RESUMO
In contrast to most other malignancies, hepatocellular carcinoma (HCC), which accounts for approximately 90% of primary liver cancers, arises almost exclusively in the setting of chronic inflammation. Irrespective of etiology, a typical sequence of chronic necroinflammation, compensatory liver regeneration, induction of liver fibrosis and subsequent cirrhosis often precedes hepatocarcinogenesis. The liver is a central immunomodulator that ensures organ and systemic protection while maintaining immunotolerance. Deregulation of this tightly controlled liver immunological network is a hallmark of chronic liver disease and HCC. Notably, immunotherapies have raised hope for the successful treatment of advanced HCC. Here we summarize the roles of specific immune cell subsets in chronic liver disease, with a focus on non-alcoholic steatohepatitis and HCC. We review new advances in immunotherapeutic approaches for the treatment of HCC and discuss the challenges posed by the immunotolerant hepatic environment and the dual roles of adaptive and innate immune cells in HCC.
Assuntos
Carcinogênese/imunologia , Carcinoma Hepatocelular/imunologia , Neoplasias Hepáticas/imunologia , Humanos , Inflamação/complicações , Inflamação/imunologiaRESUMO
Hepatitis B virus (HBV) persists by depositing a covalently closed circular DNA (cccDNA) in the nucleus of infected cells that cannot be targeted by available antivirals. Interferons can diminish HBV cccDNA via APOBEC3-mediated deamination. Here, we show that overexpression of APOBEC3A alone is not sufficient to reduce HBV cccDNA that requires additional treatment of cells with interferon indicating involvement of an interferon-stimulated gene (ISG) in cccDNA degradation. Transcriptome analyses identify ISG20 as the only type I and II interferon-induced, nuclear protein with annotated nuclease activity. ISG20 localizes to nucleoli of interferon-stimulated hepatocytes and is enriched on deoxyuridine-containing single-stranded DNA that mimics transcriptionally active, APOBEC3A-deaminated HBV DNA. ISG20 expression is detected in human livers in acute, self-limiting but not in chronic hepatitis B. ISG20 depletion mitigates the interferon-induced loss of cccDNA, and co-expression with APOBEC3A is sufficient to diminish cccDNA. In conclusion, non-cytolytic HBV cccDNA decline requires the concerted action of a deaminase and a nuclease. Our findings highlight that ISGs may cooperate in their antiviral activity that may be explored for therapeutic targeting.
Assuntos
DNA Circular , Vírus da Hepatite B , Antivirais/farmacologia , Citidina Desaminase , DNA Circular/genética , DNA Viral/genética , DNA Viral/farmacologia , Exorribonucleases , Vírus da Hepatite B/genética , Humanos , Interferons , Proteínas , Replicação ViralRESUMO
Hepatitis B virus (HBV) infection is a major health threat causing 880,000 deaths each year. Available therapies control viral replication but do not cure HBV, leaving patients at risk to develop hepatocellular carcinoma. Here, we show that HBV envelope proteins (HBs)-besides their integration into endosomal membranes-become embedded in the plasma membrane where they can be targeted by redirected T-cells. HBs was detected on the surface of HBV-infected cells, in livers of mice replicating HBV and in HBV-induced hepatocellular carcinoma. Staining with HBs-specific recombinant antibody MoMab recognising a conformational epitope indicated that membrane-associated HBs remains correctly folded in HBV-replicating cells in cell culture and in livers of HBV-transgenic mice in vivo. MoMab coated onto superparamagnetic iron oxide nanoparticles allowed to detect membrane-associated HBs after HBV infection by electron microscopy in distinct stretches of the hepatocyte plasma membrane. Last but not least, we demonstrate that HBs located on the cell surface allow therapeutic targeting of HBV-positive cells by T-cells either engrafted with a chimeric antigen receptor or redirected by bispecific, T-cell engager antibodies. TAKE AWAYS: HBs become translocated to the plasma membrane. Novel, recombinant antibody confirmed proper conformation of HBs on the membrane. HBs provide an interesting target by T-cell-based, potentially curative therapies.
Assuntos
Antígenos de Superfície da Hepatite B , Hepatite B , Animais , Membrana Celular , Hepatite B/terapia , Vírus da Hepatite B , Humanos , Camundongos , Proteínas do Envelope ViralRESUMO
BACKGROUND & AIMS: Hepatitis B virus (HBV) infection persists because the virus-specific immune response is dysfunctional. Therapeutic vaccines might be used to end immune tolerance to the virus in patients with chronic infection, but these have not been effective in patients so far. In patients with chronic HBV infection, high levels of virus antigens might prevent induction of HBV-specific immune responses. We investigated whether knocking down expression levels of HBV antigens in liver might increase the efficacy of HBV vaccines in mice. METHODS: We performed studies with male C57BL/6 mice that persistently replicate HBV (genotype D, serotype ayw)-either from a transgene or after infection with an adeno-associated virus that transferred an overlength HBV genome-and expressed HB surface antigen at levels relevant to patients. Small hairpin or small interfering (si)RNAs against the common 3'-end of all HBV transcripts were used to knock down antigen expression in mouse hepatocytes. siRNAs were chemically stabilized and conjugated to N-acetylgalactosamine to increase liver uptake. Control mice were given either entecavir or non-HBV-specific siRNAs and vaccine components. Eight to 12 weeks later, mice were immunized twice with a mixture of adjuvanted HBV S and core antigen, followed by a modified Vaccinia virus Ankara vector to induce HBV-specific B- and T-cell responses. Serum and liver samples were collected and analyzed for HBV-specific immune responses, liver damage, and viral parameters. RESULTS: In both models of HBV infection, mice that express hepatocyte-specific small hairpin RNAs or that were given subcutaneous injections of siRNAs had reduced levels of HBV antigens, HBV replication, and viremia (1-3 log10 reduction) compared to mice given control RNAs. Vaccination induced production of HBV-neutralizing antibodies and increased numbers and functionality of HBV-specific, CD8+ T cells in mice with low, but not in mice with high, levels of HBV antigen. Mice with initially high titers of HBV and knockdown of HBV antigen expression, but not mice with reduced viremia after administration of entecavir, developed polyfunctional, HBV-specific CD8+ T cells, and HBV was eliminated. CONCLUSIONS: In mice with high levels of HBV replication, knockdown of HBV antigen expression along with a therapeutic vaccination strategy, but not knockdown alone, increased numbers of effector T cells and eliminated the virus. These findings indicate that high titers of virus antigens reduce the efficacy of therapeutic vaccination. Anti-HBV siRNAs and therapeutic vaccines are each being tested in clinical trials-their combination might cure chronic HBV infection.
Assuntos
Antígenos de Superfície da Hepatite B/genética , Vacinas contra Hepatite B/imunologia , Vírus da Hepatite B/imunologia , Hepatite B Crônica/terapia , RNA Interferente Pequeno/administração & dosagem , Animais , Linfócitos B/imunologia , Portador Sadio/imunologia , Portador Sadio/virologia , Terapia Combinada/métodos , Modelos Animais de Doenças , Feminino , Técnicas de Silenciamento de Genes , Antígenos de Superfície da Hepatite B/imunologia , Vacinas contra Hepatite B/administração & dosagem , Vírus da Hepatite B/genética , Vírus da Hepatite B/isolamento & purificação , Hepatite B Crônica/diagnóstico , Hepatite B Crônica/imunologia , Hepatite B Crônica/virologia , Hepatócitos/virologia , Humanos , Imunização Secundária , Imunogenicidade da Vacina , Masculino , Camundongos , Linfócitos T Citotóxicos/imunologia , Replicação Viral/genética , Replicação Viral/imunologiaRESUMO
BACKGROUND: Chronic hepatitis B develops more frequently in countries with high prevalence of helminth infections. The crosstalk between these 2 major liver-residing pathogens, Schistosoma mansoni and hepatitis B virus (HBV), is barely understood. METHODS: We used state-of-the-art models for both acute and chronic HBV infection to study the pathogen-crosstalk during the different immune phases of schistosome infection. RESULTS: Although liver pathology caused by schistosome infection was not affected by either acute or chronic HBV infection, S mansoni infection influenced HBV infection outcomes in a phase-dependent manner. Interferon (IFN)-γ secreting, HBV- and schistosome-specific CD8 T cells acted in synergy to reduce HBV-induced pathology during the TH1 phase and chronic phase of schistosomiasis. Consequently, HBV was completely rescued in IFN-γ-deficient or in TH2 phase coinfected mice demonstrating the key role of this cytokine. It is interesting to note that secondary helminth infection on the basis of persistent (chronic) HBV infection increased HBV-specific T-cell frequency and resulted in suppression of virus replication but failed to fully restore T-cell function and eliminate HBV. CONCLUSIONS: Thus, schistosome-induced IFN-γ had a prominent antiviral effect that outcompeted immunosuppressive effects of TH2 cytokines, whereas HBV coinfection did not alter schistosome pathogenicity.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Hepatite B Crônica/complicações , Hepatite B Crônica/imunologia , Esquistossomose mansoni/complicações , Esquistossomose mansoni/imunologia , Animais , Citocinas/imunologia , Modelos Animais de Doenças , Feminino , Vírus da Hepatite B/fisiologia , Interferon gama/imunologia , Fígado/parasitologia , Fígado/patologia , Fígado/virologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Contagem de Ovos de Parasitas , Schistosoma mansoni , Células Th2/imunologia , Replicação ViralRESUMO
BACKGROUND & AIMS: Hepatic innate immune control of viral infections has largely been attributed to Kupffer cells, the liver-resident macrophages. However, hepatocytes, the parenchymal cells of the liver, also possess potent immunological functions in addition to their known metabolic functions. Owing to their abundance in the liver and known immunological functions, we aimed to investigate the direct antiviral mechanisms employed by hepatocytes. METHODS: Using lymphocytic choriomeningitis virus (LCMV) as a model of liver infection, we first assessed the role of myeloid cells by depletion prior to infection. We investigated the role of hepatocyte-intrinsic innate immune signaling by infecting mice lacking canonical NF-κB signaling (IkkßΔHep) specifically in hepatocytes. In addition, mice lacking hepatocyte-specific interferon-α/ß signaling-(IfnarΔHep), or interferon-α/ß signaling in myeloid cells-(IfnarΔMyel) were infected. RESULTS: Here, we demonstrate that LCMV activates NF-κB signaling in hepatocytes. LCMV-triggered NF-κB activation in hepatocytes did not depend on Kupffer cells or TNFR1 signaling but rather on Toll-like receptor signaling. LCMV-infected IkkßΔHep livers displayed strongly elevated viral titers due to LCMV accumulation within hepatocytes, reduced interferon-stimulated gene (ISG) expression, delayed intrahepatic immune cell influx and delayed intrahepatic LCMV-specific CD8+ T cell responses. Notably, viral clearance and ISG expression were also reduced in LCMV-infected primary hepatocytes lacking IKKß, demonstrating a hepatocyte-intrinsic effect. Similar to livers of IkkßΔHep mice, enhanced hepatocytic LCMV accumulation was observed in livers of IfnarΔHep mice, whereas IfnarΔMyel mice were able to control LCMV infection. Hepatocytic NF-κB signaling was also required for efficient ISG induction in HDV-infected dHepaRG cells and interferon-α/ß-mediated inhibition of HBV replication in vitro. CONCLUSIONS: Together, these data show that hepatocyte-intrinsic NF-κB is a vital amplifier of interferon-α/ß signaling, which is pivotal for strong early ISG responses, immune cell infiltration and hepatic viral clearance. LAY SUMMARY: Innate immune cells have been ascribed a primary role in controlling viral clearance upon hepatic infections. We identified a novel dual role for NF-κB signaling in infected hepatocytes which was crucial for maximizing interferon responses and initiating adaptive immunity, thereby efficiently controlling hepatic virus replication.
Assuntos
Hepacivirus/genética , Hepatite C Crônica/genética , Hepatite C Crônica/imunologia , Hepatócitos/imunologia , Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/fisiologia , Subunidade p50 de NF-kappa B/genética , Polimorfismo de Nucleotídeo Único , Fator de Transcrição RelA/metabolismo , Replicação Viral/genética , Adulto , Animais , Células Cultivadas , Modelos Animais de Doenças , Feminino , Técnicas de Inativação de Genes , Genótipo , Hepatite C Crônica/virologia , Humanos , Quinase I-kappa B/deficiência , Quinase I-kappa B/genética , Coriomeningite Linfocítica/virologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Transdução de Sinais , Adulto JovemRESUMO
Recent studies have shown that tissue macrophages (MΦ) arise from embryonic progenitors of the yolk sac (YS) and fetal liver and colonize tissues before birth. Further studies have proposed that developmentally distinct tissue MΦ can be identified based on the differential expression of F4/80 and CD11b, but whether a characteristic transcriptional profile exists is largely unknown. Here, we took advantage of an inducible fate-mapping system that facilitated the identification of CD45(+)c-kit(-)CX3CR1(+)F4/80(+) (A2) progenitors of the YS as the source of F4/80(hi) but not CD11b(hi) MΦ. Large-scale transcriptional profiling of MΦ precursors from the YS stage to adulthood allowed for building computational models for F4/80(hi) tissue macrophages being direct descendants of A2 progenitors. We further identified a distinct molecular signature of F4/80(hi) and CD11b(hi) MΦ and found that Irf8 was vital for MΦ maturation. Our data provide new cellular and molecular insights into the origin and developmental pathways of tissue MΦ.
Assuntos
Diferenciação Celular , Perfilação da Expressão Gênica , Fatores Reguladores de Interferon/metabolismo , Macrófagos/fisiologia , Células-Tronco/fisiologia , Saco Vitelino/citologia , Animais , Simulação por Computador , Feminino , Imunofenotipagem , Masculino , CamundongosRESUMO
Data on the prevalence of resistance-associated substitutions (RASs) and their implications for treatment with direct-acting antivirals (DAAs) are sparse in European patients with HCV genotype 4. This study investigated RASs before and after DAA failure in different genotype 4 subtypes and evaluated retreatment efficacies. Samples of 195 genotype 4-infected patients were collected in the European Resistance Database and investigated for NS3, NS5A and NS5B RASs. Retreatment efficacies in DAA failure patients were analysed retrospectively. After NS5A inhibitor (NS5Ai) failure, subtype 4r was frequent (30%) compared to DAA-naïve patients (5%) and the number of NS5A RASs was significantly higher in subtype 4r compared to 4a or 4d (median three RASs vs no or one RAS, respectively, P < .0001). RASsL28V, L30R and M31L pre-existed in subtype 4r and were maintained after NS5Ai failure. Typical subtype 4r RASs were located in subdomain 1a of NS5A, close to membrane interaction and protein-protein interaction sites that are responsible for multimerization and hence viral replication. Retreatment of 37 DAA failure patients was highly effective with 100% SVR in prior SOF/RBV, PI/SOF and PI/NS5Ai failures. Secondary virologic failures were rare (n = 2; subtype 4d and 4r) and only observed in prior NS5Ai/SOF failures (SVR 90%). In conclusion, subtype 4r harboured considerably more RASs compared to other subtypes. A resistance-tailored retreatment using first- and second-generation DAAs was highly effective with SVR rates ≥90% across all subtypes and first-line treatment regimens.
Assuntos
Hepatite C Crônica , Antivirais/farmacologia , Antivirais/uso terapêutico , Farmacorresistência Viral/genética , Genótipo , Hepacivirus/genética , Hepatite C Crônica/tratamento farmacológico , Humanos , Estudos Retrospectivos , Falha de Tratamento , Proteínas não Estruturais Virais/genéticaRESUMO
BACKGROUND & AIMS: We performed an integrated analysis to identify microRNAs (miRNAs) and messenger RNAs (mRNAs) with altered expression in liver tumors from 3 mouse models of hepatocellular carcinoma (HCC) and human tumor tissues. METHODS: We analyzed miRNA and mRNA expression profiles of liver tissues from mice with diethylnitrosamine-induced hepatocarcinogenesis, conditional expression of lymphotoxin alpha and lymphotoxin beta, or inducible expression of a Myc transgene (Tet-O-Myc mice), as well as male C57BL/6 mice (controls). miRNA mimics were expressed and miRNAs and mRNAs were knocked down in human (Huh7, Hep3B, JHH2) hepatoma cell lines; cells were analyzed for viability, proliferation, apoptosis, migration, and invasion. Cells were grown as xenograft tumors in nude mice and analyzed. We combined in silico target gene prediction with mRNA profiles from all 3 mouse models. We quantified miRNA levels in 146 fresh-frozen tissues from patients (125 HCCs, 17 matched nontumor tissues, and 4 liver samples from patients without cancer) and published human data sets and tested correlations with patient survival times using Kaplan-Meier curves and the log-rank test. Levels of NUSAP1 mRNA were quantified in 237 HCCs and 5 nontumor liver samples using the TaqMan assay. RESULTS: Levels of the miRNA 193a-5p (MIR193A-5p) were reduced in liver tumors from all 3 mouse tumor models and in human HCC samples, compared with nontumor liver tissues. Expression of a MIR193A-5p mimic in hepatoma cells reduced proliferation, survival, migration, and invasion and their growth as xenograft tumors in nude mice. We found nucleolar and spindle-associated protein 1 (NUSAP1) to be a target of MIR193A-5p; HCC cells and tissues with low levels of MIR193A-5p had increased expression of NUSAP1. Increased levels of NUSAP1 in HCC samples correlated with shorter survival times of patients. Knockdown of NUSAP1 in Huh7 cells reduced proliferation, survival, migration, and growth as xenograft tumors in nude mice. Hydrodynamic tail-vein injections of a small hairpin RNA against NUSAP1 reduced growth of Akt1-Myc-induced tumors in mice. CONCLUSIONS: MIR193A-5p appears to prevent liver tumorigenesis by reducing levels of NUSAP1. Levels of MIR193A-5p are reduced in mouse and human HCC cells and tissues, leading to increased levels of NUSAP1, associated with shorter survival times of patients. Integrated analyses of miRNAs and mRNAs in tumors from mouse models can lead to identification of therapeutic targets in humans. The currently reported miRNA and mRNA profiling data have been submitted to the Gene Expression Omnibus (super-series accession number GSE102418).
Assuntos
Apoptose , Carcinogênese/genética , Proteínas de Ciclo Celular/metabolismo , Neoplasias Hepáticas/prevenção & controle , MicroRNAs/metabolismo , Proteínas Nucleares/metabolismo , Animais , Apoptose/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/prevenção & controle , Linhagem Celular Tumoral , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Fígado/metabolismo , Neoplasias Hepáticas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Proteínas Associadas aos Microtúbulos/metabolismo , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
UNLABELLED: Chronic hepatitis B virus (HBV) infection remains the most common risk factor for hepatocellular carcinoma (HCC). Efficient suppression of HBV viremia and necroinflammation as a result of nucleos(t)ide analogue treatment is able to reduce HCC incidence; nevertheless, hepatocarcinogenesis can occur in the absence of active hepatitis, correlating with high HBV surface antigen (HBsAg) levels. Nuclear factor κB (NF-κB) is a central player in chronic inflammation and HCC development. However, in the absence of severe chronic inflammation, the role of NF-κB signaling in HCC development remains elusive. As a model of hepatocarcinogenesis driven by accumulation of HBV envelope polypeptides, HBsAg transgenic mice, which show no HBV-specific immune response, were crossed to animals with hepatocyte-specific inhibition of canonical NF-κB signaling. We detected prolonged, severe endoplasmic reticulum stress already at 20 weeks of age in NF-κB-deficient hepatocytes of HBsAg-expressing mice. The unfolded protein response regulator binding immunoglobulin protein/78-kDa glucose-regulated protein was down-regulated, activating transcription factor 6, and eIF2α were activated with subsequent overexpression of CCAAT/enhancer binding protein homologous protein. Notably, immune cell infiltrates and liver transaminases were unchanged. However, as a result of this increased cellular stress, insufficient hepatocyte proliferation due to G1 /S-phase cell cycle arrest with overexpression of p27 and emergence of ductular reactions was detected. This culminated in increased DNA damage already at 20 weeks of age and finally led to 100% HCC incidence due to NF-κB inhibition. CONCLUSION: The role of canonical NF-κB signaling in HCC development depends on the mode of liver damage; in the case of HBsAg-driven hepatocarcinogenesis, NF-κB in hepatocytes acts as a critical tumor suppressor by augmenting the endoplasmic reticulum stress response.
Assuntos
Carcinoma Hepatocelular/prevenção & controle , Antígenos de Superfície da Hepatite B/fisiologia , Hepatócitos/metabolismo , Neoplasias Hepáticas/prevenção & controle , NF-kappa B/fisiologia , Transdução de Sinais/fisiologia , Proteínas Supressoras de Tumor/fisiologia , Resposta a Proteínas não Dobradas , Animais , Pontos de Checagem do Ciclo Celular , Hepatite B Crônica/complicações , Humanos , Regeneração Hepática , Camundongos , Camundongos Endogâmicos C57BL , Fator de Transcrição CHOP/fisiologiaRESUMO
Chronic hepatitis B virus (HBV) infection remains the number one risk factor for hepatocellular carcinoma (HCC), accounting for more than 600 000 deaths/year. Despite highly effective antiviral treatment options, chronic hepatitis B (CHB), subsequent end-stage liver disease and HCC development remain a major challenge worldwide. In CHB, liver damage is mainly caused by the influx of immune cells and destruction of infected hepatocytes, causing necro-inflammation. Treatment with nucleoside/nucleotide analogues can effectively suppress HBV replication in patients with CHB and thus decrease the risk for HCC development. Nevertheless, the risk of HCC in treated patients showing sufficient suppression of HBV DNA replication is significantly higher than in patients with inactive CHB, regardless of the presence of baseline liver cirrhosis, suggesting direct, long-lasting, predisposing effects of HBV. Direct oncogenic effects of HBV include integration in the host genome, leading to deletions, cis/trans-activation, translocations, the production of fusion transcripts and generalized genomic instability, as well as pleiotropic effects of viral transcripts (HBsAg and HBx). Analysis of these viral factors in active surveillance may allow early identification of high-risk patients, and their integration into a molecular classification of HCC subtypes might help in the development of novel therapeutic approaches.
Assuntos
Biomarcadores Tumorais/sangue , Carcinoma Hepatocelular/virologia , Vírus da Hepatite B/patogenicidade , Hepatite B Crônica/virologia , Neoplasias Hepáticas/virologia , Animais , Antivirais/uso terapêutico , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/prevenção & controle , DNA Viral/sangue , Genótipo , Vírus da Hepatite B/efeitos dos fármacos , Vírus da Hepatite B/genética , Vírus da Hepatite B/imunologia , Hepatite B Crônica/sangue , Hepatite B Crônica/complicações , Hepatite B Crônica/diagnóstico , Hepatite B Crônica/tratamento farmacológico , Hepatite B Crônica/imunologia , Interações Hospedeiro-Patógeno , Humanos , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/prevenção & controle , Valor Preditivo dos Testes , Fatores de Risco , Carga Viral , Proteínas Virais/sangue , Proteínas Virais/genética , Virulência , Integração ViralRESUMO
UNLABELLED: Alternatively polarized macrophages (MÏ) shape the microenvironment of hepatocellular carcinoma (HCC) and temper anticancer immune responses. We investigated if sorafenib alters the HCC microenvironment by restoring classical macrophage polarization and triggering tumor-directed natural killer (NK) cell responses. In vivo experiments were conducted with sorafenib (25 mg/kg)-treated C57BL/6 wildtype as well as hepatitis B virus (HBV) and lymphotoxin transgenic mice with and without HCC. Monocyte-derived MÏ or tumor-associated macrophages (TAM) isolated from HCC tissue were treated with sorafenib (0.07-5.0 µg/mL) and cocultured with autologous NK cells. MÏ and NK cell activation was analyzed by flow cytometry and killing assays, respectively. Cytokine and growth factor release was measured by enzyme-linked immunosorbent assay. Short-term administration of sorafenib triggered activation of hepatic NK cells in wildtype and tumor-bearing mice. In vitro, sorafenib sensitized MÏ to lipopolysaccharide, reverted alternative MÏ polarization and enhanced IL12 secretion (P = 0.0133). NK cells activated by sorafenib-treated MÏ showed increased degranulation (15.3 ± 0.2% versus 32.0 ± 0.9%, P < 0.0001) and interferon-gamma (IFN-γ) secretion (2.1 ± 0.2% versus 8.0 ± 0.2%, P < 0.0001) upon target cell contact. Sorafenib-triggered NK cell activation was verified by coculture experiments using TAM. Sorafenib-treated MÏ increased cytolytic NK cell function against K562, Raji, and HepG2 target cells in a dose-dependent manner. Neutralization of interleukin (IL)12 or IL18 as well as inhibition of the nuclear factor kappa B (NF-κB) pathway reversed NK cell activation in MÏ/NK cocultures. CONCLUSION: Sorafenib triggers proinflammatory activity of TAM and subsequently induces antitumor NK cell responses in a cytokine- and NF-κB-dependent fashion. This observation is relevant for HCC therapy, as sorafenib is a compound in clinical use that reverts alternative polarization of TAM in HCC. (HEPATOLOGY 2013;57:2358-2368).
Assuntos
Antineoplásicos/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Ativação de Macrófagos/efeitos dos fármacos , Niacinamida/análogos & derivados , Compostos de Fenilureia/farmacologia , Animais , Antineoplásicos/uso terapêutico , Apoptose , Carcinoma Hepatocelular/imunologia , Citocinas/metabolismo , Avaliação Pré-Clínica de Medicamentos , Humanos , Células Matadoras Naturais/efeitos dos fármacos , Neoplasias Hepáticas/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Niacinamida/farmacologia , Niacinamida/uso terapêutico , Compostos de Fenilureia/uso terapêutico , SorafenibeRESUMO
INTRODUCTION: The identification of risk factors for precursor lesions of colorectal cancer (CRC) holds great promise in the context of prevention. With this study, we aimed to identify patient characteristics associated with colorectal polyps (CPs) and polyp features of potential malignant progression. Furthermore, a potential association with gut microbiota in this context was investigated. METHODS: In this single-center study, a total of 162 patients with CPs and 91 control patients were included. Multiple variables including information on lifestyle, diet, serum parameters, and gut microbiota, analyzed by 16S-rRNA gene amplicon sequencing and functional imputations (Picrust2), were related to different aspects of CPs. RESULTS: We observed that elevated serum alkaline phosphatase (AP) levels were significantly associated with the presence of high-grade dysplastic polyps. This association was further seen for patients with CRC. Thereby, AP correlated with other parameters of liver function. We did not observe significant changes in the gut microbiota between patients with CP and their respective controls. However, a trend toward a lower alpha-diversity was seen in patients with CRC. Interestingly, AP was identified as a possible clinical effect modifier of stool sample beta diversity. DISCUSSION: We show for the first time an increased AP in premalignant CP. Furthermore, AP showed a significant influence on the microbial composition of the intestine. Relatively elevated liver enzymes, especially AP, may contribute to the detection of precancerous dysplastic or neoplastic changes in colorectal lesions. The association between elevated AP, premalignant CP, and the microbiome merits further study.
Assuntos
Pólipos do Colo , Neoplasias Colorretais , Microbioma Gastrointestinal , Humanos , Neoplasias Colorretais/genética , Pólipos do Colo/diagnóstico , Pólipos do Colo/patologia , Bactérias , Fezes , Microbioma Gastrointestinal/genética , HiperplasiaRESUMO
Hepatocellular carcinoma (HCC) ranks as the third leading cause of cancer-related death worldwide with currently limited treatment options. Chronic hepatitis B virus (HBV) infection accounts for HCC development in more than 50% of cases. The lifetime risk of HBV carriers to develop cirrhosis, liver failure or HCC is estimated to be as high as 15-40%. Although several pathways and triggers contributing to HCC development have been described, many features of hepatocellular carcinogenesis and the attributed direct role of viral factors remain elusive. Host genetic factors, the geographic area and epidemiologic factors, as well as the direct risk related to chronic HBV and hepatitis C virus (HCV) infections, account for geographical and gender differences of HCC prevalence. There is growing evidence that hepatocarcinogenesis is a multistep process. Human HCC is typically preceded by chronic inflammation and apoptotic and nonapoptotic cell death with compensatory liver proliferation. However, we still lack a thorough understanding of the common underlying molecular mechanisms. High levels of HBV replication and chronicity of inflammation are known to independently increase the risk for HCC. A direct carcinogenic role of viral factors is very likely to contribute to liver cancer since HCC is known to also occur in noncirrhotic livers of individuals with an inactive chronic or even with occult HBV infection with no significant histological signs of inflammation or cytopathic effects. Furthermore, synergistic or independent viral risk factors for primary liver cancer development have been described, such as HBV genotype, integration of viral DNA into the host genome and direct effects of viral proteins. A broader understanding of these viral factors in hepatocarcinogenesis might give rise to new diagnostic and therapeutic means in the future. We review the current state of research in liver cancer development and focus on the role of direct viral factors in HBV infection.
Assuntos
Vírus da Hepatite B/fisiologia , Hepatite B Crônica/virologia , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/virologia , Proteínas Virais/metabolismo , Animais , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/virologia , Modelos Animais de Doenças , Vírus da Hepatite B/genética , Hepatite B Crônica/epidemiologia , Hepatite B Crônica/transmissão , HumanosRESUMO
BACKGROUND: Liver biopsies to determine the cause of unclear hepatopathy and acute liver failure represent a diagnostic hallmark and require close cooperation between clinicians and pathologists. The commonly acute presentation of hepatic dysfunction warrants a rapid diagnosis. Infectious causes of hepatitis may be identified by the pathologist, supporting further diagnostic and therapeutic steps. OBJECTIVES: Rare infectious causes of hepatic dysfunction with distinct histomorphological features are presented. MATERIALS AND METHODS: Retrospective cases of liver biopsies for evaluation of hepatic dysfunction with infection confirmed by laboratory tests were selected from the archive of the institute of pathology of TUM and evaluated for morphologic diagnostic criteria. RESULTS AND CONCLUSIONS: Infections with adenovirus, Herpes simplex virus, Hepatitis A virus, and coxiella burnettii are rare findings in liver core biopsies but able to cause hepatic dysfunction that present with distinct histomorphologic alterations that can contribute to the identification of the causative agent. Rare infectious causes should be considered in any patient presenting with hepatic dysfunction of unknown etiology. Knowledge of the histomorphologic criteria by the pathologist is crucial to initiate further diagnostic testing and treatment.
Assuntos
Hepatite A , Hepatite , Falência Hepática Aguda , Fígado , Humanos , Diagnóstico Diferencial , Hepatite/diagnóstico , Estudos Retrospectivos , Fígado/patologiaRESUMO
Nivolumab is a promising monoclonal antibody inhibitor of programmed death-1, a protein on the surface of T-cells. As such, it is approved for use in patients with multiple advanced malignancies and can significantly elongate progression-free survival. However, monoclonal antibody inhibitors can lead to adverse hepatic reactions, which in rare cases result in further hepatic damage. Herein, we present a case of a patient with locally advanced gastric carcinoma treated with fluorouracil, oxaliplatin, docetaxel and the checkpoint inhibitor nivolumab. Five months after her first dosage of nivolumab and without a preexisting liver disease, she presented with transaminitis. During the course of her stay, the patient developed status epilepticus, which required mechanical ventilation followed by fulminant hepatic failure. A subsequent liver biopsy revealed severe liver damage with extensive confluent parenchymal necrosis corresponding to checkpoint-inhibitor-induced hepatitis. Alternative reasons for this hepatic failure were ruled out. Despite aggressive therapeutic interventions including corticosteroids and plasma exchange, the patient died due to liver failure. Although hepatic failure is rarely seen in patients with checkpoint inhibitor therapy, it requires early awareness and rapid intervention.
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Hepatic steatosis is characterized by intrahepatic lipid accumulation and may lead to irreversible liver damage if untreated. Here, we investigate whether multispectral optoacoustic tomography (MSOT) can offer label-free detection of liver lipid content to enable non-invasive characterization of hepatic steatosis by analyzing the spectral region around 930 nm, where lipids characteristically absorb. In a pilot study, we apply MSOT to measure liver and surrounding tissues in five patients with liver steatosis and five healthy volunteers, revealing significantly higher absorptions at 930 nm in the patients, while no significant difference was observed in the subcutaneous adipose tissue of the two groups. We further corroborated the human observations with corresponding MSOT measurements in high fat diet (HFD) - and regular chow diet (CD)-fed mice. This study introduces MSOT as a potential non-invasive and portable technique for detecting/monitoring hepatic steatosis in clinical settings, providing justification for larger studies.
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Ascitic fluid infection is a serious complication of liver cirrhosis. The distinction between the more common spontaneous bacterial peritonitis (SBP) and the less common secondary peritonitis in patients with liver cirrhosis is crucial due to the varying treatment approaches. This retrospective multicentre study was conducted in three German hospitals and analysed 532 SBP episodes and 37 secondary peritonitis episodes. Overall, >30 clinical, microbiological, and laboratory parameters were evaluated to identify key differentiation criteria. Microbiological characteristics in ascites followed by severity of illness and clinicopathological parameters in ascites were the most important predictors identified by a random forest model to distinguish between SBP and secondary peritonitis. To establish a point-score model, a least absolute shrinkage and selection operator (LASSO) regression model selected the ten most promising discriminatory features. By aiming at a sensitivity of 95% either to rule out or rule in SBP episodes, two cut-off scores were defined, dividing patients with infected ascites into a low-risk (score ≥ 45) and high-risk group (score < 25) for secondary peritonitis. Overall, the discrimination of secondary peritonitis from SBP remains challenging. Our univariable analyses, random forest model, and LASSO point score may help clinicians with the crucial differentiation between SBP and secondary peritonitis.
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BACKGROUND & AIMS: A single hepatitis B virus (HBV) particle is sufficient to establish chronic infection of the liver after intravenous injection, suggesting that the virus targets hepatocytes via a highly efficient transport pathway. We therefore investigated whether HBV uses a physiological liver-directed pathway that supports specific host-cell targeting in vivo. METHODS: We established the ex vivo perfusion of intact human liver tissue that recapitulates the liver physiology to investigate HBV liver targeting. This model allowed us to investigate virus-host cell interactions in a cellular microenvironment mimicking the in vivo situation. RESULTS: HBV was rapidly sequestered by liver macrophages within 1 hour after a virus pulse perfusion but was detected in hepatocytes only after 16 hours. We found that HBV associates with lipoproteins in serum and within machrophages. Electron and immunofluorescence microscopy corroborated a co-localization in recycling endosomes within peripheral and liver macrophages. Recycling endosomes accumulated HBV and cholesterol, followed by transport of HBV back to the cell surface along the cholesterol efflux pathway. To reach hepatocytes as final target cells, HBV was able to utilize the hepatocyte-directed cholesterol transport machinery of macrophages. CONCLUSIONS: Our results propose that by binding to liver targeted lipoproteins and using the reverse cholesterol transport pathway of macrophages, HBV hijacks the physiological lipid transport pathways to the liver to most efficiently reach its target organ. This may involve transinfection of liver macrophages and result in deposition of HBV in the perisinusoidal space from where HBV can bind its receptor on hepatocytes.
Assuntos
Vírus da Hepatite B , Hepatite B , Humanos , Vírus da Hepatite B/fisiologia , Hepatócitos/metabolismo , Colesterol/metabolismo , Lipoproteínas/metabolismo , LipídeosRESUMO
Hepatitis B virus (HBV) infection is a prominent cause of hepatocellular carcinoma (HCC) but the underlying molecular mechanisms are complex and multiple pathways have been proposed such as the activation of the Wnt-/ß-catenin-signalling and dysregulation of E-cadherin/ß-catenin adherens junctions. This study aimed to identify mechanisms of how HBV infection and replication as well as HBV X protein (HBx) gene expression in the context of an HBV genome influence Wnt-/ß-catenin-signalling and formation of adherens junctions and to which extent HBx contributes to this. Regulation of E-cadherin/ß-catenin junctions and ß-catenin-signalling as well as the role of HBx were investigated using constructs transiently or stably inducing replication of HBV+/-HBx in hepatoma cell lines. In addition, HCC and adjacent non-tumorous tissue samples from HBV-infected HCC patients and drug interference in HBV-infected cells were studied. Although HBV did not alter overall expression levels of E-cadherin or ß-catenin, it diminished their cell surface localization resulting in nuclear translocation of ß-catenin and activation of its target genes. In addition, HBV gene expression increased the amount of phosphorylated c-Src kinase. Treatment with Src kinase inhibitor Dasatinib reduced HBV replication, prevented adherens junction disassembly and reduced ß-catenin-signalling, while Sorafenib only did so in cells with mutated ß-catenin. Interestingly, none of the HBV induced alterations required HBx. Thus, HBV stimulated ß-catenin-signalling and induced disassembly of adherens junctions independently of HBx through Src kinase activation. These pathways may contribute to hepatocellular carcinogenesis and seem to be more efficiently inhibited by Dasatinib than by Sorafenib.