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©2007 Schattauer GmbH,Stuttgart AnniversaryIssueContribution Thrombosis and haemostasis research: 1957–2007) Stimulating, hardwork andfun y( Margareta Blombäck Department of Molecular Medicine and Surgery, Karolinska Institutet, Karolinska University Hospital,Stockholm, Sweden Anniversar odoresearch is to be inquisitive,tobeprepared to setout The first attemptatclinical useoffraction I-0 prepared from th in unexpected directions in response to newfindings –be humanplasma wasmade on apatient with acquired fibrinogen 50 T theynegativeorpositive. Research is likeanincurable in- deficiency; however, the patient went into averysevere fibrino- fectious disease –once you’ve got it, you’ve got it for life. A lytic state.Welater understood thatpyrogens cangiverise to fi- negative result or aresultthat contradicts what youpreviously brinolysis (wehad,for the preparation, used waterwhich wasnot believedoften leads to newideasand inventions.Research also pyrogen free).Weknew, however, thatCohn’sfraction Icon- givesyou friends all over the world, both collaboratorsand com- tained an antihaemophilic component, so working with Inga petitors. BelowImention just some colleagues with whom we Marie Nilsson from Malmö, we testedfraction I-0 for the pres- performed the basic and clinicalresearch,but rest assured: Ihave ence of factor VIII (FVIII). Thefirst sample testedgaveavery not forgotten anyone. high yield. Later it wasfound out that we had used haemophilia Bplasma in testing the activity of this sample.Wehad notyet characterized Stockholm’shaemophiliacs in termsofhaemophi- 1950s: Fibrinogen, factorVIII, vonWillebrand lia Aand B. However, it did indeed have 100% yield of the FVIII factor(VWF) activity of the original plasma when testedproperly. Dr.IkuoYamashina (3),watching me and Inga Marie sitting After afew years studying literatureand socialwork, respect- on eitherside of awaterbathtilting tubestostudy clot formation, ively, BirgerBlombäck and I, then24years old, started studying said “Maybe or maybe not”. Ihope he meant this as ajoke. We medicine at KarolinskaInstitutet, where we are still doing re- continuedtopurify fibrinogen and FVIII from fraction I-0(2–8). search.In1951 Birger Blombäck wasasked to join the research Birger also continuedwith structural analyses of fibrinogen (3). group of Erik Jorpes, the "heparin man". As Birger’swifeIwas At thetime we were fortunate enoughtohavegood financial also allowedtojoin the group (1).Jorpes had acommission from supportfrom ErikJorpes, whohad very high royalty incomes Vitrum (a private firm laterincorporated with Kabi/Pharmacia/ from Vitrum (donated tax-free to research). He alsoreceivedbig Pfizer) to test heparin activity and the analysis required purified grantsfrom the NationalInstitute of Health (NIH). Generous fibrinogen. This wasprepared from bovine blood,which we supportwas also providedbyJorpes’friends, the thoraxsurgeon fetched from the slaughterhouse, travelling by tram and laterby ClarenceCrafoord,and the headofStockholm Blood Centre, motorbike, lugging the blood around in large glass bottles. The ErikSköld,who provided the human blood free of charge. Cra- fibrinogen (preparedaccording to the methodsthen in use) was, foordneeded fraction I-0 for thosepatients whowere bleeding however, stable onlyfor afew hours. Birger,who has always read duetofibrinogendeficency(fibrinolysis due to pyrogens?) (3, the literaturemore carefullythan I, wasintrigued by Edward 4). In order to test the heparinactivity in the blood,wepartici- Cohn’sidea of plasma fractionation and demonstration of fibri- patedinthe second heart-lungoperation ever performed in nogen in fraction I. We found thatwhen fraction Iwas extracted Stockholm, in 1955. This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited. in the presence of high concentrations of glycine,the solubility In 1956 IngaMarie Nilssonhad apatient, Birgitta, with severe of fibrinogen wasreduced butmost of the impurities went into pseudohaemophilia (in1957 identified as being vonWillebrand ´s solution. The stable fibrinogen fraction thus obtained we called disease [VWD] type 3) whonolongertoleratedbloodtransfusions fraction I-0 (2, 3). Theclinical studiesdescribed in the following andwhosemenstrualbleedingwas life-threatening. We decided to would probablynot have beenallowedtoday, buttherewere no trytostop the bleedingwith fraction I-0 (containing FVIII). It had ethical committees at thattime. to be prepared in arush. Thetrains andaeroplanesenlisted to Correspondence to: Received May12, 2007 MargaretaBlombäck Accepted May21, 2007 Dept. of MolecularMedicine and Surgery Karolinska Institutet Prepublished onlineJune 12, 2007 Clin. Chem. Build. L205 doi:10.1160/TH07–05–0343 Karolinska University Hospital Stockholm, 17177Sweden ThrombHaemost 2007; 98: 8–15 Te l.: +4685177 4437, Fax: +46 8312 438 E-mail: [email protected] 8 Blombäck:Thrombosisand haemostasis research transportthe freshly preparedfraction I-0 from Stockholm hadto of Åland,who had aformofpseudohemophilia, describedin wait until we got it to them. But it did have adramatic effect: bleed- 1926 by the Finnish doctor Erik vonWillebrand (13).Most of the ing stopped, the FVIII levelincreased andthe bleedingtime was population living in the archipelago of Åland,situatedverynear normalized (4, 8–12). In different studieswetriedtofigure out Stockholm, are of Swedish origin and speak Swedish. Prompted whythe bleedingtime wascorrected.Birger and ourteam pre- by Erik Jorpes, whowas born and raisedthere, we took atripto paredthe fractions,and IngaMarie wasthe one mainlyincharge these islands,tostudy the surviving members of the originalvon of treating the patients. Purifiedfibrinogen and purified FVIII had Willebrand family(for results seebelow). Becauseofdifferences 1957–2007) no normalizingeffect on the bleedingtime, whereas fraction I-0 in voltage,the large centrifuge we had brought from Stockholm devoid of FVIII or prepared fromhaemophiliaAplasma corrected to spin down platelets did not work, forcing ayoung doctor to y( it. We also observedthatthe half-lives of FVIIIwerequitediffer- makedaily flights back and forthtoStockholm. Everyday he entinpatients with haemophilia Aversus thosewith VWD (4, 8) flewout with patient blood samples (and duty-free liquor) and (Fig. 1). In this wayweshowedthe existence of anew “anti-bleed- worked with Birgertoprepare the platelets for the assays (4,11). ing”factorabsentinVWD (byusfirst called the bleedingtime fac- The investigation showedthat the findings in the original family tor) andwenamed it “von Willebrand factor” (4, 12). were the same as thoseinthe patients in Sweden. Beforetravel- Anniversar Most of the fraction I-0 thatwas prepared wasgiven to pa- ling to Åland,ErikJorpes had met Prof. RudolfJürgens on the th tients with haemophilia AorVWD.For the first tenyears,itwas streetand revealed ourplans to him, so Jürgens, whowas on his 50 prepared aseptically (as we lost too muchofthe FVIII in filters waytothe islands to hunt seals, of course also brought ateam to used at thattime), by the devotedstaff in alaboratorywheredur- investigatethe same individuals. Theyfound adecreased levelof ing the first years insulin and peptideswere also prepared from FVIII, just as we did. We could confirm our suspicion that the animalintestines (3, 4). Swedish patients had the same disease and the same heredity as In 1956–1957 IngaMarieNilsson and I, accompaniedbya those from the Åland archipelago (11, 14). geneticist,travelledaround central Sweden in acar thatgaveus In 1958 IngaMarie, Birgerand Iwere going to present ourre- headaches. (Halfwaythrough the trip we discovered the exhaust sults at the VII th InternationalSociety of Hematology Congress system wasbroken.)Our mission wastostudy different families in Rome. We travelledinour car, whichhad alot of problems withpseudohaemophilia,aswecalleditthen. We visited high with its brakes. En route,wepassedthrough Vienna, where we and low, rich and poor alike (10).At thattime heredity could only visited anothermeeting. Laterwestayedinasmall place on the be analysedbyblood group determination; nonetheless, in two Italian east coast, happilydrinking the Italian wine there and in out of ten familieswefound a(healthy) child whose purported othersmall places.Arriving in Rome one daylate, we found that fatherwas notthe real father. IngaMarie’sboss (JanWaldenström) and ErikJorpes had given The clinicalfeatures,heredity and some laboratoryfindings us up for lost and were discussing whoshould present our find- in our patients were similartothose of patients from the islands ings. The congress participants were invitedtomeet the Pope Figure1:Difference in sur- This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited. vival time forfactorVIII in apatient withseverevWD type3(left)and apatient withmoderate haemophi- lia (right) in connection withhysterectomy and ap- pendectomy, respectively. Theadministered factorVIII is expressedasU/g protein. At the first occasion, thus3.9 x 300 U= 1,170 Uoffactor VIII were given.The broken arrow represents freshblood admin- istration.(printed withper- mission from Haemostasis 1992; 22: 239.) 9 Blombäck:Thrombosisand haemostasis research Pius XII, whotold us that he wasnot against abortions if the child In Pehr Edman’slaboratorywestudiedthe amino acid se- wasdiseased (a haemophiliac). quenceofthe human fibrinopeptides A(FPA) and B(FPB). Allthrough this timewehad to earnaliving, in addition to When we had returned to
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