Figure 2
From: Increased sphingosine-1-phosphate improves muscle regeneration in acutely injured mdx mice
Dystrophic pathology following muscle injury is improved with THI treatment. ( A ) Experimental schematic of THI (0.075 μg/day) and PBS (vehicle)-treated mdx mice injected IP twice daily for the first 72 hours following CTX injury. Muscles from aged mdx4cv mice (n = 7, THI-treated: 3 × 11-MO females, 4 × 16-MO males; n = 6 vehicle-treated: 3 × 11-MO females, 3 × 16-MO males) were harvested for histopathology analysis 18 days post CTX injury. ( B ) Histological quantification of picrosirius red staining indicates lower fibrotic accumulation following injury in both TA and quadriceps (quads) muscles from mice treated with THI. For CTX-injected muscles, damaged regions of muscle (for example fields with the greatest accumulation of sirius red staining) were quantified for both THI and vehicle-treated mice. The level of fibrosis was not significantly different between treated and control (vehicle) uninjured quadriceps; however, uninjured TA muscles from 11-MO THI-treated mice had lower fibrosis compared to control TA muscles. For each muscle, three separate sections (200 μm apart in longitudinal distance) were analyzed. ( C ) Representative photographs of injured quadriceps stained with picrosirius red and fast green depict collagen deposition (red staining), while muscle morphology and organization is depicted with hematoxylin and eosin staining. Scale bars = 50 μm. ( D ) Oil Red O staining depicts fat deposits (arrows) over the entire CSA of THI-treated and vehicle-injured quadriceps from 16-MO males. Scale bars = 500 μm. ( E ) The ratio of fat deposition in injured TAs over uninjured contralateral TAs quantified from Oil Red O staining was significantly reduced in THI-treated versus control animals in 11-MO (*) but not 16-MO mdx4cv mice. In contrast, the ratio of injured over uninjured fat deposits in quadriceps was significantly reduced in 16-MO (#) but not in 11-MO mdx mice. *P <0.05, **P <0.01 by student’s t-test. Error bars represent SEM. CSA, cross-sectional area; CTX, cardiotoxin; IP, intraperitoneal; MO, month-old; SEM, standard error of the mean; TA, tibialis anterior; THI, 2-acetyl-4(5)-tetrahydroxybutyl imidazole.