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. 2020 Mar 10;39(18):3754–3773. doi: 10.1038/s41388-020-1251-2

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 5 — a Flow cytometry sorting of AT2 cells (Sca-1⁺CD31⁻CD45⁻) (P4) and further sorting of ABCG1⁺ cells (Sca-1⁺Abcg1⁺) (P5) from P4 from the lungs of 6-month aged WT or KO mice; b IF identification of AT2 and Sca-1⁺Abcg1⁺cells as P4 and P5 subset, respectively, (Bar = 10 µm); Comparison of the positive rates (c) and percentages (d) of AT2 and Sca-1⁺Abcg1⁺cells isolated from WT or KO mice; Colony formation (e) and quantitative analysis (f) of Sca-1⁺Abcg1⁺cells isolated from WT or KO mice, (Bar = 500 µm); g Confocal analysis of SPA/ABCG1 and α6/β4 in formed colony of Sca-1⁺Abcg1⁺cells isolated from KO mice, (Bar = 20 µm); Comparison of 3D spheroid formation (h) in limiting dilutions of 1, 10, 100, or 1000 (i) Sca-1⁺Abcg1⁺cells isolated from WT or KO mice, (Bar = 1000 µm); j Confocal analysis of SPA/ABCG1 and α6/β4 in spheroid of Sca-1⁺Abcg1⁺cells isolated from KO mice, (Bar = 10 µm); k Flowchart of the isolation of cells from lung tissue samples from mice or from patients with lung cancer, followed by the injection of the cells into mouse lungs via the tail vein and harvest of lung tissue that was subjected to further embedding, sectioning and staining; l Flow cytometry sorting of AT2 cells (P4) from 6-month aged KO mice and patients with lung cancer, and further sorting of ABCG1⁺ cells (Sca-1⁺Abcg1⁺ or SPA⁺ABCG1⁺) (P5) from P4, the remaining cells (Sca-1⁺Abcg1⁻ or SPA⁺ABCG1⁻) were marked as Not P5; Tumor incidence (m) and IF staining of Sca-1/Abcg1 or SPA/ABCG1 in the S/TB region or tumor region after the injection of Sca-1⁺Abcg1⁻ or Sca-1⁺Abcg1⁺cells from KO mice, and SPA⁺ABCG1⁻ or SPA⁺ABCG1⁺cells from lung tissue samples via the tail vein (n), (Bar = 100 µm); Data were collected from three independent experiments with triplicate samples. ***P < 0.001.

Fig. 5 — a Flow cytometry sorting of AT2 cells (Sca-1⁺CD31⁻CD45⁻) (P4) and further sorting of ABCG1⁺ cells (Sca-1⁺Abcg1⁺) (P5) from P4 from the lungs of 6-month aged WT or KO mice; b IF identification of AT2 and Sca-1⁺Abcg1⁺cells as P4 and P5 subset, respectively, (Bar = 10 µm); Comparison of the positive rates (c) and percentages (d) of AT2 and Sca-1⁺Abcg1⁺cells isolated from WT or KO mice; Colony formation (e) and quantitative analysis (f) of Sca-1⁺Abcg1⁺cells isolated from WT or KO mice, (Bar = 500 µm); g Confocal analysis of SPA/ABCG1 and α6/β4 in formed colony of Sca-1⁺Abcg1⁺cells isolated from KO mice, (Bar = 20 µm); Comparison of 3D spheroid formation (h) in limiting dilutions of 1, 10, 100, or 1000 (i) Sca-1⁺Abcg1⁺cells isolated from WT or KO mice, (Bar = 1000 µm); j Confocal analysis of SPA/ABCG1 and α6/β4 in spheroid of Sca-1⁺Abcg1⁺cells isolated from KO mice, (Bar = 10 µm); k Flowchart of the isolation of cells from lung tissue samples from mice or from patients with lung cancer, followed by the injection of the cells into mouse lungs via the tail vein and harvest of lung tissue that was subjected to further embedding, sectioning and staining; l Flow cytometry sorting of AT2 cells (P4) from 6-month aged KO mice and patients with lung cancer, and further sorting of ABCG1⁺ cells (Sca-1⁺Abcg1⁺ or SPA⁺ABCG1⁺) (P5) from P4, the remaining cells (Sca-1⁺Abcg1⁻ or SPA⁺ABCG1⁻) were marked as Not P5; Tumor incidence (m) and IF staining of Sca-1/Abcg1 or SPA/ABCG1 in the S/TB region or tumor region after the injection of Sca-1⁺Abcg1⁻ or Sca-1⁺Abcg1⁺cells from KO mice, and SPA⁺ABCG1⁻ or SPA⁺ABCG1⁺cells from lung tissue samples via the tail vein (n), (Bar = 100 µm); Data were collected from three independent experiments with triplicate samples. ***P < 0.001.