Tanabe Ikuya, Ishikawa Shinkichi
Scientific Product Assessment Center, R&D Group, Japan Tobacco Inc., 6-2 Umegaoka, Aoba-ku, Yokohama, Kanagawa, 227-8512, Japan.
Sci Rep. 2025 Jul 2;15(1):22995. doi: 10.1038/s41598-025-07219-8.
Lung epithelia are exposed to various substances in the atmosphere through breathing. To investigate the mechanisms and treatments for respiratory diseases caused by these substances, robust in vitro lung epithelial models are essential. This study aimed to develop an in vitro alveolar epithelial model using primary human pulmonary alveolar epithelial cells (HPAEpiCs). HPAEpiCs were cultured at an air-liquid interface (ALI) for 28 days in a medium supplemented with three small molecules: Y-27632, A-83-01, and CHIR99021. The characteristics of the ALI-cultured cells were then analyzed. Immunostaining revealed that many cells expressed alveolar type 2 (AT2) cell markers, such as surfactant protein B and prosurfactant protein C. Single-cell gene expression analysis further confirmed that the majority of the cells expressed genes reported to be highly expressed in AT2 cells. The apical surface of the ALI-cultured HPAEpiCs was covered with a liquid containing a variety of lipids and proteins known to be present in lung surfactant in vivo. Our data demonstrate that 28 days of ALI culture promoted the differentiation of HPAEpiCs into AT2 cells capable of secreting lung surfactant components. This ALI culture model containing differentiated AT2 cells could be valuable for investigating the mechanisms and treatments for respiratory diseases.
肺上皮细胞通过呼吸接触大气中的各种物质。为了研究由这些物质引起的呼吸系统疾病的发病机制和治疗方法，强大的体外肺上皮模型至关重要。本研究旨在利用原代人肺肺泡上皮细胞（HPAEpiCs）建立一种体外肺泡上皮模型。将HPAEpiCs在气液界面（ALI）培养28天，培养基中添加三种小分子：Y-27632、A-83-01和CHIR99021。然后分析ALI培养细胞的特性。免疫染色显示许多细胞表达肺泡II型（AT2）细胞标志物，如表面活性蛋白B和前表面活性蛋白C。单细胞基因表达分析进一步证实，大多数细胞表达据报道在AT2细胞中高表达的基因。ALI培养的HPAEpiCs的顶端表面覆盖着一种液体，其中含有多种已知存在于体内肺表面活性物质中的脂质和蛋白质。我们的数据表明，28天的ALI培养促进了HPAEpiCs分化为能够分泌肺表面活性物质成分的AT2细胞。这种含有分化的AT2细胞的ALI培养模型对于研究呼吸系统疾病的发病机制和治疗方法可能具有重要价值。
