Verticinone induces cell cycle arrest and apoptosis in immortalized and malignant human oral keratinocytes.
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.
Yun
Young-Gab
YG
Department of Prescription, College of Oriental Medicine, Wonkwang University, Iksan, South Korea.
Jeon
Byung-Hun
BH
Lee
Jong-Hyun
JH
Lee
Sun-Kyung
SK
Lee
Hwa-Jeong
HJ
Jung
Kyung-Hee
KH
Jun
Chang-Duk
CD
Lee
Suk-Keun
SK
Kim
Eun-Cheol
EC
D000906:Antibodies / Q000378:metabolism; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000032:analysis / Q000096:biosynthesis; D002277:Carcinoma / Q000473:pathology; D002453:Cell Cycle / Q000187:drug effects*; D018797:Cell Cycle Proteins / Q000032:analysis / Q000096:biosynthesis; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002595:Cevanes / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004365:Drugs, Chinese Herbal / Q000494:pharmacology; D031410:Fritillaria / Q000737:chemistry; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D015603:Keratinocytes / Q000187:drug effects*; D009062:Mouth Neoplasms / Q000473:pathology; D013997:Time Factors
D000906:Antibodies; D000972:Antineoplastic Agents, Phytogenic; D051017:Apoptosis Regulatory Proteins; D018797:Cell Cycle Proteins; D002595:Cevanes; D004365:Drugs, Chinese Herbal; C014242:verticine