Treatment of resting zone chondrocytes with bone morphogenetic protein-2 induces maturation into a phenotype characteristic of growth zone chondrocytes by downregulating responsiveness to 24,25(OH)(2)D-3 and upregulating responsiveness to 1,25-(OH)(2)D-3

To determine if bone morphogenetic protein-2 (BMP-2) can induce the endocho ndral maturation of resting zone (RC) chondrocytes, confluent fourth-passag e cultures of these cells were pretreated for 24, 36, 48, 72, or 120 h with recombinant human BMP-2. At the end of pretreatment, the media were replac ed with new media containing 10(-10)-10(-8) M1,25-(OH)(2)D-3 or 10(-9)-10(- 7) M 24,25-(OH2)D-3 and the cells incubated for an additional 24 h. This se cond treatment was chosen, because prior studies had shown that the more ma ture growth zone (GC) chondrocytes and RC cells respond to 1,25-(OH)(2)D-3 and 24,25-(OH)(2)D-3 in distinctly different ways with respect to the param eters examined. The effect of BMP-2 pretreatment on cell maturation was ass essed by measuring alkaline phosphatase specific activity (ALPase). In addi tion, changes in matrix protein production were assessed by measuring colla gen synthesis, as well as [S-35]-sulfate incorporation into proteoglycans. When RC cells were pretreated for 72 or 120 h with BMP-2, treatment with 1, 25-(OH)(2)D-3 caused a dose-dependent increase in ALPase specific activity and collagen synthesis, with no effect on proteoglycan sulfation. RC cells pretreated with 1,25-(OH)(2)D-3 responded like RC cells that had not receiv ed any pretreatment. RC cells normally respond to 24,25(OH)(2)D-3; however, RC cultures pretreated for 72 or 120 h with BMP-2 lost their responsivenes s to 24,25(OH)(2)D-3. These results indicate that BMP-2 directly regulates the differentiation and maturation of RC chondrocytes into CC chondrocytes. These observations support the hypothesis that BMP-2 plays a significant r ole in regulating chondrocyte maturation during endochondral ossification.