Pathogenesis of human cholesterol cholelithiasis.

The pathogenesis of cholesterol cholelithiasis in humans has been studied by means of three techniques. The cholesterol-solubilizing capacity of bile may be determined by estimation of the relative composition of the three major lipid constituents of bile. Consistent reduction in the cholesterol-carrying capacity of gallbladder bile of persons with gallstones when compared with normal subjects has not been shown. Normal subjects frequently have supersaturated bile. Secretion rates of biliary lipids have been estimated by two methods; with the method that appears to be more physiologic no change in lipid secretion rates was found in gallstone patients. Bile acid pool size has been measured by isotope dilution techniques; it is reduced in patients with gallstones. It is not clear whether this reduction is important in the pathogenesis of cholesterol cholelithiasis, for the bile acid secretion rate is normal because of an increased rate of cycling of the pool through the enterohepatic circulation. The role of the gallbladder in the genesis of cholesterol cholelithiasis may be more important than has been realized.     

成石前后胆囊粘膜上皮多聚核蛋白体RNA含量变化—胆液糖蛋白和蛋白质增加的机制探讨

胆囊淤泥是胆囊结石的前身。测定胆囊粘膜上皮游离(FPR)和结合多聚核蛋白体(MBPR)的RNA含量及胆液粘液糖蛋白(MGP)的结果表明:胆囊淤泥患者胆囊粘膜合成和分泌MGP明显亢进,MBPR与相应GB中MGP的相关性比较提示此时尚有胆液淤积存在,且FPR的RNA含量也显著增高,提示细胞增殖加快。荧光胺法测定胆液蛋白质的含量发现,胆囊淤泥和胆固醇性结石患者胆囊液GB蛋白质含量显著高于色素性结石患者及“正常”对照,但蛋白质的GB浓度与HB浓度的比值显著低于胆固醇GB/HB比值,且GB中MGP与蛋白质含量呈显著正相关。说明蛋白质含量的增加与GB中过高的MGP阻止胆囊中蛋白质的清除过程有关。成石前GB中MGP和蛋白质含量的同时增高及其机制的确立,对认识及预防体内结石均有意义。     

Comparison of the Coomassie brilliant blue, bicinchoninic acid and Lowry quantitation assays, using non-glycosylated and glycosylated proteins.

The concentrations of several non-glycosylated and glycosylated recombinant and native proteins were determined by three widely used colorimetric methods: Coomassie brilliant blue, bicinchoninic acid and Lowry, and, for comparison, by amino acid composition analysis. The colorimetric methods gave results differing from the values derived from the amino acid analysis, in some cases by up to 60%. For the non-glycosylated recombinant proteins, the results were in relatively good agreement with each other and with the values determined on the basis of the amino acid analysis. The Coomassie blue method was strongly dependent on the hydrophobicity of the individual protein. The bicinchoninic acid method gave results closest to those of the amino acid analysis. For the glycosylated proteins, both recombinant and native, the Coomassie blue assay gave values lower, whereas the two other methods gave values higher than those determined on the basis of the amino acid analysis. The concentration of a recombinant interferon gamma receptor produced in two differently glycosylated forms was underestimated by the Coomassie blue assay and overestimated by the bicinchoninic acid and Lowry methods, while for the non-glycosylated form of the same protein, the three colorimetric methods delivered comparable values. The results suggest a potential interference of protein glycosylation with the colorimetric assays.     

Hepatic cholesterol metabolism in cholesterol gallstone disease

Hepatic cholesterol metabolism was examined in 27 Swedish patients with cholesterol gallstone disease and in 13 patients free of gallstones operated for roentgenographically suspect polyps in the gallbladder. All 40 patients underwent cholecystectomy, and a liver biopsy and gallbladder bile were obtained at surgery. The cholesterol saturation of gallbladder bile was significantly higher in patients with gallstones compared to the gallstone-free controls (131 +/- 13 vs. 75 +/- 5%, P less than 0.001). Microsomal 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity, governing cholesterol synthesis, did not differ between gallstone and gallstone-free patients (104 +/- 11 vs. and 109 +/- 22 pmol/min per mg protein, respectively). The activity of cholesterol 7 alpha-hydroxylase, catalyzing the catabolism of cholesterol to bile acids, was not significantly decreased in gallstone patients (6.2 +/- 1.1 vs. 8.0 +/- 2.0 pmol/min per mg protein). The capacity to esterify cholesterol, judged by the activity of acyl coenzyme A:cholesterol acyltransferase (ACAT), was similar in gallstone and gallstone-free patients (5.4 +/- 0.4 vs. 6.7 +/- 1.1 pmol/min per mg protein). In the presence of exogenous cholesterol, ACAT activity increased by more than fourfold in both groups. No correlation was found between the saturation of gallbladder bile and any of the mentioned enzyme activities in gallstone patients. It is concluded that distinct abnormalities in cholesterol metabolizing enzymes are not of major importance for development of gallstones in Swedish patients with cholesterol gallstone disease. The results support the contention that the etiology of cholesterol gallstones is multifactorial.     

Cholesterol gallstones in alloxan-diabetic mice

Normal and alloxan-diabetic male mice (Crj-ICR) were fed a diet containing 0.5% cholesterol for 5 and 10 weeks, and gallbladder bile was analyzed for cholesterol, phospholipids and bile acids, feces for sterols and bile acids, and plasma and liver for cholesterol, phospholipids, and triglycerides. Normal mice developed no gallstones but the diabetic mice developed cholesterol gallstones with an incidence of 70% by 5 weeks and 80% by 10 weeks after feeding of the cholesterol diet. Diabetic mice fed the ordinary diet also developed stones (23%) by 10 weeks. In the diabetic mice, the gallbladder was enlarged about threefold, and biliary lipid concentration, diet intake, and fecal excretion of sterols and bile acids increased but body weight decreased. Cholic acid and beta-muricholic acid comprised over 40% each of the total biliary bile acids in normal mice, but cholic acid increased to about 80% and beta-muricholic acid decreased to a few percent in the diabetic mice. Fecal excretion of bile acids increased after cholesterol feeding in both normal and diabetic mice, but the increased bile acid in the normal animals was beta-muricholic acid and that in the diabetic mice was deoxycholic acid. The mice that developed gallstones showed a marked increase in biliary cholesterol value and decreases in gallbladder bile and bile acid concentration, but no difference in biliary and fecal bile acid composition, bile acid synthesis, fecal sterols, or plasma and liver lipid levels. Cholesterol absorption was increased in the diabetic mice when examined by plasma 14C/3H ratio and fecal 14C-labeled sterol excretion after a single oral administration of [14C]cholesterol and a simultaneous intravenous injection of [3H]cholesterol. These data led to the conclusion that cholesterol gallstones developed in alloxan-diabetic mice fed excess cholesterol, due to the hyperphagia and the enhancement of cholesterol absorption caused by increases in the synthesis and secretion of cholic acid.     

Characterization of the bile acid profile in developing male and female hamsters in response to dietary cholesterol challenge

The Syrian golden hamster is a frequently used model to study cholesterol and bile acid metabolism as well as cholesterol-induced cholelithiasis. However, diet-induced gallstones seem limited to young male hamsters of certain strains that develop depressed cholate/chenodeoxycholate bile acid ratios. To further elucidate gender and age specific aspects of cholesterol and bile acid metabolism, i.e. a possible age-related bile acid/gallstone relationship, plasma and biliary lipids and bile acid composition were analyzed in male and female hamsters under various physiological conditions of age and diet, the latter formulated with and without dietary cholesterol. During normal development (no cholesterol challenge) the percentage of cholic acid decreased while chenodeoxycholate increased, the shift being more pronounced in males. Furthermore, female hamsters had higher total plasma cholesterol than in males, while hepatic and biliary lipids did not differ. When challenged with excessive dietary cholesterol, female hamsters again developed significantly higher total plasma and hepatic cholesterol concentrations. Biliary lipids and cholesterol gallstone incidence revealed a significant gender effect with male hamsters developing a higher lithogenic index and more gallstones (cholesterol and pigment stones) than females. Female hamsters revealed a lower percentage of chenodeoxycholate and a higher percentage of cholate resulting in a more protective, higher cholate/cheno ratio (1.5 +/- 1.0) than in males (1.0 +/- 0.2). In summary, the bile acid pattern in developing and cholesterol-fed hamsters renders females less susceptible to gallstones, in part because they maintain more favorable biliary lipid and bile acid profiles, characterized by lower molar percentages of biliary cholesterol and chenodeoxycholate.     

Nucleation of cholesterol crystals from native bile and the effect of protein hydrolysis

Nucleation time represents the terminal step in in vitro studies examining bile lithogenicity. Because of the concern that residual microcrystals, left after ultracentrifugation, may be responsible for the rapid nucleation time of gallbladder bile from patients with cholesterol gallstones, we have included a final filtration step. However, we found this procedure to considerably lengthen the nucleation time of abnormal biles. In view of the central importance of the nucleation assay we compared the effect of three commonly used gallbladder bile pre-treatment regimes (designed to remove endogenous crystals) on nucleation time. They were: a) immediate filtration of bile (0.22 micron filter); b) ultracentrifugation; and c) ultracentrifugation followed by filtration. The respective nucleation times were: a) 9.3 +/- 3.7 days, n = 6; b) 2.9 +/- 0.4 days, n = 10; c) 12.8 +/- 2.3 days, n = 11. To determine whether the dramatic change in nucleation time was due to the removal of components other than seed crystals, we examined the mucus content, the total lipid composition of bile, and that of its cholesterol transport components following the different pre-treatments. No significant difference in total lipid, percentage cholesterol carried by the transport components, or their cholesterol/phospholipid ratio were found. Ultracentrifugation alone was sufficient to removal all detectable large molecular weight mucus glycoprotein. Although nucleation time of the abnormal gallbladder samples was extended in the ultracentrifuged/filtered biles, it was still significantly different (P less than 0.01) from that of normal gallbladder biles, confirming an intrinsic difference between abnormal and normal biles, in cholesterol metastability. We also examined the effect of protein digestion on the nucleation time of native biles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immunoglobulins as nucleating proteins in the gallbladder bile of patients with cholesterol gallstones.

The gallbladder bile of patients with cholesterol gallstones contains pronucleating proteins which accelerate precipitation of cholesterol crystals from bile. In this study we have improved the purification procedure developed earlier for these nucleating proteins and have now identified the nature of these proteins. Gallbladder bile from patients with cholesterol gallstones was applied to concanavalin A affinity columns. The ConA-binding glycoprotein fractions containing the nucleating proteins were then separated by FPLC (fast protein liquid chromatography) using a Superose 12 gel filtration column. Nucleating activity was detected in the high molecular weight (FPLC-1) as well as in the low molecular weight fractions (FPLC-3). Investigation of the high molecular weight fraction by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by electroelution and amino acid sequencing suggested that these proteins were immunoglobulins. Immunostaining of Western blots with specific monoclonal antibodies identified the presence of immunoglobulin (Ig) M and IgA in the FPLC-1 fraction. These immunoglobulins were further purified by affinity chromatography employing an antibody exchanger (ABx) column which specifically binds immunoglobulins. There was no reduction in the cholesterol nucleating activity in the Abx-bound fraction compared to FPLC-1. Additional studies showed that the FPLC-1 fraction was significantly more potent than the ConA glycoproteins from either rapid and slow nucleating biles. Also the number of crystals formed was significantly greater in the FPLC-1 fraction isolated from cholesterol gallstone biles than from the FPLC-1 fraction from control patient biles. Commercially obtained IgM and IgA had no effect on nucleation, but IgM isolated from the serum of patients with Waldenstrom's macroglobulinemia did accelerate the nucleation of cholesterol. We conclude that the IgM and possibly IgA are pronucleating proteins and may be important in the pathogenesis of cholesterol gallstones in man.     

Effect of dietary garlic and onion on biliary proteins and lipid peroxidation which influence cholesterol nucleation in bile

Formation of cholesterol gallstones in gallbladder is controlled by procrystallizing and anticrystallizing factors present in bile. Dietary garlic and onion have been recently observed to possess anti-lithogenic potential in experimental mice. In this investigation, the role of biliary proteins from rats fed lithogenic diet or garlic/onion-containing diet in the formation of cholesterol gallstones in model bile was studied. Cholesterol nucleation time of the bile from lithogenic diet group was prolonged when mixed with bile from garlic or onion groups. High molecular weight proteins of bile from garlic and onion groups delayed cholesterol crystal growth in model bile. Low molecular weight (LMW) proteins from the bile of lithogenic diet group promoted cholesterol crystal growth in model bile, while LMW protein fraction isolated from the bile of garlic and onion groups delayed the same. Biliary LMW protein fraction was subjected to affinity chromatography using Con-A and the lectin-bound and unbound fractions were studied for their influence on cholesterol nucleation time in model bile. Major portion of biliary LMW proteins in lithogenic diet group was bound to Con-A, and this protein fraction promoted cholesterol nucleation time and increased cholesterol crystal growth rate, whereas Con-A unbound fraction delayed the onset of cholesterol crystallization. Biliary protein from garlic/onion group delayed the crystallization and interfered with pronucleating activity of Con-A bound protein fraction. These data suggest that apart from the beneficial modulation of biliary cholesterol saturation index, these Allium spices also influence cholesterol nucleating and antinucleating protein factors that contribute to their anti-lithogenic potential.     

Cholesterol, bile acid, and lipoprotein metabolism in two strains of hamster, one resistant, the other sensitive (LPN) to sucrose-induced cholelithiasis

A comprehensive study of cholesterol, bile acid, and lipoprotein metabolism was undertaken in two strains of hamster that differed markedly in their response to a sucrose-rich/low fat diet. Under basal conditions, hamsters from the LPN strain differed from Janvier hamsters by a lower cholesterolemia, a higher postprandial insulinemia, a more active cholesterogenesis in both liver [3- to 4-fold higher 3-hydroxy 3-methylglutaryl coenzyme A reductase (HMG-CoAR) activity and mRNA] and small intestine, and a lower hepatic acyl-coenzyme A:cholesterol acyltransferase activity. Cholesterol saturation indices in the gallbladder bile were similar for both strains, but the lipid concentration was 2-fold higher in LPN than in Janvier hamsters. LPN hamsters had a lower capacity to transform cholesterol into bile acids, shown by the smaller fraction of endogenous cholesterol converted into bile acids prior to fecal excretion (0.34 vs. 0.77). In LPN hamsters, the activities of cholesterol 7alpha-hydroxylase (C7OHase) and sterol 27-hydroxylase (S27OHase), the two rate-limiting enzymes of bile acid synthesis, were disproportionably lower (by 2-fold) to that of HMG-CoAR. When fed a sucrose-rich diet, plasma lipids increased, dietary cholesterol absorption improved, hepatic activities of HMG-CoA reductase, C7Ohase, and S27OHase were reduced, and intestinal S27OHase was inhibited in both strains. Despite a similar increase in the biliary hydrophobicity index due to the bile acid enrichment in chenodeoxycholic acid and derivatives, only LPN hamsters had an increased lithogenic index and developed cholesterol gallstones (75% incidence), whereas Janvier hamsters formed pigment gallstones (79% incidence).These studies indicate that LPN hamsters have a genetic predisposition to sucrose-induced cholesterol gallstone formation related to differences in cholesterol and bile acid metabolism.     

Protein-blotting on Polybrene-coated glass-fiber sheets. A basis for acid hydrolysis and gas-phase sequencing of picomole quantities of protein previously separated on sodium dodecyl sulfate/polyacrylamide gel

A procedure has been developed which allows the immobilization on glass-fiber sheets coated with the polyquaternary amine, Polybrene, of proteins and protein fragments previously separated on sodium-dodecylsulfate-containing polyacrylamide gels. The transfer is carried out essentially as has been used for protein blotting on nitrocellulose membranes [Towbin, H., Staehelin, T. and Gordon, J. (1979) Proc. Natl Acad. Sci. USA 76, 4350-4354], but is now used to determine the amino acid composition and partial sequence of the immobilized proteins. Protein transfer could be carried out after staining the proteins in the gels with Coomassie blue, by which immobilized proteins are visible as blue spots, or without previous staining, after which transferred proteins are detected as fluorescent spots following reaction with fluorescamine. The latter procedure was found to be more efficient and yielded binding capacities of +/- 20 micrograms/cm2. Fluorescamine detection was of equal or higher sensitivity than the classical Coomassie staining of proteins in the gel. Immobilized proteins could be hydrolyzed when still present on the glass fiber and reliable amino acid compositions were obtained for various reference proteins immobilized in less than 100 pmol quantities. In addition, and more importantly, glass-fiber-bound proteins could be subjected to the Edman degradation procedure by simply cutting out the area of the sheet carrying the immobilized protein and mounting the disc in the reaction chamber of the gas-phase sequenator. Results of this immobilization-sequencing technique are shown for immobilized myoglobin (1 nmol) and two proteolytic fragments of actin (+/- 80 pmol each) previously separated on a sodium-dodecylsulfate-containing gel.     

Identification of a phosphatidylcholine active phospholipase C in human gallbladder bile

This study describes the identification of a phospholipase C activity against phosphatidylcholine in delipidated human gallbladder bile. All biles were obtained from cholesterol gallstone patients and were negative on bacterial culture. The biliary enzyme was inhibited by EDTA and had a pH optimum of between 7-8. All of the 15 gallbladders examined contained significant phospholipase C activity (32.85 +/- 8.37 nmol/h/mg delipidated protein). The finding of a phospholipase C in gallbladder bile of patients with cholesterol gallstones may be one of the factors responsible for or related to the rapid in vitro nucleation seen in these biles.     

Extraction of fixed, stained protein bands from gels for micro amino acids analysis using o-phthaldialdehyde.

A method is presented for extraction of fixed, stained protein bands from polyacrylamide gels suitable for automated fluorescence analysis of amino acids using o-phthaldialdehyde. Bands, containing microgram quantities of protein and stained with Coomassie blue, are extracted from homogenized gel slices with sodium dodecyl sulfate. The Coomassie blue and sodium dodecyl sulfate do not interfere with the amino acid determination, and contamination by ammonia from the gels is low. The method has been applied to the analysis of human carbonic anhydrase C, and the amino acid composition is found to be similar to that obtained by other methods requiring larger amounts of protein.     

Determination of proteins with the Coomassie brilliant blue G 250 method. III. Behavior of various pure proteins and comparative analysis with the biuret and Lowry methods

Some proteins were tested by the Bradford's method with Coomassie Brilliant Blue G 250. The findings were compared to those obtained by the Lowry's and the biuret methods. Coomassie, as the other methods, has the inconvenient of giving different absorbaces according to the nature of single protein. The use of standards of analogous composition in assaying proteins by the Bradford's method is suggested.     

Comparative analysis of cholesterol transport in bile from patients with and without cholesterol gallstones

Aggregation of cholesterol-phospholipid vesicles in supersaturated biles precedes cholesterol crystal formation. In this study we examined the relationship between the percentage of cholesterol carried by vesicles and/or their composition and the propensity to form cholesterol crystals (nucleation time). Bile (common bile duct, gallbladder and T-tube) was obtained from patients with and without gallstones. Gel filtration chromatography resolved three peaks, a void volume vesicle, a smaller vesicle (identified by electron microscopy and of distinct composition compared to the larger void volume vesicle), and the mixed micelle. The void volume vesicle was present in 11 of 28 abnormal gallbladder biles, but in none of the 10 normal gallbladder biles. Despite this difference, no correlation between the nucleation time of whole bile with either the percentage of cholesterol carried by or cholesterol/phospholipid ratio of the void volume vesicle was found. Nucleation time was, however, found to correlate with the composition of the small-vesicular transport form. No significant difference in the composition or percentage of the small-vesicular form or the combined vesicular forms was found between normal and abnormal gallbladder biles, although the latter nucleated significantly more rapidly. Our results confirm the importance of vesicles in the nucleation process but suggest that other factors, not yet identified, appear to be responsible for the more rapid nucleation seen in abnormal gallbladder biles.     

Chemical Characterization of Gallstones: An Approach to Explore the Aetiopathogenesis of Gallstone Disease in Sri Lanka

Introduction

Records on gallstones and associated ailments in Sri Lankan community are scarce, despite frequent detection of gallstone disease. Identification of the chemical composition of gallstones in the local setting is important in defining aetiopathogenic factors which in turn are useful in implementing therapeutic and preventive strategies. This study aimed to describe the chemical composition of gallstones and the socio-demographic factors of a cohort of Sri Lankan patients with gallstone disease.

Materials and Methods

Data on clinical and socio-demographic factors, and gallstones removed at surgery were collected from patients with cholelithiasis admitted to Teaching Hospital, Peradeniya, Sri Lanka from May 2011 to December 2012. External and cross sectional morphological features of gallstones were recorded by naked eye observation. Compositional analysis was carried out by Fourier Transform Infrared Spectroscopy, X - ray Powder Diffraction, and Atomic Absorption Spectrophotometry. Scanning Electron Microscopy was used to identify the microstructure of gallstones.

Results

Data of 102 patients were analyzed. Of them majority (n = 77, 76%) were females with a female: male ratio of 3:1. Mean age of the study group was 46.1±11.6 years. All the patients had primary gallbladder stones. According to the physical and chemical analysis, majority (n = 54, 53%) were pigment gallstones followed by mixed cholesterol gallstones (n = 38, 37%). Only 10 (9%) had pure cholesterol gallstones. Calcium bilirubinate, calcium carbonate and calcium phosphate were the commonest calcium salts identified in pigment gallstones and core of mixed cholesterol gallstones.

Conclusion

Presence of a pigment nidus in gallstones is a common feature in majority of Sri Lankan patients denoting the possible role of elevated unconjugated bilirubin in bile on the pathogenesis of GS. Hence it is imperative to explore this further to understand the aetiopathogenesis of GS among Sri Lankans.     

Contraceptive steroids increase cholesterol in bile: mechanisms of action

Contraceptive steroids increase the risk of acquiring cholesterol gallstones. The factors responsible include an increase in cholesterol saturation of bile and an increase in rate of secretion of cholesterol into bile. The goal of this study was to investigate the mechanism(s) of these increases in biliary cholesterol. During the use of contraceptive steroids, cholesterol saturation of gallbladder bile and the amount of cholesterol secreted per mole of bile acid increased (P less than 0.05 and P less than 0.02, respectively). Cholesterol absorption, cholesterol synthesis, chylomicron remnant clearance, and the concentration of plasma and lipoprotein lipids were not altered by contraceptive steroids. Despite this apparent lack of effect, important correlations were present during steroid use. LDL (low density lipoprotein) cholesterol increased as dietary cholesterol increased (r = 0.58, P less than 0.025). Cholesterol synthesis correlated directly with VLDL cholesterol concentration (r = 0.64, P less than 0.01), biliary cholesterol secretion (r = 0.68, P less than 0.01) and with molar percent cholesterol in bile (r = 0.49, P = 0.06). Chylomicron remnant clearance also correlated with cholesterol secretion (r = 0.85, P less than 0.001). As either remnant uptake or synthesis increased, the effect of the other source of hepatic cholesterol on biliary cholesterol secretion diminished. These relationships were not observed in the same subjects when they were not taking the hormones. The findings suggest that both newly synthesized and dietary cholesterol contribute to the cholesterol secreted in bile. This is consistent with the hypothesis that cholesterol for secretion into bile and VLDL is derived from a common metabolic pool of free cholesterol. It is proposed that contraceptive steroids exert their effect on biliary cholesterol by increasing cholesterol entering the pool and/or by inhibiting hepatic ACAT (acylcoenzyme A:cholesterol acyltransferase) activity, a known effect of progesterone, so that an increase in free cholesterol entering the pool leads to an increase in output.     

Cholecystectomy prevents expansion of the bile acid pool and inhibition of cholesterol 7alpha-hydroxylase in rabbits fed cholesterol

To study the effect of cholecystectomy on the regulation of classic and alternative bile acid syntheses, gallbladder-intact (n = 20) and cholecystectomized (n = 20) New Zealand White rabbits were fed either chow or chow with 2% cholesterol (3 g/day). After 10 days, bile fistulas were constructed in half of each rabbit group to recover and measure the bile acid pool and biliary bile acid flux. After cholesterol feeding, the bile acid pool size increased from 268 +/- 55 to 444 +/- 77 mg (P < 0.01) with a 2-fold rise in the biliary bile acid flux in intact rabbits but did not expand the bile acid pool (270 +/- 77 vs. 276 +/- 62 mg), nor did the biliary bile acid flux increase in cholecystectomized rabbits. Ileal apical sodium-dependent bile acid transporter protein increased 46% from 93 +/- 6 to 136 +/- 23 units/mg (P < 0.01) in the intact rabbits but did not change in cholecystectomized rabbits (104 +/- 14 vs. 99 +/- 19 units/mg) after cholesterol feeding. Cholesterol 7alpha-hydroxylase activity was inhibited 59% (P < 0.001) while cholesterol 27-hydroxylase activity rose 83% (P < 0.05) after cholesterol feeding in the intact rabbits but neither enzyme activity changed significantly in cholesterol-fed cholecystectomized rabbits. Fecal bile acid outputs reflecting bile acid synthesis increased significantly in the intact but not in the cholecystectomized rabbits fed cholesterol.Removal of the gallbladder prevented expansion of the bile acid pool after cholesterol feeding as seen in intact rabbits because ileal bile acid transport did not increase. As a result, cholesterol 7alpha-hydroxylase was not inhibited.     

Variation in Hepatic Bile Composition Following Cholecystectomy in Patients with Previous Gallstones

The composition of fasting hepatic bile was analyzed in 63 samples from 8 patients following cholecystectomy to determine if bile was lithogenic in patients with previous cholesterol gallstones after removal of the gallbladder. Bile specimens were obtained from t-tubes over a 7-20 day study period following re-establishment of the enterohepatic circulation. Bile composition varied on a day to day basis in each patient. 18 of 63 samples were lithogenic according to criteria of Admirand and Small while 35 of 63 samples were lithogenic according to criteria of Hegardt and Dam. Variations in the composition of hepatic bile appeared related to changes in the excretion rate of bile acids. These studies demonstrate that hepatic bile may be lithogenic after cholecystectomy and indicate that factors other than sequestration of the bile acid pool in the gallbladder influence the enterohepatic circulation of bile acids and the lithogenicity of bile.     

Antilithiasic effect of beta-cyclodextrin in LPN hamster: comparison with cholestyramine

Beta-Cyclodextrin (BCD), a cyclic oligosaccharide that binds cholesterol and bile acids in vitro, has been previously shown to be an effective plasma cholesterol lowering agent in hamsters and domestic pigs. This study examined the effects of BCD as compared with cholestyramine on cholesterol and bile acid metabolism in the LPN hamster model model for cholesterol gallstones. The incidence of cholesterol gallstones was 65% in LPN hamsters fed the lithogenic diet, but decreased linearly with increasing amounts of BCD in the diet to be nil at a dose of 10% BCD. In gallbladder bile, cholesterol, phospholipid and chenodeoxycholate concentrations, hydrophobic and lithogenic indices were all significantly decreased by 10% BCD. Increases in bile acid synthesis (+110%), sterol 27-hydroxylase activity (+106%), and biliary cholate secretion (+140%) were also observed, whereas the biliary secretion of chenodeoxycholate decreased (-43%). The fecal output of chenodeoxycholate and cholate (plus derivatives) was increased by +147 and +64%, respectively, suggesting that BCD reduced the chenodeoxycholate intestinal absorption preferentially. Dietary cholestyramine decreased biliary bile acid concentration and secretion, but dramatically increased the fecal excretion of chenodeoxycholate and cholate plus their derivatives (+328 and +1940%, respectively). In contrast to BCD, the resin increased the lithogenic index in bile, induced black gallstones in 34% of hamsters, and stimulated markedly the activities of HMG-CoA reductase (+670%), sterol 27-hydroxylase (+310%), and cholesterol 7alpha-hydroxylase (+390%). Thus, beta-cyclodextrin (BCD) prevented cholesterol gallstone formation by decreasing specifically the reabsorption of chenodeoxycholate, stimulating its biosynthesis and favoring its fecal elimination. BCD had a milder effect on lipid metabolism than cholestyramine and does not predispose animals to black gallstones as cholestyramine does in this animal model.