RESUMEN
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.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Enfermedad del Hígado Graso no Alcohólico , Oncostatina M , Animales , Carcinogénesis , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Ratones , Enfermedad del Hígado Graso no Alcohólico/genética , Enfermedad del Hígado Graso no Alcohólico/patologíaRESUMEN
The most common form of primary liver malignancy is hepatocellular carcinoma (HCC) [...].
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/terapia , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/terapia , Neoplasias Hepáticas/patologíaRESUMEN
Primary liver cancers (PLC), including hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA), are among the leading causes of cancer-related mortality worldwide. Bi-dimensional in vitro models are unable to recapitulate the key features of PLC; consequently, recent advancements in three-dimensional in vitro systems, such as organoids, opened up new avenues for the development of innovative models for studying tumour's pathological mechanisms. Liver organoids show self-assembly and self-renewal capabilities, retaining essential aspects of their respective in vivo tissue and allowing modelling diseases and personalized treatment development. In this review, we will discuss the current advances in the field of liver organoids focusing on existing development protocols and possible applications in regenerative medicine and drug discovery.
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Neoplasias de los Conductos Biliares , Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/patología , Hígado/patología , Organoides/patología , Neoplasias de los Conductos Biliares/patología , Conductos Biliares Intrahepáticos/patologíaRESUMEN
Chronic liver disease (CLD) is a major global health threat and has emerged as a leading cause of human death [...].
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Hepatopatías , Humanos , Enfermedad Crónica , Hepatopatías/diagnóstico , Hepatopatías/etiología , Hepatopatías/terapiaRESUMEN
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.
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Carcinoma Hepatocelular/tratamiento farmacológico , Sistemas de Liberación de Medicamentos , Ácido Hialurónico/farmacología , Liposomas/administración & dosificación , Macrófagos/efectos de los fármacos , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Polietilenglicoles/química , Animales , Carcinoma Hepatocelular/inmunología , Carcinoma Hepatocelular/patología , Humanos , Ácido Hialurónico/química , Liposomas/química , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/inmunología , Neoplasias Hepáticas/patología , Macrófagos/inmunología , Ratones , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/inmunología , Enfermedad del Hígado Graso no Alcohólico/patologíaRESUMEN
This observational study aims to assess the outcome and safety of O2-therapy by high-flow nasal cannula (HFNC) in 28 consecutive patients with severe hypoxemic acute respiratory failure (hARF) consequent to SARS-CoV-2 infection, unresponsive to conventional O2-therapy. Nineteen patients had a positive response. Nine patients required escalation of treatment to non-invasive ventilation (five subsequently intubated). None of the staff had a positive swab testing during the study period and the following 14 days. Severity of hypoxemia and C reactive protein level were correlated with HFNC failure. These data suggest HFNC to be a safe treatment for less severe patients with SARS-CoV-2 hARF and efficacy will need to be assessed as part of a clinical trial.
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Betacoronavirus , Infecciones por Coronavirus/complicaciones , Infecciones por Coronavirus/terapia , Hipoxia/terapia , Terapia por Inhalación de Oxígeno , Neumonía Viral/complicaciones , Neumonía Viral/terapia , Insuficiencia Respiratoria/terapia , Adulto , Anciano , Anciano de 80 o más Años , COVID-19 , Cánula , Estudios de Cohortes , Femenino , Humanos , Hipoxia/virología , Masculino , Persona de Mediana Edad , Ventilación no Invasiva , Pandemias , Insuficiencia Respiratoria/virología , SARS-CoV-2 , Resultado del TratamientoRESUMEN
Squamous cell carcinoma antigen-1 (SCCA1) overexpression is associated with poor prognosis and chemoresistance in several tumor types, however, the underlying mechanisms remain elusive. Here, we report SCCA1 in relation to the immune and peritumoral adipose tissue microenvironment in early and advanced esophageal adenocarcinoma (EAC). In our series of patients with EAC, free SCCA1 serum levels were associated with significantly worse overall survival, and SCCA1-IgM serum levels showed a trend to a worse overall survival. Serum SCCA1 and intratumoral SCCA1 were inversely correlated with immune activation markers. In agreement with these findings, SCCA1 induced the expression of the immune checkpoint molecule programmed death ligand-1 on monocytes and a direct correlation of these 2 molecules was observed in sequential tumor sections. Furthermore, SCCA1 mRNA expression within the tumor was inversely correlated with stem cell marker expression both within the tumor and in the peritumoral adipose tissue. In vitro, in EAC cell lines treated with different chemotherapeutic drugs, cell viability was significantly modified by SCCA1 presence, as cells overexpressing SCCA1 were significantly more resistant to cell death. In conclusion, poor prognosis in EAC overexpressing SCCA1 is due to reduced tumor chemosensitivity as well as intratumoral immunity impairment, likely induced by this molecule.
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Adenocarcinoma/metabolismo , Antígenos de Neoplasias/sangre , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Resistencia a Antineoplásicos , Neoplasias Esofágicas/metabolismo , Serpinas/sangre , Adenocarcinoma/genética , Anciano , Antígenos de Neoplasias/genética , Antineoplásicos/farmacología , Línea Celular Tumoral , Supervivencia Celular , Neoplasias Esofágicas/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Prospectivos , Serpinas/genética , Análisis de Supervivencia , Regulación hacia ArribaRESUMEN
Objectives: The real incidence of pneumomediastinum (PNM) in adult patients with severe acute asthma exacerbation continues to be unknown. The current study aims to investigate the occurrence of PNM in an adult population of patients presenting a severe asthma attack and to evaluate the risk factors associated to its development. Methods: The 45 consecutive subjects who were admitted to our Division between January 1, 2015 and December 31, 2016 for severe acute asthma exacerbation underwent a diagnostic protocol including a standard chest X-ray and continuous monitoring of arterial oxygen saturation (SaO2) during the first 24 hours following admission. The patients showing persistence or deterioration of oxyhemoglobin desaturation were prescribed a chest Computed Tomographic (CT) scan. Results: Five out of the 45 patients (11.1%) with severe acute asthma exacerbation were diagnosed with PNM, in one case on the basis of an X-ray image and in four on the basis of a chest CT scan. Data analysis showed that the PNM patients were younger [21 (17-21) vs 49.5 (20-73) yrs; p < 0.001] and more likely to show sensitization to Alternaria (2/5 vs 0/40; p = 0.0101) with respect to their non-PNM counterparts. The duration of hospital stay was similar in the two groups [8 (4-12) vs 7 (3-15) days; p = 0.6939]. Conclusions: PNM is a common clinical entity in young adults with severe acute asthma exacerbation, particularly in those with unsatisfactory response to initial medical therapy. Although generally benign, patients with suspected PNM should be closely monitored because of the risk of developing severe hypoxemia.
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Asma/epidemiología , Enfisema Mediastínico/epidemiología , Adolescente , Adulto , Factores de Edad , Anciano , Alternariosis/epidemiología , Antiasmáticos/uso terapéutico , Asma/tratamiento farmacológico , Humanos , Hipersensibilidad Inmediata/epidemiología , Tiempo de Internación , Persona de Mediana Edad , Oxígeno/sangre , Estudios Prospectivos , Radiografía Torácica , Índice de Severidad de la Enfermedad , Factores Sexuales , Fumar/epidemiología , Factores Socioeconómicos , Estado Asmático/epidemiología , Tomografía Computarizada por Rayos X , Adulto JovenRESUMEN
UNLABELLED: In cirrhosis, 11,12-epoxyeicosatrienoic acid (EET) induces mesenteric arterial vasodilation, which contributes to the onset of portal hypertension. We evaluated the hemodynamic effects of in vivo inhibition of EET production in experimental cirrhosis. Sixteen control rats and 16 rats with carbon tetrachloride-induced cirrhosis were studied. Eight controls and eight rats with cirrhosis were treated with the specific epoxygenase inhibitor N-(methylsulfonyl)-2-(2-propynyloxy)-benzenehexanamide (MS-PPOH; 20 mg/kg/day) for 3 consecutive days. Portal blood flow and renal and splenic resistive indexes were calculated through echographic measurements, while portal and systemic pressures were measured through polyethylene-50 catheters. Small resistance mesenteric arteries were connected to a pressure servo controller in a video-monitored perfusion system, and concentration-response curves to phenylephrine and acetylcholine were evaluated. EET levels were measured in tissue homogenates of rat liver, kidney, and aorta, using an enzyme-linked immunosorbent assay. Urinary Na(+) excretion function was also evaluated. In rats with cirrhosis, treatment with MS-PPOH significantly reduced portal blood flow and portal pressure compared to vehicle (13.6 ± 5.7 versus 25.3 ± 7.1 mL/min/100 g body weight, P < 0.05; 9.6 ± 1.1 versus 12.2 ± 2.3 mm Hg, P < 0.05; respectively) without effects on systemic pressure. An increased response to acetylcholine of mesenteric arteries from rats with cirrhosis (50% effect concentration -7.083 ± 0.197 versus -6.517 ± 0.73 in control rats, P < 0.05) was reversed after inhibition of EET production (-6.388 ± 0.263, P < 0.05). In liver, kidney, and aorta from animals with cirrhosis, treatment with MS-PPOH reversed the increase in EET levels. In both controls and rats with cirrhosis, MS-PPOH increased urinary Na(+) excretion. CONCLUSION: In rats with cirrhosis, in vivo inhibition of EET production normalizes the response of mesenteric arteries to vasodilators, with beneficial effects on portal hypertension. (Hepatology 2016;64:923-930).
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Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Amidas/uso terapéutico , Cirrosis Hepática Experimental/tratamiento farmacológico , Circulación Esplácnica/efectos de los fármacos , Resistencia Vascular/efectos de los fármacos , Ácido 8,11,14-Eicosatrienoico/antagonistas & inhibidores , Ácido 8,11,14-Eicosatrienoico/metabolismo , Acetilcolina , Amidas/farmacología , Animales , Aorta/metabolismo , Evaluación Preclínica de Medicamentos , Hipertensión Portal/tratamiento farmacológico , Riñón/metabolismo , Hígado/metabolismo , Cirrosis Hepática Experimental/fisiopatología , Masculino , Arterias Mesentéricas/efectos de los fármacos , Ratas Wistar , Sodio/metabolismoRESUMEN
One of the most daunting challenges of nanomedicine is the finding of appropriate targeting agents to deliver suitable payloads precisely to cells affected by malignancies. Even more complex is the ability to ensure that the nanosystems enter those cells. Here, we use 2 nm (metal core) gold nanoparticles to target human hepatocellular carcinoma (HepG2) cells stably transfected with the SERPINB3 (SB3) protein. The nanoparticles were coated with a 85:15 mixture of thiols featuring, respectively, a phosphoryl choline (to ensure water solubility and biocompatibility) and a 28-mer peptide corresponding to the amino acid sequence 21-47 of the hepatitis B virus-PreS1 protein (PreS1(21-47)). Conjugation of the peptide was performed via the maleimide-thiol reaction in methanol, allowing the use of a limited amount of the targeting molecule. This is an efficient procedure also in the perspective of selecting libraries of new targeting agents. The rationale behind the selection of the peptide is that SB3, which is undetectable in normal hepatocytes, is overexpressed in hepatocellular carcinoma and in hepatoblastoma and has been proposed as a target of the hepatitis B virus (HBV). For the latter, the key recognition element is the PreS1(21-47) peptide, which is a fragment of one of the proteins composing the viral envelope. The ability of the conjugated nanoparticles to bind the target protein SB3, expressed in liver cancer cells, was investigated by surface plasmon resonance analysis and in vitro via cellular uptake analysis followed by atomic absorption analysis of digested samples. The results showed that the PreS1(21-47) peptide is a suitable targeting agent for cells overexpressing the SB3 protein. Even more important is the evidence that the gold nanoparticles are internalized by the cells. The comparison between the surface plasmon resonance analysis and the cellular uptake studies suggests that the presentation of the protein on the cell surface is critical for efficient recognition.
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Carcinoma Hepatocelular/metabolismo , Oro/química , Neoplasias Hepáticas/metabolismo , Nanopartículas del Metal/química , Péptidos/química , Secuencia de Aminoácidos , Carcinoma Hepatocelular/patología , Células Hep G2 , Humanos , Neoplasias Hepáticas/patología , Microscopía Electrónica de Transmisión , Espectroscopía de Protones por Resonancia Magnética , Espectroscopía Infrarroja por Transformada de FourierRESUMEN
INTRODUCTION: Liver cirrhosis is characterized by structural and hemodynamic changes that affect mainly the liver, the kidney and the vascular system. Cytochrome P-450 (CYP) is a variegated family of enzymes that, among many other activities, metabolize arachidonic acid to the vasoactive epoxyeicosatrienoic acids (EETs). AIM: To investigate in an animal model of cirrhosis the m-RNA expression of CYPs in liver, kidney and aorta and to evaluate the effect of epoxygenase inhibition by N-(methylsulfonyl)-2-(2-propynyloxy)-benzenehexanamide (MS-PPOH). METHODS: In aorta, liver and kidney from 3 control, 3 cirrhotic and 6 cirrhotic rats treated with MS-PPOH, quantitative real-time PCR reactions were performed and the m-RNA expression of CYP2J3, CYP2J4, CYP2J10, CYP2C11, CYP2C12 and CYP2C23 was calculated. RESULTS: In cirrhotic rats, the gene expression of hepatic CYP2C11 and CYP2J10 was increased, of aortic CYP2J4 was increased, of aortic CYP2C12 was reduced and of renal CYP2C11 was increased. In cirrhotic rats, MS-PPOH reduced CYP2J10 hepatic and CYP2C11 renal gene expression to levels similar to the ones of control rats. CONCLUSIONS: Changes in CYPs gene expression may contribute to the hemodynamic alterations typical of cirrhosis. The altered gene expression of CYPs can, in some cases, be reversed by epoxygenase inhibition.
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Aorta/metabolismo , Sistema Enzimático del Citocromo P-450/genética , Regulación Enzimológica de la Expresión Génica , Riñón/metabolismo , Cirrosis Hepática/genética , Hígado/metabolismo , Amidas/farmacología , Animales , Aorta/efectos de los fármacos , Citocromo P-450 CYP2J2 , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Riñón/efectos de los fármacos , Hígado/efectos de los fármacos , Cirrosis Hepática/enzimología , Masculino , Especificidad de Órganos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas WistarRESUMEN
AIMS/HYPOTHESIS: Chronic foot ulceration is a severe complication of diabetes, driving morbidity and mortality. The mechanisms underlying delaying wound healing in diabetes are incompletely understood and tools to identify such pathways are eagerly awaited. METHODS: Wound biopsies were obtained from 75 patients with diabetic foot ulcers. Matched subgroups of rapidly healing (RH, n = 17) and non-healing (NH, n = 11) patients were selected. Proteomic analysis was performed by labelling with isobaric tag for relative and absolute quantification and mass spectrometry. Differentially expressed proteins were analysed in NH vs RH for identification of pathogenic pathways. Individual sample gene/protein validation and in vivo validation of candidate pathways in mouse models were carried out. RESULTS: Pathway analyses were conducted on 92/286 proteins that were differentially expressed in NH vs RH. The following pathways were enriched in NH vs RH patients: apoptosis, protease inhibitors, epithelial differentiation, serine endopeptidase activity, coagulation and regulation of defence response. SerpinB3 was strongly upregulated in RH vs NH wounds, validated as protein and mRNA in individual samples. To test the relevance of serpinB3 in vivo, we used a transgenic mouse model with α1-antitrypsin promoter-driven overexpression of human SERPINB3. In this model, wound healing was unaffected by SERPINB3 overexpression in non-diabetic or diabetic mice with or without hindlimb ischaemia. In an independent validation cohort of 47 patients, high serpinB3 protein content was confirmed as a biomarker of healing improvement. CONCLUSIONS/INTERPRETATION: We provide a benchmark for the unbiased discovery of novel molecular targets and biomarkers of impaired diabetic wound healing. High serpinB3 protein content was found to be a biomarker of successful healing in diabetic patients.
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Antígenos de Neoplasias/metabolismo , Biomarcadores/metabolismo , Pie Diabético/metabolismo , Pie Diabético/fisiopatología , Serpinas/metabolismo , Cicatrización de Heridas/fisiología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Antígenos de Neoplasias/genética , Femenino , Humanos , Masculino , Persona de Mediana Edad , Serpinas/genética , Adulto JovenRESUMEN
BACKGROUND: In the setting of liver injury hepatic progenitor cells are activated, counterbalancing the inhibited regenerative capacity of mature hepatocytes. Chronic activation of this compartment may give rise to a subset of liver tumours with poor prognosis. SerpinB3, a serpin over-expressed in injured liver and in primary liver cancer, has been shown to induce apoptosis resistance, epithelial to mesenchymal transition and to increase TGF-beta and Myc expression. Aim of the present study was to explore the presence of SerpinB3 in hepatic progenitor cells in human livers and in a mouse model of liver stem/progenitor cell activation.Hepatic progenitor cells were analysed in foetal and adult livers at protein and transcriptional levels. To induce experimental activation of the liver stem/progenitor compartment, C57BL/6J mice were injected with lipopolysaccharide plus D-galactosamine and were sacrificed at different time points. Liver cDNA was amplified using specific primers for mouse-homologous SerpinB3 isoforms and automatically sequenced. RESULTS: The presence of SerpinB3 in the progenitor cell compartment was detected in sorted human foetal and adult epithelial cell adhesion molecule (EpCAM) positive liver cells. By immunohistochemistry SerpinB3 was found in human cirrhotic livers in portal areas with progenitor cell activation showing ductular proliferation. CK-7, CK-19, EpCAM and CD-90 positive cell were also positive for SerpinB3. In the animal model, time course analysis in liver specimens revealed a progressive increase of SerpinB3 and a parallel decrease of activated caspase 3, which was barely detectable at 20 hours. Transcription analysis confirmed the presence of SerpinB3-homologous only in the liver of injured mice and sequence analysis proved its belonging to mouse Serpinb3b. CONCLUSION: SerpinB3 is highly expressed in hepatic stem/progenitor cell compartment of both foetal and adult livers.
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Antígenos de Neoplasias/metabolismo , Hígado/citología , Serpinas/metabolismo , Células Madre/metabolismo , Animales , Secuencia de Bases , Caspasa 3/metabolismo , Moléculas de Adhesión Celular/metabolismo , Modelos Animales de Enfermedad , Molécula de Adhesión Celular Epitelial , Humanos , Inmunohistoquímica , Queratina-19/metabolismo , Queratina-7/metabolismo , Cirrosis Hepática/metabolismo , Cirrosis Hepática/patología , Ratones , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , Alineación de Secuencia , Células Madre/citología , Antígenos Thy-1/metabolismoRESUMEN
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.
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Hepatopatías , Factor de Crecimiento Transformador beta1 , Humanos , Progresión de la Enfermedad , Cirrosis Hepática/genética , Cirrosis Hepática/metabolismo , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
The voltage-dependent anion-selective channel isoform 1 (VDAC1) is a pivotal component in cellular metabolism and apoptosis with a prominent role in many cancer types, offering a unique therapeutic intervention point. Through an in-silico-to-in-vitro approach we identified a set of VA molecules (VDAC Antagonists) that selectively bind to VDAC1 and display specificity toward cancer cells. Biochemical characterization showed that VA molecules can directly interact with VDAC1 with micromolar affinity by competing with the endogenous ligand NADH for a partially shared binding site. NADH displacement results in mitochondrial distress and reduced cell proliferation, especially when compared to non-cancerous cells. Experiments performed on organoids derived from intrahepatic cholangiocarcinoma patients demonstrated a dose-dependent reduction in cell viability upon treatment with VA molecules with lower impact on healthy cells than conventional treatments like gemcitabine. VA molecules are chemical entities representing promising candidates for further optimization and development as cancer therapy strategies through precise metabolic interventions.
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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.
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Enfermedad del Hígado Graso no Alcohólico , Ratones , Animales , Humanos , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Receptor PAR-2 , Proteína beta Potenciadora de Unión a CCAAT , Cirrosis Hepática/tratamiento farmacológico , Ratones Transgénicos , InflamaciónRESUMEN
Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease. NAFLD can evolve from simple fatty liver to non-alcoholic steatohepatitis (NASH), and ultimately, to cirrhosis. Inflammation and oxidative stress, promoted by mitochondrial dysfunction, play a crucial role in the onset and development of NASH. To date, no therapy has been approved for NAFLD and NASH. The aim of this study is to evaluate if the anti-inflammatory activity of acetylsalicylic acid (ASA) and the mitochondria-targeted antioxidant effect of mitoquinone could hinder the progression of non-alcoholic steatohepatitis. In mice, fatty liver was induced through the administration of a deficient in methionine and choline and rich in fat diet. Two experimental groups were treated orally with ASA or mitoquinone. Histopathologic evaluation of steatosis and inflammation was performed; the hepatic expression of genes associated with inflammation, oxidative stress, and fibrosis was evaluated; the protein expression of IL-10, cyclooxygenase 2, superoxide dismutase 1, and glutathione peroxidase 1 in the liver was analyzed; a quantitative analysis of 15-epi-lipoxin A4 in liver homogenates was performed. Mitoquinone and ASA significantly reduced liver steatosis and inflammation by decreasing the expression of TNFα, IL-6, Serpinb3, and cyclooxygenase 1 and 2 and restoring the anti-inflammatory IL-10. Treatment with mitoquinone and ASA increased the gene and protein expression of antioxidants, i.e., catalase, superoxide dismutase 1, and glutathione peroxidase 1, and decreased the expression of profibrogenic genes. ASA normalized the levels of 15-epi-Lipoxin A4. In mice fed with a deficient in methionine and choline and rich in fat diet, mitoquinone and ASA reduce steatosis and necroinflammation and may represent two effective novel strategies for the treatment of non-alcoholic steatohepatitis.
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SerpinB3 is a serine protease inhibitor that plays a relevant role in disease progression and cancer by increasing fibrosis, cell proliferation, and invasion, besides conferring resistance to apoptosis. The mechanisms underlying these biological activities are not yet fully understood. The aim of this study was to generate antibodies directed against different SerpinB3 epitopes to better investigate their biological role. Five exposed epitopes were identified using the software DNASTAR Lasergene and the corresponding synthetic peptides were used for NZW rabbit immunization. Anti-P#2 and anti-P#4 antibodies were able to recognize both SerpinB3 and SerpinB4 by ELISA. Anti-P#5 antibody, produced against the reactive site loop of SerpinB3, showed the greatest specific reactivity for human SerpinB3. This antibody was able to recognize SerpinB3 at nuclear level, while anti-P#3 antibody recognized SerpinB3 only at cytoplasmic level, both by immunofluorescence and by immunohistochemistry. The biological activity of each antibody preparation was assessed in HepG2 cells overexpressing SerpinB3 and anti-P#5 antibody reduced proliferation by 12% cell and cell invasion by 75%, while trivial results were obtained with the other antibody preparations. These findings indicate that the reactive site loop of SerpinB3 is essential for the invasiveness features induced by this serpin and it could become a novel druggable target.
Asunto(s)
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Animales , Conejos , Células Hep G2 , Progresión de la Enfermedad , Ensayo de Inmunoadsorción EnzimáticaRESUMEN
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.
RESUMEN
We have investigated the change in SerpinB3 during hepatic ischemia and the potential role of its antiprotease activity in cell protection by the administration of wild-type SerpinB3 (SerpinB3-WT) or active loop-deleted recombinant SerpinB3 protein (SerpinB3-D) in a rat model of ischemia (60 min)/reperfusion (60 min) (I/R). A time-dependent increase of SerpinB3, both at transcription and protein level, was found in ischemic livers after 60, 120 and 180 min. SerpinB3-WT decreased polymorphonuclear cell infiltration and serum enzymes and increased ATP when compared with I/R group. These events were not obtained using SerpinB3-D. No significant changes in both liver SerpinB3 mRNA and protein were found in all I/R groups considered. The present data show that the administration of SerpinB3-WT reduced the I/R injury and this effect appears to be dependent on its anti-protease activity.