Fig. 3
Loss of ALKBH5 impairs wound re-epithelialization in mice. a Schematic ideogram showing the strategy for the deletion of the ALKBH5 gene using CRISPR/Cas9 technology. b, c The loss of ALKBH5 expression in the skin and epidermis of Alkbh5‒/‒ mice was confirmed by WB (b) and IF (c), respectively. The full-length blots are presented in Additional file 8: Fig. S1. d The global m6A level of skin at the wound edge of WT and Alkbh5‒/‒ mice was measured by m6A DB assays. Total RNA was determined by methylene blue staining (as a loading control). The experiments were performed in triplicate, and the relative m6A modification contents are shown as the mean ± SD. t test, two-sided *P < 0.05. e IF revealed the loss of ALKBH5 expression in the epidermis at the wound edge of Alkbh5‒/‒ mice. Scale bar: 100 μm. Dotted lines denote epidermal boundaries. Epi epidermis. f, g Representative images of cutaneous wounds (f) of WT and Alkbh5‒/‒ mice on PWD0, PWD2, PWD4, PWD6, and PWD8. Scale bar: 2 mm. The black dashed circle delineates the original wound of 5 mm width. Rates of wound closure (g) were quantified by using ImageJ software and are expressed as the percentage of the nonhealing area. Twelve wounds of six mice were included in the analysis. The relative percentages of wound closure are shown as the mean ± SD. t test, two-sided **P < 0.01, ***P < 0.001. h, i Histological analysis (h) of wound re-epithelialization in WT and Alkbh5‒/‒ mice at PWD8. The line with arrowheads indicates the 5 mm width of the original wound gap. The line with blunt ends delineates the epithelial gap, which represents the nonepithelialized wound area. Scale bar: 1 mm. The percentage of re-epithelialization was quantified (i) in WT and Alkbh5‒/‒ mice at PWD8. Twelve wounds of six mice were included in the analysis. The relative percentages of re-epithelialized wound area are shown as the mean ± SD. Mann–Whitney test, two-sided ****P < 0.0001. Scale bar: 1 mm. Dotted lines denote epidermal boundaries