Effects of β-lactam antibiotics on intestinal microflora and bile acid metabolism in rats

Wistar male rats were treated for six days with broad spectrum β-lactam antibiotics, latamoxef, and cefotaxime. On the seventh day, the number of fecal anaerobic microbes decreased, total fecal bile acids decreased, and bile acid pools increased. Secondary bile acids such as β-hyocholic, hyodeoxycholic, lithocholic, and deoxycholic acids decreased in the feces while the primary bile acids, cholic, β-muricholic, and chenodeoxycholic acids, became predominant. Coprostanol, a microbial metabolite of cholesterol, also disappeared from the feces during the treatment. The cecum enlarged to almost twice the size of that in control rats, whereas the liver weight was not significantly changed. After treatment was stopped, the number of fecal microbes returned to the initial counts within a week, but restoration of bile acid and cholesterol metabolism required at least three weeks.     

Effect of Peucedanum japonicum Thunb on the expression of obesity-related genes in mice on a high-fat diet

The present study describes the effect of Peucedanum japonicum Thunb (PJT) intake on the expression of obesity-related genes in mice fed a high-fat diet. To explore the mechanism underlying the effect of PJT, This study focused on the expression of genes, especially those related to obesity and metabolism syndrome, in the liver and adipose tissues. In agreement with our previous observations, intake of 10 % PJT for 4 weeks significantly reduced serum triglyceride (TG), leptin, abdominal fat, and adipocyte size. PJT also significantly increased fecal excretion of TG, decreased that of bile acid, and tended to increase the fecal excretion of total cholesterol. Microarray analysis was used to monitor changes in 324 metabolic syndrome-related genes in the liver. Statistically significant upregulation of PPP1R10, RORC, and PBEF1 genes and downregulation of DUSP1, INSIG2, and SERPINA12 genes were noted and confirmed by real-time RT-PCR. These changes were indicative of increased fatty acid oxidation in the maintenance of lipid homeostasis and insulin sensitivity in the livers of PJT-fed mice. PJT increased the expression of PPARγ, FXRα, DGAT1, and ATGL genes, suggesting an enhancement of adipocyte differentiation and normalization of functionality of adipose tissue.     

Bile acid metabolism in young-old parabiotic rats

Serum cholesterol, triglyceride and phospholipid levels, liver cholesterol concentration, bile flow, biliary cholesterol, phospholipid and bile acid secretion rates, fecal sterol and bile acid levels and their bile acid compositions were examined in young-old parabiotic rats and compared with those in young and old control rats and young-young parabiotic rats. Bile acid composition was expressed in terms of the cholic acid group/chenodeoxycholic acid group (CA/CDCA) ratio. Body weight (BW) gain decreased after parabiosis especially in old rats, but the liver weight (g/100 g BW), diet-intake, feces dry weight, liver cholesterol concentration and fecal sterol level were almost the same in all the groups. The biliary bile acid secretion rate was higher and the fecal bile acid level was lower in old rats than those in young rats but both the levels became comparable with those in young rats after parabiosis of old rats with young rats. Young rats, however, showed no changes in these levels after parabiosis. The serum cholesterol level and the biliary and fecal CA/CDCA ratios in old rats were higher than those in young rats but decreased after parabiosis with young rats, although they were still higher than those in young rats. The serum cholesterol level in young rats increased after parabiosis with old rats, but not after parabiosis with young rats, and the fecal bile acid level and the CA/CDCA ratio were not changed in either case. It is concluded from these findings that the serum cholesterol level and the CA/CDCA ratio increased with age and that these increases were prevented after parabiosis with young rats, while young rats, although their serum cholesterol level was increased, showed no increase in the CA/CDCA ratio after parabiosis with old rats.     

Effects of cholestyramine and squalene feeding on hepatic and serum plant sterols in the rat

Hepatic and serum phytosterol concentrations were compared in the rat under basal conditions and during activated cholesterol and bile acid production due to squalene and cholestyramine feeding. Both treatments consistently decreased hepatic and serum levels of sitosterol and campesterol and, unlike esterified cholesterol, esterified plant sterols were not increased in liver during squalene feeding. Serum levels of phytosterols were decreased quite proportionately to those in the liver. The hepatic levels of sitosterol and campesterol closely correlated with each other, but not with cholesterol levels. The percentage esterification of both phytosterols was lower than that of cholesterol. The results indicate that activation of hepatic sterol production leads to depletion of hepatic plant sterols. It is suggested that poor esterification of plant sterols may contribute to this decrease.     

Effects of triton WR 1339 and orotic acid on lipid metabolism in rats

In order to investigate the effect of hepatic cholesterol flux on biliary bile acids, Triton WR 1339 and orotic acid were administered to rats, and the biliary cholesterol, phospholipids and bile acids were analyzed together with serum lipoproteins and hepatic lipids. Triton, which raised serum very low density lipoprotein and lipid levels and decreased serum high density lipoprotein liver lipid levels, increase the biliary cholic acid group/chenodeoxycholic acid group ratio (CA/CDCA) in the bile without affecting the total amount of bile acids and the other biliary lipids. Orotic acid, which decreased serum lipid and lipoprotein concentrations and increased liver lipid levels, increased the biliary excretion of cholesterol and phospholipids, but produced no significant change in the total amount of bile acids and in the CA/CDCA ratio in bile.     

Absorption and metabolic fate of dietary3H-squalene in the rat

The absorption and metabolic fate of dietary squalene were investigated on the rat by administering a single oral dose of³H-squalene and¹⁴C-cholesterol. Experiments on rats with cannulated thoracic duct revealed that³H-squalene was, like¹⁴C-cholesterol, absorbed through the lymphatic vessels and that ca. 20% of absorbed³H-squalene was cyclized to sterols during the transit through the intestinal wall. Feces contained³H-sterols, indicating that newly synthesized mucosal sterols had been secreted into the gut lumen. In intact animals,³H-squalene appeared in the circulation more rapidly than¹⁴C-cholesterol and did not persist to any significant extent in the squalene-rich adipose and muscle tissues. The increase in dietary squalene load (8–48 mg) decreased the absorption percentage of³H-squalene (45–26%) but did not affect the absorption of¹⁴C-cholesterol (47%). Determination of fecal steroids revealed that during the first days absorbed³H-squalene was eliminated to a significantly higher extent than¹⁴C-cholesterol as fecal bile acids (34% vs 11%). The experiments indicate that the rat intestine has a marked capacity for absorbing dietary squalene and that the absorbed squalene is preferentially converted to bile acids in the liver.     

Muricholic Acids Promote Resistance to Hypercholesterolemia in Cholesterol-Fed Mice

Background and aims: Hypercholesterolemia is a major risk factor for atherosclerosis and cardiovascular diseases. Although resistant to hypercholesterolemia, the mouse is a prominent model in cardiovascular research. To assess the contribution of bile acids to this protective phenotype, we explored the impact of a 2-week-long dietary cholesterol overload on cholesterol and bile acid metabolism in mice. Methods: Bile acid, oxysterol, and cholesterol metabolism and transport were assessed by quantitative real-time PCR, western blotting, GC-MS/MS, or enzymatic assays in the liver, the gut, the kidney, as well as in the feces, the blood, and the urine. Results: Plasma triglycerides and cholesterol levels were unchanged in mice fed a cholesterol-rich diet that contained 100-fold more cholesterol than the standard diet. In the liver, oxysterol-mediated LXR activation stimulated the synthesis of bile acids and in particular increased the levels of hydrophilic muricholic acids, which in turn reduced FXR signaling, as assessed in vivo with Fxr reporter mice. Consequently, biliary and basolateral excretions of bile acids and cholesterol were increased, whereas portal uptake was reduced. Furthermore, we observed a reduction in intestinal and renal bile acid absorption. Conclusions: These coordinated events are mediated by increased muricholic acid levels which inhibit FXR signaling in favor of LXR and SREBP2 signaling to promote efficient fecal and urinary elimination of cholesterol and neo-synthesized bile acids. Therefore, our data suggest that enhancement of the hydrophilic bile acid pool following a cholesterol overload may contribute to the resistance to hypercholesterolemia in mice. This work paves the way for new therapeutic opportunities using hydrophilic bile acid supplementation to mitigate hypercholesterolemia.     

Egg ovomucin attenuates hypercholesterolemia in rats and inhibits cholesterol absorption in Caco-2 cells

This experiment was designed to evaluate the effect of casein or ovomucin (OV) on the micellar solubility of cholesterol and the taurocholate binding capacity in vitro. We also evaluated the effects of casein or OV on cholesterol metabolism in rats and Caco-2 cells. OV has a significantly greater bile acid-binding capacity than that of casein in vitro. Micellar cholesterol solubility in vitro was significantly lower in the presence of OV compared to casein. The cholesterol micelles containing OV significantly suppressed cholesterol uptake by Caco-2 cells compared to the cholesterol micelles containing casein. Consistent with these in vitro findings, OV-feeding significantly increased the fecal excretion of bile acids or cholesterol compared with casein-feeding. Serum total cholesterol was significantly lower in rats fed OV than in those fed casein. The concentrations of total lipids in liver were significantly lower in the OV-fed group compared with the casein group. These results suggest that the suppression of cholesterol absorption by direct interaction between cholesterol mixed micelles and OV in the jejunal epithelia is part of the mechanism underlying the hypocholesterolemic action of OV. OV may also inhibit the reabsorption of bile acids in the ileum, thus lowering the serum cholesterol level.     

Cholesterol metabolism in gnotobiotic gerbils

Germfree gerbils were associated with a murine-derived hexaflora which produced only minor changes in the primary bile acid pattern of rats. These hexaflora-associated gerbils had relatively small ceca (4% of body weight) and reproduced well. Although serum cholesterol levels of both conventional and hexaflora-associated gerbils increased in response to dietary cholesterol, the hexaflora-associated gerbil showed a greater elevation in serum cholesterol than the conventional gerbil when maintained on a diet containing 0.1% cholesterol. This increase in serum cholesterol manifested itself almost totally in the very low density lipoprotein and low density lipoprotein fractions. The fecal bile acids of the hexaflora-associated gerbil were largely deconjugated, but very little further modification of either cholic or chenodeoxycholic acid had taken place. The data suggest that in the absence of elements of the intestinal microflora that can express a bile acid-modifying potential, and particularly a 7-α-dehydroxylating capacity, catabolism of cholesterol to bile acids is reduced, and cholesterol accumulates in the very low density and low density serum lipoprotein fractions.     

Amaranth Oil Increased Fecal Excretion of Bile Acid but Had No Effect in Reducing Plasma Cholesterol in Hamsters

Hamsters were fed for 4 weeks on four different diets: control (C) (balanced diet containing 20 % corn oil as the lipid source), hypercholesterolemic (H) (identical to C but containing 12 % coconut oil, 8 % corn oil and 0.1 % cholesterol as the lipid source), amaranth oil (A) (identical to H without corn oil but with amaranth oil), and squalene (S) (identical to H but admixed with squalene in the ratio found in amaranth oil). There were no significant differences in lipid profile, and in the cholesterol excreted in the animals’ feces from amaranth oil (A) and squalene (S) groups. Fecal excretion of bile acids was greater in the amaranth oil (A) and squalene groups (S) as compared to the other groups. The scores of steatosis and parenchymal inflammation observed in the amaranth oil (A) and squalene groups (S) were superior to the ones observed in the other groups. Our findings demonstrated that amaranth oil, and its component squalene, increased the excretion of bile acids but did not have a hypocholesterolemic effect in hamsters fed on a diet containing high amounts of saturated fat and cholesterol.     

Effects of dietary methionine and cystine on lipid metabolism in hepatoma-bearing rats with hyperlipidemia

Abnormal lipid metabolism and its restoration by dietary methionine (Met) and cystine (Cys) were studied in Donryu rats subcutaneously implanted with an ascites hepatoma cell line of AH109A. The hepatoma-bearing rats exhibited byperlipidemia characterized by rises in serum triglyceride and cholesterol levels. Decreased lipoprotein lipase (LPL) activities in epididymal adipose tissue, cardiac muscle, and gastrocnemius as well as increased fatty acid mobilization from adipose tissue were considered to be responsible for the hepatoma-induced hypertriglyceridemia, while increased hepatic cholesterogenesis and decreased steroid excretion into feces were thought to be responsible for the hepatoma-induced hypercholesterolemia. Dietary-supplemented Met or Cys reduced the AH109A-induced hypertriglyceridemia with suppression of fatty acid synthesis in the host liver. Met restored the fall of LPL activities, while Cys did not. Dietary Met or Cys also reduced the hypercholesterolemia with restoration of decreased bile acid excretion into feces. These results suggest that dietary Met or Cys is hypolipidemic in the hepatoma-bearing rats with slight differences in their modes of action.     

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.     

Effects of different levels of dietarytrans-octadecenoate on steroid metabolism in rats

Rats were fed semipurified diets containing olive oil or partially hydrogenated corn oil at the 5 or 20% level for ca. 30 days. These fat diets contained the same amount of octadecenoate but differed in the geometry with respect to each fat level. Contents oft-18∶1 were 26% and 41% of total fatty acids, respectively. The linoleic acid content was also made equivalent (3.8 energy %). After feeding on cholesterol-free diets, rats ontrans fat, compared to those oncis fat, showed: (a) no changes in serum cholesterol and apolipoprotein levels, (b) no effects on the bile flow and concentrations of biliary cholesterol or bile acids, (c) a trend toward increased fecal excretion of neutral and acidic steroids, (d) a lesser extent of transformation of cholesterol to coprostanol in the gut, and (e) no changes in the composition of biliary and fecal bile acids. Observations (c) and (d) were more marked with a hightrans fat regimen. These observations, except for serum apolipoproteins and fecal steroid excretion, were practically reproducible even when rats were fed cholesterol-enriched diets. A preliminary part of this study was presented at the 74th annual AOCS meeting, Chicago, 1983.     

Alteration of cholesterol metabolism by 4-0-methylascochlorin in rats

The effect of 4-0-methylascochlorin (MAC), an experimental hypocholesterolemic agent, on cholesterol metabolism was investigated in rats in two separate experiments. The administration of MAC for 2 and 6 consecutive weeks at daily doses of 100–135 mg/kg resulted in reduction in serum cholesterol levels of 16% after 2 weeks of treatment in the first experiment, and 13% after 6 weeks in the second experiment in comparison to the corresponding controls. MAC administered at a daily dose of 100 mg/kg for 2 weeks showed a significant increase in the biliary excretion of bile acids and cholesterol in bile-duct cannulated rats with or without the administration of taurocholate. In the second experiment, MAC treatment for 6 weeks produced a marked increase in the fecal output of acidic sterols during a 2 to 6-week period. MAC treatment also further enhanced hepatic cholesterol 7α-hydroxylase in the rats. Therefore, it appears that the mechanism of serum cholesterol lowering due to MAC is related to the enhancement of hepatic bile acid synthesis and the increase in biliary and fecal excretion of bile acids.     

Comparative hypocholesterolemic effects of capybara (Hydrochoerus hydrochaeris dabbenei) oil, horse oil, and sardine oil in cholesterol-fed rats

The hypocholesterolemic efficacy of various polyunsaturated fatty acids was compared in rats given cholesterol-enriched diets. Capybara oil (CO, linoleic+α-linolenic acids), horse oil (HO, α-linolenic acid), and sardine oil (SO, eicosapentaenoic+docosahexaenoic acids) were added to diets at 50 g/kg. The weight gain, food intake, and liver weight in the CO-fed group were significantly higher than those in other groups during the 6-wk experimental period. The serum total and very low density lipoprotein (VLDL)+intermediate density lipoprotein (IDL)+low density lipoprotein (LDL) cholesterol concentrations of the CO-fed and SO-fed groups were significantly lower than in the HO-fed group after 6 wk. The serum high density lipoprotein cholesterol concentration in the SO-fed group was significantly higher than that in the CO-fed and HO-fed groups. The fecal neutral sterol concentration in the CO-fed group was reduced significantly compared with the other groups, and the fecal bile acid concentration in the HO-fed group was significantly higher than that in the SO-fed group. The results of this study demonstrate that CO lowers the serum total cholesterol and VLDL+IDL+LDL-cholesterol concentrations in the presence of excess cholesterol in the diet as well as SO.     

Cholesterol metabolism in swine fed diets containing either casein or soybean protein

Diets containing casein produce higher concentrations of serum cholesterol in swine than those containing soybean protein, but only when such diets contain high levels of cholesterol. We suggest that dietary casein causes an increase in the absorption of cholesterol and bile acids, thus explaining the observed decrease in fecal excretion of neutral steroids and bile acids when compared to soybean protein. The mechanism in molecular terms by which dietary proteins influence steroid absorption remains to be established.     

Effect of chitosan feeding on intestinal bile acid metabolism in rats

The effect of chitosan feeding (for 21 days) on intestinal bile acids was studied in male rats. Serum cholesterol levels in rats fed a commercial diet low in cholesterol were decreased by chitosan supplementation. Chitosan inhibited the transformation of cholesterol to coprostanol without causing a qualitative change in fecal excretion of these neutral sterols. Increased fiber consumption did not increase fecal excretion of bile acids, but caused a marked change in fecal bile acid composition. Litcholic acid increased sigificantly, deoxycholic acid increased to a leasser extent, whereas hyodeoxycholic acid and the 6β-isomer and 5-epimeric 3α-hydroxy-6-keto-cholanoic acid(s) decreased. The pH in the cecum and colon became elevated by chitosan feeding which affected the conversion of primary bile acids to secondary bile acids in the large intestine. In the cecum, chitosan feeding increased the concentration of α-,β-, and ω-muricholic acids, and lithocholic acid. However, the levels of hyodeoxycholic acid and its 6β-isomer, of monohydroxy-monoketo-cholanoic acids, and of 3α, 6ξ, 7ξ-trihydroxy-cholanoic acid decreased. The data suggest that chitosan feeding affects the metabolism of intestinal bile acids in rats.     

Effects of a single ingestion of sodium taurocholate on esterified cholesterol concentration in liver and cholesterol turnover in the rat

The overall response of the rat’s cholesterol metabolism to a single ingestion of taurocholate (80 mg) was studied with the isotopic equilibrium method. The bile acid production, measured by the daily¹⁴CO₂ output of rats in isotopic equilibrium of [26-¹⁴C]-cholesterol, initially decreased and then increased. Conversely, the hepatic concentration of esterified cholesterol first increased and then decreased. Moreover, the ingestion of taurocholate increasing the intestinal absorption coefficient of dietary cholesterol increased the abosprtion and decreased the fecal excretion and the intestinal biosynthesis of cholesterol. The balance of these last effects is an excess cholesterol inflow. The classical hypothesis of negative feedback regulation of bile acid production fails to explain the observed biphasic effect of taurocholate. This compound, when its origin is exogenous, appears to stimulate the storage of esterified cholesterol in the liver, at the expense of bile acid synthesis. This accumulation rate takes into account not only the decrease in cholesterol transformation into bile acids but also the excess inflow of cholesterol. As the exogenous taurocholate was eliminated from the body, cholesteryl ester hydrolysis occurred and provided a supplementary source of free cholesterol for bile acid synthesis.     

Effects of phenobarbital upon bile acid synthesis in two strains of rats

Rats of the Wistar and Sprague-Dawley strains were injected with sodium phenobarbital (100 mg/kg body wt/day) for 8 days. Fecal bile acid excretion was measured on days 6 and 8 of the experiment, and biliary bile acid composition, hepatic microsomal cholesterol, 7α-hydroxylase, and 7α-hydroxy-4-cholesten-3-one 12α-hydroxylase were determined at the end of the study. In the Wistar rat, injection of phenobarbital produced a doubling of fecal bile acid output (controls, 5.3 mg/rat/day; treated rats, 10.6 mg/rat/day) and a two-three fold increase in cholesterol 7α-hydroxylase. The fecal bile acid output of Sprague-Dawley rats increased 20% in response to phenobarbital (controls, 9.5 mg/rat/day; treated rats, 11.6 mg/rat/day). The activity of cholesterol 7α-hydroxylase remained unchanged. In both strains, phenobarbital treatment produced a decrease in the proportion of cholic acid in total biliary bile acids (controls, 85%; treated groups, 65%). This was associated with a decrease of 7α-hydroxy-4-cholesten-3-one 12α-hydroxylase activity by ca. 50%. Biliary cholesterol concentrations were reduced in phenobarbital treated rats of both strains, but liver cholesterol concentrations remained unchanged. The drug produced a 25% increase in liver wt, on the average.     

Sterol synthesis from biliary squalene in the jejunal mucosa of the rat in vivo

Because bile contains substantial amounts of cholesterol precursors, e.g., squalene and differnet methyl sterols, the fate of biliary squalene was studied by incubating isolated jejunal loops of the rat in vivo with bile containing³H-squalene and¹⁴C-cholesterol. After 90 min, no radioactivity was found in plasma lipids. Closer analysis of gut epithelium revealed that both labeled compounds were preferentially taken up by the villous cells. Biliary³H-squalene was absorbed almost completely and was further cyclized to free and esterified methyl sterols and cholesterol. Whereas squalene not cyclized to sterols stayed in the mucosa, the newly synthesized sterols were transferred to lumen. The lipid patterns of gut lumen and mucosal cells were quite different, suggesting that the transfer of the newly synthesized lipid to intestinal lumen was not due to the desquamation of epithelial cells alone. The results suggest that biliary cholesterol precursors can contribute to the cholesterol production of the jejunal villous cells bypassing the rate-limiting step of the cholesterol synthesis pathway, and to the “nonexchanging” fecal neutral sterols of the rat.