Plasma oxyphytosterols most likely originate from hepatic oxidation and subsequent spill-over in the circulation

We evaluated oxyphytosterol (OPS) concentrations in plasma and various tissues of two genetically modified mouse models with either increased cholesterol (apoE KO mice) or increased cholesterol and plant sterol (PS) concentrations (apoExABCG8 dKO mice). Sixteen female apoE KO and 16 dKO mice followed the same standard, low OPS-chow diet. Animals were euthanized at 36 weeks to measure PS and OPS concentrations in plasma, brain, liver and aortic tissue. Cholesterol and oxysteml (OS) concentrations were analyzed as reference for sterol oxidation in general. Plasma campesterol (24.1 +/- 4.3 vs. 11.8 +/- 3.0 mg/dL) and sitosterol (67.4 +/- 12.7 vs. 4.9 +/- 1.1 mg/dL) concentrations were severely elevated in the dKO compared to the apoE KO mice (p < 0.001). Also, in aortic and brain tissue, PS levels were significantly elevated in dKO. However, plasma, aortic and brain OPS concentrations were comparable or even lower in the dKO mice. In contrast, in liver tissue, both PS and OPS concentrations were severely elevated in the dKO compared to apoE KO mice (sum OPS: 7.4 +/- 1.6 vs. 4.1 +/- 0.8 ng/mg, p < 0.001). OS concentrations followed cholesterol concentrations in plasma and all tissues suggesting ubiquitous oxidation. Despite severely elevated PS concentrations, OPS concentrations were only elevated in liver tissue, suggesting that OPS are primarily formed in the liver and plasma concentrations originate from hepatic spill-over into the circulation

Effects of plant sterol- or stanol-enriched margarine on fasting plasma oxyphytosterol concentrations in healthy subjects.

AbstractBackgroundConsumption of plant sterols and plant stanols reduces low-density lipoprotein cholesterol (LDL-C) concentrations. At the same time, plasma plant sterol concentrations will increase after plant sterol consumption, but decrease after plant stanol consumption. In contrast to plant stanols, plant sterols can undergo oxidation and form oxyphytosterols. Findings from in vitro and animal studies suggest that oxyphytosterols might be atherogenic.ObjectiveThe objective was to examine whether plant sterol and stanol consumption changes fasting plasma oxyphytosterol concentrations.DesignA randomized, double blind, cross-over study was performed in which 43 healthy subjects (18–70 years) consumed for 4 weeks a plant sterol-enriched (3.0 g/d of plant sterols), a plant stanol-enriched (3.0 g/d of plant stanols), and a control margarine separated by wash-out periods of 4 weeks. Oxyphytosterol concentrations were determined in BHT-enriched plasma via GC–MS.ResultsCompared to control, serum LDL-C concentrations were reduced after plant sterol (−8.1%; p < 0.001) and plant stanol consumption (−7.8%; p < 0.001). Plant sterol consumption did not change plasma oxyphytosterol concentrations. On the other hand, intake of the plant stanol margarine reduced 7β-OH-campesterol by 0.07 ng/mL (∼14%; p < 0.01) and by 0.07 ng/mL (∼15%; p < 0.01) compared with the control and sterol margarines, respectively. When standardized for serum cholesterol, effects on these oxyphytosterols were comparable. In addition, plant stanol intake reduced cholesterol-standardized 7-keto-campesterol levels compared with plant sterol intake (p < 0.05).ConclusionsDaily consumption of a plant sterol-enriched margarine does not increase oxyphytosterol concentrations, while plant stanol consumption may reduce the concentrations of the oxidative plant sterol metabolites 7β-OH-campesterol and 7-keto-campesterol. This trial is registered at clinicaltrials.gov as NCT01559428

Plasma oxyphytosterols most likely originate from hepatic oxidation and subsequent spill-over in the circulation

We evaluated oxyphytosterol (OPS) concentrations in plasma and various tissues of two genetically modified mouse models with either increased cholesterol (apoE KO mice) or increased cholesterol and plant sterol (PS) concentrations (apoExABCG8 dKO mice). Sixteen female apoE KO and 16 dKO mice followed the same standard, low OPS-chow diet. Animals were euthanized at 36 weeks to measure PS and OPS concentrations in plasma, brain, liver and aortic tissue. Cholesterol and oxysterol (OS) concentrations were analyzed as reference for sterol oxidation in general. Plasma campesterol (24.1 ± 4.3 vs. 11.8 ± 3.0 mg/dL) and sitosterol (67.4 ± 12.7 vs. 4.9 ± 1.1 mg/dL) concentrations were severely elevated in the dKO compared to the apoE KO mice (p < 0.001). Also, in aortic and brain tissue, PS levels were significantly elevated in dKO. However, plasma, aortic and brain OPS concentrations were comparable or even lower in the dKO mice. In contrast, in liver tissue, both PS and OPS concentrations were severely elevated in the dKO compared to apoE KO mice (sum OPS: 7.4 ± 1.6 vs. 4.1 ± 0.8 ng/mg, p < 0.001). OS concentrations followed cholesterol concentrations in plasma and all tissues suggesting ubiquitous oxidation. Despite severely elevated PS concentrations, OPS concentrations were only elevated in liver tissue, suggesting that OPS are primarily formed in the liver and plasma concentrations originate from hepatic spill-over into the circulation

Oxyphytosterol formation in humans: Identification of high vs. low oxidizers.

Animal studies suggest that oxyphytosterols are atherogenic. However, we previously shown that plasma oxyphytosterol concentrations did not consuming a diet enriched in plant sterol esters (3g/day), while minor were seen after consuming a plant stanol ester-enriched diet. Large oxyphytosterol concentrations between individuals however existed. The this study was to identify factors that may explain inter-individual in plasma oxyphytosterol concentrations. For this, 43 subjects consumed weeks in random order a plant sterol, stanol and control margarine. oxyphytosterol concentrations were determined in butylated (BHT)-enriched EDTA plasma via GC-MS and serum oxidized low-density (oxLDL) concentrations were analyzed via ELISA. Trolox equivalent capacity (TEAC) values, alpha-tocopherol concentrations and iron/copper were determined to assess plasma oxidative and anti-oxidative capacity. (non-oxidized) sitosterol and campesterol concentrations did not plasma oxysitosterol and oxycampesterol concentrations during any of the dietary interventions. Moreover, plasma oxyphytosterol concentrations relatively stable over time. Six subjects could be arbitrarily having consistent low or high plasma oxyphytosterol concentrations, also reflected in oxLDL concentrations. However, oxidative and anti- capacity markers, such as iron/copper status, alpha-tocopherol TEAC values, could not explain these differences. In conclusion, have consistent plasma oxyphytosterol concentrations, which resulted in identification of 'low and high oxidizers'. Differences, however, could attributed to the oxidative and anti-oxidative capacity marker

The influence of consuming an egg or an egg-yolk buttermilk drink for 12 wk on serum lipids, inflammation, and liver function markers in human volunteers

OBJECTIVES: Dietary cholesterol elevates serum total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) concentrations modestly. There are indications that the cholesterol-raising effect depends on the food matrix, that is, sphingolipids and lactic acid bacteria are suggested to influence cholesterol metabolism. Traditional buttermilk is rich in both sphingolipids and lactic acid bacteria. Therefore, the aim of this study was to evaluate whether effects on cholesterol metabolism depend on food matrix (e.g., cholesterol provided as egg [yolk] or incorporated into traditionally prepared buttermilk drink). METHODS: Participants (N = 97) took part in a 12-wk intervention study. The controls (n = 20) continued their regular egg consumption of one to two eggs a week. The other two groups consumed either one extra egg per day (n = 57) or a buttermilk drink containing one egg yolk (n = 20). Blood was sampled at day 1 and at the end of the experimental period (day 90) to analyze serum lipids, lipoproteins, and markers reflecting cholesterol metabolism, low-grade systemic inflammation, endothelial activity, and liver function. RESULTS: Serum TC and LDL-C concentrations increased significantly by respectively 0.63 mmol/L (P < 0.05) and 0.59 mmol/L (P < 0.05) in women consuming one additional egg per day compared with controls. There were no effects on markers for inflammation, endothelial activity, or liver function. The increase in serum TC and LDL-C concentration was no longer significant in women consuming the same egg yolk incorporated in a buttermilk drink (0.33 mmol/L [P = 0.66] and 0.31 mmol/L [P = 0.55], respectively). CONCLUSION: Daily egg consumption for 12 wk increases serum TC and LDL-C concentrations in women but not markers for inflammation, endothelial activity, and liver function. Interestingly, the rise in serum LDL-C concentrations is less pronounced when egg yolk is incorporated into a buttermilk drink, indeed suggesting that fractions in the buttermilk might influence dietary cholesterol absorption

7beta-Hydroxysitosterol crosses the blood-brain barrier more favored than its substrate sitosterol in ApoE-/- mice

In this study, we compare the distribution of intraperitoneally injected sitosterol, 7beta-hydroxysitosterol or vehicle only (control) for 28days in male ApoE-/- mice. Furthermore we examine its impact on surrogate markers of cholesterol biosynthesis and sterol absorption rate in plasma, brain and liver tissues from these animals. Injection of sitosterol revealed a 32.1% (P=0.013) lower plasma total cholesterol compared with control. Cholesterol corrected plasma and absolute brain and liver levels of sitosterol are 4.1-, 1.7-, and 7.2-fold (P<0.001 for all) higher, respectively. This is in accordance with a reduced plasma campesterol to cholesterol ratio (-16.2%; P=0.018) together with a 24.1% (P=0.047) lower concentration of hepatic lathosterol. After injection of 7beta-hydroxysitosterol the concentrations of 7beta-hydroxysitosterol in plasma, brain and liver are 21.0-, 65.8- and 42.7-fold (P<0.001 for all) higher, respectively, compared with control. Injection of 7beta-hydroxysitosterol revealed significantly lower plasma cholesterol corrected cholestanol and campesterol (-44.2%; P=0.001 and -24.5; P=0.004) as well as lower absolute liver cholestanol levels (-31.9%; P<0.001) compared with control. Intraperitoneally injected sitosterol and 7beta-hydroxysitosterol differently influence cholesterol metabolism in plasma and liver. We conclude that the polar 7beta-hydroxysitosterol compound can pass the blood brain barrier with higher efficacy than its substrate, sitosterol. Though present in higher amounts in the brain, both, sitosterol and 7beta-hydroxysitosterol do not influence cholesterol metabolism in the brain as proven by our surrogate markers

The relationships of phytosterols and oxyphytosterols in plasma and aortic valve cusps in patients with severe aortic stenosis.

Phytosterols such as campesterol and sitosterol are susceptible to oxidation by reactive oxygen species. We hypothesize that the plant sterols (PS) campesterol and sitosterol and their 7-oxygenated metabolites (POPs) correlate within and between human plasma and aortic valve cusps tissues. Plasma and tissue concentrations of PS and POPs were analyzed by gas chromatography-mass spectrometry-selected ion monitoring. Prior to analysis valve cusps tissue was mechanically separated from the calcified parts. PS and POP levels per dry cusps tissue weight were significantly higher compared with the concentrations in the calcified part. Against our hypothesis we found that despite the fact that there is a high correlation between plant sterols in and between plasma and valves cusps tissue, as well as a high correlation between plant sterols and oxyphytosterols and oxyphytosterols themselves within the valve cusps tissue, there was hardly any correlation in the amount of oxyphytosterols in plasma and between plasma and valves. Because plasma samples are easily accessible for large scale population based studies, we have to understand in more detail what the analysis of POPs implies in terms of CVD risk for the future

Increased plant sterol deposition in vascular tissue characterizes patients with severe aortic stenosis and concomitant coronary artery disease

The aim of the study was to evaluate the relationship between phytosterols, oxyphytosterols, and other markers of cholesterol metabolism and concomitant coronary artery disease (CAD) in patients with severe aortic stenosis who were scheduled for elective aortic valve replacement. Markers of cholesterol metabolism (plant sterols and cholestanol as markers of cholesterol absorption and lathosterol as an indicator of cholesterol synthesis) and oxyphytosterols were determined in plasma and aortic valve tissue from 104 consecutive patients with severe aortic stenosis (n=68 statin treatment; n=36 no statin treatment) using gas chromatography-flame ionization and mass spectrometry. The extent of CAD was determined by coronary angiography prior to aortic valve replacement. Patients treated with statins were characterized by lower plasma cholesterol, cholestanol, and lathosterol concentrations. However, statin treatment did not affect the sterol concentrations in cardiovascular tissue. The ratio of campesterol-to-cholesterol was increased by 0.46+/-0.34mug/mg (26.0%) in plasma of patients with CAD. The absolute values for the cholesterol absorption markers sitosterol and campesterol were increased by 18.18+/-11.59ng/mg (38.8%) and 11.40+/-8.69ng/mg (30.4%) in the tissues from patients with documented CAD compared to those without concomitant CAD. Campesterol oxides were increased by 0.06+/-0.02ng/mg (17.1%) in the aortic valve cusps and oxidized sitosterol-to-cholesterol ratios were up-regulated by 0.35+/-0.2ng/mg (22.7%) in the plasma of patients with CAD. Of note, neither cholestanol nor the ratio of cholestanol-to-cholesterol was associated with CAD. Patients with concomitant CAD are characterized by increased deposition of plant sterols, but not cholestanol in aortic valve tissue. Moreover, patients with concomitant CAD were characterized by increased oxyphytosterol concentrations in plasma and aortic valve cusps