Use Of Bile Acids Research Articles

Site-Directed Mutagenesis and Use of Bile Acid−MTS Conjugates to Probe the Role of Cysteines in the Human Apical Sodium-Dependent Bile Acid Transporter (SLC10A2)

The residues involved in substrate interaction of the human apical sodium-dependent bile acid transporter (hASBT, SLC10A2) remain undefined. Biochemical modification of conserved cysteine residues has suggested their direct involvement in hASBT function. In the present study, we developed novel methanethiosulfonyl (MTS)-bile salt derivatives and describe their reactivity toward hASBT and its mutants. Endogenous Cys residues were subjected to Ala/Thr scanning mutagenesis and subsequent exposure to affinity inactivators. We show that C51A/T, C105A/T, C144A, and C255A/T are loss-of-function mutations. Additionally, C74A/T cell surface expression was abolished suggesting a role in protein folding and/or trafficking. C270A remained largely unaffected in the presence of 1.0 mM polar and charged MTS reagents (MTSEA, MTSES, and MTSET) and retained function similar to wt-hASBT control. However, in the presence of synthetic cholyl- and chenodeoxycholyl-MTS analogues, C270A displayed a significant decrease in K(T) and J(max). Our findings demonstrate that Cys270 is a highly accessible extracellular residue susceptible to thiol modification in its native form that remains largely unaffected upon mutation to Ala. Consequently, C270A provides an ideal scaffold for cysteine scanning mutagenesis studies. Furthermore, the substantial decrease in ligand affinity and maximal transport capacity of C270A suggest that C270 may potentially impact, although not critically, a putative substrate binding domain of hASBT. Overall, bile acid-MTS conjugates can serve as novel and powerful tools to probe the role of endogenous as well as engineered Cys residues and, ultimately, aid in defining their role in the bile acid binding region(s) of hASBT.

Bile acid derivatives as enantiodifferentiating host molecules in inclusion processes

A review on the use of bile acids as hosts for molecular recognition and enantioresolution is presented. The optical resolution of different classes of organic derivatives, i.e., lactones, aliphatic alcohols, sulfoxides, epoxides, ketones, cyclic amides, cyclic carbonates, and sulfites, may be obtained by host-guest enantioselective inclusion complexation. The salient aspects of the enclathration process, including X-ray diffraction results, are discussed.

Use of Bile Acids in Pharmacological and Supramolecular Applications

AbstractFor Abstract see ChemInform Abstract in Full Text.

Use of Bile Acids in Pharmacological and Supramolecular Applications

AbstractBile acid‐based structures have become increasingly important in different fields of chemistry over recent years, having found applications in pharmacology, supramolecular chemistry and nanoscience. Some interesting studies concerning these applications are reviewed, together with the latest developments in synthetic and analytical methods for bile acid‐derived frameworks. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)

Current aspects in pharmacologic use of bile acids

It has been established that combination of bile acids with amphotericin B has potential Leishmanicideal effect and combination with ciprofloxacine has improved its antibacterial activity against Pseudomonas aeruginosa in vitro. PHARMACADYNAMIC EFFECTS: Bile acids have analgesic and hypoglycemic effect. They also have anti-HIV effect probably suppressing virus transmission from cell to cell. New studies of natural bile acids and new synthetic bile acids have revealed that they are not only adjuvants to existing active principles in pharmaceutical forms, but they can act as new therapeutic agents. However, it is necessary to study their possible mechanisms, but they are not crucial for their therapeutic application. Toxicological and pharmacological studies will determine the role of newly synthesized bile acids and their salts in current therapy.

Combined Analysis of Bile Acids and Sterols/Stanols from Riverine Particulates To Assess Sewage Discharges and Other Fecal Sources

This investigation aimed to demonstrate the combined use of bile acids, stanols, and sterols to assess fecal matter inputs into aquatic environments. Bile acids, stanols, and sterols were determined in suspended particulates in water samples collected from sites in the vicinity of discharges from sewage treatment works along the course of the Avon River, Bristol, U.K. The concentrations of the major fecal bile acids [lithocholic (LCA) and deoxycholic (DOCA)] were determined using GC and GC/MS and found to increase along the course of the river. These results agreed with those obtained for coprostanol, the traditional indicator of fecal pollution and other related sterols and stanols. In contrast, sterols and stanols not originating from feces, i.e., 24-ethylcholesterol and 24-ethylcholestanol, tended to decrease in concentration as compared to coprostanol and other fecal markers in the lower reaches of the river. The increasing concentration of bile acids downstream of sewage discharges correlates with the coprostanol/(coprostanol + 5α-cholestanol) ratio of >0.7, thus supporting the use of bile acids as sewage pollution markers. Overall, it is demonstrated that a combined multimolecular approach involving bile acids, stanols, and sterols provides an enhanced means of assessing fecal matter inputs into aquatic environments.

Delta 4-3-oxosteroid 5 beta-reductase deficiency: failure of ursodeoxycholic acid treatment and response to chenodeoxycholic acid plus cholic acid.

In some infants with liver disease, 3-oxo-delta 4 bile acids are the major bile acids in urine, a phenomenon attributed to reduced activity of the delta 4-3-oxosteroid 5 beta-reductase required for synthesis of chenodeoxycholic acid and cholic acid. These patients form a heterogeneous group. Many have a known cause of hepatic dysfunction and plasma concentrations of chenodeoxycholic acid and cholic acid that are actually greater than those of the 3-oxo-delta 4 bile acids. It is unlikely that these patients have a primary genetic deficiency of the 5 beta-reductase enzyme. To document the bile acid profile, clinical phenotype, and response to treatment of an infant with cholestasis, increased plasma concentrations of 3-oxo-delta 4 bile acids, low plasma concentrations of chenodeoxycholic acid and cholic acid, and no other identifiable cause of liver disease. This infant was compared with normal infants and infants with cholestasis of known cause. Analysis of bile acids by liquid secondary ionisation mass spectrometry and gas chromatography-mass spectrometry. The plasma bile acid profile of the patient was unique. She had chronic cholestatic liver disease associated with malabsorption of vitamins D and E and a normal gamma-glutamyltranspeptidase when the transaminases were increased. The liver disease failed to improve with ursodeoxycholic acid but responded to a combination of chenodeoxycholic acid and cholic acid. Treatment of primary 5 beta-reductase deficiency requires the use of bile acids that inhibit cholesterol 7 alpha-hydroxylase.

Chromatographic (TLC) Differentiation of Grizzly Bear and Black Bear Scats

While past work concluded that thin-layer chromatography (TLC) was inadequate for the separation of grizzly (Ursus arctos horribilis) and black bear (U. americanus) scats, our study found differences adequate for species separation. A key was constructed using 19 of 40 data points recorded on each (N = 356 profiles of 178) known-species scat. Accuracy was best for late summer scats (94%). Methods for specimen preparation, analysis, and reading the TLC profiles are discussed. Factors involved in scat variation were tested. Int. Conf. Bear Res. and Manage. 9(1):497-501 The lack of an effective method to discriminate the species of bear seats in areas where grizzly bears and black bears are sympatric is a major impediment in studies of these species. Field sign of the species is often absent when seats are collected days or weeks after deposition. While seat diameter has been widely used as an indicator of species, it inherently confuses black bears with small grizzly bears (Hamer et al 1981). Calder (1984) indicates that the allometric slope of intestinal diameter is low in mammals suggesting that the larger body size of the grizzly would be reflected in only a small increase in intestinal diameter. Seat diameter is also affected by the nature and quantity of food consumed. Seat volume also does not appear to be a reliable character for species separation (Hamer et al. 1981). Since grizzly bears dig more than black bears it might be expected that their seats would contain more soil and mineral matter than those of black bears. However, this expectation was not supported by an exploratory study of the mineral ash content of seats as a method of species discrimination. Electrophoretic methods of detecting species from the proteins shed from the intestine into the seat appear to be possible but have not been used for bears. These fecal proteins can be expected to be degraded by moist conditions and exposure to the elements (Schribner and Warren 1984). DNA has been extracted from bear seats by E. Vyse (per. commun., January 1989 and 1992) but it is not clear if bear DNA from the intestinal cells can be separated from the large amounts of bacterial DNA present in fecal material. Bile acids are very stable compounds having been extracted from 2,000-year-old human coprolites (Lin et al. 1978) and apparently from dinosaur coprolites (per. commun., J. Horner, January 1992). Major et al. (1980) and Johnson et al. (1984) reported that speciesspecific separation of carnivore seats could be obtained by use of thin-layer chromatography (TLC) to identify the array of bile acids contained in the seats. Goodwin (1984) in Alaska and Picton (1986) in Montana attempted to apply this technique to the separation of grizzly and black bear scats without success. Thinlayer chromatography bile acid profiles were obtained from both scats and bile from gall bladders (Welsh and Picton 1983). Both study groups agreed that TLC assessment of bile acids did not provide a reliable means of differentiating scats of the 2 species. The removal of plant pigments that obscured the desired TLC profiles was a major problem. Another possible problem with the use of bile acids is that bears may eat the livers of prey animals and thus bile acids in the scats may represent this exogenous source rather than those of the bear itself. Chemical extraction procedures used in chromatogram preparation extract many neutral lipids and related materials in addition to bile acids. Johnson et al. (1979) reported finding steroid sex hormones in scats and being able to distinguish sexes. Evaluation of data from Goodwin's (1984) and Picton's (1986) studies suggested that discrimination of scats might be possible if all spots on the chromatograms were used instead of just the presumed bile acids. Here we report an evaluation of the use of TLC to identify bear scats to species. This study was funded by the National Park Service at Glacier National Park and the Gabooney Foundation. We are indebted to the members of the Interagency Grizzly Bear Study Team and the Montana Fish, Wildlife and Parks Department for providing scats of known identity. The Montana Agricultural Chemistry Analytical Laboratory provided chemical advice and carried out all of the chemical TLC procedures used in the study. We also wish to thank the 6 individuals who prepared scats for analysis and assisted in reading the TLC plates.

Protein hypersecretory Trichoderma reesei mutant RUT C-30 displays increased ethanol and polyene resistance

Addition of low amounts of ethanol (0.5–2%, v/v) or of polyene antibiotics (nystatine, amphotericin B 0.5–2 mg l −1, respectively) to cultures of T. reesei QM 9414 in the early phase of mycelial growth, arrested the secretion of three cellulases (cellobiohydrolase I and II, endoglucanase I) as well as of a xylanase. Enzyme formation by the enzyme-hyperproducing strain T. reesei RUT C-30, in contrast, was less sensitive to these agents. By the use of [U- 14C]- l-valine addition, this blockage of enzyme formation was shown to be due to a general inhibition of protein secretion by ethanol. Ethanol and amphotericin B also arrested the formation of intracellular precursers of cellobiohydrolase I, and the formation of the cbh1-mRNA, hence proving that blockage occurs at a pre-translational level. The higher sensitivity of the lower producing mutant T. reesei QM 9414 correlated with an increased synthesis of a 36 kDa stress protein. The data are discussed with respect to a plasma membrane located resistance to ethanol or polyenes in the hyperproducing mutant T. reesei RUT C-30, probably caused by the use of bile acids as colony restrictor in the course of mutant selection.

Effect of the cyanobacterial (blue-green algal) toxins from Microcystis aeruginosa on isolated enterocytes from the chicken small intestine

Livestock deaths, and clinical reports of human injury, follow the consumption of toxic blue-green algae. The experiments described show that isolated intestinal enterocytes from chicks are both deformed and killed by exposure to Microcystis toxins, in a dose-dependent and time-dependent manner. The enterocytes were protected from toxicity by deoxycholate, bromosulphothalein and rifampicin. It was concluded that the gastroenteritis clinically associated with accidental Microcystis ingestion is likely to reflect enterocyte injury by Microcystis toxins, and that the therapeutic use of bile acids or transport inhibitors may be of value in treatment.

A Practical Guide to the Nonsurgical Treatment of Gallstones

Until recently, cholecystectomy was the only treatment available for symptomatic gallstone disease. During the past 20 years, better understanding of the pathogenesis of cholesterol gallstone disease has led to alternative nonsurgical methods for treating gallstones in selected groups of patients. Use of 2 naturally occurring bile acids, chenodeoxycholic acid (CDCA) and ursodeoxycholic acid (UDCA), was reported in 1972 and 1975, respectively, for successful dissolution of cholesterol gallstones in humans. Both these bile acids act by reducing cholesterol secretion in bile, thus enabling it to solubilise more cholesterol from the stone surface. Micellar solubilisation is involved, together with liquid crystal formation in the case of UDCA. Having been extensively studied in clinical trials to assess efficacy and safety, both these compounds are now available for general use. The efficacy of CDCA can be enhanced by single bedtime dose administration and by taking a low cholesterol diet. Bedtime administration also enhances the effect of a suboptimal dose of UDCA. CDCA induces dose-related diarrhoea and hypertransaminaemia, and UDCA can induce calcification of gallstones, thus rendering them resistant to medical dissolution. A combination of the 2 bile acids at half the recommended dose for each has become an accepted practice for reducing adverse effects, and this may also enhance efficacy. One of the main problems of bile acid therapy is that dissolution of gallstones is a very slow process. Use of extracorporeal shockwave lithotripsy (ESWL) to break the stones into smaller fragments, with concurrent use of bile acids, has been shown to speed dissolution rate and to achieve complete gallstone dissolution in 78% of selected cases within 12 months.(ABSTRACT TRUNCATED AT 250 WORDS)

Review article: bile acid therapy

Over the past 3 years there has been a renewed interest in bile acid therapy not only because of the promising results obtained by combining this therapy with extracorporeal shock-wave lithotripsy for rapid gallstone dissolution, but also because of its novel use as a treatment for primary biliary cirrhosis. This article reviews the use of bile acids for both these indications.

Quantitative measurement of biliary excretion and of gall bladder concentration of drugs under physiological conditions in man.

Gall bladder storage of hepatic bile prevents complete recovery of biliary excretion of drugs to be obtained under physiological conditions in man. The aim of this study was to develop and validate a method for simultaneous measurement of gall bladder storage of a cholephilic drug, and of its duodenal excretion and t1/2 in bile. Duodenal perfusion using polyethylene glycol as intestinal recovery marker for measurement of drug duodenal excretion, with an iv bolus of 99mTc HIDA for measurement of drug mass within the gall bladder was used. Gall bladder volume was measured by ultrasonography. T1/2 in bile was measured by relating drug duodenal excretion to that of bile acid used as an endogenous bile marker. The use of bile acid as biliary marker was validated in two subjects receiving simultaneous iv infusion of indocyanine green. Seven healthy subjects were studied using a beta-lattam antibiotic, Cefotetan 1 g iv, as test drug. Median values during the study period (seven hours) were 51.1 mg for Cefotetan duodenal excretion, 45.2 mg for gall bladder mass and 2.8 mg/ml for concentration within the gall bladder. T1/2 of the drug in bile was 100 minutes. This technique enables measurement of mass and concentration of drugs within the gall bladder to be carried out, in addition to measurements of t1/2 of drugs in bile. These measurements may have specific application for assessment of potential efficacy of antibiotics in biliary tract infections, as well as general application for assessment of biliary excretory kinetics of drugs.