Lipid metabolism in human platelets

Lewis, Nancy M.; Majerus, Philip W.

doi:10.1172/jci106178

Cited by 95 publications

(16 citation statements)

References 25 publications

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“…The composition of the phospholipid fraction is given in Table II. Our data agree reasonably well with data in the literature (31)(32)(33)(34)(35)(36)(37)(38)(39). A small amount of phosphatidylinositol was detectable but was not quantified.…”

Section: Introductionsupporting

confidence: 91%

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The effect of phospholipase C on human blood platelets.

Otnaess¹,

Holm²

1976

J. Clin. Invest.

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A B S T R A C T The effect of phospholipase C (EC 3.1.4.3) on human blood platelets has been studied. Phospholipase C from Bacillus cereus was purified to homogeneity as judged by analytical and sodium dodecyl sulphate disc gel electrophoresis and by immunoelectrophoresis. Human platelets isolated from platelet-rich plasma by gel filtration or by centrifugation and washing were incubated with phospholipase C.A loss of 20-45% of the total platelet phospholipid was observed, whereas 88% was hydrolyzed when platelet homogenates were submitted to identical enzyme treatment. Intact platelets lost 50-75% phosphatidylethanolamine, 20-50% phosphatidylcholine, and 20-25% phosphatidylserine. Sphingomyelin was not a substrate for the enzyme under the conditions used. The platelets contained no detectable endogenous phospholipase C activity.The loss of phospholipid was not accompanied by aggregation of the platelets, nor did the platelets lose their ability to aggregate with ADP or thrombin. Total platelet factor 3 releasable by freezing and thawing was reduced. Measurements of releasable platelet factor 4 and the efflux of serotonin showed that no release reaction was triggered even when up to 45% of the total phospholipid in the platelets was hydrolyzed. When sphingomyelinase was added together with, before, or after phospholipase C, aggregation occurred. Sphingomyelinase alone gave no aggregation. The gel-filtered platelets also aggregated upon addition of purified phospholipase C from Clostridium perfringens. The distribution of phospholipids in the platelet membrane is discussed.

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Section: Introductionsupporting

confidence: 91%

“…Immunoelectrophoresis was carried out as described by Scheidegger (27) Phospholipids. In good accordance with data of others (30,31), the platelets were found to contain 10.0-13.9 ug lipid phosphorus/109 platelets. The composition of the phospholipid fraction is given in Table II.…”

Section: Introductionsupporting

confidence: 91%

The effect of phospholipase C on human blood platelets.

Otnaess¹,

Holm²

1976

J. Clin. Invest.

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“…Previous work done in our laboratory (2) has shown that thrombin causes a 7-fold increase in the rate of incorporation of glycerol into phosphatidyl serine in the platelet membrane. This burst in the rate of lipid synthesis is maximal within 2 min of the addition of thrombin and such synthesis returns to the pre-thrombin rate by 20 (8).…”

mentioning

confidence: 76%

“…Platelets are anucleate circulating particles, with a diameter of [2][3][4] Mum in man, which function in hemostasis. These "cells" are metabolically active and catalyze the formation of fatty acids, phospholipids, glycogen, and even protein (1).…”

mentioning

confidence: 99%

A Thrombin-Sensitive Protein of Human Platelet Membranes

Baenziger

Brodie

Majerus

1971

Proc. Natl. Acad. Sci. U.S.A.

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323

201

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The action of thrombin on intact human platelets has been studied with the aid of polyacrylamide gel electrophoresis in sodium dodecylsulfate. A single major membrane protein band with a molecular weight of 190,000 disappears after thrombin treatment, while a new membrane protein with a molecular weight of 107,000 appears. This may represent hydrolysis of the thrombinsensitive protein. When platelets are disrupted or when the thrombin-sensitive protein is solubilized from membranes prior to thrombin treatment, no hydrolysis occurs. The effect of thrombin on the platelet membrane protein is complete within 2 min which suggests that hydrolysis of this membrane protein may trigger the physiological effects of thrombin on platelets.Platelets are anucleate circulating particles, with a diameter of 2-4 Mum in man, which function in hemostasis. These "cells" are metabolically active and catalyze the formation of fatty acids, phospholipids, glycogen, and even protein (1). Both glycolytic and Krebs cycle oxidative pathways are active in these cells.Blood platelets are rapidly converted from free floating cells to an aggregated mass of intact platelets during the process of hemostasis or of thrombus formation. This obvious change in the platelet surface is accompanied by a variety of morphological and biochemical changes which have been termed "viscous metamorphosis" (1). The most potent agent known to induce this process is the proteolytic enzyme thrombin, which is generated from reactions of the coagulation pathway. Characterization of the role of thrombin in producing viscous metamorphosis is integral to understanding thrombus formation and may also provide some insight into the membrane specification of cell-cell interactions.Previous work done in our laboratory (2) has shown that thrombin causes a 7-fold increase in the rate of incorporation of glycerol into phosphatidyl serine in the platelet membrane. This burst in the rate of lipid synthesis is maximal within 2 min of the addition of thrombin and such synthesis returns to the pre-thrombin rate by 20 min. Studies of the mechanism of this thrombin effect indicate that proteolysis is involved, since the effect can be mimicked by trypsin and inhibited by prior incubation of trypsin with soybean trypsin inhibitor. Human platelets, collected and isolated as described previously (2), were washed twice in a pH 6.5 isotonic buffer containing 0.113 M NaCl, 0.0043 M K2HPO4, 0.0043 M\I Na2HPO4, 0.0244 M NaH2PO4, and 1 mg/ml glucose. After washing, l)latlets were resuspended in either 0.154 M NaCl-0.154 A Tris HCl (pH 7.4) 9:1 with 1 mg/ml glucose, or in a pH 7.5 buffer containing 0.109 M NaCl, 0.043 M K2HP04, 0.0106 M Na2HPO4, 0.0083 MI NaH2PO4, and 1 mg/ml glucose.Platelets were incubated at 370C at 1-2 X 109 cells/ml in resuspension buffer with or without 1 U/ml thrombin and 2.5 mM CaCl2. The cells were disrupted by no-clearance-pestle homogenization (5), glycerol loading, and hypotonic lysis (6) Samples were solubilized in 1-3%o SDS-1%o 0-mercaptoethanol -0.1 AM so...

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“…Although considerable information has been accumulated regarding the biochemistry and physiology of platelet-plug formation (1)(2)(3)(4)(5)(6)(7), little is known about initiation of platelet plug formation, namely platelet adhesion. Early work, employing light microscopy, (8)(9)(10) had suggested that platelets adhered to traumatized mesenteric fibers which were of "collagen" nature.…”

Section: Introductionmentioning

confidence: 99%

Biochemical and biophysical aspects of human platelet adhesion to collagen fibers

Lyman¹,

Rosenberg²,

Karpatkin³

1971

J. Clin. Invest.

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A B S T R A C T A method has been developed for measuring the adhesion of platelets to purified collagen fibers obtained from bovine tendon. This method differs from others in that: (a) platelet adhesion is measured in the absence of platelet aggregation; (b) platelet-rich plasma collected in ACD (acid citrate dextrose) or EDTA, or washed platelets can be employed; (c) adherent platelets are enumerated directly; (d) erythrocytes and leukocytes do not adhere.Washed platelets suspended in human Ringer solution exhibit negligible adhesion (at the platelet concentrations employed) in contrast to washed platelets suspended in plasma. Addition of purified human fibrinogen (95% clottable, 2-4 mg/ml) to human Ringer solution completely restores the ability of washed platelets to adhere to collagen fibers. Albumin (fatty acid free, 50 mg/ml) is also capable of restoring adhesion. Albumin and seven other proteins at concentrations of 5-10 mg/ml, with varying molecular weights, isoelectric points, and frictional coefficients are incapable of supporting the adhesion of washed platelets. The proteins tested were human globulin, hexokinase, hemoglobin, cytochrome-C, insulin, thyroglobulin, and muramidase. Platelet adhesion is proportional to both platelet concentration and fibrinogen concentration, but is independent of temperature or glycogen stores.Modification of fibrinogen by acylation of amino groups or removal of sialic acid has no effect on its ability to support platelet adhesion. Degradation of fibrinogen with purified plasmin results in decreased support of platelet adhesion. This accompanied formation of early breakdown products with clottability ranging from 84-0%. Formation of fibrinogen degradation products was monitored by SDS-polyacrylamide gel electrophoresis of the corresponding fibrins after reduction of Received for publication 21 January 1971 and in revised form 17 February 1971. disulfide bonds (a method capable of distinguishing a-, P-and gamma-chains). Decreased support of platelet adhesion is associated with the disappearance of intact a-chains and early modification of the a-chains.Purified proteinpolysaccharide macromolecules obtained from bovine nasal and humeral cartilage, and from nucleosus pulposus are as effective as fibrinogen on a weight basis and ten to thirty times more effective on a molar basis in supporting platelet adhesion. The purified mucopolysaccharide side chains: chondroitin-4-sulfate, chondroitin-6-sulfate, and keratan-sulfate are incapable of supporting platelet adhesion. INTRODUCTIONAlthough considerable information has been accumulated regarding the biochemistry and physiology of platelet-plug formation (1-7), little is known about initiation of platelet plug formation, namely platelet adhesion. Early work, employing light microscopy, (8)(9)(10)

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Lipid metabolism in human platelets

Cited by 95 publications

References 25 publications

The effect of phospholipase C on human blood platelets.

The effect of phospholipase C on human blood platelets.

A Thrombin-Sensitive Protein of Human Platelet Membranes

Biochemical and biophysical aspects of human platelet adhesion to collagen fibers

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