Inhibition of Nitrotyrosine Formation Reduces Endotoxin-Induced Liver Injury Irrespective of TNF-α

The hypotheses that nitrotyrosine formation is a downstream mechanism following TNF-α production in lipopolysaccharide (LPS)-induced liver injury and its inhibition reduces the injury were examined. Under anesthesia, experiments were performed 6h after the intravenous administration of LPS (10mg/kg) or saline using male Sprague-Dawley rats (300-400g). Liver injury was evaluated in 4 groups: control (n=4), LPS alone (n=5), LPS+quercetin (n=5), LPS+clodronate (n=5) by plasma alanine aminotransferase (ALT) level, nitrotyrosine concentration in liver and histological changes. Quercetin, an inhibitor of nitrotyrosine formation was injected at a dose of 50mg/kg i.p. 24h before the LPS injection and clodronate, a Kupffer cell remover was injected i.v. at a dose of 0.2ml (0.23M) 24h before the LPS injection. Nitrotyrosine was measured by enzyme-linked immunosorbent assay and high performance liquid chromatography-electrochemical detection methods. In the LPS group, the liver contained 1, 403±240ng nitrotyrosine/g protein (control: 21.2±1.7ng/g) and plasma ALT was elevated (1, 781±455IU/liter vs. control 55±5IU/liter). Nitrotyrosine formation was decreased in the LPS+quercetin and LPS+clodronate groups (240.6±47.6 and 301.4±26.2ng/g protein, respectively) and the increase of plasma ALT was attenuated (486±20.7 and 79.8±9.4IU/liter, respectively). The plasma TNF-α level was markedly elevated in the LPS group (1, 124.1±67.6pg/ml). It remained high in quercetin groups (1, 292.4±159.9pg/ml) but was very low in the clodronate group (34.8±5.3pg/ml). LPS-induced liver injury involves nitrotyrosine formation and treatments to decrease its formation attenuated the injury. The protection was observed without affecting TNF-α level; suggesting that the nitrotyrosine formation is a downstream mechanism for the injury following TNF-α production.