Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
[Preprint]. 2024 Jan 25:2024.01.24.577041.
doi: 10.1101/2024.01.24.577041.

Wnt7a is Required for Regeneration of Dystrophic Skeletal Muscle

Uxia Gurriaran-Rodriguez  1   2 , Kasun Kodippili  1   2 , David Datzkiw  1   2 , Ehsan Javandoost  1   2 , Fan Xiao  1   2 , Maria Teresa Rejas  3 , Michael A Rudnicki  1   2
Affiliations

Wnt7a is Required for Regeneration of Dystrophic Skeletal Muscle

Uxia Gurriaran-Rodriguez et al. bioRxiv. .

Abstract

Intramuscular injection of Wnt7a has been shown to accelerate and augment skeletal muscle regeneration and to ameliorate dystrophic progression in mdx muscle, a model for Duchenne muscular dystrophy (DMD). However, loss-of-function studies to investigate the requirement for Wnt7a in muscle regeneration has not been evaluated. Here, we assessed muscle regeneration and function in wild type (WT) and mdx mice where Wnt7a was specifically deleted in muscle using a conditional Wnt7a floxed allele and a Myf5-Cre driver. We found that both WT and mdx mice with deletion of Wnt7a in muscle, exhibited marked deficiencies in muscle regeneration at 21 d following cardiotoxin (CTX) induced injury. Unlike WT, deletion of Wnt7a in mdx resulted in a marked decrease in specific force generation prior to CTX injury. However, both WT and mdx muscle lacking Wnt7a displayed decreased specific force generation following CTX injection. Notably the regeneration deficit observed in mdx mice lacking Wnt7a in muscle was rescued by a single tail vein injection of an extracellular vesicle preparation containing Wnt7a (Wnt7a-EVs). Therefore, we conclude that the regenerative capacity of muscle in mdx mice is due to the upregulation of endogenous Wnt7a following injury, and that systemic delivery of Wnt7a-EVs represents a therapeutic strategy for treating DMD.

Keywords: Duchenne Muscular Dystrophy; Extracellular Vesicles; Regeneration; Skeletal muscle; Wnt7a.

PubMed Disclaimer

Figures

Figure 1:
Figure 1:. Lack of Wnt7a expression impairs muscle regeneration.
(A-left) H&E stained sections of injured and non-injured TA muscle of mdx mice at 3 dpi. (A-right) Wnt7a immunostained (green) sections of injured and non-injured TA upon 3 dpi. Laminin (red) delineates myofibers. Nuclei are stained with DAPI. (B) Mouse strains used and experimental design. (C and D) Quantification of Pax7-expressing cells in regenerating WT muscle. (E and F) Quantification of Pax7-expressing cells in regenerating mdx TA muscle. (G and H) Myofiber caliber distribution at 21 d following regeneration in WT and mdx TA muscle with and without Wnt7a deletion. (I and J) H&E stained sections at 21 d following regeneration in WT and mdx TA muscle with and without Wnt7a deletion. TA (Tibialis Anterior), dpi (days post injury). n≥4 mice, mean ± s.e.m., p values determined by two-sided Student’s t-test (*p<0.05, n.s.=not significant).
Figure 2:
Figure 2:. Reduced force generation in regenerated TA muscles lacking Wnt7a.
(A) Schematic of mouse strains used and experimental design. (B) Specific force frequency curve in uninjured WT TA muscle with (dark blue) and without (light blue) Wnt7a expression. (C) Specific force frequency curve in uninjured mdx TA muscle with (dark green) and without (light green) Wnt7a expression. (D) Specific force frequency curve of regenerated WT TA muscle, with (dark blue) and without (light blue) Wnt7a expression, at 21 d following injury. (E) Specific force frequency curve of regenerated mdx TA muscle, with (dark green) and without (light green) Wnt7a expression, at 21 d following injury. TA (Tibialis Anterior). n≥3 mice, mean ± s.e.m., p values determined by two-way ANOVA test (*p<0.05, **p<0.01, n.s.=not significant).
Figure 3:
Figure 3:. Wnt7a-EVs exhibit enhanced bioactivity.
(A) Schematic showing the experimental protocol for ex vivo EV treatment of single myofibers isolated from FDB muscles, imaging and analysis using high-content screening. (B) Representative immunofluorescence images of symmetric (top panel) and asymmetric (bottom panel) satellite stem cell divisions. (C) Rate of muscle stem cell division after 42 h in culture is not altered by treatment with either Wnt7a-EVs or with recombinant Wnt7a. (D) Symmetric satellite stem cell divisions are significantly increased by Wnt7a-EV treatment. EVs (extracellular vesicles), FDB (Flexor Digitorum fibers). n≥5 replicates, mean ± s.e.m., p values determined by two-sided Student’s test (*p<0.05, **p<0.01, ***p<0.001, n.s.=not significant).
Figure 4:
Figure 4:. Systemic delivery of Wnt7a-EVs rescues muscle regeneration in mdx mice.
(A) Schematic of experimental design. (B) Specific Force frequency curve of uninjured mdx TA muscle lacking Wnt7a, comparing untreated (light green), Wnt7a-EV (red) injected, or control-EV (dark green) injected. (C) Specific Force frequency curve of regenerated mdx TA muscle lacking Wnt7a, comparing untreated (light green), Wnt7a-EV (red) injected, or control-EV (dark green) injected. (D) Myofiber caliber distribution of regenerated mdx TA muscle lacking Wnt7a, comparing Wnt7a-EV (red) injected, or control-EV (dark green) injected. (E) Wnt7a-EV treatment results in increased numbers of myofibers. (F) H&E stained sections of regenerated TA muscles after 21 dpi (G) Representative H&E stained sections of diaphragms from mdx mice lacking Wnt7a intravenously injected with Wnt7a-EVs or control-EVs (H) Myofiber caliber distribution of diaphragms from mdx mice lacking Wnt7a injected with Wnt7a-EVs or control-EVs (I) Minimal fiber feret average in diaphragms of injected mice. TA (Tibialis Anterior), dpi (days post injury. n≥4 mice, mean ± s.e.m., p values determined by two-way ANOVA or two-sided Student’s test (*p<0.05, **p<0.01, ****p<0.0001, n.s.=not significant).
See this image and copyright information in PMC

Similar articles

See all similar articles

References

    1. Bentzinger C. F., et al., Wnt7a stimulates myogenic stem cell motility and engraftment resulting in improved muscle strength. Journal of Cell Biology 205, 97–111 (2014). - PMC - PubMed
    1. Le Grand F., Jones A. E., Seale V., Scimè A., Rudnicki M. A., Wnt7a Activates the Planar Cell Polarity Pathway to Drive the Symmetric Expansion of Satellite Stem Cells. Cell Stem Cell 4, 535–547 (2009). - PMC - PubMed
    1. von Maltzahn J., Bentzinger C. F., Rudnicki M. A., Wnt7a-Fzd7 signalling directly activates the Akt/mTOR anabolic growth pathway in skeletal muscle. Nat Cell Biol 14, 186–91 (2012). - PMC - PubMed
    1. Dumont N. A., et al., Dystrophin expression in muscle stem cells regulates their polarity and asymmetric division. Nat Med 21, 1455–1463 (2015). - PMC - PubMed
    1. von Maltzahn J., Renaud J.-M. M., Parise G., Rudnicki M. A., Wnt7a treatment ameliorates muscular dystrophy. Proc Natl Acad Sci U S A 109, 20614–20619 (2012). - PMC - PubMed

Publication types

-