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OBJECTIVE The synthetic triterpenoid 2-cyano-3,12-dioxoolean-1,9(11)-dien-C28-methyl ester(CDDO-Me)is considered a promising anti-tumorigenic compound.In this study,we investigated the anti-cancer effect of CDDO-Me on breast cancer cells and its underlying mechanisms.METHODS To investigate the effect of CDDO-Me on various breast cancer cells,cell viability assay using calcein-AM and EthD-1 as well as MTT assay was performed.To clarify the origin of CDDO-Me-induced vacuoles,electron microscopy as well as fluorescence microscopy using YFP-ER or YFP-Mito construct was performed.To measure the changes in intracellular Ca2+and ROS levels,flow cytometry using Fluo-3 and H2DCF-DA was performed.RESULTS CDDO-Me treatment induces progressive ER-derived vacuolation and subsequent apoptosis in various breast cancer cells.CDDO-Me-induced increases in intracellular Ca2+ levels,reflecting influx from the extracellular milieu,make a critical contribution to ER-derived vacuolation and subsequent cell death.In parallel with increasing 2+ Calevels,CDDO-Me markedly increases the generation of reactive oxygen species(ROS).Interestingly,we found that there exists a reciprocal positive-regulatory loop between Ca2+ influx and ROS generation that triggers ER stress and ER dilation in response to CDDO-Me.CONCLUSION ER-derived vacuolation via Ca2+ influx and ROS generation is responsible for the potent anticancer effects of CDDOMe on breast cancer cells.
OBJECTIVE The synthetic triterpenoid 2-cyano-3,12-dioxoolean-1,9 (11) -dien-C28-methyl ester (CDDO-Me) is considered a promising anti-tumorigenic compound. In this study, we investigated the anti- cancer effect of CDDO-Me on breast cancer cells and its underlying mechanisms. METHODS To investigate the effect of CDDO-Me on various breast cancer cells, cell viability assay using calcein-AM and Eth-1 as well as MTT assay performed. To clarify the origin of CDDO-Me-induced vacuoles, electron microscopy as well as fluorescence microscopy using YFP-ER or YFP-Mito construct was performed.To measure the changes in intracellular Ca2 + and ROS levels, flow cytometry using Fluo-3 and H2DCF -DA was performed .RESULTS CDDO-Me treatment induces progressive ER-derived vacuolation and subsequent apoptosis in various breast cancer cells. CDDO-Me-induced increases in intracellular Ca 2+ levels, reflecting influx from the extracellular milieu, make a critical contribution to ER- derived vacuolation and subsequent cell d eath. In parallel with increasing 2+ Calevels, CDDO-Me markedly increases the generation of reactive oxygen species (ROS) .Interestingly, we found that there exists a reciprocal positive-regulatory loop between Ca2 + influx and ROS generation that triggers ER stress and ER dilation in response to CDDO-Me. CONCLUSION ER-derived vacuolation via Ca2 + influx and ROS generation is responsible for the potent anticancer effects of CDDOMe on breast cancer cells.