Amounts Of Sterols Research Articles

Localization of ABCG5 and ABCG8 proteins in human liver, gall bladder and intestine.

BackgroundThe molecular mechanisms that regulate the entry of dietary sterols into the body and their removal via hepatobiliary secretion are now beginning to be defined. These processes are specifically disrupted in the rare autosomal recessive disease, Sitosterolemia (MIM 210250). Mutations in either, but not both, of two genes ABCG5 or ABCG8, comprising the STSL locus, are now known to cause this disease and their protein products are proposed to function as heterodimers. Under normal circumstances cholesterol, but not non-cholesterol sterols, is preferentially absorbed from the diet. Additionally, any small amounts of non-cholesterol sterols that are absorbed are rapidly taken up by the liver and preferentially excreted into bile. Based upon the defects in sitosterolemia, ABCG5 and ABCG8 serve specifically to exclude non-cholesterol sterol entry at the intestinal level and are involved in sterol excretion at the hepatobiliary level.MethodsHere we report the biochemical and immuno-localization of ABCG5 and ABCG8 in human liver, gallbladder and intestine using cell fractionation and immunohistochemical analyses.ResultsWe raised peptide antibodies against ABCG5 and ABCG8 proteins. Using human liver samples, cell fractionation studies showed both proteins are found in membrane fractions, but they did not co-localize with caveolin-rafts, ER, Golgi or mitochondrial markers. Although their distribution in the sub-fractions was similar, they were not completely contiguous. Immunohistochemical analyses showed that while both proteins were readily detectable in the liver, ABCG5 was found predominately lining canalicular membranes, whereas ABCG8 was found in association with bile duct epithelia. At the cellular level, ABCG5 appeared to be apically expressed, whereas ABCG8 had a more diffuse expression pattern. Both ABCG5 and ABCG8 appeared to localize apically as shown by co-localization with MRP2. The distribution patterns of ABCG5 and ABCG8 in the gallbladder were very similar to each other. In the small intestine both ABCG5 and ABCG8 appear to line the brush border. However, at the level of the enterocyte, the cellular distribution patterns of ABCG5 and ABCG8 differed, such that ABCG5 was more diffuse, but ABCG8 was principally apical. Using standard deglycosylation methods, ABCG5 and ABCG8 do not appear to be glycosylated, suggesting a difference between human and mouse proteins.ConclusionWe report the distribution patterns of ABCG5 and ABCG8 in human tissues. Cell fractionation studies showed that both proteins co-fractionated in general, but could also be found independent of each other. As predicted, they are expressed apically in both intestine and liver, although their intracellular expression patterns are not completely congruent. These studies support the concept of heterodimerization of ABCG5 and ABCG8, but also support the notion that these proteins may have an independent function.

Organic composition of aerosol particulate matter during a haze episode in Kuala Lumpur, Malaysia

The solvent-extractable compounds of urban airborne particulate matter were analyzed to determine the distributions of homologous and biomarker tracers. Samples were collected by high-volume air filtration during the haze episode of 1997 around the University of Malaya campus near Petaling Jaya, a suburb of Kuala Lumpur, Malaysia. These results show that the samples contain n-alkanes, n-alkan-2-ones, n-alkanols, methyl n-alkanoates, n-alkyl nitriles, n-alkanals, n-alkanoic acids, levoglucosan, PAHs, and UCM as the dominant components, with minor amounts of terpenoids, glyceryl esters and sterols, all derived from natural biogenic sources (vascular plant wax), from burning of biomass, and from anthropogenic utilization of fossil fuel products (lubricating oil, vehicle emissions, etc.). Some compositional differences are observed in the samples and greater atmospheric concentrations were found for almost all organic components in the samples collected near a roadway. The results interpreted in terms of major sources are due to local build-up of organic contaminants from vehicular emissions, smoke from biomass burning, and natural background as a result of the atmospheric stability during the haze episodes. The organic components transported in from areas outside the region, assuming all smoke components are external to the city, amount to about 30% of the total organic particle burden.

Mechanisms of azole resistance in petite mutants of Candida glabrata.

We previously showed that resistant colonies of Candida glabrata inside the azole inhibition zones had respiratory deficiency due to mutations in mitochondrial DNA. Here, we analyzed the mechanisms of azole resistance in petite mutants of C. glabrata obtained by exposure to fluconazole or induced by ethidium bromide. The respiratory deficiency of these mutants was confirmed by oxygraphy and flow cytometric analysis with rhodamine 123, and its mitochondrial origin was demonstrated by transmission electron microscopy and restriction endonuclease analysis of the mitochondrial DNA. Flow cytometry with rhodamine 6G suggested an increased drug efflux in mutant cells, which was further supported by Northern blot analysis of the expression of the C. glabrata CDR1 (CgCDR1) and CgCDR2 genes, encoding efflux pumps. Conversely, the expression of CgERG11, which encodes the azole target, was not affected by petite mutations, and no differences were seen in the sequence of this gene between parent isolates and mutants. Moreover, sterol analysis showed similar overall amount of sterols in parent and mutant cells, but quantitative modifications were observed in the mutants, with almost undetectable biosynthesis intermediates. Further analysis performed after separation of free sterols from steryl esters revealed a defect in sterol esterification in mutant cells, with free ergosterol representing 92% of the overall sterol content. Thus, resistance or decreased susceptibility to azoles in petite mutants of C. glabrata is associated with increased expression of CgCDR1 and, to a lesser extent, of CgCDR2. In addition, the marked increase in free ergosterol content would explain their increased susceptibility to polyenes.

Intestinal steroids in rats are influenced by the structural parameters of pectin

We investigated the effects of pectin with different degrees of methylation (34.5, 70.8, and 92.6%, respectively) on the composition and concentration of intestinal and fecal bile acids and neutral sterols in conventional and germfree rats. Diets containing 6.5% pectin (galacturonan) were given for 3 weeks. High concentrations of free and secondary bile acids appeared in cecum and colon of conventional rats. With increasing degree of methylation, more bile acids were transported into lower parts of intestinal tract and excreted whereas the proportion of secondary bile acids decreased. In contrast, the composition of bile acids in intestinal contents and feces was relatively unchanged in germfree rats. Exclusively cholesterol was found as a neutral sterol in germfree rats. Coprostanol appeared in cecum of conventional rats and additionally coprostanone in colon. Amounts of neutral sterols increased with increasing degree of methylation of pectin. Additionally, concentrations of bile acids in plasma decreased if the pectin-containing diets were given. Besides the degree of methylation, the molecular weight of pectin used in the diets influenced concentration and composition of intestinal and fecal steroids in rats.

Stabilization of palm oils by using Ficus exasperata leaves in local processing methods

This work was done to ascertain the efficacy of the local use of Ficus exasperata leaves in processing palm oils to improve and stabilize the quality. Palm oils were obtained from the ripe fruits of Elaeis guineensis and Elaeis guineensis var virescens, including Ficus exasperata leaves in the traditional processing procedures––cooking and soaking the fruits in water. Phytochemical analysis of the Ficus leaves and the oils were performed. The quality of the oils were assessed in terms of saponification, acid and peroxide values, and free fatty acid content. Results showed the presence of phenolic compounds, terpenoids, alcohols and carbonyl compounds in the leaves. Oil from both oil palm varieties contained appreciable amounts of carotenoids, flavonoids and sterols, but only the Elaeis guineensis var virescens oil contained saponins. The use of the Ficus leaves eliminated saponins and the foaming tendencies of the var virescens oil, but reduced the sterol levels and increased the carotenoid levels in the oils from both oil palm varieties. Oils processed by soaking of fruits and use of the Ficus leaves had slightly greater saponification values, while oils processed by cooking of fruit and use of leaves had reduced FFA, acid and peroxide values. The use of Ficus exasperata leaves makes for a better stabilized oil with potentially good keeping qualities.

Sterol dynamics of heterotrophic Tetraselmis suecica and its nutritional implication in the bivalve aquaculture

Employment of heterotrophic culture of microalgal foods in the hatchery-based seed production of oysters is still controversial because some algae produced using the method appear to loose sterols, a key nutritional factor for bivalve growth. We traced the changes in sterol content of Tetraselmis suecica growing under photoautotrophic and heterotrophic conditions with the aid of gas chromatography (GC) and GC–mass spectrometry. The photoautotrophic T. suecica at the mid-logarithmic growth phase contained six major (cholesta-5,22-dien-3β-ol, ergost-5-en-3β-ol, cholest-5-en-3β-ol, 24-methylcholesta-5,22-dien-3β-ol, 24-methylcholesta-5,24-dien-3β-ol, and 24-ethylchlolesta-5,24-dien-3β-ol) and two minor sterols (24-methylcholesta-5-en-3β-ol and 24-ethylchlolesta-5en-3β-ol). In the comparison of algal growth and sterol level, photoautotrophic alga appeared to need higher amounts of major sterols for cell growth over heterotrophic alga. These findings, that heterotrophic alga needed less amount of sterols for growth, may have significant implications in the introduction of the method in bivalve hatcheries. We also reviewed the sterolic properties of the alga obtained from heterotrophic method with respect to bivalve aquaculture.

The Saccharomyces cerevisiae alcohol acetyl transferase Atf1p is localized in lipid particles.

The yeast alcohol acetyl transferase I, Atf1p, is responsible for the major part of volatile acetate ester production in fermenting Saccharomyces cerevisiae cells. Some of these esters, such as ethyl acetate and isoamyl acetate, are important for the fruity flavours of wine, beer and other fermented beverages. In order to reveal the subcellular localization of Atf1p and further unravel the possible physiological role of this protein, ATF1::GFP fusion constructs were overexpressed in brewer's yeast. The transformant strain showed a significant increase in acetate ester formation, similar to that of an ATF1 overexpression strain, indicating that the Atf1p-GFP fusion protein was active. UV fluorescence microscopy revealed that the fusion protein was localized in small, sphere-like organelles. These organelles could be selectively stained by the fluorescent dye Nile red, indicating that they contained high amounts of neutral lipids and/or sterols, a specific characteristic of yeast lipid particles. Purification of lipid particles from wild type and ATF1 deletion cells confirmed that the Atf1p-GFP fusion protein was located in these organelles. Furthermore, a clear alcohol acetyl transferase activity could be measured in the purified lipid particles of both wild type and transformed cells. The localization of Atf1p in lipid particles may indicate that Atf1p has a specific role in the lipid and/or sterol metabolism that takes place in these particles.

FATTY ACIDS, STEROLS, α‐TOCOPHEROL AND TOTAL CAROTENOIDS COMPOSITION OF DIACRONEMA VLKIANUM

ABSTRACT Microalgae are a large group of photosynthetic microorganisms with a great potential as a source of energy and with the ability to convert solar energy into biomass. Several conditions as well as different growing stages were found to play an important role in the biosynthesis of different molecules. These microorganisms are easily cultivated, and even under the most difficult conditions, are able to develop. In the present study the production of different biomolecules, namely sterols, fatty acids, tocopherols and total carotenoids during the life cycle with 24 h of photoperiod and 18C was followed. The major amounts of sterols, fatty acids and total carotenoids were detected in stationary phase. Higher amounts of α‐ tocopherol were present in the decay phase. Tocopherol and total carotenoids levels were higher compared to conventional foods known as rich sources of these components. A significant difference was observed between replicates of D. vlkianum for α‐tocopherol and some sterols.

Characterization of field-isolates and derived DMI-resistant strains of Cercospora beticola

Cercospora beticola strains with laboratory induced resistance to tetraconazole were compared with their parental WT sensitive strains to evaluate the effects of resistance on fitness and assess whether any change in the sterol biosynthetic pathway was associated to the reduced fungicide sensitivity. In vitro growth rate on agar media and pathogenicity were found to be negatively affected by resistance. The main functional sterols in C. beticola WT strains under investigation were ergosterol, brassicasterol and ergosta-7,22-dienol. Resistant strains showed the same qualitative sterol composition, ruling it out as, per se, a cause for resistance. On the basis of the sterols detected both in sensitive and resistant strains, a possible biosynthetic pathway to the three functional sterols is proposed. Tetraconazole treatment caused, in all sensitive strains, the immediate accumulation of 14alpha-methylated sterols, which, for inhibitor concentrations up to EC50 values, were, in order of abundance, 14alpha-methylergosta-8,24(28)-dien-3beta,6alpha-diol, eburicol and obtusifoliol. However the data do not support a critical role of the 14-methyl-3,6-diol derivative in the growth arrest of C. beticola. As main difference between sensitive and resistant strains, the formers were found to accumulate higher amounts of 14alpha-methylated sterols. Although the data do not allow to establish a specific mechanism for resistance, some molecular mechanisms such as target site alterations and sterol biosynthetic pathway can be ruled out as a possible cause for reduced sensitivity.

Combined overexpression of genes of the ergosterol biosynthetic pathway leads to accumulation of sterols in Saccharomyces cerevisiae.

Genes of the post-squalene ergosterol biosynthetic pathway in Saccharomyces cerevisiae have been overexpressed in a systematic approach with the aim to construct yeast strains that produce high amounts of sterols from a squalene-accumulating strain. This strain had previously been deregulated by overexpressing a truncated HMG-CoA reductase (tHMG1) in the main bottleneck of the early ergosterol pathway. The overexpression of the gene ERG1 (squalene epoxidase) induced a significant decrease of the direct substrate squalene, a high increase of lanosterol, and a small increase of later sterols. The overexpression of the ERG11 gene encoding the sterol-14alpha-demethylase resulted in a decrease of lanosterol and an increase of downstream sterols. When these two genes were simultaneously overexpressed, later sterols from zymosterol to ergosterol accumulated and the content of squalene was decreased about three-fold, indicating that these steps had limited the transformation of squalene into sterols. The total sterol content in this strain was three-fold higher than in a wild-type strain.

Identification of plant sterols in hexaploid and tetraploid wheats using gas chromatography with mass spectrometry.

Free sterols from hexaploid and tetraploid free-threshing wheats (Triticum aestivum L. and T. durum Desf.) and from their respective hulled wheats (T. spelta L. and T. dicoccon Schrank) were analysed by gas chromatography with mass spectrometry. The qualitative analysis of sterols showed a similar pattern either between hexaploid (T. aestivum, T. spelta) and tetraploid (T. durum, T. dicoccon) wheats or between free-threshing (T. aestivum, T. durum) and hulled (T. spelta, T. dicoccon) wheats. However, quantitative differences were found between tetraploid and hexaploid wheats, in that free sterol amounts in tetraploid wheats were 40% higher than in hexaploid ones. The mass spectra of the sterols were classified into four groups, taking into account the structural features of rings A and B. Typical mass spectral fragmentations of the four classes, and additional evidence related to the side chain of each molecule, were investigated together with their chromatographic behaviour, allowing identification of all the detected sterols.

Application of comprehensive two-dimensional gas chromatography to the quantification of overlapping faecal sterols

Standard solutions containing a mixture of seven sterols and 5α-cholestane as internal standard, and sample mixtures that comprised varying ratios of sterol and stanols from green lip mussel tissue and dried cow faeces were analysed by using comprehensive two-dimensional gas chromatography (GC×GC). Quantitative results were compared with single-column GC analysis. The latter samples included sterols of interest, but which cannot be readily obtained elsewhere. It may also mimic potential environmental samples where dairy production and aquaculture (oyster, mussel cultivation) share the same catchment; environmental sterol signatures may exhibit characteristics of both sample types comprising this mixture. Whereas single-column GC–flame ionisation detection was unable to reliably quantitate target sterols, the GC×GC experiment permitted small amounts of sterols and stanols to be detected and separated. Likewise GC–MS analysis was unable to detect some of the minor sterols which coeluted on a single column. The GC×GC mode allows complete separation of several important sterols and stanols, such as 24-ethylcoprostanol, campesterol and 24-methylenecholesterol, demonstrating the enhanced resolving power of the GC×GC system. Separation of 24-ethyl-epi-coprostanol from several algal-derived interfering components was achieved, leading to higher degree of confidence in the quantitative analysis of faecal sterols. The effects of a number of operating variables—column length, carrier flow-rate and elution temperature—on component resolution and presentation of data in the two-column analysis are described.

Physical stability of a microcrystalline beta-sitosterol suspension in oil.

Sterols have been shown to reduce plasma cholesterol by blocking the absorption of cholesterol from the gut. The physical properties of crystalline plant sterols limit their use in foods. A coarse-grained structure can be avoided by recrystallisation, a method that affords a reduction in the particle size. A previous work described how to produce a microcrystalline beta-sitosterol suspension. The present study deals with the stability of that suspension. Recrystallisation was carried out by two different methods; one based on rapid the other based on slow cooling, whereby six different compositions were made containing 5-30% of beta-sitosterol and secondly either 5 or 20% water was added. The particle size and habit were evaluated during a 16 weeks storage period (+4 or -19 degrees C) by way of optical microscopy. The crystal structure and degree of crystallinity was analysed by X-ray diffraction. Suspensions can, in most cases, be stored for 16 weeks without any changes to the size and habit. The only evidence of crystal growth came from a suspension with a low sterol concentration at a temperature of +4 degrees C. This is due to the dissolution-diffusion process which is affected by temperature and viscosity. Suspensions containing higher amounts of sterol remained stable, if stored at +4 or -19 degrees C, for 16 weeks. The suspensions included both hemihydrous and monohydrous beta-sitosterol crystals. Suspensions containing less sterol showed greater amounts of monohydrated crystals. This illustrates more water penetration into the crystals. A higher sterol concentration led to a larger number of smaller crystals creating reflections similar to hemihydrated crystals.

Crosstalk between cytosolic and plastidial pathways of isoprenoid biosynthesis in Arabidopsis thaliana.

In plants, the formation of isopentenyl diphosphate and dimethylallyl diphosphate, the central intermediates in the biosynthesis of isoprenoids, is compartmentalized: the mevalonate (MVA) pathway, which is localized to the cytosol, is responsible for the synthesis of sterols, certain sesquiterpenes, and the side chain of ubiquinone; in contrast, the recently discovered MVA-independent pathway, which operates in plastids, is involved in providing the precursors for monoterpenes, certain sesquiterpenes, diterpenes, carotenoids, and the side chains of chlorophylls and plastoquinone. Specific inhibitors of the MVA pathway (lovastatin) and the MVA-independent pathway (fosmidomycin) were used to perturb biosynthetic flux in Arabidopsis thaliana seedlings. The interaction between both pathways was studied at the transcriptional level by using GeneChip (Affymetrix) microarrays and at the metabolite level by assaying chlorophylls, carotenoids, and sterols. Treatment of seedlings with lovastatin resulted in a transient decrease in sterol levels and a transient increase in carotenoid as well as chlorophyll levels. After the initial drop, sterol amounts in lovastatin-treated seedlings recovered to levels above controls. As a response to fosmidomycin treatment, a transient increase in sterol levels was observed, whereas chlorophyll and carotenoid amounts decreased dramatically when compared with controls. At 96 h after fosmidomycin addition, the levels of all metabolites assayed (sterols, chlorophylls, and carotenoids) were substantially lower than in controls. Interestingly, these inhibitor-mediated changes were not reflected in altered gene expression levels of the genes involved in sterol, chlorophyll, and carotenoid metabolism. The lack of correlation between gene expression patterns and the accumulation of isoprenoid metabolites indicates that posttranscriptional processes may play an important role in regulating flux through isoprenoid metabolic pathways.

Some characteristics of nigella ( Nigella sativa L.) seed cultivated in Egypt and its lipid profile

Physico-chemical properties of nigella seeds cultivated in Egypt were determined. Physical and chemical analyses of crude oils extracted from the seeds, by two different methods were also performed. The results revealed that nigella seed is a good source for oil and protein. Percentage of oil extraction, as well as physical and chemical properties of the crude oil, was influenced by the extraction methods. TLC and GLC analysis indicated that nigella seed oil contained significant amounts of sterols. Linoleic (C 18:2), oleic (C 18:1) and palmitic (C 16:0), as in most of the common edible oils, are the main fatty acids. The oil was rich in β-sitosterol that inhibits the absorption of dietary cholesterol.

Identification of extractable substances from rubber nettings used to package meat products

Ten meat nettings were sampled from four different suppliers in the Netherlands. These meat nettings consisted both of natural rubber and of vegetable fibres. Nitrosamines were extractable up to 2 mg kg-1 netting, the nitrosamine being N -nitrosodibenzylamine. The nitrosatable substances found were precursors of N -nitrosodimethylamine and N -nitrosodibenzylamine and were extracted up to 0.4 mg kg-1 nettings. Considering the ratio of meat netting and foodstuff, the extractable amount of nitrosamines and nitrosatable substances does not raise concern for public health. The meat nettings were also screened for other potential migrants. Extractable amounts of several alkanes, alkenes, acids, antioxidants, plasticizers and sterols were found. Several of these extracted substances are not allowed in the Netherlands, although some are authorized in other countries. Several substances, however, have not been evaluated for use in food-contact materials and therefore the possible risk to public health is unknown.

Preparation of Salmon Roe Polar Lipid Concentrate as an HUFA Enricher of Artemia Nauplii

Polar lipids (PL) concentrate was prepared from total lipids (TL) of salmon roe in order to use for highly unsaturated fatty acids (HUFA) enricher of Artemia nauplii, an important live food for marine fish larvae. The PL were concentrated by a solvent partition procedure using a solvent system of equilibrated hexane, ethanol and water. Starting with 120 g of TL, the solvent partition procedure resulted in 29.3 g (27.2 %) of the salmon roe PL concentrate. Major components of the concentrate were phosphatidylcholines (PC; 80.0 %), phosphatidylethanolamines (PE; 7.0 %), and phosphatidylinositols (PI; 5.5 %). Lesser amounts of free fatty acids, sterols and monoacylglycerols were also found in the PL concentrate. Fatty acid analysis revealed the contents of docosahexaenoic acid (DHA; 26.9 %), icosapentaenoic acid (IPA; 14.2 %), and arachidonic acid (AA; 2.4 %). PC and PE isolated from the concentrate were rich in DHA and IPA, whereas PI were rich in AA. The salmon roe PL concentrate appeared to have a potential as an effective HUFA enricher of Artemia nauplii. The solvent partition procedure seems to be useful for convenient preparation of such fish roe PL concentrate.

Expression of a Streptomyces 3-hydroxysteroid oxidase gene in oilseeds for converting phytosterols to phytostanols

Plant sterols and their hydrogenated forms, stanols, have attracted much attention because of their benefits to human health in reducing serum and LDL cholesterol levels, with vegetable oil processing being their major source in several food products currently sold. The predominant forms of plant sterol end products are sitosterol, stigmasterol, campesterol and brassicasterol (in brassica). In this study, 3-hydroxysteroid oxidase from Streptomyces hygroscopicus was utilized to engineer oilseeds from rapeseed ( Brassica napus) and soybean ( Glycine max), respectively, to modify the relative amounts of specific sterols to stanols. Each of the major phytosterols had its C-5 double bond selectively reduced to the corresponding phytostanol without affecting other functionalities, such as the C-22 double bond of stigmasterol in soybean seed and of brassicasterol in rapeseed. Additionaly, several novel phytostanols were obtained that are not produced by chemical hydrogenation of phytosterols normally present in plants.

Dietary Fiber-Rich Barley Products Beneficially Affect the Intestinal Tract of Rats

The objective of this study was to investigate the effects of barley-rich diets in the intestinal tract of rats. Four test groups (A-D) of 10 young male Wistar rats were fed diets containing 50 g/100 g barley extrudates (A, B and D) or mixtures (C) for 6 wk; the control diet contained no barley. The barley-containing supplements in the test diets were: A = cultivar "HiAmi"; B = "HiAmi" and "Prowashonupana" (50:50); C = "Prowashonupana" and Novelose (50:50); D = "Prowashonupana" and amylose from maize (60:40). These supplements contained 7-12 g/100 g beta-glucan and 7-24 g/100 g resistant starch. Additionally, 5 g microcrystalline cellulose/100 g was present in all diets. Carbohydrate utilization (indirect calorimetry) was lower (P < 0.05) in rats fed the barley-containing diets C or D than in the controls. In the test groups, the following differences from the controls were found: greater food intake in the last 2 wk (P < 0.05); increased weight gain in wk 6 (P < 0.05); greater mass of the ceca (groups B-D; P < 0.05) and colons (P < 0.001) as well as masses of cecal (groups C and D; P < 0.01) and colon contents (P < 0.001); greater concentrations of resistant starch in cecal and most of the colon contents (P < 0.05); and more beta-glucan in the small intestine, cecum and colon (P < 0.05). The numbers of coliforms and Bacteroides were lower than in the controls in groups B-D and those of Lactobacillus were greater in all test groups (P < 0.05). Short-chain fatty acids (SCFA) were higher in the cecal contents of the test groups (> or = 800 micro mol/g DM; P < 0.001) compared with the controls ( approximately 200 micro mol/g DM). Similarly, SCFA were higher in colon and feces of the test groups. The concentrations of excreted bile acids increased up to 30% during the feeding period. The proportions of secondary bile acids were lower and the amounts of neutral sterols (P < 0.001) were greater in feces of rats fed the barley-containing diets for 6 wk than in the controls. Diets containing more soluble macromolecular dietary fibers such as beta-glucans affected the excretion of bile acids and neutral sterols the most, whereas the fermentation of dietary fiber, including resistant starch, influenced the steroids in feces. These results suggest that dietary fiber-rich barley-containing diets have beneficial physiologic effects.

Lipids and ABC transporters

The next session was dedicated to lipid transport within and/or across the membrane. SHAILESH PATEL (Charleston, South Carolina, USA) spoke on the recent discovery of the relation between mutations in the ABCG5 and ABCG8 genes in patients with sitosterolemia. These two genes encode half-transporters (named sterolin-1 and sterolin-2, respectively). Clinical symptoms of sitosterolemia are tendon xanthomas, accelerated atherosclerosis particularly affecting males at young age, hemolytic episodes, and arthritis and arthralgias. The hallmark biochemical feature of the disease is the elevated concentration of plant sterols in plasma. In different patients with this rare inherited disorder mutations were found in either the ABCG5 or the ABCG8 gene. This, together with the knowledge that several other ABC half-transporters dimerize into functional pumps, has given rise to the hypothesis that ABCG5 and ABCG8 are obligatory partners in a plant sterol pump. It is of importance to note that both genes are expressed in liver as well as in the intestine. Hence, the current hypothesis is, that in the gut the transporter couple extrudes plant sterols that enter the epithelial cells back into the gut lumen. The small amount of plant sterols that do enter the body are then efficiently pumped into bile by ABCG5 and ABCG8 in the canalicular membrane. Because these patients also have hypercholesterolemia, it is the hypothesis that this transporter couple also translocates cholesterol to some extent. However, it remains to be determined to what extent this transporter determines enteral absorption as well as biliary secretion of cholesterol. The locus has a very peculiar structure in that the two genes lay head-to-head on chromosome 2p21. Although the promoter of this gene pair has not been identified, yet it is clear that they are coordinately expressed and induced by the nuclear receptor LXR. GERD SCHMITZ (Regensburg, Germany) spoke about ABCA1, the expression of which is induced by cholesterol as is the case for a number of other ABC transporter genes of the A subfamily, such as ABCA3 and ABCA7. Mutations in the ABCA1 gene cause a complete lack of high-density lipoprotein (HDL; Tangier disease). The molecular function of ABCA1 in the assembly of nascent HDL is still unclear. He reported that, in addition to induction by the oxysterol-sensitive nuclear receptor LXR, ABCA1 transcription is repressed by the zinc finger protein ZNF202. The later transcription factor is located in a hypoalphalipoproteinemia locus on chromosome 11q23. In collaboration with the group of B. Sarkadi, G. Schmitz's lab analyzed ATP binding and ATPase activity in membranes from Sf9 cells expressing ABCA1. While they could observe ATP-binding properties of the protein, they were unable to measure ATPase activity, which led Gerd Schmitz to the assumption that ABCA1 functions as an ion (potassium) channel regulator rather than as a lipid translocator. Via this function ABCA1 might regulate the intracellular trafficking of vesicles to and from the plasma membrane. This function would be analogous to that of SUR in its complex with the potassium channel KIR6.2. ALBERT K. GROEN (Amsterdam, The Netherlands) spoke about the mechanism of biliary lipid transport. Phospholipid secretion involves phosphatidylcholine translocation by MDR3 P-glycoprotein (P-gp), but it is still unclear whether other gene products are involved in this complex process. When it was discovered that ABCA1 is involved in cholesterol transfer from cells to nascent HDL it was an interesting candidate for catalysis of cholesterol secretion into bile. Analysis of cholesterol secretion into bile of Abca1-/- mice demonstrated, however, that Abca1 plays no role in this process. In addition, the surprising observation was made that reverse cholesterol transport from peripheral tissues to the liver is not impaired in these knockout mice, indicating that the role of HDL in this process is overestimated. At least, in the absence of HDL in mice, reverse cholesterol transport is achieved via different pathways. Albert Groen reported that Abcg5 expression under conditions of different biliary cholesterol transport correlated well with the extent of cholesterol secretion into bile, which suggests that this gene product is involved.