探討數種天然化合物對Aβ1-42造成大鼠腦皮質神經細胞毒性之保護功效

失智症 (Dementia) 是因為疾病或腦部受創而導致腦部神經細胞死亡或不正常運作的一種症狀，而阿茲海默氏症 (Alzheimer’s disease, AD ) 是最為普遍的一種失智症，佔失智症患者的 60~80％。阿茲海默氏症是一種漸進式且不可逆的神經退化疾病，患者的大腦呈現萎縮的現象，就神經病理學來看，阿茲海默氏症主要有兩個病理特徵，細胞外的老化斑塊 (senile plaques) ，和 tau 微管蛋白過度磷酸化後瓦解，導致細胞內神經纖維糾結現象 (neurofibrillary tangles, NFTs) 。近年的研究認為，小分子具水溶性的一種胜肽鏈，β-類澱粉胜肽寡聚合物 (Aβ oligomers) 是引發神經細胞毒性，造成神經細胞死亡的主要因素。截至目前為止，尚未開發出有效治療阿茲海默氏症的藥物，所以預防阿茲海默氏症的發生顯得非常重要。本實驗目的是利用 Aβ1-42 類澱粉胜肽寡聚合物誘導大鼠初代皮質神經細胞毒性作為模式，並以銀杏萃出物 (EGb761) 作為正控制組，探討天然化合物保護神經細胞對抗 Aβ1-42 誘導神經細胞死毒性以達到預防阿茲海默氏症的發生，樣品包括川陳皮素 (nobiletin) ，橘皮素 (tangeretin) ，四甲氧基類黃酮 (TMF) ，芝麻素 (sesamin) ，芝麻酚 (sesamol) ，四氫薑黃素 (tetrahydrocurcumin) ，茶胺酸 (L-theanine) ，並且進一步的研究有效樣品對於神經細胞可能的保護機制。根據 MTT assay 細胞存活率的結果，七種樣品之中， tangeretin ， nobiletin 抗 Aβ1-42細胞毒性的效果較好，甚至比正控制組的效果更佳， TMF 也有不錯的效果。另外樣品清除自由基能力 (DCF-DA) assay 的結果顯示， EGb761 具有很強的清除 Aβ1-42 誘導胞內自由基的能力，但是其餘的樣品效果不佳；Western bolt 的結果顯示，EGb761 具有抑制聚集和解集 Aβ1-42 的效果，tangeretin 則可能具有促進 Aβ1-42的聚集的能力以降低其對神經細胞的毒性， nobiletin 對 Aβ1-42的聚集沒有影響，其可能透過其他機制保護神經細胞。

關鍵字

阿茲海默氏症；類澱粉胜肽寡聚合物；初代皮質神經細胞；銀杏萃出物；川橘皮素；橘皮素；四甲氧基類黃酮；芝麻素；芝麻酚；四氫薑黃素；茶胺酸

並列摘要

Alzheimer’s disease is an irreversible, progressive neurodegenerative disorder. It is also the most common type of dementia, accounts for 60 to 80 percent of cases. Neuropathologically, there are three hallmarks of Alzheimer’s disease including brain atrophy, extracellular amyloid beta plaques and intracellular neurofibrillary tangles. Recent researches suggest that amyloid beta peptide induces neurontoxicity and causes neuronal cell death. Therefore, the objective of this study is to find the potential phytochemicals that can protect the brain neuron against Aβ1-42 oligomers induced neurotoxicity and thus prevent Alzheimer’s disease. Samples under investigation include nobiletin, tengeretin, L-theanine, sesamol, sesamin, tetramethoxyflavone, tetrahydrocurcumin and Ginkgo biloba extract, (EGb761) which is the positive control. Cells were treated with various concentrations of samples for 30 min before exposed to 1 μM Aβ1-42, 24 hr at 37℃. In this research, the pure compounds were used to evaluate their potential neuroprotective effects against Aβ1-42 induced toxicity in primary rat cortical neurons by assessing the cell viability with MTT assay and further investigate their possible mechanisms. According the results of viability assessed by MTT assay, the effective compounds are EGb761, nobiletin and tengeretin. In DCFDA assay, EGb761 has strong free radical scavenging activity but tangeretin and nobiletin do not have such activity. In Western blot, EGb761 was found to prevent Aβ1-42 oligomers formation and inhibits Aβ1-42 oligomers fibrillogenesis. Tangeretin possibly promotes the assembly Aβ1-42 into fibrils but nobiletin doesn’t have any effect on Aβ1-42. it indicates that tangeretin lowering Aβ1-42 oligomers toxicity may due to the promotion of Aβ1-42 fibrils production and nobiletin protects neurons from Aβ1-42 oligomers induced toxicity by other pathway.

並列關鍵字

Alzheimer’s disease (AD) ； Amyloid beta peptide ； primary rat cortical neuron ； nobiletin ； tangeretin ； TMF ； sesamin ； sesamol ； tetrahydrocurcumin ； L-theanine

參考文獻

侯怡卉。2010。數種天然物對於Aβ1-40引起PC12及分化後PC12細胞死亡的保護效果。國立台灣大學食品科技研究所。碩士論文。台北市。

蔡依潔。2011。數種天然物對大鼠初代皮質神經細胞的保護功效。國立台灣大學食品科技研究所。博士論文。台北市。

陳逸婷。2012。探討當歸甲醇萃出物及其活性成分對大鼠腦皮質神經細胞之保護功效。國立台灣大學食品科技研究所。博士論文。台北市。

Ahmed, M., Davis, J., Aucoin, D., Sato, T., Ahuja, S., Aimoto, S., Elliott, J. I., Van Nostrand, W. E., and Smith, S. O. (2010). Structural conversion of neurotoxic amyloid-beta(1-42) oligomers to fibrils. Nat Struct Mol Biol 17, 561-7.

Allan Butterfield, D. (2002). Amyloid β-peptide (1-42)-induced Oxidative Stress and Neurotoxicity: Implications for Neurodegeneration in Alzheimer's Disease Brain. A Review. Free Radical Research 36, 1307-1313.

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探討數種天然化合物對Aβ1-42造成大鼠腦皮質神經細胞毒性之保護功效

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