Fig. 8
Etoposide induced p53 activity is dampened in chronic hypoxia. D283-MED cells were incubated in 1% O2 or 21% O2 for 1 day or 5 days, prior to etoposide treatment where indicated. Three p53 target genes, MDM2, PUMA and p21 were assessed by qPCR. (a) Basal levels of p53 target genes in hypoxia without etoposide treatment were measured. The mRNA levels were normalised with the house-keeping gene cyclophilin A. (b) Levels of MDM2, p21 and PUMA mRNA with or without etoposide treatment. Data represented as normalised to housekeeping gene (cyclophilin A) and fold change with respect to the untreated control. Data are representative of three independent experiments, error bars are SD of a single experiment. (c) Total p53 and phosphorylated p53 serine 15 levels assessed by western blot. (d) Densitometry quantification of the band intensity was analysed by ImageJ from 3 independent experiments. Plot represents p53 serine 15 over the p53 total