Figure 5.

VRAC levels affect cisplatin sensitivity also in naturally occurring cisplatin-selected cells. (a) Scheme to illustrate the establishment of cisplatin-resistant Pica_C cells by chronic exposure to cisplatin. (b,c) 2D/3D Pica_C cell models to demonstrate cisplatin resistance. Cells (b) or spheroids (c) were drug treated (cells—48 h; spheroids—72 h), and viability was normalized to untreated controls. (d) Resistant Pica_C cells show a lower number of cisplatin-induced DNA damage events (γH₂AX foci) per cell as automatically quantified by high-throughput microscopy. γH₂AX foci stained by specific fluorescent antibodies. (e) Immunoblot analysis confers downregulation of VRAC (LRRC8A) protein levels in resistant Pica_C cells. GAPDH served as loading control. Data from one representative experiment of three independent experiments shown. For immunoblot analysis of Pica_KO36 cells, refer to Figure 2b. For uncropped blots, refer to Supplementary Materials.