S. Kitchen

Guidelines for the management of hemophilia.

Hemophilia is a rare disorder that is complex to diagnose and to manage. These evidence‐based guidelines offer practical recommendations on the diagnosis and general management of hemophilia, as well as the management of complications including musculoskeletal issues, inhibitors, and transfusion‐transmitted infections. By compiling these guidelines, the World Federation of Hemophilia aims to assist healthcare providers seeking to initiate and/or maintain hemophilia care programs, encourage practice harmonization around the world and, where recommendations lack adequate evidence, stimulate appropriate studies.

Inhibition of activated protein C and its cofactor protein S by antiphospholipid antibodies

Summary. We have investigated the effects of purified IgG fractions from plasma containing the lupus anticoagulant (LAC) and/or IgG anticardiolipin antibody (ACA) on the degradation of factor Va by an activated protein C‐protein S complex. Plasma samples from 10 patients were studied. LAC was detected by a Russells Viper venom technique. ACA was determined by ELISA. IgG fractions were obtained from each plasma sample by protein A‐Sepharose fractionation. This fraction was shown to exhibit ACAILAC activity. Using purified activated protein C (APC), protein S and phosphati‐dylserine/phosphatidylcholine, factor Va degradation was assessed in the presence and absence of IgG fractions from LAC/ACA containing plasmas. After 2 min incubation the mean factor Va degradation by APC and protein S in the presence of IgG LAC/ACA fractions was 14% compared with 52% with normal IgG. A similar effect was seen when phospholipid was substituted by washed freeze‐thawed platelets. Experiments employing varying concentrations of protein S and phospholipid revealed marked differences in respect of the inhibitory specificity of the different antiphospholipid antibodies. These results indicate that antiphospholipid antibodies have an inhibitory effect on the activated protein C/protein S complex and provide some explanation for a relationship between antiphospholipid antibodies and thrombosis.

Rapid reversal of oral anticoagulation with warfarin by a prothrombin complex concentrate (Beriplex): efficacy and safety in 42 patients

Summary.  Beriplex, a prothrombin complex concentrate (PCC), was administered to 42 patients requiring immediate reversal of their oral anticoagulant therapy. The dose administered was determined using the pretreatment International Normalized Ratio (INR). Blood samples were obtained before treatment and at 20, 60 and 120 min after treatment. The following investigations were performed on all samples – INR, clotting factors II, VII, IX and X, coagulation inhibitors protein C (PC) and antithrombin (AT), and other markers of disseminated intravascular coagulation, plasma fibrinogen, d‐dimer and platelet count. Immediate reversal of the INR, the vitamin K‐dependent clotting factors and PC was achieved in virtually all patients. Reduced AT levels were present in 18 patients before treatment. Further slight AT reductions occurred in four patients, but other associated abnormalities of haemostasis were observed in only one of the four patients. One patient with severe peripheral vascular disease, sepsis and renal and cardiac failure died of a thrombotic stroke following leg amputation, 48 h after receiving Beriplex. No other arterial and no venous thromboembolic events occurred within 7 d of treatment. Beriplex is effective in rapidly reversing the anticoagulant effects of warfarin, including PC deficiency, without inducing coagulation activation. Caution should continue to be exercised in the use of these products in patients with disseminated intravascular coagulation, sepsis or liver disease.

Guidelines on fibrinogen assays

Haematology departments in the UK have traditionally performed fibrinogen assays to detect decreased levels and abnormalities of fibrinogen, and to assess haemorrhagic risk. It has also been shown that elevated fibrinogen levels are a predictor of a variety of arterial cardiovascular events, and fibrinogen assays are sometimes recommended with this in mind. The Clauss fibrinogen assay (based on the thrombin clotting time) is the most popular technique in UK hospital laboratories, although many other methods are also in use. There appears to be great variability in both the source of reagents and the exact method used for the Clauss assay. Most laboratories are now equipped with automated coagulation analysers, and many of these perform a fibrinogen estimation derived from the degree of change of light scatter or optical density during the prothrombin time (PT-Fg). A number of problems have been described in the use of the PT-Fg method: it generally gives higher values than the Clauss technique, but the exact degree of discrepancy seems to depend on a number of different variables. International and National standards are available for fibrinogen, but do not appear to be universally used. These guidelines have been prepared against this background and recommend which methods should be used in various clinical settings, as well as highlighting a variety of problems with fibrinogen assays.

Plasma D-dimer levels and their relationship to serum fibrinogen/fibrin degradation products in hypercoagulable states.

Plasma D‐dimer was measured and compared with serum fibrinogen/fibrin degradation product levels (FDPs) in patients with disseminated intravascular coagulation (DIC) and other conditions associated with a hypercoagulable state. D‐dimer (N<200 ng/ml) was elevated in all 43 patients with DIC, in 48 of 59 patients with liver disease, in 22 of 27 patients with acute leukaemia at presentation, in 17 of 23 patients with malignant disease, in 29 of 39 women in the third trimester of a complicated pregnancy, in 17 of 18 patients with deep venous thrombosis and in only four of 27 patients with acute myocardial infarction. There was a significant correlation between plasma D‐dimer and serum FDP levels (P<0.01) as follows; DIC: r =r=0.58, liver disease: r=0.57, acute leukaemia: r=0.84, malignancy: r=0.87. The frequent elevation of D‐dimer observed in liver disease, acute leukaemia, malignancy and complicated pregnancy indicates that a hypercoagulable state is a common occurrence in these conditions although in liver disease elevated levels resulting from a failure of normal clearance mechanisms cannot be excluded. The close relationship between D‐dimer and FDP levels suggests that serum FDPs predominantly arise from the interaction of plasmin with crosslinked fibrin rather than with fibrinogen in the conditions in which these were compared.

Corn trypsin inhibitor in fluorogenic thrombin-generation measurements is only necessary at low tissue factor concentrations and influences the relationship between factor VIII coagulant activity and thrombogram parameters.

The fluorogenic calibrated automated thrombin-generation assay is influenced by contact pathway activation in platelet-rich and platelet-poor plasma. This influence lessens with increasing tissue factor (TF) concentrations and is inhibited by corn trypsin inhibitor (CTI). CTI is expensive and at what TF concentration its influence becomes irrelevant is unclear. Spiking of factor VIII (FVIII)-depleted plasma with FVIII, in samples without CTI, shows a plateau of thrombin generation at low normal FVIII levels. Given the association with thrombosis at high levels, a continuing increase in thrombin generation would be expected. We studied the effect of CTI on this relation by spiking experiments up to 4.8 IU/ml at 1 pmol/l TF and compared the influence of CTI at 1 and 5 pmol/l in platelet-poor plasma. CTI significantly influences thrombin generation in platelet-poor plasma at 1 pmol/l TF (difference of means for endogenous thrombin potential of 232.5 nmol/l per min, P < 0.0001) and peak of 48 nmol/l (P < 0.0001)) but not at 5 pmol/l. Spiking experiments without CTI confirm the hyperbolic relation between FVIII coagulant activity (FVIII:C) and endogenous thrombin potential with a plateau at 0.70–1.40 IU/ml. With CTI, a near-linear response up to 1.0 IU/ml was found with a plateau at 2.4–4.8 IU/ml. For peak thrombin, no plateau was reached with CTI. The present study confirms and extends previous data on CTI and the relationship between FVIII:C and thrombin generation. CTI is not necessary at 5 pmol/l TF, and thrombin generation remains dependent on FVIII:C up to 4.8 IU/ml at 1 pmol/l with CTI. Higher levels than previously thought may be needed to normalize thrombin generation.

An evidence‐based review and guidelines for patient self‐testing and management of oral anticoagulation

There is a limited evidence base for self‐testing and ‐management for oral anticoagulation management. Available data suggest that these are credible models for a significant minority of patients if underpinned by structured training and follow‐up. The guidelines presented are necessarily consensual and outline procedures for patient selection, training, product procurement, product maintenance, quality assurance procedures, dosage adjustment and clinical supervision. The cost‐effectiveness of these models remains to be elucidated within the UK. Further data on both health economic and clinical outcomes are required from UK based studies before widespread implementation of self‐testing and management can be recommended on a wider scale.

Guidelines on the laboratory aspects of assays used in haemostasis and thrombosis

Publications known to the writing group were supplemented with additional papers identified by searching Medline/PubMed for publications in the last 12 years using keywords: coagulation assays, amidolytic assays, haemostasis assays, and thrombophilia testing, in core clinical journals and English language. Additional relevant articles were identified by screening reference lists and by publications known to the writing group. The writing group produced the draft guideline which was subsequently revised by consensus by members

Wide variability in the sensitivity of APTT reagents for monitoring of heparin dosage.

AIM: To assess the sensitivity of activated partial thromboplastin time (APTT) reagents for monitoring heparin dosage using data from the UK National External Quality Assessment Scheme (NEQAS) for blood coagulation. METHODS: Data were reviewed from four surveys using samples prepared by addition of heparin to normal plasma in vitro and from two surveys in which samples were prepared using plasma from patients receiving heparin therapy (ex vivo samples). RESULTS: For both in vitro and ex vivo samples, notable differences between APTT reagents with respect to heparin sensitivity were noted. This indicates that a uniform therapeutic range of 1.5-2.5 calculated by the APTT ratio may not be appropriate for all reagents. Reagent sensitivity in ex vivo samples was substantially different to that in in vitro samples. CONCLUSIONS: The results of this large series of laboratories clearly indicate that reagent specific therapeutic ranges may be necessary, and that samples prepared by the addition of heparin to normal plasma in vitro can be misleading and should not be used.

Problems relating to the laboratory diagnosis of factor XIII deficiency: a UK NEQAS study.

Summary.  Familial (F)XIII deficiency is an extremely rare bleeding disorder. In most laboratories the diagnosis is initially established through a clot‐solubility screening test. We report here results from a series of UK NEQAS (Blood Coagulation). Proficiency Testing investigations, in which laboratories were provided with samples from normal individuals and from various subjects with FXIII deficiency with a request to perform their usual test for this disorder and to provide an interpretation of their results. Over 95% of centers were able to diagnose severe familial FXIII deficiency in previously untreated patients and to identify samples from normal subjects. However, both quantitative and qualitative methods produced widely variable results on samples obtained from previously treated individuals with FXIII deficiency but having measurable levels of FXIII. Data generated by UK NEQAS investigations suggested that solubility tests employing thrombin show greater sensitivity to FXIII deficiency, and this was confirmed in a subsequent single‐center study. Our results lead us to recommend the use of thrombin and acetic acid in the clot‐solubility screening test. Use of sensitive screening tests, and improvement in the accuracy and precision of quantitative FXIII assays will aid study of the clinical importance of moderate FXIII deficiency.