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.

Sex differences in abdominal aortic aneurysms

“…For instance, administration of exogenous testosterone increased NOX activity and expression in adult Wistar rats as well as leukocyte migration to the adventitia (15). These effects were inhibited with the addition of flutamide (AR antagonist) or apocynin (NOX inhibitor) (15). This suggests that androgen signaling is able to induce leukocyte migration through a NOX-dependent mechanism, which may contribute to both oxidative stress and immune cell infiltration in AAA.…”

“…In opposition to estrogen signaling, androgen signaling increases NOX activity and expression. For instance, administration of exogenous testosterone increased NOX activity and expression in adult Wistar rats as well as leukocyte migration to the adventitia (15). These effects were inhibited with the addition of flutamide (AR antagonist) or apocynin (NOX inhibitor) (15).…”

“…Although these studies presented conflicting results, it is likely that androgen signaling differentially regulates COX-2 expression depending on multiple factors such as physiological serum androgen levels or inflammatory conditions. Because COX-2 is a major source of ROS production in the vasculature (15), further studies are warranted to investigate the cardiovascular implications of androgen signaling and ROS generation from COX-2 in AAA.…”

Constrictor prostanoids and uridine adenosine tetraphosphate: vascular mediators and therapeutic targets in hypertension and diabetes

“…Accordingly, long‐term treatment with testosterone, or with its non‐aromatizable androgen receptor agonist dihydrotestosterone, exacerbates endotoxin‐induced inflammation in the cerebral circulation by mechanisms that involve increased nuclear NF‐κB activation and increased levels of COX‐2 and inducible NOS (Gonzales et al ., ). Reinforcing a role for testosterone in vascular inflammation, data from our laboratory demonstrate that testosterone induces leukocyte migration by COX‐2‐dependent mechanisms (Chignalia et al ., ).…”

Reactive oxygen species: players in the cardiovascular effects of testosterone

“…Our group recently demonstrated that COX2-dependent ROS production is obligatory for testosterone-induced leukocyte migration (32), which may contribute to inflammation and vascular dysfunction. However, more studies on the role of androgens modulating COX-derived ROS and its implications for the cardiovascular system are warranted.…”