endothelin-1 and Cholestasis

We have reported previously that centrally applied ET (endothelin)-1 and ET-3 induce either choleresis or cholestasis depending on the dose. In the present study, we sought to establish the role of these endothelins in the short-term peripheral regulation of bile secretion in the rat. Intravenously infused endothelins induced significant choleresis in a dose-dependent fashion, ET-1 being more potent than ET-3. Endothelins (with the exception of a higher dose of ET-1) did not affect BP (blood pressure), portal venous pressure or portal blood flow. ET-1 and ET-3 augmented the biliary excretion of bile salts, glutathione and electrolytes, suggesting enhanced bile acid-dependent and -independent bile flows. ET-induced choleresis was mediated by ET(B) receptors coupled to NO and inhibited by truncal vagotomy, atropine administration and capsaicin perivagal application, supporting the participation of vagovagal reflexes. RT (reverse transcription)-PCR and Western blot analysis revealed ETA and ET(B) receptor expression in the vagus nerve. Endothelins, through ET(B) receptors, augmented the hepatocyte plasma membrane expression of Ntcp (Na⁺/taurocholate co-transporting polypeptide; Slc10a1), Bsep (bile-salt export pump; Abcb11), Mrp2 (multidrug resistance protein-2; Abcc2) and Aqp8 (aquaporin 8). Endothelins also increased the mRNAs of these transporters. ET-1 and ET-3 induced choleresis mediated by ET(B) receptors coupled to NO release and vagovagal reflexes without involving haemodynamic changes. Endothelin-induced choleresis seems to be caused by increased plasma membrane translocation and transcriptional expression of key bile transporters. These findings indicate that endothelins are able to elicit haemodynamic-independent biological effects in the liver and suggest that these peptides may play a beneficial role in pathophysiological situations where bile secretion is impaired.

Topics: Animals; Bile; Blood Pressure; Cholagogues and Choleretics; Cholestasis; Endothelin-1; Endothelin-3; Hemodynamics; Nitric Oxide; Rats; Rats, Sprague-Dawley; Receptor, Endothelin B; Reflex; Regional Blood Flow; Vagotomy; Vagus Nerve

Cholestasis of the liver is known to be an important risk factor for surgical morbidity and mortality after major hepatectomy. However, the mechanism of liver injury in cholestatic liver is not fully understood. The goal of this study was to investigate the process of liver injury due to hepatic ischemia/reperfusion in obstructive cholestasis.. Male C57BL/6 mice underwent common bile duct ligation and subsequently developed obstructive cholestasis. The mice were subjected to 90 min of partial hepatic ischemia followed by reperfusion.. The survival rate of the mice with cholestatic livers after hepatic ischemia/reperfusion was lower than that of the mice with normal livers. Biochemical and histological analyses showed that the cholestatic mice had a much higher degree of hepatocellular injury after reperfusion than the normal mice. Neutrophil accumulation after reperfusion was significantly decreased in the cholestatic livers; however, considerable microcirculatory disturbances were observed in cholestatic livers after reperfusion. Hepatic stellate cell activation and hepatic expression of endothelin-1 were evaluated by immunohistochemical staining in cholestatic livers after reperfusion. These observations were also associated with increased serum levels of endothelin-1.. Hepatic stellate cell activation and increased endothelin-1 production play a crucial role in hepatic ischemia/reperfusion injury in cholestatic liver.

Topics: Animals; Chemokines, CXC; Cholestasis; Common Bile Duct; Endothelin-1; Hepatic Stellate Cells; Ligation; Liver; Liver Diseases; Male; Mice; Mice, Inbred C57BL; Neutrophil Infiltration; Reperfusion Injury

Experimental hepatopulmonary syndrome (HPS) after common bile duct ligation (CBDL) in rat is accompanied by increased lung vascular endothelial endothelin B (ETB) receptor expression and increased circulating levels of endothelin-1 (ET-1). The onset of HPS is hypothesized to be triggered by ET-1/ETB receptor activation of endothelial nitric oxide synthase (eNOS)-derived NO production in the pulmonary endothelium. However, whether functional pulmonary vascular ETB receptors are required for the development of experimental HPS is not defined. We evaluated the effects of vascular ETB receptor deficiency on the development of experimental HPS. The molecular and physiological alterations of HPS were compared in 2-wk CBDL wild-type and ETB receptor-deficient (transgenic sl/sl) rats. Relative to wild-type rats, basal hepatic and plasma ET-1 levels were elevated in sl/sl controls although, unlike wild-type animals circulating ET-1 levels, did not increase further after CBDL in sl/sl animals. In contrast to wild-type animals, ETB receptor-deficient rats did not develop increased Akt and eNOS expression and activation and did not develop gas exchange abnormalities of HPS after CBDL. There was a similar degree of pulmonary intravascular monocyte accumulation in both 2-wk CBDL sl/sl and wild-type animals. In conclusion, ETB receptor deficiency inhibits lung Akt/eNOS activation and prevents the onset of experimental HPS after CBDL. This effect is independent of inhibition of pulmonary intravascular monocyte accumulation. These results demonstrate that ET-1/ETB receptor signaling plays a key role in the initiation of experimental HPS.

Topics: Animals; Animals, Genetically Modified; Cholestasis; Endothelin-1; Gene Deletion; Gene Expression Regulation; Genetic Predisposition to Disease; Heme Oxygenase-1; Hepatopulmonary Syndrome; Liver; Male; Nitric Oxide Synthase Type III; Oncogene Protein v-akt; Rats; Rats, Wistar; Receptor, Endothelin B

Endothelin-1 (ET-1) is a potent stimulator of gallbladder contractility. Its role in modulation of sphincter of Oddi (SO) motility and trans-sphincteric flow (TSF) has not been evaluated. To characterize the effects of ET-1 on SO motility and TSF, 10 anaesthetized Australian possums (in vivo, n = 6) were given graded doses of ET-1 (5-200 pmol kg-1) via closed intra-arterial injection. Blood pressure, TSF and SO motility (basal pressure, phasic amplitude, contraction frequency) were analysed. For in vitro studies, eight SO rings were subjected to 10-12-10-7 mol L-1 cumulative concentrations of ET-1 in organ bath and SO motility was measured. Data are expressed as mean +/- SEM. Statistical analysis used anova. ET-1 induced a dose-related increase in blood pressure with a maximal increase of 37.5 +/- 2.5 mmHg at 200 pmol kg-1, (P < 0.001). ET-1 also increases SO basal pressure (P < 0.001) and contraction frequency (P < 0.0001). However, the contraction amplitude was not significantly affected. ET-1 decreased TSF in a dose-related manner (P < 0.001) with cessation of TSF at the highest dose (P < 0.001). In vitro studies showed a significant increase in mean SO motility index, and frequency of contractions at higher ET-1 concentrations (10-9-10-7 mol L-1). ET-1 is a potent stimulator of SO motility resulting in a reduction in TSF.

Topics: Animals; Cholestasis; Endothelin-1; Female; Gallbladder; Gastrointestinal Motility; Male; Opossums; Organ Culture Techniques; Sphincter of Oddi

Plasma endothelin-1 (ET-1) is a potent vasoconstrictor peptide involved in the pathogenesis of several disorders. Endothelin-1 concentrations are increased in adult patients with cirrhosis. However, little is known about ET-1 concentrations in children with cirrhosis.. Radioimmune assay was used to measure plasma ET-1 concentrations in 19 children with cirrhosis (8 patients with ascites, and 11 without ascites), and 11 age- and sex-matched healthy children. The plasma ET-1 concentrations were correlated with the mean blood pressure, creatinine clearance, and severity of portal hypertension, as measured by portal flow volume and portal flow velocity.. Patients with cirrhosis and ascites had increased plasma ET-1 concentrations compared with patients who did not have ascites (6.8 pg/mL +/- 0.62 pg/mL vs. 4.6 pg/mL +/- 0.35 pg/mL; mean +/- SEM; < 0.01) and controls (3.6 pg/mL +/- 0.27 pg/mL; mean +/- SEM; < 0.0005). Plasma ET-1 concentrations were higher in patients with cirrhosis who did not have ascites compared with controls ( < 0.005). No significant differences were observed between concentrations of the patients with cholestasis and those without cholestasis (5.4 pg/mL +/- 0.52 pg/mL vs. 5.2 +/- 0.32 pg/mL; mean +/- SEM; = 0.1). Plasma ET-1 concentrations correlated positively with the mean blood pressure ( = 0.58; < 0.05) and negatively with renal function, as measured by creatinine clearance ( = -0.7; <0.005). However, no correlation was detected between ET-1 concentrations and portal flow volume ( = -0.02; = 0.4) or portal flow velocity ( = -0.16; = 0.4).. Plasma ET-1 concentrations are increased in children with cirrhosis, with or without ascites, compared with controls. Patients with cirrhosis and ascites have increased ET-1 concentrations compared with those without ascites. The degree of increase does not relate to the severity of portal hypertension. This increase tends to maintain systemic blood pressure but is associated with a decrease in renal function.

Topics: Ascites; Blood Pressure; Case-Control Studies; Child; Cholestasis; Creatinine; Endothelin-1; Female; Humans; Hypertension, Portal; Kidney; Liver Cirrhosis; Male; Radioimmunoassay; Renal Circulation

Hepatic stellate cells (HSC) are involved in the pathogenesis of liver fibrosis; although ET-1 is increased in cirrhosis, its pathophysiological role in fibrogenesis and portal hypertension remains controversial. The aim of this study was to investigate splanchnic hemodynamics and to correlate them with changes in ET-1 expression and HSC activation in bile duct ligated (BDL) rats.. Expression of the ET-1 gene was increased early as measured by quantitative reverse transcriptase-polymerase chain reaction (6-fold 3 days after BDL) whereas ET-1 peptide measured by RIA increased significantly only in the late phase (30-fold at 28 days). There was a linear correlation between portal pressure and the amount of ET-1 in the portal vein (r = 0.66; P = 0.003), as well as between ET-1 and the volume fraction of myofibroblasts (r = 0.80, P < 10(-7)) as assessed by morphometry and immunohistochemical staining using alpha-smooth muscle actin.. During chronic liver injury activation of HSCs and of preproET-1 mRNA is accentuated in the early phase after BDL. The late increase in ET-1 peptide may indicate that this peptide is only secondarily involved in HSC activation. The correlation between ET-1 in portal vein and portal pressure suggests that ET-1 may play an important role in the development of portal hypertension.

Topics: Animals; Aspartic Acid Endopeptidases; Cholestasis; Endothelin-1; Endothelin-Converting Enzymes; Hemodynamics; Hypertension, Portal; Liver; Liver Cirrhosis, Experimental; Male; Metalloendopeptidases; Rats; Rats, Sprague-Dawley; RNA, Messenger

The accumulation of myofibroblasts and fibrosis around proliferating bile ducts in cholestatic liver disease has been attributed to the proliferation and phenotypic modulation of portal fibroblasts, whereas the contribution of hepatic stellate cells remains uncertain. There is increasing evidence to indicate that bile ducts may stimulate chemoattraction of hepatic stellate cells (HSC). In the present study, we undertook dynamic tests to examine such a possibility and to investigate the role of two potential mediators: platelet-derived growth factor-BB (PDGF-BB) and endothelin-1. Cholestasis was induced by bile duct ligation in rats. HSC were isolated from normal rats and culture activated into myofibroblasts expressing PDGF-beta receptors. Migration of myofibroblastic HSC was investigated in a Transwell chemotaxis filter assay. As compared with basal conditions, PDGF-BB (100 microg/l) and endothelin-1 (10(-8) M) induced a 3-fold and 1.7-fold increase in HSC migration, respectively. Bile duct segments isolated from cholestatic rats triggered a 3-fold increase in migration. This stimulation was significantly more potent than that observed in the presence of normal bile ducts. It was inhibited by neutralizing anti-PDGF antibodies and by STI571 PDGF receptor tyrosine kinase inhibitor, by 60% and 85%, respectively, whereas Bosentan, an endothelin receptor antagonist, had no significant inhibiting effect. In bile duct segments from cholestatic rats PDGF-B chain mRNA was detected at higher levels than in controls, whereas PDGF-BB was immunolocalized in bile duct epithelial cells. The results indicate that chemotaxis of HSC towards bile duct structures may contribute to the development of periductular fibrosis in cholestatic disorders, and that PDGF-BB is the major mediator in this process. In addition, anti-liver fibrogenic properties of STI571 are suggested by potent inhibition of myofibroblastic HSC function.

Topics: Animals; Bile Ducts; Chemotaxis; Cholestasis; Endothelin-1; Liver; Liver Cirrhosis, Experimental; Male; Platelet-Derived Growth Factor; Pyrimidines; Rats; Rats, Sprague-Dawley

Endothelin 1 induces contraction, proliferation, and collagen synthesis of hepatic stellate cells in vitro, which may be mediated via the endothelin A receptor. It is unknown if specific blockade of the endothelin A receptor inhibits hepatic fibrosis in vivo.. Groups of 10-20 rats with bile duct occlusion were treated with the nonpeptide endothelin-A receptor antagonist LU 135252 at 80 mg. kg(-1). day(-1) from week 1-6 or from week 4-6, or with LU at 10 mg. kg(-1). day(-1) from week 1-6. Animals with bile duct occlusion alone and sham-operated rats without or with LU at 80 mg. kg(-1). day(-1) over 6 weeks served as controls. After 6 weeks, parameters of fibrogenesis were determined.. LU treatment led to improved histology, paralleled by a dose-dependence up to 60% reduction of liver collagen, even when administered at an advanced fibrosis stage. This was accompanied by a decreased messenger RNA of hepatic procollagen alpha1(I) and tissue inhibitor of metalloproteinase 1, 2 major effectors of fibrosis, and of serum procollagen type III, a surrogate marker of liver fibrogenesis.. Selective endothelin-A receptor blockade can dramatically reduce collagen accumulation in rat secondary biliary fibrosis, a model refractory to most potential antifibrotic agents. Endothelin-A receptor antagonists are promising antifibrotic agents in chronic liver disease.

Topics: Administration, Oral; Alanine Transaminase; Alkaline Phosphatase; Animals; Ascites; Aspartate Aminotransferases; Bilirubin; Cholestasis; Collagen; Disease Models, Animal; DNA, Complementary; Endothelin Receptor Antagonists; Endothelin-1; Female; Hydroxyproline; Hypertension, Portal; Jaundice; Liver; Liver Cirrhosis, Experimental; Organ Size; Phenylpropionates; Pyrimidines; Rats; Rats, Wistar; Receptor, Endothelin A; Receptor, Endothelin B; Receptors, Endothelin; RNA, Messenger; Tissue Inhibitor of Metalloproteinase-1

Circulating plasma endothelin-1 (ET-1) is elevated in liver cirrhosis, in a disease-stage-dependent manner. However, ET-1 exerts its effects mainly via paracrine and autocrine pathways. Therefore, the aim of the present study was to analyze the hepatic endothelin (ET) system in liver cirrhosis resulting from bile duct obstruction (BDO). Wistar rats were subjected for 6 weeks to either sham operation (control) or BDO. Thereafter, hepatic ET-1 concentrations were elevated 7.2-fold in BDO compared to control (p <0.001), whereas big ET-1 was unchanged. The density of both ET receptor subtypes was upregulated in BDO (ETA: 7.4-fold and ETB: 4.9-fold vs control, p < 0.001, respectively). The affinity of both receptor subtypes was significantly reduced in BDO. In conclusion, our data demonstrated for the first time that the hepatic ET system in liver cirrhosis is characterized by a simultaneous upregulation of both ET-1 tissue concentration as well as the density of hepatic ETA- and ETB-receptors, suggesting a synergistic activation of the hepatic ET system in rats with BDO. The increased ET-1 tissue concentration is not a result of an altered big ET-1 synthesis in biliary liver fibrosis, suggesting an increased activity of endothelin-converting enzyme (ECE) in liver cirrhosis.

Topics: Animals; Aspartic Acid Endopeptidases; Cholestasis; Endothelin-1; Endothelin-Converting Enzymes; Female; Liver; Liver Cirrhosis, Experimental; Metalloendopeptidases; Rats; Rats, Wistar; Receptor, Endothelin A; Receptor, Endothelin B; Receptors, Endothelin

Mediators responsible for renal changes in obstructive jaundice are not specified. This study is designed to study the role of endothelin-1 (ET-1) in obstructive jaundice in rats. Animals were randomly placed into five experimental groups. Group 1 (N = 3) was the sham-operated group. Group 2 (N = 8) after common bile duct (CBD) ligation, received bosentan, which is a nonselective endothelin receptor blocker, 50 mg/kg/day for seven days. Group 3 (N = 7) received 1 microg/kg/day captopril. Group 4 (N = 7) was given both drugs orally for seven days. Group 5 (N = 6) after CBD ligation, received Arabic gum as the vehicle. Blood was drawn from the infrahepatic vena cava for the determination of ET-1, bilirubin, creatinine, protein oxidation products, hyaluronic acid, and beta-N-acetyl-hexosaminase. Liver tissue samples were obtained to determine glutathione levels. ET-1, protein oxidation products, hyaluronic acid, bilirubin, and creatinine levels increased significantly in the control group when compared with sham. Bosentan effectively prevented ET-1 elevation but could not reverse creatinine or bilirubin elevation. Captopril with or without bosentan was cytoprotective but did not reverse increased creatinine levels. It is concluded that increased ET-1 in obstructive jaundice may be one of the contributing factors of renal damage.

Topics: Angiotensin-Converting Enzyme Inhibitors; Animals; beta-N-Acetylhexosaminidases; Bilirubin; Bosentan; Captopril; Cholestasis; Creatinine; Endothelin Receptor Antagonists; Endothelin-1; Hyaluronic Acid; Kidney Diseases; Liver; Male; Random Allocation; Rats; Rats, Sprague-Dawley; Sulfonamides

To investigate the cause of decrease of gastric mucosal blood flow (GMBF) in obstructive jaundice under stress.. With common bile duct ligation (CBDL) in Wistar rats under cold restraint stress, GMBF and the content of Endothelin-1, Angiotensin-II, H2, alpha 1 receptor in gastric mucosa were measured. Before stress anti-ET-1 serum, Enalapril, Cimentidine and Phetolamins were administrated, and the change of GMBF was studied.. GMBF was significantly decreased in CBDL in stress than those in control subjects. The content of ET1 and Ang-II was significantly increaced, the density of H2 and alpha 1 receptor was significantly decreased. Before stress antagonist was administrated, and GMBF was significantly increased.. GMBF was decreased by increased ET, Ang-II and decreased H2, alpha 1 receptor in CBDL, under stress. Antagonist improved gastric mucosal blood flow. They had protection from gastric mucosa.

Topics: Angiotensin II; Animals; Cholestasis; Cold Temperature; Endothelin-1; Gastric Mucosa; Male; Rats; Rats, Wistar; Receptors, Histamine H2; Regional Blood Flow; Stomach Ulcer; Stress, Physiological