Figure 2.

Exposure to α-toxin triggers release of eATP that is inhibited by IFNα-pretreatment. A549 cells were treated with the indicated concentrations of α-toxin (A) or diluted conditioned medium from cultures of Hla⁺ or Hla^- S. aureus (B). The curves show normalized mean luminescence intensity kinetics measured every minute within 1 h after α-toxin. Data in (A) are mean of quadruplicate cultures and are representative of two independent experiments. Data in (B) are mean±SEM of triplicate cultures and are representative of three independent experiments. (C): SAEC were pretreated with IFNα for 24 h and exposed to 0.1 μg/ml α-toxin for 30 min. Cell-free conditioned medium was used to measure eATP. The data are mean±SD of quadruplicate cultures and are representative of three independent experiments. (D): Exogenous ATP was added to medium- or IFNα-pretreated A549 cells for 30 min. Cell-free conditioned medium was used to measure remaining ATP (% input, the data are mean±SD of quadruplicate cultures and are representative of three experiments). (E): A549 cells were pretreated with oxidized ATP for 2 h prior to exposure to 2.5 μg/ml α-toxin. Cell death at 24 h after α-toxin is shown. The data are mean±SD of quadruplicate cultures and are representative of three independent experiments. (F): C57BL6/ mice (8-wk old, females) were administered α-toxin or α-toxin with oxidized ATP diluted in 50 μl sterile PBS via intranasal route. Body temperature was measured at the indicated time points. The data are mean±SD, n=5. Control mice received 50 μl sterile PBS. Asterisks indicate the time points when the body temperature in mice treated with α-toxin alone was significantly lower than in other groups of mice (p<0.05).