Treatment of resting zone chondrocytes with transforming growth factor-beta 1 induces differentiation into a phenotype characteristic of growth zone chondrocytes by downregulating responsiveness to 24,25-(OH)2D3 and upregulating responsiveness to 1,25-(OH)2D3.

To determine if transforming growth factor-beta 1 (TGF-beta 1) can induce the differentiation of resting zone (RC) chondrocytes, confluent, fourth passage cultures of these cells were pretreated for 24, 36, 48, 72, and 120 h with TGF-beta 1. At the end of pretreatment, the media were replaced with new media containing 10(-10)-10(-8) mol/L 1,25-(OH)2D3 and the cells incubated for an additional 24 h. This second treatment was chosen because prior studies had shown that only the more mature growth zone (GC) chondrocytes respond to this vitamin D3 metabolite. The effect of TGF-beta pretreatment on cell maturation was assessed by measuring alkaline phosphatase (ALPase)-specific activity. In addition, changes in matrix protein synthesis were assessed by measuring collagen synthesis, as well as 35SO4 incorporation into proteoglycans. When RC cells were pretreated for 120 h with TGF-beta 1, treatment with 1,25-(OH)2D3 caused a dose-dependent increase in ALPase-specific activity and collagen synthesis, with no effect on proteoglycan production. RC cells pretreated with 1,25(OH)2D3 responded like RC cells that had not received any pretreatment. RC cells normally respond to 24,25-(OH)2D3; however, RC cultures pretreated for 120 h with TGF-beta 1 lost their responsiveness to 24,25-(OH)2D3. These results indicate that TGF-beta 1 directly regulates the maturation of RC chondrocytes into GC chondrocytes and support the hypothesis that this growth factor may play a significant role in regulating chondrocyte maturation during endochondral ossification.