1/15. Fibrinogens Bern IV, Bern V and Milano XI: three dysfunctional variants with amino acid substitutions in the thrombin cleavage site of the Aalpha-chain.Thrombin-induced cleavage of fibrinopeptide a is the initial step in the conversion of fibrinogen to fibrin. Three dysfunctional fibrinogen variants are described with an amino acid substitution at position 16 of the Aalpha-chain: the fibrinogen variants Bern IV and Milano XI having an Arg-->His substitution and the variant Bern V having an Arg-->Cys substitution. Routine coagulation studies revealed prolonged plasma thrombin and reptilase clotting times in all patients, and a discrepancy between the plasma levels of fibrinogen determined by the clotting assay and electroimmunoassay. The defect was localized by high-performance liquid chromatography analysis of fibrinopeptide release and confirmed by polymerase chain reaction and sequencing of exon 2 of the Aalpha-chain. immunoblotting analysis with a rabbit antiserum against human serum albumin indicated that albumin was linked to the additional sulfhydryl group of fibrinogen Bern V. The assay of tissue-plasminogen-activator-induced plasmic degradation revealed that the fibrinolysis of fibrin Bern V was delayed, whereas fibrin Bern IV was digested in the same way as normal fibrin.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
2/15. Williams trait. Human kininogen deficiency with diminished levels of plasminogen proactivator and prekallikrein associated with abnormalities of the Hageman factor-dependent pathways.An asymptomatic woman (Ms. Williams) was found to have a severe abnormality in the surface-activated intrinsic coagulation, fibrinolytic, and kinin-generating pathways. Assays for known coagulation factors were nromal while Fletcher factor (pre-kallikrein) was 45%, insufficient to account for the observed markedly prolonged partial thromboplastin time. plasminogen proactivator was present at 20% of normal levels and addition of highly purified plasminogen proactivator containing 10% plasminogen activator partially corrected the coagulation and fibrinolytic abnormalities but not the kinin-generating defect. This effect was due to its plasminogen activator content. In addition, Williams trait plasma failed to convert prekallilrein to lakkilrein or release kinin upon incubation with kaolin. Kininogen antigen was undetectable. When normal plasma was fractionated to identify the factor that corrects all the abnormalities in Williams trait plasma, the Williams factor was identified as a form of kininogen by its behavior on ion exchange chromatography, gel filtration, disc gel electrophoresis, and elution from an anti-low molecular weight kininogen immunoadsorbent. High molecular weight kininogen as well as a subfraction of low molecular weight kininogen, possessed this corrective activity while the bulk of low molecular weight kininogen functioned only as a kallikrein substrate. Kininogen therefore is a critical factor required for the functioning of Hageman factor-dependent coagulation and fibrinolysis and for the activation of prekallikrein.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
3/15. fibrinogen Ledyard (A alpha Arg16Cys): biochemical and physiologic characterization. fibrinogen Ledyard was discovered in a 10-year-old boy with a mild bleeding history. His father had the same defect and a bleeding history after surgery. Both patients were heterozygous. The plasma fibrinogen concentration was normal immunologically (335 mg/dL) and very low functionally (52 mg/dL). Purified fibrinogen Ledyard had a prolonged polymerization, which was somewhat corrected by addition of Ca2 ions. High performance liquid chromatography (HPLC) analyses of the fibrinopeptides released by thrombin showed 1 mol of fibrinopeptide a (FPA) and 2 mol of fibrinopeptide b (FPB) released per mole of fibrinogen Ledyard. Steady-state kinetic parameters were evaluated for release of FPA by thrombin. When the concentration of fibrinogen Ledyard was corrected to 50% of total protein, because only 50% of fibrinogen Ledyard can release FPA, the kinetic constants were similar to those of control fibrinogen (Km = 7.5 mumol/L for A alpha chain, kcat = 54 s-1). This finding indicates that the cleavage site of the A alpha chain in these abnormal molecules may not interact with the catalytic site of thrombin. The three chains of fibrinogen Ledyard were isolated on reverse-phase C4-HPLC. The sequence of the amino terminus of A alpha chain showed that Arg in position 16 was replaced by Cys in the abnormal molecules. Approximately half of fibrinogen Ledyard (52%) was clotted by reptilase, suggesting that fibrinogen Ledyard may consist of 50% normal homodimers (A alpha Arg16 . A alpha Arg16) and 50% abnormal homodimers (A alpha Cys16 . A alpha Cys16). Abnormal molecules could form disulfide bond between the A alpha Cys16 residues. Thus, the abnormal molecules have a different structure that does not bind to thrombin. Probably the abnormality of polymerization of fibrinogen Ledyard results from the interaction of the abnormal molecules with normal fibrin monomers, so that the growth of fibrin protofibrils is inhibited. This abnormal fibrinogen supports adenosine diphosphate-induced platelet aggregation in a normal manner.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
4/15. A new congenital abnormal fibrinogen Ise characterized by the replacement of B beta glycine-15 by cysteine.A new case of heterozygous dysfibrinogenemia characterized by the replacement of NH2-terminal amino acid of fibrin beta-chain was found in a 50-year-old man. Despite a prolonged thrombin time, the propositus' fibrinogen had a normal reptilase time with the normal release of fibrinopeptide a. Release of fibrinopeptide b by thrombin was strongly affected, but a very high concentration of thrombin almost completely released fibrinopeptide b with a normal elution pattern on reversed-phase high performance liquid chromatography (HPLC). Lysylendopeptidase-cleavage of purified B beta-chains analyzed on HPLC showed the decrease of one peptide compared with the normal and the appearance of an abnormal peptide peak. These peptides were treated with thrombin and further separated on HPLC. amino acid sequence analysis of the abnormal peptide demonstrated that B beta glycine-15, NH2-terminus of the fibrin beta-chain, was replaced by cysteine. These findings will be of particular importance because they strongly support the hypothesis that the NH2-terminal portion of the fibrin beta-chain is involved in the polymerization reaction by thrombin. The propositus' daughter and two sisters had the same abnormal fibrinogen. This unique inherited abnormal fibrinogen was designated as fibrinogen Ise. During these studies, we found that a very high concentration of thrombin cleaves not only the A alpha Arg19-Val20 bond but also the COOH-terminal region of alpha-chains, which results in the generation of further degraded alpha-chains with apparent molecular weights of 44,000 or less.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
5/15. Isolation and characterization of an acquired antithrombin antibody.A 68-year-old man, following mitral valve replacement, presented with a low-grade chronic consumptive coagulopathy. Laboratory analysis showed mild fibrinolysis, minimal effect of coumadin therapy, and a prolonged thrombin time (greater than 150 seconds using bovine IIa). When purified human IIa was used the thrombin time normalized to within 17 seconds of controls, suggesting a possible inhibitor of bovine IIa. An anti-IIa antibody was isolated by protein A-sepharose (Sigma, St Louis, MO) chromatography followed by affinity chromatography using a bovine IIa-sepharose column. The effects of this purified anti-IIa antibody on both bovine and human IIa procoagulant and anticoagulant functions were studied. The isolated immunoglobulin g (IgG) was observed to inhibit bovine IIa in all assays tested. This IgG was also able to slightly prolong fibrinogen clotting by human IIa. Using an enzyme-linked immunosorbent assay it was observed that the IgG bound to bovine IIa, bovine II, human IIa, but not to human II. Further, binding was detectable at approximately 50-fold lower concentrations to bovine IIa (1 nmol/L IgG concentration) than to human IIa (50 nmol/L IgG concentration). The effect of the antibody on the reaction between IIa and AT III/heparin was investigated. Human IIa was found to be protected from AT III/heparin neutralization in the presence of this antibody. These results suggest that this patient developed an antibody that strongly binds to and inhibits the bovine IIa in all assays tested. However, the antibody only significantly affects human IIa neutralization by AT III/heparin, and has little effect on the human IIa procoagulant activity. These data suggest that the decreased effect of AT III/heparin on this patient's IIa may have been a contributing factor in his coagulopathy. The exact cause of this antibody development is unclear, but the role of bovine topical thrombin used during cardiac valve replacement surgery is suspect.- - - - - - - - - - ranking = 2keywords = chromatography (Clic here for more details about this article) |
6/15. fibrinogen Birmingham: a heterozygous dysfibrinogenemia (A alpha 16 ArgHis) containing heterodimeric molecules. fibrinogen was isolated from the plasma of a 25-year-old female with a history of mild bleeding and several recent moderate to severe hemorrhagic episodes. Coagulability with thrombin approached 100% and varied directly with the time of incubation with the enzyme. High-performance liquid chromatography analysis of thrombin-induced fibrinopeptide release demonstrated retarded fibrinopeptide a (FPA) and fibrinopeptide b (FPB) release and the presence of an abnormal A peptide (FPA) amounting to 50% of the total. The same biochemical abnormalities were found in her asymptomatic father. Amino acid analysis and carboxypeptidase digestion of FPA demonstrated the substitution of His for Arg at A alpha 16. In contrast to the thrombin- and reptilase-sensitive Arg-Gly bond in the normal A alpha chain, the abnormal A alpha chain (A alpha) sequence is resistant to reptilase attack but is slowly cleaved by thrombin. To evaluate whether Birmingham A alpha and A alpha chains had been assembled nonselectively into heterodimeric (ie, 50% A alpha, A alpha) and homodimeric (ie, 25% A alpha, A alpha; 25% A alpha, A alpha) species, the clot and the clot liquor resulting from reptilase treatment of normal or Birmingham fibrinogen were separated, and each was then further incubated with thrombin to release remaining fibrinopeptides. Assuming that fibrinogen Birmingham contained heterodimeric molecules and that these and the normal molecules were completely incorporated into a reptilase clot, the expected coagulability would be 75%. In addition, subsequent thrombin treatment of the reptilase clot would release 50% of the total FPA and 75% of the total FPB present in the original sample. On the other hand, if only homodimeric fibrinogen species (50% A alpha, A alpha; 50% A alpha, A alpha) existed, the maximum reptilase coagulability would be 50%, and after thrombin treatment, 50% of the total FPB and no FPA would be recovered from the reptilase clot. We found the propositus's fibrinogen to be 68% coagulable, and we recovered 45% of the FPA and 70% of the FPB from the reptilase clot. Essentially the same coagulability and distribution of fibrinopeptides was found in the reptilase clot from her father's fibrinogen. We therefore conclude that fibrinogen Birmingham contains heterodimeric species (A alpha, A alpha) amounting to approximately 50% of the circulating fibrinogen molecules. The existence of heterodimers is consistent with a nonselective intracellular process of constituent chain assembly of dimeric plasma fibrinogen molecules.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
7/15. antithrombin iii deficiency: an etiology of budd-chiari syndrome.This report documents a unique case of budd-chiari syndrome associated with antithrombin iii deficiency and massive thrombus in the superior vena cava and right atrium. This aberration of the coagulation mechanism is proposed as an etiologic factor in the pathogenesis of hepatic venous obstruction whenever the cause is obscure. Antithrombin III was restored to a level adequate to permit thrombus extraction with cardiopulmonary bypass and relief of portal hypertension with a mesoatrial shunt. The protocol for reversing this hypercoagulable state with fresh frozen plasma and warfarin (Coumadin) is discussed, and technical innovations employed in the management of this complex problem are described.- - - - - - - - - - ranking = 0.0022983957746181keywords = extraction (Clic here for more details about this article) |
8/15. fibrinogen Kawaguchi: an abnormal fibrinogen characterized by defective release of fibrinopeptide a.A congenital dysfibrinogenemia was found in a 32-year-old asymptomatic female and her immediate family. The propositus, apparently a heterozygote for the abnormality, characteristically showed defective release of fibrinopeptide a from half of her fibrinogen molecules. No fibrinopeptide a was cleaved off from the isolated abnormal molecule by thrombin or snake venoms (Reptilase and ancrod) as evidenced by radioimmunoassay, high performance liquid chromatography and determination of the NH2-terminal amino acids. The abnormal fibrinogen formed a solid gel solely by the release of fibrinopeptide b upon incubation with thrombin. We provisionally designate this abnormal fibrinogen as "fibrinogen Kawaguchi", although possible identity with other abnormal fibrinogens is not excluded.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
9/15. Inherited fibrinolytic disorder due to an enhanced plasminogen activator level.A family with "in vitro" increased red-cell fall out from the blood clot was studied. One member of the family (JVM) had a clinical history of hemorrhages after minor trauma or dental extractions. Routine coagulation and platelet function were normal except for the fibrinogen level which was slightly low in several members. The antigenic as well as functional evaluation of factor xiii was within normal limits. No factor xiii inhibitors were present. An increase in the clot permeability index was observed in most family members. The study of the fibrinolytic system showed an enhanced lysis of euglobulins, a normal plasminogen value, normal level of fibrin/ogen degradation products, normal fibrinolytic inhibitors, and an increase in the activity of the plasminogen activator. The activity of this activator was inhibited by an antiserum against tissue-type plasminogen activator. The t-pA inhibitor was in the normal range. It is concluded that the family studied in this paper shows familial alteration in the fibrinolytic system due to an excess of plasminogen activator immunologically related to that in human tissue.- - - - - - - - - - ranking = 0.0022983957746181keywords = extraction (Clic here for more details about this article) |
10/15. factor vii survival studies in factor vii Padua abnormality.Two patients with factor vii Padua abnormality were studied during prophylactic administration of factor vii concentrates for tooth extractions. The amounts administered were 15.8 and 17.2 units/kg body weight in 30 and 20 min, respectively. The end infusion factor vii activity levels were 55 and 64%, respectively. survival times of the exogenous components were 6.5 and 6 h, respectively. Satisfactory hemolstasis was obtained in both instances.- - - - - - - - - - ranking = 0.0022983957746181keywords = extraction (Clic here for more details about this article) |
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