Zhang Keyu, Zheng Wenli, Zheng Haihong, Wang Chunmei, Wang Mi, Li Tao, Wang Xiaoyang, Zhang Lifang, Xiao Sui, Fei Chenzhong, Xue Feiqun
Key Laboratory of Veterinary Drug Safety Evaluation and Residues Research, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, No. 518 Ziyue Road, Shanghai, 200241, China,
Cell Biol Toxicol. 2014 Dec;30(6):313-29. doi: 10.1007/s10565-014-9287-0. Epub 2014 Sep 16.
Quinocetone, a new quinoxaline 1,4-dioxide derivative used in food-producing animals in China, exerts genotoxic effects on HepG2 cells. It triggers significant cytotoxicity and genotoxicity in vitro, but the detailed mechanism by which quinocetone induces adverse biological effects is not yet known. We analyzed the mechanisms behind quinocetone intoxication by investigating oxidative stress based on non-enzymatic and enzymatic antioxidant activities, and by identifying differentially regulated genes of HepG2 cells exposed to quinocetone using polymerase chain reaction (PCR)-based suppression subtractive hybridization to illustrate the toxicity mechanism of quinocetone. Meanwhile, the characteristics of oxidative stress and differentially regulated genes induced by quinocetone metabolites, 1,4-bisdesoxyquinocetone and 3-methylquinoxaline-2-carboxylic acid, were investigated too. Results showed that quinocetone damaged the antioxidant defense abilities of HepG2 cells by reducing the activities of endogenous antioxidant enzymes, lowering glutathione concentration, and elevating malondialdehyde level. We identified 160 quinocetone-responsive genes that were associated with cell proliferation, glucose metabolism, oxidative stress, and apoptosis, such as NAD(P)H dehydrogenase, quinone 1; and prolyl 4-hydroxylase, beta polypeptide. The expressions of some differentially regulated genes were confirmed by real-time reverse transcription-polymerase chain reaction. However, quinocetone metabolites showed little effects on HepG2 cells. These results showed that reactive oxygen species were the key mediators of quinocetone cytotoxicity in HepG2 cells and that c-MYC-dependent activation of the mitochondrial apoptotic pathway may be associated with quinocetone-induced toxicity.
喹烯酮是中国用于食用动物的一种新型喹喔啉 - 1,4 - 二氧化物衍生物，对HepG2细胞具有遗传毒性作用。它在体外引发显著的细胞毒性和遗传毒性，但喹烯酮诱导不良生物学效应的详细机制尚不清楚。我们通过基于非酶和酶抗氧化活性研究氧化应激，并使用基于聚合酶链反应（PCR）的抑制性消减杂交技术鉴定暴露于喹烯酮的HepG2细胞中差异表达的基因，来分析喹烯酮中毒背后的机制，以阐明喹烯酮的毒性机制。同时，还研究了喹烯酮代谢产物1,4 - 双脱氧喹烯酮和3 - 甲基喹喔啉 - 2 - 羧酸诱导的氧化应激特征和差异表达基因。结果表明，喹烯酮通过降低内源性抗氧化酶活性、降低谷胱甘肽浓度和升高丙二醛水平，损害了HepG2细胞的抗氧化防御能力。我们鉴定出160个与细胞增殖、葡萄糖代谢、氧化应激和凋亡相关的喹烯酮反应基因，如NAD(P)H脱氢酶醌1和脯氨酰4 - 羟化酶β多肽。一些差异表达基因的表达通过实时逆转录 - 聚合酶链反应得到证实。然而，喹烯酮代谢产物对HepG2细胞影响很小。这些结果表明，活性氧是喹烯酮在HepG2细胞中细胞毒性的关键介质，并且线粒体凋亡途径的c - MYC依赖性激活可能与喹烯酮诱导的毒性有关。
