Although the existence of blood platelets had been known for many years it was not until 1882 that Bizzozero (1) called attention to the role of platelets in blood coagulation. Morawitz (2), twenty years later, was among the first to consider the specific chemical contribution of the platelet to the clotting process.Howell (3), in 1912, demonstrated the presence of an unsaturated phosphatide ("kephalin") in thromboplastic substances of tissue origin. Mills (4), in 1927, showed cephalin to be present in blood platelets, and the following year Haurowitz and Sladek (5) published results of chemical analysis of horse platelets, reporting a lipid content of 12 per cent of dry weight. The first convincing data relating the cephalin content of platelets to acceleration of coagulation were supplied by Chargaff, Bancroft and Stanley-Brown (6). These workers also reported at that time what has recently been emphasized again (7,8): phosphatides from other natural sources (soy bean, yeast) also were capable of accelerating blood clotting in vitro. Erickson, Williams, Avrin and Lee (9), confirmed the results of Chargaff and associates relating to platelet phospholipid content. Wallach, Surgenor and Steele (10), identified phosphatidyl ethanolamine and phosphatidyl serine in human platelets. Subsequently the presence of inositol phosphatide was noted ( 11,12 A. Isolation of platelets from whole blood. Blood was obtained from normal adult volunteers; it was collected by gravity flow into plastic bags containing 1.5 per cent disodium dihydrogen ethylenediaminetetraacetate in 0.7 per cent NaCl,1 employing a plastic donor set.2 A proportion of 9 volumes of blood to 1 volume of anticoagulant was used. Forty ml aliquots were then gently decanted into silicone-coated3 glass tubes and these were centrifuged at room temperature at 300 G for 15 minutes. All glassware used in collection and transfer of platelet-containing plasma was similarly treated. All glassware used in the coagulation studies was acidcleaned. The supernatant platelet-rich plasma was transferred by pipette to glass centrifuge tubes. Aliquots of platelet-rich plasma were then taken for platelet and white cell counting, and aliquots to be used in coagulation studies were stored at 4°C in glass tubes. The platelet-rich plasma thus obtained had fewer than 300 leukocytes per cu mm, and was completely free of red cells. The platelet-rich plasma was then centrifuged at 2,000 G at 40 C. The supernatant platelet-poor plasma thus obtained was decanted into other centrifuge tubes and again centrifuged at 40 C for 30 minutes at 2,000 G. This supernatant plasma had fewer than 1,000 platelets per cu mm (49) and was considered to be "plateletpoor." The sedimented platelets obtained after the first 2,000 G centrifugation were pooled and washed 3 times with isotonic NaCl solution and resedimented by centrifugation at 40 C for 5 minutes at 900 G. The platelets thus obtained were then subjected to lipid extraction and analysis.B. Extractiont, separation, and measurement of platelet and...

The liver has been proposed as a site of synthesis of blood-clotting factors, in part because of the demonstrated dominant role of the liver in synthesis of all plasma protein fractions except the y-globulins (1, 2). In addition, observations after hepatectomy (3, 4) and hepatic injury (5, 6) and in hepatic disease (7,8) have provided a body of evidence consistent with hepatic synthesis of fibrinogen, prothrombin, and Factors V, VII, IX, and X. This evidence, however, has not been considered definitive, mainly because it is based on indirect observations in vivo in humans and in animals suffering from variable, often profound, metabolic abnormalities induced by hepatic dysfunction or extirpation.This report describes the use of the isolated perfused rat liver in a more direct experimental evaluation of the role of the liver in the production of blood-clotting factors. The use of the isolated perfused liver was encouraged by Lupton's observations of shortened prothrombin time during perfusions of 3 to 4 hours (9), and by Pool and Robinson's studies of changes in clotting factor activity during in vitro incubation of rat liver slices (10). Presented here is not only more detailed direct evidence of hepatic synthesis of prothrombin and Factor VII, but also the first direct evidence for hepatic synthesis of Factors V and X.

No abstract

The effects of PCMB and PCMBS on cation permeability, ADP aggregation, platelet volume and clot retraction have been studied in intact human platelets and have been compared to the effects of ouabain on the same functions. The influence of these agents on Mg+ + -dependent ATPase activity and Na+, K+ activated ATPase activity of osmotically lysed platelets also has been evaluated. (a) Ouabain, which produces inhibition of Na-, K+ dependent ATPase activity, has no influence on ADP-induced aggregation or clot retraction. (b) Both PCMB and PCMBS inhibit all ATPase activities in osmotically lysed platelets and inhibit ADP aggregation by intact platelets. Cysteine reverses the inhibition of ADP aggregation produced by both PCMB and PCMBS at a time when PCMB induced inhibition of clot retraction no longer is reversible. (c) Clot retraction is inhibited by the penetrating agent, PCMB, but is not affected by identical concentrations of PCMBS. These observations suggest that ADP aggregation and clot retraction are independent phenomena.

No abstract