真菌毒素(DON、AFB_1)对HepG2/C3A细胞联合毒性及机理研究  

Co-Effect Cytotoxicity and Mechansim Research of Deoxynivalenol and Aflatoxin B_1 in HepG2/C3A Cells

作　　者：李文竹[1] 张根义[1] 桑亚秋 LI Wenzhu;ZHANG Genyi;SANG Yaqiu(School of Food Science & Technology,Jiangnan University,Wuxi 214122,China)

机构地区：[1]江南大学食品学院,江苏无锡214122

出　　处：《食品与生物技术学报》2017年第11期1171-1179,共9页Journal of Food Science and Biotechnology

基　　金：国家农业部公益项目(201203069)

摘　　要：脱氧雪腐镰刀菌烯醇(DON)和黄曲霉毒素B_1(AFB_1)是一类常见的毒素类污染物,常共存于小麦、玉米等食品及制品中。实验采用不同浓度的DON(0.56、1.125、2.25、4.5、6.75μmol/L)和AFB_1(2.5、5、10、20、30μmol/L)单独及联合作用于HepG2/C3A细胞24 h后,分别测定细胞增殖抑制率(SRB)、双链DNA含量变化以及低浓度条件下细胞凋亡率和细胞周期分布,采用RTqPCR测定凋亡相关基因caspase-3、Bax以及Bcl-2 mRNA的相对含量。最后通过RNA-Seq测序分析2种毒素作用的毒性机理。结果显示,2种毒素单独及混合作用于HepG2/C3A细胞后使得细胞增殖抑制率增加,双链DNA含量下降,经分析后发现两者联合作用类型为加和作用。低浓度作用下均能引起细胞发生凋亡,使得细胞阻滞在S期或者S期和G2/M期,并且引起caspase-3和Bax表达量增加,Bcl-2则下降,从而引发细胞凋亡。RNA-Seq测序结果显示,经过差异基因的富集分析,AFB_1与DON的毒性作用机理并不相同,其中AFB_1以及两者的混合作用主要是通过P53信号通路影响细胞凋亡,而DON并不是。AFB_1与DON可能是通过2种不同的途径一起共同导致细胞凋亡的发生。Deoxynivalenol(DON)and Aflatoxin B1are the most prevalent mycotoxins in cereal-based ingredients or foods and they frequently coexist.HepG2/C3A cell treated with different concentrations of DON(0.56、1.125、2.25、4,5、6.75?滋mol/L)and AFB1(2.5、5、10、20、30?滋mol/L)for24h was used to analyze their toxic effects expressed by cell proliferation inhibition rate(SRB),total double-stranded DNA,apoptosis rate,cell cycle arrest and apoptosis-related gene,caspase-3,Bax,Bcl-2were analyzed by real-time quantitative PCR.The results showed that their co-effect was classified as being additive.Flow cytometry assay on the rate of apoptosis also shown an additive interaction which could indicated that these two toxins could induced apoptosis and they also induced cycle arrest in S-phase or S,G2/M-phase.The increase of expression of caspase-3and Bax as well as the decrease of Bcl-2further support their additive toxic effect.RNA-Seq analysis showed different cytotoxicity mechanism between AFB1and DON that AFB1and their combination effect induce cell apoptosis mainly through P53pathway while not for DON.We guess AFB1and DON may through different pathway to induce cell apoptosis together.

关 键 词：脱氧雪腐镰刀菌烯醇 黄曲霉毒素B1 联合毒性 RNA-Seq 

分 类 号：R99[医药卫生—毒理学]