Figure 4. p53 deficiency is required for NF-κB-driven transcription induced by doxorubicin.

A, B, graph shows the relative levels of ICAM-1, TNFAIP-3 and CXCL-1 obtained by qRT-PCR (a) and ICAM-1 determined by IHC (b) taking into account all tumors in which p53 has been considered as WT or deficient status by immunohistochemistry. C, MEFs WT and p53-/- were preincubated 1h 30 min with MLN120B 205µM and exposed to 5µM doxorubicin for 4hours. qRT-PCR analysis of ICAM-1 was performed and normalized to the RPLP0 expression in each condition. p53 expression in MEFs was determined by WB, tubulin expression was used as loading control. D, MCF-7 were infected with 3 different lentiviral particles coding for; shCT and 2 shRNA against TP53 (shTP53.1 and shTP53.2); qRT-PCR of TP53 relative to RPLP0 and Western Blot determination of p53 in cell lysates from cells exposed to 5µM doxorubicin for 24h was were used as knocking down controls. E, MCF-7 shCT and shTP53 were exposed for 4hours to doxorubicin 5µM. Relative expression levels of ICAM-1 and TNFAIP3 were determined by qRT-PCR in each condition. RPLP0 expression was used for normalization. F, MDA-MB-231 were transfected with pCDNA3CT vector and pCDNA3fagp53. RNA was extracted from both transfected cell lines and RT-PCR was done to detect fag p53 expression. RPLP0 was used as loading control. G, qRT-PCR analysis of ICAM-1, CXCL-1 and TNFAIP3 was performed in MDA-MB-231pcDNA3CT and pCDNA3 fagp53 after 4 hours of 5µM doxorubicin treatment. The relative target gene expression level was also normalized to the RPLP0 expression in each condition. The graph shows the results of the expression of the different genes in each condition relative to their expression in control condition. Mean and SE from triplicate experiments are indicated.