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1.
Nature ; 630(8015): 166-173, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38778114

RESUMO

For many adult human organs, tissue regeneration during chronic disease remains a controversial subject. Regenerative processes are easily observed in animal models, and their underlying mechanisms are becoming well characterized1-4, but technical challenges and ethical aspects are limiting the validation of these results in humans. We decided to address this difficulty with respect to the liver. This organ displays the remarkable ability to regenerate after acute injury, although liver regeneration in the context of recurring injury remains to be fully demonstrated. Here we performed single-nucleus RNA sequencing (snRNA-seq) on 47 liver biopsies from patients with different stages of metabolic dysfunction-associated steatotic liver disease to establish a cellular map of the liver during disease progression. We then combined these single-cell-level data with advanced 3D imaging to reveal profound changes in the liver architecture. Hepatocytes lose their zonation and considerable reorganization of the biliary tree takes place. More importantly, our study uncovers transdifferentiation events that occur between hepatocytes and cholangiocytes without the presence of adult stem cells or developmental progenitor activation. Detailed analyses and functional validations using cholangiocyte organoids confirm the importance of the PI3K-AKT-mTOR pathway in this process, thereby connecting this acquisition of plasticity to insulin signalling. Together, our data indicate that chronic injury creates an environment that induces cellular plasticity in human organs, and understanding the underlying mechanisms of this process could open new therapeutic avenues in the management of chronic diseases.


Assuntos
Transdiferenciação Celular , Hepatócitos , Hepatopatias , Fígado , Humanos , Sistema Biliar/citologia , Sistema Biliar/metabolismo , Sistema Biliar/patologia , Biópsia , Plasticidade Celular , Doença Crônica , Progressão da Doença , Células Epiteliais/metabolismo , Células Epiteliais/citologia , Células Epiteliais/patologia , Hepatócitos/metabolismo , Hepatócitos/citologia , Hepatócitos/patologia , Insulina/metabolismo , Fígado/patologia , Fígado/metabolismo , Fígado/citologia , Hepatopatias/patologia , Hepatopatias/metabolismo , Regeneração Hepática , Organoides/metabolismo , Organoides/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA-Seq , Transdução de Sinais , Análise de Célula Única , Serina-Treonina Quinases TOR/metabolismo
2.
Nature ; 615(7950): 134-142, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36470304

RESUMO

Preventing SARS-CoV-2 infection by modulating viral host receptors, such as angiotensin-converting enzyme 2 (ACE2)1, could represent a new chemoprophylactic approach for COVID-19 that complements vaccination2,3. However, the mechanisms that control the expression of ACE2 remain unclear. Here we show that the farnesoid X receptor (FXR) is a direct regulator of ACE2 transcription in several tissues affected by COVID-19, including the gastrointestinal and respiratory systems. We then use the over-the-counter compound z-guggulsterone and the off-patent drug ursodeoxycholic acid (UDCA) to reduce FXR signalling and downregulate ACE2 in human lung, cholangiocyte and intestinal organoids and in the corresponding tissues in mice and hamsters. We show that the UDCA-mediated downregulation of ACE2 reduces susceptibility to SARS-CoV-2 infection in vitro, in vivo and in human lungs and livers perfused ex situ. Furthermore, we reveal that UDCA reduces the expression of ACE2 in the nasal epithelium in humans. Finally, we identify a correlation between UDCA treatment and positive clinical outcomes after SARS-CoV-2 infection using retrospective registry data, and confirm these findings in an independent validation cohort of recipients of liver transplants. In conclusion, we show that FXR has a role in controlling ACE2 expression and provide evidence that modulation of this pathway could be beneficial for reducing SARS-CoV-2 infection, paving the way for future clinical trials.


Assuntos
Enzima de Conversão de Angiotensina 2 , COVID-19 , Receptores Virais , Ácido Ursodesoxicólico , Animais , Humanos , Camundongos , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/metabolismo , COVID-19/metabolismo , COVID-19/prevenção & controle , Receptores Virais/genética , Receptores Virais/metabolismo , Estudos Retrospectivos , SARS-CoV-2/metabolismo , Tratamento Farmacológico da COVID-19 , Cricetinae , Transcrição Gênica , Ácido Ursodesoxicólico/farmacologia , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Organoides/efeitos dos fármacos , Organoides/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Mucosa Nasal/efeitos dos fármacos , Mucosa Nasal/metabolismo , Sistema de Registros , Reprodutibilidade dos Testes , Transplante de Fígado
3.
Nature ; 598(7881): 473-478, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34646017

RESUMO

The progression of chronic liver disease to hepatocellular carcinoma is caused by the acquisition of somatic mutations that affect 20-30 cancer genes1-8. Burdens of somatic mutations are higher and clonal expansions larger in chronic liver disease9-13 than in normal liver13-16, which enables positive selection to shape the genomic landscape9-13. Here we analysed somatic mutations from 1,590 genomes across 34 liver samples, including healthy controls, alcohol-related liver disease and non-alcoholic fatty liver disease. Seven of the 29 patients with liver disease had mutations in FOXO1, the major transcription factor in insulin signalling. These mutations affected a single hotspot within the gene, impairing the insulin-mediated nuclear export of FOXO1. Notably, six of the seven patients with FOXO1S22W hotspot mutations showed convergent evolution, with variants acquired independently by up to nine distinct hepatocyte clones per patient. CIDEB, which regulates lipid droplet metabolism in hepatocytes17-19, and GPAM, which produces storage triacylglycerol from free fatty acids20,21, also had a significant excess of mutations. We again observed frequent convergent evolution: up to fourteen independent clones per patient with CIDEB mutations and up to seven clones per patient with GPAM mutations. Mutations in metabolism genes were distributed across multiple anatomical segments of the liver, increased clone size and were seen in both alcohol-related liver disease and non-alcoholic fatty liver disease, but rarely in hepatocellular carcinoma. Master regulators of metabolic pathways are a frequent target of convergent somatic mutation in alcohol-related and non-alcoholic fatty liver disease.


Assuntos
Hepatopatias/genética , Hepatopatias/metabolismo , Fígado/metabolismo , Mutação/genética , Transporte Ativo do Núcleo Celular/genética , Proteínas Reguladoras de Apoptose/genética , Linhagem Celular Tumoral , Doença Crônica , Estudos de Coortes , Ácidos Graxos não Esterificados/metabolismo , Feminino , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O1/metabolismo , Humanos , Resistência à Insulina , Hepatopatias Alcoólicas/genética , Hepatopatias Alcoólicas/metabolismo , Masculino , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/metabolismo , Triglicerídeos/metabolismo
4.
Nature ; 583(7815): 265-270, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32581361

RESUMO

Cancers arise through the acquisition of oncogenic mutations and grow by clonal expansion1,2. Here we reveal that most mutagenic DNA lesions are not resolved into a mutated DNA base pair within a single cell cycle. Instead, DNA lesions segregate, unrepaired, into daughter cells for multiple cell generations, resulting in the chromosome-scale phasing of subsequent mutations. We characterize this process in mutagen-induced mouse liver tumours and show that DNA replication across persisting lesions can produce multiple alternative alleles in successive cell divisions, thereby generating both multiallelic and combinatorial genetic diversity. The phasing of lesions enables accurate measurement of strand-biased repair processes, quantification of oncogenic selection and fine mapping of sister-chromatid-exchange events. Finally, we demonstrate that lesion segregation is a unifying property of exogenous mutagens, including UV light and chemotherapy agents in human cells and tumours, which has profound implications for the evolution and adaptation of cancer genomes.


Assuntos
Segregação de Cromossomos/genética , Evolução Molecular , Genoma/genética , Neoplasias/genética , Alelos , Animais , Reparo do DNA , Replicação do DNA , Receptores ErbB/metabolismo , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Mutação , Neoplasias/patologia , Seleção Genética , Transdução de Sinais , Troca de Cromátide Irmã , Transcrição Gênica , Quinases raf/metabolismo , Proteínas ras/metabolismo
5.
Nature ; 574(7779): 538-542, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31645727

RESUMO

The most common causes of chronic liver disease are excess alcohol intake, viral hepatitis and non-alcoholic fatty liver disease, with the clinical spectrum ranging in severity from hepatic inflammation to cirrhosis, liver failure or hepatocellular carcinoma (HCC). The genome of HCC exhibits diverse mutational signatures, resulting in recurrent mutations across more than 30 cancer genes1-7. Stem cells from normal livers have a low mutational burden and limited diversity of signatures8, which suggests that the complexity of HCC arises during the progression to chronic liver disease and subsequent malignant transformation. Here, by sequencing whole genomes of 482 microdissections of 100-500 hepatocytes from 5 normal and 9 cirrhotic livers, we show that cirrhotic liver has a higher mutational burden than normal liver. Although rare in normal hepatocytes, structural variants, including chromothripsis, were prominent in cirrhosis. Driver mutations, such as point mutations and structural variants, affected 1-5% of clones. Clonal expansions of millimetres in diameter occurred in cirrhosis, with clones sequestered by the bands of fibrosis that surround regenerative nodules. Some mutational signatures were universal and equally active in both non-malignant hepatocytes and HCCs; some were substantially more active in HCCs than chronic liver disease; and others-arising from exogenous exposures-were present in a subset of patients. The activity of exogenous signatures between adjacent cirrhotic nodules varied by up to tenfold within each patient, as a result of clone-specific and microenvironmental forces. Synchronous HCCs exhibited the same mutational signatures as background cirrhotic liver, but with higher burden. Somatic mutations chronicle the exposures, toxicity, regeneration and clonal structure of liver tissue as it progresses from health to disease.


Assuntos
Células Clonais/citologia , Células Clonais/patologia , Fibrose/genética , Fibrose/patologia , Fígado/citologia , Fígado/metabolismo , Mutação , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Células Clonais/metabolismo , Análise Mutacional de DNA , Hepatócitos/citologia , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Fígado/patologia , Masculino , Pessoa de Meia-Idade , Filogenia , Células-Tronco/citologia , Células-Tronco/metabolismo , Células-Tronco/patologia
6.
Clin Gastroenterol Hepatol ; 21(2): 535-537.e1, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-35032633

RESUMO

Splenomegaly in the context of liver disease is classically associated with advanced cirrhosis and portal hypertension.1 More recently, we observed an increasing number of patients with splenomegaly and nonalcoholic fatty liver disease (NAFLD), but in whom intensive work-up revealed no evidence of advanced liver disease or portal hypertension. In this study, we found no correlation between spleen size and histological stage of NAFLD, and a strong correlation between body weight, height and serum high density lipoprotein (HDL) levels.


Assuntos
Hipertensão Portal , Hepatopatia Gordurosa não Alcoólica , Humanos , Hepatopatia Gordurosa não Alcoólica/patologia , Baço/patologia , Esplenomegalia/etiologia , Fígado/patologia , Cirrose Hepática/patologia , Hipertensão Portal/complicações
7.
Dig Dis Sci ; 67(12): 5551-5561, 2022 12.
Artigo em Inglês | MEDLINE | ID: mdl-35288833

RESUMO

BACKGROUND: Mutations in the ABCB4 gene are associated with failure of bile acid emulsification leading to cholestatic liver disease. Presentations range from progressive familial intrahepatic cholestasis type 3 (PFIC3) in childhood, to milder forms seen in adulthood. AIMS: We sought to characterize adult disease with particular reference to histology which has been hitherto poorly defined. METHODS: Four unrelated adults (three female, mean age 39 years) and three sisters presenting with cholestatic liver disease in adulthood, associated with variants in the ABCB4 gene, were identified. Clinical review and detailed blinded histopathological analysis were performed. RESULTS: Two novel pathogenic ABCB4 variants were identified: c.620 T > G, p.(Ile207Arg) and c.2301dupT, p.(Thr768TyrfsTer26). Sub-phenotypes observed included low-phospholipid-associated cholelithiasis syndrome (LPAC), intrahepatic cholestasis of pregnancy (ICP), drug-induced cholestasis, idiopathic adulthood ductopenia, and adult PFIC3. Of note, 5/7 had presented with gallstone complications (4 meeting LPAC definition) and 4/6 females had a history of ICP. Considerable overlap was observed phenotypically and liver transplantation was required in 3/7 of patients. Histologically, cases generally demonstrated ductopenia of the smaller tracts, mild non-ductocentric portal inflammation, bilirubinostasis, significant copper-associated protein deposition, and varying degrees of fibrosis. CONCLUSIONS: Adults with ABCB4 mutations may harbor a spectrum of cholestatic disease phenotypes and can progress to liver transplantation. We observed a distinct histological pattern which differs from classical biliary disease and describe two novel pathogenic ABCB4 variants. ABCB4 sequencing should be considered in patients with relevant cholestatic phenotypes and/or suggestive histology; accurate diagnosis can guide potential interventions to delay progression and inform family screening.


Assuntos
Colestase Intra-Hepática , Colestase , Doenças da Vesícula Biliar , Cálculos Biliares , Feminino , Humanos , Gravidez , Colestase Intra-Hepática/diagnóstico , Colestase Intra-Hepática/genética , Mutação
8.
Transpl Int ; 34(4): 709-720, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33462839

RESUMO

Transplantation of severely steatotic donor livers is associated with early allograft dysfunction and poorer graft survival. Histology remains the gold standard diagnostic of donor steatosis despite the lack of consensus definition and its subjective nature. In this prospective observational study of liver transplant patients, we demonstrate the feasibility of using a handheld optical backscatter probe to assess the degree of hepatic steatosis and correlate the backscatter readings with clinical outcomes. The probe is placed on the surface of the liver and emits red and near infrared light from the tip of the device and measures the amount of backscatter of light from liver tissue via two photodiodes. Measurement of optical backscatter (Mantel-Cox P < 0.0001) and histopathological scoring of macrovesicular steatosis (Mantel-Cox P = 0.046) were predictive of 5-year graft survival. Recipients with early allograft dysfunction defined according to both Olthoff (P = 0.0067) and MEAF score (P = 0.0097) had significantly higher backscatter levels from the donor organ. Backscatter was predictive of graft loss (AUC 0.75, P = 0.0045). This study demonstrates the feasibility of real-time measurement of optical backscatter in donor livers. Early results indicate readings correlate with steatosis and may give insight to graft outcomes such as early allograft dysfunction and graft loss.


Assuntos
Transplante de Fígado , Sobrevivência de Enxerto , Humanos , Fígado/diagnóstico por imagem , Projetos Piloto , Índice de Gravidade de Doença , Doadores de Tecidos
9.
Hepatology ; 65(4): 1165-1180, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27863448

RESUMO

Nonalcoholic fatty liver disease (NAFLD) can progress from simple steatosis (i.e., nonalcoholic fatty liver [NAFL]) to nonalcoholic steatohepatitis (NASH), cirrhosis, and cancer. Currently, the driver for this progression is not fully understood; in particular, it is not known how NAFLD and its early progression affects the distribution of lipids in the liver, producing lipotoxicity and inflammation. In this study, we used dietary and genetic mouse models of NAFL and NASH and translated the results to humans by correlating the spatial distribution of lipids in liver tissue with disease progression using advanced mass spectrometry imaging technology. We identified several lipids with distinct zonal distributions in control and NAFL samples and observed partial to complete loss of lipid zonation in NASH. In addition, we found increased hepatic expression of genes associated with remodeling the phospholipid membrane, release of arachidonic acid (AA) from the membrane, and production of eicosanoid species that promote inflammation and cell injury. The results of our immunohistochemistry analyses suggest that the zonal location of remodeling enzyme LPCAT2 plays a role in the change in spatial distribution for AA-containing lipids. This results in a cycle of AA-enrichment in pericentral hepatocytes, membrane release of AA, and generation of proinflammatory eicosanoids and may account for increased oxidative damage in pericentral regions in NASH. CONCLUSION: NAFLD is associated not only with lipid enrichment, but also with zonal changes of specific lipids and their associated metabolic pathways. This may play a role in the heterogeneous development of NAFLD. (Hepatology 2017;65:1165-1180).


Assuntos
Eicosanoides/metabolismo , Cirrose Hepática/patologia , Regeneração Hepática/fisiologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Fosfolipídeos/metabolismo , Animais , Biópsia por Agulha , Dieta Hiperlipídica , Dieta Ocidental , Modelos Animais de Doenças , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Humanos , Imuno-Histoquímica , Cirrose Hepática/metabolismo , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Masculino , Espectrometria de Massas , Camundongos , Camundongos Endogâmicos C57BL , Prognóstico , Distribuição Aleatória , Medição de Risco , Índice de Gravidade de Doença , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
10.
Histopathology ; 69(2): 315-21, 2016 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26826338

RESUMO

AIMS: Liver pathology is a challenging subspeciality, with histopathologists frequently seeking specialist opinions. This study aims to determine the impact of specialist reviews on the final diagnosis and patient management. METHODS AND RESULTS: Agreement with the initial reporting centre in the histopathological diagnosis of 1265 liver biopsies was determined. The nature of differences was explored in more depth for 103 discrepant cases. Differences in the histopathological interpretation were present in 749 of 1265 (59%) biopsies, of which 505 of 749 (67%) were predicted at the time of reporting to impact upon patient management. Agreement was good in cases with chronic viral hepatitis, fatty liver disease, malignancy and minimal pathological changes, while diagnostic differences occurred in more than 70% with biliary disease, autoimmune hepatitis or vascular/architectural changes. A clinical review of a subset of reports with histopathological differences predicted changes in patient management in 63 of 103 (61%). CONCLUSIONS: Clinically significant differences in liver biopsy interpretation between local pathologists and subspecialists are common. Diagnoses with frequent discrepancies, such as biliary disease, may benefit from a specialist review as standard when diagnosed initially, while cases requiring specialist advice from disease subgroups where discrepancies are less common, such as chronic viral hepatitis, could be selected during the clinicopathological conference process.


Assuntos
Fígado Gorduroso/diagnóstico , Hepatite Autoimune/diagnóstico , Hepatopatias/diagnóstico , Fígado/patologia , Encaminhamento e Consulta/normas , Biópsia , Erros de Diagnóstico/prevenção & controle , Fígado Gorduroso/patologia , Hepatite Autoimune/patologia , Humanos , Hepatopatias/patologia , Especialização
11.
Liver Transpl ; 21(4): 487-99, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25545865

RESUMO

Ischemia/reperfusion injury (IRI) that develops after liver implantation may prejudice long-term graft survival, but it remains poorly understood. Here we correlate the severity of IRIs that were determined by histological grading of time-zero biopsies sampled after graft revascularization with patient and graft outcomes. Time-zero biopsies of 476 liver transplants performed at our center between 2000 and 2010 were graded as follows: nil (10.5%), mild (58.8%), moderate (26.1%), and severe (4.6%). Severe IRI was associated with donor age, donation after circulatory death, prolonged cold ischemia time, and liver steatosis, but it was also associated with increased rates of primary nonfunction (9.1%) and retransplantation within 90 days (22.7%). Longer term outcomes in the severe IRI group were also poor, with 1-year graft and patient survival rates of only 55% and 68%, respectively (cf. 90% and 93% for the remainder). Severe IRI on the time-zero biopsy was, in a multivariate analysis, an independent determinant of 1-year graft survival and was a better predictor of 1-year graft loss than liver steatosis, early graft dysfunction syndrome, and high first-week alanine aminotransferase with a positive predictive value of 45%. Time-zero biopsies predict adverse clinical outcomes after liver transplantation, and severe IRI upon biopsy signals the likely need for early retransplantation.


Assuntos
Transplante de Fígado/efeitos adversos , Traumatismo por Reperfusão/patologia , Adulto , Fatores Etários , Idoso , Alanina Transaminase/sangue , Aloenxertos , Biomarcadores/sangue , Biópsia , Isquemia Fria/efeitos adversos , Feminino , Sobrevivência de Enxerto , Humanos , Estimativa de Kaplan-Meier , Transplante de Fígado/mortalidade , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Valor Preditivo dos Testes , Modelos de Riscos Proporcionais , Traumatismo por Reperfusão/sangue , Traumatismo por Reperfusão/etiologia , Traumatismo por Reperfusão/mortalidade , Estudos Retrospectivos , Fatores de Risco , Índice de Gravidade de Doença , Fatores de Tempo , Doadores de Tecidos , Resultado do Tratamento , Adulto Jovem
12.
Am J Respir Crit Care Med ; 187(5): 502-8, 2013 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-23262512

RESUMO

RATIONALE: α1-Antitrypsin deficiency is one of the most common heritable human diseases, predisposing to liver and lung injury. Significant heterogeneity in phenotypic expression is well documented, but less is known of the prevalence, severity, and correlates of chronic liver disease among individuals presenting with lung disease. OBJECTIVES: To determine the frequency of and risk factors for severe liver fibrosis and cirrhosis among individuals with PiZZ-related lung disease. METHODS: A well-characterized cohort of 57 PiZZ adults attending a tertiary referral respiratory clinic was screened prospectively for clinical, laboratory, radiologic, and (when appropriate) histologic evidence of chronic liver disease. MEASUREMENTS AND MAIN RESULTS: Thirty-six (63.2%) of 57 had a history or clinical findings suggestive of liver disease; or had one or more abnormalities of liver function, or liver ultrasound, and 24 of these underwent liver biopsy. Ten (17.5%) had evidence of severe fibrosis or cirrhosis and were more likely to have higher body mass index (P = 0.04), alanine transaminase (P = 0.0001), alkaline phosphatase (P = 0.0009), prothrombin time (P = 0.0005), and maximal vital capacity (VCmax) (P = 0.04); lower platelet count (P = 0.007); abnormal liver echogenicity (P < 0.001); and splenomegaly (P = 0.001) at ultrasound. Screening with liver ultrasound provided a sensitivity and negative predictive value for severe fibrosis or cirrhosis of 100%, as were the specificity and positive predictive value for platelet count less than or equal to 174,000 per mm(3) and splenomegaly. Among individuals undergoing liver biopsy, fibrosis stage correlated with increasing VCmax (P = 0.02) and % predicted VCmax (P = 0.05), and decreasing residual volume/total lung capacity (TLC) (P = 0.02) and % predicted residual volume/TLC (P = 0.05). CONCLUSIONS: Significant chronic liver disease is common in PiZZ individuals with lung disease and can be screened effectively by a combination of conventional tests of liver function, platelet count, and liver ultrasound.


Assuntos
Cirrose Hepática/diagnóstico , Cirrose Hepática/epidemiologia , Fenótipo , Doença Pulmonar Obstrutiva Crônica/epidemiologia , Deficiência de alfa 1-Antitripsina/epidemiologia , Adulto , Biomarcadores , Doença Crônica , Comorbidade , Feminino , Humanos , Cirrose Hepática/diagnóstico por imagem , Testes de Função Hepática , Masculino , Pessoa de Meia-Idade , Contagem de Plaquetas , Prevalência , Estudos Prospectivos , Fatores de Risco , Sensibilidade e Especificidade , Ultrassonografia , Reino Unido/epidemiologia
13.
J Hepatol ; 58(3): 549-56, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23142622

RESUMO

BACKGROUND & AIMS: Models of non-alcohol-related fatty liver disease (NAFLD) reveal features of accelerated ageing, such as impaired regeneration, and an increased risk of hepatocellular carcinoma. The relation between accelerated ageing, disease progression and clinical outcome has not been previously investigated and is the subject of the current study. METHODS: Liver sections from 70 patients with NAFLD (105 biopsies) and 60 controls were studied for telomere length, nuclear area, DNA damage and cell cycle phase markers, using quantitative fluorescent in situ hybridization and immunohistochemistry. RESULTS: Hepatocyte telomeres were shorter in NAFLD than controls (p <0.0001). Hepatocytes in NAFLD demonstrated lack of cell cycle progression beyond G1/S phase and high-level expression of p21, the universal cell cycle inhibitor (p=0.001). γ-H(2)AX expression increased with steatosis (p=0.01), indicating DNA damage, and was associated with shorter hepatocyte telomeres (p <0.0001). Hepatocyte p21 expression correlated with fibrosis stage and diabetes mellitus, independently (p <0.001 and p=0.002, respectively). Further analysis revealed that an adverse liver-related outcome was strongly associated with higher hepatocyte p21 expression and greater hepatocyte nuclear area (p=0.02 and p=0.006), but not with telomere length. In paired biopsies, changes in hepatocyte p21 expression and nuclear area mirrored changes in fibrosis stage (p=0.01 and p=0.006, respectively). CONCLUSIONS: These findings are consistent with hepatocyte senescence and permanent cell cycle arrest in NAFLD. Hepatocyte senescence correlated closely with fibrosis stage, diabetes mellitus, and clinical outcome. Hepatocyte p21 expression could be used as a prognostic marker and for stratification in clinical studies.


Assuntos
Senescência Celular , Fígado Gorduroso/patologia , Hepatócitos/patologia , Adulto , Idoso , Biópsia , Núcleo Celular/patologia , Inibidor de Quinase Dependente de Ciclina p21/análise , Progressão da Doença , Feminino , Histonas/análise , Humanos , Masculino , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica , Telômero
15.
Gastroenterology ; 140(7): 2056-63, 2063.e1, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21396368

RESUMO

BACKGROUND & AIMS: Patients with ferroportin iron overload due to loss-of-function mutations in SLC40A1 have macrophage iron overload, hyperferritinemia, and normal transferrin saturation. In contrast, hepatocellular iron storage, hyperferritinemia, and increased saturation of transferrin are a distinct clinical presentation of ferroportin iron overload that results from SLC40A1 mutations that confer resistance of ferroportin to hepcidin-mediated inactivation. METHODS: SLC40A1 was sequenced in patients from 2 independent pedigrees affected by hepatic iron overload unrelated to HFE. Functions of the ferroportin variants were tested in vitro. RESULTS: A patient heterozygous for the variant p.W158C in SLC40A1 presented with macrophage iron overload, hyperferritinemia, and normal transferrin saturation. A patient with hepatocellular iron storage, hyperferritinemia, and increased transferrin saturation was heterozygous for p.H507R. Expression of the p.W158C form of ferroportin in 293T cells resulted in defective trafficking to the plasma membrane and reduced iron export activity; the iron export activity of cells that expressed the p.H507R form of ferroportin did not differ from cells that expressed ferroportin without this mutation. The p.H507R of ferroportin localizes normally to the plasma membrane but is resistant to hepcidin-mediated inactivation. Addition of a synthetic peptide derived from ferroportin without these mutations (amino acids 500-518) decreased the inhibitory activity of hepcidin in cells, whereas a peptide from the same region, with p.H507R, had no effect on hepcidin activity. CONCLUSIONS: The variant p.W158C in SLC40A1 impairs intracellular trafficking of ferroportin, resulting in reduced iron export. The variant p.H507R does not bind hepcidin in vitro and results in apparent hepcidin resistance.


Assuntos
Proteínas de Transporte de Cátions/genética , Sobrecarga de Ferro/genética , Ferro/metabolismo , Fígado/metabolismo , Mutação , Adulto , Peptídeos Catiônicos Antimicrobianos/metabolismo , Biópsia , Estudos de Casos e Controles , Proteínas de Transporte de Cátions/sangue , Proteínas de Transporte de Cátions/metabolismo , Análise Mutacional de DNA , Ferritinas/metabolismo , Predisposição Genética para Doença , Células HEK293 , Hepcidinas , Humanos , Ferro/sangue , Sobrecarga de Ferro/sangue , Sobrecarga de Ferro/metabolismo , Macrófagos/metabolismo , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Linhagem , Fenótipo , Transporte Proteico , Transfecção , Transferrina/metabolismo
16.
Pancreatology ; 12(1): 8-15, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22487467

RESUMO

OBJECTIVE: Current practice to diagnose pancreatic cancer is accomplished by endoscopic ultrasound guided fine needle aspiration (EUS-FNA) using a cytological approach. This method is time consuming and often fails to provide suitable specimens for modern molecular analyses. Here, we compare the cytological approach with direct formalin fixation of pancreatic EUS-FNA micro-cores and evaluate the potential to perform molecular biomarker analysis on these specimen. METHODS: 130 specimens obtained by EUS-FNA with a 22G needle were processed by the standard cytological approach and compared to a separate cohort of 130 specimens that were immediately formalin fixed to preserve micro-cores of tissue prior to routine histological processing. RESULTS: We found that direct formalin fixation significantly shortened the time required for diagnosis from 3.6 days to 2.9 days (p<0.05) by reducing the average time (140 vs 33 min/case) and number of slides (9.65 vs 4.67 slides/case) for histopathological processing. Specificity and sensitivity yielded comparable results between the two approaches (82.3% vs 77% and 90.9% vs 100%). Importantly, EUS-FNA histology preserved the tumour tissue architecture with neoplastic glands embedded in stroma in 67.89% of diagnostic cases compared to 27.55% with the standard cytological approach (p < 0.001). Furthermore, micro-core samples were suitable for molecular studies including the immunohistochemical detection of intranuclear Hes1 in malignant cells, and the laser-capture microdissection-mediated measurement of Gli-1 mRNA in tumour stromal myofibroblasts. CONCLUSIONS: Direct formalin fixation of pancreatic EUS-FNA micro-cores demonstrates superiority regarding diagnostic delay, costs, and specimen suitability for molecular studies. We advocate this approach for future investigational trials in pancreatic cancer patients.


Assuntos
Biomarcadores Tumorais/análise , Biópsia por Agulha Fina/métodos , Endossonografia/métodos , Neoplasias Pancreáticas/diagnóstico por imagem , Adulto , Idoso , Idoso de 80 Anos ou mais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/análise , Feminino , Fixadores , Formaldeído , Proteínas de Homeodomínio/análise , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/patologia , Estudos Retrospectivos , Sensibilidade e Especificidade , Fatores de Tempo , Fatores de Transcrição HES-1 , Ultrassonografia de Intervenção
17.
Eur Radiol ; 22(12): 2790-7, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22752441

RESUMO

OBJECTIVES: Conventional imaging techniques are insensitive to liver fibrosis. This study assesses the diagnostic accuracy of MR elastography (MRE) stiffness values and the ratio of phosphomonoesters (PME)/phosphodiesters (PDE) measured using (31)P spectroscopy against histological fibrosis staging. METHODS: The local research ethics committee approved this prospective, blinded study. A total of 77 consecutive patients (55 male, aged 49 ± 11.5 years) with a clinical suspicion of liver fibrosis underwent an MR examination with a liver biopsy later the same day. Patients underwent MRE and (31)P spectroscopy on a 1.5 T whole body system. The liver biopsies were staged using an Ishak score for chronic hepatitis or a modified NAS fibrosis score for fatty liver disease. RESULTS: MRE increased with and was positively associated with fibrosis stage (Spearman's rank = 0.622, P < 0.001). PME/PDE was not associated with fibrosis stage (Spearman's rank = -0.041, p = 0.741). Area under receiver operating curves for MRE stiffness values were high (range 0.75-0.97). The diagnostic utility of PME/PDE was no better than chance (range 0.44-0.58). CONCLUSIONS: MRE-estimated liver stiffness increases with fibrosis stage and is able to dichotomise fibrosis stage groupings. We did not find a relationship between (31)P MR spectroscopy and fibrosis stage. KEY POINTS: Magnetic resonance elastography (MRE) and MR spectroscopy can both assess the liver. MRE is superior to ( 31 ) P MR spectroscopy in staging hepatic fibrosis. MRE is able to dichotomise liver fibrosis stage groupings. Gradient-echo MRE may be problematic in genetic haemochromatosis.


Assuntos
Técnicas de Imagem por Elasticidade/métodos , Cirrose Hepática/patologia , Espectroscopia de Ressonância Magnética/métodos , Biópsia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Curva ROC
18.
Mol Metab ; 48: 101210, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33722690

RESUMO

OBJECTIVE: Non-alcoholic fatty liver disease (NAFLD) is a silent pandemic associated with obesity and the metabolic syndrome, and also increases cardiovascular- and cirrhosis-related morbidity and mortality. A complete understanding of adaptive compensatory metabolic programmes that modulate non-alcoholic steatohepatitis (NASH) progression is lacking. METHODS AND RESULTS: Transcriptomic analysis of liver biopsies in patients with NASH revealed that NASH progression is associated with rewiring of metabolic pathways, including upregulation of de novo lipid/cholesterol synthesis and fatty acid remodelling. The modulation of these metabolic programmes was achieved by activating sterol regulatory element-binding protein (SREBP) transcriptional networks; however, it is still debated whether, in the context of NASH, activation of SREBPs acts as a pathogenic driver of lipotoxicity, or rather promotes the biosynthesis of protective lipids that buffer excessive lipid accumulation, preventing inflammation and fibrosis. To elucidate the pathophysiological role of SCAP/SREBP in NASH and wound-healing response, we used an Insig1 deficient (with hyper-efficient SREBPs) murine model challenged with a NASH-inducing diet. Despite enhanced lipid and cholesterol biosynthesis, Insig1 KO mice had similar systemic metabolism and insulin sensitivity to Het/WT littermates. Moreover, activating SREBPs resulted in remodelling the lipidome, decreased hepatocellular damage, and improved wound-healing responses. CONCLUSIONS: Our study provides actionable knowledge about the pathways and mechanisms involved in NAFLD pathogenesis, which may prove useful for developing new therapeutic strategies. Our results also suggest that the SCAP/SREBP/INSIG1 trio governs transcriptional programmes aimed at protecting the liver from lipotoxic insults in NASH.


Assuntos
Colesterol/biossíntese , Progressão da Doença , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lipogênese/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Animais , Biomarcadores/metabolismo , Dieta Ocidental , Feminino , Humanos , Resistência à Insulina/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Cirrose Hepática/genética , Cirrose Hepática/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica/genética , Transcriptoma
19.
Science ; 371(6531): 839-846, 2021 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-33602855

RESUMO

Organoid technology holds great promise for regenerative medicine but has not yet been applied to humans. We address this challenge using cholangiocyte organoids in the context of cholangiopathies, which represent a key reason for liver transplantation. Using single-cell RNA sequencing, we show that primary human cholangiocytes display transcriptional diversity that is lost in organoid culture. However, cholangiocyte organoids remain plastic and resume their in vivo signatures when transplanted back in the biliary tree. We then utilize a model of cell engraftment in human livers undergoing ex vivo normothermic perfusion to demonstrate that this property allows extrahepatic organoids to repair human intrahepatic ducts after transplantation. Our results provide proof of principle that cholangiocyte organoids can be used to repair human biliary epithelium.


Assuntos
Doenças dos Ductos Biliares/terapia , Ductos Biliares Intra-Hepáticos/fisiologia , Ductos Biliares/citologia , Terapia Baseada em Transplante de Células e Tecidos , Células Epiteliais/citologia , Organoides/transplante , Animais , Bile , Ductos Biliares/fisiologia , Ductos Biliares Intra-Hepáticos/citologia , Ducto Colédoco/citologia , Células Epiteliais/fisiologia , Vesícula Biliar/citologia , Regulação da Expressão Gênica , Humanos , Fígado/fisiologia , Transplante de Fígado , Transplante de Células-Tronco Mesenquimais , Camundongos , Organoides/fisiologia , RNA-Seq , Obtenção de Tecidos e Órgãos , Transcriptoma
20.
J Hepatol ; 53(2): 252-60, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20462651

RESUMO

BACKGROUND & AIMS: Increasing age is associated with impaired immune function and in chronic HCV infection specifically, with progressive fibrosis, liver failure, HCC and impaired responses to antiviral therapy. T-lymphocyte telomere length declines with age. We hypothesised that shorter T-lymphocyte telomere length would be associated with poor clinical outcome in HCV infection. METHODS: Circulating T-lymphocyte telomere length, an objective measure of immune senescence, was measured by flow-FISH in 135 HCV-RNA-positive, treatment-naïve patients and 41 healthy controls in relation to clinical outcome. RESULTS: Shorter CD4+CD45RO+ T-lymphocyte telomeres were associated with severe fibrosis (p=0.003), independent of male sex (p=0.04), CMV positivity (p=0.003), previous HBV infection (p=0.007), and age (p=ns) in viraemic patients compared to controls. There were inverse correlations between CD4+CD45RO+ telomere length and fibrosis stage (p<0.001), portal tract inflammatory grade (p=0.035), prothrombin time (p<0.001) and bilirubin (p=0.001). One hundred and twenty-four viraemic individuals were followed prospectively to a composite endpoint of death, hepatic decompensation or HCC. Independent of age, those with shorter CD4+CD45RO+ telomeres were less likely to be complication free after 2-years than those with longer telomeres (86% versus 96%, p=0.009) with an age-adjusted hazard ratio of 0.93 (0.90-0.96). In addition, CD4+CD45RO+ telomere length predicted successful antiviral therapy (p=0.001) independent of other factors. CONCLUSIONS: CD4+ T-lymphocyte telomere length, independent of age, was related to inflammatory grade, fibrosis stage, laboratory indices of severity, subsequent hepatic decompensation and treatment outcome in patients with chronic HCV infection.


Assuntos
Antivirais/uso terapêutico , Linfócitos T CD4-Positivos/patologia , Hepacivirus , Hepatite C Crônica/tratamento farmacológico , Hepatite C Crônica/patologia , Cirrose Hepática/patologia , Telômero/patologia , Adulto , Biópsia , Estudos de Casos e Controles , Feminino , Hepatite C Crônica/diagnóstico , Humanos , Fígado/patologia , Masculino , Pessoa de Meia-Idade , Fenótipo , Prognóstico , Análise de Regressão , Índice de Gravidade de Doença , Resultado do Tratamento
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