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| 題名: | 台灣香檬陳皮萃取物對阿茲海默症細胞之分析比較
The comparison of fermented peel extracts of Citrus depressa in Alzheimer’s disease model cells |
| 作者: | 鄭宇彤
CHENG, YU-TUNG |
| 貢獻者: | 醫學檢驗暨生物技術學系碩士班
林詠? |
| 關鍵詞: | 阿茲海默症;類澱粉蛋白;濤蛋白;台灣香檬;陳皮
Alzheimer’s disease;amyloid-β;Tau;Citrus depressa;fermented peel |
| 日期: | 2023-06-08 |
| 上傳時間: | 2024-01-22 13:23:36 (UTC+8) |
| 摘要: | 目的：阿茲海默症(Alzheimer’s disease; AD)是目前全球需迫切面臨的疾病之一，但目前臨床上的治療藥物皆為症狀治療，無法根治或預防AD。台灣香檬(Citrus depressa)是一種柑橘類水果，本研究的主旨在評估從台灣香檬萃取出的化合物是否對AD細胞具有保護作用，降低類澱粉蛋白的產生及其毒性。
方法：在此研究中，我們將台灣香檬的陳皮(Fermented peel)經過乙醇萃取，初步分離成乙酸乙酯(ethyl acetate; EA)層、丁醇(butanol; BuOH)層和水(H2O)層；另外，將小鼠神經母細胞瘤N2a細胞大量表達突變的類澱粉前驅蛋白(amyloid precursor protein; APP) (APP-Swe/Ind)的方式建立AD細胞模型。透過細胞計數試劑CCK8評估台灣香檬陳皮萃取物是否提高AD細胞存活度，再利用免疫螢光染色、酵素結合免疫吸附分析法和西方墨點法探討台灣香檬陳皮萃取物是否會影響類澱粉蛋白生成路徑、濤(Tau)蛋白磷酸化、原肌球蛋白受體激?A (TrkA)路徑、細胞外訊號調節激?(ERK)路徑和蛋白激?B (Akt)路徑等。
結果：我們使用100 μg/ml作為處理AD細胞的濃度，因為這是細胞存活實驗的共同安全濃度。初步結果顯示，在EA層處理後的AD細胞中，細胞存活度上升且類澱粉蛋白生成路徑的中間產物APP-羧端片段(C-terminal fragment；CTF)-β有下降的趨勢，在細胞培養液中也觀察到Aβ1-42有下降的情形，並且也降低了Tau蛋白磷酸化的程度。於是我們進一步純化EA層，選出了六個毒性較低的成份作用於AD細胞，發現有部分成份能有效使AD細胞存活度上升，並抑制APP-CTFβ和胞外Aβ的產生，也使Tau蛋白磷酸化程度下降。
結論：台灣香檬陳皮以乙酸乙酯萃取出的成分可以提高AD模型細胞的存活度並且降低AD病理標誌Aβ和磷酸化Tau蛋白，未來可發展為治療AD的潛在藥物。
Objective: Alzheimer’s disease (AD) is one of the severe public health problems. Because of the complex mechanism, no cure would ever successfully treat the people living with this disease. In this study, we evaluated whether Citrus depressa extracts can inhibit amyloidogenic pathway and promote the survival of AD model cells.
Methods: The components of the ethanol extract of Citrus depressa fermented peel were liquid-liquid partitioned using ethyl acetate (EA), butanol (BuOH) and water (H2O). To induce the pathology of AD in cells, we transfected neuroblastoma N2a cells with pCAX-APP-Swe/Ind, establishing AD model cell. The AD cells were treated with these extracts, and the cell viability was evaluated using cell counting kit CCK-8. We then investigated the effects of these extracts on amyloidogenic pathway, Tau protein phosphorylation, and the pathways of TrkA, ERK and Akt.
Results: From the cell viability test, we determined the minimal toxic concentration of the three layers. For safety, we used 100 μg/ml for following experiments. The results showed that cells treated with the EA layer increased AD cell viability. Further experiments showed anti-amyloidogenic activities of the EA layer with decreased levels of amyloid precursor protein-C-terminal fragment (APP-CTF)-β, the intermediate of amyloidogenic pathway, in the cell lysates, and amyloid-β in the cell culture media. Furthermore, we found that the EA layer decreased phosphorylation of Tau. Finally, we fractionalized the EA layer and found that a few fractions displayed good anti-amyloidogenic activities.
Conclusions: The ingredients extracted from Citrus depressa fermented peel with EA can improve the survival of AD model cells and reduce AD pathological markers Aβ and phosphorylated Tau, which can be developed into potential treatment of AD in the future. |
| 描述: | 碩士
指導教授:林詠?
委員:林詠?
委員:高淑慧
委員:李慶國 |
| 資料類型: | thesis |
| 顯示於類別: | [醫學檢驗暨生物技術學系所] 博碩士論文
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