锰致大鼠神经元中IncRNA与mRNA表达谱的改变及其生物信息学的分析
发布时间：2019-07-04 07:16
【摘要】：目的检测原代海马神经元经不同浓度锰染毒后其lncRNA及mRNA的表达情况,通过生物信息学的方法分析、预测并初步验证由哪些基因、lncRNA以及micro RNA可能形成ceRNA调控模型通路,为进一步研究lncRNA在锰致神经毒性中的可能作用提供线索。方法培养SD大鼠的原代海马神经元随机分成4组,至第4天分别以0、100、400、800μM浓度的氯化锰溶液染毒24小时,芯片检测海马神经元中lncRNA以及mRNA的表达情况,使用从聚法分析不同浓度染锰后原代海马神经元中mRNA和lncRNA的总体表达谱;火山图用于表示筛选出统计学差异大于等于2倍和p小于等于0.05的mRNA和lncRNA差异表达情况;GO分析用于将差异表达的mRNA进行功能归类注释;Pathway分析用于研究差异表达mRNA的代谢通路;交叉分析用于找出锰暴露后海马神经元中共同上调或下调的mRNA和lncRNA;利用lncRNA与mRNA的共表达网络并通过target scan、blast比对等工具预测可能的ceRNA通路,并通过Q-PCR检测相应基因、lncRNA及microRNA在不同浓度染锰后的大鼠神经元中的表达水平。结果(1)芯片测试结果表明,100μM组与0μM组相比,表达差异的mRNA共1848个,其中972个上调,876个下调;表达差异的lncRNA共566个,其中337个上调,229个下调(差异均大于2倍且p0.05)。400μM组与0μM组相比,表达差异的mRNA共3328个,其中1435个上调,1793个下调;表达差异的lncRNA共1161个,其中589个上调,572个下调(差异均大于2倍且p0.05)。800μM组与0μM组相比,表达差异的mRNA共4022个,其中1828个上调,2194个下调;表达差异的lncRNA共1474个,其中839个上调,635个下调(差异均大于2倍且p0.05)。(2)GO分析结果表明,差异表达的mRNA参与到生物途径、细胞定位以及分子功能中;pathway分析结果表明,100μM组与0μM组相比,最富集的信号通路是胰岛素分泌和细胞周期;400μM组与0μM组相比,最富集的信号通路是类风湿性关节炎和DNA复制;800μM组与0μM组相比,最富集的信号通路是胰岛素分泌和DNA复制。(3)交叉分析结果表明,和0μM组相比,100、400、800μM组海马神经元中共同上调的有135个lncRNA和373个mRNA,共同下调的有150个lncRNA和560个mRNA。(4)lncRNA与mRNA共表达分析结果表明,Pearson相关系数达98%以上的共表达相关组合共9个,选取基因Arsi并使用target scan检索到与其一致性分值最高的microRNA-125b,通过blast比对及预测工具发现基因Arsi,microRNA-125b,lncRNA-BC089928三者可能形成ce RNA调控通路。(5)Q-PCR验证结果表明,随着染锰浓度的升高,基因Arsi与lncRNA-BC089928的表达显著下调(p0.05),而microRNA-125b的表达则显著上调(p0.05),三者很可能形成ceRNA调控模型。结论不同浓度染锰后的海马神经元中存在不同数量表达差异的mRNA和lncRNA,并存在一定数量共同上调或下调的mRNA和lncRNA,且lncRNA-BC089928可能通过与microRNA-125b共同调控基因Arsi形成ceRNA调控通路,提示lncRNA可能在锰致海马神经元神经毒性中发挥了重要作用。
[Abstract]:Objective to detect the expression of lncRNA and mRNA in primary hippocampal neurons exposed to different concentrations of manganese, and to predict and preliminarily verify which genes, lncRNA and micro RNA may form ceRNA regulatory model pathway by bioinformatics analysis, so as to provide clues for further study on the possible role of lncRNA in manganese-induced neurotoxicity. Methods the primary hippocampal neurons of SD rats were randomly divided into 4 groups and exposed to 0 100400800 渭 M manganese chloride solution for 24 hours. The expression of lncRNA and mRNA in hippocampal neurons was detected by microarray. The overall expression profiles of mRNA and lncRNA in primary hippocampal neurons exposed to different concentrations of manganese were analyzed by polymerase chain reaction. The volcanic map was used to show the differential expression of mRNA and lncRNA with statistical difference greater than or equal to 2 times and p less than or equal to 0.05; GO analysis was used to classify and annotate the differentially expressed mRNA; Pathway analysis was used to study the metabolic pathway of differentially expressed mRNA; and cross analysis was used to identify mRNA and lncRNA; co-upregulated or down-regulated in hippocampal neurons after manganese exposure. The co-expression network of lncRNA and mRNA was used to predict the possible ceRNA pathway by target scan,blast ratio, and the expression levels of lncRNA and microRNA in rat neurons exposed to manganese at different concentrations were detected by Q-PCR. Results (1) the results of chip test showed that there were 1848 differentially expressed mRNA in 100 渭 M group compared with 0 渭 M group, of which 972 were up-regulated and 876 were down-regulated, of which 566 were up-regulated and 229 were down-regulated (the difference was more than 2 times and p0.05). Compared with 0 渭 M group, there were 3328 mRNA expression differences in 400 渭 M group, of which 1435 were up-regulated and 1793 were down-regulated. There were 1161 differentially expressed lncRNA, of which 589 were up-regulated and 572 were down-regulated (the difference was more than 2 times and p0.05). Compared with 0 渭 M group, there were 4022 mRNA expression differences in 800 渭 M group, of which 1828 were up-regulated and 2194 were down-regulated. There were 1474 differentially expressed lncRNA, of which 839 were up-regulated and 635 were down-regulated (the difference was more than 2 times and p0.05). (2). The results of GO analysis showed that the differentially expressed mRNA was involved in biological pathway, cell localization and molecular function, and the results of pathway analysis showed that insulin secretion and cell cycle were the most abundant signal pathways in 100 渭 M group compared with 0 渭 M group. Compared with 0 渭 M group, the most abundant signal pathways in 400 渭 M group were rheumatoid arthritis and DNA replication. Compared with 0 渭 M group, insulin secretion and DNA replication were the most abundant signaling pathways in 800 渭 M group. (3) the results of cross analysis showed that there were lncRNA and 373 mRNA, co-down-regulated mRNA. and mRNA. (4) lncRNA and mRNA co-expression in hippocampal neurons in 100400800 渭 M group compared with 0 渭 M group. The results showed that there were 9 co-expression related combinations with Pearson correlation coefficient above 98%. The gene Arsi was selected and the microRNA-125b, with the highest consistency score was found by blast comparison and prediction tools. (5) the results of Q-PCR verification showed that the expression of gene Arsi and lncRNA-BC089928 was significantly down-regulated (p0.05), while the expression of microRNA-125b was significantly up-regulated (p0.05) with the increase of manganese exposure concentration, and the results of Q-PCR verification showed that the expression of gene Arsi and lncRNA-BC089928 was significantly down-regulated (p0.05), while the expression of microRNA-125b was significantly up-regulated (p0.05). The three are likely to form ceRNA regulation model. Conclusion there are different quantities of mRNA and lncRNA, in hippocampal neurons exposed to different concentrations of manganese, and there are a certain number of mRNA and lncRNA, up-regulated or down-regulated by different concentrations of manganese. LncRNA-BC089928 may form ceRNA regulatory pathway by co-regulating gene Arsi with microRNA-125b, suggesting that lncRNA may play an important role in neurotoxicity of hippocampal neurons induced by manganese.
【学位授予单位】：广西医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R114
本文编号：2509731
[Abstract]:Objective to detect the expression of lncRNA and mRNA in primary hippocampal neurons exposed to different concentrations of manganese, and to predict and preliminarily verify which genes, lncRNA and micro RNA may form ceRNA regulatory model pathway by bioinformatics analysis, so as to provide clues for further study on the possible role of lncRNA in manganese-induced neurotoxicity. Methods the primary hippocampal neurons of SD rats were randomly divided into 4 groups and exposed to 0 100400800 渭 M manganese chloride solution for 24 hours. The expression of lncRNA and mRNA in hippocampal neurons was detected by microarray. The overall expression profiles of mRNA and lncRNA in primary hippocampal neurons exposed to different concentrations of manganese were analyzed by polymerase chain reaction. The volcanic map was used to show the differential expression of mRNA and lncRNA with statistical difference greater than or equal to 2 times and p less than or equal to 0.05; GO analysis was used to classify and annotate the differentially expressed mRNA; Pathway analysis was used to study the metabolic pathway of differentially expressed mRNA; and cross analysis was used to identify mRNA and lncRNA; co-upregulated or down-regulated in hippocampal neurons after manganese exposure. The co-expression network of lncRNA and mRNA was used to predict the possible ceRNA pathway by target scan,blast ratio, and the expression levels of lncRNA and microRNA in rat neurons exposed to manganese at different concentrations were detected by Q-PCR. Results (1) the results of chip test showed that there were 1848 differentially expressed mRNA in 100 渭 M group compared with 0 渭 M group, of which 972 were up-regulated and 876 were down-regulated, of which 566 were up-regulated and 229 were down-regulated (the difference was more than 2 times and p0.05). Compared with 0 渭 M group, there were 3328 mRNA expression differences in 400 渭 M group, of which 1435 were up-regulated and 1793 were down-regulated. There were 1161 differentially expressed lncRNA, of which 589 were up-regulated and 572 were down-regulated (the difference was more than 2 times and p0.05). Compared with 0 渭 M group, there were 4022 mRNA expression differences in 800 渭 M group, of which 1828 were up-regulated and 2194 were down-regulated. There were 1474 differentially expressed lncRNA, of which 839 were up-regulated and 635 were down-regulated (the difference was more than 2 times and p0.05). (2). The results of GO analysis showed that the differentially expressed mRNA was involved in biological pathway, cell localization and molecular function, and the results of pathway analysis showed that insulin secretion and cell cycle were the most abundant signal pathways in 100 渭 M group compared with 0 渭 M group. Compared with 0 渭 M group, the most abundant signal pathways in 400 渭 M group were rheumatoid arthritis and DNA replication. Compared with 0 渭 M group, insulin secretion and DNA replication were the most abundant signaling pathways in 800 渭 M group. (3) the results of cross analysis showed that there were lncRNA and 373 mRNA, co-down-regulated mRNA. and mRNA. (4) lncRNA and mRNA co-expression in hippocampal neurons in 100400800 渭 M group compared with 0 渭 M group. The results showed that there were 9 co-expression related combinations with Pearson correlation coefficient above 98%. The gene Arsi was selected and the microRNA-125b, with the highest consistency score was found by blast comparison and prediction tools. (5) the results of Q-PCR verification showed that the expression of gene Arsi and lncRNA-BC089928 was significantly down-regulated (p0.05), while the expression of microRNA-125b was significantly up-regulated (p0.05) with the increase of manganese exposure concentration, and the results of Q-PCR verification showed that the expression of gene Arsi and lncRNA-BC089928 was significantly down-regulated (p0.05), while the expression of microRNA-125b was significantly up-regulated (p0.05). The three are likely to form ceRNA regulation model. Conclusion there are different quantities of mRNA and lncRNA, in hippocampal neurons exposed to different concentrations of manganese, and there are a certain number of mRNA and lncRNA, up-regulated or down-regulated by different concentrations of manganese. LncRNA-BC089928 may form ceRNA regulatory pathway by co-regulating gene Arsi with microRNA-125b, suggesting that lncRNA may play an important role in neurotoxicity of hippocampal neurons induced by manganese.
【学位授予单位】：广西医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R114
【参考文献】
相关期刊论文 前1条
1 XIANG MengQing;LI ShengGuo;;Foxn4:A multi-faceted transcriptional regulator of cell fates in vertebrate development[J];Science China(Life Sciences);2013年11期
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