Figure 1

Differentiation induction and cells treated with the differentiation-inducing agent NaBu. (a) MCF-7 cells were grown on glass cover slips and treated with increasing amounts of sodium butyrate (NaBu), as indicated, for 72 hours. Morphological changes were evaluated by fluorescent microscopy (×63). Induction of biochemical differentiation was determined by using Nile Red stain for lipid vesicles in cells (red). Nuclear DNA was stained with 4',6-diamidino-2-phenylindole (DAPI) (blue). (b) Quantitative analysis of lipid induction. MCF-7 cells, as above, were stained with Nile Red and analyzed by flow cytometry as described in the Materials and methods. Standard deviations are shown as error bars (n = 6). (c) Quantitative (Q) PCR of ß-casein mRNA expression following treatment with increasing doses of NaBu. The induction of ß-casein is normalized against that seen in untreated cells. (d) Q-PCR of MYB mRNA from MCF-7. MCF-7 cells treated with the increasing dosage of NaBu show a dose dependent decrease in expression of MYB mRNA. Data from (c) and (d) represent mean values (n = 6), and standard deviations are shown as error bars. All Q-PCR data were normalized against cyclophilin A controls. (e) Western blot analysis of MYB from total cell lysates as above. A ß-tubulin loading control is also shown. Results are indicative of two independent experiments.