Effects of pregnenolone-16α-carbonitrile on the metabolism of cholesterol in rat liver microsomes

The effects of pregnenolone-16α-carbonitrile (PCN) on hepatic metabolism of cholesterol were studied in rat liver microsomes in order to clarify the underlying mechanisms of the PCN-induced biliary hypersecretion of cholesterol. Male Sprague-Dawley rats were fed a diet supplemented with 0.05% of PCN for one week. The microsomal activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase, regulating cholesterol biosynthesis, decreased from 577 ± 46 (SEM) to 367 ± 38 pmol/min/mg protein compared to the controls. Cholesterol 7 α-hydroxylase activity, governing bile acid synthesis, was 9.0 ± 1.1 pmol/min/mg protein in the treated group and 34.8 ± 7.4 pmol/min/mg protein in the controls, a reduction of 74% (P<0.01). The acyl CoA:cholesterol acyltransferase (ACAT) activity, catalyzing the esterification of cholesterol, remained unchanged, as did the levels of total and free cholesterol in liver homogenates and microsomes. The results of this study provide evidence that the increase in biliary cholesterol secretion during PCN treatment is not caused by a change in ACAT activity, but can be explained by a decreased catabolism of cholesterol to bile acids.     

Oxidation of 3-oxygenated Δ4- and Δ5-C27 steroids by soybean lipoxygenase and rat liver microsomessteroids by soybean lipoxygenase and rat liver microsomes

The formation of dioxygenated metabolites of cholesterol, epicholesterol (5-cholesten-3α-ol) 4-cholesten-3β-ol, 4-cholesten-3α-ol, 4-cholesten-3-one and 4-stigmasten-3-one was studied after incubations with soybean lipoxygenase and linoleic acid. From cholesterol and epicholesterol were formed the 7α-hydroxy-, 7α-hydroperoxy-, 7β-hydroxy-, 7β-hydroperoxy-, 7-oxo and 5,6-epoxyderivatives as well as 6β-hydroxy-4-cholesten-3-one. All Δ⁴-steroids were hydroxylated in the 6α- and 6β-positions. The ratios between the yields of 6β- and 6α-hydroxylated metabolites varied between 3∶1 and 2∶1. Incubations with 4-cholesten-3α-ol and 4-cholesten-3β-ol also afforded the 4,5-epoxides of these steroids. The ratios between the yields of the 4β,5β- and 4α,5α-epoxides were ca. 4∶1 for 4-cholesten 3β-ol and ca. 3∶2 for 4-cholesten-3α-ol. With iron-supplemented microsomes from rat liver, the compounds formed were qualitatively and quantitatively the same as with soybean lipoxygenase, whereas with 18,000 × g rat liver supernatant fractions the yields of all products formed—except 7α-hydroxycholesterol and 6β-hydroxy-4-cholesten-3-one—were markedly decreased. The results indicate the presence of a rat liver microsomal 6β-hydroxylase which can use 4-cholesten-3-one as a substrate and extend previous findings of similarities between soybean lipoxygenase and a nonspecific lipoxygenase in rat liver microsomes.     

Studies on the 12α and 26-hydroxylation of bile alcohols by rabbit liver microsomes

12α-Hydroxylation of two C₂₇-steroids by rabbit liver microsomes was studied. Optimal assay conditions were determined with 7α-hydroxy-4-cholesten-3-one and 5β-cholestane-3α,7α-diol as substrates. The rate of 12α-hydroxylation of 7α-hydroxy-4-cholesten-3-one was found to be greater than that of 5β-cholestane-3α,7α-diol by ca. 60%. Microsomal 26-hydroxylation of 5β-cholestane-3α7α-diol was also measured, and the ratio of 26-hydroxylation to 12α-hydroxylation of 5β-cholestane-3α,7α-diol was found to be ca. 0.4. Rabbit liver 12-αhydroxylase was more active than that of three other species (man, rat, monkey), explaining in part the predominance of cholic acid in rabbit bile.     

The cholesterol-lowering effect of guar gum is not the result of a simple diversion of bile acids toward fecal excretion

The effects of partially hydrolyzed, nonviscous, guar gum (PHGG) on cholesterol metabolism and digestive balance have been compared with those of native guar gum (GUAR) in rats adapted to 0.4% cholesterol diets. Both types of guar gum elicited acidic fermentations in the large intestine, but only GUAR effectively lowered plasma cholesterol (P<0.001), chiefly in the triglyceride-rich lipoprotein fraction. The biliary bile acid excretion was significantly enhanced in rats fed GUAR (P<0.05), as well as the intestinal and cecal bile acid pool (P<0.001). In rats fed GUAR and to a lesser extent in those fed PHGG, the fecal excretion of bile acids and neutral sterol was higher than in controls (P<0.01). The digestive balance (cholesterol intake-steroid excretion) was positive in control rats (+47 μmol/d), whereas it was negative in rats fed GUAR (−20 μmol/d), which could involve a higher rate of endogenous cholesterol synthesis. In rats fed PHGG, the steroid balance remained slightly positive. Liver 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase activity was very low (22 pmol/min/mg protein), owing to cholesterol supplementation, in control rats or in rats fed PHGG, whereas it was markedly higher (+463%) in rats fed GUAR. In conclusion, even if PHGG does alter some parameters of the enterohepatic cycle of cholesterol and bile acids, its effects are not sufficient to elicit a significant cholesterol-lowering effect. The intestinal (ileal or cecal) reabsorption of bile acids was not reduced, but rather increased, by GUAR; nevertheless the intestinal capacities of reabsorption were overwhelmed by the enlargement of the digestive pool of bile acids. In the present model, induction of HMG-CoA reductase probably takes place in the presence of elevated portal bile acid concentrations.     

γ-tocotrienol as a hypocholesterolemic and antioxidant agent in rats fed atherogenic diets

This study was designed to determine whether incorporation of γ-tocotrienol or α-tocopherol in an atherogenic diet would reduce the concentration of plasma cholesterol, triglycerides and fatty acid peroxides, and attenuate platelet aggregability in rats. For six weeks, male Wistar rats (n=90) were fed AIN76A semisynthetic test diets containing cholesterol (2% by weight), providing fat as partially hydrogenated soybean oil (20% by weight), menhaden oil (20%) or corn oil (2%). Feeding the ration with menhaden oil resulted in the highest concentrations of plasma cholesterol, low and very low density lipoprotein cholesterol, triglycerides, thiobarbituric acid reactive substances and fatty acid hydroperoxides. Consumption of the ration containing γ-tocotrienol (50 μ/kg) and α-tocopherol (500 mg/kg) for six weeks led to decreased plasma lipid concentrations. Plasma cholesterol, low and very low density lipoprotein cholesterol, and triglycerides each decreased significantly (P<0.001). Plasma thiobarbituric acid reactive substances decreased significantly (P<0.01), as did the fatty acid hydroperoxides (P<0.05), when the diet contained both chromanols. Supplementation with γ-tocotrienol resulted in similar, though quantitatively smaller, decrements in these plasma values. Plasma α-tocopherol concentrations were lowest in rats fed menhaden oil without either chromanol. Though plasma α-tocopherol did not rise with γ-tocotrienol supplementation at 50 mg/kg, γ-tocotrienol at 100 mg/kg of ration spared plasma α-tocopherol, which rose from 0.60±0.2 to 1.34±0.4 mg/dL (P<0.05). The highest concentration of α-tocopherol was measured in plasma of animals fed a ration supplemented with α-tocopherol at 500 mg/kg. In response to added collagen, the partially hydrogenated soybean oil diet without supplementary cholesterol led to reduced platelet aggregation as compared with the cholesterol-supplemented diet. However, γ-tocotrienol at a level of 50 mg/kg in the cholesterol-supplemented diet did not significantly reduce platelet aggregation. Platelets from animals fed the menhaden oil diet released less adenosine triphosphate than the ones from any other diet group. The data suggest that the combination of γ-tocotrienol and α-tocopherol, as present in palm oil distillates, deserves further evaluation as a potential hypolipemic agent in hyperlipemic humans at atherogenic risk.     

Age-related changes in the metabolism of cholesterol in rat liver microsomes

The effects of aging on the hepatic metabolism of cholesterol were studied in 1-, 6- and 24-month-old male Sprague-Dawley rats. Microsomal 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity, which regulates cholesterol biosynthesis, decreased from 835±144 (SEM) pmol/min/mg protein in the youngest group to 219±34 and 205±53 pmol/min/mg protein (p<0.001) in the 6- and 24-month-old groups, respectively. Cholesterol 7α-hydroxylase activity, which governs bile acid synthesis, was gradually reduced from 70±14 pmol/min/mg protein in the 1-month-old group to 32±7 and 16±3 pmol/min/mg protein (p<0.05) in the 6- and 24-month-old groups, respectively. Acyl coenzyme A:cholesterol acyltransferase activity, which catalyzes the esterification of cholesterol, averaged 431±47 and 452 ±48 pmol/min/mg protein in the 1- and 6-month-old groups, respectively, and was increased to 585±55 pmol/min/mg protein (p<0.05) in the 24-month-old group. The level of total cholesterol showed an age-related increase from 1.56±0.16 mg/g liver in the 1-month-old group to 1.70±0.15 and 2.20±0.19 mg/g liver (p<0.05) in the 6- and 24-month-old groups, respectively. The increase was mainly caused by an accumulation of esterified cholesterol. We conclude that a marked decrease in HMG-CoA reductase occurs between 1 and 6 months of age; thereafter the enzyme activity stays unchanged. The activity of cholesterol 7α-hydroxylase decreases progressively and drastically with age, whereas the capacity for esterifying cholesterol increases slightly. We speculate that the reduced conversion of cholesterol to bile acids may be one explanation of the age-related increase of plasma cholesterol seen in rats.     

Inhibition of cholesterol synthesis and hepatic 3-hydroxy-3-methylglutaryl-CoA reductase in rats by simvastatin and pravastatin

In this communication we attempt to provide one possible explanation for the observed differences regarding kinetics and distribution between simvastatin and pravastatin. Rats treated with simvastatin or pravastatin exhibited a reduction in the incorporation of [2-¹⁴C]acetate into liver cholesterol and displayed lower plasma mevalonate levels as compared to control animals. Moreover, both the total and dephosphorylated 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase (EC 1.1.1.34) activities, particularly 1 h after treatment, were greatly reduced in liver microsomes obtained from simvastatin-treated as compared to control rats. During the same time frame, these parameters were actually elevated with pravastatin treatment. It is known that HMG-CoA reductase synthesis and activity increase following their competitive inhibition. Our results suggest that pravastatin, at 1 h following treatment, was no longer bound to the enzyme; however, it had entered the liver because its inhibitory effect on cholesterol synthesis was manifest at early times after administration. These data provide a plausible rationale for the earlier observation that activity of simvastatin persists longer in plasma than does that of pravastatin.     

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.     

Effect of dietary cholesterol on low density lipoprotein-receptor, 3-hydroxy-3-methylglutaryl-CoA reductase, and low density lipoprotein receptor-related protein mRNA expression in healthy humans

We investigated the possibility that dietary cholesterol downregulates the expression of low density lipoprotein (LDL) receptor and 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase genes of circulating mononuclear cells in vivo in healthy humans. We also studied the variations of the LDL receptor-related protein (LRP) gene in the same conditions. Dieters (n=5) were submitted to a 4-d fat restriction (mean cholesterol intake: 6±4 mg/d), followed by a 7-d cholesterol (a mean of 791±150 mg/d) supplementation. Controls (n=3) did not change their diet. During fat restriction, serum total and LDL cholesterol decreased significantly (P<0.05), and LDL receptor and HMG-CoA reductase mRNA copy numbers in mononuclear cells increased by 57 and 147%, respectively (P<0.05). After reintroducing cholesterol, serum cholesterol was stable whereas LDL receptor and HMG-CoA reductase mRNA decreased by 46 and 72% (P<0.05) and LRP mRNA increased by 59% (P<0.005). The changes in LDL receptor and HMG-CoA reductase mRNA abundance were correlated (r=+0.79, P=0.02) during cholesterol reintroduction as were LDL receptor and LRP mRNA levels, but negatively (r=−0.70, P=0.05). Also, 70% of the variability in LRP mRNA (P<0.005) was explained by dietary cholesterol. Thus, the basic mechanisms regulating cellular cholesterol content, the coordinate feedback repression of genes governing the synthesis and uptake of cholesterol, are operating in vivo in humans. However, serum cholesterol did not increase in response to dietary cholesterol, suggesting that these mechanisms may not play as predominant a role as previously believed in the short-term control of serum cholesterol in vivo in humans. A new finding is that LRP gene is also sensitive to dietary cholesterol, suggesting that it may participate in the control of serum cholesterol. Further in vivo studies in humans are warranted to explore the molecular mechanisms of the physiological response to dietary cholesterol in humans.     

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.     

Evidence that leptin contributes to intestinal cholesterol absorption in obese (ob/ob) mice and wild-type mice

In the present study, the effect of leptin on intestinal cholesterol absorption was investigated in C57 BL/6 OlaHsd Lep^ob/Lep^ob obese (ob/ob) mice and lean C57 BL/6 (wild-type) mice. Animals were treated either with or without recombinant leptin for 2 wk. Cholesterol absorption was measured by the constant isotope feeding method and indirectly by the ratio of campesterol to cholesterol in serum. In ob/ob mice, cholesterol absorption was significantly higher compared to wild-type mice [83.4±2.3% (SD) vs. 77.6±1.5%, P<0.01]. Treatment with leptin significantly reduced cholesterol absorption in both ob/ob and wild-type mice by 8.5 (P<0.001) and 5.2% (P<0.05), respectively. Serum concentrations of campesterol and the ratio of campesterol to cholesterol in ob/ob mice were significantly higher compared to wild-type mice (2.2±0.3 mg/dL vs. 1.2±0.3 mg/dL, P<0.001; and 36.8±2.8 μg/mg vs. 28.0±3.3 μg/mg, P<0.001). After treatment of ob/ob mice with leptin, concentrations of campesterol and its ratio to cholesterol were significantly lower (2.2±0.3 mg/dL vs. 1.0±0.2 μg/mg, P<0.001; and 36.8±2.8 μg/mg vs. 13.2±2.2 μg/mg, P<0.001, respectively). In wild-type mice, the ratio of campesterol to cholesterol in serum was also significantly lower after treatment with leptin (28.0±3.3 μg/mg vs. 22.6±5.0 μg/mg, P<0.05). A significant positive correlation (r=0.701, P<0.01) between cholesterol absorption and the ratio of campesterol to cholesterol, in serum was found. It is concluded that leptin contributes to intestinal cholesterol absorption in ob/ob mice and lean wild-type mice.     

Vitamin E supplementation increases circulating vitamin E metabolites tenfold in end-stage renal disease patients

Vitamin E supplementation could elevate circulating vitamin E metabolites while modulating oxidative and inflammatory status in end-stage renal failure patients undergoing hemodialysis. Plasma concentrations of carboxyethyl-hydroxychromanols (α-and γ-CEHC), ascorbic acid, α-and γ-tocopherols, E₂-isoprostanes, and inflammatory biomarkers [tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), ferritin, and C-reactive protein (CRP)] were measured in blood samples obtained from patients (n=11) before and after dialysis on two occasions prior to, and at 1 and 2 mon of daily vitamin E supplementation (400 IU RRR-α-tocopherol). Supplementation nearly doubled plasma α-tocopherol concentrations (from 18±0.5 to 31±1.7 μM, P<0.0001), whereas γ-tocopherol concentrations decreased (from 2.8±0.3 to 1.7±0.2 μM, P=0.001). Serum α-CEHC increased 10-fold from 68±3 to 771±175 nM (P<0.0001), and γ-CEHC increased from 837±164 to 1136±230 nM (P=0.008). Vitamin E supplementation also increased postdialysis hematocrits from 38±1% to 41±1% (P<0.001). Dietary antioxidant intakes (vitamins E and C) were low in most subjects; plasma ascorbic acid levels (88±27 μM) decreased significantly with dialysis (33±11 μM, P=0.01). Plasma Il-6, CRP, TNF-α, and free F₂-isoprostane concentrations were elevated throughout the study. There is a complex relationship between chronic inflammation and oxidative stress that is not mitigated by short-term vitamin E supplementation. Importantly, serum vitamin E metabolite concentrations that increased 10-fold within 30 d of supplementation did not increase further, suggesting routes other than urine for removal of metabolites.     

Octacosanol Administration to Humans Decreases Neutral Sterol and Bile Acid Concentration in Feces

To investigate octacosanol (OC) metabolism in humans and its influence on cholesterol metabolism, two studies were conducted. In the first study ten healthy women received daily 30 mg OC for a period of 4 weeks. Blood and feces samples were collected at baseline and after the intervention. Serum concentrations of total cholesterol, LDL cholesterol, and HDL cholesterol were not altered following OC administration. Concentrations of excreted cholesterol end products decreased with the intervention (neutral sterols: 24.6 ± 9.7 mg/g vs. 20.3 ± 7.5 mg/g dry matter, P < 0.05; bile acids: 6.47 ± 3.89 mg/g vs. 4.03 ± 2.26 mg/g dry matter, P < 0.05). OC was not detected in serum samples, but the fecal OC concentration increased after the intervention period (11 ± 7 μg/g vs. 817 ± 179 μg/g dry matter, P < 0.05). In the second kinetic study on three participants, OC was identified in serums after oral application of 50 mg OC within 8 h. The decrease in the concentration of fecal cholesterol end products may underline a systemic effect of OC on cholesterol metabolism, even though the serum cholesterol levels were not influenced. Dual first authorship. S. Keller and F. Gimmler having contributed equally to the basic science presented.     

Oryzanol decreases cholesterol absorption and aortic fatty streaks in hamsters

Oryzanol is a class of nonsaponifiable lipids of rice bran oil (RBO). More specifically, oryzanol is a group of ferulic acid esters of triterpene alcohol and plant sterols. In experiment 1, the mechanisms of the cholesterol-lowering action of oryzanol were investigated in 32 hamsters made hypercholesterolemic by feeding chow-based diets containing 5% coconut oil and 0.1% cholesterol with or without 1% oryzanol for 7 wk. Relative to the control animals, oryzanol treatment resulted in a significant reduction in plasma total cholesterol (TC) (28%, P<0.01) and the sum of IDL-C, LDL-C, and VLDL-C (NON-HDL-C) (34%, P<0.01). In addition, the oryzanol-treated animals also exhibited a 25% reduction in percent cholesterol absorption vs. control animals. Endogenous cholesterol synthesis, as measured by the liver and intestinal HMG-CoA reductase activities, showed no difference between the two groups. To determine whether a lower dose of oryzanol was also efficacious and to measure aortic fatty streaks, 19 hamsters in experiment 2 were divided into two groups and fed for 10 wk chow-based diets containing 0.05% cholesterol and 10% coconut oil (w/w) (control) and the control diet plus 0.5% oryzanol (oryzanol). Relative to the control, oryzanol-treated hamsters had reduced plasma TC (44%, P<0.001), NON-HDL-C (57%, P<0.01), and triglyceride (TG) (46%, P<0.05) concentrations. Despite a 12% decrease in high density lipoprotein cholesterol (HDL-C) (P<0.01), the oryzanol-treated animals maintained a more optimum NON-HDL-C/HDL-C profile (1.1±0.4) than the contorl (2.5±1.4; P<0.0075). Aortic fatty streak formation, so defined by the degree of accumulation of Oil Red O-stained macrophage-derived foam cells, was reduced 67% (P<0.01) in the oryzanol-treated animals. From these studies, it is concluded that a constituent of the nonsaponifiable lipids of RBO, oryzanol, is at least partially responsible for the cholesterol-lowering action of RBO. In addition, the cholesterol-lowering action of oryzanol was associated with significant reductions in aortic fatty streak formation.     

Quantitation of cholesterol oxidation products in unirradiated and irradiated meats

A method to detect 7-ketocholesterol, cholesterol-5β,6β-epoxide, cholesterol-5α,6α-epoxide, 4-cholesten-3-one, 4,6-cholestadien-3-one and 4-cholestene-3,6-dione in unirradiated and irradiated beef, pork and veal was developed by use of chloroform-methanol-water extraction, solid-phase extraction, column separation, thin-layer chromatography and gas chromatography. This method recovered 78–88% of the cholesterol oxidation products and detected the cholesterol oxidation products at 10 ppb or higher. Irradiation of the meats to a dose of 10 kGy increased these compounds, except 4,6-cholestadien-3-one for all three types of meat, over unirradiated, and except cholesterol-5α,6α-epoxide and 4-cholesten-3-one for the pork. All the cholesterol oxidation products in the unirradiated meats increased during storage at 0–4°C for 2 wk with some exceptions for the pork. The increases of cholesterol oxidation products in stored irradiated meats were greater than those in the unirradiated.     

Fecal steroids in diarrhea: IV. Cholera

Fecal bile acid and neutral sterol patterns were studied in eight healthy adult volunteers who were challenged withVibrio cholerae classical Ogawa 395 strain in the course of vaccine development studies. Bacterial 7α-dehydroxylation of cholic and chenodeoxycholic acids was not altered during experimentally induced cholera diarrhea, despite the fact that fecal weight in g/day (wet wt) was increased greatly during diarrhea (1913±390 vs 161±11 in controls, p<0.005). Consistent with the findings on bile acids, no significant changes in the production of coprostanol, epicoprostanol, or coprostanone were observed although the percentage of unmodified cholesterol was increased during the diarrheal episode (20.7±3.3% vs 11.9±2.3, p<0.02). Total concentrations of both bile acids and cholesterol in mg/g of feces (wet wt) were decreased considerably as a result of diarrhea). However, total bile acid and neutral steroid excretions in mg/kg/day in subjects with and without diarrhea do not appear to be different. Intestinal transit times, measured in eight subjects by the use of carmine red dye, were found to be shortened in diarrhea (5.8±1.1 hr vs 23.4±4.1 hr in controls, p<0.001). The results from this study are similar to those observed in experimentally induced travellers' diarrhea associated with toxigenicEscherichia coli, but they are in striking contrast to the changes in gastrointestinal steroid metabolism observed in acute shigello sis, an invasive intestinal infection.     

Effects of dietary phenolic compounds on tocopherol, cholesterol, and fatty acids in rats

The effects of the phenolic compounds butylated hydroxytoluene (BHT), sesamin (S), curcumin (CU), and ferulic acid (FA) on plasma, liver, and lung concentrations of α- and γ-tocopherols (T), on plasma and liver cholesterol, and on the fatty acid composition of liver lipids were studied in male Sprague-Dawley rats. Test compounds were given to rats ad libitum for 4 wk at 4 g/kg diet, in a diet low but adequate in vitamin E (36 mg/kg of γ-T and 25 mg/kg of α-T) and containing 2 g/kg of cholesterol. BHT significantly reduced feed intake (P<0.05) and body weight and increased feed conversion ratio; S and BHT caused a significant enlargement of the liver (P<0.001), whereas CU and FA did not affect any of these parameters. The amount of liver lipids was significantly lowered by BHT (P<0.01) while the other substances reduced liver lipid concentrations but not significantly. Regarding effects on tocopherol levels, (i) feeding of BHT resulted in a significant elevation (P<0.001) of α-T in plasma, liver, and lung, while γ-T values remained unchanged; (ii) rats provided with the S diet had substantially higher γ-T levels (P<0.001) in plasma, liver, and lung, whereas α-T levels were not affected; (iii) administration of CU raised the concentration of α-T in the lung (P<0.01) but did not affect the plasma or liver values of any of the tocopherols; and (iv) FA had no effect on the levels of either homolog in the plasma, liver, or lung. The level of an unknown substance in the liver was significantly reduced by dietary BHT (P<0.001). BHT was the only compound that tended to increase total cholesterol (TC) in plasma, due to an elevation of cholesterol in the very low density lipoprotein + low density lipoprotein (VLDL+LDL) fraction. S and FA tended to lower plasma total and VLDL+LDL cholesterol concentrations, but the effect for CU was statistically significant (P<0.05). FA increased plasma high density lipoprotein cholesterol while the other compounds reduced it numerially, but not significantly. BHT, CU, and S reduced cholesterol levels in the liver TC (P<0.001) and percentages of TC in liver lipids (P<0.05). With regard to the fatty acid composition of liver lipids, S increased the n-6/n-3 and the 18∶3/20∶5 polyunsaturated fatty acids (PUFA) ratios, and BHT lowered total monounsaturated fatty acids and increased total PUFA (n−6+n−3). The effects of CU and FA on fatty acids were not highly significant. These results suggest some in vivo interactions between these phenolic compounds and tocopherols that may increase the bioavailability of vitamin E and decrease cholesterol in rats.     

Dietary phospholipid alters biliary lipid composition in formula-fed piglets

Plasma cholesterol, arachidonic acid (AA, 20∶4n−6), and docosahexaenoic acid (DHA, 22∶6n−3) are higher in breast-fed infants than in infants fed formula without cholesterol, AA, or DHA. This study investigated differences in plasma, hepatic, and bile lipids and phospholipid fatty acids, and expression of hepatic proteins involved in sterol metabolism that result from feeding formula with cholesterol with egg phospholipid to provide AA and DHA. For this study, three groups of piglets were evaluated: piglets fed formula with 0.65 mmol/L cholesterol, the same formula with 0.8% AA and 0.2% DHA from egg phospholipid, and piglets fed sow milk. Piglets fed the formula with phospholipid AA and DHA had higher plasma high density lipoprotein, but not apoprotein (apo) B cholesterol or triglyceride; higher bile acid and phospholipid concentrations in bile; and higher liver and bile phospholipid AA and DHA than piglets fed formula without AA and DHA (P<0.05). Hydroxy methylglutaryl (HMG)-CoA reductase and 7-α-hydroxylase, the rate-limiting enzymes of cholesterol and bile acid synthesis, respectively, and low density lipoprotein receptor mRNA levels were not different between piglets fed formula without and with phospholipid AA and DHA, but HMG-CoA reductase and 7α-hydroxylase mRNA were higher, and plasma apo B containing lipoprotein cholesterol was lower in all piglets fed formula than in piglets fed milk. These studies show that supplementing formula with AA and DHA from egg phospholipid alters bile metabolism by increasing the bile AA and DHA, and bile acid and phospholipid.     

Enzyme-resistant fractions of beans lowered serum cholesterol and increased sterol excretions and hepatic mRNA levels in rats

Feeding rats beans with resistant starch reduces their serum cholesterol concentration; however, the mechanism by which this occurs is not fully understood. We examined the effects of enzyme-resistant fractions of adzuki (Vigna angularis) and tebou (Phaseolus vulgaris, var.) beans on serum cholesterol and hepatic mRNA in rats. Rats were fed a cholesterol-free diet with 50 g of cellulose powder (CP)/kg, 50 g of an enzyme-resistant fraction of adzuki starch (AS)/kg, or 50 g of an enzyme-resistant fraction of tebou starch (TS)/kg diet for 4 wk. There were no significant differences in body weight, liver weight, and cecum contents among the groups, nor was there a significant difference in food intake among the groups. The levels of serum total cholesterol, VLDL + intermediate density lipoprotein + LDL-cholesterol, and HDL cholesterol in the AS and TS groups were significantly (P<0.05) lower than in the CP group throughout the feeding period. Total hepatic cholesterol in the CP group was significantly (P<0.05) lower than in the AS and TS groups, fecal cholesterol excretion in the TS group was significantly (P<0.05) greater than in the CP and AS groups, and the fecal total bile acid concentrations in the AS and TS groups were significantly (P<0.05) higher than in the CP group. Cecal acetate, propionate, and n-butyrate concentrations in the AS and TS groups were significantly (P<0.05) higher than in the CP group. The level of hepatic scavenger receptor class B1 (SR-B1) mRNA in the TS group was significantly (P<0.05) higher than in the CP group, and the levels of hepatic cholesterol 7α-hydroxylase mRNA in the AS and TS groups were significantly (P<0.05) higher than in the CP group. These results suggest that AS and TS have a serum cholesterol-lowering function due to the enhanced levels of hepatic SR-B1 and cholesterol 7α-hydroxylase mRNA.     

Response of plasma lipids to dietary cholesterol and wine polyphenols in rats fed polyunsaturated fat diets

This experiment was designed to evaluate the effects of dietary red wine phenolic compounds (WP) and cholesterol on lipid oxidation and transport in rats. For 5 wk, weanling rats were fed polyunsaturated fat diets (n−6/n−3=6.4) supplemented or not supplemented with either 3 g/kg diet of cholesterol, 5 g/kg diet of WP, or both. The concentrations of triacylglycerols (TAG, P<0.01) and cholesterol (P<0.0002) were reduced in fasting plasma of rats fed cholesterol despite the cholesterol enrichment of very low density lipoprotein + low density lipoprotein (VLDL+LDL). The response was due to the much lower plasma concentration of high density lipoprotein (HDL) (−35%, P<0.0001). In contrast, TAG and cholesteryl ester (CE) accumulated in liver (+120 and +450%, respectively, P<0.0001). However, the cholesterol content of liver microsomes was not affected. Dietary cholesterol altered the distribution of fatty acids mainly by reducing the ratio of arachidonic acid to linoleic acid (P<0.0001) in plasma VLDL+LDL (−35%) and HDL (−42%) and in liver TAG (−42%), CE (−78%), and phospholipids (−28%). Dietary WP had little or no effect on these variables. On the other hand, dietary cholesterol lowered the α-tocopherol concentration in VLDL+LDL (−40%, P<0.003) and in microsomes (−60%, P<0.0001). In contrast, dietary WP increased the concentration in microsomes (+21%, P<0.0001), but had no effect on the concentration in VLDL+LDL. Cholesterol feeding decreased (P<0.006) whereas WP feeding increased (P<0.0001) the resistance of VLDL+LDL to copper-induced oxidation. The production of conjugated dienes after 25 h of oxidation ranged between 650 (WP without cholesterol) and 2,560 (cholesterol without WP) μmol/g VLDL+LDL protein. These findings show that dietary WP were absorbed at sufficient levels to contribute to the protection of polyunsaturated fatty acids in plasma and membranes. They could also reduce the consumption of α-tocopherol and endogenous antioxidants. The responses suggest that, in humans, these substances may be beneficial by reducing the deleterious effects of a dietary overload of cholesterol.