Abstract
Clonal culture of stem cells is crucial for their identification, and the characterization of the cellular and molecular mechanisms that regulate their proliferation and differentiation. In the adult mouse lung, epithelial stem/progenitor cells are defined by the phenotype CD45neg CD31neg EpCAMpos CD104pos CD24low. Here we describe a tissue dissociation and flow cytometry strategy for the detection and isolation of adult mouse lung epithelial stem/progenitor cells, and a three-dimensional colony-forming assay for their clonal culture in vitro.
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Rawlins EL, Okubo T, Xue Y et al (2009) The role of Scgb1a1+ Clara cells in the long-term maintenance and repair of lung airway, but not alveolar, epithelium. Cell Stem Cell 4:525–534
Hong KU, Reynolds SD, Watkins S et al (2004) In vivo differentiation potential of tracheal basal cells: evidence for multipotent and unipotent subpopulations. Am J Physiol Lung Cell Mol Physiol 286:L643–L649
Rock JR, Onaitis MW, Rawlins EL et al (2009) Basal cells as stem cells of the mouse trachea and human airway epithelium. Proc Natl Acad Sci U S A 106:12771–12775
Hegab AE, Ha VL, Gilbert JL et al (2011) Novel stem/progenitor cell population from murine tracheal submucosal gland ducts with multipotent regenerative potential. Stem Cells 29:1283–1293
Tesei A, Zoli W, Arienti C et al (2009) Isolation of stem/progenitor cells from normal lung tissue of adult humans. Cell Prolif 42:298–308
Hong KU, Reynolds SD, Watkins S et al (2004) Basal cells are a multipotent progenitor capable of renewing the bronchial epithelium. Am J Pathol 164:577–588
Giangreco A, Reynolds SD, Stripp BR (2002) Terminal bronchioles harbor a unique airway stem cell population that localizes to the bronchoalveolar duct junction. Am J Pathol 161:173–182
Reynolds SD, Giangreco A, Power JH et al (2000) Neuroepithelial bodies of pulmonary airways serve as a reservoir of progenitor cells capable of epithelial regeneration. Am J Pathol 156:269–278
Kim CF, Jackson EL, Woolfenden AE et al (2005) Identification of bronchioalveolar stem cells in normal lung and lung cancer. Cell 121:823–835
Chapman HA, Li X, Alexander JP et al (2011) Integrin alpha6beta4 identifies an adult distal lung epithelial population with regenerative potential in mice. J Clin Invest 121:2855–2862
Barkauskas CE, Cronce MJ, Rackley CR et al (2013) Type 2 alveolar cells are stem cells in adult lung. J Clin Invest 123:3025–3036
Teisanu RM, Chen H, Matsumoto K et al (2011) Functional analysis of two distinct bronchiolar progenitors during lung injury and repair. Am J Respir Cell Mol Biol 44:794–803
McQualter JL, Yuen K, Williams B et al (2010) Evidence of an epithelial stem/progenitor cell hierarchy in the adult mouse lung. Proc Natl Acad Sci U S A 107:1414–1419
Chen H, Matsumoto K, Brockway BL et al (2012) Airway epithelial progenitors are region specific and show differential responses to bleomycin-induced lung injury. Stem Cells 30:1948–1960
Lee JH, Kim J, Gludish D et al (2012) SPC H2B-GFP mice reveal heterogeneity of surfactant protein C-expressing lung cells. Am J Respir Cell Mol Biol 48:288–298
Kajstura J, Rota M, Hall SR et al (2011) Evidence for human lung stem cells. N Engl J Med 364:1795–1806
Oeztuerk-Winder F, Guinot A, Ochalek A et al (2012) Regulation of human lung alveolar multipotent cells by a novel p38alpha MAPK/miR-17-92 axis. EMBO J 31:3431–3441
McQualter JL, Brouard N, Williams B et al (2009) Endogenous fibroblastic progenitor cells in the adult mouse lung are highly enriched in the sca-1 positive cell fraction. Stem Cells 27:623–633
McQualter JL, McCarty RC, Van der Velden J et al (2013) TGF-beta signaling in stromal cells acts upstream of FGF-10 to regulate epithelial stem cell growth in the adult lung. Stem Cell Res 11:1222–1233
Wion D, Christen T, Barbier EL et al (2009) PO(2) matters in stem cell culture. Cell Stem Cell 5:242–243
Acknowledgements
This work was supported by grants from the Australian National Health and Medical Research Council (NHMRC) and the Australian Stem Cell Centre.
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McQualter, J.L., Bertoncello, I. (2015). Clonal Culture of Adult Mouse Lung Epithelial Stem/Progenitor Cells. In: Rich, I. (eds) Stem Cell Protocols. Methods in Molecular Biology, vol 1235. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1785-3_17
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DOI: https://doi.org/10.1007/978-1-4939-1785-3_17
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Publisher Name: Humana Press, New York, NY
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Online ISBN: 978-1-4939-1785-3
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