REVERSAL OF MUSCLE DIFFERENTIATION DURING URODELE LIMB REGENERATION

Recent studies suggest that maintenance of the differentiated state requires continuous regulation. Limb regeneration in urodele amphibians provides a context in which to address this issue, as limb regeneration may involve the dedifferentiation of multinucleate myotubes to yield mononucleate blastemal cells, which then proliferate and contribute to regenerate tissues. To evaluate this possibility, cultured newt limb myotubes were selectively microinjected with the lineage tracer rhodamine-dextran and introduced into regenerating limbs. In culture, such labeled myotubes were stable after 6-8 weeks, and transfer of the tracer to mononucleate cells was not observed. In contrast, after implantation of labeled myotubes under the wound epidermis of limb blastemas, strongly labeled mononucleate cells were observed after 1 week. These cells could be double-labeled with the cytoplasmic lineage tracer and [H-3]thymidine that had been incorporated into the nuclei of implanted myotubes. The number of labeled mononucleate cells increased significantly by 2-3 weeks after implantation, indicating that these cells proliferated. Although the fate of these cells at later times was uncertain, we provide evidence consistent with their subsequent differentiation. These results demonstrate reversal in the mononucleate-to-multinucleate transition of vertebrate myogenesis.