[Production of reactive oxygen species by rat alveolar macrophages. Dissociation between the intracellular and extracellular release of hydrogen peroxide].
In order to clarify the features of reactive oxygen species produced by rat alveolar macrophages (AMs), the concentrations of intracellular and extracellular hydrogen peroxide were measured under various experimental conditions. Intracellular hydrogen peroxide was measured by DCFH method using a flow cytometer, while the extracellularly released fraction was measured by scopoletin method using a spectrophotometer. The concentration of intracellular hydrogen peroxide after stimulation with opsonized zymosan (10 micrograms/ml) was significantly higher than that after stimulation with phorbol myristate acetate (PMA; 100 ng/ml). On the other hand, hydrogen peroxide released extracellularly after stimulation with PMA was significantly greater that that after stimulation with opsonized zymosan. These results indicate that the soluble membrane stimulant and the phagocytic particles have different mechanisms in activating the production of hydrogen peroxide in AMs. That is, hydrogen peroxide induced by PMA was mainly released extracellularly, while that induced by zymosan was mainly released into the intracellular environment. At rest, the concentration of intracellular hydrogen peroxide in rat AMs was high. Potassium cyanate, a known mitochondrial inhibitor, suppressed the intracellular hydrogen peroxide in AMs not only at rest but also after stimulations, indicating that most of the reactive oxygen species released into the intracellular environment in AM are produced by mitochondria. From these results, in order to gain a closer insight into the function of AMs, it is very important to distinguish the oxidative metabolites produced intracellularly which are related to bactericidal function from those of the extracellularly released fraction which give rise to lung damage.
Uwabe
Y
Y
Third Department of Internal Medicine, National Defense Medical College, Saitama, Japan.
Katsura
Y
Y
Noritake
M
M
Kiryu
T
T
Shinomiya
N
N
Kayashima
S
S
Wakiyama
H
H
Nagata
N
N
Tsuru
S
S
D000818:Animals; D002478:Cells, Cultured; D003485:Cyanates / Q000494:pharmacology; D005609:Free Radicals; D006861:Hydrogen Peroxide / Q000378:metabolism*; D016676:Macrophages, Alveolar / Q000378:metabolism*; D008297:Male; D008928:Mitochondria / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D015054:Zymosan / Q000494:pharmacology
D003485:Cyanates; D005609:Free Radicals; D015054:Zymosan; D006861:Hydrogen Peroxide; C024835:potassium cyanate; D013755:Tetradecanoylphorbol Acetate