Innovations_in_Molecular_Mechanisms_and_Tissue_Engineering_(Stem_Cell_Biology_and_Regenerative_Medicine)

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organization of distally-located proliferating chondrocytes differentiating into hyper-

trophic chondrocytes demonstrates the induction of an endochondral ossifi cation

center in response to BMP2 that is analogous to the chondrogenic organization of the

developing P2 bone (Fig. 5.2e ). While cartilaginous cells of the regenerate lacked the

customary columnar organization of a growth plate, the polarized production of

matrix, thus suggestive of columnar chondrocytes , was similar to that of the growth

plate. These data supported the conclusion that P2 bone elongation was mediated via

the regeneration of a growth plate-like structure in response to BMP2 treatment.

Moreover, and akin to bone development, the endochondral ossifi cation center of the

regenerate is comprised of chondrocytes, which function to create a template for

subsequent bone deposition, thus regeneration of the amputated bone. The BMP2 -

induced regeneration of bone was contiguous with and constrained to P2, however

with no indication of joint tissue or P3 regeneration, thus the induced regeneration

response is segment specifi c. The segment-specifi c nature of this BMP2 -induced

regenerative response was also demonstrated in an adult model of limb amputation

where patterned skeletal elements were induced to regenerate following amputation

through the shank [ 19 ].

For non-regenerating amputations of adult digits, the P2 amputation has emerged

as a model system to study both tissue repair after amputation and induced regen-

eration [ 20 – 24 ]. P2 is analogous to other long bones of the mammalian body, in that

it cannot mount a successful endogenous regeneration response after amputation.

Instead, the injured structure undergoes wound repair and ultimately scar formation,

nonetheless, the wound repair response of P2 is quite dynamic. P2 is located cen-

trally within the digit , bounded dorsally by a ligament and overlying dermis and

epidermis rich with hair follicles, and ventrally by a deep digital fl exor tendon with

associated fi brocartilage , and underlying dermis and epidermis. The P3 marrow is

distinct from the P2 marrow, in that P3 contains abundant vasculature and is rela-

tively more cell dense compared to the fatty P2 marrow. Amputation through P2

exposes the marrow cavity to the wound site and completely removes the nail organ

and digit tip (Fig. 5.3a ). P2 wound closure is achieved by forward contraction of the

dermal tissues, apparent by 6 DPA , distinct from the characteristic epidermal retrac-

tion of P3 post amputation (Fig. 5.3b ). Moreover, while osteoclast mediated bone

erosion is associated with the P2 amputation response, evident by the signifi cant dip

in bone volume at 7 DPA, the bone erosion typically does not result in expelling the

bone with concomitant wound closure as it does in P3. Instead the epidermis and

underlying soft connective tissue migrate distal to the bone stump, closing the

wound, with wound epidermis formation by 9 DPA (Fig. 5.3c ). Unlike the blastemal

intramembranous bone redifferentiation that represents P3 regeneration, the P2

response to amputation is via the formation of a periosteal-derived cartilaginous

callus , testing immunopositive for several cartilage matrix proteins, including

Collagen 2 and Aggrecan by 9 DPA (Fig. 5.3c–e outlined). Importantly, the transient

cartilaginous callus is observed exclusively along the periosteal surface proximal to

L.A. Dawson et al.