In vitro studies on the regulation of endochondral ossification by vitamin D

The research described in this article has focused on the complex autocrine, paracrine, and endocrine regulation of endochondral ossification using vitamin D metabolites and TGF-beta as models. By comparing results from a number of laboratories utilizing a diverse array of in vivo and in vitro systems, a coherent picture is beginning to emerge. Vitamin D metabolites influence cell differentiation and maturation and have direct effects on cell function. Differentiation of the mesenchymal cells into chondroblasts is regulated by both 1,25-(OH)2D3 and 24,25-(OH)2D3, as well as by TGF-beta. The resting zone chondrocytes respond primarily to 24,25-(OH)2D3 in terms of matrix synthesis and matrix vesicle biochemistry. They synthesize both metabolites and other factors that stabilize matrix vesicle enzymes like AHSG. In addition to the paracrine role these factors may play in regulating the matrix, it is possible that they may influence the cells in the growth plate itself. Growth zone chondrocytes also synthesize both metabolites, but respond primarily to 1,25-(OH)2D3 for the parameters measured in the studies described. These cells also synthesize TGF-beta which further increases alkaline phosphatase activity, perhaps via an autocrine stimulation of the cell. While cells from the calcified zone have not yet been studied directly in culture, it is likely that they respond to paracrine signals from the avascular cartilage as well as to serum-derived factors. How the signals are transferred among the cells is unknown. Certainly one can postulate information flow in both upward and downward directions. The signal transduction mechanisms for the factors at the cellular level are complex. While it is known that 1,25-(OH)2D3 stimulates gene transcription and stabilization of mRNA for proteins like alkaline phosphatase, its nongenomic effects are only beginning to emerge. Membrane effects of this metabolite have been shown in intestine and kidney in conjunction with studies on Ca flux. It is becoming increasingly evident that other steroid hormones may operate in similar ways. Studies with the rat costochondral chondrocytes are the first to show that there are specific membrane effects for at least two vitamin D metabolites and that membrane enzymes, including those involved in phospholipid metabolism, can be differentially regulated by them. Furthermore, these experiments have provided for the first time a clear hypothesis for how cells can regulate events in the extracellular matrix after the matrix vesicles are produced and incorporated into the matrix.