Fig. 7: 1,25D inhibits gefitinib resistance in NSCLC cells.
A H1975 cells were seeded into 6-well plates (n = 3 per group). The next day, the cells were treated with vehicle (control), 1,25D alone (100 nM), EB (100 nM), gefitinib (1 μM), or with a combination of gefitinib and 1,25D or EB. Treatments were repeated every 3 days. Colony formation was assessed by crystal violet staining. Colony numbers were counted by using the ImageJ software and clonal formation efficiency was calculated (mean ± SD; n = 3; ***P < 0.001). B PC9/GR and H1975 cells were exposed to various concentrations of gefitinib and gefitinib + 1,25D (100 nM) for 2 days. Cell proliferation was determined by the CCK8 assay. The IC50 value against gefitinib was analyzed. Data are presented as mean ± SD (n = 3, *P < 0.05, **P < 0.01). C PC9/GR and H1975 cells were exposed to vehicle, 1,25D (100 nM), EB (100 nM), gefitinib (1 μM), 1,25D + gefitinib, or EB + gefitinib for 48 h and stained for Edu (Scale bar: 100 μm; original magnification: ×100; representative images from three experiments). Cell proliferation rate was calculated as a percentage of Edu-positive nuclei to total nuclei (mean ± SD; n = 3; *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001). D Horizontal migration of PC9/GR and H1975 cells (gap-closing assay) was evaluated after cells had been treated with the vehicle, 1,25D (100 nM), EB (100 nM), gefitinib (1 μM), 1,25D + gefitinib, or EB + gefitinib at the indicated time points, and the migration rate was calculated (mean ± SD; n = 3; #P < 0.05, ##P < 0.01 vs 1,25D or EB; *P < 0.05, **P < 0.01 vs GEF).
