Induction of limb regeneration in lizards in relation to the expression of the signaling proteins FGF8 and Shh

After hind limb amputation in lizards, scars or short outgrowths are formed. FGFs may stimulate regeneration. Injections of FGF-1/-2 into the limb stump were made until 26 days post-amputation. This treatment gives rise to outgrowths of 1.5-3.5 mm within 40-70 days post-amputation, containing axial cartilages continuous with stump bones but missing muscles. Immunohistochemical localization in the outgrowth shows 5BrdU-labeled proliferating cells in the apical wound epidermis and perichondrium at 40-50 days post-amputation. Low to no proliferation is present in the axial cartilages and connective tissues at 60 and 70 days when growth terminates, except in scales. Immunodetection of FGF8 and Shh shows that the proteins are present at 30-40 days in the wound epidermis, apical connective tissue and perichondrium. These proteins lower to disappear in outgrowths from 50 to 70 days post-amputation, coincident with lowering of cell proliferation. The study suggests that the injection of FGF1/2 initially stimulates the growth of the limb mainly in cartilaginous axial structures, dense connective tissue, and new scales. No stimulation of muscle tissues is observed. Short cartilaginous rods of tibia, fibula or femur are produced but no autopodial elements. The presence of FGF8 and Shh in the wound epidermis and perichondrium of the initial outgrowths and their subsequent downfall reflect the initial growth and the later growth cessation. In future experiments, longer treatments with FGFs, hyaluronate and other signaling proteins, micro-injected in specific regions of the outgrowths may further enhance limb growth in this or other amniote models of limb regeneration.