Long-chain n-3 fatty acids (FA) consistently inhibit the growth of human breast cancer (BC) cells both in culture and in grafts in immunosuppressed mice. Large cohort studies have, however, failed to confirm a protective effect for fish oils rich in n-3 FA against BC risk. The present review examines new evidence on biological mechanisms which may be involved in the inhibition of mammary carcinogenesis by long-chain n-3 FA, focusing on an apoptotic effect by its lipid peroxidation products. Dietary intake of n-3 FA leads to their incorporation into cell membrane lipids. Increased apoptosis in human BC cells following exposure to long-chain n-3 FA such as eicosapentaenoic and docosahexaenoic acids is generally ascribed to their inhibition of cyclooxygenase 2 which promotes mammary carcinogenesis. In addition however, long-chain n-3 FA are particularly likely to activate peroxisome proliferator-activated receptor (PPAR)-γ, a key regulator of lipid metabolism but also capable of modulating proliferative activity in a variety of cells including mammary cells. Expression of PPAR-γ in the nucleus is activated by second messengers such as J series prostaglandins and the latter have been shown to cause apoptosis in vivo in explants of human BC cells in immunosuppressed mice. In mammary tumours, it is observed that long-chain FA not only increase apoptosis, but also increase lipid peroxidation, and the apoptotic effect can be reversed by antioxidants. The rationale for use of n-3 FA dietary supplements in counteracting BC progression needs to be tested clinically in a phase 2 pilot study, while at the same time, the effect on whole-body lipid peroxidation needs to be monitored. Dietary supplements of fish oil rich in n-3 FA are proposed for premenopausal women over the age of 40 years who are shown to be at increased BC risk. Biological markers in breast tissue of BC progression will be monitored, and observed changes related to serial plasma levels of isoprostanes as a measure of whole-body lipid peroxidation.

The influence of myristic acid in a narrow physiological range (0·5 to 2·4 % of total dietary energy) on the plasma and hepatic cholesterol metabolism was investigated in the hamster. The hamsters were fed on a diet containing 12·5 g fat/100 g and 0·05 g cholesterol/100 g with 0·5 % myristic acid (LA diet) for 3 weeks (pre-period). During the following 3 weeks (test period), they were divided into four dietary groups with 0·5 % (LA), 1·2 % (LM), 1·8 % (ML) or 2·4 % (M) myristic acid. Finally, half the hamsters in each group were again fed the LA diet for another 3 weeks (post-period). At the end of the test period, the hepatic expression of the scavenger receptor BI (SR-BI) was lower in the LM, ML and M groups than in the LA group whereas the hepatic cholesteryl ester concentration was higher. Cholesterol 7α hydroxylase activity was lower in the ML and M groups than in the LA and LM groups while the sterol 27 hydroxylase and 3-hydroxy-3-methyl glutaryl coenzyme A reductase activities were not modulated by dietary myristic acid. This is the first time a negative correlation has been observed between the HDL-cholesterol concentration and the hepatic mass of SR-BI (r -0·69; P<0·0001) under physiological conditions. An inverse linear regression was also shown between SR-BI and the percentage of myristic acid in the diet (r -0·75; P<0·0001). The hepatic mass of SR-BI in the M group had increased at the end of the post-period compared with the test-period values. The present investigation shows that myristic acid modulates HDL-cholesterol via a regulation of the SR-BI expression.

We aimed to improve the use of the human intestinal Caco-2 cell line for studying dietary lipid and cholesterol processing by using isolated pure clones (). Three clones (TC7, PD7 and PF11) were grown as monolayers on semi-permeable filters and compared for cell viability, fatty acid and cholesterol apical uptake or basolateral secretion, apolipoprotein B-48 basolateral secretion and 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase activity. The TC7 clone showed the best viability upon apical incubation with mixed micelles and should be preferred for routine work. Short-term (3·0 h) rates of apical uptake of cholesterol were not different with the three clones, whereas the rate of apical uptake of oleic acid (18 : 1) was lower (P<0·05) with PF11 (250·6 nmol/mg) and the basolateral secretion of cholesterol and oleic acid was lower with the TC7 clone (0·40 and 29·1 nmol/mg respectively). The secretion of apolipoprotein B-48 basolaterally was about 2-fold lower than from PD7 clone. The basal levels of HMG-CoA reductase activity were significantly different (P<0·05; TC7>PF11>PD7). The down-regulation of the enzyme activity was moderate (range 13·8–21·0 %) and comparable in the presence of apical micellar cholesterol, but was much marked upon basolateral incubation with LDL (range 34·0–53·6 %), especially for the PD7 clone. In conclusion, the Caco-2 clones characterized here proved to be particularly suitable for studying lipid nutrients processing. Because these three clones exhibit some different metabolic capabilities, they provide a new tool to study intestinal response to lipid nutrients.

The effect of maternal phylloquinone supplementation on vitamin K in breast milk was studied to establish: (1) if phylloquinone is the source of menaquinone-4 in breast milk; (2) the dose–effect relationship between intake and obtainable levels. Four groups of lactating mothers with a full-term healthy infant participated and took oral phylloquinone supplements of 0·0 (n 8), 0·8 (n 8), 2·0 (n 8), and 4·0 (n 7) mg/d for 12 d, starting at day 4 post-partum. Milk samples were collected on days 4, 8, 16, and 19. Blood samples were collected on days 4 and 16. Vitamin K and vitamin E concentrations, the latter for reason of comparison, were assayed. Phylloquinone and menaquinone-4 were present in all milk samples: 5·84 (SD 2·31) AND 2·98 (sd 1·51) nmol/l (n 31) respectively, in colostrum (day 4 sample). A strong correlation between the vitamers was found (r 0·78, P<0·001). Breast-milk phylloquinone levels were raised in a dose-dependent manner: 4-, 12-, and 30-fold on day 16 for the 0·8, 2·0, and 4·0 mg group respectively. In addition, menaquinone-4 levels were higher: 2·5- (P<0·05) and 7-fold (P<0·001) in the 2·0 and 4·0 mg groups respectively. Plasma of supplemented subjects contained 3-, 5-, and 10-fold higher phylloquinone levels on day 16. Detectable menaquinone-4 was found in ten of thirty-one day 4 plasma samples. All day 16 plasma samples of the 4 mg supplemented group contained the vitamin. There was no correlation between the K-vitamers in plasma. Vitamin E and phylloquinone appear to differ in their distribution in breast milk, milk:plasma concentration ratios were ≤1 and 3–5 for vitamin E and phylloquinone respectively. The milk:plasma concentration ratio of menaquinone-4 was >10. In conclusion, dietary phylloquinone is a source of menaquinone-4 in breast milk. Phylloquinone supplementation to lactating mothers may be of benefit to the newborn infant, since both phylloquinone and menaquinone-4 are raised by supplementation.

4-Desmethylsterols and -stanols reduce plasma total cholesterol (TC) and LDL cholesterol by inhibition of intestinal cholesterol absorption, while the cholesterol-lowering potential of 4,4′-dimethylsterols is less well defined. The present study aimed to compare the effects of 4-desmethylsterols, -stanols, and 4,4′-dimethylsterols on plasma and hepatic cholesterol, sterol excretion and bile acid metabolism. Male golden Syrian hamsters were fed diets containing 13 g/100 g fat, 0·08 g/100 g cholesterol and 0 (control), 0·24 or 0·48 % (w/w) esterified 4-desmethylsterols (sterols) and esterified hydrogenated 4-desmethylsterols (stanols) from common vegetable oils or esterified 4,4′-dimethylsterols from rice bran oil for 5 weeks. Sterol and stanol esters at the dose of 0·24 % were equally effective and significantly (P<0·05) lowered TC by 15 %, while 0·24 % 4,4-dimethylsterols reduced TC by 10 %. Liver total and esterified cholesterol concentrations were significantly (P<0·05) lowered by 40, 22, 43 and 31 % in hamsters fed 0·48 % sterols, 0·24 % stanols, 0·48 % stanols or 0·48 % dimethylsterols, respectively. Daily faecal bile acid excretion and hepatic cholesterol 7α-hydroxylase activity were not altered, indicating that sterols, stanols and dimethylsterols had no effect on the intestinal re-absorption of bile acids or on hepatic bile acid synthesis. Daily excretion of cholesterol was significantly higher in hamsters fed esterified sterols and stanols, but was only slightly increased in those fed dimethylsterols. The results indicate that esterified sterols and stanols were equally effective in lowering plasma TC and LDL cholesterol, while dimethylsterol esters caused a weaker cholesterol-lowering effect. Sterols and stanols achieve their cholesterol-lowering effect by stimulating faecal cholesterol excretion through inhibiting intestinal cholesterol absorption, but do not affect bile acid excretion. Other mechanisms need to be considered to explain the effect on plasma and hepatic cholesterol of dimethylsterols.

Ca may interfere with fat and cholesterol metabolism through formation of insoluble soaps with fatty and bile acids in the intestine. In the present study, we examined the effects of different dietary Ca levels on the serum lipid profile and cholesterol metabolism in obese Zucker rats fed a low-fat diet. We also tested whether dietary Ca interfered with the lipid-lowering effects of a pine oil-derived plant sterol mixture. Increase in dietary Ca intake from 0·2 to 0·8 %, and further to 2·1 % (w/w) dose-dependently decreased serum total cholesterol (r -0·565, P=0·002, n 27), LDL-cholesterol (r -0·538, P=0·006, n 25), and triacylglycerol (r -0·484, P=0·014, n 25) concentrations, and increased HDL-cholesterol (r 0·478, P=0·016, n 25) and HDL : LDL cholesterol (r 0·672, P<0·001, n 25) in rats fed a 1 % cholesterol diet. Analysis of serum campesterol : cholesterol and sitosterol : cholesterol suggested that Ca dose-dependently increased intestinal cholesterol absorption (r 0·913, P<0·001, n 18), whereas serum desmosterol : cholesterol and lathosterol : cholesterol indicated that Ca dose-dependently increased endogenous cholesterol synthesis (r 0·691, P=0·003, n 18). Therefore, the decrease of serum LDL-cholesterol appeared to be due to Ca-induced increase in the conversion of cholesterol to bile acids. The increase in Ca intake did not interfere with the beneficial effects of plant sterols on serum total cholesterol, LDL-cholesterol and HDL-cholesterol concentrations. The high-Ca diet with plant sterol supplementation further increased the HDL-cholesterol concentration and HDL : LDL cholesterol. The present findings indicate that the beneficial effects of dietary Ca on the serum lipid profile during a low-fat diet are dose-dependent, and resemble those of bile acid sequestrants. Increased dietary Ca did not impede the lipid-lowering effects of natural plant sterols.

The role of the placenta in controlling the supply of fatty acids to the fetus was investigated in term placentas from non-smokers (n 5), smokers (>ten cigarettes/d; n 5) and after addition of ethanol at 2 mg/ml (n 4). The maternal side was of the placenta was perfused ex vivo for 90 min with a physiological mixture of fatty acids and fatty acid:human albumin ratio. There was no effect of smoking on the transfer of linoleic (LA, 18 : 2 n-6), α-linolenic (αLN, 18 : 3 n-3), arachidonic (AA, 20 : 4 n-6) or docosahexaenoic acid (DHA, 22 : 6 n-3), expressed per perfused area (calculated from H218O exchange). However, the presence of ethanol in the perfusate at a concentration of 2 mg/ml significantly reduced (P<0·01) the absolute rate of transfer of the two n-3 polyunsaturated fatty acids, αLN and DHA. This specific effect of ethanol on αLN and DHA also resulted in an altered selectivity for transfer of individual fatty acids. In the non-smoking control group the placenta selectively transferred polyunsaturated fatty acids to the fetus in the order DHA>AA>αLN>LA. The order of selectivity was unaltered in placentas from smokers, but the addition of ethanol to the perfusion medium altered the order of selectivity to AA>αLN>LA>DHA. The presence of ethanol in the perfusate was also associated with a significant reduction (P<0·05) in the clearance of H218O. These results suggest that the presence of ethanol at a concentration of 2 mg/ml may reduce the availability of polyunsaturated fatty acids to the developing fetus.

The present study was designed to compare the conjugated linoleic acid (CLA) isomeric distribution pattern in the liver of suckling rats in relation to those in the milk and maternal diet. Silver-ion HPLC was used to separate individual CLA isomers. It was found that the isomeric distribution pattern in the milk was very similar to that in the maternal dietary fat. However, the CLA isomeric distribution patterns in the liver phospholipids (PL) and triacylglycerols were different from those in the diet and milk. In the liver PL, total cis/trans isomers accounted for 63·6–63·9 % of total CLA, which was in contrast to the values of 88·1–89·1 % in the milk and diet. In the liver PL, total trans/trans isomers were 20·6–20·8 % of the total CLA isomers whereas they were only 2·6–3·7 % in the milk and diet. It is concluded that trans/trans-CLA were preferentially incorporated into the liver whereas for the incorporation of cis/trans-CLA there was partial discrimination.

Dietary trials with subjects on a freely selected diet may be affected by unwanted behavioural changes. Few studies, if any, have examined changes in coffee consumption and possible concomitant changes in diet and health-related habits. The aim of the present study was to examine whether induced changes in coffee consumption lead to changes in food habits and leisure-time physical activity. Healthy, non-smoking coffee-drinkers (n 214) were asked to change their coffee habits in a controlled clinical trial on the metabolic effects of coffee. The participants were asked to maintain their usual dietary habits. Self-perceived changes in diet and physical activity during the 6-week intervention period were assessed at the end. In the analyses, the participants were rearranged into groups reflecting the difference in coffee intake during the trial as compared with habitual intake. Associations with changes in food intake or physical activity were analysed by Spearman rank correlation. Changes in intake of ‘chocolate, sweets’ (r 0·179, P<0·05), ‘cakes, sweet biscuits, pastry’ (r 0·306, P<0·001), and ‘jam’ (r 0·198, P<0·05) showed positive associations with change in coffee intake during the trial. Negative associations were found for ‘dishes with fish’ (r -0·204, P<0·01) and many of the drinks as well as with physical activity (r -0·164, P<0·05). Induced changes in coffee intake seem to alter ad libitum intake of several foods. The recognized associations between health behaviours may have physiological explanations.

Although bone metabolism is largely under genetic control, the role of nutrition is considerable. The present study evaluates the effects of chicken eggshell powder, a new source of dietary Ca, and purified CaCO3 on bone mineral density (BMD) of the lumbar spine and hip. Besides BMD we also looked at biochemical markers of bone and Ca metabolism. Both Ca sources were provided in combination with minerals and vitamins including Mg, cholecalciferol and phylloquinone. We designed a randomised, double-blind, placebo-controlled study to take place over 12 months. Healthy Caucasian women (n 85), selected by age (≥50 and <70 years), from the databases of general practitioners were recruited by telephone calls. They had to be at least 5 years post-menopausal, with lumbar spine T-score being >-2·5. At baseline, their mean habitual daily Ca intake was adequate. The women were randomly allocated to: eggshell powder-enriched (group A; n 24), purified CaCO3-enriched (group B; n 22), or a placebo product (group C; n 27). BMD was measured at baseline and then after 6 and 12 months of supplementation as were the biochemical markers bone-specific alkaline phosphatase, amino-terminal propeptide extension of type I collagen, deoxypyridinoline, calcitonin, intact parathyroid hormone, calcidiol, and urinary Ca. After 12 months of supplementation, only mean BMD of the femoral neck in group A was significantly increased (P=0·014) by 1·75 % (95 % CI 0·18, 3·32) compared with a decrease of -0·60 % (95 % CI -1·92, 0·72) in group C. This increase coincided with significant decreases in markers of bone resorption and formation. No significant changes were seen in BMD at other sites, including lumbar spine, nor in groups B and C. No differences were found between groups A and B, or B and C. The present study indicates that healthy late post-menopausal women with an adequate Ca intake at baseline may increase BMD of the hip within 12 months following supplementation with the chicken eggshell powder-enriched supplement.