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·THROMBOSIS
AND
ANTICOAGULANT THERAPY
Proceedings of a Symposium arranged by
Professor P. A. Owren, Professor R. B. Hunter,
and Dr W. Walker, and held in QUEEN'S
COLLEGE, DUNDEE, 29 and 30
September, and 1 October, 1960.
on
AMDA nt
Edited by W. Walker.
UNIVERSITY OF ST ANDREWS

·ACKNOWLEDGMENTS Financial help from the following made the Symposium possible the University Court of the University of St Andrews, J. R. Gergy, Ltd. Manchester, Nyegaard and Co, Oslo, and the privately endowed fund for Thrombosis Research in the Department of Pharmacology and Therapeutics, Queen's College

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IN GREAT BRITAIN BY
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·Members of the Symposium Professor G. H. BELL, Department of Physiology, Dr J. MARSHALL, The National Hospital, Queen Queen's College, Dundee, Square, London. Dr F. K. BELLER, Universitacts-Frauenklinik, Dr M, MATTER, Krankenanstalt Neumuenster, Tuebingen. Zurich. Dr ROSEMARY BIGGS, Blood Coagulation Dr J. MATZKE, Rigshospitalet, Copenhagen. Research Unit, Churchill Hospital, Oxford. Dr J. R. A MITCHELL, Department of Hæma- Dr ULF BLIX, Nyegaard & Co, Oslo. tology, The Radcliffe Infirmary, Oxford. Professor A. M. BOYD, Department of Surgery, Professor G. L. MONTGOMERY, Department of Royal Infirmary, Manchester. Pathology, The University, Edinburgh. Dr A. BARHAM CARTER, 114 Harley Street, Dr S NIEWIAROWSKI, Instytut Henatologul, London, Warsaw. Dr R. P. COOK, Department of Biochemistry, Professor P. A. OWREN, Rikshospitalet, Oslo. Queen's College, Dundee. Professor Sir GEORGE PICKERING, Department Professor R. E. COUPLAND, Department of of Medicine, Radcliffe Infirmary, Oxford Anatomy, Queen's College, Dundee, Dr J C. F. POOLE, Sir William Dunn School of Professor K. W. DONALD, Department of Pathology, Oxford. Medicine, University New Buildings, Edinburgh. Dr RL, RICHARDS, Southern General Hospital, Dr A. S. DOUGLAS, Department of Medicine, Glasgow. Royal Infirmary, Glasgow Dr J. F. RILEY, Department of Radiotherapy, Professor D. M. DOUGLAS, Department of Surgery, Queen's College, Dundee. Royal Infirmary, Dundee, Professor H. W. FULLERTON, Department of Mr N G. ROTHNIE, St Thomas's Hospital Medical School, London, Medicine, Royal Infirmary, Aberdeen Dr A. RAE GILCHRIST, Royal Infirmary, Dr M. SAMAMA, Hopital Broussais, Paris. Edinburgh Dr C. I SCHWARTZ, Department of Medicine, Dr R. H. GIRDWOOD, Department of Medicine, Radcliffe Infirmary, Oxford. University New Buildings, Edinburgh Dr A. A. SHARP, Department of Pathology, Dr H. B. GOODALL, Department of Pathology, Radcliffe Infirmary, Oxford Queen's College, Dundee. Dr D) A. SHAW, The National Hospital, Queen Dr R H GOSLING, Geigy Pharmaceutical Square, London, Company, Ltd, Roundthorne Estate, Wythen Mr G SMITH, Department of Surgery, Western shawe, Manchester, Infirmary, Glasgow. Dr H HARTERT, Medizinische Universitaetsklinik, Heidelberg Dr AS. TODD, Department of Pathology, Queen's College, Dundee. Professor 1. GW HILL, Department of Medicine, Queen's College, Dundee. Professor W J. TULLOCH, Department of Bacteriology, Queen's College, Dundee. Professor R B. HUNTER, Department of Pharmacology and Therapeutics, Queen's College, De A, C. TURNBULL, Department of Midwifery Dundee and Gynæcology, Queen's College, Dundee. Professor E KOWALSKI, Instytut Hematologil, Dr M VERSTRAETE, Service de Medecine Warsaw. Interne, Voes des Capucins 57, Louvain Dr M J LARRJEU, Hôpital Saint-Lous, Paris. Professor J WALKER, Department of Midwifery Dr D. LEAK, Royal Infirmary, Edinburgh. and Gynäcology, Queen's College, Dundee. Sir JAMES LEARMONTH, Broughton, Biggar, De W. WALKER, Department of Pharmacology Lanarkshire, and Therapeutics, Qucen's College, Dundee Professor A. C. LENDRUM, Department of Dr HAMISH WATSON, Department of Medicine, Pathology, Queen's College, Dundee Queen's College, Dundee Dr K. G. LOVE, Department of Medicine, Queen's Dr E. WINDSOR, Riker Laboratories, Northridge, College, Dundee California. Dr R. G MACFARLANE, Blood Coagulation Dr W ZOLLINGER, Krankenanstalt Neumuenster, Research Unit, Churchill Hospital, Oxford. Zurich, ·FOREWORD The conference on thromlinsis and anticoagulant therapy sponsored by the University of St Andrews was, to my mind, an excellent example of the value of concentration in discussion. The xuloject was timely. Though representative of a number of European countries and of the United States, the total number of delegates was small enougl to allow them lo he housed under one roof. The proceedings were conducted in one language. Tlie university and the civic autliorities were to limpitable that it was clear that neither had overlooked the original meaning of the word rymposium. Very properly, the conference began loy attempting to define the phenomena to lie di cusacel in terms of pathology, and it emergeel that information on these matters was by no means complete. Ilowever, a new method of producing ariiſicial thrombi anil a new technique for identifying plasminogen activator in the tissues lield promise of prosiling more information about ibe factors which preserve the fluiility of blood. That ilierapeutic efforts to ensure fluidity of blood were desirable in a variety of clinical conditions wat generally agreed and, on the whole, Thrombole i was accepted at the most generally useful metliod of controlling the administration of anticoagulant drugs. The ideal, a test to dieclose both tendency to tlıromlouis and tendency to bleeding, seemell as elusive as ever. Although the precise way in which anticoagulant druga produce their good results when given for long periodle remained unsettled, the cloice of ille types of patient to receive life-long anticoagulant therapy was not a matter of con. troversy, nor were priorities within these type. That such therapy might take a fulstantial contribution to any national economy was generally agreed and the administrative machinery for securing thie proseil a fascinating topic. The relatively recent use of extracorporeal circulations provided a new field for the use of anticoagulants, which was duly explored. And, finally, it was olivious that the possibility of producing new synthetic anticoagulants was by no meang exhausted. One of the advantages of a symposiu is that it is not burdened by any obligation to reacli final conclusions, or to make recommendations. What a symposium can do is to accomplish - strictly for the time being certain objectives; to confirm good methods of treatnient and good techniques in research; to agree to abandon those which have been unfriniful; to choose those wluch are promising enough to be retained after improveinent; and, above all, to suggest new avenues of approach Perusal of these proceedings will make it clear that, in the pursuit of these objectives, the symposium succeeded unconimonly well. James Leamonth.

·CONTENTS Page Current concepts of blood coagulation in relation to vascular disease. THROMBOSIS, BLOOD COAGULATION AND ATHEROMA A Critical Appraisal J. R. A. Mitchell 9 TIIROMBOSIS AND HEMORRHAGIC DIATHESIS - M. Matter 13 FACTORS INFLUENCING THE FORNIATION OF ARTIFICIAL THROMBI J. C. F. Poole 17 THE RECALCIFIED PLASMA CLOTTING TIME AS A MEASURE OF BLOOD COAGULABILITY H. W. Fullerton and G. A. McDonald 18 21 STUDIES ON FIBRINOLYSIS. Some Inter-relationships Between Clotting and Fibrinolysis E. Kowalski and S. Niewiarowski THE TISSUE ACTIVATOR OF PLASMINOGEN AND TIROMBOSIS A.S. Todd 25 DISCUSSION 32 The organisation and laboratory control of anticoagulant therapy. THE ORGANISATION OF LARGE SCALE LIFE-LONG ANTICOAGULANT THERAPY P. A. Owren 35 THE LABORATORY DETECTION OF THROMBOSING AND OF BLEEDING TENDENCIES Hellmut Hartert 40 42 COMPARATIVE STUDY OF CONTROL METHODS FOR ANTICOAGULANT THERAPY W. Zollinger THE CHOICE OF TEST FOR CONTROL J. M. Matthews, R. Shulman, A. Tuxford, W. Walker 46 DISCUSSION 50 The use of anticoagulants in myocardial infarction, in peripheral and cerebral vasculor disease, and in prophylaxis. ANTICOAGULANT THERAPY IN MYOCARDIAL INFARCTION R, B, Hunter 53 R. L. Richards 55 ANTICOAGULANT THERAPY IN ACUTE JYOCARDIAL INFARCTIOX. Clinico-Pathological Study ANTICOAGULANT THERAPY IN PERIPHERAL VASCULAR DISEASE George Smith 57 ·CONTENTS Pase PROPIIYLAXIS BY JSTICOAGULIXTS (COUMADIN) IN SURGICAL DISORDERS J.J. Larriçu, J. Samama. E. Terrier, Cl. Frileux 59 CO PROPITYLIXIS BY ANTICOAGULAYTS AFTER GYXECOLOGICAL OPERATIONS. Selection of Patients 1. C. Turnbull THE ROLE OF ANTICOAGULANT THERAPY IX CEREBRAL IXFIRCTION A. Barham Carter 02 ANTICOAGULANT THERAPY IX CEREBROVASCULAR DISE.ISE John Marshall and D. A. Shaw G3 DISCUSSION 08 Heparinoids and their place in therapy. STUDIES ON THE MECIIANISM OF TIIE ANTICOAGULANT ACTION OF JIEMARIX S. Sicwarowski and Z. Wegrzynowicz A NEW HEPARIXOID ANTICOAGULINT. F. K. Beller 72 75 A NEW ORALLY EFFECTIVE HEPARINOID: Inticoagulant and Intilipemic activity in animals and Jan After Oral and Parenteral Administration E. Windsor and G. E. Cronhcim . 78 DISCUSSION 85 Hæmatological complications of the extracorporeal circulation. THE HEMATOLOGY OF EXTRACORPOREAL CIRCULATION WITH AND WITHOUT IIYPOTHERMIA A. A. Sharp, B. Excell, E. Salzman, O. Thorup 88 ABXORMAL HÆMORRHAGE AFTER PERFUSION: The Importance of Unneutralised Heparin and Fibrinogen Depletion N, G, Rothnie and J. B. Kinmonth 94 COAGULABILITY DURING EXTRACORPOREAL CIRCULATION WITH PARTICULAR REFERENCE TO ANTIILEMOPHILIC FACTOR B (FACTOR IX.) J. Matzke, R. Smith Jensen and I. II. Rygg 99 BLOOD COAGULATION PROBLEMS DURING EXTRACORPOREAL CIRCULATION IN MAN- 1. Verstraete, A. Amery, C. Vermylen 100 DISCUSSION 103

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·Thrombosis, Blood Coagulation And Atheroma A CRITICAL APPRAISAL 3. R. A. MITCHELL -- M.R.C. Clinical Research Fellow, Department of Hematology, Radciffe Infirmary, Oxford Anticoagulants are being used with eves. offer an explanation based on tho assumptions. Firstly, that thrombosis 5 Dira-vascular clotting, and secondly, that the in vivo action of anticoagulants is the same as their in vitro action. Unfortunately, and studied the staining properties of the fibrin the two premises on which this train of thought formed. They found that unless they added 1 g. depends are not vand. Whatever thrombosis is, it is not simple intravascular cloting, and naked-eye examination of the end products of the the processes convices us of thes. An intravascular cloi is dark red, shiny, soft Amewn lo larr him firm observations), and until this has been resolved, the confidens identification of fibrın by the fluorescent antibody technique must be accepted with caution. fibrio coagulum, was a relatively late event, and chemical environment will determine the staining reaction rather than the mere presence of the blood component in question. There may be a place for the fluorescent antibody technique in plarelet ideouf. cation, and we are studying dus problem. Finally, I shalt discuss briefly the relationship L. predisposes to coronary occlusion by thrombus, resulting in myocardial infarction (Fig. 4). However, cach of these assumpungs can be assailed. The relationship between infarction and pen? enough calcium to deal with the transfused bipod and yet ir does not clot). Laboratory tests of blood coagulation may not, therefore, give any indication of the state of affairs in the living vessesls survey of atheroma, occlusion and ischemic heart disease and find that in our series, in recent infarctions 9

·Figure 1
Figure 2
10

·Figure 1-Section through clot. showing even distribution of red cells and fine fibrin strands. Mallory P.T.AH X 200. Figure 2-Section through experimental rabbit pulmonary thrombus showing fibrin lamina, white cells, amorphous placelet debris, and paucity of red cells Mallory P.T.AH x 200. Figure 3-Section through mura! thrombus showing platelet aggregates surrounded by white cells. H&E, 200 Figure 3 MYOCARDIAL CORONARY CORONARY GENERAL INFARCTION THROMBOSIS ATHEROMA ATHEROMA CLASSICAL DUGUID Figure 4 - Diagram showing supposed inter-relationship be tween cardiac infarction and vessel disease, and (dotted lines) the false analogy between human lipid findings and the results of rabbit experiments RABBIT ATHEROMA HIGH BLOOD RABBIT CHOLESTEROU POISONING CHOLESTEROL Figure 4 ·- 40 with definite cardiac necrosis, occlusion is found in over 80%, whereas in patients who have died suddenly honni.... luallis. The second equation was questioned by Morris (1951) who showed that, despite the rising tide of cardiac infarction, the amount of coronary atheroma had not increased over the years in post-mortem examinations at the London Hospital, whereas the frequency of coronary occlusion had. Robertson (1959) found that in racial groups with widely differing infarct_rates such as Jamaicans and the inhabitants of New Orleans the coronary atheroma pattern may be very similar, whereas the number creasing frequency of cardiac infarction than we st botic wall & is sug Regis mona of in virtua incren this ti 1 -how larini in low and huph informata -- uki years ui une population group, or in one racial group as compared with another, then it should leave behind in the patients who recover an increased amount of atheroma). If thrombosis is the critical factor in the in- 'The three equations shown in Figure 4 are still incomplete and even if the inter-relationships do exist, is the direction of the "reaction a forward one from atheroma to thrombosis or is it Dugud's "back reaction” from thrombosis to atheroma ? Figure 5 DEATHS FROM PULMONARY 1200 EMBOLISM Tota) 1000 REFERENCES Bizzozero, J (1882) Virchow's Arch Path Anat, 90, 261 Branwood, AW and Montgomery, G L (1956) Scot Med J , 1, 367 Chandler, A B (1958) Lab Invest, 7, ITO Dugud, J B (1954) Lancet, 1, 891 Gitlin, D and Craig, J M (1957) Am J Path , 33, 367 Gude, W D, Upton, A C and Odell, TT (1956) Lab Invest, 5. 349 Jones, T W (1850) Guy's Hosp Rep. and Series, 7, 1 Morris, J N (1951) Lancet, 1, 69 Poole, J CF (1959) Quart J Cup Physiol. 44. 377 Robertson, WB (1959) Lancet. 1, 444 WHO fechu Rer Ser (1938), 143 Zahn, FW (1875) Virchow's Arch Path Anat , 62, 81 800 Females 600 DEATHS PER YEAR Maies 400 200 6th Revision 7th 1940 1950 1960 Figure 5-Annual death rate from pulmonary embolism and pulmonary in/arction (Registrar. General's Statistical Review of England and Wales 1940-1958) YEAR 12 ·Thrombosis And Haemorrhagic Diathesis M. MATTER -Krankenanstalt Neumünster, Zurich bosis the values of r and k return to normal or cien show decreased coagulability (15). Even this method does not detect a tendency to or an existing thrombosts with certainty. As changes in the concentration of coagulation factors and variations of rand k in .... -L-- Thrombosis and honorrhagic diathesis are considered to be opposite processes. Thrombosis s certainly rare in thronbocytopernia and severe coagulopathics. This is one reason why anticoagulants are used in thrombosis. Another is the belief that thrombosis is the consequence of hypercoagulability, As thrombosis and its prevention are the main subiccts of this conference, I shall concentrate on thrombogenesis, and show that study of the coagulation mechanish and of coagulation defects has greatly contributed to the understanding of this process. I shall not discuss the possibly important role the fibrinolytic system plays in regulating physiological and pathological thrombus formation. Stress and the administration of ACTH and cortisone lead to an imbalance of this mechanism which may be of great importance in the pathogenesis of venous and arterial thrombosis. In severe hæmophula, a marked deficiency of a coagulation factor leads to a prolonged clotting ume and delayed and incomplete thrombus formation. It is casy to measure such dcfects with simple methods 'In thrombosis one would by contrast decreased coagulability. If great amounts of tissue juice enter the circulation as in premature separation of the placenta, fibrinogen, prothrombin, factor V and platelets may be so reduced by intravascular coagulation that a severe hemorrhagic disthesis follows. fig 2 shows the TEG of such a patient (A): no thrombus formation occurs ; B is the normal control, C a mixture of the patient's and normal blood. If small amounts of thomboplastin enter the circulation massive thrombosis docs not develop, but the following changes are seen in a sequence of formation is a more dynamic process and the coagulation factors may rapidly change. Only frequent measure.nents of blood sa.nples may show the sequence of events Stalc iu). Ile Sallic langes are nolcd tctore, during and two days after normal delivery. They may be the result of the entrance of small amounts of tissue juice from the placenta into the circulation (6). The same changes are often found during thronhat chatne ACARE -Lthrombosis (3). As arterial thrombosis is usually I critaneler But the whole blood clotting time marital also, TL ALL measi Fig. speec At the site of an endothelial arterial lesion, hatched area. the first visible change supposedly consists of the Hypercoagulative states show a shortening of agglutination of platelets (Fig. 4). This may take r and k. With this sensitive method it has been only a few seconds and 18 independent nf the .. possible to measure increased coagulability in many mere af vann ----L---- 13

·Figure
Hypercoagulability
normal range
Hypocoagulability
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Figure 3
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·Extrinsic System Intrinsic System Tissue Thromboplastin Platelet Factor III CO" FV Platelets - Thrombus E VII FIX FX PTA coagulation- Thrombus FX (Hogeman-F.) Active Thromboplostin Active Thromboplastin Fibrin Prothrombin >Thrombin Thrombin Prothrombin Figure 4 Fibrinogen Figure 5 PLASMA COAGULATION SERUM FZI, X, (PTA) 100% 1 ACTIVITY 50% FX, Hageman-F 0% FII, V, W TIME 15 ·(14). Platelet factor 3 reacts with Ca, the antihumophilic factors, PTA, Hageman factor and factor X. activated when the blood is exposed to wettable surfaces such as glass (12, 13). Their activation at the site of the endothelial lesion may be one of the first changes initiating coagulation in thrombosis. The experiments of Wessler favour the hypothesis that the production of serum is very important in thro.bogenesis. It agrees with the clinical observation that a major thrombotic process, particularly venous, usually follows minor thrombosis due to traumna including surgery and delivery. During coagulation fibrinogen, prothrombin, factor V and factor VIII are consumed. The activity of Ca, factor VII, factor X and PTA remains m. 1 contains thus a thrombotic accelerator, which is present also in the sera of patients with the following coagulopathies : factor V, factor VII, factor VIII and factor X deficiency (19). The serum of factor IX and Hageman factor deficiency does not contain this accelerator. The activity is decreased in PTA deficiency and in patients treated with dicoumarol for at least one week. Factors IX and Hageman, inactive in plasma, become thromboplastin in the intima Wessler found heparin efficient in preventing experimental thrombosis whereas dicoumarol and its derivatives showed decreased activity in the serum only after a week of treatment (20). This effect probably is due to factor IX deficiency, which only develops after this period of therapy. We are not yet able to produce isolated Hagerman factor deficiency for therapeutic purposes It would never lead to hæmorrhage, and, therefore, would be the ideal antithrombotic treatment. REFERENCES 1-Baer, r, Geiger, M, Koller, F (1957) Acta Hämatol, II-Varchal, C, Leroux, XL, Duhamel, G, Samama, V 18. 33 (1958) Le Sang, 29, 666 2--Delta Santa R, Faller, R (1951) Schweiz Mecl Il schr, 12---Rapaport, SI, Ass, K, Owren, PA (1954) J Clin Sr. 1242 Insest, 34, 9 --Deutsch, E (1954) I International Conference on 13-Ratnoft, OD, Rosenblum, J.M (1958) Amer J Med, Thrombosis and Embolism, Basel, p 433 25, 260 4-Fcassly, R (1954) I Tuternational Conference on Throm 14--Rosham, J (1960) Schweiz Ved IV'schr, 90, 917 bosis and Lmbolism, Basel, p 391 15-Runge, H, Hartert, H (1992) (1953) hlin Wschr. 5-llartert, 11 (1953) Munch Bed Ischr, 95, 1108 30, 395, 31, 32 6--Jamun, 33, Leroux, ME, J.ctesszer, A (1958) Gym 16-Soulier, J P. Le Bolloch, A G (1950) Rev Hematot, Obst, 57, 81 3. 148 7--Ingram, (, JC. 1188, R (1953) J Clin Path, 1, 4 17—Wessler, S (1955) J Clin Invest, 34, 647 8-Koller, 1. Loeliger, Duckert, !, Hu Wang, II (1952) 18--Wessler, S, Ballon, J D, Katz, J K (1957) N Engld Dtsch Ved ihr, 77, 528 J Sled, 256, 1223 0-1 croux, M L (1957) These Vled, Paris 19-Wessler, S, Reimer, S M (1960) J Clin Invest, 39. 262 ro-Loeliger, A (1900) Personal cominunication 20-lessler, S (1960) l'ersonal comunication ·Factors Influencing The Formation Of Artificial Thrombi J. C. F. POOLE-Sir William Dunn School of Pathology. University of Oxford 72- -A.-L.11 in the body, in short, a method for producing TILAA blood when poured into a glass tube or some such container, while thrombosis is taken to mean the If Chandler's procedure is modified slightly by deposition on the inside of a blood sessel wall of sticking the ends of the tube together to make a solid material from the blood Sone people use the smooth joint it is possible to form structures which words indiscriminately. But the distinction is in- resemble natural thrombi even more closely (Poole, portant because the microscopical appearances of 1959). In such an apparatus the blood does not a thrombus and a clot are different, clot at all in the ordinary sense. Instead, a small A thrornbus has an elaborate structure made solid body forms just behind the advancing cdge up of two different kinds of material. One of the column of blood and floats round in the blood for an indefinite period. A section through one of these bodies shows a structure simular to that of certain kinds of thrombus with a white head and a red tail. masses appear as structureless areas. That this Th. cena--.,..- consists of platelets is not immediately obvious but in fact there has never been much doubt about it. Several workers in the cightecn-eighties observed the formation of thrombi in living vessels ; they could see platelets being deposited and when they cut sections through such regions areas of a.morphous head. Then, after an interval, the red tail forms material were seen. The matter was settled by suddenly and the structure is fully formed. Levenc & Levenc (1957), who studied the structure of human thrombi with the clectron microscope, and confirmed that in these areas, where no structure is shown in ordinary sections, platelets very closely packed together can be made out. This basic structure is con:non to all naturally occurring thrornbi, but there are many variations. The proportion of the platelet and white cell material to the red cell and fibrin material varies greatly. The platelet material may comprise nearly the whole structure or only a small part. In a common type it is concentrated in a small area called the white head attached to the vessel wall while the greater part consists of a red tail of red cells and The effect of free fatty acids was investigated fibrin, extending downstream from the white head. because of the known effect of fatty acids on the The platelet and leucocyte masses may be loosely clotting of plasma (Poole, 1955). Long chain satupacked branching coralline structures or form streaks rated fatty acids in low concentration shorten the or Jayers. clotting ume of plasma, while short chain saturated The structure of a blood clot is quite different. fatty acids and long chain unsaturated fatty acids The formed elements of the blood are distributed do not. When these observations were made nobody knew whether free fatty acids occurred in human blood Robinson, Harris, Poole & Jeffres (1955) showed that the usual range of free fatty acid in the plasma of healthy young men was about 0.3 to about Rara 2 mill.quivalents per litre. Stearic acid added to plasma in that concentration gives a maximal effcct on the clotting time and some effect with smaller A PARA It is obvious that a new technical approach 1s needed to provide a bridge between studies of blood clotting in the test tube and of thrombosis 17 ·---.. concentrations in which the active fatty acids pro acid concentration in the blood, but there is much duce a shortening of the thrombus formation time controversy about whether or not fatty meals produce are again small and within the physiological fluctu- hypercoagulability. Conflicting claims have been ations. Of the straight-chain saturated fatty acids, made and it is difficult at present to hold any clear views on the matter. The effect of adrenaline injection and stress is less obscure. Cannon & Gray (1914) observed palmitic acid, C 16, there is a marked shortening shortened whole blood clotting time in cats after and with longer chain acids from stearic acid, C 18, injections of adrenaline. Connor confirmed this . . . . . - onwards there is an even more marked effect. The long chain fatty acids tested included one of the unnatural series, the odd-number acids, C 23, and this behaved much like its neighbours in the series. The four long chain unsaturated fatty acids tested are inactive or nearly so. They are oleic acid, elaidic acid, ricinoleic acid, and arachidonic acid. Of the active acıds the most abundant in nature 18 stearic acid and the investigation has started of treated rats of the same strain in exactly the same the effect of simple modifications of the molecule. Substitution of one methyl group reduces and of way and found an increase in the free fatty acid two abolishes activity. A hydroxyl group makes little content of their blood, so here again in vivo hypercoagulability and increased free fatty acids go if any difference, together. When artificial thrombi are formed in the Further speculation is unprofitable as the data presence of an active fatty acid they are not only are as yet incomplete. But when a system which formed more rapidly but are also larger. Whether in produces objects very like natural thrombi is inlife the size of a thrombus or the speed of its formation fluenced in this way by free fatty acids in concenis the more important, in this experimental situation trations which occur in the circulating blood, then the answer is the same one must take seriously the possibility that some such In vivo, a fatty meal leads to a rise of free fatty mechanism might operate in vivo. REFERENCES Cannon, W B, and Gray, I (1914) Amer J Phs sol, 34.232 Dole, V P (1956) J Clin. Invest.. 35, 150 Chandler, A B (1958) Lab Invest, 7, ITO Levene, M, and Levene, C. I (1957) J. Clin Path, ro, 200. Connor, W [ (unpublished observations) Hallov, S, and Witt, P (1960) Fed Proc, 19, 229 Connor, w E, and Poole, JCr (1961) Quart J Exp Poole, J C F (1955) Brit J Exp. Path , 36, 248. Phv 101, 46, I Poole, J.Cr (1959) Quart J Exp Phusio!. 44, 377 De Long, r, Uhlev, Il N, and Friedman, M (1959) Amer Robinson, D S, Harris, P . Poole, J C, F, and Jeffries, Phicol, 195, 429 G. HI (1955) Biochem J. 60, XITVI1 The Recalcified Plasma Clotting Time As A Measure Of Blood Coagulability H. W. FULLERTON and GA. MCDONALD- Department of Medicine, University of Aberdeen A further difficulty is that the process of coagu- One approach to the problem of the patho- .Lemaris has been to vitro by a wide he large amount these lines, the DU 111011 is Sulu Hu... fact, the results --- IVL HAVE In recent years we have carried out several studies using the recalcificd plasma clotting (R.P.C.) time as a measure of coagulability, and our first 18 ·objective was to standardise the technique and to Observations in two groups of a nbulant subjects find out whether or not it gave reproducible results. have shown distinct differences from the previous Details of the technique have been given in a previous two groups of in patients. communication (McDonald and Fullerton, 1958 a) and all the steps have always been undertaken in exactly the same way by one of us (G.A.M.). We Group C. consider this to be of great importance of accurate results are to be achieved and we would stress that This comprised 10, our patients who had had none of the work on which this communication is based has been delegated to technicians. higher (mean 126 7 seconds) than in either group of greatest difference between the two arms was only in patients, and second, that the reduction 31 hours 19 3 5 seconds. after the high-fat meal is less (mean 16.7 seconds). • shortcns the I which gives Group D. arms in 14 patients, the venepuncture being satisfactory in one This consisted of 23 students who reached arm, while in the other it was unsatisfactory in that hospital by walking short distances and by using Lissue trauma was produced inadvertently or deliber. public or private transport. After the high-fat ately. The readings in the samples obtained by breakfast they attended lectures and clinical inunsatisfactory venepuncture were consistently much struction, walking a few hundred yards in the shorter, the mean difference being 28 3 seconds. intervals between them. The findings are very Accordingly we have always discarded blood samples similar to those in Group C (Table 3). The mcan obtained by unsatisfactory venepuncture. fasting R.P.C. time is 139.7 seconds and the mean We have previously shown that a high-fat reduction 3 hours after ingestion of the meal is breakfast containing approximately 85 8. anınal-fat 18.4 seconds. (2) oz. bacon, 2 eggs, 1-11 oz. butter, tea with milk and sugar) results in a marked shortening of the Group E. R.P.C. me a few hours after the meal (McDonald and Fullerton, 1958, a and b). We have found a Comprised 16 of the 23 students of Group D. similar effect after approximately the same amount In orda of vegetable-fat mainly as margarine (McDonald start and Fullerton, 1958 a), but our observations in this only communication are confined to the results obtained after with the standard animal-fat meal and they are remaining, 16 reported to hospital as before, and converuently discussed under two headings meal was only 7.2 seconds whereas it was 23.8 1-The effect of physical actity on the acceleration of the RPC. time after ingestion of the high-fat meal. Observations have been made in two groups of in-patients, none of whom suffered from cardiac disability, or received any drug likely to affect coagulability, Group A. The findings in the first group of 11 patients time which occurred after the fatty meal when the subjects were mainly sedentary. In subjects at rest in bed (Groups A and B) the fasting R.P.C. time is short and is much accelerated after a high-fat intake. Both of these factors muid ho rannede fa Ar chnun in The Group B. The cernnd in 1953). 19 ·2-The effect of phenindione on the increase in blood coagulabılty following a high-fat intake. TABLE I This has been studied in two groups of patients as follows: ARMIS SIMULTANEOUS R.P.C. TIMES (SECS) IN BOTH IN 14 SUBJECTS: VENEPUNCTURE UNSATISFACTORY IN ONE ARM, Group F. 10 in-patients with recent cardiac infarction at rest in bed and treated with phenindione for periods varying between 4 and 18 days at the tune the experiments were conducted, Satisfactory Unsatisfactory Amm Arm 120 7 92.4 Means Difference 28.3 TABLE 2 MEAN R P.C. TIMES (SECS.) BEFORE AND AFTER HIGHI-FAT MEAL. GROUP A: I IN-PATIENTS (NON-CARDIAC). Fasting 3 hour Difference 4 hour Difference 114 75 38.7 79 35 5 TABLE 3 MEAN R.P.C. TIMES (SECS.) BEFORE AND AFTER HIGH-FAT ALEAL. Group G. lo out-patients with previous cardiac infarction who had had therapeutic doses of phenindione for periods varying between 12 and 45 months. In both groups the one-stage " prothrombin time" had been maintained at 2 to 2 times the control level and the last dose of phenindione was taken in the evening before the test day. In all the subjects in both groups blood samples were removed before and 3 hours after the breakfast containing 85 g. animal fat. It is seen that in Group F (Table 4) the mean fasting R.P.C. time is slightly prolonged (147.1 seconds) as a result of the administration of phenindione, but the interesting point is that the mean acceleration 3 hours after the high-fat meal is only 3 6 seconds. This is in marked contrast the findings in Groups A and B (non-cardiac in-patients receiving no phenindione) where the mean reduction in the R.P.C time was more than 30 seconds In Group G (Table 4) the fasung R.P.C. uume is again slightly prolonged (mean 1473 seconds) and at 34 hours after the high-fat meal the expected acceleration is not seen, the mean of the readings being only 2 3 seconds less than the mean fasting value. In respect of physical activity this group is comparable to Group C (Table 3), 1.e. the control group of out-patients with previous cardiac infarction receiving I mg. phenindione daily, in which the mean R.P.C. time 31 hours after the high-fat meal IS 16 7 seconds shorter than the mean fasting value. Therapeutic doses of phenindione virtually abolish the acceleration of the RPC. tine which occurs after the ingestion of a high-fat meal in subjects not receiving this drug. So far as we are aware this action of phenindione has not been annan Fasting 31 hour Difference 111.6 80 4 31.2 Group B to In-patients (non-cardiac). Group C 10 Out-patients (old cardiac infarction 1 mg P.). 126.7 110 16.7 Group D: 23 Students, "sedentary day. 1397 121.3 184 . Group E 16 Students, active ” day 133.1 I25.9 7.2 TABLE4 The practical importance of this action of np.Li..hat the MEAN RPC. TIMES (SECS) BEFORE AND AFTER HIG?I-FAT MEAL THERAPEUTIC DOSES OF PHENINDIONE. * wivu Fasting 31 Hour. Difference Group F REFERENCES 147.1 143.5 36 10 In-patients, recent McDonald, A, Fullertou, 11 W (1958) Lancet, II, 599 cardiac infarction, 11958) Ibxi, II, 600 Vorris, J., Headı, 1 A, Rame, PAB, Roberts, CG, Group G 147.3 145 2.3 Parks, SH (1951) bid, II, IOS 3 10 Out-patients, old Report of the Working Parts On Inticoagulant Therapy in cardiac infarction. Coronary Thrombosis to the Vedic al Research Council (1959) Brit Med J 1, 803 30 ·Studies On Fibrinolysis SOME INTER-RELATIONSHIPS BETWEEN CLOTTING AND FIBRINOLYSIS E. KOWALSKI and S NIEWIAROWSKI – Laboratory for Clinical Biochemistry. Institute of Hæmatology. Warsaw EUGLOBULIN FIBRINOLYSIS AND CONTACT FACTOR INCU DATION MIXTURE ( 30 MIN 37°c) LYSIS TIME (HOURS) The theoretical background of thrombosis and its treatment is related to the phenomena of coagu. lation and fibrinolysis. Coagulation research is the " basic science " in which progress may be of great importance for adequate prophylaxis and treatment of thrombotic discase. Coagulanon and fibrinolysis are intimately connected. Their biological role is antagonistic. An increase in coagulability favours thrombus formation, the activation of the fibrinolytic system counteracts it. The intimate mechanism of these mutual interactions is not completely known. We do not know, we only guess which particular conponents may act simultancously in both systems. The following are the significant findings concerning coagulation factors which influence the fibrinolytic system. (1) Fibrinogen and fibrin possess considerable SERVM EUGLOBULIN • SALINE 20,0 EUGLOBULIN OBTAINED FROM SERUM SHAKEN WITH KAOLIN (KAOUIN IN EUGLOBULIN), SALINE 3,25 PA..... EUGLOBVUN OBTAINED FROM SERUM SHAKEN NITH KAOLIN (KAOLIN REMOVED) SALINE 10,0 ALCALINE EIVATE FROM KAOLIN SALINE 11.0 theory of clot dissolution in vivo postulated by Sherry et al. (1959) (2) The fibrinolytic activity of thrombin in Vitro was found by Guest and Ware (1950). Kowarzyk (1952) described a "thrombin protease" connected with thrombin. Seegers and Landaburu (1957) demonstrated that one of the different molecules of thrombin (thrombin E) possesses fibrinolytic activity. Interrelationship between thrombin and the fibrinolytic system in vivo was demonstrated by Kudriaszov and Ulytina (1958). After an injection of thrombin into animals the fibrinolytic activity increases so rapidly that the blood does not clot. (3) Kowarzyk and Rechnic (1957) found that the cuglobulin fibrinolysis is activated by removal of plasmin inhubitor during thrombın formation from prothrombin (" thrombogenetic disinhibition of fibrinolysts "). (4) Calcium ions added in appropriate concentration activate the coagulation system and also fibrinolysis in euglobulin (Kowalski et al., 1959). ALCALINE ELUATE FROM KAOLIN SERUM EUGLOBULIN 2,5 Figure 1 can be seen that hie al... --- activit F evaluduun of these phenomena. After this short review of coagulation factors influencing the fibrinolytic system some data concerning the influence of plas.nin (fibrinolysın) on clotting factors may be summarised. Plasmin inhibits all coagulation stages by destroying some coagulation factors. Its action on fibrinogen and fibrın is very well known euglobulin fibrinolysis time is shown in Fig. 1. It 21 ·FIBRINOLYTIC ACTIVITY OF EUGLOBULIN PREPARED OF SERA SHAKEN WITH KAOLIN AND CELITE LYSIS TIME (MIN ) SPECIES SPONTANEOUS ACTIVITY SX - ACTIVATED CONTROL KAOLIN TREACILITE TREA CONTROL KADUN TEA CUTE TRA SERUM TED SERUM VED SERVM SERUM EDITRUM HLED SERUM Figure 2 HUMAN 300 23 34 4 1.5 4 LOVINE > 1000 >1000 1000 20 105 SHEEP > 1000 > 1000 1000 1000 > 1000 > 1000 NORSE 1000 55 180 210 20 25 RAJBIT 960 140 960 30 5 10 GUINEA PIG 7 1 2 ACTIVITY OF BLOOD CLOTTING FACTORS AFTER INCUBATION WITH PLASMIN (+37°c) 10 Figure 3 CO 40 X O 10 E 10 INCVOATION TML (VN 10 PLATELET THMONDAY76 94010 - FACTOR - AUX 50 PLASMA TROMBOPLASTIN - FACTOR EUGLOBULIN FIBRINOLYSIS AFTER INJECTION OF HEPARIN AND ITS DIURNAL VARIATIONS Op WTO $000 m CONTOL - JALO v 260 220 .. I DEROBE MED RIN 180 1 2 AUN ASTH MEA NO o WWW AFTER MEDABA Figure 4 800 Y II 22 ·- Souliet et al. (1936), PLASMA FIRRINDLYTIC SYSTEM IN PATIENTS DURING using whole plasma, reportcu LONGTERM ANICOAGULANT THERADY that in vitro plasmin destroys Factor V, Vii, prothrombin, AHG, Chnstmas Factor and complement, in order of PUDELAN MAKALLI decreasing Sensitivity, notoLTS IN ACTIVITY ( MI) Nichiarpuski and Latallo (1997), who used purified PADNUDSON factors, found that plasanın ACTITTY (ERC) masurated AHG, Factor V, and plasma thromboplastin, but does not nfluence the Christmas Factor, Factor VII and the thromboplastic activity To tu (*) of platelets (Fig. 3). There is also experimental evidence that the fibrin stabilising factor (FSF) IS destroyed by plasmin (Buluk 1959). At least two different inhibitors of coagulation Σ Τ Υ T 1 appear during the proteolysis of fibrinogen: anti-thrombin VI (Niewiarowski and Figure 5 Kowalski 1996; Niewiarowski et al. 1959) and inhibitor of thromboplastın formation, THE COAGULATION ANO FIBRINOLYTIC SYSTEM I.T.F. (Niewarowski et al. IN RABBITS AFTER A PELENTAN ADMINISTRATION 1959). They inhibit thromboplastin formauon, thrombin action on fibrinogen and fibrin %. OF INITIAL VALUE monomer polymerisation Reprotucal fonts Throun (Kowalski 1959). 190 Diath Harmor by Nestront of Thc pharmacological Schallaner Telur Stuttgart treatinent of thrombosis can be performed with three kinds of substance: (1) heparin or heparinoids ; (2) dicoumarol and its denvatives ; (3) plastrin 19 ANTIPLASMA and other fibrinolytic sub- Each stances. soc these of FLASMAGEN substances may interfere with FARINOGEN the naturally occurring clothing DRUTNR OMRIN or fibrinolyuic processes. 6 (1) Heparin. The actions EUGLOBULIN FIBRINOLYSIS of heparin on blood clotting are 40 described in a separate paper. Concerning the influence of 20 TIME AFTER ADMINISTRATION heparin on the fibrolytic OS POLENTAN (TROMETAN) 40 system, the literature is (HOURS) contradictory (von Kaulla and MacDonald, 1958). We in- Figure 6 içcted heparin in a single dose of 5000 or 10,000 units into normal human has no significant influence on the fibrinolytic system Subjects Blood samples were drawn before injection in vio. of heparin and 20 minutes or 3 bours after the micction. The fibrinolytic activity was estimated using the euglobulin method (Fig. 4). No significant (2) Dicoumarol derivatives. Their influence on hanem changes were observed 20 minutes after injection of hepartn. An activagon of fibrinolysis was noted 180 minutes after injection of sooo or 10,000 units. However, thus acuvauon us of the same order of magnitude as the diurnal variations of fibrinolysis coagulants. No activation of fibrinolssis in whole plasminogen level is not influenced by these anu- (Fearnley et al. 1957; Kanisk et al. 7959). Similarly, it was shown in these experiments plasma was observed (Halse 1948; Wurecki 1998). that heparin has no influence, at 20 and 180 Our group investigated the influence of the dicoumarol derivatives Tromesan, Sincrom, and minutes, on the plasminogen and antiplasmin levels. Marcoumar on plasma fibrinolytic activity (Nie- From these investigations it seems that heparin studied in a number of patients with coronary warowska and Wegrzynowicz 1959). They were 23 T ·DOG 1 DOG 3 3 1 » INFLUENCE OF PLASMIN (10 ml/kg. OF WEIGHTON and an increase of antiplasmin level. These two THE BLOOD COAGULATION SYSTEM IN THE DOG changes may be correlated with the prolongation of the prothrombin time (Figs. 5 and 6). It has been suggested that a substance inhibiting fibrinolysis activation increases in the plasma of DOG 2 patients treated with dicoumarol derivatives. This substance could be removed by kaolin or celite (Niewiarowska and Wegrzynowicz 1960). (3) The purpose of treatment with proteolytic Fibrinogen mg/ml 1,68 1,68 1,56 1,0 1,68 144 enzymes is the dissolution of clots. We injected SK, activated plasmın into dogs and cats and examined Fibrinolysis in Total Plasma 10% 10% 10% 100% 0% 100%| the coagulation system. Contrary to results in vitro no changes in clotting factors other than fibrinogen were observed after plasmin injection The fibrinogen Cloting Time (Lee-White) 3'40" 6'55" 1 30" 420" 3'10" 7'30" level decreases extremely fast in cats, while in dogs this phenomenon is slower (Fig. 7). Recalcination Time 50 60 46* 246 | 52* The finding of an indefinitely prolonged thrombin time and greatly increased antithrombin 31" 6 17" 176 17 26 Antithrombin VI 18° 32' 21' >6 Prothrombin Time 9 916,5' 8" 16 Foctor v 14 12" 12" 12 12 proteolytic enzymes is related to the level of antithrombin VI in the circulation. Factor VIP 18" 18" 19" 18" 20" 21" We consider the thrombin time to be the most effective test for the control of treatment with Factor VII (Ang) 100% 100% 100% 100% 100% 100% proteolytic enzymes. Deutsch and Fischer (1960) are of the same opinion. Factor IX (Christmas) 100% 110% 100% 115% 100% 95% After an excessive dose of proteolytic enzymes Thrombin Time 139 1 BEFORE HFUSION || 50 MINUTES AFTER INFUSION * AFTER 74 HOURS EX AFTER 20 MINUTES Figure 7 thrombosis on long-term anticoagulant therapy and in rabbits treated with a single dose These anticoagulants have no significant effect on fibrinogen and plasminogen levels. They all cause a prolongation of the euglobulin fibrinolysis time, merisation and that protamine sulphate and other agents may overcome thuis effect (Kopec et al. 1960). These findings are not discussed here in detail They provide a rational background for treatment with proteolytic enzymes, its appropriate control and prophylaxis of complications Hemor, REFERENCES Buluk, K, personal communication, 1959 Niewiarowska, V, Wegrzynowicz, 2, unpublished, 1960 Deutsch, E. 1 scher, M. Thromb et Dath Hæmor, 4, 482, Men jarowshı, S, Latallo, 2 Bull And Polon Sci, 5, 219, 1960 1957 Tearnley, GR, Bulmforth, G , rearntry, C Clin Set, ih, Niewiaronski, S, honala, L. VI Congrés de la Sox Lurop 645, 1997 d'Hematol, Copenhager, 1997 (harger, 1958) Guest, MV, Ware, A G Science, 112, 21, 1950 View larowski, S, Kowalsh, E Revuc d'Hématol, 33, 320, Tlake, T Fibrinolie, Freiburg, 1958 1958 hantak 1, Swish hotsrhy, VI, Glogowska, I Past Viewiarowski, S. Konutske, L, Stachurska, J Acta Bloch Uus Med Ios , 12, 299. 1959 Pol, 6, 43, 1959 Micwarowski, S, Kaulla, KN MacDonald, T S Blood, 13, 811, 1958 Prou Hartelle, 0 Thromu, et Dinth 3, 593, 1952 kopec, V, howalski, L, Stachurska, I 1hround et Diath Scegers, l'11 Pror of the IV th Tutor Congress of Bio amor (in press) chemistrs, Ienna, 1958, p 18 (Pergamon Press) howalski, I Vontreux Conference on Blood Clotting actors, Serges, H, Lindaburu, Amer J thy stol, 195, 167, 1959 1937 Kowalski, E, Latallo, 1. dieu laron shi, s, Marczak, k, Sherr, , 1 tetscher, 4 P, Alkyrrsig. Physiol Rev, Panaset 107, J Pol 11 Lek, 12, 1221, 1957 39. 343, 1959 Kowalski, L, kopec, 11, Viewsrowsht, S J Clin Pathol. Soulier, J P. Alagille, D, Larrieu, Y J La Semaine des 12, 215, 1959 Hopitaux de Paris 1 7. 32, 1, 1956 howarzyk, 11 Sature, n. 614, 1952 Haaler, BÀ Scend J Clin Lab invest IT Suppl 17. 135. Iowarzyk, 11 , Rechnic, V. Acta Physiol Polon, 7, 1, 1957 1999 Kudrtaszos, 1 A, L24 tuna, P D Nature, 182, 397, 1958 trecki, Il Suppl ad 33. 1938 #zowska, l, llegrzynowicz, Ł Thromb et Diath Hitte, S I Intern Conference on Throm lists and I inbolisti, Harmor. 3, 279. 1959 Basle, 455, 1954 2+ ·The Tissue Activator Of Plasminogen And Thrombosis A. S. TODD-Department of Pathology, Queen's College, Dundee 1. f.lloved in of a large vein and incubating them in a fibrin film. Under these conditions, endotherum caused enough fibrinolysis to account for all the activity seen in sections (Fig. 9). The fibrinolytic activity produced by, arteries is usually very much less than that scen in veins. reme Adee " Since thrombosis is a local phenomenon it may be caused by failure of a local fibrinolytic mechanism Therefore, it seemed necessary to investigate the distribution of fibrinolyuc acuvity within the tissues. The chicf fibrinolytic substance in tissues is an activator of plasminogen (Astrup and Permin, 1947). This substance can be detected and assayed by the fibrin plate method (Permin 1947, Astrup and Albrechtsen, 1957). The method is to incubate Forman of ten- or dmne nf treur extract on a thus reaction involves 3 stages :-1-lhe conversion of a plasminogen proactivator to an activator bs 2 kinase (streptokinase). 2- Conversion of plasminogen into plasmin by activator. 3-Lysis of fibrin by plasmin. method was modified for microscopic examination (Todd, 1958, 1959). in the new method, a bustological section of fresh ussue is placed on a very thin layer of fibrin, and incubated. During incubation structures con taining plasminogen activator cause lysis of the adiacent fibrin. After incubation, the preparation from the circulating plasma but this is so loosely attached that even two Washings will remove it. These results suggest that the fibrinolytic mechanism In all tissues the areas of fibrinolysis appears to in the endothelium is in the form of a latent system be related to blood vessels and in most cases the which is set in motion by change in local conditons, veins show more activity than the arteries (Fig 2). or perhaps by exposure to some humoral agent which acts as a "kinase ". In the liver and spleen the amount of fibrino- Iyuc acuvity is small and it is almost caurely related Three observations are of interest in a search to large veins both portal and systemic (Fig. 3). for the local conditions or substances which may The pulmonary arteries are very active (Figs. 4 produce activation of this fibrinolytic mechanism. and s) and so are the main pulmonary veins The first concerns the distribution of the fibrinofytic acuvity throughout the issues of the vascular Large blood vessels show most activity in relation to their vasa vasorum (Fig 6), but zones of fibrinolysis can often be seen in relation to thic 10tuna, and to fragments of endothelium lying free in the lumen (Figs. 7 and 8.) The latter observation suggested that it was the endothelium that contained the plasminogen activator. This was tested by scraping cells off the intima di celles ils liell is DegugiDIC. I 25 ) De ui ineunan- 25

·Figure Fibrinolysis autograph of
muscle. The pale areas are gaps in
the fibrin background indicating
fibrinolysis due to the presence of
plasminogen activator in the tissues
Hæmalum x 76
@
Figure 2-Salivary Gland showing
fibrinolytic activity round blood
vessels The ducts are Inactive
Hamalum X 30
Figure 3-Liver. The activity is
confined to large veins and their
Y2$a vasorum
Hamalum 30

·Figure 4-Lung. The fibrinolytie
activity is related to pulmonary
arttries.
Hamalum x 30
Figure - Lung. Showing lysis
round a pulmonary artery.
Hamalum x 76
Figure 4 Aorta, The fibrinolytic
activity is related to the Vasa
vasorum. The intima (left) is
inactive
Hamalum x 30

·Figure 7-Vena Cava showing activity in relation to the vasa
vasorum, to Intima, and to cellular fragments in the lumen
Hæmalum x 45
Figure 9-Endothelial cells from a vein. They show
marked fibrinolytic activity
Hämalum X 96
Hamalum 76
Figure 8--Pulmonary Artery showing fibrinolytic activity
round desquamating fragments of endothelium
Hæmalum 30
Figure 10—Salivary Gland showing moderate activity in
the large central artery and normal venous activity.

·Figure Il-The wall of a vein from a leg amputated under bloodless conditions Note the platelet clumps in the areas of lysis Hamalum 76 ism would explain the gradient of endothelial fibrinolytic activity from the enous to the arterial parts of the circulation. Incidentally, a similar gradient of fibrinolytic activity has already been described in the blood itself, that from the veins being more active than that from the arteries (Fearnley and Ferguson, 1997). The second set of observations related to the behaviour of blood and tissue in lumbs which have been schanic, Kwaan and McFadzcan (1956) L.L ..11 JI...1-CLO samples are taken the distribution of fibrinolytic activity is such as to suggest that the most active blood is that which has stagnated in the capillaries and small venules I was able to study tissues from a fimb amputated for sarcoma under "bloodless surgical conditions using an Esmarch bandage vam *** nafhan-na-an Amd raing of hee paralleled by an increase in the amount of plasminogen activator in the vascular endothelium ran'est cellular eet of cells ar was least indothelium uld release the latent activator mechanism It was noted in the preparations from the ischemic lub that the active endothelial cells were often accompanied by clumps of platelets, suggesting that they, too, had been damaged. Figure 12—Artery from an amputated leg. There is These observations suggest two ways in which unusual activity in the arterial contents (above) which include detached endothelium. the fibrinolytic mechanism in endothelium may be set in motion Firstly, by damage, and secondly, Hæmalum x 76 29 ·Figure 13-Thrombosed vein showing fibrinolytic activity in the vasa vasorum and in channels In the thrombus Hæmalum X 30 Figure 14-Metastatic pros. tatic carcinoma in the seminal Vesicle. The fibrinolytic activity is related to the surrounding tissue rather than to the clump of cancer cells. Hæmalum y 30 by some humoral substance whuch may come from the tissues into the venous blood. It has previously been shown by Astrup and Buluk (1960) that the I. annagre 1 fibrinolysis can produce a dangerous hæmorrhagic state, presumably because fibrin acts as a part of the blood-ussue barrier, as Nolf (1908b) suggested, These two extremes can be illustrated by histological preparation. A section of a thrombosed veint shows that where the thrombus is adherent to the vessel wall no fibrinolytic acuvity can be detected in the intima. Activity can, however, be - h. 211 LAV Brinkhous anu Suus, igyu) (Opie and Barker, 1907) in the blood normally keep the rate of these reactions at a low level (Astrup, 1956b). Alterauon in these humoral factors may determine the outcome of local damage. A balance anhanense enables and an gulation rease in ?. KL 30 ·(Fig. 14) show that it is the tissue round the tumour in a moist chamber at 4°C. For use they are placed, rather than the carcinoma itself which produces cellophane side down, on a microscope slide. the excessive activity. 3-Tissue Sections are cut on the freezing slide To SUMALARISE. 1-The fibrinolytic activity microtome using the Schultz Braun cold knife of tissues is concentrated in the endothelium of technique (Romcis, 1948). The sections are taken blood vessels. 2-The endothelium of veins is much straight on to the fibrin film. more active than that of arteries. 3-Arterial endothelium may be activated by cxposure to strepto- 4-Hautchen Preparations of endothelium are kinasc. 4-There is some evidence to suggest that made as follows: A square of tissue is cut from the thc fibrinolytic activity of endothelium may be wall of a large blood vessel and washed in salınc. increased Sy damage and perhaps by some sub- After being allowed to drain briefly it is placed, stance which accumulates in the veins from the endothelium downwards, on a microscope slide and tissues..5-It is suggested that local fibrinolytic the excess fluid at the edges is dried off by touchung mechanisms such as the one described are important with the edge of a filterpaper. The slide is then in the pathogenesis of abnormal thrombosis. placed on the chuck of the freezing microtome and the area bearing the vessel is chilled with two APPENDIX (Materials and Methods). ---L--- Le Li. I-Tissues. Biopsy and post-morten human tissues varying in age from a few minutes to 48 hours may be used. If the material is to stored it is rapidly frozen and kept in the ice chest. Fibrinogen. Bovine Plasma Traction 1 ("Armour") which is supplied as a freeze dried powder is dissolved in modified Michaelis buffer S-Incubation, 15-30 minutes incubation at ---L-I.-T.. 37°C is usually sufficient to demonstrate fibrinolytic activity. The preparations are enclosed in a moist cha:nber" during this stage. 6—Fixation The preparations are exposed to the vapour of boiling formaldehyde solution for 30 seconds and are then placed in formol saline contain 20 units per mi. solution for I hour, During this stage the fibrin Cellophane. Ordinary commercial cellophane becomes detached from the cellophane. Fibrin 15 used. It must be wettable and have neither and section are then treated together in the same way as a "floated out" section. 7-Staining. The preparations are stained for 5 minutes in full strength Harris's hematoxylin and differentiated overnight in distilled water. They are "blued" by exposure to anmonia vapour. 2-Preparation of Fibrin Film. This is made as a thin layer of fibrin on a cello- jelly on glass slides. 8--Mfounting. They are mounted in glycerine phane backing using the following technique. A sheet of plate glass is accurately levelled by Fibrinolyric Acritury of Irhole Blood. By the method of Fearnley, Balmforth and Fearnley (1956). The activity was expressed as the I. poured into the dish and spread quickly with a glass rod and left to coagulate. I ml of fibrinogen X 10,000 lysis time in minutes ACKNOWLEDGMENTS I wish to thonk the administrators of the Wellcome Trust for o Research Travel Grant and to acknowledge the encouragement given me by the continued interest of Professors JB Duguid and A Lendrum. I om grateful to Di Tage Astrup for his interest ond the opportunity to work in his laboratory, to Dr R. O K Schade for much help with the illustrations, and to Dr W W Pork for many helpful comments. R. CAR REFERENCES Albrechtsen, O K (1997), Boot J Hematology, 3, 284 Mullertz, S and Lassen, VI (1952), Proc Soc. Lxp. Bio Med, Astrup, T (1956a), Blood, 11, 781 82, 261 Astrup. T (1936b), Lancet, 1, SÓS Xoll, P (1908), Arch Internat de Physiol. 6, : (a) and Astrup, T and Albrechtsen, o K (1937), Scand J Clin Lab 306 (b) Opie, Land Barker, B 1 (1907). J L.xp Med. 9. 207. Permin, P V (1917), Nature, 160, 571 Romeis, B (1948), Mikroskopische Technik, 15th , Vunich, 1957). 123 Tagnon, HS, Whitmore W. F and Shulman, R. (1952) Cancer, 5.9 Tadi... BlacFarlane, R G (1937), Lancet, 1, 1103 MacFarlane, RG and Biggs, R (1957). Hurran Blood (1936), Coagulation and its Disorders, and Edition, Orford, 387. 31 Sci, ·DISCUSSION I- PLATELETS 'NIN ADHESION 1. TISSUE SURFACE ( CONNECTIVE TISSUE FIBERS) (SURFACE COATING) 2. PLATELET ADHESIVENESS PROPERTY 3. REO CELL TACTOR (HELLEM) RED CELLS It is certainly not arterial spasm, REYERSIBLE 4. PLAS"A ADHESIVENESS FACTOR (NILSSON-LOUBICK) for there is no stimulus for this, AGGREGATION s. CALCIUM (MAGNESIUM) Besides, arterial blocks in various sites are local, probably from a platelet buld-up, and the cause is IRREVERSIBLE THROWBIN FORMED in the vessel wall. Also, did Dr -EXTRINSIC SYSTEM I AGSREGATION IN THE PLATELET Todd's method with frozen sec- ATNIOSPHERE INTRINSIC SYSTEM tions exclude the activation of VISCOUS lysis in the processing? METAMORPHOSIS Professor OWREN-Mitchell's → PLATELET LIPOID FACTOR IMPERMEABLE investigations again emphasise the lack of correlation between PLATELET PLUG arteriosclerosis and clinical INTRINSIC SYSTEN disease. The rising mortality ITE seems not to be caused by PLASMA FIBRINA THROMB IN FORMED increasing atherosclerosis. An IN PLASMA increased tendency to thrombosis CONSOLIDATED +EXTRINSIC SYSTEM must be considered. The decline HEMOSTATIC PLUG in post-operative thrombosis and coronary disease in Norway during Figure 1-The hamostatic mechanism the war is consistent with this. The effect of diet must be studied further, and I agree that it is not simply a problem in coagulation : PLATELET TRANSFUSION factors independent of u are HEMOPHILIA A recognised. We have studied mainly hæmostasis after injury to small vessels, but certain reactions are the same in arterial thrombosis though the trigger mechanisms 30 may differ. In both the wound surface and the vessel wall are decisive. Wessler's and Poole's 25 experiments, however, are differ- SECONDARY BLEEDING TIME ent, because clot - promoting factors in the blood play the major 20 role. They may give important information for venous throm- NO CHANCE IN bosis and the problem of hyper- 15 OLOTTING TIME (20-30 min) coagulability, but it must be CEPHALIN TIXE (200.213 v. 1 applied cautiously to arterial ZACTOI VE KI per cant thrombosis. 10 PRUMAAT BLEEDING TIME (urmeu In the hainostatic mechanism (Fig 1) factors 1-4 are essential 5 since the primary bleeding time and platelet adhesiveness in vivo (Borchgrevinck, Acta Med Scand., 1960) are altered in 0 2 3 4 5 6 7 hæmorrhagic telangiectasia, thro.nbosthenua, severe anzmia DAYS and Von Willebrand's disease Figure 2-The effect on the secondary bleeding time in hzmophilia of The tissue substrate for platelet transfusion of normal platelets (Platelets from 21 of plasma were given) adhesion is the collagen and elastic fibres (Hugues). Platelets adhere loosely or not at all to cholesterol and fattreated red cells increases adhesiveness to glass relevant to the trigger mechanism of thrombosis parallel to the hæmatocrit value A protein-free secondary to athero.na. concentrate of factor R has been prepared, and causes .-.An.hu The red cell factor (factor R of Hellem, Scand. J. Clin. Lab. Invest., Suppl 12, 1960) may be an important trigger. Platelets in red cell-free plasma are not adhesive. Addition of washed or trypsin. 32 ·1 platelet thrombus. Biecding in the vessel wall may also be significant. The second stage is platelet fusion by viscous metamorphosis, presumably caused by thronbin. death of patients with grossly hypertrophicd ventricles 15 replicable by anoxia favouring ventricular fibrillation the capillary blood supply does not keep pace Hith the enlarging muscle fibres. Dr SHARP - When platelet-rich plas na clots, the platelets clump some time before fibrin forms For 1-2 minutes after clumping it can be reversed In Poole's illustrations, the platelets in the thrombus formed discreet clumps of unuform sizc. La to about is. Moreover, plasma concentrations must be reduced Sudden death from electrical instability (ven. with adiancing coronary descase. mass. Platelets on clumping increase in volume roughly fourfold. This also is of interest in thrombus formation. Dr SCHIT ARTZ - In the " sudden cardiac death group there is no coronary thrombosis and the degree of atheroma 18 unimportant (current Oxford studies). This group should be excluded from discussions of coronary occlusion in myocardial .1-. not believe that atheroma is thrombotic in origin, ..L .. Covered. injected iritant substances into the arterial lumen Quite frequently at autopsy widespread throm- While the blood flow was temporarily arrested, but botic occlusions are found in many arterics. This this resulted in thrombosis only rarely and unpre suggests a systemic thrombotic tendency. dictably. To produce thrombosis regularly, it was There is much autopsy and clinical support for necessary to reduce the rate of flow through the Mitchell's views. Coronary and femoral artery segment; thrombosis was then almost invariable. The shock of a myocardial infarct may produce the thrombosis together have been found to occur during adequatc anticoagulant treatment. A man aged 46 circumstances leading to thrombus formation 1, therefore, welcome Dr Mitchell's finding that developed mesenteric angina due to aortic thrombosis occlusive thrombus is found more often in patients he had primary polycythema with a platelet count over 3 million, and his aorta was free from atheroma living for some days after the schämic cpisode, compared with patients dying suddenly. Dr MITCHELL - What reason is there for attributing sudden death to heart disease at all? Dr SCHIV ARTZ-The low incidence of occlusive thrombus in Professor Montgomery's As in Beck's experiments, these deaths might possibly be due to embolic occlusion of a twig vessel from a series 13 probably due to the age of the infarcts. It is proximal atheromatous plague, giving ventriculour much less common in older infarcts. fibrillation Cardiac infarction cannot be due to Professor HILL - The patients who die instantly atheroma alone : the deterioration is too rapid. at the onset of myocardial necrosis are important Thrombus or embolus is the most likely explanation. Dr VERSTRAETE --- Did Professor Fullerton find a strict parallelism between the Quick ume and the R.P.C. time, and was the prolongation of the latter found even when the Quick time was not in the therapeutic range? Professor FULLERTON - Our experiments Relative acute ischiamia may explain infarction were all done on patients in the therapeutic range or without thrombosis. Similarly, the liability to sudden showing no effect at all. 33 .L.-- ·inhihi...- Dr HARTERT - Did Dr Poole find increased viscosity shortly before visible clotting started ? With a similar device, the ring viscometer, we could not distinguish between increased viscosity and the beginning of real clot formation. Dr POOLE - When fibrin forms, the level of the advancing edge becomes still lower, quite quickly, and thus is how we measure thrombus formation time. The change in the position of the column of blood is probably due to friction between the fibrin and the walls of the tube. t t 14 LUI Seegers. In Verstraete's experiments, moreover, components of the clotting system other than thrombin may have been eliminated. With regard to Dr Todd's findings, Kowalskı et al. (Blood, 1958, 8, 436), using entirely different methods, found that the activity of plasminogen or plasminogen proactivator is very high in organs rich in connective tissue. Dr MATTER -- The increased antiplasmin activity in an Пr yep стр Алт- . Professor DONALD -- With regard to Dr. Todd's experiment in which he activated separated artar.nl -.. 1 a! ULAJULUI Oi welldle or thrombin needed for thromboplastin generation : other tests fail to reveal its activity. D- ACIV nonmenta inhibit substa (throrr 2010 VI bo- 11- Lal ve volained free of IT.G, and electrophoretically pure I.T.G has been prepared by Lipinski in our group. I.T.G may, in fact, act by Lauuuudi leus ? Dr TODD -- Red cells and leucocytes lose their activity, derived from the plasma, after two washings, whereas the substance is firmly attached to arterial endothelium Serosal cells behave like arterial cells. Bladder cpithelum is inactive. In answer to Professor Pickering, the activity of the tissues decreases with age and also with fixation Only fixation in cold 70% ethanol is of any use. The activity found in tissues is inversely proportional to the amount of autolysis. It might also be suggested that bacterial activity could be responsible for the phenomenon. This is unlikely, because (1) sterile biopsy material shows it; (2) no bacteria are seen in digestion zones; (3) there is no lag phase before lysis begins ; (4) addition of antibiotics and antseptics to the system does not abolish the reaction. 34 ·The Organisation Of Large Scale Life-Long Anticoagulant Therapy P. A. OWREN— Rikshospitalet, Oslo Introduction. the first group anticoagulants have marked effect Anticoagulant therapy has been violenily debated on mortality and disablement: this is our first priority group. In the second group the effect of anticoagulant therapy is less well established. The high priority group includes five subgroups. (a) Chronic rheumatic heart disease. Figs. I and 2 show the effectiveness of prophylaxis in L-L malaki. two problems : (1) Which patient should be treated ? (2) How should the control be organised? The therapy was first given to patients with acute diseases, and was stopped after a few weeks However + rip-onl - J.L. (6) A-*. - . otren important work. Our first study of angina pectoris comprised 471 patients admitted to the University Clinic of Oslo. All patients were given anticoagulants, and the mortality was only 36% per year in the treatment period, totalling 1400 years (Owren 1954). The usual mortality in patients not so treated 13 8-10% per year in a second study of 275 patients Waaler (1957) found that patients with a short history had a mortality of less than 1% during the first year (fig 3). In the group with long duration of symptoms before therapy, averaging 3.5 years in this material, the mortality was about 6% per year, which is not significantly different from the usual mortality of 8-10° (fig. 4) Based on these results we organised a third study which has been conducted by Dr Borchgrevink (1960). In order to So far, most physicians and laboratories have experience in managing only a few hundreds of patients However, because of the incidence especially of atherosclerotic disease in the western countries, we must now face the practical problem of making permanent anticoagulant therapy available to the great number of patients who need it. This means that we must develop an organisation for large-scale therapy. Indications for Life-Long Anticoagulant Therapy. The peneral etmirha of . L... PLUL Laugy, 45 expiessed by the P & P method and consequently also by the new Thrombotest method, must be below 50% (c) Myocardial infarction in this large group We select patients with only one attack, with no 44 LO duterosclerotic disease and rheumatic angina, or with angina for less than 2 years Fig. 6 heart disease because patients with other diseases shows the survival curve for non-selected patients giving indication for life-long anticoagulant therapy on anticoagulant therapy, our own group of 308 are comparatively few. patients and Bjerkelund's group of 119 patients. We classify the patients in two main groups. In Survival curves for 3 groups of non-treated patients 35 ·OBSERVATION PER100 (YEARS! 7 6 5 TREATMEHT PERICO (YEARST 3 4 5 B 4 3 2 1 1 8 ---- Figure 1 - The effectiveness of anticoagulant treatment in patients with mitral valvular disease and recurrent em. bolism prior to therapy EVBOLIC EPISODES ! FIRST EP Soor CONTROL ARAU TIME FM YAAS COACHIMUTATHARY T/MEX YEARS FIRST EPISODE Figure 2 Prophylactic effect of anticoagulant therapy after the first embolic episode in patrents with mitral valvular disease 17 PATICNTS #1 DOSCAVATION YEARS 22 AECURRENT MAJOR [49OLIC EPISODES 19 PATIENTS CO TREATMENT VLARS O 041 PISCO ANGINA PECTORIS 100 TREATED GROUP 901 WAALER (134) 801 Figure 3 -- Hect of ancicoagu- (ant therapy in angina pectoris with short duration of symptoms prior to therapy. BLOCX ET AL. 701 60 CONTROL GROUP 50 40 301 201 101 36 1 2 3 4 5 OBSERVATION YEARS ·ANGINA PECTORIS % 100 90 TREATED GROUP 80 WMLER (132) Figure 4 -- Effect of anticoagulant therapy in angina pectoris with tong duration of symptoms (average 35 years) prior to therapy PLOTX ET AL. CONTROL GROUP 70 60 50 40 30 20 101 1 2 3 4 5 OBSERVATION YEARS P&P LEVEL 50% 20 % NO. OF PATIENTS 68 70 Figure 5 - Effect of anticoagulant therapy in patients below 70 years of age with angina pectoris of less than 2 years duration and without myocardial infarction. ASYOCARDIAL INFARCTIONS 9 1 CARDIAC DEATHS 6 1 MORTALITY IN % PER YEAR 8 1.3 are included for comparison. The number of patients protection against coronary death. A high proportion with more than one infarction before therapy was of these patients die of coronary disease or of cerebral relatively small in these series, and definite con arterial disease, At our outpatient clinic, 140 patients clusions regarding the prognosis in such cases could with thrombosing atherosclerosis in the lower not be drawn. The figures, however, suggest that extremities have been on anucoagulant therapy for new infarctions were more frequent in patients with an average of 4 years (Osvik, 1960). The total more than one infarction prior to therapy, both in mortality has been 3.75°. per year, with 2.7% of the treated groups and in the control groups. cardiovascular disease. A study at Ulleval Hospital Bjerkelund states that anticoagulant therapy does in Oslo of a large group of simular patients not not seem to have protected these patients much treated with anticoagulants showed a total mortality (Bjerkelund 1957, p. 134). It is also logical to assume of 80 per year. We must not forget that atherothat the effectiveness of anticoagulant therapy is sclerotic disease is a generalised disease, and should highest at an early stage of the disease. There is no be treated as such. reason to believe that the antithrombotic effect (e) Intermittent insufficiency of the cerebral pet se is less in advanced discase. The difference circulation, presumably due to thrombosis, is the in results is presumably caused by a higher frequency last category on our high priority list. A decisive of non-thrombotic deaths in advanced disease, study math front of menn La that is, death from congestive heart failure and is stil sudden death from acute ventricular fibrillation us th (d) Atherosclerosis in the legs without signs The 1 or symptoms of coronary or cerebral atherosclerotic however. disease. All these patients have high priority, not Our list of second priority cases includes three because of the local disease, but because they need main categories : rheumatic heart disease without 37 1.- - ·OBSERVATION PERIOS (YEAAS) 7 6 $ 4 3 TREATME 1ST PERICA (YEARS 2 5 6 8 1 7 L.--..L Figure 1 The effectiveness of anticoagulant treatment in patients with mitral valvular disease and recurrent embolism prior to therapy. .. EVECLIC EP1300-s ! FIAST EPISODE CONTAMGROUP TIME IN YCARS & WAT THERAPY TIME IN TCARS FIAST EPISODE 3 7 3 Figure 2 - Prophylactic effect of anticoagulant therapy after the first embolic episode in patients with mitral valvular disease 17 PATIENTS It CASCRIATICK PEARS 22 RCCURACHT MAJOR (19CLIC EPISODES 13 PATTCUTS O TREATMENT TEARS O CUICLIC EPISCE ANGINA PECIORIS 100 90 TREATED GROUP WAALER (134) 801 701 601 Figure 3 - Effect of anticoagulant therapy in angina pectoris with short duration of symp" toms prior to therapy. PLOCX ET AL sol CONTROL GROUP 40 30 201 101 1 36 2 3 4 5 OBSERVATION YEARS ·ANGINA PECTORIS 100 90 TREATED GROUP 80 70 WALER (132) Figure 4 - Effect of anticoagulant therapy in angina pectoris with long duration of symptoms (average 35 years) prior to therapy 60 FLOCK ET AL. CONTROL GROUP 50 40 30 20 10 1 2 3 4 5 OBSERVATION YEARS P&P LEVEL 50% 20 % NO. OF PATIENTS 68 Figure 5-Effect of anticoagu. lant therapy in patients below 70 years of age with angina pectoris of less than 2 years duration and without myocardial infarction 70 MYOCARDIAL INFARCTIONS 9 1 CARDIAC DEATHS 6 1 MORTALITY IN % PER YEAR 8 1.3 are included for comparison. The number of patients protection against coronary death. A high proportion with more than one infarction before therapy was of these patients die of coronary disease or of cerebral relatively small in these series, and definite con arterial discase. At our outpatient clinic, 140 patients clusions regarding the prognosis in such cases could with thrombosing atherosclerosis in the lower not be drawn. The figures, however, suggest that extremities have been on anticoagulant therapy for new infarctions were more frequent in patients with an average of 4 years (Osvik, 1960). The total more than one infarction prior to therapy, both in mortality has been 3.75% per year, with 2.7% of the treated groups and in the control groups. cardiovascular disease. A study at Ulleval Hospital Bjerkelund states that anticoagulant therapy does in Oslo of a large group of simular patients not not seem to have protected these patients much treated with anticoagulants showed a total mortality of 8% per year. We must not forget that atherosclerotic disease is a generalised disease, and should be treated as such (e) Intermittent insufficiency of the cerebral circulation, presumably due to thrombosis, is the last category on our high priority list. A decisive study rthat is, death from congestive heart failure and is still sudden death from acute ventricular fibrillation us th: (d) Atherosclerosis in the legs without signs The r Of symptoms of coronary or cerebral atherosclerotic however. disease. All these patients have high priority, not Our list of second priority cases includes three because of the local disease, but because they need main categories: Theumatic heart disease without 37 ·MYOCARDIAL INFARCI ION (SUAVIVAL IN PER CENT) % 100 TREATED GROUPS 90 80 OWAEN (308) BERKELUND (119) 70 Figure 6- The effect of life long anticoagulant therapy on mortality of survivors of acute myocardial infarction 60 50 WERKELUND (118) DRAMMEN HOSPITAL (126) RULLFVAL HOSPITAL (929) CONTROL GROUPS 40 30 20 10 4 5 1 2 3 OBSERVATION YEARS ernbolism, angina pectoris of more than 2 years duration and two or more myocardial infarctions. For these groups there is only suggestive evidence that anticoagulant therapy decreases mortality. Therefore, it is not justified to insist on treatment for all these patients. Each patient should be considered an individual problem However, further research using the technique of well-controlled clinical trials must explore the statistical value of anticoagulants in such patients Size of the Problem. Our question is now. how big a practical problem is it to organise anticoagulant therapy for all patients in the high priority group and for selected patients in the low priority group ? The death rate in Norway from chronic rheumauc heart disease is about ten per hundred thousand If we assume an average life expectancy of 15 to 20 years after the appearance of clinical symptoms, the morbidity would be 150-200 per 100,000 Probably not more than 25 of these are candidates for protection by anticoagulants, that is, about 50 per 100,000. The death rate for atherosclerous and degenerative heart disease in Norway 1$ 173 per 100,000 This includes also most cases with arteriosclerosis probably includes about I/s of these or about 150 per 100,000 in Norway. If all new patients receive therapy, the number will increase to 800 per 100,000, or somewhat more if one allows for increased life expectancy due to treatment, Thus, we may conclude that roughly 200 per 100,000 at present, rising to 1000 per 100,000 or Iºn of the population in the next 10 years need to be on anticoagulant therapy. There are many assumptions and approximations in this calculation, but it appears realistic to us. The morbidity of atherosclerotic disease is somewhat higher in Great Britain than in Norway, but still the size of the problem is not overwhelming This, then, will be the problem. to maintain as many as 0.2% up to 1° of the population on safe and effecuve anticoagulant therapy. How should thus be done? Basic Problems of Anticoagulant Therapy. I must first analyse the basic problems of life. long therapy. To make my points clear, I shall arbitrarily divide these problems in two groups, The Geo ir of ol and has 2 items. other forms of therapy and psychological problems. 1.1 that some doctors tend to give anticoagulants to patients who do not need the treatment, such as 38 ·patients with cardiac neurosis, with uncomplicated art. To obtain good results, the doctor must be accurate in details and have a high degree of responsibility. atamarin thanay Organisation. Whea wc started long-term anticoagulant therapy at the University Clinic in Oslo 12 years ago, we used the P & P method for control. After 10 years, with more than 100,000 tests, we decided to modify it. The P & P test has one theoretical Can- ! explanations or repeat the test at once. This is an important detail. In this way charts are not mixed up, and it constantly reminds the patient that great care is necessary. If the patient has no problems, he goes back to work at once. The same afternoon all charts are reviewed by the doctor who makes necessary adjustments in the dosage. This is again an important detail. Such adjustments must never --- come these difficulties, in order to make anticoagulant therapy simple and yet safe. The P & P test was first modified to make it sensitive to all four factors reduced by the oral .-- ! - doctor about, he can see the doctor when the test Is finished. In this organisation the technical and medical problems...are handled by the same doctor, who Cara with thousands of patients, the capillary test saves much time and is more convenient for the patient. -1-LI. oscheap. The new test is simple, accurate and takes only about 2 minutes. It has been in general use in Norway for it years. It can be done by any office nurse or less than half an hour each day reviewing the charts. The patients spend a few minutes in the laboratory every month Finally, this organisation is inexpensive: the cost of reagent is at present about 9 shillings and of technical assistance about 8 shillings a year per patient. When the laboratory is part of a hospital, it is sometimes necessary to separate the technical and medical problems. The laboratory then takes care ... (3) Administration of anticoagulants is apparently an extremely simple matter. Nevertheless, experience shows that the result is often bad; the patents frequently get too small and fluctuating doses Both patient and doctor may be responsible for this. The patient must be thoroughly instructed It is necessary to sit down with fun and explain in detail what the purpose is, what the dangers are, capacity for anticoagulant therapy in our country and that you expect him to follow directions absolutely. Never must he forget to take his pells, and Bever must be change the prescribed dosage. The doctor must know the rules of dosage. They are few and simple, but often overlooked. Personal experience is the best teacher, and most doctors need two months before they master this enced doctor. 39 ·proposals. It is not possible to build up one system for all patients, since the system must be modified to meet local needs. It is a waste of time to discuss the theory of centralisation versus decentralisation, because this is an entirely practical problem, In conclusion, I will stress that this is a large, but not overwhelming problem. It can be solved, L. Many patients cannot conveniently come to such a laboratory for control. Their problem can be solved in two ways, Either they can have their practitioner send the blood to the laboratory, or the practitioner can do the test in his own office. It is perfectly feasible for him to do this. With a little experience and cheap equipment he can obtain reliable results. This solution would also combine the technical and medical responsibility, which is an advantage. Experience shows, however, that the result of this organisation is often not good This is not because there are any difficult problems, but simply because most doctors need experience before they do the job well. This is especially true for the dosage, which needs more experience than the performance of the test. We therefore recommend the following solution. If the general practitioner has a number of such patients, and if he is willing to devote himself henn....haha REFERENCES Bjerkelund, C. J. The effect of long term treatment with dicoumarol in myocardial infarction Thesis Olo Valversity Press (1957) Acra med Scand Suppl to vol, 158 (1957) Borchgrevink, Chr T. Long-term anticorgulant therapy . LI do the whole work alone, In this discussion I have tried to outline the general principles, rather than to make specific Ouren, PA Long term anticoagulant therapy in coronary artery disease Schweiz med schr , 84, 823-827 (1951) Haaler, BA The effect of permanent anticoagulant therapy on symptoms and mortality in angina pectoris Acta med Scand, 157, 389 306, (1957) The Laboratory Detection Of Thrombosing And Of Bleeding Tendencies HELLMUT HARTERT - Med Uniy Hospital, Heidelberg, West Germany The problem with which I would like to deal With Citrated plasma we had found that arose in a very practical way in our laboratory. We l-awhahalin maridante harta mon vnlue of were used to controlling anticoagulant therapy by being thromboelastography (TEG). We believe this to value be a safe method of control, since phases of produnnounced tendency to thrombosis during therapy culty of statistical comparison, a great ditterence in favour of the oxalated full blood method. This difference in sensitivity seemed not to be caused by the addition of the red cells but by the -anta Ayalata e do not yet know We were very pleased about the increased method. sensitivity of this test for a thrombosing tendency unul we noticed that now the sensitivity for a For practical reasons we changed our method bleeding tendency had gone This was indicated by of drawing blood one year ago Before this we took several heavy bleedirls accidents, where the Quick Le cansrated test constantly showed values below 5°. The TEG on the contrary failed with the oralated blood method to indicate a bleeding tendency. It has always been our opinion that the good sensitivity of TÉG in the control of anticoagulant therapy and especially in detecting a trombosing With this change to oxalated full blood we noticed tendency was due to its special sensitt ry for the a very pronounced sensitivity for increased coagu thronboplastic factors of the plasma. Th inportance lability. This was indicated by a greater shortening of estudiating thronboplastic activity 1. anticoaguof the clotting time in thromboses, in cases with lant therapy has also been indicated by the attempts Jung embolism and also in myocardial reinfarction of several authors to include one or more hrombo- 40 ·1 plastic factors in routine determination of the prothrombin complex. . Let us assume that in the TEG study of the gives a good cross-section of the plasma proteins, but docs not reveal for example important antibodies, which can cause death. Anticoagulant therapy is a rather crude method. It does the same as bulwarks on the slopes against rolling avalanches. Would it not be better to catch the sparrow before he starts the avalanche-or prevent the snow from becoming so dangerous ? But - mason"hasen nhw na TL - na -.-) tendency can apparently not be detected, if the influence of the extrinsic clotting system, as it 13 determined in the Quick test, becomes more prevalent by changing the method. This means the impossibility of a joint determination of thrombosing and of bleeding tendencies in one test, of both are to attain the highest possible sensitivity. hand there is no explanation why the therapeutic reduction of clotting factors must be so relatively high to escape thrombosis. In contrast to this, the bleeding tendency seems to be strictly correlated with a clotting defect except where there is a toxic defect of the capillaries. It is not my intention to underminc any existing or proposed method. I should like to ask rather how a control system could be devised which would both avoid bleeding and prevent thrombo-embolism.

·Comparative Study Or Control Methods For Anticoagulant Therapy W. ZOLLINGER - Krankenhaus, Neumunster, Zurich With the technical assistance of Miss L Muller The methods used were : Quick's test, whole (d) Reproducible results by different blood clotting time, recalcification time, thrombworkers at different laboratories. elastography and the heparin tolerance test. Many (e) Same results with plasma, decalcified authors believe that it is safe to use Quick's method whole venous or capillary blood. alone, Others believe in simultaneous use of the (f) Should not give too long a clotting time. methods mentioned. (g) Moderate cost. The reliability and reproducibility of any It should be available anywhere anticoagulant particular method depend on several important therapy is practised. Such a preparation can be of factors: animal or human origin. 1-The techniques have to be simple, so that We studied the commercial preparations in any laboratory the same results can be obtained by different workers. 2-The interpretation of the results should be by an experienced physician taking into consideration the anticoagulant used, whether long or short acting. 3-The greatest problem is the choice of thromboplastin, The ideal thromboplastin should fulfil the firms. following criteria: Different thromboplastins were first compared (a) Simplicity in use. using plasma. There was a good relation between (b) Sensitivity to all factors depressed by Quick's method and the Thrombotest, especially the anticoagulant. within the therapeutic range. The mean value by (c) Insensitivity to factors not influenced. Quick's method is about 10% higher than by Thrombotest, because different_dıluents are used CLOTTING for the calibration curves, for Thrombotest Seitzfiltered plasma and for Quick's method Michaelis TIME buffer The same picture was obtained when Thrombokinase Geigy was compared with Thrombotest, When human brain thromboplastın was compared SEC. with Thrombokinase Geigy a good correlation was obtained in the therapeutic range. Thrombokinase Geigy is rather insensitive to variations in factor VII (4) Since factor VII is rapidly depressed at 70 THROMBOTEST Figure 1 50 30 HUMAN BRAIN THROMBOPLASTIN 10 % FACTOR IX 14 2 6 00 10 42 ·the beginning of therapy, Thrombokinase Geisy gives higher values at this time than human brain thromboplastin, which is very sensitive to factor VII. The same was seen with Thrombokinase .L. 1.... - in both methods. For out-patients it is of great advantage if capillary blood can be used. We used the method described by Fiechter (2) with human brain thromboplastin and Thromboplastin Roche. This method has the disadvantage that larger volumes of capulary blood are needed. Quite good results are obtained by this method, but with Thrombokinase Geigy and Thrombotest much more reproducible results are obtained, especially within . showed absolutely normal. Thrombotest values. PLASMA Figure 2 * HUMAN BRAIN THROMBOPLASTIN THROMBOKINASE GEIGY' 100- 1004 50- 50 FACTOR х FACTOR 7. 50 100 50 100 P/. 10041 THROMBOTEST i 100 THROMBOPLASTIN 'ROCHE' 5071 50 FACTOR X FACTOR X . 50 100 50 Cross – Vit. K, or after cessation of therapy Large dot = Beginning of therapy 100 ·Comparative Study Of Control Methods For Anticoagulant Therapy W. ZOLLINGER - Krankenhaus, Neumunster, Zurich With the technical assistance of Miss L. Muller The methods used were : Quick's test, whole (d) Reproducible results by different blood clotting time, recalcification time, thrombworkers at different laboratories. elastography and the heparin tolerance test. Many (e) Same results with plasma, decalcified authors believe that it is safe to use Quick's method whole venous or capillary blood. alone. Others believe in simultaneous use of the (f) Should not give too long a clotting time. methods mentioned. (g) Moderate cost. The reliability and reproducibility of any It should be available anywhere anticoagulang particular method depend on several important therapy is practised. Such a preparation can be of factors : animal or human origin. The techniques have to be simple, so that We studied the commercial preparation in any laboratory the same results can be obtained by different workers. 2-The interpretation of the results should be by an experienced physician taking into consideration the anticoagulant used, whether long or short acting. 3-The greatest problem is the choice of thromboplastin. The ideal thromboplastin should fulfil the firms. following criteria: Different thromboplastins were first compared (a) Simplicity in use, using plasma. There was a good relation between (b) Sensitivity to all factors depressed by Quick's method and the Thrombotest, especially the anticoagulant. within the therapeutic range. The mean value by (c) Insensitivity to factors not influenced Quick's method is about 10% higher than by Thrombotest, because different diluents are used GLOTTING for the calibration curves, for Thrombotest Seitzfiltered plasma and for Quick's method Michaelis TIME buffer: honnant when Thrombokinası Wher SEC. with obtair Geig! VIII 70- THROMBOTEST Figure 50 .--ViSSiSS=: 307 HUMAN BRAIN THROMBOPLASTIN 10 % IX 6 2 10 14 18 FACTOR 42 ·4 แนน the beginning of therapy, Thrombokinase Grigy in both methods. For out-patients it is of great les higher values at thus time than human brain advantage of capillary blood can be used. The used -- * Very sencitive to factor the method described by Fiechter (2) with human brain thromboplastin and Thromboplastin Roche. This method has the disadvantage that larger volumes of capillary blood are needed. Quite good results are obtained by this method, but with Thrombokinase Geisy and Thrombotcst much more tcproducible results are obtained, especially within the therapeutic range. The chief property of a thromboplastın is its sensitivity to Variations in any one of the clotting factors depressed. Thrombotcst is claimed to be sensitive to the variations in factor X activity, Several severe cases of humophilia B in Switzerland showed absolutely normal Thrombotest values PLASMA Figure 2 . • HUMAN BRAIN THROMBOPLASTIN THROMBOXINASE 'GEIGY 1004 100 SO - 50 FACTOR X FACTOR X s'o 100 Y-% 100 50 . Wa 100- THROMBOTEST 100 THROMBOPLASTIN 'ROCHE' 50 SC FACTOR X FACTOR X T' 50 100 50 Cross = Vic K, or after cessation of therapy. Large dot = Beginning of therapy. ry 100 ·Comparative Study Of Control Methods For Anticoagulant Therapy W. ZOLLINGER -- Krankenhaus, Neumunster, Zurich With the technical assistance of Miss L. Muller The methods used were : Quick's test, whole (d) Reproducible results by different blood clotting time, recalcification time, thromb. workers at different laboratories. elastography and the heparin tolerance test. Many (e) Same results with plasma, decalcified authors believe that it is safe to use Quick's method whole venous or capullary blood. alone. Others believe in simultaneous use of the (1) Should not give too long a clotting time. methods mentioned, (g) Moderate cost. The reliability and reproducibility of any It should be available anywhere anticoagulant particular mechod depend on several important therapy is pracused. Such a preparation can be of factors: 1-The techniques have to be simple, so that used, whether long or short acting. 3--The greatest problem is the choice of thromboplastin, The ideal thromboplastin should fulfil the following criteria : (a) Simplicity in use (b) Sensitivity to all factors depressed by the anticoagulant. (c) Insensitivity to factors not influenced. GLOTTING TIME over other methods by being sensitive to tactor 1X as well as prothrombin, factors VII and X (s). Tests were strictly carried out as instructed by the various firms. Different thromboplastins were first compared using plasma. There was a good relation between Quick's method and the Thrombotest, especially within the therapeutic range. The mean value by Quick's method is about 10% higher than by Thrombotest, because different duluents are used for the calibration curves, for Thrornbotest Seitzfiltered plasma and for Quick's method Michaelis buffer. The same picture was obtained when Thrombokinase Geigy was compared with Thrombotest. When human brain thromboplastin was compared with Thrombokinase Gegy a good correlation was obtained in the therapeutic range. Thrombokinase Geigy is rather insensitive to variations in factor VII (4) Since factor VII 18 rapidly depressed at SEC. 70 THROMBOTEST Figure / 50 ======: 30 HUMAN BRAIN THROMBOPLASTIN 10 NS % FACTOR IX 10 14 18 42 ·ment factor X values arc about 8 lower than those obtained by Quick's method. Different mixtures of congenital factor X-deficient plasma with normal plasma showed a similarly good sensitivity to factor X for human brain thromboplastin and Thrombokinase Geixy. The same investigations as done on factor IX were carried out on a pure congenital factor VII- Conclusions : 1-The simplest methods are Thrombotest and Thrombokinase Geigy. 2-Factor IX activity is not detectable by any of the four methods. Thrombotest shows the fastest dectcase in the Geigy. S-All four methods take up the same time. Human brain_thromboplastin, Thromboplastin Roche and Thrombokinase Geigy cost much less than Thrombotest. We hope that the cost of the Thronborcst reagent can be reduced, because ut is too crpensive for routine work, even if half of the original amount advised 19 used. REFERENCES !! These differences appear only at the beginning of therapy. Later the results are parallel. Wc used the four thromboplastins in assaying factor II and factor VII in our specific one-stage mahal. Th. not 6, I 1-1 ocliger, A. (1959) Schweiz Med Weslicer 88, 639 5-Owren, PA, (1959) Lancet 1, 754. (1959) 11, 1036, (1960) I, 436 100 THROMBOTEST Figure 4 HUMAN BRAIN THROMBOPLASTIN THROMBOKINASE 'GEIGY' THROMBOPLASTIN 'ROCHE' 50 5 10 15 20 5 ZZZZ ܬ 3 6 9 12 15 MARCUMAR mg ·Therefore we studied the sensitivity of Thrombotest portions. The mixtures were diluted 1:5 to obtain to different factor IX concentrauons. The results plasma clatung factor leveis comparable to those are shown in Fig. 1. Normal plas.na was mixed in agricoagulant therapy. The curves fron Thrombowith that of severe hæmophilia B in different protest and Quick's method are identical. Therefore Thronbotest is not sensitive to factor IX. Pauents CLOTTING with low factor IX activity during anticoagulant TIME therapy did not show especially low Thrombotest values as compared with Quick's method, Fig. 2 shows results with the four different thromboplastins plotted against the factor X values in the same samples. Factor X acts both in the SEC. intrinsic and extrinsic systems. When a method is sensitive to factor X it is sensitive to (at least) one factor of the intrinsic system. Of the four preparafionis correla at the more s... AULU AA. 1 IULA Lerill treat- 80 THROMBOTEST Figure 3 1 604 HUMAN BRAIN THROMBOPLASTIN THROMBOKINASE 'GEIGY' 40 THROMBOPLASTIN 'ROCHE' 20 10 14 2 6 18 FACTOR VII ·ment factor X values are about 8 loser than those obtained by Quick's method. Different mis tures of congenital factor X-deficient plas.Ta with normal plasma showed a similarly good sensitivity to factor X for human brain thromboplastin and Thro:ibokinase Gey. The same investigations as done on factor IX were carried out on a pure congenital factor VII- Conclusions : 1-The simplest methods are Thrombotest and Thrombokinase Geigy. 2-Factor IX actiszty is not detectable by any of the four methods. 3-It is important to have a central department to control and standardise the technique used in different laboratories. This is well organised in Switzerland by Gary. 4-All four thromboplastins give good results with capillary blood, the most reliable for capillary blood being Thronborest and Thrombokinase Geiss 5-All four methods take up the same time. 6Human brain_thromboplastin, Thromboplastin Roche and Thrombokinase Geiry cost much less than Thrombotest. We hope that the cost of the Thronbotest reagent can be reduced, because it is too expensive for routine work, even if half of the original amount advised is used. Thrombotest shows the fastest decrease in the clotting system because it is more sensitite to factor VII, whereas Thrombokinase Geigy shows the slow est. The results with human brain thromboplasun and Thromboplasıın Roche ? berheers. These differences appear only at the beginning of therapy. Later the results are parallel We used the four thronboplastins in assaying factor II and factor VII in our specific one-stage methods (3). The only variation was the change of REFERENCES 1-Bachması, F, Durhert, F and Koller, F..(1959) Thromb Duathm. 2. 24. 2-Frechter, . (1910) Schwer. Het. Weber. 70, 259. 3-Koller, Loel. v,A and Luchert F. (1951) Acta Izmal. -Lorliger, A. (1938) Schwesz. Met. Hesheer. 89, 639. s-Outer, (1959) Lancet 1, 754. (1959) 11, 103, 11960) 1, 436 100-4 THROMBOTEST Figure 4 HUMAN BRAIN THROMBOPLASTIN THROMBOKINASE 'GEIGY! THROMBOPLASTIN 'ROCHE' 50 5 10 15 20 5 3 po 9 12 15 MARCUMAR mg 45 ·The Choice Of Test For Control J. M. MATTHEWS, R. SHULMAN, A. TUXFORD and W. WALKER- Department of Pharmacology and Therapeutics, Queen's College. University of St Andrews, and Clinical Investigation Unit, Maryfield Hospital, Dundee, Most laboratories in Great Britain still use the 1.Lh which the maintenance of the most intensive therapy compatible with safety may be relatively more important. TL.7.-.,-.:. f IULHILILU. LODAS. -11-. ---LIA VAT fudgment are more important than the marginal advantages conferred by newer techniques It could be said that for twelve years the Quick test has worked well in hundreds of laboratories and in hundreds of thousands of patients. The theoretical and potential advantages of Our own experience with the Quick test for Thrombotest are obvious. Its sensitivity to all four control, obtained in a relatively small hospital, is factors depressed by coumarin drugs, under the conditions of Lire really indicated in Figs. I and 2. Of the two deaths from effective treat hæmorrhage only one could be ascribed exclusively while the use ety, able activity of the [rol Certain difficulties, however, were encountered A DECADE OF ANTICOAGULANT THERAPY MARYFIELD HOSPITAL, DUNDEE 1950-59 1957-59 Number treated 1580 617 Wies, an10 Silviu IIUL VL wuwu LV vuuu. great technical advance that Thromborest has made. Serious hæmorrhage 5 2 Further experience has eliminated the diffi- (Non-fatal) (0.32°) (0.32%) culties we found in our first trial (Matthews and Walker, 1959). The chief difficulty with capillary Fatal hæmorrhage 2 None blood was the large volume required, 0.1 ml (0.12 ) not only did some of our patients declare their preference for a venepuncture, but a free flow of Figure ! to anticoagulant therapy; the other was from cerebral hæmorrhage in a very ill patient with a finger Moreover, the present cost of the reagen! recent cerebral infarction Two of the five serious hond encficinn mok place the two y of the 2-rate in phlebothrombosis and pulmonary embolism in treated patients (Fig. 2). All three of the patients who died were very ill before treatment was started, YENOUS THROMBOSIS AND PULMONARY and two were over seventy and had pre-existing cardiac and cerebral discase. On the other hand, EMBOLISM MARYFIELD HOSPITAL, DUNDEE, 1957-59 Deep Venous thrombosis 136 cases (F 104, M 32) There is little doubt that a test possessing Pulmonary embolism 45 cases greater sensitivity and giving a better warning of (F 35, M. 10) likely hæmorrhage would permut more effective control. This has become all the more important Deaths 3 (6.696) with the increasing use of anticoagulants in arteria), especially coronary artery, disease, in wluch the Figure 2 bencficial effect has been harder to establish and in 46 IVY AL andad 1 Ann·stored in siliconed or Lusteroid tubes. Although for blood there is no doubt that some silicones and some plastic tubes are more effective in preventing contact activation than others, and that the collection of blood in a glass syringe, unless it contains citrate, may sometimes cause activation or other changes even when the blood is stored in plastic tubes (Fig. 3) We have found three methods effective in preventing such changes: (1) as suggested by Owrco, the blood is measured into a syringe already containing the appropriate volume of citrate, mixed and I. Lid the cost is halved, and for the control of an outpatient would be no greater than the cost of Clinitest tablets for urine-testing in diabetics. The results correlate well with those obtained with os ml, of reagent. With regard to the changes in stored venous shows the good correspondence between the capillary test and delayed tests on stored venous blood collected by the second method. With any other method quite large variations in the coagulation activity of stored venous blood have been found quite frequently, and although at the end of 24 hours such Figure 3 100 CORRELATION : CAPILLARY AND VENOUS BLOOD STORED 3 HRS. IN POLYSTYRENE 80 TUBES CAPILLARY 40 BLOOD 20 % 20 40 60 BO TOO VENOUS BLOOD COLLECTED IN GLASS SYRINGES ·variations in or near the therapeutic range would L1_n 1- method is preferable to measuring the volumes in the syringe, since this is more prone to inaccuracy and rather less effective in preventing activation. Cinca natin sterilisation best method blood from Our conclusion is that routine laboratories should use Thrombotest with capillary blood for control of the increasing numbers of patients on long-term outpatient therapy. This was the main purpose for which the test was designed, and for which its advantages are most fully exploited Any increased cost of reagent will be offset by the saving nel--L I 111---1 4+ CUIL Tyth an hona L.. tessor 1. G. W. Hill we had the opportunity of studying a patient who bled on a day when his Quick test was at the upper limit of the therapeutic range. The P and P result was just below and the with the precautions mentioned may be used Otherwise the Quick test will continue to be used for inpatients but unless local difficulties are great, Thrombotest I preferable. For postal samples Thrombotest on blood collected in the manner 10G CORRELATION CAPILLARY AND VENOUS BLOOD STORED OO 3-24 HOURS IN Figure 4 . o POLYSTYRENE TUBES GLASS TUBES 80 70 60 CAPILLARY SO 40 00079 o 30 o 20 000 10 OO O So 60 30 40 70 O 90 10 20 100 IN VENOUS COLLECTED NYLON SYRINGES BLOOD 48 ·Figure 5 100 QUICK TEST RATIO 90 2 2 3 17 16 2 3 17 2 2 24 IS CENT JO PEA 70 P AND P w. THROMBOTEST 60 THROMBOTEST 50 OR 40 P AND P 30 20 10 CHLORPHENINDIONE MG. 5 10 15 20 25 30 35 40 DAYS OF TREATMENT described is probably preferable to the Quick test. Certain laboratories with special facilities may prefer to continue with other techniques such as the ” and P method or (for in-patients) thromboclastography, REFERENCES Batthews, J 1, and Walker, W (1959) Lancet 11, 1159 Owren, PA (1959a) Lancet u, 754 Owren, PA (1959b) Farmakoterapi, 4. 92 The chlorphenindrone used was supptred through the courtesy of Geigy, Ltd, Manchester.

·DISCUSSION If the extrinsic system is blocked, as in congenital factor VII deficiency, the Thrombotest method becomes a test only for the intrinsic system. The canci. - Dr BIGGS Since the one real danger of anticoagulant therapy is hæmorrhage, the main duty of those controlling therapy is to ensure freedom fron hæmorrhage. There are two main problems. One is the organisation of the service, and much could be done to improve this. The exact nature of the service will depend on local custom and personnel, but, in relation to these, serious attention should be paid to co-ordination of laboratory and clinical work. The second problem is the choice of technique, Many techniques have been used į some nra hann..1... ot a normal standard plasma, the upper curve for a plasma totally deficient in factor IX. The difference in times illustrates the sensitivity to lack of factor IX. The sensitivity is low at 100% (undiluted plasma) because the normal extrinsic system at this concentration gives a rather short clotting time. Thrombo. clinicians are asking for as a wartung against a bleeding tendency during anticoagulant therapy, caused by a disproportionate reduction of factor ix. Thant-prin TYS.STY- -** - - control plasma must be assured Jup) wupi Luwu au 15. Lul wou dudpleu Zollinger's results may be caused either by to use in small laboratories The other, the Thrombo differences in technique or possibly by different test, has most of the advantages of the P and defects in the plasma of his patients and ours. It 18 method, and is based on the use of a standardised important to clarify the cause of these divergencies commercial reagent I think that this Thrombotest method is the best for general use Dr BARHAM CARTER I have found certain difficulties in the organisation of an antiby preparing correct correlation graphs for the coagulant clinic The question of clinical responsi preparations used, and Zollinger has shown that bility arises-should the pathologist accept this as good results can be obtained in expert hands The part of clinical pathology, or the physician, or, per problem is, however, to have a simple test which haps best of all, both equally ? gives reliable results in less experienced hands, and I have found recurrent cerebrovascular disease for this the Thrombotest method was devised. difficult owing to confusional states and the difficulty 1.0 --- of persuading relatives to accept responsibility for and the tablets meas I have found Quick's one-stage prothrombin for I time unreliable on many occasions, and think that We always recommend that technicians or doctors nu price is too high for increased reliability, unfamiliar with coagulation tests should have practica! I wonder whether the results of all the tests instruction before undertaking anticoagulant therapy could be expressed in time rather than percentages, and control. Many visitors have come to us for this but perhaps this would not give an accurate answer instruction, which also includes the technique of administration of anticoagulants and organisation Professor OII'REN - Zollinger's finding that When venous blood has to be used, I would recommend the method advocated by Walker, to collect the blood directly from the needlc into a plastic tube containing anticoagulant and marked for the correct amount of blood. We have found this most satisfactory. The answer to Barham Carter's question, whether the pathologist or clinician should do the test and guide thc dosage will depend on local circumstances. Our experience seems to show that the best results are obtained when both testing and dosage done by the same person, Results should always be gisen as percentages read from a correlation graph prepared for the method and thromboplastin used : scrial dilutions 50 ·120 2700! 80 "THROMBOTEST" P-P METHOD Frgure i-Sensitivity of Thrombotest to dispro. portionately excessive reductions of Factor IX. Plasma from a patient with hamophilia B (Christmas disease) was mixed with normal plasma in various proportions. The mixtures were assayed after dilution 5 with ad. sorbed human plasma In order to imitate conditions during anticoagulant therapy. The concentrations of prothrombin, Factor VII and factor X. these lore, 120% in all samples tested. CLOTTING TIME IN SEC, QUICK'S METHOD 20 4 8 12 16 20 ANTI-HEMOPHILIA-B FACTOR occurring in stored blood, is not suitable for postal samples. of a normal plasma are tested, adsorbed plasma being used as a diluent. This is the only way to compare results from different methods and thromboplastias. * Prothrombin" umcs given in seconds are mislead- 20g even if the time for "normal" plastia is also given, because there is no correlation between prothrombin ratio and the actual clotting activity A ratio of 2 (or index of so, twice the normal time) And he artela 200 HEMOPHILIA B 3100 CLOTTING TIME IN SEC. 50 NORMAL 30 be lowered as soon as increased production makes thus possible. Dr SAMTAMA - We used Quck's test with human brain thromboplastin and Owren's Thrombotest in the last year We often use thromboelastography as well in parallel Our unpression is that global coagulability is sometimes a better measure of the efficiency and safety of treatment. Two patients, for instance, may both give a Quick result of 10 but the TEG be very different, indicating a hush probability of bleeding in the one and a very remote probability of bleeding in the other. Naturally, concurtent control by TEG and Thrombotest is very expensive. Dr HARTERT -I should mention that thromboelastography, being very sensitive to changes 51 100% 10 20 30 50 PLASMA DILUTION Figure 2-Dilution curves (correlation curves) for normal plasma and hemophila B plasma (Christmas disease plasma) ·failure of Thrombokinase Geigy to measure the decrease in factor VII is taken into account, it works as well as Thrombotest. The differences found in the factor IX sensitivity of Thrombotest are very interesting, and the reasons obscure. We carried out our investigations several times with different and severe cases of Christmas disease, with the same result. Dr Loeliger has had the same experience (personal communiation), Dr WALKER - Should not the calibration curve for Thrombotest be extended below 10% to give a better indication of the degree of excessive reduction in coagulability ? Professor OWREN— Yes, this is being done. 90 80 Coagulation time in sec. A test Figure 3-Correlation graphs for Quick's prothrombin time using different thromboplastins The curves are prepared by serial dilutlons of a normal standard plasma In adsorbed plasma 30 20 B 23" 16" 13* 10 20 30 40 50 60 70 80 90 100 10 per ceny Prothrombin Prothrombin ladex 50 40 (2 x norme) (2normal) 10% 1 Acetone dehydrated rabbit brain z MO Simptastin + Human brain (untreated) 5 Soluplastia Human brain (dehydrated) 247 30% 30 % 17% 11% 20% 19% 12% 3 70 6 Figure 4-Correlation graphs for different thromboplastins prepared as In Figure 3. 60 SO 40 Clotting timu in seconds 10 20 30 40 60 10 90 90 100 *Prothrombin activity in por cent of permet 52 & ·Anticoagulant Therapy In Myocardial Infarction R. B. HUNTER-Department of Pharmacology and Therapeutics, Queen's College, Dundee Pioneer work on the use of anticoagulants in Q.-.Tud times a suggestion, that anticoagulants prevented a further myocardial infarction. What, in fact, a physician was saying, therefore, when he said he could select pauents for treatment was that he could foretell which cases were likely to develop thrombo embolic complications. Thus mortality and the incidence of thrombocrnbolism were reduced and on the basis of this alone the anticoagulant therapy of acute myocardial infarction was jusufied. At the sa.nc time one had to Conor of ... . engendered much heat, controversy and discord. orie series will 2110LICI. During the earlier years of coumarin treatment, :-Lab al Today more is known about acure, myocardial and therefore the comparisons were of no value. No one knows the therapeutically desirable action of anticoagulants and the criterion for the introduction of new drugs has been duration of action, giving some advantage in administration, or consistency of action. To find out what is the imin hospital was required. portant therapeutic effect and to embark on a We learned that reliable information is only chemical adventure to try to improve that effect is likely to result from scientific investigation, that a most urgent matter, because anticoagulants of the the numbers of patients in the controlled trials coumarın type are by no means an ideal treatment. must be large and that meaculous attention must Professor Owren has now made the control of be paid to laboratory technique. This experience therapy safe and simple and capable of being conover the past fifteen years has shown, more than any Other, that the days of opinion and impression in medicine have gone and advances depend on serious scientific study. In addition, close collaboration between physicians and laboratory workers is required to get the answer. Morcover, we did not know what caused Inyocardial infarction was it always, in fact, the result of coronary thrombosis? We did not know to what degree the clinical syndrome was a simple with safety to all who need it. Dicture due to infarcted cardiac muscle or a com Almost everyone will now accept that in acute plicated picture of which the myocardial infarction myocardial infarction the only patients that should was only a part. be denied it are those with some clear contra- When the strictly controlled trials of treatment indication when the question arose as to whether in acute myocardial infarction were first reported, long-term anucoagulant therapy mught also give the clear finding was that the incidence of thrombobenefit, it was natural for physicians at this stage embolic complications was reduced, and because in our knowledge to choose serious and sometimes the drug used had been shown to be specific for tragic cases, particularly in the young, for long-term other conditions of thrombo-embolism the results treatment, often therefore cases in which serious suggested that thrombo-embolism played a more damage to the heart and the vascular system had important part in the mortality of myocardial already occurred, and again a confused situation was infarction than had previously been thought; that created, Cases were published which appeared to many of the fatalk benefit and others which did not. occurred TV 1 clinically pulmonar disease a suggest that anticoagulants produced benefit in this way. There was so clear evidence, though some- 53 ·The At complications and lower the mortality rate though there was no clear evidence that recurrence of myocardial infarction was prevented. If, therefore, one was considering the trial of anticoagulant therapy to prevent further myocardial infarction in patients who had recovered from an acute episode ---.--. to be greater in men under the age of 55. Under the age of $5 men suffered reinfarction at one-fifth of the rate of the control group and above the age of 55 at one-half the rate of the control group. An important point is that the difference between the treated groups and the control groups is unlikely to have arisen by chance, but the difference between the age groups above ss and under 55 may well be fortuitous. Too few cases were included in the Medical Research Council trial of females for any conclusions to be drawn. One of the difficulues of a trial of this sort 15 that whereas it starts with two groups that are strictly comparable with cases randomly allocated to the high or low dosage group, after two years the groups are by no means comparable presumably because further thrombo-embolic complications would be prevented. Tar fasthana -annnoncha ..--.L. 11.341 of disa has mat con 12 from the treated group at the end of two years to trials completed in the last few years Since one must accept this evidence for certain age groups, particularly men under the age of 55, the question arises whether thus is sufficient cvidence to say that in any disease of the vascular system which is known to be complicated by myocardial infarction the pauents should have anticoagulant risk. This is a very important study which is at therapy. I believe that this is so and that, since present being carried out by the Medical Research patients with angina of effort get myocardial Council. infarction, this in itself is sufficient justification for putting these patients on this form of treatment It REFERENCES Bjerkelund, CJ (1957) Acta Med Scand, 158. Suspl 330 Tine Per Dayal Can Vted, 47, 317. 959 sider, on these grounds, whether we are ethically Report (1959) justified in denying cases of angina of effort anticoagulant therapy. Similarly, other diseases of the Owren, P A (1954) Schweiz Med Wschr. 84, 822 heart and vascular system, known to be complicated Tulloch, J A and Gilchrist, A R (1950) Brit Ved J. 14.965 by thrombo-embolism and myocardial infarction, Wright, I. S, Marple, CD, and Beck, DF (1948) Amer should also be the subject of long-term treatment Heart J 36, 8or. r Ihre 51 ·Anticoagulant Therapy In Acute Myocardial Infarction A CLINICO-PATHOLOGICAL STUDY R. L. RICHARDS - Southern General Hospital, Glasgow dying within 24 hours of admission. In this group there were three instances of rupture of the heart and one of hemopericardium without rupture. The percentage of autopsies on the fatal cases i$ much the same in the other two groups. There are more males than females in the anticoagulant group, whereas the smaller non-anticoagulant group comprises an equal number. The mean age at death in the two groups is similar. As regards duration of thrombosis or its occurrence in an artery narrowed or occluded by atherosclerosis. Although we may be impressed by reports that the mortality is less in death are similar. In both groups most deaths were due either to myocardial infarction or to cardiac 1 - lestons found at autopsy, and are impressed by the instances in which the treatment has been followed by rupture of the heart or some hæmorrhagic catastrophe. anticoagulant group there were four deaths due to this cause. Particular attention was paid to the incidence of thrombus within the heart, pulmonary infarction, systematic infarction, cardiac rupture and any major hæmorrhage. The presence or absence of a recent thrombus in the coronary arteries was also noted. Coronary thrombosis often may follow rather than precede the cardiac infarction (Branwood and Montgomery 1956), and it seemed desirable to find MATERIAL Tha pu. Am nan NAAR Table 1 mortality in the three periods is about the same; roughly one-third of the patients de during their hospital stay. This figure has remained remarkably constant during the II years It was our policy L---L- POST-MORTEM FINDINGS Anti- Non-Anti- Coagulang Coagulant Group Group Number o Number 0 Autopsies 52 100 25 100 Thrombo-embolic lesions Mural thrombus 13 25 60 Recent thrombus in coronary artery 15 29 14 56 Pulmonary Infarction 3 6 7 28 Systematic infarction 6 11.5 5 20 Cardiac rupture S 10 0 0 Hæmorrhage 8 15 0 0 15 Heparin was given zatrayenously for the first 48 to YEAH out whether the incidence of recent thrombi in the coronary arteries was influenced by anticoagulants. Unfortunately, the veins of the legs were not routinely examined, and, therefore, no satisfactory data on the incidence of venous thrombosis can be given. There is a lower incidence of 11 th . RESULTS It is improbable that anticoagulant therapy would influence the pathological findings in patients 55 ·Similarly, the relationship of the cardiac ruptures and the hæmorrhages to the therapy can be considered (Table 2). Most occurred during the first ten days of treatment, but some later. There is no evidence group and five instances of cardiac rupture. There were no such lesions in the non-anticoagulant group. Because of the few autopsies in the nonanticoagulant group, it seemed desirable to see whether the results could be regarded as representative. A similar analysis of the 24 autopsied fatal cases in McCluskie and Seaton's non-anticoagulant group (1959) give results very like Table 1. In my own series, one can consider the relationship of the thrombo-embolic lesions in the anticoagulant group to the anticoagulant protection Table 2 interesting patients were those who had hæmorrhagic lesions in the lungs. All four had the clinical and radiological picture of pulmonary infarction, but at autopsy they had hæmorrhagic, oedematous lungs .:: Murce for ; '', N'C'. 1., these .....As tases of ::.. --- CARDIAC RUPTURES AND HÆMORRHAGES LESION CONTROL Cardiac rupture TH * = Septal rupture Hæmorrhagic pericarditis, lungs and pleural effusions Hæmorrhagic lungs Cerebral hæmorrhage Melana Renal hæmorrhage Hæmorrhage into psoas Satisfactory control Control unsatisfactory prothrombin me too low One excessive prothrombin time. = More than one excessive prothrombin me incidence of mural thro:nbus in the anticoagulant group is lower than that recorded in any, and of pulmonary and systemuc infcrction than a most untreated series reported Thus increases the probability that the low incidence is due to a genuine reduction brought about by the therapy, Probably no conclusions can be based on the data regarding the incidence of recent thrombi in the coronary arteries. Although such thrombi were * 1.-c-.-d. donna + # given These lesions occurred in 30 patients. Of these - Ant. h. •LahARI This study does not help to clarify the relationship of rupture of the heart to anticoagulant therapy. Although there were five instances of cardiac rupture in the anticoagulant group and none the nonanticoagulant group, there were four cases of rupture in the untreated patients who died within twenty-four hours of admission The other hæmorrhagic lesions, however, represent a hazard of anticoagulant therapy, and major hæmorrhagic lessons occur even where the dosage has not been excessive. It is important to realise that hæmorrhagic lesions in the lungs may present a picture clinically simular to pulmonary infarction were found in cases where the control was inadequate. The most significant finding is that pulmonary infarction was found in only one instance in which the patient was adequately protected REFERENCES Вгати ооd, 4 1, and Montgomery, GL (1956), Scot McCluske, J AI, and Satur, DI (1990). Gent Vel Ved ), 1, 367 J. 4. 305 Conrad, F G., and Rothertuch, a O (1959), Arch Intern Richards, R 1. (1933), Ibid. 3. 235 Med. 103, 411 Wright, I S. Marple, ( 1), and Lech, DI (1954), Gluech, H 1. Rider, H11, and I'ascermat, P (1956), cardial infarction its clinical manifestations and treal Circulation, 13, 5$ mest with anticoagulants, New York Grute and Stratton Ler, T and O'Neal, R II (1956). Amer J Wed, 21, 555 56 ·Anticoagulant Therapy In Peripheral Vascular Disease GEORGE SMITH-University Department of Surgery, The Western Infirmary, Glasgow TYPE OF DISEASE Unilateral Bilateral Tota1 High occlusion Z 2 In peripheral vascular disease there is little argument over the use of anticoagulant drugs in the acute phase of vascular occlusion. The indications for long-term treatment are, however, much less clear. Most of the patients have atherosclerosis, with thromboangeitis obliterans next in frequency. In the latter there is threat to limb, in the former to life. In the atherosclerotic group, more than 95%. of the patients have intermittent claudication. The fate of these $ not completely known, although subject to various studies with many disparate conclusions. Thus Hines and Basker (1940) and Spaulding (1956) followed groups of just over temoro.poplitesi 32 29 41 Distal 2 A symmetrical 9 9 16 58 Table 2 ATHEROSCLEROTIC GROUP atheromatous disease of the leg Vessels By this means of preventing secondary thrombotic occlusion, the prognosis might be improved even in the presence of the underlying continuing atheroma. 32 patients were placed on long-term phenindione treatment. These are in two groups. The first 16 showed only acheroma with occlusion at the femoro-popliteal area. Endarterectomy was performed with heparin as the anticoagulant in the operative and immediate post-operative phase. Thereafter phenindione was given for periods up to one year. The second group consists of 16 consecutive patients with severe atherosclerosis and the worst prognosis based on the criteria already menuoned bilateral involvement, age over 50, bad famsly history and evidence of generalised disease. From the first group, it was hoped to answer the muretion whether or n. Analyst. of 58 Cases (5 females, 53 males). Age. Under 40 41-50 51-60 Over 60 year! 9 30 15 L- Table 1 Table 3 MORTALITY colleague Dr. Richards who is doing most of the work of thus investigation. The original five year survey concerned 58 patients with atherosclerosis and minimal or no nutritional changes in the legs who first attended with intermittent claudication. The sex, age and distribution of the disease 12. 17 of the n 13, the age and is had ad a diastolic 17 patients died during follow up period (29%) Thala --- CAUSE OF DEATH Myocardial infarction 7 Cerebrovascular accident 3 Congestive heart failure Z Diabetes, tuberculo919 and gangrene 1 The changes which occurred in the 45 patients who survived the five-year period are listed in Table 4. Of these 17 had angina of effort and 13 had myocardial infarcation 2 had congestive heart failure, 3 had cerebral vascular incidents with neurological sequelæ; 13 had a drastolic blood press sure over 100 mm Hg. 31 survivors had evidence of generalised vascular disease and in only 10 was the disease apparently still confined to the lower limbs From these results there is a good case for long-term anticoagulant treatment in such patients, particularly those over 50 years of age with bilateral 57 Carcinoma pancreas Perforation of peptic ulcer Unknown ·ary thrombosis and advancing atheromatous de Ten cases fulfil the diagnostic requirements generation in the production of arterial occlusion, In s the presentation was distal ischemia and toe The drug has been given for periods ranging from skin breakdown. In these lumbar sympathectomy 6 months to 2 years (Table 5). with local removal of dead tissue was carried out. It appears that phenindione prevents thrombosis Thereafter treatment with phenindione was begun in a medium-sized artery devoid of normal intima and continued until healing occurred. Should SURVIVORS 41 cases Unchanged or improved is slowly stopped over a period of three weeks. 27 Worse Unilateral bilateral 3 Acute arterial occlusion 3 Definite deterioration 5 Amputation 3 to liuc ue patents over to middle 11e waren wie disease is self-limiting, In acute occlusive processes heparin is the undoubted drug of choice. This group includes der af einke narinn Arad - . Are 41 Table 4 but the important change is not so much secondary thrombosis as unrelenting encroachment on the arterial lumen by the athero.natous degeneration. The efficacy of heparin in acute vascular occlusion is also underlined in six patients with phlegmasia cærulea dolens in whom the drug given intravenously with elevation of the limb was successful in re-establishing venous drainage of the leg in three of these cases, phenindione had been given previously for deep vein thrombosis and 1! would appear that the further venous occlusion had occurred in spite of this. In a further five cases seen at the stage of venous gangrene, pheandione had to 10 years. Their ages range from 18 to 35 years, 11 hain haln - Afarta T- de ha (Hutchison). Table 5 long term anticoagulant therapy (phenandoon Group Nurnber Age Crumplications to survive since the actual tissue loss is frequently less than initially appears likely. 16 39-59 Endarterectomy fur Isolated lesion at lemon-popliteal arta Reucetusjon at 2/12 Myucardial infarct death at 8/12 1 14 52-69 Severe generalised #theroscleros Anputation for gangrene (Biateral in 3) Myocardial infarct, cerebral thropsbosi.. myocardial infarct. death REFERENCES lines, CA, and Barker, AW (1910) Amer med Sci. 200), 717 Hutchison, HI 1. Department of Pathology, Western Infirmary, personal communication Spaulding, 11 B (1956 Cand Vfrd Ass 1 ,,75, tog Stammers, 1993+) J Bone Richards, KL. (1937) Drit Nec JJ1, 1091, Ss ·Prophylaxis By Anticoagulants (Coumadin) In Surgical Disorders M J. LARRIEU, M. SAMAMA, E. TERRIER and CL. FRILEUX-Saint-Louis Hospital, Paris Coumadın (Warfarin) was used for the prophy A stable therapeutic level was reached within laxis of thrombo-embolism in orthopædic operations $ to 8 days, and was casily maintained in 22 patients on 31 cases--19 Women and 12 men. but less easily in 6, who were elderly as well as poor Their age varied from 20 to 90-21 being over tisk. 60. The average length of treatment was 57 days, most being treated for 20 to 60 day's The patients were immobilised throughout. Coumadın was given in a single daily dose. The initial dose was to mgm. for the first two periods scre exceptional. days in younger patients, 5 mgm. in patients older In parallel with Quick's test and heparin than so, and 2.ş mgm, in poor risk patients or in tolerance we did 180 Thrombotests. We noted patients older than 60. The average maintenance the simplicity of the method, its reproducibility, and its extreme sensitivity. The results accord with those of Quick's test (Fig. 1), but the values of Thrombotest were always lou er. Simultaneously, we measured coagulability d using the thronboelastogram and the heparın w.piex Val Sludll Idulur dhiu ProCONVLUL) tolerance test. We feel that these techniques more assay. The last three investigations were not carried accurately anticipate the risks of hæmorrhage and out routinely. thrombo-embolism. The prophylactic range of 30 Control tests were carried out three times a 10 40%. (Quick) or 20 (Owren) corresponds to a week in the first week, twice a weck in the following moderate hypocoagulability wecks, and once a week when the treatment extended We had a thronbophlebitis in one patient, over 40 days. w owing to inadequate therapy. In 4 cases an abnormal bleeding tendency was attributed to therapy: melæna in a patient aged 63 with an undiagnosed carcinoma of the alimentary ...1 e a 30 Results TON. 49 YEARS counted for by the age of the patients and their poor state of health, and by premature anticoagulant therapy after surgery. We used Coumadın in three patients who were resistant to phenindione and/or Tromexan. A dose of 10 to 12 5 mgm. of Coumadın gave a definite hypoprothrombinæmia. 50% 30% QUICK 20% OWREN 0 10 VARBARINE 5 5 m. sms 59 DAY ·Prophylaxis By Anticoagulants After Gynaecological Operations nrand 195 SELECTION OF PATIENTS A. C. TURNBULL --Department of Obstetrics and Gynæcology, Queen's College, Dundee Pulmonary embolism is a common cause of embolism after - death after tions be prophyl vpulalionis, despite " early ambulation" and daily examination to detect and Many factors are thought to be associated with treat venous thrombosis as early as possible. increased risks of thrombo-embolism, but analysis showed finally that cases could be selected for treat- The difficulty is, of course, that the thrombosis ment on the basis of only three factors-age, body is often silent and insidious, in two-thirds of the weight, and past or present " venous complications. in the I Age. sigris we Age was found to increase very considerably transient the risk of death from embolism, although it only Marks et al (poc.) antaraincreased slightly the in WW cases of - UU ,eu ال۔ ادب . - Prophylactic Anticoagulation. A trial veinn - Aberd given post-o prothr cent; 30 per stage 1.. un mas useu. While the incidence of deep Vein thrombosis was the same in both groups (15 per cent.), there were no emboli in the 201 cases treated compared 11h * uil yuwiger group. Of the 30 cases of fatal embolism between 1937 and 1954, twenty-two were so years or more, five were between 45 and 49, and only three under 45. Obesity. The weight-height ratio was ricade om onder of 1 wh bet wit. tria. - wu Hut uevelop any thanh 1). Fifty per cent of controls were of "normal weight, compared with only 24 per cent of com plicated cases. Of the 19 complicated cases of normal weight, only four (20 per cent.) had an embolism and none was fatal, but of the 60 obese cases, 24 (40 per cent had an embolism and six died. Weight or height had not been measured in all cases, but of the four fatal emboli whera H. اداری دهه هه Y5 110 1101e biceding in treated cases than controls Severe bleeding, requiring blood transfusion de 1 Precious Thrombo-embolism and Varicose Vans. Accurate information about these fortare um llcllumage. Selection of Cases for Prophylacrc Anticoagulant The trial choquert that prophulans- The possibility of selecting " high risk" cases or the 67 cases with previous implications, was studied in three groups of cases - first, 3,040 ten (13 per cent.) developed comph sons, three times as many as in cases without pre 1 us trouble. patients who had a major synzcological operation between 1951 and 1954, of whom 119 developed Pulmonary embolism was more than twice as common thrombo-embolism ; second, 30 patients who died of and one case was fatal. Varicose veins were found in ·almost one-third of all cases, and the incidence of lactic anticoagulants after a major synacological venous thrombosis (6 per cent.) and of embolism operauon should be made only after careful assess- (4 per cent.) was higher than in the group without ment of the patient. varicosities (2.3 and 1.7 per cent. respectively). of the 370, cases treated, 33 (14 per cent.) had Morcover, of the thirty patients who died from bleeding and this was severe, necessitating blood etbolism between 1937 and 1954, only three were transfusion or further operation in 11 (3 per cent.). under 45 years of age, and all three had a history of There were no deaths from hæmorrhage. In the Previous deep venous thrombosis. untreated group, only 13 of the 465 cases i per cent.) had any bleeding at all. Criteria for Selection of Cases. If, between 1937 and 1954. prophylacuc anu. In the 1955-1956 trial, 37 per cent of pelvic foor repairs had bleeding following phenindione coagulants had been given to all women who were both 45 years or more and overweight, or who had treatment, so that an incidence of 14 per cent. in this larger series shows that the risks have not proved Varicose veins or a history of thrombo-embolism, and of the treatment had been effective, at least 25 R$ great as expected. of the 30 deaths from embolison would have been Conclusions. prevented. (Body weight was not known in the other five patients, although all were over 45 years of age.) The incidence and mortality of thrombo-embolison following gynæcological surgery can be It was decided to introduce a policy of prophy. greatly reduced by prophylactic anticoagulation lacuc anticoagulation in both units, using these with pherundione. Chalmers ct al (1960) have criteria to select patients, recently reported similar conclusions following five Preliminary Results in Anticoagulant Prophylaris in Selected Cases. This policy has been in practice since April 1957. Phenindione was given from the Ist to the ing the use of prophylacuc anticoagulants. 12th post-operative days and the prothrombin concentration kept at 25-30 per cent. The results refer to pelvic floor repair. (Details are not yet available for other operations, but it is understood that no deaths from embolism have occurred.) The incidence of thrombo-embolism is much less in the 1957-1959 series (1.5 per cent.) than in 1951.1954 (4 per cent.) when there was po prophylaxis, but the striking difference between the The policy is practical, safe and effective, two groups is in the number of fatal emboli, with only one death la 835 patients between 1957 and I om groteful to Professor Dugald Baird for permission to publish cato obta ned from investigations which were, ont are being carried out in his department. I am also indebted to Dr A Herriot, who supplied me with the details of the results of treatment in pelvic floor repair coses between 1957 and 1959 dercioped in the 465 untreated cases, of whom one died of embolism. Actually, this patient should have REFERENCES been treated prophylactually because she had severe bilateral varicose veins. Chalmers, I) C, Marks, J. Bottomlev. JC, and Lloyd, O. (1960) Lancet 1, 220 The criteria, therefore, for selecting cases for Varhs, J. Truscott, RV, Withycomb, 1 F. R. (1954) treatment appear to be adequate and satisfactory Lancet 11, 787 in practice, but the decision NOT to give prophy. Scritt, S, Gallagher, G. (1959) Lancet 11, 991 .

·The Role Of Anticoagulant Therapy In Cerebral Infarction A BARHAM CARTER --Ashford Hospital, Middlesex The role of anticoagulant therapy in cerebral over 70 years of age and unless they had a persistent infarction has been a source of controversy since diastolic blood pressure over 120 mm.Hg. Five patients were rejected on the above grounds. The patiente para avamined fix month after infrontian ate short-term measure in acute cerebral infarction to prevent the spread of the infarcted area and so conserve its marginal viable tissue, or as a long-term treatment to prevent further cerebral episodes in patients with a history of previous and recurring cerebral infarction. Current Views. The results of clinical trials have so far remained as inconclusive as that of Hedenius and owing to possible dangers in treatment views have been strongly divergent. The studies of Sibley (1957), Moyes (1957), Peterman (1959), and their colleagues showed that dicoumarol increased the hæmorrhagic, tendency In the group of patients suffering from atheromatous cerebral infarction without obvious embolism J.L-1L (1960) have shown that the routine use of antiof patients for a further trial of anticoagulants on coagulants in unselected patients with clinica) the grounds of slowness of onset and incompleteness of lesion It was soon found that speed of onset cerebral infarction not due to cerebral embolism was almost impossible to measure, and patients were therefore selected who showed progression of clinical signs while in hospital or who gave a clear history of progression of symptoms immediately before admission and whose lesion was by then complete. In addition the diastolic blood pressure had to be persistently below 120 mm of Hg., the cerebrospinal fluid clear, age below 70 with few exceptions, and the history free from symptoms of peptic ulcer or hepatic disease. In the four years 1955 to 1958, 83 patients out of 277 admitted with the diagnosis of cerebral thrombosis satisfied the above criteria, but three had to be withdrawn as a history of visceral bleeding energed within a few days of treatment of the remaining 80, four of fatal cerebral hernorrhage was greater in the were found to have been wrongly dragnosed, by chance treated than the untreated groups. This trial, how two in each group. Three were found at autopsy ever, included some hypertensive patients. The Series My own experience, some of which has been previously reported (Carter 1957, 1959, 1960) The first 10 series of patients 1on treated from The second is a its from 1935 to evious attacks of hæmic episodes Acute Cerebral Infarction. of an nationte raken from mitted to 1002 Ashfc to 1958 (Tab suffering from onsidered separately ( 1 UIC 23. coagulants-about one in seven. Anticoagulants were given to patients with clinical cerebral embolism in 1994, unless there was Recurrent Cerebral Infarction. The problem of recurring cerebral ischamia blood in the cerebrospinal fluid, unless they were 62 Anine ·478 524 with or without infarction was investigated from 1955 to the end of 1998. The series is a very small TABLE 1 onc, as only 32 patients below the age of 70 could be found who gave a history of two or more cpisodes Cases of " stroke "admitted to Ashford Hospital, of cerebral ischaemia or cerebral infarction, and 1952-58, showing sex and etiology. whose diastolic blood pressure was below 120 mm. of Hg. Suteen of these patients were selected at No. of cases 1002 random and put on long-term anticoagulant therapy, Sex: and their progress in regard to recurrence and Niale survival conpared with the 16 controls, with a Female follow-up varying from one to four years. The recurrences have been divided into tko Ætiology: groups. Minor rccurrences are those showing Embolism 85 transient neurological deficits folloned by a return Thrombosis +25 to previous disability, or by only a slight worsening Honorrhage 492 as judged by the patient's ability to get about and look after himself. Major recurrences are those showing longer and more severe neurologcal deficits TABLE 2 often producing permanent deterioration. In the control group three patients had no recurrence, one Cascs of cerebral embolism, 1952-58. had minor recurrences, seven had a major recurrence and five had both. Twelve patients, therefore, Matral Stenosis out of the 26 had one or more major recurrences, Average and Fib. Mitral and of these sit died. In the treated group, six Cases Males Females Age rillation Stenosis patients were free from recurrence, three had minor recurrences, three had a major recurrence and four 85 32 S3 S3 39 S had both. Seven patients out of the 16, therefore, had one or more major recurrences, and of these Subacute ThyrotoxicOSIS four dicd. Cardiac Bacterial and If the patients with hindbrain ischaemia are Fibrillation Infarct Endocardius Fibrillation considered separately, there were four in the control 7 26 + series and five in the treated. Of the controls one had minor recurrences and three had maior and minor recurrences, two of these dying. Of the treated TABLE 3 Statistical comparison of results in the treatment of cerebral embolism P = 0.025. 1 Patents who Patients who Percent. d) VC wurun wwe 11dUIC 6). recovered or died or were age re- Year Total were improved not improved covering Afechamsm of Treatment. 1952 16 11 31 1953 18 33 Treatment-No anticoagulants No. of & 1954 1955 1956 1997 1958 IS 9 9 9 7 II 6 6 6 ھ لیا با ما تم 57 Treatment-Anticoagulants TABLE 4 Results of a controlled trial in patients with progressive atheromatous cerebral infarction, 1955-58 Total No. of Cases 76 Anticoagulant Therapy No of cases Recovered or improved Not improved or died No Anucoagulant Therapy No, of cases 38 Recovered or improved Not improved or died ·A. 16 ineffective and occasionally harmful. One must also recognise that apparently adequate prolongation of hn en hann hinna numure TABLES prevent I in this Results of treatment of lesion is incomplete tment of and if lesion is complete. No. of Patients. 76 Dangers of Treatment. Anticoagulant Therapy The dangers of anticoagulant therapy are three Lesion Incomplete Lesion Complete in number. The first is bleeding into and around Group A Group B Group A Group B the infarcted brain; the second is major visceral 17 7 bleeding producing systemic hypotension; and the Per cent. 77 23 56 44 third is the risk of treating intracranial hæmatomata with anticoagulants because of errors in diagnosis. No Anticoagulant Therapy These dangers are very real and stress the importance Lesion Incomplete Lesion Complete of strict control, and selection of patients, and Group A Group B Group A Group B above all of correct diagnosis, which may be more TO 10 9 9 difficult than is commonly supposed, and even Per cent, SO 50 50 SO investigation by, arteriography is often unhelpful. Group A = Recovered or improved. Fortunately in this series no major visceral bleeding Group B = Uomproved or died. was encountered during the last three years, but there were 17 minor hæmorrhagic episodes in nine TABLE 6 patients. Hæmaturia accounted for seven, epistaxis Results of anticoagulant therapy in recurrent cerebral infarction. TOTAL CASES-Follow-up I to 4 years. CONTROL GROUP : No of patients No recurrence 3 Minor recurrence I Major recurrences: Surviving Died 6 TREATED GROUP : No of patients 16 No recurrence 6 Minor recurrence Major recurrences : Surviving the term subacute cerebral infarction might be used Died to describe this type of case. It is clear from the work HIND BRAIN ISCHÆMIA. of Marshall and Shaw (1960) and from my own ful.a. ha CONTROL GROUP No of patients No recurrence None Minor recurrence Major recurrences. Surviving I Died 2 TREATED GROUPS No of patients No recurrence Minor recurrence progressive cerebral infarction and recurrent cerebral Major recurrences. infarction subject to the usual contraindications. Surviving None Although this decision is not wholly supported by Died 1 my past figures, there seemed sufficient evidence to justify it REFERENCES Carter, AB (1957) Quart Wed, 26, 335 Voves, PD, Vulihan, I. Mahm, G. Syte, (P hinant, J P 11957) Tor Mayo Clin. 32, 124 (1959) ibid. 23. 125 (1990) Lancet, 1, 345 Peterman, A., Wakt, KG. Styre, G , When, JP, De Vorster, G, Tausot, R (19571 Barr Cons, I ranc Yed, Wullahan, C'H (1959) ] Neuropath, 18. 37 1, 110 Sibley, WA, Vorleder. JHLaphan, W11957) Amer Ficha CV (1958) \eurology. 8, 311.. Sibley, Wed Sci. 234. hós Symonds, C (1957) Modern Trends in Yeurningy, London rand Vastola, EF, Frugh, A (1959) Yeurology. 7. 141 11-), CE (1959) Arch eurol Psychiat, S1, hey Millikau,1 ,. Wright, I S. Vellesatt, E. (1954) Lancet, 11, 825 Stahl Meet Mayo Clin, 30, 116 6+ I P 55) Pror ·Anticoagulant Therapy In Cerebrovascular Disease JOHN MARSHALL and D. A. SHAW -- The Academic Unit, Institute of Neurology and National Hospital. Queen Square, London, W.C.1. The cerebral circulation is peculiarly complex in cerebrovascular discase, started in 1957, it was and anatomically variable: with its efficient anas decided to consider the problems as they most torxotic poresti! across the id-line, 3 pressure monly present in seneral medical practice. By gradients are highly unpredictable and vascular - hann fcycral highly favourable lesions may produce their ischemic cffects at remote vascusites and even ouru ith the territory of supply of the ra! in- Iessel affected. Unfortunately kutle of the cerebral 3gnostic vasculature 13 accessible to clinical examination. unCCI LU! fic sub- We can palpate and auscultate the carotid arteries TOUDS represent a very $92dd your cerebroin the neck and inspect the retinal vessels; we can vascular disease, it was felt that the value of anticompare the pressures in the retinal attcries on the coagulant treatment should be assessed in the broad two sides with the ophthalmodynamomerct but spectrum of cerebrovascular cases encountered in these are the limits of our clinical assessment The everyday practice. effects of ischamia cannot be accurately detected The results of two trials will be described clettrically as they can in the case of the heart briefly, Details of design and composition of the muscle, and the value of contrast radiography 15 trials have been reportd (Marshall and Shaw, At present limited by its pertaning cxa nination 1960; Hill, Afarshall and Shaw, 1960). The first only of the major vessels. In a recent study of eighty trial was designed to assess the value of anticoagulant patients submitted to angiography following an therapy in patients who had suffered an acute acuic stroke," there were forty-six in whom to ** stroke" believed to be due to non-embolic cerebral radiological abnormality was demonstrated (Bull, infarccion. All patients were under 70 years of age; Marshall and Shaw, 1980). treatment was started as soon as possible after the The diagnosis of cerebrovascular lesions de onset of the attack but 72 hours was the maximum pends largely on the indirect evidence provided by Interval permitted. In additon to clinical diagnosis, the resulung neural damage, and the same neural all sligible patients had theu spinal fud examined, pathways may be interrupted and the same clinica! principally to confirm the absence of blood, and picture produced by a wide variety of pathological were submitted to cerebral angiography. Blood 10 processes. We are particularly bandicapped by an the spinal flud, severe hypertensive retinopathy inabulity to drstinguish with certainty between in or left ventriculas fassure excluded patients from the farction and hamorrhage, a point of obvious sign trial, the latter because it might demnand the use ficance in relation to anticoagulant therapy The Anne Peptic ulceration and bleeding presence or absence of blood in the spinal fluid, mindications. although of valuc, is not pathognomaruc. the basis Besides, we cannot predict with any accuracy pairs and the probable outcome of "stroke." In the carly allotted to treatment J. Those in stages we have no means of knowing whether the the treatment group were given an mutial dose of functional deficit is due to destruction of neurones 300 mgs. phenindione and, starting twelve hours or to reversible unpairment of discharge or conduc after their angiogram, threc doses of heparin tion and hence apparently dense deficits may show (12,500 units) were injected intravenously at sux* rapid and unexpected recovery. hourly intervals From the second day onwards The implications of these facts are obvious seatment was continued with phenindione, the for the assessment of anticoagulant therapy in Lunnhirs tuge (Quick one-stage test) being cerebrovascular disease. Clearly the variability in .hr imes the control progaosis makes controlled study obligatory The days and diagnostic uncertainty is perhaps most disturbing nd control in connection with the recurrent minor impairments h regard to of function--the 80-called " transient ischamic iotherapy attacks for these, particularly in brain-stem ► the trial, Critory, are readily mimicked by non-vascular twenty-six in the ulu...' twenty-five rut probable that the conflicting experience in the control, There were tweaty-tour males and come measure to twenty-sever females with approximately even ysical symptoms distribution of the sexes to the two groups. The wshing berween anatomical sates of the vascular lesions based on thaps the most Clinical and angiographic findings were very comind it may prove parable in the two groups. use of anticoagu Survival or failure to survive for six weeks from In uncontrolled the start of treatment was the criterion on which iplication of low results were based and these were analysed by means numerical incidence may not be apparent but of a restricted sequential procedure. We were Barron and Fergusson (1959) have drawn attention indebted to Dr Peter Armitage for the detailed to the large number of fatal cerebral hæmorrhages statistical design which is embodied in the sequen- In the extensive literature on anticoagulant therapy tial chart (Fig 1). Patients were considered in pairs in cerebrovascular disease The possibility will be according to the original random allocation to treat discussed later that these complications of treat ment and control groups. If both members of a ment are in fact the result of misdiagnoses of the pair survived or both died no preference was ind- Original presenting illness, cated in respect of that pair for treatient or control In designung our trials of anticoagulant therapy and no result was plotted. If, however, menbers 63 studies web ·of a pair behaved dissimilarly a preference was plotted and the direction of the arrow indicated whether it was the treated or control member who had failed to survive. Arrows pointing downwards and to the right represent the pairs in which the treated member died while the control member survived ; arrows pointing upwards and to the right indicated pairs in which it was the treated member who survived. The horizontal axis represents the total number of preferences while the vertical axis represents the number of pairs showing preference for anticoagulation treatment minus the number showing preference for control treatment The apex of the plot thus registering a minus value (Fig. 1), indicates a trend in favour of the control treatment. Had the upper or lower boundary been reached, it who died. Three of the six patients in the treatment group who died showed evidence of cerebral hæmorr. hage. The possible implications of this will be discussed later, but the general conclusion drawn from this trial was that anticoagulants are not of value in acute strokes when used according to our criteria of selection and management. The second trial was designed to assess the value of long-term anticoagulants as a prophylactic measure in patients suffering from cerebrovascular disease. The trial was designed in collaboration with Professor Bradford Hill. The precise objective was to determine whether or not the expectation of life could be increased or the incidence of further cerebrovascular accidents decreased in patients with cerebrovascular disease. Again we did not confine ourselves to any particular diagnostic subgroups but included the whole range of cases most commonly encountered in general medical practice. Patients were considered for inclusion who were under seventy years of age and in whom there was evidence of a previous neurological disturbance lasting for more than twenty-four hours and judged by the clinician caring for the patient at the time, and by ourselves subsequently, to be due to nonhæmorrhagic cerebral, carotid or vertebro-basılar arterial disease. No distinction was made between those who had made a good functional recovery and those who had been left disabled. In all cases & minimum of fourteen days had clapsed between the last acute cerebrovascular lesion and inclusion in the trial. All patients considered for the trial attended hospital as out-patients for a day for investigation and assessment. X-rays of skull and chest, electrocardiogram and renal a point was reached when the upper boundary was no longer attainable and the trial was therefore stopped with no significant difference between the treatment and control groups demonstrated. In all there were six deaths among the twentysix patients in the treated group and three deaths among the twenty-five controls. Post mortem examinations were carried out on eight of the nine patients 35 Figure 1 10 20 TOTAL at hourly intervals with the patient supine. The usual contraindications to anticoagulant therapy were 15 observed and patients were also excluded if it was felt that PREFERENCES they would be unable or unwilling to co-operate sufficiently or to attend regularly for the necessary supervision, Patients accepted for the trial were differentiated by sex and randomly allotted in pairs 10 one of two groups. The treated group (high-dosage group) received tablets containing 50 mg, phenindione, while the control group (low-dosage group) were given apparently identical tablets containing I mg, phenındtone, an amount -15 insufficient to affect the clotting mechanism The patients did not know to which group they Restricted procedure for 0.09 with maximum nur ber of belonged and both groups were preferences, *19. managed in the same way with regular prothrombin estima- Suppose is probability of preference for anticoagulants, tions ar intervals not exceeding II 0=05 (null lıypotlicsıs), probability of teaching cach outer four weeks In the high-dosage boundary :s 0 022 (1 e., slightly lower than clic nominal 0 025). group the aim of treatment was to maintain the prothrombin If =09, probability of reaching upper boundary is o 97(10, time (Quick one-stage test) at a slightly lugher than the nominal 0 95). level of two to two-and-a-half times the control value (Points at which the boundaries can be reached are shown as circles ) 66 PREFERENCES FOR ANTICOAGULANT TREATMENT MINUS PREFERENCES FOR CONTROL TREATMENT ·Many of the criteria for selection and details of management were based on the plan of the Medical Rescarch Counal trial of anucoagulant therapy in coronary thrombosis (A.R.C. 1959), to which access was kindly granted, and we grate fully acknowledge the help thus received. One hundred and forty-two patients were admitted to the trial, 71 (47 men and 24 women) to the treatment group and 71 (47 men and 24 TABLE 1.-SUMMARY OF RISULTS. High Iosage Group Low Dosage Group Male temule Total Male Female Total Totul No of Patients +7 24 71 47 71 on fatal cerebrovascular accidents 2 $____ + Fatal cerebrovascular accidents 2 due to haemorrhage than has formerly beco supposed. We suspect, and there is supporting patho- 3 L--- • One diere of myocardial infarction, and one is alive and has had no further cerebrovascular accidents .L. ar -L-LI. TLhonorrhage, three were a nong the twenty-four hypertensives in the treatment group and one was fron the forty-seven who were not hypertensive. These figures are not significant, but in further studies we are avoiding the use of anticoagulant treatment in hypertensive subjects. group and four recurrences in three pauents in the In conclusion it must be stressed that these control group, a difference which is clcarly not results need not necessarily conflict with more significant. However, there were four fatal cerebro- favourable reports of experience with anticoagulant vascular accidents in the treatment group and none drugs in specific cerebrovascular syndrones. We in the control. Furthermore all of these deaths were have so far attcnpted only to assess the value of due to hæmorrhage and that they were related to anticoagulants in the two commonest situations in --- which the physician is concerned in the treatment of acute and chronic cerebrovascular disease. From our studies we believe that the general use of anticoagulant therapy in these situations is not to be recommended TABLE 2.- COUPLICATIONS OF TREATVEST High Dexnge Group Low Dosage Group Vale I emale Total Vale Female Total Total o of Patients 47 24 71 Hæmontagne Treatment interrupted 5 Hæmorrhage observing by chance five deaths in the one group and Treatment abandoned none in the other does not quite reach the formal lvpersensitivity AnlamlL.. C.11.11. L- Treatment abandoned Intercurrent Ilness- Treatment abandoned 47 24 70 8 13 1 --- reproduce the cables cerebral hæmorrhage were, in fact, infarcts in which bleeding had been induced seems unlikely both on REFERENCES pathological grounds and by inference from the Barron, a D and Fergusson, G. (1999). Neurology, 9, 417 absence of a similar incidence of infarction in the Bull, HD, Marshall, J and Shaw, D A (1960), Lancet control group It seems improbable, too, that 1, 562 inadequate control of therapy in our series can be Carter, A B (1957), Quart J Ved, S. 26, 335 held responsible since our complication rate in HII, Bradford, Varshall, J and Shaw, D A., (1960), other respects does not compare unfavourably with Quart J Med S29397 Varshall, J and Shar, D A (1960), Lancet, 4 995 other series (Table 2) Medical Research Council Working Party (1959). Bat Med The incidence of cerebral hæmorrhage in J, 1, 803 67 ·DISCUSSION Professor HILL - A very high proportion of the deaths in myocardial infarction occur in the first 24 hours after onset. These are in general due to causes pot amenable to anticoagulant therapy: of the disease. I should like to refer to a use of anticoagulants not mentioned hitherto, namely their value in cutting short the ingravescent cerebral thrombosis in dehydrated infants with cyanotic congenital heart disease A small venesection (20 ml) and tiny doses of heparin can save some such infants from a crippling hemiplegia, which if unchecked may render later operation for the cardiac condition unjustifiable. The experience of Mr F. R. Brown in respect of the incidence of pulmonary embolism after operation for thyrotoxicosis should be mentioned. In Hunter and Richards stressed the importance of an adequate depression of the " prothrombin level in order to obtain therapeutic effect. There seems to be no doubt that many patients are undertreated because this important fact is not correctly understood. This is often caused by incorrect assessment of the therapeutic level for the actual a xine or tri-iodothyronine pre-operatively as prophylactic? of the hæmorrhagic tendency. An absolutely stable level at a specified percentage cannot be achieved The practical result, therefore, is that we should aim at a level of 15% with 10% and 25% as the lower and upper limits. my TO.-.1.-1 1 1 is probably that the number of patients is too small A recent study in Sweden showed statistically significant effect above 60 years of age but not below The situation in myocardial infarction is very probably the same as in angina pectoris, the main factor influencing the therapeutic effect being the "age" of the disease prior to therapy and not the actual age of the patient We believe that it is not correct to consider age above 60 years as a contraindication to therapy. This would also exclude the majority of women because of the delayed onset of coronary disease in the female Richards found no direct relationshup between from patients on anticoagulant therapy have normal concentrations of these factors and, therefore, give shorter clotting times than those shown by the correlation curve for the corresponding concentrations of the 4 factors depressed. If adsorbed plasma is used as diluent, instead of salınc, Thrombotest and Quck's test give more identical results, differences occurring when factor IX is disproportionately reduced It must further be remembered that whenever methods are compared the same standard plasma must be used for all correlation curves Dr MITCHELL - I wish to stress some of the dangers inherent in statistical comparisons between various groups of patients. No investigator approaches a problem without some idea about the values and dangers of a particular treatment, and however great the care taken to avoid it, bias is inescapable. Thus, Dr Richards said that his pathologists had been struck by the incidence of various events in patients on anticoagulants ; it surely would have been better thercafter for the pathologists to be unaware of the treatment of the 'mission on or control unior staff they may the day of drur or 1 method, Li iparable 1 69 ·MYOCARDIAL INFARCTION PATIENTS < 60 YEARS ($wetu u TDT? 100 TREATED GROUPS 901 SOV (1) NOULIO (3) Figure 1 (left) -The effect of life. tong anticoagulant therapy on mortality of survivors of acute myocardial infarction. Survival curves for patlents below 60 years of age 80 70 CONTROL GROUP WEREDLUND (9) 60 50 OBSERVATION YEARS Figure 2 (below) - Survival curves for patients above 60 years of age. Dr Barham Carter's assessment of MYOCARD LAL INFARCTION PATIENTS > 60 YEARS 100 SURVIVAL IN 901 corrclauon between the patient's own assessment of his progress and the objective assessment by exercise tolerance test. n.1---L.-.-..L. - V. TREATED GROUPS 801 70 ORDA(142) placebo and dies because anticoagulant is --- WthLW91 Ant CONTROL GROUP 60 NEOCLUD 14, So! groups. 1 Dr SCHIV ARTZ-Dr Richards's 2 3 figures for myocardial rupture in treated and untreated cases of myocardial infarction OBSERVATION YEARS might be interpreted as evidence that anticoagulants predispose to rupture of the myo 107 outpatients who have been treated for 1302 cardium However, a rupture incidence of 10% of patient-months with dicoumarol derivatives Half of these patients were maintained on anticoagulant therapy for more than six months. anticoagulants delay the time of rupture, or that there are very sensitive to dicoumarol derivatives (3 to has been some unusual selection of cases. 9 mg Marcoumar per week), others on the contrary develop a progressive resistance (42-54 mg. Mar- DT VERSTRAETE - This study concerns coumar per week). 69 ·A During the 1302 months of treatment, there by anticoagulants. There were 4 reinfarctions during were 7 relapses of thrombo-embolic phenomena. the post-infarction period in hospital, and in every In 4 of these patients, the Quick test was above the one of these we found, 12 to 24 hours before che therapeutic range 12 hours after the accident; 27 minor hamotthages were observed during the accident, a marked shortening of coagulation time In 3 of these 4 cases we started anticoagulants at 39,060 days of treatment (125 years) and 3 severe ---L- 1-1--- hæmorrhages, one of these being fatal (cerebral bleeding). Besides the indications for long-term anticoagulant treatment, there is an urgent need for proper evaluation of coumarin drugs combined with vasoactive substances New and simple methods Professor DOUGLAS -- There is a big differfor early detection of minor bleeding tendencies ence in the incidence of fatal post-operative pulshould be developed as a safety measure. monary embolism between abdomunal and thoracic operations. In the former it is about 2%, while in the latter in 400 consecutive thoracotomes for cardiac disease it was nul. Hence systemic causes are unlikely to be the whole answer. Perhaps muld to determine the value of anticoagulant treatment inflammatory lesions around the large veins in the for the survival time in cerebral thrombosis and pelvis are important ætrologically. embolism. The only criterion was life or death, The value, or lack of it, of anticoagulants in occlusive arterial disease of the legs is quite unknown There were in all 66 cases of cerebral thrombosis But it could be well settled by a double blind trial 10 not treated with anticoagulants and a regular follow any vascular clinic if it were well controlled by up was obtained in 45 Nineteen patients survived arteriographic studies the initial thrombosis for an average period of 39 months Twenty-six patients with an average age Professor OIREN — The question was raised of 63 years died after Is months If the first month whether anticoagulant therapy influences angina of survival is excluded, the mean survival time was pectoris Waaler Waaler, B A (1957), Acta Med 24 months. Scand, 157, 289) found that 40-50% the patients Of the 41 treated patients with cerebral thromwere improved, a distinctly higher percentage than bosis, no recent information was obtained in 14. Of found with a placebo. Improvement occurred gradythe remaining 27, 2: (mean age 61 years) were still ally during the first 12-18 months and then persisted, alive 131 months after the initial thrombosis. Only a finding which contradicts a placebo effect A 6 patients in this group died after an average of placebo effect was also often observed, but this 33 months effect disappeared after a few weeks. In Borchgrevink's study (Acta Med Scand., in print) the Thus, 577 of the 45 patients with cerebral effect on angina pectoris was analysed by the double thrombosis not treated with anticoagulants died blind technique and the results confirmed Waaler's after 15 months compared with 22°. of the 27 anti findings. The effect may possibly be explained by coagulated patients who died after 33 months. the prevention of new thrombotic occlusions, thereby of 6 untreated patients with cerebral embolism favouring an undisturbed development of the who could be traced, there was an average survival collateral circulation Improvement of symptoms time of 35 months, and 3 deaths after 2 months Of was much more frequent in cases with duration of 8 patients with cerebral embolism treated with symptoms of less than 2 years before therapy and in anticoagulants 7 are still alive after 22 months and patients who kept up regular exercise, which may only I died after 5 months also promote improvement of the collateral circus lation. Dr MARSHALL - As regards Dr Mitchell's Professor HILL-A trial of long-term and criticism, if the series is large enough errors in diag. nosis will be distributed equally between treated coagulant therapy in angina pectoris is sound if and control groups. In any case, when both members survival of the patients is made the yardstock of of a pair die thus is a tied pair which does not success. The difficulties of assessing subjective affect the sequential analysis In our study, all deaths improvement (pain) are well known, for example in fact occurred in untied" from the critical work of Cole. Repeated testing by pairs. exercise tests, which can be very dangerous, 1$ unjustifiable and profitless if the aim is simply to measure survival Morcover, pain may disappear because the muscle is dead. Dr MITCHELL - Many people feel that Dr TURNBULL - It is true that older patients exercise tests in angina pectoris are dangcrous If share ale floor rennirs but the rupe of it is thought that the test may produce infarction, at should be remembered that Master showed that there was no correlation between physical activity and the onset of an infarct, since the incidents occurred randomly throughout the day As Waync and Laplace pointed out, one is asking the patient to do no more, while under supervision, than he repairs. does several times a day on his own Dr HARTERT - Concernung reinfarctions, we Professor BOYD - Is it neccssarily myocardial made a scrupulous follow-up by thrombo-elastography accrosis that causes death under such aircumstances ? of a number of myocardial infarctions not treated 70 ·Professor HILL I myself have seen a patient a blind tral is necessary when this type of objective die immediately after such a test, and have heard measurement is possible. of others. Professor BOYD -- 1 dislike the use of anti- Dr MITCHELL - I wonder whether it is fear coagulants in phlebitis, superfiaal or deep, of the of the event, or of the event occurring in the labora. legs, I have treated phlebitis for 30 years and never tory, that worries the doctor. Professor BOYD-Do domestic and other accidents in patients on long-term anticoagulant therapy constitute a serious hazard? that anticoagulants might disintegrate or loosen a Dr RICHARDS With regard to hamorr recent clor, hagic complications of therapy, minor domestic I myself do not treat peripheral arterial embolism accidents often mave tise to minor hematomata, with annicoagulants but have seen ad Dos Santos but these are seldom serious. of Lisbon's cases of perpheral embolism treated I agree that the overall incidence of cardiac by heparin : Asteriograms showed disintcxration or rupture is about 10 of myocardial infarctions solution of the thrombus, the patency of the arterial coming to autopsy and that a study of all cases of lumen becoming completely restored rupture does not indicate any connection with anticoagulant therapy. With regard to the duration of of the legs, but I have followed up 1440 patients with I have not used anucoagulants in atherosclerosis ilincss to death in the cases of cardiac riprure, atherosclerosis and intermittent claudication but although this is longer than usual in the ruptures in without sangenc or pregangrebous changes, in the anticoagulant group, the 4 cases of rupture in order to study the natural course of the disease. patients, dying within 24 hours of admission are included the mean time to death comes down to Aftet rs scars, 85% of the deaths were due to atherosclerosis 75°. to 2 days. coronary thrombosis" and 10° to cerebral vascular catastrophes. Of the My series does not include a high proportion survivors, 10 had a non-fatal coronary or cerebral of patients with muluple infarcts. The high death acadent in the first 5 years, rising to 20, after 10 rate is mainly due to the many patients who died years. These complscations might possibly have been soon after admission and similar in this respect prevented by anticoagulant therapy, to the series of Honey and Truclove. A. the intarin 16.-.Li. essels. Rather strict criteria in my selection of patients limit the number in whom the operation noted that and S... DY BARHAM CARTER My first series did not constitute a strictly controlled trial, but Dr Mitchell is doubtless glad to note that my later inal was randomised. Neurologists are more fortunate than cardio logists in being able to measure objectively disabilities short of death, and I think that the extent of a paraly- $1$ is objective enough I have never thought that a number of tatal cases, but at post-mortem death was found to be due to myocardial infarction. I am astounded by the high incdence of pulmonary embolism in some American papers. There seems to be a marked geographical and seasonal distnbution of this complication. 71 ·fta.ft .L, -L During the 1302 months of treatment, there by anticoagulants There were 4 reinfarctions during were 7 relapses of thrombo-embolic phenomena. In 4 of these patients, the Quick test was above the the post-infarction period in hospital, and in every therapeutic range 12 hours after the accident ; 27 one of these we found, 12 to 24 hours before the minor hæmorrhages were observed during the accident, a marked shortening of coagulation time. In 3 of these 4 cases we started anucoagulants at 39,060 days of treatment (125 years) and 3 severe hæmorrhages, one of these being fatal (cerebral bleeding). Besides the indications for long-term anticoagulant treatment, there is an urgent need for to every patient with acute myocardial infarction proper evaluation of coumarın drugs combined with vasoactive substances. New and simple methods Professor DOUGLAS -- There is a big differfor early detection of minor bleeding tendencies ence in the incidence of fatal post-operative pulshould be developed as a safety measure. monary embolism between abdominal and thoracic operations. In the former it is about 2%, while in the latter in 400 consecutive thoracotones for cardiac disease it was nil. Hence systemic causes are unlikely to be the whole answer. Perhaps mild to determine the value of anticoagulant treatment inflammatory lesions around the large veins in the for the survival time in cerebral thrombosis and pelvis are important ætiologically. ernbolism. The only criterion was life or death The value, or lack of it, of anticoagulants in occlusive arterial disease of the legs is quite unknown There were in all 66 cases of cerebral thrombosis But it could be well settled by a double blind trial in not treated with anucoagulants and a regular follow any vascular clinic if it were well controlled by up was obtained in 45. Nineteen patients survived arteriographic studies. the initial thrombosis for an average period of 39 months. Twenty-six patients with an average age Professor OREN --The question was raised of 63 years died after 15 months. If the first month whether anticoagulant therapy influences angina of survival is excluded, the mean survival time was pectoris, Waaler (Waaler, B. A. (1957), Acta Mied 24 months. Scand., 157, 289) found that 40-50% of the patients Of the 41 treated patients with cerebral throm were improved, a distinctly higher percentage than bosis, no recent information was obtained in 14 Of the remaining 27, 21 (mean age 61 years) were still alıve 131 months after the initial thrombosis Only 6 patients in this group died after an average of effect disappeared after a few weeks. In Borch- 33 months grevink's study (Acta Med. Scand, in print) the Thus, 57.7º. of the 45 patients with cerebral effect on angina pectoris was analysed by the double thrombosis not treated with anticoagulants died blind technique and the results confirmed Waaler's after 15 months compared with 22" of the 27 anti findings. The effect may possibly be explained by coagulated patients who died after 33 months. the prevention of new thrombotic occlusions, thereby Of 6 untreated patients with cerebral embolism favouring an undisturbed development of the who could be traced, there was an average survival collateral circulation. Improvement of symptoms ume of 35 months, and 3 deaths after 2 months Of was much more frequent in cases with duration of 8 pauents with cerebral embolism treated with symptoms of less than 2 years before therapy and in anticoagulants 7 are still alive after 22 months and pauents who kept up regular exercise, which may only I died after 5 months also promote improvement of the collateral circu. lation Dr MARSHALL-As regards Dr Mitchell's criticism, If the series is large enough errors in diag- Professor HILLA trial of long-term antinosis will be distributed equally between treated coagulant therapy in angina pectoris is sound 1 and control groups. In any case, then both members surtzial of the patients is made the yardsaack of success pair which does not of a pair die this is a "tied The difficulties of assessing subjective affect the sequential analysis In our study, al} deaths improvement (pain) are well known, for example * in fact occurred in untied from the critical work of Cole. Repeated testing by pairs, exercise tests, which can be very dangerous, is unjustifiable and profitless if the aim is simply to measure survival Moreover, pain may disappear because the muscle is dead. Dr MITCHELL - Alany people feel that exercise tests in angina pectoris are dangerous if it is thought that the test may produce infarction, it should be remembered that Master showed that there was no correlauon between physical activity and the onset of an infarct, since the incidents occurred randomly throughout the day. As Wayne and Laplace pointed out, one is asking the patient to do no more, while under supervision, than he repairs. does several times a day on hus own. Dr HARTERT - Concerning reinfarctions, we Professor BOYD- Is it necessarily myocardial made a scrupulous follow-up by thrombo-clastography necrosis that causes death under such circumstances ? of a number of myocardial infarctions not treated 70 31 ·- from icare A New Heparinoid Anticoagulant F. K. Beller-Universitats-Frauenklınık, Tübingen Heparinoids arc synthetic or semi-synthetic corresponds to a ratio of approximately 3.5 in compounds with biological cffects like alpha- vitro. This compares very favourably with other heparin Further information on the therapeutic heparinoids. But this in vitro effect cannot be compared from with the in vivo effect. The effects of single intrahave venous doses were measured by Howell's recalcification time (Fig. 1). Only after 0.3-0.4 8. is the initial ccular 1 natural substances, the anticoagulant effect is incrcased, but the side-effects are also increased. To obviate this vicious circle, the compromise was attempted of increasing the anticoagulant action only to that point where the side-effects were still tolerable. Thereforc, the potency of heparinoids is less than that of heparin, so that the same therapeutic effect needs morc drug and toric doses are more plastin which depends on the preparation used easily reached. Thus the therapeutic index is lower (Fig 2). than heparin. We adopted, therefore, a scheme of treatment G31, 150 (Gersy) is the calcrum complex of the (Fig. 3). The ratio of effect in the body compared sulphated oxidative breakdown products of poly- It is instead galacturonic acid methyl ester, with a calcium arinoid are content of 4.5% and an average molecular weight of ients. Most 40,000 It contains 3-4 methoxyl groups and has two from a sulphur content of 13°. recent infarction and twenty-five from so-called This product has excellent clinical tolerability congestion states in abdominal carcinoma; we Its anticoagulant cffect was tested by thrombo. previously established that these swellings remitted clastography as described by Hartcrt Heparin or under heparin and at least partially under heparinoids. heparinoid is mixed with platelet-rich and platelet- The results where heparinoid was injected for 4-6 poor plasma, with and without incubation, and the days and then the drug changed to Sintrom TEG done. In citrated platelet-rich plasma a correspond to well-known experience with heparın. heparinoid solution of 1.1000-I 1250 is equivalent Mild cases do with three daily doses, the minimum in effect to a heparin dilution of 14000, which in surgical patients. As described by Merz, C71518 The effect of different doses on the The effect of different doses on the heparin time recalcification time mean value of 5 cases mean value of 5 cases 560 520 520- 400 400- 440- 400 400 360- 360- 120- Heparin time in seconds double O-level 320- Recalcification time in seconds 280- 400 mg 400 mg 200 240 300 mg 240- 200 300-mg 2004 160- 200 mg 200 mg 160- 120 100 mg 100 mg 1204 0 VIL Figure 1 hours after injection 75 ·citrare and the eluates examined The antiheparin activity of platelet suspensions and of serum was found to be of the same order of magnitude (Fig. 4). Serum antiheparin activity is not related to Christmas Factor activity, in spite of the powerful inhibitory action of heparin on this factor, Similarly there is some evidence that serun antiheparin activity is not related to Hageman Factor. Hanns- .L PTA, Christmas Factor (IX), Proaccelerin (V), and AHG (VIII) in order of decreasing sensitivity, It also inhibited the surface activation of Hageman Factor, In other experiments various quantities of heparin (2.5, 5, 10 and 20 units) were added to 10 ml. of human oxalated blood. After 30 minutes incubation at room temperature, samples of heparirused blood were centrifuged and clorting factors were determined in plasma diluted I: 10 Tamard: tin rantaila... factopares - '; Was :,: It is well known that human serum, like platelets, shows antiheparin activity. According to O'Brien (1960) this activity is related to the presence of some platelet proteins in serum (plareler-like activity of serum) and to Christmas Factor activity. O'Brien (1960) suggests that heparin neutralises Christmas Factor and platelets giving complexes with these substances, Other findings are, however, contradictory Poller (1960) found that serum antiheparin activity is not related to the activities of Factor VII, IX and Xin the serum. We compared the antheparin activities of a platelet suspension 300,000 per cu min. with normal human serum and with Christmas serum. In the same experiment both sera were adsorbed on BaSO4. The BaSO4 precipitate was eluted with sodium spite of the great amount of work done on this subject. Our findings are that heparin does not inhibit prothrombin and complex VII, but it inhibits Factor V, VIII, IX, PTA and Hageman. These factors are pa Нер inhibitor of th On the contra considered rati blood-clotting combined ther system." therapy. ANTIHEPARIN ACTIVITY OF SERUM, SERUM FRACTIONS AND DLATELETS INCUBATON MIXTURE HUMAN CITRATED DLASMA THROMBIN CLOITING TIME 0.2 m SALINE 0,3 ml 0,1 10 0,1 mol 01 ml SALINE HEPARIN 03 0.1 60. 0,1 ml PLATELETS HEDALIN 0,3 ml 0.1 12 REFERENCES Bambell, E, Corsin, A H, Lapone, V A Amer J Med Sci (1955), 230, 276 Chargaff, C, Zuff, V, Cohen, S SJ Biol Chem. (1940), 136, 257 Deutsch, L, Johnson, S A, Seegers, WH' Circ Res (1955), 3, 110. Douglas, A. J Clin Investig (1956). 35, 533 Folemaps, R, Vermylen, c, Verstraete. M Thromb Diath, Ham (1960), #, 303 Slonch, WH, Holt, L. Amer. J Physiol (1918), 47, 328 Koller, Loeliger, A. Duckert, F Acta Hæmat (1957), 6,1 Kowaishi, E, Kopec. , Latio, 2 Roszkowski, s, Sendys, N Blood (1958), 8, 436 Hacian, RL, Brown, K. KG) Lab Clin Ned (1954), 44, 378 Nren tarou sha, M llegrzynowicz, Z Thromb Diath ilum (1959), 3, 279 O'Brien, J R Nature (19581, 181, 101 O'Brien, J R J Clin Path" (1960), 13, 19бо Poller, L. J Clin Path (1960), 13, 226 Quick, A J Amer J Physiol (1938), 123, 913 Shanberge, J N, Sarelis, A, Regan, I Journ Lab Clin Ved (1959). 54, 5o1 Soulier, JP, Larrieu, M J Sang (1952), 33, 549 Soulier, P., Larrieu, VJ Sang (1953), 24, 205 Sower, P, Larrieu, y ] Les Aatiorgulantes en Thérapeutique (1955) Doin, Ed, Paris Waaler, BA Scannt J Clin Lab Invest (1959), 11 Suppl. 17. 135. Wolf, P J Clin Path (1953), 6, 34 0.1 Q10 NORMAL SIRUM HEPARIN 0,3 0,1 12% 01 nt HEMOPHILIA B SERUM HEPARIN 0,3 01 10 h 01 ! 0,3 01 ml 0.1 M4 ELVATE (NORMAL) 0,8 HEPARIN 17 0.1 로 0,5 M 01 Be SOKELUATE (HEMOPHIUA B) HEPARIN 15 Figure 4 74 ·.harafas n.oh A New Heparinoid Anticoagulant F. X. Beller - Universitats-Frauenklinik, Tubingen Heparinoids are synthetic or semi-synthetic corresponds to a ratio of approximately 1:3.5. in compounds with bioloncal effects like alpha- vitro. This comparts very favourably with other heparın. Further informaton on the therapeutic heparinoids. action of the heranın group is derived not only from But thus in vitro cffcct cannot be compared their varying effects on coagulation, but also from with the in viso effect. The effects of single intratheir side-effects, Only a few such products have enous doses were measured by Howell's recalafica. attained clinical significance, and cien these are uon ime (Fig. 1). Only after 0 3-0.4 8. is the injual disputed. By sulphauon of certain high-molecular value doubled at three hours, the usual prerequisite natural substances, the anticoagulant effect 15 fondo ne prenn. increased, but the side-effects are also increased. To obviate this vicious arcle, the compromusc 25 attempted of increasing the anticoagulant action only to that point where the side-effects were still tolerable. Therefore, the potency of heparinoids is less than that of heparin, so that the same therapeutic effect needs more drug and toxic doses are more casily reached. Thus the therapeutic index is lower than ficparın. We adopted, therefore, a scheme of treatment G3!, 150 (Gessy) is the calcium complex of the (Fig. 3). The rauo of effect in the body compared sulphated oxidative breakdown products of polyt is instead galacturonic acid methyl ester, with a calcium warinoid are content of 4.5., and an average molecular weight of ients. Most 40,000. It contains 3.4. methoxyl groups and has two from a sulphur content of 13.- recent infarction and twenty-five from so-called This product has excellent clinical tolerability. congestion states in abdominal carcinoma ; Its anticoagulant effect was tested by thrombo previously established that these su cllings remitted clastography as described by Hartert. Heparin or under heparin and at least partially under heparinoids. heparinoid is mixed with platelet-nch and platelet- The results where heparinoid was injected for 4-6 poor plasma, with and without incubation, and the days and then the drug changed to Sintrom TEG done. In citrated platelet-rich plasma a correspond to well-known experience with heparın. heparinoid solution of 1 1000-1'1250 is equivalent Mild cases do Sith three daily doses, the minimum in cffect to a heparin dilution of 1:4000, which in surgical pauents As described by Merz, crisis The effect of different doses on the The effect of different doses on the heparin time recalcification time mean value of S cases mean value of 5 cases we 560 520 520- 480 480 440- 40 400- 400-1 360- 360- 20- Heparin time in seconds double O-level 320 + Recalcification time in seconds 200- 400 mg 200- 400 mg 300 mg 240) 2404 200+ 300-mg 200- 160- 200 mg 160- 200 mg 120- 100 mg 100 mg 1202 80 VA 3 0 Figure 1 hours after injection 75 ·PTA, Christmas Factor (IX), Proaccelerin (V), and citrate and the eluates examined The antiheparin AHG (VIII) in order of decreasing sensitivity. It also activity of platelet suspensions and of serum was inhibited the surface activation of Hageman Factor. found to be of the same order of magnitude (Fig. 4). In other experiments various quantities of Serum antiheparin activity is not related to heparin (2.5, 5, io and 20 units) were added to Christmas Factor activity, in spite of the powerful 10 ml. of human oxalated blood. After 30 minutes' inhibitory action of heparın on this factor. Similarly incubation at room temperature, samples of hepari there is some evidence that serum antiheparin nised blood were centrifuged and clotting factors activity is not related to Hageman Factor, were determined in plasma diluted 1: 10. Heparin added in vitro to blood inhibits clotting factors in a similar manner to heparin injected parenterally (Fig. 3) It should be noted that AHG was more inhibited in vitro than in vivo. spite of the great amount of work done on this It is well known that human serum, like plate- subject. lets, shows anttheparin activity. According to Inae ant inhihir O'Brien (1960) this activity is related to the presence of some platelet proteins in serum (platelet-like activity of serum) and to Christmas Factor activity, O'Brien (1960) suggests that heparin neutralises Christmas Factor and platelets giving complexes with these substances. Other findings are, however, contradictory Poller (1960) found that serum antiheparin activity is not related to the activities of Factor VII, IX and X in the serum. We compared the antheparin activities of a platelet suspension 300,000 per cu, mm. with normal human serum and with Christmas serum. In the same experiment both sera were adsorbed on BaSO4. The BaSO4 precipitate was eluted with sodium therapy. 1.- foton .be 14-nin ANTIHEPARIN ACTIVITY OF SERUM, SERUM FRACTIONS AND DLATELETS INCUBATION MIXTURE HUMAN CITRATED PLASMA THROMBIN CLOTTING TIME 0,2 ml SALINE 0,3 ml 0,1 ml 10 0,1 0.1 GALINE HETARIN 0,1 ml 60. 0,1 ml RATELETS KEPARIN 0,3 mil 01 t2. REFERENCES Branibell, C L. Corwin, A H. Lapone, VA Amer. J Med Sci (1955), 230, 276 Chargafl, L, Ziff, M, Cohen, S S J. Biol Chem (1940), 136, 257 Deutsch, CJohnson, S A, Scegers, W H Carc "Res (1955), 3. 110 Touglas, A s J Clin Investig (1956), 35.533 Holemans, R , Vermy len, c, lerstraete, M Thromb Diath Hæm (1960), 4, 303 Lonell, W 1!, fols, Amer J Physiol (1918). 47, 318 Koller, F, Loeliger, A. Duckert, F Acta Hamat (1957), 2, I Kowalsha, L, hopce, Latallo, 2 Roszhoushi, s, Sendys, N Blood (1958), 6, 430 MacMillan, RL, Brown, K. W GJ Lab Clin Ned (1954), 44, 378 View arouska, y llegrzynowicz, Z Thromb Diath H.um (1959). 3. 279 O'Brien, J R Nature (1958), 181, 1801 O'Brien, J R J Clin Path (1960), 13, 1960 Poller, L J Clin Path (1960), 13, 220 Quick, 4) Aner J Physiol (1938), 123, 712 Shanberge, J N. Sarelis, A, Regan, EE Journ Lab Chin Ved (1959), 54. Sor Soulier, JP Larrieu, MJ Sang (1952), 23, 549 Soulier, JP, Larrieu, M J Sang (1953), 21, 205 Soulier, J 1, Larrieu, V T Les Anti coagulantes en Théripeutique (1935) Doin. La, Pans 1aaler, BA Scand J Clin Lab Invest (1959). 11 Suppl. 37, 135 Woll, P J Clin Path (1933), 6, 31 0,3 m! 0.1 0,1 mol NLXML SERUM Q1 m! HEPARIN mi 12% HEMOPHILAB SERUM HEPA RIN 0,3 0 1 ml 10% 91 17 0,4 l 304 ELVATE (NORMAL) 0,1 ml HEPARIN 0,3 15 01 84 SO, ELUATE (HEMOPHILA B) HEPARIN 0,3 0.1 al Figure 4 74 ·- Example : 100 mg. - 1 ml. of the 10 solution contained in the ampoules. Ist Day and Day 3rd Day 8 ooh : 400 mg. 1.v. 8.00h : 400 mg. i.v. 8.00h 400 mg, 1.v. 13 och : 300 mg. i.x, 3 h 300 mg, iv, 13.00h 200 mg. i.x. 17.00h : 300 mg. 1.v. 17.00h : 200 mg. i.v. 200 mg. 1.v. 21.00h : 300 mg. i.v. 21.00h: 300 mg. l.v. 21.ooh 200 mg. J.V 17 ooh : : : : 4th Day sth Day 6th Day 8 ooh : 400 mg 1.4. 8.ooh 400 mg, 1.Y. 8.00h 400 mg. iv. 13.00h : 200 mg.i.v. 13 ooh 200 mg.i.v. 13 ooh : 200 mg. iv. 18 ooh : 300 mg.i.v. 18 ooh 300 mg. 1. + cumarin + cumarin In some instances various routes of application (i v., 3.C., 1m.) were successfully used for the individual doses. Figure 3 The effect of a combination of intravenous and subcutaneous injection of Heparinoid on the heparin lime 550 500 - 450 seconds 400 Depot injection 500 mg 350- 300 A. and sc. injection Heparin time 250 1. injection IV. injection 400 mg 200 Z 175 0 1 2 3 T 6 4 5 7 9 10 12 Figure 4 Time after injection in hours ·TO .. Prothrombintime (Quick) in dependence thromboplastin. Our and lysis took place at latest six days after the begin. ning of therapy, when the patients were able to get corresponding to the daily intravenous dose, but do One gram of a depot preparation may be injected, up. In six years we had only one failure: a woman absorption of therapeutic signuficance occurs withun with puerperal sepsis, who died with extensive two hours. There was no difference in the effect of thrombosis following an injection of Pyrexal; a subcutaneous and intramuscular depot injections. Schwartzman phenomenon may have occurred Fifteen pregnant women were treated only with heparinoid G 31 150, in slightly lower daily dosage. Pregnancy is the only condition where treatment was continued longer than six days, though with gradual reduction in dosage. All gave birth to living children; one child was deformed and died after three days. The rest were in no way abnormal, In three cases premature labour started during treatment. Therapy was continued to to minutes before parturition, when the effect was interrupted with The excretion is similar to that of heparin, but protamine sulphate. The post partum period was more biologically active substance is present in the normal. urine than after heparın. This is clearly seen with depot heparinoid. After injection of alpha-heparin, the remaining biological activity is approxi. mately 1/3-1/4 of the excreted on the amount, after heparinoid approximately one-half. That resembles results with other heparinoids, and these substances are probably 1, 2, 3 and 4 different thromboplastinpreparations less broken down in the body than heparin. The excretion values also confirm the findings with respect to saturation dose 400 mg. IV and thus support the dosage scheme. % In experience, the following side-effects are frequent after the heparinoids investigated 100 hitherto : after rapid injection: tachycardia, feeling of oppression in the chest, nausea, vomiting, cyanosis, later paræsthesia We observed none of these even after rapid intravention injection of G 31, 150 in doses up to 1.5 g. 80- The incidence of side-effects as So far under 1%. However, Stamm reports more frequení 70 side-effects during longer therapy. We have no experience of longterm therapy, and it seems inad- 60 visable to prolong treatment beyond eight days. A special problem is loss of hatr, which is particularly promi- 50- nent in gynecology and obstetrice, We observed it 8-12 weeks after the beginning of therapy 40 even with heparin in approxi. mately 70% of cases, and after heparinoids in almost 100%. Of 30- these again 30-40% have total alopecia After Heparinoid Geigy, counting distinctly visible reduction of hair, it occurred in ap- 20 proximately 80% and total alopecia in under 5°. In all, the hair grow again normally. 'O This product may be used where there is heparin allergy, but it will find its chicf place where heparin is prohibited or limited 0 by its price. It is of great value 0 2 3 4 5 for the initiation of thrombosis treatment, and 1s worth including hours after injection in Our therapeutic equipment. 76 90 Prothrombintime in % Figure 2 ·Lipemia Clearing Activity in Dogs ofter intravenous Injection Figure 2 Sulfopoly glucin Heporin in Ualls/kg in mg/kg 500(3 dogs) 5 (4) 3514 10 (4) 214) Cleoring Activity (CU) 1/4) 1.35141 200(4) 05(4) 0.264) 50(4) 203) 0.1(4) anna HOURS 2 "Ediol," a stabilised 50, coconut oil emulsion, when tested the al nominaf nu plnom activities we the use of (obtained t Institute for " Free fatty acids, before and after incubation of plasma wa Ediol or human low-density Beta- 1. In L--La 3.a. doses being given concurrently. SPG was similarly Injected ! v. into groups of 4 dogs at doses of 10 to 01 mg./kg. In all cases blood samples were taken before, and I, 1, 1, 3 and 5 hours after injection for determination of LCA and CT. With respect to anticoagulant effect (Fig. 1), the time effect curves for heparin and SPG were qualitatively similar, both series showing rapid peaks, and slower disappearance of effect. Heparin had a Inuch greater anticoagulant effect than SPG. On a weight basis, it was approximately five to seven times more active than SPG. This agrees very well with the figure of 21 USP units per mg, which was found for SPG in vitro LCA ume-cffect curves (Fig 2) resulting from injection of heparin and SPG differed both quantitatively and qualitatively. Like the anti- I v Mongrel dogs were fasted overnight (about 20 hours) with access to water The drug was conyined Intravenous SPG v Heparın Heparin was injected iv Into dogs in groups of 3 or 4 at doses of 500 to 20 USP units/ks, all 79 In a study in which radioactive SPG containing ·A New Orally Effective Heparinoid ANTICOAGULANT AND ANTILIPÆMIC ACTIVITY IN ANIMALS AND MAN AFTER ORAL AND PARENTERAL ADMINISTRATION E. WINDSOR and G. E. CRONHEIM - Research Division, Riker Laboratories, Inc., Northridge, California The present report describes the results with a Riker 560 is a mixture by weight of 4 parts of new heparinoid which is absorbed following oral SPG Potassium and I part of disodiun dihydrogen dosage. The drug has been successfully administered ethylenediamine-tetraacecate (“ Versenate" sodium). either parenterally or orally or both, to rars, rabbirs, Heparin Sodium USP for parenteral adminicats, dogs, monkeys, and to man, stration was " Lipo-Hepin" (Darwin), 10,000 USD Experiments designed to learn the effects of units per ml., diluted with physiological saline to a prolonged administration of heparinoids on lipid final yolume of 0.5 ml (kg. Heparin sodium USP metabolis. have been hampered in the nast hv was also used as a powder, with a potency of 130 USP units per mg. 1972.nl. The present report is limited on Ilean . Jinpin ton In and where in oplical density of enzyme-substrate mixture incubated at 20°C, and read ar 700 mu ) The only modifications were the limitation of reaction time to 10 minutes, and the use of “Ediol (SchenLab), diluted 1 to 50 with water, as a substrate, With Whole Blood Clothing Time in Dogs ofter Intravenous Injection AA Figure / Heparin in Units/kg Sulfopolyglucın in mg/kg 601 ) 505 200(4) 500 (3 dogs) 15(4) 10(4) CLOTTING TIME in minutes 5014) 214 *2018 101 Nu 3 5 HOURSI 78 ·Clotting Time in Dogs after Heparin below 1 ms. ks. Comparing the lipolytic cllects of each drug by the two modes of injection, subcutaneous SPG at 1 mg/kg or more was more cffective than i.y. SPG, while sc. heparin showed less lipolysis than 1 y. heparin at all doses. Subcutoneous Dose in Units/kg 601 509 13 dogs) 507 401 Oral Riker 560, SPG, and Horarin. SPG and Riker soo ucre gisen orally to fasted dogs at doses of 100 to 1000 mg./kg, and 25 to 200 mg./kg. respectively. Anticoagulant etfects (Fig 6) appeared with considerable dclay compared to those after 1.v. ad ministration, while LCA ne-effect curves were qualitatively simular to results after IV. injection SPG doses below 300 ms ikg, and Riker s6o doses of 100 mg./kg. or less had little or Do effect on CT. However, all doscs produced considerable increases in LCA. LCA and CT dosc-response curves at 3 hours after the drugs (Fig 7) were parallel, and indicated that the addition of Verscnate sodium to SPG increased the absorption from the gut about five-fold. Comparing the lipolytic effects of oral Riker 360 with parenteral SPG or Riker 560 one drives the value for gastrointestinal absorpbon of two to three per cent. of the oral dose. After the increased absorpuon of SPG by the addition of Versenate sodium had been established, it was found that the sa ne adjuvant would bring about the absorption of heparin from the gastro-intestinal tract, as shown by CT and LCA measurements Both LCA and CT effects were delayed, as compared with E 30 200 (3) Time in minutes 201 100 (6) 6006) & 10 35 (5) Clotting HOURS 2 --- Figure 4 SPG alone required a minimum of zo g. to produce an antılıpænic cffect The addion of Versenate (potassium in Series A, sodium in Series B) increased absorption of SPG approximately five-fold at a rauo of SPG: Versene of 4.1. This per cent of the administered dose. ratio was selected because higher ratios reduced No changes beyond control levels were seen in the utilisation of SPG and lower ratios limited the Quick prothrombin time, or “ Thrombotest” time total dose of SPG that could be administered due after peroral heparin, heparin + Versenate, Riker to side effects of Versene 560, or Versenate. Exploratory studies have indicated that Riker 560 at a dose of 4 + 1 g. induced LCA levels of the same order of magnitude as that seen in man after subcutaneous doses of heparin of 10,000 USP units, but without the concomitant anticoagulant effects The LCA responses were dose-related and the peak occurred more rapidly than had been expected after experiments with dogs Fasting conditions promoted the best absorption of the drug, a slight decrease in LCA being seen if given with a carbohydrate meal, and a considerable 81 LI ..tr ·maximum LCA was also seen within to 1 hour. However, levels were about 3 times higher than after the same heparin doses and fell gradually. At 5 hours after !. ing./kg. the LCA level was still nyant.L-- S35 was injected into dogs (Fig. 3) radioactivity, metachromas12, and prolonged anticoagulant effect all disappeared from the blood at the same rate (Levy et al. 1960), indicating that the presence of SPG was required to produce the anticoagulant effect. This had previously been shown to be true for heparın hv Eshers the highe less than the peak injected w or CT revela Subcutaneous SPG v. Heparı. Heparin was injected subcutaneously at doses of soo to 35 USP units/kg. into groups of 3 to 6 dogs. Similarly, SPG was injected at doses of S to 0.35 mg./kg. Blood was sampled before and 1, 5, 1, 3 and 5 hours after injection for LCA and CT determination No prolongation of CT after subcutaneous SPG was seen, even at the highest dose of 5 mg./kg. At the same weight of heparin (500 U/kg.) (Fig. 4), there was a fairly rapid rise in CT to more than Characterization of Lipolync Activity. In order to determine substrate specificity of the lipæmia-clearing enzyme system in plasma, post-SPG dog plasmas were incubated at 37 with either human low-density Beta-lipoproteins or with Ediol, Free fatty acid (FFA) was titrated before and after 1 hour of incubation at 37°C and the difference was calculated as microequivalents per litre per hour. For the same 19 plasma samples the levels of CU were also determined A correlation was found to exist between the two measurements, I CU being equal to 1.3 +/- 0.1 uEq/Lhr., with the Beta-lipoprotein substrate No difference was seen in CU measurements be- TUAAR Data & 3 f Suuwing LLA at 5 hours was still 50% of the maximum effect. After sc. SPG at low doses (up to 1 mg./kg.) intravenous Radioactive Sulfopoly glucin t in Dogs Figure 3 Metachromasia Clotting Time 91. ☆59 Radioactivity SPG-535 Dose in mg/kg 540 1.2 40% Radioactivity in commune plasma mens $ em Metachromosio in pg/ml plasma Clotting Time in minutes 20 06 F210 03 TOL ماس NF 3 4 HOURS 5 80 ·Clotting Time in Dogs after Heparin in minutes belon I mg. kg. Comparing the lipolytic effects of each drug by the two modes of injection, subcutaneous SPG at 1 ms. ks. or more was more effective than i.v. SPG, while s.c. heparin showed less lipolysis than 1.v. heparin at Subcutaneous Dose in Units/kg all doses. Oral Rikar 560, SPG, and Heparin. SPG and Riker 360 were given 60+ orally to fasted dogs at doses of 100 to 1000 mg./kg. and 25 to 200 mg./ks, respectively. Antcoagulant effects (Fig. 6) appeared 50913 dogs) with considerable delay compared to those after 1.v. administration, 501 while LCA time-effect curves were qualitatively sumular to results after LV. injection. SPG doses below 300 mg./kg., and Riker 360 doses of 100 mg/kg or less had little or no effect on CT. However, all 401 doses produced considerable increases in LCA. LCA and CT dose-response curves at 3 hours after the drugs (Fig. 7) were parallel, and indicated that the addition of Versenate sodium to E301 2003) SPG increased the absorpuon from the gut about five-fold. Comparing the lipolytic effects of oral Riker 560 with parenteral SPG or Riker 560 one derives the value for gastrointestinal absorp- 0204 tion of two to three per cent of 100(6) the oral dose, After the increased absorption of SPG by the addition of 60(6) Versenate Sodium had been established, it was found that the 35 (5) sa ne adjuvant would bring about the absorption of heparin from the gastro-intestinal tract, as shown by CT and LCA measurements Both LCA and CT effects were delayed, as conpared with HOURS 2 3 the almost unediate effects seen after 1 v. heparin, Quanutatively, the oral heparin at 13,000 U/kg. Figure 4 was approximately equivalent to 1v heparin at 100 U/kg. with respect to peak SPG alone required a minimum of 20 g. to anticoagulant effect, and to 200 units, kg with produce an antılıpæ.nic effect The addition of respect to lipolytic effect (comparing areas under Versenare (potassium in Series A, sodium in Serres rption B) Increased absorption of $PG approximately in two five-fold at a ratio of SPG Versene of 4:1. This ratio was selected because higher ratios reduced the utilisation of SPG and lower ratios limited the total dose of SPG that could be adninistered due Riker to side effects of Versene. Exploratory studies have indicated that Riker 560 at a dose of 4 - Ig induced LCA levels of the sane order of magnutude as that seen in man after subcutaneous doses of heparin of 10,000 USP unts, but without the concomitant anticoagulant effects. The LCA responses were dose-related and the peak occurred more rapidly than had been expected after experiments with dogs Fasting conditions pro noted the best absorption of the drug, a slight decrease in LCA being seen uf given with a carbohydrate meal, and a considerable Time clotting een in tine 81

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·belor I mg kg. Comparing the Clotting Time in Dogs ofter Heparin Lipolytic effects of cach drug by the ruo moles of injection, subcutançous SPG at I ms. ks, or more was more effective than 1.v. SPG, while sc. heparin showed less lipolysis than iv, heparin at Subcutaneous Dose in Units/kg all doses. Oral Rikar 560, SPG, and Herurin, SPG and Riker 360 ucre given 60+ orally to fasted docs at doses of 100 to 1ooo mg /ks and 25 to 200 mg./kg. respectively. Anucoagulant etfects (Fig. 6) appeared 509 13 dogs) with considerable delay compared to those after i.v. administration, 507 while LCA timc-effect cures were qualitatively $1nilar to results after LV injection. SPG doses below 300 mg/kg, and Riker 560 doses of 100 mg/kg. or less had little or no effect on CT. Howeser, all 401 doses produced considerable increases in LCA, LCA and CT dose-response curies at 3 hours after the drugs (Fig. 7) were parallel, and indicated that the addition of Versenate sodium to E 301 SPG increased the absorption 200 (3) Comparing the lipolytic cerfects of oral Riker 560 with parentera! SPG or Rixer 360 one derives the value for gastrointestinal absorp- 20 tion of two to three per cent. of 100 (6) the oral dose. After the increased absorption of SPG by the addition of 6006) Versenate sodium had been established, it was found that the 10 35 (5) Sa.Te adjuvant would bring about the absorption of heparin from the Rastro-intestinal tract, as shown by CT and LCA measuremients Both LCA and CT effects were delayed, as compared with 5 the almost inmediate effects seen HOURS 2 after i v. heparin. Quantitatively, the oral heparin at 13,000 U/kg. Figure 4 was approximately cquivalent to 19. heparin at 100 U/kg. with respect to peak SPG alone required a minimum of 20 & to desicoagulant effect, and to 200 units with produce an antılıpæ:nic effect. The addition of respect to lipolytic effect (comparing arcas under Versenate (potassium in Series A, sodrum in Series duration curves). This would result from absorption B) increased absorption of SPG approximately through the gastrointestinal tract of less than two per cent, of the administered dosc. five-fold at a ratio of SPG Versene of 41. This ratio was selected because higher ratios reduced No changes beyond control levels were seen in the uulisation of SPG and lower ratios limited the Thrombotest tine after peroral heparin, heparin + Versenatc, Riker total dose of SPG that could be administered due to side effects of Versene. Exploratory studies have indicated that Riker 560 at a dose of 4 + 1 & induced LCA levels of the same order of magnitude as that seen in man after Riker 560 and SPG were administered orally subcutaneous doses of heparin of 10,000 USP to 76 hcalthy volunteers in 132 single doscs ranging units, but without the concomitant anticoagulant from 1 to 20 g. of SPG, in 2 series of trials (Table !! effects. The LCA responses were dose-related and Only a single case was observed in which whole the peak occurred more rapidly than had been blood clotting time was prolonged it reached 15 times the control level 2 hours after a dose of 4% expected after experiments with dogs. of SPG plus 2 g. of Versenate potassium, and it Fasting conditions promoted the best absorption of the drug, a slight decrease in LCA being seen if given with a carbohydrate meal, and a considerable Time in minutes Clotting - Quick prothrombin (me, or 560, or Versenare. Human Pharmacology, Teturned to the control level at 4 hours. 81 ·S35 was injected into dogs (Fig. 3) radioactivity, metachromasia, and prolonged anticoagulant effect all disappeared from the blood at the same rate (Levy et al, 1960), indicating that the presence of SPG was required to produce the anticoagulant effect. This had previously been shown to be true Subcutaneous SPG v. Heparın. Heparin was injected subcutaneously at doses of soo to 35 USP units/kg, into groups of 3 to 6 dogs Similarly, SPG was injected at doses of 5 to 0.35 mg/kg. Blood was sampled before and 5, 1, I, 3 and 5 hours after injection for LCA and CT determination. No prolongation of CT after subcutaneous SPG was seen, even at the highest dose of 5 mg./kg. At the same weight of heparin (500 U/kg.) (Fig. 4), there was a fairly rapid rise in CT to more than 3 times maximum LCA was also seen within 1 to 1 hour. However, levels were about 3 times higher than after the same heparin doses and fell gradually. At 5 hours after I mg/kg. the LCA level was still greater than I CU. At higher does of 2 and 5 mg/kg. the LCA levels continued to rise for 5 hours The course after that time is unknown, but at 24 hours after injections LCA had returned to control values. Characterization of Lipolytic Activity. In order to determine substrate specificity of the lipæmia-clearing enzyme system in plasma, post-SPG dog plasmas were incubated at 37 with either human low-density Beta-lipoproteins or with Ediol Free fatty acid (FFA) was titrated before and after I hour of incubation at 37°C and the difference was calculated as microequivalents per litre per hour For the same 19 plasma samples the levels of CU were also determined. A correlation was found to exist between the two measurements, I CU being equal to 13 +/- 01 Eq/L/hr., with the Beta-lipoprotein substrate No difference was seen in CU measurements between Beta-lipoprotein or Ediol substrates. Becal. the LCA and hepar with one LAAN sampling period, it was assumed, for purposes of computation, that the rate of LCA decrease was the same as those observed at lower doses LCA was still slightly elevated at 5 hours and at 500 U/kg. LCA at 5 hours was still 50% of the maximum effect. After s.c. SPG at low doses (up to I mg./kg.) Intravenous Radioactive Sulfopoly glucin in Dogs Figure 3 S Metachromasia Clotting Time 607 Radioactivity SPG-335 Dose in mg/kg 510 1.2 1 X Radioactivity in com/mi_plasma X 10-3 10 yo Metachromasia in pg/ml plasma Clotting Time in minutes 2 ठ Wimhn 4 5 HOURS 2 3 3 4 5 80 ·Lipemla Clearing Activity in Dogs after Subcutaneous Injection 6. Heparin in Units/kg Sulfopolyglucin in mg/kg 5 5 12 dogs! 34 2(4) 5003 Clearing Activity cu) I (3) 06 (3) 200(3) 100(6) GOIGT 035(2) 2 HOURS ! Figure 5 Figure 6 10 EDTA No EDTA Clearing Clotting 5 + 50 4 40 Clearing Units 30 Minutes 2 20 0 10 25 50 500 1000 100 200 mg / kg ·TABLE I LIPÆMIA CLEARING ACTIVITY IN MAN AFTER SULPHOPOLYGLUCIN PLUS VERSENE lowering if given with a fatty meal, Delaying the fatty meal for 45 minutes did not improve absorption of the drug. Plasma samples drawn during series A (Table 2) were incubated with an Ediol substrate and the increase in release of free fatty acid was measured. While 4 g. of SPG alone had no effect on lipolysis, the same dose when given with Versenate increased free fatty acid by approximately eight-fold at 2 hours after the drug. Trials of Riker 560 at daily dose levels of 2 g. are now in progress. In this and other trials to be reported later it has been found that at dose levels too low to show much change in plasma LCA, significant lowering of serum low-density Betalipoproteins occurs Discussion Most of the measured differences between SPG and heparin are quantitative rather than qualitative. Thus their influence on coagulation of blood seems to be dependent on the presence of the drug, as Lipæmia Clearing Activity Dose in g. Experi- No x too (CU) mental of (mean and range) SPG sene Condition Persons o Hr. 2 Hrs. 4 Hrs. Ver- SERIES A: SPG, Potassium Versenate. 20 Fasting 4 84 (41-133) 4 Fasting 3 4 (0-8) 4 Fasting 5 284 74 (0-5) (139-267) (10-268) 4 1.33 Fasting 4 1 97 30 (0-4) (5-304) (2-108) JO 0,5 Fasting 4 22 (7-52) 2 O SERIES B: SPG,+Sodium Versenate. IO Fasting 4 II (0-22) 5 Fasting 4 2 3 (0-4) (0-1) 13 (0-28) 10 (0-17) greater degree than heparin and consequently is a much stronger anion. Nevertheless, its potency is only about one-fifth that of heparin Even the slight anticoagulant effect seen after 1 v. injection can be further reduced by slowing the rate at which SPG appears in the bloodstream, as by subcutaneous injection, or by absorption through the gut. The rates of appearance in the plasma of the to differ 4 I Fasting 12 2 1 Fasting 8 --- anima erretem ceem I Fasting 8 6 (0-50) 4 (0-17) I (0-4) 2 (0-17) IO (4-22) 76 55 (0-150) (0-175) 71 42 (4-265) (0-217) 17 0 (0-12) (0-10) 33 8 (0-100) (0-17) 14 (0-50) (0-17) 2 1 Fasting 8 Fasting 4 4 I 14 (0-0) 4 1 Carbohy- 4 drate Meal Fat 4 Meal Fat 8 Meal 45 min later. 58 (22-118) 20 (0-78) 29 (0-83) (0-57) (0-22) (0-17) (0-0) TO (0-38) 4 I amount to produce the enzyme which when exhausted reappears at a limiting rate or, SPG replaces heparin which then extracts the enzyme from depots such as fat tissue. Clearly, there is a need for further research on the mechanisms inkerf der annearance in the TABLE 2 heparın of op hy the INCREASE IN FREE FATTY ACID DURING PLASMA INCUBATION (SERIES A). Free Fatty Acid Release in MicroEq/L./hr. (mcan and range) Dose (g.) No of Sample taken at SPG Versene Persons o Hours 2 Hours 4 Hours 4 o 240 185 (65-410) 4 2 $ (9-391) 1820 890 (1530-2170) (320-1450) 1.33 4 1910 690 (490-4190)(270-1100) magnesium in the filesLilla tort where lar chelates of Versene were inactive, while the alkaline salts were active, The ability to bring about the appearance 82 ·Lipemia Cleoring Activity in Dogs after Subcutaneous Injection 6 6, Heparin in Unlts/kg Sulfopolyglucin in mg/kg 5+ 5+ 4! 5 12 dogs! 3+ 2 (4) 50013) a Clearing Activity U 2+ 1 (3) 06 (3) 6016 200(3) 100(6) 1561 A035(2) HOURS I 4 Figure 5 Figure 6 EDTA NO EDTA Clearing Clotting 5 50 40 $ Clearing Units 3 30 Minutes 2 20 10 25 50 500 100 200 mg / kg 1000 83 ·TABLE 1 LIPAMIA CLEARING ACTIVITY IN MAN AFTER SULPHOPOLYGLUCIN PLUS VERSENE. lowering if given with a fatty meal. Delaying the fatty meal for 45 minutes did not improve absorption of the drug. Plasma samples drawn during serics A (Table 2) were incubated with an Ediol substrate and the increase in release of free fatty acid was measured. While 4 g. of SPG alone had no effect on lipolysis, the same dose when given with Verscnate increased free fatty acid by approximately cight-fold at 2 hours after the drug. Trials of Riker 560 at daily dose levels of 28. are now in progress. In this and other trials to be reported later it has been found that at dose levels too low to show much change in plasma LCA, significant lowering of serum low-density Betalipoproteins occurs. Lipæmia Cleaning Activity Dose in g. Experi. No x 1OO (CU) Vermenta! of (mean and sange) SPG sene Condition Persons o Hr. 2 Hrs. 4 Hrs. Discussion. Most of the measured differences between SPG ond han- SERIES A: SPG, +/- Potassium Versenare. 20 Fasting 4 84 (41-133) 4 Fasting 3 (0-8) 4 2 T'asting 5 1 184 74 (0-5) (139-267) (10-168) 4 1.33 Fasting 4 97 30 (0-4) (5-304) (2-108) 10 0.5 Fasting 4 22 (7-52) I SERIES B: SPG, +/- Sodium Versenate. 10 Fasting 4 11 (0-22) S Fasting 4 O gealer uegree than heparin and consequently is a much stronger anion. Nevertheless, its potency is only about one-fifth that of heparın. Firan the min. 13 (0-28) 10 (0-17) 2 (1ھ 2 I (0-10) Mb SPG may Fat 4 I Fasting 12 6 76 55 (0-50) (0-150) (0-175) LIC XUL Fasting 8 4 71 42 The rates of appearance in the plasma of the (0-17) (4-265) (0-217) lipæmia-clearing enzymc system seem to differ Z 1 Fasting 8 I 17 0 following 1. injection of the two drugs. Where (0-4) (0-72) heparin injection is followed immediately by the 2 Fasting 8 33 8 appearance of LCA, SPG injection is followed by (0-17) (0-100) (0-17) LCA expressed in delayed curves, the greater the 1 Fasting 4 10 14 7 dose, the later the peak. Some of the possible inter- (4-22) (0-50) (0-17) pretations of this phenomenon are activate an inactive precursor in the tissues to form the clcaring enzyme SPG may act as a cofactor, 4 1 Carbohy- 4 O 58 14 SPG may require the presence of an unknown drate Mcal (0-0) (22-118) (0-37) substance normally present in the blood in limited 4 1 Fat 4 amount to produce the cnzyme which when ex- Meal (0-0) (0-78) (0-22) 4 I hausted reappears at a limiting rate, or, SPG 8 10 29 replaces heparin which then extracts the enzyme Meal (0-38) (0-83) (0-17) from depots such as fat tissue. Clearly, there is a 45 min. later. need for further rescarch on the mechanisms involved. However, the rate of drug appearance in the bloodstream is involved This is substantiated to some extent by the similarity in shapes of LCA TABLE 2 INCREASE IN FREE FATTY ACID DURING PLASMA INCUBATION (SERIES A). Free Fatty Acid Release in The increase in oral absorption of SPG by the microEq/L./hr. (mcan addition of Versenate sodium is a new phenomenon and range) That it is not restricted to SPG was confirmed by Dose (E) No of Sample taken at successful oral absorption of heparin, dextran SPG Versene Persons o Hours 2 Hours 4 Hours sulphate, and polyethylene sulphonate (Windsor 4 o 240 185 (65-410) 4 2 S (9-391) 1820 890 (1530-2170) (320-1450) 1.33 1910 690 (490-4190)(270-1100) alkaline salts were active The ability to bring about the appearance 87 7 ·Lipemia Cleoring Activity in Dogs ofter Subcutaneous Injection 6, 65 Heparin in Units/ko Sullopolyglucin in mg/kg 5+ 54 4 5 (2dogs) 34 2(4) 50013 Clearing Activity (CU) 21 I (3) 06 (3) 200(3) 10016) 6016 25151 0.35(2) 3 HOURS Figure 3 Figure 6 . O EDTA NO EDTA Clearing Clotting 5 t 50 40 Clearing Units 3 30 Minutes N 20 10 25 50 200 500 100 mg/kg 1000 83 ·Effect of Riker 560 in Dogs after Peroral Dosage 67 260 Clearing Activity Clotting Time 200 50 100 40 50 Clothing Time (min) 200 30 เก๋ว) 44ละ 50เมื่อ 12 1100 50 25 HOURS 2 5 5 Figure 7 Grossman, WI (1954), ) Lab and Chn Wed, 43, 445 Um WT ----- HL... 11 in the blood of a lipæmia-clearing lipase by an oral drug without the necessity for concern about increasing coagulation times should stimulare further research into various disorders human lipid metabolism REFERENCES Dole, V P (1954), J Clin Invest, 35, 150 Liber, H B and Danishefsky, I (1958), AMA Arch Int Med, 102, 789 McLean, J (1916), Am J Physiol, 41, 250 Owren, P A (1959), Lancet, 11, 754 Quick, A J (1938), J Am Ned Assoc , 110, 1698 Windsor, Land Cronheim, G (1960), Nature, in prese, and unpuhlished results ·DISCUSSION of anticoagulant. With the discovery of heparin Sir JAMIES LEARNIONTH --I shall confine my remarks to the place of anticoagulants in peripheral occlusive arterial discasc, in whuch I have most experience. First, I wish to point out that, in addition to the phenomenon of thrombosis occurring in a patient ponse to the presence of a clot--more properly perhaps a thrombus. .L A. L. appreciated that the heparins of some species have With regard to the use of anticoagulants; of clotting. la and the mula nf there for more on IN PA 1 OLVO This leads me to my only question. I wonder whether an attempt has been made to suppress the Jonization of heparin by esterifying the remaining half-esters of sulphuric acid with organic groups which would also render the molecule more fatsoluble ? Such a heparin, if absorbed, would no doubt be trapped in the fat depots. There, with the aid of a friendly esterase, the original sulphated here is the same as that determined in a specimen of systemic blood However, in any collateral system -..71 Orion Thiemowha hamrohariof ver Lilas Leus mal normally dwell in the connective . T wme MY ! offer a simple explanation for the only situation in whuch heparin is known to enter the blood in quantity, namely the blood of the dog in peptone or angnhwant hi va inertian nt.la-reb organ, .. It is haranea of N- W I would Dr RILEY - Looking at the mast cell story in other ar retmener instead merely a this species. Dr VERSTRAETE-A third alternative besides surgery and anticoagulants can be considered in the 85 ·...-1.1.. Jana T. nutar ana OP treatment of recent peripheral vascular occlusion : the administration of lytic agents. Aortograms performed on a 72-year-old man showed acute complete occlusion of one common iliac artery and the contalateral internal liac artery. He was given 6,000,000 units of streptokinase locally through a percutaneous catheter and subsequently 16,000,000 units intravenously, by continuous drip. He recovered gradually and the second aortogram showed that both closed vessels had become patent. All arterial pulses were felt and the patient walked home unaided. examined electrophoretically for the presence of sulphopolygluci. No drug could be detected in these samples. On dialysis and 10-fold concentration of the dialysed urines, however, a very faint spot corresponding to sulphopolyglucin was detected. Dr WALKER - Dr D. M. Shepherd and I made a comparative study of the lipæmia-clearing effects produced by Riker 560 (sulphopolyglucin plus EDTA adjuvant) administered orally, and sulphopolyglucin administered intravenously. Lipæmia-clearing activity was measured by the spectro-photometric method of Grossman, A group of three rats, starved for 24 hours before usc,, each received by stomach tube R560 Effect of route of administration on lipæmia-clearing activity of sulphopolyglucin 10 min. OPTICAL DENSITIES (O.D.) Intravenous Route Oral Route 2 gm + SPG Saline SPG Saline Control 200 mg. Control adjuvant Initial 0.D. 0.691 0.508 0,637 0.573 O.D afrer 10 min 0.695 0 436 0 637 0.576 OD, after 30 min. 0713 0.313 0 645 0 582 O.D. fall in 10 min. 0.072 O.D fail in 30 min. 0.195 treated rats nor the controls showed any trace of clearing activity in the blood A group of three rats each received by intravenous injection sulphopolyglucin 5 mg per kg. . Figure 2 A similar comparative experiment was carried out in human subjects (Fig. 2) No clearing activity Assuming a daily urinary output of 2 litres, we calwas produced at three hours by orally administered culate that thus would correspond to a daily urinary R$60 (2 gm. sulphiopolyclucın plus adjuvant), glucin. excretion of about 5 milligrammes of sulphopoly although strong anticoagulant activity at 15 minutes, These results indicate that R560 is very poorly absoțbed from the, gut under our conditions of Effect of route of administration on lpæmia-clearing activity of sulphopolyets. glucin in rats. OPTICAL DENSITIES (O D.). IW Intravenous Route Oral Route ch SPG ct SPG Saline 200 mg /kg. Saline gs 5 mg/kg. Control + adjuvant Control suggests that the production of venous thrombosis YARA nee tha VP9 Initial O.D. 0521 0.554 0 590 0.484 O.D. after 0.367 0.561 O 586 0489 Fall in OD. O 194 There is no evidence that heparın has any physiological role cxcept perhaps in fat transport Figure / If it doo ne might exnect that in and good clearing actsvity at two hours, were obtained when sulphopolyglucın 3 mg. per kg was given intravenously. In addition, faur humans were given orally the lant. same dose of 2 gr. sulphopolyglucın plus adjuvant, and blood was taken for examination at two and four hours; while three others received orally I gm. Afr ROTHNIE-I should like to raise two plus adjuvant six-hourly to a total of 8 gm., the blood points which have a bearing on the use of extrabeing examined 24 hours after starting treatment and corporcal circulations, It is well known from the work 86 10 min. bolem ·of Roskam and Hugues that heparin prevents the adhesion of platelets in vivo. However, in the heparinised blood used in the heart-lung machines With regard to hairfall, we believe that it is a problem, not of the duration of treatment, but of the dose. When a certain dose is reached, of the magnitude of which we have as yet no knowledge, hairfall occurs after eight to ten wecks. So far its origin is not clear, the Intal. Dr SHARP - I should like to ask Dr Windsor whether he has any evidence that EDTA given intravenously with heparin potentiates the action of the latter. 0 Dr ITINDSOR - EDTA injected intravenously simultaneously with either oral or intrapreservative in the commercial preparation ? venous sulphopolyglucin does not potentiatc anti- Should we therefore usc freshly-made solutions of lipämic or anticoagulant effects. heparin? Work is being done at present by our chemists Secondly, I should like to ask Dr Beller whether to find suitable compounds which will be less polar the alopecia following heparin administration is due and therefore more casily absorbed by mouth. to prolonged administration and is therefore If the effects I described were merely due to the dependent on duration of treatment or is dependent histamine-binding so prevalent in dogs, thereby on the total dose, whatever the time factor. We releasing heparin from the mast cells, in addition had a recent patient who underwent perfusion for to the anulipamic effects there would be concomitant about one hour, followed by complete heparin anticoagulant effects in the same ratio ay is observed neutralisation, and who developed a patch of alopecia following heparin. 81x weeks after operation. Could this be due to heparin Thc rat is an unfortunate animal for oral despite the short exposure to the drug ? heparinoids since the intestinal tract 15 always full of fibres and food which bind the strongly charged Dr BELLER - On the question of whether sulphates However, a lqud diet for 72 hours it is better to use coumarins alone or hepann and enables the rat to demonstrate its ability to absorb sulphopolyglucin, Side-effects such as intestinal cramping are associated with large doses of Riker 560. We now find anulpemic effects with smaller doses by measuring the decline in low-density beta-lipopreparations. proteins ultracentrifugally. .....

·The Haematology Of Extracorporeal Circulation With And Without Hypothermia A. A. SHARP, B. EXCELL, E. SALZMAN and O. THORUP Department of Hæmatology, The Radcliffe Infirmary, Oxford The artificial extracorporeal circulation is now San dilinn ofaron la enfatmina established as a mechanical substitute for the heart and lungs or the kidney. While such circulations have been used for many years in the treatment of uræma, it is only in recent years that more complicated machines have been used to further the scope of cardiac surgery. There is now an established literatura nn hre chiant hu... - hazard is the severe hæmorrhage from the damaged tissues in the immediate post-perfusion period. Many reasons have been advanced to explain this hæmorrhagic tendency, namely, thrombocytopania, fikrinoltre fibrinn equivalent. Blood heparin equivalent = plasma heparin 100 Hæmatocrit equivalent x 100 Antihemophilic Globulin Assay-Biggs and Macfar. lane, 1957. In Vitro Hæmolysis Modification of the method of Stewart and Surridge, 1959. -.-1 1.... data removed and the supernatant plasma examined for the amount of free hæmoglobin. MATERIALS Anticoagulant For Perfusion Blood. Heparin (Boots Crystalline Cresol-free). The extracorporeal pump used in the present Hexadimethrine Bromide (Polybrene—Abbott.) series needs 8-12 pints of blood to prime before Edglugateconnecting to the patient's circulation Originally, EDTA (Ethylene diamine tetra acetic acid) fresh heparinised blood (20 mgms heparin/500 mis. blood) collected on the morning of operation was used Stored heparinised blood (30 mgms, heparia 500 ml. blood) collected the day before operation was found to be unsatisfactory as the platelet count LIAN .. Huku pH after autoclaving 6.7 METHODS. frorning of the operation, the angcoagulant Plarelet Count-Brecher and Cronkite (1950) EDGLÚGATE was substituted for heparin. Bloou Packed Cell Volume-Hawkesley micro-method collected into this anticoagulant on the day before Thrombin Cloiting Time. operation is heparinised and recalcified before the 0.1 ml. human thrombin (Fibrındex-Ortho) pump is primed. This was found to be a suitable 20 units/ml. was added to 0.5 ml. plasma and the substitute for fresh blood and does confer the clotting time recorded. additional advantage of leaving a reasonable period Estimation of Fibrinogen-Thrombin Titre Method for adequate cross-matching of the blood 30 mgms. heparin and s ml 10% calcium gluconate ate added Sharp, Biggs, Howie and Methuen, 1958, to each pint just before the pump is primed. Plasma Fibrinolysis Biggs and Macfarlane. 1957. Plasma Hemoglobin-Dacie, 1956. RESULTS. Prothrombin-One-stage Biggs and Macfarlane, Two-stage 1957 Pre-Operative. Protamine or Polybrene Titration of Plasma Heparin All patients were screened for defects of their hæmostatic and coagulation mechanisms. In only Levels ( modification of the method of Jaques, 3 patients were any significant defects determined 1943). 88 ·PROTAMINE TITRATION OF HEPARIN LEVELS 70 (Table 1). Two patients had a long bleeding time. In on the defect was inhented as the mother also had a long bleeding time. These patients were given steroid therapy for one 21.. adharth the 60 50 40 30 bleeding time post-operatively, but surgery had not improved his defect. The other did not have a bleeding tine performed post-operatively. In ont patient thrombocytopenia was found, but the bleeding tune was normal. No preoperative treatient 5533 given, and mild post-operative bleeding occurred. This patient, who was Very cyanosed, also had marked secondary polycythamia (hematocrit 68%). No difficulty was met in the irutial stages of perfusion in spite of the obvious increased viscosity of the blood During Perfusion. Platelets-In all patients the platelet count dropped the moment the pump and pauent's blood were mixed (Fig. 2). The degree of induced thrombocytopenia varied, but the platelet count was always below 150,000/cu.mm, and usually below 100,000/cu mm. This thrombocytopenia persisted all through perfusion, and was accentuated if the blood was cooled to Thrombin clotting time. 20 10) 0 2 4 8 10 12 14 16 18 20 22 24 26 28 30 Protamine Concentration mg/100 ml Figure I-Prolemine and Polybrene Titrotron (Thrombin) The clotting times of protamine - plasma- thrombin mixtures showing by the sudden shortening of the time ( break point) the amount of antidote equivalent to the plasma heparin. TABLE Patient Bleeding Time Platelet Count Diagnosis Cyanosis Family | Post op History Bleeding Time Post op Haemorrhage 37 0 Normal ++ Nil Mitral Stenosis Nil 40 Normal V.S.D. ? NII 210 Normal 24,000/ cu.mn Fallot ++ NU ¡Normal Defects of hemostatic mechanism discovered preoperatively. 89 ·TABLE Case Sex Haemolysis Fibrinolysis Thrombin Defibrination Titre Age Fibrinogen Post - Op Bleeding Therapy 50 60 71 me to 1/32 NII NII NAI 3 Ni1 1/16 2.5 GB i N11 Ot! 5 100 37 ++ 1/16 ++ NII NII 210 5 NII 1/64 NII Nil Ni1 26 17 Nil 1/16 ++ 5 Gms Nil 28 21 Nil ++++ 1/32 Nut Nil 33 0 22 ++++ 1/64 NI1 Nil NII The inter-relationship of hæmolysis, fibrinolysis and defibrination in 7 cases. .. 50,000 PLATELET COUNTS 26 CASES (20 HYPOTHERMIA) 300,000 250,000 PLATELET COUNT / CUMM. Figure 2- Platelet counts during the sequence of perfusion in 26 cases 200,000 150,000 loopoo 59000 of wat op PATIENT PUMP BLOCO START HYPOTHESMA REWARM ENO TWO HOURS OF PERAUSION OF PEAFUSION POST OPERATION 90 ·ye fechno. hon dependineatient blood was was used. Platelet Transfusions In some instances, thus calculated dose was As thrombocytopenia was invariable during inadequate, and titration of heparin was found perfusion, platelet concentrates were prepared and to be too insensitive for detecting residual heparin in the blood after the appropriate anudote was given. Such residual levels are usually less than Osmgmm. heparin/100 ml, blood. The thrombin clotting time of plasma was found to be a sensitive indicator of such traces of Fibrinolyris. heparin (Peden and McFarland, 1959). If the In all but 4 cases some degree of fibrinolytic activity could be demonstrated. Although a relatively insensiuve technique was used, the activity was very marked in 7 cases (Table II), In 9 patients demonstrable activity was present in the patient's blood taken just before perfusion started the remainder all showed active fibrinoly 815 by the end of perfusion. No onc factor was discovered to accentuate fibrinolysis, although in one case the sudden need to return sucker blood to the pump was accompanied by a dramatic increase of activity. Fibrinolysis dis Post-Parfusion Hamorrhage. appeared immediately after the end of perfusion, In the first few cases of this series, protamine sulphate was used to neutralise heparin in a dose of Fibrinogen Depletion. I mgm. for each mgm, of heparin equivalent that was In 3 of the 7 cases with severe fibrinolysis calculated to be present in the patient at the end of evidence of fibrinogen depletion was detected pcrfusion Such doses were found to be inadequate (Table II). In all the defect was only partial, but in and severe post-perfusion capillary bleeding occurred. two patients concentrated fibrinogen was given If the heparin antidote was given in amounts prophylactically. In none of these was there any ... annan in. AfhORAexcessive post-operative ha northage. Hemolysis. Other Coagulation Factors. hæmolysis. The levels of other coagulation factors were experiments performed by the not studied after perfusion as the results of such method of Stewart et al (1959) confirmed that the tests must be prejudiced by the amounts of heparin amount of hæmolysis is influenced by the type of or heparin antidote in the plasma. Where attempts plastic tubing used Stewart's findings that Tygon were made to assay antihæmophilic globulin and tubing was the best were confirmed, but British prothrombin in patients in the post-perfusion period, Beverage tubing was found to produce more only a partial depletion of these factors was found. hæmolysis than Stewart et al. (1959) measured. The Quick prothromom mme was usually prolonged, Autoclaving and washing after use increased the as is Owren's prothrombin time, but the level of amount of hæmolysis produced by Tygon tubing prothrombin has never been below 50%. In Heparin Levels. The dose of heparin given to the patent was 3. mgms /kg of body weight. As the blood used for perfusion contains 6 mgmns / 100 ml, the resultant level of the mixed between 4-6 mgins, on the blood volume of the patient Therefore, the amount of heparin in the blood was estimated by protamine or Polybrene titation at intervals during perfusion Serotonin and ATP. to determine the level of heparin at the beginning and the end of perfusion. Sarajas, Kristoffersson and Frick (1959) have Protamine or Polybrene Neutralisation of Heparin. Protamine or Polybrene was given at the end of perfusion in a dose equivalent to the total 91 * In vitro heparin .ha. A ir ·of both these substances were performed by Dr. Blashko and Dr.G.V.R. Born on blood samples obtained in 200000 PLATELET COUNTS (CASE 30) significant amounts of these toxic substances were detected in the plasina. Post-Operative Period. The rise in platelet count that develops in the immediate postperfusion period was not sustained rner-nnarativator T-1. tsapoo, cytopenia was not accompanied by a prolonged bleeding time. Heparin was not used postrernt.vely.hu!... " 11:9 IMI: * IPN "li. 1 .. LU levels below 8(wren and Aas, 1991). These low levels in PLATELETS /CU.MM. logpoo 50000 Candahn.-l Discussion The greatest danger to the patient MOURS TOYS narfrican have OPERATION POST OP. Figure 3–Platelet counts during perfusion in the post-operative period (case 30). hemorrhage from cut surfaces This bleeding if the antidote was given in amounts equivalent to takes the form of a diffuse ooze from all cult the heparin that could be measured in the blood at surfaces rather than fron only one bleeding point. the end of perfusion, it was obviously insufficient The amount of blood lost can be a matter of pints to produce adequate neutralisation of heparin Thus within a few minutes. It is surprising that such would suggest that the fall of blood heparin was not diffuse hæmorrhage is not the invariable result of due to destruction but to its escaping from the perfusion as the composite defect induced by circulation into the extravascular tissues, whence thrombocytopenia, active fibrinolysis, partial defib it returns when the normal circulation is restored. rination, and probable partial depletion of other Normally this reappearance of heparin in the blood coagulation factors, such as prothrombin and anti is rapid, but, if delayed, it could be an explanation hrnophilic globulin would, in most hæmatologists of the "rebound pheno:nenon." opinion, be an adequate explanation Yet these Heparin is an inefficient anticoagulant for predefects can exist together without hæmorrhage servation of stored blood, but it is the only one occurring avatlable for perfusion techniques In addition, it is The factor responsible for this post-perfusion a dangerous anticoagulant to use when there are fresh wounds or partial defects of hæmostatic or coagulation factors However, it is casily and quickly neutralised by cither protamine sulphate or Polybrene and these can effectively stop heparin-induced hænorrhage once it has started. Weight for weight Poiy brene was found to be more effective than prota nine and, if either has toxic effects, then Polybrenc is the drug of choice. Otherwise there does not appear to be any advantage in one over the other. The anticoagulant effect of both drugs appears to be similar, but the safety margin 42 ·is generous (4 mg./100 ml. blood). Neither drug was found to produce toxic effects. Fibrinolysis was very active in most cases, but was not found to be deleterious to the patient. Whether it was responsible for fibrinogen depletion, in the three cases described, is uncertain. Such defibrination or defibrinogenation must be a potential danger, but, from studies of obstetrical patients (Sharp, Biggs, Howie and Methuen, 1958), such partial defects as have been encountered in these cases do not merit routine prophylactic treatment with concentrated fibrinogen as suggested by Nilsson and Swedberg (1958). ---LI. hrok --- Then reappearing platelets are for the most part damaged, non-viable, and unable to survive. The platelet count only returns to normal when the patient produces a new generation of viable platelets. Platelet transfusions are not necessary if no hæmorrhage takes place. But if it does, fresh blood containing viable platelets should be given at the end of perfusion in order to raise not only the level of platelets, but that of any other plasma factor that may be depleted Fresh blood alone cannot control hæmorrhage when heparin remains unneutralised. Hemolysis was not a major problem in the present series. When increased, it appears to be sofely an indicator of faulty perfusion technique, and was not a cause of any particular complication in the patient. The use of profound hypothermia in the sequence of perfusion was not found to alter the blood picture apart from inducing more marked thrombocytopenia. This procedure appears to reduce rather than increase the incidence of humorrhage. The majority of the above statements are based on successful perfusions of moderate durauon (1-rd hours). Longer or inadequate perfusions seriously prejudice the patient's chance of survival and increase the incidence of severe hæmorrhage. In such cases, the blood picture does not differ in any significant way from that already described. ACKNOWLEDGMENTS We are indebted to Professor P. R Allison and Mr A Gunning, of the Nuffield Department of Surgery, without whose co-operation and interest this study would not have been possible, and to Miss Ann Kimbrey for technical assistance REFERENCES Biggs, R and Vacfarlane, RG (1957) Hunian Blood l'eden, J C and McFarland, JA (1959) Blood, 14, 1230. Coagulation and its Disorders Blackwelt Srientific Terkins, II A, Osborne, J j and Cerbode, r (1958) Am. Publications, Oxford Ichin Path , 30, 397 Brecher, Gand Crookite, t. P (1950) J Appl Physiol , Terkins, 1. A. Osborne, J J and Gerbode, F (1959) Ann 3, 365 intern Sied, 31, 658 Dacie, J V. (1996) Fractical Hematology. J and A Churchill, Rathnie, X Gand kinmonth, J B (1960) Brit Med. J. London, P 139 1, 73 Drew, CE, and Anderson, I V (1959) Lancet, 1, 749 Sarajas, ISS, Kristoffersson, R and Frick, M11 (1959) Jaques, L B (1943) Biochem J, 37, 189 Nature, 184, 1127 Von Kaulla, K N and Swan, H (1958) J thorac Surg. Seening, A (1959) Acta cbur Scand, 17, 55 36, 519 Sharp, AA (1958) Brit J. Hrmat 4. 28 Nilsson, I M and Swedberg, J (1959) Acta chur Scand, Sharp, A A Biggs, R , Ilowie, B and Methuen, D (1958). 117, 47 Lanret, 11, 1309 (wren, P A and Aas, (1950) Scand J Clin lab Invest Solandt, D Y and Best, C H (1940) Lancet, i, 1043. 3, 201 Stenart, J Wand Surridge, VP (1959) Lancet, 1, 340 93 ·of both these substances were performed by Dr. Blashko and Dr G. V.R. Born on blood samples obtained in 209,000 significant amounts of these substances were detected in the plasma. TOXIC PLATELET COUNTS (CASE 30) Post-Operative Period. The rise in platelet count that develops in the immediate post. perfusion period was not sustained ISQ000 ….. . PLATELETS /CU.MM. cytopenia was not accompanied by a prolonged bleeding time. Heparin was not used postpionpoo operatively, but phenindione was given prophylactically in three cases. In all, the response to the initial dose was dramatic, the plasma “prothrombin falling to levels below 8% (Owren and Aas, 1931). These low levels in a association with the thrombocytopenia already mentioned were accompanied by a prolongation of the bleeding time. 50000 In all cases it was felt that the risk of secondary hæmorrhage was too great to justify continuation of therapy. In two cases Vitamin K was given as an antidote, and in both the bleeding time returned to normal limits. Tours Discussion. The greatest danger to the patient KUARIT OPERATION POST OP. Figure 3-Platelet counts during perfusion in the post-operative the outcome of successful surgery can period (case 30) be jeopardised by post - perfusion hemorrhage from cut surfaces. takes the form of a diffuse ooze from all cut This bleeding of the antidote was given in amounts equivalent to surfaces rather than from only one bleeding point. The anount of blood lost can be a matter of pints within a few minutes It is surprising that such diffuse hæmorrhage is not the invariable result of perfusion as the composite defect induced by thrombocytopenia, active fibrinolysis, partial defib- - Intion is restorect. rination, and probable partial depletion of other coagulation factors, such as prothrombin and antı. hemophilic globulin would, in most hanatologists' opinion, be an adequate explanation Yet these Heparin is an inefficient anticoagulant for predefects can exist together without hæmorrhage servation of stored blood, but it is the only one occurring available for perfusion techniques. In addition, it 18 The factor responsible for this post-perfusion a dangerous anticoagulant to use when there are fresh wounds or partial defects of hæmostatic or coagulation factors However, it 19 casily and quickly neutralised by either protaninc sulphate or Polybrene and these can effectively stop heparin-induced hæ norrhage once it has started. hage. 'where nepali locultan JULI Weight for weight brcnc was found to be no hemorrhage occurred apart from that 'from more effective than prota Tinc and, if either has unsecured blood vessels. toxic effects, then Polybrene is the drug of choice. The measurable loss of heparin fron the blood Otherwise there does not appear to be any advantage during perfusion was initially interpreted to be due in one over the other. The anticoagulant effect of both 10 destruction of the heparin (Seening, 1959), but drugs appears to be similar, but the safety margin 9?

·30 30 20 20 THROMBIN CLOTTING TIME (SEC) CONTROL 10 25 50 75 100 PERCENTAGE REDUCTION IN FIBRINOGEN Figure 2--Effect of reduction in fibrinogen on the thrombin clotting-time. a reduction is equivalent to 300 mg. of fibrinogen per 100 ml of plasma. in nemal af - afier perfusion, to give a dose of andote based on the heparinising dose given to the patient, and thus related to body weight, and expressed as a rado of anudote to heparin. If unneutralised heparin was detected after thes, further doses of anudote wese experimental dogs atter pertusion and to human volunccers without produang aninh Quiga of coagulauon. There is # good margin of safety, as with the newer antidore Polybrene, in the doses UKUL used nepasis The heparin cuculating after perfusion is de- Tived from the heparising dose given to the patient (2-2.5 mg./kg.) and the perfused donor blood (20-25 mg per soo ml.), a varsable amount of włuch is used during bypass it is thus difficult after bypass 10 csurgate accurately and quickly the amount of heparin requiring neutralisation, we have found 11 more rapid and convenient than assaying the beparin 95 ·Abnormal Haemorrhage After Perfusion THE IMPORTANCE OF UN-NEUTRALISED HEPARIN AND FIBRINOGEN DEPLETION N. G. ROTHNIE and J. B. KINMONTH --St Thomas's Hospital Medical School, London The last decade his scen an increasing number 2. An absorptio.nctric method for atimation of of congenital and acquired carduc dcfects corrected fibrinogen follou cd by obscrvauon of the fibrin clot by open operation using an extracorporeal circu. for 1sts. lation. One of the problems of recent cardiac surgery Thrombin clotting-time (ingrain 1955; Rothnic is a disturbance of natural himostasis leading to and Kinmonth 1960). abnormal bleeding following total body perfusion. Thus is a more sensitive indicator of heparia This is mainly duc to the trauma intricted on the activity than the other clothing tests, c... the whole blood as it passes through the external circuit and blood clotting tine (Lee and White) and one-stase to the massive transfusion of donor blood pumped prothrombin time (Quick), used in previous studies. from the heart-lung machine. The hoparın, essential Thrombin (bovine) and 18/80 C Cl2 are added during perfusion, may also cause bleeding if it is to citrated platelet-poor plasma and clot formation incompletely neutralised alteru ards, tined. The main factors affecting the thrombin Abnormal bleeding after perfusion has been clotting-t1.mc, apart heparin, are the thrombin attributed to a deficiency of one or other of the clot- and fibrinogen concentrations. The test is made ting clements of the blood (Osborn ct al 1955: suitably sensitive to heparin activity by adjusting Allen 1958; Brown and Smith 1958; Von Kaulla the thrombin concentration to give a control clotting and Swan 1958; Gollub ct al 1939; 'Perkins et al 11.Tlc of 10-12 sec. (Fox, 1). 1959 and Nilsson and Swedberg 1959). However, temminwhenfus nf fihun scant attention has previously been given to the role of unneutralised heparin and fibrinogen denietion in its causation. Under the guidance of Dr G. I. C. Ingram, we have studied the importance of these two factors, for which corrective treatment is readily available. We used simple and rapid clotting gests which reduction is grcater than that found after an average can be donc by the heart-lung Icam at any time after perfusion so that any increase in the thrombin perfusion clotting-time after operation is probably due to 1. The thrombin clotting time to detect une heparın. (Fig. 2). neutralised heparin. This is confirmed by adding toluidine blue INCOAGULABLE 10 80 THROMBIN CLOTTING TIME (SEC) APPROX: LEVEL IN HEPARINISED PATIENT 20 → CONTROL 10 I 6 2 4 8 10 20 40 60 80 100 HEPARIN CONCENTRATION (Hg/ml) figure I-Effect of different heparin concentrations on the thrombin clotting-time 94 ·30 30 20 - - 20 THROMBIN CLOTTING TIME (SEC) -- CONTROL 10 . : 25 50 75 100 PERCENTAGE REDUCTION IN FIBRINOGEN Figure 2-Effect of reduction in fibrinogen on the thrombin clotting-time. 0% reduction is equivalent to 300 mg of fibrinogen per 100 ml. of plasma after perfusion, to give a dose of antidote based on th heparinising dose given to the patient, and thu related to hade maish ind - with the plnom stration of clotting-tim blue was a times were of cach pił unneutralised heparin and not to a marked decrease in fibrinogen In our early cases, using protamine as the heparin neutraliser, troublesome cozing from the wounds and I. brdo a ratio of 1.5:1. 95 ·Abnormal Haemorrhage After Perfusion THE IMPORTANCE OF UN-NEUTRALISED HEPARIN AND FIBRINOGEN DEPLETION N. G. ROTHNIE and J. B. KINMONTHSt Thomas's Hospital Medical School, London 2. The Inet plannda hne ran finner An absorptiometric method for estimation of fibrinogen followed by observation of the fibrin clot for lysis. Thrombin clorting-time (Ingram 1955; Rothnic and Kinmonth 1960). This is a more sensitive indicator of heparia This is mainly due to the trauma inflicted on the activity than the other clotting tests, c.g the whole blood as it passes through the external circuit and blood clotting time (Lee and White) and one-stage to the massive transfusion of donor blood pumped prothrombin time (Quick), used in previous studies. from the heart-lung machune. The heparin, essential Thrombin (bovine) and M/80 Ca Cl2 are added during perfusion, may also cause bleeding if it is to citrated platelet-poor plasma and clot formation incompletely neutralised afterwards. turned. The main factors affecting the thrombin Abnormal bleeding after perfusion has been er of the cloret al 1955; ; Von Kaulla Perkins et al 9). However, scant attenuon has previously been given to the -- TOO mg. or less, is necessary before there is prolongation in the thrombin clotting-time. This reduction is greater than that found after an average can be done by the heart-lung team at any time after perfusion so that any increase in the thrombin perfusion : clotting-time after operation is probably due to 1. The thrombin clotting time to detect un. heparın. (Fig. 2). neutralised heparin. This is confirmed by adding toludine blue INCOAGULABLE 100 80 40 THROMBIN CLOTTING TIME (SEC) APPROX: LEVEL IN HEPARINISED PATIENT 20 CONTROL . 1 10 2 6 8 10 20 40 60 80 100 HEPARIN CONCENTRATION (49/ml) Figure 1-Effect of different heparin concentrations on the thrombin clotting-time. 94 ·PERFUSION TIME (min) 60 15 PATIENTS 30 DOGS X PERCENTAGE REDUCTION OF FIBRINOGEN X X XX X X GAIN IN FIBRINOGEN X 90 20 30 40 50 60 70 80 PERFUSION TIME (min) Figure 5-Scattergram relating to percentage reduction in fibrinogen to the duration of perfusion in fifteen clinical and thirty experimental perlusions. EB. AGE I3 W 33KG CLOT LYSIS + + MIL NIL NL 300 300 200 200 TH.CL, Tcsec) PLATELET COUNT (10%cu mm) CONCENTRATION (mg%) FIBRINOGEN 100 100 (9) - POLEERENE (175mg/kg) on onO OP HEPARIN (2-5mg/kg) BEFORE PERFUSION AFIBANOGEN INFUSION PERFUSION 2 3 TIME (hours) Figure & Changes in the platelet count(*) fibrinogen concentration (o) and thrombin clotting-time (TH.CLT.) after perfusion for 24 hours + indicates fibrinolysis The effect of an infusion of fibrinogen is shown 97 ·With the tests described, the detection and - elimination of resídual heparin was found to be an The citrated platelet-poor plasma is recalcified and the opacity increase due to the fibrin clot ing a violet filter. Is converted to fibrinogen con. absorptiometer calibrated with plasma containing known quantities of fibrinogen PLASMA TOL BLUE B CLINICAL STATE A.S.D. WITH ANOMALOUS PULMONARY VEINS - PRE-OPERATIVE PRE-OPERATIVE POST-PROT 2:1 POST-PROT 2:1 POST-PROT 3:1 POST-PROT 3:1 MARKED QOZING + 0 10 20 30 40 50 POST OP DRAINAGE THROMBIN CLOTTING TIME (SEC) 3500ml Figure 3-A patient with atrial septal defect (A S D.) and anomalous pulmonary veins showing the use of the thrombin clotting-time in the detection of unneutralised heparin, which caused abnormal bleeding after perfusion 350 15 PATIENTS MEAN REDUCTION 12% MEAN PERFUSION TIME 55 min 300 250 FIBRINOGEN CONCENTRATION (mgm%) 200 90 150L 10 20 30 40 50 60 70 80 PERFUSION TIME (min) Figure 4–The changes in fibrinogen concentration in fifteen clocal perfusions plotred to show the rate of change in individual cases 96 ·REFERENCES Alen, J. (1959): In Extracorporeal Circulation P. 250 Osbon, J. , Mackenzie, R, Shaw, A, Perkins, H., Hurt, Thomas, Springfield, 14.nous. R. and Gerbode, 1. (1955) Surg Forum, 6. 9h Brown, I W. and Smith, W. (1953) Ann Intern Merl. Perkins, II., Osborn, J J and Grounde, F. (1959): Ann 49, 1035 Intern Med , 31, 658 Gollub, S, 'lin, A. H. Winchell, H. S, Ehrlich, T. and Rothose, Gand kinmonth, J B (196) Brit lied J. Weiss, W. (19592) Surgery. 45, 201 1, 73. Ingram, G. I C. (1953). Biochem.31,393. Rothnse, VG, Sorman, A. G., Steele, and Kirmonth, Ingram, G 1. C. (1955) J. Clin. Path, 8, 318 ). 13. (1980). Brut J. Surg (11 pres) Nilsson, I I and Snedberg, J (1959): Acta Chir Scand, Von Kaulla, K and Swan, 11 (1959) J Thorac Surg. 117. 47 36, 519 Het normal Dla HI 50 Coagulability During Extracorporeal Circulation With Particular Reference To Antihaemophilic Factor B (Factor IX) J. MATZKE, R. SMITK JENSEN and I. H, RYGG- The University Hospital and the State Serum Institute, Copenhagen We have ined to find the reasons for the senous the following methods: (1) The cephalin time of bleeding tendency often encountered during and Waaler (1957), which is sensitive to deficiency of after cxtracorporcal arculation It is usually not cxplained by the platelet count, bleeding ume, P and P test, whole blood clotting time, fibrinogen assay and tests for fibrinolysis. Our observations refer to four patients with Fallot's tetralogy and two with atrial septal defect. The heart-lung machine used was the Rygg- (Owren and Aas, 1951) and (3) the Quck test. Kyvsgaard tyde, in which a plastic bag is used as (4) In some cases the proaccelerın (factor V) test oxygenator and discarded after use. (Hjort, 1957) was carried out. The patients' clothing factors were studied by The cephalin ume was used as a scrcening test for the factors mentioned, FALLOT after an evaluation of the other three tests When a prolonged MOT cephalın time, was found, the following additions were made in turn to the reagents: barium cupin me 10 sulphate-adsorbed plasma, containing factor VIII and PTA; old serum, containing factor IX and PTA; adsorbed serum, containing (-meonly PTA For measurement of factors VIII and IX, modifications of the methods of Waaler (1959) and Stapp (1958) were used, When a deficiency of factor IX was found, the modifications of the cephalın test menzoned were used to measure the factor IX level. In the two ASD patients, the initial tests were all normal. Inainty o 100 m During hepannisation the cephalın tests fell to under 1%. Ibnoget from a marked decline in factor IX, and the Quick time was prolonged owing to the antithrombin Wodtransfusions action; the P and P test was less sensitive to this action, being normal. Factor V was depressed, but not enough to cause this prolongation in the Quick test. After protamine sulphate the tests were all nortnal, but in one-anda-half hours a secondary fall was found, particularly a great decline tromboga 500 in the cephalın test and factor IX. ma There is at present no adequate 99 procwert prothrombone brine 70 10 hmy hawhanage NX factor lactor normal DE 14 IN 500 Qurcie w 0 200 200 • • RUND ·VH AGE 52 Wt 44 kg. - MS CLOT LYSIS + NIL NIL NIC 400 300 FIBRINOGEN CONCENTRATION (mg%) 200 100 POLYBRENE TOTAL DRAINAGE 650 830 950 1100 PERFUSION 1 6 18 BEFORE PERFUSION 2 3 4 5 TIME Chours) Figure 7-Changes in fibrinogen concentration (-o) after perfusion for 50 min Post-operative blood drainage from the chest is shown. + indicates fibrinolysis no direct relationship between the degree of reduc- fibrinogen concentration, ca. roo mg, % (Fig 6). tion of fibrinogen and the duration of perfusion The plasma clot showed rapid lysis (within 20 min.), (Fig. 5). The degree of reduction depends on many A plasma specunen half an hour later produced variable factors which differ from case to case. One scanty clot which was rapidly lysed. There was marked cannot therefore forecast accurately the changes oozing from the various incisions An infusion of due to perfusion, Changes in fibrinogen were insuffi- 4.4 g. of human fibrinogen in 200 ml, of distilled cient to produce a failure of hæmostasis in these water was given with a return to normal fibrinogen patients and none showed evidence of abnormal concentration as estimated at intervals over the oozing, or fibrinolysis. following 3 hours. There was no evidence of fibrinolytic activity after the infusion and the thrombin CL I fibrinoproduce lacement ars One showed briefer and less marked fibrinolytic activity and fibrinogen depletion, and post-operative blood drainage was not excessive. F.L-A1- circulating fibrinogen. There is at present no satisfactory method of reducing fibrinolytic activity and treatment consists of fibrinogen replacement until activity ceases and 2 normal fibrinogen concentration is attained. 98 ·hours before the operation and heparinised (75 mg. heparin for 400 ml. blood). All patients received - hoog results of a protamine or Polybrene titration test. A scries of 10 tubes, tach containing 0.7 ml. of the anh 20 Figure 1 16 12 Glotting time in minutes normal range 0 Clotting time of 10 patients before the second dose of Heparin - antidote Figure 2 78 (min) Heparin im 1 ----- Meparin yli Heparin Soyl ml 8 RECALCIFICATION TIME RECALCIFICATION TIME heparin / ml 10 Y ME PARINI ML COM PROT SULF. YIML 50 y 25y 10 2. 100 GAMMA 40 POLYBRENE / ML 20 60 101 ·IX activity is highest at a pH of 7-35, being nearly inactive at 7.0. In the fourth patient the findings were similar to the first, but we tried to avoid the secondary prolonged cephalin ume by infusing factor IX along with the second injection of protamine sulphate. This infusion was repeated, with a decreasa in the cephalin time and cessation of the heavy blood loss The common finding in all these patients was secondary S 2 explanation of this. Two hours after perfusion there was a compensatory phase of hypercoagulability. in the four pauents with Fallot's tetralogy serious bleeding occurred before, during and after perfusion and was not controlled by heparinneutralisers, blood transfusion or surgical hæmostasis. The patients were treated with a human plasma preparation made from Cohn's fraction IV. This contains several blood clotting factors, in particular factor IX. The first and second of these patients, though the initial P and P and cephalin times were above the bleeding limit, bled heavily before perfusion and after heparin a very severe hæmorrhage occurred (Fig.). The cephalın test fell to under 1%, and the Quick time was prolonged to over 100 seconds. After protamine sulphate the cephalin time was less and the Quick time normal The secondary rise in the cephalın ume was found here also, with a corresponding severe and prolonged bleeding since the spontaneous hypercoagulable phase did not appear. After infusion of the factor IX preparation, the cephalın ume was shortened, with corresponding hæmostasis. The Quck ume showed no secondary prolongation, presumably because there was no fibrinogen deficiency nor prolonged heparin effect. As expected, infusion of fibrinogen had no effect on the bleeding. The third patient also had an initially prolonped с p P e р gical value. Heparin seems particularly to influence the factor iX level, and to ensure an effective andcoagulant effect during perfusion it is advisable to prolong the Quick ume in order to uulise the anuthrombin action, The P and P test is virtually unchanged throughout, and is not useful in this context. In uncomplicated cases, where the hypercoagulable phase comes soon, it may be better to start anticoagulant treatment earlier in the postoperative period. a $11011 wine aiter the heart was entered, it **L. REFERENCES Hjort, P F Scand J Clin Lab Invest. 9. Suppl 27, 1957. Ouren, PA and Aas, K Scand. J Clin Lab Invest. 3 201, 1951 Quck, A J J Biol Chem, rog LXXIII, 1935 Haaler, B A Scand J Clin Lab Invest , 9 322, 1957 Waaler, BA Scand J Chun Lab invest, II 194, 1959 Stapp, W F Scand J. Clin Lab Invest, 10 269, 1958 enceuve' verore but not atter perfusion. The factor Blood Coagulation Problems During Extracorporeal Circulation in Man M VERSTRAETE, A AMERY and C. VERMYLEN -- Department of Medicine, University of Louvain, Belgium Bleeding has been a major problem in the use of extracorporeal circulation in dogs and in man The hæmorrhagic state, someumes found in man bepaliny 10 9 trouhland t' 0 Jun114111sou vroud heat exchanges assembly was interposed in the inflow from one group to the other, it is generally accepted that the better results are a consequence of a careful routine and greater experience on the part of the Tona hata 100 ·DISCUSSION ry Dr BIGGS - Dr Matzke is to be congratulated on his discovery of a test, the cephalın time, which appears to give results which are correlated with the tendency of the patient to hæmorrhage. It is a pits that the results of this test have been interpreted as showing factor IX deficiency. The measurement Professor Ol!REN-As stated by Sharp, the oozing cannot usually be accounted for by the moderate Ldeficiency The mech draw your may play a role at least in certain cases, namely a This time Vivo cand -corne ...... .. Indeed, by the coagulability of fibrinogen Even correction of the clotung time by aged serum will not ensure that the technique gives a measure of factor IX. Thus, while Dr Matzke's results are very helpful in rescaling a test which may correlate with the bleeding tendency it may prove misleading if regarding the required dosage. the results are interpreted too narrowly at this The thrombocytoponia is largely caused by stage as indicating a specific defect in factor IX. trapping of platelet clumps in the filict. I believe that the main reason for platclet clumping is the Dr. SAATAMA - I should like to describe release of factor R by contact of red cells with foreign surfaces. Even very small amounts of factor R a study of a patient who died from cataclysmic drastically increase platelet adhesiveness and clumpfibrinolysis during operation for tetralogy of Fallot. ing. Intravenous injection of factor R in rabbits It throws light on the failure of substitute therapy causes, as mentioned previously, platelet clumping in superacute fibrinolysis, and verifies the efficacy and thrombocytopenia, presumably because of of a new antifibrinolytic agent C 766 or Insprol. trapping of platelet clumps in the capillary bed. The pauent was a girl aged 17. Before operation However, factor R becomes inactivated by plasma, there was only a slight thrombocytopenia and no the clumping 15 reversed, and the platelets reappear fihnnnlene in the circulation. Thus observation in rabbits is much like the temporary thrombocytopenia of bypass arculation as shown by Sharp, and the mechanisms may be the same. The thrombocytopenia of by-pass circulation may be largely prevented by devising pumps and using plastic tubes which give minimal trauma to the red cells. Further, the filter could probably be discarded if its only effect is to remove small platelet clumps, because the introduction of small aggregates of platelets into the Cr 6 hours circulation seems to be without harmful effect. The Laboratory tests showed only traces of fibrinogen clumps are broken down and the platelets are in the plasma. Quick's test in the presence of fibrino- released after a temporary trapping. een was 50% (factor V 40.9.). Von Kaulla's euglobulin technique showed that lysis was almost Thrombin immediate, while a mixture of Taroa boplutio generation toot olotting patient's plasma with 3 parts of normal plasma gave a lysis time of 15 minutes (normal over 180 minutes). Thrombo-elastography Addition 1 was characteristic of superacute 2 3 4 5 6 13 fibrinolysis and showed that the patient's plasma was capable of Selip. 11 11 11 11 lysing 10 times its own volume, ie the patient's total plasma volume was capable of lysing 1/32 Polytrone | 180+ |180+| 180+/ 180+| 180+ | 130+ fibrinogen in the order of 100 gms One can readily understand the */128 polybrane 255 155 158 152 150 147 failure of the standard substitute 14 1/256 Polybrose 50 20 27 15 15 25 In vitro, with 100 units of 14 inlubitor, the lysis time increased from 45 to 150 minutes; with 200 1/512 Polybrano 14 33 12 12 12 12 13 units, lysis was practically abolished. It seems that a clinical dose of 5 million units would attenuate or clotting flae in besonde 11 14 treatment. abolish the residual lytic activity Figure 1 103 ·hit ba ala. injected. A second series of the anti-heparin agent was needed in the post-operative period (15 cases). with lower concentrations (from 2 gamma/ml. to There were to cases where two injections of pro. 20 gamma/ml. with 2 gamma intervals) was used after tamine sulphate were given (Fig. 1). In 3 cases a the first neutralisation for further detection of traces supplementary dose of protamine sulphate was of heparin, given mainly because the surgeon felt that the patient Other techniques used were thc Lec and White was bleeding excessively, although the clotting time clotting time of venous blood, fibrinogen assay by and the protamine titration test were normal. If the method of Clauss (1957) and fibrinolysis observed the amounts of protamine given are averaged, it was found that the protamine-heparin ratio was only 1.1 in these 10 cases. The fact that a second dose of heparın antidote was needed in this group suggests that the optimal protarline-heparin ratio results obtained in cases of aortic stenosis explain the growing frequency of open heart surgery with cardiac by-pass in adults. Half of the patients had a ventricular septal defect. It is to be noted that patients with pulmonary stenosis and atrialseptal defects 3 mg. heparin/3 mg. Polybrene, an immediate and sustained normalisation of the clotting ume was obtained. The results were as favourable with - Ladainh The plasma fibrinogen was also decreased at the end of the cardiac by-pass (to about 140 mg. %) and a tendency to a further slight fall was noted in some patients. If the results of all cases are averaged, there is a 10 mg. per cent difference observed immediately after the slow administration of protamine -.AL 10, 25 and so gamma heparin/ml. (Fig. 2). It can be seen that in similar experiments with protamine sulphate, a rather limited range of concentration corresponds to normal coagulation times; higher or lower concentrations give prolonged clorting times because all the heparin is not neutralised or too much protamine is added and exerts an anti- ---LA REFERENCES Clauss, A (1957), Acta Hæmat, 17, 237 Perkins, A A, Osborn, J J and Gerbode, F (1959), Ann Cru, i o, Oliveira, A C and Neis (1958) Am J Phys Int Med, 55, 658 195, 440. Rothne, N G and kinmonth, J B (1960), B VJ I 73. Holemans, R, Anery, A and Verstraete, M, to be published Van Goldsenhoven, F (1926), Ann Soc Sci Brux, 45, 219 in Cardiologia, 1960. Von Kaulla, KN and Swan, H (1958), J Thor Chir, 36, 519 Holcmans, R., Vermylen, C and Verstraete, M (1960), Med Weiner, AE, Red, D L and Roby, C. C (1953), Amer Lup 2, 294 J Obstet Gynec 66, 475 Kimura, LT, Young, PR, Stein, RJ and Richards Weiss, WA, Gelman, J S, Catenacci, A J and Osterberg, RK. (1959). Tos and Appl Pharmac, 1, 185 A E (1958), ) ANA, 166, 603 102 ·DISCUSSION DT BIGGS - Dr Mlatzke is to be congratulated on tus discovery of a test, the cephalin time, which appears to sie esults whach are correlated with Professor OREN-As stated by Shaip, the cozing cannot usually be accounted for by the mndemte thmmhyttannin halnya 15 wetuviu us Plooy LLUN Ul uie Liceuling ume and reduced platelet adhesiveness by the in VIVO technique (Borchgrevink, Chr.: Acta Med. Scand. 1960, in print). Platelets in donor blood become regarding the required dosage, The thromboxytopenia ss largely caused by frapping of platelet clumps in the filter. I believe that the main reason for platelet clumping is the Dr SAMIAMIA - I should like to describe release of factor R by contact of red cells with a study of a patient who died from cataclysmic fibrnolysis during operation for tetralogy of Fallot. I throws light on the farlure of substitute therapy 1 superacute fibnnolysis, and verifies the efficacy of a new antifibrinolytic agent C 766 or Iniprol. The patient was a girl aged 17. Before operation However, factor R becomes inactivated by plasma, Tarot alotting 13 11 L! O hours. Laboratory tests showed only traces of fibrinogen in the plasma. Quick's test in the presence of fibrino released after a temporary trapping. gen was 50% (factor V 40°.). Voa Kaulla's euglobulin technique showed that lysis was almost immediate, while a mixture of Turaboplastin neration tot patient's plasma with 3 parts of of 15 minutes (normal over 180 minutes). Thrombo-elastography 1 Latlon 2 3 4 5 6 was characterisuc of superacute fibrinolysis and showed that the patient's plasma was capable of Salipo 11 12 21 lysing 10 times its own volume, ie the patient's total plasma volume was capable of lysing '/32 Polybrose 180 1804 1804 1804 1804 180+ fibrinogen in the order of 100 gms One can readily understand the '7128 polybrane 155 155 158 157 150 147 failure of the standard substitute */256 Polybrose 50 20 17 15 15 15 In vitro, with 100 units of inhibitor, the lysis time increased from 45 to 150 minutes; with 200 1/512 Polybro. 44 33 12 12 12 units, lysis was practically abolished. It seems that a clinical dose of Smullion units would attenuate or abolish the residual lytic activity. clotting tipe la seconde 103 14 14 treatment. 14 12 23 Figure / ·injected. A second series of the anti-heparin agent with lower concentrations (from 2 gamma/ml. to 20 gamma/ml. with 2 gamma intervals) was used after the first neutralisation for further detection of traces of heparin. was needed in the post-operative period (15 cases) There were 10 cases where two injections of protaminc sulphate were given (Fig. 1). In 3 cases a . L. T AJ IVIL.. the amounts of protamine given are averaged, at was found that the protamine-heparin ratio was only 1.1 in these 10 cases. The fact that a second dose of heparin antidote was needed in this group The majority of patients were infants ; the food suggests that the optimal protamine-beparia ratio results obtained in cases of aortic stenosis explain the growing frequency of open heart surgery with cardiac by-pass in adults. Half of the patients had a ventricular septal defect. It is to be noted that patients with pulmonary stenosis and atrial, septal defects suslaucu porudisJLIUI U ule Liului www ned The results were as favourable with 5 mg. heparin/s mg. Polybrene per kg. body weight With a 2/1 Polybrene-heparin ratio, no fall ander միյսկսll vi Vietdi. Jupilat. W Illl ki, Issuts of all patients were grouped, however, no difference could be noted. Tui. ELately after the slow administration of protamine advantage of Polybrenc over protamine sulphate is the finding that in vitro a wide range of concentrations neutralises a fixed amount of heparin for 10, 25 and so gamma heparin/ml. (Fig 2). It can be seen that in similar experiments with protamine sulphate, a rather limited range of concentration corresponds to normal coagulation times; higher or lower concentrations give prolonged clotning times because all the heparin is not neutralised or too much protamine is added and exerts an anticoagulant activity by itself. A final advantage is that with Polybrene no fall in blood pressure or agglutination of red cells is observed in the doses used even after rapid intravenous administration, was based on the protamine or Polybrene titration test. On retrospecuve analysis of the quantities used, it was found that the protamine sulphate-heparın ratio was 1.5 and the Polybrene-heparin ratio 1.4 for all cases where only one dose of heparin antidote REFERENCES Clauss, A (1957), Acta Hæmat, 17, 237 Perkins, A A, Osborn, J J and Gerbode, F (1959), Ann Cruz, w o, Oliveira, A C and Meis (1958) Am J Phys Int led, 31, 658 195, 440 Rothrie, N G and Kumonth, J B (1960), B M.J 1 73 Holemans, R, Amery, A and Verstraete, M, to be published Van Goldsenboven, F (1926). Ann Soc Sci Brux , 45, 219 in Cardiologia, 1960. on kaulin, K N and Swan, H" (1958), J Thor Chir, 36, 519 Holemans, R. Vermylen, C and Verstraete, V (1960), Ved Weiner, A E, Reid, D E, and Roby, C C (1953), Amer Lep 2, 294 J 'Obstet Gynes 66, 475 Kimura, CT, Young, PR, Stein, RJ and Richards Weiss, WA, Gelman, J $, Catenacci, A J and Osterberg, RK (1959), Tor and Appl Phirmac, 1, 185 HE (1958), JANA, 166, 603 102 ·tration of factor IX he obtained in the concentrate large increase in factor IX activity after administra. tion of the concentrate NEUTRALIZATION OR HEPAI- BY >OTAMIN Menu I (HIFONDS) 43 2018 whether these assays represent the truth, in view of the poortle artefact in respect to the utbromhin VI phenomenon of Kowalski The only dissimilar- AT IS the fall in the platelet count fahing bypass whereas there is Do fall following the streptokinase hom. This can be related to the pecunial loss of platelets during the byt 255 The development of a labor tory defect as i constant finding after bass. Occasionally there is co abnormal biccding and cry casionally it " catastrophic. Hicre often there 18 abnormal kleedag as manifest by the blood coming through the chest drains, which goes on for several hours bers not of sufficient magnitude to crase enacty. A 600 1.1.1 100% 201 400g TO has passed. the use of inhibitor of plasminogen activation. Dr SHARP- I should like to ask a question about the amount of heparin used in the perfusion blood. I feel that the platclet clumping and resultant thrombocytopenia seen during perfusion are due to the heparin levels used being too low to prevent nhen we':'••••••• We k Hager: possit': Dr factor in an incro centrate the reas. Dr NT. LYCIS of heparin during perfusion. ; wf .. tritop 10 houdy explain the lack of serotonin release mentioned by Sharp. w anc $15 use aft bucplokinase and plasmın therapy. At the hepann has to be given immediately Name ume, to prevent a new thrombotic tendency, I should like to ask Dr Matzke what concen Even to the patients he described. He got quite a Dr SHARP I agree with Professor Ouren that factor R docs exist in hemolysates and does clump platelets, but we have not noticed increased clumping or thrombocytopenia in those cases where hamolysis was excessive. With regard to Dr Douglas's remarks, it has been our experience that fibrinolysis disappcars within 2 hours of perfusion and in the one case where defihrination existed but was not treated the mal in about 6 hours has passed the iing secondary of factor -L, kalcife We had or factor IX normal pla a I wonder whether the colkype of neutralising - Shanolytic drugs like epsilonibrinolysis may Itralisation may mbi. If lysis is be due to ·I I PLATELETS I PLASMA BO a BYPASS abnormality there are a number of PERCENTAGE O.--0 STREPTOKINASE collateral methods of approach Dogs can be put on the bypass without heparın or protamine and they develop a hæmorrhagic state. FIBRINOGEN The dog, of course, has a con- PLATELETS PLASMINOGEN siderable capacity under specific circumstances for endogenous 100 production of heparin, but this nevertheless as a pointer that this defect is more complex than merely a question of heparin neutralisation. Another pointer, which I think can help to disentangle the heparin/protamine issue from the 60 remainder of the problem, is the fact that there is often a defect many hours after the operation when, even if no heparin antagonist 40- had been given, one would have expected the heparin in any case to have disappeared in virtue of its usual" in vivo " survival time. 20 The fibrinogen fall may on occasion be progressive for nine hours after the bypass. More often the initial fall is sustained despite the continued transfusion of fresh 24 HOURS 24 HOURS 24 HOURS blood. This supports the concept that there 19 continuing Figure 2 abnormality, which is probably proteolytic in its nature. Most of n- NOTICIAS- Thahmapin. darant .. hinta the reports today have shown only the occasional occurrence of increased fibrinolytic activity. These determinations have, however, been made on the basis of whole-clot lysis orlysis in dilutions of plasma Before the possible importance of proteolytic activity is decided it should be studied by more modern and sensiave techniques-the use of tagged fibrinogen and plasminogen-enriched tagged fibrinogen. Fig. 2 shows the mean results from 12 bypass operations and 4 streptokinase infusions It will be seen that with the bypass procedure there is an appreciable fall in plasminogen concentration Some collateral evidence for the proteolytic As has been stressed today it is certainly correct nature of the bypass defect can be found in a comto study initially the question of proper neutralisation parison with the defect following streptokinase of heparin by protamine or Polybrene. I am not infusion (Figs. 2 and 3). I have some difficulty in certain that I know how to ensure that the heparin / protamine titration has been accurately BY PASS Figure 3 accomplished. 'The thrombin STREPTOKINASE clotting time and the whole blood coagulation time techniques have PERCENTAGE serious limitations. It can be shown that Polybrene, protamine and AHG CHRISTMAS FACTOR V FACTOR VII heparin are all inhibitory to 100 (x) thromboplastin generation at a concentration which does not interfere with the thrombin 80 clotting time of the plasma (Fig 1) The whole blood clotting time has 60 the same type of technical limitation, I think that we have to use the thrombin clotting time and 40 the whole blood clotting time in our attempts to judge the efficacy of the neutralisation effect, but it 20 is probably wise to be aware of their limitations In an attempt to sort out 24 HOURS 24 HOURS 24 HOURS 24 HOURS the nature of the hæmostatic 104 ·20 70 knowing whether these assays represent the truth, in view of NEUTRAUZATION OF HEPARIN AND ANTITHROMBIN YI the possible artefact in respect to BY PROTAMINE SULPHATE the antithrombin VI phenomenon THROMBON TIME of Kowalski. The only dissimilar (SECONDS) ity is the fall in the platelet count following bypass whereas there is no fall following the streptokinase A-MORMAL PLASMA + ANTITHROMBINY infusion. This can be related to the mechanical loss of platelets during 40 8 - NORMAL PLASMA + HEDARIN the bypass. The development of a labora- 30 tory defect is a constant finding after bypass. Occasionally there is no abnormal blecding and very B occasionally it 15 catastrophic. More often there is abnormal bleeding as manifest by the blood coming through the chest drains, which goes on for several hours but is not of sufficient magnitude to cause anxiety. PROTMINE In the therapy of this condition 16009 go 1% 40001 20004 400ng QOY anas SULPHATE It is important in the first place to CONCENTRATION do what one can to ensure adequate heparin neutralisation, and to be Figure 4 prepared to transfuse with fresh blood until the "proteolytic storm has passed. We know from Dr Niewiarowski's paper that There is a possibility for the future in the use of Hageman factor is depressed by fibrinolysis. It is epsilon - amino-caproic acid as an inhibitor of possible that the factor IX depression found by plasminogen activation. br Matzke was due to a depression of Hageman Dr SHARP - I should like to ask a question about the amount of heparin used in the perfusion blood. I feel that the platelet clumping and resultant thrombocytopenia seen during perfusion are due to the heparin levels used being too low to prevent Dr NIEVIAROITSKI - The Hagernan factor 10 gooor Pony 925 SOOLS pages 1 .. levels of heparin during perfusion. protamune titration. In an experiment, heparin and antithrombin VI were added to human plasma to T).... Orvni may explain the lack of serotonin release mentioned by Sharp. necessary than for neutralisation of heparin (Fig. 4). Dr SHARP- I agree with Professor Owren that factor R does exist in hæmolysates and does clump platelets, but we have not noticed increased clumping or thrombocytopenia in those cases where hærnolysis was excessive. With regard to Dr Douglas's remarks, it has been our experience that fibrinolysis disappears within 2 hours of perfusion and in the one case haca JACL I wonder whether the concept of neutralising fibrinolysts with antifibrinolytic drugs like epsilonamino-caproic acid is well-advised Fibrinolysis may be more often beneficial, and neutralisation may induce unwanted intravascular thrombt. If lysis is 105 ·investigations now in progress seem to show the same results. These are, I know, surprising, but I do nnt hal.ava nia ya maliar farinn minn - TVY obviously doing harm, such treatment must, of course, be given. Mr ROTHNIE - I would like to stress that abnormal bleeding may not only be due to hæmostatic defects but to surgical causes. Meticulous surgical hämostasis is as essential after perfusion as carrying out clotting tests. I am most interested to learn from Professor Owren about factor R and platelet clumping, and relieved to hear that such clumps are reversible. I assure him that we do not use the filter in the bypass circuit to remove the platelet clumps but to stop foreign material entering the patient's arterial circulation. Fortunately platelets are the only material we have found on the filter ; one day we may have the courage to dispense with it, as he suggests, and so reduce the platelet loss. However, such platelet loss as we have found does not seem to have any serious effect on hæmostasis. We have not found, as Dr Douglas has, that fibrinolysis lasts longer than a few hours after the end of perfusion. It seems to burn itself out gradually in 2-3 hours, and our object in treatment has been - Ler infusion ases. We suggested in DTD ACTO Wy have found a great cated with .myeloma of course, these patients did not have a major operation. We seem to use larger amounts of heparin (3 mg. per kg) than the other investigators. I do not know whether this has a bearing on our better results in cardiac surgery, in terms of avoidance of platelet-clumping or intravascular coagulation. In dogs undergoing a cardiac bypass anuthrombin VI may appear but also another antithrombin not adsorbed by barium sulphate. Dr MATZKE -- So far we have no adequate explanation for the phenomena i described My report is a preliminary one, but more extended

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