Effects of fatty acids on activity and calmodulin binding of Ca-ATPase of human erythrocyte membranes

Activation and inhibition of Ca²⁺-ATPase of calmodulin-depleted human erythrocyte membranes by oleic acid and a variety of other fatty acids have been measured. Low concentrations of oleic acid stimulate the enzyme activity, both in the presence and in the absence of calmodulin. Concomitantly, the affinity of the membrane bound enzyme to calmodulin progressively decreases due to competitive interactions of calmodulin and oleic acid with the enzyme. Removal of oleic acid from the membrane by serum albumin extinguishes the activating effect of oleic acid and restores the ability of the enzyme to bind calmodulin with high affinity. High concentrations of oleic acid induce an almost complete and irreversible loss of enzyme activity which cannot be abolished by removal of oleic acid. Despite a complete loss of enzyme activity, binding of calmodulin to membranes is approximately normal after removal of oleic acid. Activities of (Na⁺ + K⁺)-ATPase, Mg²⁺-ATPase and acetylcholine esterase, as well as the total protein content, show no gross changes upon treatment of membranes with increasing amounts of oleic acid, which seems to exclude that membrane solubilisation by oleic acid causes an inactivation of the enzyme.     

Effects of stereochemically different fatty acids on blood cholesterol in rats

In the present study some observations about the effects of stereochemically different fatty acids on cholesterolaemia are reported. It is well known that high in EFA diets are useful for cholesterol metabolism: in fact cholesterol is preferably esterificated by EFA. Because of the increasing intake of fats, as high in hydrogenated EFA margarines, it has been verified whether such a technological process, inducing structural modifications, affects or not the ipocholesterolemic effect of EFA, particularly linoleic acid. Actually this research has shown such an effect to fail in presence of trans fatty acids and has pointed out some implications of lipid metabolism which are related to the different stereochemical configurations.     

Effects of acute cold exposure on rectal temperature, blood glucose and plasma free fatty acids in alloxan-diabetic rats

These experiments were carried out to study the effects of acute cold exposure (0-2 degrees C/4 hr) on rectal temperature, blood glucose and plasma free fatty acids (FFA) in alloxan-diabetic rats. Male Wistar rats weighing 170-190 g were used and diabetes was induced by i.v. alloxan injection (40 mg/kg body wt). Cold exposure produced severe hypothermia in diabetic rats. After 4 hr of cold, blood glucose of diabetic rats was reduced from 296 +/- 16 to 86 +/- 12 mg/dl (P less than 0.01), and FFA increased slightly, but was not statistically different (P greater than 0.05) from the initial value. As expected, interscapular brown adipose tissue (IBAT) and retroperitoneal and epididymal white adipose tissues were significantly lower in diabetic than in control rats. Cold exposure reduced total IBAT lipids in control but not in diabetic animals. The results of this experiment suggest that diabetic rats were unable to maintain body temperature in the cold, probably because of a failure to generate an adequate amount of heat by nonshivering thermogenesis in brown adipose tissue.     

Effect of acute exercise on tissue free fatty acids in untrained rats.

The effects of a single bout of swimming on free fatty acids (FFA) in adipose tissue, heart, skeletal muscle, and serum were examined. Surprisingly, in previously untrained rats, FFA were elevated (P less than 0.001) in epididymal, inguinal, and retroperitoneal adipose depots 48 h after a 2-h swim. FFA in the three fat depots returned to resting levels 96 h after exercise. In heart, soleus, and fast-red fibers of the quadriceps, FFA remained elevated (P less than 0.01) for as long as 72 h after the 2-h swim. Serum FFA were still elevated (P less than 0.001) 96 h after swimming but not after 168 h. These results provide evidence that the rise in FFA is an acute effect of exercise and not a cellular adaptation resulting from daily episodes of lipolysis induced by exercise training. In a separate experiment, involving the adaptive response to endurance exercise, adipocytes from epididymal, inguinal, and retroperitoneal depots were reduced in size (P less than 0.001) to approximately the same degree. These results provide evidence that adipocytes from each depot contribute equally in meeting the energy needs of muscle during repeated bouts of endurance exercise.     

Effects of medium-chain fatty acids on body composition and protien metabolism in overweight rats

The lack of efficiency of classical treatments for obesity has led to propose alternative strategies. In order to obtain information about the effects of dietary fatty acid composition on body fat and protein metabolism, overweight female rats were fed on isoenergetic diets, using either medium-chain (MCT) or long-chain (LCT) triglycerides as a lipid source. After 23 days, the MCT group had mildly decreased body weight but greatly reduced adipose tissue depots. All fat depots were significantly diminished. MCT-fed rats showed a decrease in some hormones involved in energy balance, such as leptin and triiodothyronine. Feeding MCT resulted in improvements in nitrogen balance. Muscle protein content was similar in both treatments despite an increase in protein degradation in the MCT group. The present data clearly show that a diet with MCT as lipid fuel depresses weight gain and fat stores, relative to a standard LCT diet.     

Effects of dietary lead,cadmium, mercury,and selenium on fatty acid composition of blood serum and erythrocyte membranes from chicks

A series of experiments was conducted to study the effects of dietary and injected lead (as Pb acetate-3H₂O) and of dietary Cd, Hg, and Se on fatty acid composition of serum lipids of chicks as measured by gas-liquid chromatography. The effect of dietary Pb on fatty acid composition of erythrocyte membranes was measured also. Dietary Pb (1000 ppm) increased the serum concentration of arachidonic acid (20:4, first no. = no. of carbon atoms:second no. = no. of double bonds) and decreased the concentration of linoleic acid (18:2) and the ratio 18:2/20:4. Intraperitoneal injection of Pb (52 mg/100 g body weight) did not alter serum fatty acid composition by 4 h post-injection. The separate effects of 2000 ppm Pb, 60 ppm Cd, 500 ppm Hg, and 10 ppm Se added to the diet on serum fatty acids were measured in a single experiment. In comparison to controls, Pb and Cd lowered serum concentration of 18:2. Only Pb raised serum 20:4. Pb lowered the ratio 18:2/20:4, whereas Cd and Hg raised the ratio and Se was without effect. Dietary Pb (2000 ppm) raised the concentration of 20:4 and lowered the ratio 18:2/20:4 in erythrocyte membranes. The different effects of injected and dietary Pb on the serum 18:2/20:4 ratio suggest that Pb alters 20:4 synthesis from 18:2 rather than mobilization of 20:4 from tissues. The Pb-induced increase of lipid peroxida-tion in erythrocytes observed by other workers may be a reflection of increased 20:4 level in erythrocyte membranes.     

Effects of drugs on water permeability of erythrocyte membranes

The method of NMR-relaxation with the manganese doping has been applied to study changes of water permeability of red blood cell membranes affected by various concentrations of chlorhexidine digluconate and dimephosphone. It is shown that both investigated substances suppress the water permeability of the red blood cell membrane in a dose-dependent manner. Half-maximum inhibitory effect of studied substances was reached at the concentrations of 9 μM of chlorhexidine and 400 μM of dimephosphone.     

Fluidity properties and liquid composition of erythrocyte membranes in Chediak-Higashi syndrome

We have earlier shown through electron spin resonance (ESR) studies of leukocytes that membranes of cells from both Chediak-Higashi syndrome (CHS) mice and humans have abnormally high fluidity. We have extended our studied to erythrocytes. Erythrocytes were labeled with the nitroxide-substituted analogue of stearic acid, 2-(3-carboxypropyl)-4,4- dimethyl-2-tridecyl-3-oxazolidinyloxyl, and ESR spectra were obtained. Order parameter, S, at 23 degrees C, was 0.661 and 0.653 for erythrocytes of normal and CHS mice (P less than 0.001). S was 0.684 for normal human erythrocytes and 0.675 (P less than 0.001) for CHS erythrocytes at 25 degrees C. Because S varies inversely to fluidity, these results indicate that CHS erythrocytes tend to have higher fluidity than normal. In vitro treatment of both mice and human CHS erythrocytes with 10 mM ascorbate returned their membrane fluidity to normal. We prepared erythrocyte ghosts and extracted them with CHCl3:CH3OH (2:1). Gas-liquid chromatography analysis showed a greater number of unsaturated fatty acids for CHS. The average number of double bonds detected in fatty acids for mice on a standard diet was 1.77 for normal and 2.02 for CHS (P less than 0.04); comparison of human erythrocytes from one normal control and one CHS patient showed a similar trend. Our results suggest that an increased proportion of unsaturated fatty acids may contribute to increased fluidity of CHS erythrocytes. Our observation that both leukocytes and erythrocytes of CHS have abnormal fluidity indicates that CHS pathophysiology may relate to a general membrane disorder.     

Effects of fatty acids on activity and calmodulin binding of Ca2+-ATPase of human erythrocyte membranes

Activation and inhibition of Ca2+-ATPase of calmodulin-depleted human erythrocyte membranes by oleic acid and a variety of other fatty acids have been measured. Low concentrations of oleic acid stimulate the enzyme activity, both in the presence and in the absence of calmodulin. Concomitantly, the affinity of the membrane bound enzyme to calmodulin progressively decreases due to competitive interactions of calmodulin and oleic acid with the enzyme. Removal of oleic acid from the membrane by serum albumin extinguishes the activating effect of oleic acid and restores the ability of the enzyme to bind calmodulin with high affinity. High concentrations of oleic acid induce an almost complete and irreversible loss of enzyme activity which cannot be abolished by removal of oleic acid. Despite a complete loss of enzyme activity, binding of calmodulin to membranes is approximately normal after removal of oleic acid. Activities of (Na+ + K+)-ATPase, Mg2+-ATPase and acetylcholine esterase, as well as the total protein content, show no gross changes upon treatment of membranes with increasing amounts of oleic acid, which seems to exclude that membrane solubilisation by oleic acid causes an inactivation of the enzyme.     

Dietary trans fatty acids modulate erythrocyte membrane fatty acyl composition and insulin binding in monkeys

The substitution of trans- for half of the cis-monounsaturated fatty acids in the diet of Macaca fasicularis monkeys resulted in alterations in erythrocyte fatty acid composition and insulin receptor properties but not in membrane fluidity. Both cis and trans diets contained 10% fat and similar fatty acid compositions, except that approximately 50% of the cis-octadecenoate (c-18:1) in the cis diet was replaced with trans-octadecenoate isomers (t-18:1) in the trans diet. Compared with the cis diet, the trans diet resulted in the incorporation of approximately 11% t-18:1, an approximately 50% decrease in c-18:1, an approximately 16% decrease in total saturated fatty acids, and an approximately 20% increase in 18:2(n-6) in erythrocyte membrane lipids. The increase in 18:2(n-6) may reflect on homeostatic mechanisms designed to maintain overall membrane fluidity, as no diet-related changes in fluidity were observed with diphenylhexatriene steady state fluorescence polarization. Values observed for insulin binding and insulin receptor number were higher and binding affinity was lower in monkeys fed the cis diet. In the absence of an effect on overall membrane fluidity, altered receptor activity suggests that insulin receptor activity is dynamic, requiring specific fluid membrane subdomains or highly specific fatty acid-protein interactions.     

Differential effect of cold acclimation on blood composition in rats and hamsters

Male rats and hamsters were exposed to a progressively lower air temperature and shorter photoperiod to simulate the onset of winter. Normothermic hamsters had a higher haematological oxygen carrying capacity (OCC) and coagulability (shorter prothrombin time and activated partial thromboplastin time) than rats. Cold acclimation significantly increased the OCC of rats, which parallels an increased metabolic rate, while no differences were observed in hamsters. Red cell transit time through filters was faster in the acclimated rats but not in hamsters, reflecting the lower mean cell volume due to a decreased rate of clearance from the circulation. Platelet counts were significantly lower in both cold-acclimated rats and hamsters, and there was a significant leucopenia in rats, which would reduce the degree of microvascular blockade. Whole blood viscosity, plasma viscosity, and serum osmolarity showed little change in either species. However, whole blood viscosity was significantly lower in cold-acclimated hamsters than control hamsters at the lowest shear rate tested (0.95 s(-1)). Interestingly, plasma viscosity and serum osmolarity were significantly lower in hamsters exposed to low temperatures for a shorter period (4 weeks), and may reflect the development of a reduced coagulability. These data suggest that blood composition in hamsters contributes to an innate tolerance of low temperatures, maintaining tissue perfusion under hypothermic conditions and aiding arousal from hibernation.     

The fatty acid composition of 1,2-diacylglycerol and polyphosphoinositides from human erythrocyte membranes.

The fatty acid compositions of 1,2-diacylglycerol and polyphosphoinositides have been determined in human erythrocyte membranes that have been incubated in the presence or in the absence of Ca2+. The results show that the diacylglycerol that is formed in Ca2+-treated membranes has a fatty acid composition closely similar to that of the inositides, consistent with previous indications that Ca2+ stimulates the activity of a polyphosphoinositide phosphodiesterase in the membranes. In contrast with some previous results, it appears that these plasma-membrane inositides and their derived diacylglycerols are rich in stearic acid and arachidonic acid.     

Effect of cell age on erythrocyte fatty acid composition in rats on different dietary regimes

1. Rat erythrocytes were fractionated into young, mature and old cell fractions by centrifugation. The fatty acid composition of each fraction was determined by gas-liquid chromatography, under four different dietary conditions: with adequate linoleic acid in the diet, with a diet deficient in linoleic acid, and with the deficient diet supplemented with corn oil for 3 and 12 days. 2. Significant differences were observed in the fatty acid composition of cells of different ages on all diets. The pattern of fatty acid distribution depended on the particular acid in question, on its concentration in the total erythrocyte sample and on the nature of the dietary fat. 3. When corn oil was fed to rats that had been fed previously on a deficient diet, the changes in fatty acid composition that occurred depended on the acid and on the cell age. For example, young cells were more active in incorporating palmitic acid and arachidonic acid but incorporated linoleic acid at a slightly lower rate than older cells. 4. These results are believed to indicate the presence in the erythrocyte of transacylases with different specificities, and to show that transacylase activity changes as the cells age.