Increasing the Fungicidal Action of Amphotericin B by Inhibiting the Nitric Oxide-Dependent Tolerance Pathway
AmB induced accumulation of superoxide and nitric oxide radicals and membrane permeabilization in S. cerevisiae. Yeast cultures were treated with different concentrations of AmB for 3 h and subjected to flow cytometry or plating assays. (a) Levels of superoxide radical detected by dihydroethidium (DHE) fluorescence and flow cytometry. (b) Levels of nitric oxide radical detected by 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate (DAF-FM DA) fluorescence and flow cytometry. (c) Membrane permeabilization events detected by propidium iodide (PI) fluorescence and flow cytometry. (d) Number of CFU/mL in Log-scale, assessed by plating assays and CFU counting. Means and standard errors of the means (SEM) of at least 3 independent biological experiments () are presented. Two-way ANOVA followed by Dunnett multiple comparison test was performed to analyse statistically significant differences in the number of PI-, DHE-, and DAF-FM DA-positive cells and cells able to proliferate between control treatment and treatment with different concentrations of AmB. and represent and , respectively.