Air pollution may be associated with increased airway responsiveness to allergens in allergic rhinitis (AR). Ozone-aged environmental black carbon (O3BC) is an important constituent of atmospheric particulate matter (PM), for which the mechanisms underlying its effects have not been fully elucidated in AR. The objective of the present study was to determine the O3BC and pollen-induced alterations in the transcriptome in human nasal epithelial cells (hNECs) in vitro. hNECs from nasal epithelial mucosal samples of healthy individuals undergoing nasal surgery (turbinoplasty or septoplasty) were established as air–liquid interface (ALI) cultures and exposed to O3BC, pollen, or a combination of O3BC+ pollen. Changes in cell viability were analyzed by fluorescence and changes in the transcriptome by high-throughput RNA sequencing (RNA-seq). Several differentially expressed genes were verified by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Enrichment analysis, based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database, was performed to determine major biological functions and pathways involved. Exposure to ≥ 50 μg/ml O3BC or 25 μg/ml O3BC+ 200 μg /ml pollen significantly decreased cell viability of the hNECs compared to control (p < 0.05) or 25 μg/ml O3BC alone (p < 0.05); whereas exposure to pollen alone did not alter cell viability at any concentration investigated. High-throughput RNA sequencing analysis indicated that there was significant difference in gene expression between pollen or O3BC alone and O3BC+ pollen exposed cells. Exposure to 200 μg/ml O3BC was associated with hypoxia stress response GO terms, whereas exposure to 25 μg/ml O3BC+ 200 μg/ml pollen was associated with inflammatory response GO terms; including regulation of neutrophil migration and chemotaxis, macrophage differentiation and chemotaxis, mast cell activation, and phagocytosis. KEGG pathway analysis indicated the top 10 upstream regulators to be IL1B, CSF1, CCL2, TLR2, LPL, IGF8, SPP1, CXCL8, FCER1G and IL1RN; of which expressions of inflammation-related genes IL1B, CSF1 and FCER1G were significantly increased. O3BC and pollen allergen combined exposure may induce innate immune and allergic inflammation in hNECs, and therefore potentially exacerbate the symptoms of AR in affected individuals.

中文翻译：

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通过高通量转录组学评估暴露于臭氧老化的黑碳和花粉变应原的鼻上皮细胞遗传转录组的变化

空气污染可能与过敏性鼻炎（AR）中气道对过敏原的反应性增强有关。臭氧老化的环境黑碳（O3BC）是大气颗粒物（PM）的重要成分，其作用机理尚未在AR中得到充分阐明。本研究的目的是确定人鼻上皮细胞（hNECs）转录组中O3BC和花粉诱导的变化。来自健康个体经鼻外科手术（鼻成形术或隔膜成形术）的鼻上皮粘膜样本的hNECs被建立为气液界面（ALI）培养物，并暴露于O3BC，花粉或O3BC +花粉的组合中。通过荧光分析细胞活力的变化，并通过高通量RNA测序（RNA-seq）分析转录组的变化。通过逆转录定量聚合酶链反应（RT-qPCR）验证了几个差异表达的基因。进行了基于基因本体论（GO）和《京都议定书》的基因与基因组百科全书（KEGG）数据库的富集分析，以确定涉及的主要生物学功能和途径。与对照组（p <0.05）或单独25μg/ ml O3BC（p <0.05）相比，暴露于≥50μg/ ml O3BC或25μg/ ml O3BC + 200μg/ ml花粉显着降低了hNEC的细胞活力。而单独接触花粉并不能改变任何浓度的细胞活力。高通量RNA测序分析表明，单独的花粉或O3BC与暴露于O3BC +花粉的细胞之间的基因表达存在显着差异。暴露于200μg/ ml O3BC与缺氧应激反应GO项有关，而暴露于25μg/ ml O3BC + 200μg/ ml花粉与炎症反应GO术语相关；包括中性粒细胞迁移和趋化性，巨噬细胞分化和趋化性，肥大细胞活化和吞噬作用的调节。KEGG通路分析表明，排名前10位的上游调节因子为IL1B，CSF1，CCL2，TLR2，LPL，IGF8，SPP1，CXCL8，FCER1G和IL1RN。其中炎症相关基因IL1B，CSF1和FCER1G的表达显着增加。O3BC和花粉过敏原的联合暴露可能在hNEC中诱导先天性免疫和过敏性炎症，因此有可能加重受影响个体的AR症状。巨噬细胞的分化和趋化性，肥大细胞活化和吞噬作用。KEGG通路分析表明，排名前10位的上游调节因子为IL1B，CSF1，CCL2，TLR2，LPL，IGF8，SPP1，CXCL8，FCER1G和IL1RN。其中炎症相关基因IL1B，CSF1和FCER1G的表达显着增加。O3BC和花粉过敏原的联合暴露可能在hNEC中诱导先天性免疫和过敏性炎症，因此有可能加重受影响个体的AR症状。巨噬细胞的分化和趋化性，肥大细胞活化和吞噬作用。KEGG通路分析表明，排名前10位的上游调节因子为IL1B，CSF1，CCL2，TLR2，LPL，IGF8，SPP1，CXCL8，FCER1G和IL1RN。其中炎症相关基因IL1B，CSF1和FCER1G的表达显着增加。O3BC和花粉过敏原的联合暴露可能在hNEC中诱导先天性免疫和过敏性炎症，因此有可能加重受影响个体的AR症状。