Effects of phospholipids on sphingomyelin hydrolysis induced by intestinal alkaline sphingomyelinase: an in vitro study

Digestion of dietary sphingomyelin (SM) is catalyzed by intestinal alkaline sphingomyelinase (SMase) and may have important implications in colonic tumorigenesis. Previous studies demonstrated that the digestion and absorption of dietary SM was slow and incomplete and that the colon was exposed to SM and its hydrolytic products including ceramide. In the present work, we studied the influences of glycerophospholipids and hydrolytic products of phosphatidylcholine (PC; i.e., lyso-PC, fatty acid, diacylglycerol, and phosphorylcholine) on SM hydrolysis induced by purified rat intestinal alkaline SMase in the presence of 10 mM taurocholate. It was found that various phospholipids including PC, phosphatidylserine (PS), phosphatidylinositol (PI), phosphatidylethanolamine (PE), and phosphatidic acid (PA) inhibit alkaline SMase activity in a dose-dependent manner, with the degree of inhibition being in the order PA > PS > PI > PC > PE. Similar inhibition was also seen in a buffer of pH 7.4, which is close to the physiologic pH in the middle of the small intestine. When the effects of hydrolytic products of PC were studied, lyso-PC, oleic acid, and 1,2-dioleoyl glycerol also inhibited alkaline SMase activity, whereas phosphorylcholine enhanced SMase activity. However, in the absence of bile salt, acid phospholipids including PA, PS, and PI mildly stimulated alkaline SMase activity whereas PC and PE had no effect. It is concluded that in the presence of bile salts, glycerophospholipids and their hydrolytic products inhibit intestinal alkaline SMase activity. This may contribute to the slow rate of SM digestion in the upper small intestine.     

31P MRS analysis of the phospholipid composition of the peripheral blood mononuclear cells (PBMC) and bone marrow mononuclear cells (BMMC) of patients with acute leukemia (AL)

The aim of this study was to evaluate the phospholipid concentration in acute leukemia (AL) blast cells from peripheral blood (PBMC) and bone marrow (BMMC). In vitro ³¹P Nuclear Magnetic Resonance Spectroscopy (³¹P MRS) was used. The integral intensities of the resonant peaks and the phospholipid concentrations in PBMC and BMMC were analyzed. Differences in the phospholipid concentrations in cells from myeloblastic or lymphoblastic lines were also evaluated. This investigation was carried out on phospholipid extracts from PBMC and BMMC from 15 healthy volunteers and 77 patients with AL (samples taken at the moment of diagnosis). A significant decrease in sphingomyelin (SM) and phosphtidylserine (PS) was observed in the PBMC of patients with AL relative to the results for the healthy volunteers. For ALL, we found a significant decrease in the concentration of phosphatidylcholine plasmalogen (CPLAS), SM, PI+PE (phosphatidylinositol + phosphatidylethanolamine) and PS in comparison with the results for healthy volunteers and patients with AML. Experiments with BMMC cells revealed a significant decrease in the concentration of CPLAS, SM, PI+PE, and PS in ALL relative to AML. Additionally, a significant decrease in phosphatidylcholine (PC) concentration was observed in ALL compared to AML. If the phospholipid extracts were taken simultaneously from the same patient, there were no significant differences in the integral intensities and phospholipid concentrations between PBMC and BMMC.     

Cerebral Phospholipid Content and Na+, K+-ATPase Activity During Ischemia and Postischemic Reperfusion in the Mongolian Gerbil

Using bilateral carotid artery occlusion in adult gerbils we examined the effects of ischemia and ischemia/reperfusion on cerebral phospholipid content and Na+,K+-ATPase (EC 3.6.1.3) activity. In contrast to the large changes in phospholipid content and membrane-bound enzyme activity that have been observed in liver and heart tissues, we observed relatively small changes in the cerebral content of total phospholipid, phosphatidylcholine (PC), phosphatidylserine (PS), and phosphatidylethanolamine (PE) following ischemic intervals of up to 240 min. Following 15 min of ischemia the cerebral content of sphingomyelin (SM) was decreased to less than 50% of control values but returned to near-normal levels with longer ischemic periods. Significant decreases in the cerebral content of phosphatidylinositol (PI) and phosphatidic acid (PA) were observed following shorter intervals of ischemia (15-45 min). Na+,K+-ATPase activity of cerebral homogenates prepared from the brains of gerbils subjected to 30-240 min of ischemia was decreased but significantly different from control activity only after 30 min of ischemia (-29%, p less than or equal to 0.05). With the exception of PS, reperfusion for 60 min following 60 min of ischemia resulted in marked increases in cerebral phospholipid content with PC, SM, PI, and PA levels exceeding and PE levels equal to preischemic values. Longer periods of reperfusion (180 min) resulted in decreases in cerebral phospholipid content toward (PC, SM, PI, and PA) or below (PE) preischemic levels. In contrast, the cerebral content of PS significantly decreased during reperfusion (-51% at 60 min, p less than or equal to 0.05) and remained below preischemic values even after 180 min of reperfusion.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of different molecular species of sphingomyelin on phospholipase C δ1 activity

Bovine brain sphingomyelin was separated into different molecular species using a reverse phase column. PLC δ1 was inhibited by all molecular species of sphingomyelin. The extent of this inhibition was dependent on the hydrophobicity. Based on fatty acid analysis, we conclude that the inhibition of PLC δ1 depends on the chain length and degree of unsaturation of the fatty acid moiety of SM. N-palmitoyl-D-sphingomyelin and N-stearoyl-D-sphingomyelin inhibited PLC δ1 less then N-oleoyl-D-sphingomyelin. In the absence of Ca²⁺ (1 mM EGTA) all tested molecular species of SM inhibited weakly the enzyme. The sensitivity of PLC δ1 to inhibition by SM increased with increasing Ca²⁺ concentration. The shape of calcium curve differed for molecular species with saturated and unsaturated fatty acids. Inhibition of PLC δ1 by N-palmitoyl-D-sphingomyclin and N-stearoyl-D-sphingomyclin reached a maximum at 0.2 μM Ca²⁺, while inhibition by N-oleoyl-D-sphingomyclin reached maximum at 2 μM Ca²⁺. PLC δ1 is more sensitive to inhibition by SM when it is maximally activated by spermine and calcium and the extent of this inhibition depends on the length and degree of fatty acid unsaturation of the molecular species.     

Phospholipid Composition in Spinal Cord Regions after Ischemia/Reperfusion

Ischemia-reperfusion induced changes in concentration of phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), phosphatidylinositol (PI) and sphingomyelin (SM) in the gray matter taken in toto, white matter, dorsal horns, intermediate zone and ventral horns of the rabbit's spinal cord were studied and compared with neurohistopathological changes. With the exception of PI concentration in the dorsal horns, ischemia of 25 min caused significant degradation of all phospholipids. While short-lasting recirculation (1 h) did not returned the levels of phospholipids to control values, postischemic recirculation for 3 h sharply increased the resynthesis of all phospholipids, but only the concentration of PE, PS, and PI in the dorsal horns and PC in the intermediate zone significantly improved and returned close to control values. Corresponding neurohistopathological changes resulting after the same reperfusion periods are given.     

31P MRS analysis of the phospholipid composition of normal human peripheral blood mononuclear cells (PBMC)

The aim of this investigation was to characterize the phospholipid composition of normal human blood mononuclear cells using 31P NMR spectroscopy. Mononuclear cells of peripheral blood were obtained from 10 volunteers. Phospholipid extracts were prepared from 60x10(6) cells according to modified Folch's method. An AMX 300 Bruker spectrometer 7.05 T was used. The 31P spectrum of phospholipid extracts from normal human PBMC consisted of 9 peaks, with one each for phosphatidylcholine (PC), plasmalogen of phosphatidylcholine (CPLAS), lysophosphatidylcholine (LPC), sphingomyelin (SM), phosphatidylethanolamine (PE), phosphatidylinositol (PI), phosphatidylserine (PS) and cardiolipin (CL), and another one due to the external reference substance, methylenediphosphonic acid (MDPA). The concentrations of these phospholipids (PL), based on the integral intensities, were as follows: 0.398 +/- 0.078 mmole/l for PC; 0.033 +/- 0.019 mmole/l for CPLAS; 0.155 +/- 0.043 mmole/l for SM; 0.266 +/- 0.104 mmole/l for PI+PE; 0.101 +/- 0.040 mmole/l for PS, and 0.026 +/- 0.033 mmole/l for CL. The results of this study confirmed that 31P MRS is a convenient tool for measuring the phospholipid concentrations of biological samples.     

Investigation of interaction of human platelet membrane components with anticoagulant drugs Abciximab and Eptifibatide

Abciximab (Abci) and eptifibatide (Epti) are antiaggregate drugs which may reduce thrombotic complications in acute coronary syndromes. The aim of this work was the investigation of the interaction between the phospholipid-GPIIb/IIIa glycoprotein complex and Abci or Epti, and the influence of these drugs on the phospholipid ratio in the platelet membrane. The interaction between the phospholipid-GPIIb/IIIa glycoprotein complex and antiaggregate drugs were investigated using the Surface Plasmon Resonance Imaging technique (SPRI). Phospholipids phosphatidylinositol (PI), phosphatidylserine (PS), phosphatidylethanolamine (PE), phosphatidylcholine (PC) and sphingomyelin (SM) were first immobilized onto the gold chip surface. The phospholipid ratio in the platelet membrane was determined by the HPLC. Only PI, PS, PE and PC were determined. Human platelets treated 'in vitro' with Abci or Epti exhibit changes in the phospholipid ratio in the platelet membrane. The ratio of PS decreases and PC rises. The SPRI distinctly shows interactions between phospholipids and glycoprotein GPIIb/IIIa, and between the phospholipid-glycoprotein GPIIb/IIIa complex and Abci or Epti. The interaction between phospholipids and glycoprotein GPIIb/IIIa is growing in the sequence: PI相似文献   

Lipopolysaccharide-induced hydrolysis of plasmalogenic phosphatidylethanolamine in human platelets

The composition of human platelet major phospholipids-phosphatidylcholine (PC), phosphatidylinositol (PI), phosphatidylserine (PS), phosphatidic acid (PA), sphingomyelin (SM), plasmalogenic and diacyl species of phosphatidylethanolamine (PPE and APE, respectively) was quantitatively analyzed by high performance liquid chromatography. Incubation (10 min, 37 degrees C) of washed platelets with lipopolysaccharide B (LPS) of Salmonella typhimurium was found to produce (in the absence of aggregation) marked hydrolysis of PI (ca. 15%) and PPE (ca. 19%) containing the bulk of polyenic fatty acids. PC and APE were less degraded (8-9%), while the amounts of PS and SM were practically unchanged and the level of PA rose by 20%. Addition of thrombin to LPS-pretreated platelets resulted in their more rapid aggregation which was accompanied by a decreased and nearly equal hydrolysis of APE and PPE (7-8%) as compared with control platelets (10 and 12%, respectively). The extent to which PI was degraded (ca. 34%), by the action of thrombin was not affected by preliminary incubation with LPS. It is suggested that thrombin (as well as LPS) activating endogenous phospholipase(s) A2 can liberate from PPE not only arachidonic acid but also other essential polyenic fatty acids present in PPE in relatively high amounts. Besides, the agents studied may activate the intrinsic platelet system of rapid arachidonoyl transfer from diacyl PC and PE to PPE.     

Comparison of linoleate,palmitate and acetate metabolism in rat ventral prostate

Rat ventral prostate incorporated (1-¹⁴C)acetate, (1-¹⁴C)palmitate and (1-¹⁴C)linoleate into different phospholipids in a time-dependent process. The rate of incorporation into total phospholipids was higher with linoleate (10.0 nmol/g) than with either palmitate (5.8 nmol/g) or acetate (4.7 nmol/g). Predominant labelling with all the radioactive substrates assayed was found in choline glycerophospholipids (PC). The radioactive profiles for linoleate in the other ventral prostate phospholipids differed from those obtained with palmitate and acetate. Specifically linoleate was incorporated into inositol glycerophospholipids plus lysoethanolamine glycerophospholipids (PI+LPE) and not into sphingomyelin (SM), while palmitate and acetate incorporated into SM but not into PI+LPE. Acetate showed the highest oxidation to CO₂ whereas no differences were observed in the radioactivity incorporated into CO₂ from a saturated (palmitate) or an essential unsaturated fatty acid (linoleate). These studies also show zinc-dependence by the acetate to CO₂ oxidation.Abbreviations PL total phospholipids - PC choline glycerophospholipids - PE ethanolamine glycerophospholipids - PI+LPE inositol glycerophospholipids plus lysoethanolamine glycerophospholipids - PS serine glycerophospholipids - SM sphingomyelin     

Changes of phospholipid composition and superoxide dismutase activity during global brain ischemia and reperfusion in rats

Alterations in phospholipid content and Cu/Zn superoxide dismutase (SOD) activity were examined in rat brain after 15 min of global ischemia (four-vessel occlusion) followed by 2-, 24- or 48-h reperfusion. Phosphatidylcholine (PC) and phosphatidylethanolamine (PE), the main brain phospholipids, were markedly decreased in ischemic rats and remained decreased during the whole reperfusion period. Concentrations of phosphatidylinositol (PI) and sphingomyelin (SM) were also significantly reduced during ischemia but recovered during reperfusion period. In contrast, phosphatidylserine (PS) and lysophospholipids (LysoPL) were unchanged during ischemia but were elevated after 24 h of reperfusion. Significant reductions in blood plasma phospholipids were also demonstrated. 24-48 h of reperfusion markedly decreased PE, PC and PS contents, while the concentrations were almost unchanged by ischemia alone. Brain SOD activity decreased significantly during ischemia and was recovered to control value already after 2 h of reperfusion. These results suggest that ischemia/reperfusion is accompanied by a significant and selective degradation of brain phospholipids that may be attributable to oxidative stress and activation of phospholipases.     

New reagents for phosphatidylserine recognition and detection of apoptosis

The phospholipid bilayer surrounding animal cells is made up of four principle phospholipid components, phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), and sphingomyelin (SM). These four phospholipids are distributed between the two monolayers of the membrane in an asymmetrical fashion, with PC and SM largely populating the extracellular leaflet and PE and PS restricted primarily to the inner leaflet. Breakdown in this transmembrane phospholipid asymmetry is a hallmark of the early to middle stages of apoptosis. The consequent appearance of PS on the extracellular membrane leaflet is commonly monitored using dye-labeled Annexin V, a 36 kDa, Ca2+-dependent PS binding protein. Substitutes for Annexin V are described, including small molecules, nanoparticles, cationic liposomes, and other proteins that can recognize PS in a membrane surface. Particular attention is given to the use of these reagents for detecting apoptosis.     

beta 2-Glycoprotein-I (apolipoprotein H) interactions with phospholipid vesicles

The binding characteristics of the human serum protein beta 2-glycoprotein-I, also called apolipoprotein H, with multilamellar phospholipid vesicles has been studied. It was found that beta 2-G-I is not or almost not bound to the "neutral" phospholipids phosphatidylcholine (PC), phosphatidylethanolamine (PE) and sphingomyelin (SM). The negatively charged compounds phosphatidylserine (PS) and phosphatidylinositol (PI) interact strongly with beta 2-G-I. In terms of phospholipid concentration the binding to PS is about one order of magnitude greater than to PI. The binding capacity is influenced by several parameters such as the molarity of buffer, presence of mono- or divalent cations as well as ethylenediaminotetraacetic acid (EDTA). Proteins like bovine serum albumin (BSA), human serum albumin (HSA) or horse gamma-globulin (HGG) influence the binding also in a concentration dependent manner.     

Adaptive changes in lipid composition of rat liver plasma membrane during postnatal development following maternal ethanol ingestion

The fatty acid composition of constituent phospholipids and the cholesterol content of rat liver plasma membranes were determined subsequent to maternal alcohol ingestion during pregnancy and lactation. The alcoholic group was given a liquid Metrecal diet containing 37% ethanol-derived calories. The control group was pair-fed an isocaloric sucrose/Metrecal diet. Litters were killed for lipid analyses at days 5, 15 and 25 after birth. These studies revealed that the total phospholipid phosphorus was similar and increased significantly with age in both groups. Cholesterol also increased significantly with age in both groups but was greater in the alcoholic pups, resulting in a higher cholesterol/phospholipid molar ratio. While the phosphatidylethanolamine (PE) content increased with age in both groups, that of sphingomyelin decreased. Phosphatidylserine + phosphatidylinositol (PS + PI) was significantly higher in the control group at all ages studied. A consistent increase of C22:6 in phosphatidylcholine (PC), sphingomyelin, PS + PI and in the total phospholipid fraction from alcoholic pups was observed. Although other fatty acid changes were found in PC, PS + PI and sphingomyelin, PE was not affected. These results suggest that specific adaptive changes were induced in the liver plasma membrane lipids of the progeny from alcoholic rats.     

Model of a Raft in Both Leaves of an Asymmetric Lipid Bilayer

We present a theory of inhomogeneities in the plasma membrane, or rafts, that can exist in both leaves of the plasma membrane. We note that although neither of the major phospholipid components of the outer leaf, sphingomyelin (SM) nor phosphatidylcholine (PC), evinces a tendency to form phases characterized by nonzero curvature, one of the major components of the inner leaf, phosphatidylethanolamine (PE), displays a strong tendency to do so whereas the other, phosphatidylserine (PS), does not. Therefore, we posit that the concentration difference of PS and PE couples to height fluctuations of the plasma membrane bilayer. This brings about a microemulsion in the inner leaf. Coupling of the concentration difference between PS and PE in the inner leaf and SM and PC in the outer leaf propagates the microemulsion to that leaf as well. The characteristic size of the inhomogeneities is equal to the square-root of the ratio of the bending modulus of the bilayer to its surface tension, a size which is ∼100 nm for the plasma membrane. If the coupling between leaves were to be provided by the interchange of cholesterol, then our model raft would consist of SM and cholesterol in the outer leaf and PS and cholesterol in the inner leaf floating in a sea of PC and PE in both leaves.     

Model of a Raft in Both Leaves of an Asymmetric Lipid Bilayer

We present a theory of inhomogeneities in the plasma membrane, or rafts, that can exist in both leaves of the plasma membrane. We note that although neither of the major phospholipid components of the outer leaf, sphingomyelin (SM) nor phosphatidylcholine (PC), evinces a tendency to form phases characterized by nonzero curvature, one of the major components of the inner leaf, phosphatidylethanolamine (PE), displays a strong tendency to do so whereas the other, phosphatidylserine (PS), does not. Therefore, we posit that the concentration difference of PS and PE couples to height fluctuations of the plasma membrane bilayer. This brings about a microemulsion in the inner leaf. Coupling of the concentration difference between PS and PE in the inner leaf and SM and PC in the outer leaf propagates the microemulsion to that leaf as well. The characteristic size of the inhomogeneities is equal to the square-root of the ratio of the bending modulus of the bilayer to its surface tension, a size which is ∼100 nm for the plasma membrane. If the coupling between leaves were to be provided by the interchange of cholesterol, then our model raft would consist of SM and cholesterol in the outer leaf and PS and cholesterol in the inner leaf floating in a sea of PC and PE in both leaves.     

Changes in phospholipids and free fatty acids in the brains of mice preconditioned by hypoxia.

Changes in the composition and contents of phospholipids and free fatty acids were observed and compared in three groups: (A) unpreconditoned normal controls, (B) exposure to 1 run of hypoxia and (C) exposure to 4 runs of hypoxia. In group B, the content of phosphatidyl ethanolamine (PE), phosphatidyl serine (PS) and free fatty acids (FFAs) increased significantly and the content of phosphatidyl choline (PC) and sphingomyelin (SM) decreased significantly. While in group C the content of PE, PS, PC and FFAs changed significantly when compared with that of group B, all phospholipid (except SM) and FFA contents tended to decrease to the level of group A. No new FFA was seen in the brain homogenates in any of the three groups. These results suggest that the changes in the content of mouse brain phospholipids and FFAs may be adaptive and involved in the animals' tolerance to hypoxia.     

Effect of severe incomplete ischaemia and postischaemic reperfusion on phospholipids and unesterified cholesterol in rabbit spinal cord

Effect of severe incomplete ischemia, induced by occlusion of the abdominal aorta caudal to the left renal artery for 40 min, and postischemic reperfusion for 6 h, 1 and 4 days on phospholipid composition and unesterified cholesterol concentration was studied in the lumbosacral (L3-S1) spinal cord separated into the gracile fascicle (Fg), dorsal part without Fg (Dp) and ventral part (Vp). Ischemia decreased the inositol phospholipid (PI) concentration in Dp and Vp and this was recovered during reperfusion. Within 6 h following ischemia, ethanolamine (PE) and serine (PS) phospholipid concentrations decreased in Dp and PS also in Vp. During the long reperfusion intervals the concentrations of the two major phospholipids, PE and choline phospholipid (PC) declined in Fg, Dp and Vp. No changes were observed in sphingomyelin (SM). The concentration of unesterified cholesterol (UC) was lower throughout the reperfusion period in Dp and Vp, while the decrease in Fg was delayed. The molar ratio UC/TPL was reduced starting from 24 h of reperfusion. The pattern of changes, which were delayed in the white matter as compared to Dp and Vp (containing the gray matter) indicated severe damage to the membrane structures in the tissue, developed during reoxygenation, that was related to decreased tissue viability.     

Application of 31P MRS to the analysis of phospholipid changes in plasma of patients with acute leukemia

The aim of the experiment was to evaluate the changes of phospholipid concentrations in patients (n=30) with acute leukemia compared with reference group of healthy volunteers (n=21). The analysis focused on the following phospholipids (PL) collected from plasma: phosphatidylcholine (PC), plasmalogen of phosphatidylcholine (CPLAS), lysophosphatidylcholine (LPC), sphingomyelin (SM), phosphatidylethanolamine (PE), and phosphatidylinositol (PI). Phospholipid extracts were obtained by Folch's method from 4 ml of plasma. 31P MR spectra were obtained on an AMX Bruker 300 MHz (7.05 T) spectrometer. Calculation of concentration based on integral intensity of the phospholipid relative to an internal concentration standard of MDPA. For healthy volunteers, the following values of phospholipid concentrations were obtained: (5.18+/-1.615) mmol/l for PC+CPLAS; (0.364+/-0.178) mmol/l for LPC; (1.211+/-0.411) mmol/l for SM; (0.343+/-0.124) mmol/l for PI+PE. PLs of patients were assayed at least twice: at the time of diagnosis and, when appropriate, at the time of complete remission from the disease (CR). At the time of diagnosis, the mean concentrations of studied compounds were: (1.602+/-0.716) mmol/l for PC+CPLAS; (0.041+/-0.048) mmol/l for LPC; (0.398+/-0.198) mmol/l for SM; (0.045+/-0.071) mmol/l for PI+PE. After attainment of complete remission (CR), the respective values were as follows: (4.094+/-1.886) mmol/l for PC+CPLAS; (0.295+/-0.139) mmol/l for LPC; (1.123+/-0.634) mmol/l for SM; (0.230+/-0.125) mmol/l for PI+PE. All concentrations found in patients at the time of diagnosis were significantly lower than in reference group and in those benefited from complete remission (CR). By contrast the differences in concentrations of phospholipids in plasma between patients with complete remission (CR) and healthy volunteers were no statistically significant.     

Mobilization of arachidonic acid in collagen-stimulated human platelets.

Stimulation of platelets with collagen results in the mobilization of arachidonic acid (AA) from phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS) and phosphatidylinositol (PI). In this study the effect of aspirin, indomethacin, BW755C and prostaglandin H2 (PGH2) on labelled AA release in response to varied concentrations of collagen was investigated. Our results indicate that aspirin (0.56 mM) and indomethacin (5.6 microM) not only inhibited the collagen-mediated formation of cyclo-oxygenase metabolites, but also caused a significant reduction in the accumulation of free labelled AA and 12-hydroxyeicosatetraenoic acid (12-HETE) (21-64%). Aspirin and indomethacin also inhibited the release of [3H]AA from PC (37-75%) and PI (33-63%). The inhibition of AA release caused by aspirin was reversed partially by PGH2 (1 microM). In contrast, a smaller/no inhibition of collagen-stimulated labelled AA and 12-HETE accumulation (0-11%) and of collagen-stimulated AA loss from PC and PI was observed in the presence of BW755C. The results obtained in the presence of aspirin, indomethacin and BW755C at lower concentrations of collagen further demonstrate that AA release from PI (45-61% inhibition at 10 micrograms of collagen), but not from PC, was affected by the inhibition of cyclo-oxygenase. The results obtained on the effect of PGH2 further support that deacylation of phospholipids occurs independently of cyclo-oxygenase metabolites, particularly at higher concentrations of collagen. These results also demonstrate that aspirin and indomethacin, but not BW755C, cause a direct inhibition of collagen-induced [3H]AA liberation from PC as well as from PI. We also conclude that the diacylglycerol lipase pathway is a minor, but important, route for AA release from PI in collagen-stimulated human platelets. The mechanisms underlying the regulation of AA release by collagen in the absence of cyclo-oxygenase metabolites are not clear.     

Separation of phospholipid classes in sea urchin,paracentrotus lividus by high-performance liquid chromatography

A simple high-performance liquid chromatography (HPLC) method for determination of major phospholipid classes in sea urchin Paracentrotus lividus is described. The separation was performed on a Tracer Extrasil SI 5 microm 25 x 0.4 cm column and an isocratic mobile phase of acetonitrile-methanol 85%-phosphoric acid (50:50:1.8, v/v). The HPLC method utilizes UV detection at 205 nm. Five phospholipids were identified and quantified: phosphatidylinositol (PI), phosphatidylserine (PS), phosphatidylethanolamine (PE), phosphatidylcholine (PC) and sphingomyelin (SM). Fresh and canned samples were analyzed. Student's t-test showed no significant difference (P < or = 0.05) between the mean phospholipid contents of raw and canned sea urchin.