Fig 2: Evaluation of the healing of small full-thickness wounds treated with ECM scaffolds.a Workflow for evaluating skin wound healing. b The surgical process for skin excisional wound model of Ctrl_SW and ECM_SW group. c Representative H&E images of Ctrl_SW and ECM_SW samples. d Histologic quantification of de novo HFs on POD28 (Data are presented as mean ± SD, n = 5 biologically independent samples, two-tailed t-test, ****p = 0.000001). e The anatomical structure of samples. f Spatial feature plot showing the expression of Cd3d (marker gene of T cells) and Crabp1 (marker gene of papillary fibroblasts) in ST profile and corresponding quantitative analysis. p value: *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

Fig 3: Spatial atlas of cell microenvironment around biomaterials of immunodeficient mice.a The anatomical structure of each sample. b Gene enrichment analysis between WT and Rag2-/- group. c Spatial feature plot and violin plot showing the distribution and expression level of integrated PF2 subcluster in tissue sections. d Spatial feature plot and violin plot showing the distribution and expression level of Crabp1 (marker gene of PF2) in tissue sections; Representative IF images of stained PF2 (CRABP1+), white arrowheads showing the CRABP1+ cells. e Spatial feature plot and violin plot showing the distribution and expression level of integrated RF2 subcluster in tissue sections. f Spatial feature plot and violin plot showing the distribution and expression level of Mest (marker gene of RF2) in tissue sections; Representative IF images of stained RF2 (MEST+), white arrowheads showing the MEST + cells. g Spatial feature plot and violin plot showing the distribution and expression level of integrated AIM2 subcluster in tissue sections. h Spatial feature plot and violin plot showing the distribution and expression level of Mrc1 (marker gene of AIM2) in tissue sections; Representative IF images of stained AIM2(F4/80 +CD206+), white arrowheads showing the F4/80 +CD206+ cells. i Spatial feature plot and violin plot showing the distribution and expression level of integrated Treg2 subcluster in tissue sections. j Spatial feature plot and violin plot showing the distribution and expression level of Foxp3 (marker gene of Treg2) in tissue sections; Representative IF images of stained Treg2 (FOXP3+), white arrowheads showing the FOXP3+ cells.

Fig 4: Bulk RNA-seq analysis of cells sorted by surface marker.a In situ hybridization of E16.5 mouse embryo showing dermis. eYFP, Pdgfra, and Thy1 signals were observed at dermis while only DAPI (blue color) was observed at epidermis. b Section staining of E16.5 mouse embryo showing dermis. eYFP, Pdgfra, and Crabp1 signals were observed at dermis while only DAPI (blue color) was observed at epidermis. c Section staining of E16.5 mouse embryo showing muscle fiber and brown adipose tissue. While DAPI and YFP signal were observed at both locations, muscle and brown adipose tissue were respectively stained by MF20 and Pparg antibodies. d In vitro culture assay of E14.5 YFP+ cells sorted by the combination of Pdgfra and Thy1 antibodies. Scale bar, 100 µm. e Bar plot of Oil Red O intensity between cultured samples. Student’s t-test was performed between two culture conditions. Values are means ± SEM, n = 3; ns not significant, ***P < 0.001. f Culture schematic and RT-qPCR result of E14.5 Pdgfra+/Thy1− cells (LP) before and after TGFβ1 ligand treatment. Two-way ANOVA was performed followed by Dunnett’s multiple comparisons test. Values are means ± SEM, n = 3; ns, not significant; **P < 0.01,***P < 0.001, ****P < 0.0001. g Venn diagrams of highly variable gene sets between sorted and cultured samples. Highly variable genes between two populations were selected. Key genes found in the corresponding sets are listed. Genes with normalized expression data > 0.3 were kept as set in the diagram.

Fig 5: Imunostain validations of RNA-Seq predicted gene expression patterns.The tissues sections shown are of one half of the E14.5 forming palate, with the medial edge epithelium at the left side of the image. For Agtr2 the RNA-Seq strongest expression was in the lateral and oral compartments, consistent with the immunostain results. Crabp1 immunostain showed strongest expression in the lateral compartment, as predicted by RNA-Seq. Both App and Ibsp showed strongest expression in the nasal compartment (above), compared to oral compartment (below), as predicted by RNA-Seq.