Chronic hepatitis C virus (HCV) infection remains a major global health burden, responsible for substantial morbidity and mortality despite the advent of curative antiviral therapy. HCV induces hepatic injury and carcinogenesis through direct viral effects, persistent inflammation, oxidative stress, and metabolic disturbance. The introduction of direct-acting antivirals (DAAs) has revolutionized therapy, achieving sustained virologic response rates exceeding 95% and transforming HCV from a chronic, progressive disease into a curable infection. Nevertheless, viral eradication does not fully normalize hepatic or systemic risk. Patients with advanced fibrosis or cirrhosis continue to face an elevated incidence of hepatocellular carcinoma (HCC) and other complications, reinforcing the need for long-term monitoring. This review summarizes current knowledge of the molecular mechanisms underlying HCV-mediated carcinogenesis, the partial restoration of hepatic homeostasis following DAA-induced cure, and the clinical implications for surveillance and management in the post-HCV era. By integrating insights from molecular virology, immunopathogenesis, and clinical hepatology, the review highlights how persistent epigenetic and inflammatory footprints may sustain oncogenic potential even after viral clearance. A comprehensive understanding of these processes is essential for optimizing HCC prevention strategies, guiding surveillance policies, and advancing future therapeutic innovations aimed at complete hepatic recovery.

Hepatitis C virus (HCV) infection compromises the natural defense mechanisms of the liver leading to a progressive end stage disease such as cirrhosis and hepatocellular carcinoma (HCC). The hepatic stress response generated due to viral replication in the endoplasmic reticulum (ER) undergoes a stepwise transition from adaptive to pro-survival signaling to improve host cell survival and liver disease progression. The minute details of hepatic pro-survival unfolded protein response (UPR) signaling that contribute to HCC development in cirrhosis are unknown. This study shows that the UPR sensor, the protein kinase RNA-like ER kinase (PERK), mediates the pro-survival signaling through nuclear factor erythroid 2-related factor 2 (NRF2)-mediated signal transducer and activator of transcription 3 (STAT3) activation in a persistent HCV infection model of Huh-7.5 liver cells. The NRF2-mediated STAT3 activation in persistently infected HCV cell culture model resulted in the decreased expression of hepatocyte nuclear factor 4 alpha (HNF4A), a major liver-specific transcription factor. The stress-induced inhibition of HNF4A expression resulted in a significant reduction of liver-specific microRNA-122 (miR-122) transcription. It was found that the reversal of hepatic adaptive pro-survival signaling and restoration of miR-122 level was more efficient by interferon (IFN)-based antiviral treatment than direct-acting antivirals (DAAs). To test whether miR-122 levels could be utilized as a biomarker of hepatic adaptive stress response in HCV infection, serum miR-122 level was measured among healthy controls, and chronic HCV patients with or without cirrhosis. Our data show that serum miR-122 expression level remained undetectable in most of the patients with cirrhosis (stage IV fibrosis), suggesting that the pro-survival UPR signaling increases the risk of HCC through STAT3-mediated suppression of miR-122. In conclusion, our data indicate that hepatic pro-survival UPR signaling suppresses the liver-specific HNF4A and its downstream target miR-122 in cirrhosis. These results provide an explanation as to why cirrhosis is a risk factor for the development of HCC in chronic HCV infection.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 ajp.amjpathol.org 63 64 65 66 67 68 69 70 71 72 73 74 75 76 Cellular Mechanism for Impaired Hepatitis C Virus Clearance by Interferon Associated with IFNL3 77 78 79 80 81 82 83 84 Gene Polymorphisms Relates to Intrahepatic Interferon-l Expression Pauline Ferraris,* Partha K. Chandra,* Rajesh Panigrahi,* Fatma Aboulnasr,* Srinivas Chava,* Ramazan Kurt,y Jean-Michel Pawlotsky,z Ludwig Wilkens,x Pamela Osterlund,{ Rune Hartmann,k Luis A. Balart,y Tong Wu,* and Srikanta Dash*y 85 86 87 88 89 From the Departments of Pathology and Laboratory Medicine,* and Medicine, Gastroenterology, and Hepatology,y Tulane University Health Sciences Center, New Orleans, Louisiana; the Institut National de la Recherche Medicale,z Creteil, France; the Institute of Pathology,x University of Bern, Bern, Switzerland; the Department of Vaccination and Immune Protection Viral Infections,{ National Institute for Health and Welfare, Helsinki, Finland; and the Department of Molecular Biology and Genetics,k Aarhus University, Aarhus, Denmark 90 91 92 Accepted for publication

The single nucleotide polymorphism located within the IFNL3 (also known as IL28B) promoter is one of the host factors associated with hepatitis C virus (HCV) clearance by interferon (IFN)-alpha therapy; however the mechanism remains unknown. We investigated how IL28B gene polymorphism influences HCV clearance with infected primary human hepatocytes, liver biopsies, and hepatoma cell lines. Our study confirms that the rs12979860-T/T genotype has a strong correlation with ss469415590-Delta G/Delta G single nucleotide polymorphism that produces IFN-lambda 4 protein. We found that IFN-alpha and IFN-lambda 1 antiviral activity against HCV was impaired in IL28B T/T infected hepatocytes compared with C/C genotype. Western blot analysis showed that IL28B TT genotype hepatocytes expressed higher levels of IFN-lambda proteins (IL28B, IL-29), preactivated IFN-stimulated gene (ISG) expression, and impaired Stat phosphory when stimulated with either IFN-alpha or IFN-lambda 1. Furthermore, we showed that silencing IFN-lambda 1 in T/T cell line reduced basal ISG expression and improved antiviral activity. Likewise, overexpression of IFN-lambda (1 to 4) in C/C cells induced basal ISG expression and prevented IFN-alpha antiviral activity. We showed that IFN-lambda 4, produced at low level only in T/T cells induced expression of IL28B and IL-29 and prevented IFN-alpha antiviral activity in HCV cell culture. Our results suggest that IFN-lambda 4 protein expression associated with the IL28B-T/T variant preactivates the Janus kinase-Stat signaling, leading to impaired HCV clearance by both IFN-alpha and IFN-lambda.

Background Hepatic steatosis is a risk factor for both liver disease progression and an impaired response to interferon alpha (IFN-α)-based combination therapy in chronic hepatitis C virus (HCV) infection. Previously, we reported that free fatty acid (FFA)-treated HCV cell culture induces hepatocellular steatosis and impairs the expression of interferon alpha receptor-1 (IFNAR1), which is why the antiviral activity of IFN-α against HCV is impaired. Aim To investigate the molecular mechanism by which IFNAR1 expression is impaired in HCV cell culture with or without free fatty acid-treatment. Method HCV-infected Huh 7.5 cells were cultured with or without a mixture of saturated (palmitate) and unsaturated (oleate) long-chain free fatty acids (FFA). Intracytoplasmic fat accumulation in HCV-infected culture was visualized by oil red staining. Clearance of HCV in FFA cell culture treated with type I IFN (IFN-α) and Type III IFN (IFN-λ) was determined by Renilla luciferase activity, and the expression of HCV core was determined by immunostaining. Activation of Jak-Stat signaling in the FFA-treated HCV culture by IFN-α alone and IFN-λ alone was examined by Western blot analysis and confocal microscopy. Lysosomal degradation of IFNAR1 by chaperone-mediated autophagy (CMA) in the FFA-treated HCV cell culture model was investigated. Results FFA treatment induced dose-dependent hepatocellular steatosis and lipid droplet accumulation in HCV-infected Huh-7.5 cells. FFA treatment of infected culture increased HCV replication in a concentration-dependent manner. Intracellular lipid accumulation led to reduced Stat phosphorylation and nuclear translocation, causing an impaired IFN-α antiviral response and HCV clearance. Type III IFN (IFN-λ), which binds to a separate receptor, induces Stat phosphorylation, and nuclear translocation as well as antiviral clearance in FFA-treated HCV cell culture. We show here that the HCV-induced autophagy response is increased in FFA-treated cell culture. Pharmacological inhibitors of lysosomal degradation, such as ammonium chloride and bafilomycin, prevented IFNAR1 degradation in FFA-treated HCV cell culture. Activators of chaperone-mediated autophagy, including 6-aminonicotinamide and nutrient starvation, decreased IFNAR1 levels in Huh-7.5 cells. Co-immunoprecipitation, colocalization and siRNA knockdown experiments revealed that IFNAR1 but not IFNLR1 interacts with HSC70 and LAMP2A, which are core components of chaperone-mediated autophagy (CMA). Conclusion Our study presents evidence indicating that chaperone-mediated autophagy targets IFNAR1 degradation in the lysosome in FFA-treated HCV cell culture. These results provide a mechanism for why HCV induced autophagy response selectively degrades type I but not the type III IFNAR1.

ABSTRACT Ribavirin (RBV) continues to be an important component of interferon-free hepatitis C treatment regimens, as RBV alone does not inhibit hepatitis C virus (HCV) replication effectively; the reason for this ineffectiveness has not been established. In this study, we investigated the RBV resistance mechanism using a persistently HCV-infected cell culture system. The antiviral activity of RBV against HCV was progressively impaired in the persistently infected culture, whereas interferon lambda 1 (IFN-λ1), a type III IFN, showed a strong antiviral response and induced viral clearance. We found that HCV replication in persistently infected cultures induces an autophagy response that impairs RBV uptake by preventing the expression of equilibrative nucleoside transporter 1 (ENT1). The Huh-7.5 cell line treated with an autophagy inducer, Torin 1, downregulated membrane expression of ENT1 and terminated RBV uptake. In contrast, the autophagy inhibitors hydroxychloroquine (HCQ), 3-methyladenine (3-MA), and bafilomycin A1 (BafA1) prevented ENT1 degradation and enhanced RBV antiviral activity. The HCV-induced autophagy response, as well as treatment with Torin 1, degrades clathrin heavy chain expression in a hepatoma cell line. Reduced expression of the clathrin heavy chain by HCV prevents ENT1 recycling to the plasma membrane and forces ENT1 to the lysosome for degradation. This study provides a potential mechanism for the impairment of RBV antiviral activity in persistently HCV-infected cell cultures and suggests that inhibition of the HCV-induced autophagy response could be used as a strategy for improving RBV antiviral activity against HCV infection. IMPORTANCE The results from this work will allow a review of the competing theories of antiviral therapy development in the field of HCV virology. Ribavirin (RBV) remains an important component of interferon-free hepatitis C treatment regimens. The reason why RBV alone does not inhibit HCV replication effectively has not been established. This study provides a potential mechanism for why RBV antiviral activity is impaired in persistently HCV-infected cell cultures and suggests that inhibition of the HCV-induced autophagy response could be used as a strategy to increase RBV antiviral activity against HCV infection. Therefore, it is anticipated that this work would generate a great deal of interest, not only among virologists but also among the general public.

BACKGROUND:HCV replication in persistently infected cell culture remains resistant to IFN-α/RBV combination treatment, whereas IFN-λ1 induces viral clearance. The antiviral mechanisms by which IFN-λ1 induces sustained HCV clearance have not been determined.AIM:To investigate the mechanisms by which IFN-λ clears HCV replication in an HCV cell culture model.METHODS:IFN-α sensitive (S3-GFP) and resistant (R4-GFP) cells were treated with equivalent concentrations of either IFN-α or IFN-λ. The relative antiviral effects of IFN-α and IFN-λ1 were compared by measuring the HCV replication, quantification of HCV-GFP expression by flow cytometry, and viral RNA levels by real time RT-PCR. Activation of Jak-Stat signaling, interferon stimulated gene (ISG) expression, and miRNA-122 transcription in S3-GFP and R4-GFP cells were examined.RESULTS:We have shown that IFN-λ1 induces HCV clearance in IFN-α resistant and sensitive replicon cell lines in a dose dependent manner through Jak-Stat signaling, and induces STAT 1 and STAT 2 activation, ISRE-luciferase promoter activation and ISG expression. Stat 3 activation is also involved in IFN-λ1 induced antiviral activity in HCV cell culture. IFN-λ1 induced Stat 3 phosphorylation reduces the expression of hepatocyte nuclear factor 4 alpha (HNF4α) through miR-24 in R4-GFP cells. Reduced expression of HNF4α is associated with decreased expression of miR-122 resulting in an anti-HCV effect. Northern blot analysis confirms that IFN-λ1 reduces miR-122 levels in R4-GFP cells. Our results indicate that IFN-λ1 activates the Stat 3-HNF4α feedback inflammatory loop to inhibit miR-122 transcription in HCV cell culture.CONCLUSIONS:In addition to the classical Jak-Stat antiviral signaling pathway, IFN-λ1 inhibits HCV replication through the suppression of miRNA-122 transcription via an inflammatory Stat 3-HNF4α feedback loop. Inflammatory feedback circuits activated by IFNs during chronic inflammation expose non-responders to the risk of hepatocellular carcinoma.

INTRODUCTION:Myocardial infarction (MI) as a life-threatening event, carrying high risk of recurrence and chronic disabling complications, increases the risk of developing acute stress disorder (ASD), posttraumatic stress disorder (PTSD), or both. The aim of this study was to investigate the relationship between illness perceptions and having ASD, PTSD, or both in patients after MI.METHOD:Seventy-six patients diagnosed with acute MI were enrolled into our prospective study. We evaluated patients during the first week and six months after MI. Patients were assessed by using the Clinician Administered PTSD Scale (CAPS), the Hamilton Depression Rating Scale (HDRS), the Hamilton Anxiety Rating Scale (HARS), the Brief Illness Perception Questionnaire (BIPQ), and a semi-structured interview for socio-demographic characteristics during both the first and second evaluations.RESULTS:Acute stress disorder (ASD) developed in 9.2% of patients and PTSD developed in 11.9% of patients with MI. Illness perception factors of 'consequences, identity and concern' predicted the occurrence of both ASD and PTSD, whereas 'emotion' predicted only PTSD.CONCLUSION:The factors of illness perceptions predicted the induction of ASD and PTSD in patients who had acute MI.

Aims and background Nonalcoholic fatty liver disease (NAFLD) and hepatocellular carcinoma (HCC) are two major causes of liver disease worldwide. Epidemiological and clinical data have clearly demonstrated that NAFLD and its associated metabolic abnormalities are a risk factor for HCC. Traditionally, the mechanisms whereby NAFLD acts as a risk for HCC are believed to include replicative senescence of steatotic hepatocytes and compensatory hyperplasia of progenitor cells as a reaction to chronic hepatic injury. Recent years have witnessed significant advances in our understanding of the mechanisms underlying the link between NAFLD and HCC. Methods In the present review, we provide an update on the pathophysiological pathways linking NAFLD and its associated metabolic derangements to malignant hepatic transformation, with a special focus on insulin resistance, adipokines, inflammation, and angiogenesis. We will also discuss the potential therapeutic implications that such molecular links carry. Results Although treating NAFLD could reduce the risk of malignant hepatic transformation, no long-term studies focusing on this issue have been conducted thus far. Insulin resistance, inflammation as well as derangements in adipokines and angiogenic factors associated with NAFLD are closely intertwined with the risk of developing HCC. Conclusions Traditional therapeutic approaches in NAFLD including metformin and statins may theoretically reduce the risk of HCC by acting on common pathophysiological pathways shared by NAFLD and HCC.

Hepatitis B virus (HBV) infections are a major public health problem. It is estimated that there are more than 350 million HBV carriers in the world, of whom one million die annually from HBV-related liver disease [1]. Chronic hepatitis B may cause cirrhosis, hepatic decompensation, and hepatocellular carcinoma (HCC) resulting in significant morbidity and mortality. For this reason the main goals of chronic hepatitis B treatment are a sustained reduction of HBV DNA, biochemical remission, and histological improvement in order to prevent disease progression and long-term complications such as HCC [2]. Tenofovir disoproxil fumarate (tenofovir DF), the oral pro-drug of tenofovir, is a nucleotide reverse transcriptase inhibitor that shows potent in vitro activity against both HBV and immumodeficiency virus (HIV-1) [3]. Tenofovir DF was recently approved by the United States Food and Drug Administration (FDA) for the treatment of chronic HBV infections in adults. Tenofovir has a high genetic barrier and potent viral suppressive effect [4]. Its viral suppression ability in the first weeks after the initial treatment may be important in conditions where urgent viral reduction is necessary. The present study investigated the viral suppression rate sequentially after starting tenofovir treatment. The factors affecting the rapid viral response are also evaluated. This was a prospective trial that recruited patients from the Department of Gastroenterology, School of Medicine, Marmara University between 2008 and 2009. Sixteen patients (Male (n=10), Female (n=6), mean age: 46.9 ± 12.5) with chronic hepatitis B were enrolled in the study and treated with tenofovir (Table I). The median modified histological activity index (HAI) score for the initial liver biopsy was 7.166±5.166. The basal fibrosis stage of the patients was 0 (n=7), 1 (n=7) and 3 (n=2). Is tenofovir a drug of choice for patients needing a rapid viral response?

PurposeAlthough chronic hepatitis C virus (HCV) infection has been treated with the combination of interferon alpha (IFN-α) and ribavirin (RBV) for over a decade, the mechanism of antiviral synergy is not well understood. We aimed to determine the synergistic antiviral mechanisms of IFN-α and RBV combination treatment using HCV cell culture. MethodsThe antiviral efficacy of IFN-α, RBV alone and in combination was quantitatively measured using HCV infected and replicon cell culture. Direct antiviral activity of these two drugs at the level of HCV internal ribosome entry site (IRES) mediated translation in Huh-7 cell culture was investigated. The synergistic antiviral effect of IFN-α and RBV combination treatment was verified using both the CalcuSyn Software and MacSynergy Software. ResultsRBV combination with IFN-α efficiently inhibits HCV replication cell culture. Our results demonstrate that IFN-α, interferon lambda (IFN-λ) and RBV each inhibit the expression of HCV IRES-GFP and that they have a minimal effect on the expression of GFP in which the translation is not IRES dependent. The combination treatments of RBV along with IFN-α or IFN-λ were highly synergistic with combination indexes <1. We show that IFN-α treatment induce levels of PKR and eIF2α phosphorylation that prevented ribosome loading of the HCV IRES-GFP mRNA. Silencing of PKR expression in Huh-7 cells prevented the inhibitory effect of IFN-α on HCV IRES-GFP expression. RBV also blocked polyribosome loading of HCV-IRES mRNA through the inhibition of cellular IMPDH activity, and induced PKR and eIF2α phosphorylation. Knockdown of PKR or IMPDH prevented RBV induced HCV IRES-GFP translation. ConclusionsWe demonstrated both IFN-α and RBV inhibit HCV IRES through prevention of polyribosome formation. The combination of IFN-α and RBV treatment synergistically inhibits HCV IRES translation via using two different mechanisms involving PKR activation and depletion of intracellular guanosine pool through inhibition of IMPDH.

Low-grade fibromyxoid sarcoma (LGFMS) is a bland spindle cell neoplasm that typically arises in the deep soft tissues of the proximal extremities or trunk of young adults. The majority of LGFMS are characterized by a recurrent (7;16)(q34;p11) translocation, resulting in the FUS-CREB3L2 fusion gene, which generates a chimeric protein with transcriptional regulatory activity. Small numbers harbor a FUS-CREB3L1 fusion resulting from t(11;16)(p11;p11), whilst rare cases harbor the EWSR1-CREB3L1 fusion. LGFMS is of low to moderate cellularity and consists of bland spindle cells with small, angulated nuclei and scant, wispy cytoplasm, arranged in a whorled growth pattern and typically showing abrupt transition from myxoid to fibrous areas. Immunohistochemical expression of MUC4 is a consistent finding. Hyalinized spindle cell tumor with giant rosettes (HSCTGR) is a morphological variant of LGFMS that shares the same balanced translocation, and is also immunoreactive for MUC4. A potential relationship between LGFMS and sclerosing epithelioid fibrosarcoma (SEF), a rare fibroblastic neoplasm that most commonly arises in the deep soft tissues of the lower extremities, limb girdles or trunk, has also been suggested. SEF is classically composed of nests and cords of epithelioid cells with clear or eosinophilic cytoplasm embedded within densely sclerotic stroma. In some cases, areas indistinguishable from LGFMS are present, and these have been shown to contain FUS-CREB3L2 fusion transcripts. The majority of pure SEF tumors harbor EWSR1 rearrangements, with EWSR1-CREB3L1 and more rarely EWSR1-CREB3L2 gene fusions more common than those involving FUS. MUC4 immunoreactivity is also seen in approximately 70% of SEF. Surgical resection of these tumors with clear margins is the treatment of choice. Correct diagnosis is important because of the significant potential for recurrence and late metastatic spread. We review LGFMS and SEF, discussing morphology and immunohistochemistry, genetics and molecular findings, and the differential diagnosis.

Objectives: We sought to determine whether serum concentrations of fibroblast growth factor 19 (FGF19) - an ileum-derived enterokine which plays a role in the control of glucose and lipid homeostasis - are altered in patients with biopsy-proven nonalcoholic fatty liver disease (NAFLD).Design and methods: Serum levels of FGF19 were measured using enzyme-linked immunosorbent assay in 91 patients with biopsy-proven NAFLD and 74 controls.Results: FGF19 levels were significantly lower in patients with biopsy-proven NAFLD (median: 130 pg/mL) than in controls (median: 210 pg/mL, P<0.001). Serum FGF19 levels were significantly but modestly associated with hepatocyte ballooning scores in univariate analysis (r = -0.25. P<0.05) but not after adjustment for potential confounders (beta = -0.18; t = 1.78, P = 0.08).Conclusions: This pilot study suggests that serum FGF19 levels are decreased in patients with NAFLD but are not independently associated with liver histology findings. (C) 2012 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.