916 Abstracts inhibit::>r capacity (BIC); the calculated regression line was used to predict lowering of BIC resulting from i. v. infusion of brinase in man. A total of 355 i. v. brinase infusions (single doses of 50- 200 mg) were given to 36 patients suffering from advanced peripheral arterial disease. Anticoagulant treatment (dicoumarolor heparin) was given concomitantly to 34/36 patients. BIC was determined by the azocollagen method before and after infusion of the enzyme. Good correlation was found between calculated lowering of BIC and determined post-infusion values. Individual dose require- ments could be calculated and free proteolytic activity by depletion of inhibitors was avoided. the Changes of ()( 1-antitrypsin and ()( 2 -macroglobulin induced by brinase infusion showed same trends as those recorded for BIC; there was no quantitative correlation. Fibrinogen was lowered by infusion of the enzyme, subnormal values were determined at 4 occasions. There was a moderate increase of fibrinogen degradation products and thrombin time was prolonged. Positive ethanol gelation t est following brinase infusion was a constant finding. Brinase induced slight lowering of Thrombotest values and prolongation of the activated part-ial thromboplastin time; dosage of anticoagulants was not influenced. Other coagulation tests showed insignificant chauges only. In two patients, not on anticoagulants, with a history of renal disease, transient renal failure occurred subsequent to brinase infusion. Laboratory data indicated intravascular coagulation as a possible cause. Bleeding complications were not observed.

Caroline 1vlcKillop, W. Edgar, G. D. Forbes and G. R. M. Prentice (University Department of Medicine, Royal Infirmary, Glasgow, Scotland): Formation of Soluble Complexes of Fibrinogen Relateancrod infusion were studied. Blood samples were obtained before treatment and after 6 and 24 hours ancrod infusion. Fibrinogen and its derivatives were precipitated with beta-alanine and separated by 6 per cent agarose gel filtration. A range of soluble complexes were demonstrated after 6 hours infusion. Polyacrylamide gel electrophoresis in SDS showed that the soluble complexes were largely composed of units with molecular weight similar to a minimally degraded early Fragment X. Polyacrylamide gel electrophoresis in SDS and mercaptoethanol showed a marked loss of intact alphachain in the soluble complexes when compared with the uncomplexed material, suggesting that the soluble complexes had undergone preferential fibrinolytic digestion. It is suggested t-hat, during ancrod therapy, FDP may be produced directly from soluble complexes rather than insoluble micro-thrombi as has been suggested previously.

G. Lawrence Slade and W. Abe Andes (United States Army Institute of Surgical Research, Fort Sam Houston, Texas 78234): Platelet Aggregation Following Defibrination with Ancrod. ( 431) A transient defect in platelet aggregation was observed following defibrination with Ancrod. Adult mongrel dogs were defibrinated using a slow intravenous infusion of Ancrod. Defibrination was maintained for ninety-six hours. Immediately following d e fibrination there was a complete ablation of the normal platelet response to thrombin or ADP with a gradual return toward normal aggregation over the ninety-six hour period. Fibrin This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited. d egradation product titers were highest at the time of maximum inhibition of platelet aggregation. The return to normal aggregation paralleled the fall in the fibrin degradation product titer.

M. Bielawiec, A. Perzanowslci and M. Mysliwiec (Haemoatology Clinic, Institute of Internal Medicine, Me dical School, Bialystok, Poland): Long Term Therapy of Patients Suffering from Occlusive Arterial Diseases with Combined Therapy with Phenformin and Stanozolol. (432) Phenformin (Dibotin-Winthrop) 100 mg daily and Stanozolol (Stromba-Winthrop) 7,5 mg daily w e re given for at least half a year and longer periods to 7 5 patients suffering Abstracts 917 from coronary heart disease and obliterative arteriosclerosis of the lower limbs. Clinical improvement was observed in above 80% of the pa.tients. Statisticaly si gnificant increase of fibrinolytic activity and decrease of platelet function, fibrinogen, cholesterol, lipids, and beta-lipoproteins levels as well as platelet - l eucocyte aggregates number in blood during the therapy were found. Readministration of phenformin and stanozolol after some time of cessation of the herapy caused the similar effects. The results of our investigations show that combined t herapy with phenformin and stanozolol may be of value in prophylaxis and treatment of thrombotic conditions and occlusive arterial diseases.

N . Crawford and A. G. Castle (Department of Biochemistry, Univereity of Birmingham, Birmingham B 15 2TT, U.K.): Isolation and Characterisation of Platelet "Tubulin", a Sub-Unit Protein of the Microtubules. ( 433) From studies with whole platelets using microtubule binding agents such as colchicine a nd vinblastine, it appears that the microtubule have a cytoskeletal function and may be involved in shape changes, intracellular transport and other forms of cell motile activity. A microtubule subunit protein, " tubulin", has been isolated from pig platelets and shown to have properties similar to tubulins from cilia , flagella and mammalian brains. The protein binds 3H-colchicine, has a sedimentation constant as the dimer of about 6 S and the monomer (MW ± 55,000) coelectrophoreses with rat, rabbit and guineapig brain tubulins, running in acrylamide gels as a closely spaced doublet similar to the Ci and ~ components of brain tubulin. Some higher molecular weight presumptive dynein com- ponents are also present. The polymer formed during a temperature dependent " in vitro" assembly procedure has been studied by electron microscopy. Long linear aggregates with side to side association can be seen, resembling the microtubule subfilamentous structures seen in whole platelet preparations.

D. C . B . 1 l1ills and D. E. Macfarlane (Specialized Cent er for Thrombosis Research, Temple University Hospital, P hiladelphia, Pa., 19140, U . S.A.): Cytochalasins: Effects ~n Platelet Aggregation, Adenylate Cyclase and 1\:Iembrane Transport Processes. (434). Cytochalasins A and B (CA and CB) are fungal metabolites that inhibit cellular motility,. platelet aggregation and clot retraction. CA differs from CB only in the substitution of a. carbonyl inCA for a hydroxy in CB. This confers on CA reactivity towards thiol groups .. We have studied t he effects of CA and CB on various membrane functions of intact. human platelets in citrated plasma. CA inhibited the uptake of adenosine and 5 HT without affecting intracellular nucleotide m e tabolism. CA (56 uM) but n ot CB increased t he rate of adenine uptake from 11.7 ±0.2 to 80.1 ± 4.4 pmolesfmin/108 platelets, while the affinity of the uptake m echanism was r educed (Km apparent increased from 0.12 ± 0.0 l to 1.31 ±0.12 u.M) . CA inhibited platelet aggregation induced by ADP at concentrations lower than those that blocked the shape change. CA (20-50 u.M) abolished the inhibition by ADP of cyclic AMP accumulation in platelets exposed to PGE1 and the phosphodies- terase inhibitor RA 233, whereas CB (170 uM) was inactive. This effect of CA was prevented but not revers ed by 1 mM cystein e. This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited. The observed action s of CA are similar to those of other thiol reagents but occurred at concentrations below t h at of plasma protein thiol, indicating a prefere ntial reactivity of CA towards the platelet membrane. (Supported by N. I. H. HL 14217.)

P . Massini (Theodor Kocher Institute, Univ ersity of Berne, Postfach 99 , CH-3000 Bern 9, Switzerland): The Role of Calcium Ions in the ActiYation of Blood Platelets. (435) The plasma membrane of the r e sting platelet is only slightly p e rmeable to Ca2+-ions. Stimulation of platelets with thrombin or other a ctivators induces an increased influx o£