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Cilj: Stečena hemofi lija A (SHA) je rijetki stečeni poremećaj zgrušavanja zbog pojave autoprotutijela, najčešće na FVIII i potencijalno smrtonosan ukoliko se rano ne prepozna i liječi. Cilj je prikazati rezultate liječenja stečene hemofi lije tijekom desetogodišnjeg razdoblja u KBC Zagreb. Metode: Analizirani su retrospektivno rezultati liječenja bolesnika sa SHA u Zavodu za hematologiju, KBC Zagreb od 2010–2020. Rezultati: Liječeno je 19 bolesnika, 11 muškaraca (58%) i 8 žena (42%). Medijan dobi iznosio je 68 godina (raspon 33–81 godina), medijan aktivnosti FVIII 5% (raspon 1–15%), inhibitora 6, 8 BIU/dL (raspon 1– 2000 IU/dL). Transfuziju eritrocita trebalo je 14 bolesnika (73, 7%). Kod 58% bolesnika bio je poznat uzrok SHA (oligodendrogliom, B-CLL i AIHA toplih protutijela, AIHA toplih protutijela, AIHA hladnih protutijela, reumatoidni artritis kod 2 bolesnika te pemfi gus kod jedne bolesnice, polimijalgija reumatika u 3 bolesnika, i jedna postpartalna SHA). Hemostatsku terapiju primilo je 17 bolesnika (89, 4%). APCC (FEIBA) primalo je 12 bolesnika (70, 6%), rFVIIa (NovoSeven) 5 bolesnika (29, 4%). Svi su bolesnici primali eradikacijsku terapiju, ciklofosfamid i kortikosteroid 17 bolesnika (89, 4%), 2 bolesnika kortikosteroid (10, 6%), a rituksimab 3 bolesnika u 2. liniji (15, 8%). Medijan eradikacije inhibitora iznosio je 18 dana (raspon 3–300), a za postizanje KR 32, 5 dana (raspon 15–300). Medijan preživljenja od trenutka dijagnoze je 21, 5 mjeseci (1–90 mj). Prva linija liječenja dovela je do KR u 18 bolesnika (94, 4%) bolesnika. Troje bolesnika (15, 8%) dožvjelo je relaps (dvoje bolesnika 1. relaps, a jedna bolesnica 2. relaps) te su ponovno postigli KR kombinacijom kortikosteroida i ciklofosfamida (2 bolesnika) a jedna bolesnica na rituksimab. U praćenju je umrlo 6 bolesnika (31, 5%). Nitko nije umro od posljedica krvarenja. Uzroci smrti su sepsa kod 2 bolesnika, pneumonija, kardiogeni šok, kardiorespiratorni arest te infarkt miokarda kod ostala 4 bolesnika. Viša aktivnost inhibitora bila je povezana s poznatim predležećim uzrokom te je uz dob, komorbiditete nepovoljno utjecala na preživljenje, a infekcije su uz komorbiditet (kardiovaskularne bolesti) glavni neposredni uzrok smrtnosti. Zaključak: Rezultati 10-godišnjeg liječenja stečene hemofi lije u KBC Zagreb u skladu su s rezultatima europskih registara uz visoku stopu kompletnog odgovora (>80%) ali i uspješno liječenje relapsa.

Bakgrund - Hemofili A och B är X-kromosombundna blödarsjukdomar, som beror på genetiska avvikelser i de gener som kodar för koagulationsfaktor VIII respektive IX. I dagsläget förlitar sig hemofilipatienter på kontinuerliga intravenösa injektioner med faktorkoncentrat, för att förhindra att potentiellt livshotande blödningar uppstår. Genterapi med rekombinanta adeno-associerade virus (AAV) skulle kunna erbjuda ett kurativt behandlingsalternativ, genom införandet av friska arvsanlag i hepatocyter. Syfte - Syftet med den här litteraturstudien var att undersöka huruvida genterapi medierad av AAV-vektorer är en effektiv och säker behandling mot hemofili A och B ur ett långsiktigt perspektiv. Metod - Studien är genomförd som en systematisk litteraturstudie och är baserad på sex originalartiklar framsökta via databasen PubMed, med sökorden "hemophilia AND gene therapy". Specificerade sökkriterier användes för att underlätta relevansbedömning och valet av artiklar. Resultat - En ökad endogen koagulationsfaktorproduktion kunde påvisas hos majoriteten av studiedeltagarna efter genterapibehandlingarna. Sammantaget observerades också en väsentlig blödningsreducering och en minskad faktorkoncentratanvändning. Störst förbättring noterades i de kohorter som erhållit högre genterapidoser eller den muterade faktor IX Padua-genen. Ingen immunrespons mot transgenprodukten detekterades i någon studie. Däremot sågs ett humoralt immunsvar mot AAV-kapsiden hos samtliga studiedeltagare. En mycket stor variation i T-cellssvar mot AAV-kapsiden kunde noteras. Förhöjda nivåer av alaninaminotransferas (ALAT) var den vanligast förekommande incidenten, men samtliga fall kunde framgångsrikt behandlas med glukokortikoidpreparat. Slutsats - Genterapibehandling med rekombinanta AAV-vektorer mot hemofili A och B förefaller effektiv och säker. Förhöjda ALAT-nivåer återstår dock som en behandlingsproblematik. Längre uppföljningar av fler genterapibehandlade hemofilipatienter krävs, för att kunna dra några definitiva slutsatser, väga risker mot nytta, samt optimera och individanpassa doser. Background - Hemophilia A and B are X-linked bleeding disorders, resulting from defects in the genes encoding coagulation factors VIII and IX respectively. The current treatment for hemophiliacs entails frequent intravenous injections of coagulation factor concentrates, to prevent potentially life-threatening hemorrhages. Gene therapy utilizing recombinant adeno-associated viruses (AAV) could offer a potentially curative treatment option through the introduction of healthy genes into hepatocytes. Aim - The aim of this literature study was to investigate the long-term efficacy and safety of AAV vector-mediated gene therapy for the treatment of hemophilia A and B. Methods - The study is conducted as a systematic literature study and is based on six original articles retrieved from the search engine PubMed, using the key words "hemophilia AND gene therapy". Specific search criteria were used to facilitate the relevance assessment and selection of articles. Results - An increased endogenous coagulation factor synthesis was noted in the majority of the study participants after the gene therapy. Overall, a significant reduction in bleeding episodes and the use of factor concentrates were observed. The greatest improvements were noted in the cohorts that received the higher gene therapy doses or the mutated factor IX Padua gene. None of the study participants had an immunologic response to the transgene product. A humoral immune response against the AAV capsid was seen in all participants though. Large differences in AAV capsid-specific T-cell activation were observed. The most common adverse event was an elevation in the alanine aminotransferase (ALT) level. However, these events could be controlled with glucocorticoids. Conclusions - AAV vector-mediated gene therapy for the treatment of hemophilia A and B had a positive efficacy and safety profile. Although increased ALT levels remain a concern. Monitoring of larger numbers of study participants for longer follow-up periods is necessary for any definite conclusions to be drawn, to weigh risks against benefits and to optimize individual dosing.

Das hochmolekulare, lineare multimere Glykoprotein von Willebrand Faktor (VWF) ist ein unverzichtbarer Bestandteil des Bluts von S��ugern und Menschen. Es erf��llt eine Reihe von wichtigen Funktionen in der H��mostase, des Tumorwachstums und der Angiogenese doch eine andere wichtige Seite seines Wirkungsbereichs ist sein thrombogenes und entz��ndungsf��rderndes Potenzial. VWF ist der Schl��ssel zu zum Verst��ndnis des konterintuitiven Ph��nomens der durch hydrodynamische Kr��fte verst��rkten prim��ren H��mostase (das ist die Bildung eines prim��ren Thrombozytenaggregats). Scherstr��mung und andere divergente Str��mungsformen verursachen ein Entwirren und teilweises Entfalten der multimeren VWF Ketten, die sonst als lose Kn��ul zirkulieren. Ein Strecken von VWF ist verbunden mit einer Aktivierung von kritischen Funktionen, wie die Bindung an Kollagen und an Blutpl��ttchenrezeptoren (prim��re H��mostase) und die Zug��nglichkeit f��r proteolytische Spaltung, was wiederum das thrombogene Potenzial reguliert. Diese mechanische Regulation macht VWF zum Scherratensensor des Blutes. In dieser Dissertation wurde die Rasterkraftmikroskopie (AFM) im sogenannten 'tapping' Modus und ein eigens konstruiertes 2-Platten-rheometerartiges Ger��t verwendet, um auf Einzelmolek��lebene, die Reaktionen von gestreckten rekombinat hergestellten, multimeren VWF Ketten zu studieren. Dazu wurde eine Methode entwickelt mit der einzelne multimere VWF Molek��le an einer Tr��geroberfl��che lokalisiert und auch nach Interaktionsstudien wiedergefunden werden k��nnen, um davon AFM Bilder vor und nach einer Bindung mit einem rekombinanten Faktor VIII (FVIII) aufzunehmen. FVIII ist essentiell an der Aktivierung der sekund��ren H��mostase beteiligt, doch seine geringe Stabilit��t in humanem Plasma verlangt nach der stabilisierenden Komplexbildung mit VWF. Als n��chstes wurde ein mikrofluidisches, rheometerartiges Ger��t verwendet, mit dem einzelne VWF Molek��le bei exakt definierten Scherraten (mechanisch) gestreckt werden k��nnen. Kombination dieser beiden Methoden und die Interaktion der Protease ADAMTS13 in L��sung mit dem Glykoprotein als Kontrolle, f��hrte zum unerwarteten Ergebnis, dass FVIII nur bei geringen bis mittleren Scherraten an VWF bindet, w��hrend bei hohen Scherraten FVIII freigesetzt wird, was den selektiven Transport von FVIII zu Bereichen mit besch��digtem Endothel erkl��rt. Schlie��lich wurde auch reiner FVIII mit AFM untersucht und ein Ca2+-abh��ngiger Voraktivierungs- und Selbststabilisierungs-Mechanismus gezeigt., The high molecular mass linear chain multimeric glycoprotein von Willebrand factor (VWF) is an indispensable component of mammalian (including human) blood. It serves multiple beneficial roles in hemostasis, cancer suppression and angiogenesis but in turn has also thrombogenic and pro-inflammatory potential. VWF is the key to understand the counter-intuitive observation that primary hemostasis (that is, platelet recruitment at sites of vascular injury to form a preliminary plug and quickly stop bleeding) is not disturbed but enhanced by hydrodynamic flow. Shear-flow and other divergent (elongational) flow conditions can cause an unravelling and partial unfolding (-stretching-) of the VWF multimeric chains which are otherwise loosely coiled in circulation. Stretching VWF goes along with an activation of crucial functions such as binding to collagens of vessel walls and platelet receptors (primary hemostasis) and susceptibility to proteolytic cleavage which downregulates its own thrombogenic potential. This mechanical activation makes VWF the flow sensor of blood. In this thesis, atomic force microscopy (AFM) in the tapping mode was used and a homebuilt 2-plate-rheometer-like device was applied to study reactions of stretched VWF chains at single molecular level. First, a method was developed to retrieve an individual single glycoprotein molecule enabling before-and after imaging of a recombinant VWF-Factor VIII (FVIII) binding. FVIII is essential for the activation of secondary hemostasis but its stability in the circulation is dependent on its complex formation with VWF. Next, a microfluidic rheometer device was developed which is capable to stretch single molecule VWF chains at well-defined shear-rates. Combining these two devices, and introducing the protease ADAMTS13 as a control, provided the unexpected result that FVIII only binds FVIII at low/moderate shear-rates whereas high shear-stretching elicit a release of FVIII, explaining a targeting mechanism of FVIII to sites of injured endothelium. Finally, also FVIII as such was investigated by AFM revealing a Ca2+-linked pre-activation and self-stabilizing mechanism.

Hemostasis is essential to life; it is the ability of organism to stop bleeding and to maintain the fluidity of the blood in an intact vascular bed at one time. Factor VIII is called antihemophilic globulin or antihemophilic factor. It is a plasma glycoprotein composed of two noncovalently associated chains. One chain is a heavy one, comprising domains A1- A2- B and the light chain composed of the domains A3- C1-C2. FVIII activity was measured in two ways from historical reasons a) a one-phase method, where a mixture of FVIII deficient plasma and patient plasma is analyzed using APTT assay; the absence of FVIII in the patient plasma leads to lengthening of the time, b) two-phase method, where the first step leads to formation of FVIIIa and FXa and in the second phase there are thrombin and fibrin created. Two-phase method is difficult to implement in routine laboratory, and therefore it was stopped using during time. This method was substituted by the chromogenic method after development of the chromogenic substrates, where there is in the first step created FVIIIa and FXa in the presence of FIXa, phospholipids and Ca2+, and in the second step there is formed a yellow coloration by the addition of the chromogenic substrate. FVIII deficiency causes a severe bleeding disorder, hemophilia A. I performed measurements from May to September 2013 at Coagulation laboratory at the Institute of Hematology and Blood Transfusion in Prague, where I was employed. I examined in total 76 patients, hemophiliacs A, who were at least 8 days without any treatment or substitution. I assigned numbers to patient's samples to ensure anonymity of patients. I worked with the automatic coagulation analyzer STA- R Evolution? from Diagnostica Stago, which works on the principle of photometry and chronometry. I determined the factor VIII by one-phase method and two-phase method and I compared the results. The genetic part of the work was analyzed in the genetic laboratory, which is part of our department. I worked up the results obtained from both methods in 76 patients to the table and the graph. The group included 14 (18 %) moderate and 56 (74 %) of mild hemophiliacs, then 6 (8%) hemophiliacs who did not meet the criterion of a mild hemophilia A, but clinically they belonged into mild hemophiliacs. Based on the stated criteria, we found out that 15 (20 %) patients had a ratio of FVIII: C1st/FVIII: Chr or FVIII: Chr / FVIII: C1st 0.6, they differed significantly in their values set by one-stage clotting FVIII and FVIII set by the chromogenic method. A total of 11 patients with FVIII activity were higher in the single-phase method. At three patients FVIII: C1st was even on the upper limit of the normal value, while FVIII chromogenic method gave on average 16%. We managed to find a causal mutation in the FVIII in 14 patients with "the different results", we could not investigate 1 patient genetically because of the missing genetic material. Mutations in patients with lower activity of FVIII set up by the chromogenic two-phase method were concentrated predominantly into the A3 domain; mutations in patients with FVIII a lower activity set up by one-phase clotting assay were concentrated in the A2 domain. The results presented show, that diagnostic of any patient with mild or moderate hemophilia A should include determination of FVIII by both methods; FVIII: C1st and FVIII: Chr.

Cjelovitost I multidisciplinarnost u nastanku venske tromboze kao I specificnosti rizika u bolesnika s upalnom bolesti crijeva. Prevencija I rana dijagnostika tromboembolijske bolesti u populaciji IBD bolesnika.

Objektive: Darstellung anhand von Gerinnnungsphysiologischen Analysen im venösen Blut von Patienten, in wie weit es zu Veränderungen im Gerinnungssystem und zur Aktivierung mit Erhöhung des Thromboserisikos kommen kann. Die Perkutane Transluminale Coronarangioplastie wurde der diagnostischen Coronarangiographie gegenübergestellt. Methode: Die Gerinnungsparameter wurden anhand von Fibrinogen, Faktor VIII und vWF in einem Zeitrahmen von direkt präinterventionell und 24h postinterventionell, unter Blutabnahmen zu 5 verschiedenen Zeitpunkten, miteinander verglichen. Dies erfolgte auch unter dem Aspekt des Vergleichs der Geschlechter. Ergebnis: Deutlicher Anstieg der Plasmaspiegel, insbesondere des Fibrinogen und vWF unter durchgeführter Coronarangioplastie. Geschlechtsdifferenziert erhöhte Plasmaspiegel im weiblichen Patienten- und Kontrollkollektiv. Zusammenfassung: Erhöhtes Thrombose- bzw. Restenose- Risiko der Patienten die sich einer Coronarangioplastie unterzogen anhand der erhöhten Konzentration insbesondere von Fibrinogen und vWF. Zusätzlich kann von einer erhöhten Thrombogenität des weiblichen Geschlechts ausgegangen werden, anhand von präinterventionell signifikant erhöhten Plasmaspiegeln der untersuchten Gerinnungsparameter. Objective: Activation of the coagulation system with higher plasmalevels of hemostatic parameters in patients undergoing a coronary angioplasty vs. a coronary angiography with differences in gender Methods: Concentration or activity was measured for three hemostatic factors before intervention and directly afterwards, 2-, 6- and 24 hours later in venous blood of 95 patients and compared two 35 patients undergoing a coronary angiography only. The results were also compared in gender. Results: Higher levels were found in the female group of patients before angioplasty and angiography. Levels of Fibrinogen and vWF were higher after 24hours than before intervention in the patient group. Factor VIII showed a fall of the plasmalevel after angioplasy under the typical dose of heparin due to the antithrombotic effect and presumably because of a higher usage. Conclusion: Patients with active ischemic syndromes showed a higher risk of thrombosis after coronary angioplasty by significant alterations of their coagulation system. In the female gender a higher level of the hemostatic parameters was found before intervention with a higher risk of thrombosis compared to the male gender.

Evaluiert wurde die Rolle des familiär erhöhten Faktor VIII bei Kindern mit einer ersten Entstehung venöser Thrombosen und ob eine mögliche Verbindung zum Auftreten vom postthrombotischen Syndrom oder rezidivierender vT vorhanden ist. FVIII:Ag-, FVIIIC- und von Willebrand Faktor Antigen- Spiegel wurden zusammen mit bekannten Thrombophilien nach dem akuten thrombotischen Ereignis bei 103 kaukasischen Kindern im Alter von 0-18 Jahren berechnet, die an einer ersten vT leiden. Die medianen Werte von FVIII:Ag und FVIII:C- Werte waren bei den Patienten im Vergleich zu der Kontrollgruppe erhöht . Der Vergleich von FVIII-Werten mit FVIII-Werten unterhalb des altersabhängigen cut-offs,zeigte eine signifikant erhöhte Odds Ratio und einen 95%igen Konfidenzintervall bei Kindern mit vT. Innerhalb von 12 Monaten trat bei 19 von 59 Kindern ein PTS auf, bei fünf Individuen persistierte diese bei den Nachuntersuchungen: PTS war signifikant mit der Ausdehnung der ersten Thrombosebelastung assoziiert.

Im Rahmen dieser Arbeit etablierte, optimierte und testete ich zwei molekulargenetische Methoden zur Mutationsdetektion im Faktor VIII-Gen. Hierbei handelt es um die SSCP und die ddF. Die SSCP erwies sich als zuverlässige, zeit- und kostensparende Methode, deren Detektionsrate 82% betrug. Demgegenüber zeigte sich die ddF als zeit-, kosten- und arbeitsintensiv, stach aber durch eine bemerkenswert hohe Detektionsrate von 97% hervor. Zur Erfolgskontrolle der optimalen Etablierung wurden Hämophilie A-Patienten mit noch nicht charakterisierten Mutationen untersucht, wobei drei bis dahin unbekannte Mutationen im Faktor VIII-Gen identifiziert werden konnten. Die effektivste Vorgehensweise für die zukünftige Diagnostik bestünde darin, zunächst 80% der Mutationen zügig und kostengünstig durch Einsatz der SSCP zu detektieren. Die verbleibenden 20% der Mutationen könnten anschließend durch den Einsatz der aufwändigeren, aber sehr effektiven ddF ergänzt werden.

http://www.ester.ee/record=b4338686~S58*est

11-14 Ekim 1989 tarihlerinde İstanbul'da yapılan XXI. Ulusal Hematoloji Kongresi ve 1. Ulusal Kemoterapi Kongresinde tebliğ edilmiştir. Bu çalışmada, kliniğimize Faktör VIII ve Faktör IX'a karşı inhibitölere bağlı durdurulamayan burun kanaması ile başvuran ve yapılan incelemeler sonunda SLE tanısı konan bir hasta takdim edilerek ilgili literatür gözden geçirilmiştir. In this report, a patient who had applied to our clinic with a complaint of unceaseble nose bleeding due to inhibitors against factors VIII and IX and who had SLE diagnosis after clinical and laboratory evaluations had been presented with the literature available being reviewed.

Pozvano predavanje o diferencijalnoj dijagnozi mogućih uzroka nastanka stečenog inhibitora faktora VIII, krvarenja, liječenja i preživljenja.

Agregacija trombocita, koncentracija faktora VIII i fibrinogena ispitivana je u 20 zdravih osoba i 70 dijabetičkih bolesnika podijeljenih u tri skupine: 20 novootkrivenih dijabetičkih bolesnika, 20 bolesnika s kraćim trajanjem dijabetesa i 30 bolesnika s dužim trajanjem dijabetesa i razvijenim komplikacijama. Agregacija trombocita nije se značajno razlikovala ni između pojedinih skupina dijabetičkih bolesnika, niti u odnosu na kontrolnu skupinu. Faktor VIII bio je povišen u novootkrivenih dijabetič­ kih bolesnika u odnosu na bolesnike s kraćim trajanjem dijabetesa (p0,05). Fibrinogen je bio povišen u dijabetičkih bolesnika s dugotrajnim dijabetesom i razvijenim komplikacijama i u odnosu na novootkrivene (p, Platelet aggregation, concentration of factor VIII and fibrinogen were analysed in 20 healthy persons and 70 diabetics devided into three groups: 20 newly diagnosed diabetics, 20 patients with shorter duration of diabetes and 30 patients with longer duration of diabetes and developed chronic complications. Platelet aggregation did not differ significantly in newly diagnosed diabetics both compared to the control group and to the other diabetics. Concentration of factor VIII was increased in newly diagnosed diabetics as compared to the patients with shorter duration of diabetes (p0.05). Fibrinogen was increased in diabetics with longer duration of diabetes both compared to the newly detected patients (p