spacer gif spacer gif spacer gif spacer gif spacer gif
QUICK SEARCH:   [advanced]


spacer gif
     Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents    

First published online 6 June 2006
doi: 10.1242/jcs.03000

Journal of Cell Science 119, 2679-2687 (2006)
Published by The Company of Biologists 2006
This Article
Right arrow Summary Freely available
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Yokota, T.
Right arrow Articles by Partridge, T. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Yokota, T.
Right arrow Articles by Partridge, T. A.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati   Add to Twitter  
What's this?

Expansion of revertant fibers in dystrophic mdx muscles reflects activity of muscle precursor cells and serves as an index of muscle regeneration

Toshifumi Yokota1,2, Qi-Long Lu3,*, Jennifer E. Morgan4, Kay E. Davies5, Rosie Fisher5, Shin'ichi Takeda6 and Terence A. Partridge1

1 Muscle Cell Biology Group, Medical Research Council Clinical Science Centre, Hammersmith Hospital Campus, Imperial College School of Medicine, London University, Du Cane Road, London, W12 0NN, UK
2 Center for Genetic Medicine Research, Children's Research Institute, Children's National Medical Center, 111 Michigan Ave, NW, Washington, DC 20010, USA
3 McColl Lockwood Laboratory for Muscular Dystrophy Research, Neuromuscular/ALS Center, Carolinas Medical Center, 1000 Blythe Blvd, Charlotte, NC 28231, USA
4 Department of Paediatrics, Imperial College London, The Dubowitz Neuromuscular Centre, Hammersmith Hospital, Du Cane Road, London, W12 ONN, UK
5 Department of Human Anatomy and Genetics, University of Oxford, South Parks Road, Oxford, OX1 3QX, UK
6 Department of Molecular Therapy, National Institute of Neuroscience, National Center of Neurology and Psychiatry (NCNP), Ogawa-Higashi 4-1-1, Kodaira, Tokyo 187-8502, Japan


Figure 1
View larger version (25K):

[in a new window]
  		Fig. 1. Micro-dystrophin transgene structures in CS1, AX11 and M3 transgenics. The schematic view of micro-dystrophin transgenes shows that CS1 (4.9 kb), AX11 (4.4 kb) and M3 (3.9 kb) encode 4, 3 and 1 rod domains, and 3, 2 and 2 hinge domains, respectively. The size of the normal muscle dystrophin transcript is 14 kb and contains 24 rod domains and 4 hinges.

 

Figure 2
View larger version (94K):

[in a new window]
  		Fig. 2. Immunohistochemistry analysis of dystrophin expression in muscles of micro-dystrophin transgenics. Immunohistochemistry of RFs in muscles of M3, AX11 and CS1 micro-dystrophin-transgenic mice, and the mdx mouse, with P7 antibody and MANDRA1 antibody at 5 weeks (5W) and 10 weeks (10W) of age. The P7 antibody recognizes RFs only, whereas MANDRA1 recognizes revertants and micro-dystrophin (top). Bar, 100 µm.

 

Figure 3
View larger version (19K):

[in a new window]
  		Fig. 3. The number of RFs and clusters are decreased in micro-dystrophin-transgenic mice. (A) The maximal number of RFs. (B) The maximal number of revertant clusters containing more than one RF. (C) The maximal number of RFs in one cluster. Values are mean ± s.e.m. for 4–10 TA muscles per group. *Significant difference (P<0.05) from mdx muscles of the same age. 5W, 5 weeks; 10W, 10 weeks.

 

Figure 4
View larger version (22K):

[in a new window]
  		Fig. 4. Muscle degeneration and regeneration is required for RF expansion. (A) The percentage of central nucleated fibers in each strain of mice. The number of regenerating fibers decreases from mdx, to M3, to AX11 to CS1 mice aged 5 weeks and 10 weeks. (B) Correlation between the number of RFs and the percentage of centrally nucleated fibers in each strain at 10 weeks of age. (C) Correlation between the number of clusters and the percentage of central nucleated fibers in each strain at 10 weeks of age. The direct correlation between the number of RFs or clusters and centrally nucleated fibers suggests that the muscle degeneration and regeneration is an essential step in the expansion of RFs. Values are mean ± s.e.m. for 4-10 muscles per group. *Significant difference (P<0.05) from mdx muscles of the same age.

 

Figure 5
View larger version (47K):

[in a new window]
  		Fig. 5. Immunohistochemistry analysis of dystrophin expression in muscles of utrophin-transgenic mdx mice. (A) Immunohistochemistry with anti-utrophin and anti-dystrophin antibodies in mdx mice and transgenic mdx mice expressing full-length utrophin (utro/mdx) at 1 year of age (note that a proportion of fibers are centrally nucleated in the muscle of mice of the same age that are transgenic for full-length utrophin). Only a single RF is seen in the transgenic mouse compared with a large cluster in the control mdx mouse. (B) The number of RFs in mdx mice (black columns), mdx mice expressing mini-utrophin (gray columns) and mdx mice expressing full-length utrophin (white columns), at 1 month, 6 months and 1 year of age. Each point is the mean ± s.e.m. of 3-8 muscles per group. *Significant difference (P<0.05) from mdx muscles of the same age. Bar, 100 µm.

 

Figure 6
View larger version (30K):

[in a new window]
  		Fig. 6. Reduced expression of revertant dystrophin in irradiated mdx muscles. (A) Mice were irradiated at the age of 3 weeks with 4 Gy, 10 Gy, or 18 Gy, and sacrificed at 3 months after the irradiation. The irradiated muscles show reduced number of RFs compared with untreated control mdx muscle (A), suggesting that muscle regeneration is crucial for RF expansion. (B) The number of RF clusters, total number of RFs, and maximal number of RFs in one cluster in mdx TA muscle at 4 weeks after irradiation at 3 weeks of age. The mice were irradiated with 4 Gy (gray bars), 10 Gy (dashed bars) or 18 Gy (white bars), or were non-irradiated (black bars). (C) Comparison of the level of suppression of RF expansion at 1 month or 1 year after 18 Gy irradiation at the age of 3 weeks. Black bars, non-irradiated muscles; white bars, irradiated muscles. Each point is the mean ± s.e.m. of 3-8 muscles per group. *Significant difference (P<0.05) from mdx muscles of the same age. Bar, 100 µm.

 

Figure 7
View larger version (36K):

[in a new window]
  		Fig. 7. Notexin injection after irradiation restores RF expansion. (A) Experimental design of irradiation and notexin treatments in TA muscles of mdx mice. (1) Irradiation only; (2) notexin treatment only; and (3) notexin treatment 4 weeks after 18 Gy irradiation at 3 weeks of age. IHC, immunohistochemistry. (B) The irradiated TA muscle shows a single RF, in contrast to the untreated control TA muscle and notexin-injected muscles without irradiation (notexin only). A regenerating RF cluster reappears after notexin injection in pre-irradiated muscles (18 Gy + notexin). Bar, 100 µm. (C) The total number of RFs and the maximal number of RFs per cluster in TA muscle after notexin treatment with or without pre-irradiation. *Significant difference (P<0.05) was obtained between control mice treated with 18 Gy irradiation only and the mice treated with notexin after 18 Gy pre-irradiation. Each point is the mean ± s.e.m. of 3-4 muscles per group.

 

Figure 8
View larger version (57K):

[in a new window]
  		Fig. 8. nNOS expression is only found in RFs in micro-dystrophin-transgenic muscles and mdx mice. The expression of nNOS and dystrophin RFs in M3 and AX11 transgenic mice, and in mdx mice, aged 5 weeks. nNOS is expressed only in RFs in these transgenics, suggesting that the dystrophin rod domain has an important role in targeting nNOS at the sarcolemma. Bar, 50 µm.

 

Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati   Add to Twitter Twitter    What's this?



© The Company of Biologists Ltd 2006