Abstract 5509: Arginase Activity Mediates Vascular Inflammation
We have shown before that activity of NOX2 NADPH oxidase has a critical role in causing vascular inflammation. We have also shown that diabetes-induced vascular endothelial cell (EC) dysfunction is mediated by increases in arginase activity which decreases availability of L-arginine to EC nitric oxide synthase (eNOS). While the specific role of arginase activity in vascular inflammation is not yet clear, increases in arginase I levels have been described in conditions of elevated oxidative stress and inflammation. We have now used a model of endotoxin-induced retinal inflammation to assess the hypothesis that increased arginase activity is involved in the development of vascular inflammation. Mice were injected with lipopolysaccharide (LPS, 4 mg/kg, ip, 3–16 hrs). Measurement of urea formation showed that retinal arginase activity was increased in the LPS-treated mice (1.9±0.3-fold, n=5). Quantitative PCR showed that arginase I (AI) mRNA was also increased with a peak at 12 hrs (2.2±0.1-fold, n=3). Expression of the AII isoform was unchanged. Western blotting, immunohistochemistry and flow cytometry confirmed an increase in AI protein expression, which was localized to vascular cells, glia and microglia. AI expression and arginase activity were also increased in LPS-treated glia and microglia. The LPS-induced increases in AI expression were correlated with increases in mRNA for MCP-1 (575.1±84.8-fold, n=5), TNF-alpha (55.6±10.3-fold, n=4) and iNOS (103.3±1.5-fold, n=3) and with increased leukocyte adhesion to the vessel wall (5.5±1.5 fold, n=3–5). Studies using knockout mice deficient in one copy of the AI gene and both copies of AII or mice treated with the selective arginase inhibitor ([S]-[2-boronoethyl]-L-cysteine-HCl, 20 mg/kg, iv) showed that LPS-induced increases in inflammatory gene expression and leukostasis were blocked by knocking down or inhibiting arginase. Furthermore, the LPS-induced increase in AI was abrogated by NOX2 deletion or apocynin treatment, indicating involvement of NADPH oxidase activity. In conclusion, LPS-induced vascular inflammation is mediated by NADPH oxidase-dependent increases in arginase activity.
This research has received full or partial funding support from the American Heart Association, AHA Greater Southeast Affiliate (Alabama, Florida, Georgia, Louisiana, Mississippi, Puerto Rico & Tennessee).