Beta 2 adrenergic receptor polymorphisms: association with factor VIII and von Willebrand factor levels and the risk of venous thrombosis.

Several studies have shown that elevated plasma levels of coagulation factor VIII (FVIII) are a risk factor of venous thrombosis. This risk remained after correction for the main determinants of FVIII levels, blood group and vonWillebrand factor (VWF) [1–3]. FVIII activity levels (FVIII:C) ‡150 IU dL increase the risk of a first venous thrombosis fivefold when compared to levels >100 IU dL. The prevalence of FVIII:C ‡ 150 IU dL among thrombosis patients is 25%. Since these levels are found in 10% of the population, the contribution of elevated FVIII levels to all thrombotic events in the population is considerable [2]. Several lines of evidence support the idea that high FVIII levels are indeed causative to thrombosis and not a consequence of the thrombotic event, such as a dose-dependent relationship with risk [2], the persistence of elevated levels over time [3] and familial clustering [4]. The last of these, familial clustering, supports the hypothesis that FVIII levels are, at least in part, determined genetically. Because no variations have been found in the FVIII and VWF genes that are associated with thrombosis [1,5], it is likely that genes encoding proteins regulating plasma levels of FVIII and VWF are involved. A candidate regulator of FVIII levels is the b2-adrenergic receptor (b2AR). It is well-known that epinephrine infusion causes a significant rise in FVIII levels. This effect can be blunted by prior administration of a b-blocker. Hoppener et al. [6] showed that in patients with venous thromboembolism and FVIII levels >175 IU dL, FVIII:C levels could be effectively lowered by treatment with propranolol. FVIII:C returned to its initial elevated levels within 2 months after discontinuation of treatment. However, Schönauer et al. [7] reported that in patients with venous thromboembolism and FVIII levels >170 IU dL, propranolol administration could not lower FVIII levels significantly. We approached this issue from a different angle by determining the possible association of single nucleotide polymorphisms (SNP) in the b2AR gene with FVIII and VWF levels and thrombotic risk. We studied three coding b2AR SNPs, that have previously been implicated in clinically relevant effects [8–10], in a large population-based, case–control study, the Leiden Thrombophilia Study (LETS). The LETS consists of 474 consecutive patients and 474 controls. All the patients were referred for anticoagulant treatment after a first objectively confirmed episode of deep vein thrombosis. Patients with underlying malignancies were excluded. The controls were matched for sex and age. DNA samples are available for 469 cases and 470 controls. FVIII:C was measured in the plasma of all these participants by a one-stage clotting assay. VWF antigen and FVIII antigen (FVIII:Ag) were measured by enzymelinked immunosorbent assay in the plasma of 301 patients and 301 controls. The design of this study has previously been described in more detail [11]. Using polymerase chain reaction–restriction fragment length polymorphism analyses the three b2AR SNPs, Arg16Gly, Glu27Gln and Thr164Ile, were determined. The distributions of genotypes for the three SNPs studied were inHardy–Weinberg equilibrium in the controls. The allele frequencies in the controls were consistent with those found in previous studies [12,13]. No differences in allelic distributions were observed between the cases and controls (Table 1). There was a protective effect on the occurrence of venous thrombosis for the rare allele of Thr164Ile, however, this effect was weak, with wide confidence limits around the odds ratio. Based on the frequencies of the SNPs, four different haplotypes were identified in our study population (Table 1) with the help of special software, ARLEQUIN [14]. These haplotypes corresponded to those reported in previous publications [10,12]. No significant differences were found in the distribution of the haplotypes between cases and controls. The trend we observed for the rare allele of Thr164Ile was recovered in haplotype 4. In addition, homozygotes for haplotype 2 showed an odds ratio of 1.67 (95% confidence interval: 0.75–3.74). Combining this with the results for the individual SNPs, it is unlikely that the gene for the b2AR contains a common polymorphism that is associated with the risk of venous thrombosis. Correspondence: A. Yaël Nossent, Department of Haematology, Haemostasis and Thrombosis Research Center, Leiden University Medical Center, C2-R, PO Box 9600, 2300 RCLeiden, the Netherlands. Tel.: +31 71 5266689/ +31 71 5261648; e-mail: [email protected]