Fig. 7.

RA treatment (differentiating conditions) protects differentiated cells from neuronal cell death. Human SH-SY5Y neuroblastoma cells were grown to 80% confluency onto cell culture Petri dishes in normal serum. Then, cells were subjected to medium replacement containing low serum and supplemented with 15 μM RA as described in “Materials and methods” section. Chronic RA treatment was performed by replacing the medium with fresh medium supplemented with 15 μM RA every 2 days. On the experimental day 5, cells were pretreated with 0.5 μM KN-62 for 20 min followed by 300 μM BzATP for 48 h. After treatments, trypan blue exclusion assay was performed as described in “Materials and methods” section. Non-RA treated cells (undifferentiated cells) and RA-treated cells (differentiated cells) without additional treatments served as controls. Dashed line represents cultured cells treated with 4 μM staurosporine for 2 h as positive control (n = 15; 18.2 ± 2.6). Data are the means ± SEM of values from at least three independent experiments. BzATP- and KN-62-treated cells were compared to non-RA treated cells (undifferentiated cells) and RA-treated cells (differentiated cells) without additional treatments (*** P < 0.001 one-way ANOVA)