Phytosterols: Potential Metabolic Modulators in Neurodegenerative Diseases.

Phytosterols constitute a class of natural products that are an important component of diet and have vast applications in foods, cosmetics, and herbal medicines. With many and diverse isolated structures in nature, they exhibit a broad range of biological and pharmacological activities. Among over 200 types of phytosterols, stigmasterol and ß-sitosterol were ubiquitous in many plant species, exhibiting important aspects of activities related to neurodegenerative diseases. Hence, this mini-review presented an overview of the reported studies on selected phytosterols related to neurodegenerative diseases. It covered the major phytosterols based on biosynthetic considerations, including other phytosterols with significant in vitro and in vivo biological activities.

A network-based pharmacology study of active compounds and targets of Fritillaria thunbergii against influenza.

Seasonal and pandemic influenza infections are serious threats to public health and the global economy. Since antigenic drift reduces the effectiveness of conventional therapies against the virus, herbal medicine has been proposed as an alternative. Fritillaria thunbergii (FT) have been traditionally used to treat airway inflammatory diseases such as coughs, bronchitis, pneumonia, and fever-based illnesses. Herein, we used a network pharmacology-based strategy to predict potential compounds from Fritillaria thunbergii (FT), target genes, and cellular pathways to better combat influenza and influenza-associated diseases. We identified five compounds, and 47 target genes using a compound-target network (C-T). Two compounds (beta-sitosterol and pelargonidin) and nine target genes (BCL2, CASP3, HSP90AA1, ICAM1, JUN, NOS2, PPARG, PTGS1, PTGS2) were identified using a compound-influenza disease target network (C-D). Protein-protein interaction (PPI) network was constructed and we identified eight proteins from nine target genes formed a network. The compound-disease-pathway network (C-D-P) revealed three classes of pathways linked to influenza: cancer, viral diseases, and inflammation. Taken together, our systems biology data from C-T, C-D, PPI and C-D-P networks predicted potent compounds from FT and new therapeutic targets and pathways involved in influenza.

An ultra high performance liquid chromatography with tandem mass spectrometry method for simultaneous determination of thirteen components extracted from Radix Puerariae in rat plasma and tissues: Application to pharmacokinetic and tissue distribution study.

A rapid and sensitive ultra high performance liquid chromatography with tandem mass spectrometry method was established and validated for simultaneous determination of thirteen bioactive components (gallic acid, protocatechuic acid, puerarin, p-hydroxycinnamic acid, daidzin, ononin, daidzein, naringenin, genistein, apigenin, formononetin, biochanin A, and ß-sitosterol) of Radix Puerariae extract in rat plasma and tissues. The plasma and tissues samples were pretreated by protein precipitation extraction, and umbelliferone and rutin were used as internal standards. Sample separation was performed on a ZORBAX RRHD Eclipse plus C18 column (2.1 mm × 50 mm, 1.8 µm, Agilent) with a mobile phase consisting of methanol-water (containing 0.1% formic acid). The mass spectrometry analysis was conducted in positive and negative ionization modes with multiple reaction monitoring. The lower limit of quantitation range for the 13 analytes was 0.2-35 ng/mL. The intra- and inter-day precision of all the analytes were less than 10.92%, with an accuracy ranging from -13.10 to 11.96%. Both the recovery and matrix effect were within acceptable limits. This method was successfully applied to pharmacokinetic and tissue distribution study of the 13 bioactive components in rats after oral administration of R. Puerariae extract.

Phytosterol containing diet increases plasma and whole body concentration of phytosterols in apoE-KO but not in LDLR-KO mice.

Phytosterol metabolism is unknown in the hypercholesterolemia of genetic origin. We investigated the metabolism of phytosterols in a cholesterol-free, phytosterol-containing standard diet in hypercholesterolemic mice knockouts for low density lipoprotein receptor (LDLR) and apolipoprotein E (apoE) mice compared to wild-type mice (controls). Phytosterols were measured in mice tissues by GCMS. ApoE-KO mice absorbed less phytosterols than LDLR-KO and the latter absorbed less phytosterols than control mice, because the intestinal campesterol content was low in both KO mice, and sitosterol was low in the intestine in apoE-KO mice as compared to LDLR-KO mice. Although the diet contained nine times more sitosterol than campesterol, the concentration of sitosterol was lower than that of campesterol in plasma in LDLR-KO, and in the liver in controls and in LDLR-KO, but only in apoE-KO. On the other hand, in the intestine sitosterol was higher than campesterol in controls, and in LDLR-KO but with a tendency only in apoE-KO. Because of the high dietary supply of sitosterol, sitosterol was better taken up by the intestine than campesterol, but the amount of sitosterol was lower than that of campesterol in the liver, while in the whole body the amounts of these phytosterols do not differ from each other. Therefore, via intestinal lymph less sitosterol than campesterol was transferred to the body. However, as compared to controls, in apoE-KO mice, but not in LDLR-KO mice, the increase in campesterol and sitosterol in plasma and in the whole body indicating that apoE-KO mice have a marked defect in the elimination of both phytosterols from the body.

Sitosterol prevents obesity-related chronic inflammation.

The physiological roles of phytosterols in chronic inflammation, which are believed to be involved in the underlying mechanisms for metabolic diseases, have yet to be elucidated. Therefore, in the present study, we aimed to elucidate the physiological roles of phytosterols in both clinical studies and animal experiments. We observed the existence of rather specific negative correlations between the serum sitosterol level and the serum IL-6 and the TNF-α levels in both diabetic subjects (n=46) and non-diabetic subjects (n=178). Multiple regression analyses also revealed that the serum IL-6 and TNF-α levels exhibited strong negative correlations with the serum sitosterol levels. When ABCG5/8 KO mice with markedly elevated plasma sitosterol levels and ABCG5/8 hetero mice were fed a high-fat diet, we observed that the increase in body weight, the fatty liver changes, and the expansion of perigonadal adipose tissues were suppressed in ABCG5/8 KO mice without any modulation of food intake. We also observed that the plasma IL-6 and TNF-α levels, the expressions of TNF-α and PAI-1 in the liver and the expressions of the IL-6, TNF-α, and MCP-1 levels in the adipose tissue were lower in ABCG5/8 KO mice. These results suggest that sitosterol might suppress obesity-related chronic inflammation and might be applicable to the treatment of metabolic diseases.

Synthesis and characterization of redox-sensitive heparin-ß-sitosterol micelles: Their application as carriers for the pharmaceutical agent, doxorubicin, and investigation of their antimetastatic activities in vitro.

Although there are several clinical attempts to treat tumors at the primary site, only few therapies can inhibit the spread of metastatic cancers. In this study, we synthesized redox-sensitive heparin-ß-sitosterol micelles that show antimetastatic activity. Proton nuclear magnetic resonance and Fourier transform infrared analyses confirmed the formation of bioreducible heparin-ß-sitosterol (bHSC) conjugates, whereas dynamic light scattering was used to measure the particle size and zeta potential. Both 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and flow cytometry assays confirmed the low toxicity of the synthesized micelles. Doxorubicin (Dox) was encapsulated via the dialysis method, and its loading and encapsulation efficiencies were 16.49±1.2% and 58.47±1.87%, respectively. An in vitro release study showed that approximately 89% and 52% of Dox were released after 48h in the presence and absence of reduced glutathione, respectively. The hemocompatibility and antimetastatic effects of bHSC were evaluated using the hemolysis and scratch assays, respectively. F-Actin fluorescence microscopy showed that heparin- and bHSC-treated HeLa cells had poorly oriented stress fibers. In summary, the synthesized bHSC micelles are good candidates as drug delivery systems owing to their low toxicity, excellent hemocompatibility, and antimetastatic effects.

Lipid nanovehicles with adjustable surface properties for overcoming multiple barriers simultaneously in oral administration.

Lipid nanoparticles (LNs) are widely investigated for oral drug delivery, and for achieving significant advantages in colloidal stability, biocompatibility and scaled-up possibility. However, researchers face challenge of developing methods to improve the ability of LNs in overcoming multiple barriers (i.e., mucus and epithelium barrier) in gastrointestinal (GI) tract because of the contradictory requirement of nanoparticle (NP) surface properties in the two processes. Therefore, we designed novel LNs with adjustable surface properties by coating lipid core with hydrophobic substitutes grafting N-(2-hydroxypropyl) methacrylamide copolymer (pHPMA). In the present study, different substitutes (i.e., monocyclic, polycyclic, and linear segments) were grafted on pHPMA backbone. Screening studies demonstrated that type and grafting degree of substitutes both influenced hydrophilic-hydrophobic properties of NP surface and improved penetration through mucus. When a hydrophilic-hydrophobic balance was achieved, NPs showed elevated mucus permeability compared with bare LNs; this phenomenon subsequently contributed to higher cellular uptake. Moreover, ß-sitosterol (SITO)-modified pHPMA-coated (grafting degree: 5%) LNs (5% SITO-LNs) exhibited the highest mucus permeability, transepithelial transport, and in situ absorption. Interestingly, even with the highest surface hydrophilicity, 5% SITO-LNs with Caco-2 cells did not show impaired membrane affinity, which was not observed in other groups. Further investigations of mechanism demonstrated that membrane affinity was significantly enhanced by ß-SITO-mediated interaction with Niemann-Pick C1-like 1 (NPC1L1) protein on cell membranes. These results proved that hydrophobic substitutes play a critical role in altering hydrophilic-hydrophobic property of particle surface and improving penetration through multiple barriers. ß-SITO-induced specific interaction can provide additional benefits to efficiency of oral delivery of LNs.

Simultaneous determination of ß-sitosterol, campesterol, and stigmasterol in rat plasma by using LC-APCI-MS/MS: Application in a pharmacokinetic study of a titrated extract of the unsaponifiable fraction of Zea mays L.

A liquid chromatography with atmospheric pressure chemical ionization tandem mass spectrometry method was developed and validated to investigate the pharmacokinetic properties of ß-sitosterol, campesterol, and stigmasterol in rat plasma. Cholesterol-d6 was used as an internal standard. To avoid interference of the three phytosterols in rat plasma and minimize matrix effects, a small volume (10 µL) of 4% bovine serum albumin was used as a surrogate matrix for making calibrators and quality control samples. Rat plasma (10 µL) samples were extracted by liquid-liquid extraction with methyl tert-butyl ether and separated on a Kinetex C18 column. The detection was performed on a triple quadrupole tandem mass spectrometer in selected reaction monitoring mode using positive atmospheric pressure chemical ionization. This assay was linear over concentration ranges of 250-5000 ng/mL (ß-sitosterol), 250-5000 ng/mL (campesterol), and 50-2000 ng/mL (stigmasterol). Additionally, a second set of quality controls made in rat plasma was also evaluated against calibration curves made using the surrogate matrix. All the validation data, including the specificity, precision, accuracy, recovery, matrix effect, stability, and incurred sample reanalysis conformed to the acceptance requirements. Our method was successfully applied to study the pharmacokinetics of three phytosterols in rats.

Approaches to intravenous clinical pharmacokinetics: Recent developments with isotopic microtracers.

Obtaining pharmacokinetic data from the intravenous route for drugs intended for oral administration has traditionally been expensive and time consuming because of the toxicology requirements and challenges in intravenous formulations. Such studies are necessary, however, particularly when regulator agencies request absolute bioavailability data. A method has emerged whereby the drug administered intravenously is isotopically labeled and dosed at a maximum of 100 µg concomitantly with an oral administration given at a therapeutically relevant level. The intravenous administration has been termed a microtracer and obviates intravenous toxicology requirements as well as simplifying formulations. The study design also essentially removes issues of nonlinear pharmacokinetics that may occur when oral and intravenous doses are administered separately. This review examines the methodology and the literature to date, including those studies intended for regulatory submission. The method has been extended to the study of prodrug-to-active drug kinetics and to obtaining clearance, volume of distribution, and absolute bioavailability at steady-state conditions.

Acute intake of plant stanol esters induces changes in lipid and lipoprotein metabolism-related gene expression in the liver and intestines of mice.

The kinetics of plant stanol uptake and routing in 8-week-old C57BL/6J mice were determined after a plant stanol ester gavage. In addition, acute changes in intestinal and hepatic gene expression were investigated. Mice were fed a plant sterol/stanol poor diet from weaning. At the age of 8 weeks, they received an oral gavage consisting of 0.25 mg cholesterol + 50 mg plant stanol esters dissolved in olive oil. Animals were euthanized at different time points. In a second comparable set-up, mesenteric lymph-cannulated versus sham-operated mice received the same oral gavage, which was now deuterium labeled. Intestinal and hepatic sitostanol concentrations increased within 15 min post-gavage. This rapid hepatic appearance was absent in lymph-cannulated mice, suggesting a very fast lymph-mediated uptake. Hepatic mRNA expression of SREBP2 and its target genes rapidly decreased, whereas expression of LXR target genes increased. The intestinal SREBP2 pathway was increased, whereas the expression of LXR target genes hardly changed. The fivefold and sixfold increased expression of intestinal LDLr and PCSK9 is suggestive of TICE activation. We conclude that in C57BL/6J mice plant stanol kinetics are fast, and affect intestinal and hepatic gene expression within 15 min postprandial after lymph-mediated uptake.

Lecithin-linker formulations for self-emulsifying delivery of nutraceuticals.

Lecithin-linker microemulsions are formulations produced with soybean lecithin in combination with a highly lipophilic (lipophilic linker) and highly hydrophilic (hydrophilic linkers) surfactant-like additives. In this work, lecithin-linker systems were formulated to produce self-emulsifying delivery systems for ß-carotene and ß-sitosterol. The concentration of the lipophilic linker, sorbitan monooleate, was adjusted to minimize the formation of liquid crystals. The concentration of hydrophilic linkers, decaglyceryl caprylate/caprate and PEG-6-caprylic/capric glycerides, was gradually increased (scanned) until single phase clear microemulsions were obtained. For these scans, the oil (ethyl caprate) to water ratio was set to 1. The single phase, clear microemulsions were diluted with fed-state simulated intestinal fluid (FeSSIF) and produced stable emulsions, with drop sizes close to 200 nm. Using pseudo-ternary phase diagrams to evaluate the process of dilution of microemulsion preconcentrates (mixtures of oil, lecithin and linkers with little or no water) with FeSSIF, it was determined that self-emulsifying systems are obtained when the early stages of the dilution produce single phase microemulsions. If liquid crystals or multiple phase systems are obtained during those early stages, then the emulsification yields unstable emulsions with large drop sizes. An in vitro permeability study conducted using a Flow-Thru Dialyzer revealed that stable emulsions with drop sizes of 150-300 nm produce large and irreversible permeation of ß-carotene to sheep intestine. On the other hand, unstable emulsions produced without the linker combination separated in the dialyzer chamber.

Impaired hepatic and intestinal ATP-binding cassette transporter G5/8 was associated with high exposure of ß-sitosterol and the potential risks to blood-brain barrier integrity in diabetic rats.

OBJECTIVES: Plant sterols are thought to treat hypercholesterolemia via inhibiting intestinal cholesterol absorption. The aim of this study was to evaluate the contribution of impaired ATP-binding cassette transporter G5/8 (ABCG5/8) expression by diabetes to the increased ß-sitosterol (BS) exposure and impact of increased BS on integrity of blood-brain barrier (BBB). METHODS: Basal BS level in tissues of streptozotocin-inducted rats and ABCG5/8 protein levels in liver and intestine were investigated; pharmacokinetics of BS was studied following oral dose; and primarily cultured rat brain microvessel endothelial cells (rBMECs) were used to study BS transportation across BBB and effect of BS on BBB integrity. KEY FINDINGS: Diabetic rats showed greatly upgraded basal levels of BS in plasma, intestine, cerebral and hippocampus, accompanied by impairment of ABCG5/8 protein expression in liver and intestine. Pharmacokinetics studies demonstrated higher AUC0-48 and Cmax , and lower faecal recoveries of BS after oral administration, indicating enhancement of absorption or efflux impairment. In-vitro data showed increased ratio of BS/cholesterol in high levels BS-treated rBMECs was associated with increased BBB permeability of some biomarkers including BS itself. CONCLUSIONS: Impaired ABCG5/8 protein expression by diabetes led to increase in BS exposure, which may be harmful to BBB function.

Incorporation of ß-sitosterol into mitochondrial membrane enhances mitochondrial function by promoting inner mitochondrial membrane fluidity.

Recent findings suggest that mitochondrial membrane fluidity could influence mitochondrial energy metabolism. ß-sitosterol (BS) is a common plant sterol that is prevalent in plant oils, nuts, cereals and plant food products. Its chemical structure is very similar to that of cholesterol. As a cholesterol analog, BS is highly lipid soluble and largely resides in the membranes of cells or organelles where it may have an influence on the membrane fluidity. The present study reports that, with the cholesterol chelator 2-hydroxypropyl-ß-cyclodextrin (HPßCD) as its carrier, BS is able to increase the fluidity of the inner mitochondrial membrane (IMM) without affecting the fluidity of the outer mitochondrial membrane (OMM), and consequently to increase the mitochondrial membrane potential (∆Ψm) and mitochondrial ATP content. It has been previously proposed that a therapeutical boost in adenosine triphosphate (ATP) levels in mitochondria may be beneficial for neurodegenerative diseases such as Alzheimer's disease (AD). Given that dietary administration of plant sterols could increase brain BS concentrations, these results may provide a better understanding of the beneficial effects of plant sterol-enriched nutrients on neurodegenerative diseases such as AD.

Absolute oral bioavailability and metabolic turnover of ß-sitosterol in healthy subjects.

The metabolic turnover, absolute oral bioavailability, clearance, and volume of distribution for ß-sitosterol were measured in healthy subjects. [(14)C]ß-Sitosterol was used as an isotopic tracer to distinguish pulse doses from dietary sources and was administered by both oral and intravenous routes. The administered doses of [(14)C]ß-sitosterol were in the region of 3 to 4 µg, sufficiently low as not to perturb the kinetics of ß-sitosterol derived from the diet. Because the plasma concentrations of [(14)C]ß-sitosterol arising from such low doses were anticipated to be very low, the ultrasensitive isotope ratio analytical method of accelerator mass spectrometry was used. The limit of quantification for [(14)C]ß-sitosterol was approximately 0.1 pg/ml, the oral absolute bioavailability was just 0.41%, clearance was 85 ml/h, volume of distribution was 46 L, and the turnover was 5.8 mg/day. Given the steady-state concentrations of ß-sitosterol (2.83 µg/ml), then the dietary load was calculated to be approximately 1400 mg/day.

Development and characterization of phyto-vesicles of ß-sitosterol for the treatment of androgenetic alopecia.

Alopecia is a psychologically distressing phenomenon. Androgenetic alopecia (AGA) is the most common form of alopecia, which affects millions of men and women worldwide, and is an androgen driven disorder. To study the effect of ß-sitosterol phyto-vesicles on AGA, the testosterone-induced alopecia model was used. For the study, the albino rats were used and the period of study was 21 days. ß-Sitosterol is a phytosterol which is chemically similar to cholesterol. This compound was found suitable for the preparation of phyto-vesicles by the process involving its complexation with phosphatidyl choline. Pharmacokinetic studies of ß-sitosterol reveal its poor absorption through the intestine. The objective of the present study is to enhance the bioavailability of ß-sitosterol by its complexation with phosphatidyl choline and then to formulate it as phyto-vesicles for the treatment of alopecia. The complex of ß-sitosterol was prepared with phosphatidyl choline and characterized on the basis of solubility, melting point, TLC, UV, IR and NMR spectroscopy. This complex was then formulated as phyto-vesicles and then characterized. The results revealed that effect on alopecia is better in case of phyto-vesicles as compared to the complex, physical mixture and the ß-sitosterol itself. Enhanced bioavailability of the ß-sitosterol complex may be due to the amphiphilic nature of the complex, which greatly enhance the water and lipid solubility of the compound. The present study clearly indicates the superiority of phyto-vesicles over the complex and ß-sitosterol, in terms of better absorption and improved activity for the treatment of alopecia.

ß-sitosterol inhibits high cholesterol-induced platelet ß-amyloid release.

Recently, increasing evidence has linked high cholesterol to the pathogenesis of Alzheimer's disease (AD), suggesting that cholesterol may be a target for developing new compounds to prevent or treat AD. Plant sterols, a group of sterols enriched in plant oils, nuts, and avocados, have the structure very similar to that of cholesterol, and have been widely used to reduce blood cholesterol. Due to their cholesterol-lowering property, plant sterols such as ß-sitosterol may also influence cholesterol-depending functions including its role in AD development. Using human platelets, a type of peripheral blood cells containing the most circulating amyloid precursor protein (APP), this study investigated the effect of ß-sitosterol on high cholesterol-induced secretion of ß amyloid protein (Aß). It was found that ß-sitosterol effectively inhibited high cholesterol-driven platelet Aß release. In addition, ß-sitosterol prevented high cholesterol-induced increase of activities of ß- and γ-secretase, two APP cleaving enzymes to generate Aß. Additional experiments showed that high cholesterol up-regulated lipid raft cholesterol. This effect of cholesterol could be suppressed by ß-sitosterol. These findings suggest that ß-sitosterol is able to inhibit high cholesterol-induced Aß release probably through maintenance of membrane cholesterol homeostasis. Given that dietary plant sterols have the potential of penetrating the blood-brain barrier (BBB), these data suggest that plant sterols such as ß-sitosterol may be useful in AD prevention.

Action of plant sterol intervention on sterol kinetics in hypercholesterolemic men with high versus low basal circulatory plant sterol concentrations.

BACKGROUND: The relationship between plant sterol (PS) absorption and circulatory concentrations with cholesterol absorption and biosynthesis during PS consumption has yet to be clearly elucidated in humans. It is therefore essential to examine campesterol, ß-sitosterol, and cholesterol absorption and cholesterol fractional synthesis rate (FSR) following PS consumption in individuals with high versus low basal circulatory PS concentrations. DESIGN: A randomized, crossover trial was conducted in 82 hypercholesterolemic men consuming spreads with or without 2 g/d of PS for two 4-week periods, each separated by a 4-week washout. Endpoint tracer enrichments after ingestion of (2)H-labeled campesterol or ß-sitosterol and (13)C-labeled cholesterol were determined by isotope ratio mass spectrometry. RESULTS: For both phases of dietary intervention, the endpoint cholesterol absorption index was positively correlated with campesterol (r = 0.5864, p < 0.0001) and ß-sitosterol (r = 0.4676, p < 0.0001) absorption indices; inversely, endpoint cholesterol FSR correlated negatively with the absorption indices of campesterol (r = -0.5004, p < 0.0009), ß-sitosterol (r = -0.4154, p < 0.05), and cholesterol (r = -0.4056, p < 0.0001). PS intervention reduced absorption indices of campesterol, ß-sitosterol, and cholesterol by 36.5% ± 2.7%, 39.3% ± 2.9%, and 34.3% ± 1.9%, respectively, but increased cholesterol FSR by 33.0% ± 3.3% relative to control. Endpoint circulatory PS levels (cholesterol adjusted) were positively associated with endpoint absorption indices of campesterol (r = 0.5586, p < 0.0001, for placebo; r = 0.6530, p < 0.0001, for PS intake) and cholesterol (r = 0.3683, p < 0.001 for placebo; r = 0.3469, p < 0.002, for PS intake) and were negatively associated with cholesterol FSR (r = -0.3551, p < 0.002, for placebo; r = -0.3643, p < 0.001, for PS intake). The cholesterol-lowering effect of PS was most pronounced among individuals falling within the 50th-75th percentiles of basal PS concentrations. CONCLUSION: These data suggest that basal PS concentrations indicate not only sterol absorption efficiency but also the extent of PS-induced cholesterol reduction and thus might be clinically useful to predict the extent of cholesterol response to PS intervention within a given individual.

Therapies targeting exogenous cholesterol uptake: new insights and controversies.

Exogenous cholesterol uptake involves a complex process in the intestines for the absorption of cholesterol and bile acids. This process is regulated by intestinal nuclear transcription factors such as LXR that affect sterol transporters NPC1L1, ABCG5/G8, and ABCG1, and enzymes such as ACAT-2. Plant sterol/stanols, ezetimibe, and bile acid sequestrants have a variety of effects on these various transporters, and new insights into their mechanism(s) of action have provided a plethora of exciting targets for metabolic diseases, dyslipidemia, and atherosclerosis.

Missense mutation of Abcg5 in stroke-prone spontaneously hypertensive rats does not influence lymphatic sitosterol absorption regardless of the dose: comparison with Wistar rats.

The lymphatic recovery of radiolabeled sitosterol administered in various amounts to the stomach was almost the same between stroke-prone spontaneously hypertensive rats (SHRSPs), a strain having a missense mutation in ATP binding cassette transporter g5 (Abcg5), and Wistar rats, a normal strain. The results suggest that the mutation of Abcg5 in SHRSPs, compared with Wistar rats, did not influence the ability for intestinal sitosterol absorption regardless of the dose.

Chemoprevention of tumor metastasis by liposomal beta-sitosterol intake.

To investigate chemopreventive effect of liposomal beta-sitosterol on tumor metastasis, we prepared liposomal beta-sitosterol composed of egg yolk phosphatidylcholine for oral delivery. Although orally administered beta-sitosterol (4 micromol as beta-sitosterol/mouse) was not absorbed into plasma, the amount of immune response cytokines such as IL-12 and IL-18 was increased in the small intestine after the liposome intake. Moreover, after daily oral administration of the liposome for 7 d, natural killer (NK) cell activity in the mice was increased, suggesting that the immune surveillance activity of mice was enhanced by the liposomal beta-sitosterol intake. Thus, we examined metastatic potential of B16BL6 melanoma cells, which were intravenously injected into mice after sequential administration of liposomal beta-sitosterol for 7 d. The number of metastatic colonies in the lungs was significantly less than that of control group two weeks after the injections of the cells. These results suggest that daily liposomal beta-sitosterol intake prevents tumor metastasis may be due to enhancement of gut immune surveillance systems.