Immunopharmacology and Inflammation Differential involvement of cyclooxygenase isoforms in neutrophil migration in vivo and in vitro Gustavo Batista Menezes a , Rafael Machado Rezende a , Pedro Elias Marques Pereira-Silva a , André Klein a , Denise Carmona Cara b , Janetti Nogueira Francischi a, ⁎ a Laboratório de Inflamação e Dor - Instituto de Ciências Biológicas, UFMG, Brazil b Laboratório de Neuroimunopatologia Experimental, Instituto de Ciências Biológicas, UFMG, Brazil abstract article info Article history: Received 15 April 2008 Received in revised form 29 July 2008 Accepted 21 August 2008 Available online 19 September 2008 Keywords: Leukocyte recruitment Cyclooxygenase inhibitors Boyden chamber Intravital microscopy Pretreatment using celecoxib, a cyclooxygenase (COX) 2 inhibitor, or indomethacin, a nonselective COX inhibitor, reduced lypopolyssaccharide (LPS)-induced leukocyte migration to the rat peritoneal cavity. The effect of celecoxib (12 mg/kg) or indomethacin (2 mg/kg) on neutrophil chemotaxis induced by formyl- methionyl-leucyl-phenylalanine (FMLP) in an in vitro chemotactic assay (Boyden chamber) was investigated. Celecoxib and indomethacin inhibited chemotaxis induced by FMLP (Control= 26.6 ± 1.45, Celecoxib = 12.8 ± 3.04, Indomethacin = 6.26 ± 2.19 cells/field). When observed under intravital microscopy, a mouse cremaster preparation was used to assess the microvasculature to further investigate which step of cell recruitment was affected by these drugs. Celecoxib and indomethacin inhibited leukocyte migration induced by 0.05 μg/kg LPS injected into the cremaster muscle. However, the effect of celecoxib was associated with reduced cell rolling and adhesion, whereas indomethacin was only effective at inhibiting cell adhesion. Furthermore, SC560 pretreatment (a COX-1 selective inhibitor) of normal or LPS-challenged tissues did not alter leukocyte migration or cell adhesion, but it did enhance leukocyte rolling activity in both cases. Taken together, these results indicate that: 1) COX-1 activity is mainly related to leukocyte traffic under physiological conditions, and 2) COX-2 activity is mainly related to cell traffic under inflammatory conditions in vascular beds, suggesting a possible effect of selective COX-2 inhibitors on the expression of adhesion molecules. © 2008 Elsevier B.V. All rights reserved. 1. Introduction Leukocyte recruitment is a prominent feature of inflammatory processes. The sequence involved in this phenomenon is a multistep process that includes an initial endothelium–leukocyte interaction (known as a tethering and rolling process), firm adhesion and subsequent transmigration out of the blood vessels (Kubes et al., 1991). After diapedesis, leukocytes migrate toward the site of the lesion in response to locally derived chemotactic factors. The molecules present in leukocytes and the endothelial lining that orchestrates this process belong to different families, which include selectins (E, P and L series) and α 4 -integrins (Kelly et al., 2007), among others. Selectins are important at the rolling stage, while integrins (mainly β 2 -integrins), lymphocyte function-associated antigen-1 (LFA-1) and macrophage associated chemokine (MAC-1) promote firm adhesion that is important for the subsequent transmigration event (Issekutz and Issekutz, 1992). Infiltrated neutrophils are first line defense cells and the importance of these cells in the initiation and resolution phases of the inflammatory process is well documented (Serhan, 2007). Despite the importance of the presence of leukocytes at the site of lesions and infection in relation to defending the host, the accumula- tion of these cells in tissues and organs is also a hallmark of the development of inflammatory conditions, such as atherosclerosis, severe pulmonary and renal diseases and acute myocardial infarction (Alvarez et al., 2001). In the last case, patients submitted to surgical reperfusion can present extensive lesioning of the cardiac muscle due the massive inflammatory process mediated by neutrophils, which are attracted by mediators released under hypoxic conditions (Abdel- Rahman et al., 2007). In this paradigm, modulation of the recruitment of these cells could provide a new way to regulate the inflammatory response. Nonsteroidal antiinflammatory drugs (NSAIDs) are widely used to control pain, fever and edematogenic conditions (Ferreira et al., 1973). The most accepted mechanism of action of these drugs is the inhibition of cyclooxygenase (COX) isoenzymes, which reduce the biosynthesis of eicosanoids, including prostaglandins and tromboxane (Vane, 1971). Prostaglandins are responsible for the majority of the primary symptoms of the inflammatory process (Ferreira et al., 1973; European Journal of Pharmacology 598 (2008) 118–122 ⁎ Corresponding author. Departamento de Farmacologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Avenida Antônio Carlos, 6627 - Campus da Pampulha Belo Horizonte, MG, CEP: 31270-901, Brazil. Tel.: +55 313409 27 15; fax: +55 31 3409 26 95. E-mail address: janettif@icb.ufmg.br (J.N. Francischi). 0014-2999/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.ejphar.2008.08.037 Contents lists available at ScienceDirect European Journal of Pharmacology journal homepage: www.elsevier.com/locate/ejphar