Figure 9

Activated BMP signaling during muscle regeneration. Parallel section ((A-D) and (E-H) represent images of 2 parallel sections) of the regenerating gastrocnemius muscle 4 days after injury were analyzed for Lama (A,E), MyoD (B) and p-Smad (F). Nuclei are stained with DAPI (C,G). Boxed areas in the overlay images (D,H) are enlarged in (D',H'). Both, MyoD (A-D,D') and p-Smad-positive nuclei (E-H,H') are located beneath the basement lamina of regenerating myofibers (white arrows in D',H'). (J) Double immunostaining of MyoD, detected with a goat anti-MyoD antibody (green), and p-Smad, detected with a rabbit anti-p-Smad antibody (red), revealed co-localization of both proteins in a subset of nuclei (yellow, arrows) in the regenerating tissue. Nuclei were counterstained with DAPI (blue). Magnification is 400×. (I) Analysis of Chordin expression by RT-PCR shows highest expression levels after 10 days of injury (n = 6 control animals, n = 2 animals at day 1 and n = 3 animals at day 4 and day 10).