RETINOIC ACID IS A MAJOR REGULATOR OF CHONDROCYTE MATURATION AND MATRIX MINERALIZATION

During the process of endochondral bone formation, chondrocytes underg o a series of complex maturational changes. Our recent studies indicat e that this maturational process is influenced by the vitamin A deriva tive retinoic acid (RA). To learn how this agent regulates chondrocyte development, we characterized matrix gene expression during maturatio n of cartilage cells in chick sternum. RNAs were isolated from the cep halic portion of day 13, 14, 16, 18, and 20 chick embryo sternum and a nalyzed via northern blots. Type II collagen RNA levels remained fairl y constant during this developmental period. In contrast, expression o f type X collagen and alkaline phosphatase (APase) genes was first det ected at day 16, followed by that of osteonectin (ON) and osteopontin (OF). To explore the mechanisms triggering these changes, chondrocytes were isolated from the cephalic portion of day 17-18 sternum (US cell s) and grown in monolayer in standard serum-containing medium. After 3 weeks in culture, most of the cells enlarged and became type X collag en-positive, but they exhibited low APase activity and contained only trace amounts of ON and OP mRNAs. Treatment of parallel 3-week-old cul tures with RA (10-100 nM) rapidly increased expression of the APase, O N, and OF genes severalfold. In concert with a significant increase in APase activity, there was abundant calcium accumulation in the RA-tre ated cultures. Electron microscopy confirmed the formation of large ma trix-associated mineral crystals and the presence of numerous matrix v esicles. The effects of RA were also studied in cultures of immature c hondrocytes isolated from the caudal portion of sternum (LS cells). In these cells, RA failed to induce high levels of APase activity, ON an d OP gene expression, and mineralization; instead, it greatly promoted cell proliferation. Thus RA appears to have major, stage-specific eff ects on the maturation program of chondrocytes. The retinoid rapidly i nduces expression of late maturation genes and activates mineralizatio n of the cartilage matrix. (C) 1994 Wiley-Liss, Inc.