傳統中藥抑肝散過去常用於治療神經退行性疾病、精神官能症、失眠。而阿茲海默氏症是失智症裡各類型中最常見的，主要是以澱粉樣蛋白β (Aβ) 的異常堆積和Tau蛋白的過度磷酸化造成腦部細胞損傷有關。本文利用DPPH檢測中藥抑肝散的自由基清除與抗氧化能力；再透過細胞存活率分析法(MTT assay)檢視中藥抑肝散對於SH-SY5Y神經細胞的保護作用，實驗結果顯示中藥抑肝散具有相當高的自由基清除與抗氧化能力，對於SH-SY5Y神經細胞也有顯著的保護作用。本文利用轉基因果蠅表現Aβ42和hTauR406W毒性蛋白質作為AD動物模式，研究中藥抑肝散對人類Aβ 和人類Tau蛋白過度累積所引發的神經毒性在果蠅上是否具有保護作用。由於人類Aβ蛋白過度表現的Aβ42 會產生複眼退化；因此利用綠螢光蛋白觀察Aβ42 表現所造成的複眼退化，以及計算hTauR406W的背甲剛毛數目，可以評估中藥抑肝散對於Aβ和Tau蛋白過度表現果蠅的神經保護作用，實驗結果顯示中藥抑肝散可以緩解Aβ42 複眼的退化；另外也可以顯著緩解hTauR406W果蠅的背甲剛毛退化。此外，透過高靈敏的免疫磁減量(ImmunoMagnetic Reduction，IMR)技術可以評估Aβ42的Aβ蛋白表現，以及評估hTauR406W的Tau蛋白表現，結果顯示中藥抑肝散可以降低Aβ蛋白以及Tau蛋白的表現。歸納研究結果，可以說明中藥抑肝散對於緩解阿茲海默氏症可能具有替代醫療效果。

Yi-Gan-San (YGS), a traditional herbal medicine, has been used to manage neurodegenerative disorders and treat neurosis, insomnia, and dementia. Alzheimer’s disease (AD), a main cause of dementia, is the most common neurodegenerative disease that is related to the abnormal accumulation of amyloid β (Aβ) and Tau proteins. This master's thesis aims to use transgenic AD fly models overexpressing Aβ42 and Tau proteins to evaluate whether the traditional Chinese medicine Yigansan (YGS) exhibits a neuroprotective effect against the toxicity of accumulated Aβ and Tau proteins in Drosophila. In this study. 1,1-diphenyl-2-picrylhydrazyl (DPPH) was used to detect the free radical scavenging and anti-oxidant ability of YGS; then the cell viability (MTT) assay was used to examine the protective effect of YGS on SH-SY5Y neuroblastoma cells. Our results showed that YGS exhibited high free radical scavenging and antioxidant capacities and has a significant protective effect on SH-SY5Y cells. Next, we observed the green fluorescent protein expression of Aβ42 flies and calculated the number of bristles of hTauR406W flies to evaluate the neuroprotective effect of the YGS on the over-expression of Aβ42 and Tau toxicity in Drosophila. Our results showed that YGS can significantly alleviate degeneration but enhance green fluorescent protein expression in the eyes of Aβ42 flies; also, YGS can significantly alleviate bristles degradation in hTauR406W flies. In addition, through highly sensitive immunomagnetic reduction (IMR) technology, the Aβ42 and Tau protein expressions of Drosophila can be evaluated. Our results showed that YGS can significantly reduce the Aβ42 protein expression of Aβ42 flies and the Tau protein expression of hTauR406W flies. Our findings suggest that YGS may have a beneficial alternative therapy for AD via alleviating Aβ and Tau neurotoxicity that might help to alleviate the symptoms of neurodegeneration and dementia in the elderly.

Arriagada, P.V., Growdon, J.H., Hedley-Whyte, E.T., Hyman, B.T. (1992). Neurofibrillary tangles but not senile plaques parallel duration and severity of Alzheimer's disease. Neurology, 42(3 Pt 1), 631-639. https://doi.org/10.1212/wnl.42.3.631

Ballard, C.G., Gauthier, S., Cummings, J.L., Brodaty, H., Grossberg, G.T., Robert, P., Lyketsos, C.G. (2009). Management of agitation and aggression associated with Alzheimer's disease. Nature reviews. Neurology, 5(5), 245-255. https://doi.org/10.1038/nrneurol.2009.39

Bjerke, M., Engelborghs, S. (2018). Cerebrospinal Fluid Biomarkers for Early and Differential Alzheimer's Disease Diagnosis. Journal of Alzheimer's disease: JAD, 62(3), 1199-1209. https://doi.org/10.3233/JAD-170680

Braak, H., Del Tredici, K. (2015). The preclinical phase of the pathological process underlying sporadic Alzheimer's disease. Brain: a journal of neurology, 138(Pt 10), 2814-2833. https://doi.org/10.1093/brain/awv236

Burr, A.A., Tsou, W.L., Ristic, G., Todi, S.V. (2014). Using membrane-targeted green fluorescent protein to monitor neurotoxic protein-dependent degeneration of Drosophila eyes. Journal of neuroscience research, 92(9), 1100-1109. https://doi.org/10.1002/jnr.23395

Cerejeira, J., Lagarto, L., Mukaetova-Ladinska, E.B. (2012). Behavioral and psychological symptoms of dementia. Frontiers in neurology, 3, 73. https://doi.org/10.3389/fneur.2012.00073

Chieh, J.J., Huang, K.W., Chuang, C.P., Wei, W.C., Dong, J.J., Lee, Y.Y. (2016). Immunomagnetic Reduction Assay on Des-Gamma-Carboxy Prothrombin for Screening of Hepatocellular Carcinoma. IEEE transactions on bio-medical engineering, 63(8), 1681-1686. https://doi.org/10.1109/TBME.2015.2478845

Chiu, M.J., Yang, S.Y., Horng, H.E., Yang, C.C., Chen, T.F., Chieh, J.J., Chen, H.H., Chen, T.C., Ho, C.S., Chang, S.F., Liu, H.C., Hong, C.Y., Yang, H.C. (2013). Combined plasma biomarkers for diagnosing mild cognitive impairment and Alzheimer's disease. ACS chemical neuroscience, 4(12), 1530-1536. https://doi.org/10.1021/cn400129p

Doo, A.R., Kim, S.N., Park, J.Y., Cho, K.H., Hong, J., Eun-Kyung, K., Moon, S.K., Jung, W.S., Lee, H., Jung, J.H., Park, H.J. (2010). Neuroprotective effects of herbal medicine, Yi-Gan San on MPP+/MPTP-induced cytotoxicity in vitro and in vivo. Journal of ethnopharmacology, 131(2), 433-442. https://doi.org/10.1016/j.jep.2010.07.008

Gao, L., Li, X., Meng, S., Ma, T., Wan, L., Xu, S. (2020). Chlorogenic Acid Alleviates Aβ25-35-Induced Autophagy and Cognitive Impairment via the mTOR/TFEB Signaling Pathway. Drug design, development, and therapy, 14, 1705-1716. https://doi.org/10.2147/DDDT.S235969

Gong, W.X., Zhou, Y.Z., Qin, X.M., DU, G.H. (2019). Involvement of mitochondrial apoptotic pathway and MAPKs/NF-κ B inflammatory pathway in the neuroprotective effect of atractylenolide III in corticosterone-induced PC12 cells. Chinese journal of natural medicines, 17(4), 264-274. https://doi.org/10.1016/S1875-5364(19)30030-5

Hardy, J.A., Higgins, G.A. (1992). Alzheimer's disease: the amyloid cascade hypothesis. Science (New York, N.Y.), 256(5054), 184-185. https://doi.org/10.1126/science.1566067

He, Z., Guo, J.L., McBride, J.D., Narasimhan, S., Kim, H., Changolkar, L., Zhang, B., Gathagan, R.J., Yue, C., Dengler, C., Stieber, A., Nitla, M., Coulter, D.A., Abel, T., Brunden, K.R., Trojanowski, J.Q., Lee, V.M. (2018). Amyloid-β plaques enhance Alzheimer's brain tau-seeded pathologies by facilitating neuritic plaque tau aggregation. Nature medicine, 24(1), 29-38. https://doi.org/10.1038/nm.4443

Heitman, E., Ingram, D.K. (2017). Cognitive and neuroprotective effects of chlorogenic acid. Nutritional neuroscience, 20(1), 32-39. https://doi.org/10.1179/1476830514Y.0000000146

Ikarashi, Y., Mizoguchi, K. (2016). Neuropharmacological efficacy of the traditional Japanese Kampo medicine yokukansan and its active ingredients. Pharmacology & therapeutics, 166, 84-95. https://doi.org/10.1016/j.pharmthera.2016.06.018

Iwasaki, K., Satoh-Nakagawa, T., Maruyama, M., Monma, Y., Nemoto, M., Tomita, N., Tanji, H., Fujiwara, H., Seki, T., Fujii, M., Arai, H., Sasaki, H. (2005). A randomized, observer-blind, controlled trial of the traditional Chinese medicine Yi-Gan San for improvement of behavioral and psychological symptoms and activities of daily living in dementia patients. The Journal of clinical psychiatry, 66(2), 248-252. https://doi.org/10.4088/jcp.v66n0214

Jia, S.L., Wu, X.L., Li, X.X., Dai, X.L., Gao, Z.L., Lu, Z., Zheng, Q.S., Sun, Y.X. (2016). Neuroprotective effects of liquidity on cognitive deficits induced by soluble amyloid-β1-42 oligomers injected into the hippocampus. Journal of Asian natural products research, 18(12), 1186-1199.
https://doi.org/10.1080/10286020.2016.1201811

Kong, Z.H., Chen, X., Hua, H.P., Liang, L., Liu, L.J. (2017). The Oral Pretreatment of Glycyrrhizin Prevents Surgery-Induced Cognitive Impairment in Aged Mice by Reducing Neuroinflammation and Alzheimer's-Related Pathology via HMGB1 Inhibition. Journal of molecular neuroscience: MN, 63(3-4), 385-395. https://doi.org/10.1007/s12031-017-0989-7

Lawlor, B.A. (2004). Behavioral and psychological symptoms in dementia: the role of atypical antipsychotics. The Journal of clinical psychiatry, 65 Suppl 11, 5-10.
Lee, T.K., Kang, I.J., Kim, B., Sim, H.J., Kim, D.W., Ahn, J.H., Lee, J.C., Ryoo, S., Shin, M.C., Cho, J.H., Kim, Y.M., Park, J.H., Choi, S.Y., Won, M.H. (2020). Experimental Pretreatment with Chlorogenic Acid Prevents Transient Ischemia-Induced Cognitive Decline and Neuronal Damage in the Hippocampus through Anti-Oxidative and Anti-Inflammatory Effects. Molecules (Basel, Switzerland), 25(16), E3578. https://doi.org/10.3390/molecules25163578

Leinonen, V., Koivisto, A.M., Savolainen, S., Rummukainen, J., Tamminen, J.N., Tillgren, T., Vainikka, S., Pyykkö, O.T., Mölsä, J., Fraunberg, M., Pirttilä, T., Jääskeläinen, J.E., Soininen, H., Rinne, J., Alafuzoff, I. (2010). Amyloid and tau proteins in cortical brain biopsy and Alzheimer's disease. Annals of neurology, 68(4), 446-453. https://doi.org/10.1002/ana.22100

Lin, C.M., Lin, Y.T., Lee, T.L., Imtiyaz, Z., Hou, W.C., Lee, M.H. (2020). In vitro and in vivo evaluation of the neuroprotective activity of Uncaria hirsuta Haviland. Journal of food and drug analysis, 28(1), 147-158. https://doi.org/10.1016/j.jfda.2019.10.004

Mena, R., Edwards, P., Pérez-Olvera, O., Wischik, C.M. (1995). Monitoring pathological assembly of tau and beta-amyloid proteins in Alzheimer's disease. Acta neuropathologica, 89(1), 50-56. https://doi.org/10.1007/BF00294259

Mizukami, K., Asada, T., Kinoshita, T., Tanaka, K., Sonohara, K., Nakai, R., Yamaguchi, K., Hanyu, H., Kanaya, K., Takao, T., Okada, M., Kudo, S., Kotoku, H., Iwakiri, M., Kurita, H., Miyamura, T., Kawasaki, Y., Omori, K., Shiozaki, K., Odawara, T., et al. (2009). A randomized cross-over study of a traditional Japanese medicine (kampo), yokukansan, in the treatment of the behavioural and psychological symptoms of dementia. The international journal of neuropsychopharmacology, 12(2), 191-199. https://doi.org/10.1017/S146114570800970X

Nagata, K., Yokoyama, E., Yamazaki, T., Takano, D., Maeda, T., Takahashi, S., Terayama, Y. (2012). Effects of yokukansan on behavioral and psychological symptoms of vascular dementia: an open-label trial. Phytomedicine : international journal of phytotherapy and phytopharmacology, 19(6), 524-528. https://doi.org/10.1016/j.phymed.2012.02.008

Nakatani, Y., Tsuji, M., Amano, T., Miyagawa, K., Miyagishi, H., Saito, A., Imai, T., Takeda, K., Ishii, D., Takeda, H. (2014). Neuroprotective effect of yokukansan against cytotoxicity induced by corticosterone on mouse hippocampal neurons. Phytomedicine : international journal of phytotherapy and phytopharmacology, 21(11), 1458-1465. https://doi.org/10.1016/j.phymed.2014.06.004

Okahara, K., Ishida, Y., Hayashi, Y., Inoue, T., Tsuruta, K., Takeuchi, K., Yoshimuta, H., Kiue, K., Ninomiya, Y., Kawano, J., Yoshida, K., Noda, S., Tomita, S., Fujimoto, M., Hosomi, J., Mitsuyama, Y. (2010). Effects of Yokukansan on behavioral and psychological symptoms of dementia in regular treatment for Alzheimer's disease. Progress in neuro-psychopharmacology & biological psychiatry, 34(3), 532-536. https://doi.org/10.1016/j.pnpbp.2010.02.013

Olsson, B., Lautner, R., Andreasson, U., Öhrfelt, A., Portelius, E., Bjerke, M., Hölttä, M., Rosén, C., Olsson, C., Strobel, G., Wu, E., Dakin, K., Petzold, M., Blennow, K., Zetterberg, H. (2016). CSF and blood biomarkers for the diagnosis of Alzheimer's disease: a systematic review and meta-analysis. The Lancet. Neurology, 15(7), 673-684. https://doi.org/10.1016/S1474-4422(16)00070-3

Sun, X., Zeng, H., Wang, Q., Yu, Q., Wu, J., Feng, Y., Deng, P., Zhang, H. (2018). Glycyrrhizin ameliorates inflammatory pain by inhibiting microglial activation-mediated inflammatory response via blockage of the HMGB1-TLR4-NF-kB pathway. Experimental cell research, 369(1), 112-119. https://doi.org/10.1016/j.yexcr.2018.05.012

Tan, F.H.P., Azzam, G. (2017). Drosophila melanogaster: Deciphering Alzheimer's Disease. The Malaysian journal of medical sciences : MJMS, 24(2), 6-20. https://doi.org/10.21315/mjms2017.24.2.2

Teunissen, C.E., Chiu, M.J., Yang, C.C., Yang, S.Y., Scheltens, P., Zetterberg, H., Blennow, K. (2018). Plasma Amyloid-β (Aβ42) Correlates with Cerebrospinal Fluid Aβ42 in Alzheimer's Disease. Journal of Alzheimer's disease : JAD, 62(4), 1857-1863. https://doi.org/10.3233/JAD-170784

Wang, N., Zhou, Y., Zhao, L., Wang, C., Ma, W., Ge, G., Wang, Y., Ullah, I., Muhammad, F., Alwayli, D., Zhi, D., Li, H. (2020). Ferulic acid delayed amyloid β-induced pathological symptoms by autophagy pathway via a fasting-like effect in Caenorhabditis elegans. Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association, 146, 111808. https://doi.org/10.1016/j.fct.2020.111808

Wittmann, C.W., Wszolek, M.F., Shulman, J.M., Salvaterra, P.M., Lewis, J., Hutton, M., Feany, M.B. (2001). Tauopathy in Drosophila: neurodegeneration without neurofibrillary tangles. Science (New York, N.Y.), 293(5530), 711-714. https://doi.org/10.1126/science.1062382

Yang, S.Y., Chiu, M.J., Chen, T.F., Horng, H.E. (2017). Detection of Plasma Biomarkers Using Immunomagnetic Reduction: A Promising Method for the Early Diagnosis of Alzheimer's Disease. Neurology and therapy, 6(Suppl 1), 37-56. https://doi.org/10.1007/s40120-017-0075-7

Yang, S.Y., Liu, H.C., Chen, W.P. (2020). Immunomagnetic Reduction Detects Plasma Aβ1-42 Levels as a Potential Dominant Indicator Predicting Cognitive Decline. Neurology and therapy, 9(2), 435-442. https://doi.org/10.1007/s40120-020-00215-2

Yeh, P.A., Chien, J.Y., Chou, C.C., Huang, Y.F., Tang, C.Y., Wang, H.Y., Su, M.T. (2010). Drosophila notal bristle as a novel assessment tool for pathogenic study of Tau toxicity and screening of therapeutic compounds. Biochemical and biophysical research communications, 391(1), 510-516. https://doi.org/10.1016/j.bbrc.2009.11.089

Zhai, K.F., Duan, H., Cui, C.Y., Cao, Y.Y., Si, J.L., Yang, H.J., Wang, Y.C., Cao, W.G., Gao, G.Z., Wei, Z.J. (2019). Liquiritin from Glycyrrhiza uralensis Attenuating Rheumatoid Arthritis via Reducing Inflammation, Suppressing Angiogenesis, and Inhibiting MAPK Signaling Pathway. Journal of agricultural and food chemistry, 67(10), 2856-2864. https://doi.org/10.1021/acs.jafc.9b00185

Zhang, J.G., Geng, C.A., Huang, X.Y., Chen, X.L., Ma, Y.B., Zhang, X.M., Chen, J.J. (2017). Chemical and biological comparison of different sections of Uncaria rhynchophylla (Gou-Teng). European journal of mass spectrometry (Chichester, England), 23(1), 11-21. https://doi.org/10.1177/1469066717694044

Zhu, W.L., Zheng, J.Y., Cai, W.W., Dai, Z., Li, B.Y., Xu, T.T., Liu, H.F., Liu, X.Q., Wei, S.F., Luo, Y., Wang, H., Pan, H.F., Wang, Q., Zhang, S.J. (2020). Ligustilide improves aging-induced memory deficit by regulating mitochondrial related inflammation in SAMP8 mice. Aging, 12(4), 3175-3189. https://doi.org/10.18632/aging.102793