Basal cells as stem cells of the mouse trachea and human airway epithelium

Department of Cell Biology, Duke University Medical Center, Durham, NC 27710, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 08/2009; 106(31):12771-5. DOI: 10.1073/pnas.0906850106
Source: PubMed


The pseudostratified epithelium of the mouse trachea and human airways contains a population of basal cells expressing Trp-63 (p63) and cytokeratins 5 (Krt5) and Krt14. Using a KRT5-CreER(T2) transgenic mouse line for lineage tracing, we show that basal cells generate differentiated cells during postnatal growth and in the adult during both steady state and epithelial repair. We have fractionated mouse basal cells by FACS and identified 627 genes preferentially expressed in a basal subpopulation vs. non-BCs. Analysis reveals potential mechanisms regulating basal cells and allows comparison with other epithelial stem cells. To study basal cell behaviors, we describe a simple in vitro clonal sphere-forming assay in which mouse basal cells self-renew and generate luminal cells, including differentiated ciliated cells, in the absence of stroma. The transcriptional profile identified 2 cell-surface markers, ITGA6 and NGFR, which can be used in combination to purify human lung basal cells by FACS. Like those from the mouse trachea, human airway basal cells both self-renew and generate luminal daughters in the sphere-forming assay.

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    • "Cell-surface antigens including CD44, MUC1, and nerve growth factor receptor (NGFR) and intracellular antigens CK19, p63, CK13, and CK4 were all expressed, but variably , in the surface epithelium, where their location could be associated with an increasingly differentiated state from the basal layer to the outer surface. For example, CK19 and NGFR were exclusive to the basal layer of the surface epithelium (Figure 1I, center row), whereas p63, a widely annotated marker of basal epithelial cells (Pellegrini et al., 2001; Rock et al., 2009; Senoo et al., 2007), and CD44, a receptor for hyaluronic acid, were seen not only in the basal layers but also in the parabasal layers, extending up to the mid-layers of the surface epithelium (Figure 1I, center row). CK13, CK4, involucrin , and cell-surface MUC1 were detected only in the parabasal and upper layers of the surface epithelium and not in any basal cells (Figure 1J, center row). "
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    ABSTRACT: Human palatine tonsils are oropharyngeal lymphoid tissues containing multiple invaginations (crypts) in which the continuity of the outer surface epithelium is disrupted and the isolated epithelial cells intermingle with other cell types. We now show that primitive epithelial cells detectable in vitro in 2D colony assays and in a 3D culture system are CD44 + NGFR + and present in both surface and crypt regions. Transcriptome analysis indicated a high similarity between CD44 + NGFR + cells in both regions, although those isolated from the crypt contained a higher proportion of the most primitive (holo)clonogenic cells. Lentiviral transduction of CD44 + NGFR + cells from both regions with human papillomavirus 16-encoded E6/E7 prolonged their growth in 2D cultures and caused aberrant differentiation in 3D cultures. Our findings therefore reveal a shared, site-independent, hierarchical organization, differentiation potential, and transcrip-tional profile of normal human tonsillar epithelial progenitor cells. They also introduce a new model for investigating the mechanisms of their transformation.
    Stem Cell Reports 12/2015; 7:1-16. DOI:10.1016/j.stemcr.2015.09.020 · 5.37 Impact Factor
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    • "Thus, epithelial YY1 controls the condensation of SOX9-positive mesenchymal cells into precartilage nodules. We assessed the expression of p63, a marker of basal cells, which are known to generate ciliated, club and goblet cells (Rock et al., 2009). Basal cells were distributed irregularly along the proximal airways of Yy1 flox/flox ;Shh +/Cre embryos (Fig. 3H,I). "
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    ABSTRACT: Yin Yang 1 (YY1) is a multifunctional zinc-finger-containing transcription factor that plays crucial roles in numerous biological processes by selectively activating or repressing transcription, depending upon promoter contextual differences and specific protein interactions. In mice, Yy1 null mutants die early in gestation whereas Yy1 hypomorphs die at birth from lung defects. We studied how the epithelial-specific inactivation of Yy1 impacts on lung development. The Yy1 mutation in lung epithelium resulted in neonatal death due to respiratory failure. It impaired tracheal cartilage formation, altered cell differentiation, abrogated lung branching and caused airway dilation similar to that seen in human congenital cystic lung diseases. The cystic lung phenotype in Yy1 mutants can be partly explained by the reduced expression of Shh, a transcriptional target of YY1, in lung endoderm, and the subsequent derepression of mesenchymal Fgf10 expression. Accordingly, SHH supplementation partially rescued the lung phenotype in vitro. Analysis of human lung tissues revealed decreased YY1 expression in children with pleuropulmonary blastoma (PPB), a rare pediatric lung tumor arising during fetal development and associated with DICER1 mutations. No evidence for a potential genetic interplay between murine Dicer and Yy1 genes during lung morphogenesis was observed. However, the cystic lung phenotype resulting from the epithelial inactivation of Dicer function mimics the Yy1 lung malformations with similar changes in Shh and Fgf10 expression. Together, our data demonstrate the crucial requirement for YY1 in lung morphogenesis and identify Yy1 mutant mice as a potential model for studying the genetic basis of PPB. © 2015. Published by The Company of Biologists Ltd.
    Development 09/2015; 142(17). DOI:10.1242/dev.120469 · 6.46 Impact Factor
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    • "The mouse trachea contains three major cell types: TRP63 + , KRT5 + basal cells (BCs); luminal secretory cells (SecCs, mostly Scgb1a1 + Club/Clara-like cells); and luminal ciliated cells (CCs) (Rock et al., 2010). Previous population-level lineage tracing using transgenic Tg(KRT5-CreER) mice demonstrated that BCs include self-renewing stem cells involved in tracheal growth, homeostasis (at least for up to 16 weeks), and repair (Rock et al., 2009). However, it is not known if BCs are a functionally heterogeneous population. "
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    ABSTRACT: Epithelial lineages have been studied at cellular resolution in multiple organs that turn over rapidly. However, many epithelia, including those of the lung, liver, pancreas, and prostate, turn over slowly and may be regulated differently. We investigated the mouse tracheal epithelial lineage at homeostasis by using long-term clonal analysis and mathematical modeling. This pseudostratified epithelium contains basal cells and secretory and multiciliated luminal cells. Our analysis revealed that basal cells are heterogeneous, comprising approximately equal numbers of multipotent stem cells and committed precursors, which persist in the basal layer for 11 days before differentiating to luminal fate. We confirmed the molecular and functional differences within the basal population by using single-cell qRT-PCR and further lineage labeling. Additionally, we show that self-renewal of short-lived secretory cells is a feature of homeostasis. We have thus revealed early luminal commitment of cells that are morphologically indistinguishable from stem cells. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
    Cell Reports 06/2015; 12(1). DOI:10.1016/j.celrep.2015.06.011 · 8.36 Impact Factor
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