Fig 2: Topical ASA Creams Can Improve Adult HI Skin Differentiation(A) Overview of experimental design.(B) Survival percentage graph based on health-euthanasia thresholds as an estimate of treatment tolerance (n = 3–10).(C) Relative urine salicylic acid content from topical treatments determined from Trinder assay (n = 3–5).(D) Day 11 body weight as a percentage of day 5 treatment starting weight (n = 3–9); unbolded. The p values are relative to the AD cream-treated mice of matched genotype, while bolded p values are relative to the untreated mice of the matched genotype.(E) Graph of the total epidermal thickness from treatments and genotypes indicated (n = 3–12). The p values are relative to the AD cream-treated mice of the matched genotype, with the exception of the AD cream itself, which is relative to the untreated mice.(F) H&E staining of Abca12?/? and Abca12f/f mice treated with AD creams as indicated (n = 3–8).(G) Immunostaining of Abca12?/? and Abca12f/f skins (treated as indicated) for KRT14 and/or PPL counterstained with DAPI, as indicated (n = 3–5).(H) Immunostaining of Abca12?/? and Abca12f/f skins (treated as indicated) for KRT10 and/or KRT6a counterstained with DAPI, as indicated (n = 3–5).All data are from day 11 of treatment. All of the mice are 4OHT treated unless indicated. 5ASA + acitretin refers to 3% 5ASA + 0.02% combination acitretin treatment. The error bars are SEMs. Scale bars, 50 µm.

Fig 3: Primed cell-state is peculiar of infundibular Lrig1 GL progeny and maintained in ageing.a, Whole-mounts of Lgr5 GL HF triplets at the indicated time point. b-f, Single cell data from Lgr5 GL cells. UMAP of cells coloured by time points (b), clusters (c), replicates (d) or cell-cycle (e). f, Glut1 and Krt6a expressions plotted on UMAP. Scale in log(counts). g, Whisker plots of the average expression of each GO term enriched in Fig. 4b. Data are median with 25th and 75th percentiles. n = 46 (0w_cl5), n = 39 (1w pw1_cl7), n = 17 (1w pw1_cl0), n = 16 (8w pw1_cl5), n = 26 (1w pw2_cl7), n = 16 (1w pw2_cl0) cells. h, Left panel: Smoothed Relative Expression (SRE) of deregulated genes in the indicated cluster for Lrig1 GL (up) and Lgr5 GL (down) cells. Cell plasticity genes in the black rectangle. Right panel: percentage of cell plasticity genes are shown for ‘A’, ‘B’ or ‘C’ cells. Wound-primed cells at 8w pw1 in Transition Cluster (‘C’) only exists in Lrig1 GL cells. i, GO analysis for the deregulated genes in cell subsets ‘B’ and ‘C’, as -log10 of adjusted p-value (-log10(AdjP)). Dotted lines represent significance. j, Venn diagram of the genes in ‘A’ and ‘B’ for Lgr5 and Lrig1 GL populations. k, Ex vivo migration assay. Exit area (mm2) of Lrig1 GL cells from explants collected at 40w pw1 from memory (Distal- 2 to 7 mm from wound bed) and control areas (Ctrl- > 2 cm from wound bed), related to Fig. 6e. n = 4 (Ctrl), n = 8 (Distal). l, m, scRNA-seq of Lrig1 GL cells at the two aged homeostasis: 40w and 40w pw1. UMAP of cells coloured by replicate (l) or by cell cycle phase (m). n, Krt6a expression levels. Scale in log(counts). o, Quantification of Krt6 (left) in the infundibulum from epidermal whole-mount (right). n = 3 (40w), n = 10 (40w pw1_Distal). p, Epidermal whole-mount of pS6 staining (green) at 40w or 40w pw1_Distal, related to Fig. 6k. q, MitoTracker staining in whole-mount infundibulum and quantification. n = 5 (40w), n = 12 (40w pw1_Distal) wounds. P-value (P) from a two-tailed t-test. Data are mean ± SD, if not differently indicated. Scale bars: 50 μm (a, o, p, q). scRNA-seq data (b-j) are the integration two independent experiments, each of them based on 4 biological replicates. Source data

Fig 4: Injury selectively induces the proliferation of wild-type cells in HrasG12V/+ mosaic skin.a, Representative two-photon images of the epidermal preparation, immunostained for p-histone H3 (highlighted by white dashed circle). b, Quantification of p-histone H3-positive (p-H3+) cells in tdTomato+ and tdTomato− populations in injured (I) and uninjured (U) skin. n = 4 mice. c,d, Quantification of mitotic cells in uninjured (c) and uninjured (d) ears. n = 4 mice. e, Representative two-photon images of the epidermal preparation, immunostained for keratin-10 (white). f, Quantification of keratin-10-positive (KRT10+) cells in tdTomato+ and tdTomato− populations in injured and uninjured skin. n = 3 mice. b–d,f, Paired or unpaired two-tailed t-test for comparison between tdTomato+ and tdTomato− populations in the same group or in different groups of mice. At least three independent areas of approximately 300 µm2 were analysed for each mouse (Methods). KRT10, keratin-10. g, Uniform manifold approximation and projection (UMAP) showing epidermal keratinocyte clusters from uninjured and injured conditions. h, Violin plots (left) showing gene expression in clusters from g, together with the gene expression superimposed on the UMAP (right). i, Cell classification into homeostatic and injury-responsive populations based on clustering and gene expression, overlaid on the UMAP. g–i, n = 12 mice. j, Quantification of wild-type and HrasG12V/+ cells as a proportion of tdTomato+ cells in basal non-committed and basal committed groups for homeostatic and injury-responsive cells. Data are averaged results for each biological replicate. One-sided t-test with Holm–Sidak correction. k, Violin plots showing Krt6a and Krt10 expression in cells grouped as in j. In h,k, internal box plots denote the 25th, 50th and 75th centiles, with whiskers depicting minima and maxima, excluding outliers that are beyond 1.5× the interquartile range. j,k, Homeostatic cells from n = 12 mice and injury-responsive cells from n = 6 mice. P values are shown. Data are mean ± s.d.Source Data