Histidine-rich glycoprotein in metabolic dysfunction-associated steatohepatitis-related disease progression and liver carcinogenesis.

Background: Metabolic dysfunction-associated steatotic liver disease (MASLD), previously non-alcoholic fatty liver disease (NAFLD), is a leading cause of chronic liver disease worldwide. In 20%-30% of MASLD patients, the disease progresses to metabolic dysfunction-associated steatohepatitis (MASH, previously NASH) which can lead to fibrosis/cirrhosis, liver failure as well as hepatocellular carcinoma (HCC). Here we investigated the role of histidine-rich glycoprotein (HRG), a plasma protein produced by hepatocytes, in MASLD/MASH progression and HCC development. Methods: The role of HRG was investigated by morphological, cellular, and molecular biology approaches in (a) HRG knock-out mice (HRG-/- mice) fed on a CDAA dietary protocol or a MASH related diethyl-nitrosamine/CDAA protocol of hepatocarcinogenesis, (b) THP1 monocytic cells treated with purified HRG, and (c) well-characterized cohorts of MASLD patients with or without HCC. Results: In non-neoplastic settings, murine and clinical data indicate that HRG increases significantly in parallel with disease progression. In particular, in MASLD/MASH patients, higher levels of HRG plasma levels were detected in subjects with extensive fibrosis/cirrhosis. When submitted to the pro-carcinogenic protocol, HRG-/- mice showed a significant decrease in the volume and number of HCC nodules in relation to decreased infiltration of macrophages producing pro-inflammatory mediators, including IL-1ß, IL-6, IL-12, IL-10, and VEGF as well as impaired angiogenesis. The histopathological analysis (H-score) of MASH-related HCC indicate that the higher HRG positivity in peritumoral tissue significantly correlates with a lower overall patient survival and an increased recurrence. Moreover, a significant increase in HRG plasma levels was detected in cirrhotic (F4) patients and in patients carrying HCC vs. F0/F1 patients. Conclusion: Murine and clinical data indicate that HRG plays a significant role in MASLD/MASH progression to HCC by supporting a specific population of tumor-associated macrophages with pro-inflammatory response and pro-angiogenetic capabilities which critically support cancer cell survival. Furthermore, our data suggest HRG as a possible prognostic predictor in HCC patients with MASLD/MASH-related HCCs.

The protease activated receptor 2 - CCAAT/enhancer-binding protein beta - SerpinB3 axis inhibition as a novel strategy for the treatment of non-alcoholic steatohepatitis.

OBJECTIVE: The serine protease inhibitor SerpinB3 has been described as critical mediator of liver fibrosis and it has been recently proposed as an additional hepatokine involved in NASH development and insulin resistance. Protease Activated Receptor 2 has been identified as a novel regulator of hepatic metabolism. A targeted therapeutic strategy for NASH has been investigated, using 1-Piperidine Propionic Acid (1-PPA), since this compound has been recently proposed as both Protease Activated Receptor 2 and SerpinB3 inhibitor. METHODS: The effect of SerpinB3 on inflammation and fibrosis genes was assessed in human macrophage and stellate cell lines. Transgenic mice, either overexpressing SerpinB3 or carrying Serpinb3 deletion and their relative wild type strains, were used in experimental NASH models. Subgroups of SerpinB3 transgenic mice and their controls were also injected with 1-PPA to assess the efficacy of this compound in NASH inhibition. RESULTS: 1-PPA did not present significant cell and organ toxicity and was able to inhibit SerpinB3 and PAR2 in a dose-dependent manner. This effect was associated to a parallel reduction of the synthesis of the molecules induced by endogenous SerpinB3 or by its paracrine effects both in vitro and in vivo, leading to inhibition of lipid accumulation, inflammation and fibrosis in experimental NASH. At mechanistic level, the antiprotease activity of SerpinB3 was found essential for PAR2 activation, determining upregulation of the CCAAT Enhancer Binding Protein beta (C/EBP-ß), another pivotal regulator of metabolism, inflammation and fibrosis, which in turn determined SerpinB3 synthesis. CONCLUSIONS: 1-PPA treatment was able to inhibit the PAR2 - C/EBP-ß - SerpinB3 axis and to protect from NASH development and progression, supporting the potential use of a similar approach for a targeted therapy of NASH.

The polymorphic variant of SerpinB3 (SerpinB3-PD) is associated with faster cirrhosis decompensation.

BACKGROUND: SerpinB3 is a cysteine protease inhibitor involved in liver disease progression due to its proinflammatory and profibrogenic properties. The polymorphic variant SerpinB3-PD (SB3-PD), presents a substitution in its reactive centre loop, determining the gain of function. AIMS: To disclose the clinical characteristics of a cohort of patients with cirrhosis in relation to the presence of SB3-PD and to assess the effect of this genetic variant on fibrogenic and inflammatory cytokines in vitro. METHODS: We assessed SB3 polymorphism in 90 patients with cirrhosis, prospectively followed up in our referral centre. We used HepG2 and HuH-7 cells transfected to overexpress either wild-type SB3 (SB3-WT) or SB3-PD to assess their endogenous effect, while LX2 and THP-1 cells were treated with exogenous SB3-WT or SB3-PD proteins. RESULTS: Patients carrying SB3-PD had more severe portal hypertension and higher MELD scores, than patients carrying SB3-WT. In multivariate analysis, SB3-PD was an independent predictor of cirrhosis complications. Patients with SB3-PD polymorphism presented with more severe liver fibrosis and inflammatory features. Hepatoma cells overexpressing SB3-PD showed higher TGF-ß1 expression than controls. The addition of recombinant SB3-PD induced an up-regulation of TGF-ß1 in LX2 cells and a more prominent inflammatory profile in THP-1 cells, compared to the effect of SB3-WT protein. CONCLUSIONS: The polymorphic variant SB3-PD is highly effective in determining activation of TGF-ß1 and inflammation in vitro. Patients with cirrhosis who carry SB3-PD polymorphism may be more prone to develop severe liver disease progression. However, further validation studies are warranted to support the in vivo relevance of this polymorphism.

Hepatocellular Carcinoma: Latest Research in Pathogenesis, Detection and Treatment.

The most common form of primary liver malignancy is hepatocellular carcinoma (HCC) [...].

Chronic Liver Disease: Latest Research in Pathogenesis, Detection and Treatment.

Chronic liver disease (CLD) is a major global health threat and has emerged as a leading cause of human death [...].

SerpinB3 Upregulates Low-Density Lipoprotein Receptor-Related Protein (LRP) Family Members, Leading to Wnt Signaling Activation and Increased Cell Survival and Invasiveness.

Abnormal activation of the Wnt-ß-catenin signaling cascade is involved in tumor growth and dissemination. SerpinB3 has been shown to induce ß-catenin, and both molecules are overexpressed in tumors, particularly in those with poor prognoses. The aim of this study was to evaluate the ability of SerpinB3 to modulate the Wnt pathway in liver cancer and in monocytic cells, the main type of inflammatory cells in the tumor microenvironment. The Wnt cascade, Wnt co-receptors, and low-density lipoprotein receptor-related protein (LRP) members were analyzed in different cell lines and human monocytes in the presence or absence of SerpinB3. The Wnt-ß-catenin axis was also evaluated in liver tumors induced in mice with different extents of SeprinB3 expression. In monocytic cells, SerpinB3 induced a significant upregulation of Wnt-1/7, nuclear ß-catenin, and c-Myc, which are associated with increased cell lifespan and proliferation. In liver tumors in mice, the expression of ß-catenin was significantly correlated with the presence of SerpinB3. In hepatoma cells, Wnt co-receptors LRP-5/6 and LRP-1, implicated in cell survival and invasiveness, were upregulated by SerpinB3. The LRP pan-inhibitor RAP not only induced a decrease in LRP expression, but also a dose-dependent reduction in SerpinB3-induced invasiveness. In conclusion, SerpinB3 determines the activation of the Wnt canonical pathway and cell invasiveness through the upregulation of LRP family members.

Metabolic Reprogramming of HCC: A New Microenvironment for Immune Responses.

Hepatocellular carcinoma is the most common primary liver cancer, ranking third among the leading causes of cancer-related mortality worldwide and whose incidence varies according to geographical area and ethnicity. Metabolic rewiring was recently introduced as an emerging hallmark able to affect tumor progression by modulating cancer cell behavior and immune responses. This review focuses on the recent studies examining HCC's metabolic traits, with particular reference to the alterations of glucose, fatty acid and amino acid metabolism, the three major metabolic changes that have gained attention in the field of HCC. After delivering a panoramic picture of the peculiar immune landscape of HCC, this review will also discuss how the metabolic reprogramming of liver cancer cells can affect, directly or indirectly, the microenvironment and the function of the different immune cell populations, eventually favoring the tumor escape from immunosurveillance.

Inflammatory processes involved in NASH-related hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the fourth leading cause of cancer-related death worldwide. In the recent years nonalcoholic fatty liver disease (NAFLD) is becoming a growing cause of HCCs and the incidence of NAFLD-related HCCs is expected to further dramatically increase by the next decade. Chronic inflammation is regarded as the driving force of NAFLD progression and a key factor in hepatic carcinogenesis. Hepatic inflammation in NAFLD results from the persistent stimulation of innate immunity in response to hepatocellular injury and gut dysbiosis as well as by the activation of adaptive immunity. However, the relative roles of innate and adaptive immunity in the processes leading to HCC are still incompletely characterized. This is due to the complex interplay between different liver cell populations, which is also strongly influenced by gut-derived bacterial products, metabolic/nutritional signals. Furthermore, carcinogenic mechanisms in NAFLD/NASH appear to involve the activation of signals mediated by hypoxia inducible factors. This review discusses recent data regarding the contribution of different inflammatory cells to NAFLD-related HCC and their possible impact on patient response to current treatments.

Oncostatin M: From Intracellular Signaling to Therapeutic Targets in Liver Cancer.

Primary liver cancers represent the third-most-common cause of cancer-related mortality worldwide, with an incidence of 80-90% for hepatocellular carcinoma (HCC) and 10-15% for cholangiocarcinoma (CCA), and an increasing morbidity and mortality rate. Although HCC and CCA originate from independent cell populations (hepatocytes and biliary epithelial cells, respectively), they develop in chronically inflamed livers. Evidence obtained in the last decade has revealed a role for cytokines of the IL-6 family in the development of primary liver cancers. These cytokines operate through the receptor subunit gp130 and the downstream Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathways. Oncostatin M (OSM), a member of the IL-6 family, plays a significant role in inflammation, autoimmunity, and cancer, including liver tumors. Although, in recent years, therapeutic approaches for the treatment of HCC and CCA have been implemented, limited treatment options with marginal clinical benefits are available. We discuss how OSM-related pathways can be selectively inhibited and therapeutically exploited for the treatment of liver malignancies.

Hepatic Myofibroblasts: A Heterogeneous and Redox-Modulated Cell Population in Liver Fibrogenesis.

During chronic liver disease (CLD) progression, hepatic myofibroblasts (MFs) represent a unique cellular phenotype that plays a critical role in driving liver fibrogenesis and then fibrosis. Although they could originate from different cell types, MFs exhibit a rather common pattern of pro-fibrogenic phenotypic responses, which are mostly elicited or sustained both by oxidative stress and reactive oxygen species (ROS) and several mediators (including growth factors, cytokines, chemokines, and others) that often operate through the up-regulation of the intracellular generation of ROS. In the present review, we will offer an overview of the role of MFs in the fibrogenic progression of CLD from different etiologies by focusing our attention on the direct or indirect role of ROS and, more generally, oxidative stress in regulating MF-related phenotypic responses. Moreover, this review has the purpose of illustrating the real complexity of the ROS modulation during CLD progression. The reader will have to keep in mind that a number of issues are able to affect the behavior of the cells involved: a) the different concentrations of reactive species, b) the intrinsic state of the target cells, as well as c) the presence of different growth factors, cytokines, and other mediators in the extracellular microenvironment or of other cellular sources of ROS.

SerpinB3 as a Pro-Inflammatory Mediator in the Progression of Experimental Non-Alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) is becoming the most common chronic liver disease worldwide. In 20-30% of patients, NAFLD can progress into non-alcoholic steatohepatitis (NASH), eventually leading to fibrosis, cirrhosis and hepatocellular carcinoma development. SerpinB3 (SB3), a hypoxia-inducible factor-2α dependent cysteine protease inhibitor, is up-regulated in hepatocytes during progressive NAFLD and proposed to contribute to disease progression. In this study we investigated the pro-inflammatory role of SB3 by employing phorbol-myristate acetate-differentiated human THP-1 macrophages exposed in vitro to human recombinant SB3 (hrSB3) along with mice overexpressing SB3 in hepatocytes (TG/SB3) or knockout for SB3 (KO/SB3) in which NASH was induced by feeding methionine/choline deficient (MCD) or a choline-deficient, L-amino acid defined (CDAA) diets. In vivo experiments showed that the induction of NASH in TG/SB3 mice was characterized by an impressive increase of liver infiltrating macrophages that formed crown-like aggregates and by an up-regulation of hepatic transcript levels of pro-inflammatory cytokines. All these parameters and the extent of liver damage were significantly blunted in KO/SB3 mice. In vitro experiments confirmed that hrSB3 stimulated macrophage production of M1-cytokines such as TNFα and IL-1ß and reactive oxygen species along with that of TGFß and VEGF through the activation of the NF-kB transcription factor. The opposite changes in liver macrophage activation observed in TG/SB3 or KO/SB3 mice with NASH were associated with a parallel modulation in the expression of triggering receptor expressed on myeloid cells-2 (TREM2), CD9 and galectin-3 markers, recently detected in NASH-associated macrophages. From these results we propose that SB3, produced by activated/injured hepatocytes, may operate as a pro-inflammatory mediator in NASH contributing to the disease progression.

G protein-coupled receptor 21 in macrophages: An in vitro study.

GPR21 is an orphan and constitutively active receptor belonging to the superfamily of G-Protein Coupled Receptors (GPCRs). GPR21 couples to the Gq family of G proteins and is expressed in macrophages. Studies of GPR21 knock-out mice indicated that GPR21 may be involved in promoting macrophage migration. The aim of this study was to evaluate the role of GPR21 in human macrophages, analyzing (i) its involvement in cell migration and cytokine release and (ii) the consequence of its pharmacological inhibition by using the inverse agonist GRA2. THP-1 cells were activated and differentiated into either M1 or M2 macrophages. GPR21 expression was evaluated at gene and protein level, the signalling pathway was investigated by an IP1 assay, and cytokine release by ELISA. Cell migration was detected by the Boyden chamber migration assay, performed on macrophages derived from both the THP-1 cell line and human peripheral blood monocytes. In addition, we compared the effect of the pharmacological inhibition of GPR21 with the effect of the treatment with a specific GPR21 siRNA to downregulate the receptor expression, thus confirming that GRA2 acts as an inverse agonist of GPR21. GRA2 does not affect cell viability at the tested concentrations, but significantly reduces the release of TNF-α and IL-1ß from M1 macrophages. The analysis of the migratory ability highlighted opposite effects of GRA2 on M1 and M2 macrophages since it decreased M1, while it promoted M2 cell migration. Therefore, the pharmacological inhibition of GPR21 could be of interest for pathological conditions characterized by low grade chronic inflammation.

Hyaluronated and PEGylated Liposomes as a Potential Drug-Delivery Strategy to Specifically Target Liver Cancer and Inflammatory Cells.

Hepatocellular carcinoma (HCC) is the most frequent primary liver cancer and is characterized by poor clinical outcomes, with the majority of patients not being eligible for curative therapy and treatments only being applicable for early-stage tumors. CD44 is a receptor for hyaluronic acid (HA) and is involved in HCC progression. The aim of this work is to propose HA- and PEGylated-liposomes as promising approaches for the treatment of HCC. It has been found, in this work, that CD44 transcripts are up-regulated in HCC patients, as well as in a murine model of NAFLD/NASH-related hepatocarcinogenesis. Cell culture experiments indicate that HA-liposomes are more rapidly and significantly internalized by Huh7 cells that over-express CD44, compared with HepG2 cells that express low levels of the receptor, in which the uptake seems due to endocytic events. By contrast, human and murine macrophage cell lines (THP-1, RAW264.7) show improved and rapid uptake of PEG-modified liposomes without the involvement of the CD44. Moreover, the internalization of PEG-modified liposomes seems to induce polarization of THP1 towards the M1 phenotype. In conclusion, data reported in this study indicate that this strategy can be proposed as an alternative for drug delivery and one that dually and specifically targets liver cancer cells and infiltrating tumor macrophages in order to counteract two crucial aspect of HCC progression.

Oncostatin M is overexpressed in NASH-related hepatocellular carcinoma and promotes cancer cell invasiveness and angiogenesis.

Oncostatin M (OSM) is a pleiotropic cytokine of the interleukin (IL)-6 family that contributes to the progression of chronic liver disease. Here we investigated the role of OSM in the development and progression of hepatocellular carcinoma (HCC) in non-alcoholic fatty liver disease (NAFLD)/non-alcoholic steatohepatitis (NASH). The role of OSM was investigated in (1) selected cohorts of NAFLD/NASH HCC patients, (2) liver cancer cells exposed to human recombinant OSM or stably transfected to overexpress human OSM, (3) murine HCC xenografts, and (4) a murine NASH-related model of hepatic carcinogenesis. OSM was found to be selectively overexpressed in HCC cells of NAFLD/NASH patients, depending on tumor grade. OSM serum levels, barely detectable in patients with simple steatosis or NASH, were increased in patients with cirrhosis and more evident in those carrying HCC. In this latter group, OSM serum levels were significantly higher in the subjects with intermediate/advanced HCCs and correlated with poor survival. Cell culture experiments indicated that OSM upregulation in hepatic cancer cells contributes to HCC progression by inducing epithelial-to-mesenchymal transition and increased invasiveness of cancer cells as well as by inducing angiogenesis, which is of critical relevance. In murine xenografts, OSM overexpression was associated with slower tumor growth but an increased rate of lung metastases. Overexpression of OSM and its positive correlation with the angiogenic switch were also confirmed in a murine model of NAFLD/NASH-related hepatocarcinogenesis. Consistent with this, analysis of liver specimens from human NASH-related HCCs with vascular invasion showed that OSM was expressed by liver cancer cells invading hepatic vessels. In conclusion, OSM upregulation appears to be a specific feature of HCC arising on a NAFLD/NASH background, and it correlates with clinical parameters and disease outcome. Our data highlight a novel pro-carcinogenic contribution for OSM in NAFLD/NASH, suggesting a role of this factor as a prognostic marker and a putative potential target for therapy. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Hepatocyte-Specific Deletion of HIF2α Prevents NASH-Related Liver Carcinogenesis by Decreasing Cancer Cell Proliferation.

BACKGROUND & AIMS: Hypoxia and hypoxia-inducible factors (HIFs) are involved in chronic liver disease progression. We previously showed that hepatocyte HIF-2α activation contributed significantly to nonalcoholic fatty liver disease progression in experimental animals and human patients. In this study, using an appropriate genetic murine model, we mechanistically investigated the involvement of hepatocyte HIF-2α in experimental nonalcoholic steatohepatitis (NASH)-related carcinogenesis. METHODS: The role of HIF-2α was investigated by morphologic, cellular, and molecular biology approaches in the following: (1) mice carrying hepatocyte-specific deletion of HIF-2α (HIF-2α-/- mice) undergoing a NASH-related protocol of hepatocarcinogenesis; (2) HepG2 cells stably transfected to overexpress HIF-2α; and (3) liver specimens from NASH patients with hepatocellular carcinoma. RESULTS: Mice carrying hepatocyte-specific deletion of HIF-2α (hHIF-2α-/-) showed a significant decrease in the volume and number of liver tumors compared with wild-type littermates. These effects did not involve HIF-1α changes and were associated with a decrease of cell proliferation markers proliferating cell nuclear antigen and Ki67. In both human and rodent nonalcoholic fatty liver disease-related tumors, HIF-2α levels were strictly associated with hepatocyte production of SerpinB3, a mediator previously shown to stimulate liver cancer cell proliferation through the Hippo/Yes-associated protein (YAP)/c-Myc pathway. Consistently, we observed positive correlations between the transcripts of HIF-2α, YAP, and c-Myc in individual hepatocellular carcinoma tumor masses, while HIF-2α deletion down-modulated c-Myc and YAP expression without affecting extracellular signal-regulated kinase 1/2, c-Jun N-terminal kinase, and AKT-dependent signaling. In vitro data confirmed that HIF-2α overexpression induced HepG2 cell proliferation through YAP-mediated mechanisms. CONCLUSIONS: These results indicate that the activation of HIF-2α in hepatocytes has a critical role in liver carcinogenesis during NASH progression, suggesting that HIF-2α-blocking agents may serve as novel putative therapeutic tools.

GPR21 Inhibition Increases Glucose-Uptake in HepG2 Cells.

GPR21 is a constitutively active, orphan, G-protein-coupled receptor, with in vivo studies suggesting its involvement in the modulation of insulin sensitivity. However, its precise contribution is not fully understood. As the liver is both a major target of insulin signalling and critically involved in glucose metabolism, the aim of this study was to examine the role of GPR21 in the regulation of glucose uptake and production in human hepatocytes. In particular, HepG2 cells, which express GPR21, were adopted as cellular models. Compared with untreated cells, a significant increase in glucose uptake was measured in cells treated with siRNA to downregulate GPR21 expression or with the GPR21-inverse agonist, GRA2. Consistently, a significantly higher membrane translocation of GLUT-2 was measured under these conditions. These effects were accompanied by an increased ratio of phAKT(Ser473)/tot-AKT and phGSK-3ß(Ser9)/tot-GSK-3ß, thus indicating a marked activation of the insulin signalling pathway. Moreover, a significant reduction in ERK activation was observed with GPR21 inhibition. Collectively, these results indicate that GPR21 mediates the negative effects on glucose uptake by the liver cells. In addition, they suggest that the pharmacological inhibition of GPR21 could be a novel strategy to improve glucose homeostasis and counteract hepatic insulin resistance.

Hypoxia, Hypoxia-Inducible Factors and Liver Fibrosis.

Liver fibrosis is a potentially reversible pathophysiological event, leading to excess deposition of extracellular matrix (ECM) components and taking place as the net result of liver fibrogenesis, a dynamic and highly integrated process occurring during chronic liver injury of any etiology. Liver fibrogenesis and fibrosis, together with chronic inflammatory response, are primarily involved in the progression of chronic liver diseases (CLD). As is well known, a major role in fibrogenesis and fibrosis is played by activated myofibroblasts (MFs), as well as by macrophages and other hepatic cell populations involved in CLD progression. In the present review, we will focus the attention on the emerging pathogenic role of hypoxia, hypoxia-inducible factors (HIFs) and related mediators in the fibrogenic progression of CLD.

Liver fibrogenesis: un update on established and emerging basic concepts.

Liver fibrogenesis is defined as a dynamic and highly integrated process occurring during chronic injury to liver parenchyma that can result in excess deposition of extracellular matrix (ECM) components (i.e., liver fibrosis). Liver fibrogenesis, together with chronic inflammatory response, is then primarily involved in the progression of chronic liver diseases (CLD) irrespective of the specific etiology. In the present review we will first offer a synthetic and updated overview of major basic concepts in relation to the role of myofibroblasts (MFs), macrophages and other hepatic cell populations involved in CLD to then offer an overview of established and emerging issues and mechanisms that have been proposed to favor and/or promote CLD progression. A special focus will be dedicated to selected issues that include emerging features in the field of cholangiopathies, the emerging role of genetic and epigenetic factors as well as of hypoxia, hypoxia-inducible factors (HIFs) and related mediators.

SerpinB3 Differently Up-Regulates Hypoxia Inducible Factors -1α and -2α in Hepatocellular Carcinoma: Mechanisms Revealing Novel Potential Therapeutic Targets.

BACKGROUND: SerpinB3 (SB3) is a hypoxia and hypoxia-inducible factor (HIF)-2α-dependent cysteine-protease inhibitor up-regulated in hepatocellular carcinoma (HCC), released by cancer cells and able to stimulate proliferation and epithelial-to-mesenchymal-transition. Methods: In the study we employed transgenic and knock out SerpinB3 mice, liver cancer cell line, human HCC specimens, and mice receiving diethyl-nitrosamine (DEN) administration plus choline-deficient L-amino acid refined (CDAA) diet (DEN/CDAA protocol). Results: We provide detailed and mechanistic evidence that SB3 can act as a paracrine mediator able to affect the behavior of surrounding cells by differentially up-regulating, in normoxic conditions, HIF-1α and HIF-2α. SB3 acts by (i) up-regulating HIF-1α transcription, facilitating cell survival in a harsh microenvironment and promoting angiogenesis, (ii) increasing HIF-2α stabilization via direct/selective NEDDylation, promoting proliferation of liver cancer cells, and favoring HCC progression. Moreover (iii) the highest levels of NEDD8-E1 activating enzyme (NAE1) mRNA were detected in a subclass of HCC patients expressing the highest levels of HIF-2α transcripts; (iv) mice undergoing DEN/CDAA carcinogenic protocol showed a positive correlation between SB3 and HIF-2α transcripts with the highest levels of NAE1 mRNA detected in nodules expressing the highest levels of HIF-2α transcripts. Conclusions: These data outline either HIF-2α and NEDDylation as two novel putative therapeutic targets to interfere with the procarcinogenic role of SerpinB3 in the development of HCC.

Oncostatin M, A Profibrogenic Mediator Overexpressed in Non-Alcoholic Fatty Liver Disease, Stimulates Migration of Hepatic Myofibroblasts.

BACKGROUND: Hepatic myofibroblasts (MFs) can originate from hepatic stellate cells, portal fibroblasts, or bone marrow-derived mesenchymal stem cells and can migrate towards the site of injury by aligning with nascent and established fibrotic septa in response to several mediators. Oncostatin M (OSM) is known to orchestrate hypoxia-modulated hepatic processes involving the hypoxia-inducible factor 1 (HIF-1). METHODS: In vivo and in vitro experiments were performed to analyze the expression of OSM and OSM-receptor (OSMR) in three murine models of non-alcoholic-fatty liver disease (NAFLD) and -steatohepatitis (NASH) and in human NASH patients as well as the action of OSM on phenotypic responses of human MFs. RESULTS: Hepatic OSM and OSMR levels were overexpressed in three murine NASH models and in NASH patients. OSM stimulates migration in human MFs by involving early intracellular ROS generation and activation of Ras/Erk, JNK1/2, PI3K/Akt as well as STAT1/STAT3 pathways and HIF-1α. OSM-dependent migration relies on a biphasic mechanism requiring early intracellular generation of reactive oxygen species (ROS) and late HIF1-dependent expression and release of VEGF. CONCLUSION: OSM is overexpressed in experimental and human progressive NAFLD and can act as a profibrogenic factor by directly stimulating migration of hepatic MFs.