Figure 4.

Implanted decellularised skeletal muscles allow functional muscle regeneration in a mouse model of VML. (a) Schematic diagram showing the surgical strategy used to generate VML model and implantation of acellular scaffolds. (b) Representative images showing the surgical procedure used for scaffold implantation. EDL muscle was exposed (A) and a first anastomosis was performed between the proximal edge of EDL muscle and the scaffold (B). The second anastomosis was carried out to the distal tendon of the EDL, then the scaffold length adapted to the implant size and the native EDL muscle resected between the two anastomosis (C). No-absorbable stitches are blue. Implant was relocated in the original anatomic position (D) and skin sutured (E). (c) Representative images showing macroscopic appearance of untreated contralateral (A) and implanted (B) mouse hindlimb 2 months after implantation. Arrow points at the proximal stitch used to anchor the scaffold to the native EDL muscle (blue). (d) Representative images showing macroscopic appearance of contralateral untreated EDL muscle (A), negative control muscle (B) and implanted scaffold (C) 2 months after implantation and after tibialis anterior dissection. (e) Representative images showing macroscopic appearance of contralateral untreated (UNTD, left) and LatB-, DET- and SDS-implanted EDL muscles (right) 2 months after implantation. (f) Mean weight of contralateral untreated EDL muscles (UNTD) and LatB-, DET- or SDS-implanted muscles two months after implantation. Data are shown as mean ± s.e.m.; n = 8-7, each group. No significant differences were observed among the groups. (g) Ex vivo force measurement of untreated EDL muscles, and LatB-, DET- and SDS-implants two months after implantation. Data are shown as mean ± s.e.m.; n = 6, each group. No significant differences were observed among the groups. (h) Ratio of the maximal force generated ex-vivo from regenerated implants (LatB, DET and SDS) and contralateral untreated EDL muscles two months after implantation. Data are shown as mean ± s.e.m.; n = 6, each group. No significant differences were observed among the groups. (i) Representative confocal images of cross-sections from contralateral untreated EDL muscles, and LatB-, DET- and SDS-implanted scaffolds stained with bungarotoxin (BTX - red) and synaptophysin (SYP - green) two months after implantation. Nuclei were stained with dapi (blue) and phase contrast (PhC) is shown to highlight staining localization (scale bar: 10 µm). (j) Percentage of neuromuscular junctions in untreated and implanted scaffolds calculated as percentage of SYP/BTX double-positive cells. **P < 0.01; ***P < 0.001 each decellularised sample compared to untreated muscles.