Unbiased Quantitation of Alveolar Type II to Alveolar Type I Cell Transdifferentiation during Repair after Lung Injury in Mice
Unbiased Quantitation of Alveolar Type II to Alveolar Type I Cell Transdifferentiation during Repair after Lung Injury in Mice
复制标题
DOI：
10.1165/rcmb.2017-0037ma
复制
发表时间：
2017-11-01
影响因子：
6.4
通讯作者：
Zemans, Rachel L.
中科院分区：
文献类型：
--
作者：
Jansing, Nicole L.;McClendon, Jazalle;Zemans, Rachel L.
肺泡上皮由鳞状肺泡型(AT)I型和立方ATII型细胞组成。ATI细胞覆盖95-98%的肺泡表面，因此在屏障完整性方面发挥关键作用，并且非常薄，因此允许有效的气体交换。在肺损伤期间，ATI细胞死亡，导致上皮通透性增加。ATII细胞通过增殖和转分化为ATI细胞，使肺泡表面重新上皮化。转分化的特征是ATII细胞标志物下调，ATI细胞标志物上调，细胞扩散，导致形态由立方变为鳞状，从而恢复正常的肺泡结构和功能。ATII向ATI细胞转分化的机制在体内尚未得到很好的研究。机械调查的前提是一个严格的、公正的方法来量化这一过程。在这里，我们使用了表达绿色荧光蛋白(GFP)的ATII细胞及其后代的SPCCreERT2；mTmG小鼠，并应用体视学技术测量了脂多糖诱导的损伤修复过程中的转分化。转分化以表达ATI而不表达ATII的细胞标志物所覆盖的肺泡表面积百分比来定量。利用这种方法，确定了修复过程中转分化的时间进程和程度。我们发现ATI细胞丢失和上皮通透性在第4天发生，ATII到ATI细胞的转分化从第7天开始并持续到第16天。值得注意的是，转分化和屏障恢复是时间相关的。这种方法可用于研究转分化的分子机制，最终揭示加速肺损伤修复的新的治疗靶点。
The alveolar epithelium consists of squamous alveolar type (AT) I and cuboidal ATII cells. ATI cells cover 95-98% of the alveolar surface, thereby playing a critical role in barrier integrity, and are extremely thin, thus permitting efficient gas exchange. During lung injury, ATI cells die, resulting in increased epithelial permeability. ATII cells re-epithelialize the alveolar surface via proliferation and transdifferentiation into ATI cells. Transdifferentiation is characterized by down-regulation of ATII cell markers, up-regulation of ATI cell markers, and cell spreading, resulting in a change in morphology from cuboidal to squamous, thus restoring normal alveolar architecture and function. The mechanisms underlying ATII to ATI cell transdifferentiation have not been well studied in vivo. A prerequisite for mechanistic investigation is a rigorous, unbiased method to quantitate this process. Here, we used SPCCreERT2; mTmG mice, in which ATII cells and their progeny express green fluorescent protein (GFP), and applied stereologic techniques to measure transdifferentiation during repair after injury induced by LPS. Transdifferentiation was quantitated as the percent of alveolar surface area covered by ATII-derived (GFP1) cells expressing ATI, but not ATII, cell markers. Using this methodology, the time course and magnitude of transdifferentiation during repair was determined. We found that ATI cell loss and epithelial permeability occurred by Day 4, and ATII to ATI cell transdifferentiation began by Day 7 and continued until Day 16. Notably, transdifferentiation and barrier restoration are temporally correlated. This methodology can be applied to investigate the molecular mechanisms underlying transdifferentiation, ultimately revealing novel therapeutic targets to accelerate repair after lung injury.