Azithromycin, a lysosomotropic antibiotic, impairs fluid-phase pinocytosisin cultured fibroblasts

The dicationic macrolide antibiotic azithromycin inhibits the uptake of hor seradish peroxidase (HRP) by fluid-phase pinocytosis in fibroblasts in a ti me- and concentration-dependent fashion without affecting its decay (regurg itation and/or degradation). The azithromycin effect is additive to that of nocodazole, known to impair endocytic uptake and transport of solutes alon g the endocytic pathway. Cytochemistry (light and electron microscopy) show s a major reduction by azithromycin in the number of HRP-labeled endocytic vesicles at 5 min (endosomes) and 2 h (lysosomes). Within 3 h of exposure, azithromycin also causes the appearance of large and light-lucent/electron- lucent vacuoles, most of which can be labeled by lucifer yellow when this t racer is added to culture prior to azithromycin exposure. Three days of tre atment with azithromycin result in the accumulation of very large vesicles filled with pleiomorphic content, consistent with phospholipidosis. These v esicles are accessible to fluorescein-labeled bovine serum albumin (FITC-BS A) and intensively stained with filipin, indicating a mixed storage with ch olesterol. The impairment of HRP pinocytosis directly correlates with the a mount of azithromycin accumulated by the cells, but not with the phospholip idosis induced by the drug. The proton ionophore monensin, which completely suppresses azithromycin accumulation, also prevents inhibition of HRP upta ke. Erythromycylamine, another dicationic macrolide, also inhibits HRP pino cytosis in direct correlation with its cellular accumulation and is as pote nt as azithromycin at equimolar cellular concentrations. We suggest that di cationic macrolides inhibit fluid-phase pinocytosis by impairing the format ion of pinocytic vacuoles and endosomes.