Ossification of the mouse metatarsal: Differentiation and proliferation in the presence/absence of a defined growth plate

There is significant diversity in growth plate behavior among sites within an individual skeleton and between skeletons of different species. This variation within wild-type animals is an underutilized resource for studying skeletal development. One bone that potentially exhibits the most diverse behavior is the metatarsal. While one end forms a growth plate with an epiphyseal secondary center of ossification as in other long bones, the opposite end undergoes direct ossification in a manner more similar to short bones. Although descriptions of human metatarsal/metacarpal ossification are available, a detailed comparative analysis has yet to be conducted in an animal model amenable to biomolecular analysis. Here we report an analysis of proximal and distal ossification in an age series of mouse metatarsals. Safranin O staining was used for qualitative and quantitative histology, and chondrocyte differentiation and proliferation were analyzed using immunohistochemistry for type X collagen and proliferative cell nuclear antigen expression. We establish that, as in the human, both growth plate formation and direct ossification occur in the mouse metatarsal, with chondrocyte populations showing distinct differentiation patterns at opposite ends of the bone. In addition, growth plate formation is characterized by a peak of proliferation in reserve zone chondrocytes that distinguishes it from both established growth plates and direct ossification. Our analysis demonstrates that the mouse metatarsal is a productive model for investigating natural variation in ossification that can further understanding of vertebrate skeletal development and evolution.