15d-PGJ2 Induces Apoptosis of MCF-7 and MDA-MB-231 Cells via Increased Intracellular Calcium and Activation of Caspases, Independent of ERα and ERβ.

Reports indicate that 15deoxydelta12,14prostaglandinJ2 (15dPGJ2) has anticancer activities, but its mechanisms of action have yet to be fully elucidated. We therefore investigated the effects of 15dPGJ2 on the human breast cancer cell lines, MCF7 (estrogen receptor ERα+/ERβ+) and MDAMB231 (ERα/ERβ+). Cellular proliferation and cytotoxicity were determined using the 3(4,5dimethylthiazol2yl)2,5diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays while apoptosis was determined by fluorescence microscopy and flow cytometry using annexin Vpropidium iodide (PI) staining. ER expression was determined by Western blotting. Intracellular calcium was stained with Fluo4 AM while intracellular caspase activities were detected with CaspaseFLICA® and measured by flow cytometry. We showed that 15dPGJ2 caused a significant increase in apoptosis in MCF7 and MDAMB231 cells. ERα protein expression was reduced in treated MCF7 cells but preincubation with the ERα inhibitor' ICI 182 780' did not affect the percentage of apoptotic cells. The expression of ERβ was unchanged in both cell lines. In addition, 15dPGJ2 increased intracellular calcium (Ca²+) staining and caspase 8, 9 and3/7 activities. We therefore conclude that 15dPGJ2 induces caspasedependent apoptosis that is associated with an influx of intracellular Ca²+ with no involvement of ER signaling.