黑木耳多糖（SNAAP）辐射防护作用的microRNA调控机制研究

MicroRNA is a class of small non-coding RNA playing important roles in various biological processes including cellular response to radiation as a powerful regulator of messenger RNA. Epidemiological, clinical, and animal studies support a role of polysaccharides in the prevention of various chronic diseases. Antioxidant and anti-radiation activity for SNAAP (Sulfate of neutral Auricularia auricular polysaccharides) are mainly concentrated on biological effects in vitro or in vivo. Our results showed that SNAAP can improve the expression of many important enzymes involved in oxidative stress response such as superoxide dismutase, catalase, and at the same time reduce the expression of marrow peroxidase. Likewise, SNAAP can increase the amount of reduced glutathione and reduce the amount of malondialdehyde in tested organs. Consequently, SNAAP can activate the anti-oxidase system, inhibit the oxidase system, alleviate lipid peroxidation, reduce damage to cell membrane, and decrease the mutation of the micronucleus rate and of chromosomes aberrations. However, the regulatory functions of microRNA in the radioprotective effects of SNAAP, so far, lacks research. Basing on our preliminary findings, we are going to investigate the differential expression profiles of microRNAs in 60Co γ-ray irradiated mice(Non-irradiated control, Irradiated control, SNAAP, SNAAP+Irradiated）and screen the specific microRNAs involved in the protective response of SNAAP against ionizing radiation. The results are expected to reveal the mechanisms underlying the protective effects of SNAAP against radiation by identifying the microRNAs and their target genes involved in this process.

microRNA是信使RNA强有力的调节子。流行病学和动物研究支持多糖在预防各种慢性病中起作用。多糖通过调控miRNA而发挥生理功能已有文献报道。黑木耳中性多糖片段硫酸酯（SNAAP）抗辐射活性研究主要集中在体外或体内生物效应，基于miRNA表达调控的SNAAP辐射防护作用机制研究未见报道。基于前期SNAAP激活机体内抗氧化酶系，提高机体免疫系统，调控细胞周期等途径而发挥辐射服防护作用这一研究结果，本研究采用深度测序技术对实验小鼠（对照组、辐射组、SNAAP组、SNAAP+辐射组）脾组织中microRNA进行高通量筛查，采用减法杂交，比较去掉表达量高的miRNA，找到影响抗辐射效果最明显的miRNA；实时定量PCR方法进行miRNA表达验证，通过靶基因预测软件，采用Western blot、qRT-PCR、双荧光素报告基因确定候选miRNA的靶基因，从分子水平阐释SNAAP抗辐射效应机理。

microRNA是信使RNA强有力的调节子。流行病学和动物研究支持多糖在预防各种慢性病中起作用。多糖通过调控miRNA而发挥生理功能已有文献报道。本研究采用深度测序技术对实验组样品同时进行miRNA-seq和mRNA-seq测序。首先制备完整、高纯度小鼠脾细胞RNA，对各实验组提取合格的小鼠脾脏总RNA进行测序，然后对得到的数据进行质控和筛选，并与相关参考基因组、miRNA数据库比对，进行分类注释，除去rRNA、tRNA等其他种类RNA，最终得到总miRNA。其中对于不同样品已经分析鉴定出已知miRNA 600种左右，并且预测出新miRNA 180种左右。且已完成对不同类型的miRNA进行差异表达分析、靶基因预测，进行靶基因GO富集分析，最终结合KEGG数据库进行靶基因代谢通路分析，构建miRNA-mRNA调控网络数据库，结合生物信息学靶标预测，获得黑木耳糖通过miRNA调控的mRNA，进行靶基因GO富集分析，结合KEGG数据库进行靶基因代谢通路分析，初步阐释SNAAP提高机体免疫系统而发挥辐射防护作用。.日常辐射防护将是我国面临的一个严重的公共卫生问题。食用菌不仅能满足人们对食物的要求，而且营养均衡、药食同源，是未来辅助辐射防护保健食品消费升级的重要方向。“十三五”期间，保健食品将是我国食品行业重要的支柱产业之一，因此开展高值、新型的高端食用菌保健品的高效加工关键技术，既有利于延长食用菌产业链、促进食用菌产业健康发展，又是促进我国“大健康”产业发展的重要组成部分。