Salamander Limb Regeneration
Salamanders and many other related amphibians have a remarkable aptitude for the regeneration of various body structures when compared to other vertebrates. These structures include the jaws, spinal cord, heart ventricles, some eye structures, and most notably their limbs (1). Limb regeneration itself is not a unique feature of salamanders as all other organisms possess some degree of regenerative capability. What sets salamanders apart from the rest is that fact that they can fully regenerate amputated limbs at any time during their life cycle (2).
Limb regeneration in salamanders occurs in several overlapping steps. A few hours to a day following limb amputation, epidermal cells around the wound area are recruited in order to cover the wound (3). This process requires a sufficient amount of nerves to be present in the wound area (Why??). If this condition is met, it results in the formation of a structure called the wound epidermis. The purpose of the wound epidermis is to prevent the entry of debris into the wound site (4).
As the wound epidermis develops, it eventually forms another structure called the apical epithelial cap (AEC) which resembles a bud on the surface of the site of amputation. The AEC is essential to regeneration as it secretes various growth factors which aid in limb outgrowth (2). These factors include fibroblast growth factors (FGFs) which are found in many organisms including humans, and are typically involved in tissue repair in adult organisms (3,5).
Tissues below the wound epidermis such as local cartilage, muscle, and Schwann cells then undergo histolysis which leads to cell dedifferentiation (3). Cell dedifferentiation is a process in which cells that have already changed into a specialized cell type are reverted back to an undifferentiated form, the opposite of differentiation. In the case of salamander regeneration, they are reverted into mesenchymal stem cells which can differentiate into a number of cell types including bone, fat, and cartilage cells (6). Cell dedifferentiation results in a population of mesenchymal stem cells which migrate to the wound surface and form a cone-shaped mass of cells known as the regeneration blastema (2).
The process of dedifferentiation is not yet well understood due to its complexity and the fact that it does not follow the same process for each tissue type. However, common factors seen in all dedifferentiation events includes the down-regulation of differentiation-promoting genes and the upregulation of embryonic and regeneration-specific genes (7). In addition, histolysis of these cells prior to dedifferentiation is triggered by matrix metalloproteinases, (MMPs) (2). MMPs are enzymes that have the ability to degrade proteins (e.g. collagen) of the extracellular matrix which provides structural support to surrounding cells (8). MPPs are also involved in the prevention of scar formation, and contribute to the overall maintenance and growth of the blastema (1).
The blastema grows distally over time via the proliferation of mesenchymal stem cells until the limb has fully regenerated. This is mediated with the help of factors secreted by the AEC (2). During this process, cells of the blastema produce neurotrophic factors involved in the regeneration of sensory and motor nerves (3). Simultaneous to the development of the blastema, its cells begin to re-differentiate into tissue cells specific to the regenerating limb and limb structural repatterning proceeds. The original limb cells at the site of injury are thought to possess positional memory of their placement along the 3 axes of the limb which are inherited by cells of the blastema as they re-differentiate, allowing them to migrate to the appropriate position in the growing limb. These 3 axes include the proximal-distal, anterior-posterior, and dorsal ventral axes. The re-expression of various developmental genes including HOX genes helps to direct the regeneration process of the limb structure, ensuring proper differentiation of blastemal cells until it is completely rebuilt (9).
With the knowledge of this ability, one might ask if human limb regeneration may be feasible in the future. Many simple organisms such as Hydra have a high regenerative ability. This suggests that a higher regenerative capacity was an ancestral trait which was eventually lost in mammals (11). When a human limb is amputated, the end result is scar formation rather than the initiation of limb regeneration (12). Mammals including humans have poor limb regenerative capacity as adults, but during the early stages of life such as the embryonic or fetal stages, they have a limited ability to regenerate the digit tips (2). In addition, human tissue in the early stages of life responds to injury with regeneration rather than scarring (1). While it is currently not possible to induce limb regeneration in humans, by studying the genetic and molecular mechanisms at work during injury repair in early mammalian life and in other organisms which can fully regenerate limbs, we can make strides towards tapping into this suppressed ability in the future.
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