Production of angiogenesis inhibitors and stimulators is modulated by cultured growth plate chondrocytes during in vitro differentiation: dependence on extracellular matrix assembly.

Secretion of angiogenesis inhibitors and stimulators is modulated during in vitro differentiation of embryonic chick growth plate chondrocytes. Supernatants from dedifferentiated cells undergoing maturation to hypertrophic chondrocytes in suspension progressively inhibited vascular cell random migration and invasion of basement membrane matrix by endothelial cells. Maximal inhibition was exhibited by conditioned medium from hypertrophic chondrocytes. The same medium also repressed vascular cell migration induced by highly angiogenic Kaposi's sarcoma cell supernatants and prevented formation of an anastomosed network of tube-like structures by endothelial cells plated on matrigel. On the contrary, when the suspension culture of hypertrophic chondrocytes was supplemented with ascorbic acid, a condition leading to the formation of a mineralized tissue similar to calcified cartilage, a dramatic switch to production of angiogenic activity was observed. Medium conditioned by osteoblast-like cells derived from hypertrophic chondrocytes also induced vascular cell migration and invasion of basement membrane matrix. The presence of angiogenic activity in the conditioned medium was assessed also by an in vivo assay in mice using reconstituted basement membrane associated with heparin. Therefore, interactions of chondrocytes with their extracellular matrix are an absolute requirement for the expression of angiogenic activities by hypertrophic chondrocytes at late developmental stages.