DIC related to APL (3 cases, No. 1-3), AML (No. 4), gastric cancer (No. 5), prostate cancer (No. 6), progressive systemic sclerosis (No. 7), and mitral stenosis & insufficiency (No. 8), were described.
Most of them showed, more or less, remarkable hemorrhagic diathesis.
In the terminal stage of Case No. 1, 2, 4 and 7, shock, oliguria and renal failure that might be due to DIC were observed in clinical and some laboratory findings. FDP and thrombocytopenia were demonstrable in all.
Fibrinogen was reduced in most of them, however, in cases already complicated with some infections when the diagnosis of DIC was made, it revealed higher level than in the other.
All cases were intravenously treated with heparin from 6, 000 to 20, 000U/day. In case No. 2, t-AMCHA was used because of negative paracoagulation test and few data suggesting DIC, and in case No. 1, 5, the same was done because of no improvement of hemorrhagic diathesis.
It was impressed that in Case No. 1 t-AMCHA was effective and Case No. 6 recovered dramatically by heparin.
However, it may be suggested that the effect of treatments on DIC might be affected by the improvement of each basic diseases, as they recovered simultaneously from each basic ones.
Case No. 8 showed no change in clinical and hematological findings with or without therapy. In addition, from echocardiogram, thrombus, which was found in left atrium, might be a cause of DIC findings.
Six cases (No. 1, 2, 3, 4, 5 and 7) ended lethally and autopsy was done in all.
In three cases (No. 2, 3 and 7) of them, microthrombi were pathologically demonstrable in some tissues.
As microthrombi are indispensable to pathological diagnosis of DIC, it was just identified in above three cases.
From 1966 to 1975, microthrombi were observed in all of 23 DIC cases which were examined in our department of pathology.
In comparison to it, less microthrombi were seen in this presentation. This discrepancy between two papers may depend on intensity of fibrinogenolysis and/or fibrinolysis, effects of treatments, degree of postmorten changes and tissue staining methods.
As heparin was used in all cases, it may be suggested that it's use is related to one of the causes of decreased detection rate of microthrombi.
It has been reported by some authors that DIC may not be always based on primary coagulation-secondary fibrinolysis, because sometimes antiplasmin therapy may be effective in DIC.
As the mechanism of DIC has not been completely recognized at present, it may be suggested that “Defibrination Syndrome” may be prefered to DIC.

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Various factors affecting coagulation and fibrinolysis, such as factor VIII procoagulant activity, factor VIII related antigen, von Willebrand factor activity, fibrinogen, plasminogen-plasmin, plasmin inhibitor and fibrin degradation product (F. D. P.) in blood were studied together with plasminogen activator, plasminogen-plasmin and F. D. P. in urine for a total of 45 cases of Schönlein-Henoch purpura which include 25 male and 20 female Japanese children of 2 to 15 years of age.
These children were classified into 4 groups, according to clinical manifestations: Group 1, which included 20 patients with no renal complications; Group 2, which included 16 patients with acute glomerulonephritis; Group 3, which included 9 patients with nephrotic syndrome; Group 4, which included no patient in this study and in whom chronic nephritis was evident.
Group 1 showed a high level of serum F. D. P. in 13 cases, at the initial stage, but the other fibrinolytic factors, factor VIII procoagulant activity, factor VIII related antigen and von Willebrand factor activity fell within normal range. The high level of serum F. D. P. returned to almost normal after 2 to 4 weeks.
Group 2 showed, at the initial stage, high levels (over 150%) of factor VIII procoagulant activity, factor VIII related antigen and von Willebrand factor activity in 5-8 cases and normal ranges were reverted to 3 weeks to 3 months later. A moderate increase was observed in fibrinogen content in 8 cases and a high level of serum F. D. P. in 15. Urinary levels of plasminogen activator, plasminogen-plasmin and F. D. P. were increased in 10-20 cases. Normal ranges were reverted to 6 weeks to 3 months later.
Group 3 showed, at the initial stage, high levels of factor VIII procoagulant activity, factor VIII related antigen and von Willebrand factor activity, in 6-7 cases and all levels returned to normal except for a few cases. An increase was observed in plasma fibrinogen, plasminogen-plasmin and serum F. D. P. in most cases. Levels of plasminogen activator, plasminogen-plasmin and F. D. P. in the urine were increased significantly in all cases.
Renal biopsy was performed on 17 children with Schönlein-Henoch purpura accompained with renal symptoms and histological findings and deposition of fibrin or factor VIII related antigen in the glomerular lesion were investigated.
Histological observations revealed with minimal change, 7 with proliferative glomerulonephritis, 2 with focal segmental glomerulonephritis and 5 with focal lobular glomerulonephritis.
The degree of glomerular fibrin deposits correlated closely with plasma fibrinogen, plasminogen-plasmin in plasma and urine, and F. D. P. in serum and urine.
The degree of glomerular deposition of factor VIII related antigen showed a close correlation not only with factor VIII procoagulant activity, factor VIII related antigen and von Willebrand factor activity, but with fibrin deposition.

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Factor VIII procoagulant activity (VIII: C), factor VIII related antigen (VIII R: AG) and von Willebrand factor (VIII R: WF) were measured in patients with liver disease (33 cases of cirrhosis, 8 cases of acute hepatitis, 10 cases of chronic hepatitis, 3 cases of fulminant hepatitis and 3 cases of primary liver cancer) and in 30 normal individuals.
Level of the three subunits of factor VIII complex were increased in cirrhotic patients as compared to those in normal individuals. In normal individuals there was a high degree of correlation between the three subunits, whilst this correlation was low in cirrhotic patients. In most cases with liver cirrhosis, high levels of VIII R: AG were not followed by a proportional increase in VIII R: WF or VIII: C. Similar results were obtained in the other liver diseases. These data suggest a functionally or qualitatively different factor VIII complex molecule from that found in normal plasma.
The two dimensional crossed immunoelectrophoresis (CIE) of the plasmas from patients with liver disease in whom disproportionately important elevation of VIII R: AG was found as compared with VIII R: WF, revealed abnormalities of the shape and migration of the precipitin arcs with more anodal electrophoretic mobility. The elution profile of the cryoprecipitate on Sepharose 4B of the plasma of liver cirrhosis differed from normal in respect that the partially purified factor VIII protein had markedly reduced VIII R: WF. These studies indicate that some patients with liver disease may comprise both quantitative and qualitative defects of factor VIII complex, presenting discrepancy between VIII R: AG and VIII R: WF.
Although the reason for the abnormality is not clear it may be related to the abnormal synthesis of factor VIII protein or to the accumulation of partially degraded protein due to normal or abnormal catabolism, or to disorder of joining or activating VIII: C. An alternative possibility is that it could be due to degradation of the protein by plasmin as the authers previously reported.

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Female carriers of hemophilia are usually asymptomatic. Female hemophilia B or symptomatic carriers of hemophilia B are extremely rare and so far only 16 cases have been reported.
We report a girl with hemorrhagic diathesis in a family of mild hemophilia B, who was suspected female hemophilia B. The patient was referred to KCMC at the age of 8 years because of easy bruising, which had first been noted at the age of 3 years. The family history showed that father and brother were mild hemophilia B, whose factor IX activity was 36% (4.5%) and 15%, and that her material paternal grandmother, uncle and her cousin had mild bleeding tendency. Mother had no bleeding tendency with normal activity of factor IX, but she was suspected a carrier of hemophilia B because her son was mild hemophilia B. The coagulation findings were as follows: Prothrombin time; 11.9sec., Kaolin activated PTT; 55-63.5sec., Factor IX activity; 4-16%, Factor II, V, VII, VIII, X, XI, XII activity and Fibrinogen level were normal.
Platelet function tests: Bleeding time; 3min. 30sec., Platelet retention test; 24%, and Platelet aggregation tests (ADP, Collagen and Ristocetin induced) were normal. The karyotype was found to be 46, XX, although sex chromatin was 4%.
If mother is a silent carrier, it is possible for this patient to be female hemophilia B. If mother is not a carrier, we must postulate hemophilia B transmitted by autosomal dominant pattern of inheritance or new disease with low activity of factor IX.
We will study on immunological determination of factor IX and more precise family history.

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The authors have reported the abnormalities of the blood coagulation and fibrinolysis in patients with renal diseases. Recently, antithrombin III (AT III) is emphasized as the most important of all the clot inhibitory factors.
The purpose of this study is to evaluate the significance of plasma AT III in patients with renal diseases.
Plasma AT III level, soluble fibrin monomer complex (SFMC) level and other factors relating to the blood coagulation and fibrinolysis were estimated in 31 patients with renal diseases and 14 normal control subjects. The patients consisted of 17 cases with nephrotic syndrome (9 in acute stage, 8 in latent stage), 4 with chronic glomerulonephritis and 10 with chronic renal failure. Plasma AT III was measured by single radial immunodiffusion method, and SFMC by Lipinski's method.
The mean levels of plasma AT III (mg/dl) and SFMC (O. D.) were 27.9mg/dl and 0.79 O. D. in controls, 17.6mg/dl and 0.39 O. D. in acute stage, 30.3mg/dl and 0.93 O. D. in latent stage, 30.0mg/dl and 0.80 O. D. in chronic glomerulonephritis, 24.3mg/dl and 1.19 O. D. in chronic renal failure.
Plasma AT III levels were lower in the acute stage of nephrotic syndrome and in chronic renal failure than those in controls. SFMC levels were higher in the acute stage of nephrotic syndrome and in chronic renal failure than those in controls. Though no correlation was found between plasma AT III and SFMC or plasma AT III and fibrinogen in controls, a negative correlation existed between plasma AT III and SFMC (r=-0.61, p<0.01) or plasma AT III and fibrinogen (r=-0.49, p<0.01) in renal diseases.
No correlation was found between plasma AT III and serum FDP (fibrin/fibrinogen degradation products) in renal diseases. However, plasma AT III levels were significantly lower in patients with abnormally high serum FDP level (≥8μg/ml) than those in patients with normal serum FDP (<8μg/ml) (p<0.01). In renal diseases no correlation was found between plasma AT III and the activity or the antigenicity of factor XIII.
Low plasma AT III with high SFMC has been often reported in the disseminated intravascular coagulation, which is apt to arise in renal diseases. And the author's results concerning coagulation and fibrinolysis can be regarded as pathophysiological features of the disseminated intravascular coagulation phenomenon in renal diseases.

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We have reported that the patients with Behcet's disease are generally in hypercoagulable state. The present paper concerns our investigation into the platelet survival time and the level of natural inhibitors of blood coagulation and fibrinolysis such as anti-thrombin III (AT-III), α₂-macroglobulin (α₂-MG), and α₁-antitrypsin (α₁-AT), in 23 patients with ocular manifestation of Behcet's disease. The platelet survival study was performed in 21 patients by the method of arachidonate-induced lipid peroxidation in aspirin loaded platelets and the inhibitors were measured by radial immunodiffusion technic. Seventeen of them were males and 6 were females, and their ages ranged from 18 to 52 years, 14 of whom had been treated with anticoagulant drug.
The level of AT-III (28.5±4.6mg/dl) was significantly lower than in normal subjects (p<0.001). On the other hand, that of α₁-AT (230.4±50.2mg/dl) was increased (p<0.001). That of α₂-MG (187.0±39.2mg/dl) was slightly decreased without significance. There was no significant difference in these inhibitor levels between the patients in remission and in attack of ocular inflammatory manifestations, and anticoagulant treatment of 6 months' duration had neither significantly affected these levels.
The platelet half-life (T1/2) in vivo was 3.96±0.44 days in 5 patients in attack, while it was 5.36±1.09 days in 16 patients in remission and 4.75±0.02 days in normal subjects. There was a significant shortening of platelet survival in attack stage (p<0.01), which may suggest that there is increased platelet consumption in exacerbated stage of Behcet's disease.
Although there were no significant changes in the inhibitor levels and platelet half-life after administration of anticoagulant (Warfarin), anticoagulant therapy seemed to be clinically effective in 10 out of 14 patients and the increased levels of fibrinogen, AHF procoagulant activity and AHF-like antigen were normalized on Warfarin treatment.
From these results the combination of oral anticoagulant and antiplatelet drugs may be more beneficial treatment for the patients with Behcet's disease.

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Changes in coagulation findings before and after development of acute myocardial infarction and stroke were determined in patients who admitted in Tokyo Metropolitan Geriatric Hospital. The patients were restricted to autopsied cases in whom coagulation analysis was carried out within 24 hours following the attack of acute myocardial infarction or apoplexy. These cases consisted of 28 of acute myocardial infarction, 28 of acute cerebral infarction (excluding cases of hemorrhagic infarction), 7 of acute cerebral hemorrhagic infarction and 10 of acute cerebral hemorrhage.
There were no changes in APTT and prothrombin time before and after the onset of the myocardial infarction or the stroke. A slight increase in plasma fibrinogen content was occasionally observed before the attack of the myocardial infarction or the apoplexy, however levels of fibrinogen in plasma tended to increase markedly following the attack. A slight increase in FDP was frequently observed in patients with poor prognosis. A remarkable prolongation of euglobulin lysis time was observed in cases of acute myocardial infarction. Euglobulin lysis times were prolonged in these cases preceding the development of the infarction. Generally, euglobulin lysis times in cases of acute cerebral hemorrhage or acute cerebral hemorrhagic infarction were slightly shorter than in cases of acute cerebral infarction (excluding hemorrhagic infarction). There was a trend toward a transient shortening of euglobulin lysis time after the attack of stroke, followed by a marked prolongation accompanied with a decrease in plasminogen content in plasma. A decrease in levels of antithrombin III in plasma was observed before and after the development of acute myocardial infarction. This most important inhibitor of blood coagulation also decreased slightly in cases of acute cerebral infarction, while concentration of α₂-macroglobulin increased transiently following the stroke.

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A 3-year-old girl underwent radical operation for common A-V canal with the aid of extracorporeal circulation. Hematological and serological tests prior to the operation gave normal results. After cessation of extracorporeal circulation which lasted for 276 minutes, hemodynamics was stable. However, plasma free hemoglobin increased up to 248mg/dl on the next day. Hemoglobinuria was continuously seen for a few days. From anemia, thrombocytopenia, hyperbilirubinemia and fragmentation of erythrocyte, DIC was suggested. Heparin and aprotinin were given intravenously. Besides these drugs human plasma haptoglobin which combines with free hemoglobin in plasma was given for prevention of acute renal failure. The patient was treated with 350ml of blood, 6000 unit of heparin, 1.8 million unit of aprotinin, and 400ml of haptoglobin by the fifth postoperative day. After administration of these drugs, hemoglobinuria disappeared and plasma free hemoglobin decreased. No renal complication occurred. In two weeks after operation, serological findings gave normal results. The patient returned home after three months.
Open-heart surgery with extracorporeal circulation is one of the causes of DIC and usually heparin is used for therapy. In our case, administration of aprotinin and haptoglobin had a good effect on improvement of this syndrome.

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Changes of coagulability, platelet aggregation, serum lipids and lipoperoxide before and after exercise were studied on 20 normal males. After Master's double two step exercise test, increase in coagulability, platelet aggregation and decrease in free fatty acid were resulted. Prothrombin time and activated partial thromboplastin time were shortened. Platelet aggregation induced by collagen was increased. Lipoperoxide had no changing but correlation with platelet aggregation. These increase in coagulability and platelet aggregation were postulated to result from sympathetic cathecolamine release.

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Thromboembolism is a major cause of morbidity and mortality in patients with a prosthetic heart valve in the follow-up period after surgery.
Warfarin, as one of potent oral anticoagulants, has been used generally to prevent this complication. On the other hand, concerning platelet kinetics in the thrombus formation, Dipyridamole is known to decrease platelet adhesiveness and aggregation, and to reduce the incidence of thromboembolism when it is administered in addition to an oral anticoagulant.
In this paper, effect of Dipyridamole on thromboembolic complication was evaluated from clinical point in connection with the relationship between the designs of Starr-Edwards ball valve and anticoagulations.
Total 186 patients with the non-cloth and cloth coverd Starr-Edwards ball valve including 108 mitral, 41 aortic, 24 combined mitral and aortic, and 13 combined mitral and tricuspid regions were assessed from thromboembolic complication. The incidence of thromboembolism was shown in the emboli per patient years from the average time of follow-up and the number of the emboli happened after counting the duration of anticoagulation, withdrawal or restart of anticoagulants in individual patient.
In these groups, the patients with Dipyridamole alone were evaluated, and 3 episodes of cerebral embolism were observed in 2 patients with the non-cloth covered mitral valve and 1 patient with the cloth covered aortic valve respectively. Four episodes of T. I. A. were also noted in each patient with the non-cloth and cloth covered mitral, cloth covered aortic and combined mitral and aortic valve. However, when Dipyridamole was given to the patients with Warfarin or Warfarin with Bucolome, only 3 episodes of T. I. A. were observed through these groups.
On the other hand, the results of thromboembolic complication in the patients treated with Warfarin or Warfarin with Bucolome showed much higher incidences of T. I. A. and cerebral embolism than those with the combination of Dipyridamole and Warfarin or Warfarin with Bucolome.
As a conclusion, 1) Dipyridamole should be given as an adjuvant in addition to an potent oral anticoagulant. But in patients with Starr-Edwards aortic valve. Dipyridamole alone can be allowed to use only after a definite time of strict anticoagulation. 2) In this series, the patients with the combination of Dipyridamole with Warfarin or Warfarin with Bucolome did not experience cerebral embolism. It may be advisable to supplement Dipyridamole to the patients with Warfarin alone. 3) Daily dosage of Dipyridamole is 300 to 400mg from theoretical reason, however, in this series, 200 to 250mg will also prevent serious thromboembolic complication.

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In the management of the new born, the so-called hyperviscosity syndrome is becoming problematic and in the high hematocrit group of over 65% circulatory obstruction and cyanose appear. We recognized symptoms of hypercoagulability namely an increase of fibrinogen and SFMC (soluble fibrin monomer complex) and followed the changes of coagulation fibrinolysis system after heparin therapy and partial exchange transfusion.
Method In 20 cases of newborn with a 65% or over hematocrit value and an increase in SFMC, 10 cases were treated with heparin and 10 cases were given partial exchange transfusion with fresh frozen plasma. Measurement on the following were made: (i) fibrinogen volume, (ii) SFMC, (iii) platelet aggregation (iv) PT. PTT, (v) TEG, (vi) antithrombin III, (vii) erythrocytoblast, (viii) ELT, , and (ix) FDP.
Results In the partial exchange transfusion group, in 3 cases which showed hematocrit values of over 75%, a remarkable increase in erythrocytoblast, and a strong hemolysis and a high anemia was seen. Partial exchange transfusion was done but a sudden shortening of ELT was seen during operation. Post operatively with a lowering of SFMC and hematocrit values improvement of clinical signs were seen.
In the heparin treated group, as a result of a 100-150 units/kg heparin injection a decrease in SFMC was seen and antithrombin III showed a 60% decrease of that prior to treatment.

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The dose and the method of administration of urokinase for patients with thrombo-embolic diseases has not yet been established.
Plasma fibrinolytic activity was evaluated by means of euglobulin lysis time and standard fibrin plate method with euglobulin fraction in the subjects. SK activated euglobulin lysis time and FDP were also determined.
Twenty-three subjects with either myocardial infarction or cerebral thromboembolic disease were divided into four groups. A dose of 3×10⁴ IU of urokinase was infused for two hours twice a day in group I and 6×10⁴ IU was given in the same way in group II. In groups III and IV, boluses of 2.4×10⁴ IU and 3.6×10⁴ IU respectively of urokinase were added prior to the same infusion as that given in group II.
Significant shortening of euglobulin lysis time was seen in the plasma of group IV patients, but not in the other groups, although standard fibrin plate method showed too few change to evaluate the fibrinolytic activity.
It is debatable and needs further study to decide which method is more preferable for the clinical evaluation of fibrinolytic activity.
Previous determination of whole plasmin content in plasma by the method of SK activated euglobulin lysis time was helpful for predicting the fibrinolytic activity after urokinase administration.
Since no significant changes were seen in the serum FDP level after infusion of urokinase in all 4 groups, determination of FDP was not an appropriate way of evaluating fibrinolytic activity in our experiment.

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Because the plasminogen value in the new born is extremely low, there is a possibility that administration of urokinase (UK) and streptokinase (SK) singly may not have a sufficient effect on the respiratory distress syndrome (RDS).
Thus, we administered urokinase and plasminogen to cases of RDS to improve the distress and to study the coagulation-fibrinolytic system in an attempt to clarify problematic points.
12 cases of RDS and 30 healthy new born infants were studied. Urokinase 300 units+plasminogen were given with 12 hours as a cycle and the following changes of the coagulation-fibrinolytic system, namely, (i) Fibrinogen, (ii) FDP, (iii) blood platelet count, (iv) plasminogen, (v) SFMC, (vi) α₂-macroglobulin, (vii) antithrombin III, (ix) platelet aggregation were studied at the same time. Blood gas analysis of actual pH, PCO₂, PO₂ was conducted.
Results The RDS infants, showed an increase in fibrinogen at the early stages and a remarkable increase in SFMC (healthy new born 3.1±0.8, diseased infants 4.1±1.4%). The diseased infants showed a especially low α₁-antitrypsin value.
In the treated group, the clinical reticular glanular pattern disappeared, a lowering of pCO₂ was recognized and a decrease to 3.5±0.6% in SFMC was seen.
While a slight lowering of platelet aggregation wit ADP was seen in RDS infants (Prior to treatment: healthy infants 25.0±4.5%, RDS 21.8±2.1%), no influence of urokinase was seen (after treatment: healthy infants 27.0±3.8%, RDS 21.0±2.4%)

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Platelet aggregation test was originated from Breddin which determined degree of platelet aggregation in vitro by mechanical stimuli such as contact of platelets on siliconized glass surface. We have carried out the PAT test using 45 subjects including 6 patients with ITP, 5 with uremia, 7 with hepatitis, 6 with carcinoma, 13 with other miscellaneous diseases and 9 healthy volunteers. In the result of PAT test, degrees of platelet aggregation were graded to I-V scales. At the same time, platelet spreadability (PSA) test following Breddin and ADP-induced platelet aggregation test were performed. We have modified PSA test to use a pair of clean plastic slide arranging parallel with one cm in distance in which the specimen was filled. Using this modified method, a reasonably high reproducibility was obtained. The degree of PSA was recorded as mean diameters of spreading platelets on plastic surface. In ADP-induced aggregation, 10μM ADP was added into platelet rich plasma using Evans aggregometer. In 8 out of 9 healthy subjects, grade III PAT was observed, whereas in 5 ITP patients, markedly lowered ability of platelets to aggregate grade I was observed. The PAT was found enhanced in the cases of uremia. Most patient with carcinoma displayed an enhanced PAT while it was depressed in the few remainders. Statistical analysis of the data revealed no significant interrelation between PAT and PSA. However, the combination with both the test, characteristic changes were noted in each of diseases. There was a statistical evidence of significant correlatation between PAT and ADP-induced platelet aggregation, with the equation of regression:
Y=1.72+0.28X, r=0.492, P=0.02
The above results would suggest that PAT and PSA test may be useful to determine some clinical conditions, when both the methods are used in a combination.

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Aggregation of isolated platelet membranes (PM) was observed under phase-contrastmicroscopy. ADP at concentration above 5mM induced an apparent aggregation of PM, while UDP, GDP or AMP at the same concentration was without effects. Light transmission in PM suspension increased by adding ADP (10mM), reaching maximum within 2 to 3 ninutes. This was unexpected observation since light transmission of intact platelet suspension usually decreases when the cells are aggregated. Decrease in light transmission of PM suspension induced by ADP occurred at any wave lengths between 350 and 800nm. There was no change in light transmission in the supernatant of suspension of PM once aggregated by ADP.
Light scattering intensity of PM suspension determined by Lasernepherometer promptly increased with the addition of ADP (10mM) and reached plateau within 2 to 3 minutes. This indicates that increase in particle size, probably aggregation of PM can proceed in the suspension.
Treatment of PM with neuraminidase (0.1u/ml), trypsin (20μg/ml) or LaCl₃ (1mM) inhibited both the aggregation and light transmission change of PM.
The addition of ADP (10mM) to PM suspension resulted in the reduction in electrophoretic mobility (0.301±0.057μ·cm/sec·V, control: 1.194±0.087) and the increase in microviscosity of PM (P value, 0.277, control: 0.255).
Changes in protein profiles in aggregated PM by ADP (5mM) were examined by SDS polyacrylamide gel disc electrophoresis. One protein band (M. W. 200, 000) on the gel disappeared and instead, there observed increased staining intensity at the top of the gel.
These results suggest that the following changes may be responsible for the aggregation of platelet membrane vesicles by ADP; 1) Reduction in surface negative charge of membranes, possibly via change in sialic acid, 2) Reduction in membrane fluidity, 3) Increase in membrane turbidity and 4) Polymerization of membrane protein with molecular weight of approximately 200, 000.

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Increasing attention has been paid to the surface electrical charge of platelets in relation to their functions. However there are several criticisms in methodological respects. In the present investigation the effect of neuraminidase on platelets was examined utilizing newly developed instruments of electrophoresis. These are Free-Flow Electrophoresis model Vap-5 (Bender and Hobein, West Germany) and Laser-Zee System 3000 (Pen-Kem Inc., U. S. A.). Rabbits were catheterized in the carotid artery and blood was obtained with EDTA as an anticoagulant. Platelet rich plasma was washed twice with Tris-HCl buffered saline and platelets were finally suspended in pH 5.3 acetate buffered saline. Washed platelets added with various concentrations of neuraminidase or control saline were incubated at 37°C for an hour. After incubation platelets were washed and suspended in 3mM triethanolamine acetate or 0.001M HEPES-NaOH, 0.15M NaCl buffer for electrophoresis with Free-Flow or Laser-Zee system respectively.
Results: Electrophoretic distribution of neuraminidase-treated platelets was shifted to the cathode side as compared with the control. The magnitude in shift of electrophoretic distribution depended upon neuraminidase concentration and there was a correlation between them (Fig. 1A, B). Relation between electrophoretic mobility and volume of platelets before and after the neuraminidase treatment was shown in Fig. 2A, B and 3. In general larger platelets were distributed in the anode side and smallers in the cathode side. With lower concentration of neuraminidase treatment this trend as shown by a regression line got greater. This result represents that the sensitivity of platelets to neuraminidase is various in accordance with the platelet volume and smaller platelets are more sensitive to neuraminidase. The platelet sensitivity to neuraminidase may have some role in platelet turn-over as smaller platelets have been believed as older ones.

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Platelets have been known to produce a labile but potent vasoactive substance as well as to release ADP, ATP and serotonin (5-HT) on stimulation. This substance reported as rabbit aorta contracting substance (RCS) by Piper and Vane² has been identified as mixture of prostaglandin (PG) endoperoxides (PGG₂, PGH₂) and thromboxane A₂.³ We have reported that; 1) intravascular platelet aggregation induced by ADP produced profound constriction and resultant injury of the pulmonary artery in rabbits; 2) intravenous injection of ADP elicited significantly attenuated cardiorespiratory disturbance and thrombus formation in spontaneously hypertensive rats (SHR) as compared with normal Wistar rats. In this paper in vitro responsiveness of the pulmonary artery of rabbit and the aorta of SHR and vitamine E deficient rat (VEDR) to RCS and 5-HT was investigated.
Materials and Methods: 1) The right or left pulmonary artery and thoracic aorta of 9 rabbits, 2) thoracic aorta of 7 pairs of 24-26 weeks old SHR and NWR and 6 pairs of 6 weeks old SHR and NWR, and 3) thoracic aorta of 3 pairs of 11 weeks old VEDR and NWR were excised promptly after death. Each vessels were cut spirally to produce 35-40×2.5-3.0mm strips and the strips were suspended in gassed Krebs medium at 37°C. RCS was generated by agitation of washed rabbit platelets with 5 units of thrombin or 6.25μg of sodium arachidonate for 40sec at 37°C.
Results: 1) The contractile response of pulmonary artery to RCS was about 50% of the maximum tension developed by 40mM K⁺ and 1μM 5-HT. The pulmonary artery response to RCS was as twice greater as the aorta and one to 5-HT was 1.5 times greater (Fig. 1A, B). 2) The contraction of 24-week SHR aorta by RCS was not different from NWR but one by 5-HT was significantly increased. However, the contraction of 6-week SHR aorta was not different from NWR (Fig. 2A, B). 3) Aorta responsiveness of VEDR was not different from NWR (Fig. 3).
Comment: Rabbit pulmonary artery contracted in great extent by RCS as well as by 5-HT. Vascular responsiveness of SHR was almost same to NWR or rather increased. So the mechanism of attenuated cardiorespiratory disturbance by intravascular platelet aggregation in SHR may not be on the vessels but on the platelet. Vascular responsiveness of VEDR will be further studied in connection with PGI₂.

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A man (66 years old) having plasma which showed an inhibitory activity of platelet aggregation induced by ADP, thrombin, epinephrine or collagen was found by Funahara, et al. Other abnormal findings observed were an increase of blood platelets number (10⁶/μl) and a loss of epinephrine induced aggregation activity of platelets. It was uncertain whether the inhibitor was newly formed as a result of thrombocythemia or an increase of the inhibitor production was accompanied by thrombocythemia. In order to clarify this question, an existence of the anti-platelet aggregating activity in plasma from healthy subjects was examined by the following experiments as a first step. The study of change of free platelets number in platelet suspension medium by Coulter-Counter ZBI showed that an activity to block platelet aggregation by ADP, thrombin or epinephrine existed in normal plasma. Though this activity was shown to reside in the outer fluid of bag by plasma dialysis, it was observed in void volume fraction when the outer fluid was passed through Sephadex G10 column, showing molecular weight of the inhibitor was small but somewhat lager than about 400 molecular weight. The activity was not affected by the treatment of plasma in boiling water for 20 minutes. Changes of the activity were not observed in the presence of aspirin, suggesting that the inhibiting site of the plasma inhibitor did not coincide with that of aspirin. The inhibitor showing very similar property to ours was reported by Mason, et al. (Am. J. Path. 74: 91a, 1974), but our inhibitor seems to be different from theirs in the behavior in Sephadex G10 column.

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Effects of colchicine and cytochalasin B on uptake of latex by platelets as well as on the ADP-induced aggregation of platelets were studied. Citrated platelet rich plasma (c-PRP) prepared from blood of healthy adults was used at room temperature within 1 hour after separation.
Experiment I: In one group, each test tube containing 0.8ml of c-PRP and 0.1ml of colchicine (10mg, 1mg, 0.1mg, 0.01mg or 0.001mg) was incubated at 37°C for 30min. In the other group, the mixture of 0.1ml of cytochalasin B (100μg, 50μg, 25μg, 12.5μg or 6.25μg) and 0.8ml of c-PRP was incubated at 37°C for 15min. After incubation, 0.1ml of latex was added to each sample in both groups, and then further incubation continued. The samples were obtained 30min. later.
The uptake was inhibited by both colchicine (>1mg/ml) and cytochalasin B (>50μg/ml). As seen in control platelets, the uptake was found when platelets were treated with low concentration of colchicine (<0.1mg/ml) or cytochalasin B (<25μg/ml). When the uptake had started, microfilaments assembled in the cytoplasm around the latex particles or in pseudopods of the platelets. In such occasion, the apperance of microfilaments and the uptake were dependent on the concentrator of the drugs. The platelets treated with colchicine (<1mg/ml) showed the uptake regardless of complete absence of intact microtubules. Platelets treated with cytochalasin B at higher concentration (>50μg/ml), however, did not take the particles in the presence of intact microtubules.
Experiment II: The effects of these drugs on the aggregation of platelets induced by ADP (final concentration: 0.02mM) were studied by using an aggregometer and the Chandler loops at 37°C for 15min. The drugs equally inhibited the aggregation at higher concentration (colchicine>1mg/ml, cytochalasin B>50μg/ml).
In conclusion, the microfilaments of platelets appear to play an important role in the uptake of latex without intact microtubules. As far as latex particles themselves are concerned in the “release reaction II”, they may participate in causing secondary aggregation of the platelet. The drugs, therefore, inhibit both the ADP-induced aggregation of platelets and the uptake as well.

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In order to understand the mechanism by which positively charged polylysine induces platelet release reaction in platelet rich plasma, the effects of neutral macromolecule, dextran, on platelets were studied in the presence or abscence of plasma. In the present study, three types of dextrans (DX 40, DX 250, DX 2000) were tested in platelet rich plasma (PRP) and in washed platelet suspension (WPS) for the ability to cause platelet release reaction and the effect of various inhibitors on the aggregation.
We observed that all three types of dextrans could induce platelet aggregation in PRP and WPS, but could not elicit the release reaction not only in WPS but also in PRP. Various inhibitors such as adenosine, aspirin, NEM, PGE₁ did not inhibit the platelet aggregation in both systems. Heparin, negatively charged acid mucopolysaccharide, also did not inhibit the dextran-induced platelet aggregation in both systems.
Earlier we investigated the effects of polylysine on platelets. The study demonstrated that in platelet rich plasma, polylysine elicited the release reaction of serotonin in contrast to dextran and polylysine formed more tightly packed platelet aggregate than dextran which was inhibited by negatively charged heparin. It was suggested that polylysine induced aggregation more effectively than dextran by reducing the negative surface charge and the possible mechanism by which polylysine elicited the release reaction was the formation of more tightly packed platelet aggregate than that by dextran in the presence of low calcium ion concentration in citrated platelet rich plasma.

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Superoxide dismutase (S. O. D.) is the enzyme to protect from destructive effect of superoxide (O₂^-) produced in many metabolic pathways related to oxygen. The purpose of this study was to investigate the possibility that S. O. D. may play an important role in the platelet function.
The cytoplasmic and mitochondrial S. O. D. has been investigated spectrophotometrically and gel electrophoretically in human platelets from eleven patients of chronic myelogenous leukemia (CML) and three patients of primary thrombocythemia (P. Th.). Neither deficiency nor abnormality of cytoplasmic and mitochondria. S. O. D. has been found electrophoretically in any case compared to normal platelets. However, the total activity from three of the CML patients and one of the P. Th. patients were above 3 unit/mg platelet protein (normal subjects: 2.11-2.70 unit/mg protein), suggesting the possibility either that more O₂^- production occurs in the platelets or that rather little O₂- production due to much O₂^- deprivation by the increased S. O. D.. The platelets from two CML patients and those from one P. Th. were separated into three different platelet density fractions. The cytoplasmic S. O. D. from two of the three patients was different in each three fractions, while that from normal subjects was almost equal, suggesting the qualitative heterogeniety in the platelets from these patients.
The S. O. D. activity of human platelets has been also investigated in several conditions, where much O₂^- generation might occur in platelets. Sodium fluoride (2mM), which increases platelet O₂^- production about 3 fold, had no effect on platelet S. O. D. The aggregated platelets induced by ADP (10^-5M), epinephrine (50μg/ml), ristocetin (1.5mg/ml) or collagen (1-20μg/ml) had no increase of S. O. D. activity compared to that from non aggregated platelets. X-ray irradiation (1, 000-20, 000R) had not induced its activity increase or decrease. These findings indicated the induction of platelet S. O. D. was not brought about under these conditions.

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The action mechanism of sulfinpyrazone (SUL) was examined with rabbit platelets by testing its influence on the aggregations induced by ADP and collagen, on platelet content of cyclic AMP, on the generation of labile aggregation-stimulating substance (LASS) by platelets, and on platelet aggregation induced by LASS. The generation of LASS was examined by incubating 20μg/ml of arachidonic acid (AA) with washed platelets in the presence or absence of SUL, and measuring platelet aggregation when the incubated mixture was added to platelet-rich plasma (PRP) containing 20μM indomethacin. SUL was administered in a dose of 200mg or 400mg to healthy adults, and plasma concentrations and aggregations induced by ADP, collagen, and epinephrine were measured before and 1, 2, 4, and 6 hours after administration.
At 3mM concentration, SUL exhibited little inhibitory effect on ADP-induced aggregation of rabbit platelets; but at concentrations of 1mM and above SUL inhibited collagen-induced aggregation, showing suppression of the release reaction. At 10mM, SUL elicited no change in platelet levels of cyclic AMP, and did not promote the PGE₁-induced increase in cyclic AMP. LASS-induced aggregation was slightly inhibited by 3mM SUL. LASS formation was markedly inhibited by 0.3mM SUL, and at higher concentrations the inhibition increased dose-dependently. After administration of SUL in a dose of 200mg, the plasma level of the drug reached a maximum (average 10.5μg/ml) after 1 hour and after 6 hours had fallen to one-third of that level (3.0μg). With a does of 400mg, the plasma levels reached a maximum (35.5μg/ml) after 2 hours and fell to approximately one-third of that level (12.7μg/ml) after 6 hours. The maximum inhibition of platelet aggregation was seen between 4 and 6 hours after administration, showing a discrepancy with the peak plasma level.
SUL inhibited the secondary but not the primary aggregation induced by ADP, only slightly inhibited collagen-induced aggregation, but strongly suppressed epinephrine-induced aggregation. SUL also inhibited the biosyntheses of PG endoperoxides and thromboxane A₂, and at high concetration showed a direct inhibitory effect against these mediators. The results can be interpreted in terms of the mechanism of inhibition of LASS generation, in view of the report that SUL, unlike aspirin, does not inhibit the activity of cyclo-oxygenase at concentrations that inhibit platelet aggregation.

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The present paper reports that EG-626 (7-ethoxycarbonyl-6, 8-dimethyl-4-hydroxymethyl-1(2H)-phthalazinone) (Nippon Kayaku Co., Tokyo) prevented aggregation of rabbit platelets in vitro by ADP, collagen and arachidonic acid, and potentiated remarkably an antiaggregatory action of prostacyclin (PGI₂).
Platelet rich plasma (PRP) was prepared by centrifugation (200xg, 15min) of rabbit blood, collected under ether anesthesia through the carotid artery into plastic tubes, containing 1/10 volume of 3.8% sodium citrate. Platelet poor plasma (PPP) was obtained by centrifugation (2, 000xg, 15min) of PRP. Aggregation of platelets (4-7×10⁵/μl) was recorded by the light transmission (PRP=0%, PPP=100%) using an aggregometer (RAM, Rikadenki Kogyo, Ltd., Tokyo).
EG-626 (100-300μM) inhibited does-dependently an aggregation by arachidonic acid (100-500μM), collagen (5-20μg/ml) or ADP (0.3-100μM). ID₅₀ values against the maximum doses of the aggregating agents were about 88, 110 and 300μM, respectively. The aggregation by arachidonic acid (400μM), collagen (20μg/ml) or ADP (30μM) was also inhibited by increasing doses (0.1-50nM) of PGI₂·Na (Ono Pharmaceutical Co., Osaka) and ID₅₀ values were about 22, 10 and 7nM, respectively.
Combination of EG-626 and PGI₂·Na in threshold doses considerably potentiated the inhibitory effects of each inhibitor on the aggregation by ADP or collagen. The aggregation by ADP (30μM) was inhibited by 72% of the control by 1.4nM PGI₂+50μM EG-626, 67% by 0.4nM PG₁+100μM EG-626 and 100% by 1.4nM PGI₂+100μM EG-626. Smaller dose (50μM) of EG-626, when added to PGI₂ (3nM), also completely prevented the aggregation by collagen (20μg/ml). The potentiating effect of EG-626 on PGI₂ may be due to an inhibition of phosphodiesterase, since PGI₂ is known to activate adenylate cyclase.

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Previously we have reported that an intravenous injection of ADP into normal Wistar rats (NWR) induced pulmonary thromboembolism, while this change was scarecely seen in spontaneously hypertensive rats (SHR) and that platelet count, volume and ADP·ATP contents were not different between SHR and NWR before the ADP injection.
In this paper, in vitro platelet function, platelet aggregation and release reaction, and in vivo ADP release reaction were analyzed. Similar study was also performed in young SHR in which hypertension had not been established to determine whether hypertension plays influence in platelet function in SHR.
Utilizing 30 mature SHRs (20-30 weeks of age) and 5 young SHRs (5-7 weeks) and same number of age matched NWRs, citrated blood samples were taken from carotid artery canulla for an analysis of platelet aggregability and ¹⁴C-serotonine (5-HT) release under pentobarbital anesthesia. ADP (1mg/kg) was injected into the jugular vein. After the injection, blood samples were taken at 3, 10 and 30min. for determination of platelet ADP and ATP contents.
Maximum intensity of ADP (100μM) induced platelet aggregation was significantly lower in SHR (46.5±11.0%, Mean±S. D.) than in NWR (59.3±5.6%) (p<0.05). ¹⁴C-5-HT release at 5min. after the addition of ADP (100μM) was significantly smaller in SHR (9.2±3.5%) than in NWR (21.5±5.1%) (p<0.05). Decrement in platelet ADP content after ADP injection was smaller in SHR (before: 2.1±0.3, at 30min.: 1.9±0.6μmoles/10¹¹pl.) as compared with NWR (before: 2.1±0.7, at 30min.: 1.1±0.3) (p<0.05). In young SHR, ADP-induced platelet aggregability showed decreased tendency as compared with age matched NWR but this difference was not statistically significant. ¹⁴C-5-HT release was significantly decreased in young SHR than NWR.
These results suggest that the lower incidence in pulmonary microthromboembolism in SHR might be explained by in vitro platelet hypoaggregability and decreased platelet release reaction. Decreased platelet reactivity seen in young SHR seems to represent inherited defence mechanism against thrombus formation in SHR.

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In pernicious anemia, mild or moderate thrombocytopenia is widely known, but qualitative platelet defects were scarcely reported.
We have reported two cases of pernicious anemia with abnormal platelet function.
Case 1: 68-year-old man who complained of palpitation and dyspnea on exertion. Laboratory data as follows: Hb 5.5g/dl, RBC 156×10⁴/μl, Ht 19%, Plts. 6.9×10⁴/μl, WBC 3, 000/μl, LDH 3, 508u, Vitamin B₁₂ 108pg/ml (N. 460-1, 140), positive Scilling test. Bone marrow aspirates disclosed hyperplasia of megaloblastic erythroid precursors and giant metamyelocytes. Platelet function as follows: Bleeding time (Duke method) 3′30″, Clot retraction 68%, PF-3 availability (TGT method) 11.0″, Platelet retention (Hellem II method) 32%, Platelet aggregation with ADP (1×10^-5M)20%, Epinephrine (1×10^-4M) 12.7% with lacked secondary aggregation, Collagen (10mcg/ml) 5.1%.
Case 2: 68-year-old woman who was admitted because of paresthesis of fingers and GI bleeding. Hb 4.7g/dl, RBC 118×10⁴/μl, Ht 15%, Plts. 7.3×10⁴/μl, WBC 3, 600/μl, direct bilirubin 2.9mg/dl, LDH 2, 152u, Vitamin B₁₂ 148pg/ml, Scilling test was positive. Megaloblasts and giant metamyelocytes were increased in bone marrow aspirates. Platelet function tests as follows: Bleeding time 1′, Clot retraction 60%, PF-3 availability 10.8″, Platelet retention 73%, Platelet aggregation with ADP 54%, Epinephrine 51%, Collagen 49%.
Impaired platelet function returned to normal after parenteral administration of Vitamin B₁₂ in both cases, and platelet aggregation was accelerated in one case.
Mechanisms of abnormal platelet function in pernicious anemia are still unknown, but we speculate that qualitative platelet defects may be associated with abnormal constituents of lipid in platelets of Vitamin B₁₂ deficiency.

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Platelet aggregation is an important phenomenon in the first step of thrombogenesis. When platelets aggregate, ADP and other platelet aggregating and vasoactive substances may release. We reported that an injection of ADP caused pulmonary thromboembolism in rats associated with transient effects including thrombocytopenia, apnea, arrhythmia, rise in central venous pressure and fall in arterial blood pressure, while these changes were less severe in SHRs. In this study, platelet aggregability and thrombogenic tendency in response to ADP were analysed in SHRs in comparison with those in the control rats. 30 Wistar rats (control) and 30 SHRs (20-30 weeks old) were used. Blood samples were taken from carotid artery cannula for an analysis of platelet count, platelet volume and platelet aggregation by ADP. ADP (1mg/kg) was injected rapidly into the jugular vein of anesthetized rats. After ADP injection, blood samples were taken at 30sec, 3, 10, 20 and 30min. ECG and respiration were recorded continuously. Before ADP injection, platelet count (control; 53.1, SHR; 53.0×10⁴/μl), the mode related platelet volume (control; 2.6, SHR; 2.7μ³) and platelet ADP content (control; 2.4, SHR; 2.7μmoles/10¹¹pl.) were not different between the two groups. Maximum intensity of ADP (10μM) aggregation was significantly lower in SHR (39.4±4.8%) than in the control (50.3±5.1%) (p<0.05). After the injection, platelet count decreased to 59.7% of preinjection level at 30sec (p<0.01) and returned to 93.8% at 3min in the control; 90.5% at 30sec (p<0.01) and 74.9% at 3min in SHR. Platelet ADP content decreased to 61% at 10min in the control, while little change in SHR. Arrhythmia and apnea observed after ADP injection were significantly longer continued in the control (p<0.01). Histological study revealed that pulmonary microvasculature was frequently filled with platelet thrombi in the control and seldom in SHR. The lower incidence in pulmonary microthromboembolism in SHR might be related to its in vitro hypoaggregability of platelet due to the insufficient release reaction. The lower response of platelet might be explained by exhaustion under hypertensive stress, though further investigation is required.

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The effect of ticlopidine (fig. 1), a new platelet aggregation inhibitor, was studied in 15 patients with Björk-Shiley prosthetic heart valves (6 aortic, 6 mitral, 3 aortic and mitral). The patients were 9 men and 6 women, aged 22 to 55 years old, who had had valve prostheses placed 2 to 43 months (mean 23.7 months) prior to entry into the study. They received 200mg of ticlopidine, divided into two oral doses, daily, and were followed up for more than 3 months. Platelet adhesiveness and aggregation were examined 1, 2, and 4 weeks, and 3 months after the treatment, and the values were compared to those obtained before the treatment. Platelet adhesiveness was determined by a modified technique of Salzman. A Sienco dual sample aggregation meter was used to measure changes of platelet aggregation through light transmission of citrated platelet-rich plasma to which adenosine diphosphate (ADP) had been added. The platelet count of the plasma sample was adjusted to a range of 250-350, 000/mm³. Student's paired t-test was used for statistical significance.
Platelet adhesiveness decreased significantly at 1 week, 2 weeks, and 3 months after the treatment, but no significant change was found at 4 weeks (fig. 2).
In every patient, platelet aggregation induced by 4μM ADP (final concentration) showed an obvious reduction one week after the treatment, and this effect continued for more than 3 months (fig. 3). A similar reduction in the aggregation caused by 20μM ADP (final concentration) was observed. In many cases, the aggregation caused by 20μM ADP, which showed the second phase of aggregation, changed after the treatment to show only the first phase of aggregation (fig. 4). Ticlopidine inhibited both the first and the second phase of ADP-induced aggregation. This dosage seemed to cause a reduction in platelet aggregation about 3 days after the start of the treatment.
We did not observe any interaction between ticlopidine and warfarin in this study.
None of the patients had clinical symptoms or side effects due to ticlopidine and no episodes of thromboembolism occurred during the examination period. Bleeding time increased slightly at 4 weeks after the treatment. No abnormal alteration was observed in the white blood cell counts, coagulation time, fibrinogen level, fasting blood sugar, serum electrolites, total protein, hepatic and renal function tests at 4 weeks and 3 months after the treatment.
Ticlopidine seems to be a useful platelet function suppressing drug for patients with prosthetic heart valves.

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Platelet abnormality in cirrhosis of the liver has been studied in our laboratory from biochemical, functional and morphological standpoint.
In this paper we studied the causes of shortening of platelet life-span in cirrhosis of the liver in relation to the splenomegaly, protein, sialic acid and glycoprotein contents of platelet and platelet plasma membrane.
[Result]
1) The volume of the spleen in cirrhosis of the liver measured from ^99mTcscinti gram increased to 648cm³ against 307cm³ of normal control.
There was no significant correlation between the platelet counts and the spleen volume (r=0.02).
2) The protein content of the platelet increased to 2.0mg/10⁹ plat. against 1.2mg/10⁹ plat. of normal control.
3) The sialic acid content of the platelet decreased to 7.2μg/mg prot. against 9.6μg/mg prot. of normal control. And there was a good correlation between the platelet count and the platelet sialic acid content (r=0.65).
4) On 7.5% SDS polyacrylamide gel electrophoresis (SDS-PAGE) of the solubilized platelet protein, in cirrhosis of the liver, a new band of MW 46, 000 appeared, while a band of MW 41, 000 almost disappeared.
5) On SDS-PAGE of the solubilized platelet plasma membrane protein, the band of MW 156, 000 disappeared in both cirrhosis of the liver and normal control pretreated with neuraminidase.
6) On 5.6% SDS-PAGE of the solubilized platelet plasma membrane glycoprotein, three major peaks (I, II, III) were observed in both normal and hepatic cirrhosis, but in the latter, peak III of MW 88, 000 was smaller than that of the former. The simillar change was observed in normal platelet plasma membrane glycoprotein pretreated with neuraminidase.
From these findings, it may be suggested that in cirrhosis of the liver, the reduced platelet sialic acid or sialoglycoprotein may promote platelet break-down in reti culoendothelial system and shorten platelet life-span.

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Spontaneous platelet aggregation (SPA) in primary thrombocythaemia was examined using Bryston aggregometer by stirring at 37°C for 10min. of citrated original PRP (platelet count 87-296×10⁴/μl) or diluted PRP (3×10⁵/μl). SPA was found in 7 out of 8 patients in original PRP but in none in diluted PRP. The critical platelet levels for positive SPA were around 10⁷/μl. SPA occurred according to all or none law; as far as SPA occur, the maximum aggregation rate did not change whereas the lag time was increased with the decrease in platelet concentration. No correlation was found between SPA and clinical symptoms or platelet lipoxygenase or cyclo-oxygenase levels. Among EDTA-2K, citrate and heparin, EDTA was the strongest to inhibit SPA. SPA was inhibited with EDTA over 0.075% at final concentration. SPA was also inhibited by dipyridamole (5×10^-5-5×10^-1mg/ml at f. c.), VCR (10^-4mg/ml) and adenosine (10^-5-5×10^-5M). Oral administration of aspirin (10mg/kg) inhibited SPA for 24 hours in all cases and did not 96 hours after the administration in all cases. This result was almost coincident with that of suppresion of aggregation induced by collagen (1μg/ml). The surface morphology of SPA was almost equal to that of irreversible aggregates with release reaction by collagen or ADP. Clinically the administration of small dose of aspirin once a day or two days may be sufficient to prevent SPA and aggregation by collagen.

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