Reduced Bile Flow Research Articles

Comparative study analyzing effects of sirolimus-cyclosporin and sirolimus-tacrolimus combinations on bile flow in the rat.

The new immunosuppressive agent sirolimus is combined in transplant patients with the cholestatic substances cyclosporin and tacrolimus. Nothing is known about possible cholestatic effects of these combinations. Therefore, we compared their effects on bile flow and on important bile parameters in an acute bile fistula model in rats. Cyclosporin reduced bile flow, biliary excretion of bile salts, cholesterol, and GSH to 20-40% of basal values. Sirolimus decreased bile flow to 50% and excretion of GSH to 30% of the initial conditions but had no effect on cholesterol and bile salt excretion. In contrast, tacrolimus increased bile flow to 120% and GSH excretion to 220% of the basal levels. Sirolimus/cyclosporin decreased bile flow and bile parameters to the same extent as cyclosporin alone. Sirolimus/tacrolimus reversed sirolimus-induced reduction of bile flow and GSH excretion and resulted in a normal bile salt and cholesterol excretion, thus it may be the better alternative in cholestatic patients.

Enhanced plasma availability of moxidectin in fasted horses

The influence of fasting prior to treatment on plasma availability and kinetic disposition of moxidectin was studied in horses. Eight adult crossbred saddle horses were allocated to two experimental groups of four horses. One group was fasted for 24 hours before treatment and the other group received their usual feed. Both groups were treated with an oral gel formulation of moxidectin at the manufacturers recommended therapeutic dose of 0.4 mg/kg. Blood samples were collected by jugular puncture at different times between 0.5 and 50 days. After plasma extraction and derivatization, samples were analyzed by HPLC with fluorescence detection. Computerized pharmacokinetic analyses were done. Fasting induced marked modifications in the pharmacokinetic behavior of moxidectin in the horse. An extended absorption process, and significantly higher area under the curve (AUC) values were obtained in the fasted animals compared to those fed their usual rations. It is suggested that fasting decreases intestinal transit time, and may have prolonged time for absorption of moxidectin. Since, biliary excretion and intestinal secretion are major routes of moxidectin elimination, fasting may have reduced bile flow and intestinal secretion. Fasting before treatment may be a useful tool for improved anthelmintic treatment in horses.

Influence of curcumin on cyclosporin-induced reduction of biliary bilirubin and cholesterol excretion and on biliary excretion of cyclosporin and its metabolites.

We investigated the ability of curcumin, which can be extracted from different Curcuma species, to prevent cyclosporin-induced reduction of biliary bilirubin and cholesterol excretion, and its influence on biliary excretion of cyclosporin (CS) and its metabolites in the bile fistula model in rats. I.v. injection of curcumin (25 and 50 mg/kg) after 30 min increased dose-dependently basal bile flow (30 microliters/kg/min) up to 200%, biliary bilirubin excretion (3000 pmol/kg/min) up to 150%, and biliary cholesterol excretion (22 nmol/kg/min) up to 113%. CS (30 mg/kg) reduced bile flow to 66% and biliary excretion of bilirubin and of cholesterol to 33% of the basal value 30 min after i.v. injection. I.v. administration of curcumin (25 and 50 mg/kg) 30 min after CS increased bile flow dose dependently again to 130% for 1 hour and biliary excretion of cholesterol and of bilirubin to 100% of the basal value for 30 and 150 min, respectively. Injection of curcumin 15 min before CS prevented the CS-induced drop of bile flow at 50 mg/kg and reduction of biliary bilirubin excretion already at 25 mg/kg until the end of the experiment (180 min). The CS-induced reduction of biliary cholesterol excretion, however, was not prevented by curcumin. Finally, the biliary excretions of CS (1200 ng/kg/min) and its metabolites (1200 ng/kg/min) were slightly reduced by curcumin at a dose of 50 mg/kg (to 83% of the initial values). The clinical importance of these controversial effects remains to be shown.

Interleukin-1β Suppresses Retinoid Transactivation of Two Hepatic Transporter Genes Involved in Bile Formation

Cytokines have been implicated in the pathogenesis of inflammatory cholestasis. This is due to transcriptional down-regulation of hepatic transporters including the Na(+)/bile acid cotransporter, ntcp, and the multispecific organic anion exporter, mrp2. We have recently shown that ntcp suppression by lipopolysaccharide in vivo is caused by down-regulation of transactivators including the previously uncharacterized Footprint B-binding protein. Both the ntcp FpB element and the mrp2 promoter contain potential retinoid-response elements. We hypothesized that retinoic acid receptor (RAR) and retinoid X receptor (RXR) heterodimers would activate these two genes and that cytokines that reduce bile flow might do so by suppressing nuclear levels of these transactivators. Retinoid transactivation and interleukin-1beta down-regulation of the ntcp and mrp2 promoters were mapped to RXRalpha:RARalpha-response elements. Gel mobility shift assays demonstrated specific binding of RXRalpha:RARalpha heterodimers to the ntcp and mrp2 retinoid-response elements. The RXRalpha:RARalpha complex was down-regulated by IL-1beta in HepG2 cells. An unexpected finding was that an adjacent CAAT-enhancer-binding protein element was required for maximal transactivation of the ntcp promoter by RXRalpha:RARalpha. Taken together, these studies demonstrate regulation of two hepatobiliary transporter genes by RXRalpha:RARalpha and describe a mechanism which likely contributes to their down-regulation during inflammation.

Increased Sinusoidal Pressure Is Associated with Early Liver Weight Gain in Ischemia–Reperfusion Injury in Isolated Perfused Rat Liver

Background. Hepatic ischemia–reperfusion (I/R) is accompanied by liver weight gain and ascites formation. This could be caused by an increase in sinusoidal pressure, a determinant of hepatic transvascular fluid movement. We determined the role of sinusoidal pressure, assessed by triple vascular occlusion pressure (Pto), in the I/R injury in isolated rat livers perfused with leukocyte-free diluted blood bivascularly via the portal vein and hepatic artery.Materials and methods. Ischemia was induced at room temperature by occlusion of either the inflow lines of the hepatic artery and portal vein (the open outflow group, n = 10) or both the inflow and the outflow (hepatic venous) lines (the closed outflow group, n = 10) for 1 h, followed by 1-h reperfusion in a recirculating manner.Results. Liver weight in both groups increased biphasically after reperfusion; the initial peak occurred at 3 min and the second peak at 60 min. Immediately after reperfusion, Pto peaked, followed by a gradual decline. The initial weight increase in groups combined was significantly and positively correlated with an increase in Pto (r = 0.716, P = 0.0002), but the second peak was independent of Pto. Liver injury, assessed by perfusate levels of hepatic enzymes and reduced bile flow rate, was observed at 60 min after reperfusion in both groups.Conclusions. These findings suggest that increased sinusoidal pressure contributes to only the early liver weight gain after reperfusion in isolated perfused rat livers. The late weight gain may be presumably due to liver injury.

Gabexate mesilate, a synthetic protease inhibitor, reduces ischemia/reperfusion injury of rat liver by inhibiting leukocyte activation.

To investigate whether gabexate mesilate, a synthetic protease inhibitor with anticoagulant properties, prevents hepatic damage by inhibiting leukocyte activation, we examined its effect on ischemia/reperfusion injury of rat liver in which activated leukocytes play a critical role. Prospective, randomized, controlled study. Research laboratory at a university medical center. Male Wistar rats weighing 220 to 280 g. Hepatic damage was evaluated by changes in bile flow and serum transaminase concentrations after ischemia/ reperfusion. Rats received continuous intravenous infusions of gabexate mesilate (10 mg/kg/hr) or intravenous administration of an inactive derivative of activated factor X (Xa), a selective inhibitor of thrombin generation (3 mg/kg), immediately before the induction of ischemia in the median and left lobes of the liver. To determine whether gabexate mesilate inhibits leukocyte activation, we examined the effects of gabexate mesilate on hepatic concentrations of tumor necrosis factor-alpha and rat interleukin-8 and on hepatic myeloperoxidase activity after ischemia/reperfusion. Hepatic dysfunction, observed after 60 mins of ischemia/reperfusion, showed a reduction in bile flow. The ischemia/reperfusion-induced decrease in bile flow was prevented by administration of gabexate mesilate. Serum transaminase concentrations increased after hepatic ischemia/reperfusion, peaking 12 hrs after reperfusion. Gabexate mesilate significantly inhibited the ischemia/reperfusion-induced increase in serum transaminase levels seen 12 hrs after reperfusion. Although an inactive derivative of factor Xa inhibited the increases in serum levels of fibrin and fibrinogen degradation products 6 hrs after reperfusion, it did not prevent ischemia/ reperfusion-induced liver injury. Hepatic levels of tumor necrosis factor-alpha, rat interleukin-8, and myeloperoxidase were significantly increased after ischemia/reperfusion. These increases were significantly inhibited by gabexate mesilate but unaffected by an inactive derivative of factor Xa. Gabexate mesilate reduced ischemia/reperfusion-induced hepatic injury not by inhibiting coagulation, but by inhibiting leukocyte activation.

Microvascular consequences of Kupffer cell modulation in rat liver fibrogenesis.

The process of Ito cell activation, which is thought to be the central pathogenic mechanism in liver fibrogenesis, may involve distinct interactions with Kupffer cells (KCs) mediated by various cytokines and growth factors. The aim of this study was to determine whether targeting KC function using gadolinium chloride (GdCl(3)) interferes with the manifestation of carbon tetrachloride (CCl(4))-induced hepatic fibrosis, placing special emphasis on the process of microvascular remodelling. Using in vivo fluorescence microscopy, characteristic microvascular features of CCl(4)-induced liver fibrosis, progressively observed within the 8-week period of toxin exposure, were the significant reduction in sinusoidal density; the increase of venular vascular space; the perivenular accumulation of Ito cells, with concomitant collagen deposition; and the collapse of parenchymal tissue. GdCl(3) effectively attenuated sinusoidal rarefaction and delayed, but did not prevent, the process of Ito cell activation-associated collagen deposition. Strikingly, the 8-week modulation of KC function by GdCl(3) exhibited sustained hepatocellular fatty vacuolation with organ weight increase, liver enzyme release, and bile flow reduction. Thus, GdCl(3) treatment attenuates the hepatic microvascular response, but favours fatty change and only delays the development of liver fibrosis following CCl(4)-exposure.

Impact of ischemia-reperfusion injury on dimensional changes of hepatic microvessels.

Dimensional alteration of hepatic microvessels was demonstrated during reperfusion after normothermic hepatic ischemia. Using a specially designed cover glass, it was possible to relocate selected sites of observation and microvessels repeatedly throughout the whole reperfusion time. Twenty minutes of hepatic ischemia resulted in a decrease of sinusoidal diameter (mean +/- SEM; 10.0 +/- 0.3 microns at baseline, 8.2 +/- 0.2 microns after ischemia) and diameter of postsinusoidal venules (26.4 +/- 1.2 at baseline, 23.0 +/- 1.0 after ischemia). In the control group (no ischemia induced) no changes of these parameters were observed. Thus, the reduction of hepatic microvascular cross section was present during the early phase of reperfusion. Hepatic dysfunction was characterized by increased serum activity of liver enzymes and reduction of bile flow in the ischemia-exposed animals. It has been suggested that postischemic dimensional microvascular changes are involved in postischemic liver dysfunction.

Taurohyodeoxycholic acid protects against taurochenodeoxycholic acid-induced cholestasis in the rat.

The prevention of the hepatotoxic effects produced by intravenous infusion of taurochenodeoxycholic acid (TCDCA) by coinfusion with taurohyodeoxycholic acid (THDCA) was evaluated in bile fistula rats; the hepatoprotective effects of the latter were also compared with those of tauroursodeoxycholic acid (TUDCA). Rats infused with TCDCA at a dose of 8 micromol/min/kg showed reduced bile flow and calcium secretion, as well as increased biliary release of alkaline phosphatase (AP) and lactate dehydrogenase (LDH). This was associated with a very low biliary secretion rate of TCDCA (approximately 1 micromol/min/kg). Simultaneous infusion of THDCA or TUDCA at the same dose preserved bile flow and almost totally abolished the pathological leakage of the two enzymes into bile. The effect was slightly more potent for THDCA. The maximum secretion rate of TCDCA increased to the highest value (8 micromol/min/kg) when coinfused with either of the two hepatoprotective bile acids (BA), which were efficiently and completely secreted in the bile, without metabolism. Calcium output was also restored and phospholipid (PL) secretion increased with respect to the control saline infusion. This increase was higher in the THDCA study. These data show that THDCA is highly effective in the prevention of hepatotoxicity induced by intravenous infusion of TCDCA by facilitating its biliary secretion and reducing its hepatic residence time; this was associated with selective stimulation of PL biliary secretion.

Liver complications of pediatric parenteral nutrition--epidemiology.

Total parenteral nutrition (TPN)-induced liver disease develops in 40-60% of infants who require long-term TPN for intestinal failure. The clinical spectrum includes cholestasis, cholelithiasis, hepatic fibrosis with progression to biliary cirrhosis, and the development of portal hypertension and liver failure in a significant number of children who are totally parenterally fed. The pathogenesis is multifactorial and is related to prematurity, low birth weight, and duration of TPN. The degree and severity of the liver disease is related to recurrent sepsis including catheter sepsis, bacterial translocation, and cholangitis. Lack of enteral feeding leading to reduced gut hormone secretion, reduction of bile flow, and biliary stasis may be important mechanisms in the development of cholestasis, biliary sludge, and cholelithiasis. Although it is unlikely that modern TPN solutions have a major role in the etiology of TPN liver disease, manganese toxicity recently has been recognized in children with hepatic dysfunction on TPN. Although there is a definite relationship with the degree of manganese toxicity and hepatic decompensation, it is not yet clear whether this is a primary mechanism or whether the high levels are related to reduced biliary excretion of manganese. The management strategies for the prevention of TPN-induced liver disease include early enteral feeding, a multidisciplinary approach to the management of parenteral nutrition, and aseptic catheter techniques to reduce sepsis. The administration of ursodeoxycholic acid may improve bile flow and reduce gall bladder and intestinal stasis. As survival from isolated intestinal transplantation improves, this therapeutic option should be considered before TPN liver disease becomes irreversible and combined liver and small bowel transplantation is required.

The role of phytosterols in the pathogenesis of liver complications of pediatric parenteral nutrition.

Long-term parenteral nutrition of infants who have had major gut resections is associated with a high incidence of cholestatic liver disease. Affected infants have high plasma concentrations of phytosterols--compounds that resemble cholesterol but have an alkylated side chain. The phytosterols that accumulate in patients receiving parenteral nutrition are derived from the soya oil and/or soya lecithin used to make the intravenous lipid emulsion. There is a striking association between phytosterolemia and cholestatic liver disease. This has led us to put forward the hypothesis that phytosterols can cause cholestasis in susceptible infants. Experiments using neonatal piglets indicate that phytosterols (given without any of the other components of parenteral nutrition) can indeed reduce bile flow. We suggest that increasing the content of phytosterols in cell membranes may interfere with the function of important transport proteins involved in the secretion of bile. Other factors that might contribute to cholestasis (such as inhibition of cholesterol 7 alpha-hydroxylase) are discussed.

Calcitonin gene-related peptide (CGRP), a potent regulator of biliary flow.

This study was designed to investigate the effect of porcine calcitonin gene-related peptide (CGRP) on the motility of the porcine biliary tract in vivo. We measured the pressure in the gallbladder and sphincter of Oddi and, in separate experiments, the biliary flow into the duodenum during local intraarterial infusions of CGRP. To determine if the observed effect could be caused by release of cholecystokinin (CCK), we measured the CCK release. The basal pressure in the sphincter of Oddi increased dose-dependently from 5.9 +/- 0.5 mmHg to 11.5 +/- 2.1 mmHg and the motility index of phasic contractions (amplitude x frequency) from 47 +/- 8 to 347 +/- 64 mmHg s-1, at an infusion rate of 32.6 pmol kg-1 min-1. No effect was observed on the gallbladder pressure. CGRP at 6.5 pmol kg-1 min-1 significantly reduced the biliary flow into the duodenum to 47.7 +/- 6% of the basal level. Atropine, injected intravenously, completely abolished the contractile effect of CGRP. CGRP had no effect on the release of CCK. We conclude that CGRP increases biliary motility and hereby reduces bile flow, an effect which involves cholinergic but not cholecystokininergic mechanisms.

Reevaluation of cyclosporine induced hepatotoxicity in the isolated perfused rat liver

Livers of male rats were perfused for 120 min in a recirculating hemoglobin-free system with different concentrations of cyclosporine (CS 2, 10, 50, 150 and 200 mg/l). CS produced damage to the livers in a dose dependent manner. The first sign of hepatotoxicity was a reduction of bile flow amounting to 50% already at 50 mg/l CS. At concentrations of 150 mg/l and 200 mg/l, CS lead to a nearly complete suppression of bile flow, furthermore to a release of cytosolic (GPT, glutamate–pyruvate transaminase, LDH, lactate dehydrogenase) and mitochondrial (GLDH, glutamate dehydrogenase) enzymes into the perfusate and to a decrease in hepatic oxygen consumption (30% at 200 mg/l CS). As a consequence of the reduced aerobic energy supply, hepatic ATP concentration declined (70% at 200 mg/l CS). The hepatic concentrations of reduced glutathione (GSH) were not changed but those of oxidized glutathione (GSSG) increased up to 5-fold by CS. Malondialdehyde (MDA) concentrations in the liver and in the perfusate were not affected consistently by CS. The toxic actions of CS in the isolated rat liver were not influenced (a) by the feeding status of the rats (fed or fasted before surgery) or (b) by addition of superoxide dismutase (SOD, 20 mg/l) and catalase (20 mg/l) to the perfusate 30 min before CS. On the other hand, CS-induced hepatic injury could be attenuated or inhibited completely by addition to the perfusate of (1) 2 mmol/l GSH; (2) 12 mmol/l serine; (3) 12 mmol/l glycine; (4) 0.09 mmol/l deferoxamine (DFO). Conclusions: CS induces cholestasis at lower concentrations, probably by another mechanism(s) than the other signs of hepatotoxicity (enzyme release, ATP depletion). Several lines of evidence indicate a probable participation of reactive oxygen species in CS-induced hepatotoxicity. GSH, DFO, glycine and serine could provide therapeutic opportunities to prevent CS-induced hepatotoxicity in patients treated with high doses of CS.

Effects of nitric oxide on leukotriene D4 decreased bile secretion in the perfused rat liver

Nitric oxide (NO) and leukotrienes are potent vasoactive agents that are involved in the control of portal blood flow. The present study investigated the role of leukotriene D4 and NO in a non-recirculating constant pressure rat liver perfusion model to analyse their interchanges on portal flow and bile secretion. The addition of leukotriene D4 (20 nM) to the perfusate for 5 minutes resulted in a decrease in portal blood flow (−55.3%), in bile flow (−24.4 %) as well as bile acid release (−35.2%). In parallel, leukotriene D4 increased glucose output. The administration of a lower dose of leukotriene D4 (5 nM) reduced the respective parameters to a lesser degree, indicating dose-dependence. The addition of NO via the infusion of sodium nitroprusside (0,05 mM, 1 mM) reduced the effect of leukotriene D4 on portal flow, bile flow and bile acid secretion whereas the leukotriene D4 effects on hepatic glucose output remained unaffected. Correlation coefficient between decrease in portal flow and reduction of bile flow by infusing leukotriene D4 was R = 0.91, while in the presence of sodium nitroprusside R = 0.85. These results suggest that the leukotriene D4-induced cholestasis is dependent on portal flow. In contrast, hepatic vasoconstriction does not contribute to glycogenolyis stimulated by leukotriene D4 in the perfused liver.

Hepatoprotective effect of picroliv against rifampicin-induced toxicity

Picroliv, the active constituent of the plant Picrorhiza Kurroa, showed significant hepatoprotective as well as anticholestatic activity against rifampicin-induced hepatic damage. Rifampicin (50 mg/kg ip × 6 days) resulted in the reduction of bile flow as well as its contents (bile salts and bile acids) in the conscious rat and anesthetized guinea pig. Further, it also caused a decrease in the viability and rate of oxygen consumption in isolated rat hepatocytes. Picroliv treatment significantly reversed the altered parameters of bile and hepatocytes. The hepatoprotective drug silymarin on comparison was found to be less active than picroliv. Drug Dev. Res. 40:299–303, 1997. © 1997 Wiley-Liss, Inc.

Bile secretion by the rat liver during synchronized regeneration.

Simultaneous coexistence of differentiated, proliferating and redifferentiated hepatocytes occurs during normal liver regeneration (LR). The aim of the present work was to study the time course of the capacity of the liver to form bile during synchronized LR. Following two-thirds partial hepatectomy (PH) in rats, i.v. administration of the ribonucleotide reductase reversible inhibitor hydroxyurea (HU) was used to transiently block liver cells at G1/S boundary. Experiments were performed at 0 and 4 hours, and 1, 3 or 7 days after releasing HU-induced inhibition. Bile acid pool size was determined by collecting bile samples over 24 hours. Initial (first hour) bile flow and bile acid output were increased early on during synchronized LR as compared with the values found in non-hepatectomized control animals. These values were thereafter (1 day) reduced, but increased again at 3 days after halting HU infusion. The time course of bile acid depletion and changes in bile flow were very similar in control and synchronized LR, except that in the latter a more important early reduction in bile flow and bile acid output was found. Shortly after PH, part of the bile acid pool was lost, but this was quickly restored, soon (1 day) reaching a net bile acid pool size very similar to that found in control rats. The highest pool size relative to liver weight was found on day 1, when bile acid output and bile flow reached their lowest values. Additional experiments were performed using in situ perfused regenerating rat livers in which stepwise infusion of taurocholate (TC) was carried out. PH alone modified neither the bile acid-independent (BAIF) nor the bile acid-dependent fraction of bile flow (BADF). However, in normal LR, the BAIF decreased on day 1 and recovered at 7 days, while in synchronized LR it remained depressed up to 7 days. The BADF was only reduced during the early phase of normal LR and did not change significantly in synchronized LR. The maximal secretion rate (SRmax) for TC, as expressed per gram of remaining liver tissue, was not affected immediately after PH, but a marked reduction was observed on day 1 in both normal and synchronized LR. Afterwards, SRmax was quickly restored in both synchronized LR and, although in a slower way, normal LR. These results suggest that synchronization of LR involves changes in the time required to the recovery of specific liver functions such as bile formation.

Dietary vitamin E does not protect from endotoxin-induced hepatic microvascular dysfunction.

Starting from the concept that lipopolysaccharide (LPS)-associated hepatotoxicity involves the action of reactive oxygen species, the present study was conducted to test whether vitamin E, a lipophilic antioxidant, prevents LPS-induced hepatic microvascular dysfunction and liver injury. Fifty-two rats were divided into three groups and fed diets containing 0 (n = 16), 75 (n = 18) or 8000 mg (n = 18) alpha-tocopherol acetate/kg food for four weeks. At 1 h and 6 h after intravenous LPS-exposure (10 mg/kg E. coli LPS) hepatic microvascular response and liver injury were assessed by the analysis of Kupffer cell phagocytic activity, leukocyte-endothelial cell interaction and nutritive sinusoidal perfusion (intravital fluorescence epi- illumination technique) as well as bile flow, serum liver enzyme activities and tissue histomorphology. In animals fed with 75 mg vitamin E/kg (standard diet), LPS caused hepatic Kupffer cell activation (increased phagocytic activity) and hepatic microvascular leukocyte activation, with stasis in sinusoids and adherence in postsinusoidal venules (1 h) followed by leukocytic infiltration into tissue (6 h) and progredient sinusoidal perfusion failure (6 h). Hepatic microvascular injury was accompanied by reduced bile flow and enhanced liver enzyme release. Vitamin E-enriched diet (8000 mg/kg) and even vitamin E-deficient diet did not significantly affect LPS-induced hepatic microvascular cell activation and perfusion failure. Thus, we conclude, that vitamin E is not effective to protect from endotoxin-induced hepatic microvascular dysfunction.

Improvement of Estradiol- 17β-D-Glucuronide-Induced Cholestasis by Sodium Tauroursodeoxycholate Therapy in Rats

Background: Estradiol-17β-D-glucuronide (E-17G), a metabolite of natural estrogen, is well known to cause intrahepatic cholestasis in humans. We therefore investigated the effect of sodium tauroursodeoxycholate (T-UDCA), on E-17G-induced cholestasis in female rats. Methods: For die evaluation of the drug, animals given E-17G (10 µmol/kg) were divided into three groups, and T-UDCA was administered intravenously at various doses after E-17G treatment. Results: T-UDCA significantly prevented a marked reduction of bile flow in E-17G-treated rats in all experimental schedules. Furthermore, T-UDCA significantly increased the biliary E-17G excretion rate at an early stage after E-17G treatment in rats. However, this drug caused no significant change in the biliary excretion rate of estradiol-3-sulfate-17β-D-glucuronide (E-3S-17G), which is identified as the major biliary metabolite with E-17G throughout me recovery periods. Conclusion: These results suggest that T-UDCA can improve E-17G-induced acute cholestasis by rapidly increasing the biliary E-17G excretion rate. Thus our finding may provide a useful approach for attempts to prevent drug-induced acute cholestasis in humans.

The canalicular multidrug resistance protein, cMRP/MRP2, a novel conjugate export pump expressed in the apical membrane of hepatocytes

The conjugate export pump in the hepatocyte canalicular membrane is, together with the ATP-dependent bile salt export pump, one of the two major pumps determining canalicular anion secretion and bile flow. The so-called bile salt-independent bile flow is largely driven by the cmrp/cmoat geneencoded conjugate export pump, as indicated by the markedly reduced bile flow in the GY TR − (11, 13–16) and the EHBR mutant rats (18–20). The importance of conjugation with glutathione (52, 53), glucuronate (11, 21), and sulfate (11, 16) for transfer of endogenous and xenobiotic substances from blood into bile has long been known. The molecular identification (7, 26, 54) and cloning (9, 10, 30) of the ATP-dependent export pump for these conjugates in the canalicular membrane was, at least in part, a consequence of the elucidation of the substrate specificity of the multidrug resistance protein (MRP) which is very similar to that of its canalicular isoform (3–6, 49). The broad substrate specificity of the conjugate export pump enables the terminal excretion of a multitude of conjugates and amphiphilic anions which are formed by a large number of relatively specific monooxygenases and transferases in phase I and phase II metabolism of endogenous and xenobiotic substances in the hepatocyte. The predominant expression of the conjugate export pump encoded by the cmrp/cmoat gene in the canalicular membrane does not exclude overexpression of this transporter in other cells and tissues when exposed to drugs and toxins that can be excreted by this pump. The apical conjugate export pump (8–10) may thus confer multidrug resistance to tumor cells in a similar manner as MRP1 (55). The observation that mRNA encoding rat cMrp/cMoat (10, 12) and its rabbit homolog (35) is not only detected in hepatocytes but also in small intestine and the kidneys suggests that the cmrp/cmoat gene-encoded conjugate export pump may function in the apical membrane domain of various epithelial cells.

In vivo assessment of hepatic alterations following gadolinium chloride-induced Kupffer cell blockade

Background/Aims: In recent years, Gadolinium chloride (GdCl 3), a rare earth metal, has frequently been used to study the role and function of Kupffer cells under physiological and pathological conditions. This study was performed to elucidate the consequences of GdCl 3-induced Kupffer cell blockade for hepatic microcirculation, hepatocellular function and integrity. Methods/Results: Using intravital fluorescence microscopy, we studied the hepatic microcirculation of rats pretreated with either GdCl 3 ( n=12; 10 mg/kg; 1 ml i.v. for 2 d) or saline ( n=9; 1 ml). The GdCl 3-treated animals revealed a significantly lower phagocytic activity of Kupffer cells when compared to controls. Concomitantly, GdCl 3-treatment resulted in a pronounced rise of serum cytokine activity (tumor necrosis factor-α; interleukin-6). The hepatic microvascular perfusion was characterized by a moderate increase in the number of non-perfused sinusoids accompanied by a reduction of bile flow. In addition, GdCl 3-treatment caused a slight increase in liver enzyme activity (<200 U/l) aspartate aminotransferase and alanine aminotransferase) with no substantial parenchymal tissue injury (light microscopy). The groups did not differ in concentrations of circulating endotoxin (GdCl 3-treatment: 0.044±0.042 ng/ml; controls: 0.052±0.014 ng/ml). Conclusions: We conclude that hepatic alterations following Kupffer cell blockade with GdCl 3 may possibly be the consequence of cytokine release as a response to the phagocytic challenge of GdCl 3-aggregates. If used for Kupffer cell blockade, the hepatic alterations following GdCl 3-treatment described in the present study should be taken into consideration.