Bone formation during forelimb regeneration: A microtomography (microCT) analysis

In our study of bone regeneration in the forelimbs of mature newts (Notophthalmus viridescens), we used noninvasive X-ray microtomography (microCT) to image regenerating limbs from 37 to 85 days and matching (contralateral) controls. We compared the patterns of regenerated and existing (nonregenerated) bone, investigating in particular the onset of mineralization of specific bones, the level of mineral present, and the lengths of the different bones. Overall, we find that the missing limb skeletal elements are restored in a proximal-to-distal direction, which reiterates the developmental patterning program. However, in contrast to this proximal-distal sequence, the portion of the humerus distal to the amputation site fails to ossify in synchrony with the regenerating radius and ulna. This finding suggests that the replacement of cartilage with mineralized bone close to the amputation site is delayed with respect to other regenerating skeletal elements.