Distinct pattern of Connexin gene expression during skeletal muscle regeneration in the adult rat.

Abstract

Aim: The aim of present work was to test the hypothesis that Cx37, Cx39, Cx40, Cx43 and Cx45 expression could be regulated in adult regenerating skeletal muscle in response to injury promoting activation of satellite cells involved in myofibers repair and regeneration. Methods: Using in situ hybridization and immunohistochemistry procedures we examined the spatial and temporal expression pattern of above listed connexins in the regenerating gastrocnemious muscle following a mechanical injury. Results: Cx43 and Cx45 mRNA were up-regulated very early, by 3 hour following muscle injury, and were localised in satellite cells, M-cadherin positive cells, distributed around the area of lesion. Three days after lesion a large number of Cx43 and Cx45 mRNA labelled cells were found inside the area of lesion where they become myoblasts, myogenin positive, and participate to muscle regeneration. By contrast, Cx39, not expressed in control muscle, appears in myogenin positive cells 3 days following muscle lesion and its expression was restricted to the area of lesion. Cx39 mRNA was mainly localised in myoblast myogenin positive cells forming clusters or rows of closely apposed cell nuclei committed to form myotubes. Cx40 mRNA labelled cells were observed within 24 hours from injury in the endothelial cells of blood vessels around the area of lesion. After 3 days these cells were localised inside the area of lesion where they co-express myogenin and start to form myotubes. By contrast, Cx37 mRNA expression appeared by 24 hrs from lesion and never was colocalised in myogenin positive cells, but it was involved in new blood vessels formation. All transcripts examined reached a peak of expression between 7-9 days from injury and then progressively declined, being undetectable at 4-5 weeks from injury. Conclusion: Taken together these results support the suggestion that, in regenerating skeletal muscle several connexins may be differentially involved in communication of myogenic committed cells during the process of cell proliferation, aggregation and fusion to form new myotubes.