kaolinite and Hemophilia-A

: Thrombin generation test (TGT) is well established tool to research blood coagulation in plasma of hemophilia patients. Traditionally coagulation in this test is triggered by a tissue factor (TF), an extrinsic coagulation pathway activator. However, it is known that disorders of the intrinsic pathway are most important for coagulation in hemophilia. In this study, we hypothesized that triggering coagulation via the intrinsic pathway could increase a sensitivity of the TGT to monitor hemophilia treatment. The aim of this study was to compare thrombin generation in hemophilia A patients with inhibitors to factor VIII before and after infusion of bypassing agent [recombinant-activated factor VIIa (rVIIa)] using standard activation of coagulation by TF or by kaolin, an activator of coagulation by intrinsic pathway. Endogenous thrombin potential (ETP) in nine patients was measured. ETP before (ETP0) and 60 min after rVIIa infusion (ETP60) were compared. It was shown that ETP0 and ETP60 were significantly different when using any coagulation activator (paired Student's t test, P = 0.017 and 3.7 × 10 for clotting activation by TF and kaolin, respectively). The ratios of ETP60/ETP0 were 1.2 ± 0.2 or 30.0 ± 22.4 (mean ± SD, n = 9) for coagulation activated by TF or kaolin, respectively, and were significantly different (paired Student's t test, P < 0.005). The TGT clearly distinguished between ETP0 and ETP60 in the case of any coagulation activator, but ETP increasing after rVIIa infusion was significantly higher when activated with kaolin. This provided increased sensitivity of this method for monitoring hemophilia therapy.

Topics: Adult; Antidiarrheals; Female; Hemophilia A; Humans; Kaolin; Male; Middle Aged; Thrombin; Young Adult

Hemophilia A (HA) patients with similar factor VIII levels can demonstrate varying bleeding tendencies. In particular, 10-15% of all severe HA patients (FVIII:C<1IUdL(-1)) do not require regular replacement therapy. Modern global coagulation assays can help to detect and study this "mild" bleeding phenotype. Here, we investigated the coagulation status of different bleeding phenotypes using various types of global coagulation assays.. Ten HA patients with severe phenotype and eleven patients with mild phenotypes were included in the study. For each patient, thromboelastography (TE), thrombodynamics (TD), and kaolin- or tissue factor-induced thrombin generation (TG) were measured. TG in platelet-rich plasma (PRP) was investigated using our original modification when the thrombin generation curve showed two peaks, previously shown to depend on platelet activity. We also utilized TG and TD with the addition of thrombomodulin.. The second peak amplitude and ETP of PRP TG were the only parameters that were significantly higher in mild bleeders (peak 41.6 ± 3.5 nM, ETP 1966 ± 169 nM*min) than in patients with severe bleeding (peak 28.3 ± 3.3 nM, ETP 1359 ± 130 nM*min).. Our results suggest that severe and mild HA phenotypes could be distiguished by TG assay in PRP suggesting that difference in platelet activity can be involved in the phenotype formation. According to our previous results we can suppose that the mechanism of the phenotypic heterogeneity is linked with TG mediated by PS-expressing platelets.

Topics: Adult; Aged; Blood Coagulation Tests; Hemophilia A; Humans; Kaolin; Middle Aged; Platelet-Rich Plasma; Thrombelastography; Thrombin; Thromboplastin; Young Adult

A limitation of bypassing agent therapy for haemophilia patients with inhibitors is the absence of a laboratory assay, which predicts the clinical response to treatment. Recent investigations have demonstrated the potential for thromboelastography to assess the effects of bypassing agent therapy in this patient population. While tissue factor activation has been used in several prior studies, a recent multicentre study failed to demonstrate an expected concentration-response effect of rFVIIa and called into question the tissue factor activation methods that have been employed. A comparison of kaolin to two concentrations of tissue factor as the activation method for thromboelastography was investigated in patients with haemophilia. We performed kaolin and tissue factor activated thromboelastography on blood from inhibitor and non-inhibitor patients with and without addition of rFVIIa and rFVIII. The results demonstrate that kaolin leads to a longer R, K and angle than the higher dilution of tissue factor (1:17 000) at baseline (no factor) and after addition of rFVIIa for both the inhibitor and non-inhibitor patients. Kaolin led to a longer R and K in comparison to a low dilution of tissue factor (1:42 000) following the addition of rFVIIa in the inhibitor patients. The longer R and K allows for better discrimination of the effects of rFVIIa thus making kaolin the most sensitive activation method in this setting. Thus kaolin activated thromboelastography should be considered an effective, perhaps the most effective, activator when utilizing thromboelastography to assess the effects of rFVIIa in haemophilia patients with inhibitors.

Topics: Adolescent; Adult; Blood Coagulation; Drug Interactions; Factor VIIa; Factor VIII; Hemophilia A; Humans; Kaolin; Male; Recombinant Proteins; Thrombelastography; Young Adult

To gain greater insight into the nature of the bleeding tendency in hemophilia, we compared the spatial dynamics of clotting in platelet-free plasma from healthy donors and from patients with severe hemophilia A or B (factor VIII:C or IX:C<1%). Clotting was initiated via the intrinsic or extrinsic pathway in a thin layer of nonstirred plasma by bringing it in contact with the glass or fibroblast monolayer surface. The results suggest that clot growth is a process consisting of two distinct phases, initiation and elongation. The clotting events on the activator surface and the preceding period free of visible signs of clotting are the initiation phase. In experiments with and without stirring alike, this phase is prolonged in hemophilic plasma activated by the intrinsic, but not the extrinsic pathway. Strikingly, both hemophilia A and B are associated with a significant deterioration in the elongation phase (clot thickening), irrespective of the activation pathway. The rate of clot growth in hemophilic plasma is significantly lower than normal and declines quickly. The resulting clots are thin, which may account for the bleeding disorder.

Topics: Blood Coagulation; Cells, Cultured; Factor IX; Factor VIII; Factor XI; Glass; Hemophilia A; Hemophilia B; Humans; In Vitro Techniques; Kaolin; Polyethylene Terephthalates; Thrombin; Time Factors

We have developed a Silica Clotting Time (SCT) test suitable to screen patients with lupus anticoagulants (LA) and compatible with photo-optical instruments. The SCT results were considered to be positive for LA whenever the clotting times were longer than the upper normal limit at low phospholipid concentration and to be confirmed when the prolonged clotting times were corrected to normal by high phospholipid concentration. We studied plasmas from healthy subjects, patients with known diagnoses of LA, patients with acquired deficiencies of blood coagulation and hemophiliacs with anti-factor VIII antibodies. The test was positive for all LA patients, and negative for all non-LA patients except 7 hemophiliacs with anti-factor VIII antibodies. Our data indicate that the SCT is a sensitive test, suitable for screening patients suspected of having LA. Its compatibility with photo-optical instruments makes it a suitable candidate to replace the kaolin clotting time. The contemporaneous performance of SCT at low and high phospholipid concentrations provides screening and confirmation in a single procedure.

Topics: Anticoagulants; Autoantibodies; Blood Coagulation Disorders; Blood Coagulation Tests; Factor VIII; Hemophilia A; Humans; Kaolin; Liver Cirrhosis; Lupus Coagulation Inhibitor; Mass Screening; Phospholipids; Sensitivity and Specificity; Silicon Dioxide

Comparative analysis of high molecular weight kininogen (HK) in various commercial congenital and immunodepleted deficiency plasmas was performed by immunoblotting of HK. It was found, that some artificially depleted deficiency plasmas contained proteolytically cleaved, kinin-free kininogen. In contrast, in all congenitally deficient plasmas, HK was present in the intact, single chain form. Thus, cleavage of kininogen could have been triggered by or during the immunodepletion procedure. It was seen, that the degree of proteolytic cleavage and degradation of HK in depleted plasmas differed among various manufacturers. E.g. depleted products of one company contained only trace amounts of cleaved HK, in contrast to products of another one, in which HK was completely degraded. The immunoblot analysis of HK reflects the occurrence of proteolytic events during the production of artificially deficient plasmas and can therefore serve as a quality control method.

Topics: Blood Coagulation Tests; Electrophoresis, Polyacrylamide Gel; Hemophilia A; Hemophilia B; Humans; Immunoblotting; Kaolin; Kininogens; Molecular Weight; Plasma; Protein C Deficiency

A simple microtechnique for carrying out partial thromboplastin time with kaolin tests with 2 microliter or less of test plasma is described. For single stage factor assays, less than 1 microliter of test solution may be used. Reagents and test plasma are loaded in sequence into a 10 microliter, long needle syringe and introduced into a micro test-tube immobilized in a water bath. The end-point is taken as a positive clearing of kaolin turbidity from the mixture while stirring. Correlation with normal techniques has been excellent.

Topics: Blood Coagulation Tests; Factor IX; Factor VIII; Hemophilia A; Hemophilia B; Humans; Kaolin; Partial Thromboplastin Time

Topics: Afibrinogenemia; Blood Coagulation Disorders; Blood Coagulation Tests; Factor V Deficiency; Factor VII Deficiency; Factor X Deficiency; Factor XI Deficiency; Factor XII Deficiency; Factor XIII Deficiency; Hemophilia A; Hemophilia B; Humans; Hypoprothrombinemias; Kaolin; Phosphatidylethanolamines; Prekallikrein; Prothrombin Time; Thrombin; Thromboplastin; von Willebrand Diseases

Amphoteric polyelectrolytes with alkaline isoelectric points (for example Ampholine 9-11) intended for use in isoelectric focussing were found to shorten the prolonged partial thrombosplastin clotting time with kaolin (PTTK) of plasma deficient in factor VIII. The response of this PTTK to Ampholine 9-11 was linear when plotted on double log paper with a slope slightly greater than that of factor VIII itself. Ampholine 9-11 at 2% concentration had factor VIII and factor IX activity equal to that of normal plasma as well as somewhat less activity as factor XI. It had no significant correcting effect on the clotting of plasmas deficient in factors V, VII, X, II OR XII. Ampholine 9-11 inhibited low concentrations of contact activators and its effect in correcting factor VIII deficient plasma was found to be related to the degree of contact activation.

Topics: Amino Acids; Blood Coagulation; Blood Coagulation Disorders; Dose-Response Relationship, Drug; Hemophilia A; Humans; In Vitro Techniques; Isoelectric Point; Kaolin

Reagents may be prepared from normal plasma and used with the prothrombin time and partial thromboplastin time tests to distinguish isolated defects of factors I, II, VII, VIII, IX, X, XI, or XII.

Topics: Adolescent; Adult; Aluminum Hydroxide; Blood Coagulation Disorders; Blood Coagulation Tests; Child; Child, Preschool; Factor XI Deficiency; Female; Hemophilia A; Hemophilia B; Humans; In Vitro Techniques; Kaolin; Male; Middle Aged; Phospholipids; Plasma; Prothrombin Time; Thromboplastin; Tromethamine; von Willebrand Diseases

Topics: Adult; Blood Coagulation Tests; Blood Protein Electrophoresis; Blood Transfusion; Factor IX; Factor VIII; Freeze Drying; Hemophilia A; Hemophilia B; Humans; Kaolin; Male; Thromboplastin

Topics: Anhydrides; Animals; Anticoagulants; Antigens; Chromatography; Chromatography, Gel; Factor IX; Factor V; Factor VIII; Factor X; Factor XII; Fibrinogen; Hemophilia A; Humans; Immunoelectrophoresis; Kaolin; Manganese; Rabbits; Succinates; Swine; Thrombin; von Willebrand Diseases

Topics: Blood Coagulation Disorders; Blood Coagulation Tests; Hemophilia A; Hemophilia B; Humans; Kaolin; Thromboplastin

Topics: China; Ethnicity; Female; Hemophilia A; Humans; Kaolin; Male; Prothrombin Time

Topics: Blood Coagulation Tests; Factor IX; Factor V; Factor VIII; Factor X; Factor XI; Factor XII; Fibrinogen; Hemophilia A; Humans; Kaolin; Prothrombin

Topics: Adsorption; Blood Platelets; Calcium Chloride; Centrifugation, Zonal; Citrates; Edetic Acid; Factor VIII; Glucose; Hemophilia A; Humans; In Vitro Techniques; Kaolin; Sodium Chloride

Topics: Adsorption; Arthritis, Rheumatoid; Factor XI; Factor XII; Gout; Hemophilia A; Humans; Kaolin; Stimulation, Chemical; Synovial Fluid

Topics: Adenine Nucleotides; Adenosine Triphosphate; Afibrinogenemia; Blood Coagulation; Blood Coagulation Factors; Blood Platelets; Cell Membrane; Epinephrine; Glass; Hemophilia A; Hemostasis; Humans; Kaolin; Norepinephrine; Phagocytosis; Reserpine; Serotonin

Topics: Blood Coagulation Tests; Factor VIII; Hemophilia A; Humans; Indicators and Reagents; Kaolin; Methods

Deficiencies of factor VIII (in haemophilia) and factor IX (in Christmas disease) prolong the partial thromboplastin time. If normal plasma is treated with alumina, the factor VIII remains but the factor IX is removed and can subsequently be recovered by elution of the alumina. If a long partial thromboplastin time is found on investigating a male patient whose history suggests a life-long bleeding disorder, the plasma may be retested after adding either alumina-adsorbed normal plasma or eluate. If the patient's partial thromboplastin time is shortened (relative to the control) by adding adsorbed normal plasma the patient is likely to be a haemophiliac; but if it is shortened by adding eluate then he is likely to have Christmas disease. Practical details for carrying out these manoeuvres are given and experiments on the validity of the test described.

Topics: Blood Coagulation Tests; Hemophilia A; Hemophilia B; Kaolin; Thromboplastin; Time Factors

Topics: Adenine Nucleotides; Blood Coagulation Disorders; Blood Coagulation Factors; Blood Coagulation Tests; Blood Platelet Disorders; Blood Platelets; Edetic Acid; Erythrocytes; Factor XII; Hemophilia A; Hemophilia B; Kaolin; Lipoproteins; Microscopy; Microscopy, Phase-Contrast; Pharmacology; Platelet Factor 3; Platelet-Rich Plasma; Research; von Willebrand Diseases

The partial thromboplastin time test provides a convenient and sensitive screening procedure for deficiencies of thromboplastic factors, especially factors VIII and IX. The test is carried out after preincubating the plasma for 10 minutes with kaolin, and Inosithin is used as a platelet substitute. The ;normal range' of the test has been estimated in terms of the differences encountered between random normal plasmas tested in pairs, because individual patients are usually tested against single control subjects. A patient's partial thromboplastin time should be regarded as abnormal if it is more than six seconds longer than the control time. In the diagnosis of haemophilia, patients' plasmas with concentrations of factor VIII as low as about 20% might be regarded as being within the range of normal, if the selected control subject's factor VIII happened to lie near the lower end of the normal range. When mild haemophilia is suspected, discrimination may be improved by diluting both the patient's and the control plasmas 1 in 20 in haemophilic plasma. With the test modified in this way the clotting time is prolonged, though the range of differences among normal subjects is unaltered, and plasmas with factor VIII concentrations below about 30%, i.e., in undiluted plasma, would be unlikely to be regarded as normal. The partial thromboplastin time may be similarly modified as a screening test for factor IX deficiency.Some clinical examples are reported.

Topics: Aged; Blood Coagulation Tests; Diagnosis; Factor VIII; Hemophilia A; Hemophilia B; Humans; Indicators and Reagents; Kaolin; Partial Thromboplastin Time; Plasma; Reference Values; Thromboplastin

Topics: Blood Coagulation; Hemophilia A; Humans; Kaolin