Biosciences Institute, University Medical School, Newcastle University, Framlington Place, Newcastle upon Tyne NE2 4HH, UK.
Bioinformatics Support Unit, Faculty of Medical Sciences, Newcastle University, Framlington Place, Newcastle upon Tyne NE2 4HH, UK.
Cells. 2020 Sep 21;9(9):2137. doi: 10.3390/cells9092137.
In vitro cultures of primary human airway epithelial cells (hAECs) grown at air-liquid interface have become a valuable tool to study airway biology under normal and pathologic conditions, and for drug discovery in lung diseases such as cystic fibrosis (CF). An increasing number of different differentiation media, are now available, making comparison of data between studies difficult. Here, we investigated the impact of two common differentiation media on phenotypic, transcriptomic, and physiological features of CF and non-CF epithelia. Cellular architecture and density were strongly impacted by the choice of medium. RNA-sequencing revealed a shift in airway cell lineage; one medium promoting differentiation into club and goblet cells whilst the other enriched the growth of ionocytes and multiciliated cells. Pathway analysis identified differential expression of genes involved in ion and fluid transport. Physiological assays (intracellular/extracellular pH, Ussing chamber) specifically showed that ATP12A and CFTR function were altered, impacting pH and transepithelial ion transport in CF hAECs. Importantly, the two media differentially affected functional responses to CFTR modulators. We argue that the effect of growth conditions should be appropriately determined depending on the scientific question and that our study can act as a guide for choosing the optimal growth medium for specific applications.
在体外培养原代人呼吸道上皮细胞（hAECs），在气液界面生长，已成为研究正常和病理条件下气道生物学以及囊性纤维化（CF）等肺部疾病药物发现的有价值的工具。现在有越来越多不同的分化培养基可供选择，这使得很难在研究之间比较数据。在这里，我们研究了两种常见的分化培养基对 CF 和非 CF 上皮细胞表型、转录组和生理特征的影响。细胞结构和密度受到培养基选择的强烈影响。RNA 测序显示气道细胞谱系发生了转变；一种培养基促进向 club 和 goblet 细胞分化，而另一种培养基则富集了离子细胞和多纤毛细胞的生长。通路分析鉴定了参与离子和液体转运的基因的差异表达。生理测定（细胞内/细胞外 pH 值、Ussing 室）特别表明，ATP12A 和 CFTR 功能发生改变，影响 CF hAECs 的 pH 值和跨上皮离子转运。重要的是，这两种培养基对 CFTR 调节剂的功能反应有不同的影响。我们认为，生长条件的影响应根据科学问题适当确定，我们的研究可以作为选择特定应用最佳生长培养基的指南。
