利用转录组测序筛选低切应力作用下人脐静脉内皮细胞的差异表达基因

Screening of Differentially Expressed Genes in hUVECs under Low Shear Stress by RNA-Seq

目的:通过转录组测序分析获得不同切应力作用下人脐静脉内皮细胞的基因的表达谱,为进一步探索切应力影响内皮细胞形态和功能的机制提供依据。方法:以人脐静脉内皮细胞为材料,通过Streamer系统建立6通道可调控切应力的流体动力学细胞模型,以层流切应力(15 dynes/cm^2)为对照,以低切应力(0.1 dynes/cm^2)为实验组,分别加载细胞18 h。提取总RNA逆转录合成cDNA,建立文库,以二代测序平台Illumina HiSeq中进行扩增和测序。结果:序列比对结果显示,有19986个基因比对上,新转录本分析显示各组新转录本数约占总转录本数的50%。基因表达差异分析显示,较对照组,低切应力组表达上调基因983个,表达下调基因701个。GO分析显示,有18499个基因得到了归类注释,绝大多数基因富集到生物学过程。KEGG分析显示,富集Top20的信号通路与细胞周期、DNA复制和细胞分裂、细胞应激和凋亡等生物学过程相关。结论:低切应力作用不仅仅激活内皮细胞中细胞的增殖相关基因,同时也涉及到DNA损伤修复和凋亡相关基因。

Objective:This study is to obtain the gene expression profiles of human umbilical vein endothelial cells(hUVECs)under different shear stress by RNA-Sequencing(RNA-Seq),and provide evidence for further exploring the mechanism on morphological and functional changes in endothelial cells affected by flow shear stress.Methods:A 6-channel streamer system was used to establish the hydrodynamic cell model with adjustable shear stress in hUVECs.Laminar shear stress(15 dynes/cm^2)was used as control group and low shear stress(0.1 dynes/cm^2)was used as experimental group.All cells were loaded for 18 h respectively;total RNA was extracted and synthesized by reverse transcription.The library was constructed and sequenced using Illumina Hi Seq.Results:Sequence alignment showed that there were 19986 gene alignments,and the number of new transcripts in each group was about 50%of the total transcripts.Differential expression analysis of genes showed that,compared with control group,there were 983 up-regulated genes and 701 down-regulated genes in low shear stress group.18499 genes were classified and annotated,GO analysis showed that most of the genes were enriched into biological process.KEGG analysis showed that Top20 enrichment signal pathway is related to cell cycle,DNA replication,cell division,cell stress and apoptosis.Conclusion:Low shear stress not only activates proliferation-related genes in endothelial cells,but also involves DNA damage repair and apoptosis-related genes.