Nitration of Bax and Bcl-2 proteins during hypoxia in cerebral cortex of newborn piglets and the effect of nitric oxide synthase inhibition.
Previous studies have shown that cerebral tissue hypoxia results in increased expression of Bax protein, thereby altering the ratio of Bax to Bcl-2 or formation of Bax/Bcl-2 heterodimer. Hypoxia also induces the generation of nitric oxide free radicals in the cerebral cortex of newborn animals. The present study tests the hypothesis that tissue hypoxia will result in nitration of Bax and Bcl-2 proteins in the neuronal nuclei of newborn piglets. Studies were performed in 22 piglets, 3-5 days old, divided into normoxic (n = 7), hypoxic (n = 9) and hypoxic + NNLA (n = 6) groups. Hypoxia was induced by decreasing the FiO(2) (5-7%) for 60 min and cerebral hypoxia documented by determining tissue ATP and phosphocreatine (PCr) levels. The density of protein bands was expressed as absorbance (OD x mm(2)). PCr levels were 3.03 +/- 0.85 micromol/g brain in the normoxic group and 0.88 +/- 0.32 micromol/g brain in the hypoxic group (p < 0.001 vs. normoxia) and 0.55 +/- 0.13 (p < 0.001 vs. normoxia) in the NNLA-treated hypoxic group. There was increased nitration of Bax protein in hypoxic neuronal nuclei as compared to normoxic and NNLA-treated-hypoxic group nuclei: 211.61 +/- 25.93 versus 124.8 +/- 14.88 and 133.86 +/- 7.42 OD x mm(2), respectively (p < 0.001 vs. normoxia). Nitration of Bcl-2 was not altered significantly in either group. We conclude that there is increased nitric oxide-mediated nitration of Bax in cortical neuronal nuclei during hypoxia and that this increase correlates inversely with the decrease in tissue energy levels. We speculate that, during hypoxia, nitration of Bax and Bcl-2 proteins may regulate heterodimer formation and activation of programmed cell death mechanisms.
Ashraf
Qazi M
QM
Department of Pediatrics, MCP Hahnemann University, Room #701, 3300 Henry Ave., Philadelphia, PA 19129, USA. qazi@drexel.edu
Zubrow
Alan B
AB
Mishra
Om P
OP
Delivoria-Papadopoulos
Maria
M
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn / Q000378:metabolism*; D002540:Cerebral Cortex / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D002534:Hypoxia, Brain / Q000378:metabolism*; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors*; D009584:Nitrogen / Q000378:metabolism*; D010725:Phosphocreatine / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D013552:Swine; D051028:bcl-2-Associated X Protein
D004791:Enzyme Inhibitors; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D010725:Phosphocreatine; D000255:Adenosine Triphosphate; D019001:Nitric Oxide Synthase; D009584:Nitrogen; D019331:NG-Nitroarginine Methyl Ester