Endotoxin (lipopolysaccharide)-induced nitric oxide production in 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated Fischer rats: detection of nitrosyl hemoproteins by EPR spectroscopy.

Electron paramagnetic resonance (EPR) spectroscopy was used to study the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on endotoxin (lipopolysaccharide)-induced nitric oxide (NO) production in Fischer rats. We found that rats treated with 50 mug/kg TCDD had increased sensitivity to endotoxin, resulting in an approximately 2-fold increase in the level of NO production detected as nitrosylhemoglobin (HbNO) in venous blood. At lower concentrations (less than or equal to5 mug/kg), TCDD did not affect the endotoxin-induced NO production. The TNF-alpha serum concentration was found to parallel that of NO. TCDD alone did not induce the production of detectable HbNO or TNF-alpha. We found that TCDD induced a dose-dependent increase in the EPR signal intensity of (Fe3+) low-spin methemoprotein complexes found in the liver and kidney. These species with EPR resonance at g = 2.43, 2.26, and 1.92 are attributed to low-spin Fe3+ in cytochromes P450 and P420. Our data confirm previous studies that have shown that TCDD induces a dose-dependent increase in the production of some cytochrome P450 enzymes. However, in rats that were subsequently challenged with endotoxin, a smaller increase in the EPR intensity of these species was observed. The decrease in the low-spin Fe3+ cytochrome P450 EPR signal in endotoxin-challenged rats could be due to one or more of the following occurring: (1) cytochrome destruction, (2) reduction of the ferric to the ESR-silent ferrous oxidation state of cytochromes by nitric oxide, and/or (3) formation of ferrous nitrosyl cytochrome complexes that contribute, in part, to the characteristic five-coordinate nitrosyl hemoprotein triplet also observed in these tissues. Since low concentrations of endotoxin can leak from the gut lumen into the systemic circulation, this investigation explores the possibility that endotoxin interaction with TCDD may be, in part, responsible for the effects of TCDD observed in these tissues.