Review: Regeneration of the tail in lizards appears regulated by a balanced expression of oncogenes and tumor suppressors

doi:10.1016/j.aanat.2021.151824

Annals of Anatomy - Anatomischer Anzeiger

Volume 239, January 2022, 151824

https://doi.org/10.1016/j.aanat.2021.151824 Get rights and content

Abstract

Background

Tail regeneration in lizards is the only case of large multi-tissue organ regeneration in amniotes.

Methods

The present Review summarizes numerous immunolocalization and gene-expression studies indicating that after tail amputation in lizards the stump is covered in 7–10 days by the migration of keratinocytes. This allows the accumulation of mesenchymal-fibroblasts underneath the wound epidermis and forms a regenerative blastema and a new tail.

Results

During migration keratinocytes transit from a compact epidermis into relatively free keratinocytes in a process of “Epithelial Mesenchymal Transition” (EMT). While EMT has been implicated in carcinogenesis no malignant transformation is observed during these cell movements in the regenerative blastema. Immunolabeling for E-cadherin and snail shows that these proteins are present in the cytoplasm and nuclei of migrating keratinocytes. The basal layer of the wound epithelium of the apical blastema express onco-proteins (wnt2b, egfr, c-myc, fgfs, fgfr, rhov, etc.) and tumor suppressors (p53/63, fat2, ephr, apc, retinoblastoma, arhgap28 etc.). This suggests that their balanced action regulates proliferation of the blastema.

Conclusions

While apical epidermis and mesenchyme are kept under a tight proliferative control, in more proximal regions of the regenerating tail the expression of tumor-suppressors triggers the differentiation of numerous tissues, forming the large myomeres, axial cartilage, simple spinal cord and nerves, new scales, arteries and veins, fat deposits, dermis and other connective tissues. Understanding gene expression patterns of developmental pathways activated during tail regeneration in lizards is useful for cancer research and for future attempts to induce organ regeneration in other amniotes including humans.

Section snippets

Regeneration evolved in lizards provides clues for regeneration in other amniotes

The ability to repair tissue or organ injury or loss varies broadly among vertebrates, from an almost complete regeneration of various organs in some urodele amphibians, lungfishes and actinopterigian fish to a complete failure in the other species, particularly in terrestrial forms, the amniotes (Goss, 1969, Reichman, 1984, Tsonis, 2000, Carlson, 2007). Mankind has been fascinated and at the same time wondering while only these few low vertebrates and not mammals are gifted with such an

Immunohistochemical analysis indicates cell control in the formation of the regenerative tail blastema in lizards

Histological, ultrastructural and expression studies have indicated that a process of EMT occurs during early stages of tail regeneration, the wound healing period (Alibardi, 2010a, Alibardi, 2010b, Alibardi, 2012a, Alibardi, 2012b, Gilbert et al., 2013). Keratinocytes at the borders of the stump take an elongate shape and migrate in sheets underneath the scab formed at 1–3 days post-amputation/autotomy (Fig. 1 A; Alibardi and Toni, 2005; Alibardi, 2010b; McLean and Vickaryous, 2011; Delorme et

Transcriptome and immunolocalization analyses of the regenerating blastema in the lizard P. muralis

Once the blastema is formed a regulated growth into a new tail occurs but how is this process controlled in lizards? The study trying to address this problem were empirical up to 2017 since various proteins and growth factors known to be involved in amphibians regeneration were also tested in lizards for comparison (Alibardi, 2010a, Alibardi, 2014). The situation dramatically changed when, in 2016–2017, the first transcriptome of P. muralis, was obtained and the search for specific genes and

Conclusions and the future of regeneration research

The immunolabeling studies have shown that the proteins coded by numerous key genes are distributed with the highest intensity in the same tissues known from experimental and microscopical studies to be essential for the regeneration of the tail, namely the apical wound epidermis and the ependyma of regenerating spinal cord. The regulated regeneration of the tail is likely obtained by the balanced action of onco-genes and tumor-suppressors within apical epithelial and ependymal cells and three

Ethical statement

I confirm that the study reported in this Review MS results from my personal work and that I have not submitted the MS to other Journals.

Declaration of Competing Interest

Hereby I declare that I have no conflicts of interest in my MS.

Acknowledgments

The transcriptome of P. muralis and the following immunohistochemical studies here presented have been self-supported (payment as a service from CRIBRI, University of Padova, Italy), elaborated and made by the author. I thank the skillful bioinformatics ability of Dr. Nicola Vitulo (University of Padua, Italy).

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Cited by (5)

Protein extracts from regenerating lizard tail show an inhibitory effect on human cancer cells cultivated in-vitro
2023, Annals of Anatomy
accumulating evidence indicates that during tail regeneration in lizards the initial stage of regenerative blastema is a tumor-like proliferative outgrowth that rapidly elongates into a new tail composed of fully differentiated tissues. Both oncogenes and tumor-suppressors are expressed during regeneration, and it has been hypothesized that an efficient control of cell proliferation avoids that the blastema is turned into a tumor outgrowth.
in order to determine whether functional tumor-suppressors are present in the growing blastema we have utilized protein extracts collected from early regenerating tails of 3–5 mm that have been tested for a potential anti-tumor effect on in-vitro culture by using cancer cell lines from human mammary gland (MDA-MB-231) and prostate cancer (DU145).
at specific dilutions, the extract determines a reduction of viability in cancer cells after 2–4 days of culture, as supported by statistical and morphological analyses. While control cells appear viable, treated cells result damaged and produce an intense cytoplasmic granulation and degeneration.
this negative effect on cell viability and proliferation is absent using tissues from the original tail supporting the hypothesis that only regenerating tissues synthesize tumor-suppressor molecules. The study suggests that the regenerating tail of lizard at the stages here selected contains some molecules that determine inhibition of cell viability on the cancer cells analyzed.
Immunohistochemistry indicates that persistent inflammation determines failure of tail, limb and finger regeneration in the Lizard Podarcis muralis
2022, Annals of Anatomy
Citation Excerpt :
Although basophil/eosinophil granulocytes may also be present, they are difficult to be discriminated and only in few cases they show a polymorphic nucleus (arrow in the inset of Fig. 3 C). Aside these limitations, the present observations further support the idea that activation of pro-inflammatory immune cells is one of the main causes that impede organ regeneration in lizards, and terrestrial vertebrates generally (Koh and DiPietro, 2013; Simkin et al., 2017; Mescher, 2017; Mescher et al., 2017; Iismaa et al., 2018; Alibardi, a-c, 2021; He et al., 2021). In the first experimental case analyzed, the light and electron microscopy studies indicate that the stump front in these non-regenerating tails is covered with a coagulated pus-corneous material.
The presence of white blood inflammatory cells in injured tissues and their effect on the process of organ regeneration in lizards has been assessed on tail, limb and digits.
The present immunohistochemical survey analyzes the occurrence of CD68-labeled cells in lizard organs uncapable of regenerating tissues that exhibit strong inflammatory activity.
This marker mainly identifies macrophages and mast cells present in large number within tissues of injured limbs and digits. Also a high inflammation is associated with amputated tails that do not regenerate, derived from cauterization or infection of tissues of the tail stump. In the healing limbs and fingers at 12–20 days post-amputation, numerous CD68-labeled cells, most likely macrophages, are seen among superficial connective tissues and injured muscles and bones. These cells likely stimulate and give rise to scarring tissues and no regeneration of limb and fingers occurs. In the cauterized or in the infected tail stump a strong accumulation of CD68-positive mast cells and macrophages is observed, where they likely evoke epidermal coagulation, formation of scarring connective tissue, and loss of regeneration.
The present observations provide further cytological evidence that support the notion that a strong and lasting inflammatory condition impedes organ regeneration in specifically lizards and, more generally other vertebrates as well.
Review. Limb regeneration in lizards under natural and experimental conditions with considerations on the induction of appendages regeneration in amniotes
2022, Annals of Anatomy
Citation Excerpt :
Among amniotes lizards show the highest regenerative capability in the tail but only limited healing ability and most scarring in other injured organs, like in mammals and birds (Bellairs and Bryant, 1985; Alibardi, 2010a, 2014, 2018a, 2021a; Gilbert et al., 2013; Jacyniak et al., 2017).
Study on the failure of limb regeneration in lizards evidences the difficult problems met from amniotes to regenerate organs. Contrary to the tail, limb loss in terrestrial environment is generally fatal and no selection for its regeneration occurred during lizard evolution.
Experimentally amputated limbs were fixed and embedded for microscopy.
After limb loss an intense inflammatory reaction occurs and immune cells are recruited underneath a wound epidermis, forming a vascularized granulation tissue. The regenerating epidermis takes 2–3 weeks to cover the limb stump since degenerating long bones must be excised first while a dense connective tissue is formed and no limb growth occurs. Cell proliferation occurs in granulation tissues and wound epidermis during the initial 2–3 weeks of wound healing but disappears later determining the arrest of growth. Transcriptome data indicates that the limb, contrary to the tail, activates numerous genes involved in inflammation, immunity and fibroplasia while down-regulates some proliferative and most myogenic genes. Attempts to stimulate limb regeneration, by implants of nervous tissues or growth factors such as FGFs only maintain proliferation for few weeks but eventually the scarring program prevails and only short outgrowths missing of autopodial elements are regenerated.
While lizard limbs show the typical scarring outcome of mammals, the comparison of genes activated in the regenerating tail has allowed identifying key genes implicated in organ regeneration in amniotes.
Manipulating Myc for reparative regeneration
2024, Frontiers in Cell and Developmental Biology
The salamander blastema within the broader context of metazoan regeneration
2023, Frontiers in Cell and Developmental Biology

View full text

Invited reviewReview: Regeneration of the tail in lizards appears regulated by a balanced expression of oncogenes and tumor suppressors

Abstract

Background

Methods

Results

Conclusions

Section snippets

Regeneration evolved in lizards provides clues for regeneration in other amniotes

Immunohistochemical analysis indicates cell control in the formation of the regenerative tail blastema in lizards

Transcriptome and immunolocalization analyses of the regenerating blastema in the lizard P. muralis

Conclusions and the future of regeneration research

Ethical statement

Declaration of Competing Interest

Acknowledgments

Progr. Histoch. Cytoch.

Acta Histoch

Ann. Anat.

Acta Histoch

Ann. Anat.

Zoology

Ann. Anat.

J. Biol. Chem.

Mech. Dev.

Dev. Biol.

J. Immunol. Regen. Med.

Morphological and cellular aspects of tail and limb regeneration in lizard: a model system with implications for tissue regeneration in mammals

Adv. Anat. Embryol. Cell Biol.

Ultrastructural features of the process of wound healing after tail and limb amputation in lizard

Acta Zool.

Observations on FGFs immunoreactivity in the regenerating tail blastema and in the limb and tail scars of lizard suggests that FGFs are required for regeneration

Belg. J. Zool.

Observations on FGFs immunoreactivity in the regenerating tail blastema and in the limb and tail scars of lizard suggests that FGFs are required for regeneration

Belg. J. Zool.

Immunolocalization of the telomerase-1 component in cells of the regenerating tail, testis, and intestine of lizards

J. Morphol.

Immunolocalization of a p53/p63-like protein in the regenerating tail epidermis of the wall lizard (Podarcis muralis) suggests it is involved in the differentiation of the epidermis

Acta Zool.

Immunocalization of c-myc positive cells in lizard tail after amputation suggests cell activation and proliferation for tail regeneration

Acta Zool.

Review: Biological and molecular differences between tail regeneration and limb scarring in lizard, an inspiring model addressing limb regeneration in amniotes

J. Exp. Zool. B

Immunolocalization of Sonic Hedgehog and Patched in the regenerating tail of lizard suggests they are involved in cell proliferation and epidermal differentiation

BAOJ Dermatol.

Perspective: appendages regeneration in amphibians and some reptiles derived from specific evolutionary histories

J. Exp. Zool. B

Ultrastructural immunolocalization of hyaluronate in regenerating tail of lizards and amphibians supports an immune-suppressive role to favor regeneration

J. Morphol.

Msx immunolocalization in the regenerating tail of a lizard but not in the scarring limb suggests its involvement in the process of regeneration

Acta Zool.

Immunolocalization of matrix metalloproteinases in regenerating lizard tail suggests that an intense remodeling activity allows for apical tail growth

Acta Zool.

Immunodetection of high mobility group proteins (HMGs) in the regenerating tail of lizard mainly indicates activation for cell proliferation

Acta Zool.

Review: the regenerating tail blastema of lizard as a model to study organ regeneration and tumor growth regulation in amniotes

Anat. Rec.

Epidermal growth factor and EGFreceptors are mainly expressed in the wound epidermis and proliferating ependymal of the regenerating tail of lizards

Acta Zool.

Immunohistochemical localization of a proto-cadherin fat tumor-suppressor homolog in the regenerating tail of lizard suggests a role in apical growth control

Acta Zool.

Immunodetection of Ephrin-like Receptors in the regenerating tail of the lizard Podarcis muralis suggests stimulation of differentiation and muscle segmentation

Zool. Res.

Immunolocalization of Wnts in the lizard blastema supports a key role of these signaling proteins for tail regeneration

J. Morphol.

Appendage regeneration in anamniotes utilizes genes active during larval-metamorphic stages that have been lost or altered in amniotes: the case for studying lizard tail regeneration

J. Morphol.

Regeneration in anamniotes was replaced by regengrow and scarring in amniotes after land colonization and the evolution of terrestrial biological cycles

Dev. Dyn.

Immunolocalization of cell adhesion molecules during tail regeneration in the lizardPodarcis muralisindicates coordinated control of epithelial differentiation

Acta Zool.

Immunolabeling for RhoV and actin in early regenerating tail of the lizard Podarcis muralis suggests involvement in epithelial and mesenchymal cell motility

Acta Zool.

Immunoreactivity for Dab2 and Foxp3 suggests that immune-suppressive cells are present in the regenerating tail blastema of lizard

Acta Zool.

Wound keratins in the regenerating epidermis of lizard suggest that the wound reaction is similar in the tail and limb

J. Exp. Zool.

Invited review
Review: Regeneration of the tail in lizards appears regulated by a balanced expression of oncogenes and tumor suppressors