ABSTRACT
BACKGROUND: The transition from steatosis to non-alcoholic steatohepatitis (NASH) is a key issue in non-alcoholic fatty liver disease (NAFLD). Observations in patients with obstructive sleep apnea syndrome (OSAS) suggest that hypoxia contributes to progression to NASH and liver fibrosis, and the release of extracellular vesicles (EVs) by injured hepatocytes has been implicated in NAFLD progression. AIM: To evaluate the effects of hypoxia on hepatic pro-fibrotic response and EV release in experimental NAFLD and to assess cellular crosstalk between hepatocytes and human hepatic stellate cells (LX-2). METHODS: HepG2 cells were treated with fatty acids and subjected to chemically induced hypoxia using the hypoxia-inducible factor 1 alpha (HIF-1α) stabilizer cobalt chloride (CoCl2). Lipid droplets, oxidative stress, apoptosis and pro-inflammatory and pro-fibrotic-associated genes were assessed. EVs were isolated by ultracentrifugation. LX-2 cells were treated with EVs from hepatocytes. The CDAA-fed mouse model was used to assess the effects of intermittent hypoxia (IH) in experimental NASH. RESULTS: Chemical hypoxia increased steatosis, oxidative stress, apoptosis and pro-inflammatory and pro-fibrotic gene expressions in fat-laden HepG2 cells. Chemical hypoxia also increased the release of EVs from HepG2 cells. Treatment of LX2 cells with EVs from fat-laden HepG2 cells undergoing chemical hypoxia increased expression pro-fibrotic markers. CDAA-fed animals exposed to IH exhibited increased portal inflammation and fibrosis that correlated with an increase in circulating EVs. CONCLUSION: Chemical hypoxia promotes hepatocellular damage and pro-inflammatory and pro-fibrotic signaling in steatotic hepatocytes both in vitro and in vivo. EVs from fat-laden hepatocytes undergoing chemical hypoxia evoke pro-fibrotic responses in LX-2 cells.
Subject(s)
Extracellular Vesicles/metabolism , Hypoxia/pathology , Liver Cirrhosis/pathology , Non-alcoholic Fatty Liver Disease/pathology , Sleep Apnea, Obstructive/complications , Animals , Cell Communication , Cell Hypoxia/drug effects , Cobalt/pharmacology , Culture Media/metabolism , Disease Models, Animal , Disease Progression , Fatty Acids, Nonesterified/metabolism , Hep G2 Cells , Hepatic Stellate Cells/metabolism , Hepatocytes/metabolism , Humans , Hypoxia/blood , Hypoxia/etiology , Hypoxia-Inducible Factor 1, alpha Subunit/agonists , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Liver Cirrhosis/blood , Liver Cirrhosis/etiology , Male , Mice , Non-alcoholic Fatty Liver Disease/blood , Non-alcoholic Fatty Liver Disease/etiology , Oxidative Stress , Sleep Apnea, Obstructive/bloodABSTRACT
BACKGROUND: Obstructive sleep apnea syndrome (OSAS) is associated to intermittent hypoxia (IH) and is an aggravating factor of non-alcoholic fatty liver disease (NAFLD). We investigated the effects of hypoxia in both in vitro and in vivo models of NAFLD. METHODS: Primary rat hepatocytes treated with free fatty acids (FFA) were subjected to chemically induced hypoxia (CH) using the hypoxia-inducible factor-1 alpha (HIF-1α) stabilizer cobalt chloride (CoCl2). Triglyceride (TG) content, mitochondrial superoxide production, cell death rates, cytokine and inflammasome components gene expression and protein levels of cleaved caspase-1 were assessed. Also, Kupffer cells (KC) were treated with conditioned medium (CM) and extracellular vehicles (EVs) from hypoxic fat-laden hepatic cells. The choline deficient L-amino acid defined (CDAA)-feeding model used to assess the effects of IH on experimental NAFLD in vivo. RESULTS: Hypoxia induced HIF-1α in cells and animals. Hepatocytes exposed to FFA and CoCl2 exhibited increased TG content and higher cell death rates as well as increased mitochondrial superoxide production and mRNA levels of pro-inflammatory cytokines and of inflammasome-components interleukin-1ß, NLRP3 and ASC. Protein levels of cleaved caspase-1 increased in CH-exposed hepatocytes. CM and EVs from hypoxic fat-laden hepatic cells evoked a pro-inflammatory phenotype in KC. Livers from CDAA-fed mice exposed to IH exhibited increased mRNA levels of pro-inflammatory and inflammasome genes and increased levels of cleaved caspase-1. CONCLUSION: Hypoxia promotes inflammatory signals including inflammasome/caspase-1 activation in fat-laden hepatocytes and contributes to cellular crosstalk with KC by release of EVs. These mechanisms may underlie the aggravating effect of OSAS on NAFLD. [Abstract word count: 257].
Subject(s)
Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Hypoxia/genetics , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Non-alcoholic Fatty Liver Disease/genetics , Sleep Apnea, Obstructive/genetics , Animals , Caspase 1/genetics , Choline Deficiency/genetics , Choline Deficiency/metabolism , Choline Deficiency/pathology , Cobalt/toxicity , Disease Models, Animal , Extracellular Vesicles/genetics , Extracellular Vesicles/metabolism , Extracellular Vesicles/pathology , Fatty Acids, Nonesterified/pharmacology , Gene Expression Regulation/drug effects , Hepatocytes/metabolism , Hepatocytes/pathology , Humans , Hypoxia/chemically induced , Hypoxia/metabolism , Hypoxia/pathology , Inflammasomes/genetics , Inflammation/chemically induced , Inflammation/genetics , Inflammation/metabolism , Inflammation/pathology , Interleukin-1beta/genetics , Kupffer Cells/metabolism , Kupffer Cells/pathology , Mice , Non-alcoholic Fatty Liver Disease/complications , Non-alcoholic Fatty Liver Disease/metabolism , Non-alcoholic Fatty Liver Disease/pathology , Rats , Sleep Apnea, Obstructive/etiology , Sleep Apnea, Obstructive/metabolism , Sleep Apnea, Obstructive/pathology , Triglycerides/geneticsABSTRACT
Se presenta la casuística de la colecistectomía laparoscópica (CL) del Hospital Militar en la patología vesicular, revisando los resultados obtenidos en 1.102 pacientes en un lapso de 23 meses. El 22 por ciento de los pacientes son portadores de un cuadro vesicular agudo. El 10 por ciento de las CL recibieron una colangiografía endoscópica retrógrada (CER) preoperatoria en búsqueda de coledocolitiasis, encontrándola en un tercio de ellos. Durante la intervención se detecta un 1,8 por ciento de coledocolitiasis, llegando a un 5,3 por ciento de cálculos coledocianos en toda la serie. En el 1,6 por ciento de la serie se realizó exploración coledociana trancística, lo que permite resolver el 40 por ciento de los hallazgos de coledocolitiasis intraopereratoria. Se convirtió la técnica en el 6 por ciento de las CL, existiendo 3,8 por ciento de complicaciones médicas, 3,5 por ciento de complicaciones quirúrgicas, con 0,7 por ciento de reoperaciones, 06 por ciento rehospitalizaciones 0,18 por ciento de sección del colédoco y 0,09 por ciento de mortalidad. Las cifras refuerzan sólidamente el concepto de que la CL es la técnica quirúrgica para tratar la patología vesicular no neoplásica, con un adecuado margen de seguridad, buena recuperación postoperatoria y rápida rehabilitación
