Tumour necrosis factor-alpha induces superoxide anion generation in mitochondria of L929 cells.
Within a few minutes after addition to L929 cells, tumour necrosis factor-alpha (TNF alpha) induced an increase in lucigenin-enhanced chemiluminescence that could be inhibited by superoxide dismutase. The generation of superoxide anion (O2.-) was sensitive to treatment with rotenone, antimycin A and cyanide, indicating that the signal originated from mitochondria. The mechanism of production of O2.- was shown to be independent of ATP synthesis, as uncoupling of this event from mitochondrial electron transport did not alter the generation of O2.- induced by TNF alpha. Chemiluminescence was further dependent on the presence of extracellular calcium, suggesting a role for this cation as a second messenger. This hypothesis was supported by the finding that inhibition of mitochondrial calcium uptake by Ruthenium Red exerted a protective effect on TNF alpha-treated L929 cells. Increased O2.- generation was followed by a marked decrease in mitochondrial dehydrogenase activity and cellular ATP levels, while cell membrane permeability was moderately increased. A role for mitochondrial O2.- generation in TNF alpha cytotoxicity was further supported by the finding that resistant L929 cells had decreased ability to produce O2.- in response to TNF alpha. In addition, we detected a decreased activity of the mitochondrial enzyme succinate dehydrogenase in these cells, suggesting that this component of the respiratory chain might be an important contributor to the TNF alpha-induced generation of O2.-.
Hennet
T
T
Institute of Veterinary Virology, University of Berne, Switzerland.
Richter
C
C
Peterhans
E
E
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D004140:Dinitrophenols / Q000494:pharmacology; D004579:Electron Transport / Q000187:drug effects; D007700:Kinetics; D007739:L Cells; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008163:Luminescent Measurements; D051379:Mice; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D010101:Oxygen Consumption / Q000187:drug effects; D011994:Recombinant Proteins / Q000494:pharmacology; D012430:Ruthenium Red / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D004140:Dinitrophenols; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D013481:Superoxides; D012430:Ruthenium Red; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase