Fibroblasts are the Critical Cells in Salamander Limb Regeneration

Written by David M Gardiner

I – Fibroblasts form the early blastema, and control the growth and pattern of the regenerated limb
1.Lineage analysis shows that more than half of the early blastema cells are progeny of the dermal fibroblasts. Most of the other blastema cells likely come from the connective tissue fibroblasts of the non-dermal stump tissues. In cell migration studies, dermal fibroblasts are seen to migrate toward the center of the wound and then give rise to the early blastema.
2.The proportion of blastema cells derived from dermal fibroblasts is much higher than the proportion of dermal fibroblasts in the stump. This means that dermal fibroblasts proliferate during early regeneration to give rise to blastema cells. Other cells from the mature limb (bone/cartilage and muscle) under-contribute to the blastema. This finding is contrary to the classical and erroneous assumption that all tissues in the stump contribute equally and in proportion to their availability.
3.You can only change the structure of the regenerated limb by changing the way in which fibroblasts interact, and you can induce a range of responses from complete inhibition of regeneration to the formation of supernumerary limb structures. Dermal fibroblasts have an overriding influence compared to fibroblasts from the interior of the limb stump. Non-fibroblast containing tissues do not influence the outcome of regeneration.
4.Grafts of only skin (epidermis plus dermis) can rescue the regeneration of limbs in which regeneration has been blocked by xirradiation. The only source of cells are dermal fibroblasts (the epidermis does not contribute to the blastema), and the skeletal, vascular, nervous and connective tissue patterns are normal. These complete limbs lack muscle, which means that you do not need muscle to get limb regeneration.
5.Dermal fibroblasts induce the formation of ectopic limbs de novo from wounds that have been stimulated by a nerve graft. Only dermal fibroblasts are needed to induce formation of a new limb. Conversely, you only get a new limb when you provide appropriate dermal fibroblasts.

II – Behavior of the other cells types is regulated by the limb blueprint created by fibroblasts
1.Other cell/tissue types (blood vessels, peripheral nerves, and muscle) can generate in mammals as well as in salamanders, and can participate in regeneration if there is a regeneration-permissive environment. Thus they are not the cause of regenerative failure in mammals. In contrast, mammalian fibroblasts form scars rather than a blueprint for regeneration. Fibroblasts do not form scars in salamander limb wounds.
2.The most extensively studied non-fibroblast cells have been muscle cells. Older, non-genetic studies suggested that these cells can transdifferentiate during regeneration to form other tissue types. Recent genetic studies have demonstrated that regenerated muscle in amphibians is derived only from adult muscle, and that adult muscle only gives rise to muscle during regeneration. This finding is the same as in all other vertebrates (including mammals) in which there are adult myogenic stem cells (satellite cells) that are the same as salamander myogenic stem cells (post-satellite cells, the source of the newt myogenic A1 cell line).
3.The blueprint function of fibroblasts is demonstrated most directly by the induction of ectopic limbs. The initial wound involves only the skin (epidermis and dermis), but the final limb is composed of all the normal limb tissues. Thus dermal fibroblasts are able to provide the signals that recruit nerves, blood vessels and muscle from tissues and adult stem cells in the stump.
4.As in the case of ectopic limbs, when the blastema forms on an amputated limb stump it provides a permissive environment that stimulates the regeneration of nerves, blood vessels and muscle either through migration of progenitor cells (e.g. myoblasts that give rise to the regenerated muscle) though regrowth from the severed structure in the stump (e.g. preexisting blood vessels and nerves).