Abstract
Purpose
To examine the effects of dantrolene on nitric oxide (NO) production and on the activity and protein expression of inducible nitric oxide synthase (INOS) induced by lipopolysaccharide (LPS) plus interferon-γ (IFNγ) in rat alveolar macrophages.
Methods
Pulmonary alveolar macrophages isolated from Sprague-Dawley rats were used. After incubation of macrophages with dantrolene (1 to 100 μM) and LPS (1 μg·mml−1) and IFN-γ ( 100 u·ml−1) for 24 hr, the cell-free medium was removed for measuring the nitrite and tumour necrosis factor-α (TNF-α) levels by Griess reaction and ELISA kit, respectively. The harvested macrophages were also used to determine the activity of iNOS by using the conversion of [3H]-L-arginine to [3H]-L-citrulline method. Protein expression of iNOS was detected by Western blot analysis.
Results
In rats alveolar macrophages, (1) dantrolene (1 to 100 μM) caused a dose-dependent suppression of the production of nitrite and TNF-α induced by LPS (1 μg·ml−1) plus IFN-γ (100 u·ml−1) and (ii) dantrolene (100 μM) inhibited the activity (by 37 ± 5%,P < 0.01 ) and protein expression (by 39 ± 12%,P < 0.01 ) of iNOS in response to LPS plus IFN-γ.
Conclusion
Dantrolene inhibits NO production as well as the activity and expression of iNOS in alveolar macrophages treated with LPS plus IFN-γ, which may be associated with the reduction of TNF-α production.
Résumé
Objectif
Évaluer les effets du dantrolène sur la production d’oxyde nitrique (NO) et sur l’activité et l’expression protéique de l’enzyme inductible synthétase de l’oxyde nitrique (iNOS) tel qu’induit par une lipopolysaccharide (LPS) associée à l’interféron-γ (IFN-γ) chez des macrophages alvéolaires de rat.
Méthodes
Des macrophages alvéolaires pulmonaires isolés chez des rats Sprague-Dawley ont été utilisés. Après incubation durant 24h des macrophages avec du dantrolène (1–100·μm), de la LPS (1 μg·ml−1) et de l’IFN-γ (1 μ·ml−1), le liquide d’incubation a été retiré pour mesurer les nitrites et le facteur de nécrose tumorale-α (TNF-α) par la réaction de Griess et un kit Elisa, respectivement. Les macrophages récoltés après incubation ont aussi été utilisés pour déterminer l’activité de iNOS par la méthode utilisant la conversion de la3H-L-arginine en3H-L-citrulline. L’expression protéique de iNOS a été détectée par l’analyse Western blot.
Résultats
Chez des macrophages alvéolaires de rat, le dantrolène (1–100 μm) cause une suppression proportionnelle à la dose de la production des nitrites et du TNF-α induit par LPS plus IFN-γ; le dantrolène (100 μm) inhibe aussi l’activité (par 37 ± 5%,P< 0,0l) et l’expression protéique (par 39 ± 12%,P< 0,0l) de iNOS ’ en réponse à LPS plus IFN-γ.
Conclusion
Le dantrolène inhibe la production de NO ainsi que l’activité et l’expression de iNOS chez des macrophages alvéolaires traités avec LPS associé à IFN-γ, et ceci pourrait être associé à la réduction de la production de TNF-α.
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Supported by a grant of the Tri-service General Hospital, Taipei, Taiwan, R.O.C. (TSGH-C87-49).
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Li, CY., Chou, T.C., Wu, C.C. et al. Dantrolene inhibits nitric oxide synthase in rat alveolar macrophages treated with lipopolysaccharide and interferon-γ. Can J Anaesth 45, 246–252 (1998). https://doi.org/10.1007/BF03012910
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DOI: https://doi.org/10.1007/BF03012910