Integrin α8 is a useful cell surface marker of alveolar lipofibroblasts

Recent advances in comprehensive gene analysis revealed the heterogeneity of mouse lung fibroblasts. However, direct comparisons between these subpopulations are limited due to challenges in isolating target subpopulations without gene-specific reporter mouse lines. In addition, the properties of lung lipofibroblasts remain unclear, particularly regarding the appropriate cell surface marker and the niche capacity for alveolar epithelial cell type 2 (AT2), an alveolar tissue stem cell.

Using cell surface markers applicable even into wild-type mouse lungs, we could classify PDGFRα⁺ total lung resident fibroblasts into at least two major distinct subpopulations: integrin α8 (ITGA8)⁺ and SCA-1⁺ fibroblasts. We analyzed their characteristics, including lipid content, transcriptome profiles, and alveolar stem cell niche capacity. ITGA8⁺ fibroblasts showed higher positivity of intracellular lipid droplets compared to SCA-1⁺ fibroblasts (91.0 ± 1.5% vs. 5.0 ± 0.5% in LipidTOX staining; 91.3 ± 1.4% vs. 7.1 ± 1.7% in Oil Red O staining). The fluorescence intensity of LipidTOX in the ITGA8⁺ fibroblasts was highest in newborn compared to adult or aged lungs. The transcriptome profile of ITGA8⁺ fibroblasts in adult mouse lungs, evaluated through two independent single-cell RNA-seq datasets, consistently showed higher expression of Tcf21 and Plin2, which are canonical markers of lipofibroblasts. ITGA8⁺ fibroblasts were primarily located in the alveolar area, particularly in the neighborhood of AT2. Compared to SCA-1⁺ fibroblasts, ITGA8⁺ fibroblasts showed higher mRNA expression of potential AT2-supportive factors, Fgf10, Fgf7, and Wnt2, but unexpectedly, exhibited lower efficiency in alveolar organoid formation.

ITGA8⁺ lung fibroblasts correspond to alveolar lipofibroblasts, but the alveolar niche capacity may be lower than SCA-1⁺ lung fibroblasts. Further studies are necessary for the functional distinction between lung fibroblast subpopulations.