Phospholipid and cholesterol levels in the tumor cell plasma membranes with different sensitivity to doxorubicin

The investigation is aimed to study qualitative and quantitative composition of phospholipids, cholesterol content and lipids unsaturation index in plasma membranes of Guerin's carcinoma cells sensitive or resistant to doxorubicin. The comparison of infrared spectra and phospholipids unsaturation index showed that the unsaturation level of fatty acids in plasma membrane from resistant cells was lower than that from sensitive carcinoma cells. 31P-NMR spectroscopy of plasma membranes phospholipids shows the increase of phosphatidylserine and sphingomyeline content in plasma membrane isolated from resistant tumor as compared with sensitive tumor. The levels of phosphatidylcholine and phosphatidylethanolamine were equal in drug-resistant and drug-sensitive carcinoma strains. Changes in plasma membrane from resistant cells result in elevation of plasma membrane microviscosity and phosphatidylserine level increase can suggest the activation of P-glycoprotein-mediated efflux of doxorubicin.     

The reactivity of plasma phospholipids with lecithin:cholesterol acyltransferase is decreased in fish oil-fed monkeys

The size of low density lipoproteins (LDL) is strongly correlated with LDL cholesteryl ester (CE) content and coronary artery atherosclerosis in monkeys fed cholesterol and saturated fat. African green monkeys fed 11% (weight) fish oil diets have smaller LDL and less CE per LDL particle than lard-fed animals. We hypothesized that this might be due to a lower plasma lecithin:cholesterol acyltransferase (LCAT) activity in fish oil-fed animals. Using recombinant particles made of egg yolk lecithin-[14C]cholesterol-apoA-I as exogenous substrate, we found no difference in plasma LCAT activity (27 versus 28 nmol CE formed per h/ml) of fish oil- versus lard-fed animals, respectively; furthermore, no diet-induced difference in immunodetectable LCAT was found. However, plasma phospholipids from fish oil-fed animals were over 4-fold enriched in n-3 fatty acids in the sn-2 position compared to those of lard-fed animals. Additionally, the proportion of n-3 fatty acid-containing CE products formed by LCAT, relative to the available n-3 fatty acid in the sn-2 position of phospholipids, was less than one-tenth of that for linoleic acid. The overall rate of LCAT-catalyzed CE formation with phospholipid substrates from fish oil-fed animals was lower (5-50%) than with phospholipid substrates from lard-fed animals. These data show that n-3 fatty acids in phospholipids are not readily utilized by LCAT for formation of CE; rather, LCAT preferentially utilizes linoleic acid for CE formation. The amount of linoleic acid in the sn-2 position of plasma phospholipids is reduced and replaced with n-3 fatty acids in fish oil-fed animals. As a result, LCAT-catalyzed plasma CE formation in vivo is likely reduced in fish oil-fed animals contributing to the decreased cholesteryl ester content and smaller size of LDL particles in the animals of this diet group.     

Biochemical changes of rat brain membranes with aging

Modification of membrane composition and enzymatic activities both in total brain homogenate and purified synaptic plasma membrane of 3 and 24 month old rats has been investigated. Protein, cholesterol and phospholipid content and (Na+, K+)ATPase and 2',3' cyclic nucleotide phosphohydrolase activities were determined. The major changes occurred in the whole homogenate where a general increase in total protein and cholesterol content with age and a significant increase of the cholesterol/phospholipids molar ratio has been detected. In S.P.M. aging process induced a decrease of protein, cholesterol and phospholipids content associated with an increased membrane viscosity and a decrease of delta E. These data are consistent with a change in the structural organization and in the distribution pattern of different cell population in the aging brain. A possible artifactual effect of freezing on the reported parameter is also discussed.     

Phospholipid unsaturation and plasma membrane organization.

A comparison has been made between the unsaturation of plasma-membrane phospholipids,present in the human erythrocyte, rat liver, mouse liver and a rapidly growing rat hepatoma. Of the double bonds present in the hydrocarbon chains of the membrane phospholipids,onethird is contributed by sphingomyelin plus phosphatidyl choline and the remainder by phosphatidyl serine, ethanolamine and inositol. Assuming that the phospholipids are asymmetrically distributed in the two leaflets of the bilayer in general, the consequences of this asymmetry in combination with cholesterol content and fatty acid distribution on plasma membrane organization and function are discussed. It is suggested, that the organizational disposition of plasma membrane components other than phospholipids is at least related if not dependent upon the latter's asymmetric distribution in the bilayer.     

Changes of membrane phospholipid composition of human erythrocytes in hyperlipidemias. I. Increased phosphatidylcholine and reduced sphingomyelin in patients with elevated levels of triacylglycerol-rich lipoproteins.

The composition of red blood cell membrane and plasma phospholipids has been analyzed in patients with hyperlipidemias. In red cells of patients with elevated levels of triacylglycerol-rich lipoproteins, phosphatidylcholine (PC) was raised and sphingomyelin (SM) reduced, resulting in a 20% increase of the membrane PC/SM ratio. In plasma phospholipids of these patients PC and SM levels were also higher and lower, respectively and the plasma PC/SM ratio was elevated by more than 50%. Close positive correlations between plasma and membrane phospholipids were obtained for PC, SM and the PC/SM ratio in normolipidemic and hyperlipidemic donors. Plasmalogen phosphatidylethanolamine (PE), a supposed endogenous protector against lipid oxidation, was reduced by about 20% in red cell membrane lipids in hyperlipidemic patients. Also plasmalogen-PE in plasma tended to be reduced in hyperlipidemic donors. Plasma HDL levels were positively related to the content of plasmalogen PE in the red cell membrane. In conclusion, there are closely related increases in PC/SM ratios in plasma and the red cell membrane in patients with elevated levels of triacylglycerol-rich lipoproteins. It is speculated that decreases in red cell membrane plasmalogen-PE in hyperlipidemic patients could be related to impaired antioxidant protection, possibly as a consequence of reductions in plasma HDL levels.     

The effect of phosphate deficiency on membrane phospholipid composition of bean (Phaseolus vulgaris L.) roots

Plasma membranes were isolated from roots of bean (Phaseolus vulgaris L.) plants cultured on phosphate sufficient or phosphate deficient medium. The phospholipid composition of plasma membranes was analyzed and compared with that of the microsomal fraction. Phosphate deficiency had no influence on lipid/protein ratio in microsomal as well as plasma membrane fraction. In phosphate deficient roots phospholipid content was lower in the plasma membrane, but did not change in the microsomal fraction. Phosphatidylcholine and phosphatidylethanolamine were two major phospholipids in plasmalemma and microsomal membranes (80 % of the total). After two weeks of phosphate starvation a considerable decrease (about 50 %) in phosphatidylcholine and phosphatidylethanolamine in microsomal membranes was observed. The decline in two major phospholipids was accompanied by an increase in phosphatidic acid and lysophosphatidylcholine content. The effect of alterations in plasma membrane phospholipids on membrane function e.g. nitrate uptake is discussed.     

Essential fatty acids in erythrocyte phospholipids during pregnancy and at delivery in mothers and their neonates: comparison with plasma phospholipids

Evidence that the essential fatty acid (EFA) status during pregnancy and at birth may not be optimal is mainly based on fatty acid profiles of maternal and neonatal plasma phospholipids. However, erythrocyte phospholipids may be more reliable than plasma phospholipids to reflect the EFA status of an individual. Therefore, the present study compares the levels of EFA and of their derivatives (LCPUFA) in erythrocyte and plasma phospholipids collected during pregnancy and at delivery of 184 women and of their infants at birth. In general, the relative concentrations of erythrocyte and plasma phospholipid fatty acids (% of total fatty acids) were strongly correlated, but not at early pregnancy. The overall changes in fatty acid concentrations during pregnancy were qualitatively comparable between erythrocytes and plasma, although the comparability became less towards the end of pregnancy. The changes in absolute amounts (mg/l) of fatty acids in erythrocyte and plasma phospholipids also compare quite well till 32 weeks of gestation, but not thereafter. Most maternal-neonatal differences in relative fatty acid concentrations are qualitatively comparable for erythrocyte and plasma phospholipids. However, significant differences were observed for the absolute amounts of arachidonic and docosahexaenoic acids. No matter these differences, plasma and erythrocyte phospholipids seem equally suitable to reliably quantify the more functional EFA and LCPUFA status based on fatty acid ratios. Correlations between neonatal and maternal fatty acid values at delivery/birth are highly significant in erythrocyte as well as plasma phospholipids. Neonatal erythrocyte (but not plasma) values also correlated strongly with maternal values at early pregnancy. Therefore, the neonatal EFA and LCPUFA status might be predicted on the basis of EFA and LCPUFA concentrations of maternal erythrocyte phospholipids at early pregnancy.     

Plasma metabolite profiles: effects of dietary phospholipids in a migratory passerine (Zonotrichia leucophrys gambelii)

Plasma metabolites, including triglycerides, beta -hydroxybutyrate, and glycerol, can be used to estimate mass change in birds. Although dietary fatty acids can be ingested and absorbed as phospholipids, they have been largely overlooked as a potential indicator of mass change. The plasma ratio of triglyceride to phospholipid could also provide insight into diet quality because a high ratio in food items indicates high relative energy content. Variability in dietary phospholipid content and triglyceride : phospholipid may also affect the relationships between metabolites and mass change. We fed Gambel's white-crowned sparrows (Zonotrichia leucophrys gambelii) diets differing in phospholipid content and triglyceride : phospholipid and measured metabolites during mass loss and mass gain. Plasma phospholipids were higher and triglyceride : phospholipid was lower in birds fed a diet higher in phospholipid content and lower in triglyceride : phospholipid. Contrary to our expectations, plasma phospholipids were negatively related to mass change. Plasma triglyceride levels were positively related to mass change and unaffected by diet. The relationships between mass change and both plasma beta -hydroxybutyrate and glycerol were affected by diet. Plasma triglyceride appears to be the most reliable metabolite predicting body mass changes, but inclusion of plasma phospholipids and triglyceride : phospholipid into metabolite profiles may provide additional information on diet quality.     

Modification of the fatty acid composition of Ehrlich ascites tumor cell plasma membranes

The fatty acyl group composition of Ehrlich ascites tumor cell plasma membranes was modified by feeding the tumor-bearing mice diets rich in either coconut or sunflower oil. When coconut oil was fed, the oleate content of the membrane phospholipids was elevated and the linoleate content reduced. The opposite occurred when sunflower oil was fed. Qualitatively similar changes were observed in the plasma membrane phosphatidylethanolamine, phosphatidylcholine and mixed phosphatidylserine plus phosphatidylinositol fractions. These diets also produced differences in the sphingomyelin fraction, particularly in the palmitic and nervonic acid contents. Unexpectedly, the saturated fatty acid content of the plasma membrane phospholipids was somewhat greater when the highly polyunsaturated sunflower oil was fed. The small quantities of neutral lipids contained in the plasma membrane exhibited changes in acyl group composition similar to those observed in the phospholipids. These fatty acyl group changes were not accompanied by any alteration in the cholesterol or phospholipid contents of the plasma membranes. Therefore, the lipid alterations produced in this experimental model system are confined to the membrane acyl groups.     

Pattern of neutral and phospholipids in the semen of normospermic, oligospermic and azoospermic men

Total lipid concentration was elevated in the seminal plasma of oligo- and azoospermic men. The total cholesterol content was comparatively more in the seminal plasma of azoospermic men than in that of normo- and oligospermic men. In general, infertility was associated with increased seminal concentrations for most of the neutral lipid classes. However, total phospholipids and most of the phospholipid classes were diminished in the seminal plasma of oligo- and azoospermic men and in the spermatozoa of oligospermic men. We suggest that there is a positive correlation between seminal phospholipids and fertility and a negative correlation between seminal neutral lipids and fertility.     

Modification of the fatty acid composition of Ehrlich ascites tumor cell plasma membranes.

The fatty acyl group composition of Ehrlich ascites tumor cell plasma membranes was modified by feeding the tumor-bearing mice diets rich in either coconut or sunflower oil. When coconut oil was fed, the oleate content of the membrane phospholipids was elevated and the linoleate content reduced. The opposite occurred when sunflower oil was fed. Qualitatively similar changes were observed in the plasma membrane phosphatidylethanolamine, phosphatidylcholine and mixed phosphatidylserine plus phosphatidylinositol fractions. These diets also produced differences in the sphingomyelin fraction, particularly in the palmitic and nervonic acid contents. Unexpectedly, the saturated fatty acid content of the plasma membrane phospholipids was somewhat greater when the highly polyunsaturated sunflower oil was fed. The small quantities of neutral lipids contained in the plasma membrane exhibited changes in acyl group composition similar to those observed in the phospholipids. These fatty acyl group changes were not accompanied by any alteration in the cholesterol or phospholipid contents of the plasma membranes. Therefore, the lipid alterations produced in this experimental model system are confined to the membrane acyl groups.     

Fatty acid spectrum in the blood plasma, intestine, liver, and kidney tissues of calves with dyspepsia and method of its correction

The possibility of correction of fatty acid content of the blood plasma, intestine, liver and kidneys in 1 month-old calves which have suffered from dyspepsia by application of biologically active additive which contained a mixture of sunflower oil fatty acids and phospholipids is shown.     

An early childhood erythrocyte phospholipid distribution as further indication of persistence of neonatal erythrocyte characteristics in Diamond-Blackfan anemia

Comparisons of erythrocyte and plasma phospholipids made among adults, newborns, and a female patient with Diamond-Blackfan anaemia (DBA) revealed some indications for the continual existence of a neonatal phospholipid distribution in DBA. The relative percentage of phospholipids in erythrocytes and plasma were similar in newborns and in the female patient. The other peculiarities characteristic of newborns, such as deviations in the absolute phospholipid content, typical fatty acid patterns of phospholipids, could not be identified in DBA.     

Asymmetric distribution of arachidonic acid in the plasma membrane of human platelets. A determination using purified phospholipases and a rapid method for membrane isolation.

1. Non-lytic degradation of human platelet phospholipids have been performed using a combination of bee venom phospholipase A2 (phosphatide 2-acyl-hydrolase, EC 3.1.1.4) and Staphylococcus aureus sphingomyelinase C (sphingomyelin choline phosphohydrolase). Under these conditions, 25.4% of total phospholipds are degraded and 6.4% of total platelet arachidonic acid is released. 2. A new method for rapid isolation of platelet plasma membrane is described, based on the use of [3H]concanavalin A as a membrane marker and of self-generating gradients of Percoll. Plasma membranes are enriched 5.2 fold in lectin marker and 0.43 in N-acetyl-beta-D-glucosaminidase, the main contaminant. This method allows to estimate that 57% of the total cell phospholipids and 61% of the total arachidonic acid content are located in the plasma membrane. 3. The distribution of phospholipids and arachidonic acid between the two leaflets of the plasma membrane has been deduced by using these values and those obtained from non-lytic treatment of intact platelets by phospholipases. It is concluded that 45% of plasma membrane phospholipids, comprising 93% of sphingomyelin, 45% of phosphatidylcholine, 9% of phosphatidylserine, 16% of phosphatidylinositol and 20% of phosphatidylethanolamine form the outer half of the human platelet plasma membrane. The phospholipids appear to bear only 10% of the total membrane arachidonic acid.     

Induced alteration of rat erythrocyte membrane with effect of pyrethroid based compounds

The effects of tetramethrin and prallethrin exposure on plasma total proteins, free amino acids, albumins, urea, urea nitrogen, uric acid, creatinine were tested. Serum SGOT, SGPT and lipid profile, antioxidants super oxide dismutase (SOD), catalase, GSH, G-Px, phospholipids, cholesterol, C/P ratio in membranes of erythrocyte and membrane fluidity were analyzed. The reason of the study were analyzed to examine the possessions of mosquito repellent pyrethroid (MRP) based compounds tetramethrin and prallethrin exposure on plasma profile, antioxidant status of erythrocyte membrane, membrane fluidity in male Wistar rats. We tested chronically for three months exposure every day (continuously for 8–10 h per day by inhalation) of tetramethrin and prallethrin markedly available (MRP) repellents treated on male Wistar rats. Our results confirmed that tetrarmethrin and prallethrin treatment effect of plasma profile alterations, and lipid homeostasis mechanism in Red Blood cells (RBCs). Tetramethrin and prallethrin treatment significantly increased in erythrocyte membrane phospholipids and decreased levels of cholesterol with no change of protein content, increased C/P ration levels. Inhalation of tetramethrin and prallethrin stimulate plasma biophysical and biochemical modify SGOT, SGPT, erythrocyte membrane cholesterol and phospholipid levels, individual phospholipids and membrane fluidity of exposure rats compared to controls.     

Purified protein S contains multimeric forms with increased APC-independent anticoagulant activity

Protein S, the cofactor of activated protein C (APC), also expresses anticoagulant activity independent of APC by directly inhibiting prothrombin activation via interactions with factor Xa, factor Va, and phospholipids. In different studies, however, large variations in APC-independent anticoagulant activities have been reported for protein S. The investigation presented here shows that within purified protein S preparations different forms of protein S are present, of which a hitherto unrecognized form (<5% of total protein S) binds with high affinity to phospholipid bilayers (K(d) < 1 nM). The remaining protein S (>95%) has a low affinity (K(d) = 250 nM) for phospholipids. Using their different affinities for phospholipids, separation of the forms of protein S was achieved. Native polyacrylamide gel electrophoresis demonstrated that the form of protein S that binds to phospholipids with low affinity migrated as a single band, whereas the high-affinity protein S exhibited several bands that migrated with reduced mobility. Size-exclusion chromatography revealed that the slower-migrating bands represented multimeric forms of protein S. Multimeric protein S (<5% of total protein S) appeared to have a 100-fold higher APC-independent anticoagulant activity than the abundant form of protein S. Comparison of purified protein S preparations that exhibited a 4-fold difference in APC-independent anticoagulant activity showed that the ability to inhibit prothrombin activation correlated with the content of multimeric protein S. Multimeric protein S could not be identified in normal human plasma, and it is therefore unlikely that this form of protein S contributes to the APC-independent anticoagulant activity of protein S that is observed in plasma.     

Lipids of bovine adrenal plasma membranes.

The lipid composition of a membrane fraction from bovine adrenal cortex was determined. This preparation has the capacity to bind adrenocorticotropic hormone and is enriched in adenylate cyclase and 5'-nucleotidase. The adrenal plasma membranes have a significantly higher lipid content (54.8%) than bovine liver plasma membranes and a surprisingly low proportion of this lipid is cholesterol (4.2%). The phospholipids comprise 76.4% of the total lipids and their composition if very similar to that of bovine liver membranes with the exception of sphingomyelin which comprises only 4.5% of the phospholipids in the adrenal preparation.     

Isolation of very low density lipoprotein phospholipids enriched in ethanolamine phospholipids from rats injected with Triton WR 1339

Phospholipids carried by very low density lipoprotein (VLDL) are hydrolysed in circulation by lipoprotein and hepatic lipases and lecithin-cholesterol acyltransferase. We have previously demonstrated [J.J. Agren, A. Ravandi, A. Kuksis, G. Steiner, Structural and compositional changes in very low density lipoprotein triacylglycerols during basal lipolysis, Eur. J. Biochem. 269 (2002) 6223-6232] that the infusion of Triton WR 1339 (TWR), which inhibits these lipases, leads in 2 h to five-fold increase in VLDL triacylglycerol concentration along with major differences in the composition of their molecular species. The present study demonstrates that the accumulation of triacylglycerols is accompanied by major changes in the content of the VLDL phospholipids, of which the most significant is the enrichment of phosphatidylethanolamine (PtdEtn). This finding coincides with the enrichment in PtdEtn demonstrated in the VLDL of a hepatocytic Golgi fraction but it had not been demonstrated that the Golgi VLDL, along with its unusual phospholipid composition, can be directly transferred to plasma. Aside from providing an easy access to nascent plasma VLDL, the TWR infusion demonstrates that lipoprotein and hepatic lipases are also responsible for the degradation of plasma VLDL PtdEtn, as independently demonstrated for plasma phosphatidylcholine. Our results indicate also, with the exception of lysophosphatidylcholine, that preferential basal hydrolysis no dot lead to major differences in molecular species composition between circulating and newly secreted VLDL phospholipids. The comparison of the molecular species composition of VLDL and liver phospholipids suggests a selective secretion of PtdEtn and sphingomyelin molecular species during VLDL secretion.     

Comparative study of lipids and fatty acids in blood plasma of river lamprey Lampetra fluviatilis and brown frog Rana temporaria at the periods of elimination of exogenous feeding

Lipids of blood plasma of lampreys and frogs are composed of phospholipids, triglycerides, free fatty acids (FA), cholesterol, cholesterol esters, and waxes. The lipids content in plasma of frogs is markedly lower as compared with that of lampreys. However, the percentage of lipid components is represented by close values. Fluidity of triglycerides and phospholipids in lampreys is determined predominantly by monoenic acids and polyenoic acids of the ω-3-type, whereas that in frogs—by monoenic acids and polyenic acids of the ω-6-type. Free FA are represented mainly by saturated and monoenic acids.     

Decreased content of docosahexaenoate and arachidonate in plasma phospholipids in Usher's syndrome

Docosahexaenoate and arachidonate were found to be significantly decreased in plasma phospholipids from Usher's syndrome patients. The fatty acid content of plasma triacylglycerols was not changed in these patients. Usher's syndrome, an autosomal recessive disorder, involves an inherited visual cell degeneration. Photoreceptor membranes are richly endowed with docosahexaenoate and arachidonate, and a metabolic defect affecting these polyunsaturated fatty acids may occur. Moreover, blindness may be due, at least partially, to an alteration in the unsaturated phospholipids of photoreceptor membranes.