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Total of 5649 unselected adult patients were enrolled to identify impaired hemostasis prior to surgical interventions. Each patient was asked to answer a standardized questionnaire of bleeding history. Activated partial thromboplastin time (aPTT), prothrombin time (PT) and platelet counts (PC) including PFA-100 (platelet function analyzer): collagen-epinephrine (C/E) and collagen-ADP (C/ADP) were routinely done in all patients. Additional tests, bleeding time (BT) and von Willebrand factor (vWF: Ag) were performed only in patients with a positive bleeding history and/or evidence of impaired hemostasis, e.g. drug ingestion. The bleeding history was negative in 5021 patients (88.8%) but positive in the remaining 628 (11.2%). Impaired hemostasis could be verified only in 256 (40.8%) of these patients. The vast majority were identified by PFA-100: C/E (n = 250; 97.7%). The only abnormality found among patients with a negative bleeding history was a prolonged aPTT due to lupus anticoagulant in 9 patients (0.2%). The sensitivity of the PFA-100: collagen-epinephrine was the highest (90.8%) in comparison to the other screening tests (BT, aPTT, PT, vWF: Ag). The positive predictive value of the PFA-100: collagen-epinephrine was high (81.8%), but the negative predictive value was higher (93.4%).We identified 254 out of 5649 unselected patients scheduled for surgery at our hospital as having either acquired (n = 182) or inherited (n = 72) impaired primary hemostasis (platelet dysfunction including von Willebrand disease). All patients were initially pretreated with desmopressin (DDAVP) and further anti-bleeding drugs in case of DDAVP-non-response. Response to DDAVP or subsequent treatment(s) was defined as correction of any one of the abnormal PFA-100 platelet function tests. The non-responders were additionally treated with tranexamic acid or aprotinin; those with von Willebrand disease (vWD) received factor VIII concentrates with von Willebrand factor (vWF). Those still unresponsive to therapy received conjugated estrogens and, as a last attempt, a platelet transfusion. The administration of DDAVP led to a correction of platelet dysfunction in 229 of the 254 patients treated (90.2%). Tranexamic acid was effective in 12 of 16, aprotinin in 3 of 5 and factor VIII concentrates with vWF in all 4 patients with unresponsive to DDAVP. The remaining 6 patients were pretreated with conjugated estrogens, and 2 of these patients were additionally treated with platelet transfusion. The frequency of blood transfusion was lower, but not statistically significant (9.4% vs. 12.2%: p = 0.202) in preoperatively treated patients with impaired hemostasis than in patients without impaired hemostasis. In a retrospective group, the frequency of blood transfusion was statistically significant higher (89.3% vs. 11.3%: p < 0.001) in patients without preoperative correction of impaired than in patients without impaired hemostasis. Preoperative identification and correction of impaired primary hemostasis is possible in nearly all patients affected, and results in a reduction of homologous blood transfusions. [ABSTRACT FROM AUTHOR]

In joint bleeds not responding to substitution of factor VIII to normal factor VIII levels angiographic embolization might be considered as a therapeutic option, especially if surgical treatment is not possible. Embolization with embolization fluid might be more promising than use of microparticles, because microparticles are resolving after 8 weeks and there is the danger of insufficient thrombosis and reopening of the vessel in patients with severe hemophilia. [ABSTRACT FROM AUTHOR]

Despite identical calibration procedures, FVIII:C values are significantly greater than FVIII:AM and FVIII:Ag. This may be due to the fact that FVIII:C but not FVIII:AM and FVIII:Ag overestimates activated FVIII. The lower limits of reference ranges for FVIII entities derived from frozen and thawed plasma samples are markedly greater than reported in previous studies: FVIII:C, 73 IU/dl; FVIII:AM, 68 IU/dl; FVIII:Ag, 67 IU/dl Freezing and thawing of plasma samples reduces FVIII:C and FVIII:AM by 27% and 12%, respectively. Freezing and thawing does not influence FVIII:Ag. Fibrinogen but not FVIII plasma levels are markedly influenced by acute phase reaction as determined by hsCRP. Blood group non-0 or non-A2 individuals have substantially greater FVIII plasma levels than subjects with blood groups 0 or A2. Diagnosis of mild FVIII deficiency or increased FVIII levels as a risk factor for venous thromboembolism requires carefully established references ranges and standardized pre-analytical conditions with respect to AB0 blood group and freezing/ thawing of plasma samples. Before measuring FVIII, plasma samples should be frozen and thawed to obtain reliable results. [ABSTRACT FROM AUTHOR]

Mutations in VKORC1 cause coumarin resistance and are also responsible for differing dosing requirements close to the upper therapeutic range. Here we present the data in 16 patients from 14 families revealing 11 different missense mutations, six of which have not been reported previously. Our data may contribute in identifying further functional epitopes of VKORC1 and may provide a basis for a rational design of novel anticoagulants targeting VKOR in the future. [ABSTRACT FROM AUTHOR]

This highlights the usefulness of the mRNA analysis for routine diagnosis of such ambiguous cases including the presence of potential cryptic splice sites and clearly demonstrates the causality of the mutations which is also important for the genetic counseling of those families. [ABSTRACT FROM AUTHOR]

We investigated the influence of the factor V haplotype R2 (FVHR2), defined by the mutation FVH1299R, on the thrombin generation in plasma as well as on the clinical expression of bleeding symptoms in patients homozygous for FVIIA294V. Due to its impaired interaction with activated protein C (APC) FVHR2 increases the thrombin generation. Measurements of the endogenous thrombin potential in presence of high APC concentrations showed significant increased ETP values in plasma containing FVHR2 compared to references with factor V wild type. In our measurements this effect is comparable to the increase of thrombin generation caused by FVLeiden. This suggests FVHR2 to be a risk factor for thrombosis similar to factor VLeiden. On the basis of these findings, we investigated the influence of FVHR2 on the clinical expression of bleeding symptoms in factor VII deficient patients. A moderating effect of FVLeiden to bleeding symptoms in hemophilia A and B as well as in case of FIILazio has already been reported by other authors. This effect is explained by the increased APC resistance of FVLeiden. Based on the APC resistance we expected a similar effect for FVHR2. This was confirmed for four patients homozygous for the mutation FVIIA294V. Two of these patients did not carry FVHR2 and show bleeding symptoms. The two asymptomatic patients are heterozygous or homozygous for FVHR2 respectively. This indicates a moderating effect of FVHR2 to bleeding symptoms in patients homozygous for the mutation FVIIA294V. [ABSTRACT FROM AUTHOR]

The molecular basis of a factor VII deficiency was investigated in a Venezuelan patient. The five-year-old boy with a residual FVII:C of 1.7% and FVII:Ag < 8% suffered from epistaxis and easy bruising since birth. By sequence analysis a mutation <60T>C in the 5’ flanking region of the FVII gene and the novel mutation at the IVS7 donor splice site IVS7+1G>A were detected. The mutated splice site is located within the first repeat of a minisatellite region. Each repeat in this region contains a normally silent pseudo splice site. By an electrophoretic mobility shift assay (EMSA) the decrease of the binding of the nuclear factor HNF-4 by the promoter mutation -60T>C could be shown. To characterize the consequence of the mutated splice site IVS7+1G>A we constructed wildtype and mutated minigens spanning exons 7 to 8. HEK 293 cells were transfected with these constructs. In cells transfected with the mutant transcript no normal splicing occurred. Three different cryptic splice sites were used in vitro, all predicting a premature termination. [ABSTRACT FROM AUTHOR]