- 學位論文
玉米赤黴烯酮對秀麗隱桿線蟲誘導帕金森氏症毒性及相關機制之探討
Evaluation of Zearalenone Toxicity on Parkinson’s Disease and the Underlying Mechanisms in Caenorhabditis elegans
摘要
玉米赤黴烯酮 (zearalenone) 為一種由鐮刀菌屬所產生的黴菌毒素，常見於燕麥、小麥、大麥與玉米中，因其結構與雌二醇相似，故過去研究多探討其生殖毒性；然而，對於其與神經退化性疾病的關聯與機制探討等相關研究則較缺乏。且臺灣在2018年由食品藥物管理署公布之《食品中污染物質及毒素衛生標準》中，亦新增zearalenone在食品裡的限量濃度，可見為亟需關注的食品安全議題之一。因此本研究以模式生物秀麗隱桿線蟲 (Caenorhabditis elegans, C. elegans)，探討暴露不同濃度zearalenone是否會造成生物體產生神經退化性疾病相關的病徵與其背後調控的機制為何。研究結果顯示，暴露1.25、10及50 μM zearalenone，會誘發C. elegans產生帕金森氏症相關的兩大病徵，一為顯著提高C. elegans體內α-突觸核蛋白 (α-synuclein) 的累積量，二為顯著增加C. elegans多巴胺神經元之退化情形；進一步觀察C. elegans與多巴胺神經元相關之食物敏感行為與游泳誘導麻痺等運動行為發現，在暴露1.25、10及50 μM zearalenone的條件下，會影響食物敏感行為；在10及50 μM zearalenone暴露下會提升游泳誘導麻痺的比例，證明zearalenone的暴露會產生與多巴胺神經元相關之神經毒性。本研究進一步分析細胞凋亡和細胞自噬途徑相關基因之表達量，顯示50 μM zearalenone的暴露下會活化細胞凋亡並抑制細胞自噬機制，同時亦觀察到zearalenone會造成C. elegans體內粒線體含量的累積與粒線體分裂比例提高。推測為細胞凋亡機制中的drp-1活化導致粒線體斷裂並加速細胞凋亡的發生，因此更進一步使用drp-1突變種C. elegans 進行探討，發現50 μM zearalenone暴露並不會對drp-1突變種造成食物敏感行為的改變；此外，應用drp-1 RNAi技術於轉基因C. elegans亦發現50 μM zearalenone暴露不會加重多巴胺神經元的損傷，顯示drp-1與zearalenone造成之神經毒性有直接關係。綜合本研究之結果，zearalenone的暴露會對C. elegans造成神經毒性，且會影響細胞自噬與細胞凋亡途徑基因的表達，並透過drp-1加速多巴胺神經元損傷的神經毒性效應。
並列摘要
Zearalenone is a mycotoxin produced by fungi such as the genus of Fusarium and has been detected in cereals, such as rice, wheat, and corn. Because its structure is similar to estrogens, previous research mainly focused on its reproductive toxicity. However, studies related to neurodegenerative diseases and underlying mechanisms are still limited. In addition, the maximum levels of zearalenone in foods are set in the “Sanitation Standard for Contaminants and Toxins in Food” by Taiwan Food and Drug Administration in 2018, so we are now paying more attention to it and regarding it as a global public health issue. Therefore, Caenorhabditis elegans (C. elegans) was used as an in vivo model organism to investigate whether exposure to zearalenone causes the hallmarks of Parkinson's disease and its related mechanisms in this present study. The results showed 1.25, 10 and 50 μM zearalenone would induce the aggregation of α-synuclein and enhance the degeneration of dopaminergic neurons in transgenic C. elegans. Further evidence indicated that zearalenone affected two dopamine-dependent behaviors, including basal slowing response after 1.25, 10 and 50 μM zearalenone exposure and swimming-induce paralysis after 10 and 50 μM zearalenone exposure. Moreover, we analyzed the expression of apoptosis and autophagy-related genes. Under effective concentration 50 μM zearalenone exposure, the apoptosis pathway was activated while the autophagy pathway was suppressed. Also, the accumulation of mitochondrial content and the higher percentage of mitochondrial fission were observed under 50 μM zearalenone exposure. We supposed that the activation of apoptosis-related gene, drp-1, would cause mitochondrial fragmentation and accelerate apoptosis pathway. We further found that the zearalenone induced deficit in basal slowing was completely abolished in drp-1 mutant strain and zearalenone did not cause further dopaminergic neurodegeneration after drp-1 RNAi treatment. This suggests the relationship between zearalenone induced neurotoxicity and drp-1. In conclusion, this study demonstrates that exposure to zearalenone results in neurotoxicity and affects the autophagy and apoptosis related genes expression, and accelerates the deficit of dopaminergic neurons via drp-1.