Adherence to the Healthy Nordic Food Index and the incidence of acute myocardial infarction and mortality among patients with stable angina pectoris.

BACKGROUND: The Healthy Nordic Food Index (HNFI) has been associated with beneficial effects on markers of cardiovascular disease (CVD). Whether such effects are present among patients with established coronary heart disease is unknown. In the present study, we investigated the association between adherence to the HNFI and the risk of acute myocardial infarction (AMI) (fatal or nonfatal) and death among patients with stable angina pectoris. METHODS: In the Western Norway B-vitamin Intervention Trial, participants completed a 169-item semi-quantitative food frequency questionnaire. The HNFI was calculated from six food groups (fish, cabbage, apples/pears, root vegetables, whole grain bread and oatmeal), scoring 0-6. Three adherence groups were defined: 0-1 points (low), 2-3 points (medium) or 4-6 points (high). Cox regression analyses investigated associations between adherence to the HNFI and outcomes. RESULTS: Among 2019 men (79.7%) and women with mean age of 61.7 years, 307 patients experienced an AMI event during a median (25th and 75th percentiles) follow-up of 7.5 (6.3 and 8.7) years. Median follow-up for total mortality was 10.5 (9.3 and 11.7) years; 171 patients died from CVD and 380 from any cause. No association between HNFI and the risk of AMI was detected. However, the HNFI was associated with a reduced risk of all-cause death, both by linear estimates [hazard ratio (95% confidence interval = 0.91 (0.84-0.98)] and by comparison of the highest with the lowest adherence group [hazard ratio (95% confidence interval = 0.70 (0.52-0.95)]. CONCLUSIONS: The results of the present study suggest that a Healthy Nordic diet may reduce mortality in patients with established CVD.

A gene variation (rs12691) in the CCAT/enhancer binding protein α modulates glucose metabolism in metabolic syndrome.

BACKGROUND AND AIMS: CCAAT/enhancer-binding protein alpha (CEBPA) is a transcription factor involved in adipogenesis and energy homeostasis. Caloric restriction reduces CEBPA protein expression in patients with metabolic syndrome (MetS). A previous report linked rs12691 SNP in CEBPA to altered concentration of fasting triglycerides. Our objective was to assess the effects of rs12691 in glucose metabolism in Metabolic Syndrome (MetS) patients. METHODS AND RESULTS: Glucose metabolism was assessed by static (glucose, insulin, adiponectin, leptin and resistin plasma concentrations) and dynamic (disposition index, insulin sensitivity index, HOMA-IR and acute insulin response to glucose) indices, performed at baseline and after 12 weeks of 4 dietary interventions (high saturated fatty acid (SFA), high monounsaturated fatty acid (MUFA), low-fat and low-fat-high-n3 polyunsaturated fatty acid (PUFA)) in 486 subjects with MetS. Carriers of the minor A allele of rs12691 had altered disposition index (p = 0.0003), lower acute insulin response (p = 0.005) and a lower insulin sensitivity index (p = 0.025) indicating a lower insulin sensitivity and a lower insulin secretion, at baseline and at the end of the diets. Furthermore, A allele carriers displayed lower HDL concentration. CONCLUSION: The presence of the A allele of rs12691 influences glucose metabolism of MetS patients.

The effect of strength training volume on satellite cells, myogenic regulatory factors, and growth factors.

The aim of this work was to study the effect of training volume on activation of satellite cells. Healthy untrained men were randomly assigned into two groups. The 3L-1UB group (n = 10) performed three-set leg exercises and single-set upper body exercises, and the 1L-3UB group (n = 11) performed single-set leg exercises and three-set upper body exercises. Both groups performed three sessions (80-90 min) per week for 11 weeks. Biopsies were taken from m. vastus lateralis and m. trapezius. The number of satellite cells, satellite cells positive for myogenin and MyoD, and the number of myonuclei were counted. Homogenized muscle was analyzed for myogenin and MyoD, and extracted ribonucleic acid (RNA) was monitored for selected growth factor transcripts. Knee extensor strength increased more in the 3L-1UB group than in the 1L-3UB group (48 ± 4% vs 29 ± 4%), whereas the strength gain in shoulder press was similar in both training groups. The number of satellite cells in m. vastus lateralis increased more in the 3L-1UB group than in the 1L-3UB group. The number of myonuclei increased similarly in both groups. The messenger RNA expression of growth factors peaked after 2 weeks of training. In conclusion, increasing training volume enhanced satellite cell numbers in the leg muscle, but not in the upper body muscle.

Effects of dietary fat modification on insulin sensitivity and on other risk factors of the metabolic syndrome--LIPGENE: a European randomized dietary intervention study.

BACKGROUND: Excessive energy intake and obesity lead to the metabolic syndrome (MetS). Dietary saturated fatty acids (SFAs) may be particularly detrimental on insulin sensitivity (SI) and on other components of the MetS. OBJECTIVE: This study determined the relative efficacy of reducing dietary SFA, by isoenergetic alteration of the quality and quantity of dietary fat, on risk factors associated with MetS. DESIGN: A free-living, single-blinded dietary intervention study. SUBJECTS AND METHODS: MetS subjects (n = 417) from eight European countries completed the randomized dietary intervention study with four isoenergetic diets distinct in fat quantity and quality: high-SFA; high-monounsaturated fatty acids and two low-fat, high-complex carbohydrate (LFHCC) diets, supplemented with long chain n-3 polyunsaturated fatty acids (LC n-3 PUFAs) (1.2 g per day) or placebo for 12 weeks. SI estimated from an intravenous glucose tolerance test (IVGTT) was the primary outcome measure. Lipid and inflammatory markers associated with MetS were also determined. RESULTS: In weight-stable subjects, reducing dietary SFA intake had no effect on SI, total and low-density lipoprotein cholesterol concentration, inflammation or blood pressure in the entire cohort. The LFHCC n-3 PUFA diet reduced plasma triacylglycerol (TAG) and non-esterified fatty acid concentrations (P < 0.01), particularly in men. CONCLUSION: There was no effect of reducing SFA on SI in weight-stable obese MetS subjects. LC n-3 PUFA supplementation, in association with a low-fat diet, improved TAG-related MetS risk profiles.

Dietary supplementation of tetradecylthioacetic acid increases feed intake but reduces body weight gain and adipose depot sizes in rats fed on high-fat diets.

AIM: The pan-peroxisome proliferator-activated receptor (PPAR) ligand and fatty acid analogue tetradecylthioacetic acid (TTA) may reduce plasma lipids and enhance hepatic lipid metabolism, as well as reduce adipose tissue sizes in rats fed on high-fat diets. This study further explores the effects of TTA on weight gain, feed intake and adipose tissue functions in rats that are fed a high-fat diet for 7 weeks. METHODS: The effects on feed intake and body weight during 7 weeks' dietary supplement with TTA ( approximately 200 mg/kg bw) were studied in male Wistar rats fed on a lard-based diet containing approximately 40% energy from fat. Adipose tissue mass, body composition and expression of relevant genes in fat depots and liver were measured at the end of the feeding. RESULTS: Despite higher feed intake during the final 2 weeks of the study, rats fed on TTA gained less body weight than lard-fed rats and had markedly decreased subcutaneous, epididymal, perirenal and mesenteric adipose depots. The effects of TTA feeding with reduced body weight gain and energy efficiency (weight gain/feed intake) started between day 10 and 13. Body contents of fat, protein and water were reduced after feeding lard plus TTA, with a stronger decrease in fat relative to protein. Plasma lipids, including Non-Esterified Fatty Acids (NEFA), were significantly reduced, whereas fatty acid beta-oxidation in liver and heart was enhanced in lard plus TTA-fed rats. Hepatic UCP3 was expressed ectopically both at protein and mRNA level (>1900-fold), whereas Ucp1 mRNA was increased approximately 30-fold in epididymal and approximately 90-fold in mesenteric fat after lard plus TTA feeding. CONCLUSION: Our data support the hypothesis that TTA feeding may increase hepatic fatty acid beta-oxidation, and thereby reduce the size of adipose tissues. The functional importance of ectopic hepatic UCP3 is unknown, but might be associated with enhanced energy expenditure and thus the reduced feed efficiency.

Healthy minds from 0-100 years: Optimising the use of European brain imaging cohorts ("Lifebrain").

The main objective of "Lifebrain" is to identify the determinants of brain, cognitive and mental (BCM) health at different stages of life. By integrating, harmonising and enriching major European neuroimaging studies across the life span, we will merge fine-grained BCM health measures of more than 5,000 individuals. Longitudinal brain imaging, genetic and health data are available for a major part, as well as cognitive and mental health measures for the broader cohorts, exceeding 27,000 examinations in total. By linking these data to other databases and biobanks, including birth registries, national and regional archives, and by enriching them with a new online data collection and novel measures, we will address the risk factors and protective factors of BCM health. We will identify pathways through which risk and protective factors work and their moderators. Exploiting existing European infrastructures and initiatives, we hope to make major conceptual, methodological and analytical contributions towards large integrative cohorts and their efficient exploitation. We will thus provide novel information on BCM health maintenance, as well as the onset and course of BCM disorders. This will lay a foundation for earlier diagnosis of brain disorders, aberrant development and decline of BCM health, and translate into future preventive and therapeutic strategies. Aiming to improve clinical practice and public health we will work with stakeholders and health authorities, and thus provide the evidence base for prevention and intervention.

Global mRNA sequencing of human skeletal muscle: Search for novel exercise-regulated myokines.

OBJECTIVE: Skeletal muscle is an important secretory organ, producing and releasing numerous myokines, which may be involved in mediating beneficial health effects of physical activity. More than 100 myokines have been identified by different proteomics approaches, but these techniques may not detect all myokines. We used mRNA sequencing as an untargeted approach to study gene expression of secreted proteins in skeletal muscle upon acute as well as long-term exercise. METHODS: Twenty-six middle-aged, sedentary men underwent combined endurance and strength training for 12 weeks. Skeletal muscle biopsies from m. vastus lateralis and blood samples were taken before and after an acute bicycle test, performed at baseline as well as after 12 weeks of training intervention. We identified transcripts encoding secretory proteins that were changed more than 1.5-fold in muscle after exercise. Secretory proteins were defined based on either curated UniProt annotations or predictions made by multiple bioinformatics methods. RESULTS: This approach led to the identification of 161 candidate secretory transcripts that were up-regulated after acute exercise and 99 that where increased after 12 weeks exercise training. Furthermore, 92 secretory transcripts were decreased after acute and/or long-term physical activity. From these responsive transcripts, we selected 17 candidate myokines sensitive to short- and/or long-term exercise that have not been described as myokines before. The expression of these transcripts was confirmed in primary human skeletal muscle cells during in vitro differentiation and electrical pulse stimulation (EPS). One of the candidates we identified was macrophage colony-stimulating factor-1 (CSF1), which influences macrophage homeostasis. CSF1 mRNA increased in skeletal muscle after acute and long-term exercise, which was accompanied by a rise in circulating CSF1 protein. In cultured muscle cells, EPS promoted a significant increase in the expression and secretion of CSF1. CONCLUSION: We identified 17 new, exercise-responsive transcripts encoding secretory proteins. We further identified CSF1 as a novel myokine, which is secreted from cultured muscle cells and up-regulated in muscle and plasma after acute exercise.

Dietary intake and nutritional indicators are transiently compromised during radiotherapy for rectal cancer.

OBJECTIVE: Patients with rectal cancer receive curative radiotherapy towards the pelvis for 5 weeks. Little is known about the impact of radiotherapy on dietary intake and nutritional status. The objective was to examine whether curative radiotherapy for rectal cancer promoted altered intake of energy and nutrients, and change in nutritional indicators. DESIGN: Prospective study. SETTING: Department of Oncology in a tertiary care hospital. SUBJECTS: A total of 31 consecutive patients receiving radiotherapy for rectal cancer (50 Gray). INTERVENTIONS: A 7-day food intake registration, body weight, upper arm circumference, and analyses of blood samples were performed at the start and the end of radiotherapy, and at follow-up 4-6 weeks and 1 year after the end of radiotherapy. RESULTS: At the end of 5 weeks of radiotherapy, the mean daily energy intake was reduced by 15% from 8.9 to 7.6 MJ as compared with baseline (P = 0.002), and the intake of several nutrients was reduced (P < 0.01). The percentages of energy derived from fat, protein, and carbohydrates did not change, nor did the nutrient density. A transient body weight reduction of < 1 kg was observed (P = 0.009). Serum concentrations of vitamin A and 25-OH vitamin D did not change during radiotherapy. The daily intake of energy and nutrients, and body weight, had increased towards pretreatment values 4-6 weeks after radiotherapy. CONCLUSIONS: Radiotherapy for rectal cancer caused a transient reduction in energy intake and nutritional indicators. The nutritional quality of the diet was unchanged during radiotherapy.

Fat-soluble vitamins in breast-fed preterm and term infants.

OBJECTIVE: To examine the supply and status of fat-soluble vitamins in very low birth weight (VLBW) infants compared to a reference group of normal birth weight (NBW) infants. DESIGN: A longitudinal study of VLBW infants in the early neonatal period. Blood samples were drawn at 1 week of age and at discharge from hospital. Plasma was analyzed for the fat-soluble vitamins: retinol, 25-OH-vitamin D, alpha-tocopherol and phylloquinone (vitamin K(1)) using high-performance liquid chromatography. SUBJECTS: A total of 40 VLBW infants were included in the study. A reference group of 33 NBW infants was randomly selected from one of our previous studies. RESULTS: The VLBW infants received fortified human milk, and daily oral vitamin supplement (Multibionta). In VLBW infants, plasma retinol concentrations decreased and plasma 25-OH-vitamin D increased during the study period. VLBW infants had significantly lower plasma retinol (0.3 vs 0.7 mu M) and higher plasma 25-OH-vitamin D (166 vs 25 nM) at discharge compared to NBW infants. Plasma phylloquinone concentration in VLBW infants was very high (53 ng/ml) at one week of age, especially in the youngest infants (192 ng/ml), but decreased rapidly during the study period resulting in low/normal plasma concentrations (0.9 ng/ml) at discharge. CONCLUSIONS: We observed alterations in plasma concentration of retinol and 25-OH-vitamin D in VLBW infants in the early neonatal period, resulting in marked differences between VLBW at discharge and NBW. Further trials are needed to evaluate whether changes in vitamin supplementation may improve clinical outcome in VLBW infants.

Biomarkers for nutrient intake with focus on alternative sampling techniques.

Biomarkers of nutrient intake or nutrient status are important objective measures of foods/nutrients as one of the most important environmental factors people are exposed to. It is very difficult to obtain accurate data on individual food intake, and there is a large variation of nutrient composition of foods consumed in a population. Thus, it is difficult to obtain precise measures of exposure to different nutrients and thereby be able to understand the relationship between diet, health, and disease. This is the background for investing considerable resources in studying biomarkers of nutrients believed to be important in our foods. Modern technology with high sensitivity and specificity concerning many nutrient biomarkers has allowed an interesting development with analyses of very small amounts of blood or tissue material. In combination with non-professional collection of blood by finger-pricking and collection on filters or sticks, this may make collection of samples and analyses of biomarkers much more available for scientists as well as health professionals and even lay people in particular in relation to the marked trend of self-monitoring of body functions linked to mobile phone technology. Assuming standard operating procedures are used for collection, drying, transport, extraction, and analysis of samples, it turns out that many analytes of nutritional interest can be measured like metabolites, drugs, lipids, vitamins, minerals, and many types of peptides and proteins. The advantage of this alternative sampling technology is that non-professionals can collect, dry, and mail the samples; the samples can often be stored under room temperature in a dry atmosphere, requiring small amounts of blood. Another promising area is the potential relation between the microbiome and biomarkers that may be measured in feces as well as in blood.

Myostatin in relation to physical activity and dysglycaemia and its effect on energy metabolism in human skeletal muscle cells.

AIM: Some health benefits of exercise may be explained by an altered secretion of myokines. Because previous focus has been on upregulated myokines, we screened for downregulated myokines and identified myostatin. We studied the expression of myostatin in relation to exercise and dysglycaemia in skeletal muscle, adipose tissue and plasma. We further examined some effects of myostatin on energy metabolism in primary human muscle cells and Simpson-Golabi-Behmel syndrome (SGBS) adipocytes. METHODS: Sedentary men with or without dysglycaemia underwent a 45-min acute bicycle test before and after 12 weeks of combined endurance and strength training. Blood samples and biopsies from m. vastus lateralis and adipose tissue were collected. RESULTS: Myostatin mRNA expression was reduced in skeletal muscle after acute as well as long-term exercise and was even further downregulated by acute exercise on top of 12-week training. Furthermore, the expression of myostatin at baseline correlated negatively with insulin sensitivity. Myostatin expression in the adipose tissue increased after 12 weeks of training and correlated positively with insulin sensitivity markers. In cultured muscle cells but not in SGBS cells, myostatin promoted an insulin-independent increase in glucose uptake. Furthermore, muscle cells incubated with myostatin had an enhanced rate of glucose oxidation and lactate production. CONCLUSION: Myostatin was differentially expressed in the muscle and adipose tissue in relation to physical activity and dysglycaemia. Recombinant myostatin increased the consumption of glucose in human skeletal muscle cells, suggesting a complex regulatory role of myostatin in skeletal muscle homeostasis.

Comparison of the portion size and frequency of consumption of 156 foods across seven European countries: insights from the Food4ME study.

There are no standardised serving/portion sizes defined for foods consumed in the European Union (EU). Typical serving sizes can deviate significantly from the 100 g/100 ml labelling specification required by the EU legislation. Where the nutritional value of a portion is specified, the portion size is determined by the manufacturers. Our objective was to investigate the potential for standardising portion sizes for specific foods, thereby ensuring complementarity across countries. We compared portion size for 156 food items measured using a food frequency questionnaire across the seven countries participating in the Food4me study. The probability of consuming a food and the frequency of consumption differed across countries for 93% and 58% of the foods, respectively. However, the individual country mean portion size differed from the average across countries in only 16% of comparisons. Thus, although dietary choices vary markedly across countries, there is much less variation in portion sizes. Our results highlight the potential for standardisation of portion sizes on nutrition labels in the EU.

The exercise-regulated myokine chitinase-3-like protein 1 stimulates human myocyte proliferation.

AIM: Chitinase-3-like protein 1 (CHI3L1) is involved in tissue remodelling and inflammatory processes. Plasma levels are elevated in patients with insulin resistance and T2DM. We recently showed that CHI3L1 and its receptor protease-activated receptor 2 (PAR-2) are expressed in skeletal muscle. Activation of PAR-2 by CHI3L1 protects against TNF-α-induced inflammation and insulin resistance. However, the effect of exercise on CHI3L1 and PAR-2 signalling remains unknown. The aim of this work was to study the impact of exercise on CHI3L1 production and the effect of CHI3L1/PAR-2 signalling on skeletal muscle growth and repair. METHODS: Three human exercise studies were used to measure CHI3L1 plasma levels (n = 32). In addition, muscle and adipose tissue CHI3L1 mRNA expression was measured in response to acute and long-term exercise (n = 24). Primary human skeletal muscle cells were differentiated in vitro, and electrical pulse stimulation was applied. In addition, myoblasts were incubated with CHI3L1 protein and activation of MAP kinase signalling as well as proliferation was measured. RESULTS: Circulating CHI3L1 levels and muscle CHI3L1 mRNA were increased after acute exercise. In addition, CHI3L1 mRNA expression as well as CHI3L1 secretion was enhanced in electrically stimulated cultured myotubes. Incubation of cultured human myoblasts with CHI3L1 protein leads to a strong activation of p44/42, p38 MAPK and Akt as well as enhanced myoblast proliferation. CONCLUSION: Our findings suggest that CHI3L1 is induced by acute exercise and that CHI3L1/PAR-2 signalling activates myocyte proliferation, which is important for restructuring of skeletal muscle in the response to exercise training.

Demonstration of intact intracellular cholesterol esterification and normal low-density lipoprotein pathway in fibroblasts from a patient with lecithin:cholesterol acyltransferase deficiency.

There was a 6-46-fold increase in intracellular cholesterol esterification in response to 25-hydroxycholesterol and low-density lipoproteins in normal and lecithin:cholesterol acyltransferase-deficient fibroblasts. Uptake and degradation of 125I-labelled low-density lipoproteins were similar in the two cell lines. Low-density lipoproteins caused a doubling of the mass of cholesteryl ester in the mutant cells. These findings indicate that: (a) acyl-CoA:cholesterol acyltransferase exhibits normal activity in mutant cells; (b) lecithin:cholesterol acyltransferase and acyl-CoA:cholesterol acyltransferase are different enzymes and are probably not products of the same gene; (c) the low-density lipoprotein-pathway is intact in fibroblasts from a patient with lecithin:cholesterol acyltransferase deficiency; (d) acyl-CoA:cholesterol acyltransferase is probably responsible for the small amount of cholesteryl ester found in plasma from patients with lecithin:cholesterol acyltransferase deficiency.

Serum half-life, distribution, hepatic uptake and biliary excretion of alpha-tocopherol in rats.

The serum clearance of alpha-[3H]tocopherol has been studied after intravenous injection of intestinal lymph labeled in vivo with radioactive alpha-tocopherol. The half-life of the injected alpha-[3H]tocopherol was approx. 12 min. Fractionation of plasma by ultracentrifugation 10 min after injection of lymph showed that 91% of the radioactive alpha-tocopherol remaining in plasma was located in chylomicrons (d less than 1.006 g/ml) and 7.8% in high-density lipoproteins (HDL, 1.05 less than d less than 1.21 g/ml). 2 h after administration of alpha-tocopherol, about 35% of the radioactivity recovered in plasma was associated with chylomicrons and approx. 51% with HDLs. alpha-[3H]Tocopherol was initially taken up by the liver, which contained more than 50% of the injected radioactivity after 45-60 min. Separation of parenchymal and nonparenchymal cells demonstrated a preferential uptake of alpha-[3H]tocopherol by the parenchymal liver cells. After 24 h about 11% of the injected dose was recovered in the liver. Considering whole organs the liver, adipose tissue and skeletal muscle had the highest content of radioactivity after 24 h. Furthermore, about 14% of the administered dose was recovered in bile during 24 h draining.

Dissociation of the lipid-enzyme complex of hormone-sensitive lipase using high density lipoprotein or apolipoprotein A-I.

Hormone-sensitive lipase in homogenates of adipose tissue occurs as a large, lipid-rich complex including several acylhydrolase activities that emerge quantitatively in the void volume on gel filtration chromatography (2% agarose). Incubation with intact human plasma high density lipoprotein or with lipid-free apolipoprotein A-I, however, disrupted the lipid-rich complex almost completely and most of the enzyme activity eluted from a 2% agarose column at about Ve = 2.3 x Vo. This use of the detergent-like properties of apolipoprotein A-I may be of value for dissociation of other lipid-associated or membrane-bound enzymes.

Uptake and degradation of cholesterol ester-labelled rat plasma lipoproteins in purified rat hepatocytes and nonparenchymal liver cells.

1. A new method for isolation and purification of rat liver hepatocytes and nonparenchymal cells by differential centrifugation is described. 2. Cholesterol ester-labelled lipoproteins (prepared by the action of lecithin: cholesterol acyltransferase) intravenously injected were taken up by hepatocytes and nonparenchymal cells. 3. Hepatocytes and nonparenchymal cells in suspension were able to take up and hydrolyse the cholesterol ester portion of lipoproteins. 4. Uptake of cholesterol ester labelled whole rat plasma and high density lipoproteins (HDL) increased with increasing concentrations until a distinct saturation level was reached in hepatocytes. In nonparenchymal cells there was no saturation of lipoprotein uptake. 5. Concanavalin A inhibited cholesterol ester-labelled lipoprotein uptake in hepatocytes, indicating that the uptake at least partially depends on carbohydrate sites on the cell surface. The uptake in nonparenchymal cells was unaffected of concanavalin A. 6. The specific activity of the acid cholesterol ester hydrolase was the same in homogenates from hepatocytes and nonparenchymal cells while acyl-CoA: cholesterol acyltransferase was found almost exclusively in hepatocytes.

Fatty acid uptake and metabolism in CaCo-2 cells: eicosapentaenoic acid (20:5(n-3)) and oleic acid (18:1(n-9)) presented in association with micelles or albumin.

In vivo, long-chain fatty acids are incorporated into bile salt micelles, which solubilize the hydrophobic fatty acids before they are transported across the unstirred water layer to the intestinal brush border membrane. In the present study we have used CaCo-2 cells, cultured on filter membranes as a model of human enterocytes, and compared presentation of fatty acids bound to albumin with a micellar form. Absorption of eicosapentaenoic acid and oleic acid from micellar solutions was 4-times faster than from fatty acid-albumin complexes after 5 h incubation, and resulted in a corresponding increase in triacylglycerol synthesis and secretion. Mass determination of newly synthesized, cell-associated triacylglycerol after 5 h incubation, indicated a 5-fold increase in cells exposed to a micellar solution versus albumin-complexed fatty acids. A 2-fold larger fraction of the absorbed fatty acids was incorporated into triacylglycerol than into phospholipids when the fatty acids were presented as micelles. Analysis by resistive pulse technique showed that secreted lipoproteins of density less than 1.006 g/ml were in the same size-range as chylomicrons derived from human plasma. In spite of an increased amount of secreted triacylglycerol from cells supplemented with micellar fatty acids, there was no increase in the mean size of these particles. Synthesis and secretion of triacylglycerol in cells exposed to eicosapentaenoic acid and oleic acid were similar regardless of whether the fatty acids were presented to the cells associated with albumin or micelles, although the total amount of triacylglycerol synthesized and secreted was highest with micelles. When incubating CaCo-2 monolayers with eicosapentaenoic acid or oleic acid bound to albumin, a similar amount of radioactivity was released as CO2 and acid soluble products into the medium with less than 3% of the lipids being oxidized after 5 h of incubation. The oxidation rate of fatty acids in cells incubated with micelles was increased 40 to 100%. In conclusion, micellar fatty acids are absorbed, metabolized and influence secretion of lipoprotein particles to a higher extent than albumin-bound fatty acids in CaCo-2 cells, and there is no major difference between eicosapentaenoic acid and oleic acid.

Acute and chronic effects of ethanol on secretion of alpha-tocopherol from primary cultures of rat hepatocytes.

Primary cultures of rat hepatocytes were used to study the effect of acute and chronic ethanol exposure on alpha-tocopherol content in cells and media. Cells treated acutely with 60 mM ethanol secreted 74.5 +/- 18.0% (P less than 0.05), and their cellular alpha-tocopherol content was 85.7 +/- 15.4% (not significant) of controls after 20 h incubation. At this time total recovery of alpha-tocopherol was significantly reduced in ethanol-exposed cells (43.1 +/- 8.4%) as compared to control cells (52.8 +/- 5.0%, P less than 0.05). Hepatocytes isolated from chronic ethanol-fed rats (35% of total energy intake as ethanol for 5 weeks) secreted 41.9 +/- 12.7% less alpha-tocopherol than did cells of pair-fed controls during 20 h incubation (P less than 0.05). The amount of alpha-tocopherol secreted was then 15.6 +/- 4.2 and 19.8 +/- 3.8% of cell-associated alpha-tocopherol at start of incubation for chronic ethanol-fed and control rats, respectively (P less than 0.05). When 60 mM ethanol was added to the incubation medium, hepatocytes of control rats secreted significantly less alpha-tocopherol (about 30%, P less than 0.05), whereas alpha-tocopherol secretion was not significantly reduced in hepatocytes of chronic ethanol-fed rats. We conclude that both acute and chronic ethanol exposure reduce alpha-tocopherol secretion from rat hepatocytes.

Inhibition of very-low-density lipoprotein secretion by chloroquine, verapamil and monensin takes place in the Golgi complex.

The effects of chloroquine, verapamil and monensin on secretion of very-low-density lipoproteins (VLDLs) were studied in cultured rat hepatocytes. Maximum inhibition of VLDL-triacylglycerol secretion by 50-90% of control was reached at 200 microM chloroquine, 200 microM verapamil and 5 microM monensin, whereas no effect on cellular triacylglycerol synthesis was observed. The inhibition could be seen within 15 min and was reversible after washout of the drugs. Chloroquine and verapamil inhibited both cellular protein synthesis and protein secretion, whereas monensin reduced protein secretion without any effect on protein synthesis. Control experiments with cycloheximide revealed that intact protein synthesis was not necessary for secretion of VLDL-triacylglycerol during 2 h. Electron micrographs of cells treated with chloroquine, verapamil or monensin showed swollen Golgi cisternae containing VLDL-like particles. By morphometry, a more than 2-fold increase in volume fractions and size indices of Golgi complexes and secondary lysosomes was observed, except that monensin had no significant effect on these parameters of secondary lysosomes. These results suggest that the inhibition of VLDL secretion by chloroquine, verapamil and monensin which takes place in the Golgi complex might be due to disruption of trans-membrane proton gradients. An increase in pH of acidic Golgi vesicles may cause swelling and disturb sorting and membrane flow through this organelle.