Fig 4. Effect of isotretinoin on polyamine transport and parasites growth.

(A) Putrescine transport was measured using epimastigote cultures (10⁷ parasites) and 100 μM [³H]-putrescine, as indicated under Methods, in presence of isotretinoin 0–100 μM (plotted in log scale). Transport values are given as the percentage of the transport rate in untreated parasites (11.7 pmol.min^-1). The IC₅₀ value was calculated by non-linear regression using the GraphPad Prism software. (B) CHO-K1 cells were infected with trypomastigote forms (in a proportion 1:50) for 4 h. Infected cells were treated with different concentrations of isotretinoin between 0 and 30 μM for 24 h. After treatment, plates were incubated at 33°C and trypomastigotes were collected from the extracellular medium on the sixth day. Parasite density is expressed as the percentage of untreated parasites (1.23 x10⁶ cells.mL^-1). (C) 10⁷ exponential phase epimastigotes were treated with different concentrations of isotretinoin between 0 and 300 μM for 72 h. Cell density is expressed as the percentage of untreated parasites (6.6x10⁷ cells.mL^-1). All the IC50 calculations were carried as described in “Trypanocidal activity assays” under the Methods section.