[The Research on the Anti-glioma Effect and Mechanism of Cinobufagin]

Abstract

Objective: To study the effect of cinobufagin (CB) on the proliferation inhibition and induction of apoptosis in glioblastoma cell lines U87 and its molecular mechanism.

Methods: A gradient concentration (0-20 μmol/L) of CB was used to treat the U87 glioma cells for 6 h,12 h,24 h and 48 h,respectively. Cell viabilities were determined by CCK-8 assay to discover the effects of different concentrations of CB on the proliferation of glioma cells. Different concentrations (1-20 μmol/L) of CB were used to treat the U87 glioma cells for 12 h and 24 h,hochest33342 staining assay was used to assess the apoptosis levels. Immunofluorescence staining was used to determine the expression of growth related proteins phospho-protein kinase B(T308)[ p-AKT(T308)] in U87 glioma cells after being treated with CB for 24 h. Western blot was used to determine the apoptotic related proteins (BAX,cleaved-caspase 3,cleaved-caspase 9) and growth related proteins [phospho-inositide 3-kinase (p-PI3K),p-AKT(T308),p-AKT(S473),phospho-ribosomal protein S6 kinase (PS6),phospho-4E-binding protein 1 (p-4EBP1)].

Results: A significant effect of CB on the proliferation inhibition and induction of apoptosis in U87 glioma cells in a time- and dose-dependent manner was observed. Treatment with CB induced the expression levels of apoptosis-related protein,cleaved-caspase 3 and BAX,and the PI3K-AKT-4EBP1 signaling pathway related proteins p-AKT(T308) and p-4EBP1 were decreased.

Conclusion: CB can inhibit U87 glioma cells growth and induce apoptosis,which may involve the PI3K-AKT-4EBP1 and BAX-caspase signaling pathways.

Keywords: Apoptosis; Cinobufagin; Glioma; Growth inhibition.