*4.3. Cell Handling*

A549 [40] and THP-1 [41] cells were grown in DMEM (1X) and RPMI-1640 medium (1X) (Gibco, Paisley, UK), respectively, and were both supplemented with FBS (10%) and penicillin-streptomycin (1%) (Gibco, Grand Island, NY, USA) at 37 ◦C in a 5% CO<sup>2</sup> humidified atmosphere. Sub-cultures were performed according to the manufacturer's instructions. To activate THP-1 in macrophage-like cells (A-THP-1), cells were resuspended in fresh media, and phorbol 12-myristate 13-acetate (PMA) was added (final concentration 200 µg/L) to the THP-1-containing medium [65]. A549 and THP-1 cells were then seeded in culture microplates, dishes or flasks depending on the experiment and incubated until sub-confluent (A549, ~80%) or 24 h (THP-1) at 37 ◦C in a 5% CO<sup>2</sup> humidified atmosphere. For experiments, test molecules were dissolved in 1% FBS-supplemented medium, unless otherwise mentioned. When prior cell cholesterol or sphingomyelin depletion was required, cells were preincubated for 30 min with 5 mM methyl-β-cyclodextrin (MβCD) or 50 mU/mL of sphingomyelinase from *Bacillus cereus* [66] in 1% FBS-supplemented medium, respectively.
