天津医药

天津医药 ›› 2020, Vol. 48 ›› Issue (1): 1-7.doi: 10.11958/20192370

• 细胞与分子生物学 •    下一篇

基于转录组测序探究PM2.5损伤肺上皮细胞 BEAS-2B的相关机制

胡玲娟 1，何翔 2，张雷 2，冉琴 2，李国平 1,2△   

  1. 基金项目：国家自然科学基金资助项目（81970026）；四川省卫生健康委员会重点科研课题（19ZD002）；四川省卫生和计划生育委员会科研 课题（18PJ012）；四川省科技计划项目（2018JY0380） 作者单位：1西南医科大学临床医学院（邮编646000）；2西南交通大学附属医院，成都市第三人民医院呼吸内科 作者简介：胡玲娟（1992），女，硕士研究生在读，主要从事PM2.5对呼吸系统疾病影响的相关研究 △通讯作者 E-mail: lzlgp@163.com
  • 收稿日期:2019-08-13 修回日期:2019-11-13 出版日期:2020-01-15 发布日期:2020-01-15
  • 通讯作者: 李国平 E-mail:lzlgp@163.com
  • 基金资助:
    四川省卫生健康委员会科研课题;四川省卫生和计划生育委员会科研课题;四川省科技计划项目

The mechanism of PM2.5 induced lung epithelial cell BEAS-2B injury based on RNA-sequencing

HU Ling-juan1, HE Xiang2, ZHANG Lei2, RAN Qin2, LI Guo-ping1,2△   

  1. 1 Department of Clinical Medicine, Southwest Medical University, Luzhou 646000, China; 2 Department of Respiratory Medicine, Chengdu Third People’s Hospital, the Affiliated Hospital, Southwest Jiaotong University △Corresponding Author E-mail: lzlgp@163.com
  • Received:2019-08-13 Revised:2019-11-13 Published:2020-01-15 Online:2020-01-15

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摘要： 目的 研究 PM2.5 对肺上皮细胞 BEAS-2B 基因表达的影响，并探讨其可能作用的信号通路。方法 BEAS-2B细胞分为对照组（PBS处理）和PM2.5组（200 mg/L PM2.5处理），干预24 h后提取各组细胞总RNA进行高通 量测序。所得的序列经质控，与参考基因组比对，获得用于后续转录本组装、表达量计算的 mapped data（reads）后，比 对到 6 大数据库［NR、Swiss-Prot、Pfam、COG（Evolutionary genealogy of genes）、GO（Gene Ontology）、KEGG（Kyoto Encyclopedia of Genes and Genomes）］注释，利用差异分析软件DESeq2筛选出差异表达基因（DEGs），运用生物信息学 分析方法对DEGs进行GO和KEGG生物功能等分析，采用qRT-PCR法检测5个关键基因（FOSB、FOSL1、MUC5AC、 CSF2、IL-6）进行验证。结果 PM2.5组和对照组与转录组的数据比较后共获得961个DEGs，其中PM2.5组上调表达 453个，下调表达508个。通过GO和KEGG功能分析后发现这些DEGs主要参与了细胞蛋白质的磷酸化、免疫调节过 程、信号转导途径的调节以及凋亡途径的调控过程，并显著富集于IL-17信号通路。qRT-PCR结果显示，PM2.5组 FOSB、FOSL1与IL-6的相对表达量显著上调（P＜0.05），这与RNA-seq测序结果一致。结论 PM2.5可能通过调控 “IL-17信号通路”中关键基因的表达，加重了细胞的炎症反应，促进了细胞凋亡等生命进程。

关键词: 转录组测序, PM2.5, 肺上皮细胞BEAS-2B, 差异表达基因, IL-17信号通路

Abstract: Objective To investigate the effect of PM2.5 on gene expressions of lung epithelial cell BEAS-2B and explore the signaling pathway. Methods BEAS-2B cells were divided into control group (PBS treatment) and experimental group (PM2.5 treatment). Total RNA of each group was extracted 24h after RNA-sequencing. After quality control, the sequence was compared with the reference genome. The mapped data (reads) were used for the assembly of subsequent transcripts, and the expression quantity calculation was obtained. And the results were analyzed in the following terms: NR, Swiss-Prot, Pfam, COG (Evolutionary genealogy of genes), GO (Gene Ontology), KEGG (Kyoto Encyclopedia of genes and genes) annotations in the six major databases. Differential expression genes (DEGs) were screened by DESeq2. Then, bioinformatics analysis was used to analyze the biological functions of DEGs, including GO and KEGG databases, and five key genes (FOSB, FOSL1, MUC5AC, CSF2 and IL-6) were detected by qRT-PCR for verification. Results The transcriptome data were compared between PM2.5 group and control group, and a total of 961 differentially expressed genes were obtained. Among them 453 genes were up-expressed and 508 genes were down-expressed. Through functional analysis of GO and KEGG, it was found that these differentially expressed genes were mainly involved in the regulation of protein phosphorylation, immune system process, positive regulation of signal transduction and apoptosis pathways. And they were significantly enriched in the IL-17 signaling pathway. qRT-PCR results showed that the relative expressions of key genes (FOSB, FOSL1 and IL-6) were significantly up-regulated in PM2.5 treatment group (P＜0.05), which was consistent with the results of RNA-seq. Conclusion It is possible that PM2.5 aggravates inflammatory response in beas-2b cells through regulating the key genes of the "IL-17 signaling pathway and promotes apoptosis.

Key words: RNA-seq, PM2.5, lung epithelial cell BEAS-2B, differential genes, IL-17 signaling pathway