Skeletal myogenic progenitors originating from embryonic dorsal aorta coexpress endothelial and myogenic markers and contribute to postnatal muscle growth and regeneration - PubMed (original) (raw)

Skeletal myogenic progenitors originating from embryonic dorsal aorta coexpress endothelial and myogenic markers and contribute to postnatal muscle growth and regeneration

L De Angelis et al. J Cell Biol. 1999.

Abstract

Skeletal muscle in vertebrates is derived from somites, epithelial structures of the paraxial mesoderm, yet many unrelated reports describe the occasional appearance of myogenic cells from tissues of nonsomite origin, suggesting either transdifferentiation or the persistence of a multipotent progenitor. Here, we show that clonable skeletal myogenic cells are present in the embryonic dorsal aorta of mouse embryos. This finding is based on a detailed clonal analysis of different tissue anlagen at various developmental stages. In vitro, these myogenic cells show the same morphology as satellite cells derived from adult skeletal muscle, and express a number of myogenic and endothelial markers. Surprisingly, the latter are also expressed by adult satellite cells. Furthermore, it is possible to clone myogenic cells from limbs of mutant c-Met-/- embryos, which lack appendicular muscles, but have a normal vascular system. Upon transplantation, aorta-derived myogenic cells participate in postnatal muscle growth and regeneration, and fuse with resident satellite cells.The potential of the vascular system to generate skeletal muscle cells may explain observations of nonsomite skeletal myogenesis and raises the possibility that a subset of satellite cells may derive from the vascular system.

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Figures

Figure 1

Figure 1

Top, Morphology of embryonic structures isolated from E9.5 mouse embryos after pancreatin digestion. Bottom, RT-PCR revealed the medial markers MyoD and Pax3 were expressed in dissected somites (different ratio of Pax3 to MyoD in different lanes depends upon isolation of somites at different cranio–caudal level); Pax3, but not MyoD, in the neural tube; and none of the markers was detectable in dissected dorsal aorta.

Figure 2

Figure 2

Morphology of typical clones derived from explant cultures of E9.5 limb bud (A), somites (B), and dorsal aorta (C), cultured in growth medium. Immunofluorescence analysis with antisarcomeric myosin antibody of a clone from dorsal aorta, after three days of culture in differentiation medium is shown in D. Bar, 15 μm.

Figure 3

Figure 3

A, Quantitative analysis of satellite cell-like clones (shown in Fig. 1) from explant cultures of different anlagen of E9.5 embryos. Each bar is the average of at least three separate experiments, each performed in triplicate. B, Time course of the appearance of satellite cell-like clones from explants of vessels (___) or limb buds (- - -) at successive periods of development. The time course of the appearance of satellite cell-like clones, directly cloned from limb buds in vivo is also shown (– · –).

Figure 4

Figure 4

Myogenic and endothelial markers are expressed in aorta-derived myogenic cells and in satellite cells. Immunofluorescence analysis with antibodies against MyoD (A and B), c-Met (C and D), Desmin (E and F), VE-cadherin (G and H), and β-3 integrin (I and J) of clones derived from E9.5 dorsal aorta (A, C, E, G, and I) or P10 satellite cells (B, D, F, H, and J). Bar, 10 μm.

Figure 5

Figure 5

MyoD and VE-cadherin are coexpressed in aorta-derived myogenic cells and in satellite cells. Double immunofluorescence analysis with antibodies against MyoD (red) and VE-cadherin (green) of clones derived from E9.5 dorsal aorta (A) and of adult satellite cells (B). Bar, 10 μm.

Figure 6

Figure 6

The message for VE-cadherin is expressed in adult satellite cells. Northern blot analysis of VE-cadherin expression in E 9.5 embryonic hearts (H), adult satellite cells (CS), and primary fibroblasts (fb).

Figure 7

Figure 7

Myogenic clones are present in the limbs of c-Met–deficient embryos. Phase-contrast microscopy of wt (A) and MetD (B) E14 embryos showing normal vasculature in mutant embryos (arrow). β-galactosidase staining of wt (C) and MetD (D) crossed to MLC3F-nLacZ embryos showing complete absence of muscle (β-gal+) cells in the limb of E15 mutant embryos. Immunofluorescence analysis with antibodies against MyoD of clones derived from E13 limb buds of wt (E) or MetD (F and G) embryos. Nuclear staining (Hoechst) is shown in H, J, and K. Bar, 10 μm.

Figure 8

Figure 8

Aorta-derived myogenic cells undergo myogenesis in vitro and in vivo. A, Cocultures of clones from E9.5 dorsal aorta from MLC3F-nLacZ embryos and P10 wt satellite cells. Arrows indicate one β-gal+ and one β-gal− nucleus within the same myotube. Bar, 10 μm. B, Cross-section of a regenerating TA of a SCID/bg injected with pooled clones of satellite cell-like clones, showing a cluster of β-gal+ nuclei (arrow) within small regenerating fibers, labeled with an antibody against laminin in C. Bar, 25 μm.

Figure 9

Figure 9

Host-derived myogenic cells are present in fetal limbs transplanted under the skin of newborn MLC3F-nLacZ transgenic mice and vascularized by the host. A, Whole-mount stain reveals a cluster of β-gal+ nuclei in the transplanted limb. B–D, Cross-section of the same sample stained for β-galactosidase activity revealed the β-gal+ nuclei (B) inside myosin positive muscle fibers (D) adjacent to VE-cadherin positive vessels (C). Bar, 25 μm.

Figure 10

Figure 10

Aorta-derived myogenic cells contribute to growing fibers and also circulate. A, Whole-mount stain reveals clusters of β-gal+ nuclei (arrow) in the TA of P14 SCID/bg mice, two weeks after transplantation of embryonic aortas from E9 MLC3F-nLacZ embryos. B, Whole-mount stain of the contralateral TA of the same SCID/bg reveals several β-gal+ nuclei (arrow) dispersed throughout the whole muscle. Bar, 50 μm.

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