Chronic exposure to cigarette smoke (CS) causes chronic inflammation, oxidative stress, and apoptosis of epithelial cells, which results in destruction of the lung matrix. However, the mechanism by which the lung fails to repair the CS-induced damage, thereby succumbing to emphysema, remains unclear. Alveolar type 2 (AT2) cells comprise the stem cells of the alveolar compartments and are responsible for repairing and maintaining lung tissues. In this study, we examined the effect of chronic CS on AT2 stem cells. Adult mice expressing GFP in their AT2 cells were exposed to CS for > 3 months. Histological assessment showed that CS not only induced emphysematous changes but also increased the number of AT2 cells compared with that of air-exposed lungs. Assessment of sorted GFP⁺/AT2 cells via the stem cell three-dimensional organoid/colony-forming assay revealed that the number and size of the colonies formed by the CS-exposed AT2 stem cells were significantly higher than those of air-exposed control AT2 cells. Although CS-exposed lungs had more apoptotic cells, examination of the surviving AT2 stem cells in two-dimensional in vitro culture revealed that they developed a higher ability to resist apoptosis. Microarray analysis of CS-exposed AT2 stem cells revealed the upregulation of genes related to circadian rhythm and inflammatory pathways. In conclusion, we provide evidence that AT2 stem cells respond to chronic CS exposure by activating their stem cell function, thereby proliferating and differentiating faster and becoming more resistant to apoptosis. Disturbances in expression levels of several circadian rhythm–related genes might be involved in these changes.

长期接触香烟烟雾（CS）会导致慢性炎症，氧化应激和上皮细胞凋亡，从而导致肺基质破坏。然而，肺不能修复CS引起的损伤从而屈服于肺气肿的机制仍不清楚。肺泡2型（AT2）细胞包括肺泡隔室的干细胞，负责修复和维持肺组织。在这项研究中，我们检查了慢性CS对AT2干细胞的影响。将在其AT2细胞中表达GFP的成年小鼠暴露于CS> 3个月。组织学评估表明，与空气暴露的肺相比，CS不仅诱导了气肿的改变，而且增加了AT2细胞的数量。评估排序的GFP ⁺/ AT2细胞通过干细胞三维类器官/菌落形成试验显示，暴露于CS的AT2干细胞形成的集落的数量和大小显着高于暴露于空气的对照AT2细胞。尽管CS暴露的肺具有更多的凋亡细胞，但是在二维体外检查存活的AT2干细胞培养表明，它们具有更高的抗凋亡能力。对CS暴露的AT2干细胞进行的微阵列分析显示，与昼夜节律和炎症途径相关的基因上调。总之，我们提供的证据表明AT2干细胞通过激活干细胞功能来响应慢性CS暴露，从而使其增殖和分化更快，并且对凋亡的耐受性更高。这些昼夜节律相关基因的表达水平紊乱可能与这些变化有关。