Cartilage and bone cells do not participate in skeletal regeneration in Ambystoma mexicanum limbs

The Mexican Axolotl is one of the few tetrapod species that is capable of regenerating complete skeletal elements in injured adult limbs. Whether the skeleton (bone and cartilage) plays a role in the patterning and contribution to the skeletal regenerate is currently unresolved. We tested the induction of pattern formation, the effect on cell proliferation, and contributions of skeletal tissues (cartilage, bone, and periosteum) to the regenerating axolotl limb. We found that bone tissue grafts from transgenic donors expressing GFP fail to induce pattern formation and do not contribute to the newly regenerated skeleton. Periosteum tissue grafts, on the other hand, have both of these activities. These observations reveal that skeletal tissue does not contribute to the regeneration of skeletal elements; rather, these structures are patterned by and derived from cells of non-skeletal connective tissue origin. [Display omitted] •Periosteum induces the formation of ectopic limb structures.•Skeletal grafts have minimal pattern inducing activity on regenerating limbs.•Heparin-sulfate chains are required for inductive activity of periosteum.•Periosteum contributes to the regenerated skeletal elements.•Skeletal tissue does not contribute to the regenerate.