臺灣博碩士論文加值系統

根據世界衛生組織於2012年指出，全球約三億人口罹患憂鬱症的盛行率會影響了整個社會的經濟結構。而治療過程中服用藥物引起的副作用及不適感會導致患者服藥順從性不佳，使得憂鬱症的治療更加困難，因此利用食補觀念加強疾病預防是當務之急。冬蟲夏草是中國歷史中傳統的名貴中藥材，但由於採集不易等因素而產量逐年下降。北冬蟲夏草（Cordyceps militaris）則被認為是冬蟲夏草良好的取代物，研究發現蟲草素可延長小鼠在水中泳動時間、延緩疲勞。本研究旨在利用類憂鬱症動物模式中，探討北冬蟲夏草水萃物是否可藉由調控發炎激素之濃度，影響Rho-associated kinase protein（ROCK2）及Akt蛋白質表現量，預防神經細胞凋亡並維持單胺類神經傳導物質濃度而達到抗憂鬱之功效。動物實驗利用不可預期慢性壓力（Unpredictable chronic mild stress）持續五週來誘導動物產生類憂鬱傾向，並同時管餵北冬蟲夏草水萃物125 mg/kg bw、250 mg/kg bw、500 mg/kg bw，後續進行蔗糖偏好試驗（Sucrose preference test, SFT）及開場試驗（Open field test, OFT）評估北冬蟲夏草水萃物抗憂鬱功效。研究結果發現，北冬蟲夏草水萃物125 mg/kg bw、250 mg/kg bw可有效減少腦部皮質區Rho-associated protein kinase（ROCK）蛋白質表現量進而調節Akt蛋白質濃度；減少血清細胞激素濃度，並顯著提升腦部皮質區血清素及多巴胺轉換率，也顯著提升動物糖水偏好程度；在開放空間試驗中提升動物總移動距離及各象限跨越次數。綜合上述結果證實北冬蟲夏草水萃物可經由調控發炎反應及ROCK2/Akt蛋白質路徑達到維持單胺類神經傳導物濃度，預防動物在壓力模式下產生憂鬱傾向。

According to world health organization, depression may be the first contribution of global burden disease in 2030. Due to the side effect in treatment and the low reception of drug, the new strategy of preventing depression is required. Cordyceps militaris has been known as a precious Chinese herb in Asia. Researchers had found amounts of neuroprotection effects in recent years such as it can significantly decrease the immobility time in forced swimming test in mice model. Modern scientists have confirmed that the mainly active compound in Cordyceps militaris including cordycepin, poly-saccharides, nucleoside and sterols. The aim of this study is to evaluate the anti-depression effect of Cordyceps militaris precisely. In unpredictable chronic mild stress animal model, Cordyceps militaris maintained the amount of water intake and food intake to prevent weight loss in stress condition. It was observed that Cordyceps militaris (125 mg/kg bw、250 mg/kg bw、500 mg/kg bw) may significantly increase the crossing number and total traveled distance in open field test. Besides, it also increased the sucrose preference. Low dosage of Cordyceps militaris (125 mg/kg bw) significantly decreased the IL-1β in serum. In brain cortex, low dosage of Cordyceps militaris (125 mg/kg bw) might altered the serotonin turnoverate and dopamine turnoverate. In western blotting, the data showed that Cordyceps militaris (125 mg/kg bw、250 mg/kg bw、500 mg/kg bw) may decreased the ROCK2 protein expression and up-regulated the Akt protein expression in cortex. In conclusion, we found that Cordyceps militaris might regulate the IL-1β in serum and further regulated the ROCK2/Akt pathway in anti-depression effect.

第一章、前言 1
第二章、文獻回顧 2
第一節、憂鬱症 2
一、憂鬱症流行病學 2
二、憂鬱症致病機轉 3
三、憂鬱症治療方式 14
第二節、北冬蟲夏草 16
一、北冬蟲夏草簡介 16
二、北冬蟲夏草栽培方式 16
三、北冬蟲夏草功效 17
第三節、ROCK2/Akt路徑 23
第三章、研究假說及目的 26
第一節、研究假說 26
第二節、研究目的 26
第四章、實驗架構 27
第五章、實驗材料與方法 29
第一節、實驗材料 29
一、北冬蟲夏草水萃物 29
二、動物實驗相關材料 29
三、化學藥品 30
第二節、動物實驗方法 32
一、動物實驗之飼養方法 32
二、動物實驗之實驗排程 32
三、不可預期慢性輕度壓力(unpredictable chronic mild stress, UCMS)動物模式 33
四、糖水偏好試驗（sucrose preference test, SPT） 33
五、開放空間試驗（open field test, OFT） 34
六、動物犧牲其器官採集 34
七、腦部單胺類神經傳導物質檢測 34
八、血清壓力賀爾蒙–皮質固醇檢測 35
第六章、實驗結果 36
一、飲水量 36
二、攝食量 36
三、體重變化 36
四、行為試驗-開放空間試驗（open field test） 37
(一)跨越各象限次數（number of crossing） 37
(二)總移動距離（total traveled distance） 37
五、蔗糖偏好試驗（sucrose preference test） 37
六、單胺類神經傳導物質濃度變化 38
(一)血清素轉換率 38
(二)多巴胺轉換率 38
七、血清IL-1β濃度結果 38
八、ROCK2蛋白質表現量. 39
九、PTEN蛋白質表現量 39
十、Akt蛋白質表現量 39
第七章、討論 41
一、飲水量、攝食量及體重結果討論 41
二、開放空間試驗（open field test）結果討論 42
三、蔗糖偏好試驗結果討論 43
四、單胺類神經傳導物質結果討論 44
(一)血清素（serotonin） 44
(二)多巴胺（dopamine） 45
五、血清中interleukin -1β（IL-1β）結果討論 46
六、ROCK2蛋白質表現量結果討論 48
七、PTEN蛋白質表現量結果討論 49
八、Akt蛋白質表現量結果討論 49
九、劑量效應討論 51
第八章、結論 64
第九章、附錄 66
第十章、參考文獻 69

Depression chapter103.
Ahn, J. Y., Neuroprotection signaling of nuclear akt in neuronal cells. Exp Neurobiol 2014, 23, 200-206.
Belmaker, R.; Agam, G., Major depressive disorder. New England Journal of Medicine 2008, 358, 55-68.
Bhatia, V.; Tandon, R. K., Stress and the gastrointestinal tract. Journal of gastroenterology and hepatology 2005, 20, 332-339.
Budzyn, K.; Marley, P. D.; Sobey, C. G., Targeting Rho and Rho-kinase in the treatment of cardiovascular disease. Trends in pharmacological sciences 2006, 27, 97-104.
Cassidy, F.; Weiner, R. D.; Cooper, T. B.; Carroll, B. J., Combined catecholamine and indoleamine depletion following response to ECT. The British Journal of Psychiatry 2010, 196, 493-494.
Chen, C.; Wang, M.-L.; Jin, C.; Chen, H.-J.; Li, S.-H.; Li, S.-Y.; Dou, X.-F.; Jia, J.-Q.; Gui, Z.-Z., Cordyceps militaris polysaccharide triggers apoptosis and G0/G1 cell arrest in cancer cells. Journal of Asia-Pacific Entomology 2015, 18, 433-438.
Das, S. K.; Masuda, M.; Sakurai, A.; Sakakibara, M., Medicinal uses of the mushroom Cordyceps militaris: current state and prospects. Fitoterapia 2010, 81, 961-968.
Del Grande da Silva, G.; Wiener, C. D.; Barbosa, L. P.; Goncalves Araujo, J. M.; Molina, M. L.; San Martin, P.; Oses, J. P.; Jansen, K.; Dias de Mattos Souza, L.; Azevedo da Silva, R., Pro-inflammatory cytokines and psychotherapy in depression: Results from a randomized clinical trial. J Psychiatr Res 2016, 75, 57-64.
Dunlop, B. W.; Nemeroff, C. B., The role of dopamine in the pathophysiology of depression. Archives of general psychiatry 2007, 64, 327-337.
Felger, J. C.; Lotrich, F. E., Inflammatory cytokines in depression: neurobiological mechanisms and therapeutic implications. Neuroscience 2013, 246, 199-229.
Hall, C. S., Emotional behavior in the rat. I. Defecation and urination as measures of individual differences in emotionality. Journal of Comparative Psychology 1934, 18, 385-403.
Heinrichs, S. C.; Cole, B. J.; Pich, E. M.; Menzaghi, F.; Koob, G. F.; Hauger, R. L., Endogenous corticotropin-releasing factor modulates feeding induced by neuropeptide Y or a tail-pinch stressor. Peptides 1992, 13, 879-884.
Herskowitz, J. H.; Feng, Y.; Mattheyses, A. L.; Hales, C. M.; Higginbotham, L. A.; Duong, D. M.; Montine, T. J.; Troncoso, J. C.; Thambisetty, M.; Seyfried, N. T., Pharmacologic inhibition of ROCK2 suppresses amyloid-β production in an Alzheimer''s disease mouse model. Journal of Neuroscience 2013, 33, 19086-19098.
Hsu, C.-C.; Huang, Y.-L.; Tsai, S.-J.; Sheu, C.-C.; Huang, B.-M., In vivo and in vitro stimulatory effects of Cordyceps sinensis on testosterone production in mouse Leydig cells. Life Sciences 2003, 73, 2127-2136.
Inan, S. Y.; Soner, B. C.; Sahin, A. S., Infralimbic cortex Rho-kinase inhibition causes antidepressant-like activity in rats. Progress in Neuro-Psychopharmacology and Biological Psychiatry 2015, 57, 36-43.
Jin, M. L.; Park, S. Y.; Kim, Y. H.; Oh, J. I.; Lee, S. J.; Park, G., The neuroprotective effects of cordycepin inhibit glutamate-induced oxidative and ER stress-associated apoptosis in hippocampal HT22 cells. Neurotoxicology 2014, 41, 102-111.
Jurado, S.; Benoist, M.; Knafo, S.; Petrok, C. N.; Esteban, J. A., PTEN is recruited to the postsynaptic terminal for NMDA receptor‐dependent long‐term depression. The EMBO journal 2010, 29, 2827-2840.
Katz, R. J.; Roth, K. A.; Carroll, B. J., Acute and chronic stress effects on open field activity in the rat: implications for a model of depression. Neuroscience & Biobehavioral Reviews 1981, 5, 247-251.
Kim, H. G.; Shrestha, B.; Lim, S. Y.; Yoon, D. H.; Chang, W. C.; Shin, D. J.; Han, S. K.; Park, S. M.; Park, J. H.; Park, H. I.; Sung, J. M.; Jang, Y.; Chung, N.; Hwang, K. C.; Kim, T. W., Cordycepin inhibits lipopolysaccharide-induced inflammation by the suppression of NF-kappaB through Akt and p38 inhibition in RAW 264.7 macrophage cells. Eur J Pharmacol 2006, 545, 192-199.
Kim, Y. K.; Na, K. S.; Myint, A. M.; Leonard, B. E., The role of pro-inflammatory cytokines in neuroinflammation, neurogenesis and the neuroendocrine system in major depression. Prog Neuropsychopharmacol Biol Psychiatry 2016, 64, 277-284.
Knafo, S.; Esteban, J. A., PTEN: Local and Global Modulation of Neuronal Function in Health and Disease. Trends Neurosci 2017, 40, 83-91.
Labandeira-Garcia, J. L.; Rodriguez-Perez, A. I.; Villar-Cheda, B.; Borrajo, A.; Dominguez-Meijide, A.; Guerra, M. J., Rho Kinase and Dopaminergic Degeneration: A Promising Therapeutic Target for Parkinson''s Disease. Neuroscientist 2015, 21, 616-629.
Leff, M. J.; Roatch, J. F.; Bunney Jr, W. E., Environmental factors preceding the onset of severe depressions. Psychiatry 1970, 33, 293-311.
Lesch, K.-P.; Bengel, D.; Heils, A.; Sabol, S. Z., Association of anxiety-related traits with a polymorphism in the serotonin transporter gene regulatory region. Science 1996, 274, 1527-1531.
Liu, Z.; Li, P.; Zhao, D.; Tang, H.; Guo, J., Anti-inflammation effects of Cordyceps sinensis mycelium in focal cerebral ischemic injury rats. Inflammation 2011, 34, 639-644.
Lo, H.-C.; Tu, S.-T.; Lin, K.-C.; Lin, S.-C., The anti-hyperglycemic activity of the fruiting body of Cordyceps in diabetic rats induced by nicotinamide and streptozotocin. Life sciences 2004, 74, 2897-2908.
Lohoff, F. W., Overview of the genetics of major depressive disorder. Current psychiatry reports 2010, 12, 539-546.
Nishizawa, K.; Torii, K.; Kawasaki, A.; Katada, M.; Ito, M.; Terashita, K.; Aiso, S.; Matsuoka, M., Antidepressant-like effect of Cordyceps sinensis in the mouse tail suspension test. Biological and Pharmaceutical Bulletin 2007, 30, 1758-1762.
World Health Orgnization .DEPRESSION,A Global Public Health Concern. 2012.
Pariante, C. M.; Miller, A. H., Glucocorticoid receptors in major depression: relevance to pathophysiology and treatment. Biological psychiatry 2001, 49, 391-404.
Prasain, J. K., Pharmacological Effects of Cordyceps and Its Bioactive Compounds. 2013, 40, 453-468.
Qin, T.; Fang, F.; Song, M.; Li, R.; Ma, Z.; Ma, S., Umbelliferone reverses depression-like behavior in chronic unpredictable mild stress-induced rats by attenuating neuronal apoptosis via regulating ROCK/Akt pathway. Behavioural brain research 2017, 317, 147-156.
Ressler, K. J.; Bradley, B.; Mercer, K. B.; Deveau, T. C.; Smith, A. K.; Gillespie, C. F.; Nemeroff, C. B.; Cubells, J. F.; Binder, E. B., Polymorphisms in CRHR1 and the serotonin transporter loci: gene× gene× environment interactions on depressive symptoms. American Journal of Medical Genetics Part B: Neuropsychiatric Genetics 2010, 153, 812-824.
Rogers, J.; Li, S.; Lanfumey, L.; Hannan, A. J.; Renoir, T., Environmental enrichment reduces innate anxiety with no effect on depression-like behaviour in mice lacking the serotonin transporter. Behav Brain Res 2017, 332, 355-361.
Sapkota, K.; Kim, S.; Park, Y.; Choi, B.-S.; Park, S.-E.; Kim, S.-J., Enhancement of tyrosine hydroxylase expression by Cordyceps militaris. Open Life Sciences 2010, 5.
Shin, S.; Moon, S.; Park, Y.; Kwon, J.; Lee, S.; Lee, C. K.; Cho, K.; Ha, N. J.; Kim, K., Role of Cordycepin and Adenosine on the Phenotypic Switch of Macrophages via Induced Anti-inflammatory Cytokines. Immune Netw 2009, 9, 255-264.
Singh, R.; Wang, B.; Shirvaikar, A.; Khan, S.; Kamat, S.; Schelling, J.; Konieczkowski, M.; Sedor, J. R., The IL-1 receptor and Rho directly associate to drive cell activation in inflammation. The Journal of clinical investigation 1999, 103, 1561-1570.
Tianzhu, Z.; Shihai, Y.; Juan, D., Antidepressant-like effects of cordycepin in a mice model of chronic unpredictable mild stress. Evid Based Complement Alternat Med 2014, 2014, 438-506.
Torres, S. J.; Nowson, C. A., Relationship between stress, eating behavior, and obesity. Nutrition 2007, 23, 887-894.
Vesterinen, H. M.; Currie, G. L.; Carter, S.; Mee, S.; Watzlawick, R.; Egan, K. J.; Macleod, M. R.; Sena, E. S., Systematic review and stratified meta-analysis of the efficacy of RhoA and Rho kinase inhibitors in animal models of ischaemic stroke. Systematic reviews 2013, 2, 33-42.
Wang, J.; Dunn, A. J., Mouse interleukin-6 stimulates the HPA axis and increases brain tryptophan and serotonin metabolism. Neurochemistry international 1998, 33, 143-154.
Willner, P.; Muscat, R.; Papp, M., Chronic mild stress-induced anhedonia: a realistic animal model of depression. Neuroscience & Biobehavioral Reviews 1992, 16, 525-534.
Zhang, X.; Gainetdinov, R. R.; Beaulieu, J.-M.; Sotnikova, T. D.; Burch, L. H.; Williams, R. B.; Schwartz, D. A.; Krishnan, K. R. R.; Caron, M. G., Loss-of-function mutation in tryptophan hydroxylase-2 identified in unipolar major depression. Neuron 2005, 45, 11-16.
Zheng, L. P.; Gao, L. W.; Zhou, J. Q.; Sima, Y. H.; Wang, J. W., Antioxidant activity of aqueous extract of a Tolypocladium sp. fungus isolated from wild Cordyceps sinensis. African Journal of Biotechnology 2008, 7. 3004-3010