Bile Acid-Induced Suicidal Erythrocyte Death.

BACKGROUND/AIMS: In nucleated cells, bile acids may activate cation channels subsequently leading to entry of Ca2+. In erythrocytes, increase of cytosolic Ca2+ activity triggers eryptosis, the suicidal death of erythrocytes characterized by phosphatidylserine exposure at the cell surface and cell shrinkage. Eryptosis is triggered by bile duct ligation, an effect partially attributed to conjugated bilirubin. The present study explored, whether bile acids may stimulate eryptosis. METHODS: Phosphatidylserine exposing erythrocytes have been identified utilizing annexin V binding, cell volume estimated from forward scatter, cytosolic Ca2+ activity determined using Fluo-3 fluorescence, and ceramide abundance at the erythrocyte surface utilizing specific antibodies. RESULTS: The exposure of human erythrocytes to glycochenodesoxycholic (GCDC) and taurochenodesoxycholic (TCDC) acid was followed by a significant decrease of forward scatter and significant increase of Fluo-3 fluorescence, ceramide abundance as well as annexin V binding. The effect on annexin V binding was significantly blunted, but not abolished by removal of extracellular Ca2+. CONCLUSION: Bile acids stimulate suicidal cell death, an effect paralleled by and in part due to Ca2+ entry and ceramide. The bile acid induced eryptosis may in turn lead to accelerated clearance of circulating erythrocytes and, thus, may contribute to anemia in cholestatic patients.

The unfolded protein response mediates reversible tau phosphorylation induced by metabolic stress.

The unfolded protein response (UPR) is activated in neurodegenerative tauopathies such as Alzheimer's disease (AD) in close connection with early stages of tau pathology. Metabolic disturbances are strongly associated with increased risk for AD and are a potent inducer of the UPR. Here, we demonstrate that metabolic stress induces the phosphorylation of endogenous tau via activation of the UPR. Strikingly, upon restoration of the metabolic homeostasis, not only the levels of the UPR markers pPERK, pIRE1α and BiP, but also tau phosphorylation are reversed both in cell models as well as in torpor, a physiological hypometabolic model in vivo. Intervention in the UPR using the global UPR inhibitor TUDCA or a specific small-molecule inhibitor of the PERK signaling pathway, inhibits the metabolic stress-induced phosphorylation of tau. These data support a role for UPR-mediated tau phosphorylation as part of an adaptive response to metabolic stress. Failure to restore the metabolic homeostasis will lead to prolonged UPR activation and tau phosphorylation, and may thus contribute to AD pathogenesis. We demonstrate that the UPR is functionally involved in the early stages of tau pathology. Our data indicate that targeting of the UPR may be employed for early intervention in tau-related neurodegenerative diseases.

Defective canalicular transport and toxicity of dietary ursodeoxycholic acid in the abcb11-/- mouse: transport and gene expression studies.

The bile salt export pump (BSEP), encoded by the abcb11 gene, is the major canalicular transporter of bile acids from the hepatocyte. BSEP malfunction in humans causes bile acid retention and progressive liver injury, ultimately leading to end-stage liver failure. The natural, hydrophilic, bile acid ursodeoxycholic acid (UDCA) is efficacious in the treatment of cholestatic conditions, such as primary biliary cirrhosis and cholestasis of pregnancy. The beneficial effects of UDCA include promoting bile flow, reducing hepatic inflammation, preventing apoptosis, and maintaining mitochondrial integrity in hepatocytes. However, the role of BSEP in mediating UDCA efficacy is not known. Here, we used abcb11 knockout mice (abcb11-/-) to test the effects of acute and chronic UDCA administration on biliary secretion, bile acid composition, liver histology, and liver gene expression. Acutely infused UDCA, or its taurine conjugate (TUDC), was taken up by the liver but retained, with negligible biliary output, in abcb11-/- mice. Feeding UDCA to abcb11-/- mice led to weight loss, retention of bile acids, elevated liver enzymes, and histological damage to the liver. Semiquantitative RT-PCR showed that genes encoding Mdr1a and Mdr1b (canalicular) as well as Mrp4 (basolateral) transporters were upregulated in abcb11-/- mice. We concluded that infusion of UDCA and TUDC failed to induce bile flow in abcb11-/- mice. UDCA fed to abcb11-/- mice caused liver damage and the appearance of biliary tetra- and penta-hydroxy bile acids. Supplementation with UDCA in the absence of Bsep caused adverse effects in abcb11-/- mice.

Progressive stages of mitochondrial destruction caused by cell toxic bile salts.

The cell-toxic bile salt glycochenodeoxycholic acid (GCDCA) and taurochenodeoxycholic acid (TCDCA) are responsible for hepatocyte demise in cholestatic liver diseases, while tauroursodeoxycholic acid (TUDCA) is regarded hepatoprotective. We demonstrate the direct mitochondrio-toxicity of bile salts which deplete the mitochondrial membrane potential and induce the mitochondrial permeability transition (MPT). The bile salt mediated mechanistic mode of destruction significantly differs from that of calcium, the prototype MPT inducer. Cell-toxic bile salts initially bind to the mitochondrial outer membrane. Subsequently, the structure of the inner boundary membrane disintegrates. And it is only thereafter that the MPT is induced. This progressive destruction occurs in a dose- and time-dependent way. We demonstrate that GCDCA and TCDCA, but not TUDCA, preferentially permeabilize liposomes containing the mitochondrial membrane protein ANT, a process resembling the MPT induction in whole mitochondria. This suggests that ANT is one decisive target for toxic bile salts. To our knowledge this is the first report unraveling the consecutive steps leading to mitochondrial destruction by cell-toxic bile salts.

Prevention of acute kidney injury by tauroursodeoxycholic acid in rat and cell culture models.

BACKGROUND: Acute kidney injury (AKI) has grave short- and long-term consequences. Often the onset of AKI is predictable, such as following surgery that compromises blood flow to the kidney. Even in such situations, present therapies cannot prevent AKI. As apoptosis is a major form of cell death following AKI, we determined the efficacy and mechanisms of action of tauroursodeoxycholic acid (TUDCA), a molecule with potent anti-apoptotic and pro-survival properties, in prevention of AKI in rat and cell culture models. TUDCA is particularly attractive from a translational standpoint, as it has a proven safety record in animals and humans. METHODOLOGY/PRINCIPAL FINDINGS: We chose an ischemia-reperfusion model in rats to simulate AKI in native kidneys, and a human kidney cell culture model to simulate AKI associated with cryopreservation in transplanted kidneys. TUDCA significantly ameliorated AKI in the test models due to inhibition of the mitochondrial pathway of apoptosis and upregulation of survival pathways. CONCLUSIONS: This study sets the stage for testing TUDCA in future clinical trials for prevention of AKI, an area that needs urgent attention due to lack of effective therapies.

Differential effects of silymarin and its active component silibinin on plasma membrane stability and hepatocellular lysis.

Silymarin (SIL) is a natural extract with hepatoprotective properties composed mainly of flavonolignans, with silibinin (SB) being its principal constituent. SB is thought to be the main responsible for SIL hepatoprotective properties, although this has not been corroborated systematically. We analysed comparatively the effects of SIL and SB on hepatocellular plasma membrane stability. SIL (500 microM concentration in SB) protected significantly the plasma membrane disruption induced by Triton X-100 (TX-100) and taurochenodeoxycholate (TCDC), both in isolated plasma membrane (assessed by recording the plasma membrane transition from bilayer to micelle using the R18 self-quenching assay) and in isolated rat hepatocytes (assessed by the release into the incubation medium of the cytosolic enzymes lactate dehydrogenase and alanine aminotransferase). Contrarily, SB (500 microM) exacerbated plasma membrane disruption induced by TX-100 in both systems at detergent concentrations relevant to induce hepatocellular lysis, although it displayed some stabilizing properties at higher concentrations. SB showed a lower stabilizing effect against TCDC-induced plasma membrane disruption when assayed in both models. In addition, SB exposure made the plasma membrane more labile to disruption induced by osmotic lysis. These results show that SIL and SB have differential effects on membrane stability; whereas SIL shows consistently stabilizing effects, SB exacerbates hepatocellular lysis or exerts only minimal stabilizing effects. This differential behaviour should be taken into account when considering the pros and cons of using purified SB vs. the whole SIL extract in medicinal formulations for liver diseases.

Phosphatidylinositol 3-kinase-dependent signaling modulates taurochenodeoxycholic acid-induced liver injury and cholestasis in perfused rat livers.

Taurochenodeoxycholic acid (TCDCA), but not glycochenodeoxycholic acid (GCDCA), activates a phosphatidylinositol 3-kinase (PI3-K)-mediated survival pathway in vitro. Here, the effects of PI3-K inhibition on TCDCA- and GCDCA-induced hepatocellular injury, apoptosis, and bile secretion were examined in the intact liver. In isolated perfused rat livers, bile flow was determined gravimetrically. Hepatovenous lactate dehydrogenase and alanine aminotransferase efflux as markers of liver integrity and biliary secretion of 2,4-dinitrophenyl-S-glutathione (DNP-GS) were determined photometrically. Apoptosis was assessed by immunohistochemistry of active caspase-3 and cytokeratin 18 in liver tissue. Phosphorylation of protein kinase B (PKB/Akt) as a readout of PI3-K activity was determined by immunoblot analysis. Bile acid concentrations were determined by gas chromatography. TCDCA (25 muM) induced moderate liver injury by hepatocellular apoptosis and distinctly reduced bile flow and DNP-GS secretion. In contrast, GCDCA (25 muM) induced severe liver injury by extensive hepatocyte apoptosis. TCDCA strongly activated PI3-K, whereas GCDCA did not markedly affect PI3-K activity. Inhibition of PI3-K by 100 nM wortmannin enhanced TCDCA-induced liver injury and apoptosis and tended to aggravate the cholestatic effect of TCDCA. In contrast, wortmannin reduced GCDCA-induced liver injury and apoptosis. Bile acid uptake tended to be reduced by wortmannin. The cholestatic effect of GCDCA was aggravated by wortmannin. Inhibition of PI3-K markedly aggravated TCDCA-induced but not GCDCA-induced liver damage and hepatocyte apoptosis. Thus TCDCA appears to block its inherent toxicity by a PI3-K-dependent survival pathway in the intact liver.

Role of PGE2 on gallbladder muscle cytoprotection of guinea pigs.

H2O2 and taurochenodeoxycholic acid (TCDC) impair the contraction induced by CCK-8, ACh, and KCl without affecting the actions of PGE2 and damage functions of membrane proteins except for PGE2 receptors. The aim of this study was to examine whether the preserved PGE2 actions contribute to cytoprotective mechanisms against reactive oxygen species. Muscle cells from guinea pig gallbladder were obtained by enzymatic digestion. Levels of lipid peroxidation and activities of SOD and catalase were determined by spectrophotometry. Pretreatment with PGE2 prevented the inhibition of H2O2 or TCDC on agonist (CCK-8, ACh, and KCl)-induced contraction and reduced the expected increase in lipid peroxidation and activities of catalase and SOD caused by H2O2 and TCDC. Incubation with CCK-8 for 60 min desensitized CCK-1 receptors up to 30 min, whereas no receptor desensitization was observed after PGE2 pretreatment. Cholesterol-rich liposome treatment enhanced the inhibition of H2O2 and TCDC on agonists-induced contraction, including that of PGE2. Pretreatment with PGE2 before H2O2 and TCDC did not completely block their inhibition on agonist-induced contraction. Cholesterol-rich liposome treatment impaired the expected increase in catalase activities in response to PGE2. We conclude that pretreatment with PGE2 prevents the muscle cell damage caused by H2O2 and TCDC due to the resistance of PGE2 receptors to agonist-induced desensitization. The preservation of PGE2 receptors may be designed to conserve these cytoprotective functions that are, however, impaired by the presence of excess cholesterol in the plasma membrane.

Comparative cytotoxic and cytoprotective effects of taurohyodeoxycholic acid (THDCA) and tauroursodeoxycholic acid (TUDCA) in HepG2 cell line.

This study was performed to compare the effects of two hydrophilic bile acids, taurohyodeoxycholic acid (THDCA) and tauroursodeoxycholic acid (TUDCA), on HepG2 cells. Cytotoxicity was evaluated at different times of exposure by incubating cells with increasing concentrations (50-800 micromol/l) of either bile acid, while their cytoprotective effect was tested in comparison with deoxycholic acid (DCA) (350 micromol/l and 750 micromol/l)-induced cytotoxicity. Culture media, harvested at the end of each incubation period, were analyzed to evaluate aspartate transaminase (AST), alanine transaminase and gamma-glutamyltranspeptidase release. In addition, the hemolytic effect of THDCA and TUDCA on human red blood cells was also determined. At 24 h of incubation neither THDCA nor TUDCA was cytotoxic at concentrations up to 200 and 400 micromol/l. At 800 micromol/l both THDCA and TUDCA induced a slight increase in AST release. At this concentration and with time of exposure prolonged up to 72 h, THDCA and TUDCA induced a progressive increase of AST release significantly (P<0.05) higher than that of controls being AST values for THDCA (2.97+/-0.88 time control value (tcv) at 48 h and 4.50+/-1.13 tcv at 72 h) significantly greater than those of TUDCA (1.50+/-0.20 tcv at 48 h and 1.80+/-0.43 tcv at 72 h) (P<0.01). In cytoprotection experiments, the addition of 50 micromol/l THDCA decreased only slightly (-5%) AST release induced by 350 micromol/l DCA, while the addition of 50 micromol/l TUDCA was significantly effective (-23%; P<0.05). Higher doses of THDCA or TUDCA did not reduce toxicity induced by 350 micromol/l DCA, but were much less toxic than an equimolar dose of DCA alone. At the concentration used in this experimental model neither THDCA nor TUDCA was hemolytic; however at a very high concentration (6 mmol/l) both bile acids induced 5-8% hemolysis. We conclude that bile acid molecules with a similar degree of hydrophilicity may show different cytotoxic and cytoprotective properties.

Micronuclei induction, cell cycle delay and apoptosis as markers of cellular stress caused by ursodeoxycholic acid in human lymphocytes.

Ursodeoxycholic acid (UDCA) is a bile acid (BA) used for cholesterol gallstone dissolution. Since epidemiological evidence indicates that BAs can be involved in the etiology of colorectal cancer, we investigated the effects of UDCA and its physiologically produced taurine conjugate tauroursodeoxycholic acid (TUDCA) on human lymphocyte cultures in terms of genetic damage in the form of micronuclei (MN) production, cell cycle modifications and induction of apoptosis. With respect to controls, treatment with UDCA (from 10 microg/ml) caused a dose-related increase in MN, whereas TUDCA caused no significant increase (up to 1000 microg/ml). Fluorescence in situ hybridization (FISH) analysis using pancentromeric probes suggested that UDCA exerts aneugenic activity. Bromodeoxyuridine/Hoechst flow cytometry showed that both BA significantly inhibit cell cycle progression (UDCA at 100 microg/ml, and TUDCA, more markedly at 300-1000 microg/ml). Neither UDCA nor TUDCA affected induction of apoptosis, as evaluated by the Annexin-V-Fluos assay. We conclude that UDCA is potentially genotoxic. However, taking into account the characteristics of other physiological BA, our findings are in line with the concept that long-term UDCA treatment may be safely administered. The multi-assay approach reported here could be useful in the toxicological evaluation of newly developed BA analogs as candidates for pharmacological use.

Mechanism for the prevention of cholestasis involving cytochrome P4503A overexpression.

BACKGROUND: To clarify the preventive effect of taurohyodeoxycholic acid on liver cholestasis induced by toxic bile acids in rats, we evaluated whether modulation of cytochrome P4503A-linked oxidases is involved in the hepatic bile acid retention and secretion mechanism. We investigated whether the safe or the toxic taurochenodeoxycholic acid, administered singly or together, affects cytochrome P450-catalyzed drug metabolism or biliary parameters. We also considered whether the inhibition of the P-glycoprotein export pump by vinblastine might be related to cytochrome P4503A overexpression. METHODS: Hydroxylation of testosterone and N-demethylation of aminopyrine were studied in subcellular rat liver preparations after intravenous infusion of hepatoprotective and toxic bile acids administered singly or together. Bile flow, calcium secretion, biliary enzymes activity, and secretion rates of the endogenous and administrated bile acids were determined. CYP3A-dependent monooxygenases were also measured in the same coinfusion model in the presence of vinblastine. RESULTS: Although wide modulation of the activities of different P450 subfamily of isoenzymes was seen, P4503A-associated monooxygenases showed similar patterns in the various situations, i.e., induction by taurohyodeoxycholic acid, reduction by taurochenodeoxycholic acid, and protection (intermediate induction) in the coinfusion experiments. This correlates well with biliary parameters demonstrating the hepatoprotective ability of taurohyodeoxycholic acid. Coadministration of bile acids and vinblastine significantly modifies CYP3A-linked activities. CONCLUSIONS: Bile acid structure seems to be linked with hepatotoxicity/hepatoprotection and P4503A modulation. Taurohyodeoxycholic acid could be therapeutic in cholestatic liver disease by inducing P4503A; we can hypothesize that an associated P-glycoprotein expression might facilitate biliary excretion of toxic taurochenodeoxycholic acid accumulated in the liver during cholestasis.

The effect of idebenone, a coenzyme Q analogue, on hydrophobic bile acid toxicity to isolated rat hepatocytes and hepatic mitochondria.

Oxidant stress induced by hydrophobic bile acids has been implicated in the pathogenesis of liver injury in cholestatic liver disorders. We evaluated the effect of idebenone, a coenzyme Q analogue, on taurochenodeoxycholic acid (TCDC)-induced cell injury and oxidant stress in isolated rat hepatocytes and on glycochenodeoxycholic acid (GCDC)-induced generation of hydroperoxides in fresh hepatic mitochondria. Isolated rat hepatocytes in suspension under 9% oxygen atmosphere were preincubated with 0, 50, and 100 micromol/l idebenone for 30 min and then exposed to 1000 micromol/l TCDC for 4 h. LDH release (cell injury) and thiobarbituric acid reactive substances (measure of lipid peroxidation) increased after TCDC exposure but were markedly suppressed by idebenone pretreatment. In a second set of experiments, the addition of 100 micromol/l idebenone up to 3 h after hepatocytes were exposed to 1000 micromol/l TCDC resulted in abrogation of subsequent cell injury and markedly reduced oxidant damage to hepatocytes. Chenodeoxycholic acid concentrations increased to 5.15 nmol/10(6) cells after 2 h and to 7.05 after 4 h of incubation of hepatocytes with 1000 micromol/l TCDC, and did not differ in the presence of idebenone. In freshly isolated rat hepatic mitochondria, when respiration was stimulated by succinate, 10 micromol/l idebenone abrogated the generation of hydroperoxides during a 90-minute exposure to 400 micromol/l GCDC. These data demonstrate that idebenone functions as a potent protective hepatocyte antioxidant during hydrophobic bile acid toxicity, perhaps by reducing generation of oxygen free radicals in mitochondria.

Effect of high concentrations of bile acids on cultured hepatocytes.

High concentrations of bile acids have been reported as injurious to hepatocytes. We report the influence of various combinations of bile acids on the liver-specific function of cultured rat hepatocytes. Using 4 bile acids (glycocholate [GC], taurocholate [TC], glycohenodeoxycholate [GCDC], and taurochenodeoxycholate [TCDC]), we obtained 6 bile-acid mixtures, each containing equal amounts of 2 bile acids (total bile acids [TBA], 2 mM). Changes in gluconeogenesis, ureagenesis, DNA contents, medium alanine aminotransferase, and morphologies were compared among the paired bile acid compositions by measuring the C/CDC ratio ([GC + TC]/[GCDC + TCDC]) of each. In terms of their relative impairments of ureagenesis from greatest to least, the acids were GCDC, TCDC, and GC, which was almost the same as TC. When the C/CDC ratio was 0, the values of all parameters measured deteriorated. When the C/CDC ratio was 1 in the presence of 1 mM GCDC, only the rate of ureagenesis was diminished. When the C/CDC ratio was infinite, no hepatocellular injury was observed. GCDC and TCDC, together or separately, showed significant hepatocellular injury when the TBA concentration was 2 mM.

Vitamin E reduces oxidant injury to mitochondria and the hepatotoxicity of taurochenodeoxycholic acid in the rat.

BACKGROUND & AIMS: Hydrophobic bile acids have been implicated in the pathogenesis of cholestatic liver injury. The hypothesis that hydrophobic bile acid toxicity is mediated by oxidant stress in an in vivo rat model was tested in this study. METHODS: A dose-response study of bolus intravenous (i.v.) taurochenodeoxycholic acid (TCDC) in rats was conducted. Rats were then pretreated with parenteral alpha-tocopherol, and its effect on i.v. TCDC toxicity was evaluated by liver blood tests and by assessing mitochondrial lipid peroxidation. RESULTS: Four hours after an i.v. bolus of TCDC (10 mumol/100 g weight), serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels peaked, hepatic mitochondria showed evidence of increased lipid peroxidation, and serum bile acid analysis was consistent with a cholestatic injury. Liver histology at 4 hours showed hepatocellular necrosis and swelling and mild portal tract inflammation. Treatment with parenteral alpha-tocopherol was associated with a 60%-70% reduction in AST and ALT levels, improved histology, and a 60% reduction in mitochondrial lipid peroxidation in rats receiving TCDC. CONCLUSIONS: These data show that hepatocyte injury and oxidant damage to mitochondria caused by i.v. TCDC can be significantly reduced by pretreatment with the antioxidant vitamin E. These in vivo findings support the role for oxidant stress in the pathogenesis of bile acid hepatic toxicity.

Synthesis and physicochemical, biological, and pharmacological properties of new bile acids amidated with cyclic amino acids.

New analogs of cyclic amino acid-conjugated bile acids were synthesized, and their physicochemical and biological properties were compared with those of natural analogs. Ursodeoxycholic acid was amidated with D-proline, L-proline, 4-hydroxy-L-proline, and 4-methoxy-L-proline. Hyocholic and hyodeoxycholic acids were amidated with L-proline. The physicochemical properties were similar to those of the natural analogs. All of them were highly stable toward enzymatic C-24 amide bond hydrolysis and 7-dehydroxylation. Their transport, metabolism, and effect on biliary lipid secretion were evaluated in bile fistula rat after intravenous infusion. All the analogs were secreted in bile unmodified. The 4-methoxy-L-proline derivative produced the highest secretion rate, much higher than those of all the other natural and synthetic analogs. This was associated with a selective reduction of cholesterol secretion with normal phospholipid secretion and choleresis. When coinfused, all the analogs were able to prevent the hepatotoxicity induced by intravenous taurochenodeoxycholic acid, as revealed by normal choleresis, alkaline phosphatase, and lactate dehydrogenase values in bile. Considering the overall data, 4-methoxy-L-proline, 4-hydroxy-L-proline, and L-proline derivatives of ursodeoxycholic acid were more potent than the natural analogs.

Tauro alpha-muricholate is as effective as tauro beta-muricholate and tauroursodeoxycholate in preventing taurochenodeoxycholate-induced liver damage in the rat.

Male Wistar rats were infused intravenously with taurochenodeoxycholate (0.4 mumol/min/100 gm) alone (group A) or with one of the three bile salts (tauroursodeoxycholate [group B], tauro beta-muricholate [group C] or tauro alpha-muricholate [group D]) at a rate of 0.2 mumol/min/100/gm for 1 hr. One-hour bile flow and bile salt excretion rates were significantly lower in group A than in the other three coinfused (B, C, D) groups. Biliary 1-hr outputs of lactate dehydrogenase and albumin in the bile, on the other hand, were significantly higher in group A than in the other groups. Plasma concentrations of lactate dehydrogenase at the time of killing (1 hr) were two to three times higher in group A than in the other groups. Although tauro alpha-muricholate does not possess a 7 beta-hydroxy group, the 6 beta-hydroxy group that tauro alpha-muricholate possesses thus appears to be as effective as a 7 beta-hydroxy group in reducing the liver damage caused by toxic bile salts such as taurochenodeoxycholate. The so-called hepatoprotective effects of tauroursodeoxycholate and tauro beta-muricholate found in previous studies may require explanation(s) other than the presence of a 7 beta-hydroxy group in their molecular structures.

Tauro beta-muricholate is as effective as tauroursodeoxycholate in preventing taurochenodeoxycholate-induced liver damage in the rat.

Cholestasis and enhanced biliary leakage of proteins such as lactate dehydrogenase (LDH) and albumin are known to be induced by infusions of relatively toxic bile salts such as taurocholate (TC) and taurochenodeoxycholate (TCDC). Tauroursodeoxycholate (TUDC) was previously shown to prevent these bile abnormalities when simultaneously infused (1-5). In the present study, we examined whether tauro beta-muricholate (T beta-MC) has a similar effect. The enhanced biliary excretion of LDH and albumin induced by the infusion of TCDC at a rate of 0.4 mumol/min/100 g was markedly prevented by the simultaneous infusion of T beta-MC or TUDC at a rate one-fourth that of TCDC. Increased LDH level in plasma and hemolysis caused by the infusion of TCDC were also reduced by either T beta-MC or TUDC. These results indicate that T beta-MC has a preventive effect on TCDC-induced hepatobiliary changes, which is as efficient as that of TUDC as shown previously, suggesting that the 7 beta-hydroxy group is important for this hepatoprotective effect. Furthermore, our results suggest that beta-muricholic acid may also have clinical value since current reports demonstrate a beneficial effect of ursodeoxycholic acid on a variety of cholestatic conditions, including primary biliary cirrhosis.

Determinants of biliary secretory pressure: the effects of two different bile acids.

Biliary secretory pressure represents the force generated to deliver bile through the biliary system. Bile acid-induced toxicity may decrease canalicular bile formation and (or) induce back diffusion causing cholestasis. To determine if biliary secretory pressure is a sensitive indicator of bile toxicity, taurocholate was compared with a less cytotoxic bile acid, tauroursodeoxycholate. In fasted male Sprague-Dawley rats, the common bile duct was cannulated and the endogenous bile salt pool was removed by enteroclysis. Taurocholate (n = 35) or tauroursodeoxycholate (n = 35) in saline was infused for 1 h. Maximal biliary secretory pressure was then measured by attaching the biliary cannula to a column monometer and recording the maximum height to which bile rose. With taurocholate administration, bile flow and bile salt secretion linearly rose to a maximum infusion of 0.5 mumol/(min.g liver), above which hemolysis and death occurred. In contrast, tauroursodeoxycholate could be infused at higher rates with bile salt secretion plateauing at 1.25 mumol/(min.g liver] Both had similar choleretic potencies. Mean biliary secretory pressure at low (less than 0.15 mumol/(min.g liver] infusions was lower with taurocholate (22.5 cm bile) than tauroursodeoxycholate (25.2 cm). Further, increasing the taurocholate infusion decreased the biliary secretory pressure; yet for taurousodeoxycholate, pressure remained unchanged even at higher infusions. Thus, taurocholate but not tauroursodeoxycholate decreases biliary secretory pressure at high infusion rates, likely a reflection of its toxicity to the hepatobiliary epithelium.