Lipids of Erythrocyte Membranes of COPD Patients: A Quantitative and Qualitative Study

Abstract

Chronic obstructive pulmonary disease (COPD) is characterized by inflammation of lung parenchyma and pulmonary hypoxemia with a proven systemic component. Tobacco smoke is the most important risk factor and plasma membrane plays a major role in the disease pathology and progression. The properties of biological membranes are a function of their lipid composition. Any change in its composition may lead to the pathophysiology. In COPD research, erythrocytes are emerging as a new therapeutic venture, as their shape and properties change in the disease. Therefore we studied the lipid composition of the erythrocyte membranes of COPD patients. The study included 30 patients having COPD, 10 healthy smokers and 10 non-smokers. Erythrocytes were separated from peripheral blood and their membranes prepared, followed by estimation of proteins, cholesterol and phospholipids. Individual phospholipids were identified and separated by TLC and fatty acid composition determined by gas chromatography. The data were analyzed statistically and P < 0.05 was considered significant. Our results demonstrate that in very severe COPD, proteins decrease, whereas phospholipids and cholesterol contents increase significantly, which showed a consistent negative correlation with FEV1%. The fatty acid analysis showed preponderance towards saturated fatty acids mainly arachidic and behenic acid, suggesting a decrease in membrane fluidity or a closer packing of lipid rafts. We are the first to report about preponderance of saturated fatty acids in plasma membrane of erythrocytes of COPD patients which may decrease the membrane fluidity and possibly impair the functions of the plasma membrane in the disease.     

Effect of oral treatment with pantethine on platelet and plasma phospholipids in IIa hyperlipoproteinemia

In a single-blind, crossover, completely randomized study, the effects of oral treatment with pantethine or placebo on fatty acid composition of plasma and platelet phospholipids were investigated in 10 IIa hyperlipoproteinemic patients. A significant decrease of total cholesterol and total phospholipids was observed both in plasma and in platelets after a twenty-eight-day treatment. In plasma, pantethine induced a decrease of the ratio sphingomyelin/phosphatidylcholine. Moreover, a relative increase of n3-polyunsaturated fatty acids both in plasma and in platelet phospholipids and a decrease of arachidonic acid in plasma phospholipids were observed. These results indicate that pantethine can affect plasma and platelet lipid composition with possibly favorable influences on the determinants of cell membrane fluidity.     

The composition of plasma lipoproteins and erythrocyte membranes in cholesterol-fed pigs with partial ileal bypass

The composition of plasma lipoproteins and erythrocyte membranes was studied in cholesterol-fed pigs with a partial ileal bypass. Cholesterol feeding caused marked increases in the plasma concentrations of cholesterol and phospholipids. In spite of continuation of cholesterol feeding, PIB reduced plasma concentrations of cholesterol and phospholipids towards basal values. PIB completely counteracted the dietary cholesterol induced alterations in the lipid composition of the apoprotein B containing plasma lipoproteins, but not in the HDL2 fraction. It is suggested that PIB specifically influences the metabolism of the atherogenic, apoprotein B containing lipoproteins. Dietary cholesterol caused significant increases in the ratios of cholesterol:phospholipids and phosphatidylcholine: sphingomyelin in erythrocytes. The high-cholesterol diet also increased the content of linoleic acid in erythrocyte phosphatidylcholine. PIB completely nullified the cholesterol-induced increase in the cholesterol:phospholipid ratio, but not the increase in the phosphatidylcholine:sphingomyelin ratio. The percentage of linoleic acid in erythrocyte phosphatidylcholine was unaffected by PIB. Neither cholesterol feeding nor PIB had an effect on the lipid fluidity of erythrocyte membranes, as measured by fluorescence polarization, using the probe diphenylhexatriene. Possible compensatory mechanisms operating to control homeostasis of lipid fluidity of erythrocyte membranes are discussed.     

Erythrocyte lipids in triose-phosphate isomerase deficiency.

Marked hypoalphalipoproteinemia was found together with relatively low serum cholesterol, triacylglycerol, and LDL levels in a triose-phosphate isomerase (TPI; D-glyceraldehyde-3-phosphate ketol-isomerase, EC 5.3.1.1)-deficient Hungarian family, especially in the two compound-heterozygote brothers. Apart from a slight increase in palmitic and stearic acids together with a slight decrease in oleic and linoleic acids, no other changes were found in the fatty acid composition of the erythrocyte phospholipids. Anisotropy measurements with n-(9-anthroyloxy) stearic and -palmitic acid fluorophores revealed increased motional freedom of the fatty acid chains in the external lipid layers of the intact erythrocytes from all members of the TPI-deficient family as compared with normal age-matched controls. This asymmetric increase in membrane fluidity was found to be significantly higher in the propositus than in his compound-heterozygote brother without any neurological disorders. The change in membrane fluidity may result from as-yet-unresolved aspects of the lipid composition of the plasma membrane. Our findings that the differences between the TPI-deficient individuals and normal controls and the differences between the two compound-heterozygote brothers were all absent in the phospholipid extracts of the same erythrocytes favor the assumption that the increased motional freedom of the fatty acid chains in the external surface of the bilayer is caused by the binding of the mutant TPI molecule to the N-terminal sequence of band 3 protein.     

Alteration of erythrocyte membrane lipid fluidity in human obesity

The lipophilic probe 1,6-diphenyl-1,3,5-hexatriene was incorporated into erythrocyte ghosts of either normal or obese humans, and the polarization of fluorescence was measured between 0 and 40 C. The membrane lipid fluidity, evaluated by fluorescence polarization, was consistently higher in the ghosts from obese subjects. A strong correlation was found between increased 1,6-diphenyl-1,3,5-hexatriene fluorescence polarization and excess body weight. Measurements of cholesterol and phospholipids indicated increased cholesterol and decreased phospholipids in erythrocyte ghosts from obese subjects. These data suggest that alterations in lipid composition in erythrocytes of obese subjects are responsible for abnormal physical properties of plasma membranes, which, in turn, may cause altered enzymatic activities.     

Comparison of changes in erythrocyte and platelet phospholipid and fatty acid composition and protein oxidation in chronic obstructive pulmonary disease and asthma

OBJECTIVE: To analyse and compare the phospholipid and fatty acid composition of total lipids and the occurrence of lipid peroxidation and protein oxidation directly in erythrocytes or platelets from chronic obstructive pulmonary disease (COPD) and asthma patients. PATIENTS: Fifteen consecutive outpatients with COPD (all smokers) and asthma (non-smokers) recruited during a moderate-to-severe (COPD) or moderate (asthma) exacerbation. Fifteen subjects with smoking habits similar to those of COPD patients were studied as a control group. METHODS: Phospholipid and total fatty acid compositions were analysed by two-dimensional thin layer chromatography or gas chromatography-mass spectrometry, respectively. The lipid fluorescence of lipid extracts was measured by spectrofluorimetry. Protein carbonyl contents and profiles were measured by immunoblot detection. RESULTS: No differences were found either in erythrocyte or platelet cholesterol or phospholipid levels. Only a decrease in the content of phosphatidylserine + phosphatidylinositol (P<0.003) was detected in platelets from the asthma patients. In erythrocytes, the fatty acid profile changed in both lung pathologies, especially as regards polyunsaturated fatty acids (decreases in arachidonic and 22:4 fatty acid contents). Other observed changes were: COPD, an increase in palmitic fatty acid; asthma, an increase in oleic and decreases in eicosapentaenoic and 22:6 + 24:1 fatty acids. In platelets, the fatty acid profiles revealed many differences between both lung pathologies: COPD, a decrease in 18:1 and increases in 20:5 and 22:5 + 24:0; asthma, a decrease in 20:4 and increase in 22:6 + 24:1. In COPD vs. asthma patients, fatty acid changes were mainly detected in platelets, especially in 18-carbon species, with decreases in stearic and 18:1 fatty acids in the COPD patients. Protein oxidation levels were increased in both lung pathologies in both erythrocytes and platelets. CONCLUSIONS: COPD and asthma are associated with common or specific changes in the lipid composition of erythrocytes and/or platelets. The data point to lipid peroxidation and protein oxidation phenomena in both types of blood cell, although platelets would be more susceptible to stress.     

Decreased membrane fluidity in erythrocytes from patients with Crohn's disease

In erythrocytes from patients with Crohn's disease, lower levels of membrane fluidity were demonstrated by means of electron spin resonance with 16-stearic acid spin label (SAL) but not 5- or 12-SAL, suggesting more apparent abnormality in the deeper portion of lipid bilayer. Membrane lipid analysis showed a significant increase of sphingomyelin, a decrease of phosphatidylcholine and decreased contents of polyunsaturated acyl-chains, particularly C18:2, of phospholipid, without alteration in the cholesterol to phospholipid molar ratio. The alteration in membrane fluidity appeared to be closely related to changes in membrane phospholipid classes and their acyl-chains. Among the plasma lipids, contents of esterified cholesterol, phospholipid, very low- and low-density lipoproteins and cholesterol of high-density lipoprotein significantly decreased. Decreased contents of polyunsaturated acyl-chains, particularly C18:2, in the plasma phospholipid were also noted. Changes in plasma and erythrocyte membrane lipids remained even in the inactive stage of the disease, except for the normalized sphingomyelin and phosphatidylcholine content in the erythrocyte membrane. Since membrane fluidity is closely involved in various cell membrane functions, the decreased membrane fluidity and altered lipid composition in erythrocyte membrane evident in patients with Crohn's disease may have important effects on the pathological status in this disease.     

Peripheral Blood Mononuclear Cell Membrane Fluidity and Disease Outcome in Patients with Multiple Sclerosis

Immune cell membrane lipids are important determinants of membrane fluidity, eicosanoid production and phagocytosis and fatty acid metabolic abnormalities have been reported in immune cells from patients with multiple sclerosis. The aim of this study was to investigate the relationship between peripheral blood mononuclear cell membrane fluidity, permeability status, and disease outcome as measured by the Kurtzke expanded disability status scale. Phospholipids, fatty acids and cholesterol composition in peripheral blood mononuclear cells from 26 patients diagnosed with multiple sclerosis and 25 healthy control subjects were determined by colorimetric assay, gas chromatography and enzymatic assays, respectively. Membrane fluidity was calculated according to previously established formulae and correlated with C-reactive protein and the Kurtzke expanded disability status scale. There were no significant differences in membrane lipids in peripheral blood mononuclear cells from patients and controls. However, correlation studies showed lipid metabolic abnormalities, which were reflected in significant correlations between membrane fluidity as measured by both its fatty acid and phospholipid compositions, and the functional system scores. C-reactive protein showed positive correlations with phosphatidylcholine, phosphatidylserine, phosphatidylinositol and total phospholipids in membranes from control subjects. Metabolic abnormalities, as well as correlations between membrane fluidity and the functional system scores, suggested the involvement of these immune cell membranes in the disease progression. Furthermore, the changed relationship between membrane phospholipids and C-reactive protein, which has been shown to correlate with infectious episodes and clinical relapse, could be an indication of immune cell dysfunction in patients with multiple sclerosis.     

Decreased membrane fluidity in erythrocytes from patients with Crohn’s disease

In erythrocytes from patients with Crohn’s disease, lower levels of membrane fluidity were demonstrated by means of electron spin resonance with 16-stearic acid spin label (SAL) but not 5- or 12-SAL, suggesting more apparent abnormality in the deeper portion of lipid bilayer. Membrane lipid analysis showed a significant increase of sphingomyelin, a decrease of phosphatidylcholine and decreased contents of polyunsaturated acyl-chains, particularly C18:2, of phospholipid, without alteration in the cholesterol to phospholipid molar ratio. The alteration in membrane fluidity appeared to be closely related to changes in membrane phospholipid classes and their acyl-chains. Among the plasma lipids, contents of esterified cholesterol, phospholipid, very low- and low-density lipoproteins and cholesterol of high-density lipoprotein significantly decreased. Decreased contents of polyunsaturated acyl-chains, particularly C18:2, in the plasma phospholipid were also noted. Changes in plasma and erythrocyte membrane lipids remained even in the inactive stage of the disease, except for the normalized sphingomyelin and phosphatidylcholine content in the erythrocyte membrane. Since membrane fluidity is closely involved in various cell membrane functions, the decreased membrane fluidity and altered lipid composition in erythrocyte membrane evident in patients with Crohn’s disease may have important effects on the pathological status in this disease.     

Impaired fetal erythrocytes' filterability: relationship with cell size, membrane fluidity, and membrane lipid composition.

The lipid composition of erythrocytes (red blood cells [RBCs]) plays a significant role in determining certain membrane biophysical properties. We have found that fetal RBCs showed a dramatically low filterability compared with adult RBCs and questioned whether this could be a consequence of their membrane lipid composition. We therefore studied fetal RBCs at two different gestational ages, neonatal RBCs and adult RBCs. Biophysical parameters were studied using two different techniques, filterability and membrane fluidity. The latter was measured by fluorescence polarization using three different probes. The membrane lipid composition was examined by measuring cholesterol and phospholipids. After extraction of the phospholipids, followed by high performance thin-layer chromatography, the fatty acids in the phospholipid subfractions were analyzed by gas-liquid chromatography. The fetal RBCs' filterability was found to be correlated with both the larger size and the higher hemoglobin content of the cells, but there was no correlation between RBC filterability and fluidity or membrane lipid composition. In adult RBCs, compared with neonatal RBCs, the slight increase of unsaturated fatty acids in phosphatidylcholine and phosphatidylethanolamine should have increased the membrane fluidity. However, in RBCs, no change was observed in the fluidity parameters measured by fluorescence polarization.     

Lipid composition, fluidity and enzymatic activities of rat liver plasma and mitochondrial membranes in dietary obese rats

Total lipids and fatty acid composition were determined in liver plasma and mitochondrial membranes from control and dietary obese rats after 4 weeks of the experimental period. The lipid composition of liver plasma and mitochondrial membranes showed an increase of triacylglycerols in obese rats. The liver plasma membranes showed a decrease of saturated/unsaturated fatty acid ratio and an increase of (n-6) polyunsaturated fatty acids, whereas the (n-3) polyunsaturated acids were decreased. Contrary to what occurs with plasma membranes, few modifications were observed in mitochondrial membranes. Changes of the fatty acid composition of the phospholipid bilayer are of potentially great importance in structural and functional parameters of membrane. Fluidity of liver plasma membranes of dietary obese rats was highly increased, while the mitochondrial ones remained unchanged. These results can be well explained by the decreased saturated/unsaturated fatty acid ratio. A significant decrease of (Na+-K+) ATPase activity (a membrane bound enzyme) was found in plasma membranes of dietary obese rats. Mitochondrial enzymatic activities and oxidative phosphorylation showed few changes except a small, but significant decrease of state 3 respiratory rate. In this study we also determined the fatty acid composition of all the foods offered to animals and their daily intakes in order to discuss their possible influence on changes in structural and functional membrane parameters.     

Effect of gemfibrozil on erythrocyte membrane lipids in geriatric patients

Twenty geriatric patients with primary or secondary hyperlipidemia and suffering from various other diseases received for three weeks once daily 900 mg gemfibrozil. The hyperlipidemia had not been treated before, and a cholesterol-reduced diet did not succeed in lowering total cholesterol below 6.75 mmol/l (260 mg/100 ml) and serum triglycerides below 1.97 mmol/l (175 mg/100 ml). The purpose of this study was to analyze the lipid composition of the erythrocyte membrane, serum lipids and rheological parameters before and after the therapy. Mean serum total cholesterol and triglyceride content decreased significantly by 16.3% (p less than 0.05) and 35.2% (p less than 0.01) on average, respectively. Aggregation of thrombocytes and of erythrocytes, thrombin time and partial thromboplastin time slightly varied during the three weeks' treatment, but without statistical significance. The content of total long-chain saturated fatty acids in the phospholipid fraction of the erythrocyte membrane decreased slightly from 41.3% to 40.9% (p less than 0.05), whereas the total w6-unsaturated fatty acids without the precursor linoleic acid increased by about the same extent from 15.66% to 16.0% (p less than 0.05). The molar ratio of phospholipid to cholesterol content decreased significantly (p less than 0.01) due to a reduced phospholipid content at the end of the therapy. In conclusion, in addition to reducing the serum lipids, gemfibrozil slightly effects the lipid composition of erythrocytes, but the effects of the varied concentrations of long-chain saturated and long-chain w6-unsaturated fatty acids in the phospholipid fraction on membrane fluidity might be compensated, at least partly, by the decrease of the ratio of membrane phospholipid to cholesterol.     

Lipid composition of cell membranes and its relevance in type 2 diabetes mellitus

Identifying the causative relationship between the fatty acid composition of cell membranes and type 2 diabetes mellitus fundamentally contributes to the understanding of the basic pathophysiological mechanisms of the disease. Important outcomes of the reviewed studies appear to support the hypotheses that the flexibility of a membrane determined by the ratio of (poly)unsaturated to saturated fatty acyl chains of its phospholipids influences the effectiveness of glucose transport by insulin-independent glucose transporters (GLUTs) and the insulin-dependent GLUT4, and from the prediabetic stage on a shift from unsaturated towards saturated fatty acyl chains of membrane phospholipids directly induces a decrease in glucose effectiveness and insulin sensitivity. In addition, it has become evident that a concomitant increase in stiffness of both plasma and erythrocyte membranes may decrease the microcirculatory flow, leading ultimately to tissue hypoxia, insufficient tissue nutrition, and diabetes-specific microvascular pathology. As to the etiology of type 2 diabetes mellitus, a revised hypothesis that attempts to accommodate the reviewed findings is presented.     

Effects of type 2 diabetes mellitus on plasma fatty acid composition and cholesterol content of erythrocyte and leukocyte membranes

Abstract Insulin resistance is a major factor in the pathogenesis of type 2 diabetes mellitus (T2DM) and is related to the fatty acid profile of the plasma membranes. The purpose of the present study was to investigate fatty acid composition and cholesterol content of cell membranes in patients with type 2 diabetes and, thus, to evaluate the possible factors leading to the alteration of plasma membrane fluidity. The study was performed in 20 healthy control subjects and 32 patients with type 2 diabetes. The fatty acid profiles and cholesterol content of the erythrocyte (RBC) and leukocyte (WBC) membranes were determined by a gas chromatographic method. When one considers the membrane constituents increasing fluidity and the ones decreasing it, the diabetics had a membrane composition decreasing fluidity compared to controls. On the other hand, when compared to control subjects, type 2 diabetic patients showed a significantly higher proportion of C16:0 components in erythrocyte and leukocyte membranes and plasma samples (25.4±3.1% vs. 31.1±4%; 23.3±2.4% vs. 29.3±5.2%; 27.6±3.9% vs. 34.5±5.7%; p<0.005, p<0.01 and p<0.005, respectively). Our results suggest that the ratio of saturated:unsaturated fatty acids changes in plasma and cell membranes of patients with type 2 diabetes. This situation may cause, at least in part, RBC–WBC function abnormalities and insulin resistance because of inconvenient membrane fluidity.     

Modulation of (Na,K)-ATPase activity by membrane fatty acid composition: therapeutic implications in human hypertension

Oxidative stress (OS) plays a key role in the pathophysiology of essential hypertension and is associated with changes in the cell membrane fatty acid composition and fluidity. As (Na,K)-ATPase is modulated by the surrounding lipid microenvironment, lipid peroxidation could alter the interactions of this enzyme with the membrane components. Thus, modifications in the membrane fatty acid profile will translate into effects on (Na,K)-ATPase activity. Accordingly, a decrease in this enzyme activity has been reported in hypertensive patients. The aim of this study was to evaluate the relationship between membrane fluidity and fatty acid composition and (Na,K)-ATPase activity in erythrocytes of essential hypertensive patients supplemented with antioxidant vitamins C and E. A double-blind, randomized, placebo-controlled study was conducted in 120 men with essential hypertension assigned to receive vitamin C (1?g/day)?+?E (400?IU/day) or placebo for 8 weeks. Measurements included OS related parameters: GSH/GSSG ratio, F2-isoprostanes and antioxidant capacity of plasma, (Na,K)-ATPase activity and erythrocytes membrane fatty acid composition (PUFA, polyunsaturated fatty acids; SAFA, saturated fatty acids). Associations were assessed by Pearson correlation and the differences by Student t-test (p?相似文献   

Electron spin resonance studies on interaction of complement proteins with erythrocyte membranes.

Sheep erythrocytes have been spin labeled with 5-, 12-, and 16-nitroxystearic acid in order to investigate complement-induced changes in the physical state of the lipid bilayer. Formation of osmotic lesions in the membrane causes an increase in the fluidity of the membrane which overcomes the decrease in membrane fluidity caused by the interaction of the complement proteins. A decrease in membrane fluidity is observed only when complement-lysed membranes are resealed or when complement proteins react with isosmolar ghosts that do not undergo osmotic lysis. The decrease in bulk fluidity of the membrane is first observed when C8 binds to the membranes bearing C5b67 and is enhanced upon the subsequent binding of C9. The decrease in membrane fluidity shown by the electron spin resonance spectra of spin-labeled fatty acids suggests that certain of the complement proteins penetrate the membrane and interact with hydrophobic regions of the lipid bilayer.     

Effect of cholestasis induced by organic anion on the lipid composition of hepatic membrane subfractions and bile in rats

Several organic anions inhibit the secretion of cholesterol and phospholipid into bile without affecting total bile acid secretion (uncoupling). The uncoupling induced by sulphobromophthalein (BSP) alters the fatty acid composition of biliary lecithin. The purpose of this study was to investigate the relationship between the lipid composition of bile and of liver subcellular membrane fractions during BSP-induced uncoupling. After depletion of the bile salt pool, rats fitted with a bile duct cannulus were infused with sodium taurocholate given either alone or with BSP. Bile was collected and liver microsomes and canalicular membranes were isolated for analysis of lipid composition. In bile, uncoupling increased the cholesterol/phospholipid ratio (C/P ratio) and the saturated/unsaturated fatty acid ratio (S/U ratio) in phosphatidylcholine. The C/P ratio was increased in the canalicular membrane, but the membrane phosphatidylcholine S/U ratio decreased during uncoupling. In microsomes, the S/U ratio of membrane phosphatidylcholine was slightly increased, but the C/P ratio was unaffected during uncoupling. These results support the hypothesis that an increased secretion of hydrophobic phosphatidylcholine species from the canalicular membrane into bile reduces the proportion of hydrophobic phosphatidylcholine species in the canalicular membrane during uncoupling. The decreased contribution of hydrophobic phosphatidylcholine species may ameliorate the decrease in membrane fluidity resulting from the accumulation of cholesterol in the canalicular membrane and stimulate the synthesis of hydrophobic phosphatidylcholine species in the microsomes.     

Inhibition of nitric oxide synthesis increases erythrocyte membrane fluidity and unsaturated fatty acid content

Changes in the lipid composition of the membrane affect its fluidity and function. These variables are altered in various forms of hypertension. Our hypothesis was that the rapid increase in blood pressure (BP) caused by inhibition of nitric oxide production would lead to alterations in membrane fluidity similar to those observed in genetic hypertension. We used Nomega-nitro L-arginine methyl ester (L-NAME) and vehicle-treated (3 weeks) Wistar-Kyoto rats to study the effects of nitric oxide synthase (NOS) inhibition on membrane fluidity and lipid composition. Erythrocyte membrane fluidity was measured by fluorescence anisotropy. Membrane lipids were separated using Sep-Pak and thin-layer chromatography. Fatty acid methyl esters were produced and analyzed by gas chromatography-mass spectrometry. Nomega-nitro L-arginine methyl ester treatment increased BP and erythrocyte membrane fluidity. The phospholipid and unsaturated fatty acid levels in the membranes from the L-NAME-treated rats were consistent with the increase in fluidity (ie, more unsaturated fatty acid, in particular, arachidonic and docosahexaenoic acid) and a reduction in membrane sphingomyelin content. Fatty acid analysis of individual lipid groups suggested the changes in membrane fatty acid composition may be asymmetric, with the majority of the changes occurring in the outer leaflet. Inhibition of NOS results in changes in membrane composition that may explain the concurrent changes in fluidity. The increased membrane fluidity observed here contrasts with the reduced fluidity observed in genetic hypertension or unchanged fluidity in secondary hypertension. The effects could be related to NOS inhibition or may be a direct effect of L-NAME.     

Biliary excretory function is regulated by canalicular membrane fluidity associated with phospholipid fatty acyl chains in the bilayer: implications for the pathophysiology of cholestasis

BACKGROUND AND AIMS: Bile canalicular membrane fluidity is modulated by phospholipid molecular species within membrane lipid bilayers. Thus, organellar membrane lipid composition is a determinant of canalicular function. In this study, the effect of phalloidin-induced cholestasis on bile lipid composition and liver subcellular membrane fraction composition in rats was examined to clarify the relationship between cholestasis and hepatic lipid metabolism. METHODS AND RESULTS: Each rat received one phalloidin dose (400 microg/kg, i.v.). After the bile was collected, liver microsomes and canalicular membranes were analysed. The bile flow rate decreased by 50% 3.5 h after phalloidin administration. Although the bile acid output remained almost the same, the phospholipid and cholesterol output were significantly decreased (by 40.3+/-5.97% and 76.9+/-5.56%, respectively). Thus, the cholesterol:phospholipid (C:P) ratio in bile was significantly decreased by 80.4+/-10.1%. Phalloidin administration also increased the saturated: unsaturated fatty acid ratio (S:U) in bile for phosphatidylcholine by 25.5+/-3.2%. In the canalicular membrane, the C:P and S:U ratios for phosphatidylcholine were increased (24.8+/-4.2% and 34.4+/-6.9%, respectively), while the S:U for sphingomyelin was decreased by 61.0+/-6.2%. In microsomes, the C:P was decreased by 41.0+/-6.0%, but the S:U for both phosphatidylcholine and sphingomyelin were unaffected. Canalicular membrane fluidity, assayed by 1,6-diphenyl-1,3,5-hexatriene fluorescence depolarization, decreased significantly. Therefore, increased secretion of hydrophobic phosphatidylcholine into bile was associated with more hydrophobic canalicular membrane phosphatidylcholine, while sphingomyelin in the canalicular membrane was less hydrophobic. CONCLUSIONS: These results indicate that phalloidin uncouples secretion of cholesterol and phospholipids, which causes a redistribution of fatty acyl chain species among canalicular membrane phospholipids that alters membrane fluidity. These changes may be a homeostatic response mediated by the phospholipid translocator in the canalicular membrane, although direct evidence for this is unavailable.     

Consumption of virgin olive oil influences membrane lipid composition and regulates intracellular signaling in elderly adults with type 2 diabetes mellitus

We aimed to define changes in membrane fatty acids and signaling proteins induced by virgin olive oil (VOO) consumption in elderly persons with type 2 diabetes (n = 16) compared to a control group (n = 28). The fatty acid composition was determined by gas chromatography and G-protein subunits and protein kinase C alpha (PKCalpha) by immunoblotting. VOO consumption increased the monounsaturated fatty acid content in phospholipids and cholesterol esters in both groups. In contrast, saturated fatty acids were decreased only in phospholipids. The levels of Galphao, Gbeta, and PKCalpha were significantly lower in diabetics than in controls. However, whereas VOO consumption reduced Galphas, Gbeta, and PKCalpha in both groups, reduction in Galphai was observed only in diabetics. These results indicate that long-term VOO consumption modifies the fatty acid composition of plasma membrane, which influences the association of G proteins and PKCalpha with the lipid bilayer. These combined effects probably account for the positive effects of VOO on glycemic homeostasis.