Figure 3.

The proliferative state of the myofibroblast (αSMA-expressing fibroblast) changes during alveolarization. Representative FACS histograms from freshly isolated mouse lung fibroblasts (A-C). Fibroblasts were stained with phycoerythrin (PE)-α-smooth muscle actin antibody (x-axis) plus anti-Ki67 followed by a secondary antibody conjugated to Alexa Fluor 647 (A647, y-axis). Some samples were stained with only the A647 secondary antibody (A), non-immune IgG control for ki67 + secondary antibody and non-immune IgG control for α-smooth muscle actin (B), or with the anti-ki67 and anti-αSMA antibodies (C). Proportions of fibroblasts that stained positively or negatively for either of the antibodies was determined, averaged and summarized in column graphs for P4 (D) and P12 (E). The fibroblast populations represent a mixture of PDGF-Rα-GFP-positive and negative subpopulations. We determined that the majority of αSMA-containing fibroblasts expressed the PDGF-Rα at both P4 and P12 (F). The shaded bars represent the αSMA-containing fibroblasts. Background compensation was determined from the non-immune IgG controls prior to calculating the proportions of the different fibroblast populations. For the mice ages P4, n = 5 groups of 2-4 mice, and for P12, n = 7 groups of 2-4 mice. #, P < 0.05 and *, P < 0.001. For the FACS histograms, on the x-axis, "PE-A" = PE Area and on the y-axis, "A647-A" = A647 Area.