Lizard tail skeletal regeneration combines aspects of fracture healing and blastema-based regeneration

Lizards are amniotes with the remarkable ability to regenerate amputated tails. The early regenerated lizard tail forms a blastema, and the regenerated skeleton consists of a cartilage tube (CT) surrounding the regenerated spinal cord. The proximal, but not distal, CT undergoes hypertrophy and ossifies. We hypothesized that differences in cell sources and signaling account for divergent cartilage development between proximal and distal CT regions. Exogenous spinal cord implants induced ectopic CT formation in lizard (Anolis carolinensis) blastemas. Regenerated spinal cords expressed Shh, and cyclopamine inhibited CT induction. Blastemas containing vertebrae with intact spinal cords formed CTs with proximal hypertrophic regions and distal non-hypertrophic regions, whereas removal of spinal cords resulted in formation of proximal CT areas only. In fate-mapping studies, FITC-labeled vertebra periosteal cells were detected in proximal, but not distal, CT areas. Conversely, FITC-labeled blastema cells were restricted to distal CT regions. Proximal cartilage formation was inhibited by removal of periosteum and could be recapitulated in vitro by periosteal cells treated with Ihh and BMP-2. These findings suggest that proximal CTs are directly derived from vertebra periosteal cells in response to BMP and Ihh signaling, whereas distal CTs form from blastema cells in response to Shh signals from regenerated spinal cords.