Brummel et al1 reported interesting results on different thrombin functions during tissue factor–induced whole blood coagulation. Thrombin generation, platelet activation, and activation of fibrinogen, factor V, and factor XIII (FXIII) were analyzed. FXIII activation was reported to occur slightly prior to fibrinopeptide A (FPA) release at a rate of 10.3 ± 0.9 nM/min and at a thrombin concentration of 0.84 ± 0.28 nM. However, these results by Brummel et al1 must be interpreted with caution.
FXIII activation is known to be strongly influenced by a common polymorphism in the FXIII A-subunit gene (FXIIIVal34Leu),2,3 which has been shown to be protective against myocardial infarction,4 ischemic stroke,5 and deep vein thrombosis.6This common G>T point mutation in codon 34, exon 2 of the A-subunit gene, which codes for the Val→Leu change, is only 3 amino acids from the thrombin activation site. The Leu allele is associated with increased cross-linking activity determined by an incorporation assay3,7 and a reduced clot formation time measured by thrombelastography.8 Kinetic studies on the activation reaction of FXIII by thrombin revealed increased catalytic efficiency (kcat/Km) for FXIII Leu34 compared with Val34.9,10Activation of FXIII Leu34 occurred at a similar rate as FPA release; FXIII Val34 was activated slower.9 Based on these findings, altered fibrin structures9 and wasteful, premature FXIII activation10 have been proposed as mechanisms of the protective effect of FXIIIVal34Leu.
Knowledge of the FXIIIVal34Leu genotype in these 6 individuals taking part in the study by Brummel et al1 is essential. Because of the high allele frequency of this polymorphism (approximately 0.25 for the Leu allele4) in the general population and the significant kinetic differences between genotypes, the Val34Leu polymorphism has to be taken into account when FXIII activation rates are studied.
Factor XIII genotypes and phenotypes
Drs Schroeder and Kohler raise an interesting point regarding the activation of factor XIII (FXIII) in whole blood concerning the Val34Leu polymorphism. They cite the work of Ariëns et al1-1 in which a 2.2- to 2.5-fold increase in the catalytic efficiency of thrombin toward homogenous factor XIII Leu34 over Val34 was observed in an in vitro assay system using purified thrombin and fibrinogen. We have observed that FXIII activation in Leu34 carriers (12 heterozygotes, 2 homozygotes) is faster (∼1.4 fold) than in Val34/homozygotes (23 individuals) in a Simplate (Organon Teknika, Durham, NC) bleeding-time blood model.1-2 Ariëns et al1-1 also observed that the Val34 release proceeded more slowly than fibrinopeptide A (FPA) release, whereas Leu34 release was at a rate similar to FPA. Drs Schroeder and Kohler seem to be implying that given the allelic frequency in the population (0.25) of Leu34 versus Val34 we should have observed, in a random population, a slower activation of FXIII than FPA release. Although it is conceivable that some of the 6 individuals studied are Leu34 variants (probability 0.32), the difference between the alleles with respect to FPA release and factor XIII activation would most likely fall within the error observed.1-3 The contributions of Leu34 FXIII versus Val34 FXIII in whole blood is a complicated issue because the plasma contribution from each of the alleles and their turnover rates are not known. Thus, genotyping alone would not be sufficient to establish the plasma concentrations of the 2 products.
Caution must be taken when individual genetic factors are used without considering the entire environment in which the biologic reaction takes place. Heterogeneity in the healthy population is a consequence of many genetic and environmental confounders and these variations do not necessarily lead to hemostatic changes. In healthy individuals, alterations in plasma coagulation factor levels range over ± 50% of the mean plasma value.1-4 The genotype and environment may alter both the qualitative and quantitative properties for every reactant in blood. This is illustrated by experiments in which FV Leiden is combined with reduced levels of tissue factor pathway inhibitor (TFPI). From in vitro measurements, van't Veer et al1-5 hypothesized that reduced expression of TFPI would enhance the prothrombotic effect of FV Leiden. Eitzman et al1-6 demonstrated that reduced expression of TFPI in FV Leiden transgenic mice is associated with thrombosis.
Predicting how coagulation should proceed in vivo from results obtained from purified systems has proven problematic. The order of events of fibrin formation in nonanticoagulated whole blood is not sequential, occurring as an integrated process involving FPA release, virtually simultaneous FXIII activation, and partial FPB release.1-7This result is not consistent with results obtained in anticoagulated plasmas1-8,1-9 and purified protein systems.1-10Our study1-3 involves the complex system of minimally altered whole blood and is thus a composite of all influences (genotype/phenotype and qualitative/quantitative). It is remarkable that healthy volunteers are, by and large, consistent in their response to tissue factor.
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