AMPK-PGC1α-TFAM通路在赭曲霉毒素A诱导胃粘膜上皮细胞氧化应激损伤及恶性转化中的作用

Ochratoxin A (OTA), a carcinogenic toxin mainly produced by ubiquitous Aspergillus and Penicillium, is one of the most abundant food-contaminating mycotoxins in the high incidence area of gastric carcinoma in China. Our previous study showed that long-term OTA exposure could induce the malignant transformation in human gastric epithelial cell line (GES-1). And the chronic oxidative stress damage plays an important role in gastric tumorigenesis. However, there were no available data regarding the molecular mechanism of OTA-induced oxidative stress damage up to now. In this study, we firstly observed the mitochondrial structure and respiratory function, and in order to evaluate the mitochondrial damage and the level of ROS of GES-1 cells after long-term OTA exposure. Furthermore, choose TFAM as the key points, we deeply explored that the relationship between TFAM and mitochondrial respiratory function in the process of OTA-induced chronic oxidative stress damage and malignant transformation of GES-1 cells by immunofluorescence technique and so on. In addition, we further explored the molecular regulation mechanism of AMPK-PGC-1α-TFAM signaling pathway in OTA-induced chronic oxidative stress of GES-1 cells by biological molecular techniques. Finally, we observed that the effect of OTA-induced chronic oxidative stress damage on gastric epithelial cell and the possible molecular mechanism by the establishment of animal moldel. The study will provide a scientific basis for revealing the possible mechanism of gastric carcinogenesis in the OTA-exposed populations.

赭曲霉毒素A（OTA）是曲霉菌和青霉菌等产生的致癌性真菌毒素, 是我国胃癌高发区粮食中一种主要污染真菌毒素。我们前期研究发现暴露OTA可诱发人胃粘膜上皮细胞（GES-1) 恶性转化，而长期氧化应激损伤在这个过程中起着重要的作用。但是OTA引起细胞氧化应激损伤的发生机制并不清楚。本项目拟通过对线粒体结构和氧化呼吸功能的观察，评价OTA暴露对线粒体的损伤及ROS生成情况。然后选择线粒体转录因子A （TFAM）作为研究重点，利用免疫荧光等技术探讨OTA诱导GES-1细胞恶性转化过程中TFAM与线粒体氧化呼吸功能及氧化应激损伤之间的关系。进一步利用分子生物学方法探讨AMPK-PGC-1α-TFAM信号通路对OTA诱导细胞氧化应激损伤的分子调控机制；最后建立动物模型从整体水平观察OTA对胃粘膜上皮细胞的氧化应激损伤作用及分子机制。本研究为进一步揭示OTA与胃癌发生的关系及可能机制提供科学依据。

赭曲霉毒素A（Ochratoxin A, OTA）是一种世界范围内常见的由青霉菌属和曲霉菌属产生的真菌毒素，是一种可能的人类致癌物。我们团队前期研究显示OTA长期作用可以诱导人胃黏膜上皮细胞（GES-1）发生恶性转化。然而，这其中可能的分子机制并不清楚。通过本项目的研究，我们发现OTA作用于GES-1细胞，造成细胞内ROS升高，抗氧化酶表达下降，线粒体膜电位下降，ATP生成减少。同时，OTA可以激活线粒体凋亡途径。非常有趣的是，我们的研究还发现OTA作用于GES-1细胞后可以激活AMPK/PGC1α/NRF1/TFAM通路，促进了线粒体生物合成。另外我们的研究还发现OTA作用于GES-1细胞后可以改变细胞的代谢途径，引起碳代谢途径相关酶的变化，进而使氧化磷酸化向无氧糖酵解转变。最后，我们通过2-D电泳技术进一步发现 Annexin A3在OTA诱导的GES-1细胞恶性转化中发挥了重要的作用。本项目丰富和加深了人们对OTA生物效应的认识，为揭示胃癌高发区居民胃癌发生机制以及寻找合理防治措施提供了科学依据。