title: Testosterone induces leucocyte migration by NADPH oxidase-driven ROS- and COX2-dependent mechanisms.
abstract: The mechanisms whereby testosterone increases cardiovascular risk are not clarified. However, oxidative stress and inflammation seem to be determinants. Herein, we sought to determine whether exogenous testosterone, at physiological levels, induces leucocyte migration, a central feature in immune and inflammatory responses and the mediating mechanisms. We hypothesized that testosterone induces leucocyte migration via NADPH oxidase (NADPHox)-driven reactive oxygen species (ROS) and cyclooxygenase (COX)-dependent mechanisms. Sixteen-week-old Wistar rats received an intraperitoneal injection (5 ml) of either testosterone (10(-7) mol/l) or saline. Rats were pre-treated with 5 ml of sodium salicylate (SS, non-selective COX inhibitor, 1.25 × 10(-3) mol/l, 1 h prior to testosterone or saline), flutamide (androgen receptor antagonist, 10(-5) mol/l), apocynin (NADPHox inhibitor, 3 × 10(-4) mol/l), N-[2-Cyclohexyloxy-4-nitrophenyl]methanesulfonamide (NS398, COX2 inhibitor, 10(-4) mol/l) or saline, 4 h before testosterone or saline administration. Leucocyte migration was assessed 24 h after testosterone administration by intravital microscopy of the mesenteric bed. Serum levels of testosterone were measured by radioimmunoassay. NADPHox activity was assessed in membrane fractions of the mesenteric bed by dihydroethidium (DHE) fluorescence and in isolated vascular smooth muscle cells (VSMC) by HPLC. NADPHox subunits and VCAM (vascular cell adhesion molecule) expression were determined by immunoblotting. Testosterone administration did not change serum levels of endogenous testosterone, but increased venular leucocyte migration to the adventia, NADPHox activity and expression (P < 0.05). These effects were blocked by flutamide. SS inhibited testosterone-induced leucocyte migration (P<0.05). Apocynin and NS398 abolished testosterone-induced leucocyte migration and NADPHox activity (P<0.05). Testosterone induces leucocyte migration via NADPHox- and COX2-dependent mechanisms and may contribute to inflammatory processes and oxidative stress in the vasculature potentially increasing cardiovascular risk.
authors.affiliation: *Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP 05508, Brazil.
authors.forename: Andreia Z
authors.initials: AZ
authors.lastname: Chignalia
authors.affiliation: *Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP 05508, Brazil.
authors.forename: Maria Aparecida
authors.initials: MA
authors.lastname: Oliveira
authors.affiliation: ‡Heart Institute, School of Medicine, University of São Paulo, São Paulo, SP 01246, Brazil.
authors.forename: Victor
authors.initials: V
authors.lastname: Debbas
authors.affiliation: §College of Medicine, University of Illinois at Chicago, Chicago, IL 60612, U.S.A.
authors.forename: Randal O
authors.initials: RO
authors.lastname: Dull
authors.affiliation: ‡Heart Institute, School of Medicine, University of São Paulo, São Paulo, SP 01246, Brazil.
authors.forename: Francisco R M
authors.initials: FR
authors.lastname: Laurindo
authors.affiliation: ∥Institute of Cardiovascular and Medical Sciences, University of Glasgow, G128TA, U.K.
authors.forename: Rhian M
authors.initials: RM
authors.lastname: Touyz
authors.affiliation: *Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP 05508, Brazil.
authors.forename: Maria Helena C
authors.initials: MH
authors.lastname: Carvalho
authors.affiliation: *Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP 05508, Brazil.
authors.forename: Zuleica B
authors.initials: ZB
authors.lastname: Fortes
authors.affiliation: †Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, SP 14049, Brazil.
authors.forename: Rita C
authors.initials: RC
authors.lastname: Tostes
mesh_terms: D000098:Acetophenones / Q000494:pharmacology; D000728:Androgens / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002465:Cell Movement / Q000187:drug effects*; D002478:Cells, Cultured; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007274:Injections, Intraperitoneal; D007962:Leukocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008642:Mesenteric Veins / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D018715:Microscopy, Video / Q000379:methods; D009131:Muscle, Smooth, Vascular / Q000166:cytology; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009578:Nitrobenzenes / Q000494:pharmacology; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism; D013739:Testosterone / Q000008:administration & dosage / Q000494:pharmacology*
chemical_list: D000098:Acetophenones; D000728:Androgens; D004791:Enzyme Inhibitors; D009578:Nitrobenzenes; D017382:Reactive Oxygen Species; D013449:Sulfonamides; D013481:Superoxides; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D013739:Testosterone; C056165:acetovanillone; D051546:Cyclooxygenase 2; D019255:NADPH Oxidases