The dietary regulation of acyltransferase and desaturase activities in microsomal membranes of rat liver

Dietary manipulation produces marked alterations in desaturase activities of rat liver microsomes with no concomitant changes in acyltransferase activities. Desaturation of stearoyl-CoA (Δ9-desaturase), linoleoyl-CoA (Δ6-desaturase), eicosatrienoyl-CoA (Δ5-desaturase) and eicosatrienoyl-phosphatidylcholine (Δ5-desaturase) was elevated in animals fed a corn oil diet and lowered in those fed a coconut oil diet compared to control animals. The Δ5-desaturase activities were also lowered in starved animals and elevated in starved animals refed a fat-free diet. However, no changes in acyl-CoA:1-acylsn-glycero-3-phosphocholine acyltransferase activity were observed in the membranes of animals maintained on any of the dietary regimens studied. These observations suggest that the desaturases of rat liver microsomes are regulated independently of the acyltransferases and that desaturation of eicosatrienoyl-phosphatidylcholine is regulated at the level of the desaturase itself and not by availability of the phospholipid substrate.     

The influence of dietary lipids on the composition and membrane fluidity of rat hepatocyte plasma membrane

Weanling male Wistar rats were fed for five weeks on standard rat chow (23 g fat/kg diet) or one of four synthetic diets with butterfat, coconut oil, corn oil, or fish oil as the main lipid source (100 g fat/kg diet). In all diets, 10% of the fat was provided as corn oil to prevent essential fatty acid deficiency. Significant differences were observed in the saturated, monounsaturated, and polyunsaturated fatty acid composition, and in the ratio of cholesterol to phospholipid, in the hepatocyte membranes. The fluidity of hepatocyte plasma membranes was assessed using the fluorescence recovery after photobleaching technique and steady-state fluorescence anisotropy of diphenylhexatriene. No significant differences were found in the fluidity of plasma membranes between animals on the different fat diets, despite diet-induced changes in their fatty acid composition. However, the proportion of lipid free to diffuse in the plasma membrane varied with diet, being significantly greater (P<0.05) in animals fed chow (63.7%), coconut oil (61.5%), and butterfat (57.6%) diets than in those fed the corn oil (47.3%) diet. Animals fed fish oil showed an intermediate (50.0%) proportion of lipid free to diffuse. The data support the hypothesis that dietary lipids can change both the chemical composition and lateral organization (lipid domain structure) of rat hepatocyte plasma membranes.     

Synthesis of α-hydroxy stearyl coenzyme A

Synthesis of α-hydroxy stearyl CoA from N-hydroxysuccinimide ester of a α-hydroxy stearic acid and coenzyme A is reported. The CoA derivative has been isolated, purified and characterized from its spectral data and chemical properties.     

Effects of dietary proteins on linoleic acid desaturation and membrane fluidity in rat liver microsomes

The effect of dietary protein, casein (CAS) and soybean protein (SOY), on linoleic acid desaturation in liver microsomes was studied in rats. The activity of Δ6 desaturase in total and rough endoplasmic reticula (ER and RER) was significantly higher in the CAS group than in the SOY group. In ER and smooth endoplasmic reticulum, the steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene, when incorporated into the membrane, was decreased in the SOY group and accompanied by a reduction in the cholesterol/phospholipid (CHOL/PL) ratio, consistent with an increase in membrane fluidity. In a separate study, the effect of varying dietary proteins, CAS, milk whey protein, egg albumin, SOY, potato protein and wheat gluten, on the relationship between the Δ6 desaturase activity and microsomal membrane fluidity was also examined. The results indicated that the dietary protein-dependent change in the liver microsomal CHOL/PL ratio affected membrane fluidity, and subsequently the activity of Δ6 desaturase in liver microsomes. However, since dietary protein influenced the Δ6 desaturase activity in RER without influencing membrane fluidity, it is possible that some regulation might have taken place at the level of enzyme synthesis.     

The effects of dietary protein on the fatty acid composition and Δ6 desaturase activity of rat hepatic microsomes

Recently, significant differences between rats fed a casein diet and rats fed a soybean protein diet have been observed in hepatic phospholipid fatty acid patterns (Sjöblom, L., and Eklund, A.,Biochim. Biophys. Acta 1004, 187–192, 1990). The influence of these two diets on the Δ6 desaturase activity was investigated in the present study because the hepatic desaturase system is a source of unsaturated fatty acids. The rats fed a casein diet showed higher desaturase activity than those fed soybean protein when using either linoleic acid (P<0.005) or oleic acid (P<0.05) as substrates. The phosphatidylcholine fraction of hepatic microsomes showed increases in oleic acid (P<0.005) and 20∶3ω9 (P<0.001) levels as well as decreases in stearic acid (P<0.001), linoleic acid (P<0.005) and arachidonic acid (P<0.005) levels in rats which were fed casein rather than soybean protein. Similar differences between the two groups were also observed in the phosphatidylethanolamine and phosphatidylinositol fractions. These data indicate that the qualitative properties of the dietary protein source may influence the fatty acid pattern of rat hepatic microsomes by interfering with Δ6 desaturase activity.     

Effect of dietary fatty acids on Δ5 desaturase activity and biosynthesis of arachidonic acid in rat liver microsomes

The effect of different fatty acids supplemented to a fat-free diet on the activity of Δ5 desaturase was studied. Fat-free diet produces a reduction in the conversion of eicosa-8,11,14-trienoic acid to arachidonic acid. The addition of thecis-ω6 acids, linoleic, γ-linolenic or arachidonic to the diet produces an increase of eicosatrienoic acid desaturation, shifting Δ5 desaturase activity towards the controls on a balanced diet. This reactivation is apparently produced by induction of enzyme biosynthesis since linoleate effect was suppressed by simultaneous cycloheximide injection. On the contrary, no changes in Δ5 desaturation activity were found when the diet was supplemented with palmitic or 9-trans,12-trans-linoleic acid. The changes on the activity of Δ5 desaturase were compared with the fatty acid composition of plasma and liver microsomes.     

Adaptive changes in Δ9 desaturase activity in rat liver

The Δ9 desaturase activity and the¹⁴C radioactivity of the de novo synthesized fatty acids incorporated into microsomal lipids and serum triglycerides were measured under different nutritional conditions. The results obtained indicate a correlation between the values of the three parameters studied after starvation or after refeeding Purina chow or either a high carbohydrate or a high protein diet. These data suggest that liver lipogenesis and Δ9 desaturase activities respond to the same regulatory factors.     

Stearoyl-coenzyme A desaturase activity in novikoff hepatoma

The activities of microsomal stearoyl-CoA desaturation, NADH-cytochrome b₅ reductase, NADH-cytochrome c reductase, and the content of cytochrome b₅ were similar in livers of normal and host rats. On the other hand, stearoyl-CoA desaturation activity was absent in Novikoff hepatoma. The activities of NADH-cytochrome b₅ and NADH-cytochrome c reductases in the hepatoma microsomes were 4.8% and 2.2%, respectively, of those in normal liver. Furthermore, in hepatoma microsomes, cytochrome b₅ was absent. An active stearoyl-CoA desaturation was reconstituted only on addition of both cytochrome b₅ and the terminal desaturase enzyme to the hepatoma microsomes. These results indicated that a complete absence of cytochrome b₅ and terminal desaturase is responsible for the lack of stearoyl-CoA desaturation in Novikoff hepatoma microsomes.     

Effects of dietary palm oil on arterial thrombosis,platelet responses and platelet membrane fluidity in rats

Wistar rats were fed a control diet containing 5 energy % (en %) sunflowerseed oil or diets containing 50 en % of either palm oil, rich in saturated fatty acids, or sunflowerseed oil, high in linoleic acid, for at least eight weeks. Arterial thrombosis tendency, measured by the aorta loop technique, tended to be lowered by the palm oil diet and was lowered significantly by the sunflowerseed oil diet, compared with the control. Aggregation of platelets in whole blood activated with collagen was not altered by palm oil feeding, but was enhanced in the sunflowerseed oil group, compared with the control. The concomitant formation of thromboxane A₂ was decreased by palm oil feeding, although formation of prostacyclin did not change; the ratio of thromboxane/prostacyclin formed was decreased significantly in the palm oil group. Compared with the control diet, platelet membrane fluidity, measured by fluorescence polarization, was not altered in the palm oil group and was significantly increased only by sunflowerseed-oil feeding. Thus, although palm oil contains about 50% saturated fatty acids, it did not increase arterial thrombosis tendency and tended to decrease platelet aggregation, as compared with highly polyunsaturated sunflowerseed oil.     

Thermal acclimation and dietary lipids alter the composition,but not fluidity,of trout sperm plasma membrane

The effect of a long-term adaptation of rainbow trout to 8 and 18°C combined with a corn oil-or a fish oil-supplemented diet on the characteristics of the spermatozoan plasma membrane was investigated. The experiment lasted up to 22 mon during which spermatozoa were collected from the mature males. Spermatozoan plasma membranes were isolated by nitrogen cavitation, and the cholesterol content, phospholipid composition and fatty acid pattern were investigated. Membrane viscosity was assessed on whole cells by electron spin resonance using spin-labeled phospholipids. Neither diet nor rearing temperature influenced the cholesterol content of the plasma membrane nor the phospholipid class distribution. The rearing temperature of the broodstock only slightly affected the phospholipid fatty acids. A minor decrease in 18∶0 and increase in monounsaturated fatty acids was observed for the cold-adapted fish. These modifications were not sufficient to affect membrane fluidity, and we conclude that trout spermatozoa do not display any homeoviscous adaptations in these conditions. On the contrary, the dietary fatty acid intake greatly modified the fatty acid profile of plasma membrane phospholipids. The fish oil-fed trout displayed a much higher n−3/n−6 fatty acid ratio than did the corn oil-fed ones, but the 22∶6n−3 levels remained unchanged. Modifications in plasma membrane composition by the diet were obtained although neither of the two diets was deficient in essential fatty acids. The enrichment in n−3 fatty acids, however, did not affect plasma membrane fluidity which was unchanged by the diets.     

The effect of dietary zinc deficiency on the lipid composition of the rat erythrocyte membrane

The effect of dietary zinc deficiency in the rat on the lipid composition of the erythrocyte membrane was determined. Weanling male Wistar rats were fed an egg whitebased diet containing <1.0 mg Zn/kg dietad libitum. Control rats were either pair-fed orad libitum-fed the basal diet suppelemented with 100 mg Zn/kg diet. A zinc refed group was fed the −Zn diet until day 18 and then pairfed the +Zn diet until day 21. The voluntary feed restriction associated with dietary zinc deficiency resulted in erythrocyte membranes that had depressed phospholipid/protein and elevated cholesterol/phospholipid ratios. Similarly, all feed restricted groups had elevated 22-carbon n−3 polyunsaturated fatty acid (PUFA) and depressed 22-carbon n−6 PUFA concentrations in alkenylacyl and diacyl glycerophosphoethanolamine, phosphatidylserine and phosphatidylcholine; they also had depressed 24∶2n−6 levels in sphingomyelin. The relative concentrations of phospholipids in the membrane was similar between −Zn and +Zn (ad libitum) groups; however, the −Zn group had significantly less phosphatidylserine relative to +Zn (pair-fed) controls.     

Liver Δ5 and Δ6 desaturase activity differs among laboratory rat strains

This study was designed to examine the variations among rat strains in hepatic fatty acid desaturase activities and to determine the correlations between the activities of these enzymes and the levels of each microsomal fatty acid. Wistar rats from two different sources as well as Long-Evans and Sprague-Dawley rats were selected to assess, under standard and identical experimental conditions, the liver Δ5 and Δ6 desaturase activities. Both desaturase activities were significantly reduced by 56% in Sprague-Dawley rats when compared to BB-Wistar control rats, whereas intermediate reduced values were detected in Wistar (CR) and Long-Evans strains. The activities of Δ5 and Δ6 desaturases were significantly and positively correlated with each other. However, no significant correlations were detected between either Δ5 or Δ6 desaturase activities and levels of any of their fatty acid substrates or any other of the major microsomal fatty acids. Fatty acid composition of microsomal total lipids showed strain dependency. A positive correlation was detected between the microsomal levels of the two major final products of both desaturases, namely 20∶4n−6 and 22∶6n−3. In general, the sum of n−3 or n−6 fatty acids but not the ratio of one to the other, varied among rat strains. The study demonstrated that Δ6 and Δ5 desaturase activities are strain-related. The data also suggested that (i) the desaturation activity should be measured and not predicted from the fatty acid composition and (ii) different rat strains should be used for lipid metabolic studies before conclusions are drawn for rats in general.     

The source of dietary fatty acids alters the activity of secretory sphingomyelinase in the rat

Secretory sphingomyelinase (sSMase) has been suggested to be involved in the development of cardiovascular diseases as well as other human pathologies. To deduce whether dietary fatty acid composition affects the circulating activity of this enzyme, we have compared its activity in serum from rats that had been given a diet containing either butter or a highly n‐6 polyunsaturated [grapeseed oil (GSO)] fat source for 14 wk. The results show that intake of GSO increases the activity of this ceramide‐producing enzyme by about 45%, when compared with intake of butter (387 ± 16 pmol/mL·h vs. 266 ± 15 pmol/mL·h; p <0.001). Furthermore, there was a strong negative correlation between sSMase activity and n‐3 PUFA concentration in serum (p <0.001). Despite the substantial increase in activity, there was no difference in either the circulating substrate (sphingomyelin) or product (ceramide) in the serum. However, since the sSMase activity in the endothelial wall has been implicated to be involved in both atherogenesis and thrombosis, these findings are of interest in the interpretation of dietary fatty acid effects on cardiovascular health.     

Effect of dietary fat on the fluidity of platelet membranes

Dietary fat type was reflected in the phospholipid fatty acid composition of the plasma membrane of rabbit platelets and apparently controlled the fluidity of these membranes. Rabbits were maintained for 6 months on diets that varied in stearic and polyunsaturated fatty acids and thus had different potentials for thrombosis. Microviscosities at 37 C, calculated from the anisotropy of fluorescence from the probe 1,6-diphenyl-1,3,5-hexatriene, were 3.5, 3.4, 2.8 and 2.2. poise for platelet membranes isolated from rabbits whose only source of dietary fat was cocoa butter, milkfat, coconut oil, or corn oil, respectively. The relative fluidities of the membrane isolates were correlated with the polyunsaturated fatty acid contents of the membrane phospholipids.     

Effects of estradiol and environmental temperature changes on rat liver Δ6 microsomal desaturase activity

The regulation of Δ6 desaturase activity by environmental temperature changes was studied in the microsomal membranes from female and ovariectomized female rat liver. Female rats adapted at 30–32 C for 20–25 days and then shifted to 13–15 C for 5 days showed an increased Δ6 desaturase system. Ovariectomized rats adapted under the same conditions did not show significant changes in this enzyme. The fatty acid compositions of microsomal phosphatidylcholine showed a decrease in arachidonic acid in female rats at 30 C compared to females at 15 C and ovariectomized rats at both temperatures. These results suggest that a modification of ovaric sex hormone levels might be responsible for the different Δ6 desaturase activity in female rats acclimated at both temperatures. In this regard, serum estradiol radioimmunoassay yielded slight differences between the two groups of female rats, suggesting that estradiol could play a role in the regulation of the Δ6 desaturase. The administration of a pharmacological dose of 17-β estradiol to female and ovariectomized rats kept at 30 and 15 C decreased the Δ6 microsomal desaturase activity. These data suggest that estradiol levels are involved in the regulation of the Δ6 desaturase during cold adaptation.     

Homeostatic control of membrane fatty acid composition in the rat after dietary lipid treatment

Diets in which both the lipid content and composition (polyunsaturated to saturated fatty acid ratio) were varied were fed to rats for 20 weeks, and the effects on the tissue lipid profiles were determined. The fatty acid profile of the plasma lipids, and the phospholipid fatty acids of the mitochondrial and microsomal fractions of liver, heart, kidney and brain, as well as erythrocyte membranes were determined. Despite large differences in the level and type of lipid present in the experimental diets and in the proportion of saturated fatty acids in the plasma lipids in response to the various diets, there was little effect on the proportion of saturated to unsaturated fatty acids in the phospholipids of the various membranes examined. The major effect of altering the dietary level of polyunsaturated to saturated fatty acids was on the ratio of the ω6/ω3 series of unsaturated fatty acids in the membrane lipids. This change occurred in all tissues except the brain, in which only a small response to altered dietary lipid intake was observed. The ω6/ω3 ratio was elevated upon feeding a diet rich in ω6 polyunsaturated fatty acids, but decreased when a diet rich in saturated fatty acids was fed. The failure to significantly alter membrane lipid saturation/unsaturation in the tissues examined would suggest that a homeostatic mechanism is operative in biological membranes and may act to buffer membranes from the effects of changes in the nature of the dietary lipid intake.     

Stearoyl-CoA desaturase activity in adipose tissue and liver of the cardiomyopathic hamster

Stearoyl-CoA desaturase activity and the fatty acid composition of lipids of adipose tissue and liver were determined in 35- and 180-day-old cardiomyopathic hamsters and age-matched normal controls. Enzyme activity was unchanged in the adipose tissue of 35-day-old animals but was significantly depressed in the 180-day-old cardiomyopathic hamsters. In the liver, stearoyl-CoA desaturase activity was significantly lower in the 35-day-old disease animals but was unchanged in the 180-day-old animals. The analysis of the fatty acid composition of the lipids isolated from adipose tissue showed an increase in the relative percentage of saturated fatty acids accompanied by a decrease in the relative percentage of unsaturated fatty acids in both age groups of the cardiomyopathic hamsters. However, linoleic acid content was increased in the diseased animals. Similar changes in fatty acid composition of lipids from the livers of 35-day-old cardiomyopathic hamsters were observed, but no significant differences in the fatty acid composition between 180-day-old cardiomyopathic hamsters and normal controls were observed. The changes in the fatty acid composition appear to be related to the observed changes in desaturase activity. In is concluded that such changes in desaturase activity and fatty acid composition could affect the normal structure and functions of membranes and membrane-related processes.     

Effect of dietary fats on desaturase activities and the biosynthesis of fatty acids in rat-liver microsomes

Four groups of rats were fed diets containing 15% (w/w) high-oleic safflower oil (SFO, rich incis-18∶1 acids), a mixture of 80% partially hydrogenated soybean oil plus 20% corn oil (H+CO, rich intrans-18∶1 acids), lard (L, rich in saturated fatty acids) and corn oil (Co, rich in 18∶2ω6). Fatty acid composition of liver microsomes and activities of the Δ⁵, Δ⁶ and Δ⁹ desaturases were determined. Microsomal Δ⁶ desaturase activity and arachidonic acid were lower in the H+CO group compared with SFO of L. No difference was found in the Δ⁵ or Δ⁶ desaturase activity of CO and SFO groups. Thus, the oleic-acid level of the SFO diet had no effect on the metabolism of 18∶2ω6. Fluorescent polarization studies, usingtrans-parinaric acid as a probe, showed no differences between the physical states of phospholipid vesicles made from lipids isolated from each group. We concluded that thetrans-18∶1 acids in partially hydrogenated soybean oil have a more inhibitory effect than saturated acids on EFA metabolism, even in the presence of adequate amounts of essential fatty acid.     

Effect of magnesium deficiency on Δ6 desaturase activity and fatty acid composition of rat liver microsomes

Experimental Mg²⁺ deficiency was induced in a group of rats by feeding them a Mg²⁺-deficient diet for 23 days. They were pair-fed to compare with a control group of rats fed a Mg²⁺-sufficient diet. In the Mg²⁺-deficient group the plasma total cholesterol and triglyceride levels were increased while HDL-cholesterol was decreased. In the Mg²⁺-deficient group the plasma level of thiobarbituric acid reacting substances (TBARS) used as a measure for lipid peroxidation was increased. The increase was attributed to the increased cytosolic Ca²⁺ in Mg²⁺-deficiency which can cause: 1) increase of hydro and endoperoxide levels as a consequence of the increase of arachidonic acid release and eicosanoid synthesis in Mg²⁺-deficiency, and 2) inhibition of the mitochondrial respiratory activity and activation of Ca²⁺-dependent proteases which may activate the conversion of xanthine dehydrogenase to xanthine oxidase which generates active O₂ species. In the Mg²⁺-deficient group, the fatty acid composition of the liver microsomes indicated a slower rate of conversion of linoleic acid to arachidonic acid which was consistent with the decrease of Δ6 desaturase activity in liver microsomes of Mg²⁺-deficient rats as measuredin vitro. The decrease of Δ6 desaturase activity was attributed to the lower concentration of actual enzyme molecules as a result of the decreased rate of protein synthesis in Mg²⁺-deficiency. The possible effects of the increased catecholamine release in Mg²⁺-deficiency are discussed.     

Stearoyl-coenzyme a desaturase activity in the mammary gland and liver of lactating rats

Stearoyl-CoA desaturase activity in microsomes from lactating rat mammary gland is very low (0.05–0.15 nmol/min/mg of protein) regardless of lactating time. In such microsomes, reductase activities and content of cytochrome b₅ are several-fold lower than in normal rat liver microsomes. Preincubation of the mammary microsomes with purified terminal desaturase gives a 55-fold stimulation of stearoyl-CoA desaturase activity, whereas preincubation with cytochrome b₅ has no effect. However, preincubation of mammary microsomes with both cytochrome b₅ and terminal desaturase results in a 200-fold stimulation of overall desaturation. These observations suggest that negligible stearoyl-CoA desaturase activity in lactating rat mammary microsomes is due to a low cytochrome b₅ content and the absence of terminal enzyme. The hepatic stearoyl-CoA desaturase activity increases 9-fold during lactation. There is little or no change in the NADH-cytochrome c reductase activity or in the concentration of cytochrome b₅ during this period, but the activity of the terminal desaturase increases with the increase of overall desaturation. These results suggest that liver is one of the more important sources of oleic acid for milk triglycerides.