摘要
目的探讨共生菌群对小鼠肺泡巨噬细胞长链非编码RNA(long non-coding RNA,LncRNA)表达的调控作用。方法分离纯化共生菌缺失小鼠及正常小鼠肺泡巨噬细胞(alveolar macrophages,AMs),利用LncRNA芯片技术筛选差异表达的LncRNA并进行生物信息学分析,利用荧光原位杂交(FISH)、RNA干扰等技术检测LncRNA-30162的亚细胞定位及其对RAW264.7细胞基因表达的调控。结果分选后的AMs纯度大于95%,共生菌缺失小鼠与正常小鼠相比较,其差异表达LncRNA共有634个(变化倍数≥2),上调363个(57.26%)、下调271个(42.74%);差异LncRNA的靶基因与免疫系统调控、细胞分化、趋化作用等密切相关;干扰LncRNA-30162的表达可以降低RAW264.7细胞中CCL24及Arg1的表达水平,干扰前CCL24及Arg1的表达水平分别为(420.23±56.25)μg/L、(2.63±0.31)×103U/L,与干扰后表达水平(218.70±31.45)μg/L、(1.24±0.21)×103U/L比较差异有统计学意义(t=5.416、6.409,P=0.006、0.003)。结论共生菌可以调控小鼠AMs LncRNA的表达,差异表达的LncRNA与多个基因功能分析分类和基因信号通路密切相关,LncRNA-30162可以调控RAW264.7细胞CCL24、Arg1的表达。
Objective To explore the role of commensal microbiota in the regulation of long non-coding RNA(LncRNA)expression in mouse alveolar macrophages(AMs).Methods AMs were separated from antibiotics-treated mice and normal mice and then were purified.LncRNA microarray technology was used to screen differentially expressed LncRNAs and conduct bioinformatics analysis.Fluorescence in situ hybridization(FISH)was used to detect the subcellular localization of LncRNA-30162.RNA interference technology was used to knock out the expression of LncRNA-30162 in RAW264.7 cells>and reverse transcription polymerase chain reation(RT-PCR)was used to detect the regulation of gene expression by LncRNA-30162 in RAW264.7 cells.Results The purity of the separated AMs was greater than 95%.Compared with normal mice,there were 634 differentially expressed LncRNAs with changes greater than 2 folds in the AMs from antibiotics-treated mice,363 of which were upregulated and 271 were downregulated.The target genes of differentially expressed LncRNAs were closely associated with immune system regulation,cell differentiation and chemotaxis.The expression levels of CCL24 and Argl in RAW264.7 macrophages were decreased after interference with LncRNA-30162 expression[(218.70±31.45)μg/L vs.(420.23±56.25)μg/L,(l.24±0.21)×10^3U/L vs.(2.63±0.31)×10^3U/L,i=5.416 and 6.409,P=0.006 and 0.003].Conclusions Commensal microbiota can regulate the expression of LncRNAs in AMs.Differentially expressed LncRNAs are associated with a variety of gene ontology(GO)biological processes and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathways.LncRNA-30162 can regulate the expression levels of CCL24 and ARG1 in RAW264.7 cells.
作者
母英杰
陈稳
胡世莲
程民
Mu Yingjie;Chen Wen;Hu Shilian;Cheng Min(Department of Geriatrics,Anhui Province Hospital,Anhui Medical University,Gerontology Institute of Anhui Province,Anhui Provincial Key Laboratory of Tumor Immunotherapy and Nutrition Therapy,Hefei 230001,China)
出处
《中华老年医学杂志》
CAS
CSCD
北大核心
2020年第8期941-945,共5页
Chinese Journal of Geriatrics
基金
安徽省自然科学基金(2008085MH277)
国家自然科学基金(81471552)
肿瘤免疫与营养治疗安徽省重点实验室绩效补助(2019b12030026)。
关键词
共生菌
巨噬细胞
肺泡
RNA
Commensal microbiota
Macrophages,alveolar
RNA
