虎杖對 EB 病毒潛伏期第一型巴克氏淋巴瘤細胞的影響

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標題:	虎杖對 EB 病毒潛伏期第一型巴克氏淋巴瘤細胞的影響 Effects of Polygonum cuspidatium on EBV Latency I Burkitt's Lymphoma Cells
作者:	周哲銘
貢獻者:	保健營養系林翠品
關鍵字:	計畫性壞死虎杖 EB 病毒 Necroptosis EBV Polygonum cuspidatum
日期:	2012
上傳時間:	2013-01-07 16:42:13 (UTC+8)
摘要:	EB 病毒是普遍存在於人類的皰疹病毒，與許多的惡性腫瘤有相關性。虎杖是一種在亞洲地區廣泛使用的中草藥。在先前研究發現虎杖乙酸乙酯分離物會增加 EB 病毒潛伏期第三型淋巴母細胞潛伏期膜蛋白質1(LMP1)、EBNA1及溶裂蛋白質的表現，進而使細胞內 Reactive oxygen species (ROS) 產生，而導致細胞產生凋亡。而本研究將進一步探討 F2a 是否也能誘發 EB 病毒潛伏期第一型巴克氏淋巴瘤細胞凋亡。實驗以 trypan blue 染色方法與MTT 方法測定 F2a 對EB病毒潛伏期第一型巴克氏淋巴瘤細胞 Akata細胞的存活率；西方墨點法測定凋亡相關蛋白質的表現；使用與 DNA 結合的螢光染劑， DAPI ，觀察細胞核形態；以流式細胞儀測定細胞內凋亡及壞死百分比。結果顯示 F2a 處理 Akata EBV (+) 細胞 24 與 48 小時後有明顯毒殺細胞能力，細胞毒性 CC50 分別為 10.8 及 9.7g/ml ；而且也發現 Akata 細胞以 F2a 處理 24小時，會使 Akata 細胞凋亡及壞死的細胞數增加，而處理 48 小時則會使 Akata 大量走上壞死;進一步以計劃性壞死（necroptosis）抑制劑，Nec-1 (Necrostatin-1) 與 F2a 一起處理 Akata 細胞發現壞死比例降低，細胞凋亡數增加，由此結果推論 F2a 使 Akata 細胞走向計畫性壞死；而當 F2a 與計劃性壞死的抑制劑 Nec-1一起作用時會使 Akata 細胞走向凋亡，所以 F2a 有潛能被發展成為治療 EB 病毒相關腫瘤的藥劑。 Epstein-Barr virus is commonly found in human herpes virus, related with the many malignant tumors. The Polygonum cuspidatium is a widely used in Asia herbal medicine. Our previous study showed that the ethyl acetate subfraction F2a from Polygonum cuspidatum increased the expression of the EBV latent membrane protein 1 (LMP1), EBNA1 and lytic proteins which induced high level of reactive oxygen species (ROS) and cause EBV latency III cells apoptosis. This study will further explore the F2a whether can also induce EBV latency I Burkitt's lymphoma to apoptosis. Trypan blue staining and MTT assay were used to the viability of EBV latency I Burkitt's lymphoma, Akata cells. The expression of apoptosis-related proteins was determined by western blotting. Using the DNA-binding fluorescent dye, DAPI to observe nuclear morphology; Flow cytometry was used to determine ROS, cell apoptosis and necrosis rates. Results showed that Akata cells treated F2a for 24 or 48 hr, the cell viability was decreased, CC50 were 10.8 and 9.7 g/ml, respectively. Furthermore, The percentage of apoptotic and necrotic cells were increased when Akata cells treated with F2a for 24 hr. Meanwhile, The F2a treatment for 48 hours caused the necrotic Akata cells increasing. Further to use necroptosis inhibitors, Nec-1 (Necrostatin-1) and F2a treatment in Akata cells found the percentage of necrotic cells reducing, apoptotic cells increasing. These results suggested that the F2a may induce Akata cell to necroptosis; when the treatment of F2a and necroptosis inhibitor Nec-1 promoted the Akata cell apoptosis, Therefore, F2a may be used as a useful therapeutic drug in the treatment of EB virus related tumor.
關聯:	校內一年後公開，校外永不公開，學年度：100，52頁
显示于类别:	[保健營養系(所) ] 博碩士論文

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