Silibinin induces apoptosis via calpain-dependent AIF nuclear translocation in U87MG human glioma cell death

U87MG cells were grown on well tissue culture plates and cell viability was measured by MTT assay. ROS generation and △ψm were estimated using the fluorescence dyes. PKC activation and Bax expression were measured by Western blot analysis. AIF nuclear translocation was determined by Western blot and immunocytochemistry.Silibinin induced activation of calpain, which was blocked by EGTA and the calpain inhibitor Z-Leu-Leu-CHO. Silibinin caused ROS generation and its effect was inhibited by calpain inhibitor, the general PKC inhibitor GF 109203X, the specific PKCδ inhibitor rottlerin, and catalase. Silibinin-induce cell death was blocked by calpain inhibitor and PKC inhibitors. Silibinin-induced PKCδ activation and disruption of △ψm were prevented by the calpain inhibitor. Silibinin induced AIF nuclear translocation and its effect was prevented by calpain inhibitor. Transfection of vector expressing microRNA of AIF prevented the silibinin-induced cell death.Silibinin induces apoptotic cell death through a calpain-dependent mechanism involving PKC, ROS, and AIF nuclear translocation in U87MG human glioma cells.Glioblastoma is the most lethal and frequent primary brain tumors [1]. It is comprised of poorly differentiated heterogeneous neoplastic astrocytes with aggressive proliferation and highly invasive properties. After diagnosis of glioblastoma, the median survival time of 9-12 months has remained unchanged despite aggressive treatment including surgery, radiation, and chemotherapy [2,3]. Thus, new effective strategies for controlling glioblastoma are required. Because glioblastoma cells avoid differentiation and apoptosis, the induction of differentiation and apoptosis in glioblastoma cells may be considered as a potential treatment strategy.Silibinin, a natural polyphenolic flavonoid, is a major bioactive component of silymarin which is isolated from the plant milk thistle (Silybum marianum), and has been extensively used for its hepatoprotective effects in Asia an