Dexamethasone-loaded Nanovesicles for Prostate Cancer Treatment

iPSC-MSCs were pre-incubated with 5 μg/mL Dxl (Sigma, PHR1883) for 24 h or not pretreated, and then broken down by serial extrusion in the presence of 50, 100, or 200 μg/mL Dxl to make Dxl-loaded NVs. The amount of Dxl loaded into NVs was determined by UV spectrometry at 230 nm as reported [25 (link)] using a spectrofluorometer (ThermoFisher). After incubation in 37 °C PBS containing 10% pooled human serum (Sigma) or 4 °C PBS for a series of periods, the supernatant was isolated by ultra-centrifugation at 100,000×g for 90 min at 4 °C using Sorvall WX Floor Ultra Centrifuge (Thermo) to measure Dxl release from NV-Dxl by UV spectrometry. Dxl-resistant PC3 cells were established as reported [26 (link)]. 5 × 10³/well parent PC3 cells, Dxl-resistant PC3 cells, or THP-1 human myeloid cells were seeded into 96-well plates, incubated with empty NVs, NV-Dxl, or free Dxl at a series of concentrations for 72 h, and then analyzed with PrestoBlue Cell Viability Assay (ThermoFisher).