請用此 Handle URI 來引用此文件:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/58885
標題: | 從中草藥萃取物庫中篩選具調控細胞自噬作用的調節劑 Screening for autophagy modulators from an herbal extract library |
作者: | Shi-Zhen Liu 劉士禎 |
指導教授: | 潘建源(Chien-Yuan Pan) |
關鍵字: | 草藥萃取物,細胞自噬,p62,LC3,篩選平台,母草屬, herbal extract,autophagy,p62,LC3,Lindernia,screening platform, |
出版年 : | 2013 |
學位: | 碩士 |
摘要: | 許多傳統漢醫使用的草藥萃取物,具有開發調控細胞自噬新藥的潛力。細胞自噬是種依靠溶酶體(lysosome)的降解機制。在缺乏養分,逆境與細胞凋亡時扮演重要的角色。本研究使用 Irp62/HEK(接合水母冷光酶之複合蛋白)細胞株,建立篩選細胞自噬調控因子的平台,來研究草藥萃取物對細胞自噬的影響。以雷帕黴素(rapamycin)作為促進劑;6-氨基-3-甲基嘌呤(3-methyladenine)作為抑制劑。在497個草藥萃取物中,有14個可以促進細胞自噬。再使用MTT assay測試此14個潛力萃取物的毒性,挑選萃取自母草屬的HE717,來進行更進一步的測試。以臺盼藍細胞存活率分析法來驗證MTT assay的結果。而活化細胞自噬的能力,則是分別透過西方墨點法對p62、流式細胞儀與螢光顯微鏡對 GFP-LC3的影響來確認。綜合以上結果,由於 HE717具細胞自噬的活化能力,因此HE717具有製成藥物的潛力。本研究開發的生物篩選平台,是第一個用來篩選草藥對細胞自噬影響的平台。另外,研究結果也表示,母草屬的萃取物可能具有活化細胞自噬的效果。 Several herbal extracts (HEs), used in traditional Chinese medicine, have potential for the purpose of autophagy regulation. Autophagy is a lysosome-dependent degradation pathway. It plays important roles in starvation, stress response and apoptosis. To establish the platform to validate the modulators of autophagy, Irp62/HEK, which contain the constitutively expressed p62-luciferase for detection cells were used to detect the effects of herbal extracts on autophagy. Rapamycin was used as the activator control, and 3-methyladenine (3-MA) was used as the inhibitor control. Among the 497 HEs screened by the luciferase assay, 14 could serve as the activator of autophagy. Among 14 HEs, MTT assay was used to test the toxicity by measure the viability of the cells, and HE717 was chosen for the further study. The low toxicity of HE717 was carried out by trypan blue exclusion assay. The effect of activators was validated by Western blot of p62, flow cytometer measurement and fluorescence puncta of GFP-LC3. Taken together, HE717, isolated from whole plant of Lindernia sp., has the potential for drug development due to its activator effect of autophagy. This was the first platform to screen autophagy modulators from herbal library and the first report that Lindernia sp. could activate autophagy through p62 and LC3. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/58885 |
全文授權: | 有償授權 |
顯示於系所單位: | 生命科學系 |
文件中的檔案:
檔案 | 大小 | 格式 | |
---|---|---|---|
ntu-102-1.pdf 目前未授權公開取用 | 3.32 MB | Adobe PDF |
系統中的文件,除了特別指名其著作權條款之外,均受到著作權保護,並且保留所有的權利。