Figure 8: Measurement of host muscle fibre size and tissue vascularization after intramuscular transplantation with young WT-MDSPCs.

(a) The gastrocnemius muscle of 12-day-old Ercc1−/− mice (n=3 mice) was injected with young WT-MDSPCs. Tissues were isolated 5 days later (17 d) and sections were immunostained for dystrophin (green), CD31 (red), and DAPI (blue) and compared with untreated 15-day-old WT and Ercc1−/− mice (from Fig. 7). The white arrow denotes a centronucleated fibre. Scale bar=100 μm. (b) Quantification of microvasculature and myofibre size in transplanted and untreated mice. Images from 4–8 sections from 3 animals per group were analysed. Reported is the ratio of CD31+ cells to dystrophin-positive muscle fibres (left), §P<0.05 between untreated and treated Ercc1−/− mice, Tukey's test. On the right is the average muscle fibre size (cross-sectional area), §P<0.05 between untreated and treated Ercc1−/− mice, Dunn's test. Error bars indicate s.e.m. (c) Seventeen day-old progeroid Ercc1−/− mice (n=10) were injected IM with WT-MDSPCs expressing nuclear LacZ to determine whether donor cells integrate into muscle fibres and contribute to blood vessels. Five days later, the mice were euthanized and the gastrocnemius muscles were isolated for analysis. Muscle sections at the injection site were stained with X-gal, to detect donor cells, and eosin. Scale bar=50 μm. (d) Histogram of the cross-sectional area distribution of myofibres in the treated gastrocnemius muscles (black bars) compared the contralateral non-injected muscle (white bars). Data were collected from 6,000 fibres from multiple sections of four mice per group. The distribution of fibre size is indicated on the x-axis, representing increasingly more mature fibres with increased size (P<0.001, Mann–Whitney rank sum test at 151–300 and 301–450). (e) Sections from the injected muscles stained for CD31 to identify blood vessels (dark brown) and X-gal to identify donor cells (blue). Red arrowheads indicate CD31+ capillary structures. Sections from dystrophic (mdx/SCID) mice treated the same way are included as a positive control (right panels). Scale bars=50 μm.