ENDOTHELIAL CELL ACTIVATION MEDIATED BY BERYTHRACTIVASE

Chudzinski-Tavassi, A.M.; Negrotto, S.; D’Atri, L.P.; Bezerra da Silva, M.; Pozner, R.G.; Lazzari, M.A.; Schattner, M.

Department of Thrombosis and Hemostasis, Hematological Research Institute, National Academy of Medicine, National Research Council (CONICET), Buenos Aires, Argentina.

The venom of Bothrops erythromelas, a species which occurs in northeastern Brazil, has a much higher procoagulant activity than other Bothrops species. Berythractivase is a 78 kDa metalloproteinase purified from B. erythromelas venom and its primary structure has been deduced from cDNA. In vitro, berythractivase is a prothrombin activator. Since Bothrops snake venoms induce local inflammatory lesions and disseminated intravascular coagulation (DIC), we have evaluated the effect of erythractivase on human umbilical vein endothelial cells (HUVEC). Morphological alterations were observed in HUVEC after 1 h of incubation with berythractivase(5 g/ml). Flow cytometry showed that berythractivase (Be) increased the expression of ICAM-1 (Control 12±1 vsBe 21±3 arbitrary fluorescence units (AFU), p<0.05, n=6) and E-selectin (Control 7±1 vsBe 25±1 AFU, p<0.05, n=4), without modifying the levels of VCAM-1 (Control 4±1 vs Be 4±0.5 AUF, n=4). Berythractivase increased the release of von Willebrand Factor (vWF) by 153% (p<0.05, n=4) and up-regulated prostacyclin (Control 1.3±0.2 vsBe 2.8±0.7 ng/ml, n=5, p<0.05) and IL-8 (Control 1.0±0.2 vs Be 3.1±0.1 ng/ml, n=6) levels (all measured by ELISA). vWF secretion was blocked by pretreating berythractivase with bothropic antiserum or EDTA, indicating a direct action of the proteinase. The prothrombotic and proinflammatory endothelial cell responses induced by berythractivase may be involved in the local lesions and systemic effects observed in humans envenomed by Bothrops snakes.

CORRESPONDENCE TO:

SCHATTNER MIRTA ANA, Pacheco de Melo 3081, Buenos Aires 1425 Argentina, Email: mschattner@hematologia.anm.edu.ar