Background: Histologic delineation of the events
involved in the development of long bones and the developmental
age at which these events occur is needed to elucidate the genetic
and molecular mechanisms associated with these events. This report describes
the sequence of histologic events involved in the formation of long
bones and their epiphyses in the New Zealand White rabbit.
Methods: Prenatal studies were performed on twelve,
fourteen, fifteen, sixteen, eighteen, twenty-one, twenty-four, and
twenty-seven-day-old rabbit embryos, and postnatal studies were
performed on newborn rabbits and on three-to-four-day-old; one,
two, four, and six-week-old; and two, three, four, six, and eight-month-old
rabbits. Histologic specimens from embryos were embedded in plastic
and stained with toluidine blue or safranin O-fast green, and specimens
from postnatal rabbits were embedded in paraffin and stained with
hematoxylin and eosin or safranin O-fast green.
Results: Studies of twelve-day-old embryos demonstrated
upper and lower limb buds filled with undifferentiated mesenchymal cells,
and studies of fourteen-day-old embryos showed mesenchymal condensation
and beginning cartilage formation outlining major long bones. Long-bone
and epiphyseal development progressed through sixteen structural
stages, and the developmental age at which these stages occurred
was determined. These stages included limb-bud formation with uniform
distribution of mesenchymal cells and formation of an apical ectodermal
ridge (stage 1); mesenchymal condensation (stage 2); cartilage differentiation
(stage 3); formation of a primary center of ossification (stage
4a); epiphyseal cartilage vascularization with formation of cartilage
canals (stage 7); vascular invasion of the developing secondary
ossification center (stage 9); bone formation and marrow cavitation
in the secondary ossification center with formation of hematopoietic marrow
(stage 10); fullest relative extent of secondary-ossification-center
development in epiphyseal cartilage (stage 14); thinning of the
physis (stage 15); and resorption of the physis with establishment
of continuity between epiphyseal and metaphyseal circulations (stage
16).
Clinical Relevance: The detailed classification
system presented here will allow for correlations between genetic
and molecular mechanisms and histologic events in normal and abnormal
development of long bones and their epiphyses. Many of the nonosseous structures
formed during long-bone and epiphyseal development in the fetus,
infant, and child are amenable to assessment with sonography and
magnetic resonance imaging. An understanding of the histopathological
features of developmental abnormalities of the long bones and their
epiphyses revealed with newer imaging techniques should greatly
improve management by allowing earlier diagnosis.