Transcription factor FOXF1 identifies compartmentally distinct mesenchymal cells with a role in lung allograft fibrogenesis
Russell R. Braeuer, Natalie M. Walker, Keizo Misumi, Serina Mazzoni-Putman, Yoshiro Aoki, Ruohan Liao, Ragini Vittal, Gabriel G. Kleer, David S. Wheeler, Jonathan Z. Sexton, Carol F. Farver, Joshua D. Welch, Vibha N. Lama
Russell R. Braeuer, Natalie M. Walker, Keizo Misumi, Serina Mazzoni-Putman, Yoshiro Aoki, Ruohan Liao, Ragini Vittal, Gabriel G. Kleer, David S. Wheeler, Jonathan Z. Sexton, Carol F. Farver, Joshua D. Welch, Vibha N. Lama
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Concise Communication Cell biology Pulmonology

Transcription factor FOXF1 identifies compartmentally distinct mesenchymal cells with a role in lung allograft fibrogenesis

Abstract

In this study, we demonstrate that forkhead box F1 (FOXF1), a mesenchymal transcriptional factor essential for lung development, was retained in a topographically distinct mesenchymal stromal cell population along the bronchovascular space in an adult lung and identify this distinct subset of collagen-expressing cells as key players in lung allograft remodeling and fibrosis. Using Foxf1-tdTomato BAC (Foxf1-tdTomato) and Foxf1-tdTomato Col1a1-GFP mice, we show that Lin–Foxf1+ cells encompassed the stem cell antigen 1+CD34+ (Sca1+CD34+) subset of collagen 1–expressing mesenchymal cells (MCs) with a capacity to generate CFU and lung epithelial organoids. Histologically, FOXF1-expressing MCs formed a 3D network along the conducting airways; FOXF1 was noted to be conspicuously absent in MCs in the alveolar compartment. Bulk and single-cell RNA-Seq confirmed distinct transcriptional signatures of Foxf1+ and Foxf1– MCs, with Foxf1-expressing cells delineated by their high expression of the transcription factor glioma-associated oncogene 1 (Gli1) and low expression of integrin α8 (Itga), versus other collagen-expressing MCs. FOXF1+Gli1+ MCs showed proximity to Sonic hedgehog–expressing (Shh-expressing) bronchial epithelium, and mesenchymal expression of Foxf1 and Gli1 was found to be dependent on paracrine Shh signaling in epithelial organoids. Using a murine lung transplant model, we show dysregulation of epithelial-mesenchymal SHH/GLI1/FOXF1 crosstalk and expansion of this specific peribronchial MC population in chronically rejecting fibrotic lung allografts.

Authors

Russell R. Braeuer, Natalie M. Walker, Keizo Misumi, Serina Mazzoni-Putman, Yoshiro Aoki, Ruohan Liao, Ragini Vittal, Gabriel G. Kleer, David S. Wheeler, Jonathan Z. Sexton, Carol F. Farver, Joshua D. Welch, Vibha N. Lama

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Figure 2

In situ niche of FOXF1-expressing Col1+ MCs.

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In situ niche of FOXF1-expressing Col1+ MCs. (A–C) Adult lungs of Col1GF...
(A–C) Adult lungs of Col1GFP mice were stained with FOXF1 (red) and CCSP (white). Representative high-resolution wide-field image of entire lung (A) and 4 locations along the bronchial tree (B). Scale bars: 50 μm (B). Arrows indicate FOXF1+Col1+ MCs along the BVB. (C) Quantification of nuclear FOXF1 intensity in Col1GFP+ cells in the bronchovascular space (Bronchi) and the alveolar interstitium (Alveoli). n = 3. (D) Confocal images and quantification of PDGFRα-GFP mouse lung tissue stained with anti-FOXF1 antibodies. Images show colocalization of FOXF1 (red) and PDGFRα (green) in the peribronchial region (cyan arrows), with no costaining noted in alveoli. n = 3. Scale bars: 20 μm. (E) Adult human lung tissue sections stained with FOXF1 (green) and the marker anti–smooth muscle actin (αSMA; red). Scale bars: 50 μm. AW, airway; BV, blood vessel. Data indicate the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001, by 1-way ANOVA with Bonferroni’s test C) and unpaired t test (D).