中草藥及天然或合成化合物對阿茲海默氏症細胞與動物模式的治療效用

Therapeutic Benefits of Chinese Herbal Medicine Extract and Natural/Synthetic Compounds on β-amyloid Induced Cell and Animal Models of Alzheimer’s Disease

邱雅貞(Chiu Ya-Jen)

指導教授 ： 李桂楨

國立臺灣師範大學/理學院/生命科學系/博士(2018年)

https://doi.org/10.6345/DIS.NTNU.SLS.025.2018.D01

摘要

阿茲海默氏症(AD)是有關漸進性認知衰退和記憶力喪失的疾病， 也是最常見的一種老人痴呆症，病理學上病徵之一是由 β-澱粉樣 (Aβ)沉積物組成的老年斑。研究發現，腦中的 Aβ 堆積會造成氧化壓 力和發炎損傷，進而導致神經細胞凋亡及認知功能失調。有鑑於 此，尋找可能減少 Aβ 聚集的方法，可能有效治療 AD。本研究檢視 中藥脹果甘草(G. inflata)及其活性成分甘草查爾酮 A (Licochalcone A)、甘草素(Liquiritigenin)，和本校化學系姚清發老師提供的合成化 合物 NC009-1，抑制 Aβ 聚集的情形及神經保護性。誘導性 Aβ-GFP 293 細胞、硫黃素 T (Thioflavin T)或 DPPH 清除自由基試驗結果顯 示，脹果甘草/活性成分和 NC009-1，皆可抑制 Aβ 蛋白聚集和相關 聯的氧化壓力。此外，LPS/IFN-γ 刺激發炎的小鼠 BV-2 微膠細胞試 驗顯示，脹果甘草/活性成分可減少 BV-2 的發炎反應，來達到抗發 炎的效果。另外，誘導性 Aβ-GFP SH-SY5Y 細胞試驗結果顯示，脹 果甘草/活性成分和 NC009-1 可抑制乙醯膽鹼酶(acetylcholinesterase) 活性、增強 SOD2 表現、及/或促進神經突生長。為瞭解脹果甘草/活 性成分和 NC009-1 對細胞保護的分子機制，以抗體或 PCR 矩陣，檢 測 Aβ-GFP SH-SY5Y 細胞中，與細胞凋亡相關蛋白或阿茲海默氏症 相關基因的表現。結果發現，脹果甘草/活性成分可減緩 BCL2 的下 降，並減低 IGFBP2 的上升、凋亡蛋白酶 3 的切割、BAD 及 BAX 的量，來保護 Aβ-GFP SH-SY5Y 細胞免於 BV-2 制約培養液誘導的 細胞死亡。然而 NC009-1 在 Aβ-GFP SH-SY5Y 細胞中，可正調控 APOE 和 TRKA 的表現。在鏈脲佐菌素(Streptozocin)誘導高血糖的 PS1M146V、APPSwe 和 TauP301L 三基因(3×Tg) AD 轉殖鼠的實驗中， NC009-1 並可挽救 APOE 和 TRKA 的減少，降低海馬迴及腦皮層中 Aβ 及 Tau 的量，及減緩認知缺失。這些研究結果指出，脹果甘草/ 活性成分與 NC009-1 可能作為 AD 的治療策略。

關鍵字

阿茲海默氏症 ； β-澱粉樣 ； 脹果甘草 ； 甘草查爾酮 A ； 甘草素 ； 合成化合物

並列摘要

Alzheimer’s disease (AD) is the most prevalent form of dementia associated with progressive cognitive decline and memory loss. One of the pathological hallmarks of AD is senile plaques consisting of β- amyloid (Aβ) deposits. Studies have proposed that Aβ deposition causes oxidative stress and inflammatory damage which lead to apoptotic cell death and eventual cognitive deficits. Therapeutic approaches to identify novel Aβ aggregate reducers could be an effective treatment for the disease. In this study, Chinese herbal medicine G. inflata and its bioactive constituents licochalcone A and liquiritigenin, and synthetic compound NC009-1 provided by Professor Ching-Fa Yao from Department of Chemistry of NTNU, were examined for Aβ aggregation reduction and neuroprotection. By using inducible Aβ-GFP 293 cells, biochemical thioflavin T or DPPH free radical scavenging assays, G. inflata/constituents and NC009-1 reduced Aβ aggregation and associated oxidative stress. Besides, G. inflata/constituents showed anti- inflammatory effect by attenuating the inflammatory response of BV-2 microglia under LPS/IFN-γ stimulation. In addition, G. inflata/constituents and NC009-1 displayed acetylcholinesterase inhibition, SOD2 up-regulation, and/or neurite outgrowth promotion in inducible Aβ-GFP SH-SY5Y cells. To reveal the molecular mechanisms underlying protective effects of G. inflata/constituents and NC009-1, antibody or PCR array was used to assess expression changes of apoptosis-associated proteins or AD-related genes in the Aβ-GFP SH- SY5Y cells. G. inflata/bioactive constituents protected Aβ-GFP SH- SY5Y cells from BV-2 conditioned media-induced cell death by ameliorating reduced BCL2 and attenuating increased IGFBP2, cleaved CASP3, BAD and BAX, whereas NC009-1 up-regulated the expression of APOE and TRKA in Aβ-GFP SH-SY5Y cells. NC009-1 further rescued the down-regulated APOE and TRKA and reduced Aβ and Tau levels in hippocampus and cortex, and ameliorated cognitive deficits in streptozocin-induced hyperglycemic PS1M146V, APPSwe, and tauP301L 3×Tg-AD mice. These results indicate that G. inflata/constituents and NC009-1 could be possible treatment strategies for AD.

並列關鍵字

Alzheimer’s disease ； β-amyloid ； Glycyrrhiza inflata ； licochalcone A ； liquiritigenin ； Indole compound

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