Abstract 182: Reduction in Interleukin-10 Level in Plasma and in Renal Tissue During Systemic Inhibition of Nitric Oxide Synthase in Anesthetized Mice

Abstract

Systemic inhibition of nitric oxide synthase (NOS) in mice increased tumor necrosis factor-alpha (TNFα) level in plasma. The present study examined the hypothesis that an inhibition of interleukin-10 (IL-10; anti-inflammatory cytokine) in response to NOS inhibition facilitates such enhancement in TNFα production. Levels of IL-10 and TNFα were measured (using ELISA kits) in plasma and in renal tissues obtained from anesthetized mice (C57BL6; ∼10 wks age) infused with or without NOS inhibitor, nitro-L-arginine methyl ester (L-NAME; 200 μg/min/kg, iv). Protein expressions of IL-10 and TNFα in the renal tissues were also measured by immuno-staining. L-NAME infusion alone for 130 min (n=6) resulted in decreased IL-10 levels in plasma (0.3±0.1 vs 2.6±0.6 ng/mL) and in renal tissue (0.5±0.1 vs 1.3±0.1 ng/mg protein) which were associated with increased TNFα levels in plasma (432±82 pg/mL vs undetected) and in renal tissue (58±7 vs 6±5 pg/mg protein) as compared to vehicle treated control mice (n=6). There was marked down-regulation of IL-10 protein expression (present only in the distal tubules in control mice) and up-regulation of TNFα protein expression (distal tubules and collecting ducts) in the renal tissues in L-NAME treated mice. Infusion of NO donor, S-nitroso-N-acetyl-penicillamine (SNAP; 25 μg/min/kg for 75 min) in a separate group of L-NAME treated mice (n=6) restored the IL-10 level in plasma (1.0±0.1 ng/mL) and in renal tissue (1.5±0.1 ng/mg protein) closer to the values in control mice indicating that L-NAME induced IL-10 responses are specific to NO inhibition. SNAP infusion in L-NAME treated mice also reduced TNFα level in plasma (to an undetected level) and in renal tissue (20±4 pg/mg protein) along with its protein immune-expression. Infusion of mouse IL-10 (0.075 ng/min/g for 75 min) in another separate group of L-NAME treated mice (n=7) also attenuated TNFα level in plasma (50±16 pg/mL) and in renal tissue (21±7 pg/mg protein) along with its protein immune-expression indicating that L-NAME induced TNFα responses are specific to reduction in IL-10 level. The results demonstrate that systemic NOS inhibition decreases IL-10 production and thus, minimizes its immune down-regulating action on the production of TNFα, particularly in the kidney.