Synergistic anti-glioma effect of Hydroxygenkwanin and Apigenin in vitro

Apigenin (AP) and Hydroxygenkwanin (HGK) are two natural flavonoid compounds. Previous studies have already demonstrated the anti-tumor capability of AP. However, it is not clear whether HGK has such property. In the current study, the anti-glioma activities of HGK and its synergistic anti-glioma effects with AP on C6 glioma cells were investigated. In addition, the possible mechanisms were also studied. MTT assay and morphologic analysis including acridine orange/ethidium bromide (AO/EB) and 4',6-diamidino-2-phenylindole (DAPI) staining were used in the research, and the results indicated that the treatment with AP or HGK could inhibit C6 glioma cell proliferation respectively. Moreover, when AP was administrated simultaneously, the anti-glioma effect of HGK was dramatically enhanced in a dose-dependent manner, which is obviously better than that of carmustine (BCNU) at the concentration 25μM for treating of 24h. Compared with control, mitochondrial membrane potential (MPP) loss and mitochondrion damage were detected by JC-1 fluorescence probes (JC-1) and transmission electron microscopy (TEM) after treatment. Obvious DNA damage and cell cycle S phase arrest were detected by alkaline comet assay and flow cytometric analysis (FCM). Additionally, up regulation of TNF-α level, activations of Caspase-3, -8, over expressions of BID and Bak protein and Bcl-xL protein down expression were also observed after treatment by the combination of AP and HGK. The results indicate that HGK may be an effective natural product to treat glioma, and the combination of AP and HGK may be a promising method for glioma chemotherapy.