Table 1.
Summary and references of the primary antibodies used.
Fig 1.
Number of satellite cells and their behaviour in the 4 different injury models.
(A) Number of satellite cells in uninjured, 18h, 1 month and 3 months post-injury; n = 124 animals (ancillary and new data). The figure also displays the number of satellite cells after re-injury (i.e. after two successive lesions carried out 28 days apart; displayed in black histograms) n = 5 animals. (B-E) Percentage of remaining Pax7 positive cells (B) 18h, (C) 2 days, (D) 4 days and (E) 1 month post-injury on TA sections. (F-I) Percentage of cycling Ki67 positive satellite cells (F) 18h, (G) 2 days, (H) 4 days and (I) 1 month after injury. Data are represented as means±s.d. *p < 0.05; **p < 0.01; ***p < 0.001; no star, statistically non significant.
Fig 2.
Muscle histology at different time points after injury.
Haematoxylin and eosin staining on cryosections. (A) 18h, (B) 2 days, (C) 4 days (D) 12 days and (E) one month post freeze injury. (F) 18h, (G) 2 days, (H) 4 days (I) 12 days and (J) one month post NTX injury. (K) 18h, (L) 2 days, (M) 4 days (N) 12 days and (O) one month post CTX injury. (P) 18h, (Q) 2 days, (R) 4 days (S) 12 days and (T) one month post BaCl2 injury. Insets represent whole muscle scan (HMR). Scale bar represents 50 μm.
Fig 3.
Fibre quantification and vascularization at different time points post injury in the 4 injury models.
(A) Fibre diameters (expressed in μm) 1 month after injury in all 4 injury models (B), 6 months after injury in all 4 injury models (C,D), 1 and 6 months, respectively, in all injury models. (E) Vessel numbers per fibre 1 month after injury in all injury models. (F) Vessel numbers per fibre 6 month after injury for all injury models. Data are represented as means±s.d. *p < 0.05; **p < 0.01; ***p < 0.001; ns, statistically non significant.
Table 2.
Qualitative and semi-quantitative summary of the histological study for the four injury models at the different time points.
Table 3.
Summary of the inflammatory infiltrate in the different injury models (cell number per 10 microscopic fields).
Fig 4.
Characterization of inflammation after injury in the 4 injury models.
(A,D,G,J) Neutrophil (GR1+ cells) quantifications (expressed as number of cells per ten microscopic fields at 40X) in the FI (A), NTX (D), CTX (G) and BaCl2 (J). (B,E,H,K) Macrophage (F4/80+ cells) quantifications (expressed as number of cells per field) in the FI (B), NTX (E), CTX (H) and BaCl2 (K). Data are represented as means±s.d. *p < 0.05; **p < 0.01; ***p < 0.001; no star, statistically non significant. (C,F,I,L) Luminex (multiplex assay) measuring the levels of cytokines in pg/g of muscle tissue, 18h, 4 days, 12 days and 1 month post-injury in the freeze injury model (C), the NTX (F), the CTX (I), and the BaCl2 (L). Selected cytokines (IL6; IL10; MCP1; MIP1a and MIG) are displayed for each injury model. Data are represented as means±s.d.
Fig 5.
Three dimensional analysis of vessels at different time points in all 4 injury models.
Images show blood vessel organisation in 3D after z-stack reconstitutions of scanned sectioned TA from Flk1GFP/+ mouse. (A-F) Vessel organisation in the freeze injury, 18h (A), 2 days (B), 4 days (C), 12 days (D), 1 month (E) and 3 months (F) post injury. (G-L) Vessel organisation in the NTX injury, 18h (G), 2 days (H), 4 days (I), 12 days (J), 1 month (K) and 3 months (L) post injury. (M-R) Vessel organisation in the CTX injury, 18h (M), 2 days (N), 4 days (O), 12 days (P), 1 month (Q) and 3 months (R) post injury. (S-X) Vessel organisation in the BaCl2 injury, 18h (S), 2 days (T), 4 days (U), 12 days (V), 1 month (W) and 3 months (X) post-injury. Arrows pointing anastomoses. Scale bars represents 10 μm.
Fig 6.
Characterization of fibrosis after injury.
(A-D) Sirius Red staining (collagen deposits)1 month after injury in all 4 injury models. (E) Percentage of fibrosis per section 1 month after injury compared with non-injured control. No statistically significant differences detected among the 4 models. ns; non significant.