Hydroxyurea Therapy Reduces The Hypercoagulability State In a Sickle Cell Mouse Model

Several lines of evidence show that sickle cell anemia (SCA) is characterized by a hypercoagulable state. SCA patients present an elevated rate of thrombotic complications and increased biological markers of coagulation activation. In a previous study we demonstrated that therapy with hydroxyurea (HU) in SCA patients was associated with reductions in hypercoagulability markers, in addition to reductions in hemolysis, inflammation and endothelial activation markers (Colella et al Journal of Thrombosis and Haemostasis 2012). In the present study, we attempted to further investigate whether the effect of HU on hemostatic activation is dependent on fetal haemoglobin (HbF) expression, hemolysis and/or inflammation. To do so, we used the BERK murine model of SCA, a homozygous model of SCA, incapable of expressing HbF. In this BERK model, treatment with HU results in no improvement in hemolysis, due to the lack of HbF induction (Lebensburger et al Haematologica 2010). The murine SCA model was generated by transplantation of nucleated bone marrow cells from BERK mice into lethally-irradiated C57BL6 mice. Only mice expressing > 97% of human hemoglobin S (HbS) at 10 weeks after transplantation were used in the study. HU therapy at the dose of 50mg/kg (HU group) or saline alone (control mice) was administered through intraperitoneal injections 5 times per week, initiated at 11 weeks after transplantation (approximate time of sickle marrow engraftment). After a treatment period of 16 weeks, blood samples drawn from the inferior cava vein were collected and submitted to evaluation of plasma levels of thrombin-antithrombin III (TAT), a final marker of thrombin generation, in both animal groups. We also evaluated plasma levels of the inflammation marker interleukin 6 (IL6), and the endothelial marker soluble vascular cell adhesion molecule-1 (sVCAM-1). TAT, IL6 and sVCAM-1 were all measured by commercially-available ELISA kits. Statistical analyses were performed using Mann-Whitney’s U test. Treatment with HU resulted in a significant reduction of TAT plasma levels (106.7 μg/L vs 138.5 μg/L; P = 0.05). Plasma levels of IL6 (7.1pg/mL vs 12.4pg/mL; P = 0.18) and sVCAM-1 (765.8 ng/mL vs 848.4 ng/mL; P= 0.43) presented non-significant reductions with HU treatment. Our results show that, in this murine SCA model, long-term HU treatment results in an improvement in the hypercoagulability state, even in the absence of HbF induction, or of a significant improvement of inflammation or endothelial activation. This indicates that HU may have a direct effect on inhibiting the activation of coagulation in SCA.