Silibinin induces cell death through reactive oxygen species-dependent downregulation of notch-1/ERK/Akt signaling in human breast cancer cells.
The present study was undertaken to determine the underlying mechanism of silibinin-induced cell death in human breast cancer cell lines MCF7 and MDA-MB-231. Silibinin-induced cell death was attenuated by antioxidants, N-acetylcysteine (NAC) and 6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid, suggesting that the effect of silibinin was dependent on generation of reactive oxygen species (ROS). Western blot analysis showed that silibinin induced downregulation of extracellular signal-regulated kinase (ERK) and Akt. When cells were transiently transfected with constitutively active (ca) mitogen-activated protein kinase (MEK), an upstream kinase of ERK and caAkt, they showed resistance to silibinin-induced cell death. Silibinin decreased the cleavage of Notch-1 mRNA and protein levels. Notch-1-overexpressed cells were resistant to the silibinin-induced cell death. Inhibition of Notch-1 signaling was dependent on ROSgeneration. Overexpression of Notch-1 prevented silibinin-induced inhibition of ERK and Akt phosphorylation. Silibinin-induced cell death was accompanied by increased cleavage of caspase-3 and was prevented by caspase-3 inhibitor in MDA-MB-231 cells but not in MCF7 cells. Silibinin induced translocation of apoptosis-inducing factor (AIF), which was blocked by NAC, and transfection of caMEK and caAkt. Silibinin-induced cell death was prevented by silencing of AIF expression using small interfering AIF RNA in MCF7 cells but not in MDA-MB-231 cells. In conclusion, silibinin induces cell death through an AIF-dependent mechanism in MCF7 cells and a caspase-3-dependent mechanism in MDA-MB-231 cells, and ROS generation and Notch-1 signaling act upstream of the ERK and Akt pathway. These data suggest that silibinin may serve as a potential agent for induction of apoptosis in human breast cancer cells.
Department of Physiology (T.H.K., J.S.W., Y.K.K.) and Department of Obstetrics and Gynecology (K.H.K.), Pusan National University School of Medicine, Yangsan, Republic of Korea; and Pusan National University Hospital Biomedical Research Institute and Pusan Cancer Center (K.H.K.), Pusan, Republic of Korea.
Thae Hyun
TH
Kim
Jae Suk
JS
Woo
Yong Keun
YK
Kim
Ki Hyung
KH
Kim
D021581:Active Transport, Cell Nucleus; D000970:Antineoplastic Agents / Q000494:pharmacology*; D000975:Antioxidants / Q000494:pharmacology; D051033:Apoptosis Inducing Factor / Q000378:metabolism; D001943:Breast Neoplasms; D053148:Caspase 3 / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D015536:Down-Regulation; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism*; D005260:Female; D006801:Humans; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D051881:Receptor, Notch1 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D000077385:Silybin; D012838:Silymarin / Q000494:pharmacology*
C494829:AIFM1 protein, human; D000970:Antineoplastic Agents; D000975:Antioxidants; D051033:Apoptosis Inducing Factor; C497761:NOTCH1 protein, human; D017382:Reactive Oxygen Species; D051881:Receptor, Notch1; D012838:Silymarin; D000077385:Silybin; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D053148:Caspase 3