Inhibitors in Patients with Congenital Bleeding Disorders Other Than Hemophilia

The most worrying complication of replacement therapy for severe hemophilia A and B is currently the occurrence of inhibitory alloantibodies against infused factor VIII and factor IX, respectively. Inhibitors compromise the management of hemorrhage in affected patients, with a considerable increase in complications, disability, and costs. While these alloantibodies have been extensively studied in the past years in hemophilia A and B, those occurring in patients with other inherited bleeding disorders are less well characterized and still poorly understood, mostly due to the rarity of these hemorrhagic conditions. This narrative review will deal with inhibitors arising in patients with inherited bleeding disorders other than "classical" hemophilia, focusing in particular on those developing in patients with congenital deficiency of coagulation factor V, factor VII, factor XI, and factor XIII

A véralvadás XIII-as faktora = Blood coagulation factor XIII.

I. A XIII-as faktor (FXIII) struktúrája és funkciója Az aktivációs peptid nélkül a FXIII-A instabil. A FXIII aktivációt a plazmában a fibrin polimerizáció determinálja, a FXIII-A Val34Leu polimorfizmusnak csak moduláló hatása van. A FXIII A és B alegységének kapcsolódását gátló monoklonális antitestek előállítása. A celluláris FXIII szerepet játszik a monocyták/macrophagok phagocytosisában. Az alvadékban aktiválódó granulocytákból felszabadult proteázok lebontják a FXIIIa-t. II. Klinikai FXIII kutatások Új típusú módszer a FXIII (ill. más transzglutaminázok) mérésére, ill. a FXIII-A Val34Leu polimorfizmus kimutatására. Az intracelluláris FXIII-A áramlásos citometriás kimutatására új módszer, mely kiválóan alkalmazható az acut myleoid leukemiák diagnosztikájában. Csontvelő abláció során szignifikánsan csökken a plazma FXIII szintje, magas thrombocyta számmal járó myeloproliferatív megbetegedésekben viszont emelkedik. Nőkben az emelkediett FXIII szint több mint kétszeresére növeli a myocardiális infarctus rizikóját nőkben. Krónikus bronchoalveoláris megbetegedésekben jelentősen emelkedik a bronchoalveoláris mosófolyadékban a FXIII mennyisége. 4 új mutációt írtunk le FXIII hiányos betegekben, s analizáltuk ezek fehérje biokémiai következményeit. FXIII hiányíos transzgén egereken igazoltuk, hogy a FXIII szükséges a normális sebgyógyuláshoz. | I. Structure and function of factor XIII (FXIII) The absence of activation peptide makes FXIII-A instable. The activation of FXIII in the plasma is determined by fibrin polymerization; FXIII-A Val34Leu polymorphism only modulates activation. Production of monoclonal antibodies with inhibitory effect on the association of FXIII A and B subunits. Cellular FXIII plays a role in the phagocytosis by monocytes/macrophages. Proteases released from granulocytes in the clot break down activated FXIII. Clinical studies on FXIII New methods on the measurement of FXIII activity, and on the detection of FXIII-A Val34Leu polymorphism. Method on the detection of intracellular FXIII-A and its application to the diagnosis of acute myeloid leukemias. Bone marrow ablation results in the significant decrease of plasma FXIII level; in myeloproliferative diseases with high platelet count plasma FXIII is elevated. In women elevated FXIII level increases the risk for myocardial infarction by more than two-folds. In chronic bronchoalveolar inflammation the amount of FXIII in the lavage fluid is significantly increased. Description of four new mutations in FXIII deficient patients, and protein structural analysis of their consequences. It was demonstrated on FXIII deficient transgene mice that FXIII is required for normal wound healing

Statistical methods of SNP data analysis with applications

Various statistical methods important for genetic analysis are considered and developed. Namely, we concentrate on the multifactor dimensionality reduction, logic regression, random forests and stochastic gradient boosting. These methods and their new modifications, e.g., the MDR method with "independent rule", are used to study the risk of complex diseases such as cardiovascular ones. The roles of certain combinations of single nucleotide polymorphisms and external risk factors are examined. To perform the data analysis concerning the ischemic heart disease and myocardial infarction the supercomputer SKIF "Chebyshev" of the Lomonosov Moscow State University was employed

A véralvadás XIII-as faktora: strukturális és funkcionális vonatkozások, jelentősége különböző kórképekben = Blood coagulation FXIII: structural, functional aspects, its involvement in various pathological conditions

Új megállapításokat tettünk a FXIII plazmában történő aktivációjára, a két alegység (FXIII-A és FXIII-B) egymáshoz kapcsolódásának strukturális elemeire és a FXIIIa-glutamin szubsztrát kapcsolatra vonatkozóan. Új módszereket dolgoztunk ki a FXIII aktivitás mérésére, a FXIII-A intracelluláris detektálására. Utóbbi bevezetésre került a leukémiák diagnosztikájában. Immunoassay-t dolgoztunk ki az a2 plazmin inhibitor két izoformájának mennyiségi meghatározására. 10 FXIII-A hiányos betegen derítettük fel a háttérben álló mutációkat és ezek következményeit a fehérje strukturájára-funkciójára. Kísérletesen bizonyítottuk, hogy a plazma FXIII hiánya sebgyógyulási zavart okoz. Kimutattuk, hogy myocardiális infarctuson (MI) átesett nőkben emelkedett a plasma FXIII szintje és az emelkedett FXIII szint 2,5-3,0 szorosra fokozza az MI rizikóját, ami kizárólag nőkön érvényesül. 16 cikk metaanalízisiével a FXIII-A L34 allél szignifikáns védőhatását lehetett kimutatni a coronaria betegség ellen, a polimorfizmus a nagy rizikóju magyar populációban azonban csak emelkedett fibrinogén szint esetén védő hatású. A L/L homozigóták FXIII szintje MI-ben szignifikánsan alacsonyabb a vad típusúakénál. Csontvelő abláció után csökken, magas thrombocyta számmal járó myeloproliferatív betegségben emelkedik a FXIII szintje. Bronchoalveoláris mosófolyadékban kimutatható az alveoláris macrophagokból származó FXIII-A, chronicus bronchitisben ennek szintje emelkedik, s esetenként megjelenik a plazma FXIII is. | New results were reported on the activation of factor XIII (FXIII) in plasma, on the structural elements involved in the association of FXIII subunits (FXIII-A and FXIII-B) and on the interaction of activated FXIII (FXIIIa) with its glutamine substrate. Methods were developed for the determination of FXIII activity and the intracellular detection of FXIII-A by flow cytometry. The latter was introduced in the diagnostics of leukemias. Immunoassay was developed for the determination of the two isoforms of a2 plasmin inhibitor. The mutations causing FXIII-A deficiency were identified in 10 patients and their consequences were explored at the protein level. The involvement of FXIII in would healing was proven. It was shown that in women with the history of myocardial infarction (MI) FXIII level was elevated and elevated FXIII level represented a 2.5-3.0-fold increased risk of MI in women, but not in men. A protective effect of the FXIII-A L34 allele against MI was demonstrated by metaanalysis of 16 articles. In the Hungarian population this protective effect prevailed only at high fibrinogen level. FXIII level was decreased in L/L homozygotes with the history of MI. Bone marrow ablation decreased plasma FXIII level, while myeloproliferative diseases increased it. In the bronchoalveolar lavage fluid FXIII-A derived from alveolar macrophages was detected, in inflammatory bronchoalveolar diseases FXIII-A level increased and occasionally plasma FXIII was also be present

Molecular Analysis of Prothrombotic Gene Variants in Venous Thrombosis: A Potential Role for Sex and Thrombotic Localization

Background: Requests to test for thrombophilia in the clinical context are often not evidence-based. Aim: To define the role of a series of prothrombotic gene variants in a large population of patients with different venous thromboembolic diseases. Methods: We studied Factor V Leiden (FVL), FVR2, FII G20210A, Methylenetetrahydrofolate reductase (MTHFR) C677T and A1298C, beta-fibrinogen -455 G>A, FXIII V34L, and HPA-1 L33P variants and PAI-1 4G/5G alleles in 343 male and female patients with deep vein thrombosis (DVT), 164 with pulmonary embolism (PE), 126 with superficial vein thrombosis (SVT), 118 with portal vein thrombosis (PVT), 75 with cerebral vein thrombosis (CVT) and 119 with retinal vein thrombosis (RVT), and compared them with the corresponding variants and alleles in 430 subjects from the general population. Results: About 40% of patients with DVT, PE and SVT had at least one prothrombotic gene variant, such as FVL, FVR2 and FII G20210A, and a statistically significant association with the event was found in males with a history of PE. In patients with a history of PVT or CVT, the FII G20210A variant was more frequent, particularly in females. In contrast, a poor association was found between RVT and prothrombotic risk factors, confirming that local vascular factors have a key role in this thrombotic event. Conclusions: Only FVL, FVR2 and FII G20210A are related to vein thrombotic disease. Other gene variants, often requested for testing in the clinical context, do not differ significantly between cases and controls. Evidence of a sex difference for some variants, once confirmed in larger populations, may help to promote sex-specific prevention of such diseases

Implications of the large OVI columns around low-redshift L∗L_* galaxies

Observations reveal massive amounts of OVI around star-forming $L_*$ galaxies, with covering fractions of near unity extending to the host halo's virial radius. This OVI absorption is typically kinematically centered upon photoionized gas, with line widths that are suprathermal and kinematically offset from the galaxy. We discuss various scenarios and whether they could result in the observed phenomenology (cooling gas flows, boundary layers, shocks, virialized gas, photoionized clouds in thermal equilibrium). If predominantly collisionally ionized, as we argue is most probable, the OVI observations require that the circumgalactic medium (CGM) of $L_*$ galaxies holds nearly all the associated baryons within a virial radius ($\sim 10^{11}M_\odot$) and hosts massive flows of cooling gas with $\approx30[nT/30{\rm~cm^{-3}K}]~M_\odot~$yr$^{-1}$, which must be largely prevented from accreting onto the host galaxy. Cooling and feedback energetics considerations require $10 <\langle nT\rangle<100{\rm~cm^{-3}K}$ for the warm and hot halo gases. We argue that virialized gas, boundary layers, hot winds, and shocks are unlikely to directly account for the bulk of the OVI. Furthermore, we show that there is a robust constraint on the number density of many of the photoionized $\sim10^4$K absorption systems that yields upper bounds in the range $n<(0.1-3)\times10^{-3}(Z/0.3)$cm$^{-3}$, where $Z$ is the metallicity, suggestive that the dominant pressure in some photoionized clouds is nonthermal. This constraint, which requires minimal ionization modeling, is in accord with the low densities inferred from more complex photoionization modeling. The large amount of cooling gas that is inferred could re-form these clouds in a fraction of the halo dynamical time, as some arguments require, and it requires much of the feedback energy available from supernovae and stellar winds to be dissipated in the CGM.Comment: 16 pages, matches version accepted to Ap

Interaction of factor XIII subunits

Coagulation factor XIII (FXIII) is a heterotetramer consisting of 2 catalytic A subunits (FXIII-A2) and 2 protective/inhibitory B subunits (FXIII-B2). FXIII-B, a mosaic protein consisting of 10 sushi domains, significantly prolongs the lifespan of catalytic subunits in the circulation and prevents their slow progressive activation in plasmatic conditions. In this study, the biochemistry of the interaction between the 2 FXIII subunits was investigated. Using a surface plasmon resonance technique and an enzyme-linked immunosorbent assay-type binding assay, the equilibrium dissociation constant (Kd) for the interaction was established in the range of 10(-10) M. Based on the measured Kd, it was calculated that in plasma approximately 1% of FXIII-A2 should be in free form. This value was confirmed experimentally by measuring FXIII-A2 in plasma samples immunodepleted of FXIII-A2B2. Free plasma FXIII-A2 is functionally active, and when activated by thrombin and Ca(2+), it can cross-link fibrin. In cerebrospinal fluid and tears with much lower FXIII subunit concentrations, >80% of FXIII-A2 existed in free form. A monoclonal anti-FXIII-B antibody that prevented the interaction between the 2 subunits reacted with the recombinant combined first and second sushi domains of FXIII-B, and its epitope was localized to the peptide spanning positions 96 to 103 in the second sushi domain

Allosteric Inhibition of Factor XIIIa. Non-Saccharide Glycosaminoglycan Mimetics, but Not Glycosaminoglycans, Exhibit Promising Inhibition Profile

Factor XIIIa (FXIIIa) is a transglutaminase that catalyzes the last step in the coagulation process. Orthostery is the only approach that has been exploited to design FXIIIa inhibitors. Yet, allosteric inhibition of FXIIIa is a paradigm that may offer a key advantage of controlled inhibition over orthosteric inhibition. Such an approach is likely to lead to novel FXIIIa inhibitors that do not carry bleeding risks. We reasoned that targeting a collection of basic amino acid residues distant from FXIIIa’s active site by using sulfated glycosaminoglycans (GAGs) or non-saccharide GAG mimetics (NSGMs) would lead to the discovery of the first allosteric FXIIIa inhibitors. We tested a library of 22 variably sulfated GAGs and NSGMs against human FXIIIa to discover promising hits. Interestingly, although some GAGs bound to FXIIIa better than NSGMs, no GAG displayed any inhibition. An undecasulfated quercetin analog was found to inhibit FXIIIa with reasonable potency (efficacy of 98%). Michaelis-Menten kinetic studies revealed an allosteric mechanism of inhibition. Fluorescence studies confirmed close correspondence between binding affinity and inhibition potency, as expected for an allosteric process. The inhibitor was reversible and at least 9-fold- and 26-fold selective over two GAG-binding proteins factor Xa (efficacy of 71%) and thrombin, respectively, and at least 27-fold selective over a cysteine protease papain. The inhibitor also inhibited the FXIIIa-mediated polymerization of fibrin in vitro. Overall, our work presents the proof-of-principle that FXIIIa can be allosterically modulated by sulfated non-saccharide agents much smaller than GAGs, which should enable the design of selective and safe anticoagulants

Polymorphisms of plasminogen activator inhibitor-1, angiotensin converting enzyme and coagulation factor XIII genes in patients with recurrent spontaneous abortion

We investigated polymorphisms of plasminogen activator inhibitor-1 (PAI-1), angiotensin converting enzyme (ACE ) and coagulation factor XIII (FXIII) genes and their association with recurrent spontaneous abortion (RSA) in Iranian patients and normal healthy controls. Ten (18.5%) patients were homozygote (4G/4G) for PAI-1 polymorphism, in contrast with two (2%) controls (p = 0.001). Patients with homozygote 4G mutation were significantly more prone to RSA in contrast to others (odds ratio: 11.0, 95% CI: 2.3-52.4). Nineteen (30.2%) patients and 25 (26.6%) controls were homozygote (DD) for ACE polymorphism. We observed only two patients and one control with homozygosity (34leu) for FXIII polymorphism. 4G/4G polymorphism for PAI-1 gene could be a thrombophilic mutation leading to abortion in Iranian population

Plasma factor XIII level variations during menstrual cycle

Factor XIII (FXIII) has an important role in the control of bleeding through fibrin cross-linking; however, its effect within the menstrual cycle is not fully understood. The aim of this study was to examine changes in FXIII activity during the normal menstrual cycle and correlate FXIII activity with menstrual blood loss. A total of 32 healthy normal women of reproductive age were recruited. Menstrual blood loss was measured using the pictorial blood-assessment chart (PBAC). A bleeding score questionnaire was also completed. Blood samples were taken during the menstrual, proliferative, periovulatory, secretory and premenstrual phase for assessment of FXIII level. The mean ± SD FXIII level was lowest during menstrual and periovulatory phases (114 ± 23 and 114 ± 21 IU/dl, respectively). Mean FXIII level during the secretory and premenstrual phases were higher than the menstrual phase (P = 0.036). Mean secretory phase FXIII was also significantly higher compared with the periovulatory phase (P = 0.02). There was no significant correlation between FXIII level during the menstrual phase and age (P = 0.53) or PBAC score (P = 0.53). There were no significant differences in FXIII level during the menstrual phase between women with PBAC scores of at least 100 (n = 14; mean 116 IU/dl) and women with PBAC scores less than 100 (n = 18; mean 113 IU/dl). There was no correlation between FXIII level and bleeding score. FXIII activity was lower during menstrual and periovulatory phases of the cycle. However, the small difference between mean values (8 IU/dl) would be unlikely to have a significant impact on diagnosis of FXIII deficiency and clinical management