Type 2 alveolar cells are stem cells in adult lung
Christina E. Barkauskas, … , Paul W. Noble, Brigid L.M. Hogan
Christina E. Barkauskas, … , Paul W. Noble, Brigid L.M. Hogan
Published June 10, 2013
Citation Information: J Clin Invest. 2013;123(7):3025-3036. https://doi.org/10.1172/JCI68782.
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Research Article Pulmonology

Type 2 alveolar cells are stem cells in adult lung

Abstract

Gas exchange in the lung occurs within alveoli, air-filled sacs composed of type 2 and type 1 epithelial cells (AEC2s and AEC1s), capillaries, and various resident mesenchymal cells. Here, we use a combination of in vivo clonal lineage analysis, different injury/repair systems, and in vitro culture of purified cell populations to obtain new information about the contribution of AEC2s to alveolar maintenance and repair. Genetic lineage-tracing experiments showed that surfactant protein C–positive (SFTPC-positive) AEC2s self renew and differentiate over about a year, consistent with the population containing long-term alveolar stem cells. Moreover, if many AEC2s were specifically ablated, high-resolution imaging of intact lungs showed that individual survivors undergo rapid clonal expansion and daughter cell dispersal. Individual lineage-labeled AEC2s placed into 3D culture gave rise to self-renewing “alveolospheres,” which contained both AEC2s and cells expressing multiple AEC1 markers, including HOPX, a new marker for AEC1s. Growth and differentiation of the alveolospheres occurred most readily when cocultured with primary PDGFRα+ lung stromal cells. This population included lipofibroblasts that normally reside close to AEC2s and may therefore contribute to a stem cell niche in the murine lung. Results suggest that a similar dynamic exists between AEC2s and mesenchymal cells in the human lung.

Authors

Christina E. Barkauskas, Michael J. Cronce, Craig R. Rackley, Emily J. Bowie, Douglas R. Keene, Barry R. Stripp, Scott H. Randell, Paul W. Noble, Brigid L.M. Hogan

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Figure 5

Self renewal and differentiation of AEC2 cells in 3D organoid culture.

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Self renewal and differentiation of AEC2 cells in 3D organoid culture. (...
(A) Sftpc-CreER;Rosa-Tm mice were dosed ×4 with Tmx (0.2 mg/g) and at least 4 dpi lungs were dissociated and sorted by FACS. The Tm+ fraction (P3, top left) was seeded at a density of 5,000 cells in 90 μl of 50% Matrigel in a 24-well Transwell insert (top right) together with 1 × 106 PDGFRA-GFPhi stromal cells freshly sorted from lungs of a Pdgfra-H2B:GFP transgenic mouse (P3, bottom panels). (B and C) After 14 days, spheres of lineage-labeled cells are present in various sizes. (D) CFE is 2.3% ± 0.3% for primary cultures (n = 8 experiments, ≥ 2 replicates per experiment), 6.2% ± 0.7% after passage 1, and 5.1% ± 0.7% after passage 2. (E–I) Histology and immunohistochemistry of sections of spheres after 16–17 days shows (E) alveolus-like areas (asterisk) with more elongated cells, (F) that all cells are lineage labeled and those on the periphery express SFTPC, while cells in the interior express T1a, AQAPORIN 5 (Aqp5) (G and H), and HOPX (I) — markers of AEC1s (see also Supplemental Figure 7). (J and K) TEM of spheres at day 16 shows many cells with lamellar bodies at different stages of maturation and apical membranes with dense microvilli (mv). The cells release surfactant into the interior of the spheres where it accumulates in large amounts. Scale bars: 100 μm (B); 50 μm (C); 50 μm (E); 50 μm (F–I); 2 μm (J and K). See also Supplemental Figures 6 and 7.