Cellular uptake

THP-1 cells were cultured as described above and seeded at 1 × 10⁴ cells/well/0.5 mL in 8-well chamber slides (Greiner Bio-One; Kremsmünster, Austria) containing 25 mg/mL PMA. After 72 h, the cells were incubated for 2 h, which dissolved either NP or MP samples at a final BTZ043 concentration of 42 µg/mL in the respective formulation. Treated cells were then washed twice with PBS and fixed with 4% paraformaldehyde (PFA) in PBS for 10 min at RT. For actin staining, cells were incubated for 30 min with 100 µL of 1:1000 Alexa Fluor 488-Phalloidin stock solution (Thermo Fisher Scientific, Waltham, MA, USA) after permeabilization and were blocked for 20 min with 1% w/v BSA and 0.1% w/v saponin in PBS. Nuclei were counterstained with 200 µL of a 1-µg/mL 4′,6-diamidino-2-phenylindole (DAPI) solution (Sigma-Aldrich) for 15 min. Images were taken with a Leica DMi8 Confocal Microscope equipped with a × 63 water immersion objective (HC APO CS2 63 × /1.20), and image analysis was performed with LAS X software (Leica Application Suite X).