Figure 2.
Vacuolin-1 does not inhibit the secretion of β-hexosaminidase induced by ionomycin. NRK (A) or HeLa (B) cells that were treated or not with 5 μM vacuolin-1 for 1 h were exposed or not to 5 μM ionomycin for 10 min, followed by collection of the supernatant and assay for β-hexosaminidase activity. The data are expressed as a percentage of the total cellular content of the enzyme, and correspond to the average of triplicates ±s.d. (C–E) Monolayers of HeLa cells were treated or not with 5 μM vacuolin-1 for 1 h and stimulated or not with 5 μM ionomycin for 10 min. β-Hexosaminidase activity was assayed in the supernatant collected after 10 min (C), in the 1 min wash (D) and in the subsequent 10 min incubation in fresh media (E). The data are expressed as a percentage of the total cellular content of the enzyme, and correspond to the average of triplicates ±s.d. (F) Immunofluorescence with anti-Lamp-1 antibodies of the cells used for the experiment shown in (C–E), before addition of ionomycin. The arrows point to the enlarged Lamp-1-positive vesicles induced by vacuolin-1. DMSO, dimethylsulphoxide.