Cytochemical localization of lysosomal enzymes in rat megakaryocytes and platelets.

Bentfeld, Mary E.; Bainton, Dorothy F.

doi:10.1172/jci108246

Cited by 109 publications

(44 citation statements)

References 43 publications

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“…Further separation of the granule fraction by isopycnic centrifugation on a sucrose gradient revealed factor V and LA-PF4 to be maximal in a denser fraction of the gradient than the acid hydrolases. The acid hydrolases are not located in the a granules (23,24), whereas the heparin-neutralizing proteins (25,26), fibrinogen (27), fibronectin (28), von Willebrand factor (29), and platelet-deprived growth factor (26) are. Our data suggest that factor V is also an a granule constituent.…”

Section: Discussionmentioning

confidence: 99%

Subcellular localization and secretion of factor V from human platelets.

Chesney¹,

Pifer²,

Colman³

1981

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

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ABSTRACT' Factor V, a plasma protein cofactor necessary for optimal conversion of prothrombin to thrombin, is also present in considerable concentration in blood platelets (9.9 units per 109 platelets). Subcellular fractionation by. two methods has localized factor V in the a granules of-unstimulated platelets. ADP and epinephrine cause release of4.6% and 6.4%, respectively, ofthe total factor V, a process completely inhibited by cyclooxygenase alkylation by aspirin. In contrast, collagen causes release of 25% of platelet factor V, a process only partially suppressed by aspirin. Secretion of factor V depends on the availability of metabolic energy, because antimycin A,. an inhibitor of aerobic metabolism, and 2-deoxyglucose, an inhibitor of anaerobic glycolysis, together almost totally inhibited the secretion of factor V induced by collagen. The data establish that factor V is not normally available on unstimulated platelets but can be secreted from a granules upon stimulation with physiological agents such as ADP, epinephrine, and collagen. Because factor V is known to serve as a receptor for factor Xa, the exposure of factor V on platelets consequent to release would accelerate the process of blood coagulation.One year after the discovery of factor V in plasma, Ware et al.(1) recognized that platelets also contain an accelerator of prothrombin conversion. Though this coagulant activity was more stable to heat inactivation and storage than was plasma factor V, it was generally assumed to be the plasma factor adsorbed to the platelet surface (2, 3). In 1975, Breederveld et aL (4), using gel filtration to isolate platelets from plasma, found minimal activity of factor V associated with platelets but were able to increase the V activity by freeze-thawing. 0sterud et aL (5) confirmed this observation and showed that platelets contain an activated form offactor V as well as a platelet activator ofplasma factor V. Kane et aL (6) have presented convincing evidence that factor Xa binds to the platelet membrane at a binding site that is identical to or requires the presence of factor V.Despite these important observations, several questions remain in evaluating the influence of platelet factor V on the hemostatic and possible thrombotic processes. What is the sub, cellular localization offactor V? Are prostaglandin synthesis and metabolic energy required for its release? Do collagen, ADP, and epinephrine release factor V as thrombin does? This study attempts to answer these questions. MATERIALS AND METHODSBovine serum albumin (fraction V), ADP, epinephrine, and bovine fibrinogen (fraction 1, type IV, 93% clottable) were supplied by Sigma. Human fibrinogen (grade L, 95%

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Section: Discussionmentioning

confidence: 99%

Subcellular localization and secretion of factor V from human platelets.

Chesney¹,

Pifer²,

Colman³

1981

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

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“…Decker (28) has demonstrated both enzyme activities in GERL of neurons. Bentfeld and Bainton (29) have shown arylsulfatase activity in GERL of megakaryocytes.…”

Section: Ingsmentioning

confidence: 98%

The endoplasmic reticulum: a cytochemist's view (a review).

Novikoff¹

1976

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

363

126

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Enzyme cytochemistry has been used, at the light and electron microscope levels, to "mark" cytoplasmic organelles of mammalian cells. Catalase cytochemistry permitted identification of microperoxisomes, apparently ubiquitous organelles that are attached by numerous slender coflnections to the endoplasmic reticulum. Thiamine pyrophosphatase and acid phosphatase cytochemistry can be used to distinguish between the Golgi apparatus and a specialized acid-phosphatase-rich region of smooth endoplasmic reticulum (ER) that appears to be involved in: (a) the formation of lysosomes and melanin granules: (b) the processing and ackaging of secretory materials in endocrine and exocrine cells; and (c) the metabolism of lipid. The acronym GERL has been given to this region of smooth ER because it is located at the inner or "trans" aspect of the Golgi apparatus and because it appears to produce various types of Lysosomes.

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“…FICURE 5 Ultrathin cryosections of freshly fixed platelets, immunostained with PAG for platelet factor 4. Immunoreactivity occurs in a-granules.…”

Section: Resultsmentioning

confidence: 99%

“…Using morphologic criteria three possible secretory granule classes have been recognized: alpha granules, dense granules, and lysosomelike granules (5,6).…”

Section: Introductionmentioning

confidence: 99%

Immunocytochemical localization of fibrinogen, platelet factor 4, and beta thromboglobulin in thin frozen sections of human blood platelets.

Sander¹,

Slot²,

Bouma³

et al. 1983

J. Clin. Invest.

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A B S T R A C T Affinity-purified monospecific antibodies against human fibrinogen and the platelet-specific proteins platelet factor 4 and beta thromboglobulin were used to localize these antigens in thin and ultrathin frozen sections of mildly fixed, washed human blood platelets. By immunofluorescent double-labeling experiments the distribution of fibrinogen was compared to that of platelet factor 4 and beta thromboglobulin. All three antigens occurred in virtually all platelets and showed an identical, dotlike distribution. For immunoelectron microscopy we used protein Acolloidal gold on ultrathin frozen sections to visualize the specific reaction indirectly. The staining for platelet factor 4, beta thromboglobulin, and fibrinogen localized exclusively over a-granules of washed platelets. Within the granules, platelet factor 4 was localized preferentially in the electron dense, a-granule nucleoid, whereas fibrinogen was more predominant in the electron-lucent granule periphery. Beta thromboglobulin localization did not show a preferential intragranular distribution.

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Cytochemical localization of lysosomal enzymes in rat megakaryocytes and platelets.

Cited by 109 publications

References 43 publications

Subcellular localization and secretion of factor V from human platelets.

Subcellular localization and secretion of factor V from human platelets.

The endoplasmic reticulum: a cytochemist's view (a review).

Immunocytochemical localization of fibrinogen, platelet factor 4, and beta thromboglobulin in thin frozen sections of human blood platelets.

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