Phospholipid Cholesterol Research Articles

A compensatory mechanism improving red cell membrane fluidity in hemodialysed patients

A fluorescence polarization technique with 1,6-diphenyl 1,3,5-hexatriene as a fluorescent probe was used to determine the fluidity of red cell membranes from hemodialysed patients before and after dialysis. After dialysis, there was a decrease in fluidity and activation energy values revealed a significantly changed distribution. The membrane lipid composition showed a significant increase in cholesterol after dialysis ( p < 0.001) and a significant reduction in saturated fatty acids ( p < 0.01) with an increase in unsaturated fatty acids ( p < 0.05). A compensatory mechanism could be suggested involving a reduction of saturated fatty acids in response to the increase in the cholesterol phospholipid ratio. This could lead to an optimization of membrane fluidity.

The role of membrane composition in ATPase activities of cirrhotic rat liver: Effect of silymarin

The activities of Ca2(+)- and Na+, K(+)-ATPases were studied in liver plasma membranes from CCl4-cirrhotic rats and from livers of rats treated with silymarin in addition to CCl4. CCl4 chronic treatment produced significant decreases in Na+, K(+)- and Ca2(+)-ATPase activities; however, the animals treated with silymarin along with CCl4 showed no differences in ATPase activities as compared to controls. The lipid analysis performed in plasma membranes revealed increases in the cholesterol/phospholipid (CH/PL) and sphingomyelin/phosphatidylcholine (SM/PC) ratios in the cirrhotic group. Again, the membranes isolated from rats receiving CCl4 + silymarin showed normal CH/PL and SM/PC values. Considering that CH/PL and SM/PC ratios are related to membrane microviscosity, this study suggests that a lower fluidity of the membrane may be responsible for the observed decreases in ATPase activities in the cirrhotic group. Additionally, the role of silymarin to improve liver function in CCl4-cirrhosis can be attributed partially to its action at membrane level by preventing the increases in CH/PL and SM/PC ratios.

Alteration in 1,2-diacylglycerol level and its fatty acid composition in hearts during the growth of hamsters

We measured the amount of 1,2-diacylglycerol (DAG) and its fatty acid composition in hamster hearts of various ages because 1,2-DAG has been recognized to be one of the intracellular second messengers. The highest level in 1,2-DAG was detected in 30-day-old hamster hearts. The heart weight increased about 2.5 times between 30 and 90 days of age. Hamster hearts at 90 days of age had 72% of the 1,2-DAG content at 30 days of age. The amount of 1,2-DAG at 240 days of age was a little lower than that in 90-day-old and 160-day-old hamsters, whereas there was no significant increase in heart weight and only a little gain in body weight between 90 and 240 days of age. The analysis of the fatty acid composition of 1,2-DAG showed an increase in the percentage of 18:0 accompanied by a change in the percentages of other fatty acids with the growth of hamsters. The largest amount of arachidonic acid expressed as 20:4 (n-6) was found at 30 days of age. On the other hand, triglyceride, cholesterol, and major phospholipid components of hamster hearts increased in accordance with growth. In particular, the increase in triglyceride content was remarkable, indicating that myocardial lipidosis may be induced by aging. These results demonstrate that 1,2-DAG may be involved in the synthesis of protein during heart growth and that the alteration of its fatty acid composition is related to the growth of hamsters.

Differences in lipid composition between isolated growth cones from the forebrain and those from the brainstem in the fetal rat

The lipid composition of nerve growth cone membranes isolated from rat fetal forebrain or brainstem by the sucrose density gradient method was analyzed biochemically and immunochemically. In the forebrain, growth cone membrane (GCM) contained lower levels of gangliosides than those from other heavier fractions, but it was not the case in the fetal brainstem at the same developmental stage. The distinctive features in the ganglioside composition of GCM are the predominance of GD3 and the presence of c-series gangliosides that are due to fetal expression in mammals. A unique acidic glycolipid, sulfoglucuronylparagloboside (SGPG), which is not present in adult brains, was first detected in both forebrain and brainstem GCM. Including such minor species, the ganglioside composition in forebrain or brainstem GCM was almost identical to other membrane fractions from the forebrain or brainstem. The compositional ratios of the major lipid classes in membranes, cholesterol and phospholipids, seemed to be common to forebrain GCM and brainstem GCM, as indicated by the identical values of phospholipid-to-protein (PL/Pr), cholesterol-to-protein (Ch/Pr), and cholesterol-to-phospholipid (Ch/PL) ratios for both. This study has revealed that GCM isolated from forebrain which is supposed to be at an earlier stage of neuronal differentiation than brainstem has less amounts of total gangliosides, high proportion of GD3 to GD1a and enriched c-series gangliosides as compared to brainstem GCM.

Effects of antithrombin III on glomerulonephritis induced by antibasement membrane antibody in rats.

Antithrombin III (AT III), a plasma inhibitor of coagulation factors, was evaluated for its inhibitory effects on the development of glomerulonephritis induced by antibasement membrane antibody in rats. AT III was administered intravenously at 250 and 1000 Units/kg to rats once a day for 14 days after intravenous injection of rabbit antiserum against the rat glomerular basement membrane. In the rat model, the high dose of AT III reduced the urinary total protein, plasma total cholesterol and plasma phospholipid, and tended to lower the incidence of fibrin deposits in Bowman's space, cell proliferation in the glomeruli, and PAS-positive granules (absorption droplets) in the glomerular epithelium. The above results suggest that AT III is effective in inhibiting the development of glomerulonephritis.

PLASMA MEMBRANE LIPIDS MODULATE THE RESPONSE TO WATER DEPRIVATION IN RAT KIDNEY

The lipid composition of biological membranes determines many of the cellular responses to stimuli such as hormones and drugs. It is known that the neonate has differences in renal function compared to the adult individual, which determine the characteristics of pharmacokinetics and pharmacodynamics at this age. We studied the lipid composition of renal plasma membranes, the renal response to water deprivation, and the activity of plasma membrane gamma-glutamyl transpeptidase (GGTP) in newborn, 21-d-old, and adult rat. It was found that the cholesterol/phospholipid (CH/PL) ratio of newborn membranes was significantly higher than that of membranes from 21-d-old and adult rats, which were similar. These findings paralleled low ability to concentrate urine in the newborn and higher sensitivity of the neonatal kidney to water deprivation. Kidney membranes from 21-d-old and adult rats showed similar lipid composition, and the response to water deprivation was also similar. The changes observed in GGTP activity at different ages varied with the cholesterol content of kidney plasma membranes. These findings suggest participation of the lipid composition of kidney membranes in modulation of the renal response to water deprivation.

The ultraviolet derivative spectrophotometric determination of neutral liposome-entrapped β-lactam antibiotics

A simple, non-invasive spectrophotometric assay to measure the concentration of some β-lactam antibiotics in turbid solutions containing liposomes was carried out. Since zero-order spectra gave strong interference because of sample turbidity, derivative spectrophotometry was used to enhance the spectral details. Derivative spectra showed bands in the ultraviolet region due to the presence of the cephalosporin and penicillin β-lactams. A linear relationship between derivative amplitudes and antibiotic concentration was found when antibiotic-containing liposome solutions were measured. A saturative trend in the liposome-encapsulation was observed. The antibiotic entrapment was lowered by increasing the cholesterol—phospholipid ratio in the mixture used for liposome preparation. After treatment of antibiotic-loaded liposomes with β-lactamase, a hydrolytic enzyme specific for β-lactams, the remaining antibiotic concentration decreased significantly, showing that some of the antibiotic was retained on the outer surface of the vesicles.

The evolution of membranes

Consideration of the influence of cholesterol on the physical properties of biological membranes leads to the conclusion that cholesterol increases the thickness of fluid membrane bilayers without appreciably increasing the microviscosity component of membrane fluidity. At sufficiently high cholesterol concentrations, the gel–liquid crystalline phase transition is completely eliminated in phospholipids–cholesterol mixtures and the system has the properties of a two-dimensional liquid over a wide range of temperatures. It is proposed that with the evolution of cholesterol and related sterols in an oxygen-rich atmosphere, the resulting modification of physical constraints on membrane properties made it possible for new evolutionary driving forces to manifest themselves leading to the peculiar properties of plasma membranes of eucaryotic cells.

The effects of l-cycloserine on membrane fluidity and lipids of mouse brain

l-Cycloserine can inhibit the synthesis of some sphingolipids, which may play a role in determining membrane fluidity. To evaluate the effects of l-cycloserine on neural cell membranes, adult mice were treated with 100 mg/kg of this drug twice daily for 1, 2, 4, 6, 13 and 17 days. Body weights in the treated mice decreased significantly. Cholesterol and total phospholipid per mass total lipid significantly increased with time. Fluorescence depolarization after the incorporation of diphenylhexatriene into membranes was measured in order to assess membrane fluidity. it increased significantly starting at treatment day 4, with a maximum increase in polarization value of 8%. These results show that l-cycloserine significantly changes the fluidity of brain membrane, but this does not correlate with changes in major lipid classes of the brain.

Diets alter jejunal morphology and brush border membrane composition in streptozotocin-diabetic rats.

Previous studies have demonstrated enhanced active and passive uptake of many nutrients in animals with experimental diabetes. These changes in absorption cannot be explained by differences in intestinal morphology, although the brush border membrane (BBM) phospholipids do change in diabetes. Manipulation of diet produces alterations in intestinal uptake of lipids and glucose. This study was undertaken to determine the effect of diet and diabetes on jejunal morphology and BBM lipid composition. Rats were rendered hyperglycemic with streptozotocin and were fed for 2 weeks on a diet that was high or low in carbohydrate, essential fatty acids, cholesterol, or protein. In both control and diabetic rats, these diets produced changes in villus height and BBM sucrase and alkaline phosphatase activities. In both control and diabetic rats, BBM phospholipids were unaffected by changes in the dietary content of essential fatty acids, cholesterol, or protein, but total BBM phospholipid content was reduced in animals fed low as compared with high carbohydrate diet. Total BBM phospholipid content was higher in diabetic than in control animals fed the low protein diet, whereas BBM phospholipid content was lower in diabetic than in control animals fed the high carbohydrate diet, and was even lower in diabetic animals fed the low as compared with the high carbohydrate diet. These changes in total phospholipids were due to alterations in the BBM content of phospholipids containing choline. In control animals, BBM cholesterol was higher in rats fed the low as compared with the high cholesterol diet, or the low as compared with the high protein diet.(ABSTRACT TRUNCATED AT 250 WORDS)

Chronic triiodothyronine supplementation does not improve the lipoprotein disorders of mildly uremic rats.

The present study was performed to appreciate the potential role that thyroid deficiency could play in the energy homeostasis and lipid metabolism of experimental chronic renal failure. For this purpose, 12 uremic rats that were supplemented with T3 (0.4 microgram/100 g body weight/day) during 5 weeks by means of osmotically driven minipumps were compared to 12 unsupplemented uremic rats and 12 control rats. The chronic supplementation of uremic rats with T3 induced no significant change in body weight gain or in the serum concentration of insulin, glucose, glycerol, nonesterified fatty acids, total triglycerides (TG), total cholesterol, and total choline phospholipids. Similarly, the metabolism of TG-rich lipoproteins was not affected by the supplementation with T3 in these uremic rats as appreciated by TG production or TG degradation (adipose tissue lipoprotein lipase activity). T3 administration induced a significant decrease in serum beta-hydroxybutyrate concentration and an increase in serum lactate concentration. Furthermore, heparin-releasable hepatic TG lipase activity as expressed per total liver mass was decreased in uremic rats treated with T3. The latter changes were observed in the absence of modifications of serum glucose or TG concentration. We conclude from these observations that rats with a moderate degree of chronic uremia do not seem to have a cellular thyroid deficiency sufficient to disturb their energy or lipid metabolism.

Plasma and red cell lipids in alcoholics with macrocytosis

The cholesterol and phospholipid content and the fatty acid composition in plasma and red cell membranes was determined in 10 alcoholics with macrocytic erythrocytes. None of the patients had anemia. Red cells exhibited macrocytosis up to 108 fl in all patients. Bilirubin, albumin, prothrombin, and cholinesterase were in the normal range, whereas transaminases and gamma-glutamyl transpeptidase activities in serum were elevated in most of the patients. The molar ratio cholesterol phospholipids in red cells was not altered in alcoholics. An abnormally high ratio of saturated unsaturated fatty acids was found in plasma as well as in red cell phospholipids from alcoholics. Linoleic acid was substantially decreased in plasma of alcoholics (controls 32.3%, alcoholics 21.8%). This fatty acid abnormality was reflected by a decrease of linoleic acid in red cell phosphatidylcholine. The present data may suggest that fatty acid changes taking place in membranes of macrocytes were a consequence of changes in the plasma and reflect plasma membrane exchanges rather than direct effects of ethanol on red cell membranes. Lipid alterations of red cell membranes may be involved in the development of macrocytosis in chronic alcoholism.

Effect of chronic ethanol and food deprivation on intestinal villus morphology and brush border membrane content of lipid and marker enzymes.

Brush border membranes (BBM) were isolated from the jejunum and ileum of control, ad libitum (CAL); control, food-restricted (CFR); control, weight gain (CWG); and ethanol-fed (EF) rabbits. Jejunal alkaline phosphatase activity was similar among control groups, but higher in CAL than EF animals. Sucrase activity was higher in EF and CWG animals than in CAL and CFR. The alkaline phosphatase/sucrase ratio was lower in EF than control animals. Ileal enzyme marker activity was similar among EF and control animals. Sucrase (S) activity was lower in the ileum than in the jejunum. Jejunal free fatty acid and phospholipid/cholesterol (PL/C) were lower in EF than control animals, whereas ileal lipid content was generally similar among all animal groups. Total phospholipid content was similar between sites, but the cholesterol and free fatty acid content were lower in the ileum than the jejunum. The phospholipid/cholesterol ratio was increased only in the ileum of EF animals. The amount of lecithin was decreased in the jejunal BBM of EF animals resulting in a decreased choline/amine phospholipid ratio as compared with control animals. The ileal phospholipid composition was similar among all groups. A large increase in villus height is observed in the jejunum of EF animals. Villus surface area and mucosal surface area are altered with ethanol feeding and food deprivation. Thus, (i) there is a gradient of S and cholesterol between the BBM of jejunum and ileum; (ii) changes in food intake are associated with changes in the morphology as well as the enzyme marker and lipid content of BBM.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylate cyclase and a fatty acid spin probe detect changes in plasma membrane lipid phase separations induced by dietary manipulation of the cholesterol:phospholipid ratio

Rats fed with a cholesterol supplement to their diet exhibited an increase in their plasma membrane cholesterol phospholipid ( C/P)-lipid molar ratio from 0.72 to 0.98, whereas those fed the hypocholesterolaemic drug clofibrate in their diet exhibited a decrease in this ratio to 0.62. The properties of these membranes were analysed with regard to ligand-stimulated adenylate cyclase activity and the mobility of a fatty acid spin probe which allowed lipid phase separations to be identified. Membranes with elevated C/P ratios exhibited two distinct lipid phase separations, one at around 36°C that was attributed to the external half of the bilayer and one at around 22°C which was attributed to the inner half of the bilayer. Membranes with lowered C/P ratios exhibited a single lipid phase separation occurring at around 21°C which was attributed to the lipids of the inner half of the bilayer. These results were compared with those obtained by manipulation of C/P ratios in vitro using liposome-cholesterol exchange techniques. Dietary manipulation of the C/P ratio of plasma membranes in vivo led to alterations in the fold stimulation of adenylate cyclase by various stimulatory ligands. Adenylate cyclase Cholesterol Membrane fluidity Glucagon Dietary manipulation Clofibrate

Lipid composition and monooxygenase relationship in liver microsomal membranes from rats fed unbalanced diets

The cholesterol and phospholipid content of microsomal membrane from rats fed either a high lipid (Hl) (30% lard) or a low protein (Lp) diet (6% casein) have been compared with those from rats fed a standard (St) diet (22% casein, 5% lard). For each diet, half of the group was treated with Phenochlor DP6. A significant increase in the phospholipid cholesterol ratio was observed in rats fed a high lipid diet or treated with DP6. These effects tend to increase the microsomal membrane fluidity. The protein deficiency decreased the phospholipid/cholesterol ratio and then the fluidity of the endoplasmic reticulum membrane. The specific activity of cytochrome P-450 to hydroxylate aniline which is independent to the enzymic form of cytochrome P-450 was closely correlated with the viscosity status of the microsomal membrane.

Interaction of hapten-sensitized liposomes with cells bearing hapten-specific receptors.

The binding of liposomes sensitized with 2,4-dinitrophenyl-6-N-aminocaproylphosphatidylethanolamine (DNP-Cap-PE) to MOPC-315 cells, which secrete and bear on their surfaces anti-DNP immunoglobulins, was studied. The binding was affected by cholesterol content, phospholipid composition and hapten density of liposomes: The binding of distearoylphosphatidylcholine liposomes sensitized with 5 mol% hapten to the cells increased with increasing cholesterol content in liposomes. The amount of liposomes composed of phospholipid with a higher transition temperature (such as distearoylphosphatidylcholine), which bound to MOPC-315 cells, was much higher than that of liposomes composed of phospholipid with a lower transition temperature (such as egg yolk phosphatidylcholine). The amount of distearoylphosphatidylcholine liposomes containing equimolar cholesterol, which bound to the cells at 0 degrees C, increased with increasing amount of the hapten in liposomes up to 2.5 mol%. The binding became maximum at 2.5 mol% and decreased with higher hapten density in liposomes. The immunogenicity of hapten-sensitized liposomes is known to be affected by the liposomal composition such as cholesterol content, phospholipid composition and hapten density. This model study suggests that the binding of liposomes to cells is important for expressing the immunogenicity of hapten-sensitized liposomes.

Alteration of synaptic membrane cholesterol/phospholipid ratio using a lipid transfer protein. Effect on gamma-aminobutyric acid uptake.

A procedure was developed to vary the cholesterol-to-phospholipid (Ch/PL) ratio of synaptic plasma membranes and synaptosomes using a nonspecific lipid transfer protein so that membrane lipid composition could be correlated with presynaptic function. In synaptic plasma membranes, Ch/PL molar ratios from 0.21 to 1.19 were produced from a normal value of 0.52 +/- 0.01 by incubation with the transfer protein and an excess of either phosphatidylcholine or cholesterol/phosphatidylcholine liposomes for 60 min at 32 degrees C. In synaptosomes, Ch/PL ratios from 0.16 to 0.81 were similarly produced from a normal value of 0.38 +/- 0.04. Cholesterol loading or depletion of the membranes was accompanied by a decrease or increase, respectively in the phospholipid-to-protein ratio. The fluidity of the synaptic plasma membrane, as estimated by 1,6-diphenylhexatriene anisotropy measurements, was increased by lowering the Ch/PL ratio and decreased by raising the Ch/PL ratio. Decreasing the Ch/PL ratio of synaptosomes and synaptic plasma membrane vesicles resulted in loss of sodium-dependent gamma-aminobutyric acid (GABA) uptake (70-100% loss at Ch/PL ratios decreased to 40% of normal) and reduction in the number of accessible GABA-binding sites. Choline uptake was not affected in these same preparations. GABA uptake was restored by reinserting cholesterol into the membrane. Synaptosomal membrane potential and synaptic plasma membrane sodium permeability were not affected by changing the Ch/PL ratio. Increase in the Ch/PL ratio above normal had no effect on either choline or GABA uptake. Both the decrease in the Ch/PL ratio and the increase in the lipid-to-protein ratio increase membrane "fluidity," lipid-to-protein ratio increase membrane "fluidity," which may modulate the vertical displacement and motional characteristics of the GABA transporter.

56] Use of a nonspecific lipid transfer protein to modify the cholesterol content of synaptic membranes

This chapter describes the use of a nonspecific lipid transfer protein to vary the cholesterol-to-phospholipid (Ch/PL) ratio of membranes. The method is illustrated with synaptosomes and synaptic plasma membranes, where loss of sodium-dependent γ-aminobutyric acid uptake is correlated with decreased Ch/PL ratio in the membrane. The method consists of incubating the membranes with the transfer protein in the presence of lipid vesicles containing either cholesterol and phosphatidylcholine (PC) or PC alone, resulting in net transfer of cholesterol to the membranes or net removal of cholesterol from the membranes, respectively. The nonspecific lipid transfer protein is purified from beef liver. The transfer protein is prepared without the heat treatment and octylagarose column chromatography steps. This shortened procedure increases the yield of transfer activity. Assay of nonspecific lipid transfer activity is based on measurement of transfer of radiolabeled cholesterol to heat-treated beef heart mitochondria. A trace of radiolabeled triolein is included in the liposomes as a nontransferable marker.

Band 3 protein—cholesterol interactions in erythrocyte membranes: Possible role in anion transport and dependency on membrane phospholipid

Band 3 protein of the human erythrocyte membrane, the anion transport protein, possesses a high affinity steroid binding site. In mixed phospholipid—cholesterol monolayers, the state of occupancy of this site is positively correlated with their cholesterol and sphingomyelin content and negatively with their glycerophospholipid content. We suggest that, in the erythrocyte membrane, the binding site is an inhibitory site of anion transport and that the modulation of its state of occupancy by the membrane lipid is responsible for the negative correlation of anion transport with the membrane's content of cholesterol and sphingomyelin and the positive correlation with the phosphatidylcholine content

Effects of the type of dietary fat at two levels of vitamin E in Wistar male rats during development and aging. IV. Biochemical and morphometric parameters of the heart

To study the possible influence of the type of dietary fat at two extreme levels of vitamin E on several biochemically determined changes in the whole heart of rats, as well as on a number of morphometrically analyzed structural and ultrastructural variations of fight and left ventricular myocytes (RVM and LVM), six groups of weanling Wistar male rats were fed ad libitum isoenergetic diets containing similar amounts (15 g per 100 g of diet) of saturated fat (coconut oil), unsaturated fat (safflower oil) or a combination of both at two levels of dl- α-tocopherol (2 or 200 mg per 100 g of diet). Most determinations were performed at 3, 6, 12, 18 and 24 months. While the cardiac concentrations of total tocopherols directly correlated with the dietary levels of vitamin E and were also influenced by the type of dietary fat, neither of these two dietary factors had any consistent effect on the other biochemical parameters studied (DNA, RNA, total protein, triglycerides, cholesterol phospholipids or total collagens). Although, irrespective of the type of diet, the levels of several of the biochemical parameters fluctuated with time, comparisons of the results at different periods showed as the most significant changes increases in cardiac DNA, RNA and total collagens between 12 and 24 months, as well as decreases in phospholipids and total protein between 3 and 24 months. The in vitro lipoperoxidative potentials (malonaldehyde production) fluctuated with time and reached peak values at 18 months, but in vivo lipoperoxidation (diene conjugation) was never detected in the cardiac lipids of rats from the different groups. The morphometric studies indicated that, with few exceptions, the diameters of LVM and RVM, the numerical and volumetric densities of mitochondria and the volumetric density of myofilaments in RVM and LVM remained practically unchanged with age. On the other hand, the volumetric density of lipofuscin pigment increased significantly with time and at the same ratein RVM and LVM. While the nutrition data suggested that the consumption of the safflower-oil diet with high levels of vitamin E was associated with a significant increment in the numerical density of mitochondria in LVM, the type of diet did not significantly influence the other age-related ultrastructural variations.