Effect of Arginase Inhibition on Pulmonary L-Arginine Metabolism in Murine Pseudomonas Pneumonia

Rationale: Infection of the lung with Pseudomonas aeruginosa results in upregulation of nitric oxide synthases (NOS) and arginase expression, and both enzymes compete for L-arginine as substrate. Nitric oxide (NO) production may be regulated by arginase as it controls L-arginine availability for NOS. We here studied the effect of systemic arginase inhibition on pulmonary L-arginine metabolism in Pseudomonas pneumonia in the mouse. Methods: Mice (C57BL/6, 8-10 weeks old, female) underwent direct tracheal instillation of Pseudomonas (PAO-1)-coated agar beads and were treated by repeated intra-peritoneal injections of the arginase inhibitor 2(S)-amino-6-boronohexanoic acid (ABH) or PBS until lungs were harvested on day 3 of the infection. L-arginine metabolites were quantified using liquid chromatography-tandem mass spectrometry, NO metabolites nitrate and nitrite by Griess reagent and cytokines by ELISA. Results: NO metabolite concentrations (48.5 +/- 2.9 vs. 10.9 +/- 2.3 mu M, p<0.0001), as well as L-ornithine (29.6 +/- 1.7 vs 2.3 +/- 0.4 mu M, p<0.0001), the product of arginase activity, were increased in Pseudomonas infected lungs compared to naive controls. Concentrations of the NOS inhibitor asymmetric dimethylarginine (ADMA) were also increased (0.44 +/- 0.02 vs. 0.16 +/- 0.01 mu M, p<0.0001). Arginase inhibition in the infected animals resulted in a significant decrease in L-ornithine (14.6 +/- 1.6 mu M, p<0.0001) but increase in L-arginine concentration (p<0.001), L-arginine/ADMA ratio (p<0.001), L-arginine availability for NOS (p<0.001), and NO metabolite concentrations (67.3 +/- 5.7 mu M, p<0.05). Arginase inhibitor treatment also resulted in an increase in NO metabolite levels in animals following intratracheal injection of LPS (p=0.015). Arginase inhibition was not associated with an increase in inflammatory markers (IFN-gamma, IL-1 beta, IL-6, MIP-2, KC or TNF-alpha) in lung. Concentrations of the L-ornithine-dependent polyamines putrescine, spermidine and spermine were increased in Pseudomonas infected lungs (p<0.001, respectively) but were unaffected by ABH treatment. Conclusions: Systemic arginase inhibition with ABH during Pseudomonas pneumonia in mice results in an increase in pulmonary NO formation but no pro-inflammatory effect.