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Our previous studies showed that silibinin promoted activation of caspases to induce apoptosis in human breast cancer MCF-7 cells by down-regulating the protein expression level of estrogen receptor (ER) α and up-regulating ERβ. Recently, it has been reported that silibinin-induced apoptosis also involved nuclear translocation of apoptosis-inducing factor (AIF). Here we report that silibinin induces nuclear translocation of AIF through the down-regulation of ERα and up-regulation of ERβ in a concentration dependent manner in MCF-7 cells. AIF knockdown with siRNA significantly reverses silibinin-induced apoptosis. The nuclear translocation of AIF is enhanced by treatment with MPP, an ERα antagonist, and blocked with PPT, an ERα agonist. In contrast to ERα activity, the nuclear AIF is increased with an ERβ agonist, DPN and blocked with an ERβ antagonist, PHTPP. Autophagy, negatively regulated by ERα, positively controls AIF-mediated apoptosis, as evidenced by the preventive effect of autophagy inhibitor 3-MA and siRNA targeting LC3, on the nuclear translocation of AIF and cell death induced by silibinin co-treatment with or without MPP. In sum we conclude that AIF in nuclei is involved in silibinin-induced apoptosis, and the nuclear translocation of AIF is increased by down-regulated ERα pathway and/or up-regulated ERβ pathway in MCF-7 cells, accompanying up-regulation of autophagy. Image 1 • Silibinin induces apoptosis-inducing factor-dependent apoptosis in MCF-7 cells. • Estrogen receptor α reduced by silibinin inhibits AIF nuclear translocation. • Estrogen receptor β increased by silibinin promotes AIF nuclear translocation. • Autophagy under the negative control of estrogen receptor α increases nuclear AIF. [ABSTRACT FROM AUTHOR]

We previously reported that silibinin induced a loss of cell viability in breast cancer (MCF-7) cells by ERα down-regulation. But whether this cytotoxicity depends on another estrogen receptor, ERβ, has yet to be elucidated. Therefore, we sought to explore the effects of ERβ modulation on cell viability by using an ERβ-selective agonist (Diarylprepionitrile, DPN) and an antagonist (PHTPP). Our data demonstrated that ERβ served as a growth suppressor in MCF-7 cells, and the incubation of silibinin, elevated ERβ expression, resulting in the tumor growth inhibition. The cytotoxic effect of silibinin was diminished by PHTPP and enhanced by DPN. Silencing of ERβ by siRNA confirmed these results. Apoptotic cascades, including the sequential activation of caspase-9 and -6, and finally the cleavage of caspase substrates, PARP and ICAD, caused by treatment with silibinin, were all repressed by PHTPP pre-treatment but exacerbated by DPN. Unlike ERα, ERβ did not involve autophagic process in the regulation, since neither autophagic inhibitor (3-MA) nor the inducer (rapamycin) affected the cell survival rates regardless ERβ activity. Taken together, silibinin induced apoptosis through mitochondrial pathway by up-regulating ERβ pathways in MCF-7 cells without the involvement of autophagy. [ABSTRACT FROM AUTHOR]