Development of the microcirculation of the secondary ossification center in rat humeral head

This work investigated the origin and development of microcirculation in the rat humeral head and the expression of vascular endothelial growth factor (VEGF) as a factor supporting the vascular growth and the development of the secondary ossification centers. Sixty rats aging 1, 3-4, 6-8, 11, and 21 days, 5 weeks, and 4 and 8 months were used. Samples of humeral head were collected for histology and immunohistochemistry for VEGF. Some animals were perfused with Mercox resin in order to obtain vascular corrosion casts (vcc) observed by scanning electron microscopy (SEM). No cartilage canals were present at birth. At 6 days postnatal, blood vessels coming from the perichondrium and the region near the capsule attachment invaded the cartilage; at 11 days postnatal, signs of calcification were present and within the third week some bone trabeculae were formed. Just before the vascular invasion of the epiphysis, a positive reaction for VEGF was localized in chondrocytes of the epiphyseal cartilage close to the capsule insertion. During the development and expansion of the secondary ossification center, VEGF expression was higher in chondrocytes but decreased when epiphysis was diffusely ossified. VEGF was expressed also by mesenchymal cells present in and around the fibrous tissue where the secondary ossification center will develop. SEM vcc confirmed that vessels penetrating into the epiphysis arose merely from the periosteal and the capsular networks, and vascular connections with the diaphyseal circulation were not evident. These observations demonstrated that VEGF production by chondrocytes begun some days after birth, supported the rapid vascular growth from the surrounding soft tissues, and was chronologically related to the development of the secondary ossification center in rat proximal humerus. Finally, the possible role of VEGF as mediator of angiogenesis and, at least indirectly, as a trigger factor also in the ossification and the bone remodeling of the secondary ossification centers has been discussed.