Effects of etamsylate on equine platelets: In vitro and in vivo studies q Dı ´dac Segura a , Luis Monreal a, * , Sı ´lvia Pe ´rez-Pujol b , Antonio Alonso b , Maribel Dı ´az-Ricart b , Rosa Brugue ´s b , Antonio Ordinas b , Gine ´s Escolar b a Servei de Medicina Interna Equina, Departament de Medicina i Cirurgia Animals, Facultat de Veterina ` ria, Universitat Auto ` noma de Barcelona, 08193 Bellaterra, Barcelona, Spain b Servicio de Hemoterapia y Hemostasia, Hospital Clı ´nic, IDIBAPS, Universitat de Barcelona, Barcelona, Spain Accepted 1 June 2006 Abstract The aim of this study was to investigate whether etamsylate produces equine platelet activation. In vitro and in vivo studies were designed in which seven and eight adult healthy horses were included, respectively. In the in vitro study, citrated blood was incubated with different concentrations of etamsylate, and P-selectin expression and annexin V binding were determined by flow cytometry. In the in vivo study, blood was collected before and 1 and 2 h after IV administration of etamsylate, and P-selectin expression was evaluated. In the in vitro study, a significant increase in P-selectin expression, leukocyte–platelet aggregate formation and annexin V binding were observed. In the in vivo study, a marked increase in P-selectin expression and heterotypic aggregate formation was seen in two and five horses, respectively, although no significant differences were detected when analyzing results from all the animals together. The results of the in vitro study indicate that etamsylate produces a pre-activation state in equine platelets, but this fact could be con- firmed by the in vivo study. Ó 2006 Elsevier Ltd. All rights reserved. Keywords: Horse; Etamsylate; P-selectin; Flow cytometry; Platelet activation 1. Introduction Etamsylate is an anti-haemorrhagic, non-thrombogenic drug that has been used in human and veterinary medicine for over 30 years in Europe. The first studies on etamsylate in both human and veterinary medicine demonstrated an action on primary haemostasis, resulting in a reduction in bleeding time in rabbits (Laporte, 1961), pigs (Deacock and Birley, 1969) and humans (Vinazzer, 1980). Later, etamsylate was shown to be effective in reducing blood loss and preventing intra- or peri-ventricular haemorrhage in efficacy studies performed in humans (Morgan et al., 1981; Cooke and Morgan, 1984; Benson et al., 1986; Ren- nie et al., 1986) and dogs (Ment et al., 1984), although other reports do not support these results (Elbourne et al., 2001; Schulte et al., 2005). In horses, the haemostatic action of etamsylate could be useful in reducing bleeding in several clinical conditions, such as exercise-induced pul- monary haemorrhage and during some surgical procedures (e.g., upper respiratory tract interventions). In considering etamsylate’s mode of action, Vinazzer (1980) postulated an improvement in platelet adhesiveness and restoration of capillary resistance, which could be mediated by the inhibition of prostacyclin (PGI 2 ). This inhibitory effect on arachidonic acid metabolism has also been demonstrated in other studies (Kovacs and Falkay, 1981; Ment et al., 1984; Rennie et al., 1986) and is indepen- dent of cyclooxygenase (Hutton et al., 1986). Further work has revealed some platelet-activating actions of etamsylate. Sack and Dujovne (1973) reported platelet aggregation in 1090-0233/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.tvjl.2006.06.002 q This manuscript represents a portion of a thesis submitted by the first author at the Autonomous University of Barcelona as partial fulfilment of the requirements for a PhD program. * Corresponding author. Tel.: +34 93 581 1968; fax: +34 93 581 2006. E-mail address: lluis.monreal@uab.es (L. Monreal). www.elsevier.com/locate/tvjl The Veterinary Journal 174 (2007) 325–329 The Veterinary Journal