Salamanders display amazing regenerative capacities that include the ability to regrow new limbs. Previous results have indicated that blastema cells are originated mainly by the dedifferentiation of pre-existing tissues into cells that re-enter the cell cycle and proliferate to form the missing structures. However, and because salamanders possess muscle satellite cells positive for Pax7 it was not clear the relative contribution of mature myofibers and those satellite cells into the regenerated limb skeletal muscle. Now, a recent paper from the laboratories of András Simon and Elly Tanaka (http://www.ncbi.nlm.nih.gov/pubmed/24268695) provides more clear evidence of the cellular origin of the regenerated muscle fibers in two different salamanders: Notophtalmus viridescens (newt) and Ambystoma mexicanun (axolotl).
Remarkably, by using a Cre–loxP-based genetic fate mapping of muscle fibers during regeneration they found fundamental differences in the cellular origin of the regenerated muscle in these two species of salamanders. Two weeks after co-electroporating the upper arm of newts with the different constructs needed to specifically label the mature muscle fibers (positive for myosin heavy chain (MHC)) with YFP (without labelling the Pax7+ satellite cells), those limbs were amputated. The authors found YFP+ cells all along the regenerated limb except the digit tips, indicating a direct contribution of the pre-existing muscle into the regenerated myofibers. Detailed analyses of those blastemas showed that in their distal regions they contained YFP+ cells that were negative for MHC pointing out to a dedifferentiation of the muscle cells to form mononuclear blastema cells. Moreover those blastema cells had re-entered the cell cycle as indicated by labelling with PCNA and the incorporation of the nucleotide analog Edu. Importantly, Edu was never incorporated in YFP+/MHC+ or MHC+ cells suggesting that cell cycle re-entry occurred after the fragmentation and dedifferentiation of muscle fibers into blastema cells.
In contrast, when they used the same strategy to label the mature muscle fibers of the axolotl and trace them upon amputation, no YFP+ cells were found within the blastema or in the regenerated limb. Although some morphological changes of the pre-existing fibers were seen at the amputation plane, these results suggested that the pre-existing muscle fibers did not contribute to muscle regeneration in axolotls. Previous results from the laboratory of Elly Tanaka had shown that GFP+ muscle fibers and satellite cells contribute to muscle regeneration in axolotls, but without being able to distinguish their relative contributions. Here, they found that of 834 GFP+ positive blastema cells, 809 expressed Pax7. Also, these GFP+ were cycling as they incorporated EdU. Taking in account these two results (lack of YFP+ cells and presence of GFP+/Pax7+ cells within the blastema) the authors conclude that were the Pax7+ satellite cells from the mature limb the ones that gave rise to the proliferative muscle progenitors required for regeneration.
Finally, they analysed whether those differences between newts and axolotls could be explained by the neotenic nature of axolotls. They metamorphosed axolotls and analysed limb regeneration in them, following the same methodological approach. Similarly, muscle regeneration in this context did not seem to depend on the dedifferentiation of pre-existing muscle fibers. Also, and complementarily, Pax7+ cells were not found either in larval newt limb blastemas (in agreement with the results obtained in adult newt blastemas).
In summary, this study characterizes at the cellular level the origin of the regenerated muscle fibers in two different species of salamanders. Remarkably, these two species use very different strategies to achieve the same final goal: skeletal muscle regeneration. These results are a beautiful and clear example of how different animals (even relatively closed phylogenetically) use different strategies to form the same cell type within the regenerative blastema. This flexibility and diversity of successful strategies may be of special relevance for the field of regenerative medicine.