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Background Inflammation plays an important role in the pathogenesis of coronary artery disease. We examined whether dietary supplementation with alpha-linolenic acid (ALA, 18:3n-3) affects the levels of inflammatory markers in dyslipidaemic patients. Methods We recruited 76 male dyslipidaemic patients (mean age=51+/-8 years) following a typical Greek diet. They were randomly assigned either to 15 ml of linseed oil (rich in ALA) per day (n=50) or to 15 ml of safflower oil (rich in linoleic acid (LA, 18:2n-6)) per day (n=26). The ratio of n-6:n-3 in linseed oil supplemented group was 1.3:1 and in safflower oil supplemented group 13.2:1. Dietary intervention lasted for 3 months. Blood lipids and C-reactive protein (CRP), serum amyloid A (SAA), and interleukin-6 (IL-6) levels were determined prior and after intervention. CRP and SAA were measured by nephelometry and IL-6 by immunoassay. Results Dietary supplementation with ALA decreased significantly CRP, SAA and IL-6 levels. The median decrease of CRP was 38% (1.24 vs. 0.93 mg/l, P=0.0008), of SAA 23.1% (3.24 vs. 2.39 mg/l, P=0.0001) and of IL-6 10.5% (2.18 vs. 1.7 pg/ml, P=0.01). The decrease of inflammatory markers was independent of lipid changes. Dietary supplementation with LA did not affect significantly CRP, SAA and IL-6 concentrations but decreased cholesterol levels. Conclusions Dietary supplementation with ALA for 3 months decreases significantly CRP, SAA and IL-6 levels in dyslipidaemic patients. This anti-inflammatory effect may provide a possible additional mechanism for the beneficial effect of plant n-3 polyunsaturated fatty acids in primary and secondary prevention of coronary artery disease.

The purpose of this study was to examine the effects of various dietary fats on low density lipoprotein (LDL) binding in an in vitro system where receptor number is not regulated. Cynomolgus monkeys were fed diets containing 37% of energy from fat, with various degrees of saturation, and 0.4 mg/kcal cholesterol or low-fat (13% of energy), low cholesterol (0.03 mg/kcal) chow. Plasma LDL was isolated after 16 weeks. The fatty acid composition of LDL showed enrichment corresponding to the dietary fats consumed, and the high fat, high cholesterol diets produced marked hypercholesterolemia compared to chow feeding. Of those fed the high fat diets, monkeys fed the fish oil diet had the highest LDL cholesterol concentrations, 13.25 +/- 0.77 mmol/l, while those fed the safflower oil diet had the lowest, 7.51 +/- 3.31. LDL from chow fed monkeys had the lowest binding affinity; the Kd was 26.2 +/- 8.7 microg/ml, nearly twice that of the high fat diets (P = 0.003). No significant differences in binding were found between the different high fat diets, although there was a trend toward lower affinity in the diets enriched in polyunsaturated fat. LDL size was affected by diet with chow fed monkeys having the smallest average LDL, 259.3 +/- 1.7 A compared to the other groups (P = 0.03). Monkeys fed the fish oil diet tended to have smaller LDL, but this was not significantly different from the other high fat diets. Binding affinity was correlated with LDL size, r = 0.54, P0.01. LDL composition, as measured by apo B/cholesterol ratio, was altered by feeding a high fat, high cholesterol diet. The ratio was reduced in the LDL samples from monkeys fed the high fat diets compared to those fed chow, but this ratio was not significantly correlated with binding. Thus, it appears that increasing dietary fat and cholesterol intake increases LDL size and binding affinity, such that LDL metabolism may be altered independently from effects on receptor number; the type of dietary fat does not seem to influence this process when fat and cholesterol content is very high.

Department of Human Biology, Maastricht University, Netherlands. G.Hornstra@HB.Unimaas.NLTo study the influence of dietary fatty acids on arterial thrombosis tendency 65 groups of male rats were fed diets containing 50% of their digestible energy as fat from 32 different oils and fats. After 8 weeks their arterial thrombosis tendency was assessed by measuring the obstruction time (OT) of a loop-shaped polythene cannula inserted into the abdominal aorta. Using multiple regression analysis log10 OT was modelled as a function of the relative amounts of the various dietary fatty acids and their combinations. The best fit (R2 = 0.79) was obtained for the sums of all monoenoic and (n-6) and (n-3) polyenoic fatty acids, which appeared antithrombotic. The fit for the sum of all saturated fatty acids, which had a prothrombotic effect, was almost as good (R2 = 0.76). The ratio between dietary polyunsaturated and saturated fatty acids (P:S ratio) appeared a strong predictor of arterial thrombosis tendency (R2 = 0.77). Marine oils did not have a more powerful antithrombotic effect than could be expected on the basis of their P:S ratios. Using stepwise regression analysis myristic acid, 14:0, was shown to be the strongest prothrombotic fatty acid whereas linoleic acid, 18:2(n-6), was the strongest antithrombotic fatty acid. Since the number of marine oils was very limited the effects of the 'fish fatty acids' eicosapentaenoic acid, 20:5(n-3) and docosahexaenoic acid, 22:6(n-3), on arterial thrombus formation could not be tested reliably. The same appeared true for gamma-linolenic acid, 18:3(n-6), and stearidonic acid, 18:4(n-3), present in a few vegetable oils only.

Oxidation of LDL-linoleic acid (LDL-LA), a major substrate for lipid peroxidation, may be counteracted by the antioxidant vitamin E. In a 3-month randomized double-blind placebo-controlled trial in 83 apparently healthy Dutch elderly, aged 67-85 years, the direct protective effect of 100 IU vitamin E on the rate of oxidized LDL-LA was studied. The oxidation of LDL-LA was measured by its disappearance after a 5-h in vitro Cu-oxidation of LDL isolated from 1 ml plasma. In the vitamin E group, the decrease in oxidized LDL LA of 10.4, (p < 0.05) was significantly different (p < 0.05) from the smaller 4.6% p< 0.01) decrease in the control group. Moreover, within the vitamin E group the decrease was even more marked over tertiles of alpha-tocopherol to LDL-LA ratio with a significant difference in decrease (p < 0.05) from the lowest compared to the highest tertile of, respectively, 18.4% [-24; -2%] (median and range) and 2.0% [-16: 34%]. In conclusion, supplementation with 100 IU vitamin E in elderly is beneficial in lowering the rate of oxidation of LDL LA. The protective effect of vitamin E might best be monitored by using the ratio of alpha-tocopherol to LDL-LA as this reflects the degree of alpha-tocopherol available to protect the amount of LDL-LA present.

The oxidative modification of human low density lipoprotein (LDL) has been widely investigated. However, there are no data concerning the oxidation susceptibility of combined very low density lipoprotein (VLDL), intermediate density lipoprotein (IDL) and low density lipoprotein fraction, although all of them are atherogenic and contain antioxidants such as alpha-tocopherol. We investigated the oxidation susceptibility and oxidation resistance of VLDL + LDL (including IDL) fraction by induction with CuCl2 and its relation to plasma alpha-tocopherol concentration and lipid standardised alpha-tocopherol concentration in 406 non-vitamin E-supplemented men from eastern Finland. Even thought we did not give oral vitamin E or any other antioxidant supplementation to our study participants, we observed a significant, consistent relationship between measurements of oxidation resistance and plasma content of vitamin E. In the multivariate regression model, a high plasma content of vitamin E or lipid standardised vitamin E concentration were the most important determinants of lag time to maximal oxidation rate (standardised regression coefficient = 0.244, P < 0.0001 for vitamin E and 0.211, P < 0.0001 for lipid standardised vitamin E). After statistical adjustment for age, use of cigarettes, hypolipidemic medication (yes vs. no), month of the measurements, plasma concentrations of total ascorbic acid (ascorbic acid + dehydroascorbic acid), beta-carotene and phospholipids, serum concentrations of LDL cholesterol and triglycerides and dietary intake of linoleic acid, the lag time to maximal oxidation rate was 10% (95% C.I. 6.0-13.5%) longer in men in the highest fifth than in the lowest fifth of plasma vitamin E content (P < 0.0001 for trend). When the fifths of lipid standardised vitamin E were compared, the lag time to maximal oxidation rate was 6% (95% C.I. 1.8-10.1%) longer in men in the highest than in the lowest fifth (P < 0.0001 for trend). Our data suggest that alpha-tocopherol is an important antioxidant preventing the in vitro oxidation of VLDL + LDL fraction even in non-supplemented subjects.

The present study demonstrates for the first time that iron ions can induce lipid peroxidation in intact macrophages without causing cell death. Macrophage lipid peroxidation increases cell-mediated oxidation of LDL, enhances the release of interleukin 1 and inhibits the release of apolipoprotein E from the macrophages. When cultured macrophages were exposed to ferrous ions (50 microM FeSO4) for 4 h at 37 degrees C, cellular lipid peroxidation (measured by analyses of malondialdehyde (MDA), conjugated dienes (CD), and lipid peroxides (PD)) increased 2-4-fold in comparison with non-treated cells. This process was iron-dose dependent, reached its maximum after 4 h of incubation, and was accompanied by 68% and 53% reductions in the content of the cellular linoleic (18:2), and arachidonic acid (20:4), respectively, and by 29% and 36% reductions of cellular vitamin E and vitamin A, respectively. Cell viability (measured by trypan blue exclusion, by [3H]thymidine incorporation into DNA, by analysis of the release of lactate dehydrogenase (LDH) or [3H]adenine), and cell morphology (studied by scanning electron microscopy) were not significantly affected by the iron-induced oxidative stress. Manitol and dimethylthiourea (DMTU), but not catalase or superoxide dismutase (SOD), significantly inhibited iron-induced cellular lipid peroxide formation, suggesting that hydroxyl radical, but not superoxides or hydrogen peroxides, mediated the iron-induced cellular lipid peroxidation. Incubation of LDL (0.2 mg of protein/ml) with oxidized macrophages resulted in LDL lipids peroxidation, as evidenced by an 8-fold increase in the LDL associated MDA in comparison with LDL that was incubated under similar conditions with non-oxidized macrophages. Furthermore, oxidation of LDL by oxidized macrophages in the presence of copper ions (10 microM CuSO4) was 2-fold higher in comparison with oxidation of LDL by non-oxidized macrophages. The release of apolipoprotein E from oxidized macrophages decreased by 50%, whereas macrophage release of beta-glucuronidase and of interleukin-1 beta increased by 83% and by a factor of 6, respectively. This study demonstrates for the first time that iron ions induce oxidation of the cellular polyunsaturated fatty acids in intact macrophages and that this cellular lipid peroxidation can subsequently induce LDL oxidation.

Eicosapentaenoic acid, which is one of the n-3 polyunsaturated fatty acids (PUFA), is reported to exert its antithrombotic and anti-atherogenic effect partly through the modulation of vascular cell functions. Vascular smooth muscle cell (VSMC) proliferation plays an important role in the pathogenesis of atherosclerosis. We reported the differential effect of various PUFA on VSMC proliferation. First we established a method for preparing PUFA rich cells in culture to mimic the in vivo situation using PUFA triacylglycerol emulsion. Using these fatty acid rich cells, we found that only EPA and docosahexaenoic acid, although less potent than EPA, inhibited the proliferation of VSMC among the fatty acids tested. This effect of EPA was reversed by the addition of anti-oxidants. It is suggested that production of the oxidized species at a low concentration from EPA inhibited the proliferation of VSMC. This anti-proliferative effect of EPA and DHA on VSMC could partly explain the anti-atherosclerotic effect of marine lipids.

Certain fatty acids induce changes in endothelial barrier function which may be mediated by alterations in normal proteoglycan synthesis/metabolism. To test this hypothesis, pulmonary artery derived endothelial cells were treated with media supplemented with linoleic acid (18:2), and/or a known proteoglycan synthesis inhibitor, beta-D-xyloside. Independent exposure to 1 mM beta-D-xyloside or 90 microM 18:2 increased albumin transfer, i.e., decreased barrier function, when compared with control cultures. 18:2 and beta-D-xyloside increased albumin transfer additively, suggesting that the mechanisms by which 18:2 and beta-D-xyloside alter the proteoglycan metabolism are different. Compared with the control group, treatment with 18:2 inhibited proteoglycan synthesis, decreased anionic properties of heparan sulfate proteoglycans in the cell monolayers and caused the release of a unique chondroitin sulfate proteoglycan into the culture media. Treatment with beta-D-xyloside caused an increased incorporation of radioactive sulfate into glycosaminoglycans but inhibited proteoglycan synthesis. These results suggest that the fatty acid- and beta-D-xyloside-induced impairment in endothelial barrier function may involve changes in the synthesis, release and physicochemical properties of proteoglycans.

Fatty acid composition of lipid classes and NMR spectra of lipoproteins were compared in 6 young (24-35-year-old) and 6 elderly (79-90-year-old) women. Cholesteryl ester, triglyceride and protein content of LDL in elderly women were significantly higher (+52-57% and +20% for lipids and proteins, respectively) than those observed in young women. HDL lipid levels were similar in the two groups. The proportion of linoleic acid (mainly in cholesteryl esters and phospholipids) of each lipoprotein species was always lower in octogenarians when compared with young females (lowering of 13-28% and 27-46% for cholesteryl esters and phospholipids, respectively). Conversely, the proportions of mono-unsaturated fatty acids (mainly oleic acid) increased in all lipid classes, although this was only significant in cholesteryl esters from each lipoprotein species. NMR spectra of lipoproteins showed a restricted mobility of acyl chain terminal CH3 groups in old women which was significant only in VLDL and HDL3. This suggests that the decrease of linoleic acid could affect the lipid mobility in lipoproteins of elderly women.

Aims Imaging mass spectrometry (IMS) enables the visualization of individual molecules present on tissue sections. We attempted to identify and visualize specific markers for aortic atherosclerotic lesions. Methods and results Atherosclerotic lesions were obtained from aortic roots of apolipoprotein E (ApoE)-deficient mice at 60 weeks of age and from femoral arteries of humans with peripheral artery occlusive disease. IMS was performed with a matrix-assisted laser desorption/ionization mass spectrometry time-of-flight (TOF)/TOF-type instrument. The molecular ions at m / z 671.6 and 673.6 were found to be specific molecules in the mouse and human lipid-rich regions. These molecules were assigned as cholesterol linoleate (CE 18:2) and cholesterol oleate (CE 18:1). In the case of the human samples, triacylglycerol was also localized in the lipid-rich regions. The distributions of the molecular ions at m / z 804.5 and 832.5 were the same as the distribution of both the mouse and the human SMCs. These molecules were assigned as phosphatidylcholine (PC) (diacyl 16:0/20:4) and PC (diacyl 18:0/20:4). The molecular ion at m / z 566.9 was localized in the mouse calcified regions, and the molecular ions at m / z 539.0 were localized in the human calcified regions. Conclusions The IMS-based histopathologic examination (IbHE) revealed the characteristic peaks of lipid-rich regions, SMCs, and calcified regions in the atherosclerotic lesions. In addition, IbHE revealed the characteristic distribution of lipids in human atherosclerotic lesions. These data indicate that an IMS-based pathologic approach is of considerable value as a new histopathologic examination.

The effect of linoleic acid hydroperoxide on replication of adenovirus type 2 DNA in cultured endothelial cells of bovine aorta was studied to elucidate the deleterious effect of lipid peroxides on replication of DNA in the arterial endothelial cells. When, just after the infection, the cells were incubated with 1 nmol (measured by the thiobarbituric acid reaction and expressed in terms of malondialdehyde of the hydroperoxide, the replication of adenovirus type 2 DNA was markedly reduced. The level of the messenger RNA for adenovirus type 2 DNA polymerase was also reduced. In accordance with the reduction of the messenger RNA, the synthesis of the polymerase in the cells was reduced. The inhibition of DNA replication in the arterial endothelial cells by lipid peroxides was discussed in relation to atherogenesis.

Compared to cholesterol or linoleic acid (18:2), oxidized lipids such as cholestan-3 beta, 5 alpha, 6 beta-triol (triol) and hydroperoxy linoleic acid (HPODE) markedly impair endothelial barrier function in culture [Hennig and Boissonneault, 1987; Hennig et al. 1986]. Because proteoglycans contribute to vascular permeability properties, the effects of cholesterol and 18:2 and their oxidation products, triol and HPODE, on endothelial proteoglycan metabolism were determined. While cholesterol was without effect, a concentration-dependent decrease in cellular proteoglycans (measured by 35S incorporation) was observed after exposure to triol. Compared to control cultures, cholesterol reduced mRNA levels for the proteoglycans, perlecan and biglycan. Triol had a similar effect on biglycan but not an perlecan mRNA levels. Compared to 18:2, 1,3 and 5 microM HPODE depressed cellular proteoglycans. Perlecan mRNA levels were reduced more by HPODE when compared to 18:2. Biglycan mRNA levels were reduced by 3 microM, but not by 5 microM HPODE. These data demonstrate that oxidized lipids such as triol and HPODE can decrease cellular proteoglycan metabolism in endothelial monolayers and alter mRNA levels of major specific proteoglycans in a concentration-dependent manner. This may have implications in lipid-mediated disruption of endothelial barrier function and atherosclerosis.

We investigated the differences between the hypocholesterolemic effects induced by dietary linoleic acid and those induced by oleic acid in hamsters. Addition of 5% linoleic acid or oleic acid to a 0.1% cholesterol-supplemented diet diminished the increases in plasma total and low density lipoprotein (LDL) cholesterol induced by cholesterol alone. Linoleic acid decreased high density lipoprotein (HDL) cholesterol in comparison with cholesterol alone, whereas oleic acid did not. As compared with a standard diet or a cholesterol-supplemented diet, linoleic acid and oleic acid each prevented hepatic LDL receptor suppression, although linoleic acid was more effective. Oleic acid prevented the increase in plasma cholesteryl ester transfer protein (CETP) activity induced by dietary cholesterol, whereas linoleic acid did not. Neither linoleic acid nor oleic acid altered hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase activity. Only oleic acid increased hepatic cholesterol 7 alpha-hydroxylase activity. These results suggest that dietary linoleic and oleic acids diminish the cholesterol-induced increases in plasma total and LDL-cholesterol by preventing hepatic LDL receptor suppression, and in the case of oleic acid by also preventing the increase in the plasma CETP activity. These effects on cholesterol 7 alpha-hydroxylase activity may influence bile lipid metabolism.

Conjugated linoleic acid (CLA) consists of a series of positional and geometric dienoic isomers of linoleic acid that occur naturally in foods. CLA exhibits antioxidant activity in vitro and in vivo. To assess the effect of CLA on atherosclerosis, 12 rabbits were fed a semi-synthetic diet containing 14% fat and 0.1% cholesterol for 22 weeks. For 6 of these rabbits, the diet was augmented with CLA (0.5 g CLA/rabbit per day). Blood samples were taken monthly for lipid analysis. By 12 weeks total and LDL cholesterol and triglycerides were markedly lower in the CLA-fed group. Interestingly, the LDL cholesterol to HDL cholesterol ratio and total cholesterol to HDL cholesterol ratio were significantly reduced in CLA-fed rabbits. Examination of the aortas of CLA-fed rabbits showed less atherosclerosis.

The fatty acid composition of adipose tissue from 4114 men and women across 22 districts of Scotland is described. A biopsy of subcutaneous adipose tissue was obtained from the upper arm using a skin biopsy punch. Overall the proportion of saturated fatty acids was lower in women and those of monounsaturated and polyunsaturated higher than in men. There were significant effects of age on adipose tissue fatty acid composition, particularly in women, where the proportion of saturated fatty acids decreased and that of monounsaturated fatty acids increased with advancing age. The fatty acid composition of adipose tissue varied among the districts studied. The district standardised mortality ratio for coronary heart disease was positively correlated with the district mean oleic acid value and inversely correlated with the district mean linoleic acid value. This paper supports the importance of adipose tissue fatty acids as indicators of risk factor status for coronary heart disease.

Intraindividual comparisons of diets supplemented with sunflowerseed oil (rich in linoleic acid, LA, C18:2n-6), linseed oil (enriched with alpha-linolenic acid, LNA, C18:3n-3) and canned mackerel (rich in eicosapentaenoic acid, EPA, C20:5n-3 and docosahexaenoic acid, DHA, C22:6n-3) were made in 30 patients with primary hyperlipoproteinemia (HLP) of phenotypes IIa (n = 9), IIb (n = 7), IV (n = 7) and V (n = 7). The lipid- and blood pressure-lowering effects of polyunsaturated fatty acids (PUFA), particularly those of the EPA- and DHA-rich diet, were confirmed irrespective of the type of HLP. Apolipoproteins A-I and B remained unchanged. The most remarkable finding was a substantial depression of free fatty acids (FFA) within a standardized glucose tolerance test (GTT) associated with the fall of serum triglycerides after diets enriched with n-6 and especially after those supplemented with n-3 PUFA. It was suggested that the decrease of FFA indicates reduced peripheral lipolysis, which might be a hitherto ignored factor involved in the triglyceride-lowering action of n-6 and, more pronounced, of n-3 PUFA.

Low levels of essential polyunsaturated fatty acids of the n-6 series are associated with coronary heart disease. Linoleic acid, but not gamma-linolenic acid requires the activity of delta 6-desaturase for its conversion to dihomo-gamma-linolenic and arachidonic acid. Evening primrose oil (EPO) and safflower oil (SO) are rich in linoleic acid, but EPO contains also 9% gamma-linolenic acid. The effect of EPO (10, 20 and 30 ml/day) and SO (20 ml/day) for 4 months on the deposition of linoleic acid metabolites in adipose tissue of 4 groups of 6-9 men with low adipose dihomo-gamma-linolenic acid was examined. EPO but not SO increased adipose dihomo-gamma-linolenic acid level from 0.080 +/- 0.005% to 0.101 +/- 0.005% (P less than 0.01; 20 ml/day for 4 months). Adipose dihomo-gamma-linolenic/linoleic acid ratio increased with EPO from 0.99 +/- 0.16 X 10(2) to 1.13 +/- 0.14 X 10(2) and fell on SO from 1.04 +/- 0.10 X 10(2) to 0.90 +/- 0.07 X 10(2) (P less than 0.01). Similar qualitative changes in the relative amount of dihomo-gamma-linolenic acid in serum triglyceride and cholesteryl ester fractions were observed. At the dose of 20 ml/day, SO and EPO did not differ in their effect on serum cholesterol (7.13 +/- 0.43 vs. 7.33 +/- 0.42 mmol/l (NS)), LDL-cholesterol (5.10 +/- 0.32 vs. 4.88 +/- 0.46 mmol/l (NS)) nor did the 2 oils differ in their effect on HDL-cholesterol. These results suggest that linoleic acid is not readily converted to dihomo-gamma-linolenic acid due to a low activity of delta 6-desaturase in these highly selected men. EPO was not an effective hypocholesterolaemic agent in this study.

We studied the effects of addition of physiological concentrations (0.5 mM) of fatty acids i.e., palmitic (16:0), stearic (18:0), oleic (18:1) and linoleic acid (18:2) on lipoprotein secretion by polarized Caco-2 cells. With saturated fatty acids, secreted lipoproteins were at IDL/LDL density, 1.009 < d < 1.068 g/ml. The numbers of secreted lipoproteins, expressed as secreted apolipoprotein (apo) B, were comparable to control without fatty acid (palmitic acid, 551 +/- 185; stearic acid, 629 +/- 304 and control, 504 +/- 176 ng apo B/4.7 cm2 filter). With unsaturated fatty acids, apo B containing lipoproteins were secreted at chylomicron/VLDL density (d < 1.006 g/ml). Oleic acid caused a two-fold higher secretion of apo B than control (1058 +/- 87 vs. 504 +/- 176 ng/4.7 cm2 filter, P < 0.001). The increase in apo B secretion was attributed to a specific increase in apo B-48. Unsaturated fatty acid caused a two-fold higher secretion of triglyceride than saturated fatty acids but incorporation of newly synthesized lipid into the secreted lipoproteins, measured by incorporation of a fatty acid marker, was 10- to 20-fold higher, indicating preferential translocation of unsaturated triglycerides into lipoproteins. Mixtures rich in either polyunsaturated, monounsaturated, or saturated fatty acids, resembling nutritional fat and oils, were capable of a two-fold stimulation of secretion of apo B containing triglyceride-rich lipoproteins. The triglyceride/apo B ratio in the basolateral medium was higher with the monounsaturated 'olive oil' mixture (12 250 +/- 2000 mol/mol) than with the polyunsaturated 'corn oil' mixture (7830 +/- 2480 mol/mol) and incorporation of newly synthesized lipid into the secreted lipoproteins was 1.5-fold higher as well. In conclusion, unsaturated fatty acids were most potent in stimulating the secretion of apo B by specifically increasing apo B-48 secretion. Unsaturated triglycerides, that contain mainly oleic acid, were more efficiently incorporated into lipoproteins than saturated triglycerides, suggesting preferential translocation by microsomal triglyceride transfer protein.

Serum lipids and lipoproteins were investigated in 10 young quadriplegic patients. All received intermittently a liquid formula diet via a naso-gastric tube, consisting of 2900 calories/day. The polyunsaturated/saturated fatty acids ratio was 1.1 : 1. Their mean body weight was 70% of the ideal body weight. The results were compared to a control group receiving a diet containing 2600 calories per day, and the polyunsaturated/saturated fatty acids ratio was 0.9 : 1. Their mean body weight was 95% of the ideal body weight. In the quadriplegic group the serum HDL-cholesterol levels and linoleic acid content of cholesterol ester were reduced. Similar findings have been observed in patients with coronary disease.

Although the stated requirement for linoleic acid in humans is less than 2% of the dietary calories, recently there has been considerable emphasis on the necessity to substitute dietary polyunsaturates for saturates in order to reduce serum cholesterol levels. In this study we have sought to determine the nutritional consequences of feeding a very high level of linoleate to rats. Three groups of thirty adult animals each were fed a semipurified diet consisting by weight of casein 17%; mineral mixture 5.5%; vitamin mixture in glucose 2.2%; cellulose fiber 3.0%; and corn oil 0% (group A), 10% (group B) or 40% (group C), which was provided at the expense of glucose. At the end of four weeks on the diets, blood was obtained in the fasting state from 16 rats in each group. The serum was ultracentrifugally fractionated into six classes of lipoproteins and analyzed for lipid composition and protein content. Disc gel electrophoresis using lipid and protein stains established that the various lipoprotein subclasses were reasonably free of adjacent density fractions. Although the total serum cholesterol levels were practically the same in the three groups, the cholesterol moiety of the major low density lipoproteins, LDL2 (d 1.019–1.050), but not of very low density lipoproteins, VLDL (d 1.006) or low density lipoproteins, LDL1 (d 1.006–1.019), was substantially and very significantly increased in rats fed the high level of corn oil as compared to the other groups. The concentration of the very low density lipoproteins was significantly lower in group C than in the groups A and B. The LDL2 concentration but not that of LDL1 was significantly greater in group C as compared to group A. The cholesterol/total lipid ratio was significantly greater in both LDL2 and LDL1 but not in VLDL of group C as compared with group A. The serum high density lipoproteins were relatively less influenced by the ingestion of an excessive level of corn oil at this time period. The serum lipoprotein levels as well as their lipid composition were generally similar in groups A and B and suggested that a moderate level (5%) of dietary linoleate did not cause any untoward changes in rats. On the basis of current information on the metabolism of lipoproteins, it has been proposed that the increase in rat serum LDL2 of group C reflects the status of the liver and that a large portion of the cholesterol moiety of LDL2 is perhaps derived from the liver while the protein and phospholipid portions may represent remnants of VLDL catabolism. In view of the magnitude of the changes observed in LDL2-cholesterol as well as in the liver cholesterol and triglycerides due to the ingestion of a 40% corn oil diet in a usually resistant species, namely the rat, further work along these lines with other species including human and nonhuman primates merits our attention.

An atherogenic diet (AD) consisting entirely of normal foods for westernised people was fed to female Vervet monkeys for 4 years. The plasma low density lipoprotein (LDL) cholesterol pool was increased and progression of atherosclerosis was enhanced by the AD compared to a more prudent Western diet. The increased LDL-cholesterol was carried by a 3-fold increase in particles of relatively normal composition and not by packing cholesterol esters into the cores of enlarged LDL particles, as has been reported after feeding semisynthetic diets loaded with extra cholesterol. Nevertheless, these LDL particles were atherogenic. The AD changed the fatty acid composition of LDL-cholesterol esters and triacylglycerol, notably by increasing arachidonic and reducing linoleic acid. Multivariate analysis showed that measures and scores of atherosclerosis were significantly dependent on sphingomyelin and phosphatidylcholine in LDL and on arachidonic acid in LDL-triacylglycerol. Although apolipoprotein B, free cholesterol, esterified cholesterol and lysophosphatidylcholine in plasma LDL and atherosclerosis were significantly positively correlated in bivariate analysis they were not selected by multivariate analysis as the strongest determinants of atherogenesis. Cholesterol in plasma high density lipoprotein was not changed by the AD and lecithin:cholesterol acyltransferase activity in plasma was inversely linked to atherosclerosis. Subcutaneous fatty acids reflected dietary fatty acids.

This study examines the effect of prostaglandin E2 and 6-keto F1 alpha on the cholesteryl ester metabolism of cells grown in tissue culture. When 3T3 mouse fibroblasts were incubated with cationized low density lipoprotein (LDL) and 3H-labelled oleic and 14C-labelled linoleic acids a marked increase in cholesteryl ester content of cells was observed. Oleic acid was the preferred substrate for cholesterol esterification. However, the presence of prostaglandin E2 or 6-keto F1 alpha (up to 10 micrograms/ml) did not affect the cholesteryl ester content or the uptake of labelled fatty acids into cellular lipids. Following preincubation with cationized LDL and labelled fatty acids the cells were reincubated in normal medium with or without prostaglandins. The presence of PGE2 or 6KPGF1 alpha (up to 10 micrograms/ml) did not appreciably change the rate of removal of cholesteryl ester or labelled lipids. This indicates that these prostaglandins even when present in relatively large doses in the incubation medium do not affect lipid metabolism of cells grown in tissue culture.

Linoleate hydroperoxide administered to rats in a total dose of 353 mg by repeated subcutaneous injections increased the incidence of myocardial fibrosis. When administered subcutaneously to rabbits in doses of 700–1000 mg, it produced myocardial fibrosis and aortic lesions. Neither linoleate nor the degradation products of linoleate hydroperoxide produced these cardiovascular lesions in rabbits when given by repeated subcutaneous injections in a total dose of 750 mg. The aortic lesions in rabbits given injections of linoleate hydroperoxide were associated with an increased peroxide value in the vessel wall but were not attributable to raised levels of cholesterol or β-lipoprotein in the serum. Lipid peroxides did not accumulate in the heart of either species after administration of linoleate hydroperoxide, although the lipid peroxide content of the liver was raised. The feeding of cholesterol to rabbits was followed by the typical cardiovascular lesions and by an elevation of serum cholesterol and β-lipoprotein levels. There was no increase in the peroxide value of the aorta or heart in these rabbits.

The fatty acid compositions of 4 serum lipid fractions were analysed from 244 randomly selected 30-59-year-old Finnish men from 4 areas involved in a population survey ('Mini-Finland') in 1979-80. Men in eastern Finland had significantly lower mean percentages of linoleate (18:2) in CE, TG, FFA and PL (45.1, 10.3, 9.3 and 18.8%, respectively) than men in the western part of the country (48.4, 12.5, 10.6 and 20.2%, respectively). Very low values of 18:2 were encountered in the North Karelian community of Ilomantsi, especially in men aged 50-59 (40.9, 8.0, 7.5 and 16.8%, respectively). The percentage of alpha-linolenate tended also to be lower and those of saturated and monounsaturated fatty acids higher in the east, but there were no or only inconsistent differences in the contents of the prostaglandin precursors dihomo-gamma-linolenate, arachidonate and eicosapentaenoate. Eighteen men were studied in November and the following April. Only minor changes in the mean composition of serum fatty acids took place during this period and the correlation coefficients between the percentages of 18:2 recorded at the two time points ranged from 0.70 to 0.81. The low concentrations of 18:2 in serum lipids in Finnish men obviously reflect a low dietary P/S ratio and may contribute to the high prevalence of IHD in Finland and to its regional differences.

Fifteen volunteers were treated daily during a week with 250 mg of sodium linoleate supply as suppository. A day before and a day after treatment venous blood samples were drawn, high density lipoproteins (HDL) fractionated and 3-sn phosphatidylcholine isolated by thin-layer chromatography from total serum and HDL and their fatty acid composition analyzed by gas-liquid chromatography. The positional esterification of linoleic acid in the phosphatidylcholine molecule was determined by enzymatic hydrolysis. Increases of linoleic acid of 1.8–136.3% in the serum phosphatidylcholine were observed in 14 out of 15 volunteers and an increment of 6.3–32.2% was also detected in the lipid fraction from HDL. In both fractions about 98% of the linoleic acid is located in the C 2 position of the phosphatidylcholine molecule. The results reported in the present paper show that it is possible to promote the enrichment of linoleic acid in the phosphatidylcholine fractions of both serum and HDL from humans subjected to low doses of sodium linoleate supply as suppository.

Four-day primary coverglass cultures of rat heart muscle (M) and endothelioid (E) cells were treated for 30 min with stearic, oleic or linoleic acids in a FFA/albumin ratio of 6:1 at concentrations from 5 × 10 −6 M to 5 × 10 −4 M . Labilization of lysosomes and mitochondria was measured by acid phosphatase and succinic dehydrogenase staining activity respectively. Stearate or linoleate, but not oleate, labilized M cell lysosomes at 5 × 10 −6 M . Lysosomes of E cells were not significantly affected by any of the FFA at 5 × 10 −4 M . The order of activity of these FFA for M cell lysosomes was linoleate=stearate > oleate. Both E and M cell mitochondria were significantly labilized by oleate or linoleate, 5 × 10 −6 M , and by stearate, 5 × 10 −5 M . The order of activity was linoleate > oleate > stearate. Treatment of cultures for 24 hr with 50 μg/ml of hydrocortisone before the FFA at 5 × 10 −5 M provided significant protection only against stearate-induced lysosomal labilization.

Lecithin : cholesterol acyl transfer (LCAT) and relative fatty acid composition of serum lecithin and cholesterol esters were studied in 20 young male survivors of myocardial infarction (MI). Comparisons were made with controls matched for serum cholesterol. There was no difference in LCAT rate between MI patients and controls. The relative content of arachidonic acid in cholesterol esters was higher in MI patients. The fatty acid composition of lecithin and cholesterol esters suggests an equal transfer of linoleic and oleic acids from lecithin to cholesterol. Furthermore negative correlations were found between LCAT and linoleic acid content of lecithin (r = --0.43, P less than 0.01) and cholesterol esters (r = --0.45, P less than 0.01). This inverse relationship does not seem to be linked to substrate specificity, but rather to be mediated by influences in common on serum lipid content and turnover.

Summary Lipid deposits have been identified histologically in various tendons and fascia in ageing man; the tendons at the body surface contained more lipid deposits than did those in more protected sites. The earliest deposits were seen at the age of 15 years. Histochemical and chemical investigation showed that the predominant lipids were cholesterol esters. The fatty acid pattern in these esters strongly resembled that in the cholesterol esters from fatty and fibrofatty atherosclerotic lesions. Both patterns resembled that in plasma, except that linoleate was significantly lower. Histological assessment of the severity of lipid deposition in the Achilles' tendon showed no correlation with deposition in the aorta, but did correlate with the extent of lipid deposition in the coronary artery. The amount of Achilles' lipid did not, however, correlate with the occurrence of myocardial infarction. Avascularity, “wear-and-tear” and binding by acidic mucosubstances are briefly discussed in relation to lipoprotein-binding by connective-tissue.

1.(1) When isocaloric diets high in saturated fat or polyunsaturated fat were alternately fed to hyperlipidaemic and normal men, serum triglyceride was 35% lower (P 2.(2) The magnitude of the reduction of both serum triglyceride and cholesterol due to the unsaturated fat was greatest in subjects with highest baseline levels. 3.(3) The reduction of triglyceride levels during the unsaturated fat period was due mainly to a change in the very low density lipoprotein (VLDL) triglyceride. The fall in cholesterol, however, was due to changes both in VLDL and low density lipoprotein (LDL) cholesterol levels. The triglyceride/cholesterol ratio of VLDL was significantly lowered by the unsaturated fat diet, but the ratio of triglyceride to apoproteins A, B and C was not significantly altered. 4.(4) Postprandial serum triglyceride levels were higher during saturated fat feeding than when unsaturated fat was fed. The difference was due to higher fasting levels, the increment due to alimentary lipaemia being similar during the two diets. 5.(5) There was no change in post-heparin lipolytic activity (PHLA) nor in fractional removal rate of triglyceride as measured by the intravenous fat tolerance test (IVFTT); these indices of triglyceride clearance from plasma suggested that the fall of triglyceride concentration induced by polyunsaturated fat could not be accounted for by enhanced uptake from plasma. 6.(6) Albumin-bound [9, 10- 3 H]palmitic acid and [1- 14 C]linoleic acid were simultaneously infused, and their relative rates of incorporation into very low density lipoprotein triglyceride (VLDL-TG) was measured in six normolipidaemic men. The results suggested that the saturated fatty acid was preferentially incorporated, and may imply that dietary polyunsaturated fat decreases the rate of secretion of VLDL-TG into plasma.

Summary o (1) Analysis of the sera of more than 100 wild baboons, Papio ursinus , revealed that the cholesteryl esters had the following fatty acid composition: 22.7 palmitic, 5.8 palmitoleic, 10.4 stearic, 24.8 oleic, 31.7 linoleic, 1.0 linolenic, and 5.0% arachidonic acids. The total unsaturated cholesteryl fatty acids comprised 68.4% of the total cholesteryl esters. (2) The average fatty acid composition of the phospholipids was as follows: 29.1 palmitic, 3.1 palmitoleic, 21.5 stearic, 16.9 oleic, 20.0 linoleic, 1.1 linolenic and 7.2 arachidonic acids. The total phospholipid contained 48.3% unsaturated acids. (3) The fatty acid composition of the triglycerides was the following, i.e. 34.7 palmitic, 7.5 palmitoleic, 11.2 stearic, 32.0 oleic, 14.3 linoleic, 1.3 linolenic and 2.5% arachidonic acids. (4) Very low density lipoproteins of baboon sera contained a higher proportion of the saturated fatty acids in cholesteryl esters, phospholipids and triglycerides than the high density lipoproteins. (5) In baboon sera the high density lipoprotein fractions contained about 55%, 59% and 80% of the total cholesterol, fatty acids, and phospholipids, respectively. Of the total triglyceride content of baboon sera, about 64% was found in the very low density fraction.

Cholesterol loading of diabetic rats is known to induce marked hyperlipoproteinaemia, and we have reported that enhancement of the activity of intestinal acyl-CoA:cholesterol acyltransferase (ACAT), one of the key enzymes involved in cholesterol absorption, might play an important role in the development of hypercholesterolaemia in these animals. In the present study, we have shown that treatment with N -(α-methylbenzyl)linoleamide (melinamide), a new hypocholesterolaemic drug, caused a substantial decrease of the enhanced intestinal ACAT activity in diabetic rats, but did not affect intestinal cholesterol esterase activity. Furthermore, marked improvement of hypercholesterolaemia in cholesterol-fed diabetic rats occurred concomitantly with the drug treatment. These results suggest that intestinal ACAT activity is closely related to the serum cholesterol level in diabetic rats, and show that melinamide lowers intestinal ACAT activity.

The fatty acid composition of the plasma lipid esters has been studied during lipid-lowering treatment of 95 patients with atherosclerotic disease. During the first two months of the trial only a diet was prescribed. During the ensuing two months either clofibrate or niceritrol, a nicotinic acid ester, was added in a randomized order. During the last two months the second drug was added. The combined treatment with diet, clofibrate and niceritrol caused highly significant serum lipid reductions. The fatty acid composition in the plasma lipid esters was determined in samples from each trial period to measure the degree of dietary adherence. During dietary treatment the relative content of saturated and monounsaturated ratty acids decreased and the polyunsaturated fatty acids increased with an increasing ratio between polyunsaturated and saturated fatty acids (P/S ratio) in the cholesterol esters and triglycerides. Only minor changes were seen in the phospholipids. The changes caused by the diet were partly reversed by clofibrate while niceritrol did not cause any major changes of the fatty acid composition. Clofibrate treatment coincided with increasing amounts of monounsaturated fatty acids, especially oleate (18 : 1), in the cholesterol esters, triglycerides and phospholipids while there were significant reductions of the content of linoleic (18 : 2) acid in both the cholesterol esters and triglycerides. The 18 : 2/18 : 1 ratio decreased significantly in all the lipid esters analyzed. However, the P/S ratio was not significantly affected, partly because the relative content of saturated fatty acids also tended to decrease during clofibrate braatment.. It is concluded that addition of clofibrate treatment to patients who are on a diet enriched with polyunsaturated fats is associated with a change from polyunsaturated to monounsaturated fatty acids in the plasma lipid esters but does not significantly affect the ratio between polyunsaturated and saturated fatty acids. The fatty acid changes caused by clofibrate treatment are counteracted by an increased amount of polyunsaturated fat in the diet.

Male rabbits (10 weeks old) were fed, for 20 weeks, purified diets rich in fat (45% of calories) containing saturated fats (butter), polyunsaturated fats (evening primrose oil + butter or sunflower oil + butter) for 20 weeks. Linoleic acid represented respectively 3.6, 33 and 34% of the dietary fatty acids, while gamma-linolenic acid was present (4.4%) solely in the second group. A significant increase in di-homo-gamma-linolenic acid in plasma, platelets and aorta was noted only in the animals fed evening primrose oil. Despite this, the results of the platelet aggregation to thrombin and ADP, the recalcification plasma-clotting time (platelet-rich plasma) and the severity of atherosclerosis were not significantly different from those observed in the group fed sunflower oil. In contrast, in comparison to the butter-fed animals, the two groups fed the polyunsaturated fats showed remarkable improvements in the clotting time (P less than 0.01) and in the severity of atherosclerotic lesions (evening primrose oil P less than 0.001; sunflower oil P less than 0.05). However, the response to thrombin-induced aggregation was significantly decreased (P less than 0.05) only in the evening primrose oil-fed animals. In these long-term studies in young rabbits, dietary gamma-linolenic acid did not seem to have marked beneficial effects, additional to those induced by linoleic acid, on platelet functions or on atherosclerosis.

The fatty acid patterns of serum triglycerides and FFA in SHR and in normotensive controls aged 4, 8 and 26 weeks were estimated by gas-liquid chromatography. In serum triglycerides of SHR, the percentage of linoleic acid (C 18:2 ) was lower and the content of arachidonic acid (C 20:4 ) higher than in age-matched control animals. A continuous increase in palmitic (C 16 ) and linoleic acids as well as a decrease in arachidonic acid has been found with advancing age, the most striking differences existing between 4- and 8-week-old animals, i.e. before onset of arterial hypertension in SHR. In the pre-hypertensive stage, the percentage of arachidonic acid was about 3 times as high as in later stages in SHR. This gradation was, however, even more pronounced in normotensive control rats. The C18:2C20:4-ratio of triglycerides was lower in SHR but increased with age in both groups reaching a 5–8-fold level. Similar behavior of the FFA fatty acid pattern was less marked. Alterations in levels of linoleic and arachidonic acids are of interest because of their pathogenic role as precursors of prostaglandins in the development of genetically spontaneous hypertension in rats. The results are discussed in connection with the hypotensive effect of a linoleic acid-rich diet recently reported in hypertensive rats.

The ultrastructural effects of a single brief intra-arterial infusion of palmitic, linoleic and acetoacetic acid on the arterial endothelium of the rat were investigated, and the following results obtained: (1) Palmitic acid, infused at a concentration of 4 mM/l, damaged the arterial lining by producing large cytoplasmic clefts and occasional blebbing and lysis of the endothelial cells. By contrast, linoleic acid, infused at the same concentration, had no damaging effects on arterial endothelium. (2) Acetoacetic acid damaged the arterial wall when infused at concentrations of 0.2 mM/l or higher by inducing extreme swelling and loss of cristae of the mitochondria in arterial endothelium and myocytes. The above results raise the possibility that (a) high saturated fatty acid diets may promote atherosclerosis not only by inducing hypercholesterolemia but also by injuring the arterial lining, and (b) diabetes may promote atherosclerosis not only by inducing hyperlipemia but also by damaging the arterial wall during periods of uncontrolled ketoacidosis.

A mixture of doubly labelled cholesterol linolenate and non-radioactive cholesterollinoleate was injected subcutaneously in rats. The lipid implants were separated into predominantly extracellular and intracellular phases by progressive solvent extractions. Chromatography and scintillation counting of the fractions showed that little esterified cholesterol had been hydrolysed in the intracellular phase and little randomization of fatty acids had occurred between radioactive and non-radioactive cholesterol. It is concluded that foam cells (macrophages) can take up cholesterol in esterified form and that preliminary hydrolysis to free cholesterol is not obligatory for uptake by these cells.

The fatty acid composition of adipose tissue from normal, atherosclerotic and diabetic men and women was studied. We found a difference in the fatty acid composition between both sexes. A decrease in linoleic acid and an increase in palmitic and stearic acids in adipose tissue from atherosclerotic men was shown, while there was an increase in palmitic acid in adipose tissue from atherosclerotic women. An increase in oleic acid was found in diabetic men. This alteration in the fatty acid content of adipose tissue may be related to some modification in the metabolism of this tissue due either to atherosclerosis or to diabetes which cause an alteration of the lipid metabolism.

1. (1) Seven healthy normolipaemic men aged 28–46 years (mean 33 years) were studied for four weeks on their usual diet, the mean polyunsaturated to saturated fat (P : S) ratio of which was approximately 0.23 and the cholesterol content 667 mg/day, and for four weeks on an isocaloric diet, containing similar proportions of protein, carbohydrate and fat, but with a mean P : S ratio of approximately 2.82 and a cholesterol content of 112 mg/day. 2. (2) Total serum cholesterol levels decreased by 18.7 ± 5.2% (mean ± SEM) on the polyunsaturated fat (PUF) diet. This was attributable to a similar reduction in serum low density lipoprotein (LDL) cholesterol. Serum very low density lipoprotein (VLDL) cholesterol showed no clear change. Total serum triglyceride levels decreased by 19.7 ± 7.1% due to a reduction in both serum VLDL and LDL triglyceride concentration. 3. (3) Total serum apolipoprotein B concentration fell by 23 ± 2.6% during the PUF diet. There was a decrease of similar magnitude in serum LDL apolipoprotein B, but no sustained effect on serum VLDL apolipoprotein B. 4. (4) Serum VLDL and LDL lipid to apolipoprotein B ratios did not change significantly during the study. 5. (5) Serum triglyceride fatty acid composition significantly altered during the PUF diet. The percentage of linoleic acid rose from 16 ± 0.90% to 36 ± 0.89%, whilst the percentage of combined saturated and monounsaturated acids fell correspondingly. These changes took place within 2 days of starting the PUF diet.

An organ culture technique was used to investigate the removal of fatty acids incorporated into various lipid fractions in intimal segments of atherosclerotic rabbit aorta. Segments of aorta were pulse-labelled in vitro with [9,10- 3 H]-oleic acid and [1- 14 C]linoleic acid for 24 h. The incubation medium was replaced with non-radioactive medium and the removal of the endogenously labelled phospholipid, triglyceride and cholesterol ester was followed for a further period of 10 days. Fatty acids incorporated into phospholipid and triglyceride were removed rapidly from the aortic intima, with fractional half-times of 4–5 days. Cholesterol ester fatty acids were removed more slowly from the aortic intima with fractional half-times of 9–12 days. When the removal rate of oleic acid incorporated into phospholipid, triglyceride and cholesterol ester was compared with that of linoleic acid incorporated into the same lipid fractions, no difference between removal of the two fatty acids could be detected with respect to any of the 3 lipid fractions.

Groups of metabolically normal (controls) and alloxan-diabetic adult female rabbits were fed semi-synthetic diets containing 40 cal % palm-kernel oil (PKO) or sunflower-seed oil (SSO) for 54 weeks. In contrast to control rabbits fed PKO-diet, the alloxan-diabetic rabbits on this diet, developed no or only a negligible degree of atherosclerosis, although the serum levels of all lipid classes had increased in the diabetic rabbits above that of the controls during almost the whole experimental period. The diabetic rabbits and the controls fed SSO-diet were both free from any significant atherosclerotic involvement in spite of the fact that the SSO-diet appeared unable to suppress the very high levels of the various serum lipid classes induced by the diabetic state. On both diets, the diabetic rabbits showed a significantly higher cholesteryl linoleate/oleate ratio than the controls, which was caused by an increase in the cholesteryl linoleate level in the diabetics. No serious aorta atherosclerosis was found in rabbits with a cholesteryl linoleate/oleate ratio higher than 0.6, although no correlation was found between the atherosclerosis indices and these ratios at values lower than 0.6. Rabbits with cholesteryl linoleate/oleate ratios below 0.6 seemed to run a greater risk of developing atherosclerosis. It is suggested that insulin might be required for atherogenesis in addition to hyperlipemia and hypercholesterolemia.

Cholesterol acyltransferase and cholesterol esterase activities of protein extracts from pig aortas have been examined and the rations of synthesis/hydrolysis rates in the presence of substrates with different fatty acids estimated. The values obtained were in the numerical order: cholesteryl oleate greater than palmitate greater than or equal to linoleate greater than linolenate greater than staerate. The results are discussed in relation to the known different accumulation of cholesterol esters in the arterial wall.

The incorporation of 3 H-labelled oleic acid and of 14 C-labelled linoleic acid into phospholipid, triglyceride and cholesterol ester in smooth muscle cells grown in incubation medium supplemented with either 5% normal or 5% hyperlipemic serum has been studied. Both fatty acids were incorporated into cholesterol esters to a greater extent when cells grown in incubation medium containing hyperlipemic serum. Oleic acid was incorporated into cholesterol esters in preference to linoleic acid. The addition of hyperlipemic serum to the incubation medium did not increase the incorporation of either 3 H-labelled oleic acid or of 14 C-labelled linoleic acid into phospholipid or triglyceride. The removal of labelled lipid fractions has also been followed for four days in cells pulse labelled for 24 hours with 3H-labelled oleic acid and 14 C-labelled linoleic acid. Both 3 H- and 14 C-labelled cholesterol esters were removed more rapidly when the smooth muscle cells were grown in medium containing normal serum than in medium containing hyperlipemic serum. The removal of both phospholipid and triglyceride was similar in normal and hyperlipemic serum. Comparison of the 3H14C ratio indicated that the cholesterol oleate and cholesterol linoleate were removed at similar rates.

Zinc deficiency mimics many of the clinical features of essential fatty acid (EFA) deficiency in rats. Since zinc appears to be needed for the Δ-6-desaturase step in EFA metabolism, experiments were conducted to determine whether bypassing this step with γ-linolenic acid (18 : 3ω6) would alleviate the biological effects induced by a zinc-deficient diet. In pair-fed rats over a period of 5 weeks the deficient diet impaired growth and changed the relative weights of internal organs. It also induced hypolipidemia but had little effect on the fatty acid composition of tissue lipids. Daily subcutaneous injection of primrose oil containing 10% 18: 3ω6 reversed most of the effects of zinc deficiency on tissue weights, growth and plasma lipids. In contrast, injection of safflower oil, which has a similar content of linoleic acid (18 : 2ω6) but is devoid of 18 : 3ω6, had only a partial effect on some tissue weight changes. Neither oil affected the plasma fatty acid pattern, but both of them increased liver triglyceride concentrations. They also elevated the proportion of 18 : 2ω6 in liver, skin and epididymal fat. The latter effects were not observed in the absence of zinc deficiency. Supplementing the diet of EFA-deficient animals with an excess of zinc in their drinking water did not affect the typical tissue fatty-acid pattern of EFA deficiency. It is suggested that several of the manifestations of zinc deficiency are mediated by a relative state of EFA deficiency attributable, at least in part, to a reduced conversion of 18 : 2ω6 to 18 : 3ω6 resulting in an accumulation of 18 : 2ω6 in tissues. These findings are consistent with a role of zinc as a cofactor in the Δ-6-desaturase enzyme reaction. The suggested role of zinc in essential fatty acid metabolism may be of significance in understanding how essential fatty acids protect against cardiovascular disease in general.

Dietary linoleate was effective to increase LDL fluidity in adult men but did not significantly influence VLDL or HDL fluidities. Lipoproteins were isolated ultracentrifugally from plasma of sixteen healthy, free living male volunteers consuming controlled diets formulated from typical U.S.A. foods to have 35 energy % fat with 10 g (diet L) or 30 g (diet H) linoleate per day, 30–50 g saturated fatty acids/day and the balance mainly monounsaturated fatty acids. Calculated cholesterol intakes were 500 mg/day at each calorie level. Changes in LDL fluidity were detected as differences in diphenylhexatriene (DPH) fluorescence polarization upon crossover between the two controlled diets. Thermotropic measurement of DPH fluorescence anisotropy and compositional analyses indicated that LDL and HDL fluidities were dependent upon phospholipid and triacylglycerol concentrations, respectively, and were modulated by the presence of cholesteryl esters. Fatty acid analyses of the major lipid classes of the isolated lipoproteins indicated that changes, upon diet crossover, in DPH fluorescence anisotropy, were a linear function of the incremental change in LDL phospholipid linoleate. The fluorescent probe described an environment corresponding to the fatty acyl moieties of the phospholipids on the LDL periphery, which composition is apparently under dietary control. It is suggested that the diet induced fluidity changes may affect the conformation of the apoprotein moiety on the LDL surface and thus the potential for LDL interaction with cellular LDL receptors.

The fatty acid composition of the cholesteryl esters in serum was measured in 7- and 8-year-old boys in groups from 16 countries. The ratio of esterified cholesterol : total cholesterol was also measured. All sample collections and analyses were carried out under standardized conditions. The proportion of palmitic acid in the cholesteryl esters was high in the groups from Asia and Africa (0.17–0.26) compared with that in the groups from the U.S.A. and Europe (0.14–0.18). The proportion of linoleic acid in the cholesteryl esters was low in the groups from Asia and Africa (0.39–0.48) and high in the groups from the U.S.A. and Europe (0.45–0.58). The proportion of oleic acid, arachidonic acid, palmitoleic acid and stearic acid showed little variation between the groups. The proportion of linoleic acid in the cholesteryl esters was positively correlated with the concentration of total cholesterol (r = 0.75, n = 26, P < 0.005).

Fatty acids of plasma lipids, red cells and platelets were analyzed from 38 demented patients (age 53–88 years), comprising 11 patients with Alzheimer's disease (AD), 19 with multi-infarct dementia (MID) and 8 with probable vascular dementia (PVD). The mean age, body mass index, duration of dementia and content of triglycerides, total cholesterol and HDL-cholesterol in plasma were similar in AD and MID. The patients with PVD were older. As compared to AD, in MID and PVD the linoleic acid (LA) and other n − 6 and n − 3 polyunsaturated fatty acids (PUFA) were significantly lower in red cells and tended to be lower also in serum triglycerides, cholesterol esters (CHE) and phospholipids (PL), and platelets. The LA content of red cells was significantly correlated with that of serum CHE and PL, and n − 6 PUFA (including arachidonic acid) of red cells. The low LA content of red cells was associated with old age, coronary heart disease and heart failure, but not with the severity of dementia.

The effect of linoleic acid on the induction of fatal ventricular fibrillation by intravenous CaCl2 (10%), was studied in rats fed for a month from weaning on a diet with either a high or low content of linoleic acid. Studies were performed in the basal state and after pretreatment with noradrenaline, which increased the sensitivity to CaCl2 equally in animals from both diet groups. Despite considerable differences in the linoleic acid levels in the plasma and myocardium, the two groups did not differ in the incidence of fatal ventricular fibrillation. Our conclusions concerning the effect of linoleic acid on cardiac arrhythmias, and sudden death in particular, are compared with those from other studies.

Serum and aortic lipids and atherosclerotic lesions were studied in five spider monkeys (Ateles sp.) and three rhesus monkeys ( Macaca mulatta ) after changing from a commercial chow diet to a high fat (40% of calories), high cholesterol (1 mg/cal), semisynthetic diet containing egg yolk and butter. There was a large species difference in response of serum cholesterol. After 12 weeks on the diet the mean elevation of serum cholesterol was 285 mg/100 ml for the rhesus and 17 mg/100 ml for the spider monkeys. Changes in serum cholesterol ester fatty acid patterns were also very much greater for the rhesus than for the spider monkey. Aortas and coronary arteries of the rhesus were extensively involved with sudanophilic lesions. The cholesterol and cholesterol ester content of the aortic intima was much greater in the rhesus than in the spider monkey. The proportional contribution of aortic cholesterol oleate was greater in rhesus than spider monkeys and the palmitate was lower. There were only a few sudanophilic lesions in the coronary arteries of the spider monkeys. The coronary arteries of one older rhesus monkey were extensively involved with fibrous lesions. There was no appreciable difference between species in the histologic appearance of the sudanophilic lesions.