Background

Epidemiologic evidence suggests that PM2.5 exposure aggravates asthma, but the molecular mechanisms are not fully discovered.

Methods

Ovalbumin (OVA)-induced mice exposed to PM2.5 were constructed. Pathological staining and immunofluorescence were performed in in vivo study. 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 exploration in vitro.

Results

PM2.5 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-cadherin and Occludin, 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.

Conclusions

By 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.

中文翻译：

---

全转录组分析发现：在 OVA 诱导的模型中，PM2.5 通过斯钙素 2 调节的上皮屏障损伤加重气道功能障碍

背景

流行病学证据表明，PM2.5 暴露会加重哮喘，但分子机制尚未完全发现。

方法

构建了暴露于 PM2.5 的卵清蛋白 (OVA) 诱导的小鼠。在体内研究中进行病理染色和免疫荧光。通过使用 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途径。进行慢病毒转染、实时 qPCR、免疫荧光染色和跨上皮电阻 (TEER) 测量以进行体外机制探索。

结果

PM2.5 暴露加剧了 OVA 诱导小鼠的气道炎症和粘液分泌。基于人支气管活检的轻度至重度哮喘的转录组分析，基因集富集分析 (GSEA) 显示上调的活性氧 (ROS) 通路基因集和下调的顶端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细胞内ROS通路上调，顶端连接下调。通过使用 qPCR 验证了参与特定基因集的基因的表达水平。连接基因、ZO-1、E-cadherin和Occludin的 mRNA 水平, 在暴露于 PM2.5 的细胞中显着减少。此外，它证实了 ROS 的抑制恢复了 E-cadherin、Occludin 和 ZO-1 的表达水平，并改善了 OVA 诱导的暴露于 PM2.5 的小鼠的气道炎症和粘液分泌。同时，ROS水平被PM2.5升高。通过检查跨上皮电阻 (TEER) 值，我们还发现 PM2.5 暴露后上皮屏障受损。重要的是，斯钙素 2 (STC2)被鉴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，而 NAC 也恢复了 PM2.5 的表达。STC2的过表达可降低ZO-1、Occludin和E-cadherin的表达水平。相反，STC2 的抑制可以增加被 PM2.5 降低的 ZO-1、Occludin 和 E-cadherin 的表达水平。

结论

通过使用转录组分析，我们发现 STC2 通过调节 OVA 诱导的小鼠的上皮屏障，在 PM2.5 加重气道功能障碍中发挥关键作用。