Fig. 3.
SCs deficient in Slug exhibit intrinsic defects in self-renewal in vivo. a Ratio of SCs between SlugcKO and Ctrl mice under indicated conditions. ***p < 0.001 by student’s t-test. b IHC for Pax7+ SCs in TAs of SlugcKO and Ctrl mice (n = 3 mice per group) at day 30 post single injury. Laminin indicates the boundaries of myofibers. Scale bar, 100 μm. Three independent experiments were performed with similar results. c Quantification of Pax7+ SC numbers in b **p < 0.01 by student’s t-test. d Scheme of SC transplantation. 3000 SCs of indicated genotype were transplanted into each side of pre-injured TA muscles of mdx recipients (n = 6), respectively. Donor-derived SCs (GFP+) in total recipient MuSCs (CD45−CD31−Sca1−Vcam I+) were analyzed 4 weeks after transplantation. e Representative flow cytometric analysis of the frequency of donor-derived SCs (GFP+) within total recipient MuSCs. Similar results were seen in three independent transplantation experiments. f Percent of donor-derived SCs (GFP+) in total recipient MuSCs. *p < 0.05 by student’s t-test. g Diagram of competitive MuSC repopulation assay. SCs from Slug+/+GFP- and Slug−/−GFP-transgenic mice were equally mixed for genomic DNA extraction and transplantation, respectively. The remaining cells were then transplanted into pre-injured TA muscles of mdx recipients. Four weeks after transplantation, GFP+ cells were sorted from total recipient MuSCs for genotyping PCR analysis. h Determination of relative ratio of repopulated MuSCs. Left, representative flow cytometric plotting of GFP+ cells from total recipient MuSCs. Right, PCR determining the ratio of SCs before and after transplantation. M DNA ladder, MBT mixed SCs before transplantation, MAT mixed SCs after transplantation, Ng negative control. i Scheme of AraC treatment indicating self-renewal. j Pax7 and Ki67 immunostaining in cultured myofibers as treated in i. Scale bar, 50 µm. Similar results were seen from three independent experiments. k Quantification of the quiescent daughter SCs stained in j (n = 3 mice, >20 myofibers per condition). ***p < 0.001 by student’s t-test. Data are shown as mean ± SEM of three independent experiments. Unprocessed gel blots are provided in the Supplementary Fig. 1. Also see Supplementary Fig. 4-6. Source data are provided as a Source Data file