Responses of denervated adult newt limb stumps to reinnervation and reinjury

Salley, Judith D.; Tassava, Roy A.

doi:10.1002/jez.1402150208

Cited by 30 publications

(27 citation statements)

References 21 publications

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“…It is known that in adult newts the amputated limbs must have an adequate number of nerve fibers for regeneration to occur (1, 3,6) except in the case of anomalous examples of aneurogenic limbs (16,17). Nerves are also needed for regeneration of adult Xenopus limbs and denervation inhibits regeneration (1 3, 14).…”

Section: Discussionmentioning

confidence: 99%

Denervation Effects on Limb Regeneration in Postmetamorphic Xenopus laevis

Kurabuchi

Inoue

1983

Dev Growth Differ

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The effects of denervation on limb regeneration of postmetamorphic Xenopus laevis in the early to late stages of regeneration were studied. Limbs that were denervated immediately after amputation did not shbw any signs of regeneration. Moreover regenerating limbs denervated 20, 30, 40 and 60 days after amputation showed significant regression of regenerates. After this regression of regenerates, the regenerative capacity was not restored during a long observation period. Denervation reduced both protein synthesis (3H leucine autoradiography) and mitotic activity of regenerating limbs even in later stages of regeneration. These results on the role of the nerve in limb regeneration of Xenopus were compared with' results in Xenopus and urodeles reported previously.The effects of denervation on regeneration have been studied mainly in urodelen amphibia and results have shown that nerves in the limbs are important in the initiation of regeneration (1). Denervation performed concomitantly with amputation of limbs prevented regeneration in adult newts and Ambystoma larvae (1-8). In a'dvanced stages of regeneration, the nerve was not required for morphogenesis and regeneration proceeded in the absence of nerves, although the regeneration process was slower (7, 8).In anuran species, the role of nerves in limb regeneration has not been studied extensively. Denervation of regenerating limbs of larval Runa spp. caused regression, as in urodeles in early stages of regeneration (5-9). The froglets and adults of Xenopus laevis employed in our experiments, are known to possess the capacity to regenerate limbs in heteromorphic forms (10-13). The role of the nervous system in limb regeneration in this frog has infrequently been studied (1 3-1 5), but results have not yet been conclusive. In the present experiments, we examined the effects of denervation at various regenerative stages and found that limb regeneration in this frog showed characteristic nerve dependency. MATERIALS AND METHODSFroglets of Xenopus laevis of 4.04.5 cm in snout-vent length were selected from the colony at Gunma University for experiments. They were anesthetized with 0.1 % MS 222 solution (Sandoz), and their forelimbs were unilaterally amputated at the zeugopodia. At the base of the stylopodium, all visible branches of the brachial nerves were excised for a length of 2-5 mm. Denervation was performed at the same time as amputation and 20, 30, 40 and 60 days after amputation of limbs. Groups of 12-15 animals were subjected to each denervation treatment. Unilaterally amputated non-denervated limbs were used as controls. The animals were kept in well water at 22*2"C and fed pellet grower for trout (Oriental Kobo Kogyo, Tokyo). The lengths of the regenerated limbs in each experimental series were measured every 10 days. The regenerates excised were fixed in BOUIN'S 463

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Section: Discussionmentioning

confidence: 99%

Denervation Effects on Limb Regeneration in Postmetamorphic Xenopus laevis

Kurabuchi

Inoue

1983

Dev Growth Differ

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“…These studies have shown that when nerves are severed prior to limb amputation, a blastema fails to develop and regeneration does not occur (Todd, 1823;Schotté and Butler, 1941;Singer, 1951;Brockes, 1987). The effect of denervation, however, is not permanent since axons regrow into the limb stump a short time after the procedure and re-amputation of this previously denervated limb will lead to normal regeneration (Salley and Tassava, 1981). Innervation is required during blastema formation to promote the formation of a functional wound epidermis that secretes mitogenic factors to stimulate blastemal cell proliferation (Globus and Vethamany-Globus, 1977;Globus et al, 1980).…”

Section: Introductionmentioning

confidence: 99%

Beryllium nitrate inhibits fibroblast migration to disrupt epimorphic regeneration

Cook

Seifert

2016

Development

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Epimorphic regeneration proceeds with or without formation of a blastema, as observed for the limb and skin, respectively. Inhibition of epimorphic regeneration provides a means to interrogate the cellular and molecular mechanisms that regulate it. In this study, we show that exposing amputated limbs to beryllium nitrate disrupts blastema formation and causes severe patterning defects in limb regeneration. In contrast, exposing full-thickness skin wounds to beryllium only causes a delay in skin regeneration. By transplanting full-thickness skin from ubiquitous GFP-expressing axolotls to wild-type hosts, we demonstrate that beryllium inhibits fibroblast migration during limb and skin regeneration in vivo. Moreover, we show that beryllium also inhibits cell migration in vitro using axolotl and human fibroblasts. Interestingly, beryllium did not act as an immunostimulatory agent as it does in Anurans and mammals, nor did it affect keratinocyte migration, proliferation or re-epithelialization, suggesting that the effect of beryllium is cell type-specific. While we did not detect an increase in cell death during regeneration in response to beryllium, it did disrupt cell proliferation in mesenchymal cells. Taken together, our data show that normal blastema organogenesis cannot occur without timely infiltration of local fibroblasts and highlights the importance of positional information to instruct pattern formation during regeneration. In contrast, non-blastemal-based skin regeneration can occur despite early inhibition of fibroblast migration and cell proliferation.

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“…Previous histological examination of denervated adult newt limb stumps between days 7 and 14 after amputation revealed no signs of overt tissue healing (cartilaginous callus, dermal pad, cicatrix) 12 at the distal tip. Instead, dedifferentiated cells and a wound epithelium were present 13,14 . This commitment to wound healing must therefore occur covertly.…”

Section: Discussionmentioning

confidence: 99%

“…In an attempt to define the factors that prevent limb stumps of adult newts from regenerating upon reinnervation, Salley and Tassava 10 reinjured the ends of previously amputated, denervated, now reinnervated limb stumps on day 35 after amputation/denervation in various ways. Regeneration capability was restored by reamputation of the distal 1 mm of the healed stump (100% of the cases), a single razor incision (60% of the cases), and removal by dissection of the healed tissues from the end of the stump (70% of the cases).…”

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confidence: 99%

“…Regeneration capability was restored by reamputation of the distal 1 mm of the healed stump (100% of the cases), a single razor incision (60% of the cases), and removal by dissection of the healed tissues from the end of the stump (70% of the cases). It was concluded that these experimental manipulations reestablished the opportunity for interactions between the important prerequisites for regeneration: injury, wound epithelium, and nerves 10 …”

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confidence: 99%

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Timing the commitment to a wound‐healing response of denervated limb stumps in the adult newt, Notophthalmus viridescens

Salley‐Guydon

Tassava

2006

Wound Repair Regeneration

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Adult newt limbs that are denervated 1 day after amputation undergo a wound-healing response and, although they become reinnervated, will not regenerate unless reinjured. Experiments were designed to determine when denervated limb stumps of adult newts become committed to a wound-healing response. In Experiment I, limbs were amputated and denervated 1 day after amputation. On days 7, 14, 21, 28, and 35 days after the initial amputation and denervation, stumps were reamputated to remove the distal tip. This design varied the time the distal stump was devoid of nerves before reamputation. None of the limbs reamputated at 7 days regenerated. About half of the limbs reamputated at 14 days regenerated and almost all of those reamputated at days 21, 28, and 35 regenerated. In Experiment II, limbs were denervated and then amputated on day 7 or 14, at two different levels. Limbs with a short stump became innervated earlier and regeneration occurred more frequently at both levels of amputation. The results of these experiments show that denervated limb stumps become committed to a wound-healing response between days 7 and 14 after amputation/denervation. If sufficient nerves arrive before day 7, a regeneration response is initiated. If the stump is denervated for 14 days or longer, commitment to wound healing occurs and ingrowing nerves cannot initiate a regeneration response.

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Responses of denervated adult newt limb stumps to reinnervation and reinjury

Cited by 30 publications

References 21 publications

Denervation Effects on Limb Regeneration in Postmetamorphic Xenopus laevis

Denervation Effects on Limb Regeneration in Postmetamorphic Xenopus laevis

Beryllium nitrate inhibits fibroblast migration to disrupt epimorphic regeneration

Timing the commitment to a wound‐healing response of denervated limb stumps in the adult newt, Notophthalmus viridescens

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