Transcriptome-wide profiling discover: PM2.5 aggravates airway dysfunction through epithelial barrier damage regulated by Stanniocalcin 2 in an OVA-induced model.
Transcriptome-wide profiling discover: PM2.5 aggravates airway dysfunction through epithelial barrier damage regulated by Stanniocalcin 2 in an OVA-induced model.
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DOI：
10.1016/j.ecoenv.2021.112408
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发表时间：
2021-06
影响因子：
6.8
通讯作者：
Lei Zhang;Xiang He;Ying Xiong;Q. Ran;Anying Xiong;Junyi Wang;Dehong Wu;Bin Niu;Guoping Li
中科院分区：
文献类型：
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作者：
Lei Zhang;Xiang He;Ying Xiong;Q. Ran;Anying Xiong;Junyi Wang;Dehong Wu;Bin Niu;Guoping Li
研究背景流行病学证据表明PM2.5暴露会加重哮喘，但其分子机制尚未完全阐明。方法构建卵清蛋白(OVA)诱导的PM2.5暴露小鼠。在体内研究中进行病理染色和免疫荧光。使用 U-BIOPRED 数据（人支气管活检）和 RNA-seq 数据（用 PM2.5 处理的 Beas-2B 细胞）进行基因集富集分析 (GSEA)，以确定与哮喘严重程度相关的通路。通过慢病毒转染、实时定量PCR、免疫荧光染色和跨上皮电阻（TEER）测量进行体外机制探索。结果PM2.5暴露加重了OVA诱导的小鼠气道炎症和粘液分泌。基于对人类支气管活检中的轻度至重度哮喘的转录组分析，基因集富集分析（GSEA）表明，上调的活性氧（ROS）途径基因集和下调的心尖连接基因集与哮喘严重程度相关。与轻至重度哮喘的分析一致，PM2.5暴露后，Beas-2B细胞中ROS通路上调，顶端连接下调。使用 qPCR 验证特定基因组中涉及的基因的表达水平。在暴露于 PM2.5 的细胞中，连接基因 ZO-1、E-cadherin 和 Occludin 的 mRNA 水平显着降低。此外，研究还证实，在暴露于 PM2.5 的 OVA 诱导小鼠中，抑制 ROS 可恢复 E-cadherin、Occludin 和 ZO-1 的表达水平，并改善气道炎症和粘液分泌。同时，PM2.5 升高了 ROS 水平。通过检查跨上皮电阻（TEER）值，我们还发现PM2.5暴露后上皮屏障受到破坏。重要的是，斯钙素2（STC2）被确定为调节上皮屏障的关键基因。结果表明，PM2.5 上调了 STC2 的表达，NAC 也恢复了 PM2.5 的表达。 STC2的过表达可以降低ZO-1、Occludin和E-cadherin的表达水平。相反，抑制STC2可以增加PM2.5降低的ZO-1、Occludin和E-cadherin的表达水平。结论通过转录组分析，我们发现STC2通过调节OVA诱导的小鼠上皮屏障，在PM2.5加重气道功能障碍中发挥关键作用。
BackgroundEpidemiologic evidence suggests that PM2.5 exposure aggravates asthma, but the molecular mechanisms are not fully discovered.MethodsOvalbumin (OVA)-induced mice exposed to PM2.5 were constructed. Pathological staining and immunofluorescence were performed inin vivostudy. Gene set enrichment analysis (GSEA) was performed to identify the pathway involved in asthma severity by using U-BIOPRED data (human bronchial biopsies) and RNA-seq data (Beas-2B cells treated with PM2.5). Lentiviruses transfection, Real-time qPCR, immunofluorescence staining and trans-epithelial electrical resistance (TEER) measurement were performed for mechanism explorationin vitro.ResultsPM2.5 exposure aggravated airway inflammation and mucus secretion in OVA-induced mice. Based on transcriptome analysis of mild-to-severe asthma from human bronchial biopsies, gene set enrichment analysis (GSEA) showed that up-regulated reactive oxygen species (ROS) pathway gene set and down-regulated apical junction gene set correlated with asthma severity. Consistent with the analysis of mild-to-severe asthma, after PM2.5 exposure, the ROS pathway in Beas-2B cells was up-regulated with the down-regulation of apical junction. The expression levels of genes involved in the specific gene sets were validated by using qPCR. The mRNA levels of junction genes,ZO-1,E-cadherinandOccludin, were significantly decreased in cells exposed to PM2.5. Moreover, it confirmed that inhibition of ROS recovered the expression levels of E-cadherin, Occludin and ZO-1, and ameliorated inflammation and mucus secretion in airway in OVA-induced mice exposed to PM2.5. Meanwhile, ROS level was elevated by PM2.5. By checking trans-epithelial electrical resistance (TEER) value, we also found that epithelial barrier was damaged after PM2.5 exposure. Importantly,Stanniocalcin 2 (STC2)was identified as a key gene in regulation of epithelial barrier. It showed that STC2 expression was up-regulated by PM2.5, which was recovered by NAC as well. Over-expression of STC2 could decrease the expression levels of ZO-1, Occludin and E-cadherin. Contrarily, suppression of STC2 could increase the expression levels of ZO-1, Occludin and E-cadherin reduced by PM2.5.ConclusionsBy using transcriptome analysis, we revealed that STC2 played a key role in PM2.5 aggravated airway dysfunction through regulation of epithelial barrier in OVA-induced mice.