Eimeria bovis modulates adhesion molecule gene transcription in and PMN adhesion to infected bovine endothelial cells Carlos Hermosilla * , Horst Zahner, Anja Taubert Institute of Parasitology, Justus Liebig University Giessen, Rudolf-Buchheim-Str. 2, D-35392 Giessen, Germany Received 27 September 2005; received in revised form 3 January 2006; accepted 4 January 2006 Abstract Eimeria bovis is an important coccidian parasite of cattle causing severe diarrhea in young animals. Its first schizogony takes place in endothelial cells of the ileum resulting in the formation of macroschizonts 14–18 days p.i. This longlasting development suggests a particular immune evasion strategy of the parasite. Here, we analyse early innate immune reactions to E. bovis by determining the adhesion of polymorphonuclear neutrophils (PMN) to infected endothelial cell layers under flow conditions and the transcription of adhesion molecule genes in infected host cells. Bovine umbilical vein endothelial cells (BUVEC) were infected with E. bovis sporozoites. Sporozoites invaded BUVEC within 1 h and the first mature macroschizonts occurred 14 days p.i. PMN adhesion was enhanced in E. bovis-infected BUVEC layers as early as 8 h p.i.; maximum adhesion occurred 48 h p.i. Increased adhesion rates persisted until the end of the observation period at 14 days p.i. PMN adhered to both infected and uninfected cells within monolayers, suggesting paracrine cell activation. E. bovis infection upregulated the transcription of genes encoding for P-selectin, E-selectin, vascular cellular adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1). Most marked effects concerned E-selectin followed by P-selectin, VCAM-1 and ICAM-1. Increased transcript levels were found beginning 30 min p.i. and maximum values occurred 1–2 h p.i. (P-selectin) and 2–4 h p.i. (E-selectin, VCAM-1, ICAM-1). By 12–24 h p.i. levels had decreased to those of uninfected controls. Tumor necrosis factor a (TNFa)-induced PMN adhesion was significantly reduced in infected vs. uninfected BUVEC. Eimeria bovis also had suppressive effects on TNFa-mediated upregulation of adhesion molecule gene transcription. The data presented here suggest that infection of BUVEC with E. bovis on one hand induces proinflammatory reactions resulting in enhanced PMN adhesion mediated by upregulated adhesion molecule gene transcription but on the other downregulates TNFa-induced cell activation. q 2006 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved. Keywords: Eimeria bovis; Endothelial cells; PMN adhesion; Flow conditions; Adhesion molecules; Realtime PCR 1. Introduction Eimeria bovis is an enteropathogen of cattle causing clinical disease in calves and high economic losses worldwide (Fitzgerald, 1980; Daugschies et al., 1998). Instead of infecting enterocytes like other Eimeria species in cattle, free E. bovis sporozoites invade endothelial cells of the central lymph capillaries of the lacteals in the villi of the ileum, where they develop within a period of 14–18 days to large (up to 250 mm) first generation macroschizonts (Hammond, 1946). Eimeria infections in general are under immunological control and a variety of studies have been performed on adaptive immunity to E. bovis (Speer et al., 1985; Hughes et al., 1988, 1989; Fiege et al., 1992; Hermosilla et al., 1999). There is, however, little data available on the innate immune reactions to this parasite. Host endothelial cells could play a critical role in these reactions. They are highly immunoreactive cells, able to produce a broad range of adhesion molecules, cytokines and proinflammatory chemokines upon activation, thereby initiating leukocyte trafficking, e.g. by recruiting polymorphonuclear neutrophils (PMN), natural killer (NK) cells, T lymphocytes and monocytes to the site of infection (for reviews, see Tedder et al., 1995; Ebnet and Vestweber, 1999; Wagner and Roth, 2000). The endothelium-derived adhesion molecules, such as E-selectin, P-selectin, intercellular adhesion molecule 1 (ICAM-1) or vascular cellular adhesion molecule 1 (VCAM-1) are important in these processes (Tedder et al., 1995; Wagner and Roth, 2000). They regulate signaling between the cells and monitor their movement. E-selectin and P-selectin mediate the reversible process of tethering and International Journal for Parasitology 36 (2006) 423–431 www.elsevier.com/locate/ijpara 0020-7519/$30.00 q 2006 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.ijpara.2006.01.001 * Corresponding author. Tel.: C49 641 9938475; fax: C49 641 9938469. E-mail address: carlos.r.hermosilla@vetmed.uni-giessen.de (C. Hermosilla).