Liver and pancreatic-targeted interleukin-22 as a therapeutic for metabolic dysfunction-associated steatohepatitis.

Metabolic dysfunction-associated steatohepatitis (MASH) is the most prevalent cause of liver disease worldwide, with a single approved therapeutic. Previous research has shown that interleukin-22 (IL-22) can suppress ß-cell stress, reduce local islet inflammation, restore appropriate insulin production, reverse hyperglycemia, and ameliorate insulin resistance in preclinical models of diabetes. In clinical trials long-acting forms of IL-22 have led to increased proliferation in the skin and intestine, where the IL-22RA1 receptor is highly expressed. To maximise beneficial effects whilst reducing the risk of epithelial proliferation and cancer, we designed short-acting IL-22-bispecific biologic drugs that successfully targeted the liver and pancreas. Here we show 10-fold lower doses of these bispecific biologics exceed the beneficial effects of native IL-22 in multiple preclinical models of MASH, without off-target effects. Treatment restores glycemic control, markedly reduces hepatic steatosis, inflammation, and fibrogenesis. These short-acting IL-22-bispecific targeted biologics are a promising new therapeutic approach for MASH.

Single-nucleus RNA sequencing of pre-malignant liver reveals disease-associated hepatocyte state with HCC prognostic potential.

Current approaches to staging chronic liver diseases have limited utility for predicting liver cancer risk. Here, we employed single-nucleus RNA sequencing (snRNA-seq) to characterize the cellular microenvironment of healthy and pre-malignant livers using two distinct mouse models. Downstream analyses unraveled a previously uncharacterized disease-associated hepatocyte (daHep) transcriptional state. These cells were absent in healthy livers but increasingly prevalent as chronic liver disease progressed. Copy number variation (CNV) analysis of microdissected tissue demonstrated that daHep-enriched regions are riddled with structural variants, suggesting these cells represent a pre-malignant intermediary. Integrated analysis of three recent human snRNA-seq datasets confirmed the presence of a similar phenotype in human chronic liver disease and further supported its enhanced mutational burden. Importantly, we show that high daHep levels precede carcinogenesis and predict a higher risk of hepatocellular carcinoma development. These findings may change the way chronic liver disease patients are staged, surveilled, and risk stratified.

The ancestral haplotype markers HLA -A3 and B7 do not influence the likelihood of advanced hepatic fibrosis or cirrhosis in HFE hemochromatosis.

Advanced hepatic fibrosis occurs in up to 25% of individuals with C282Y homozygous hemochromatosis. Our aim was to determine whether human leukocyte antigen (HLA)-A3 and B7 alleles act as genetic modifiers of the likelihood of advanced hepatic fibrosis. Between 1972 and 2013, 133 HFE C282Y homozygous individuals underwent clinical and biochemical evaluation, HLA typing, liver biopsy for fibrosis staging and phlebotomy treatment. Hepatic fibrosis was graded according to Scheuer as F0-2 (low grade hepatic fibrosis), F3-4 (advanced hepatic fibrosis), and F4 cirrhosis. We analysed associations between the severity of fibrosis and HLA-A3 homozygosity, heterozygosity or absence, with or without the presence of HLA-B7 using categorical analysis. The mean age of HLA-A3 homozygotes (n = 24), heterozygotes (n = 65) and HLA-A3 null individuals (n = 44) was 40 years. There were no significant differences between the groups for mean(± SEM) serum ferritin levels (1320 ± 296, 1217 ± 124, 1348 ± 188 [Formula: see text]g/L), hepatic iron concentration (178 ± 26, 213 ± 22, 199 ± 29 [Formula: see text]mol/g), mobilizable iron stores (9.9 ± 1.5, 9.5 ± 1.5, 11.5 ± 1.7 g iron removed via phlebotomy), frequency of advanced hepatic fibrosis (5/24[12%], 13/63[19%], 10/42[19%]) or cirrhosis (3/24[21%], 12/63[21%], 4/42[24%]), respectively. The presence or absence of HLA-B7 did not influence the outcome. Thus, HLA-A3 and HLA-B7 alleles are not associated with the risk of advanced hepatic fibrosis or cirrhosis in C282Y hemochromatosis.

Risk factors for more rapid progression of severe liver fibrosis in children with cystic fibrosis-related liver disease: A multi-center study validated by liver biopsy.

BACKGROUND AND AIMS: Early identification of risk factors for the development of severe fibrosis in children with cystic fibrosis-related liver disease (CFLD) is crucial as promising therapies emerge. METHODS: This multi-center cohort study of children with a priori defined CFLD from 1999 to 2016, was designed to evaluate the clinical utility of CF-specific characteristics and liver biomarkers assessed years prior to liver biopsy-proven CFLD to predict risk of developing severe fibrosis (F3-4) over time. Fibrosis was staged by Metavir classification. RESULTS: The overall study cohort of 42 patients (F0-2 (n = 22) and F3-4 (n = 20)) was 57% male (n = 24) with median age of 7.6 years at baseline visit versus 10.3 years at biopsy. Median FEV1 % predicted was lower in F3-4 participants at baseline versus F0-2 (59% vs. 85%; p = .002), while baseline FIB-4, APRI and GGT were higher in F3-4. Median splits for FIB-4 (≥.13), APRI (≥.36), GPR (≥.09), GGT (≥25.5), and FEV1 % (<64%) were associated with more rapid progression to F3-4 (p < .01 for all). Using a combination of change/year in FIB-4, APRI, and GPR to predict F3-4, the AUROC was .81 (95% CI, .66, .96; p < .0001). For up to 5.8 years prior, thresholds for GPR were met 6.5-fold more rapidly, and those for APRI and FIB-4 were met 2.5-fold more rapidly, in those who progressed to F3-4 than those that did not. CONCLUSIONS: This study suggests mild-moderate pulmonary dysfunction and higher liver biomarker indices at baseline may be associated with faster progression of CFLD.

TWEAK/Fn14 Signalling Regulates the Tissue Microenvironment in Chronic Pancreatitis.

Chronic pancreatitis increases the risk of developing pancreatic cancer through the upregulation of pathways favouring proliferation, fibrosis, and sustained inflammation. We established in previous studies that the ligand tumour necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) signals through its cognate receptor fibroblast growth factor-inducible 14 (Fn14) to regulate these underlying cellular processes in the chronic liver injury niche. However, the role of the TWEAK/Fn14 signalling pathway in pancreatic disease is entirely unknown. An analysis of publicly available datasets demonstrated that the TWEAK receptor Fn14 is upregulated in pancreatitis and pancreatic adenocarcinoma, with single cell RNA sequencing revealing pancreatic ductal cells as the main Fn14 producers. We then used choline-deficient, ethionine-supplemented (CDE) diet feeding of wildtype C57BL/6J and Fn14 knockout littermates to (a) confirm CDE treatment as a suitable model of chronic pancreatitis and (b) to investigate the role of the TWEAK/Fn14 signalling pathway in pancreatic ductal proliferation, as well as fibrotic and inflammatory cell dynamics. Our time course data obtained at three days, three months, and six months of CDE treatment reveal that a lack of TWEAK/Fn14 signalling significantly inhibits the establishment and progression of the tissue microenvironment in CDE-induced chronic pancreatitis, thus proposing the TWEAK/Fn14 pathway as a novel therapeutic target.

Characterization of CFTR mutations in people with cystic fibrosis and severe liver disease who are not eligible for CFTR modulators.

Cystic-fibrosis-related liver disease (CFLD) is a variable phenotype of CF. The severe CFLD variant with cirrhosis or portal hypertension has a poor prognosis and life expectancy. CFTR modulator therapies are now available for people with CF and eligibility for such treatment is based on their CFTR genotype. We evaluated the genetic eligibility for elexacaftor, tezacaftor, ivacaftor (ETI), and ivacaftor (IVA) monotherapy in a previously reported CF cohort of 1591 people with CF of whom 171 with severe CFLD. Based on their CFTR mutations, 13% (N=184/1420) of subjects without CFLD and 11% (N=19/171) of those with severe CFLD are not eligible for either ETI or IVA therapy. The non-eligible patients without CFLD or with severe CFLD can currently not take advantage of the potential benefits of these new treatments. Although this study cannot provide any data regarding the effect of ETI or IVA on the progression of severe CFLD, the consequences for ineligibility of patients with extreme liver phenotype may be even more significant because of their poorer disease risk profile.

The structural conformation of the tachykinin domain drives the anti-tumoural activity of an octopus peptide in melanoma BRAFV600E.

BACKGROUND AND PURPOSE: Over past decades, targeted therapies and immunotherapy have improved survival and reduced the morbidity of patients with BRAF-mutated melanoma. However, drug resistance and relapse hinder overall success. Therefore, there is an urgent need for novel compounds with therapeutic efficacy against BRAF-melanoma. This prompted us to investigate the antiproliferative profile of a tachykinin-peptide from the Octopus kaurna, Octpep-1 in melanoma. EXPERIMENTAL APPROACH: We evaluated the cytotoxicity of Octpep-1 by MTT assay. Mechanistic insights on viability and cellular damage caused by Octpep-1 were gained via flow cytometry and bioenergetics. Structural and pharmacological characterization was conducted by molecular modelling, molecular biology, CRISPR/Cas9 technology, high-throughput mRNA and calcium flux analysis. In vivo efficacy was validated in two independent xerograph animal models (mice and zebrafish). KEY RESULTS: Octpep-1 selectively reduced the proliferative capacity of human melanoma BRAFV600E -mutated cells with minimal effects on fibroblasts. In melanoma-treated cells, Octpep-1 increased ROS with unaltered mitochondrial membrane potential and promoted non-mitochondrial and mitochondrial respiration with inefficient ATP coupling. Molecular modelling revealed that the cytotoxicity of Octpep-1 depends upon the α-helix and polyproline conformation in the C-terminal region of the peptide. A truncated form of the C-terminal end of Octpep-1 displayed enhanced potency and efficacy against melanoma. Octpep-1 reduced the progression of tumours in xenograft melanoma mice and zebrafish. CONCLUSION AND IMPLICATIONS: We unravel the intrinsic anti-tumoural properties of a tachykinin peptide. This peptide mediates the selective cytotoxicity in BRAF-mutated melanoma in vitro and prevents tumour progression in vivo, providing a foundation for a therapy against melanoma.

Arthritis Prediction of Advanced Hepatic Fibrosis in HFE Hemochromatosis.

OBJECTIVE: To evaluate whether arthritis predicts the likelihood of advanced hepatic fibrosis in HFE hemochromatosis. PATIENTS AND METHODS: We conducted a retrospective, cross-sectional analysis of 112 well-characterized patients with HFE hemochromatosis and liver biopsy-validated fibrosis staging recruited between January 1, 1983, and December 31, 2013. Complete clinical, biochemical, hematologic, and noninvasive serum biochemical indices (aspartate aminotransferase to platelet ratio index [APRI] and fibrosis 4 index [FIB4]) were available. Scheuer fibrosis stages 3 and 4, APRI greater than 0.44, or FIB4 greater than 1.1 were used to define advanced hepatic fibrosis. Comparisons between groups were performed using categorical analysis, unpaired or paired t test. RESULTS: Male (n=76) and female (n=36) patients were similar in age. Nineteen patients had advanced hepatic fibrosis, and 47 had hemochromatosis arthritis. Arthritis was significantly associated with the presence of advanced hepatic fibrosis as determined by liver biopsy (sensitivity, 84%, [95% CI, 62% to 95%]; negative predictive value, 95% [95% CI, 87% to 99%]; relative risk, 7.4 [95% CI, 2.5 to 23]; P<.001), APRI (sensitivity, 75% [95% CI, 55% to 88%]; negative predictive value, 91% [95% CI, 81% to 96%]; relative risk, 4.5 [95% CI, 2.0 to 10.2]; P<.001), or FIB4 (sensitivity, 61% [95% CI, 41% to 78%]; negative predictive value, 67% [95% CI, 68% to 90%]; relative risk, 2.2 [95% CI, 1.1 to 4.6]; P=.03). Mean cell volume values were significantly higher pretreatment in patients with F3-4 fibrosis (96.7±1.1 fL) compared with F0-2 fibrosis (93.4±0.5 fL; P=.004) and declined following treatment (F3-4, 93.2±0.9 fL, P=.01; F0-2, 91.7±0.6 fL, P=.01). CONCLUSION: Advanced hepatic fibrosis is strongly associated with arthritis in HFE hemochromatosis. The absence of arthritis predicts a low likelihood of advanced hepatic fibrosis, supporting its use as a clinical marker for advanced hepatic fibrosis in HFE hemochromatosis.

Therapeutic potential of macrophage colony-stimulating factor in chronic liver disease.

Resident and recruited macrophages control the development and proliferation of the liver. We have previously shown in multiple species that treatment with a macrophage colony stimulating factor (CSF1)-Fc fusion protein initiated hepatocyte proliferation and promoted repair in models of acute hepatic injury in mice. Here, we investigated the impact of CSF1-Fc on resolution of advanced fibrosis and liver regeneration, using a non-resolving toxin-induced model of chronic liver injury and fibrosis in C57BL/6J mice. Co-administration of CSF1-Fc with exposure to thioacetamide (TAA) exacerbated inflammation consistent with monocyte contributions to initiation of pathology. After removal of TAA, either acute or chronic CSF1-Fc treatment promoted liver growth, prevented progression and promoted resolution of fibrosis. Acute CSF1-Fc treatment was also anti-fibrotic and pro-regenerative in a model of partial hepatectomy in mice with established fibrosis. The beneficial impacts of CSF1-Fc treatment were associated with monocyte-macrophage recruitment and increased expression of remodelling enzymes and growth factors. These studies indicate that CSF1-dependent macrophages contribute to both initiation and resolution of fibrotic injury and that CSF1-Fc has therapeutic potential in human liver disease.

Ursodeoxycholic acid and liver disease associated with cystic fibrosis: A multicenter cohort study.

BACKGROUND: The efficacy and safety of ursodeoxycholic acid (UDCA) for the treatment of liver disease associated with cystic fibrosis (CF) are under discussion, and clinical practice varies among centers. The study aimed at evaluating if the incidence of severe liver disease differs between CF centers routinely prescribing or not prescribing UDCA. METHODS: We carried out a retrospective multicenter cohort study including 1591 CF patients (1192 patients from UDCA-prescribing centers and 399 from non-prescribing centers) born between 1990 and 2007 and followed from birth up to 31 December 2016. We computed the crude cumulative incidence (CCI) of portal hypertension (PH) at the age of 20 years in the two groups and estimated the subdistribution hazard ratio (HR) through a Fine and Gray model. RESULTS: Over the observation period, 114 patients developed PH: 90 (7.6%) patients followed-up in UDCA prescribing centers and 24 (6.0%) in non-prescribing centers. The CCI of PH at 20 years was 10.1% (95% CI: 7.9-12.3) in UDCA-prescribing and 7.7% (95% CI: 4.6-10.7) in non-prescribing centers. The HR among patients followed in prescribing centers indicated no significant difference in the rate of PH either in the unadjusted model (HR: 1.21, 95% CI: 0.69-2.11) or in the model adjusted for pancreatic insufficiency (HR: 1.28, 95% CI: 0.77-2.12). CONCLUSIONS: CF patients followed-up in UDCA prescribing centers did not show a lower incidence of PH as compared to those followed in centers not prescribing UDCA. These results question the utility of UDCA in reducing the occurrence of severe liver disease in CF.

Advanced but not mild liver disease is a predictor of decreased survival in children with cystic fibrosis, with far greater impact in females: A 27-year real-life cohort study.

BACKGROUND: Improved survival of children with CF has increased our need to understand the relevance of cystic fibrosis-associated liver disease (CFLD). We assessed the impact of liver disease and disease severity on the survival of children with cystic fibrosis. METHODS: A real life, single center cohort study with 27 years follow up was conducted. Mild CFLD was diagnosed as children with abnormal serum liver function tests and abnormal ultrasound. Advanced CFLD was established by detection of cirrhosis or portal hypertension. A directed acyclic graph, Kaplan-Meier (KM) and Cox regression analysis were used to model survival. RESULTS: 290 patients were enrolled, 48 (16.5%) had mild CFLD and 55 (19%) had advanced CFLD. Ten children with advanced CFLD and 1 with mild CFLD died. Based on the KM analysis, the mean (SE) overall survival age of all CF children was 29.1 years (0.50). The mean (SE) survival among females with advanced CFLD was 24.7 years (1.58) compared to 30.4 years (0.66) for females without advanced CFLD (p = 0.0027). Advanced CFLD was a predictor of decreased survival when adjusted for sex and diabetes (HR 2.54, 95%CI 1.05-6.15, p = 0.039). Mild CFLD was not associated with decreased survival. The effect of advanced CFLD on survival was mainly borne by females (HR = 6.37, 95%CI 1.62-25.06 vs. males, HR = 1.00, 95%CI 0.25-4.01). CONCLUSION: Advanced but not mild CFLD was associated with an increased risk of death when adjusted for sex and diabetes, and resulted in premature death in females with cystic fibrosis by approximately 6 years.

Gamma-glutamyl transpeptidase-to-platelet ratio as a biomarker of liver disease and hepatic fibrosis severity in paediatric Cystic Fibrosis.

BACKGROUND: Cystic fibrosis (CF)-associated liver disease (CFLD) causes significant morbidity and mortality in children with CF. Diagnosis of liver disease prior to development of cirrhosis or portal hypertension (PHT) is challenging. While imaging modalities using Elastography show great promise they are still not widely available to all clinicians. This study investigated gamma-glutamyl transpeptidase-to-platelet ratio (GPR) as a non-invasive biomarker to detect liver disease and stage fibrosis severity in children with CF. METHODS: 237 children were enroled including 76 with CFLD and 161 with CF and no detectable liver disease (CFnoLD). CFLD was diagnosed using standard clinical, biochemical and imaging practice guidelines. Hepatic fibrosis was staged on liver biopsies available from 54 children with CFLD. Serum liver biochemistry was used to calculate GPR (median, [IQR]) and receiver operating characteristics (ROC) analysis assessed utility to detect liver disease and stage fibrosis severity. RESULTS: GPR was significantly increased in CFLD versus CFnoLD (0.33 [0.19-0.96] vs. 0.15 [0.11-0.21], P<0.0001). GPR demonstrated good diagnostic utility for detecting CFLD with an area under the curve (AUC) of 0.81 (95% confidence Interval [CI] [0.75-0.87]; P<0.0001), with sensitivity of 74% and specificity of 73%, using a cut-off of 0.20. GPR increased with increasing hepatic fibrosis stage. GPR discriminated both moderate-advanced (F2-F4) fibrosis vs. F0-F1 (AUC=0.82; 95%CI [0.71-0.94]; P<0.0001) and advanced (F3-F4) fibrosis vs. F0-F2 (AUC=0.77; 95%CI [0.64-0.90]; P = 0.004), with a cut-off 0.32 and 0.61, respectively. An elevated GPR of >0.84 was predictive of PHT at diagnosis of CFLD (AUC=0.81; 95%CI [0.67-0.95]; P = 0.0003). CONCLUSIONS: GPR demonstrates good diagnostic utility for assessing the presence of liver disease, PHT and hepatic fibrosis severity in children with CF. These findings will aid in better identification of patients at risk for CF-related liver involvement and the potential for more targeted and timely follow-up and treatment.

Cost-Effectiveness of a Serum Biomarker Test for Risk-Stratified Liver Ultrasound Screening for Hepatocellular Carcinoma.

OBJECTIVES: Risk-stratified ultrasound screening for hepatocellular carcinoma (HCC), informed by a serum biomarker test, enables resources to be targeted to patients at the highest risk of developing cancer. We aimed to investigate the cost-effectiveness of risk-stratified screening for HCC in the Australian healthcare system. METHODS: A Markov cohort model was constructed to test 3 scenarios for patients with compensated cirrhosis: (1) risk-stratified screening for high-risk patients, (2) all-inclusive screening, and (3) no formal screening. Probabilistic sensitivity analyses were undertaken to determine the impact of uncertainty. Scenario analyses were used to assess cost-effectiveness in Australia's Aboriginal and Torres Strait Islander peoples and to determine the impact of including productivity-related costs of mortality. RESULTS: Both risk-stratified screening and all-inclusive screening programs were cost-effective compared with no formal screening, with incremental cost-effectiveness ratios of A$39 045 and A$23 090 per quality-adjusted life-year (QALY), respectively. All-inclusive screening had an incremental cost-effectiveness ratio of A$4453 compared with risk-stratified screening and had the highest probability of being cost-effective at a willingness-to-pay (WTP) threshold of A$50 000 per QALY. Risk-stratified screening had the highest likelihood of cost-effectiveness when the WTP was between A$25 000 and A$35 000 per QALY. Cost-effectiveness results were further strengthened when applied to an Aboriginal and Torres Strait Islander cohort and when productivity costs were included. CONCLUSIONS: Cirrhosis population-wide screening for HCC is likely to be cost-effective in Australia. Risk-stratified screening using a serum biomarker test may be cost-effective at lower WTP thresholds.

Androgens alter the heterogeneity of small extracellular vesicles and the small RNA cargo in prostate cancer.

Proliferation and survival of prostate cancer cells are driven by the androgen receptor (AR) upon binding to androgen steroid hormones. Manipulating the AR signalling axis is the focus for prostate cancer therapy; thus, it is crucial to understand the role of androgens and AR on extracellular vesicle (EV) secretion and cargo. In this study, we report that plasma-derived circulating vesicles consisting of CD9 and double-positive for CD9 and Prostate Specific Membrane Antigen (PSMA) are increased in patients with advanced metastatic prostate cancer, whereas double positives for CD9 and CD63 small extracellular vesicles (S-EVs) are significantly higher in patients with localised prostate cancer. Androgen manipulation by dihydrotestosterone (DHT) and the clinical antagonist enzalutamide (ENZ) altered the heterogeneity and size of CD9 positive S-EVs in AR expressing prostate cancer cells, while assessment of the total number and protein cargo of total S-EVs was unaltered across different treatment groups. Furthermore, hormone stimulation caused strong and specific effects on the small RNA cargo of S-EVs. A total of 543 small RNAs were found to be regulated by androgens including miR-19-3p and miR-361-5p. Analysis of S-EVs heterogeneity and small RNA cargo may provide clinical utility for prostate cancer and be informative to understand further the mechanism of resistance to androgen targeted therapy in castration-resistant prostate cancer.

Extracellular vesicles in the development of organ-specific metastasis.

Distant organ metastasis, often termed as organotropic metastasis or metastatic organotropism, is a fundamental feature of malignant tumours and accounts for most cancer-related mortalities. This process is orchestrated by many complex biological interactions and processes that are mediated by a combination of anatomical, genetic, pathophysiological and biochemical factors. Recently, extracellular vesicles (EVs) are increasingly being demonstrated as critical mediators of bi-directional tumour-host cell interactions, controlling organ-specific infiltration, adaptation and colonization at the secondary site. EVs govern organotropic metastasis by modulating the pre-metastatic microenvironment through upregulation of pro-inflammatory gene expression and immunosuppressive cytokine secretion, induction of phenotype-specific differentiation and recruitment of specific stromal cell types. This review discusses EV-mediated metastatic organotropism in visceral (brain, lung, liver, and lymph node) and skeletal (bone) metastasis, and discusses how the pre-metastatic education by EVs transforms the organ into a hospitable, tumour cell-friendly milieu that supports the growth of metastatic cells. Decoding the organ-specific traits of EVs and their functions in organotropic metastasis is essential in accelerating the clinical application of EVs in cancer management.

Utility and limitations of Hepascore and transient elastography to detect advanced hepatic fibrosis in HFE hemochromatosis.

Aspartate aminotransferase-to-platelet ratio index (APRI) and Fibrosis-4 Index (Fib4) have been validated against liver biopsy for detecting advanced hepatic fibrosis in HFE hemochromatosis. We determined the diagnostic utility for advanced hepatic fibrosis of Hepascore and transient elastography compared with APRI and Fib4 in 134 newly diagnosed HFE hemochromatosis subjects with serum ferritin levels > 300 µg/L using area under the receiver operator characteristic curve (AUROC) analysis and APRI- (> 0.44) or Fib4- (> 1.1) cut-offs for AHF, or a combination of both. Compared with APRI, Hepascore demonstrated an AUROC for advanced fibrosis of 0.69 (95% CI 0.56-0.83; sensitivity = 69%, specificity = 65%; P = 0.01) at a cut-off of 0.22. Using a combination of APRI and Fib4, the AUROC for Hepascore for advanced fibrosis was 0.70 (95% CI 0.54-0.86, P = 0.02). Hepascore was not diagnostic for detection of advanced fibrosis using the Fib4 cut-off. Elastography was not diagnostic using either APRI or Fib4 cut-offs. Hepascore and elastography detected significantly fewer true positive or true negative cases of advanced fibrosis compared with APRI and Fib4, except in subjects with serum ferritin levels > 1000 µg/L. In comparison with APRI or Fib4, Hepascore or elastography may underdiagnose advanced fibrosis in HFE Hemochromatosis, except in individuals with serum ferritin levels > 1000 µg/L.

LXR stimulates a metabolic switch and reveals cholesterol homeostasis as a statin target in Tasmanian devil facial tumor disease.

Devil facial tumor disease (DFTD) and its lack of available therapies are propelling the Tasmanian devil population toward extinction. This study demonstrates that cholesterol homeostasis and carbohydrate energy metabolism sustain the proliferation of DFTD cells in a cell-type-dependent manner. In addition, we show that the liver-X nuclear receptor-ß (LXRß), a major cholesterol cellular sensor, and its natural ligand 24S-hydroxycholesterol promote the proliferation of DFTD cells via a metabolic switch toward aerobic glycolysis. As a proof of concept of the role of cholesterol homeostasis on DFTD proliferation, we show that atorvastatin, an FDA-approved statin-drug subtype used against human cardiovascular diseases that inhibits cholesterol synthesis, shuts down DFTD energy metabolism and prevents tumor growth in an in vivo DFTD-xenograft model. In conclusion, we show that intervention against cholesterol homeostasis and carbohydrate-dependent energy metabolism by atorvastatin constitutes a feasible biochemical treatment against DFTD, which may assist in the conservation of the Tasmanian devil.

Growth Hormone Stops Excessive Inflammation After Partial Hepatectomy, Allowing Liver Regeneration and Survival Through Induction of H2-Bl/HLA-G.

BACKGROUND AND AIMS: Growth hormone (GH) is important for liver regeneration after partial hepatectomy (PHx). We investigated this process in C57BL/6 mice that express different forms of the GH receptor (GHR) with deletions in key signaling domains. APPROACH AND RESULTS: PHx was performed on C57BL/6 mice lacking GHR (Ghr-/- ), disabled for all GH-dependent Janus kinase 2 signaling (Box1-/- ), or lacking only GH-dependent signal transducer and activator of transcription 5 (STAT5) signaling (Ghr391-/- ), and wild-type littermates. C57BL/6 Ghr-/- mice showed striking mortality within 48 hours after PHx, whereas Box1-/- or Ghr391-/- mice survived with normal liver regeneration. Ghr-/- mortality was associated with increased apoptosis and elevated natural killer/natural killer T cell and macrophage cell markers. We identified H2-Bl, a key immunotolerance protein, which is up-regulated by PHx through a GH-mediated, Janus kinase 2-independent, SRC family kinase-dependent pathway. GH treatment was confirmed to up-regulate expression of the human homolog of H2-Bl (human leukocyte antigen G [HLA-G]) in primary human hepatocytes and in the serum of GH-deficient patients. We find that injury-associated innate immune attack by natural killer/natural killer T cell and macrophage cells are instrumental in the failure of liver regeneration, and this can be overcome in Ghr-/- mice by adenoviral delivery of H2-Bl or by infusion of HLA-G protein. Further, H2-Bl knockdown in wild-type C57BL/6 mice showed elevated markers of inflammation after PHx, whereas Ghr-/- backcrossed on a strain with high endogenous H2-Bl expression showed a high rate of survival following PHx. CONCLUSIONS: GH induction of H2-Bl expression is crucial for reducing innate immune-mediated apoptosis and promoting survival after PHx in C57BL/6 mice. Treatment with HLA-G may lead to improved clinical outcomes following liver surgery or transplantation.

TWEAK/Fn14 signalling promotes cholangiocarcinoma niche formation and progression.

BACKGROUND & AIMS: Cholangiocarcinoma (CCA) is a cancer of the hepatic bile ducts that is rarely resectable and is associated with poor prognosis. Tumour necrosis factor-like weak inducer of apoptosis (TWEAK) is known to signal via its receptor fibroblast growth factor-inducible 14 (Fn14) and induce cholangiocyte and myofibroblast proliferation in liver injury. We aimed to characterise its role in CCA. METHODS: The expression of the TWEAK ligand and Fn14 receptor was assessed immunohistochemically and by bulk RNA and single cell transcriptomics of human liver tissue. Spatiotemporal dynamics of pathway regulation were comprehensively analysed in rat and mouse models of thioacetamide (TAA)-mediated CCA. Flow cytometry, qPCR and proteomic analyses of CCA cell lines and conditioned medium experiments with primary macrophages were performed to evaluate the downstream functions of TWEAK/Fn14. In vivo pathway manipulation was assessed via TWEAK overexpression in NICD/AKT-induced CCA or genetic Fn14 knockout during TAA-mediated carcinogenesis. RESULTS: Our data reveal TWEAK and Fn14 overexpression in multiple human CCA cohorts, and Fn14 upregulation in early TAA-induced carcinogenesis. TWEAK regulated the secretion of factors from CC-SW-1 and SNU-1079 CCA cells, inducing polarisation of proinflammatory CD206+ macrophages. Pharmacological blocking of the TWEAK downstream target chemokine monocyte chemoattractant protein 1 (MCP-1 or CCL2) significantly reduced CCA xenograft growth, while TWEAK overexpression drove cancer-associated fibroblast proliferation and collagen deposition in the tumour niche. Genetic Fn14 ablation significantly reduced inflammatory, fibrogenic and ductular responses during carcinogenic TAA-mediated injury. CONCLUSION: These novel data provide evidence for the action of TWEAK/Fn14 on macrophage recruitment and phenotype, and cancer-associated fibroblast proliferation in CCA. Targeting TWEAK/Fn14 and its downstream signals may provide a means to inhibit CCA niche development and tumour growth. LAY SUMMARY: Cholangiocarcinoma is an aggressive, chemotherapy-resistant liver cancer. Interactions between tumour cells and cells that form a supportive environment for the tumour to grow are a source of this aggressiveness and resistance to chemotherapy. Herein, we describe interactions between tumour cells and their supportive environment via a chemical messenger, TWEAK and its receptor Fn14. TWEAK/Fn14 alters the recruitment and type of immune cells in tumours, increases the growth of cancer-associated fibroblasts in the tumour environment, and is a potential target to reduce tumour formation.