Non-canonical Hedgehog signaling mediates profibrotic hematopoiesis-stroma crosstalk in myeloproliferative neoplasms.

The role of hematopoietic Hedgehog signaling in myeloproliferative neoplasms (MPNs) remains incompletely understood despite data suggesting that Hedgehog (Hh) pathway inhibitors have therapeutic activity in patients. We aim to systematically interrogate the role of canonical vs. non-canonical Hh signaling in MPNs. We show that Gli1 protein levels in patient peripheral blood mononuclear cells (PBMCs) mark fibrotic progression and that, in murine MPN models, absence of hematopoietic Gli1, but not Gli2 or Smo, significantly reduces MPN phenotype and fibrosis, indicating that GLI1 in the MPN clone can be activated in a non-canonical fashion. Additionally, we establish that hematopoietic Gli1 has a significant effect on stromal cells, mediated through a druggable MIF-CD74 axis. These data highlight the complex interplay between alterations in the MPN clone and activation of stromal cells and indicate that Gli1 represents a promising therapeutic target in MPNs, particularly that Hh signaling is dispensable for normal hematopoiesis.

Survival after combined resection and ablation is not inferior to that after resection alone, in patients with four or more colorectal liver metastases.

PURPOSE: Colorectal liver metastases (CRLM) are the predominant factor limiting survival in patients with colorectal cancer. Multimodal treatment strategies are frequently necessary to achieve total tumor elimination. This study examines the efficacy of liver resection combined with local ablative therapy in comparison to liver resection only, in the treatment of patients with ≥ 4 CRLM. METHODS: This retrospective cohort study was conducted at the University Hospital RWTH Aachen, Germany. Patients with ≥ 4 CRLM in preoperative imaging, who underwent curative resection between 2010-2021, were included. Recurrent resections and deaths in the early postoperative phase were excluded. Ablation modalities included radiofrequency or microwave ablation, and irreversible electroporation. Differences in overall- (OS) and recurrence-free-survival (RFS) between patients undergoing combined resection-ablation vs. resection only, were examined. RESULTS: Of 178 included patients, 46 (27%) underwent combined resection-ablation and 132 (73%) resection only. Apart from increased rates of adjuvant chemotherapy in the first group (44% vs. 25%, p = 0.014), there were no differences in perioperative systemic therapy. Kaplan-Meier and log-rank test analyses showed no statistically significant differences in median OS (36 months for both, p = 0.638) or RFS (9 months for combined resection-ablation vs. 8 months, p = 0.921). Cox regression analysis showed a hazard ratio of 0.891 (p = 0.642) for OS and 0.981 (p = 0.924) for RFS, for patients undergoing resection only. CONCLUSION: For patients with ≥ 4 CRLM, combined resection-ablation is a viable option in terms of OS and RFS. Therefore, combined resection-ablation should be considered for complete tumor clearance, in patients with multifocal disease.

Tissue Inhibitor of Metalloproteinases-1 Interacts with CD74 to Promote AKT Signaling, Monocyte Recruitment Responses, and Vascular Smooth Muscle Cell Proliferation.

Tissue inhibitor of metalloproteinases-1 (TIMP-1), an important regulator of matrix metalloproteinases (MMPs), has recently been shown to interact with CD74, a receptor for macrophage migration inhibitory factor (MIF). However, the biological effects mediated by TIMP-1 through CD74 remain largely unexplored. Using sequence alignment and in silico protein-protein docking analysis, we demonstrated that TIMP-1 shares residues with both MIF and MIF-2, crucial for CD74 binding, but not for CXCR4. Subcellular colocalization, immunoprecipitation, and internalization experiments supported these findings, demonstrating that TIMP-1 interacts with surface-expressed CD74, resulting in its internalization in a dose-dependent manner, as well as with a soluble CD74 ectodomain fragment (sCD74). This prompted us to study the effects of the TIMP-1-CD74 axis on monocytes and vascular smooth muscle cells (VSCMs) to assess its impact on vascular inflammation. A phospho-kinase array revealed the activation of serine/threonine kinases by TIMP-1 in THP-1 pre-monocytes, in particular AKT. Similarly, TIMP-1 dose-dependently triggered the phosphorylation of AKT and ERK1/2 in primary human monocytes. Importantly, Transwell migration, 3D-based Chemotaxis, and flow adhesion assays demonstrated that TIMP-1 engagement of CD74 strongly promotes the recruitment response of primary human monocytes, while live cell imaging studies revealed a profound activating effect on VSMC proliferation. Finally, re-analysis of scRNA-seq data highlighted the expression patterns of TIMP-1 and CD74 in human atherosclerotic lesions, thus, together with our experimental data, indicating a role for the TIMP-1-CD74 axis in vascular inflammation and atherosclerosis.

Value of prognostic scoring systems in the era of multimodal therapy for recurrent colorectal liver metastases.

BACKGROUND: Various predictive scoring systems have been developed to estimate outcomes of patients undergoing surgery for colorectal liver metastases (CRLM). However, data regarding their effectiveness in recurrent CRLM (recCRLM) are very limited. METHODS: Patients who underwent repeat hepatectomy for recCRLM at the University Hospital RWTH Aachen, Germany from 2010 to 2021 were included. Nine predictive scoring systems (Fong's, Nordlinger, Nagashima, RAS mutation, Tumor Burden, GAME, CERR, and Glasgow Prognostic score, Basingstoke Index) were evaluated by likelihood ratio (LR) χ2, linear trend (LT) χ2 and Akaike Information Criterion (AIC) for their predictive value regarding overall survival (OS) and recurrence free survival (RFS). RESULTS: Among 150 patients, median RFS was 9 (2-124) months with a 5-year RFS rate of 10%. Median OS was 39 (4-131) months with a 5-year OS rate of 32%. For RFS and OS, the Nagashima score showed the best prognostic ability (LT χ2 3.00, LR χ2 9.39, AIC 266.66 and LT χ2 2.91, LR χ2 20.91, 290.36). DISCUSSION: The Nagashima score showed the best prognostic stratification to predict recurrence as well as survival, and therefore might be considered when evaluating patients with recCRLM for repeat hepatectomy.

Intermittent Fasting Primes the Tumor Microenvironment and Improves Nanomedicine Delivery in Hepatocellular Carcinoma.

Fasting has many health benefits, including reduced chemotherapy toxicity and improved efficacy. It is unclear how fasting affects the tumor microenvironment (TME) and tumor-targeted drug delivery. Here the effects of intermittent (IF) and short-term (STF) fasting are investigated on tumor growth, TME composition, and liposome delivery in allogeneic hepatocellular carcinoma (HCC) mouse models. To this end, mice are inoculated either subcutaneously or intrahepatically with Hep-55.1C cells and subjected to IF for 24 d or to STF for 1 d. IF but not STF significantly slows down tumor growth. IF increases tumor vascularization and decreases collagen density, resulting in improved liposome delivery. In vitro, fasting furthermore promotes the tumor cell uptake of liposomes. These results demonstrate that IF shapes the TME in HCC towards enhanced drug delivery. Finally, when combining IF with liposomal doxorubicin treatment, the antitumor efficacy of nanochemotherapy is found to be increased, while systemic side effects are reduced. Altogether, these findings exemplify that the beneficial effects of fasting on anticancer therapy outcomes go beyond modulating metabolism at the molecular level.

Study protocol of an open-label, single arm phase II trial investigating the efficacy and safety of Trifluridine/Tipiracil combined with irinotecan as a second line therapy in patients with cholangiocarcinoma (TRITICC).

BACKGROUND: The prognosis of patients with advanced biliary tract cancer (BTC) who have progressed on gemcitabine plus cisplatin is dismal. Trifluridine/tipiracil (FTD/TPI) and irinotecan have proven efficacy in different gastrointestinal malignancies. We therefore hypothesized that this combination might improve the therapeutic outcome in patients with BTC after failure of first line treatment. METHODS: TRITICC is an interventional, prospective, open-label, non-randomised, exploratory, multicentre, single-arm phase IIA clinical trial done in 6 sites with expertise in managing biliary tract cancer across Germany. A total of 28 adult patients (aged ≥ 18 years) with histologically verified locally advanced or metastatic biliary tract cancer (including cholangiocarcinoma and gallbladder or ampullary carcinoma) with documented radiological disease progression to first-line gemcitabine based chemotherapy will be included to receive a combination of FTD/TPI plus irinotecan according to previously published protocols. Study treatment will be continued until disease progression according to RECIST 1.1 criteria or occurrence of unacceptable toxicity. The effect of FTD/TPI plus irinotecan on progression-free survival will be analyzed as primary endpoint. Safety (according to NCI-CTCAE), response rates and overall survival are secondary endpoints. In addition, a comprehensive translational research program is part of the study and might provide findings about predictive markers with regard to response, survival periods and resistance to treatment. DISCUSSION: The aim of TRITICC is to evaluate the safety and efficacy of FTD/TPI plus irinotecan in patients with biliary tract cancer refractory to previous Gemcitabine based treatment. TRIAL REGISTRATION: EudraCT 2018-002936-26; NCT04059562.

The non-selective Rho-kinase inhibitors Y-27632 and Y-33075 decrease contraction but increase migration in murine and human hepatic stellate cells.

BACKGROUND: The Rho-kinase ROCK II plays a major role in the activation of hepatic stellate cells (HSC), which are the key profibrotic and contractile cells contributing to the development of chronic liver disease. Inhibition of ROCK II ultimately blocks the phosphorylation of the myosin light chain (MLC) and thus inhibits stress fibre assembly and cell contraction. We investigated the effects of the ROCK inhibitors Y-33075 as well as Y-27632 in murine and human hepatic stellate cells. METHODS: Primary isolated HSC from FVB/NJ mice and the immortalized human HSC line TWNT-4 were culture-activated and incubated with Y-27632 and Y-33075 (10nM to 10µM) for 24h. Protein expression levels were analyzed by Western Blots and transcriptional levels of pro-fibrotic markers and proliferative markers were evaluated using real-time qPCR. Migration was investigated by wound-healing assay. Proliferation was assessed by BrdU assay. Contraction of HSC was measured using 3D collagen matrices after incubation with Y-27632 or Y-33075 in different doses. RESULTS: Both Rho-kinase inhibitors, Y-27632 and Y-33075, reduced contraction, fibrogenesis and proliferation in activated primary mouse HSC (FVB/NJ) and human HSC line (TWNT-4) significantly. Y-33075 demonstrated a 10-times increased potency compared to Y-27632. Surprisingly, both inhibitors mediated a substantial and unexpected increase in migration of HSC in FVB/NJ. CONCLUSION: ROCK inhibition by the tested compounds decreased contraction but increased migration. Y-33075 proved more potent than Y27632 in the inhibition of contraction of HSCs and should be further evaluated in chronic liver disease.

Janus kinase 2 inhibition by pacritinib as potential therapeutic target for liver fibrosis.

BACKGROUND AND AIMS: Janus kinase 2 (JAK2) signaling is increased in human and experimental liver fibrosis with portal hypertension. JAK2 inhibitors, such as pacritinib, are already in advanced clinical development for other indications and might also be effective in liver fibrosis. Here, we investigated the antifibrotic role of the JAK2 inhibitor pacritinib on activated hepatic stellate cells (HSCs) in vitro and in two animal models of liver fibrosis in vivo . APPROACH AND RESULTS: Transcriptome analyses of JAK2 in human livers and other targets of pacritinib have been shown to correlate with profibrotic factors. Although transcription of JAK2 correlated significantly with type I collagen expression and other profibrotic genes, no correlation was observed for interleukin-1 receptor-associated kinase and colony-stimulating factor 1 receptor. Pacritinib decreased gene expression of fibrosis markers in mouse primary and human-derived HSCs in vitro . Moreover, pacritinib decreased the proliferation, contraction, and migration of HSCs. C 57 BL/6J mice received ethanol in drinking water (16%) or Western diet in combination with carbon tetrachloride intoxication for 7 weeks to induce alcoholic or nonalcoholic fatty liver disease. Pacritinib significantly reduced liver fibrosis assessed by gene expression and Sirius red staining, as well as HSC activation assessed by alpha-smooth muscle actin immunostaining in fibrotic mice. Furthermore, pacritinib decreased the gene expression of hepatic steatosis markers in experimental alcoholic liver disease. Additionally, pacritinib protected against liver injury as assessed by aminotransferase levels. CONCLUSIONS: This study demonstrates that the JAK2 inhibitor pacritinib may be promising for the treatment of alcoholic and nonalcoholic liver fibrosis and may be therefore relevant for human pathology.

Blood Endotoxin Levels as Biomarker of Nonalcoholic Fatty Liver Disease: A Systematic Review and Meta-analysis.

BACKGROUND & AIMS: Growing evidence supports a role of gut-derived metabolites in nonalcoholic fatty liver disease (NAFLD), but the relation of endotoxin levels with gut permeability and NAFLD stage remains unclear. This systematic review with meta-analysis aims to provide further insights. METHODS: PubMed, Embase, and Cochrane Library were searched for studies published until January 2022 assessing blood endotoxins in patients with NAFLD. Meta-analyses and univariate/multivariate meta-regression, as well as correlation analyses, were performed for endotoxin values and potential relationships to disease stage, age, sex, parameters of systemic inflammation, and metabolic syndrome, as well as liver function and histology. RESULTS: Forty-three studies were included, of which 34 were used for meta-analyses. Blood endotoxin levels were higher in patients with simple steatosis vs liver-healthy controls (standardized mean difference, 0.86; 95% confidence interval, 0.62-1.11) as well as in patients with nonalcoholic steatohepatitis vs patients with nonalcoholic fatty liver/non-nonalcoholic steatohepatitis (standardized mean difference, 0.81; 95% confidence interval, 0.27-1.35; P = .0078). Consistently, higher endotoxin levels were observed in patients with more advanced histopathological gradings of liver steatosis and fibrosis. An increase of blood endotoxin levels was partially attributed to a body mass index rise in patients with NAFLD compared with controls. Nevertheless, significant increases of blood endotoxin levels in NAFLD retained after compensation for differences in body mass index, metabolic condition, or liver enzymes. Increases in blood endotoxin levels were associated with increases in C-reactive protein concentrations, and in most cases, paralleled a rise in markers for intestinal permeability. CONCLUSION: Our results support blood endotoxin levels as relevant diagnostic biomarker for NAFLD, both for disease detection as well as staging during disease progression, and might serve as surrogate marker of enhanced intestinal permeability in NAFLD. Registration number in Prospero: CRD42022311166.

Chemokine CXCL10 Modulates the Tumor Microenvironment of Fibrosis-Associated Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) constitutes a devastating health burden. Recently, tumor microenvironment-directed interventions have profoundly changed the landscape of HCC therapy. In the present study, the function of the chemokine CXCL10 during fibrosis-associated hepatocarcinogenesis was analyzed with specific focus on its impact in shaping the tumor microenvironment. C57BL/6J wild type (WT) and Cxcl10 knockout mice (Cxcl10-/-) were treated with diethylnitrosamine (DEN) and tetrachloromethane (CCl4) to induce fibrosis-associated HCCs. Cxcl10 deficiency attenuated hepatocarcinogenesis by decreasing tumor cell proliferation as well as tumor vascularization and modulated tumor-associated extracellular matrix composition. Furthermore, the genetic inactivation of Cxcl10 mediated an alteration of the tumor-associated immune response and modified chemokine/chemokine receptor networks. The DEN/CCl4-treated Cxcl10-/- mice presented with a pro-inflammatory tumor microenvironment and an accumulation of anti-tumoral immune cells in the tissue. The most striking alteration in the Cxcl10-/- tumor immune microenvironment was a vast accumulation of anti-tumoral T cells in the invasive tumor margin. In summary, our results demonstrate that CXCL10 exerts a non-redundant impact on several hallmarks of the tumor microenvironment and especially modulates the infiltration of anti-tumorigenic immune cells in HCC. In the era of microenvironment-targeted HCC therapies, interfering with CXCL10 defines a novel asset for further improvement of therapeutic strategies.

Extracellular Vesicles from Steatotic Hepatocytes Provoke Pro-Fibrotic Responses in Cultured Stellate Cells.

Hepatic steatosis and chronic hepatocyte damage ultimately lead to liver fibrosis. Key pathophysiological steps are the activation and transdifferentiation of hepatic stellate cells. We assessed the interplay between hepatocytes and hepatic stellate cells under normal and steatotic conditions. We hypothesized that hepatocyte-derived extracellular vesicles (EVs) modify the phenotype of stellate cells. By high speed centrifugation, EVs were isolated from conditioned media of the hepatocellular carcinoma cell line HepG2 under baseline conditions (C-EVs) or after induction of steatosis by linoleic and oleic acids for 24 h (FA-EVs). Migration of the human stellate cell line TWNT4 and of primary human stellate cells towards the respective EVs and sera of MAFLD patients were investigated using Boyden chambers. Phenotype alterations after incubation with EVs were determined by qRT-PCR, Western blotting and immunofluorescence staining. HepG2 cells released more EVs after treatment with fatty acids. Chemotactic migration of TWNT4 and primary hepatic stellate cells was increased, specifically towards FA-EVs. Prolonged incubation of TWNT4 cells with FA-EVs induced expression of proliferation markers and a myofibroblast-like phenotype. Though the expression of the collagen type 1 α1 gene did not change after FA-EV treatment, expression of the myofibroblast markers, e.g., α-smooth-muscle-cell actin and TIMP1, was significantly increased. We conclude that EVs from steatotic hepatocytes can influence the behavior, phenotypes and expression levels of remodeling markers of stellate cells and guides their directed migration. These findings imply EVs as operational, intercellular communicators in the pathophysiology of steatosis-associated liver fibrosis and might represent a novel diagnostic parameter and therapeutic target.

JAM-A is a multifaceted regulator in hepatic fibrogenesis, supporting LSEC integrity and stellate cell quiescence.

BACKGROUND AND AIMS: Leukocyte infiltration is a hallmark of hepatic inflammation. The Junctional Adhesion Molecule A (JAM-A) is a crucial regulator of leukocyte extravasation and is upregulated in human viral fibrosis. Reduced shear stress within hepatic sinusoids and the specific phenotype of liver sinusoidal endothelial cells (LSEC) cumulate in differing adhesion characteristics during liver fibrosis. The aim of this study was to define the functional role of cell-specific adhesion molecule JAM-A during hepatic fibrogenesis. METHODS: Complete, conditional (intestinal epithelial; endothelial) and bone marrow chimeric Jam-a knockout animals and corresponding C57Bl/6 wild-type animals were treated with carbon tetrachloride (CCl4 , 6 weeks). For functional analyses of JAM-A, comprehensive in vivo studies, co-culture models and flow-based adhesion assays were performed. RESULTS: Complete and bone marrow-derived Jam-a-/- animals showed aggravated fibrosis with increased non-sinusoidal, perivascular accumulation of CD11b+ F4/80+ monocyte-derived macrophages in contrast to wild-type mice. Despite being associated with disturbed epithelial barrier function, an intestinal epithelial Jam-a knockout did not affect fibrogenesis. In endothelial-specific Jam-a-/- animals, liver fibrosis was aggravated alongside sinusoid capillarization and hepatic stellate cell (HSC) activation. HSC activation is induced via Jam-a-/- LSEC-derived secretion of soluble factors. Sinusoid CD31 expression and hedgehog gene signalling were increased, but leukocyte infiltration and adhesion to LSECs remained unaffected. CONCLUSIONS: Our models decipher cell-specific JAM-A to exert crucial functions during hepatic fibrogenesis. JAM-A on bone marrow-derived cells regulates non-sinusoidal vascular immune cell recruitment, while endothelial JAM-A controls liver sinusoid capillarization and HSC quiescence.

Atezolizumab and bevacizumab with transarterial chemoembolization in hepatocellular carcinoma: the DEMAND trial protocol.

The combination of the anti-PD-L1 antibody atezolizumab and the anti-VEGF bevacizumab is the first approved immunotherapeutic regimen for first-line therapy in patients with unresectable hepatocellular carcinoma (HCC), currently approved in more than 80 countries. The efficacy and tolerability of this regimen suggest that the use of atezolizumab + bevacizumab could be extended to the treatment of patients with intermediate-stage HCC in combination with transarterial chemoembolization (TACE). The authors describe the rationale and design of the DEMAND study. This investigator-initiated, multicenter, randomized phase II study is the first trial to evaluate the safety and efficacy of atezolizumab + bevacizumab prior to or in combination with TACE in patients with intermediate-stage HCC. The primary end point is the 24-month survival rate; secondary end points include objective response rate, progression-free survival, safety and quality of life. Clinical Trial Registration: NCT04224636 (ClinicalTrials.gov).

A Radiomics Approach to Predict the Emergence of New Hepatocellular Carcinoma in Computed Tomography for High-Risk Patients with Liver Cirrhosis.

Liver cirrhosis poses a major risk for the development of hepatocellular carcinoma (HCC). This retrospective study investigated to what extent radiomic features allow the prediction of emerging HCC in patients with cirrhosis in contrast-enhanced computed tomography (CECT). A total of 51 patients with liver cirrhosis and newly detected HCC lesions (n = 82) during follow-up (FU-CT) after local tumor therapy were included. These lesions were not to have been detected by the radiologist in the chronologically prior CECT (PRE-CT). For training purposes, segmentations of 22 patients with liver cirrhosis but without HCC-recurrence were added. A total of 186 areas (82 HCCs and 104 cirrhotic liver areas without HCC) were analyzed. Using univariate analysis, four independent features were identified, and a multivariate logistic regression model was trained to classify the outlined regions as "HCC probable" or "HCC improbable". In total, 60/82 (73%) of segmentations with later detected HCC and 84/104 (81%) segmentations without HCC were classified correctly (AUC of 81%, 95% CI 74-87%), yielding a sensitivity of 72% (95% CI 57-83%) and a specificity of 86% (95% CI 76-96%). In conclusion, the model predicted the occurrence of new HCCs within segmented areas with an acceptable sensitivity and specificity in cirrhotic liver tissue in CECT.

Transient elastography in adult patients with cryptic dyskeratosis congenita reveals subclinical liver fibrosis: a retrospective analysis of the Aachen telomere biology disease registry.

BACKGROUND: Telomere biology disorders (TBD) such as dyskeratosis congenita (DKC) lead to progressive multi-organ failure as impaired telomere maintenance disturbs cellular proliferative capacity. A wide range of hepatic manifestations from asymptomatic liver enzyme elevation to overt liver fibrosis/cirrhosis can be observed in TBD patients. However, the incidence of hepatic involvement remains unknown. Non-invasive transient elastography (TE) predicts early fibrosis by measuring liver stiffness and may uncover subclinical liver damage in TBD patients. METHODS: Liver screening procedures of nine TBD patients from the Aachen TBD Registry are being presented retrospectively. Following clinical suspicion, TBD was diagnosed using flow-FISH with telomere length (TL) below the 1% percentile and confirmed by next-generation sequencing (NGS) detecting pathogenic mutations in telomere maintenance genes TERC or TERT. RESULTS: In all patients, TBD was first diagnosed in adulthood. Patients showed normal to slightly elevated liver function test parameters. Hepatic ultrasound revealed inhomogeneous parenchyma in seven (77.7%) and increased liver echogenicity in four patients (44.4%). Median liver stiffness was 10.7 kilopascal (kPa) (interquartile range 8.4, 15.7 kPa). Using 7.1 kPa as cut-off, 88.8% of patients were classified as moderate fibrosis to cirrhosis. CONCLUSION: Subclinical chronic liver involvement is frequent in patients with adult-onset TBD. TE could have a valuable role in the routine work-up of patients with telomere disorders including DKC for early detection of patients at risk for liver function impairment.

Role of circulating angiogenin levels in portal hypertension and TIPS.

BACKGROUND: Pathogenesis of portal hypertension is multifactorial and includes pathologic intrahepatic angiogenesis, whereby TIPS insertion is an effective therapy of portal hypertension associated complications. While angiogenin is a potent contributor to angiogenesis in general, little is known about its impact on TIPS function over time. METHODS: In a total of 118 samples from 47 patients, angiogenin concentrations were measured in portal and inferior caval vein plasma at TIPS insertion (each blood compartment n = 23) or angiographic intervention after TIPS (each blood compartment n = 36) and its relationship with patient outcome was investigated. RESULTS: Angiogenin levels in the inferior caval vein were significantly higher compared to the portal vein (P = 0.048). Ten to 14 days after TIPS, inferior caval vein angiogenin level correlated inversely with the portal systemic pressure gradient (P<0.001), measured invasively during control angiography. Moreover, patients with TIPS revision during this angiography, showed significantly lower angiogenin level in the inferior caval vein compared to patients without TIPS dysfunction (P = 0.01). CONCLUSION: In cirrhosis patients with complications of severe portal hypertension, circulating levels of angiogenin are derived from the injured liver. Moreover, angiogenin levels in the inferior caval vein after TIPS may predict TIPS dysfunction.

Macrophage migration inhibitory factor exerts pro-proliferative and anti-apoptotic effects via CD74 in murine hepatocellular carcinoma.

BACKGROUND AND PURPOSE: Macrophage migration inhibitory factor (MIF) is an inflammatory and chemokine-like protein expressed in different inflammatory diseases as well as solid tumours. CD74-as the cognate MIF receptor-was identified as an important target of MIF. We here analysed the role of MIF and CD74 in the progression of hepatocellular carcinoma (HCC) in vitro and in vivo. EXPERIMENTAL APPROACH: Multilocular HCC was induced using the diethylnitrosamine/carbon tetrachloride (DEN/CCl4 ) model in hepatocyte-specific Mif knockout (Mif Δhep ), Cd74-deficient, and control mice. Tumour burden was compared between the genotypes. MIF, CD74 and Ki67 expression were investigated in tumour and surrounding tissue. In vitro, the effects of the MIF/CD74 axis on the proliferative and apoptotic behaviour of hepatoma cells and respective signalling pathways were assessed after treatment with MIF and anti-CD74 antibodies. KEY RESULTS: DEN/CCl4 treatment of Mif Δhep mice resulted in reduced tumour burden and diminished proliferation capacity within tumour tissue. In vitro, MIF stimulated proliferation of Hepa 1-6 and HepG2 cells, inhibited therapy-induced cell death and induced ERK activation. The investigated effects could be reversed using a neutralizing anti-CD74 antibody, and Cd74-/- mice developed fewer tumours associated with decreased proliferation rates. CONCLUSION AND IMPLICATIONS: We identified a pro-tumorigenic role of MIF during proliferation and therapy-induced apoptosis of HCC cells. These effects were mediated via the MIF cognate receptor CD74. Thus, inhibition of the MIF/CD74 axis could represent a promising target with regard to new pharmacological therapies aimed at HCC.

Macrophage migration inhibitory factor predicts an unfavorable outcome after transarterial chemoembolization for hepatic malignancies.

Transarterial chemoembolization (TACE) is a therapeutic option for patients with intermediate-stage hepatocellular carcinoma (HCC) or metastatic liver cancers. Identifying those patients who particularly benefit from TACE remains challenging. Macrophage migration inhibitory factor (MIF) represents is an inflammatory protein described in patients with liver cancer, but no data on its prognostic relevance in patients undergoing TACE exist. Here, we evaluate MIF serum concentrations as a potential biomarker in patients undergoing TACE for primary and secondary hepatic malignancies. MIF serum concentrations were measured by multiplex immunoassay in 50 patients (HCC: n = 39, liver metastases: n = 11) before and 1 day after TACE as well as in 51 healthy controls. Serum concentrations of MIF did not differ between patients and healthy controls. Interestingly, in the subgroup of patients with larger tumor size, significantly more patients had increased MIF concentrations. Patients with an objective tumor response to TACE therapy showed comparable concentrations of serum MIF compared to patients who did not respond. MIF concentrations at day 1 after TACE were significantly higher compared to baseline concentrations. Importantly, baseline MIF concentrations above the optimal cutoff value (0.625 ng/ml) turned out as a significant and independent prognostic marker for a reduced overall survival (OS) following TACE: patients with elevated MIF concentrations showed a significantly reduced median OS of only 719 days compared to patients below the cutoff value (median OS: 1430 days, p = 0.021). Baseline MIF serum concentrations are associated with tumor size of intrahepatic malignancies and predict outcome of patients with liver cancer receiving TACE.

Balance between macrophage migration inhibitory factor and sCD74 predicts outcome in patients with acute decompensation of cirrhosis.

BACKGROUND & AIMS: Macrophage migration inhibitory factor (MIF) is an inflammatory cytokine and an important regulator of innate immune responses. We hypothesised that serum concentrations of MIF are associated with disease severity and outcome in patients with decompensated cirrhosis and acute-on-chronic liver failure (ACLF). METHODS: Circulating concentrations of MIF and its soluble receptor CD74 (sCD74) were determined in sera from 292 patients with acute decompensation of cirrhosis defined as new onset or worsening of ascites requiring hospitalisation. Of those, 78 (27%) had ACLF. Short-term mortality was assessed 90 days after inclusion. RESULTS: Although serum concentrations of MIF and sCD74 did not correlate with liver function parameters or ACLF, higher MIF (optimum cut-off >2.3 ng/ml) and lower concentrations of sCD74 (optimum cut-off <66.5 ng/ml) both indicated poorer 90-day transplant-free survival in univariate analyses (unadjusted hazard ratio [HR] 2.01 [1.26-3.22]; p = 0.004 for MIF; HR 0.59 [0.38-0.92]; p = 0.02 for sCD74) and after adjustment in multivariable models. Higher MIF concentrations correlated with surrogates of systemic inflammation (white blood cells, p = 0.005; C-reactive protein, p = 0.05) and were independent of genetic MIF promoter polymorphisms. Assessment of MIF plasma concentrations in portal venous blood and matched blood samples from the right atrium in a second cohort of patients undergoing transjugular intrahepatic portosystemic shunt insertion revealed a transhepatic MIF gradient with higher concentrations in the right atrial blood. CONCLUSIONS: Serum concentrations of MIF and its soluble receptor CD74 predict 90-day transplant-free survival in patients with acute decompensation of cirrhosis. This effect was independent of liver function and genetic predispositions, but rather reflected systemic inflammation. Therefore, MIF and sCD74 represent promising prognostic markers beyond classical scoring systems in patients at risk of ACLF. LAY SUMMARY: Inflammatory processes contribute to the increased risk of death in patients with cirrhosis and ascites. We show that patients with high serum levels of the inflammatory cytokine macrophage migration inhibitory factor (MIF) alongside low levels of its binding receptor sCD74 in blood indicate an increased mortality risk in patients with ascites. The cirrhotic liver is a relevant source of elevated circulating MIF levels.

Unexpected Pro-Fibrotic Effect of MIF in Non-Alcoholic Steatohepatitis Is Linked to a Shift in NKT Cell Populations.

Macrophage migration inhibitory factor (MIF) is a pleiotropic inflammatory cytokine with anti-fibrotic properties in toxic liver injury models and anti-steatotic functions in non-alcoholic fatty liver disease (NAFLD) attributed to the CD74/AMPK signaling pathway. As NAFLD progression is associated with fibrosis, we studied MIF function during NAFLD-associated liver fibrogenesis in mice and men by molecular, histological and immunological methods in vitro and in vivo. After NASH diet feeding, hepatic Mif expression was strongly induced, an effect which was absent in Mif∆hep mice. In contrast to hepatotoxic fibrosis models, NASH diet-induced fibrogenesis was significantly abrogated in Mif-/- and Mif∆hep mice associated with a reduced accumulation of the pro-fibrotic type-I NKT cell subpopulation. In vitro, MIF skewed the differentiation of NKT cells towards the type-I subtype. In line with the murine results, expression of fibrosis markers strongly correlated with MIF, its receptors, and markers of NKT type-I cells in NASH patients. We conclude that MIF expression is induced during chronic metabolic injury in mice and men with hepatocytes representing the major source. In NAFLD progression, MIF contributes to liver fibrogenesis skewing NKT cell polarization toward a pro-fibrotic phenotype highlighting the complex, context-dependent role of MIF during chronic liver injury.