树莓花色苷对氨基甲酸乙酯的防控效应及机理研究

Ethyl carbamate (EC), a chemical contaminant widely presented in fermented foods, has been classified as Group 2A (probably carcinogenic to humans) by International Agency for Research on Cancer (IARC). Currently, with regard to toxicological study of EC, major attention have been paid to the carcinogenicity of EC in rodents. However, the toxicity of EC on human cells and its underlying mechanism remain elusive. Recently, some studies showed that EC-induced toxicity was associated with oxidative stress. Therefore, searching for effective and safe functional ingredients from foods, which can specifically target to EC-induced oxidative stress damage, might be a promising strategy to reduce the toxicity and potential health risk triggered by EC. Therefore, combined with the international food safety status and our preliminary data, this project is aimed to investigate the underlying mechanism of pelargonidin-3-O-glucoside (Pg3G) from raspberry in attenuating EC-induced toxicity, and to elucidate the protective effect of Pg3G against EC-induced oxidative stress damage through autophagy-lysosomal pathway. Further illuminating the molecular target of Pg3G on regulating autophagy-lysosomal pathway and finding the potential interaction site. The implementation of this project contributes to a better understanding of the protective mechanism of Pg3G against EC-induced toxicity as well as to improving the food safety theory of EC. Moreover, the completion of this project will also provide novel idea for the prevention of EC.

氨基甲酸乙酯（EC）是广泛存在于发酵食品中的化学污染物，已被国际癌症研究机构列为2A等级（很可能令人类致癌的物质）。目前关于EC的毒理学研究，主要集中在啮齿类动物的致癌研究，然而关于EC对人体细胞的毒性及毒理机制尚未完全阐明。近年来的研究发现EC对人体细胞的毒性与氧化应激损伤有密切的关系。因此，挖掘安全有效的食源性功效因子，靶向干预EC引发的氧化应激损伤，对于降低EC的毒性，减少EC对人体健康的潜在危害，将是行之有效的策略。鉴于此，结合当前国际食品安全形势与本课题的前期基础，本项目拟以树莓花色苷Pg3G防控EC毒性为核心内容，阐明Pg3G通过自噬-溶酶体途径防护EC所致氧化应激损伤的功效，进一步解析Pg3G调控自噬-溶酶体途径的分子靶标，探明其相互作用的位点。本课题的实施有助于人们更好地了解Pg3G防护EC毒性的作用机制，在一定程度上完善EC的食品安全理论，同时也可为EC的防控提供新思路。

氨基甲酸乙酯（Ethyl carbamate, EC）是发酵食品中常见的化学污染物，已被国际癌症研究机构（IARC）归类为Group2A等级，即很可能令人类致癌的物质。本课题组前期研究发现，EC能够引起肝脏细胞损伤，然而其具体的毒性效应与作用机制尚不明确。从果蔬中挖掘安全有效的食源性功能因子，对防控发酵食品中EC的潜在毒性及减少EC对人体健康的危害具有重要意义。基于此，本项目系统开展了EC诱导的肝脏毒性及花色苷防控EC诱导的毒性机制研究。首先采用人正常肝脏细胞L02构建氨基甲酸乙酯诱导的细胞氧化损伤模型，采用该模型研究发现EC能够诱导肝细胞氧化损伤，并激活TFEB介导的自噬过程缓解氧化还原稳态失衡；进一步结合细胞模型和动物模型阐明了EC通过抑制谷胱甘肽（GSH）合成途径和Nrf2活性进而诱导肝脏细胞和小鼠肝脏发生铁死亡。在明确EC毒性机制的基础上，进一步开展了花色苷防控EC毒性的机制研究。首先运用固相萃取、逆流色谱和制备液相等技术，从树莓、草莓、桑葚、蔓越莓、蓝莓等分离得到20种高纯度花色苷单体。采用EC诱导的细胞氧化损伤模型，筛选发现锦葵素-3-O-阿拉伯糖苷（M3A）能够显著降低EC处理后细胞内的氧化应激水平并提高GSH含量和细胞存活率，机理研究发现花色苷M3A通过激活腺苷酸活化蛋白激酶AMPK介导的自噬过程缓解EC引发的氧化损伤，并揭示了M3A与AMPK的互作机制。本项目的研究成果为食品或环境来源的EC暴露引发的健康问题及可能的防治措施提供了理论依据，也为后续开展EC的安全防控提供了新思路。