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研究生:簡民惠
研究生(外文):Min-Hui Chien
論文名稱:木質酚K8保護神經細胞對抗氧化壓力機制與抗老化作用之探討
論文名稱(外文):The investigation of the neuroprotection mechanism of K8 against hydrogen peroxide injury and the anti-aging effect.
指導教授:陳怡伶陳怡伶引用關係
指導教授(外文):Yi-Lin Chen
學位類別:碩士
校院名稱:國立宜蘭大學
系所名稱:生物技術研究所碩士班
學門:農業科學學門
學類:畜牧學類
論文種類:學術論文
論文出版年:2009
畢業學年度:97
語文別:中文
論文頁數:68
中文關鍵詞:神經保護抗氧化延緩老化木質酚第二型環氧酶
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前人的研究結果指出細胞中過氧化反應物的累積為許多神經退化性疾病及中風致病過程中常可觀察到的現象,這些累積的過氧化反應物質會開始氧化蛋白質、核酸及脂質,並且活化細胞凋亡路徑。同時,氧化壓力也會刺激細胞中第二型環氧酶 (COX-2) 的表現量促使神經細胞走向死亡。因此,若能透過降低細胞的氧化壓力來減少神經細胞的死亡,將可達到保健腦力之效果。異柴胡內酯 (isochaihulactone, K8) 為中草藥南柴胡之萃取物,結構上屬於木質酚類,在先前研究已發現其具有抗肺癌細胞生長之效果,而同時有許多的研究也指出木質酚類這類分子,如芝麻素或芝麻林素等,都具有很好的抗氧化效果亦有保護神經之能力。此外,本實驗室也曾於 primary cortical neuron cell 上,發現 K8 能有效對抗缺氧傷害。因此本論文以添加高劑量的過氧化氫於分化的 PC12 細胞,體外模擬高氧化壓力傷害,並事先給予K8 觀察有無神經保護效果。由結果顯示, K8 能有效提高細胞存活率並降低過氧化氫的細胞毒性,並抑制細胞凋亡,同時預先給予 K8 ,可以有效地降低細胞內活性氧物質 (ROS) 及脂質過氧化反應終產物丙二醛 (MDA) 的含量,並且會提高超氧歧化酶 (SOD) 與穀胱甘肽過氧化酶 (GPx) 之活性,同時 K8 亦可透過抑制 NF-B 的表現量來降低 COX-2 之 mRNA 及蛋白質表現量。另ㄧ方面,自由基的產生與老化的過程亦有明顯的關聯。因此,使用慢性半乳糖注射導致生物體過氧化之老化體內模式,觀察活體中 K8 有無減緩衰老之功效。由結果顯示半乳糖老化模式明顯提高血清中老化相關的生化指標,而 K8 則可以延緩這些老化指標的出現。綜合以上結果顯示 K8 可能透過其抗氧化能力來達到神經保護,且可透過調控 NF-B 的表現量來降低 COX-2 基因表現,同時K8 也可延緩半乳糖導致的老化現象,因此,期望能於未來發展成延緩老化及保健腦力之健康產品。
A number of studies indicate that reactive oxygen species (ROS) are involved in neurodegenerative diseases, such as Alzheimer’s disease (AD) and Parkinson’s disease (PD). Oxidative stress may through the activation of apoptosis and inflammatory reaction to induce neuron death. Besides ROS could through the induction of cyclooxygenase-2 (COX-2) to activate inflammatory reaction. Here we investigated the protective role of isochaihulactone (K8), a lignan compound of Bupleurum scorzonerifolium, against the cytotoxicity induced by hydrogen peroxide (H2O2) in the differentiated PC12 cell line. The protective effects of K8 (5, 10 µM) on H2O2 -induced cytotoxicity may be ascribed to its anti-apoptotic effect and the antioxidative properties by reducing the intracellular reactive oxygen species (ROS), decreasing malondialdehyde (MDA) production, a common index of lipid peroxidation, and enhancing the antioxidant enzymatic activities of superoxide dismutase and glutathione peroxidase. Additionally, K8 could decrease the expression of COX-2 through the down - regulation of NF-B to reduce lipid peroxidation after H2O2 exposure. These findings suggested that K8 may be a potential neuroprotective agent.In other concept, Chronic systemic exposure of mice, rats, and Drosophila to D-galactose (D-gal) causes the acceleration of senescence and has been used as an aging model. The aging mice model was induced by subcutaneous injection of D-gal (100 mg/kg) once daily for 6 weeks. K8 (10 mg/kg) was administered subcutaneous once daily for 6 weeks along with D-gal injection. The results indicated that subcutaneous injection of D-gal decreased SOD and GPx activities, and increased MDA contents in plasma and, subcutaneous injection of K8 (10 mg/kg) for 6 weeks significantly elevated T-SOD and GPx activities, decreased MDA contents in plasma. The results suggest that K8 has potent anti-aging effects on aging mice induced by D-gal through antioxidative mechanisms.
目錄
中文摘要………………………………………………………………Ⅰ
英文摘要………………………………………………………………Ⅱ
目錄……………………………………………………………………Ⅲ
圖表目錄………………………………………………………………Ⅴ
縮寫表…………………………………………………………………Ⅵ
第一章、前言…………………………………………………………1
第一節、活性氧物質 (ROS)…………………………………………1
第二節、抗氧化防禦系統……………………………………………2
第三節、氧化壓力與神經損傷………………………………………7
第四節、第二型環氧酶與神經傷害…………………………………13
第五節、氧化壓力與老化……………………………………………16
第六節、異柴胡內酯 (isochaihulactone, K8)……………………19
第二章、實驗目的……………………………………………………22
第三章、實驗儀器與藥品試劑………………………………………23
第一節、實驗儀器……………………………………………………23
第二節、藥品試劑……………………………………………………24
第四章、實驗方法……………………………………………………27
第一節、實驗流程……………………………………………………27
第二節、PC12 離體神經細胞模式…………………………………28
第三節、半乳糖老化模式……………………………………………39
第四節、統計分析……………………………………………………40
第五章、結果…………………………………………………………41
第一節、PC12 之神經分化流程……………………………………41
第二節、細胞存活率及細胞毒性分析結果…………………………41
第三節、細胞凋亡分析結果…………………………………………41
第四節、活性氧物質及丙二醛含量分析結果………………………42
第五節、抗氧化酵素活性分析結果…………………………………42
第六節、環氧酶及其上游基因之表現量……………………………43
第七節、半乳糖老化小鼠血漿中老化生理指標分析結果…………43
第六章、討論…………………………………………………………44
第七章、結論與未來展望……………………………………………50
第八章、結果圖表……………………………………………………51
參考文獻………………………………………………………………60
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