Journal Article
Research Support, Non-U.S. Gov't
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Autophagy inhibition enhances sulforaphane-induced apoptosis in human breast cancer cells.

Anticancer Research 2010 September
AIM: Sulforaphane (SFN), which is present in cruciferous vegetables, induces growth arrest and/or cell death in cancer of various organs. The involvement of autophagy in the SFN-induced apoptotic death of human breast cancer cells was investigated.

MATERIALS AND METHODS: Cell proliferation and viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and trypan blue exclusion assay. Flow cytometry, immunofluorescence, electron microscopy, and Western blot analysis were used for detection of apoptosis and autophagy, and the role of autophagy was assessed using autophagy inhibitors.

RESULTS: SFN dose- and time-dependently retarded the growth and induced cell death in MCF-7 and MDA-MB-231 human breast cancer cells. In MDA-MB-231 cells, 30 μM SFN caused S and G2/M cell-cycle arrest associated with increased p21WAF1 and p27KIP1 levels and decreased cyclin A, cyclin B1 and CDC2 levels. Cell death was due to apoptosis with increased caspase-3 and lowered BCL-2 levels. In addition, the SFN-treated cells exhibited autophagy, as characterized by the appearance of autophagic vacuoles by electron microscopy, the accumulation of acidic vesicular organelles by flow cytometry, and the punctuate patterns of microtubule-associated protein 1 light chain 3 (LC3) by fluorescein microscopy. The levels of LC3-I and -II proteins (processed forms of LC3-I) and LC3 mRNA were increased. Treatment with autophagy inhibitor bafilomycin A1 (but not 3-methyladenine) with SFN significantly enhanced apoptosis, which was accompanied by increases in the level of BAX and the cleavage of caspase-3 and poly(ADP-ribose)polymerase (PARP)-1 and decreases in the mitochondrial membrane potential (ΔΨm).

CONCLUSION: These results indicate a cytoprotective role of autophagy against SFN-induced apoptosis and that the combination of SFN treatment with autophagy inhibition may be a promising strategy for breast cancer control.

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