Liu Ya-Nan, Wang Yue-Xia, Liu Xiao-Fang, Jiang Li-Ping, Yang Guang, Sun Xian-Ce, Geng Cheng-Yan, Li Qiu-Juan, Chen Min, Yao Xiao-Feng
Department of Preventive Medicine, Dalian Medical University, 9 W Lvshun South Road, Dalian, 116044, PR China; Liaoning Anti-Degenerative Diseases Natural Products Engineering Research Center, 9 W Lvshun South Road, Dalian, 116044, PR China.
Department of Preventive Medicine, Dalian Medical University, 9 W Lvshun South Road, Dalian, 116044, PR China.
Toxicon. 2015 Mar;95:30-7. doi: 10.1016/j.toxicon.2014.12.014. Epub 2014 Dec 29.
Citreoviridin (CIT) is one of toxic mycotoxins derived from fungal species in moldy cereals. Whether CIT exerts hepatotoxicity and the precise molecular mechanisms of CIT hepatotoxicity are not completely elucidated. In this study, the inhibitor of autophagosome formation, 3-methyladenine, protected the cells against CIT cytotoxicity, and the autophagy stimulator rapamycin further decreased the cell viability of CIT-treated HepG2 cells. Knockdown of Atg5 with Atg5 siRNA alleviated CIT-induced cell death. These finding suggested the hypothesis that autophagic cell death contributed to CIT-induced cytotoxicity in HepG2 cells. CIT increased the autophagosome number in HepG2 cells observed under a transmission electron microscope, and this effect was confirmed by the elevated LC3-II levels detected through Western blot. Reduction of P62 protein levels and the result of LC3 turnover assay indicated that the accumulation of autophagosomes in the CIT-treated HepG2 cells was due to increased formation rather than impaired degradation. The pretreatment of HepG2 cells with the ROS inhibitor NAC reduced autophagosome formation and reversed the CIT cytotoxicity, indicating that CIT-induced autophagic cell death was ROS-dependent. In summary, ROS-dependent autophagic cell death of HpeG2 cells described in this study may help to elucidate the underlying mechanism of CIT cytotoxicity.
黄绿青霉素（CIT）是一种源自发霉谷物中真菌物种的有毒霉菌毒素。CIT是否具有肝毒性以及CIT肝毒性的确切分子机制尚未完全阐明。在本研究中，自噬体形成抑制剂3-甲基腺嘌呤可保护细胞免受CIT细胞毒性的影响，而自噬刺激剂雷帕霉素则进一步降低了CIT处理的HepG2细胞的活力。用Atg5 siRNA敲低Atg5可减轻CIT诱导的细胞死亡。这些发现提示了自噬性细胞死亡导致CIT对HepG2细胞产生细胞毒性这一假说。在透射电子显微镜下观察到CIT增加了HepG2细胞中的自噬体数量，通过蛋白质免疫印迹检测到的LC3-II水平升高证实了这一效应。P62蛋白水平的降低以及LC3周转分析结果表明，CIT处理的HepG2细胞中自噬体的积累是由于形成增加而非降解受损。用活性氧（ROS）抑制剂NAC预处理HepG2细胞可减少自噬体形成并逆转CIT细胞毒性，表明CIT诱导的自噬性细胞死亡是ROS依赖性的。总之，本研究中描述的HepG2细胞的ROS依赖性自噬性细胞死亡可能有助于阐明CIT细胞毒性的潜在机制。
