Subcellular localization of the heparin‐neutralizing factor in blood platelets.

Prada, M. Da; Jakábová, Milica; Lüscher, E. F.; Pletscher, A.; Richards, John

doi:10.1113/jphysiol.1976.sp011380

Cited by 30 publications

(13 citation statements)

References 21 publications

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“…It has been suggested that antiheparin activity is located in the alpha granules of platelets (19,20). Holmsen et al (21) have reported that higher concentrations of thrombin are needed for alpha granule release than for dense body secretion.…”

Section: Resultsmentioning

confidence: 99%

“…Several recent observations indicate that the interaction of platelets with clotting factors during thrombin formation is more complex than initially postulated. Osterud et al (2) have shown that platelets contain an activated form of Factor V and have postulated Received for publication 19 December 1977 and in revised form 23 january 1978. that its release may be important in prothrombin activation. Experiments by Miletich et al (3) demonstrate that the rate of thrombin generation is 10 to 100-fold faster when platelets are used in place of phospholipid and Factor V in the activation of prothrombin by Factor Xa.…”

Section: Introductionmentioning

confidence: 99%

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Thrombin generation and secretion of platelet Factor 4 during blood clotting.

Shuman¹,

Levine²

1978

J. Clin. Invest.

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A B S T R A C T We have studied the platelet release reaction and thrombin generation during the spontaneous clotting of whole blood in vitro. Both thrombin formation and secretion of platelet Factor 4 were detected at least 12 min before clotting (clotting time, 22-26 min). Initially, at low thrombin concentrations (2-5 ng/ml), there is a small increase in plasma platelet Factor 4 (<1% of the amount present in serum). This is followed by a gradual increase in both platelet Factor 4 and thrombin concentrations over a 12 to 20-min interval. Finally, 5 min before clotting, there is a rapid increase in both thrombin generation and platelet secretion.Thus, we have shown that the release of platelet Factor 4 is a prolonged reaction and the extent to which it occurs parallels thrombin generation. It is only when thrombin concentrations are high (45-90 nglml)-during the period of clot formation-that the major part of platelet Factor 4 secretion occurs. Release of platelet Factor 4, like fibrin formation, occurs in the last step of in vitro coagulation. INTRODUCTIONPlatelets have been assigned a dual role in hemostasis. Initially, after vascular injury, a platelet aggregate or plug is formed on the subendothelial surface. During plug formation, platelets undergo changes that result in the exposure of a phospholipid surface on which Factors X and II are converted to their active forms (1). Several recent observations indicate that the interaction of platelets with clotting factors during thrombin formation is more complex than initially postulated. Osterud et al. (2) have shown that platelets contain an activated form of Factor V and have postulated

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Thrombin generation and secretion of platelet Factor 4 during blood clotting.

Shuman¹,

Levine²

1978

J. Clin. Invest.

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“…Methods for isolation of human e-granules have been reported in the past [8,11] but the fact that improvements on these methods are still being introduced [19] is an indication that they are only partially satisfactory. In this report, pig e-granules prepared by a modification of pre-existing procedures [8,16,40] were analyzed biochemically and morphologically.…”

Section: Discussionmentioning

confidence: 99%

“…The latter include platelet factor 4, fl-thromboglobulin, a cell growth factor, fibronectin, factor V, thrombospondin, 0%-antiplasmin and fibrinogen. Each group of exported molecules is thought to be stored in a distinct secretory organelle: serotonin, Ca + + and the adenine nucleotides are localized in the dense granules [12,13,16]; hydrolases are thought to be stored in lysosomes [5,11] and the polypeptide macromolecules are found in a third type of organelle, the 0~-granule [11,27,28,48,49]. Studies on patients with the "gray platelet" syndrome and with storage pool disease have shed some light on the compartmentation of the released products.…”

Section: Introductionmentioning

confidence: 99%

Isolation and partial characterization of platelet α-granule membranes

1983

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Porcine alpha-granules, prepared by a modification of pre-existing methods, were found to be essentially homogeneous by transmission electron microscopy. Freeze-fractured samples of isolated granules revealed numerous intramembranous particles on the EF (exoplasmic fracture) surface and to a lesser extent on the PF (protoplasmic fracture) surface whereas the PS (protoplasmic) surface was relatively smooth. The granules appear to be sealed, as evidenced by: a) the retention of their electron dense core material; b) the inability of impermeant labels to react with the granule contents, and c) the finding that the intragranular proteins are refractory to mild hydrolysis by externally added proteases. Membranes were isolated by alkali extraction of the granules and used for biochemical characterization. Approximately 87% of the protein, but only insignificant amounts of phospholipid were removed by this procedure, which yielded membrane vesicles devoid of the dense core. The membranes contain one major and several minor polypeptides of molecular weights ranging from 28,000 to 230,000, as determined by polyacrylamide gel electrophoresis. The major polypeptide contains carbohydrate residues. The exposure of specific proteins on the cytoplasmic surface of the granule membrane was determined by a combination of surface-specific labeling and proteolysis of intact granules, followed by membrane isolation and analysis. In sealed granules, only a limited number of bands are modified by the reagents whereas most of them are affected following granule lysis, indicating asymmetry in their transmembrane disposition. The fraction eluted by alkali extraction was also analyzed and found to contain nine major polypeptides of molecular weights ranging from 230,000 to 43,000. These are compared to the weights of the macromolecules believed to be secreted from alpha-granules, as determined by radioimmunological techniques.

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“…In fact, the possibility that proelastase and its trypsin-like acti vator be localized in different light granules should not be discarded and is supported by the possible existence in platelets of different types of a-granules as suggested by Morgenstern et al [27] and Salganicoff et al [31]: these last authors have clearly demonstrated the wide distribution of lyso somal enzymatic markers in various subfractions separated in particular conditions from a platelet homogenate. A heterogeneity of the dense bodies has also been shown by Da Prada et al [8] since platelet factor IV is located in granules distinct from the serotonin and adenine nucleotides storage granules. Whatever the precise localization of proelastase and its activator be, an important point has been raised by Nachman and H arpel [28] who have recently shown the presence of a 2-macroglobulin and oq-antitrypsin in fractions obtained from solubilized platelet membranes and granules: the trypsin-like enzyme could thus be inhibited in an enzyme-inhibitor complex which could be directly or indirectly dissociated after adhesion of the platelet to collagen.…”

Section: Discussionmentioning

confidence: 70%

Human Blood Platelet Elastase and Proelastase

Legrand¹,

Pignaud²,

Caen³

1977

Pathophysiol Haemos Thromb

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After an in vitro incubation of platelets with fibrillar collagen, their elastase activity is markedly and rapidly increased while proelastase decreases: proelastase is activated in situ into elastase which is released in its active form from the platelet. The activation of proelastase is likely due to the action of a trypsin-like enzyme present in the platelet. This protease has the same type of localization as proelastase and elastase: their highest activity is associated with light granules but part of these enzymes (or precursor) is also associated with the membranes. The mechanism of the arterial elastolysis induced by the platelets probably involves their adhesion to intimal thrombogenic surfaces (collagen) followed by a reaction during which proelastase would become available to the trypsin-like enzyme and would be activated into elastase directly released in the vessel wall.

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Subcellular localization of the heparin‐neutralizing factor in blood platelets.

Cited by 30 publications

References 21 publications

Thrombin generation and secretion of platelet Factor 4 during blood clotting.

Thrombin generation and secretion of platelet Factor 4 during blood clotting.

Isolation and partial characterization of platelet α-granule membranes

Human Blood Platelet Elastase and Proelastase

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