Center for Clinical Single-Cell Biomedicine, Henan Provincial People's Hospital, People's Hospital of Zhengzhou University, School of Clinical Medicine, Henan University, Zhengzhou, China.
Department of Oncology, Henan Provincial People's Hospital, People's Hospital of Zhengzhou University, School of Clinical Medicine, Henan University, Zhengzhou, China.
FASEB J. 2020 Sep;34(9):12466-12480. doi: 10.1096/fj.202001518. Epub 2020 Jul 24.
The role of type II alveolar epithelial stem cells (AEC II) for alveolar repair in radiation-induced lung fibrosis (RILF) remains largely unknown, mainly because of AEC II phenotype's spontaneous change in vitro. Cell differentiation status is determined by Lin28 and let-7 miRNAs in see-saw-pattern. Lin28, a repressor of let-7 and a stem cell marker, is activated by β-catenin. The expression of β-catenin is regulated by GSK-3β/TGF-β1 signaling. To understand the true role of AEC II in RILF, we freshly isolated primary AEC II directly from thoracically irradiated lungs. We then explored the expressions of cell phenotype markers and differentiation regulators in these isolated AEC II to analyze the correlation between GSK-3β/TGF-β1/β-catenin signaling pathway, lin28/let-7 balance, and AEC II phenotypes at different injury phases following irradiation. Results showed that isolated single primary cells displayed AEC II ultrastructural features and proSP-C positive. The gene expressions of prosp-c (an AEC II biomarker) and hopx (an AEC I marker) significantly increased in isolated AEC II during injury repair phase (P < .001 and P < .05) but decreased at end-stage of injury, while mesenchymal markers increased in both isolated AEC II and irradiated lungs. mRNA levels of gsk-3β, tgf-β1, and β-catenin increased in all irradiated AEC II, but more pronounced in the second half of injury phase (P < .05-P < .001). Similarly, the expression of lin28 was also significantly elevated in isolated AEC II at the late phase (P < .05-P < .001). Four let-7 miRNAs were significantly upregulated in all irradiated AEC II groups (P < .05-P < .001). The time-dependent and highly consistent uptrends for four lin28/let-7 ratios in sorted AEC II contrasted to downtrends in irradiated lungs. In conclusion, RILF occurred when GSK-3β/TGF-β1 signaling increased β-catenin levels, which led to the augmentation of AEC II population by elevated lin28/let-7 ratio and the transcription of profibrotic cytokines and factors, thereby inducing AEC II to undergo transdifferentiation into mesenchymal cells.
在放射性肺纤维化 (RILF) 中，II 型肺泡上皮干细胞 (AEC II) 对肺泡修复的作用在很大程度上尚不清楚，主要是因为 AEC II 表型在体外会自发改变。细胞分化状态由 Lin28 和 let-7 miRNAs 以跷跷板模式决定。Lin28 是 let-7 的抑制剂和干细胞标志物，被 β-catenin 激活。β-catenin 的表达受 GSK-3β/TGF-β1 信号通路调节。为了了解 AEC II 在 RILF 中的真正作用，我们直接从胸部照射的肺中新鲜分离原代 AEC II。然后，我们探索了这些分离的 AEC II 中细胞表型标志物和分化调节剂的表达，以分析 GSK-3β/TGF-β1/β-catenin 信号通路、Lin28/let-7 平衡与照射后不同损伤阶段 AEC II 表型之间的相关性。结果表明，分离的单个原代细胞显示出 AEC II 的超微结构特征和 proSP-C 阳性。在损伤修复阶段，分离的 AEC II 中 prosp-c（AEC II 标志物）和 hopx（AEC I 标志物）的基因表达显著增加（P<.001 和 P<.05），但在损伤末期减少，而间充质标志物在分离的 AEC II 和照射的肺中均增加。所有照射的 AEC II 中，GSK-3β、TGF-β1 和 β-catenin 的 mRNA 水平均增加，但在损伤后半期更为明显（P<.05-P<.001）。同样，分离的 AEC II 中 lin28 的表达在晚期也显著升高（P<.05-P<.001）。所有照射的 AEC II 中，四个 let-7 miRNAs 均显著上调（P<.05-P<.001）。在分离的 AEC II 中，四个 lin28/let-7 比值的时间依赖性和高度一致的上升趋势与照射肺中的下降趋势形成对比。总之，当 GSK-3β/TGF-β1 信号增加 β-catenin 水平时，会导致 AEC II 群体增加，这是通过升高的 lin28/let-7 比值和转录成纤维细胞因子和因子引起的，从而诱导 AEC II 向间充质细胞发生转分化。
