Post‐transcriptional regulation of inducible nitric oxide synthase mRNA in murine macrophages by doxycycline and chemically modified tetracyclines

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Abstract
Chemically modified tetracyclines [CMT‐3 (IC50 ∼6–13 μM = ∼2.5–5 μg/ml) and CMT‐8 (IC50 ∼26 μM = 10 μg/ml), but not CMT‐1, ‐2 or ‐5], which lack anti‐microbial activity, inhibited nitrite production in LPS‐stimulated macrophages. Unlike competitive inhibitors of l‐arginine which inhibited the specific activity of inducible nitric oxide synthase (iNOS) in cell‐free extracts, CMTs exerted no such direct effect on the enzyme. CMTs could, however, be shown to inhibit both iNOS mRNA accumulation and protein expression in LPS‐stimulated cells. Tetracyclines (doxycycline and CMT‐3) unlike hydrocortisone had no significant effect on murine macrophages transfected with iNOS promoter (tagged to a luciferase reporter gene) in the presence of LPS. However, doxycycline and CMT‐3 augmented iNOS mRNA degradation, in LPS‐stimulated murine macrophages. These studies show a novel mechanism of action of tetracyclines which harbours properties to increase iNOS mRNA degradation and decrease iNOS protein expression and nitric oxide production in macrophages. This property of tetracyclines may have beneficial effects in the treatment of various diseases where excess nitric oxide has been implicated in the pathophysiology of these diseases.