Figure 1.

Chd4 is required for the proliferative expansion of SCs during muscle regeneration

(A) Chd4 mRNA relative expression in QSCs, and in SCs under proliferative (in growth medium [GM]) or differentiation (in differentiation medium [DM]) conditions at the indicated time points in culture from WT mice. Data were normalized to housekeeping gene Rpl7 with the QSCs set to 1 (from left to right, n = 13, 9, 6, 7, 9, or 7 mice, respectively).

(B) Representative pictures and quantification of Chd4 in SCs (Pax7⁺) from non-injured muscles from WT mice, or at 72 h after muscle injury with CTX (n = 33 or 60 cells, respectively, from four mice). The sum of the Chd4 fluorescence intensity per cell is represented. Scale bar, 5 μm.

(C) Chd4 mRNA expression in freshly isolated SCs from non-injured muscles and at 72 h after muscle injury with CTX. Data were normalized to housekeeping gene Rpl7, with the QSCs set to 1 (n = 12 or 9 mice, respectively).

(D) Representative pictures of sections of regenerating muscles from TMX-treated Chd4^fl/f and Chd4^ΔPax7ER mice at day 7 after injury, stained with hematoxylin and eosin and antibodies against eMHC. Scale bar, 10 μm (top), 5 μm (bottom). Frequency distribution of cross-sectional area (μm²) of regenerating fibers from TMX-treated Chd4^fl/f and Chd4^ΔPax7ER mice (n = 4 and 5 mice, respectively) (bottom).

(E) Average time until first division during live time-lapse microscopy, and percentage of dividing SCs from TMX-treated Chd4^fl/f and Chd4^ΔPax7ER mice (138 or 58 individual cells from three different mice, respectively).

(F) Percentage of BrdU⁺ SCs cultured in GM for either 72 or 120 h (left), the number of SC colonies with more than five cells counted throughout 4 days after isolation and culture in GM (middle), and the number of SCs per well after seeding 50 cells/well at time 0 (right), from TMX-treated Chd4^fl/f and Chd4^ΔPax7ER mice (n = 4 independent experiments).

(G) Relative number of total and proliferating SCs after FACS purification from TMX-treated Chd4^fl/f or Chd4^ΔPax7ER mice at 3 days after injury (n = 4 mice/group).

(H) Representative pictures and quantification of proliferating (Pax7⁺/Ki67⁺; n = 4 mice/group) or differentiating cells (Myog; n = 5 mice/group) in muscles of TMX-treated Chd4^fl/f or Chd4^ΔPax7ER mice at 7 days after injury. Scale bar, 50 μm.