Structure of biliary phosphatidylcholine in cholesterol gallstone patients

The fatty acid composition of biliary phosphatidylcholine was analyzed in 13 patients with radiolucent gallstones undergoing elective cholecystectomy, and in 11 normolipemic patients without gallstones undergoing abdominal surgery. The only difference in the percentage fatty acid composition between the two groups was a significantly (p<0.05) higher percentage arachidonic acid in the first group. This acid was exclusively located in thesn-2 position of phosphatidylcholine (PC), accounting for 13.0±4.9% in the first group and 8.2±4.9% in the second (p<0.05). The percentage arachidonic acid of PC was negatively correlated (p<0.001) with the percentage biliary chenodeoxycholate in gallstone patients, but not in controls. Explanation of these findings is, at present, only speculative.     

Increased biliary calcium in cholesterol and pigment gallstone disease: The role of altered bile acid composition

The present study was undertaken to define the relationship between calcium metabolism and bile acid composition in animal models of diet induced cholesterol and pigment gallstones. Groups of prairie dogs were fed either a control non-lithogenic chow (N=12), a 1.2% cholesterol enriched chow (N=6, XOL) for two weeks, or a high carbohydrate diet deficient in iron (N=6, CHO-FeD), or a high carbohydrate diet with normal iron levels (N=6, CHO) for eight weeks. Hepatic (HB) and gallbladder (GB) bile samples were analyzed for total calcium, cholesterol, phospholipids, total bile acids (TBA), and individual bile acid composition. In each of the four groups, TBA concentrations were essentially similar and taurine conjugates accounted for approximately 90% of TBA in HB bile and about 98% in GB bile. In the control group, cholic acid (CA) was the predominant bile acid and comprised 76% of TBA and chenodeoxycholic (CDCA) accounted for about 13% of the total. Feeding a diet rich in cholesterol caused a significant change in the relative concentrations of individual bile acids of hepatic bile—such that CA decreased significantly (p<0.001) while CDCA increased by 300% (p<0.001). The changes in secondary bile acids were insignificant. An identical shift in individual bile acid composition was noted in animals maintained on high carbohydrate diet, irrespective of iron content. Similar changes were observed in the GB in the experimental groups. Calcium concentrations of GB bile with or without gallstone formation showed a positive linear relationship with TBA (y=4.35+0.14X, p<0.001) and taurochenodoxycholic acid (TCDCA) (y=15.04+0.46X, p<0.001), but an inverse relationship with taurocholic acid (TCA) (Y=55.16−0.41X, p<0.008). However, such relationships were absent in hepatic bile. These data indicate that diet-induced alterations in bile acid composition may modify calcium solubility or GB function, thereby contributing to the increased GB calcium observed during cholesterol and pigment gallstone formation.     

Oxidized cholesterol modulates age-related change in lipid metabolism in rats

For three weeks, male Sprague-Dawley rats at either four weeks (young) or eight months (adult) of age were pairfed one of the purified diets free of or containing either 0.2% of oxidized cholesterol mixture (cholesterol oxidation products) or 0.2% of cholesterol. Although the food intake was similar, dietary oxidized cholesterol lowered body weight gain in young rats, but did not increase relative liver weight, in contrast to the enlargement seen with dietary cholesterol. Oxidized cholesterol, compared to cholesterol, tended to reduce the activity of hepatic 3-hydroxy-3-methylglutaryl CoA reductase and cholesterol 7α-hydroxylase, particularly the latter in aged rats, and prevented the rise in the concentration of liver cholesterol at both ages. It also tended to increase the activity of hepatic δ6 desaturase, particularly in young rats. Moreover, oxidized cholesterol in relation to cholesterol influenced liver and serum lipid concentrations in different ways, and increased lipid peroxidation at both ages. The ratio of splenic CD4⁺/CD8⁺ T-lymphocytes increased with age, but the influence of cholesterol and oxidized cholesterol was comparable. Thus, oxidized cholesterol may specifically disturb growth and age-related changes in the lipid metabolism in rats.     

The effects of a dietary oxidized oil on lipid metabolism in rats

This study was carried out to investigate the effects of a dietary oxidized oil on lipid metabolism in rats, particularly the desaturation of fatty acids. Two groups of rats were fed initially for a period of 35 d diets containing 10% of either fresh oil or thermally treated oil (150°C, 6d). The dietary fats used were markedly different for lipid peroxidation products (peroxide value: 94.5 vs. 3.1 meq O₂/kg; thiobarbituric acid-reactive substances: 230 vs. 7 μmol/kg) but were equalized for their fatty acid composition by using different mixtures of lard and safflower oil and for tocopherol concentrations by individual supplementation with dl-α-tocopherol acetate. In the second period which lasted 16 d, the same diets were supplemented with 10% linseed oil to study the effect of the oxidized oil on the desaturation of α-linolenic acid. During the whole period, all the rats were fed identical quantities of diet by a restrictive feeding system in order to avoid a reduced food intake in the rats fed the oxidized oil. Body weight gains and food conversion rates were only slightly lower in the rats fed the oxidized oil compared to the rats fed the fresh oil. Hence, the effects of lipid peroxidation products could be studied without a distortion by a marked reduced food intake and growth. To assess the rate of fatty acid desaturation, the fatty acid composition of liver and heart total lipids and phospholipids was determined and ratios between product and precursor of individual desaturation reactions were calculated. Rats fed the oxidized oil had reduced ratios of 20∶4n−6/18∶2n−6, 20∶5n−3/18∶3n−3, 20∶4n−6/20∶3n−6, and 22∶6n−3/22∶5n−3 in liver phospholipids and reduced ratios of 20∶4n−6/18∶2n−6, 22∶5n−3/18∶3n−3, and 22∶6n−3/18∶3n−3 in heart phospholipids. Those results suggest a reduced rate of desaturation of linoleic acid and α-linolenic acid by microsomal Δ4-, Δ5-, and Δ6-desaturases. Furthermore, liver total lipids of rats fed the oxidized oil exhibited a reduced ratio between total monounsaturated fatty acids and total saturated fatty acids, suggesting a reduced Δ9-desaturation. Besides those effects, the study observed a slightly increased liver weight, markedly reduced tocopherol concentrations in liver and plasma, reduced lipid concentrations in plasma, and an increased ratio between phospholipids and cholesterol in the liver. Thus, the study demonstrates that feeding an oxidized oil causes several alterations of lipid and fatty acid metabolism which might be of great physiologic relevance.     

The effect of neomycin on cholesterol metabolism

Oral neomycin lowers serum cholesterol in man, whereas intramuscular doses are ineffective. Probably it exerts its effect in the gastrointestinal tract. It has been suggested that neomycin specifically involves alteration of the intestinal flora. An increased excretion of bile acids and the failure of the conversion of cholic acid to deoxycholic acid has been demonstrated, which supports this hypothesis. It has also been suggested that the decrease in serum cholesterol is related to a malabsorption syndrome. Although large doses of neomycin do cause a malabsorption syndrome there is no agreement as to the occurrence or severity of a malabsorption syndrome related to therapeutic doses of neomycin.     

Dietary lipid,fatty acid synthesis and cholesterol metabolism in aging rats

Male and female rats were fed diets containing 2% of calories as corn oil or that plus 40% of calories as beef tallow or corn oil. After 3, 6, 12 and 18 months groups were given 4-¹⁴C-cholesterol ip, and feces were collected for 9 days. Just prior to necropsy³H-acetate was administered ip. Samples of serum, liver, heart and carcass were obtained for analysis. Concentrations of fatty acids and cholesterol, synthesis of those and recovery of ring-labeled steroid are reported. Mortality from acute respiratory disease was very high in male rats fed beef tallow or low fat diets and very low in those fed the corn oil diet. In females, only beef tallow diet resulted in a high mortality rate, and this was lower and at a later age than in males. The most notable effects of age were in relation to fatty acid synthesis and presence of¹⁴C-acidic steroid in the carcass. In 3-month-old rats both fats depressed fatty acid synthesis in comparison to the low fat diet. At later ages beef fat ceased to depress fatty acid synthesis in both sexes. Corn oil continued to depress fatty acid synthesis up to 12 months in males and 18 months in females. The presence of¹⁴C-acidic steroid in carcass was substantial in 6-month-old rats and constituted ca. 40% of recovered¹⁴C in 18-month-old rats. The possibility that the increase in acetate incorporation into fatty acids with age in fat feeding is related to chain elongation rather than de novo synthesis is discussed. Both the presence and amount of acidic steroid in the carcass are notable and may be of importance in constructing models of cholesterol turnover. Presented in part at the AOCS Sterol Symposium, April 1970, and the Federation of American Societies for Experimental Biology, April 1971. Scientific Series Paper No. 1536, Colorado Agricultural Experiment Station.     

The effects of clofibrate and bezafibrate on cholesterol metabolism in the liver of the male rat

Fibric acid derivatives are used to treat hyperlipidemia and have wide ranging effects on lipid metabolism. The action of these compounds on cholesterol esterification, catalyzed by acyl-coenzyme A:cholesterol acyltransferase (ACAT), has been quite widely studied, but their effect on cholesteryl ester hydrolysis and the enzyme neutral cholesteryl ester hydrolase (nCEH) has been largely ignored. Male rats were therefore fed for 10 d on a standard chow diet supplemented with either clofibrate or bezafibrate, to study their effects on plasma lipid levels and hepatic cholesterol metabolism. Plasma triacyglycerols were not significantly altered by these diets, but bezafibrate significantly lowered plasma cholesterol levels (29.7%,P<0.01). When expressed per unit weight of DNA, both fibrates reduced the hepatic content of triacylglycerol, cholesterol and cholesteryl esters (40, 18.7, 16.5 and 66.7, 28.6, 34.2% for clofibrate and bezafibrate, respectively). ACAT activity was significantly reduced by both drugs, but clofibrate (65% inhibition) was more effective than bezafibrate (35% inhibition). The most dramatic effect of the diets was a marked increase in the activity of both the microsomal and the cytosolic nCEH. When expressed on a whole liver basis, the effect of bezafibrate on the cytosolic enzyme (13.6-fold increase in activity) was much greater than that of clofibrate (4.8-fold increase). Increases in the activity of a cytosolic protein that inhibits the activity of nCEH were also noted, but these changes were relatively small. The results suggest that the activation of nCEH, in combination with the inhibition in ACAT activity, contributes to a decrease in the cholesteryl ester content of the liver which may influence the secretion of very low density lipoprotein.     

The combined effects of dietary proteins and fish oil on cholesterol metabolism in rats of different ages

Male Sprague-Dawley rats at the ages of four weeks and nine months were fed purified diets containing 20% proteins either as casein (CAS), milk whey protein (WHY), or soybean protein (SOY) with 5% sardine oil for four weeks. The hypocholesterolemic effect of SOY was not statistically evident as compared to milk proteins at both ages, although serum cholesterol tended to be low in the SOY groups. A significant agedependent increase in serum cholesterol was observed in all dietary groups. Liver cholesterol concentrations were comparable in young rats, whereas in adults they were significantly lower in the SOY than in the CAS or WHY groups. At both ages, the activity of liver 3-hydroxy-3-methylglutaryl coenzyme A reductase tended to be higher in the SOY than in the other groups. Fecal steroid excretion was significantly higher in rats fed SOY than those fed either CAS or WHY, especially in adult rats. Significant age- and dietary protein-effects were observed in fatty acid profiles of liver microsomal phospholipids. Thus, the effects of dietary proteins on various lipid parameters were essentially maintained even when fish oil served as the source of dietary fat.     

Palmitic acid enhances cholesterol gallstone incidence in sasco hamsters fed cholesterol enriched diets

In an established hamster model of cholesterol cholelithiasis, a semipurified lithogenic diet containing 4% butterfat and 0.3% cholesterol leads to the production of cholesterol gallstones in only 50–60% of animals after a 6-wk feeding period. The purpose of this study was to investigate whether gallstone incidence could be increased while feeding a nutritionally adequate diet of moderate cholesterol content. The semipurified lithogenic diet was modified as follows: (i) substitution of 1.2% palmitic acid for 4% butterfat, and (ii) varying the amount of dietary cholesterol from 0.0 to 0.3% with either butterfat or palmitic acid as the lipid component of the diet. Substitution of palmitic acid for butterfat produced a significantly higher incidence of cholesterol gallstones (94%vs. 53%). Palmitic acid also raised the incidence of gallstones when added to the 0.1% and 0.2% cholesterol diets as compared to butterfat: 0%vs. 44% and 50%vs. 81%, respectively. Gallstone incidence increased from 0% to nearly 100% when the cholesterol content of the palmitic acid diets was raised from 0.0% to 0.3%, indicating a dose response effect with respect to dietary cholesterol. Hamsters fed cholesterol-free diets did not form gallstones. Increased dietary cholesterol led to increased liver weight associated with a significant increase in liver cholesterol concentration. However, the palmitic acid groups had significantly lower liver cholesterol values than the corresponding butterfat groups. Serum and biliary cholesterol concentrations increased with increasing dietary cholesterol intake, but there were no differences between the butterfat and palmitic acid groups. The cholesterol saturation index increased from 0.56 to 1.32 in the butterfat groups and from 0.56 to 1.30 in the palmitic acid groups upon raising the dietary cholesterol from 0.0 to 0.3%. Biliary total bile acid concentration did not vary significantly within all groups; however, the addition of cholesterol produced an increase in the ratio of chenodeoxycholic acid to cholic acid. It is concluded that in Sasco hamsters the saturated fatty acid, palmitic acid, when substituted for butterfat in a nutritionally adequate lithogenic diet, is capable of increasing gallstone incidence to almost 100% during a 6-wk feeding period.     

Dietary protein effects on gallstone formation

Semipurified lithogenic diets for hamsters contain casein as the protein source. Since substitution of soy protein isolate for casein reduces serum cholesterol concentrations in several species, we studied replacement of casein by soy protein for effects on gallstone formation. Feeding soy protein consistently resulted in a significantly reduced incidence of gallstones. Switching to a soy-based diet after induction of gallstones resulted in dissolution of a significant percentage of the stones. Partial substitution of soy for casein gave results intermediate between 100% casein and 100% soy. The lysine/arginine ratio of the proteins may be responsible for the observed differences in cholelithiasis. The reduction in lithogenicity associated with feeding soy protein appears to be mediated primarily through decreased secretion of cholesterol into bile.     

Apple polyphenols influence cholesterol metabolism in healthy subjects with relatively high body mass index

We performed a randomized double-blind, placebo-controlled study on moderately obese male and female subjects (71 subjects) with a body mass index ranging from 23 to 30 to evaluate the efficacy of 12-week intake of polyphenols extracted from apples and hop bract (600 mg/day). We confirmed that 12-week ingestion of polyphenol-containing capsules significantly decreased total cholesterol and LDL-cholesterol levels. The effects of the apple polyphenol-containing capsules were more marked than those of the hop bract polyphenol-containing capsules. The visceral fat area and the level of adiponectin in the group administered apple polyphenols improved in comparison with the control group. Blood and physical examinations revealed on clinical problems, and no adverse reactions were observed during the ingestion period. These results demonstrate that apple polyphenols regulate fat metabolism in healthy subjects with relatively high body mass index.     

Differential effects of ursodeoxycholic acid and ursocholic acid on the formation of biliary cholesterol crystals in mice

The preventive effect of 3α,7β,12α-trihydroxy-5β-cholanoic acid (ursocholic acid) and ursodeoxycholic acid on the formation of biliary cholesterol crystals was studied in mice. Cholesterol crystals developed with 80% incidence after feeding for five weeks a lithogenic diet containing 0.5% cholesterol and 0.25% sodium cholate. When 0.25% ursocholic acid or ursodeoxycholic acid was added to the lithogenic diet, the incidence as well as the grade (severity) of the gallstones were reduced. Plasma and liver cholesterol levels were decreased by ursodeoxycholic acid but not by ursocholic acid. Gallbladder cholesterol and phospholipid levels were decreased by both bile acids. The biliary bile acid level was decreased by ursocholic acid but not by ursodeoxycholic acid. After feeding ursocholic acid, its level in the bile was about 25% and the levels of cholic acid and β-muricholic acid decreased. Fecal sterol excretion was not changed by ursocholic acid, but was increased by ursodeoxycholic acid. After feeding ursocholic acid, fecal excretion of deoxycholic acid, cholic acid, and ursocholic acid increased. No differences were found between mice, with or without gallstones, in plasma and liver cholesterol levels, biliary phospholipid and bile acid levels, fecal sterol and bile acid levels, and biliary and fecal bile acid composition. The results suggest that the lower incidence of crystal formation after treatment with ursocholic acid is probably by a different mechanism than with ursodeoxycholic acid. In the mouse model, ursodeoxycholic acid exerts its effect at least partially, by decreasing cholesterol absorption. Ursocholic acid is well absorbed and excreted into bile and transformed into deoxycholic acid by the intestinal microflora in mice.     

The prostaglandins and their influence on lipid metabolism

Isolation, characterization, occurrence, and physiological action of the prostaglandins are reviewed.     

The effect of 5,8,11,14-eicosatetraynoic acid on lipid metabolism

The purpose of this presentation is to review the current state of knowledge regarding 5,8,11,14-eicosatetraynoic acid (ETYA, Ro 3-1428) and its effects on lipid metabolism. Accordingly, the topics discussed include hypocholesterolemic and dermatological studies involving ETYA in both animals and man, as well as the effects of ETYA on desaturate enzymes. Metabolic studies involving ETYA are also noted. Primary interest is focused on the effects of ETYA on selected processes of arachidonate metabolism, and the effect of ETYA on inflammation, platelet aggregation and tumor growth are discussed, keeping in mind the relevance of arachidonate metabolism to these processes.     

The effects of simvastatin and cholestyramine,alone and in combination,on hepatic cholesterol metabolism in the male rat

The influence of dietary simvastatin, cholestyramine, and the combination of simvastatin plus cholestyramine on hepatic cholesterol metabolism has been investigated in male rats. Recovery from the effects of the drugs was also investigated by refeeding normal chow for 24 h. Both drugs, alone and in combination, increased 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase activityin vitro, but activity returned toward control values, after drug withdrawal. Acyl-CoA:cholesterol acyltransferase (ACAT) was significantly reduced (P<0.001) by simvastatin (−75%), cholestyramine (−71%), and by the drug combination (−81%), due both to a decrease in microsomal cholesterol and to nonsubstrate-dependent modulation of enzyme activity. Refeeding control diet increased ACAT activity but not to control levels. The enhanced activity arose partly from higher microsomal cholesterol and partly from increases in total enzyme activity. Cytosolic neutral cholesteryl ester hydrolase (CEH) activity was substantially elevated by simvastatin (3-fold) and by the drug combination (6-fold), whereas the effect of cholestyramine was smaller (1.5-fold). Normal chow for 24 h only partially returned cytosolic CEH activity to control values. Microsomal CEH activity was increased by simvastatin, alone and in combination with cholestyramine (1.4 to 1.7-fold), and was also enhanced, in the cholestyramine-treated animals, following drug withdrawal. Removal of simvastatin did not allow recovery of this enzyme activity, while withdrawal of the drug combination led to values 29% below controls. The results indicate that in the rat, simvastatin and cholestyramine alter both ACAT and CEH activity, as well as inhibiting HMG-CoA reductase activity.     

The effect of short-term lipid infusion on liver function and biliary secretion in rats

This study was undertaken to determine the effect of various lipid emulsions on the hepato-biliary system in rats. Rats were randomly divided into six groups and infused continuously for 48 hr with either long-chain triglycerides (LCT), medium-chain triglycerides (MCT) or a mixture of MCT and LCT. One group infused with physiological saline solution served as controls. Throughout this period the rats received a fat free dietad libitum. During the last hour of lipid infusion bile was collected for determination of bile flow and composition. Subsequently, the rats were sacrificed and the morphology and lipid content of the liver determined. Only LCT lipid emulsions induced morphological changes and increased liver cholesterol content. In two rats infused with radiolabeled LCT, no labeled cholesterol was found in the liver, indicating that the excess hepatic cholesterol level may originate from enhanced cholesterol mobilization to the liver. Biliary cholesterol and phospholipid concentrations in LCT-treated rats were also elevated, as was the lithogenic index, whereas the other emulsions had no such effects. None of the emulsions affected the plasma liver function tests or bile flow. We therefore conclude that the lithogenicity of the bile in rats is directly related to the lipid components of the total parenteral nutrition and the type of triglyceride infused.     

The effects of polyoxyethylated cholesterol on fecal bile acids and nitrogen and on cholesterol balance in rats

Polyoxyethalated cholesterol (POEC) is a water soluble derivative of cholesterol which decreases cholesterol absorption in rats without affecting body weight, fatty acid excretion, or intestinal histology. In the present study rat feces were analyzed for cholic, deoxycholic, chenodeoxycholic, muricholic and lithocholic acid following 3 months of feeding a standard or a 2% enriched cholesterol diet with or without 1.5% POEC. In rats maintained on the cholesterol free diet, POEC increased total bile acids (mg/day) by 50% from 14±3 to 21±3 (mean ±SEM) but only the increase in chenodeoxycholic acid was significant (P<0.05). The corresponding POEC effect in the 2% cholesterol diet was 31% (70±8 to 93±3, P<0.01). Fecal nitrogen and serum cholesterol did not vary among groups. Comparing these data with neutral steroid excretion previously determined showed that POEC in the cholesterolfree diet increased the negative cholesterol balance more than three-fold (34±7vs 118±13 P<0.01). In rats fed 2% cholesterol, POEC caused a negative cholesterol balance of 222±8 compared to the control of 27±52 (P<0.01). The data indicate that POEC exerts complex effects in the intestinal tract which increase both bile acid and cholesterol excretion.     

Origin,metabolism, and adverse health effects of cholesterol oxidation products

Cholesterol oxidation products (COP), also called oxysterols, enter the organism by intake of COP-containing foodstuff or are generated in vivo by enzymatic or non-enzymatic oxidation (peroxidation) of cholesterol. COP are transported in lymph and blood with lipoproteins (mainly fatty acyl monoesters at the 3β-position) or bound to albumin. As being substrates for acyl CoA: cholesterol acyltransferase (ACAT), lecithin:cholesterol acyltransferase (LCAT), and cholesteryl ester transfer protein (CETP), COP are esterified and exchanged between lipoproteins and are distributed in the whole organism. Kupffer cells and hepatocytes are the target cells for COP transformation and excretion via bile secretion. Regarding biological activities, COP reveal widespread effects on cholesterol metabolism as well as on structure and function of cellular membranes. Throughout their actions, COP were shown to be cytotoxic but they were also suggested to be regulatory agents. Their detrimental part in atherogenesis seems to be still underestimated, while valid data on the role of COP in carcinogenesis are still missing.     

Differential effects of dietary linoleic and α-linolenic acid on lipid metabolism in rat tissues

Comparative effects of feeding dietary linoleic (safflower oil) and α-linolenic (linseed oil) acids on the cholesterol content and fatty acid composition of plasma, liver, heart and epididymal fat pads of rats were examined. Animals fed hydrogenated beef tallow were used as isocaloric controls. Plasma cholesterol concentration was lower and the cholesterol level in liver increased in animals fed the safflower oil diet. Feeding the linseed oil diet was more effective in lowering plasma cholesterol content and did not result in cholesterol accumulation in the liver. The cholesterol concentration in heart and the epididymal fat pad was not affected by the type of dietary fatty acid fed. Arachidonic acid content of plasma lipids was significantly elevated in animals fed the safflower oil diet and remained unchanged by feeding the linseed oil diet, when compared with the isocaloric control animals fed hydrogenated beef tallow. Arachidonic acid content of liver and heart lipids was lower in animals fed diets containing safflower oil or linseed oil. Replacement of 50% of the safflower oil in the diet with linseed oil increased α-linolenic, docosapentaenoic and docosahexaenoic acids in plasma, liver, heart and epididymal fat pad lipids. These results suggest that dietary 18∶2ω6 shifts cholesterol from plasma to liver pools followed by redistribution of 20∶4ω6 from tissue to plasma pools. This redistribution pattern was not apparent when 18∶3ω3 was included in the diet.     

The effect of alfalfa-corn diets on cholesterol metabolism and gallstones in prairie dogs

Cholesterol gallstones were present in prairie dogs fed alfalfa plus corn with and without exogenous cholesterol (0.4%). The diets fed to the animals for eight weeks contained alfalfa plus corn in fixed proportions of 50∶50, 85∶15 and 15∶85 (w/w). At sacrifice, all animals were healthy but had not gained weight; no deaths occurred during the experiment. Cholesterol gallstones were present in all groups. In the absence of exogenous cholesterol, the highest stone incidence was found in the animals which received the lowest fiber (highest corn) diets (alfalfa plus corn, 50∶50, 67%; alfalfa plus corn, 15∶85, 83%). Cholesterol gallstone incidence was 100% when exogenous cholesterol was added to the alfalfa plus corn diets (50∶50 and 15∶85). No pigment gallstones were detected in any animal. Liver and plasma cholesterol concentrations were highest in the animals receiving alfalfa plus corn (15∶85) plus 0.4% cholesterol (4.29 mg/g, and 356 mg/dl, respectively). These values were lowest in animals receiving 85% alfalfa plus 15% corn without cholesterol (2.19 mg/g and 88 mg/dl, respectively). Lithogenic indices were below 1.00 in all groups. Biliary bile acids were mainly amidates of cholic acid and chenodeoxycholic acid, with the former predominating. Thus, gallstones can be formed in prairie dogs in the absence of exogenous cholesterol; gallstone incidence is reduced by dietary fiber.