Culture media strongly influences primary human bronchial epithelial cells' transcriptomic response to ozone exposure

Exposure to the ambient air pollutant ozone induces acute and chronic
respiratory health effects in part by causing inflammation of the airways.
Several aspects of the inflammatory response to ozone can be modeled in
vitro using primary human bronchial epithelial cells (HBECs) cultured at
an air-liquid interface. We tested two commonly used HBEC culture media
systems, one proprietary and one non-proprietary, to identify which system
yielded the most in vivo-like pro-inflammatory response to acute ozone
exposure as indicated by gene expression. Cells from six donors were grown
in each culture system in parallel followed by examination of epithelial
morphology and cell type proportions prior to ozone exposure. Cultures
grown in the proprietary system were notably thicker and contained more
ciliated and secretory cells, as well as internal cyst-like structures.
The transcriptomic response to acute ozone exposure (0.5 parts per million
ozone x 2 hours) was strongly affected by media. HBECs grown in the
proprietary system exhibited minimal changes after ozone, with only 7
differentially expressed genes (DEGs). In contrast, HBECs grown in the
non-proprietary system exhibited a more dynamic response with 128 DEGs,
including hallmark response genes indicative of inflammation (CXCL8) and
oxidative stress (HMOX1). Gene set enrichment analysis using the 128 DEGs
further corroborated upregulation of oxidative stress and inflammation
pathways. In total, our results indicate that the choice of HBEC culture
media should be carefully considered to best model the in vivo response to
ozone.

暴露于环境空气污染物臭氧可引发急慢性呼吸系统健康损害，其部分机制为诱发气道炎症。臭氧诱导的炎症反应的多个关键环节，可通过体外（in vitro）培养原代人支气管上皮细胞（primary human bronchial epithelial cells, HBECs）并维持气液界面（air-liquid interface）进行建模。本研究测试了两种常用的HBEC培养基体系，分别为专有体系与非专有体系，旨在通过基因表达水平判断哪种体系能最逼真地模拟急性臭氧暴露后的体内（in vivo）促炎反应。我们将6名供体来源的细胞分别在两种培养基体系中平行培养，并在臭氧暴露前检测上皮细胞形态与细胞类型占比。在专有培养基中培养的细胞层显著更厚，且含有更多纤毛细胞与分泌细胞，同时还形成了内部囊状结构。急性臭氧暴露（0.5 ppm 暴露2小时）后的转录组响应显著受培养基类型影响：在专有培养基中培养的HBECs在臭氧暴露后仅出现7个差异表达基因（differentially expressed genes, DEGs），转录组变化幅度极小；与之相反，非专有培养基培养的HBECs则呈现出更动态的转录组响应，共检出128个差异表达基因，其中包括提示炎症反应的标志性基因CXCL8与氧化应激相关基因HMOX1。针对这128个差异表达基因进行基因集富集分析，进一步证实了氧化应激与炎症通路的上调。综上，本研究结果表明，若要精准模拟臭氧暴露的体内响应，需谨慎选择HBEC培养用培养基体系。