Common Bile Duct Ligation Animals Research Articles

Role of splenic reservoir monocytes in pulmonary vascular monocyte accumulation in experimental hepatopulmonary syndrome.

Background and AimPulmonary monocyte infiltration plays a significant role in the development of angiogenesis in experimental hepatopulmonary syndrome (HPS) after common bile duct ligation (CBDL). Hepatic monocytes are also increased after CBDL, but the origins remain unclear. Splenic reservoir monocytes have been identified as a major source of monocytes that accumulate in injured tissues. Whether splenic monocytes contribute to monocyte alterations after CBDL is unknown. This study evaluates monocyte distributions and assesses effects of splenectomy on monocyte levels and pulmonary vascular and hepatic abnormalities in experimental HPS.MethodsSplenectomy was performed in CBDL animals. Monocyte levels in different tissues and circulation were assessed with CD68. Pulmonary alterations of HPS were evaluated with vascular endothelial growth factor‐A (VEGF‐A) levels, angiogenesis, and alveolar–arterial oxygen gradient (AaPO2). Liver abnormalities were evaluated with fibrosis (Sirius red), bile duct proliferation (CK‐19), and enzymatic changes.ResultsMonocyte levels increased in the lung and liver after CBDL and were accompanied by elevated circulating monocyte numbers. Splenectomy significantly decreased monocyte accumulation, VEGF‐A levels, and angiogenesis in CBDL animal lung and improved AaPO2 levels. In contrast, hepatic monocyte levels, fibrosis, and functional abnormalities were further exacerbated by spleen removal.ConclusionsSplenic reservoir monocytes are a major source for lung monocyte accumulation after CBDL, and spleen removal attenuates the development of experimental HPS. Liver monocytes may have different origins, and accumulation is exacerbated after depletion of splenic reservoir monocytes. Tissue specific monocyte alterations, influenced by the spleen reservoir, have a significant impact on pulmonary complications of liver disease.

Alveolar type II epithelial cell dysfunction in rat experimental hepatopulmonary syndrome (HPS).

The hepatopulmonary syndrome (HPS) develops when pulmonary vasodilatation leads to abnormal gas exchange. However, in human HPS, restrictive ventilatory defects are also observed supporting that the alveolar epithelial compartment may also be affected. Alveolar type II epithelial cells (AT2) play a critical role in maintaining the alveolar compartment by producing four surfactant proteins (SPs, SP-A, SP-B, SP-C and SP-D) which also facilitate alveolar repair following injury. However, no studies have evaluated the alveolar epithelial compartment in experimental HPS. In this study, we evaluated the alveolar epithelial compartment and particularly AT2 cells in experimental HPS induced by common bile duct ligation (CBDL). We found a significant reduction in pulmonary SP production associated with increased apoptosis in AT2 cells after CBDL relative to controls. Lung morphology showed decreased mean alveolar chord length and lung volumes in CBDL animals that were not seen in control models supporting a selective reduction of alveolar airspace. Furthermore, we found that administration of TNF-α, the bile acid, chenodeoxycholic acid, and FXR nuclear receptor activation (GW4064) induced apoptosis and impaired SP-B and SP-C production in alveolar epithelial cells in vitro. These results imply that AT2 cell dysfunction occurs in experimental HPS and is associated with alterations in the alveolar epithelial compartment. Our findings support a novel contributing mechanism in experimental HPS that may be relevant to humans and a potential therapeutic target.

Caspase-3 inhibition prevents the development of hepatopulmonary syndrome in common bile duct ligation rats by alleviating pulmonary injury.

Common bile duct ligation (CBDL) rats is an accepted experimental model of hepatopulmonary syndrome (HPS), defined as liver disease and intrapulmonary vascular dilatation and hypoxaemia. Pulmonary Akt and ERK activation followed by angiogenesis in the later stages of CBDL, contribute to the pathogenesis of HPS. However, the mechanisms behind Akt and ERK activation remain undefined. Pulmonary injury induced by increased bilirubin, endotoxin and inflammatory mediators occurs in the early stages of CBDL. We assessed the effects of relieving pulmonary injury on Akt and ERK activation and on the development of HPS following CBDL. Pulmonary injury, angiogenesis, arterial oxygenation, cell proliferation and, phospho-Akt and ERK1 were evaluated in CBDL animals with or without caspase-3 inhibition (Z-DEVD-FMK). Pulmonary injury was assessed by histology and quantifying apoptosis and aquaporin-1 (AQP1) levels. Lung angiogenesis was assessed by quantifying AQP1 level, vWF-positive cells and microvessel count. Pulmonary apoptosis and caspase-3 activation were markedly increased in the early stages of CBDL. Caspase-3 inhibition alleviated apoptosis, the reduction in AQP1, phospho-Akt and ERK1 levels and pulmonary injury 1 week after CBDL. Caspase-3 inhibition also reduced AQP1, phospho-Akt and ERK1 levels, vWF-positive cells, cell proliferation, microvessel count, and microvascular dilatation and improved arterial oxygenation 3 weeks following CBDL. Caspase-3 inhibition alleviates pulmonary injury, thereby preventing angiogenesis as well as the development of HPS in CBDL rats. These effects are related to the regulation of the Akt and ERK1 pathways.

Endothelin-1 Activation of the Endothelin B Receptor Modulates Pulmonary Endothelial CX3CL1 and Contributes to Pulmonary Angiogenesis in Experimental Hepatopulmonary Syndrome

Hepatic production and release of endothelin-1 (ET-1) binding to endothelin B (ETB) receptors, overexpressed in the lung microvasculature, is associated with accumulation of pro-angiogenic monocytes and vascular remodeling in experimental hepatopulmonary syndrome (HPS) after common bile duct ligation (CBDL). We have recently found that lung vascular monocyte adhesion and angiogenesis in HPS involve interaction of endothelial C-X3-C motif ligand 1 (CX3CL1) with monocyte CX3C chemokine receptor 1 (CX3CR1), although whether ET-1/ETB receptor activation influences these events is unknown. Our aim was to define if ET-1/ETB receptor activation modulates CX3CL1/CX3CR1 signaling and lung angiogenesis in experimental HPS. A selective ETB receptor antagonist, BQ788, was given for 2 weeks to 1-week CBDL rats. ET-1 (±BQ788) was given to cultured rat pulmonary microvascular endothelial cells overexpressing ETB receptors. BQ788 treatment significantly decreased lung angiogenesis, monocyte accumulation, and CX3CL1 levels after CBDL. ET-1 treatment significantly induced CX3CL1 production in lung microvascular endothelial cells, which was blocked by inhibitors of Ca(2+) and mitogen-activated protein kinase (MEK)/ERK pathways. ET-1-induced ERK activation was Ca(2+) independent. ET-1 administration also increased endothelial tube formation in vitro, which was inhibited by BQ788 or by blocking Ca(2+) and MEK/ERK activation. CX3CR1 neutralizing antibody partially inhibited ET-1 effects on tube formation. These findings identify a novel mechanistic interaction between the ET-1/ETB receptor axis and CX3CL1/CX3CR1 in mediating pulmonary angiogenesis and vascular monocyte accumulation in experimental HPS.

TNF‐α neutralization improves experimental hepatopulmonary syndrome in rats

TNF-α is increased in hepatopulmonary syndrome (HPS). Pentoxifylline (PTX) mitigated experimental HPS through the inhibition of TNF-α. However, PTX has pleiotropic effects besides the inhibition of TNF-α. This study is to neutralize TNF-α with specific monoclonal antibody to TNF-α (TNF-α McAb) to investigate the effect of TNF-α on HPS. Hepatopulmonary syndrome was induced by common bile duct ligation (CBDL); controls were sham operated. The endpoints were 1, 2, 3, 4 and 5 weeks after surgery. (99m) Technetium-macroaggregated albumin (Tc-MAA) was to evaluate intrapulmonary arteriovenous shunts; Portal venous pressure, cardiac output and mean blood pressure (MAP) were also measured. Serum was for Alanine transaminase (ALT), endotoxin, TNF-α and nitric oxide (NO) measurements, liver for histology, lung for histology and iNOS, PI3K/Akt expression assay. Portal vein pressure was significantly elevated and MAP decreased in CBDL rats. Tc-MAA was mainly located in lung and very weak in brain in sham group and mainly in brain of CBDL rats. TNF-α McAb significantly decreased the radioactivity in the brain, reduced cardiac output, increased MAP and systemic vascular resistance (SVR) in CBDL animals. Serum ALT, endotoxin, TNF-α and NO were significantly increased. TNF-α McAb significantly decreased these serum indices in CBDL rats. TNF-α McAb significantly alleviated liver damage, decreased alveolar-arterial gradient and inhibited iNOS, PI3K/Akt and p-Akt expression in lung tissue. Furthermore, TNF-α McAb significantly attenuated the inflammatory response in lung. TNF-α McAb improves HPS in cirrhotic rats; this effect is likely mediated through the inhibition of TNF-α PI3K/Akt-NO pathway.

The role of CX3CL1/CX3CR1 in pulmonary angiogenesis and intravascular monocyte accumulation in rat experimental hepatopulmonary syndrome

Hepatopulmonary syndrome (HPS), classically attributed to intrapulmonary vascular dilatation, occurs in 15-30% of cirrhotics and causes hypoxemia and increases mortality. In experimental HPS after common bile duct ligation (CBDL), monocytes adhere in the lung vasculature and produce vascular endothelial growth factor (VEGF)-A and angiogenesis ensues and contribute to abnormal gas exchange. However, the mechanisms for these events are unknown. The chemokine fractalkine (CX(3)CL1) can directly mediate monocyte adhesion and activate VEGF-A and angiogenesis via its receptor CX(3)CR1 on monocytes and endothelium during inflammatory angiogenesis. We explored whether pulmonary CX(3)CL1/CX(3)CR1 alterations occur after CBDL and influence pulmonary angiogenesis and HPS. Pulmonary CX(3)CL1/CX(3)CR1 expression and localization, CX(3)CL1 signaling pathway activation, monocyte accumulation, and development of angiogenesis and HPS were assessed in 2- and 4-week CBDL animals. The effects of a neutralizing antibody to CX(3)CR1 (anti-CX(3)CR1 Ab) on HPS after CBDL were evaluated. Circulating CX(3)CL1 levels and lung expression of CX(3)CL1 and CX(3)CR1 in intravascular monocytes and microvascular endothelium increased in 2- and 4-week CBDL animals as HPS developed. These events were accompanied by pulmonary angiogenesis, monocyte accumulation, activation of CX(3)CL1 mediated signaling pathways (Akt, ERK) and increased VEGF-A expression and signaling. Anti-CX(3)CR1 Ab treatment reduced monocyte accumulation, decreased lung angiogenesis and improved HPS. These events were accompanied by inhibition of CX(3)CL1 signaling pathways and a reduction in VEGF-A expression and signaling. Circulating CX(3)CL1 levels and pulmonary CX(3)CL1/CX(3)CR1 expression and signaling increase after CBDL and contribute to pulmonary intravascular monocyte accumulation, angiogenesis and development of experimental HPS.

S-nitroso-N-acetylcysteine attenuates liver fibrosis in cirrhotic rats

This study was aimed to investigate the molecular mechanisms underlying prevention of hepatic fibrosis by S-nitroso-N-acetylcysteine (SNAC), a nitric oxide donor that inhibits lipid peroxidation. Secondary biliary cirrhosis was induced by 4 weeks of common bile duct ligation (CBDL). Both sham-operated and CBDL animals received SNAC (6.0 micromol/kg/day) starting 2 weeks after surgery. SNAC treatment reduced the increase in blood enzyme activities (alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase), induced by CBDL. Histological changes were attenuated and there was a significant decrease in the area of liver fibrosis and in the activation of stellate cells measured by alpha-smooth muscle actin (alpha-SMA) immunostaining. The increase in TBARS concentration and hydroperoxide-induced chemiluminescence were also reduced by SNAC treatment. SNAC down-regulated expression of collagen 1 alpha, alpha-SMA, tumor necrosis factor-alpha, tumor growth factor-beta, metalloproteinase-2, metalloproteinase inhibitor 1, platelet-derived growth factor (PDGF), and PDGF receptor in CBDL rats. These effects were accompanied by inhibited activation of extracellular signal-regulated kinases, Jun amino-terminal kinases, p38 and Akt. Antifibrotic effects were more efficient than those of the free thiol NAC administered at a dose of 60 mumol/kg. In conclusion, results obtained indicate that SNAC, beyond its antioxidant capacity, exerts antifibrotic effects in rats with secondary biliary cirrhosis by down-regulating increased expression of genes and modulating intracellular signaling pathways that contribute to the accumulation of matrix proteins. Thus, SNAC may be an interesting candidate for the treatment of human fibrosis and cirrhosis.

Altered Kupffer cell function in biliary obstruction

An altered Kupffer cell (KC) response is thought to be responsible for the characteristic phenotype observed after biliary obstruction: a phenotype marked by a defect in the hepatic reticuloendothelial system and a hypersensitivity to endotoxin. Few studies, however, have directly examined KC function. We have sought to define the specific alterations in function and phenotype that occur in the KC after biliary obstruction. KCs were isolated from female C57BL/6 mice 4 days after a sham or common bile duct ligation (CBDL) operation. Phagocytosis, oxidative burst potential, and intracellular bacterial killing were measured as markers of reticuloendothelial system function. The KC response to endotoxin was assessed by measuring tumor necrosis factor alpha and interleukin 6 levels in the media after stimulation with lipopolysaccharide (LPS) or with LPS plus LPS-binding protein (LBP). CBDL KCs demonstrated a significant increase in phagocytic ability and significantly decreased baseline oxidative stress, compared with Shams. The oxidative burst potential, however, was equivalent or higher for CBDL KCs. CBDL KCs also demonstrated increased numbers of viable intracellular bacteria after infection; however, it is unclear if this finding represents impaired intracellular bacterial killing or increased phagocytosis of bacteria. With respect to the KC response to endotoxin, CBDL KCs were found to be less sensitive to the stimulatory effects of LPS alone but were exquisitely sensitive to the effects of LBP. LBP levels were found to be significantly elevated in CBDL animals, and CBDL KCs demonstrated a dose-dependent, exaggerated tumor necrosis factor alpha and interleukin 6 response to LPS administered with LBP. KC function is clearly altered after biliary obstruction. Phagocytic ability is actually increased, although the ability of CBDL KCs to kill bacteria within the phagosome remains ill defined. CBDL KCs are exquisitely sensitive to the effects of LBP, and LBP levels are elevated after biliary obstruction. LBP may be responsible for the increased proinflammatory response observed after endotoxin challenge in animals with biliary obstruction.

Analysis of pulmonary heme oxygenase-1 and nitric oxide synthase alterations in experimental hepatopulmonary syndrome

Background & Aims: Cirrhosis and portal hypertension due to chronic common bile duct ligation reproduce the features of human hepatopulmonary syndrome, whereas portal hypertension alone due to partial portal vein ligation does not. Nitric oxide contributes to experimental hepatopulmonary syndrome, but the nitric oxide synthase forms involved remain controversial. Recently, increased pulmonary heme oxygenase-1 expression and carbon monoxide production have also been found after common bile duct ligation. Our aim was to explore the role of the heme oxygenase-1/carbon monoxide pathway in the pathogenesis of experimental hepatopulmonary syndrome. Methods: Pulmonary heme oxygenase-1 expression and distribution were assessed in sham; 3-week partial portal vein ligation; and 1-, 2-, 3-, 4-, and 5-week common bile duct ligation animals by Northern, Western and immunohistochemical analysis relative to endothelial and inducible nitric oxide synthase levels and to hepatopulmonary syndrome development. In vivo heme oxygenase enzyme inhibition with tin protoporphyrin IX in common bile duct ligation animals was used to define effects on intrapulmonary vasodilatation and arterial blood gases. Results: Heme oxygenase-1 expression in pulmonary intravascular monocytes/macrophages and arterial carboxyhemoglobin levels increased progressively from 3 to 5 weeks after common bile duct ligation relative to controls (5-week protein levels were 15.94 ± 1.75-fold those of sham animals; P < 0.001). Inducible nitric oxide synthase increased transiently in pulmonary intravascular monocytes/macrophages in 3-week common bile duct ligation animals, whereas pulmonary microvascular endothelial nitric oxide synthase increases began at 2 weeks and correlated with the onset of hepatopulmonary syndrome. Tin protoporphyrin treatment normalized carboxyhemoglobin and improved arterial blood gases and intrapulmonary vasodilatation, reflecting partial reversal of hepatopulmonary syndrome. Conclusions: The heme oxygenase-1/carbon monoxide system is an important contributor to the progression of experimental hepatopulmonary syndrome in addition to alterations in the endothelial nitric oxide synthase/nitric oxide pathway.

Increased pulmonary vascular endothelin B receptor expression and responsiveness to endothelin-1 in cirrhotic and portal hypertensive rats: a potential mechanism in experimental hepatopulmonary syndrome

Background/Aims: In experimental hepatopulmonary syndrome (HPS), hepatic endothelin-1 (ET-1) release during common bile duct ligation (CBDL) and ET-1 infusion in pre-hepatic portal hypertension after portal vein ligation (PVL) initiate vasodilatation through an endothelin B receptor mediated increase in pulmonary endothelial nitric oxide synthase (eNOS). We evaluated if pulmonary ET receptor expression changes in experimental cirrhosis and portal hypertension and confers susceptibility to HPS. Methods: In normal, PVL and CBDL animals, lung ET receptor expression and localization were assessed and ET receptor levels and functional analysis of ET-1 effects on eNOS levels were evaluated in intralobar pulmonary artery (PA) and aortic (AO) segments. Normal rats underwent evaluation for HPS after ET-1 infusion. Results: There was a selective increase in ET B receptor expression in the pulmonary vasculature from PVL and CBDL animals. ET-1 stimulated NO production and an ET B receptor mediated increase in eNOS levels in PA segments from PVL and CBDL animals, but not normal animals. ET-1 did not alter lung eNOS levels or cause HPS in normal rats. Conclusions: ET B receptor expression and ET-1 mediated eNOS and NO production are enhanced in the lung vasculature in cirrhotic and portal hypertensive animals and correlate with in vivo susceptibility to ET-1 mediated HPS.

Activity of brush border membrane enzymes in proximal jejunum of portal hypertensive rats

Malabsorption accompanies portal hypertension, especially when associated with chronic liver disease. The development of portal hypertension is accompanied by significant alterations in the splanchnic microcirculation. In this study, the effect of extrahepatic and intrahepatic portal hypertension on brush border membrane enzymes was estimated. Portal hypertension was induced in rats by partial portal vein ligation (PPVL) (n = 6) and common bile duct ligation (CBDL) (n = 6), and the activity of sucrase, lactase, alkaline phosphatase, and leucine aminopeptidase (LAP) in the intestinal homogenate was measured. Intrasplenic pulp pressure (ISPP) (in cm of saline) was found to be elevated in PPVL (21.3 +/- 1.47) and CBDL animals (21.5 +/- 1.79) as compared with findings in their respective sham-operated controls (12.74 +/- 0.86, 11.83 +/- 1.04). Only sucrase and LAP activity was significantly elevated (P < 0.05) in the PPVL group. No changes were observed in the CBDL group. Only sucrase and LAP activities were increased in PPVL rats.

Biliary cyst fluid from common bile duct-ligated rats stimulates endothelial nitric oxide synthase in pulmonary artery endothelial cells: a potential role in hepatopulmonary syndrome.

The hepatopulmonary syndrome (HPS) results from pulmonary microvascular dilatation in cirrhosis and is associated with increased pulmonary endothelial nitric oxide synthase (eNOS) levels. In the common bile duct ligation (CBDL) model, endothelin-1 (ET-1) released from the liver contributes to the rise in pulmonary eNOS and intrapulmonary vasodilatation. Whether substances, including ET-1, are found in the biliary tree and selectively enter the circulation after CBDL to influence the pulmonary vasculature is unknown. We assessed if control bile and fluid obtained from the obstructed biliary tree in CBDL animals contains ET-1 and alters eNOS expression and activity in bovine pulmonary artery endothelial cells (BPAECs). Control bile and biliary cyst fluid contained concentrations of ET-1 25- to 42-fold normal plasma levels, and hepatic venous concentrations of ET-1 were selectively increased after CBDL. Biliary cyst fluid caused a dose-dependent induction of eNOS messenger RNA (mRNA) (1.9-fold control), protein (2.5-fold control), and enzyme activity (2.2-fold control) maximal at a 1:10 dilution. The increases were associated with enhanced nitric oxide (NO) production (3.1-fold control) and were inhibitable with an ET(B) receptor antagonist. Bile from sham and portal vein-ligated animals did not increase eNOS expression and at dilutions of 1:100 and 1:10 caused cell toxicity. These results show that bile and biliary cyst fluid contain high concentrations of ET-1 that are specifically increased in hepatic venous blood after CBDL. Biliary cyst fluid increases eNOS expression and activity in an ET(B) receptor-dependent manner in BPAECs. The findings suggest a novel mechanism for the susceptibility of CBDL animals to the HPS.

Effect of Acalculous Cholecystitis on Gallbladder Neuromuscular Transmission and Contractility

Background. Impaired smooth muscle contractility is important in the pathophysiology of acalculous cholecystitis. Common bile duct ligation (CBDL) is a model of acalculous cholecystitis, producing acute inflammatory changes and decrease in gallbladder smooth muscle contractility. The aim of this study was to determine whether there is coexistent dysfunction of neural efferent motor pathways of the gallbladder after CBDL.Materials and methods. Gallbladder muscle contractility was studied in vitro in normal, CBDL, and sham-operated guinea pigs. Electric field stimulation (EFS; 2–16 Hz) was used to activate intrinsic nerves and exogenous acetylcholine (ACh) was used to directly stimulate the muscle. H&E-stained slides of muscle strips were scored for inflammatory changes.Results. After CBDL, there was a progressive increase in the inflammation score and decrease in gallbladder muscle contractility to ACh. There was also a progressive decline in EFS-induced contractility when expressed as absolute force or normalized to the maximal muscle contractile response to ACh. The nitric oxide synthase inhibitor l-NNA (10 μM) increased EFS-induced contractions by 50 ± 25% (P = 0.05) in CBDL animals but had no effect in sham surgical controls.Conclusions. CBDL with its acute gallbladder inflammation affects gallbladder contractility by two mechanisms: (1) decreased smooth muscle contractility, and (2) decreased neurally mediated contractions. The neurally mediated alterations result from dysfunction of cholinergic excitatory nerves and upregulation of nitric-oxide-mediated inhibition of smooth muscle contractility.

Allopurinol reduces bacterial translocation, intestinal mucosal lipid peroxidation, and neutrophil-derived myeloperoxidase activity in chronic portal hypertensive and common bile duct-ligated growing rats.

Bacterial translocation (BT) from the gastrointestinal tract has been thought to play a role in the pathogenesis of septic complications in patients with chronic portal hypertension (PH) and obstructive jaundice. The purpose of this study was to investigate the incidence of BT and to assess the role of intestinal mucosal malondialdehyde (MDA) levels as an indicator of lipid peroxidation and polymorphonuclear neutrophil-derived myeloperoxidase (MPO) in chronic portal hypertensive and common bile duct-ligated rats. Twenty male rats were subjected to sham laparotomy (SL), 20 rats to calibrated portal vein constriction (PH), 20 rats to common bile duct ligation (CBDL), and 10 rats served as a nonoperated control group (NOP). After 4 wk, 10 animals of each operated group received 50 mg/kg allopurinol intraperitoneally, at 24 h, and again 2 h prior to estimation of BT, intestinal mucosal MDA, and MPO activities. In the NOP and SL groups, BT to the mesenteric lymph nodes (MLN) and spleen was present. In PH and in CBDL rats, BT to liver, portal vein, peritoneum, and caval vein occurred. Allopurinol treatment attenuated the frequence of BT in PH and decreased BT in CBDL rats significantly (p < 0.05). Ileal mucosal MDA levels (nanomoles/g) in untreated rats increased from 45.1 +/- 7.9 in SL to 98.2 +/- 9.1 in PH and to 102.2 +/- 11 in CBDL rats (p < 0.01). In the allopurinol groups the increase of MDA to 49.1 +/- 1.3 in PH, and 66.2 +/- 2.2 in CBDL was significantly lower (p < 0.01). MPO activity (units/g) in the ileal mucosa increased in untreated rats from 319 +/- 129 after SL to 866 +/- 104 after PH and to 1016 +/- 104 after CBDL (p < 0.01). Allopurinol significantly attenuated MPO activity to 369 +/- 44 in PH, and to 372 +/- 60 in CBDL animals (p < 0.01). In PH and CBDL rats significant BT, intestinal mucosal lipid peroxidation, and polymorphonuclear neutrophil-derived MPO activity occurred. Allopurinol reduced BT and improved intestinal mucosal MDA and MPO activities, suggesting that there might be an association between BT and intestinal mucosal lipid peroxidation.

Bacterial translocation in chronic portal hypertensive and common bile duct-ligated growing rats

Spontaneous bacteremias and infectious complications occur in patients with chronic portal hypertension (PH) and obstructive jaundice, and most of these infections are caused by indigenous intestinal bacteria translocating to regional lymph nodes and the systemic circulation. The aim of this study was to investigate bacterial translocation (BT) at two stages: (1) isolated chronic PH; and (2) obstructive jaundice. Four-week-old male rats were either subjected to sham laparotomy (SL) or to pH or common bile duct ligation (CBDL). After 4 weeks, animals were weighed and the portal pressure was measured. Samples from the portal vein (PV), vena cava, liver, spleen, mesenteric lymph nodes (MLN), and ileum were obtained for bacteriologic cultures. Specimens from the liver, jejunum, and ileum were taken for histologic examination. Portal pressure increased from 7.4±0.3 to 20.5±0.6 mmHg in PH and CBDL animals (P <0.01). Bacterial cultures obtained from the ileum showed significant bacterial overgrowth (P <0.01) in pH and CBDL rats (1.3±0.8 × 10(4) after SL; 1.2±0.6 × 10(5) in PH and 1.9±0.6 × 10(6) in CBDL). BT occured in 10% of SL animals to the MLN and spleen. In PH animals 23% positive cultures were found, almost all due to BT to the PV, vena cava, and liver. CBDL resulted in a BT rate of 47%, mainly by translocation to the PV, liver, and MLN (P <0.05 vs. SL). Histomorphologically, the jejunum and ileum were normal in all three groups. These results suggest that in growing PH and CBDL rats intestinal bacterial overgrowth with significant BT occurs, and the incidence of BT seems to be related to the amount of bacteria colonizing the intestinal tract.

Altered expression and function of hepatocyte gap junctions after common bile duct ligation in the rat.

Gap junction channels allow intercellular exchange of ions and small molecules between adjacent cells. Such communication coordinates cellular and organ function in tissues, although it is unclear if altered gap junction expression and communication contribute to organ dysfunction in pathological states. We examined the immunofluorescent (IF) localization and mRNA and protein levels of the two hepatocyte gap junction proteins connexin 32 and connexin 26, after hepatic injury induced by common bile duct ligation (CBDL) in the rat. Intercellular communication was measured by comparing gap junction-mediated coordination of hormone-induced Ca2+ signals in isolated rat hepatocyte couplets from control and CBDL animals. Connexin 32 plasma membrane IF, protein, and mRNA levels decreased markedly early after CBDL and remained low at 14 days. Connexin 26 plasma membrane IF and protein levels also decreased markedly after CBDL, but mRNA levels rose, and a partial return in membrane IF and protein levels was noted at 9 and 14 days. Coordination of vasopressin-induced Ca2+ signals between cells in isolated rat hepatocyte couplets 1 day after CBDL was significantly impaired compared with controls. These results demonstrate that hepatocyte gap junction communication is impaired early after CBDL because of decreased connexin protein levels. Disruption of gap junctions after CBDL may contribute to loss of hepatic functions that depend on gap junction communication.

Role of cirrhosis in the hemodynamic response to hemorrhage in portal hypertension

We studied hemodynamic alterations in normal and three models of portal hypertension at rest, after hemorrhage, and after resuscitation to determine the role of hepatic dysfunction in the splanchnic vascular response to hemorrhage in portal hypertension. One noncirrhotic and two cirrhotic models of portal hypertension were produced in rabbits: partial prehepatic portal vein ligation, common bile duct ligation, and carbon tetrachloride-induced cirrhosis. Animals were subjected to isovolemic hemorrhage followed by reinfusion of shed blood. Portal, central, and aortic pressures, superior mesenteric artery blood flow, and portosystemic shunt were measured. Histologic examination showed parenchymal damage was absent in normal and portal vein ligation, severe in common bile duct ligation, and moderate in carbon tetrachloride-induced cirrhosis. All portal hypertensive animals exhibited diminished splanchnic vasoconstrictive response to hemorrhage compared with normal. The carbon tetrachloride cirrhosis group had severe cirrhotic changes, minimal portosystemic shunt, and mildly diminished constrictive response. In contrast, the portal vein ligation and common bile duct ligation animals had larger portosystemic shunts, markedly diminished constrictive response, and less severe parenchymal damage. A direct correlation existed between magnitude of rise in portal venous pressure or degree of portosystemic shunt and the fall in mesenteric resistance or diminution of vasoconstrictive response to hemorrhage. We concluded that the abnormal splanchnic vascular response in portal hypertension is relatively independent of the degree of hepatic parenchymal injury, but it is related to the degree of portal hypertension and possibly to splanchnic hyperemia.

Gentamicin nephrotoxicity in extrahepatic cholestasis: modulation by dietary calcium.

The present study was designed to test the hypothesis that the presence of a specific hepatobiliary disease, namely common bile duct obstruction, in the absence of other risk factors will exacerbate gentamicin nephrotoxicity. Furthermore, since bile duct ligation decreases urinary calcium excretion, we studied the role of calcium supplementation in the prevention of gentamicin nephrotoxicity in this model. Male Sprague-Dawley rats were allocated to sham groups and common bile duct ligation groups. Gentamicin at 40 and 100 mg per kg per day for 5 days induced a more severe azotemia in common bile duct ligation animals than in sham controls. Furthermore, higher levels of renal gentamicin were found in common bile duct ligation rats than in sham rats early in the course of therapy, at its termination and during the recovery period. Pretreatment of common bile duct ligation animals with dietary calcium supplementation significantly attenuated gentamicin nephrotoxicity and the increased renal gentamicin accumulation, whereas initiation of calcium supplementation concurrent with gentamicin administration had no salutary effect. We conclude that experimental extrahepatic cholestasis in the rat, in the absence of any other factor, potentiates gentamicin nephrotoxicity. The effect is prevented by pretreatment with dietary calcium supplementation but is not modified by concurrent administration of a high-calcium diet.