旋转细胞培养系统模拟微重力环境对小鼠成纤维细胞lncRNA表达的影响

目的研究旋转细胞培养系统（RCCS）模拟微重力环境对小鼠成纤维细胞株L929 lncRNA表达的影响。方法体外培养L929细胞,随机分为模拟微重力组（SMG组）和正常重力组（NG组）,每组3个样本。SMG组回转器轴心与地面平行旋转,NG组回转器轴心与地面垂直旋转,两组转速一致。RCCS培养7d,收集样本,提取样本总RNA,进行荧光标记和芯片杂交。利用Agilent Mouse lncRNA芯片分别检测SMG组和NG组L929细胞的lncRNA和mRNA表达,筛选差异表达显著的lncRNA,RT-q PCR验证芯片结果;利用GO和Pathway分析差异表达lncRNA的功能分布,结合mRNA差异...

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Published inJie fang jun yi xue za zhi Vol. 42; no. 10; pp. 876 - 882
Main Author 王占宇 姜福全 徐冰心 宋向卫 司少艳 周金莲 杨鹤鸣 李成林 崔彦
Format Journal Article
LanguageChinese
Published Beijing People's Military Medical Press 2017
100101,北京 安徽医科大学解放军306医院临床学院普通外科%100101,北京 解放军306医院普通外科%100101,北京 解放军306医院特种医学实验研究中心%100101,北京 解放军306医院病理科%100101 北京 安徽医科大学解放军306医院临床学院普通外科
100101 北京 解放军306医院普通外科
Subjects
Cell culture
Fibroblasts
lncRNA芯片
成纤维细胞
模拟微重力
lncRNA芯片
模拟微重力
fibroblasts
lncRNA chip
simulated microgravity
成纤维细胞
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ISSN0577-7402
DOI10.11855/j.issn.0577-7402.2017.10.07

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Summary:目的研究旋转细胞培养系统（RCCS）模拟微重力环境对小鼠成纤维细胞株L929 lncRNA表达的影响。方法体外培养L929细胞,随机分为模拟微重力组（SMG组）和正常重力组（NG组）,每组3个样本。SMG组回转器轴心与地面平行旋转,NG组回转器轴心与地面垂直旋转,两组转速一致。RCCS培养7d,收集样本,提取样本总RNA,进行荧光标记和芯片杂交。利用Agilent Mouse lncRNA芯片分别检测SMG组和NG组L929细胞的lncRNA和mRNA表达,筛选差异表达显著的lncRNA,RT-q PCR验证芯片结果;利用GO和Pathway分析差异表达lncRNA的功能分布,结合mRNA差异表达谱,进行lncRNA-mRNA联合分析。结果 lncRNA芯片检测分析发现,RCCS模拟微重力环境下小鼠成纤维细胞L929共有238条差异表达的lncRNA,其中134条表达上调,104条表达下调;差异表达的mRNA共有237条,其中53条表达上调,184条表达下调。获取差异表达lncRNA的聚类分析图,对差异表达显著的4条lncRNA芯片结果进行RT-q PCR验证,结果相吻合。GO分析结果显示差异表达的lncRNA与巨噬细胞分化、伤口愈合的负性调节等生物学过程相关,Pathway分析结果显示差异表达的lncRNA与系统性红斑狼疮、TGF-β等信号通路相关。同时成功构建了lncRNA-mRNA-TF可视化网络图。结论 RCCS模拟微重力环境显著影响L929细胞的lncRNA及mRNA表达谱,基于芯片技术的lncRNA靶基因预测和功能富集分析可为失重应激损伤机制探讨和修复措施建立提供理论依据。
Bibliography:Objective To investigate the effects of simulated microgravity by rotary cell culture system（RCCS） on expression profiles of long non-coding RNA（lncRNA） in mouse fibroblasts L929 cell line. Methods L929 cells were cultured in vitro and randomly divided into simulated microgravity（SMG） group and normal gravity（NG） group. Each group had three samples, the rotator axis of SMG group was paralleled to the ground rotation, while the rotator axis of NG group was vertical to the ground rotation, and the speed of rotation was consistent for the two groups. The samples of two groups were collected on 7 th day of culture and the total RNAs were extracted, labeled and hybridized in sequence. The lncRNA and mRNA were detected by Agilent Mouse lncRNA Chips respectively. Differentially expressed lncRNA were identified and then validated by RT-q PCR. GO and Pathway analysis were applied to determine the functional distribution of these target genes. The integration predictions of the lncRNA and mRNA co-expression had been pr
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ISSN:0577-7402
DOI:10.11855/j.issn.0577-7402.2017.10.07