Study on the apoptotic mechanisms of Ginkgetin,a biflavonoid from Ginkgo biloba L., in OVCAR-3 cells

• Main Authors: Shuo Kai Tsai, 蔡碩愷
• Other Authors: Chin Hsiang Chien
• Format: Others
• Language: zh-TW
• Published: 2000
• Online Access: http://ndltd.ncl.edu.tw/handle/92027071625377492017

碩士 === 國立陽明大學 === 生物化學研究所 === 88 === Ginkgetin, a biflavonoid, is an extract from Ginkgo biloba L. This compound is reported to have an effect on the selective inhibition on the growth of human ovarian carcinoma (OVCAR-3). Based on the MTT assay, we could demonstrate the survival rate decreased with the elevated concentration of Ginkgetin. Moreover, this effect showed a dose-response relationship. The data from electrophoresis, TUNEL assay and flow cytometry demonstrated those features of apoptosis such as DNA fragmentation, chromatin condensation as well as sub-G0/G1 peak could be observed after treating OVCAR-3 with 10 g/ml Ginkgetin for 12 hours. This result implied the cytotoxic effect may be the direct cause of apoptosis of OVCAR-3.and we could confirm Ginkgetin induces the apoptosis of OVCAR-3. Based upon the test on the caspase activity, we observed the activity of CPP32/YAMA increased with the time. In addition, the result of Western Blot showed the substrate of CPP32/YAMA, PARP, was cleaved by the enzyme more completely, when the activity of CPP32/YAMA was elevated, whereas the activity of caspase 9 did not change. Besides, the caspase inhibitor, z-VAD-fmk, was capable of effectively inhibiting the induction of apoptosis of OVCAR-3 by Ginkgetin after treating the cells with 50 M z-VAD-fmk for 2 hour and subsequently with 10 g/ml Ginkgetin for 24 hours. Ginkgetin is a kind of polyphenolic compound. The result indicated that GKT kills OVCAR-3, through intracellular oxidative stress and lead to apoptosis. In our result, the catalase was found to have the strongest antioxidant effect, when the tumor cells were pre-treated with vitamine C, N-acetylcysteine, and catalase respectively. Our preliminary result demonstrated the apoptosis of OVCAR-3 by Ginkgetin underwent the same signaling pathway through CPP32/YAMA as reported by Shiah et al., but not through caspase 9.This result implicated the apoptosis induced by Ginkgetin maybe undergo the pathway mediated by JNK still need to be investigated. 英文摘要---------------------------------3 緒言-------------------------------------5 材料與方法------------------------------12 實驗結果--------------------------------29 討論------------------------------------33 參考文獻--------------------------------36 附圖------------------------------------42