title: Molecular mechanism of cell cycle blockage of hepatoma SK-Hep-1 cells by Epimedin C through suppression of mitogen-activated protein kinase activation and increased expression of CDK inhibitors p21(Cip1) and p27(Kip1).
abstract: Reports elsewhere demonstrated that Epimedin C, a constituent isolated from the leaves of Epimedium sagittatum, possessed anti-tumor activity. However, its mechanism of action remains unresolved. Using SK-Hep-1 cells, a poorly-differentiated hepatoma subline, as an experimental model, we present evidence here that the anti-tumor activity of Epimedin C may involve cell cycle blockage. Immunoblotting analyses demonstrated that Epimedin C caused a decreased expression of hyperphosphorylated retinoblastoma (Rb) protein, cyclin D1, c-Myc, and c-Fos. In parallel, we measured the kinase activities and found that CDK2 and CDK4 were suppressed with commensurate increased levels of CDK inhibitors, p21(Cip1) and p27(Kip1). These data suggested that Epimedin C arrested the proliferation of these cells at G0/G1 phase through inhibition of CDK2 and CDK4 activities via an increased induction of p21(Cip1) and p27(Kip1). Alternatively, we investigated whether the anti-proliferative effect of Epimedin C on these cells might involve MAP kinase cascade. Using western blotting technique, we demonstrated that Epimedin C also selectively decreased ERK1/2 phosphorylation. Among the downstream effectors of ERK examined, we found that Epimedin C selectively decreased the expression of c-Fos, but not c-Jun. By EMSA assay, we further demonstrated that decreased c-Fos resulted in the downregulation of AP-1/DNA binding activity. Taken together, the molecular mechanisms of anti-tumor activity of Epimedin C may be proceeded by the combined effects of the cell cycle blockage via either the inhibition of CDK2 and CDK4 activities, with commensurate increase in their inhibitors, p21(Cip1) and p27(Kip1) or negatively modulates the ERK/c-Fos/AP-1 signaling pathway.
authors.affiliation: Center for Gerontological Research and Graduate Institute of Medical Biotechnology, Chang-Gang University, Kwei-Shan, Taoyuan, Taiwan, ROC.
authors.forename: Tsan-Zon
authors.initials: TZ
authors.lastname: Liu
authors.forename: Chung-Yi
authors.initials: CY
authors.lastname: Chen
authors.forename: Shuenn-Jiun
authors.initials: SJ
authors.lastname: Yiin
authors.forename: Ching-Hsein
authors.initials: CH
authors.lastname: Chen
authors.forename: Jiin-Tsuey
authors.initials: JT
authors.lastname: Cheng
authors.forename: Ming-Kuei
authors.initials: MK
authors.lastname: Shih
authors.forename: Yu-Shan
authors.initials: YS
authors.lastname: Wang
authors.forename: Chi-Liang
authors.initials: CL
authors.lastname: Chern
mesh_terms: D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000473:pathology*; D002317:Cardiovascular Agents / Q000627:therapeutic use*; D002453:Cell Cycle / Q000187:drug effects*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival / Q000187:drug effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000096:biosynthesis*; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000096:biosynthesis*; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors*; D004273:DNA, Neoplasm / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D005419:Flavonoids / Q000627:therapeutic use*; D015870:Gene Expression / Q000187:drug effects; D016762:Genes, fos / Q000187:drug effects; D016758:Genes, jun / Q000235:genetics; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008113:Liver Neoplasms / Q000473:pathology*; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000378:metabolism
chemical_list: C493956:CDKN1A protein, human; C436291:CDKN1B protein, human; D002317:Cardiovascular Agents; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004273:DNA, Neoplasm; D005419:Flavonoids; D047908:Intracellular Signaling Peptides and Proteins; D018808:Transcription Factor AP-1; C106054:epimedin C; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases; D020928:Mitogen-Activated Protein Kinases