EP4 emerges as a novel regulator of bile acid synthesis and its activation protects against hypercholesterolemia

Prostaglandin E receptor subtype 4 (EP4) knockout mice develops spontaneous hypercholesterolemia but the detailed mechanisms by which EP4 affects cholesterol homeostasis remains unexplored. We sought to determine the cause of hypercholesterolemia in EP4 knockout mice, focusing on the role of EP4 in regulating the synthesis and elimination of cholesterol. Deficiency of EP4 significantly decreased total bile acid levels in the liver by 26.2% and the fecal bile acid content by 27.6% as compared to wild type littermates, indicating that the absence of EP4 decreased hepatic bile acid synthesis and their subsequent excretion in stools. EP4 deficiency negatively regulate bile acid synthesis through repression of phosphorylated extracellular signal-regulated kinase 1/2 (ERK)-mediated cholesterol 7α-hydroxylase (CYP7A1) expression and that the hypercholesterolemia in EP4 knockout mice is due to a defect in cholesterol conversion into bile acids. Deficiency of EP4 also increased de novo cholesterol synthesis and altered cholesterol fluxes in and out of the liver. Treating high fat diet-challenged mice with the pharmacological EP4 agonist, CAY10580 (200?μg/kg body weight/day i.p) for three weeks effectively prevented diet-induced hypercholesterolemia, enhanced endogenous bile acid synthesis and their fecal excretion. In summary, EP4 plays a critical role in maintaining cholesterol homeostasis by regulating the synthesis and elimination of bile acids. Activation of EP4 serves as an effective novel strategy to promote cholesterol disposal in the forms of bile acids in order to lower plasma cholesterol levels.     

Lactobacillus plantarum LRCC 5273 isolated from Kimchi ameliorates diet-induced hypercholesterolemia in C57BL/6 mice

ABSTRACT

This study was designed to select potent cholesterol-lowering probiotic strains on HepG2 cell and investigate the effect of selected strain, Lactobacillus plantarum LRCC 5273 and LRCC 5279 in hypercholesterolemic mice. In the results, LP5273 group showed significantly reduced total and LDL cholesterol compared to HCD group. In addition to significantly up-regulated hepatic mRNA expression of LXR-α and CYP7A1, intestinal LXR-α and ABCG5 were significantly up-regulated in LP5273 group. With activation of hepatic and intestinal LXR-α and its target genes, fecal cholesterol and bile acid excretion were increased in LP5273 fed mice. These results suggest that LP5273 ameliorates hypercholesterolemia in mice through the activation of hepatic and intestinal LXR-α, resulting in enhancement of fecal cholesterol and bile acids excretion in the small intestine. The results of present study suggest mechanistic evidences for hypocholesterolemic effects of L. plantarum spp., and may contribute to future researches for prevention of hypercholesterolemia and cardiovascular disease.     

Effect of red wine on oxidative stress and hypercholesterolemia induced by feeding a high-cholesterol diet in rat

The effect of red wine on oxidative stress and hypercholesterolemia induced by feeding a high-cholesterol diet (supplemented with 1.65% of cholesterol (w/w) for 4 weeks) to female Wistar rats was examined. When red wine was simultaneously supplemented to high-cholesterol diet, total cholesterol, triglycerides, atherogenic index and lipid peroxidation products significantly decreased compared with the high-cholesterol diet alone, while GSH content and antioxidative enzymes activities were enhanced. In the hypercholesterolemic rat the excretion of fecal bile acids, as well as their plasma and hepatic concentrations were increased significantly. Administration of red wine enhanced these values, indicating an increase in the cholesterol degradation. These results suggest that red wine may have a protective effect against oxidative stress, hypercholesterolemia and atherogenic index induced by high-cholesterol diet.     

Identification of a Novel Hypocholesterolemic Protein,Major Royal Jelly Protein 1, Derived from Royal Jelly

Royal jelly (RJ) intake lowers serum cholesterol levels in animals and humans, but the active component in RJ that lowers serum cholesterol level and its molecular mechanism are unclear. In this study, we set out to identify the bile acid-binding protein contained in RJ, because dietary bile acid-binding proteins including soybean protein and its peptide are effective in ameliorating hypercholesterolemia. Using a cholic acid-conjugated column, we separated some bile acid-binding proteins from RJ and identified the major RJ protein 1 (MRJP1), MRJP2, and MRJP3 as novel bile acid-binding proteins from RJ, based on matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Purified MRJP1, which is the most abundant protein of the bile acid-binding proteins in RJ, exhibited taurocholate-binding activity in vitro. The micellar solubility of cholesterol was significantly decreased in the presence of MRJP1 compared with casein in vitro. Liver bile acids levels were significantly increased, and cholesterol 7α-hydroxylase (CYP7A1) mRNA and protein tended to increase by MRJP1 feeding compared with the control. CYP7A1 mRNA and protein levels were significantly increased by MRJP1 tryptic hydrolysate treatment compared with that of casein tryptic hydrolysate in hepatocytes. MRJP1 hypocholesterolemic effect has been investigated in rats. The cholesterol-lowering action induced by MRJP1 occurs because MRJP1 interacts with bile acids induces a significant increase in fecal bile acids excretion and a tendency to increase in fecal cholesterol excretion and also enhances the hepatic cholesterol catabolism. We have identified, for the first time, a novel hypocholesterolemic protein, MRJP1, in RJ. Interestingly, MRJP1 exhibits greater hypocholesterolemic activity than the medicine β-sitosterol in rats.     

Hypolipidemic effects of orally administered dextran and cellulose anion exchangers in cockerels and dogs

Various cellulose and dextran anion exchangers bind bile salts in vitro under conditions of pH and ionic strength resembling those in the lumen of the small intestine. Of these substances, diethylaminoethyl (DEAE) cellulose, guanidoethyl cellulose, and DEAE Sephadex reduced hypercholesterolemia when added to the diet of cholesterol-fed cockerels. In addition, DEAE Sephadex reduced serum sterols in normocholesterolemic cockerels and dogs, lowered serum phospholipids and triglycerides in cholesterol-fed hypercholesterolemic cockerels and in normocholesterolemic dogs, and increased fecal excretion of bile acids in hypercholesterolemic cockerels. The data indicate that these insoluble cationic polymers exert their hypocholesterolemic effects by interrupting the enterohepatic circulation of bile acids.     

Astragalus polysaccharides lowers plasma cholesterol through mechanisms distinct from statins

To determine the efficacy and underlying mechanism of Astragalus polysaccharides (APS) on plasma lipids in hypercholesterolemia hamsters. The effect of APS (0.25 g/kg/d) on plasma and liver lipids, fecal bile acids and neutral sterol, cholesterol absorption and synthesis, HMG-CoA reductase activity, and gene and protein expressions in the liver and small intestine was investigated in twenty-four hypercholesterolemia hamsters. Treatment periods lasted for three months. APS significantly lowered plasma total cholesterol by 45.8%, triglycerides by 30%, and low-density lipoprotein-cholesterol by 47.4%, comparable to simvastatin. Further examinations revealed that APS reduced total cholesterol and triglycerides in the liver, increased fecal bile acid and neutral sterol excretion, inhibited cholesterol absorption, and by contrast, increased hepatic cholesterol synthesis and HMG-CoA reductase activity. Plasma total cholesterol or low-density lipoprotein-cholesterol levels were significantly correlated with cholesterol absorption rates. APS up-regulated cholesterol-7α-hydroxylase and LDL-receptor gene expressions. These new findings identify APS as a potential natural cholesterol lowering agent, working through mechanisms distinct from statins.     

Effects of acute administration of taurocholic and taurochenodeoxycholic acid on biliary lipid excretion in the rat.

Comparison of the effects of biliary lipid excretion produced by infusion of taurochenodeoxycholate and taurocholate showed no significant difference when the bile acids were infused for a relatively short period of time. Cholesterol excretion rates measured during depletion of the bile acid pool were significantly higher than cholesterol excretion rates measured during infusion of bile acids at various rates. These data indicate that there is some mechanism in addition to bile acid excretion that is responsible for biliary excretion of cholesterol when the enterohepatic circulation is intact.     

Increase of bile acids synthesis and excretion caused by taurine administration prevents the ovariectomy-induced increase in cholesterol concentrations in the serum low-density lipoprotein fraction of Wistar rats

We examined the effect of dietary taurine on the concentrations of serum cholesterol and apolipoprotein in lipoprotein fractions of Six-month-old ovariectomized, which were used as a model of hypercholesterolemia in postmenopausal woman, or sham operated rats. Taurine significantly reduced the serum total and low-density lipoprotein cholesterol concentrations only in the ovariectomized rats. In contrast, taurine significantly lowered the serum apolipoprotein B concentration and serum very low-density lipoprotein-apolipoprotein E concentration only in the sham operated rats. The serum total and high density lipoprotein-apolipoprotein E concentrations were significantly lower in the rats fed taurine than in those fed the control diet regardless of whether they had undergone ovariectomy. The esterified cholesterol level in the liver was significantly lower and the level of hepatic cholesterol 7 alpha-hydroxylase activity was significantly higher in the rats fed taurine than in those fed the control diet. The total bile acids concentration in the feces and intestinal contents of rats fed taurine were significantly higher than those in rats fed the control diet regardless of whether they had undergone ovariectomy. In the sham-rats, taurine accelerated bile acid synthesis and excretion, thereby increasing cholesterol consumption. The increased cholesterol consumption might be compensated by accelerating cholesterol synthesis and/or reducing the synthesis and release of very low-density lipoprotein from the liver. But in the ovariectomized rats, although taurine also accelerated bile acid synthesis and excretion, cholesterol demand might be compensated by excess cholesterol in the blood.     

Alteration of biliary ursodeoxycholic acid in guinea pig during early stages of cholestyramine feeding

The effects of cholestyramine feeding on biliary ursodeoxycholic acid, fecal excretion of bile acids and neutral sterols on cholesterol 7α-hydroxylase and hepatic HMG-CoA reductase were examined in the guinea pig. In the bile there was a 57% decrease in the concentration of ursodeoxycholic acid while an increase was observed in the concentration of chenodeoxycholic acid. Cholestyramine feeding for ten days resulted in a decrease in plasma cholesterol levels and an increase in both hepatic HMG-CoA reductase and cholesterol 7α-hydroxylase activities. The fecal excretion of both bile acids and neutral sterols was significantly increased.     

Anti-cholestatic activity of HD-03, a herbal formulation in thioacetamide (TAA)-induced experimental cholestasis

In the present study HD-03, a herbal formulation was investigated for its anti-cholestatic activity in TAA-induced cholestasis in anaesthetized guinea pigs. Administration of TAA at a dose of 100 mg/kg body wt significantly reduced the bile flow, bile acid and bile salt excretion. Pretreatment with HD-03 at a dose of 750 mg/kg body wt per orally for 15 days in guinea pigs significantly prevented thioacetamide-induced changes in bile flow, bile acids and bile salts excretion. Thus, HD-03 can serve as a potent choleretic and anti-cholestatic agent.     

Medium-Chain Fatty Acids Enhanced the Excretion of Fecal Cholesterol and Cholic Acid in C57BL/6J Mice Fed a Cholesterol-Rich Diet

The objective of the present study was to investigate the cholesterol-reducing effect of medium-chain fatty acids (MCFAs) completed by elevated excretion of fecal neutral steroids and/or bile acids. Blood and liver lipid profiles, fecal neutral steroids, bile acids, and mRNA and protein expression of the genes relevant to cholesterol homeostasis were measured and analyzed in C57BL/6J mice fed a cholesterol-rich diet with 2% caprylic acid or capric acid for 12 weeks. Blood total cholesterol and low-density lipoprotein cholesterol (LDL-c) levels were reduced significantly as compared to diet with palmitic acid or stearic acid. Caprylic acid promoted the excretion of fecal neutral steroids, especially cholesterol. The excretion of fecal bile acids, mainly in the form of cholic acid was enhanced and accompanied by elevated expression of mRNA and the protein of hepatic cholesterol 7α-hydroxylase (CYP7A1). These results indicate that MCFAs can reduce blood cholesterol by promoting the excretion of fecal cholesterol and cholic acid.     

Importance of jejunum for bile acid absorption in pigeons: effect of jejunal resection on plasma cholesterol and cholesterol excretion

Previous studies from our laboratory suggested that the jejunum could be the major site of bile acid absorption in the pigeon. To confirm this observation directly, the effects of jejunal resection on the fecal excretion of bile acids and neutral sterols and on the level of plasma cholesterol were examined in the pigeon. This surgical procedure specifically increased (p less than 0.05) the excretion of bile acids, with no change in the excretion of neutral sterols. The total fecal steroid excretion in the pigeons with jejunal resection was significantly (P less than 0.01) higher than that in the control group. Jejunal resection also decreased the plasma cholesterol level in the pigeon. These results confirm our earlier suggestion that the jejunum is the major site of bile acid absorption in pigeons, unlike that in mammals, in which the ileum is the major site.     

Increased excretion of bile acids by genetically hyperlipoproteinemic Zucker rat

Fecal excretion of neutral sterols and bile acids was measured in age-matched hyperlipoproteinemic Zucker obese rats and their lean litter mates. The bile acid excretion (mg/day ± SEM) in Zucker rats was significantly higher (p<0.01) when compared to lean controls (Zucker obese rats 41.68 ± 2.86; lean controls 29.85 ± 1.50). Neutral sterol excretion in both the groups of rats was similar. Total fecal steroid excretion (mg/day ± SEM) in Zucker rats was significantly higher (p<0.01) than in lean controls (Zucker obese rats 52.33 ± 3.50; lean controls 39.23 ± 2.16. The Zucker rat thus mimics the increased bile acid excretion noted previously in human Type IV hyperlipoproteinemia and could serve as an ideal animal model for studying the interrelationship between bile acid excretion and very low density lipoprotein metabolism.     

Impact of β-cyclodextrin and resistant starch on bile acid metabolism and fecal steroid excretion in regard to their hypolipidemic action in hamsters1

To examine the impact on bile acid metabolism and fecal steroid excretion as a mechanism involved in the lipid-lowering action of β-cyclodextrin and resistant starch in comparison to cholestyramine, male golden Syrian hamsters were fed 0% (control), 8% or 12% of β-cyclodextrin or resistant starch or 1% cholestyramine. Resistant starch, β-cyclodextrin and cholestyramine significantly lowered plasma total cholesterol and triacylglycerol concentrations compared to control. Distinct changes in the bile acid profile of gallbladder bile were caused by resistant starch, β-cyclodextrin and cholestyramine. While cholestyramine significantly reduced chenodeoxycholate independently of its taurine–glycine conjugation, β-cyclodextrin and resistant starch decreased especially the percentage of taurochenodeoxycholate by ?75% and ?44%, respectively. As a result, the cholate:chenodeoxycholate ratio was significantly increased by 100% with β-cyclodextrin and by 550% with cholestyramine while resistant starch revealed no effect on this ratio. β-Cyclodextrin and resistant starch, not cholestyramine, significantly increased the glycine:taurine conjugation ratio demonstrating the predominance of glycine conjugated bile acids. Daily fecal excretion of bile acids was 4-times higher with 8% β-cyclodextrin and 19-times with 1% cholestyramine compared to control. β-Cyclodextrin and cholestyramine also induced a 2-fold increase in fecal neutral sterol excretion, demonstrating the sterol binding capacity of these two compounds. Resistant starch had only a modest effect on fecal bile acid excretion (80% increase) and no effect on excretion of neutral sterols, suggesting a weak interaction with intestinal steroid absorption. These data demonstrate the lipid-lowering potential of β-cyclodextrin and resistant starch. An impaired reabsorption of circulating bile acids and intestinal cholesterol absorption leading to an increase in fecal bile acid and neutral sterol excretion is most likely the primary mechanism responsible for the lipid-lowering action of β-cyclodextrin. In contrast, other mechanisms involving the alterations in the biliary bile acid profile or repressed hepatic lipogenesis, e.g., VLDL production, appear to be involved in the hypolipidemic effect of resistant starch.     

Hypocholesterolemic effect of rice protein is due to regulating hepatic cholesterol metabolism in adult rats

Aging is one of major risk factors for developing hypercholesterolemia. To elucidate the cholesterol-lowering mechanism exerted by rice protein (RP), the effects on hepatic cholesterol outputs and cholesterol metabolism related enzymes were investigated in adult rats, which were fed by casein (CAS) and RP without cholesterol in diets. After 2 weeks of feeding, the significant cholesterol-lowering effect was observed in adult rats fed by RP compared to CAS. The hepatic total- and VLDL-cholesterol secretions into circulation were significantly depressed in RP group, whereas biliary outputs of bile acids and cholesterol were effectively stimulated by RP-feeding, causing an increase in fecal sterol excretion compared to CAS. As a result, the apparent cholesterol absorption was significantly inhibited by RP. RP-feeding significantly increased the activity and gene expression of cholesterol 7α-hydroxylase, whereas acyl-CoA:cholesterol acyltransferase-2 activity and gene expression were significantly decreased by RP as compared with CAS. Neither activity nor gene expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase of RP did differ from CAS in the liver. The present study demonstrates that rice protein can prevent hypercholesterolemia through modifying hepatic cholesterol metabolism under cholesterol-free dietary condition. The findings suggest that hypocholesterolemic action induced by rice protein is attributed in part to the inhibition of cholesterol absorption during the adult period.     

Effect of bile acid deconjugation on the fecal excretion of steroids

The effect of microbiological deconjugation of bile acids on total bile acid and neutral sterol fecal excretion by adult male rats has been studied. A screening method utilizing mice allowed selection of a Clostridium perfringens type A strain, which accelerated cholesterol catabolism in mice. When this species of bacteria was associated with germfree rats, the fecal bile acids were excreted as free bile acids (deconjugated), however the quantities of bile acids excreted were not increased compared with those of germfree rats. Conventional rats excrete twice as much bile acids (all deconjugated) as do the germfree and C. perfringens-associated rats. It is, therefore, unlikely that the microbiological deconjugation of bile acids is responsible for the increased fecal excretion of bile acids seen in conventional rats. The C. perfringens-associated rats excreted identical kinds and quantities of fecal neutral sterols as did the germfree rats.     

Fecal steroid excretion in chickens with hereditary hyperlipidemia

Plasma lipids (cholesterol, triglycerides, and phospholipids; mg/dl) and the fecal excretion (mg/day) of neutral steroids and bile acids were studied in layers (L), hereditary nonlayer hens (NL), and roosters (R) fed a basal cholesterol-free grain diet ad libitum. Each group had significantly (P less than 0.05) different levels of plasma cholesterol, triglycerides, and phospholipids when compared to the other groups. The highest lipid values were found in the NL group (cholesterol, 798 +/- 89; triglycerides, 8914 +/- 679; phospholipids, 2458 +/- 112). There was no difference in the fecal excretion of neutral steroids between L and NL; however, fecal bile acid excretion by these two groups was significantly different (P less than 0.05) (L, 13.1 +/- 1.7 vs NL, 26.9 +/- 3.4). Fecal neutral steroid excretion by R was significantly greater (P less than 0.05) than that by either L or NL (L, 6.4 +/- 1.3; NL, 6.0 +/- 1.4; R, 14.4 +/- 1.2). While fecal excretion of bile acids by R (36.1 +/- 4.0) was also greater than that by either L or NL, only the difference between R and L was statistically significant (P less than 0.05). Since, in the steady state, fecal bile acid excretion is equal to its synthesis, these results suggest that bile acid metabolism in these animals can be affected by both sex and egg-laying status.     

Bile acids and bile alcohols in a child with hepatic 3 beta-hydroxy-delta 5-C27-steroid dehydrogenase deficiency: effects of chenodeoxycholic acid treatment

Duodenal bile, urine, plasma, and feces from a child with hepatic 3 beta-hydroxy-delta 5-C27-steroid dehydrogenase deficiency were analyzed by fast atom bombardment mass spectrometry and gas chromatography-mass spectrometry to investigate the formation and excretion of abnormal bile acids and bile alcohols. The biliary bile salts consisted of glycocholic acid (25%) and of sulfated and glycine conjugated di- and trihydroxycholenoic acids (55%), two C27 bile acids, and eleven sulfated bile alcohols (mainly tetrols, 20%), all having 3 beta,7 alpha-dihydroxy-delta 5 or 3 beta,7 alpha,12 alpha-trihydroxy-delta 5 ring structures. In plasma, sulfated cholenoic acids constituted 65% and unconjugated 3 beta,7 alpha-dihydroxy-5-cholestenoic acid 25% of the total level, 71 micrograms/ml. The urinary excretion of the former was 30.4 mg/day and that of unsaturated bile alcohol sulfates, mainly pentols, 7 mg/day. The predominant bile acid in feces was an unconjugated epimer of 3 beta,7 alpha,12 alpha-trihydroxy-5-cholenoic acid, and small amounts of cholic acid were present. The minimum total excretion was 11.3 mg/day. Treatment with chenodeoxycholic acid resulted in marked clinical improvement and normalized liver function tests. Further studies are needed to define the mechanism of action. Plasma bile acids decreased to 1.6 micrograms/ml and urinary excretion to 3.4 mg/day. Chenodeoxycholic and ursodeoxycholic acids became predominant in all samples. The fecal excretion of unsaturated cholenoic acid sulfates increased to 40 mg/day compared to 89 mg/day of saturated bile acids. The results provide further support for a defective hepatic 3 beta-hydroxy-delta 5-C27-steroid dehydrogenase deficiency, and indicate that the 3 beta-hydroxy-delta 5 bile acids are formed via 7 alpha-hydroxycholesterol. The formation of glycocholic acid may be due to an incomplete enzyme defect or to transformation of the 3 beta-hydroxy-delta 5 structure by bacterial and hepatic enzymes during an enterohepatic circulation.