Evaluating Senescent Lung Fibroblast Viability

Cell viability was evaluated using AlamarBlue Cell Viability Reagent (ThermoFisher Scientific). Senescent lung fibroblasts (3 × 10⁴ cells/well) were treated with nintedanib (Ofev, Boehringer Ingelheim, Germany; 300 nM), pirfenidone (Esbriet, Genentech, San Francisco; 2.5 mM) or dasatinib (Tocris, Bristol, UK; 20 μM) + quercetin (Sigma-Aldrich, St. Louis, MO; 15 μM) for 24 h followed by 3 h with the cell death ligand Super Fas Ligand (100 ng/ml). AlamarBlue Cell Viability Reagent was added to the cells and incubated for 4 h at 37 °C. Fluorescence values were assessed using a fluorescence excitation wavelength of 560 nm and an emission of 590 nm with a fluorescent microplate reader (Biotek). Results were expressed as fold-change compared to control cells.