Figure 8.

nSMase2 mediates growth arrest in response to doxorubicin. (a) MCF7 cells were seeded in 60 mm dishes and transfected either with control (pcDNA), WT p53 plasmid or different p53 mutants, *P<0.05 and **P<0.01. Cells were lysed either for protein extraction or RNA isolation. RNA was reverse transcribed to cDNA, and nSMase2 expression was determined by qRT-PCR. Also, protein extraction was performed and samples were immunoblotted for p53 (b), MCF7 cells were seeded in 60 mm dishes and siRNA knockdown was performed using AS or nSMase2 for 24 h. After that, vehicle or doxorubicin were added and cells were collected, stained with Trypan Blue and counted (c), MCF7 cells were seeded in 60 mm dishes and siRNA knockdown was performed using AS or nSMase2 for 22 h. A pulse of BrdU was given to cells for 30 min and they were collected and analyzed by flow cytometry