玉米赤黴烯酮對秀麗隱桿線蟲老化之毒性效應及相關機制探討

Abstract

近年來臺灣人民對於食品安全相關議題日漸關注，如何保障國人食之安全亦是相當需要重視的議題之一。而食品中黴菌毒素之汙染便是急需關注的食品安全議題。過去玉米赤黴烯酮之相關研究主要針對其生殖毒性之探討，而在生物體老化及相關機制之研究則相對較少，且過往研究主要以高濃度暴露玉米赤黴烯酮為主要研究設計，但在日常飲食中未必能暴露至如此高劑量，故本篇研究設計將貼近實際暴露情形，利用秀麗隱桿線蟲 (Caenorhabditis elegans) 為活體模式生物，探討較低濃度長期暴露玉米赤黴烯酮對生物體老化之影響，並進一步探討其分子調控相關機制。本研究分別就C. elegans之體長、運動行為 (body bends與head thrashes) 及體內活性氧化物質 (reactive oxygen species, ROS) 累積情形進行毒性評估，並依實驗結果挑選較具代表性之暴露濃度進行與老化相關之毒性效應及相關機制探討。實驗結果發現，暴露0.3、1.25、5、10及50 μM玉米赤黴烯酮至成蟲第0天，並不會影響C. elegans之體長，但上述濃度均能顯著對C. elegans之運動行為造成負面影響，且顯著增加C. elegans體內ROS之累積。當暴露1.25 及50 μM 玉米赤黴烯酮至成蟲第6或第8天，皆能顯著降低C. elegans之咽喉收縮率並對運動行為造成負面影響；暴露50 μM 玉米赤黴烯酮至成蟲第4、6或8天能顯著提高C. elegans體內ROS及脂褐素累積量、延長C. elegans之排泄週期，且玉米赤黴烯酮能明顯減少C. elegans壽命。同時，在相關機制探討部分發現，將daf-16 mutant暴露於1.25 及50 μM 玉米赤黴烯酮其咽喉收縮率之下降幅度高於野生種，且暴露1.25 及50 μM玉米赤黴烯酮會抑制DAF-16的入核。進一步的探討暴露50 μM玉米赤黴烯酮對於老化相關基因表達的影響發現，暴露至成蟲第0天時，hsp-16.1、hsp-16.49、hsp-70、daf-16及sod-3的基因表達量顯著上升，而ctl-1及ctl-2的表達量則顯著下降；暴露至成蟲第4天的結果則發現，hsp-16.49、hsp-70及sod-3的表達量顯著下降，而ctl-1及ctl-2的表達量亦顯著下降。綜合本研究之實驗結果，玉米赤黴烯酮的暴露，確實會促進C. elegans之老化，並影響insulin/IGF-1 signaling pathway轉錄因子DAF-16及其下游基因之表達。

Food safety issues have increasingly been a concerned topic in Taiwan in recent years. Among them, the contamination of mycotoxins in food needs urgent attention. Previous zearalenone related research has mainly focused on its reproductive toxicity. Studies related to its effects on aging and the underlying mechanism are relatively limited. In addition, previous studies were mostly conducted under high exposure concentrations of zearalenone. However, such high concentrations may not be a point of exposure in our daily diet. To reflect the actual exposure scenario, this present study aimed to investigate the toxic effects of long-term exposure to lower concentrations of zearalenone on aging and the underlying mechanism by using Caenorhabditis elegans as an in vivo model organism. We observed body length, locomotion (body bends and head thrashes), the accumulation of reactive oxygen species (ROS), aging markers, and investigated the aging-related mechanism after zearalenone exposure. The results showed that exposure to 0.3, 1.25, 5, 10 and 50 μM zearalenone did not affect the body length of C. elegans, but the above concentrations significantly reduced the body bends and head thrashes, and increased the accumulation of ROS in C. elegans. In addition, exposure to 1.25 and 50 μM zearalenone to day 4, 6, or 8 of adulthood significantly reduced the pharyngeal pumping rate and locomotion of C. elegans. Further, exposure to 50 μM zearalenone significantly increased the accumulation of ROS and lipofuscin, reduced the pharyngeal pumping rate, extended the defecation cycle, and shortened the lifespan of C. elegans. Moreover, exposure to 1.25 and 50 μM zearalenone further decreased the pumping rate of daf-16 mutant compared to N2, and inhibited the translocation of DAF-16 into the nucleus in C. elegans. We further explored the effects of exposure to 50 μM zearalenone on the expression of aging-related genes. The results showed that the expression levels of hsp-16.1, hsp-16.49, hsp-70, daf-16, and sod-3 genes were significantly increased, while the expression level of ctl-1 and ctl-2 were decreased on day 0 of adulthood C. elegans after 50 μM zearalenone exposure. Furthermore, the expression levels of hsp-16.49, hsp-70, and sod-3 genes were then significantly decreased on day 4 of adulthood C. elegans after 50 μM zearalenone exposure. The expression levels of ctl-1 and ctl-2 genes were also significantly decreased on day 4 of adulthood C. elegans after 50 μM zearalenone exposure. In conclusion, the exposure of zearalenone accelerated the aging process via the insulin/IGF-1 signaling pathway by inhibiting DAF-16 translocation and affecting downstream genes expression.