biochemical parameters were assessed. HSC activation, cell turn-over, hepatic NF-κB activation, pro-inflammatory and pro-fibrotic cytokines were determined. The effect of OCA was further evaluated in isolated HSC, Kupffer cells, hepatocytes and liver sinusoidal endothelial cells (LSEC). OCA decreased hepatic inflammation and fibrogenesis during TAA-administration and reversed fibrosis in established cirrhosis. Portal pressure decreased through reduced intrahepatic vascular resistance. This was paralleled by decreased expression of pro-fibrotic cytokines (transforming growth-factor ß, connective tissue growth factor, platelet-derived growth factor ß-receptor) as well as markers of hepatic cell turn-over, by blunting effects of pro-inflammatory cytokines (e.g. monocyte chemo-attractant protein-1). In vitro, OCA inhibited both LSEC and Kupffer cell activation; while HSC remained unaffected. This related to NF-κB inhibition via up-regulated IκBα. In conclusion, OCA inhibits hepatic inflammation in toxic cirrhotic rats resulting in decreased HSC activation and fibrosis.

Copeptin as an Indicator of Hemodynamic Derangement and Prognosis in Liver Cirrhosis.

BACKGROUND: Advanced liver cirrhosis is associated with systemic hemodynamic derangement leading to the development of severe complications associated with increased mortality. Copeptin is a stable cleavage product of the precursor of arginine vasopressin, a key-regulator in hemodynamic homeostasis. Copeptin is currently considered a reliable prognostic marker in a wide variety of diseases other than cirrhosis. The present study aimed to assess copeptin, both experimentally and clinically, as a potential biomarker of hemodynamic derangement and to evaluate its prognostic significance in cirrhosis. MATERIALS AND METHODS: Two studies were executed: 1) in 18 thioacetamide-induced cirrhotic rats and 5 control rats, plasma copeptin and hemodynamic measurements were performed, 2) in 61 cirrhotic patients, serum copeptin concentration was measured in samples collected at time of registration at the waiting list for liver transplantation. In 46 patients, also a second copeptin measurement was performed during follow-up while registered at the waiting list for liver transplantation. To determine the association of serum copeptin and clinical data with outcome, Cox proportional hazard regression analysis and Kaplan Meier analysis were performed. RESULTS: Plasma copeptin concentration was significantly higher in cirrhotic rats than in controls (1.6 ± 0.5 vs. 0.9 ± 0.1 pmol/L, p< 0.01) and was negatively correlated to the mean arterial blood pressure (r = -0.574, p = 0.013). In cirrhotic patients, serum copeptin concentration was high [11.0 (5.2-24.0) pmol/L] and increased significantly during the time of registration at the waiting list for liver transplantation. MELD and MELD-sodium score were significantly correlated to serum copeptin [MELD: (r = 0.33, p = 0.01), MELD-sodium: (r = 0.29, p = 0.02)], also at time of the second copeptin measurement [MELD and MELD-sodium: r = 0.39, p< 0.01]. In cirrhotic humans, serum copeptin concentration was significantly associated with outcome, independently of the MELD and MELD-sodium score. Patients with a low serum copeptin concentration at time of registration at the liver transplant waiting list had significantly better transplant-free survival rates at 3, 6 and 12 months of follow-up as compared to those with a high serum copeptin concentration (Log-rank: p< 0.01, p< 0.01 and p = 0.02 respectively). CONCLUSIONS: Circulating copeptin levels are elevated in rats and humans with cirrhosis. Copeptin is independently associated with outcome in cirrhotic patients awaiting liver transplantation.