Fig. 2. EPFs and ENFs are two distinct lineages of fibroblasts.
(A) Scatter plot depicting transcriptome-wide expression differences between uncultured FACS-isolated EPF and ENF populations from P56 En1Cre;R26mTmG mice. For each population (n = 3), the median expression of each gene is plotted. Within the scatter plot, gene density is represented as a heat map, with orange regions containing larger numbers of genes than blue regions. Outliers are shown as individual points. (B) Similar expression of known fibroblast markers and fibroblast-related genes (left) and ECM remodeling genes (right) compared between uncultured FACS-isolated EPF and ENF populations by microarray analysis. (C) Expression of top differentially regulated genes between uncultured FACS-isolated EPF and ENF populations by microarray analysis. Genes were filtered for Q value of <0.3 (30% false discovery rate) and a fold change of at least 1.5x. (D) FACS analysis showing abundance of EPFs and ENFs FACS-isolated from the dorsal dermis of E10.5 (top), E16.5 (one down from top), P1 (one up from bottom), and P30 (bottom) En1Cre;R26mTmG mice reveals a shift in population dynamics during dermal development from ENF-dominated dermis at E10.5 to EPF-dominated dermis at P1 and subsequent postnatal stages (n = 5 mice). (E) Fluorescent imaging of EPFs and ENFs cultured after FACS-sorting from the dorsal dermis of E16.5 and P30 En1Cre;R26mTmG mice. The relative abundance of EPFs and ENFs defined in the FACS analyses was consistent with the relative abundance seen after tissue culture plating at each time point. Scale bar, 100 μm. (F) Immunostaining of FACS-isolated EPFs and ENFs from P30 En1Cre;R26mTmG mice in vitro revealed that traditional fibroblast markers (vimentin and FSP-1) and secreted ECM components (col type I and fibronectin) are expressed similarly across both populations. Scale bar, 200 μm.