Positional plasticity in regenerating Amybstoma mexicanum limbs is associated with cell proliferation and pathways of cellular differentiation

Abstract

BackgroundThe endogenous ability to dedifferentiate, re-pattern, and re-differentiate adult cells to repair or replace damaged or missing structures is exclusive to only a few tetrapod species. The Mexican axolotl is one example of these species, having the capacity to regenerate multiple adult structures including their limbs by generating a group of progenitor cells, known as the blastema, which acquire pattern and differentiate into the missing tissues. The formation of a limb regenerate is dependent on cells in the connective tissues that retain memory of their original position in the limb, and use this information to generate the pattern of the missing structure. Observations from recent and historic studies suggest that blastema cells vary in their potential to pattern distal structures during the regeneration process; some cells are plastic and can be reprogrammed to obtain new positional information while others are stable. Our previous studies showed that positional information has temporal and spatial components of variation; early bud (EB) and apical late bud (LB) blastema cells are plastic while basal-LB cells are stable. To identify the potential cellular and molecular basis of this variation, we compared these three cell populations using histological and transcriptional approaches.ResultsHistologically, the basal-LB sample showed greater tissue organization than the EB and apical-LB samples. We also observed that cell proliferation was more abundant in EB and apical-LB tissue when compared to basal-LB and mature stump tissue. Lastly, we found that genes associated with cellular differentiation were expressed more highly in the basal-LB samples.ConclusionsOur results characterize histological and transcriptional differences between EB and apical-LB tissue compared to basal-LB tissue. Combined with our results from a previous study, we hypothesize that the stability of positional information is associated with tissue organization, cell proliferation, and pathways of cellular differentiation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12861-015-0095-4) contains supplementary material, which is available to authorized users.

Retinoic acid receptor regulation of epimorphic and homeostatic regeneration in the axolotl

“…This study identified one salamander-specific molecule (Geng et al, 2015), prod1, that has a PD gradient in newts and can proximalize distal blastemal cells in newts and axolotls (Echeverri and Tanaka, 2005). We did not observe an upregulation of axolotl Prod1 after RA treatment, which supports the finding that Prod1 transcripts are more abundant in distal blastemas compared with proximal blastemas in axolotls (McCusker et al, 2015). The current model is that Prod1 signals through epidermal growth factor receptor to induce Mmp9 expression (Blassberg et al, 2011).…”

“…The cellular mechanisms that impart positional memory are still unclear (McCusker et al, 2015;Phan et al, 2015;Roensch et al, 2013). Several transcription factors have been identified that presumably activate genes responsible for positional memory (Crawford and Stocum, 1988), including Meis1, Meis2 (Mercader et al, 2005), Hoxd10 (Simon and Tabin, 1993) and Hoxa13 (Gardiner et al, 1995), but our understanding of what makes a limb proximodistally duplicate, truncate, or grow the proper structure is lacking.…”

A Tissue-Mapped Axolotl De Novo Transcriptome Enables Identification of Limb Regeneration Factors

“…RNA-sequencing followed by de-novo transcriptome assembly (Haas et al, 2013; Robertson et al, 2010; Schulz et al, 2012) has offered investigators an alternative for identifying near-full-length transcripts and performing differential gene expression analyses without genome mapping. Recent axolotl transcriptome studies (Knapp et al, 2013; Li et al, 2014; McCusker et al, 2015; Monaghan et al, 2009; Stewart et al, 2013; Voss et al, 2015; Wu et al, 2013) have focused on and significantly advanced our understanding of the changes in transcription over time in the regenerating portion of the limb. However, an important missing component of the existing data sets is deep transcriptional information about each of the presumed parent tissue types within the limb, which contribute progenitors and serve as the template for the future regenerate limb.…”

A new and improved algorithm for the quantification of chromatin condensation from microscopic data shows decreased chromatin condensation in regenerating axolotl limb cells

“…In response to limb amputation, various biological processes are activated which culminate in the formation of a blastema, a population of regeneration-competent cells, at the site of injury (Fig 4A). This regeneration-specific structure is constituted by nearby mature somatic cells which migrate in, accumulate, and proliferate at the wounded site [20,21]; at early stages of regeneration this blastema is derived largely from cells of connective tissue origin [22,23]. The dense aggregate of early-stage blastema cells are visually distinct from somatic dermal cells in terms of their nuclear morphology–being larger and euchromatic in nature (compare Fig 4B and 4C).…”