Expression of FGF2 in the limb blastema of two Salamandridae correlates with their regenerative capability.

Limb regenerative potential in urodeles seems to vary among different species. We observed that Triturus vulgaris meridionalis regenerate their limbs significantly faster than T. carnifex, where a long gap between the time of amputation and blastema formation occurs, and tried to identify cellular and molecular events that may underlie these differences in regenerative capability. Whereas wound healing is comparable in the two species, formation of an apical epidermal cap (AEC), which is required for blastema outgrowth, is delayed for approximately three weeks in T. carnifex. Furthermore, fewer nerve fibres are present distally early after amputation, consistent with the late onset of blastemal cell proliferation observed in T. carnifex. We investigated whether different expression of putative blastema mitogens, such as FGF1 and FGF2, in these species may underlie differences in the progression of regeneration. We found that whereas FGF1 is detected in the epidermis throughout the regenerative process, FGF2 onset of expression in the wound epidermis of both species coincides with AEC formation and initiation of blastemal cell proliferation, which is delayed in T. carnifex, and declines thereafter. In vitro studies showed that FGF2 activates MCM3, a factor essential for DNA replication licensing activity, and can be produced by blastemal cells themselves, indicating an autocrine action. These results suggest that FGF2 plays a key role in the initiation of blastema growth

Effects of locomotor training on hindlimb regeneration in the urodele amphibian Pleurodeles waltlii

The effects of locomotor training on hindlimb regeneration were studied in the urodele amphibian Pleurodeles waltlii.After amputation of one hindlimb at mid-femur, adult animals were subjected to regular training sessions (1 h daily, 5 days a week, over 8 months) of terrestrial stepping.Eight months post-amputation, trained animals exhibited regenerated limbs of reduced size as compared to animals kept in their aquaria. Histological data showed an abnormal regeneration of both the femur and distal structures (e.g. digit muscles, metatarsi and phalanges) while medial structures (e.g. tibia and fibula) were totally re-formed. The study of the electromyographical activity in regenerated limbs during stepping and that of their reflex responsiveness to electrical stimulation showed that both motor and sensory innervations were functional in the limb stump of trained animals.The regenerative capacity of the abnormal stumps was preserved since following a second amputation a quite normal hindlimb was regenerated in 3 months, provided the re-amputated animals were not trained to terrestrial stepping.The stress due to handling, change in locomotor medium (aquatic vs. terrestrial) and the friction of the wound epidermis with the ground were not involved in the disruption of limb regeneration.The locomotor pattern, the reflex responsiveness and the muscle fibre composition were similar in supernumerary forelimbs grafted on the back and in normal forelimbs. However, the supernumerary forelimbs regenerated normally even in animals subjected to locomotor training while the hindlimb did not. It is concluded that the disrupting effects of locomotor training on limb regeneration were localized to the the limb directly involved in locomotion.The mechanisms underlying abnormal limb regeneration in animals subjected to locomotor training are discussed

Tissue engineering of replacement skin: the crossroads of biomaterials, wound healing, embryonic development, stem cells and regeneration

Advanced therapies combating acute and chronic skin wounds are likely to be brought about using our knowledge of regenerative medicine coupled with appropriately tissue-engineered skin substitutes. At the present time, there are no models of an artificial skin that completely replicate normal uninjured skin. Natural biopolymers such as collagen and fibronectin have been investigated as potential sources of biomaterial to which cells can attach. The first generation of degradable polymers used in tissue engineering were adapted from other surgical uses and have drawbacks in terms of mechanical and degradation properties. This has led to the development of synthetic degradable gels primarily as a way to deliver cells and/or molecules in situ, the so-called smart matrix technology. Tissue or organ repair is usually accompanied by fibrotic reactions that result in the production of a scar. Certain mammalian tissues, however, have a capacity for complete regeneration without scarring; good examples include embryonic or foetal skin and the ear of the MRL/MpJ mouse. Investigations of these model systems reveal that in order to achieve such complete regeneration, the inflammatory response is altered such that the extent of fibrosis and scarring is diminished. From studies on the limited examples of mammalian regeneration, it may also be possible to exploit such models to further clarify the regenerative process. The challenge is to identify the factors and cytokines expressed during regeneration and incorporate them to create a smart matrix for use in a skin equivalent. Recent advances in the use of DNA microarray and proteomic technology are likely to aid the identification of such molecules. This, coupled with recent advances in non-viral gene delivery and stem cell technologies, may also contribute to novel approaches that would generate a skin replacement whose materials technology was based not only upon intelligent design, but also upon the molecules involved in the process of regeneration