Emergence of a Wave of Wnt Signaling that Regulates Lung Alveologenesis by Controlling Epithelial Self-Renewal and Differentiation

Abstract

Alveologenesis is the culmination of lung development and involves the correct temporal and spatial signals to generate the delicate gas exchange interface required for respiration. Using a novel Wnt signaling reporter system, we demonstrate the emergence of a Wnt-responsive alveolar epithelial cell sublineage that arises during alveologenesis called the axin2+ alveolar type 2 cell or AT2Axin2. The number of AT2Axin2 cells increases substantially during late lung development, correlating with a wave of Wnt signaling during alveologenesis. Transcriptome analysis, in vivo clonal analysis, and ex vivo lung organoid assays reveal that AT2sAxin2 promote enhanced AT2 cell growth during generation of the alveolus. Activating Wnt signaling results in expansion of AT2s whereas inhibition of Wnt signaling inhibits AT2 cell development and shunts alveolar epithelial development towards the alveolar type 1 cell lineage. These findings reveal a wave of Wnt-dependent AT2 expansion required for lung alveologenesis and maturation.

Distinct Mesenchymal Lineages and Niches Promote Epithelial Self-Renewal and Myofibrogenesis in the Lung

“…While the adult lung is a normally quiescent organ with little cellular turnover at homeostasis, there is a high level of active Wnt signaling as noted by the finding that more than two-thirds of the adult lung mesenchyme exhibits Wnt-signaling pathway responsiveness as reported by the Axin2 CreERT2:tdT mouse line. This line faithfully reports Axin2 gene expression and thus Wnt signaling responsiveness in a pattern identical to the Axin2 lacZ line (Choi et al, 2013; Frank et al, 2016). The broad extent of active Wnt signaling during homeostasis in the mesenchyme of a quiescent organ such as the lung suggests that this pathway is playing a role in maintaining quiescence or priming the tissue for an injury response.…”

“…We used a recently described Axin2 CreERT2:tdT reporter line generated in our laboratory (Frank et al, 2016), along with a Wnt2 CreERT2 line (Peng et al, 2013), and a Pdgfrα EGFP reporter line (Hamilton et al, 2003), to de-convolute the complexity of the mesenchymal compartment in the adult lung by identifying the relative location and distribution of mesenchymal cells displaying Wnt or Pdgfr signaling activity and expression. Wnt-responsive, Axin2+ cells can be found throughout the adult lung mesenchyme, in the alveolar regions as well as surrounding the conducting airways and blood vessels (Fig.…”

“…4B and C). Since Axin2 expression quantitatively reports Wnt responsiveness in cells (Al Alam et al, 2011; Frank et al, 2016), we examined whether genetic activation of Wnt signaling could convert the AMP lineage into the MANC lineage which would have beneficial effects on promoting alveolar regeneration over fibrotic remodeling after injury. Treatment of AMPs and MANCs in culture with a Gsk3β inhibitor (CHIR) to activate Wnt signaling resulted in an increase in Acta2 and Notch3 expression specifically in the AMP population (Supplemental Fig.…”

“…The Axin2 CreERT2-tdT mouse line was generated as described (Frank et al, 2016). The Wnt2 CreERT2 mouse was described previously (Peng et al, 2013).…”

Plasticity in the lung: making and breaking cell identity

“…Alternatively, this potential to restore alveoli may be evoked in 'ordinary' type 2 cells after ablation . Along these lines, a type 2 cell subset, characterized by Wnt-responsive Axin2 expression, has recently been identified, and these progenitors have a particularly robust capacity to generate alveolar epithelium (Frank et al, 2016). It is likely that further subsets of type 2 epithelial cells will emerge with further scrutiny.…”