酒精依赖患者血清microRNA的差异表达及靶向调控分析

Alcohol dependence is a serious social problem, but its pathogenes is still unknown,which leads to the lack of effective means in the diagnosis and treatment. Effort is made to provide a theoretical foundation for solving such problems by conducting research on the differential expression and targeted analysis of miRNA in the serum of male alcoholics in Heilongjiang Province of China.Research methods: MiRNAs of the observed group which consists alcoholics with different clinical are extracted for the construction of expression profile and its further analysis using the llumina GA∥x sequencing instrument.The extracted MiRNAs are identified by the method of Real-time PCR(RT- polymerase chain reaction) .MiRNAs were predicted using multiple target prediction algorithms and a consensus approach,and predicted targets were matched against differentially expressed mRNAs from the same samples.It is to go through the GO analysis and KEGG pathway analysis. The regulative relation between the specific target genes and miRNA with differential expressions could contribute to a network diagram of MiRNA and genes for further evaluation.The aim of the study was to assess whether it can be as a diagnostic miRNA alcohol dependence indicator of biology, and discusses the pathogenesis of alcohol dependence, looking for more effective for the treatment of the method.

酒精依赖是一个严重的社会问题，但目前其发病机制不清，缺乏有效地诊断治疗手段。本研究拟通过对黑龙江汉族男性酒精依赖患者血清miRNA差异表达及靶向分析的研究，为解决上述问题提供理论依据。近年microRNA的研究为各种复杂疾病的研究提供了新的方向。本研究对不同临床症状的酒精依赖患者的血清中miRNA应用llumina GA∥x测序仪进行测序，构建miRNA表达谱。利用Real-time PCR方法对差异表达的miRNA进行鉴定，选取三种靶基因预测软件对差异表达的miRNA进行靶基因预测，取交集作为最终miRNA的靶基因。对特异性靶基因进行GO分析和KEGG pathway分析。跟据分析结果构建miRNA与基因的调控网络图并进行评价。本研究旨在评定miRNA是否可以作为诊断酒精依赖的生物学指标,并探讨酒精依赖的发病机制,为治疗寻找更加有效的方法。

酒精依赖是目前国际上较为重要的公共卫生问题之一。近年来，有研究证实miRNAs与酒精依赖有关，其在调节神经系统的发育、神经可塑性以及大脑内环境的稳态等方面起到了重要的作用。但是目前研究结论不一，甚至还有一些miRNAs表达相反。可能与试验设计、酒精的浓度和暴露时间有关，研究证明长时间低浓度的酒精暴露比短时间高浓度的酒精暴露对神经细胞的损害更严重，酒精依赖戒断期与非戒断期的机制也不尽相同。我们应用国家自然基金经费主要完成三部分研究，包括：①酒精依赖大鼠持续饮酒期和戒断期前额叶皮质miRNAs的差异表达谱，根据检测结果，利用生物信息分析软件分析可能的靶基因。检测酒精依赖患者血浆中miRNAs的差异表达，并进行验证。研究发现酒精依赖组大鼠与对照组大鼠前额叶皮质脑区差异表达的miRNAs65个。持续饮酒组与戒断组表达不一致。BDNF、TPH1、MAPK3和NGFr是预测的可能与酒精依赖有关的靶基因。酒精依赖患者血浆中miRNAs的差异无显著性。结论是在酒精依赖的不同时期，miRNAs的作用机制有所不同。外周血检测miRNA未发现酒精依赖的标志物。②探讨自噬是否参与神经生长因子对酒精染毒的神经细胞的保护作用。研究发现NGF显著增加酒精染毒后PC12细胞内LC3-II蛋白的表达量，这种增加呈剂量依赖性。NGF显著减弱酒精染毒后PI3K/Akt/mTOR通路的磷酸化作用，而这种作用可被LY294002(Akt阻断剂)阻断。结论是神经生长因子通过PI3K/Akt/mTOR通路预防酒精诱导的神经毒性。③研究了中国东北和俄罗斯远东地区之间酒精相关问题的流行病学特点和差别。