Dehydrocholic Acid Ameliorates Sodium Taurocholate-Induced Acute Biliary Pancreatitis in Mice.

Acute biliary pancreatitis (ABP) with a high mortality rate is an incurable digestive system disease induced by abnormal bile acid regurgitation due to the biliary obstruction. Dehydrocholic acid (DA) alleviates the severity of cholestatic hepatitis related to biliary inflammation, suggesting DA is potential to develop for the incurable ABP management. Here we identified DA potency and explored the underlying mechanism in ABP. Our data showed that DA administration not only reduced typically clinicopathological parameters including serum levels of amylase and lipase but also suppressed pancreatic tissue edema, necrosis and trypsin activation in ABP mice. We also found that DA significantly reduced the necrosis of pancreatic acinar cells induced by sodium taurocholate (NaT). Further experimental data showed the significant inhibitions of DA on mitochondrial membrane potential depolarization, ATP exhaustion, calcium overload and reactive oxygen species (ROS) erupted in acinar cells induced by NaT, indicating DA could avert acinar cell death through protecting the mitochondrial function, scavenging excessive oxidative stress and balancing calcium. The comprehensive study found DA elevated the expression of transcription factor EB (TFEB) in vitro thus to increase the functional lysosome content. Indeed, DA decreased the Microtubule-associated protein light chain 3 (LC3) II/I ratio as well as ubiquitin-binding protein p62 and Parkin expressions in vivo and in vitro, revealing autophagy restoration maybe through the improvement of TFEB-mediated lysosome biogenesis. These data indicate that DA improves ABP through the mitochondrial protection, antioxidant ability enhancement and autophagy recovery. In conclusion, our study proposes a potential therapy strategy for the incurable ABP.

Ursodeoxycholic acid is conjugated with taurine to promote secretin-stimulated biliary hydrocholeresis in the normal rat.

BACKGROUND & AIMS: Secretin induces bicarbonate-rich hydrocholeresis in healthy individuals, but not in untreated patients with primary biliary cirrhosis (PBC). Ursodeoxycholic acid (UDCA)--the first choice treatment for PBC--restores the secretin response. Compared with humans, secretin has poor effect in experimental normal-rat models with biliary drainage, although it may elicit hydrocholeresis when the bile-acid pool is maintained. In view of the benefits of UDCA in PBC, we used normal-rat models to unravel the acute contribution of UDCA (and/or taurine-conjugated TUDCA) for eliciting the biliary secretin response. METHODS: Intravascular and/or intrabiliary administration of agonists and inhibitors was performed in normal rats with biliary monitoring. Secretin/bile-acid interplay was analyzed in 3D cultured rat cholangiocytes that formed expansive cystic structures with intralumenal hydroionic secretion. RESULTS: In vivo, secretin stimulates hydrocholeresis upon UDCA/TUDCA infusion, but does not modify the intrinsic hypercholeretic effect of dehydrocholic acid (DHCA). The former effect is dependent on microtubule polymerization, and involves PKCα, PI3K and MEK pathways, as shown by colchicine (i.p.) and retrograde biliary inhibitors. In vitro, while secretin alone accelerates the spontaneous expansion of 3D-cystic structures, this effect is enhanced in the presence of TUDCA, but not UDCA or DHCA. Experiments with inhibitors and Ca(2+)-chelator confirmed that the synergistic effect of secretin plus TUDCA involves microtubules, intracellular Ca(2+), PKCα, PI3K, PKA and MEK pathways. Gene silencing also demonstrated the involvement of the bicarbonate extruder Ae2. CONCLUSIONS: UDCA is conjugated in order to promote secretin-stimulated hydrocholeresis in rats through Ae2, microtubules, intracellular Ca(2+), PKCα, PI3K, PKA, and MEK.

Roles of the outer membrane protein AsmA of Salmonella enterica in the control of marRAB expression and invasion of epithelial cells.

A genetic screen for suppressors of bile sensitivity in DNA adenine methylase (dam) mutants of Salmonella enterica serovar Typhimurium yielded insertions in an uncharacterized locus homologous to the Escherichia coli asmA gene. Disruption of asmA suppressed bile sensitivity also in phoP and wec mutants of S. enterica and increased the MIC of sodium deoxycholate for the parental strain ATCC 14028. Increased levels of marA mRNA were found in asmA, asmA dam, asmA phoP, and asmA wec strains of S. enterica, suggesting that lack of AsmA activates expression of the marRAB operon. Hence, asmA mutations may enhance bile resistance by inducing gene expression changes in the marRAB-controlled Mar regulon. In silico analysis of AsmA structure predicted the existence of one transmembrane domain. Biochemical analysis of subcellular fractions revealed that the asmA gene of S. enterica encodes a protein of approximately 70 kDa located in the outer membrane. Because AsmA is unrelated to known transport and/or efflux systems, we propose that activation of marRAB in asmA mutants may be a consequence of envelope reorganization. Competitive infection of BALB/c mice with asmA(+) and asmA isogenic strains indicated that lack of AsmA attenuates Salmonella virulence by the oral route but not by the intraperitoneal route. Furthermore, asmA mutants showed a reduced ability to invade epithelial cells in vitro.

Aqueous sodium dehydrocholate-sodium deoxycholate mixtures at low concentration.

The behavior of the sodium dehydrocholate (NaDHC)-sodium deoxycholate (NaDC) mixed system was studied by a battery of methods that examine effects caused by the different components of the system: monomers, micelles, and both components. The behavior of the mixed micellar system was studied by the application of Rubingh's model. The obtained results show that micellar interaction was repulsive when the aggregates were rich in NaDHC. The gradual inclusion of NaDC in micelles led to a structural transformation in the aggregates and the interaction became attractive. The bile salts' behavior in mixed monolayers at the air-solution interface was also investigated. Mixed monolayers are monotonically rich in NaDC, giving a stable and compact adsorbed layer. Results have shown that the interaction in both micelles and monolayer is not ideal and such behavior is assumed to be due to a structural factor in their hydrocarbon backbone.

Identification, characterization, and immunolocalization of a nucleoside triphosphate diphosphohydrolase in pig liver.

Different isoforms of nucleoside triphosphate diphosphohydrolases (NTPDases; EC 3.6.1.5), also identified as ATP diphosphohydrolases, have been previously described in mammalian tissues. We report here the biochemical characterization of NTPDases in the pig liver. Optimum pH of catalysis is more acidic for this enzyme than for NTPDases (neutral or alkaline pH) found in other mammalian tissues. It is less sensitive to bile salts than the bovine spleen NTPDase. Calculated Km values for ATP and ADP (31 and 21 microM, respectively) are slightly higher than those reported for the latter enzyme. Electrophoretograms of these enzymes also show different migration patterns. Western blots with Ringo, an antibody that recognizes the different isoforms of mammalian NTPDases, show a small but reproducible difference in estimated molecular masses (75 kDa for liver vs 78 kDa for spleen NTPDase). A second antibody, generated against a different sequence of NTPDase I, does not recognize the liver enzyme, thereby indicating some differences in primary structure. Immunolocalization produced a strong signal on hepatocytes, epithelial cells of the bile duct system, and vascular cells. Immunoreactivity was variable among hepatocytes of different lobules and among hepatocytes within a given lobule. In general, those located in the perilobular zone were more reactive than those located in the central zone and in the periphery of the centrolobular vein.

Effect of sodium tauroursodeoxycholate (UR-906) on liver dysfunction in bile duct-ligated rats.

We investigated the effect of sodium tauroursodeoxycholate (UR-906) on cholestasis in common bile duct-ligated rats in comparison with the effect of dehydrocholic acid. UR-906 (30-180 mumol/kg) and dehydrocholic acid (180 mumol/kg) were intravenously given once daily for consecutive 20 days in rats and the common bile duct was ligated for the last 10 days. On the next day after the last test drug administration, serum biochemical and plasma hemostatic variables were determined. UR-906 significantly ameliorated the elevation of serum cholesterol, phospholipid, bilirubin and bile acid concentrations in bile duct-ligated rats. UR-906 significantly suppressed the prolongation of plasma prothrombin time and activated partial thromboplastin time. Furthermore, UR-906 significantly suppressed the decreases in plasma coagulation factor II and X activities. However, dehydrocholic acid did not cause significant changes in any of the variables examined in this model. These results suggest that UR-906 has a beneficial effect against cholestasis induced by bile duct ligation in rats and that this drug may be useful in the treatment of clinical cholestatic disorders.

Avaliaçäo da atuaçäo do ácido dehidrocólico na funçäo da vesícula bibliar / Therapeutic evolution of dehydrocolic acid (B-Vesil) at gallbladder discinesy

Foi feito um acompanhamento de 10 pacientes com uso do ácido dehidrocólico (B-Vesil), que apresentavam problemas de discinesia biliar. Estes pacientes foram acompanhados com exames complementares os quais demonstraram uma boa resposta ao estímulo de esvaziamento da vesícula biliar

Reversibility of organic anion-induced cholestasis: association with compensatory hypersecretion of biliary phospholipid and protein in the dog.

The effect of a concomitant infusion of organic anions, structurally related phthaleins, on bile flow was studied in anaesthetized dogs. A combination of rose bengal and sulfobromophthalein was found to uniquely and synergistically produce an acute, reversible form of intrahepatic cholestasis (< 10% of control level). This phenomenon was not observed with the administration of those individual organic anions at concentrations previously associated with the induction of intrahepatic cholestasis. The infusion of either a micelle forming bile salt, sodium taurocholate, or a non-micelle forming bile salt, sodium dehydrocholate, rapidly reversed the intrahepatic cholestasis (within 20 min after bile salt infusion). During the choleretic phase immediately following the bile salt infusion, a transient but marked hypersecretion, a disproportionately increased output in relation to that of bile acids, of biliary phospholipid (176% of control level by taurocholate and 138% of control level by dehydrocholate), and an even more striking amount of biliary protein hypersecretion were observed (392% of control level by taurocholate and 357% of control level by dehydrocholate). Although the significance of these new post-cholestatic observations requires clarification, it is suggested that the intrahepatic cholestasis induced by organic anions reflects a reversible defect in the mechanism(s) involved in transcellular transport.

[Effects of dehydrocholic acid (Biliton) on metabolism and liver function in cows]. / Dehydrocholsäure (Biliton)-Wirkungen auf Stoffwechsel und Leberfunktion bei Kühen.

Dehydrocholic acid (Biliton) was given to 9 cows with a predisposition for the fat mobilization syndrome in daily doses of 5.5 g each. This was done two weeks after parturition and the results were compared with those from 9 untreated cows. Five other cows suffering from ketosis or indigestion symptoms were treated too. Decreased concentrations of liver lipids, free fatty acids (FFA), bilirubin, beta-OH-butyrate and urea as well as increased glucose in blood plasma indicated a favourable action of Dehydrocholic acid on metabolism and liver function. We did not observe a significant influence on milk and reproduction parameters. The use of Dehydrocholic acid is recommended for use in liver disturbances.

[The role of bile acids in the regulation of human gallbladder filling]. / Rol zhovchnykh kyslot u reguliatsii napovnennia zhovchnogo mikhura liudyny.

An artificial increase of a pool of bile acids due to administration of dehydrocholic, chenodesoxycholic acid preparations and dry bile in 22 healthy people and 179 patients with chronic cholecystitis has been studied for its effect on regulation of the gallbladder filling on an empty stomach using daily echocholecystometry. Filling of the gallbladder under artificial sequestration of bile acids during duodenal probing, enterosorption and cholatogenic diarrhea was studied in 55 patients with cholecystitis. It has been proved that an increase of the pool of bile acids induces intensification of the gallbladder filling on an empty stomach, while a decrease of the pool, vice versa, causes attenuation of its filling. It is found out a size of a pool of bile acids in the human organism is an important humoral factor in the physiological mechanism of the gallbladder filling regulation.

Importance of high-density lipoprotein-phosphatidylcholine in secretion of phospholipid and cholesterol in bile.

The purpose of this work was to evaluate biliary phosphatidylcholine (PC) secretion after intravenous infusion of high density lipoprotein (HDL)-[3H]phosphatidylcholine (HDL-[3H]PC) in rats and to study the effect of infusion of dehydrocholic and cholic acids, which, respectively, inhibit and stimulate biliary secretion of PC. The data obtained in this study showed that, in the basal state, HDL-PC accounted for 38% of biliary PC. Dehydrocholic acid infusion caused only a "residual" secretion of HDL-PC in the bile; however, cholic acid infusion stimulated the secretion of HDL-PC as well as PC from intrahepatic microsomes. The low level of radioactivity of HDL-PC in intrahepatic compartments suggests that HDL-PC taken up by the liver is predestined for the bile secretion. The correlation between the kinetics of bile secretion of HDL-cholesterol and HDL-[3H]PC suggests the importance of HDL-PC in reverse transport of cholesterol to the liver and its transport to the bile. The differences between the effects of dehydrocholic acid and cholic acid infusions can be explained by the differences in bile salts binding to the surface of HDL.

Influence of dehydrocholic and cholic acids on the biliary secretion of anionic polypeptide fraction, the major apoprotein of the biliary lipoprotein complex.

This work was undertaken to study the effect of intravenously infused dehydrocholate (DHCA) and cholate (CA) on lipid and anionic polypeptide fraction (APF) secretion in bile. APF is a small acidic amphipathic apoprotein closely associated with biliary lipids and bilirubin and involved in the control of bile-destined cholesterol. Rats were infused with increasing doses of DHCA (2 and 3 mumols/min/100 g b.w.) and then CA (1, 2, and 3 mumols/min/100 g b.w.). Each dose was infused for 30 min. As expected, intravenous DHCA inhibited biliary phospholipid (PL) and cholesterol secretion, and CA restored it. When DHCA was infused, the level of APF increased fourfold compared with controls. The APF/PL ratio also increased, but biliary albumin remained stable. When bile secretion was stimulated by infusion of CA, biliary APF returned to normal. These data indicate that biliary secretion of APF depends on the nature and the amount of bile salts returning to the liver, and consequently, APF can be considered a marker of bile secretion disorders.

Synthesis and choleretic activity of 3-[2-(3-R', 4-R'', 5-R'''-benzyl)-5-R-benzimidazol-1-yl]-butanoic acids.

On the ground of the evidentiated choleretic activity of 3-[2-benzylbenzimidazol-1-yl]butanoic acid, 28 new acids were prepared in order to evaluate the influence of suitable substitutions in either C5 of heteroring or C3', C4', C5' of benzyl group in position 2 on the choleretic activity. Pharmacological results after i.v. administration of 0.5 mmol/Kg in rats confirmed a general high choleretic activity that in eleven cases showed during the first 4 hours an increase of bile volume higher than 80%, that is superior to that produced by dehydrocholic acid. Only in a few cases the bile volume increase was less than 37% of basal value.

Biliary excretion of polyethylene glycol molecular weight 900. Evidence for a bile salt-stimulated vesicular transport mechanism.

Polyethylene glycol molecular weight 900 (PEG-900) has been used as a marker of vectorial water transport into bile canaliculus. However, the mechanisms by which this compound is excreted have not been clarified. To gain more information on this process, we studied the biliary excretion of [3H]PEG-900 in rats during choleresis induced by canalicular choleretics. In addition, the effects of the microtubule inhibitors colchicine and vinblastine, and of the acidotropic agent chloroquine, on PEG-900 excretion were studied to determine whether a vesicular pathway is involved. Continuous i.v. infusion of either dehydrocholate (DHC, a non-micelle forming bile salt choleretic) or 4-methylumbelliferone (4-MU, a non-bile salt canalicular choleretic) at stepwise-increasing rates [0.7, 1.0 and 1.2 mumol.min-1.(100 g body wt)-1] induced a gradual increment in bile flow, whereas a transient increment of [3H]PEG-900 excretion was observed only during DHC-induced choleresis. Furthermore, studies in which two consecutive i.v. injections of DHC (10 mumol/100 g body wt) were administered showed that [3H]PEG-900 excretion induced by a second administration of DHC was 54% lower than that induced by the first one, despite a similar excretion in bile flow. Finally, colchicine (0.5 mumol/100 g body wt), vinblastine (0.5 mumol/100 g body wt) and chloroquine (50 mg/kg body wt) pretreatments inhibited the DHC-induced increment in biliary [3H]PEG-900 output, while DHC-induced choleresis was almost unaffected. Conversely, excretion of [14C]sucrose, when coadministered with [3H]PEG-900, was not impaired by the treatments. These results suggest that, unlike sucrose, PEG-900 excretion is not associated with canalicular water movements. Instead, it may be related to a vesicular transport process followed by a bile acid-stimulated discharge of secretory vesicles into bile through the lysosomal compartment.

Effects of sodium taurocholate and sodium dehydrocholate on bile flow, lipid and bilirubin secretion in sheep.

The investigation was performed on 8 sheep with implanted catheters in the common bile duct and in the cystic duct. Sodium taurocholate and sodium dehydrocholate were infused into the jugular vein at the rate of 50 mumol/min for 20 min. Directly after the termination of the sodium taurocholate infusion, the volume of the secreted bile increased from 8.4-9 microliters.kg-1.min-1 to the highest mean value of 17.8 microliters.kg-1, min-1, with a simultaneous increase in the concentration of cholates from 1.71 mmol/l to 4.82 mmol/l and bilirubin from 271.1 mumol/l to 461.7 mumol/l. The concentration of cholesterol and phospholipids in the bile also increased, but did not reach statistically significant values. The infusion of sodium dehydrocholate caused an increase in the bile secretion to the highest mean value of 20.59 microliters.kg-1.min-1 with a simultaneous decrease in the concentration of bilirubin to 148.75 mumol/l, cholesterol to 233.0 micrograms/ml, phospholipids to 56.11 micrograms/ml and cholate to 1.0 mmol/l. The results show that biliary secretion of phospholipids, cholesterol and bilirubin is dependent on the secretion of sodium taurocholate rather than on dehydrocholic acid.

Studies on the origin of biliary phospholipid. Effect of dehydrocholic acid and cholic acid infusions on hepatic and biliary phospholipids.

The correlation between the secretion of biliary phospholipid (PL) and bile acid suggests a regulatory effect of bile acid on PL secretion. Bile acids may influence PL synthesis and/or the mobilization of a preformed PL pool. The objective of this study was to determine the contribution of these two sources to biliary PL, by using an experimental protocol in which dehydrocholic acid (DHCA) and cholic acid (CA) were infused to manipulate biliary PL secretion. In control rats, there was a steady state in bile flow. PL secretion and the biliary secretion of newly synthesized phosphatidylcholine (PC). The specific radioactivity of PC in bile was significantly higher than in plasma, microsomes and canalicular membranes. DHCA infusion decreased biliary PC secretion rate by 80%, and secretion returned to normal values at the transport maximum of CA. The specific radioactivity of biliary PC was decreased by 30% by DHCA infusion and reached normal values during CA infusion. There were no significant changes in the specific radioactivity of PC in plasma or cellular organelles during infusion of bile acids. These data indicate that: (1) newly synthesized PC contributes a small percentage to biliary PC; thus a preformed pool (microsomal and extrahepatic) is a major source of biliary PL; (2) the contribution of the extrahepatic pool to the biliary PL may be more important than the microsomal pool.

Biliary excretion of proteins in the rat during dehydrocholate choleresis.

Choleresis induced by dehydrocholate (DHC) stimulates the discharge into bile of lysosomes, which are implicated in the biliary excretion of proteins. Contrary to taurocholate-induced choleresis, DHC choleresis is not affected by microtubule (mt) inhibition. Therefore, the role of mt's in the biliary protein excretion during bile salt choleresis was analyzed in this study. Normal rats and rats treated with the mt poisons colchicine or vinblastine or with the acidotropic agent chloroquine (Cq) were used. The analysis of the protein component in bile was made on SDS-polyacrylamide gel, and the individual polypeptides were quantitated by densitometry. The excretion of bile polypeptides were compared with that of lysosomal acid phosphatase. Bile flow and bile salt output did not show changes on account of treatments. The biliary excretion of acid phosphatase was stimulated by DHC, and it was not affected by mt inhibitors but was markedly diminished by Cq. DHC choleresis produced different effects on the bile polypeptides. The biliary excretion of polypeptide of high molecular mass (84-140 kDa) was stimulated by DHC. Cq treatment increased their basal biliary excretions, whereas DHC-induced secretion was qualitatively and quantitatively similar to that of controls. The 69-kDa polypeptide (albumin) also increased during DHC-induced choleresis, but it showed a different excretory pattern. Cq treatment inhibited such an increase but no correlation with the excretory pattern of the lysosomal marker was found. The biliary excretion of polypeptides of low molecular mass (down to 14 kDa) suffered a transitory decrease and then a subsequent increase over basal values during the DHC choleresis.(ABSTRACT TRUNCATED AT 250 WORDS)

Estradiol-17 beta-D-glucuronide (E-17G) cholestasis in perfused rat liver: fate of E-17G and choleretic responses to bile salts.

This study was designed to test the hypothesis that increasing the infusion rate of bile salts could overcome drug-induced cholestasis. Cholestasis was induced by administration of 17.5 mumol/L estradiol-17 beta-D-glucuronide during the infusion of taurocholate, tauroursodeoxycholate or dehydrocholate at 20 nmol/min/gm liver. After 30 min, a bolus of 10 mumol of the bile salts was added to the perfusate, and the infusion rate of each bile salt was increased. Taurocholate at a rate of 62 or 125 nmol/min/gm liver, caused a prompt dose-dependent increase of the depressed bile flow and bile salt excretion. A higher rate of taurocholate infusion (180 nmol/min/gm liver) was less effective than either the 62 or 125 rate in increasing bile flow. Infusion of tauroursodeoxycholate at 250 or 390 nmol/min/gm liver also led to a dose-dependent recovery. Further increase of tauroursodeoxycholate infusion rate of 580 nmol/min/gm liver did not provide any additional recovery in bile flow. Dehydrocholate, at rates of 62 or 125 nmol/min/gm liver, gave only a slight enhancement of bile flow. Both taurocholate and tauroursodeoxycholate caused a marked removal of the estradiol-17 beta-D-glucuronide, which had accumulated in the liver. At lower taurocholate infusion rates, the estradiol-17 beta-D-glucuronide was excreted mainly in the bile. At the highest rate, however, biliary excretion of estradiol-17 beta-D-glucuronide declined significantly, and a marked back-efflux of the estrogen into the perfusate was noted. In contrast, tauroursodeoxycholate led to enhanced biliary estradiol-17 beta-D-glucuronide excretion at all increased tauroursodeoxycholate infusion rates and to only a small increase in back-efflux of estradiol-17 beta-D-glucuronide at the two highest tauroursodeoxycholate infusion rates.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of dehydrocholic acid on the secretion of bile acids and biliary lipids in rats.

This study investigated the influence of dehydrocholic acid (DHCA) infusion on the secretion of endogenous bile acids, and biliary lipids in rats in an attempt to explain the reduction of biliary lipid secretion associated with DHCA infusion. DHCA increased bile flow and the bile acids produced during the infusion were composed of three hydroxy-oxo metabolites (83-93%), and cholic acid (6-14%). Very little DHCA was secreted unchanged (less than 2%). The secretions of all the endogenous biliary bile acids were diminished within 30-60 min of infusion. DHCA furthermore reduced the secretion of exogenous cholic acid when co-infused with DHCA. Phospholipid secretion declined to an undetectable amount and cholesterol declined to 10% of the base value by the end of the infusion. The reduction of biliary lipid secretion during DHCA infusion was attributed to the diminished secretion of endogenous bile acids. These data show that DHCA infusion induces choleresis associated with reduced secretion of endogenous and/or exogenous biliary components.