Exogenous adenine nucleotides replete endothelial cell adenosine triphosphate after oxidant injury by adenosine...

We studied the ability of human umbilical vein endothelial cells to recover from oxidant-induced ATP depletion. When endothelial cell ATP levels were depressed to 0.93 +/- 0.14 pmol/micrograms protein (compared with 4.96 +/- 0.6 pmol/micrograms protein in control cells) by hydrogen peroxide generated with 25 mU/ml glucose-glucose oxidase over 45 minutes, ATP levels returned to 1.73 +/- 0.21 pmol/micrograms protein during a 3-hour recovery period after oxidant injury ceased. When 25 microM ATP, ADP, AMP, or adenosine was added to the recovery media, intracellular ATP was significantly (p less than 0.001) increased to greater than 4.4 pmol/micrograms cell protein for each metabolite. HPLC of supernatants from oxidant-injured endothelial cells incubated with ATP, ADP, and AMP demonstrated extracellular metabolism of the adenine nucleotides to adenosine. When adenosine transport was inhibited with dipyridamole and nitrobenzylthioinosine, recovery of intracellular ATP by exogenous ATP, ADP, AMP, and adenosine was significantly (p less than 0.001) inhibited. Such cells were intact, as demonstrated by lack of LDH release. When oxidant stress was prolonged to 90 minutes, ATP depletion was irreversible, regardless of exogenously supplied adenosine; such cells demonstrated loss of cell integrity as demonstrated by release of intracellular LDH. Our results demonstrated that exogenous adenine nucleotides enhance recovery of oxidant-induced ATP depletion through metabolism to adenosine and subsequent adenosine uptake. Prolonged oxidant injury resulted in irreversible ATP depletion and loss of cell integrity that was not altered by exogenously supplied adenosine.