Nitric oxide inhibits neonatal hepatocyte oxidative metabolism.
OBJECTIVE
Liver function is frequently impaired in neonates with sepsis. Nitric oxide (NO) is thought to be a mediator of organ dysfunction and liver oxidative metabolism during sepsis. The authors developed an in vitro model to investigate the effect of NO and the combined effect of NO plus H2O2 on neonatal hepatocyte oxidative metabolism.


METHODS
Hepatocytes were isolated from neonatal rats. Oxygen consumption was measured polarographically. In Study A, cells were exposed to S-Nitroso-N-acetylpenicillamine (SNAP), an NO donor, at various concentrations. In study B, myxothiazol and oligomycin, inhibitors of mitochondrial respiration, were added to investigate the site of action of NO. In study C, hepatocytes were incubated in the presence of both SNAP (300 micromol/L) and H2O2 (1.5 mmol/L). In study D, morphological alterations induced by NO and NO plus H2O2 were investigated by hepatocyte electron microscopy.


RESULTS
In study A, SNAP caused a dose-dependent decrease in oxygen consumption. A significant inhibition was reached at 300 micromol/L SNAP. In study B, the lack of further inhibition when SNAP was given together with myxothiazol indicates that NO acts intramitochondrially. Similarly, no further inhibition occurred when the NO donor was given together with oligomycin, suggesting that the effect of NO is mainly at the level of ATP synthase. In study C, concomitant addition of 300 micromol/L SNAP and 1.5 mmol/L H2O2 to hepatocytes caused further inhibition of oxygen consumption compared with either SNAP or H2O2 alone. In study D, mild alterations in hepatocyte morphology were noted in the presence of SNAP or SNAP plus H2O2.


CONCLUSIONS
In neonatal hepatocytes, NO significantly inhibits mitochondrial oxygen consumption, possibly at the level of ATP synthase. The effect of NO is additive to that of H2O2. Morphological findings were consistent with these biochemical effects and suggest that NO and H2O2 are important mediators of liver damage during sepsis.
Romeo
C
C
Institute of Child Health and Great Ormond Street Hospital for Children, London, England.
Eaton
S
S
Spitz
L
L
Pierro
A
A
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology; D066298:In Vitro Techniques; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D008689:Methacrylates; D008930:Mitochondria, Liver / Q000187:drug effects / Q000378:metabolism; D009569:Nitric Oxide / Q000494:pharmacology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D009840:Oligomycins / Q000494:pharmacology; D010084:Oxidation-Reduction / Q000187:drug effects; D010101:Oxygen Consumption / Q000187:drug effects*; D010396:Penicillamine / Q000031:analogs & derivatives / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D026423:S-Nitroso-N-Acetylpenicillamine; D013844:Thiazoles / Q000494:pharmacology
D004791:Enzyme Inhibitors; D008689:Methacrylates; D020030:Nitric Oxide Donors; D009840:Oligomycins; D013844:Thiazoles; D009569:Nitric Oxide; C030517:myxothiazol; D026423:S-Nitroso-N-Acetylpenicillamine; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D010396:Penicillamine