有機硒 N-γ-(L-glutamyl)-L-selenomethionine 改善老化、肥胖及帕金森氏症之病徵：以秀麗隱桿線蟲探索其機制

Abstract

硒為微量必須元素而無機硒的必須濃度區間十分的狹窄。日常食物中所攝取的硒主要為有機硒的形態。雖然大部分硒在生物體中之分子機制尚不明確，許多研究指出一些人類疾病都與病患體硒蛋白相關。在人類體中目前發現有25個基因負責轉譯硒蛋白，而TRXR-1是模式生物秀麗隱桿線蟲 (C. elegans) 體中唯一的硒蛋白。 本博士論文中我探討有機硒 (N-γ-(L-glutamyl)-L-selenomethionine, Glu-SeMet) 是如何參與於線蟲並調控以下三個生物效應。如何 (1) 改善老化與長壽；(2) 如何抑制肥胖；(3) 如何改善帕金森氏症 (Parkinson’s disease, PD) 之病徵且其上述之調控機制為何。 研究目標一，結果顯示經過Glu-SeMet (0.01 µM) 餵養之野生種線蟲顯著提升其抗氧化以及抗熱壓力之能力，但在延長壽命之試驗中卻沒有顯著的效果。此外， Glu-SeMet的餵養對於其長壽相關的試驗中與老化指標皆有顯著的改善，但在硒蛋白調控相關之trxr-1突變株線蟲中皆無法觀察到此一改善現象。由這些結果可以得知Glu-SeMet餵養之線蟲，在長壽的調控以及在過程中老化行為的改善機制硒蛋白TRXR-1參與於其中。 研究目標二，研究結果顯示無機硒Se(IV) 及Glu-SeMet皆能顯著的降低線蟲體內脂肪的含量。此外，在高糖飲食組別中我們可以發現，Glu-SeMet (0.01 µM) 可以降低大量誘導之脂質累積，與其他已知具減脂作用的resveratrol相比Glu-SeMet在0.01 µM濃度即可以顯著的降低線蟲體內脂質累積並顯著的降低線蟲油酸與硬脂酸之比例。然而在fat-6、fat-7與trxr-1突變株中皆無法觀察到抑制脂肪累積之效應，代表了stearoyl-CoA desaturases FAT-6與FAT-7參與Glu-SeMet抑制脂肪形成過程，此外硒蛋白TRXR-1亦參與其中。 研究目標三中，利用兩株基因轉殖線蟲進行帕金森氏症模式探討，其中BZ555攜帶綠色螢光蛋白於多巴胺神經；NL5901則表達人類α-synuclein於肌肉細胞。結果顯示Glu-SeMet (0.01 µM) 具有顯著的改善被6-OHDA或錳所誘導之多巴胺神經傷害；此外與帕金森氏症臨床用藥L-DOPA與Selegilin在相同濃度下進行比較，Glu-SeMet具有較佳之多巴胺神經保護功效。此外，當BZ555利用6-OHDA誘導，Glu-SeMet (0.01 µM) 顯著的提升GST-4與GCS-1之mRNA表達量，然而此提升效應則無法在進行skn-1 RNAi後觀察到。研究結果進一步顯示，在NL5901線蟲中，Glu-SeMet (0.01 µM) 能夠顯著的降低α-synuclein累積，然而此現象並未在缺失trxr-1基因之NL5901線蟲中觀察到。綜合以上結果，建議Glu-SeMet可能透過SKN-1與硒蛋白TRXR-1參與調控並改善帕金森氏症之效應。 總結來說，本博士論文提供有機硒Glu-SeMet對有益健康效應的新的面向，具有改善老化、肥胖及帕金森氏症的潛能。因線蟲和人類的基因具有高度的同源性，本博士論文建議硒可做為未來有益健康效應相關研究新的方向。

Selenium is an essential trace element but the essentiality window of inorganic selenium for organisms is narrow. The major daily intake of selenium is organic forms from food. Although molecular mechanisms or pathways, where selenium is involved, are little known, human diseases are known to link with selenoproteins. There are 25 genes encoding selenoproteins in human, yet TRXR-1 is the only selenoprotein identified in the nematode Caenorhabditis elegans (C. elegans). In my dissertation, I used the model organism C. elegans to investigate the mechanisms about how the organic selenium (N-γ-(L-glutamyl)-L-selenomethionine, Glu-SeMet) regulates the following three physiological events in C. elegans: how Glu-SeMet (1) ameliorates aging and improves longevity; (2) inhibits obesity, and (3) ameliorates Parkinson’s symptoms and their underlying mechanisms. In aim 1, results showed that Glu-SeMet (0.01 µM) significantly enhanced stress resistance of wild-type N2 worms under oxidative and thermal stress challenges; however, Glu-SeMet treatment did not extend the lifespan of wild-type N2 C. elegans. Glu-SeMet significantly increased the longevity of wild-type N2 C. elegans, but the phenomena were absent from the C. elegans selenoprotein trxr-1 null mutant worms. Furthermore, Glu-SeMet significantly ameliorated the aging indicators as results of improving longevity. Nevertheless, improving effects of longevity were absent from the trxr-1 null mutant worms. These findings suggest that the ameliorative longevity and aging indicators during aging process by Glu-SeMet in C. elegans are mediated via TRXR-1. In aim 2, results showed that both inorganic Se(IV) and Glu-SeMet have ability to reduce lipid droplets content in wild-type N2 C. elegans. In addition, Glu-SeMet (0.01 µM) reduced fat storage under high-glucose condition. Furthermore, comparing with reported compound (resveratrol), Glu-SeMet at 0.01 μM was enough to significantly reduce fat storage in both normal and high-glucose diet and altered the fatty acid composition. The ratio of oleic acid / stearic acid decreased in Glu-SeMet treatment group. Moreover, the anti-obesity effect was absent in fat-6, fat-7 and trxr-1 mutants, suggesting that FAT-6 and FAT-7 and TRXR-1 are involved in Glu-SeMet regulated fat storage in C. elegans. In aim 3, two transgenic strains: BZ555 expressing green fluorescent protein in dopaminergic neurons and NL5901 expressing human α-synuclein in muscle cells were employed as Parkinson’s disease (PD) model. Results showed that Glu-SeMet at 0.01 μM showed an ameliorative effect on improving dopaminergic neuron defects induced by 6-hydroxydopamine (6-OHDA) or manganese in strain BZ555. Furthermore, comparing with other PD clinical drugs (L-DOPA and Selegilin) at same treatment concentration, Glu-SeMet showed better ameliorative effects on 6-OHDA-induced DA neurons damage in strain BZ555. Glu-SeMet (0.01 μM) significantly increased mRNA level of SKN-1, GST-4, and GCS-1 in 6-OHDA-treateded BZ555 strain; however, the effects were absent with skn-1 RNA interference (RNAi). Furthermore, Glu-SeMet (0.01 μM) decreased α-synuclein accumulation in strain NL5901, whereas similar effect was not observed in the absence of trxr-1. Results from the present study suggest that anti-Parkinson’s symptoms by Glu-SeMet might be modulated via SKN-1 and TRXR-1 in C. elegans. In conclusion, this doctoral dissertation provides new aspects of organic selenium Glu-SeMet on the beneficial health effects, suggesting a potential for Glu-SeMet as a new treatment for aging, obesity, Parkinson’s disease or their complications. Since high genetically conserved pathways are shared between C. elegans and human, this study suggests selenium as a new direction in future research about beneficial health effects.