Fig 7. Sulfatase pretreatment of MDCK cells significantly prevents the cytotoxic effect of Etx but does not affect toxin oligomerization.

(A) MDCK cells were preincubated in the absence or the presence (0.1 or 0.2 U) of sulfatase from Aerobacter aerogenes for 90 minutes at 37°C. Etx was then added at a final concentration of 2 or 5 nM. After 30 minutes at 37°C, MTS assay was used to determine cell viability. Results represent mean ± SEM of cell survival compared to controls (0 U), from three experiments. ***p < 0.001 for 5 nM Etx and ###p < 0.001 for 2 nM Etx, when compared with cells without sulfatase treatment, using one-way ANOVA, followed by the Bonferroni’s post-hoc test. (B) MDCK cells preincubated in absence or presence of 2U/ml of sulfatase were incubated at 37°C with 10 or 30 nM Etx for 30 min. After washing, cells were lysated and the presence of Etx monomers and oligomers analyzed by western blot. The figure shows a representative western blot of three independent experiments.