Plant sterols may induce a Th1 shift in humans. However, whether plant stanols have similar effects as well as the underlying mechanism are unknown. We have now shown that - like sitosterol - sitostanol, which are both 4-desmethylsterols, induce a Th1 shift, when added in vitro at physiological concentrations to human PBMCs. This conclusion was based on a higher IFNgamma production, with no change in the production of IL-4 and IL-10. Alpha-amyrin, a 4.4-dimethylsterol, had comparable effects. Since 4.4-dimethylsterols cannot activate transcription factor LXR, this finding indicates that LXR activation was not involved. Sitosterol and sitostanol did not alter the production of IL-12 and IL-18 in PBMCs as well as in monocyte derived U937 cells, suggesting that plant sterols directly affect T-helper cells, without activating APCs. However, in PBMCs treated with a TLR2 blocker (T2.5), IFNgamma production was completely inhibited, while blocking TLR4 with HTA125 had no such effect. To confirm these findings, PBMCs from TLR2-/- mice were cultured in the presence of sitosterol and sitostanol. In these cells, no Th1 shift was observed. Our results therefore indicate that TLR2 activation is essential to induce a Th1 shift in human PBMCs by plant stanols and plant sterols.

Evidence is accumulating that high serum concentrations of triacylglycerols (TAG) are, like LDL cholesterol, causally related to cardiovascular disease. A recent meta-analysis has indicated that plant stanol ester (PSE) intake not only lowered LDL cholesterol, but also serum TAG concentrations, especially in subjects with high baseline TAG concentrations. We therefore evaluated the effects of PSE supplementation on lipid metabolism in a population with elevated fasting TAG concentrations. In a randomized, placebo-controlled, parallel study, 28 subjects with elevated TAG concentrations (>1.7 mmol/L) were studied. After a 1-week run-in period during which a control margarine was used, subjects consumed for 3 weeks either control or PSE-enriched margarine (2.5 g/day of plant stanols). Serum plant stanol concentrations increased in all subjects receiving the PSE-enriched margarines, demonstrating good compliance. PSE supplementation significantly decreased serum total (6.7%, P = 0.015) and LDL cholesterol (9.5%, P = 0.041). A significant interaction between baseline TAG concentrations and PSE intake was found; PSE intake lowered TAG concentrations, particularly in subjects with high baseline TAG concentrations (>2.3 mmol/L; P = 0.009). Additionally, a significant interaction between baseline total number of LDL particles (LDL-P) and PSE intake was found (P = 0.020). PSE consumption lowered LDL-P, primarily in subjects with elevated baseline values; this was mainly due to a non-significant decrease in the number of atherogenic small LDL-P. Circulating levels of hs-CRP, glucose, and insulin were not changed after PSE intake. Taken together, PSE supplementation not only lowered LDL cholesterol, but also serum TAG concentrations, especially in subjects with overt hypertriglyceridemia.

Mushrooms are known for their immune-modulating and anti-tumour properties. The polysaccharide fraction, mainly beta-glucans, is responsible for the immune-modulating effects. Fungal beta-glucans have been shown to activate leukocytes, which depend on structural characteristics of beta-glucans. As edible mushrooms come in contact with the intestinal immune system, effects on enterocytes are also interesting. Our aim was to evaluate the effect of mushroom polysaccharide extracts varying in beta-glucan structure on nitric oxide production by bone marrow-derived macrophages (BMMs) from mice and on nuclear factor-kappaB transactivation in human intestinal Caco-2 cells. We demonstrated that extracts from Agaricus bisporus stimulated nitric oxide production by BMM, whereas extracts from Coprinus comatus and spores of Ganoderma lucidum had only minor effects. Furthermore, extracts of A. blazei Murill and Phellinus linteus had no effect at all. Almost all mushroom extracts lowered nuclear factor-kappaB transactivation in Caco-2 cells. Structural analysis of A. bisporus compared with A. blazei Murill suggests that branching of the beta-glucan chain is essential for immune-stimulating activity. In conclusion, extracts from A. bisporus activate BMM, without activating enterocytes. These characteristics make A. bisporus an attractive candidate as a nutritional compound to stimulate the immune response in depressed states of immunity.

Plant stanol esters not only lower low density lipoprotein cholesterol but also have previously been shown to lower serum triacylglycerol (TAG) concentrations, especially in subjects with elevated TAG concentrations. To find a possible explanation, we explored changes in serum lipoprotein profiles, as measured with nuclear magnetic resonance. For this, serum samples from two parallel-designed controlled studies were evaluated before and 8 weeks after the consumption of plant stanol esters. In the first study, dyslipidemic metabolic syndrome subjects participated and in the second study normolipidemic subjects. In metabolic syndrome subjects, plant stanol esters lowered concentrations of large (>60 nm) and medium (35-60 nm) VLDL particles as compared to controls. In normolipidemic subjects, the serum concentration of large VLDL-1 particles was also lowered, although less pronounced. Based on these findings, we hypothesize that the effect of plant stanol esters on serum TAG concentrations origins from a lowered hepatic production of large TAG-rich VLDL-1 particles.

BACKGROUND: Enterocytes are exposed to antigens present in the intestinal lumen, like beta-glucans that are carbohydrate structures present not only in the cell wall of yeast and fungi but also in cereals. Beta-glucans are known for their immune modulating properties and we have earlier reported an increased immune response by enterocytes after addition of fecal water prepared from ileostomic contents obtained from participants consuming an oat beta-glucan diet versus a placebo diet. We hypothesized that our observation of immune stimulating effects by oat beta-glucan in enterocytes was mediated through the beta-glucan receptor dectin-1. METHODS: Presence of dectin-1 in enterocytes was examined by reverse transcriptase PCR, western blot, and flow cytometry followed by an evaluation of the functional involvement of dectin-1 by using dectin-1 inhibitors during fecal water incubations. RESULTS: Reverse transcriptase PCR and western blot analysis showed dectin-1 presence in the INT407 and Caco-2 NF-kappaB reporter enterocyte cell lines. Moreover, human enterocytes isolated from ileum or colon biopsies also contained dectin-1 protein. However, dectin-1 expression could not be confirmed by flow cytometry in INT407 cells, suggesting that in these cell lines dectin-1 is not expressed at the extracellular membrane. Furthermore, dectin-1 inhibitors did not suppress the beta-glucan containing fecal water-induced IL-8 production by INT407 cells and NF-kappaB transactivation by Caco-2 NF-kappaB reporter cells. CONCLUSION: INT407 and Caco-2 NF-kappaB reporter cells seem to express no functional dectin-1. The absence of this pattern recognition receptor may function to protect the intestine against inflammatory damage, as the dectin-1 ligand beta-glucan is largely present in the intestinal lumen.

Recent animal and human studies have shown that plant sterols and stanols, which are used as functional food ingredients to lower increased LDL cholesterol concentrations, pass the blood-brain barrier. Whether this affects neurocognitive functioning and mental well-being in humans has, to our knowledge, never been investigated. The aim of the present study was therefore to examine the effects of long-term plant sterol or stanol consumption on neurocognitive functioning and mood in a randomized, double-blind, placebo-controlled dietary intervention trial. To this end, hypercholesterolemic individuals, aged 43-69 y, receiving stable statin treatment were randomly assigned to an 85-wk supplementation with margarines enriched with plant sterol esters (2.5 g/d), plant stanol esters (2.5 g/d), or placebo. At baseline and at the end of the intervention period, all participants underwent a cognitive assessment. In addition, subjective cognitive functioning and mood were assessed by means of questionnaires (Cognitive Failure Questionnaire and depression subscale of the Symptom Checklist 90, respectively). Long-term supplementation with plant sterol or stanol esters did not affect cognitive performance (memory, simple information processing speed, complex information processing speed, Letter-Digit Substitution test performance), subjective cognitive functioning, or mood. In conclusion, the present results indicate that long-term use of plant sterols or stanols at recommended intakes of 2.5 g/d does not affect neurocognitive functioning or mood in hypercholesterolemic individuals receiving statin treatment.

BACKGROUND: Increasing HDL cholesterol concentrations by stimulating de-novo apolipoprotein A-I (apoA-I) production in the liver and/or in the small intestine is a potential strategy to reduce coronary heart disease risk. Although there is quite some knowledge concerning regulatory effects in the liver, less is known concerning potential agents that could elevate de-novo apoA-I production in the small intestine. METHODS: Therefore, we compared side-by-side effects of various peroxisome proliferator-activated receptor (PPAR)alpha, PPARgamma, retinoid-X-receptor alpha, and farnesoid-X-receptor agonists on de-novo apoA-I production in differentiated CaCo-2 and HepG2 cells. RESULTS: For PPARa agonists, we showed that GW7647 elevated apoA-I concentrations in the medium of both cell models, whereas WY14643 elevated only de-novo apoA-I concentrations in differentiated CaCo-2 cells. Unexpectedly, fenofibric acid lowered apoA-I medium concentrations in both cell lines, which could not be explained by a lack of PPAR transactivation or a lack of retinoid-X-receptor a activation. For farnesoid-X-receptor agonists, chenodeoxycholic acid strongly reduced apoA-I concentrations both in differentiated CaCo-2 and HepG2 cells, whereas GW4064 and taurocholate only lowered apoA-I in CaCo-2 cells (GW4064) or in HepG2 cells (taurocholate). However, overall effects of all individual components on apoA-I production in differentiated CaCo-2 and HepG2 cells were highly correlated (r = 0.68; P = 0.037; N=9). CONCLUSION: We conclude that differentiated CaCo-2 cells are suitable models to study de-novo small intestinal apoA-I production in vitro enabling the possibility to screen for potential bioactive dietary components. This cell model may also determine small-intestinal-specific effects, as some discrepancy was found between both cell models.

We evaluated the effects of 2 commonly available strategies (plant stanol ester drink and 10 mg simvastatin) on coronary heart disease (CHD) risk variables in participants with metabolic syndrome. Metabolic syndrome patients are at increased risk to develop CHD, partly due to high triacylglycerol (TAG) and low HDL cholesterol (HDL-C) concentrations and a low-grade inflammatory profile. Effects of plant stanol esters on TAG concentrations in these participants are unknown. After a 3-wk run-in period in which individuals consumed placebo yogurt drinks and placebo capsules, participants were randomly divided into 4 groups: placebo (n = 9), simvastatin + placebo drink (n = 10), placebo + stanol drink (n = 9), and simvastatin + stanol drink (n = 8). After 9 wk, we evaluated the effects on serum lipids, low-grade inflammation, and endothelial dysfunction markers. In metabolic syndrome patients, stanol esters (2.0 g/d), simvastatin, or the combination lowered non-HDL-C by 12.8%(P = 0.011), 30.7%(P < 0.001), and 35.4%(P < 0.001), respectively, compared with placebo. TAG were lowered by 27.5%(P = 0.044), 21.7%(P = 0.034), and 32.7%(P < 0.01), respectively. The total-:HDL-C ratio was significantly lowered in all 3 intervention groups. We found no treatment effects on the apolipoprotein CII:CIII ratio, cholesterol ester transfer protein mass, FFA concentrations, and markers for low-grade inflammation or endothelial dysfunction. This study shows that in metabolic syndrome patients, plant stanol esters lower not only non-HDL-C, but also TAG. Effects on TAG were also present in combination with statin treatment, illustrating an additional benefit of stanol esters in this CHD risk population.

ABSTRACT: Background Rosiglitazone not only improves insulin-sensitivity, but also exerts anti-inflammatory effects. We have now examined in type 2 diabetic patients if these effects are reflected by changes in mRNA expression in peripheral blood mononuclear cells (PBMCs) to see if these cells can be used to study these anti-inflammatory effects at the molecular level in vivo. Method Eleven obese type 2 diabetic patients received rosiglitazone (2x4 mg/d) for 8 weeks. Fasting blood samples were obtained before and after treatment. Ten obese control subjects served as reference group. The expression of NFkappaB-related genes and PPARgamma target genes in PBMCs, plasma TNFalpha, IL6, MCP1 and hsCRP concentrations were measured. In addition, blood samples were obtained after a hyperinsulinemic-euglycemic clamp. Results Rosiglitazone reduced plasma MCP1 and hsCRP concentrations in diabetic patients (-9.5 +/- 5.3 pg/mL, p=0.043 and -1.1 +/- 0.3 mg/L p=0.003), respectively). For hsCRP, the concentration became comparable with the non-diabetic reference group. However, of the 84 NFkappaB-related genes that were measured in PBMCs from type 2 diabetic subjects, only RELA, SLC20A1, INFgamma and IL1R1 changed significantly (p<0.05). In addition, PPARgamma and its target genes (CD36 and LPL) did not change. During the clamp, insulin reduced plasma MCP1 concentration in the diabetic and reference groups (-9.1 +/- 1.8%, p=0.001 and -11.1 +/- 4.1%, p=0.023, respectively) and increased IL6 concentration in the reference group only (23.5 +/- 9.0%, p=0.028). Conclusions In type 2 diabetic patients, the anti-inflammatory effect of rosiglitazone is not reflected by changes in NFkappaB and PPARgamma target genes in PBMCs in vivo. Furthermore, our results do not support that high insulin concentrations contribute to the pro-inflammatory profile in type 2 diabetic patients.

Observational epidemiological studies have shown that low carotenoid intake and/or low carotenoid blood levels increase the risk of degenerative diseases like age-related macular degeneration. Functional foods enriched with plant sterol or stanol esters may lower serum concentrations of fat-soluble carotenoids. Theoretically, as a result the macular pigment optical density (MPOD), a marker for eye health, may change. We carried out a double-blind placebo-controlled human intervention trial with a duration of 18 months to evaluate the possible effects of plant stanol and sterol esters on serum lutein/zeaxanthin concentration in relation to the MPOD. Forty-seven subjects were randomly assigned to one of the three treatment groups: margarine without added plant sterols or stanols, plant sterol-enriched margarine, or plant stanol-enriched margarine. Serum cholesterol and lutein/zeaxanthine concentrations and the MPOD were evaluated at baseline and at study end. Changes in lipid-adjusted serum lutein/zeaxanthine concentrations between baseline and study end differed significantly between the three groups (P = 0.001). We found no differences in the MPOD between the three treatment groups, despite the differences in both absolute and cholesterol-standardized serum lutein/zeaxanthine concentrations. This shows that the observed reduction in serum carotenoid concentrations during 18 months consumption of these functional foods does not affect MPOD.