Fatty acid specificity in the inhibition of cell proliferation and its relationship to lipid peroxidation and prostaglandin biosynthesis

Primary cultures of smooth muscle cells were established from the medial layer of guinea pig aorta. Cells at passage level 4 were treated with different series of fatty acids belonging to the n-9, n-6 and n-3 families. Lipid peroxidation was measured by the thiobarbituric acid assay and prostaglandin biosynthesis was measured by the radioimmunoassay of PGE and 6-keto-PGF_1α. Cell proliferation was estimated from the total cell number of cultures seeded at low density. 18∶1(n-9) did not form lipid peroxides and this fatty acid stimulated cell proliferation. All fatty acids which generated lipid peroxides inhibited cell proliferation, but inhibition was correlated with the degree of lipid peroxidation only in the n-9 fatty acid family. 22∶4(n-6) and 22∶6(n-3) inhibited prostaglandin biosynthesis. 18∶2(n-6), 18∶2(n-9), 18∶3(n-3), 20∶2(n-9), 20∶3(n-3) and 20∶5(n-3) had no effect on prostaglandin biosynthesis. 18∶3(n-6), 20∶3(n-6) and 20∶4(n-6) generated prostaglandins. 20∶3(n-9) generated metabolites with prostaglandin immunoreactivity. The inhibition of cell proliferation did not correlate with enhanced or inhibited prostaglandin synthesis. The inhibition of cell proliferation was related to the structures of the different polyunsaturated fatty acid families decreasing in the order n-9>n-6>n-3. Eicosatrienoic acids were the most effective inhibitors of cell proliferation in each fatty acid family and 20∶3(n-9) was the most potent eicosatrienoic acid. These data show that specific as yet unrecognized products of fatty acid metabolism are responsible for the inhibition of cell proliferation. Fatty acids are designated by the number of carbon atoms: number of double bonds and the position of the first double bond from the methyl terminus of the acyl chain is noted in parenthesis: 18∶1(n-9), 9-octadecenoic acid; 18∶2(n-9), 6,9-octadecadienoic acid; 18∶2(n-6), 9,12-octadecadienoic acid; 18∶3(n-6), 6,9,12-octadecatrienoic acid, 18∶3(n-3), 9,12,15-octadecatrienoic acid; 20∶2(n-9), 8,11-eicosadienoic acid; 20∶3(n-9), 5,8,11-eicosatrienoic acid; 20∶3(n-6), 8,11,-14-eicosatrienoic acid, 20∶4(n-6), 5,8,11,14-eicosatetraenoic acid; 20∶5(n-3), 5,8,11,14,17-eicosapentaenoic acid; 22∶4-(n-6), 7,10,13,16-docosatetraenoic acid, 22∶6(n-3), 4,7,10,13,16,19-docosahexaenoic acid. Presented at the 73rd AOCS annual meeting, Toronto, Canada, May 1982.     

Lipid composition and prostaglandin synthesis in mouse lung microsomes: Alterations following the ingestion of menhaden oil

Three groups of male mice were fed a normal diet or a semisynthetic diet containing either 10% hydrogenated coconut oil (CO group) or 10% menhaden oil (MO group) for two wk. The synthetic diet altered the fatty acid composition of lung microsomal lipids. Mice ingesting menhaden oil contained greater amounts of eicosapentaenoic acid (20∶5 n−3), docosapentaenoic acid (22∶5 n−3) and docosahexaenoic acids (22∶6 n−3) and decreased amounts of n−6 fatty acids such as arachidonic and adrenic. Synthesis of prostaglandin E₂ and prostaglandin F_2α from exogenous arachidonic acid was significantly depressed in n−3 fatty acid-enriched lung microsomes. These studies indicated that dietary fish oil not only alters the fatty acid composition of lung microsomes but also lowers the capacity of lungs to synthesize prostaglandins from arachidonic acid.     

Albumin-bound docosahexaenoic acid and collagen-induced human platelet reactivity

Anin vitro system designed to mimic the effect of various plasma nonesterified (polyunsaturated) fatty acids on platelet function and metabolism was employed. Human platelet aggregation induced by submaximal (1.8 μg/ml) collagen stimulation was significantly inhibited by 2 min preincubation with 20 μM albumin-bound docosahexaenoic acid (22∶6n−3) (DHA), but not by the other fatty acids tested. [³H]Phosphatidic acid (PA) formation, an indicator of phospholipase C activation following platelet stimulation, was moderately inhibited by eicosapentaenoic acid (20∶5n−3), 11,14,17-eicosatrienoic acid (20∶3n−3), dihomo-γ-linolenic acid (20∶3n−6), as well as DHA, but not by arachidonic acid (20∶4n−6); this inhibition of phospholipase C activation could not explain the differential effect of DHA on platelet aggregation. The decreased production of thromboxane A₂ (TxA₂), as assessed by [³H]12-hydroxy-5,8,10-heptadecatrienoic acid (HHT) formation, may account for the inhibition of collagen-induced aggregation by 20 μM DHA. Surprisingly, preincubation with 40 μM albumin-bound DHA, even though resulting in greater inhibition of collagen-induced aggregation, had less impact on HHT formation. A small but significant increase in [³H]prostaglandin D₂ (PGD₂) levels following 3-min collagen stimulation may have contributed to the greater antiaggregatory effect of 40 μM DHA. It is concluded that increased plasma nonesterified DHA may contribute to the dampened platelet activation and altered metabolism following fish oil supplementation of the diet.     

Incorporation of n−3 fatty acids of fish oil into tissue and serum lipids of ruminants

This study examines the biohydrogenation and utilization of the C₂₀ and C₂₂ polyenoic fatty acids in ruminants. Eicosapentaenoic (20∶5n−3) and docosahexaenoic (22∶6n−3) acids were not biohydrogenated to any significant extent by rumen microorganisms, whereas C₁₈ polyenoic fatty acids were extensively hydrogenated. The feeding of protected fish oil increased the proportion of 20∶5 from 1% to 13–18% and 22∶6 from 2% to 7–9% in serum lipids and there were reductions in the proportion of stearic (18∶0) and linoleic (18∶2) acids. The proportion of 20∶5 in muscle phospholipids (PL) increased from 1.5% to 14.7% and 22∶6 from 1.0% to 4.2%; these acids were not incorporated into muscle or adipose tissue triacylglycerols (TAG). In the total PL of muscle, the incorporated 20∶5 and 22∶6 substituted primarily for oleic (18∶1) and/or linoleic (18∶2) acid, and there was no consistent change in the porportion of arachidonic (20∶4) acid.     

The fatty acids ofEntomophthora coronata

The total lipids and fatty acid composition ofEntomophthora coronata were determined. The fungus was grown on a chemically defined medium and a chemically nondefined medium (Sabouraud dextrose yeast extract) for a period of 26 days. The organism contained from 16.2% to 44.6% total lipids depending upon the days of growth. The major fatty acids were 12∶0 (5.5–9.0%), 13∶0 (1.2–8.2%), 14∶0 (33.5–43.5%), 16∶0 (9.7–13.9%), 18∶1₉ (20.4–22.4%), and 18∶2_9,12 (3.5–10.5%). Lesser amounts of 15∶0, 16∶1, 16∶2, 17∶0, 18∶0, two other 18∶2 (both having conjugated double bonds), 18∶3_6,9,12, another 18∶3 (conjugated double bonds), 20∶3_8,11,14, 20∶4_5,8,11,14, another 20∶4 (conjugated double bonds), and 24∶1 acids were found. Trace amounts of 20∶0, 20∶1, 20∶2, 22∶0 and 24∶0 were also present. The relative percentage of most of the fatty acids did not vary appreciably with growth. However, 18∶2_9,12 and 20∶4_5,8,11,14 increased with age of the chemically defined culture. Peak E (18∶2, conjugated double bonds) increased and 13∶0 and 18∶3_6,9,12 decreased with age of the chemically nondefined culture. The fatty acids were predominately saturated (56.9–69.1%) and contained a high percentage of shorter chain fatty acids (C 12 to C 15). The fatty acids of the chemically defined culture were more unsaturated than the Sabouraud culture and the unsaturation increased with age of the culture.     

Formation of a thiamine disulfide complex with fatty acid: Mechanism of formation of the complex

The formation of complexes between thiamine disulfide (TDS) orO-acetyl thiamine disulfide (O-acetyl TDS) and fatty acid or fatty acid methyl ester in methanol has been studied by fluorescence quenching and¹³C NMR relaxation (T₁) measurements. The association constants (K-values) of TDS andO-acetyl TDS with fatty acids (from 11∶0 to 18∶0, and 18∶1, 18∶2, 18∶3 and 20∶4) and fatty acid methyl esters have been determined. These values do not depend on either the number of carbon atoms or the degree of unsaturation of the fatty acid. The K-values of TDS andO-acetyl TDS with fatty acid were 7.8 M⁻¹ and 5.1 M⁻¹, respectively. The K-values of TDS andO-acetyl TDS with fatty acid methyl ester were very small. These results show that the-OH moiety in TDS and the-COOH moiety in the fatty acid are necessary for formation of the complex     

Effect of dietary α-linolenic acid and its ratio to linoleic acid on platelet and plasma fatty acids and thrombogenesis

The effect of dietary α-linolenic acid (18∶3n−3) and its ratio to linoleic acid (18∶2n−6) on platelet and plasma phospholipid (PL) fatty acid patterns and prostanoid production were studied in normolipidemic men. The study consisted of two 42-d phases. Each was divided into a 6-d pre-experimental period, during which a mixed fat diet was fed, and two 18-d experimental periods, during which a mixture of sunflower and olive oil [low 18∶3n−3 content, high 18∶2/18∶3 ratio (LO-HI diet)], soybean oil (intermediate 18∶3n−3 content, intermediate 18∶2/18∶3 ratio), canola oil (intermediate 18∶3n−3 content, low 18∶2/18∶3 ratio) and a mixture of sunflower, olive and flax oil [high 18∶3n−3 content, low 18∶2/18∶3 ratio (HI-LO diet)] provided 77% of the fat (26% of the energy) in the diet. The 18∶3n−3 content and the 18∶2/18∶3 ratio of the experimental diets were: 0.8%, 27.4; 6.5%, 6.9; 6.6%, 3.0; and 13.4%, 2.7, respectively. There were appreciable differences in the fatty acid composition of platelet and plasma PLs. Nevertheless, 18∶1n−9, 18∶2n−6 and 18∶3n−3 levels in PL reflected the fatty acid composition of the diets, although very little 18∶3n−3 was incorporated into PL. Both the level of 18∶3n−3 in the diet and the 18∶2/18∶3 ratio were important in influencing the levels of longer chain n−3 fatty acid, especially 20∶5n−3, in platelet and plasma PL. Production of 6-keto-PGF_1α was significantly (P<0.05) higher following the HI-LO diet than the LO-HI diet although dietary fat source had no effect on bleeding time or thromboxane B₂ production. The present study showed that both the level of 18∶3n−3 in the diet and its ratio to 18∶2n−6 were important in influencing long-chain n−3 fatty acid levels in platelet and plasma PL and that prostanoid production coincided with the diet-induced differences in PL fatty acid patterns.     

Biosynthesis of polyunsaturated fatty acids in the developing brain: I. Metabolic transformations of intracranially administered 1-14C linolenic acid

Thirteen-day old rats were given intracranial injections of 1-¹⁴C linolenic acid (allcis 9,12,15 octa decatrienoic acid) and were sacrificed after 8 hr. Analysis of brain fatty acids showed that 16∶0, 18∶0, 18∶1, 18∶3, 20∶3, 20∶4, 20∶5, 22∶5, and 22∶6 were labeled. The total fatty acid methyl esters were separated into classes according to degree of unsaturation on a AgNO₃∶SiO₂ impregnated plate. The bands were scraped off and the eluted fatty acids were first analyzed by radiogas liquid chromatography and then subjected to reductive ozonolysis to determine double bond position. The saturated acids, 16∶0, and 18∶0, as well as the mono-unsaturated 18∶1, must have been formed from radioactive acetate produced by β oxidation of the injected linolenate. Among the polyunsaturated fatty acids, the triene fraction was characterized and identified as 18∶3 ε3 (Δ^9,12,15), the starting material, and 20∶3 ω3 (Δ^11,14,17); the tetraene fraction was identified as 20∶4 ω3 (Δ^8,11,14,17); the pentaene fraction was identified as 20∶5 ω3 (Δ^5,8,11,14,17) and 22∶5 ω3 (Δ^{7,10,13,16,19}); and, finally, the hexaene fraction was shown to be 22∶6 ω3 (Δ^{4,7,10,13,16,19}). The biosynthesis of these ω3 family fatty acids in the brain in situ is discussed.     

Comparative study of the blood pressure effects of four different vegetable fats on young,spontaneously hypertensive rats

Following the suckling period, four groups of male four-week-old spontaneously hypertensive rats (SHR) were fed semisynthetic diets with 14% (by weight) of either sunflower seed oil [46% 18∶2(n−6); linoleic acid (LA)-rich], linseed oil [62.5% 18∶3(n−3)+12.9% 18∶2(n−6); α-linolenic acid (LNA)-rich], evening primrose oil [9.2% 18∶3(n−6)+71% 18∶2(n−6); γ-linolenic acid (LNA)-rich] or hydrogenated palm kernel fat [1.5% 18∶2(n−6); polyunsaturated fatty acid (PUFA)-deficient], respectively, up to an age of 18 wk. All diets enriched with PUFA provoked an attenuation of hypertension development. The effect was lowest in the LA-rich group and highest in the γ-LNA-rich group. Differences in fatty acid composition of renal phospholipids between groups reflect the fatty acids present in the respective dietary fats. Renomedullary production of PGF_2α was significantly reduced in α-LNA-rich and slightly diminished in γ-LNA-rich fed rats. Aortic formation of 6-keto-PGF_1α and TXB₂ was increased in animals fed the γ-LNA-rich diet. Thus, the attenuation of hypertension development cannot be explained only by changes in prostanoid formation. Other mechanisms possibly involved should be pursued.     

Non-methylene-interrupted and ω4 dienoic fatty acids of the white shrimpPenaeus setiferus

The total lipid fatty acids from the white shrimpPenaeus setiferus were found to contain several unusual dienoic fatty acid species. These included two methylene-interrupted species: Δ11, 14-C_18∶2 (18∶2ω4) and δ13, 16-C_20∶2 (20∶2ω4). Also found were several non-methylene-interrupted dienoic fatty acids including δ7, 11 and Δ7, 13-C_20∶2, Δ7, 13-C_21∶2, Δ7, 13, Δ7, 15, Δ9, 13, Δ9, 15, and Δ7, 17-C_22∶2. Many minor C_20∶2 non-methylene-interrupted dienes were found but could not be unequivocally characterized.     

The effect of long-chain monoenes on prostaglandin E2 synthesis by rat skin

In order to ascertain whether the dermal lesions observed in male rats fed rapeseed oils are due to impaired prostaglandin biosynthesis, endogenous levels of prostaglandin E₂ (PGE₂) in skin and the capacity of this tissue to synthesize PGE₂ from arachidonic acid was investigated. Male Sprague-Dawley rats were fed from weaning for 8 weeks either a standard rat diet (chow) or semisynthetic diets containing 20% by weight of the following fat sources: corn oil; commercial lard; commercial lard to which was added 5.4% free erucic acid; rendered pig fat; or the following rapeseed oils:Brassica napus var. Zephyr;B. campestris var. Span;B. campestris var. Arlo (15%) and var. Echo (85%) designated HEAR (high erucic acid rapeseed). The long-chain monoene content (18∶1, 20∶1, and 22∶1) of the diets fed ranged from 30 to 71 mole % and that of skin from 27 to 74 mole %. A significant (P<0.01) correlation was found between the level of 18∶2n−6 in the diet and the endogenous PGE₂ levels in skin and the capacity of this tissue to synthesize PGE₂. No relationship was found between these two PGE₂ parameters and the level of erucic acid in the diet. The rate of turnover of PGE₂ appeared to be lower in rats fed rapeseed oil as evidenced by the relatively high endogenous PGE₂ levels when these oils were fed (96 to 105 μg/g). On the other hand, the lowest capacity for PGE₂ synthesis was found with skin from rats fed Zephyr rapeseed oil, rats which also had the most severe incidence of hair loss and dermal lesions. Significant (P<0.01) negative correlations were observed between the level of monoenes and specifically the level of oleic (18∶1n−9) acid in the diets and PGE₂ synthesis capacity of skin, possibly confirming the known inhibitory effect of 18∶1n−9 on the prostaglandin synthesizing enzyme system. Contribution No. 687, Animal Research Institute.     

Fatty acid composition of rat hearts as influenced by age and dietary fatty acids

Fatty acid composition of neutral and polar lipid fractions from rat hearts was determined in rats of different ages as their diet source changed. Piebald rats were weaned at 21 days and were fed standard lab chow. Lipids from rat hearts, mothers milk and lab chow were purified on a Sephadex G-25 fine column and separated into neutral and polar lipid fractions by silicic acid column chromatography. These lipid fractions were then hydrolyzed and methylated with BF₃ in methanol, prior to gas liquid chromatographic separation on a 1/8 in. × 10 ft aluminum column of 15% EGS on 80–100 mesh acid-washed Chromosorb W. Three major fatty acids in the neutral lipid fraction comprised 72% of total neutral lipid fatty acids from young hearts. At sexual maturity (at least 74 days old) C_18∶1 was the major fatty acid, followed by C_16∶0 and C_18∶0. The same three fatty acids comprised 83% of total polar lipid fatty acids, but C_18∶0 was the major fatty acid, followed by C_16∶0 and C_18∶1. The fatty acid composition of dietary lipids influenced the total neutral lipid fatty acid composition of the rat heart, but had little influence on the fatty acid composition of the polar lipid fraction. Presented in part at the AOCS Meeting, New Orleans, April 1970.     

Effects of dietary n−3 polyunsaturated fatty acids on phospholipid composition and calcium transport in mouse cardiac sarcoplasmic reticulum

The effects of dietary n−3 and n−6 polyunsaturated fatty acids on the fatty acid composition of phospholipid, Ca⁺⁺· Mg⁺⁺ ATPase and Ca⁺⁺ transport activities of mouse sarcoplasmic reticulum were investigated. Mice were fed a 2 weight percent fat diet containing either 0.5 weight percent ethyl esters of 18∶3n−3, 20∶5n−3 or 22∶6n−3 as a source of n−3 polyusaturated fatty acid or 0.5 weight percent safflower oil as a cource of n−6 polyunsaturated fatty acid for 10 days. Olive oil (2 weight percent) was used as a control diet. Although feeding n−6 polyunsaturated fatty acid induced very little modifications of the phospholipid sarcoplasmic reticulum fatty acid composition, feeding n−3 polyunsaturated fatty acid altered it markedly. Inclusion of 18∶−3, 20∶5n−3 or 22∶6n−3 in the diet caused an accumulation of 22∶6n−3, which replaced 20∶4n−6 and 18∶2n−6 in phospholipid sarcoplasmic reticulum. The saturated fatty acids were significantly increased with a concurrent reduction of 18∶1n−9. These changes in the fatty acid composition resulted in a decrease in the values of the n−6/n−3 polyunsaturated fatty acid ratio and a decrease in the ratio of 20 carbon to 22 carbon fatty acids esterified in the phospholipid sarcoplasmic reticulum. This was associated with a decrease in Ca⁺⁺ uptake by n−3 polyunsaturated fatty acid enriched sarcoplasmic reticulum vesicles as compared with n−6 fatty acid and control diet sarcoplasmic reticulum vesicles. However, neither the affinity for Ca⁺⁺ nor the maximal velocity of ATP hydrolysis activity of Ca⁺⁺·MG⁺⁺ ATPase were altered by the different diets. The data suggest that the incorporation of 22∶6n−3 and/or the decrease of 20∶4n−6 plus 18∶2n−6 in the phospholipid sarcoplasmic reticulum may affect the membrane lipid bilayer structure and make it more permeable to Ca⁺⁺.     

Fatty acid metabolism in marine fish: Low activity of fatty acyl Δ5 desaturation in gilthead sea bream (Sparus aurata) cells

Marine fish have an absolute dietary requirement for C₂₀ and C₂₂ highly unsaturated fatty acids. Previous studies using cultured cell lines indicated that underlying this requirement in marine fish was either a deficiency in fatty acyl Δ5 desaturase or C_18–20 elongase activity. Recent research in turbot cells found low C_18–20 elongase but high Δ5 desaturase activity. In the present study, the fatty acid desaturase/elongase pathway was investigated in a cell line (SAF-1) from another carnivorous marine fish, sea bream. The metabolic conversions of a range of radiolabeled polyunsaturated fatty acids that comprised the direct substrates for Δ6 desaturase ([1-¹⁴C]18∶2n−6 and [1-¹⁴C]18∶3n−3), C_18–20 elongase ([U-¹⁴C]18∶4n−3), Δ5 desaturase ([1-¹⁴C]20∶3n−6 and [1-¹⁴C]20∶5n−3), and C_20–22 elongase ([1-¹⁴C]20∶4n−6 and [1-¹⁴C]20∶5n−3) were utilized. The results showed that fatty acyl Δ6 desaturase in SAF-1 cells was highly active and that C_18–20 elongase and C_20–22 elongase activities were substantial. A deficiency in the desaturation/elongation pathway was clearly identified at the level of the fatty acyl Δ5 desaturase, which was very low, particularly with 20∶4n−3 as substrate. In comparison, the apparent activities of Δ6 desaturase, C_18–20 elongase, and C_20–22 elongase were approximately 94-, 27-, and 16-fold greater than that for Δ5 desaturase toward their respective n−3 polyunsaturated fatty acid substrates. The evidence obtained in the SAF-1 cell line is consistent with the dietary requirement for C₂₀ and C₂₂ highly unsaturated fatty acids in the marine fish the sea bream, being primarily due to a deficiency in fatty acid Δ5 desaturase activity.     

Fatty acids in the lipids ofDrosophila heads: Effects of visual mutants,carotenoid deprivation and dietary fatty acids

Lipids ofDrosophila heads were extracted and separated by high-performance thin-layer chromatography. Fatty acid compositions of major phospholipids as well as of triglycerides were analyzed by gas-liquid chromatography. Proportions of the major fatty acids (14∶0, 16∶0, 16∶1, 18∶0, 18∶1, 18∶2, 18∶3) varied depending on the lipid analyzed. Docosahexaenoic acid (22∶6), common in vertebrate photoreceptors and brain, and arachidonic acid (20∶4), a precursor of eicosanoids, were lacking. A comparison of the fatty acid composition of the dietvs. the head suggested thatDrosophila can desaturete but may not be able to elongate fatty acid carbon chains. Fatty acid analyses were carried out after the following visual system alterations: i) the transduction mutant whereno receptorpotential results from a deficit in phospholipase C; ii) an allele ofeyes absent; iii) the mutantouterrhabdomeresabsent which lacks visual pigment and rhabdomeres in the predominant type of compound eye receptor, rhabdomeres 1 through 6; and iv) carotenoid deprivation which reduces opsin and rhabdomere size. We also evaluated aging by comparing newly-emergedvs. aged wild-type flies. Alterations in fatty acid composition based on some of these manipulations were found. Based on comparisons between flies reared on media differing in C₁₆ and C₁₈, there is an indication that diet readily affects tissue fatty acid composition.     

Occurrence of 5c,8c,11c,15t-eicosatetraenoic acid and other unusual polyunsaturated fatty acids in rats fed partially hydrogenated calola oil

Uncommoncis andtrans fatty acids can be desaturated and elongated to produce unusual C₁₈ and C₂₀ polyunsaturated fatty acids in animal tissues. In the present study we examined the formation of such metabolites derived fromcis andtrans isomers of oleic and linoleic acids of partially hydrogenated vegetable oil origin in rats. For two months, aduut male rats were fed a partially hydrogenated canola oil diet containing moderately high levels oftrans fatty acids (9.6 energy%) and an adequate level of linoleic acid (1.46 energy%). Analysis of the phospholipid (PL) fatty acids of liver, heart, serum and brain showed no new C₁₈ polyunsaturated fatty acids, except for those uncommon 18∶2 isomers originating from the diet. However, minor levels (each <0.3% PL fatty acids) of six unusual C₂₀ polyunsaturated fatty acids were detected in the tissues examined, except in brain PL. Identification of their structures indicated that the dietary 9c,13t−18∶2 isomer, which is the majortrans polyunsaturated fatty acid in partially hydrogenated vegetable oils, was desaturated and elongated to 5c,8c,11c,15t−20∶4, possibly by the same pathway that is operative for linoleic acid. Furthermore, dietary 12c−18∶1 was converted to 8c,14c−20∶2 and 5c,8c,14c−20∶3; dietary 9c,12t−18∶2 metabolized to 11c,14t−20∶2 and 5c,8c,11c14t−20∶4, and dietary 9t,12c to 11t,14c−20∶2. These results suggested that of all the possible isomers of oleic and linoleic acids in partially hydrogenated vegetable oils, 12c−18∶1, 9c,13t−18∶2, 9c,12t−18∶2 and 9t,12c−18∶2 are the preferred substrates for desaturation and elongation in rats. However, their conversions to C₂₀ metabolites were not as efficient as that of oleic or linoleic acids.     

Octadecatrienoic acids as the substrates for the key enzymes in glycerolipid biosynthesis and fatty acid oxidation in rat liver

The activities of key enzymes in glycerolipid biosynthesis and fatty acid oxidation were compared using CoA esters of naturally occurring positional isomers of octadecatrienoic acids (18∶3) as the substrates. The trienoic acids employed were 9,12,15–18∶3 (α-18∶3), 6,9,12–18∶3 (γ-18∶3), and 5,9,12–18∶3 (pinolenic acid which is a fatty acid contained in pine seed oil, po-18∶3). The activities of microsomal glycerol 3-phosphate acyltransferase obtained with various 18∶3 were only slightly lower than or comparable with those obtained with palmitic (16∶0), oleic (18∶1), and linoleic (18∶2) acids. Mitochondrial glycerol 3-phosphate acyltransferase was exclusively specific for saturated fatty acyl-CoA. The activities of microsomal diacylglycerol acyltransferase measured with various polyunsaturated fatty acyl-CoAs were significantly lower than those obtained with 16∶0- and 18∶1-CoAs. Among the polyunsaturated fatty acids, γ-18∶3 gave the distinctly low activity. The V_max values of the mitochondrial carnitine palmitoyltransferase I were significantly higher with α-18∶3 and po-18∶3 but not γ-18∶3, than with 16∶0 and 18∶2, while the apparent K_m values were the same irrespective of the types of acyl-CoA used except for the distinctly low value obtained with γ-18∶3. The response to an inhibitor of the acyltransferase reaction, malonyl-CoA, was appreciably exaggerated with 18∶2, α-18∶3, and po-18∶3 more than with 16∶0 and 18∶1. However, the response with γ-18∶3 was the same as with 16∶0. Thus, some of glycerolipid biosynthesis and fatty acid oxidation enzymes could discriminate not only the differences in the degree of unsaturation of fatty acids but also the positional distribution of double bond among the naturally occurring 18∶3 acids.     

On the specificity of a phospholipase A2 purified from the 106,000×g pellet of bovine brain

Assessment has been made of the specificity of a purified phospholipase A₂ from the 106,000×g pellet (microsomal fraction) of bovine grey matter which shows strong activity toward phosphatidylinositol (PI). In the first series of experiments involving the utilization as substrates of PI with different¹⁴C- or³H-labeled fatty acids in the 2-position, the purified phospholipase A₂ most readily removed 16∶0 palmitic acid, followed by 18∶0 stearic acid, 18∶1 oleic acid and 20∶4 arachidonic acid. In the second series of experiments, the purified phospholipase A₂ showed preferential action toward PI (100%) compared to phosphatidylcholine (PC, 62.5%), phosphatidic acid (PA, 32.6%), phosphatidylethanolamine (PE, 25.1%) and phosphatidylserine (PS, 21.5%), where each phosphoglyceride was labeled in the 2-position with [1-¹⁴C] oleic acid. In the third series of experiments, fatty acids were shown to cause inhibition of action of the purified phospholipase A₂ on 1-acyl, 2-[1-¹⁴C] oleoyl PI in the order 20∶4>18∶1>18∶0>16∶0 which is the reverse order to that just noted. In the final series of experiments, the addition of the phosphoglycerides PC, PE, PS and PA in amounts of 5 or 10 μM caused either no inhibition (PE, 2%), slight inhibition (PC, 15%) or reasonably significant inhibition (PA, 20% and PS, 40%) of action of the purified phospholipase A₂ on 1-acyl, 2-[1-¹⁴C]-oleoyl PI. The pattern of specificity observed for the purified phospholipase A₂ combined with its microsomal location are the expected properties of a phospholipase A₂ that might function in a deacylation-reacylation cycle for modifying the fatty acid distribution in PI.     

Moderate changes in linoleate intake do not influence the systemic production of E prostaglandins

A pilot study was undertaken to determine if moderate changes in linoleate (18∶2ω6) intake would modulate the prostaglandin E turnover concurrently with, or independently of, changes in the plasma prostaglandin (PG) precursor levels. Four adult male volunteers in good health were fed two controlled diets containing 35% of energy from fat, with either 10 (diet L) or 30 g (diet H) linoleate/day, 30 to 50 g saturated fatty acids/day, and the balance mainly monounsaturated fatty acids. All four subjects were consuming sufficient amounts of polyunsaturates before the study. Protein (13–14%) and carbohydrate (51–53%) contribution to total caloric intake was kept constant. The menu cycle was 7 days, and all diets were calculated to provide adequate amounts of nutrients known to be required by man when data were available. Plasma fatty acids were determined by gas-liquid chromatography, and the turnover of E prostaglandins was assessed by measuring the urinary output of the major metabolite of PGE₁+PGE₂ (PGE-M). Whereas we found a clear correlation between 18∶2ω6 intake and 18∶2ω6 concentrations in the neutral lipid (P=0.007) and phosphoglyceride (P=0.012) fractions of plasma, arachidonate (20∶4ω6) concentrations in those same plasma fractions did not respond significantly to changes in linoleate intake. Moreover, we could not detect an influence of moderate changes in dietary levels of 18∶2ω6 on the systemic production of PGE as measured by the daily urinary output of PGE-M.     

Conjugated linoleic acids alter bone fatty acid composition and reduce ex vivo prostaglandin E2 biosynthesis in rats fed n-6 or n-3 fatty acids

This study evaluated the effects of conjugated linoleic acids (CLA) on tissue fatty acid composition and ex vivo prostaglandin E₂ (PGE₂) production in rats given diets varying in n-6 and n-3 fatty acids. Four groups of rats were given a basal semipurified diet (AIN-93G) containing 70 g/kg of added fat for 42 d. The fat treatments were formulated to contain CLA (0 vs. 10 g/kg of diet) and n-6 (soybean oil having an n-6/n-3 ratio of 7.3) and n-3 fatty acids (menhaden oil+safflower oil having an n-6/n-3 ratio of 1.8) in different ratios in a 2×2 factorial design. Fatty acids in liver, serum, muscle, heart, brain, spleen, and bone (cortical, marrow, and periosteum) were analyzed by capillary gas-liquid chromatography. The various dietary lipid treatments did not affect growth; however, CLA improved feed efficiency. The CLA isomers were found in all rat tissues analyzed although their concentrations varied. Dietary CLA decreased the concentrations of 16∶1n−7, 18∶1, total monounsaturates and n−6 fatty acids, but increased the concentrations of n−3 fatty acids (22∶5n−3 and 22∶6n−3), and saturates in the tissues analyzed. Ex vivo PGE₂ production in bone organ culture was decreased by n−3 fatty acids and CLA. We speculate that CLA reduced the concentration of 18∶1 fatty acids by inhibiting liver Δ9-desaturase activity. The fact that CLA lowered ex vivo PGE₂ production in bone organ culture suggests that these conjugated fatty acids have the potential to influence bone formation and resorption.