Transgenic overexpression of ADAM12 suppresses muscle regeneration and aggravates dystrophy in aged mdx mice - PubMed (original) (raw)

Transgenic overexpression of ADAM12 suppresses muscle regeneration and aggravates dystrophy in aged mdx mice

Louise Helskov Jørgensen et al. Am J Pathol. 2007 Nov.

Abstract

Muscular dystrophies are characterized by insufficient restoration and gradual replacement of the skeletal muscle by fat and connective tissue. ADAM12 has previously been shown to alleviate the pathology of young dystrophin-deficient mdx mice, a model for Duchenne muscular dystrophy. The observed effect of ADAM12 was suggested to be mediated via a membrane-stabilizing up-regulation of utrophin, alpha7B integrin, and dystroglycans. Ectopic ADAM12 expression in normal mouse skeletal muscle also improved regeneration after freeze injury, presumably by the same mechanism. Hence, it was suggested that ADAM12 could be a candidate for nonreplacement gene therapy of Duchenne muscular dystrophy. We therefore evaluated the long-term effect of ADAM12 overexpression in muscle. Surprisingly, we observed loss of skeletal muscle and accelerated fibrosis and adipogenesis in 1-year-old mdx mice transgenically overexpressing ADAM12 (ADAM12(+)/mdx mice), even though their utrophin levels were mildly elevated compared with age-matched controls. Thus, membrane stabilization was not sufficient to provide protection during prolonged disease. Consequently, we reinvestigated skeletal muscle regeneration in ADAM12 transgenic mice (ADAM12(+)) after a knife cut lesion and observed that the regeneration process was significantly impaired. ADAM12 seemed to inhibit the satellite cell response and delay myoblast differentiation. These results discourage long-term therapeutic use of ADAM12. They also point to impaired regeneration as a possible factor in development of muscular dystrophy.

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Figures

Figure 1

Figure 1

Sirius staining of 1-year-old normal (A), mdx (B), and ADAM12+/mdx (C) mice showing considerable fibrosis and adipose infiltration in the ADAM12+/mdx mice compared with both normal and mdx mice. Original magnifications, ×100.

Figure 2

Figure 2

Utrophin staining of 1-year-old ADAM12+/mdx (A) and mdx (B) mice. Utrophin expression remains mildly elevated in the old ADAM12+/mdx mice compared with the age-matched controls. Negative controls for ADAM12+/mdx (C) and mdx (D). Original magnifications, ×200.

Figure 3

Figure 3

H&E stainings of ADAM12+ TG mice (B and D) and their LCs (A and C). Original magnifications, ×200.

Figure 4

Figure 4

Morphometrical analysis of ki-67 protein expression pattern during regeneration of ADAM12+ and normal LCs. Black squares: ADAM12+ mice ±1 SE; open squares: LC mice ±1 SE (P = 0.1) (n = 2 to 3).

Figure 5

Figure 5

Semiquantitative analysis of mRNA expression levels of pax7 (A), myoD (B), myogenin (C), and NCAM (D) during regeneration of ADAM12+ and normal LC mice (P < 0.01 for all markers). Black bars: ADAM12+ mice; white bars: LC mice (n = 2 to 3).

Figure 6

Figure 6

Morphometrical analysis of pax7 (A), myogenin (B), and NCAM (C) protein expression patterns during regeneration of ADAM12+ and normal LC mice. Black squares: ADAM12+ mice ±1 SE; open squares: LC mice ±1 SE (Pax7, P < 0.001; myogenin, P < 0.05; NCAM, P = 0.27) (n = 2 to 3).

Figure 7

Figure 7

NCAM staining of the regenerating area at day 5 after injury reveals differing morphologies in ADAM12+ mice (A) and normal LCs (B). Original magnifications, ×400.

Figure 8

Figure 8

Sirius staining of the regeneration process after massive, acute injury in ADAM12+ (A–C) and LC (D–F) mice. Insets in C and F show dystrophin expression in the membranes at day 10 after injury to illustrate the end-point of regeneration. Original magnifications, ×200.

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