Isolation and some properties of the heparin-neutralizing factor (PF4) released from human blood platelets

Käser-Glanzmann, R.; Jakábová, Milica; Lüscher, E. F.

doi:10.1007/bf01946183

Cited by 37 publications

(12 citation statements)

References 7 publications

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“…Blunting 'or ablating of agonistevoked Cai response by cyclic nucleotides involve key elements ofCa, homeostasis, including inhibition of(a) agonist-receptor interaction (29); (b) PLC (11,12,30,31); (c) Ca mobilization from intracellular organelles (26,27,30,32); and (d) Ca uptake from the extracellular compartment (27, 30, 32). Additionally, cyclic nucleotides stimulate Ca sequestration (33). Irrespective of the specific mechanisms by which cyclic nucleotides lower the Ca,, cAMP and cGMP do not lower Ca, in platelets under basal conditions, or when the Ca1 is raised by Ca ionophores, as has been demonstrated here by us and by others (26,28,30,34).…”

Section: Discussionsupporting

confidence: 70%

Cyclic nucleotides attenuate thrombin-evoked alterations in parameters of platelet Na/H antiport. The role of cytosolic Ca.

Kimura¹,

Lasker²,

Aviv³

1992

J. Clin. Invest.

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In this work, we explored the role of cyclic nucleotides in modulating parameters of the Na/H antiport in human platelets. Sodium nitroprusside and iloprost, as well as cyclic nucleotide analogues, were used to raise cellular levels of cAMP and cGMP. Cyclic nucleotides reversed the thrombin-evoked alkaline shift in cytosolic pH set point and the activity of the Na/H antiport, concurrently with attenuation of thrombin-induced rise in cytosolic free Ca. No effect of cyclic nucleotides was observed in platelets not treated with thrombin, or platelets subjected to phorbol 12-myristate 13-acetate. cAMP did not reverse ionomycin-induced changes in the parameters of the Na/H antiport. Collectively, these observations indicate that cyclic nucleotides modulate the Na/H antiporter in human platelets through their effect on thrombin-evoked changes in cytosolic free Ca. Presumably, this effect holds for other agonists which stimulate phospholipase C, raise cytosolic-free Ca, and activate the Na/H antiport through protein kinase C dependent and protein kinase C-independent mechanisms. (J.

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Section: Discussionsupporting

confidence: 70%

Cyclic nucleotides attenuate thrombin-evoked alterations in parameters of platelet Na/H antiport. The role of cytosolic Ca.

Kimura¹,

Lasker²,

Aviv³

1992

J. Clin. Invest.

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“…Isolation o f PF4 from the supernatants of thrombin-treated, washed human blood platelets was carried out as described earlier [1,11]. This procedure includes gel filtration (Biogel A-5 m, 100-200 mesh) of the solution of a euglobulin precipitate from the super natant, whereby first a proteoglycan PF4 complex, and in a second step at an ionic strength of 0.75 fi, PF4 are isolated.…”

Section: Methodsmentioning

confidence: 99%

“…We have already reported preliminary results of this work [11], and, in particular, described the properties of the mucopolysaccharide moiety of the released PF4 com plex [1]. Here we want to report some additional properties of the heparinneutralizing factor and some observations pertaining to its disputed complex formation with fibrinogen and its derivatives.…”

mentioning

confidence: 97%

Heparin-Neutralizing Factor (Platelet Factor 4) from Human Blood Platelets and its Reactivity with Fibrinogen and Soluble Fibrin-Monomer Complexes

Käser-Glanzmann¹,

Jakábová²,

Lüscher³

1972

Pathophysiol Haemos Thromb

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The heparin-neutralizing factor (PF4) has been isolated from the supernatant of thrombin-treated, washed human blood platelets. The major part of the factor is present as a PF4-proteoglycan complex which can be dissociated at higher ionic strength. PF4 thus obtained is a basic protein of molecular weight 29,700. The carrier proteoglycan will recombíne with a maximum of 4 molecules of PF4, whereby dimerization of the resulting complex is observed. This is in contrast to the naturally occurring, released complex, which has a sedimentation constant which is rather compatible with the presence of monomeric material. Heparin displaces PF4 from its proteoglycan carrier; thereby a 1:1 heparin PF4 complex is formed. Contrary to preparations obtained by zinc precipitation, neither the PF4-carrier complex, nor purified PF4 obtained by dissociation at high ionic strength form insoluble complexes with fibrinogen or with complexes formed from fibrin-monomers and fibrinogen degradation products. Pretreatment of zinc-precipitated PF4 with EDTA abolishes its paracoagulating properties.

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“…A plasma factor with this property was first predicted inr 1948 by Conley (3) when he noticed that patients with thrombocytopenia had increased sensitivity to heparin. Partial purification of the specific, platelet-derived protein, referred to as platelet factor 4 (4), was reported by Deutsch et al (5), and, more recently, various investigators have published methods for isolation of the protein in an apparently homogeneous form (6)(7)(8)(9)(10)(11)(12)(13). Platelet factor 4 is a relatively small, heat-stable protein that is insoluble in solutions of low ionic strength, but that is soluble in high ionic strength solutions and in acidic buffers (12,13).…”

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confidence: 99%

Amino acid sequence of human platelet factor 4.

Deuel¹,

Keim²,

Farmer³

et al. 1977

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

277

121

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Human platelet factor 4, a protein that binds heparin, has been purified to apparent homogeneity and the complete amino acid sequence of the protein has been determined. The 70-residue polypeptide chain contains no methionine, tryptophan, or phenylalanine, and contains only a single tyrosyl residue. The sequence analysis demonstrates a highly negatively charged amino-terminal region. The carboxyl-terminal region of the polypeptide is unusual in that it contains a repetitive clustering of positively charged and hydrophobic pairs of amino acids; preliminary evidence suggests that this domain may play a role in the binding of heparin.Heparin is a potent anticoagulant that is used extensively in clinical medicine for the treatment of thrombotic disease (1). Platelets release-a small protein that neutralizes the anticoagulant effect of heparin (2). A plasma factor with this property was first predicted inr 1948 by Conley (3) when he noticed that patients with thrombocytopenia had increased sensitivity to heparin. Partial purification of the specific, platelet-derived protein, referred to as platelet factor 4 (4), was reported by Deutsch et al. (5), and, more recently, various investigators have published methods for isolation of the protein in an apparently homogeneous form (6-13). Platelet factor 4 is a relatively small, heat-stable protein that is insoluble in solutions of low ionic strength, but that is soluble in high ionic strength solutions and in acidic buffers (12, 13). Platelet factor 4 is released with and bound to a carrier proteoglycan molecule from which it is displaced by heparin (8, 10).Because of a possible relationship of platelet factor 4 to arterial and venous thrombosis (14)(15)(16)(17)(18)(19)(20) tained by precipitation between ammonium sulfate concentrations of 50 and 100%, by affinity chromatography on columns of heparin-E-aminocaproic acid-Sepharose prepared as described by Schmer (21), and by gel filtration through Sephadex G-100 in 1 M NaCl. These methods and the modified thrombin time used in the assay of platelet factor 4 were adapted from those reported by Levine and Wohl (12). Purified platelet factor 4 binds 27 units of heparin per mg of protein.The purified protein appears as a single band after polyacrylamide gel electrophoresis (15% acrylamide) in the presence of sodium dodecyl sulfate and is estimated to be at least 95% pure both by this criterion and by automated Edman degradation of the intact S-carboxamidomethylated protein. The purity of the preparation was also established by immunological techniques. The IgG fraction of antisera obtained from rabbits immunized with platelet factor 4 was isolated by ammonium sulfate precipitation and DEAE-cellulose chromatography (22). The purified IgG gave a single precipitin line against purified platelet factor 4 in immunodiffusion experiments, suggesting that the purified protein behaves as a single major antigenic species.

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Isolation and some properties of the heparin-neutralizing factor (PF4) released from human blood platelets

Cited by 37 publications

References 7 publications

Cyclic nucleotides attenuate thrombin-evoked alterations in parameters of platelet Na/H antiport. The role of cytosolic Ca.

Cyclic nucleotides attenuate thrombin-evoked alterations in parameters of platelet Na/H antiport. The role of cytosolic Ca.

Heparin-Neutralizing Factor (Platelet Factor 4) from Human Blood Platelets and its Reactivity with Fibrinogen and Soluble Fibrin-Monomer Complexes

Amino acid sequence of human platelet factor 4.

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