Although
verticinone, a major
alkaloid isolated from the bulbus of Fritillaria ussuriensis, has been shown to induce differentiation in human
leukemia cells, the exact mechanism of this action is not completely understood in
cancer cells.
Verticinone was used to conduct growth and apoptosis-related experiments for two stages of
oral cancer on immortalized human oral keratinocytes (IHOKs) and primary
oral cancer cells (HN4). The procedures included MTT assay, three-dimensional (3-D) raft cultures, Western blotting, cell cycle analysis, nuclear staining and
cytochrome c expression related to the apoptosis signaling pathway.
Verticinone inhibited the proliferation of immortalized and malignant oral keratinocytes in a dose- and time-dependent manner. In 3-D organotypic culture,
verticinone-treated cells were less mature than the control cells, displaying low surface keratinization and decreased epithelial thickness. The major mechanism by which
verticinone inhibits growth appears to be induced apoptosis and G(0)G(1) cell cycle arrest. This finding is supported by the results of the cell cycle analysis,
FITC-
Annexin V staining, DNA fragmentation assay and
Hoechst 33258 staining. Furthermore, the cytosolic level of
cytochrome c was increased, while the expression of Bcl-2
protein was gradually down-regulated and Bax was up-regulated, accompanied by
caspase-3 activation. The data suggests that
verticinone may induce apoptosis through a
caspase pathway mediated by mitochondrial damage in immortalized keratinocytes and
oral cancer cells.