Fig. 5: MuSC-specific Lama2 knockout slows MuSC proliferation ex vivo.
a Experimental approach: mice with MuSC-specific Lama2 knockout (MuSC-Lama2KO) were homozygous for the floxed Lama2 allele, while control littermates were wild-type for the Lama2 allele (control). Both were heterozygous for Pax7CreER and the EGFP knock-in. Mice were treated for 5 consecutive days with tamoxifen and collected 3 days later for isolation of EGFP+ cells and single fibers. b PCR on DNA isolated from sorted EGFP+ control and MuSC-Lama2KO cells. Product size after deletion of exon 3 = 433 bp; product size of wild-type allele = 755 bp. c Representative anti-laminin-α2 staining in tissue culture dishes with (upper panels, full stain) or without (lower panels, negative control) secondary antibodies (EGFP in green, laminin-α2 in magenta, DAPI in blue). d Quantification of the proportion of EdU+/Pax7+/EGFP+ after incubation with EdU in the medium from 0–6 and 6–21 h in culture. Cells were isolated from n = 5 mice. e Representative immunostaining of single EDL fibers isolated from control and MuSC-Lama2KO mice and stained after 0 (T0), 24 (T24), 42 (T42) and 72 (T72) h in culture (EGFP in green, Ki67 in orange, laminin-α2 in magenta, DAPI in blue). f Quantification of the proportion of Ki67+ and Ki67− EGFP+ cells at T0, T24, T42 and T72. g Quantification of the size of clusters formed by dividing EGFP+ cells at T0, T24, T42 and T72. Cell-cell contact between EGFP+ cells was a requisite to include them as a cluster of 2 or more cells. h Quantification of the proportion of Pax7+/Myogenin−, Pax7+/Myogenin+, Pax7–/Myogenin+ and Pax7−/Myogenin− EGFP+ cells at T0, T24, T42 and T72. In f–h, a minimum of 20 fibers were analyzed per mouse at each time point; n = 4 mice. Data are means ± SEM. In all graphs, statistical significance was determined by two-way ANOVAs with Bonferroni’s multiple comparisons test. *P < 0.05; **P < 0.01. Source data and P-values are provided as a Source Data file. a Created in BioRender. Ruegg, M. (2025) https://BioRender.com/d18khcw.
