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Fluoromicroscopic studies of bleomycin-induced intracellular oxidation in alveolar macrophages and its inhibition by taurine.
Bhat M; Rojanasakul Y; Weber SL; Ma JY; Castranova V; Banks DE; Ma JK
Environ Health Perspect 1994 Dec; 102(Suppl 10):91-96
A fluoromicroscopic method for determining bleomycin (11056067) induced intracellular oxidation in alveolar macrophages (AM) and the inhibition of the bleomycin effect by taurine was reported. Alveolar macrophages were collected from Sprague-Dawley-rats by bronchoalveolar lavage. Cell suspensions were prepared and incubated with 2',7'-dichlorofluorescin-diacetate (DCFHDA) for the quantitative evaluation of intracellular oxidation. Up to 187.5 micromolar (microM) taurine was added as pretreatment. Hydrogen- peroxide, A23187, or bleomycin were added after measurements of basal fluorescence. Cell viability was determined. Intracellular free calcium concentrations, oxygen consumption, and superoxide- anion release were measured. Bleomycin at 5 to 20 micrograms/milliliter had a moderate stimulatory effect on the secretion of superoxide-anion. At 20 micrograms/milliliter, bleomycin inhibited cellular response to zymosan induced secretion of superoxide-anion. Bleomycin at 4 micrograms/milliliter had no effect on cell membrane integrity or morphology, but caused a significant increase in intracellular oxidation. The oxidative process was found to be iron dependent and accompanied by an increase of up to 35 nanomolar intracellular calcium. Pretreatment of the cells with taurine inhibited both the intracellular oxidation and calcium rise. The inhibitory effect on intracellular oxidation was 36, 57, and 60%, respectively, for taurine at 25, 125, and 187.5microM concentrations. The authors conclude that a stimulation/inhibition relationship exists between bleomycin and taurine on cellular oxidation at subcytotoxic doses of bleomycin, and suggest that the oxidative effect of the intracellular bleomycin/iron complex is important in the initiation of the fibrotic process.
NIOSH-Publication; NIOSH-Author; NIOSH-Cooperative-Agreement; Analytical-methods; Laboratory-techniques; Lung-cells; Alveolar-cells; Cytotoxic-effects; Lung-irritants; Cell-damage; In-vitro-study; Oxidative-processes; Author Keywords: taurine; bleomycin; DCFHDA; intracellular oxidation; alveolar macrophages
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