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ABSTRACT: Urodele amphibians, such as the axolotl (Ambystoma mexicanum), have the unique faculty among vertebrates to regenerate lost appendages (limbs and tail) and other body parts (apex of the heart, forebrain and jaw) after amputation. Interestingly, axolotls never seem to form scar tissue at the site of amputation once regeneration is completed. Before now, very few studies were directly focused on the description of the events happening during wound healing after a skin injury in salamanders. In this paper, we directly investigated skin wound healing after excisional wounding which removed the epidermis, dermis and basement membrane in the axolotl. Axolotls were wounded with a 1.5-mm skin biopsy punch. Results show rapid re-epithelialization of the wound within 8 hrs after wounding. Histological analysis of wound healing confirmed the absence of tissue fibrosis throughout the process and shows that skin integrity is re-established by 90 days after wounding. Results also reveal the absence of neutrophils in the wound area, suggestive of a lack of or low inflammatory response. The expression of proteins central to wound healing seemed different than in mammals as α-smooth muscle actin was absent and transforming growth factor β-1 was only transiently expressed during wound healing in the axolotl. Finally, subcutaneous injections of bleomycin were performed to verify whether the induction of scar tissue was possible in axolotls. Surprisingly, results show that axolotls are not resistant to bleomycin-induced tissue fibrosis, but the resulting scar tissue does not seem to contain significant amounts of collagen.
Journal of Experimental Zoology Part B Molecular and Developmental Evolution 12/2010; 314(8):684-97. · 2.42 Impact Factor
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ABSTRACT: Axolotls (urodele amphibians) have the unique ability, among vertebrates, to perfectly regenerate many parts of their body including limbs, tail, jaw and spinal cord following injury or amputation. The axolotl limb is the most widely used structure as an experimental model to study tissue regeneration. The process is well characterized, requiring multiple cellular and molecular mechanisms. The preparation phase represents the first part of the regeneration process which includes wound healing, cellular migration, dedifferentiation and proliferation. The redevelopment phase represents the second part when dedifferentiated cells stop proliferating and redifferentiate to give rise to all missing structures. In the axolotl, when a limb is amputated, the missing or wounded part is regenerated perfectly without scar formation between the stump and the regenerated structure. Multiple authors have recently highlighted the similarities between the early phases of mammalian wound healing and urodele limb regeneration. In mammals, one very important family of growth factors implicated in the control of almost all aspects of wound healing is the transforming growth factor-beta family (TGF-beta). In the present study, the full length sequence of the axolotl TGF-beta1 cDNA was isolated. The spatio-temporal expression pattern of TGF-beta1 in regenerating limbs shows that this gene is up-regulated during the preparation phase of regeneration. Our results also demonstrate the presence of multiple components of the TGF-beta signaling machinery in axolotl cells. By using a specific pharmacological inhibitor of TGF-beta type I receptor, SB-431542, we show that TGF-beta signaling is required for axolotl limb regeneration. Treatment of regenerating limbs with SB-431542 reveals that cellular proliferation during limb regeneration as well as the expression of genes directly dependent on TGF-beta signaling are down-regulated. These data directly implicate TGF-beta signaling in the initiation and control of the regeneration process in axolotls.
PLoS ONE 02/2007; 2(11):e1227. · 4.09 Impact Factor
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ABSTRACT: Urodele amphibians like the axolotl are unique among vertebrates in their ability to regenerate and their resistance to develop cancers. It is unknown whether these traits are linked at the molecular level.
Blocking p53 signaling in axolotls using the p53 inhibitor, pifithrin-alpha, inhibited limb regeneration and the expression of p53 target genes such as Mdm2 and Gadd45, suggesting a link between tumor suppression and regeneration. To understand this relationship we cloned the p53 gene from axolotl. When comparing its sequence with p53 from other organisms, and more specifically human we observed multiple amino acids changes found in human tumors. Phylogenetic analysis of p53 protein sequences from various species is in general agreement with standard vertebrate phylogeny; however, both mice-like rodents and teleost fishes are fast evolving. This leads to long branch attraction resulting in an artefactual basal emergence of these groups in the phylogenetic tree. It is tempting to assume a correlation between certain life style traits (e.g. lifespan) and the evolutionary rate of the corresponding p53 sequences. Functional assays of the axolotl p53 in human or axolotl cells using p53 promoter reporters demonstrated a temperature sensitivity (ts), which was further confirmed by performing colony assays at 37 degrees C. In addition, axolotl p53 was capable of efficient transactivation at the Hmd2 promoter but has moderate activity at the p21 promoter. Endogenous axolotl p53 was activated following UV irradiation (100 j/m2) or treatment with an alkylating agent as measured using serine 15 phosphorylation and the expression of the endogenous p53 target Gadd45.
Urodele p53 may play a role in regeneration and has evolved to contain multiple amino acid changes predicted to render the human protein defective in tumor suppression. Some of these mutations were probably selected to maintain p53 activity at low temperature. However, other significant changes in the axolotl proteins may play more subtle roles on p53 functions, including DNA binding and promoter specificity and could represent useful adaptations to ensure p53 activity and tumor suppression in animals able to regenerate or subject to large variations in oxygen levels or temperature.
BMC Evolutionary Biology 02/2007; 7:180. · 3.52 Impact Factor
