Mammalian Appendages Research Articles

Antler regeneration: a dependent process of stem tissue primed via interaction with its enveloping skin

Deer antlers are unique mammalian appendages in that each year they are cast and fully regenerate from permanent bony protuberances, called pedicles. In a previous study, we found that there is a difference in the degree of association between pedicle bone and its enveloping skin: tight at the distal third and loose at the proximal two thirds of a pedicle stump. The distal part has been termed the "potentiated" region, and the proximal part the "dormant" region. In the present study, pedicle stumps were artificially created in yearling sika deer by cutting off the tissue distal to either the potentiated or the dormant region. A piece of impermeable membrane was then inserted into the space between the bone and the skin of each treated pedicle stump, while the control pedicles had the same surgery without membrane insertion. The results showed that the inserted membrane blocked pedicle skin participation in the process of antler regeneration. All three potentiated bony pedicle stumps regenerated skin-less antlers; whereas, one of the three dormant bony pedicle stumps failed to regenerate any antler tissue. The other two dormant stumps eventually regenerated normal antlers; however, this only occurred after loss of the inserted membrane. No antler tissue regenerated from the dormant stumps while the inserted membrane remained in place (up to 55 days). All control pedicle stumps regenerated normal antlers. Therefore, we conclude that it is the pedicle bone, but not pedicle skin, that gives rise to regenerating antlers, and that pedicle bone can acquire the potential to regenerate an antler only when it is primed via interaction with its enveloping skin.

Identification of key tissue type for antler regeneration through pedicle periosteum deletion

Epimorphic regeneration is the "holy grail" of regenerative medicine. Research aimed at investigating the various models of epimorphic regeneration is essential if a fundamental understanding of the factors underpinning this process are to be established. Deer antlers are the only mammalian appendages that are subject to an annual cycle of epimorphic regeneration. In our previous studies, we have reported that histogenesis of antler regeneration relies on cells resident within the pedicle periosteum (PP). The present study elaborates this finding by means of functional studies involving the deletion of PP. Four yearling and four 2-year-old stags were selected for total PP deletion or partial PP deletion experiments. Of the animals in the total PP deletion group, one showed no signs of antler regeneration throughout the antler growth season. Two showed substantial and one showed marginal delays in antler regeneration (at 34, 20 and 7 days, respectively) compared with the corresponding sham-operated sides. Histological investigation revealed that the delayed antlers were derived from regenerated PP. Unexpectedly, the regenerative capacity of the antler from the total periosteum-deleted pedicles depended on antler length at surgery. Of the four deer that had partial PP deletion, two regenerated antlers exclusively from the left-over PP on the pedicle shafts in the absence of participation from the pedicle bone proper. The combined results from the PP deletion experiments convincingly demonstrate that the cells of the PP are responsible for antler regeneration.

Programmed cell death in the regenerating deer antler

Antlers are the only mammalian appendages capable of epimorphic regeneration and thus provide a unique model for investigating the mechanisms that underlie mammalian regeneration. Antlers elongate by a modified endochondral ossification process while intramembranous ossification takes place concurrently around the antler shaft. In this study, sites of apoptosis in the growing antler tip were identified by TUNEL staining and related to cell proliferation, as determined by PCNA staining. Bcl-2 and bax were identified by RT-PCR and bax was also immunolocalized in tissue sections. The apoptotic index was high in perichondrium, undifferentiated mesenchymal cells and cellular periosteum but was low in skin. The proliferation index was high in mesenchyme, skin (specifically in hair follicles) and cellular periosteum; it was low in fibrous perichondrium and periosteum, and barely detectable in cartilage. Both bcl-2 and bax were found to be more highly expressed in the perichondrium/mesenchyme and non-mineralized cartilage than in skin and mineralized cartilage. Bax was immunolocalized in mesenchyme cells, chondroprogenitors, chondrocytes, osteoblasts, osteocytes and osteoclasts. In conclusion, this study shows that programmed cell death plays a necessary role in regenerating antlers, as it does during skeletal development, bone growth and bone remodelling. The high level of apoptosis and proliferation in mesenchymal progenitor cells confirms that this represents the antler 'growth zone'. In fact, the percentage of TUNEL-positive cells in the mesenchymal growth zone (up to 64%) is higher than that recorded in any other adult tissue. This extensive cell death probably reflects the phenomenal rate of morphogenesis and tissue remodelling that takes place in a growing antler. The local and/or systemic factors that control the balance between cell growth and apoptosis in antler tissues now need to be determined.

The newt in us

Research on regeneration is a rapidly expanding field and increasingly attracts commercial interest, but therapeutic applications are clearly not around the next corner

Recapitulation of the parathyroid hormone-related peptide-Indian hedgehog pathway in the regenerating deer antler.

Parathyroid hormone (PTH)-related peptide (PTHrP) and the PTH/PTHrP receptor (PPR) play an essential role in controlling growth plate development. The aim of the present study was to use the deer antler as a model to determine whether PTHrP and PPR may also have a function in regulating cartilage and bone regeneration in an adult mammal. Antlers are the only mammalian appendages that are able to undergo repeated cycles of regeneration, and their growth from a blastema involves a modified endochondral process. Immunohistochemistry was used to establish sites of localization of PTHrP and PPR in antlers at different stages of development. The pattern of Indian Hedgehog (IHH) and transforming growth factor-beta1 (TGF beta1) distribution was also investigated, because PTHrP expression in the developing limb is regulated by IHH and during embryonic growth plate formation TGF beta1 acts upstream of PTHrP to regulate the rate of chondrocyte differentiation. In the antler blastema (<10 days of development), PTHrP, PPR, and TGF beta1 were localized in epidermis, dermis, regenerating epithelium, and in mesenchymal cells but IHH expression was not detected. In the rapidly growing antler (weeks 4-8 of development), PTHrP, PPR, and TGF beta1 were localized in skin, perichondrium, undifferentiated mesenchyme, recently differentiated chondrocytes, and in perivascular cells in cartilage but not in fully differentiated hyperytrophic chondrocytes. IHH was restricted to recently differentiated chondrocytes and to perivascular cells in cartilage. In mineralized cartilage and bone, PTHrP, PPR, IHH, and TGF beta1 were immunolocalized in perivascular cells and differentiated osteoblasts. PTHrP and PPR were also present in the periosteum. TGF beta1 in vitro stimulated PTHrP synthesis by cells from blastema, perichondrium, and cartilage. The findings of this study suggest that molecules which regulate embryonic skeletal development and postnatal epiphyseal growth may also control blastema formation, chondrogenesis, and bone formation in the regenerating deer antler. This finding is further evidence that developmental signaling pathways are recapitulated during adult mammalian bone regeneration.

Natural protein fibers

AbstractAlpha keratin fibers (hairs, wools, quills, and other mammalian appendages) together with fibroin fibers such as silks and spiders webs are all highly extensible fibrous proteins for which the mechanical properties are of primary importance both to the animal from which they originate and their ultimate application by man. Similarly, the collagens are highly inextensible fibrous proteins, which form the major component of mammalian skin and connecting structures such as tendons. All these fibrous proteins are biological polymers of polypeptide chains for which the mechanical and allied physical properties, such as water absorption, relate to both their macrostructure and their molecular and near‐molecular structure. Because of both their commercial application and their relatively complex structure at the molecular and near‐molecular level, interpretation of the physical properties of α‐keratin fibers represents the main component of this presentation. The mechanical properties of α‐keratin fibers are primarily related to the two components of the elongated cortical cells, the highly ordered intermediate filaments (microfibrils) which contain the α‐helices, and the matrix in which the intermediate filaments are embedded. The matrix consists of globular proteins plus water, the content of the latter being dependent on the fibers environment. The Extended Two‐Phase Model (ETPM) has been developed and results in a detailed coverage of the bulk mechanical properties of α‐helical fibers in terms of their known molecular and near‐molecular structure. The inextensible protein fibers, the collagens and fibroins, are also briefly discussed in terms of the relationship between mechanical properties and the structure of these fibers. © 2002 Wiley Periodicals, Inc. J Appl Polym Sci 83: 489–507, 2002

Epidermis, basement membrane, and connective-tissue healing after amputation of mouse digits: implications for mammalian appendage regeneration.

Soft tissues from amputation sites of mice were examined at both light and electron microscope levels to determine whether features of growth buds (blastemas), which are necessary for amphibian limb regeneration, exist in nonregenerating mice. Several such features were found. A small area of the wound bed was covered by wound epithelium which, as in regenerating newt limbs, initially lacked an underlying basement membrane. Serially sectioned digits revealed blastemalike growth in the subdermal layer surrounding periosteal chondrogenic cells. Mesenchymelike cells were seen among the fibroblasts and leucocytes within the proliferating tissues. However, no evidence of dedifferentiation was seen in the dermis, which persisted as an apparent intact obstruction to growth bud formation. Existence of the essential ingredients of growth buds and soft-tissue proliferation adjacent to chondrogenic cells proximally suggest that the tissues of mammalian healing may differ quantitatively rather than qualitatively from tissues of appendage regeneration. This premise is encouraging for efforts at growth enhancement in mammals.

A Mammalian Appendage: Deer Antlers . Regeneration, Function, and Evolution. Richard J. Goss. Illustrated by Wendy Andrews. Academic Press, New York, 1983. xvi, 316 pp. $45.

A Mammalian Appendage: <i>Deer Antlers</i> . Regeneration, Function, and Evolution. Richard J. Goss. Illustrated by Wendy Andrews. Academic Press, New York, 1983. xvi, 316 pp. $45.

Prospects for Regeneration in Man

Reports of fingertip regrowth in children are interpreted in the perspective of epimorphic regeneration in lower forms and in relation to a few other examples in mammals. The latter include cases of antler replacement in deer as well as ingrowth from the margins of holes cut in bat wing membranes and in the external ears of rabbits, pikas, cats, and echolocating bats. It is suggested that the relative inadequacy of regeneration in warm-blooded vertebrates may be attributed to the precocity with which they tend to form dermal scars in healing wounds, scars that are believed to preclude blastema production. Wound healing around the margins of rabbit ear holes is uniquely characterized by the development of prominent epidermal downgrowths adjacent to the severed sheets of dermis in the integument on either side of the ear. If these downgrowths act as epidermal blockades preventing scar formation in favor of allowing blastema cells to accumulate, a logical approach to the experimental induction of regeneration in normally nonregenerating mammalian appendages would involve manipulation of the mechanisms by which epidermis heals amputation stumps.