Nitric-oxide-induced inhibition of glyceraldehyde-3-phosphate dehydrogenase may mediate reduced endothelial cell monolayer integrity in an in vitro model blood-brain barrier.
The process of nitric-oxide (NO)-induced cellular toxicity may involve energy deprivation since the radical is reported to prevent both mitochondrial oxidative phosphorylation and glycolysis. In order to determine whether these processes are important in NO-induced blood-brain barrier (BBB) dysfunction, we used a cell culture model of the BBB and compared the effects of gaseous NO, potassium cyanide (KCN, a mitochondrial respiratory chain inhibitor) and iodoacetate [IA, an inhibitor of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH)] on endothelial cell ATP content, GAPDH activity and barrier integrity. NO lead to a rapid breakdown in model barrier integrity and resulted in a reduction in endothelial cell ATP content and GAPDH activity. KCN had no effect on endothelial cell ATP content or barrier integrity, while IA, at a concentration that completely blocked endothelial cell GAPDH activity, resulted in a rapid decline in ATP content but did not lead to a decline in barrier integrity until at least 2 h of exposure. These results indicate that inhibition of endothelial cell GAPDH activity rather than mitochondrial respiration causes an energy deficiency and delayed barrier dysfunction. However, the rapid detrimental effects of gaseous NO on barrier integrity cannot be fully explained by endothelial cell energy depletion and may be related to the actions of the free radical and its products on cellular lipids.
Hurst
R D
RD
Centre for Research in Biomedicine, Faculty of Applied Sciences, University of the West of England, Bristol, UK. roger.hurst@uwe.ac.uk
Azam
S
S
Hurst
A
A
Clark
J B
JB
D000255:Adenosine Triphosphate / Q000378:metabolism; D001812:Blood-Brain Barrier / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D017097:Electric Impedance; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005609:Free Radicals / Q000378:metabolism; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007461:Iodoacetates / Q000494:pharmacology; D015227:Lipid Peroxidation / Q000502:physiology; D009569:Nitric Oxide / Q000378:metabolism*; D010085:Oxidative Phosphorylation / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology; D011190:Potassium Cyanide / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology
D004791:Enzyme Inhibitors; D005609:Free Radicals; D007461:Iodoacetates; D013481:Superoxides; D009569:Nitric Oxide; D000255:Adenosine Triphosphate; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D011190:Potassium Cyanide