黄曲霉毒素B1（AFB1）的短期投喂对凡纳滨对虾肠道黏膜屏障的影响

为探究黄曲霉毒素B1(AFB1)对凡纳滨对虾肠道黏膜屏障的影响,以投喂含有15 mg/kg AFB1饲料的凡纳滨对虾作为实验组,不含AFB1饲料投喂的凡纳滨对虾作为对照组,分别于第2、4、8、12天取肠道组织,采用实时荧光定量PCR检测mTOR信号通路中的真核细胞始动因子4E结合蛋白(eif4ebp),真核翻译起始因子1a(eif4e1a),真核翻译起始因子2(eif4e2)和核糖体s6蛋白激酶(p70s6k)基因,与免疫相关的转录因子Dorsal和Relish基因,酚氧化酶原(proPO)基因以及黏蛋白样围食膜因子(mucin-like PM)基因表达水平的变化,利用组织切片技术研究AFB1对肠道组织形态的影响。结果发现,AFB1的添加会对mTOR通路相关基因的表达产生影响,实验组对虾eif4ebp基因自第2天起发生显著上调,此后呈现先升高后降低的趋势;eif4e2和eif4e1a基因皆在第8和第12天被显著抑制;p70s6k基因在第2和第4天呈现下调趋势,并于第12天回升至初始水平。AFB1同时也刺激了免疫系统的响应,实验组Dorsal基因和Relish基因均被显著诱导,并呈现先增高后降低的趋势;proPO基因于第4和第8天显著上调并于第12天回落至初始水平;mucin-like PM基因在第2、4、8天均显著上调。AFB1的添加也破坏了凡纳滨对虾肠道正常的组织形态,出现上皮细胞核肥大、边缘模糊、上皮细胞层部分脱落等现象。研究表明,AFB1严重影响凡纳滨对虾肠道黏膜的屏障功能,不仅对肠道黏膜造成机械损伤,同时对肠道化学屏障和免疫屏障产生影响。

Effects of short term addition of aflatoxin B1(AFB1) on the intestinal mucosal barrier of Litopenaeus vannamei

In order to determine the impact of aflatoxin B1(AFB1) on the intestinal mucosa of Litopenaeus vannamei, shrimps were fed with normal diet and diet containing 15 mg/kg of AFB1 for 12 days, and the tissues of intestine were collected on the day 2, 4, 8 and day 12 of the experiment. The relative expression of genes in mTOR signal pathway including eif4ebp, eif4e1a, eif4e2 and p70s6k, the immune-related genes including Dorsal, Relish and proPO and mucin-like PM gene were quantified with RT-qPCR. The changes of intestinal morphology were explored under microscope. The results showed that, the expression of genes in mTOR signal pathway was affected by AFB1, eif4ebp gene was up-regulated from the 2 nd day, and decreased after reaching its peak on the 8th day; eif4e2 and eif4e1a genes were both inhibited on the 8th and 12th day; p70s6k gene was down-regulated from the 2rd to 8th day, and climbed to the initial level on the day 12. The immune response of the intestine was also stimulated by AFB1, Dorsal and Relish genes were induced significantly after fed with AFB1; proPO gene was up-regulated on the day 4 and day 8, followed by a fall to the initial level on the day 12; and the mucin-like PM gene was also significantly induced on the day 2, 4 and day 8. The morphology of shrimp gut was changed by AFB1, the nucleus of epithelial cells swelled, the clear margin of chromatin was no longer present, and the ecclasis of epithelial cell layer occurred. In conclusion, the function of intestinal mucosal barrier was affected by AFB1, not only the mechanical barrier was damaged, but also the chemical and immunologic barriers were impaired.