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標題: | 亞慢性大氣懸浮微粒暴露對阿茲海默症三基因轉殖小鼠神經毒性之影響 Effects of sub-chronic exposure to ambient particles on CNS toxicity of Alzheimer's disease in 3×Tg-AD mice. |
作者: | Yi-Hsuan Chen 陳怡瑄 |
指導教授: | 鄭尊仁 |
關鍵字: | 大氣細懸浮微粒,神經毒性,阿茲海默症,氧化壓力,行為實驗, Atmospheric fine aerosols,neurotoxicity,oxidative stress,behavioral experiments, |
出版年 : | 2019 |
學位: | 碩士 |
摘要: | 空氣污染是現今頗受關注之問題,許多研究顯示暴露大氣懸浮微粒 (particulate matter, PM)會對心血管及呼吸系統產生等不良健康效應。最近,有許多研究接連指出暴露PM可能會影響中樞神經系統(central nervous system, CNS),造成包括阿茲海默症(alzheimer's disease, AD)等之神經退化性疾病(neuro-degenerative diseases)。AD於病理學上主要有兩種病徵:(1) 由乙型類澱粉蛋白(amyloid-beta, Aβ)沉積所形成之類澱粉蛋白斑塊(amyloid plaque);(2) 由過度磷酸化Tau蛋白(phosphorylated Tau protein, p-Tau)聚集而成的神經纖維糾結(neurofibrillary tangles, NFTs),上述兩種病徵造成神經細胞功能異常以及壞死,且研究指出氧化壓力、微膠細胞(microglia)活化及自噬作用(autophagy)的增加也扮演一定的角色。
目前大氣PM之毒理研究多使用濃縮微粒暴露系統,雖能提供較高之暴露濃度,但無法反映出真實大氣環境之暴露實態,故本研究使用臺北空氣污染暴露系統(Taipei Air Pollution Exposure System for Health, TAPES)進行動物實驗。同時,本研究使用6個月大之阿茲海默症三基因轉殖小鼠(triple-transgenic AD mice, 3xTg-AD mice)做為實驗動物,此品系能夠發展出與人類相似之阿茲海默症病徵。因此,我們希望藉由上述實驗動物模型來暸解暴露PM後是否會加速AD相關病理變化。 本研究利用6個月3×Tg-AD母鼠進行3個月全身連續性呼吸暴露於臺北大氣微粒後,腦組織分為嗅球(olfactory bulb)、小腦(cerebellum)、海馬迴(hippocampus)及皮質(cerebral cortex)以西方墨點法(western blot)測定AD病徵相關蛋白含總Tau、p-Tau、Aβ42,及微膠細胞活化指標Iba-1、自噬作用指標LC3B等蛋白。同時以LC-MS/MS量測丙二醛(malondialdehyde, MDA)藉以評估氧化壓力。犧牲前,透過兩種行為實驗(莫氏水迷津及滾輪式跑步機試驗)以此探討空間記憶、學習能力以及運動能力之改變。 本研究因小鼠之出生日期不同,而分為兩部分作為暴露。第一部分(108年10月1日至12月31日)暴露之PM2.5平均質量濃度為11.15μg/m3,第二部分(108年12月3日至109年3月3日)暴露之PM2.5平均質量濃度為11.60μg/m3。莫氏水迷津結果顯示在知識採集階段實驗,控制組和暴露組之間於皆無顯著差異。而在空間探索實驗的部分,在參數目標停留平臺象限時間(time spent in platform quadrant) 於兩組中有差異(p<0.05),然而暴露組停留時間高於控制組,其餘參數則皆無顯著差異。滾輪式跑步機結果顯示在第二天試驗時,掉落時轉速(speed at fall) (p<0.05)及掉落時間(latency to fall) (p<0.05)於控制組皆顯著低於暴露組。氧化壓力指標結果顯示MDA於嗅球(p<0.05)及海馬迴(p<0.05)在暴露3個月後有顯著的上升,其餘腦區皆有上升趨勢然而沒有達到顯著。在阿茲海默症蛋白質指標方面,Tau及LC3B蛋白,在四個腦區皆無顯著差異。p-Tau蛋白在嗅球暴露組顯著高於控制組(p<0.05)。Aβ42蛋白,在海馬迴暴露組顯著低於控制組(p<0.05)。Iba-1蛋白,在皮質暴露組顯著低於控制組(p<0.05)。 上述研究結果顯示亞慢性呼吸暴露大氣PM會誘導腦中的氧化壓力上升與p-Tau蛋白增加,且兩者在嗅球的部分具有顯著結果,顯見嗅球可能在PM暴露大腦之途徑扮演重要的角色。然而3個月大氣PM暴露並未對6個月3xTg-AD母鼠之空間學習、記憶功能及運動能力構成顯著影響。本研究發現呼吸暴露大氣PM對CNS有不一致的影響,然無法完整解釋誘發AD可能的機制及路徑,因此未來的研究需要透過延長暴露時間及使用老年小鼠來更進一步來釐清PM對CNS的影響。 Air pollution is a problem that has received much attention today. Many studies in the past have shown that exposure to atmospheric particulate matter (PM) can produce adverse health effects such as cardiovascular disease and respiratory system. To date, numerous studies have repeatedly pointed out that exposure to PM may affect the central nervous system (CNS), which in turn causes neurodegenerative diseases including Alzheimer's disease (AD). There are two main pathological signs of AD: (1) amyloid plaque formed by deposition of amyloid-beta (Aβ); (2) Neurofibrillary Tangles (NFTs) formed by the aggregation of phosphorylated Tau protein (p-Tau). These two changes cause neuronal dysfunction and necrosis, and studies have shown that the increase of oxidative stress, microglia activation, and autophagy in central nervous system also play a role. At present, most toxicological studies use concentrated particle exposure systems to exposure atmospheric PM. Although it can provide a higher concentration, it does not reflect the actual exposure of the real world. Therefore, this study will use the Taipei Air Pollution Exposure System for Health (TAPES) as an exposure device. At the same time, this study used 6-month-old triple-transgenic AD mice (3xTg-AD mice) as experimental animals. This strain can develop similar pathology of human AD. Therefore, we hope to understand the neurotoxicity of the CNS after exposure to PM by the above experimental animal model. In this study, 6-month-old 3×Tg-AD female mice were exposed for 3 months to atmospheric PM in Taipei. We measured the markers related to the pathology of AD (Tau, p-Tau, and Aβ42), the activation of microglia (Iba-1) and the autophagy (LC3B). Malondialdehyde (MDA) was also measured to assess the degree of oxidative stress. The above indicators were measured olfactory bulb, cerebellum, hippocampus and cerebral cortex. Before the sacrifice, the changes in spatial memory, learning ability and athletic ability were determined through two behavioral experiments (morris water maze and rotarod). The average concentration of PM2.5 in the first stage (108/10/01-108/12/31) was 11.15 μg/m3, and the second stage (108/12/03-109/03/03) was 11.60 μg/m3. The results of morris water maze showed that there was no significant difference between the control group and the exposed group in the knowledge acquisition stage. In the space exploration experiment, the time spent in platform quadrant had significant difference in the two groups. However, the exposed group was higher than the control group. In the rotarod test, speed at fall (p<0.05) and latency to fall (p<0.05) on the second day were significantly different. However, the control group was lower than the exposed group. The results of oxidative stress index showed that MDA in the olfactory bulb (p<0.05) and hippocampus (p<0.05) increased significantly after 3 months of exposure, and the other brain regions showed an upward trend, but the trend was not significant. In protein analysis, there were no differences in the four brain regions for the Tau and LC3B proteins. The p-Tau protein in the exposure group was significantly higher in the olfactory bulb (p<0.05). The Aβ42 of exposure group in the hippocampus was significantly lower than the control group (p<0.05). The Iba-1 of exposure group in the cerebral cortex was significantly lower than the control group (p<0.05). The results of the above studies show that sub-chronic inhalation exposure to atmospheric PM induces an increase in oxidative stress and an increase in p-Tau protein in the brain, both of which were significant in the olfactory bulb. It is obvious that the olfactory bulb may play an role in the translation of the PM into the brain. However, exposure to PM for 3 months did not significantly affect spatial learning, memory function, and exercise capacity of 6-month-old 3xTg-AD female mice. This study provides a supporting evidence linking the exposure to atmospheric PM and CNS toxicity. To further clarify the effects of PM on the CNS, future research is needed to extend the exposure time and use the aged mice. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/7275 |
DOI: | 10.6342/NTU201901973 |
全文授權: | 同意授權(全球公開) |
顯示於系所單位: | 職業醫學與工業衛生研究所 |
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