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Scope Vegetarian diets are considered health-promoting; however, a plasma cholesterol lowering effect is not always observed. We investigate the link between vegetarian-diet-induced alterations in cholesterol metabolism. Methods and results We study male and female omnivores, lacto-ovo vegetarians, lacto vegetarians, and vegans. Cholesterol intake, absorption, and fecal sterol excretion are measured as well as plasma concentrations of cholesterol and noncholesterol sterols. These serve as markers for cholesterol absorption, synthesis, and catabolism. The biliary cholesterol secretion rate is estimated. Flux data are related to body weight. Individual vegetarian diet groups are statistically compared to the omnivore group. Lacto vegetarians absorb 44% less dietary cholesterol, synthesized 22% more cholesterol, and show no differences in plasma total and LDL cholesterol. Vegan subjects absorb 90% less dietary cholesterol, synthesized 35% more cholesterol, and have a similar plasma total cholesterol, but a 13% lower plasma LDL cholesterol. No diet-related differences in biliary cholesterol secretion and absorption are observed. Total cholesterol absorption is lower only in vegans. Total cholesterol input is similar under all vegetarian diets. Conclusions Unaltered biliary cholesterol secretion and higher cholesterol synthesis blunt the lowered dietary cholesterol intake in vegetarians. LDL cholesterol is significantly lower only in vegans.

Objective— To determine whether long-term exposure to moderate elevations in plasma plant sterol levels increases risk for atherosclerosis. Methods and Results— In Old Order Amish participants aged 18 to 85 years, with (n=110) and without (n=181) 1 copy of the ABCG8 G574R variant, we compared mean plasma levels of plant sterols and cholesterol precursors and carotid intima-media wall thickness. Carriers of a single 574R allele had increased plant sterol levels (eg, 35%–37% higher plasma levels of sitosterol, campesterol, and stigmasterol) and increased plant sterol/cholesterol ratios ( P P =0.03). Adjustment for body weight, blood pressure, and standard lipid measures (high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, and triglycerides) did not alter this association. Conclusion— Although the G574R variant is associated with moderately elevated plant sterol levels, carriers of the 574R allele had modestly lower levels of carotid wall thickness compared with noncarriers.

Feeding of margarines containing sitostanol (CAS 19466-47-8), sitostanol acetate (CAS 73052-08-1), sitostanol oleate (CAS 107615-79-2), or placebo (equivalent of 0.5 g of sitostanol t.i.d) on cholesterol absorption and serum lipids were studied in 10 normolipemic volunteers in a randomized double blind cross-over trial. The study was divided into an open one-week run-in phase and four one-week treatment periods. Each treatment week was followed by a two-week washout period. Measurements of cholesterol absorption was performed by the continuous isotope feeding method using stable isotope labeled cholesterol and sitostanol. Cholesterol absorption during placebo, sitostanol, sitostanol acetate and sitostanol oleate feeding averaged 41.6 +/- (SD) 8.0, 10.2 +/- 6.6, 17.0 +/- 6.7, and 20.5 +/- 5.3%, respectively (p < 0.001 for all against placebo). Low density lipoprotein (LDL) cholesterol was proportionally reduced by 22 (p < 0.001), 14 (p < 0.05), and 8% (ns). Absorption efficiency was significantly lower with free sitostanol than with sitostanol acetate or oleate (p < 0.01). Percent reduction in cholesterol absorption with all preparations compared to placebo correlated positively with the percent reduction in LDL cholesterol (r = 0.404; p < 0.03). The results indicate that unesterified sitostanol is more effective in inhibiting cholesterol absorption and reducing LDL cholesterol than the acetate or oleate esters.

The development of statins improved the therapy of hypercholesterolemia and atherosclerotic disease tremendously. The beneficial effects of statins were clearly demonstrated in large scale primary and secondary prevention studies. In addition to the reduction of plasma cholesterol, inhibition of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase also results in the depletion of intermediates of cholesterol biosynthesis that are important for cellular integrity. The so called pleiotrophic effects of statins and probably also their adverse events can be attributed to the inhibition of synthesis of these intermediates. The review article describes the pathogenesis of atherosclerosis, the pharmacokinetic and pharmacodynamik of statins, and their pleiotrophic effects concerning endothelial function, LDL (low density lipoprotein) oxidation, macrophages, smooth muscle cell proliferation, atherosclerotic plaque, platelets, thrombosis, proinflammatory factors, haemorheology, hypertension, venous thrombosis, bone metabolism, stroke, and the possible influence on the prevention of Alzheimer's disease.

Lifibrol is a potent lipid-lowering drug with an unknown mechanism of action. We investigated its effects on lipoprotein and sterol metabolism in normocholesterolemic male participants. Seven participants were treated for 4 weeks with 600 mg/d lifibrol and 9 with 40 mg/d pravastatin in a double-blind randomized parallel-group trial. Kinetic studies were performed at baseline and under acute and chronic treatment. Turnover of apolipoprotein B-100 was investigated with endogenous stable-isotope labeling, and kinetic parameters were derived by multicompartmental modeling. Lathosterol and cholesterol metabolism were investigated using mass isotopomer distribution analysis (MIDA) after [1-13C]acetate labeling. Carbon metabolism was investigated by calculating the total isotope incorporation into newly formed sterols and measuring the disposal of acetate by 13CO2 breath analysis. Total- and low-density lipoprotein (LDL) cholesterol decreased by 18% and 27% under lifibrol and by 17% and 28% under pravastatin, respectively, whereas very-low-density lipoprotein (VLDL) cholesterol, triglycerides, and high-density lipoprotein (HDL) cholesterol did not change. Very-low-density lipoprotein apoB fractional synthesis and production increased under lifibrol but remained unchanged under pravastatin. Low-density lipoprotein apoB fractional synthesis and production increased under pravastatin but remained unchanged under lifibrol. Mass isotopomer distribution analysis indicated that both drugs decrease endogenous sterol synthesis after acute administration, but pravastatin had more powerful effects. Carbon-13 appearance in breath was higher during pravastatin than during lifibrol treatment. Mass isotopomer distribution analysis and carbon metabolism analysis indicated compartmentalization at the site of sterol synthesis, thus suggesting differential effects of the 2 drugs. Although having comparable lipid-lowering properties, lifibrol seems to have a mechanism of action distinct from that of statins. Lifibrol could serve as a model compound for the development of new lipid-lowering agents.

This study evaluates changes in cholesterol balance in hypercholesterolemic subjects following treatment with an inhibitor of cholesterol absorption or cholesterol synthesis or coadministration of both agents. This was a randomized, double blind, placebo-controlled, four-period crossover study to evaluate the effects of coadministering 10 mg ezetimibe with 20 mg simvastatin (ezetimibe/simvastatin) on cholesterol absorption and synthesis relative to either drug alone or placebo in 41 subjects. Each treatment period lasted 7 weeks. Ezetimibe and ezetimibe/simvastatin decreased fractional cholesterol absorption by 65% and 59%, respectively (P < 0.001 for both relative to placebo). Simvastatin did not significantly affect cholesterol absorption. Ezetimibe and ezetimibe/simvastatin increased fecal sterol excretion (corrected for dietary cholesterol), which also represents net steady state cholesterol synthesis, by 109% and 79%, respectively (P < 0.001). Ezetimibe, simvastatin, and ezetimibe/simvastatin decreased plasma LDL-cholesterol by 20, 38, and 55%, respectively. The coadministered therapy was well tolerated. The decreases in net cholesterol synthesis and increased fecal sterol excretion yielded nearly additive reductions in LDL-cholesterol for the coadministration of ezetimibe and simvastatin.

Apolipoprotein E (apoE) is a regulator of peripheral cholesterol homeostasis, and the apoE-isoform E4 is a major risk factor for the development of Alzheimer's disease (AD). Accumulating evidence suggests a key role for aberrant cholesterol metabolism in AD. We hypothesized that apoE-deficiency in mice not only affects cholesterol homeostasis in the periphery, but also in the brain, and that this can be restored by astrocyte-specific expression of human apoE3, but not apoE4. Using gas-chromatography mass-spectrometry, we found that absence of apoE in mice does not affect brain cholesterol homeostasis although serum sterol levels increase dramatically, especially when the apoE-knockout mice are fed a high fat diet. We provide evidence suggesting that apoD and the ATP-binding Cassette Transporter A1 (ABCA1) play a compensatory role in the apoE-deficient brain. Surprisingly, astrocyte-specific expression of human apoE3 or apoE4 in brains of apoE-knockout mice significantly increases brain levels of cholesterol and its precursors compared to control mice, indicative of an increased cholesterol synthesis rate in the brain. This increase is independent of the apoE-isoform, suggesting that the detrimental effect of apoE4 on the pathogenesis of AD is unlikely to be due to an apoE-isoform effect on brain cholesterol homeostasis.

The consumption of products enriched with plant sterol or stanol esters lowers serum total and LDL-cholesterol concentrations, thereby most likely reducing the risk of coronary heart disease. However, using plant sterol (not plant stanol) enriched products elevates serum plant sterol concentrations in humans. This may be unwanted because health effects of elevated serum plant sterol concentrations are still controversial. Within postlaunch monitoring of functional foods, we compared serum plant sterol and plant stanol concentrations among users of plant sterol (n = 67) or plant stanol (n = 13) enriched margarines with those of matched nonusers (n = 81) in the ongoing Dutch Doetinchem cohort study. Subjects (aged 29-67 y) were examined in 1994-1998 (before the introduction of enriched margarines) and re-examined in 1999-2003. Serum concentrations of plant sterols and stanols were measured in samples from nonfasting subjects by GLC-MS. Intake of plant sterols was 1.1 +/- 0.6 g/d and was associated with a decrease of serum total cholesterol concentration of 0.25 +/- 0.91 mmol/L (4%, P < 0.05), a change that differed (P < 0.05) from the nonsignificant increase in nonusers (+2%, 0.12 +/- 0.78 mmol/L, P = 0.16). Cholesterol-standardized serum sitosterol and campesterol increased in plant sterol users by 22% (P < 0.0001) and 103% (P < 0.0001), respectively. Cholesterol-standardized serum sitostanol and campestanol increased in plant stanol users by 197% (P = 0.02) and 196% (P = 0.01). To our knowledge, these data are the first to show changes in serum cholesterol, plant sterol, and plant stanol concentrations after (long-term) consumption of plant sterol and stanol enriched margarines in a free-living population in a nonexperimental setting. Whether the increased serum sterol concentrations result in adverse side effects needs to be investigated in future postlaunch monitoring studies.

Results of previous studies have shown that ezetimibe (10 mg/day) reduces LDL cholesterol in patients with mild hypercholesterolemia on a normal-cholesterol diet (dietary intake of 200-500 mg/day) by 16-22%. However, the LDL cholesterol-lowering effect of ezetimibe in subjects with an extremely low dietary cholesterol intake (vegetarians) has not been studied. We conducted a randomized, double-blind, placebo-controlled, two-phase crossover study in 18 healthy pure vegetarians to assess the effect of ezetimibe (10 mg/day) on plasma lipids, cholesterol absorption, and its synthesis. Treatment periods lasted 2 weeks each, with an intervening 2 week washout period. Fractional cholesterol absorption was determined using the continuous dual stable isotope feeding method. Mean dietary cholesterol intake in the pure vegetarians was extremely low and averaged 29.4 +/- 16.8 and 31.4 +/- 14.4 mg/day during the placebo and ezetimibe administration phases, respectively. Fractional cholesterol absorption during the placebo phase was 48.2 +/- 8.2% and was decreased by 58% during ezetimibe treatment to 20.2 +/- 6.2% (P < 0.001). This change in intestinal cholesterol absorption was followed by a significant reduction in LDL cholesterol of 17.3%. In individuals with extremely low dietary cholesterol intake, treatment with ezetimibe (10 mg/day) leads to a significant reduction of cholesterol absorption and a clinically relevant decrease of plasma LDL cholesterol, comparable to that of subjects with a normal dietary cholesterol intake. Thus, the lipid-lowering effect of ezetimibe is mediated mainly through a reduction of the absorption of endogenous (biliary) cholesterol.

We previously identified two inbred mouse strains, C57BL/6J and CASA/Rk, with different plasma plant sterol levels. An intercross between these strains revealed a broad plasma plant sterol locus on chromosome 14, which peaked at 17 centimorgan (cM) with a maximum logarithm of the odds score of 9.9. Studies in a chromosome 14 congenic strain, 14KK, with a 4–60 cM CASA/Rk interval on the C57BL/6J background revealed that males, but not females, had decreased plasma plant sterol levels and intestinal cholesterol absorption. In two subcongenic strains, 14PKK and 14DKK, with 4–19.5 and 19.5–60 cM CASA/Rk intervals, respectively, both males and females had decreased plasma plant sterol levels and decreased intestinal cholesterol absorption. Compatible with the decreased plasma plant sterol phenotype, 14PKK mice had increased biliary plant sterol excretion, whereas 14DKK mice did not. Therefore, gender-dependent interactions of genes at the 14PKK and 14DKK intervals are likely to underlie the 14KK interval effect on plasma plant sterol levels and sterol absorption from the intestine. These studies confirm the plasma plant sterol locus on mouse chromosome 14 and provide evidence that there are at least two sets of genes operating: one set affecting intestinal sterol absorption and biliary excretion, and the other set mainly affecting intestinal sterol absorption.

Background and Objective The polymorphism of SLCO1B1 (solute carrier organic anion transporter family, member 1B1), encoding the hepatic uptake transporter organic anion transporting polypeptide 1B1, has been associated with increased pravastatin concentrations in single-dose studies. We have investigated whether this polymorphism influences the pharmacokinetics and lipid-lowering efficacy of multiple-dose pravastatin. Methods A prospective, parallel-group study of 16 healthy volunteers, including 8 carriers of an SLCO1B1 variant haplotype and 8 control subjects, was carried out. Pravastatin pharmacokinetics and reduction in total and low-density lipoprotein (LDL) cholesterol concentrations were analyzed after treatment with 40 mg pravastatin daily for 3 weeks. Results The mean area under the plasma concentration-time curve of pravastatin was 110% higher (305.0 ± 149.4 ng · h/mL [mean ± SD] versus 144.9 ± 48.2 ng · h/mL, P = .012) and the mean peak concentration in plasma was 231% higher (174.5 ± 98.6 ng/mL versus 52.7 ± 22.1 ng/mL, P = .0042) in the SLCO1B1 variant haplotype group than in the control group. Pravastatin significantly reduced the concentrations of total and LDL cholesterol in both groups. The mean percentage reduction in total cholesterol concentration was 13.1% ± 9.1% and 19.1% ± 8.3% in the variant and control groups, respectively (P = .19; 95% confidence interval of difference between groups, −15.3% to 3.3%). The mean percentage reduction in LDL cholesterol concentration was 27.7% ± 8.3% in the variant group and 32.3% ± 11.2% in the control group (P = .37; 95% confidence interval for difference, −15.1% to 6.0%). Conclusions Despite considerably higher plasma pravastatin concentrations in carriers of an SLCO1B1 variant haplotype, there was no significant difference in the lipid-lowering efficacy of pravastatin between the variant haplotype and control groups. However, this pilot study had sufficient statistical power to detect only a large difference in lipid response between the 2 groups. Further clinical studies are warranted to characterize the impact of the SLCO1B1 polymorphism on the lipid response to pravastatin. Clinical Pharmacology & Therapeutics (2006) 79, 419–426; doi: 10.1016/j.clpt.2006.01.010

We previously showed that variant SLCO1B1 haplotype *1b (A388G) accelerates and that *5 (T521C) delays hepatocellular uptake of the HMG-CoA reductase inhibitor pravastatin [Mwinyi et al. (2004): Clin Pharmacol Ther 75:415-421]. In the present study we checked for differential effects of variant SLCO1B1 haplotypes on hepatocellular cholesterol synthesis. We analyzed the serum levels of cholesterol, lathosterol, and campesterol in healthy white males which had been grouped on the basis of their SLCO1B1 haplotype: *1a (n=10), *1b (n=10), and *5 (n=8). The subjects received a single oral dose of 40 mg pravastatin. Cholesterol and lathosterol levels were lower in all subjects following pravastatin intake for up to 24. Median levels 6 h post-dosing of lathosterol decreased in each SLCO1B1 haplotype group in the rank order of *1b (-0.11 mg dl(-1); min-max: -0.20 to -0.04; p=0.005)*1a (-0.09 mg dl(-1); min-max: -0.22 to -0.05; p=0.005)*5 (-0.07 mg dl(-1); min-max: -0.17 to -0.05; p=0.012). Lathosterol median-change values were significantly greater in haplotype *1b than in haplotype *5 individuals (p=0.041, non-adjusted), which was congruent with the extent of mean changes in lathosterol-to-cholesterol ratios, although the latter did not reach statistical significance. Post-treatment serum levels of campesterol were not affected by SLCO1B1 haplotype. Interestingly, sterol basal serum levels tended to be highest in *1b carriers, followed by those in *1a and *5 individuals, with significant differences in lathosterol concentrations between the *1b and *5 (p=0.041, non-adjusted) haplotype group. Our findings suggest an association of SLCO1B1*1b and *5 haplotypes to pravastatin's inhibition of the hepatocellular HMG-CoA reductase. Furthermore, SLCO1B1 haplotypes seem to play a role in basal cholesterol homeostasis.

Sterol regulatory element binding proteins-1 and -2 (SREBPs) are transcription factors controlling lipid homeostasis in human cells. The G-allele carriers of the SREBF-1 gene C-G polymorphism in exon 18c and coding for glycine at the protein level (G952G) have shown to associate more frequently with obesity and type 2 diabetes than the C-allele carriers. However, the C-allele has suggested to be linked to dyslipidemia. Thus, our aim was to study effect of the SREBF-1 gene polymorphism (G952G) on sterol metabolism in man. Ninety-five subjects with moderate hypercholesterolemia participated in this study and 14 homozygous CC carriers of the SREBF-1 (G952G) gene were found. Plasma lathosterol concentration and lathosterol-to-cholesterol ratio, markers of endogenous cholesterol synthesis, were significantly higher in CC homozygous subject compared to others. Similarly muscle cholesterol (p=0.045) and lathosterol (p=0.054) concentrations were elevated in the CC homozygotes supporting the view that endogenous cholesterol synthesis rate is SREBF-1 genotype-dependent.

Background and Aims Ezetimibe is an inhibitor of the cholesterol uptake transporter Niemann-Pick C1-like protein (NPC1L1). Target concentrations can be influenced by intestinal uridine diphosphate–glucuronosyltransferases (UGTs) and the efflux transporters P-glycoprotein (P-gp) (ABCB1) and multidrug resistance associated protein 2 (MRP2) (ABCC2). This study evaluates the contribution of these factors to the disposition and cholesterol-lowering effect of ezetimibe before and after induction of UGT1A1, P-gp, and MRP2 with rifampin (INN, rifampicin). Methods Serum concentrations of ezetimibe, as well as its glucuronide, and the plant sterols campesterol and sitosterol (surrogate for cholesterol absorption) were studied in 12 healthy subjects before and after rifampin comedication. In parallel, duodenal expression of UGT1A1, P-gp, MRP2, and NPC1L1 was quantified by use of real-time reverse transcriptase–polymerase chain reaction and quantitative immunohistochemical evaluation. The affinity of ezetimibe and its glucuronide to P-gp and MRP2 was assessed in P-gp– overexpressing Madin-Darby canine kidney II cells and P-gp–containing or MRP2-containing inside-out vesicles. Results Up-regulation of intestinal P-gp, MRP2, and UGT1A1 (but not of NPC1L1) by rifampin was associated with markedly decreased areas under the curve of ezetimibe and its glucuronide (116 ± 78.1 ng·h/mL versus 49.9 ± 31.0 ng·h/mL and 635 ± 302 ng·h/mL versus 225 ± 86.4 ng·h/mL, respectively; both P = .002) and increased intestinal clearances (2400 ± 1560 mL/min versus 5500 ± 4610 mL/min [P = .003] and 76.6 ± 113 mL/min versus 316 ± 457 mL/min [P = .010], respectively) and nearly abolished sterol-lowering effects. Intestinal expression of UGT1A1, ABCB1, and ABCC2 was inversely correlated with the effects of ezetimibe on plant sterol serum concentrations. Parallel in vitro studies confirmed that ezetimibe glucuronide is a high-affinity substrate of MRP2 and has a low affinity to P-gp whereas ezetimibe interacts with P-gp and MRP2. Conclusions The disposition and sterol-lowering effects of ezetimibe are modified by metabolic degradation of the drug via intestinal UGT1A1 and either intestinal or hepatic secretion (or both) via P-gp and MRP2. Clinical Pharmacology & Therapeutics (2006) 79, 196–196; doi: 10.1016/j.clpt.2005.11.004

Dietary plant sterols and cholesterol have a comparable chemical structure. It is generally assumed that cholesterol and plant sterols do not cross the blood-brain barrier, but quantitative data are lacking. Here, we report that mice deficient for ATP-binding cassette transporter G5 (Abcg5) or Abcg8, with strongly elevated serum plant sterol levels, display dramatically increased (7- to 16-fold) plant sterol levels in the brain. Apolipoprotein E (ApoE)-deficient mice also displayed elevated serum plant sterol levels, which was however not associated with significant changes in brain plant sterol levels. Abcg5- and Abeg8-deficient mice were found to carry circulating plant sterols predominantly in high-density lipoprotein (HDL)-particles, whereas ApoE-deficient mice accommodated most of their serum plant sterols in very low-density lipoprotein (VLDL)-particles. This suggests an important role for HDL and/or ApoE in the transfer of plant sterols into the brain. Moreover, sitosterol upregulated apoE mRNA and protein levels in astrocytoma, but not in neuroblastoma cells, to a higher extend than cholesterol. In conclusion, dietary plant sterols pass the blood-brain barrier and accumulate in the brain, where they may exert brain cell type-specific effects. (c) 2006 Elsevier B.V. All rights reserved.

The HMG-CoA reductase inhibitors, or statins, are well established in the prevention and treatment of coronary artery disease, mainly by lowering low-density lipoprotein (LDL) cholesterol levels. These compounds are structurally similar, but differ in their lipophilicity. Several studies have indicated a link between cholesterol and Alzheimer’s disease (AD), and there is also epidemiological evidence that statin treatment may decrease the prevalence of dementias. In the present study we wanted to investigate whether pravastatin treatment affects brain cholesterol metabolism. A post hoc analysis was performed with plasma material from a clinical trial where 51 healthy men (35±4 years) were randomly assigned to receive either pravastatin (40 mg/day) or placebo for 6 months. Cholesterol, its precursor lathosterol, its brain-specific metabolite 24(S)-hydroxycholesterol (24S-OH-chol) and 27-hydroxycholesterol (27-OH-chol) were determined in plasma samples before and after treatment by using gas-liquid chromatography (GC)-flame ionization detection (GC-FID) and GC mass spectrometry (GC-MS). Besides reducing total cholesterol (−20%, P

Consumption of plant sterols and treatment with ezetimibe both reduce cholesterol absorption in the intestine. However, the mechanism of action differs between the two treatments, and the consequences of combination treatment are unknown. Therefore, we performed a double-blind, placebo-controlled, crossover study for the plant sterol component with open-label ezetimibe treatment. Forty mildly hypercholesterolemic subjects were randomized to the following treatments for 4 weeks each: 10 mg/day ezetimibe combined with 25 g/day control spread; 10 mg/day ezetimibe combined with 25 g/day spread containing 2.0 g of plant sterols; 25 g/day spread containing 2.0 g of plant sterols; and placebo treatment consisting of 25 g/day control spread. Combination treatment of plant sterols and ezetimibe reduced low density lipoprotein cholesterol (LDL-C) by 1.06 mmol/l (25.2%; P < 0.001) compared with 0.23 mmol/l (4.7%; P = 0.006) with plant sterols and 0.94 mmol/l (22.2%; P < 0.001) with ezetimibe monotherapy. LDL-C reduction conferred by the combination treatment did not differ significantly from ezetimibe monotherapy (-0.12 mmol/l or -3.5%; P = 0.13). Additionally, the plasma lathosterol-to-cholesterol ratio increased with all treatments. Sitosterol and campesterol ratios increased after plant sterol treatment and decreased upon ezetimibe and combination therapy. Our results indicate that the combination of plant sterols and ezetimibe has no therapeutic benefit over ezetimibe monotherapy in subjects with mild hypercholesterolemia.

The aim of this study was to characterize the role of the efflux transporter Mrp2 (Abcc2) in the pharmacokinetics of orally and intravenously administered pravastatin in rats. Eight Mrp2-deficient TR- rats and eight wild-type rats were given an oral dose of 20 mg/kg pravastatin. Four TR- animals and four wild-type animals were studied after intravenous administration of pravastatin (5 mg/kg). The TR(-) rats showed a 6.1-fold higher mean area under the plasma concentration-time curve (AUC) of pravastatin (p < 0.001) after oral administration and a 4.7-fold higher AUC (p < 0.01) after intravenous administration of pravastatin as compared with the wild-type animals. The mean systemic (total) clearance of pravastatin was 4.6-fold higher (39.2 versus 8.50 l/h/kg, p < 0.001) and the mean V 4.3-fold higher (14.1 versus 3.29 l/kg, p < 0.01) in the wild-type rats. The mean renal clearance of pravastatin in the TR(-) rats was 16.5-fold increased as compared with the wild-type animals (0.695 versus 0.042 l/h/kg, p < 0.05). The increased systemic exposure to oral pravastatin in the TR- rats was associated with a greater inhibitory effect on 3-hydroxy-3-methylglutaryl CoA reductase, as shown by smaller lathosterol to cholesterol concentration ratios. These results suggest that the reduced biliary pravastatin excretion in the Mrp2-deficient TR- rats is partly compensated for by increased urinary excretion of pravastatin. Furthermore, intestinal Mrp2 does not appear to play a major role in the oral absorption of pravastatin in normal rats.

Screening of 932 adults on the Pacific island of Kosrae for plasma plant sterol levels disclosed three subjects, two of them asymptomatic, with phytosterolemia. Sequencing the ATP binding cassette subfamily G member 8 (ABCG8) gene revealed a novel exon 2 mutation that causes a change in codon 24 from glutamine to histidine and a frame shift followed by a premature stop codon, precluding the formation of a functional ABCG8 protein. Genotyping of 1,090 Kosraens revealed 150 as carriers, a 13.8% carrier rate. DNA sequencing of 67 carriers revealed the same mutation as in the probands. In carriers, plasma campesterol and sitosterol levels were 55% and 30% higher, respectively, than in noncarriers. Moreover, compared with noncarriers, carriers showed 21% lower plasma levels of lathosterol, a surrogate marker for cholesterol biosynthesis. There was no difference between the groups in plasma total cholesterol, triglycerides, apolipoprotein B, or apolipoprotein A-I levels. In summary, on the island of Kosrae, a strong founder effect of a mutant ABCG8 allele results in a large number of carriers with increased plasma plant sterol levels and decreased lathosterol levels. The latter finding suggests that heterozygosity for a mutated ABCG8 allele results in a modest increase in dietary cholesterol absorption and a decrease in cholesterol biosynthesis.

Recent epidemiological studies suggest that inhibitors of 3-hydroxy-3-methyl-glutaryl CoA reductase, so-called statins, are effective in lowering the prevalence of Alzheimer's disease. Whether the effect of statins is due to a local inhibition of cholesterol synthesis in the brain or whether it is mediated by the reduced levels of cholesterol in the circulation is not known. In the present work, we tested the possibility that high doses of lipophilic and hydrophilic statins, simvastatin and pravastatin, respectively, or a diet high in cholesterol could affect cholesterol homeostasis in the brain of guinea pigs. The total brain cholesterol levels were not affected by high-dose simvastatin or pravastatin treatment. Significantly lower levels of the cholesterol precursor lathosterol and its ratio to cholesterol were found in the brains of simvastatin and pravastatin-treated animals. 24S-Hydroxycholesterol, the transportable form of cholesterol across the blood-brain barrier, was significantly lower in the brain of pravastatin-treated animals. Excessive cholesterol feeding resulted in higher serum cholesterol levels but did not affect total brain cholesterol level. However, de novo cholesterol synthesis in the brain seemed to be down-regulated, as indicated by lower absolute levels and cholesterol-related ratios of lathosterol compared with controls. The passage of deuterium-labeled cholesterol across the blood-brain barrier in one animal was found to be approximately 1%. Our results suggest that brain cholesterol synthesis in guinea pigs can be slightly, but significantly, influenced by high doses of lipophilic and hydrophilic statins as well as by high dietary cholesterol intake, while total brain cholesterol content and thus, cholesterol homeostasis is maintained.

To examine the effect of gender and polymorphisms of CYP46 and apo E on plasma levels of 24S-hydroxycholesterol in Alzheimers disease (AD) patients and to determine whether these factors contribute to the variability in responses to statin treatment. Fifty-three AD patients had measurement of plasma levels of 24S-hydroxycholesterol , plasma and lipoprotein cholesterol and genotyping of CYP46 and apo E. Thirty-nine of the subjects subsequently participated in a statin trial for 6 weeks, and had a repetition of the baseline measurements. Baseline levels of 24S-hydroxycholesterol were higher in women than in men. There was a positive and significant correlation of plasma oxysterol levels with levels of total plasma cholesterol (women: r = .72, P < .0001; men: r = .47, P = .02) and non-HDL cholesterol (women: r = .68, P < .0001; men: r = 0.51, P = .01) (and LDL cholesterol) but not HDL cholesterol levels. There was no association of CYP46 or apo E polymorphisms with plasma levels of 24S-hydroxycholesterol. AD subjects treated with statins had a similar percent reduction in lathosterol, 24S-hydroxycholesterol, total cholesterol and non-HDL (and LDL) cholesterol regardless of gender and polymorphisms of CYP46. Subjects with the e4 / 4 polymorphism had less reduction in the ratios of 24S-hydroxycholesterol-LDL cholesterol. Women with AD had higher levels of plasma 24S-hydroxycholesterol levels than men. Women also showed a very strong correlation of plasma levels of 24S-hydroxycholesterol-to-total and non-HDL cholesterol. This may suggest that the oxysterol may be an important marker of AD risk instead of total cholesterol, as suggested by others. Polymorphisms of CYP46 or apo E do not explain levels of oxysterol or non-HDL cholesterol or the responsiveness to statin treatment in this study.

Sterol regulatory element-binding proteins (SREBPs) are transcription factors involved in cholesterol and fatty acid synthesis. Recently, a polymorphism in the 5′-region of the SREBP-1a gene has been described to be correlated with alterations in the plasma levels of cholesterol. Consequently the relationship between this SREBP-1a gene polymorphism and Alzheimer’s disease (AD) alone and in combination with the apolipoprotein E (ApoE) 4 allele was evaluated. No association between SREBP-1a polymorphism alone and AD could be seen. However, in the group of healthy ApoE4 allele carriers, the number of homozygote SREBP-1a ΔG allele carriers was significantly higher than in AD patients. Cerebrospinal fluid levels of cholesterol were lower in AD patients who were carriers of the SREBP-1a ΔG allele, and the ratio of 24S-hydroxycholesterol to cholesterol was increased in these probands. Our data suggest a reduced risk of AD in carriers of an ApoE4 allele who are additionally homozygous for the SREBP-1a ΔG allele, which is possibly due to the influence of SREBP-1a polymorphism on brain cholesterol metabolism. This is the first report on a genetic factor which prevents the deleterious effect of the ApoE4 allele and thus reduces the risk of AD.

The present study investigated the role of apolipoprotein E (apoE) phenotype on intestinal cholesterol absorption and cholesterol synthesis. Studies were carried out in eight subjects homozygous for the apoE4 and 12 subjects homozygous for the E2 allele (six normocholesterolemic volunteers and six patients with type III hyperlipoproteinemia). Cholesterol absorption did not differ between the three groups of subjects and averaged 38 +/- 2% (mean +/- SEM) in normolipemic E2/2, 37 +/- 4% in type III hyperlipemic E2/2, and 41 +/- 3% in E4/4 subjects, respectively. Dietary intake of fat and cholesterol had no influence on cholesterol absorption efficiency. A positive correlation between efficiency of cholesterol absorption and the ratio of campesterol to cholesterol in plasma, an indirect marker for cholesterol absorption, was observed after combining the results of the three groups (r = 0.504; P < 0.02). Bile acid and total cholesterol synthesis were also not affected by the different apoE alleles, but the well-known relationship between body weight and cholesterol synthesis was noticed (r = 0.574; P < 0.01). Thus, the present study provides evidence that the efficiency of intestinal absorption and synthesis of cholesterol in humans are not related to the apoE phenotype.

Plasma plant sterol levels differ among humans due to genetic and dietary factors. A disease characterized by high plasma plant sterol levels, β-sitosterolemia, was recently found to be due to mutations at the ABCG5/ABCG8 locus. To detect variants at this and other loci, a genetic cross was carried out between two laboratory mouse strains. Parental C57BL/6J had almost twice the campesterol and sitosterol levels compared with parental CASA/Rk mice, and F 1 mice had levels halfway between the parentals. An intercross between F 1 s was performed and plasma plant sterol levels measured in 102 male and 99 female F 2 mice. Plasma plant sterols in F 2 s displayed a unimodal distribution, suggesting the effects of several rather a single major gene. In the F 2 mice, a full genome scan revealed significant linkages on chromosomes 14 and 2. With regard to chromosome 14, analysis showed a single peak for linkage at 17 cM with a logarithm of odds (LOD) score of 9.9, designated plasma plant sterol 14 ( Plast14 ). With regard to chromosome 2, analysis showed two significant peaks for linkage at 18 and 65 cMs with LOD scores of 4.1 and 3.65, respectively, designated Plast2a and Plast2b , respectively. Four interactions between loci, predominantly of an additive nature, were also demonstrated, the most significant between Plast14 and Plast2b (LOD 16.44). No significant linkage or gene interaction was detected for the ABCG5/ABCG8 locus on chromosome 17. Therefore, other genes besides ABCG5/ABCG8 influence plasma plant sterol levels and now become candidates to explain differences in plasma plant sterol levels between humans.

Cholesterol and other sterols exit the body primarily by secretion into bile. In patients with sitosterolemia, mutations in either of two ATP-binding cassette (ABC) half-transporters, ABCG5 or ABCG8, lead to reduced secretion of sterols into bile, implicating these transporters in this process. To elucidate the roles of ABCG5 and ABCG8 in the trafficking of sterols, we disrupted Abcg5 and Abcg8 in mice ( G5G8 −/− ). The G5G8 −/− mice had a 2- to 3-fold increase in the fractional absorption of dietary plant sterols, which was associated with an ≈30-fold increase in plasma sitosterol. Biliary cholesterol concentrations were extremely low in the G5G8 −/− mice when compared with wild-type animals (mean = 0.4 vs. 5.5 μmol/ml) and increased only modestly with cholesterol feeding. Plasma and liver cholesterol levels were reduced by 50% in the chow-fed G5G8 −/− mice and increased 2.4- and 18-fold, respectively, after cholesterol feeding. These data indicate that ABCG5 and ABCG8 are required for efficient secretion of cholesterol into bile and that disruption of these genes increases dramatically the responsiveness of plasma and hepatic cholesterol levels to changes in dietary cholesterol content.

In a randomized, placebo-controlled, double-blind study, we investigated whether statins alter cholesterol metabolites and reduce Abeta levels in the cerebrospinal fluid of 44 patients with Alzheimer's disease. Individuals were given up to 80mg simvastatin daily or placebo for 26 weeks. Overall, simvastatin did not significantly alter cerebrospinal fluid levels of Abeta40 and Abeta42. In post hoc analysis, simvastatin significantly decreased Abeta40 levels in the cerebrospinal fluid of patients with mild Alzheimer's disease. The reduction of Abeta40 correlated with the reduction of 24S-hydroxycholesterol. These changes were not observed in more severely affected patients.

Ratios of cholestanol, campesterol, and sitosterol to cholesterol in serum are known to reflect cholesterol absorption efficiency. Here, a possible link between these ratios and biliary secretion rates of cholesterol was investigated. Biliary lipid secretion rates and serum sterols were determined in 13 patients with gallstones. Seven were treated with ursodeoxycholic acid (UDCA) (1,000 mg/d). Serum cholesterol and non-cholesterol sterols were also measured in a cross over study in 20 healthy volunteers, who received either placebo or UDCA (750 mg/d). Biliary cholesterol secretion was significantly lower, whereas the non-cholesterol sterols and their ratio to cholesterol were higher in patients with gallstones treated with UDCA. A highly significant negative linear correlation between the ratios of non-cholesterol sterols to cholesterol and biliary cholesterol secretion was observed. In volunteers, administration of UDCA for 4 weeks was followed by a significant increase in non-cholesterol sterols and their ratios. Even 4 weeks after discontinuing UDCA administration, campesterol and sitosterol were still significantly higher than pretreatment levels, which was also true for the campesterol-cholesterol ratio after 8 weeks. The results suggest that the ratios of cholestanol, campesterol, and sitosterol to cholesterol can be used as indicators of changes in biliary cholesterol secretion rates.

There is mounting evidence that specific sterol precursors of cholesterol are associated with reproductive functions. Testicular meiosis-activating sterol (T-MAS) and follicular fluid meiosis-activating-sterol (FF-MAS) are 2 cholesterol precursors found in reproductive organs of mammals, which are able to overcome meiotic arrest in vitro. This study investigates the influence of simvastatin and pravastatin, 2 inhibitors of 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase, and BM 15.766, an inhibitor of 7-dehydrocholesterol reductase, on concentrations of neutral sterols in liver and testis of guinea pigs. Concentrations of T-MAS, lathosterol, and desmosterol were markedly higher in testis compared with liver. Simvastatin (150 mg/d) and pravastatin (150 mg/d and 350 mg/d) markedly reduced cholesterol precursors in liver. In contrast, T-MAS and desmosterol in testis remained unchanged, albeit other cholesterol precursors were reduced. BM 15.766 led to the accumulation of 7- and 8-dehydrocholesterol, both in liver and testis. However, concentrations of T-MAS in testis were not changed by BM 15.766. We conclude that treatment of guinea pigs with simvastatin, pravastatin, or BM 15.766, which simulates the biochemical defect of the Smith-Lemli-Opitz (SLO) syndrome, does not affect T-MAS concentrations in testis.

Dietary phytosterols have been reported to lower total and low-density lipoprotein (LDL) cholesterol. However, less is known about the influence of cholesterol and fat intake on the cholesterol-lowering effect of esterified phytosterols in mild to moderate hypercholesterolemia. Sixty-three healthy subjects (38 women, 25 men, 42 +/- 11 years, LDL cholesterol > 130 mg/dL) were investigated in a randomized, double-blind, placebo-controlled, cross-over study. A total of 20 g/d of a phytosterol ester-enriched margarine (1.82 g/d of phytosterols) was compared with a control margarine (0.06 g/d of phytosterols). After 3 weeks of intake, participants crossed over to the other margarine. A 3-day dietary recall was performed at the beginning and at the end of the study to assess cholesterol, fat, and energy intake. Phytosterol ester-enriched margarine significantly changed total cholesterol (-3.4%, P

Treatment with carbamazepine (CBZ) affects cholesterol concentrations, but little is known about the precise nature and underlying mechanisms of changes in lipoprotein metabolism. We investigated prospectively the effects of CBZ on lipid metabolism in normolipemic adults. In 21 healthy males, lipoprotein and noncholesterol sterol concentrations were measured before and during treatment with CBZ for 70 ± 18 days. Thirteen subjects underwent kinetic studies of apolipoprotein-B (ApoB) metabolism with the use of endogenous stable isotope labeling. Lipoprotein kinetic parameters were calculated by multicompartmental modeling. Significant increases in total cholesterol, in ApoB-containing lipoproteins [very-low-density lipoprotein (VLDL), intermediate density lipoprotein (IDL), and low-density lipoprotein (LDL)], and in triglycerides, but not in high-density lipoprotein (HDL), were observed. Lipoprotein particle composition remained unchanged. Mean fractional catabolic and production rates of ApoB-containing lipoproteins were not significantly different, although mean production rates of VLDL and IDL were substantially increased (+46 ± 139% and +30 ± 97%, respectively), whereas mean production of LDL remained unchanged (+2.1 ± 45.6%). Cholestanol in serum increased significantly but not the concentrations of plant sterols (campesterol, sitosterol) and the cholesterol precursors (lathosterol, mevalonic acid). There was a significant correlation between the decrease in free thyroxine and the increase in IDL cholesterol. Treatment with CBZ increases mainly ApoB-containing lipoproteins. CBZ seems not to influence endogenous cholesterol synthesis or intestinal absorption directly. The increase is neither related to increased ApoB production nor to decreased catabolism but is rather due to changes in the conversion cascade of IDL particles, most likely as an indirect effect through a decrease in thyroid hormones.

The benefits of lipid lowering therapy on coronary heart disease have been clearly established in many clinical trials on primary and secondary prevention. Despite the availability of potent lipid lowering drugs, many patients do not reach the current treatment goals. This paper reviews new therapeutic approaches in lipid lowering drugs focusing on compounds which lower cholesterol absorption. The role of plant sterols and stanols, new acyl-CoA:cholesterol O-acyl transferase (ACAT) inhibitors, microsomal triglyceride transfer protein (MTP) inhibitors, and ezetimibe are summarised. Although the lipid lowering effect of plant sterols and plant stanols is only moderate, their use as functional foods is beneficial for patients with mild hypercholesterolaemia and is able to enhance the lipid lowering effect of HMG-CoA reductase inhibitors (statins). The role of ACAT inhibitors that might also inhibit cholesterol absorption remains unclear. Avasimibe, the first oral bioavailable ACAT inhibitor, has entered phase III trials. However, the presently available data in humans do not indicate a clear clinical benefit. The role of MTP inhibitors, which exhibit remarkable effects on all plasma lipids, also remains unclear, as safety concerns must first be addressed. Ezetimibe, the first available 2-azetidinone, succeeded in phase III trials showing remarkable effects in inhibition of cholesterol absorption as well as cholesterol lowering. The synergistic effect of co-administration of ezetimibe with statins seemingly offers a new approach in reaching the therapeutic goals.