MicroRNA在AFB1致大鼠急性毒中的作用研究

发布时间：2018-05-22 14:47

本文选题：黄曲霉毒素 + AFB1　；参考：《福建农林大学》2012年硕士论文

【摘要】：黄曲霉毒素（Aflatoxin，AFT）主要由黄曲霉（Aspergillus flavel）等产生，AFT是真菌的次级代谢产物，属有毒物质。目前，已发现包括黄曲霉毒素B1（Aflatoxin B1，AFB1）在内的二十多种黄曲霉毒素，而黄曲霉毒素B1是其中含量较多且毒性较强的毒素之一。AFB1广泛存在于储存不当的食物或动物饲料中，它不仅对植物、微生物有伤害作用，对哺乳动物也有毒害作用，甚至可诱使癌症发生，如肝癌等。它对哺乳动物产生的毒害作用由剂量和作用时间共同决定，作用的主要靶器官是肝脏。AFB1的长期作用，，可能引起癌症的发生；它的短期作用则可损伤细胞、抑制细胞增殖或引起细胞凋亡。近来，人们发现微小RNA（microRNA，miRNA）是一类进化保守的非编码小RNA（non-coding smallRNA），它不仅在机体地生长、发育、分化和繁殖等正常代谢过程起作用，也在肿瘤的发生和转移等病变过程中扮演着重要的角色。我们试图研究microRNA在AFB1致大鼠急性毒中的作用。本研究将6只F344大鼠随机分为两组，每组各3只。实验组和对照组分别用AFB1和生理盐水，进行灌胃处理一次。在实验组中，AFB1的浓度为1.5mg/kg。3d后观察发现，实验组的大鼠进食减少、行动变缓、身体时而颤抖、尿液发黄、体重变轻。大鼠经乙醚麻醉后拉颈处死，收集血液，离心得到血清并进行肝功能测定。发现实验组大鼠血液中的乳酸脱氢酶、总胆红素、碱性磷酸酶、丙氨酸转氨酶和谷草转氨酶均显著上升，说明AFB1对大鼠有毒害作用。提取大鼠的肝脏总RNA，并进行小RNA高通量测序。通过小RNA高通量测序，我们发现有116个miRNAs发生显著性差异性表达，其中32个上调表达，84个下调表达。利用Stem-loop RT-qPCR验证小RNA测序结果，发现miR-34a发生显著性上调表达，与测序结果一致。最后，我们将mimic miR-34a转染到RH35细胞，可抑制其生长。说明miR-34a可能在AFB1致毒通路中起作用。
[Abstract]:Aflatoxin (AFT) is mainly produced by Aspergillus flavell et al. AFT is a secondary metabolite of fungi and is a toxic substance. At present, more than 20 aflatoxins, including aflatoxin B1(Aflatoxin B1-AFB1, have been found, and aflatoxin B1 is one of the most abundant and highly toxic toxins. AFB1 is widely found in poorly stored food or animal feed. It is not only harmful to plants and microorganisms, but also toxic to mammals, and even induces cancer, such as liver cancer. Its toxicity to mammals is determined by the dose and time of action. The main target organ is the long-term action of liver. AFB1, which may cause cancer, and its short term action can damage the cells. Inhibit cell proliferation or induce apoptosis. Recently, it has been found that microRNAs are a class of evolutionarily conserved small RNA(non-coding RNAs, which not only play a role in normal metabolic processes such as growth, development, differentiation and reproduction of the body. It also plays an important role in tumorigenesis and metastasis. We try to study the role of microRNA in acute toxicity induced by AFB1 in rats. In this study, 6 F344 rats were randomly divided into two groups, 3 rats in each group. The experimental group and the control group were treated with AFB1 and normal saline respectively. After the concentration of AFB1 in the experimental group was 1.5mg/kg.3d, it was found that the rats in the experimental group ate less, moved slowly, sometimes trembled, urine turned yellow and weight became lighter. After anesthesia with ether, the rats were killed by pulling their necks, blood was collected, serum was obtained by centrifugation and liver function was measured. It was found that the levels of lactate dehydrogenase, total bilirubin, alkaline phosphatase, alanine aminotransferase and alanine aminotransferase in blood of the experimental group were significantly increased, indicating that AFB1 had toxic effect on the rats. Total RNAs of rat liver were extracted and high throughput sequencing of small RNA was performed. By high throughput sequencing of small RNA, we found 116 miRNAs expressed differentially, including 32 up-regulated and 84 down-regulated. Stem-loop RT-qPCR was used to verify the sequencing results of small RNA, and it was found that the expression of miR-34a was up-regulated significantly, which was consistent with the result of sequencing. Finally, we transfect mimic miR-34a into RH35 cells and inhibit its growth. These results suggest that miR-34a may play a role in the toxic pathway of AFB1.
【学位授予单位】：福建农林大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R114

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