RESUMEN
PURPOSE: To establish a new rat model, the pathogenesis of which is closer to the clinical occurrence of chronic obstructive jaundice with liver fibrosis. METHODS: 90 SD rats were randomly divided into 3 groups. Group A common bile duct ligation, group B common bile duct injection compont and group C injection saline. The serum of three groups was extracted, and the liver function was detected by ELISA. HE staining, Masson staining and immunohistochemistry were used to detect liver pathology. RESULTS: Group B showed a fluctuant development of jaundice, obstructive degree reached a peak at 2 weeks, and decreased from 3 weeks. HA, LA and PCIII were significantly higher than control group. 3 weeks after surgery, liver tissue fibrosis occurred in group B, and a wide range of fiber spacing was formed at 5 weeks. Immunohistochemistry showed that hepatic stellate cells were more active than the control group. CONCLUSION: Intra-biliary injection of Compont gel is different from the classic obstructive jaundice animal model caused by classic bile duct ligation, which can provide an ideal rat model of chronic obstructive jaundice with liver fibrosis.
Asunto(s)
Conductos Biliares/efectos de los fármacos , Modelos Animales de Enfermedad , Geles/administración & dosificación , Ictericia Obstructiva/inducido químicamente , Cirrosis Hepática/inducido químicamente , Fosfatasa Alcalina/sangre , Animales , Aspartato Aminotransferasas/sangre , Compuestos Azo , Conductos Biliares/patología , Bilirrubina/análisis , Ensayo de Inmunoadsorción Enzimática , Eosina Amarillenta-(YS) , Femenino , Inmunohistoquímica , Inyecciones , Ictericia Obstructiva/patología , Cirrosis Hepática/patología , Verde de Metilo , Distribución Aleatoria , Ratas Sprague-Dawley , Valores de Referencia , Reproducibilidad de los Resultados , Albúmina Sérica/análisis , Factores de Tiempo , gamma-Glutamiltransferasa/sangreRESUMEN
Clinical conditions that result in endotoxemia, such as sepsis and alcoholic hepatitis (AH), often are accompanied by cholestasis. Although hepatocellular changes in response to lipopolysaccharide (LPS) have been well characterized, less is known about whether and how cholangiocytes contribute to this form of cholestasis. We examined effects of endotoxin on expression and function of the type 3 inositol trisphosphate receptor (ITPR3), because this is the main intracellular Ca2+ release channel in cholangiocytes, and loss of it impairs ductular bicarbonate secretion. Bile duct cells expressed the LPS receptor, Toll-like receptor 4 (TLR4), which links to activation of nuclear factor-κB (NF-κB). Analysis of the human ITPR3 promoter revealed five putative response elements to NF-κB, and promoter activity was inhibited by p65/p50. Nested 0.5- and 1.0-kilobase (kb) deletion fragments of the ITPR3 promoter were inhibited by NF-κB subunits. Chromatin immunoprecipitation (ChIP) assay showed that NF-κB interacts with the ITPR3 promoter, with an associated increase in H3K9 methylation. LPS decreased ITPR3 mRNA and protein expression and also decreased sensitivity of bile duct cells to calcium agonist stimuli. This reduction was reversed by inhibition of TLR4. ITPR3 expression was decreased or absent in cholangiocytes from patients with cholestasis of sepsis and from those with severe AH. Conclusion: Stimulation of TLR4 by LPS activates NF-κB to down-regulate ITPR3 expression in human cholangiocytes. This may contribute to the cholestasis that can be observed in conditions such as sepsis or AH.
Asunto(s)
Conductos Biliares/efectos de los fármacos , Conductos Biliares/metabolismo , Endotoxemia/metabolismo , Endotoxinas/toxicidad , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Adulto , Señalización del Calcio/efectos de los fármacos , Colestasis/etiología , Colestasis/metabolismo , Endotoxemia/complicaciones , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Hepatitis Alcohólica/metabolismo , Humanos , Receptores de Inositol 1,4,5-Trifosfato/genética , Masculino , Persona de Mediana Edad , FN-kappa B/metabolismoRESUMEN
Abstract Purpose: To establish a new rat model, the pathogenesis of which is closer to the clinical occurrence of chronic obstructive jaundice with liver fibrosis. Methods: 90 SD rats were randomly divided into 3 groups. Group A common bile duct ligation, group B common bile duct injection compont and group C injection saline. The serum of three groups was extracted, and the liver function was detected by ELISA. HE staining, Masson staining and immunohistochemistry were used to detect liver pathology. Results: Group B showed a fluctuant development of jaundice, obstructive degree reached a peak at 2 weeks, and decreased from 3 weeks. HA, LA and PCIII were significantly higher than control group. 3 weeks after surgery, liver tissue fibrosis occurred in group B, and a wide range of fiber spacing was formed at 5 weeks. Immunohistochemistry showed that hepatic stellate cells were more active than the control group. Conclusion: Intra-biliary injection of Compont gel is different from the classic obstructive jaundice animal model caused by classic bile duct ligation, which can provide an ideal rat model of chronic obstructive jaundice with liver fibrosis.
Asunto(s)
Animales , Femenino , Conductos Biliares/efectos de los fármacos , Modelos Animales de Enfermedad , Geles/administración & dosificación , Cirrosis Hepática/inducido químicamente , Aspartato Aminotransferasas/sangre , Valores de Referencia , Compuestos Azo , Factores de Tiempo , Conductos Biliares/patología , Bilirrubina/análisis , Albúmina Sérica/análisis , Ensayo de Inmunoadsorción Enzimática , Inmunohistoquímica , Distribución Aleatoria , Reproducibilidad de los Resultados , Ratas Sprague-Dawley , Eosina Amarillenta-(YS) , Ictericia Obstructiva/inducido químicamente , Ictericia Obstructiva/patología , Fosfatasa Alcalina/sangre , gamma-Glutamiltransferasa/sangre , Inyecciones , Cirrosis Hepática/patología , Verde de MetiloRESUMEN
Bile acids (BA) are key molecules in generating bile flow, which is an essential function of the liver. In the last decades there have been great advances in the understanding of the role of a number of specific transport proteins present at the sinusoidal and canalicular membrane domains of hepatocytes and cholangiocytes in generating and maintaining bile flow. Also, a clearer understanding on how BA regulate their own synthesis and the expression and/or function of transporters has been reached. This new knowledge has helped to better delineate the pathophysiology of cholestasis and the adaptive responses of hepatocytes to cholestatic liver injury as well as of the mechanisms of injury of biliary epithelia. In this context, therapeutic approaches including new hydrophilic BA such as the conjugation-resistant nor- ursodeoxycholic acid, nuclear receptors (FXR, PPAR-alpha) agonists, FGF19 analogues, inhibitors of the apical sodium-dependent bile acid transporter [ASBT] and modulators of the inflammatory cascade triggered by BAs are being studied as novel treatments of cholestasis. In the present review we summarize recent experimental and clinical data on the role of BAs in cholestasis and its treatment.
Asunto(s)
Ácidos y Sales Biliares/uso terapéutico , Conductos Biliares/efectos de los fármacos , Colestasis/tratamiento farmacológico , Fármacos Gastrointestinales/uso terapéutico , Hígado/efectos de los fármacos , Animales , Ácidos y Sales Biliares/metabolismo , Conductos Biliares/inmunología , Conductos Biliares/metabolismo , Conductos Biliares/patología , Colestasis/inmunología , Colestasis/metabolismo , Colestasis/patología , Humanos , Inmunidad Innata/efectos de los fármacos , Hígado/inmunología , Hígado/metabolismo , Hígado/patología , Transducción de Señal/efectos de los fármacosRESUMEN
Biliary atresia (BA) is a destructive bile duct disease occurring in newborn children within a few weeks after birth. In this study, the effect of miR-29c and miR-129-5p on epithelial-mesenchymal transition (EMT) in experimental BA was explored by constructing BA mouse models via Rhesus rotavirus vaccine infection. miR-29c and miR-129-5p expression was analyzed by real-time quantitative polymerase chain reaction. EMT was established by induction with transforming growth factor (TGF)-ß1. miR-29c and miR-129-5p were overexpressed and inhibited, respectively, by Lipofectamine transfection. EMT-related protein (formin-like 2, FMNL2; E-cadherin; vimentin; and cytokeratin-19, CK-19) expression was analyzed by western blot and immunofluorescent assay. The results indicated that miR-29c and miR-129-5p were downregulated and upregulated in BA mice. TGF-ß1 induction caused a time-dependent decrease and increase in miR-29c and miR-129-5p, respectively. Additionally, TGF-ß1 induced an increase in FMNL2 and vimentin expression and a decrease in E-cadherin and CK-19 expression (P < 0.05). Overexpression or suppression of miRNA-29c or miR-129-5p, respectively, induced the inhibition of FMNL2 and vimentin, and promotion of E-cadherin and CK-19 expression, in the test groups compared to the non-intervention group (P < 0.05). However, the FMNL2, vimentin, E-cadherin, and CK- 19 expression did not differ between the control and non-intervention groups (P > 0.05). Thus, miR-29c upregulation or miR-129-5p downregulation effectively prevented EMT in BA by regulating the expression of EMT pathway-related proteins. Therefore, miR-29c and miR-129-5p could be utilized as therapeutic targets for BA in the future.
Asunto(s)
Atresia Biliar/genética , Células Epiteliales/efectos de los fármacos , Transición Epitelial-Mesenquimal/efectos de los fármacos , MicroARNs/genética , Factor de Crecimiento Transformador beta1/farmacología , Animales , Animales Recién Nacidos , Conductos Biliares/efectos de los fármacos , Conductos Biliares/metabolismo , Conductos Biliares/patología , Atresia Biliar/etiología , Atresia Biliar/metabolismo , Atresia Biliar/patología , Cadherinas/genética , Cadherinas/metabolismo , Modelos Animales de Enfermedad , Células Epiteliales/metabolismo , Células Epiteliales/patología , Femenino , Forminas , Regulación de la Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Queratina-19/genética , Queratina-19/metabolismo , Ratones , Ratones Endogámicos BALB C , MicroARNs/metabolismo , Cultivo Primario de Células , Vacunas contra Rotavirus/administración & dosificación , Vacunas contra Rotavirus/efectos adversos , Transducción de Señal , Vimentina/genética , Vimentina/metabolismoRESUMEN
Schistosomiasis is a major cause of portal hypertension worldwide. It associates with portal fibrosis that develops during chronic infection. The mechanisms by which the pathogen evokes these host responses remain unclear. We evaluated the hypothesis that schistosome eggs release factors that directly stimulate liver cells to produce osteopontin (OPN), a pro-fibrogenic protein that stimulates hepatic stellate cells to become myofibroblasts. We also investigated the utility of OPN as a biomarker of fibrosis and/or severity of portal hypertension. Cultured cholangiocytes, Kupffer cells and hepatic stellate cells were treated with soluble egg antigen (SEA); OPN production was quantified by quantitative reverse transcriptase polymerase chain reaction (qRTPCR) and ELISA; cell proliferation was assessed by BrdU (5-bromo-2'-deoxyuridine). Mice were infected with Schistosoma mansoni for 6 or 16 weeks to cause early or advanced fibrosis. Liver OPN was evaluated by qRTPCR and immunohistochemistry (IHC) and correlated with liver fibrosis and serum OPN. Livers from patients with schistosomiasis mansoni (early fibrosis n=15; advanced fibrosis n=72) or healthy adults (n=22) were immunostained for OPN and fibrosis markers. Results were correlated with plasma OPN levels and splenic vein pressures. SEA-induced cholangiocyte proliferation and OPN secretion (P<0.001 compared with controls). Cholangiocytes were OPN (+) in Schistosoma-infected mice and humans. Liver and serum OPN levels correlated with fibrosis stage (mice: r=0.861; human r=0.672, P=0.0001) and myofibroblast accumulation (mice: r=0.800; human: r=0.761, P=0.0001). Numbers of OPN (+) bile ductules strongly correlated with splenic vein pressure (r=0.778; P=0.001). S. mansoni egg antigens stimulate cholangiocyte proliferation and OPN secretion. OPN levels in liver and blood correlate with fibrosis stage and portal hypertension severity.
Asunto(s)
Proliferación Celular , Hipertensión Portal/metabolismo , Cirrosis Hepática/metabolismo , Osteopontina/metabolismo , Esquistosomiasis mansoni/metabolismo , Adolescente , Adulto , Animales , Antígenos Helmínticos/farmacología , Conductos Biliares/citología , Conductos Biliares/efectos de los fármacos , Conductos Biliares/metabolismo , Línea Celular , Células Cultivadas , Femenino , Células Estrelladas Hepáticas/efectos de los fármacos , Células Estrelladas Hepáticas/metabolismo , Interacciones Huésped-Parásitos , Humanos , Hipertensión Portal/genética , Hipertensión Portal/parasitología , Inmunohistoquímica , Macrófagos del Hígado/efectos de los fármacos , Macrófagos del Hígado/metabolismo , Cirrosis Hepática/genética , Cirrosis Hepática/parasitología , Masculino , Ratones , Persona de Mediana Edad , Osteopontina/sangre , Osteopontina/genética , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Schistosoma/fisiología , Esquistosomiasis mansoni/genética , Esquistosomiasis mansoni/parasitología , Adulto JovenRESUMEN
BACKGROUND: MUC2 and MUC5AC overproduction is considered to be associated with hepatolithiasis and related to inflammation. However, mechanisms underlying MUC upregulation under inflammatory stimulation in human intrahepatic biliary epithelial cells (HIBECs) are not completely understood. MATERIAL AND METHODS: Expression of MUC2 and MUC5AC mRNA in HIBECs was detected by real-time PCR. Expression of COX-2, EP4, and phosphorylated ERK, JNK and p38MAPK protein was detected by Western blot. Concentrations of PGE2, IL-1ß and TNF-α in cell culture supernatants were measured using the Quantikine Elisa kit. RESULTS: COX-2 expression as well as PGE2 production in HIBECs was upregulated significantly by LPS, which was completely blocked by either TLR4 antagonist or NFκB inhibitor. Selective COX-2 inhibitor suppressed LPS-induced MUC2 and MUC5AC mRNA expression remarkably. Exogenous PGE2 increased MUC2 and MUC5AC mRNA expression in a dosage-dependent manner independent of IL-1ß and TNF-α. PGE2 receptor EP4 agonist elevated MUC2 and MUC5AC expression, whereas EP4 antagonist had the opposite effect. Expression of phosphorylated p38MAPK was upregulated by exogenous PGE2, and p38MAPK inhibitor reduced MUC2 and MUC5AC expression in HIBECs. In addition, it was found that levels of PGE2, MUC2 and MUC5AC in bile samples from the hepatic ducts affected by intrahepatic stones were significantly higher than those from the unaffected hepatic ducts of patients with hepatolithiasis. CONCLUSIONS: Our findings indicate that PGE2 induces MUC2 and MUC5AC expression in HIBECs via EP4-p38MAPK signaling.
Asunto(s)
Conductos Biliares/efectos de los fármacos , Dinoprostona/farmacología , Células Epiteliales/efectos de los fármacos , Mucina 5AC/metabolismo , Mucina 2/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E/agonistas , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Bilis/metabolismo , Conductos Biliares/enzimología , Conductos Biliares/inmunología , Western Blotting , Células Cultivadas , Colelitiasis/metabolismo , Colelitiasis/patología , Ciclooxigenasa 2/metabolismo , Inhibidores de la Ciclooxigenasa 2/farmacología , Dinoprostona/metabolismo , Relación Dosis-Respuesta a Droga , Activación Enzimática , Ensayo de Inmunoadsorción Enzimática , Células Epiteliales/enzimología , Células Epiteliales/inmunología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Mediadores de Inflamación/metabolismo , Interleucina-1beta/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Lipopolisacáridos/farmacología , Mucina 5AC/genética , Mucina 2/genética , FN-kappa B/metabolismo , Fosforilación , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Transducción de Señal/efectos de los fármacos , Receptor Toll-Like 4/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Regulación hacia ArribaRESUMEN
OBJECTIVE: The aim of this study was to evaluate the effect of oral tamoxifen treatment on the number of myofibroblasts present during the healing process after experimental bile duct injury. METHODS: The sample consisted of 16 pigs that were divided into two groups (the control and study groups). Incisions and suturing of the bile ducts were performed in the two groups. Tamoxifen (20 mg/day) was administered only to the study group. The animals were sacrificed after 30 days. Quantification of myofibroblasts in the biliary ducts was made through immunohistochemistry analysis using anti-alpha smooth muscle actin of the smooth muscle antibody. Immunohistochemical quantification was performed using a digital image system. RESULTS: In the animals treated with tamoxifen (20 mg/day), there was a significant reduction in immunostaining for alpha smooth muscle actin compared with the control group (0.1155 vs. 0.2021, p = 0.046). CONCLUSION: Tamoxifen reduced the expression of alpha smooth muscle actin in the healing tissue after bile duct injury, suggesting a decrease in myofibroblasts in the scarred area of the pig biliary tract. These data suggest that tamoxifen could be used in the prevention of biliary tract stenosis after bile duct surgeries.
Asunto(s)
Conductos Biliares/lesiones , Antagonistas de Estrógenos/uso terapéutico , Miofibroblastos/efectos de los fármacos , Tamoxifeno/uso terapéutico , Cicatrización de Heridas/efectos de los fármacos , Actinas/análisis , Actinas/efectos de los fármacos , Animales , Conductos Biliares/efectos de los fármacos , Recuento de Células , Femenino , Inmunohistoquímica , Músculo Liso/química , Músculo Liso/efectos de los fármacos , Reproducibilidad de los Resultados , Porcinos , Resultado del TratamientoRESUMEN
OBJECTIVE: The aim of this study was to evaluate the effect of oral tamoxifen treatment on the number of myofibroblasts present during the healing process after experimental bile duct injury. METHODS: The sample consisted of 16 pigs that were divided into two groups (the control and study groups). Incisions and suturing of the bile ducts were performed in the two groups. Tamoxifen (20 mg/day) was administered only to the study group. The animals were sacrificed after 30 days. Quantification of myofibroblasts in the biliary ducts was made through immunohistochemistry analysis using anti-alpha smooth muscle actin of the smooth muscle antibody. Immunohistochemical quantification was performed using a digital image system. RESULTS: In the animals treated with tamoxifen (20 mg/day), there was a significant reduction in immunostaining for alpha smooth muscle actin compared with the control group (0.1155 vs. 0.2021, p = 0.046). CONCLUSION: Tamoxifen reduced the expression of alpha smooth muscle actin in the healing tissue after bile duct injury, suggesting a decrease in myofibroblasts in the scarred area of the pig biliary tract. These data suggest that tamoxifen could be used in the prevention of biliary tract stenosis after bile duct surgeries.
Asunto(s)
Animales , Femenino , Conductos Biliares/lesiones , Antagonistas de Estrógenos/uso terapéutico , Miofibroblastos/efectos de los fármacos , Tamoxifeno/uso terapéutico , Cicatrización de Heridas/efectos de los fármacos , Actinas/análisis , Actinas/efectos de los fármacos , Conductos Biliares/efectos de los fármacos , Recuento de Células , Inmunohistoquímica , Músculo Liso/química , Músculo Liso/efectos de los fármacos , Reproducibilidad de los Resultados , Porcinos , Resultado del TratamientoRESUMEN
OBJECTIVE: To determine whether serum levels of unconjugated bilirubin (UCB) can be decreased by enhancing fecal fat excretion. STUDY DESIGN: Gunn rats were fed a high-fat diet (control) or the same diet mixed with the lipase inhibitor orlistat. At regular intervals, plasma UCB concentrations were determined and 72-hour fat balances were performed. RESULTS: Orlistat treatment decreased plasma UCB concentrations (at 3 weeks; 100 mg/kg, -33%+/-8%, P<.05; 200 mg/kg, -46%+/-10%, P<.01). Within days of treatment, orlistat treatment increased fecal excretion of UCB (at day 3; +220%, P<.05). During 24 weeks of orlistat treatment (200 mg/kg diet), the plasma bilirubin concentrations were continuously approximately 35% lower than in control rats. Plasma UCB concentrations were inversely correlated with the amount of fecal fat excretion (n=12, r=-0.87, P<.001). CONCLUSIONS: In Gunn rats, orlistat treatment increases the fecal excretion of fat and enhances the disposal of UCB. This approach could lead to novel strategies for prevention and treatment of unconjugated hyperbilirubinemia in patients.
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Bilirrubina/análisis , Bilirrubina/sangre , Inhibidores Enzimáticos/uso terapéutico , Heces/química , Hiperbilirrubinemia/sangre , Hiperbilirrubinemia/tratamiento farmacológico , Lactonas/uso terapéutico , Animales , Conductos Biliares/efectos de los fármacos , Conductos Biliares/metabolismo , Bilirrubina/metabolismo , Modelos Animales de Enfermedad , Hiperbilirrubinemia/metabolismo , Masculino , Orlistat , Ratas , Ratas Gunn , Factores de TiempoRESUMEN
Mujer de 47 años de edad, con regulares hábitos higiénicos-dietéticos. Menarca a los 15 años, GVI, PV, Ci; último parto hacía 8 años; fecha de la última menstruación 1o. de julio de 1982. A los 24 años de edad se le transfundió sangre después de una cesárea. En 1979 presentó neumonía lobar aguda e ictericia; fue tratada con eritromicina por ser alérgica a la penicilina. Desde 1980 se refirió elevación de las bilirrubinas séricas y bilirrubinas. En 1981 recibió HAIN, etambutol y estreptomicina por tener tos y una radiografía de tórax compatible con tuberculosis pulmonar. En 1982 se estableció el diagnóstico de probable síndrome de Sjögren