Hypoxia accelerates nitric oxide-dependent inhibition of mitochondrial complex I in activated macrophages.
Excess production of nitric oxide (NO) is implicated in the development of multiple organ failure, with a putative mechanism involving direct mitochondrial inhibition, predominantly affecting complex I. The persistent effects of NO on complex I may be mediated through S-nitrosylation and/or nitration. The temporal contribution of these chemical modifications to the inhibition of respiration and the influence of concurrent hypoxia have not been previously examined. We therefore addressed these questions using J774 macrophages activated by endotoxin and interferon-gamma over a 24-h period, incubated at 21% and 1% oxygen. Oxygen consumption and complex I activity fell progressively over time in the activated cells. This was largely prevented by coincubation with the nonspecific NO synthase inhibitor L-N5-(1-iminoethyl)-ornithine. Addition of glutathione ethyl ester reversed the inhibition at initial time points, suggesting an early mechanism involving nitrosylation. Thereafter, the inhibition of complex I became more persistent, coinciding with a progressive increase in mitochondrial nitration. Hypoxia accelerated the persistent inhibition of complex I, despite a reduction in the total amount of NO generated. Our results suggest that hypoxia amplified the mitochondrial inhibition induced by NO generated during inflammatory disease states.
Wolfson Institute for Biomedical Research, University College London, Gower St., London WC1E 6BT, UK.
Matthew T
MT
Frost
Qi
Q
Wang
Salvador
S
Moncada
Mervyn
M
Singer
D000818:Animals; D015687:Cell Hypoxia / Q000502:physiology*; D002460:Cell Line; D042967:Electron Transport Complex I / Q000502:physiology*; D008262:Macrophage Activation / Q000502:physiology*; D008264:Macrophages / Q000201:enzymology*; D008928:Mitochondria / Q000201:enzymology*; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D013997:Time Factors
D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D042967:Electron Transport Complex I; D010100:Oxygen