BMP-6 is an autocrine stimulator of chondrocyte differentiation

While parathyroid hormone-related protein (PTHrP) has been characterized as an important negative regulator of chondrocyte maturation in the growth pl ate, the autocrine or paracrine factors that stimulate chondrocyte maturati on are not well characterized. Cephalic sternal chondrocytes mere isolated from 13-day embryos, and the role of bone morphogenetic protein-6 (BMP-6) a s a positive regulator of chondrocyte maturation was examined in monolayer cultures. Progressive maturation, which was accelerated in the presence of ascorbate, occurred in the cultures. During maturation, the cultures expres sed high levels of BMP-6 mRNA which preceded the induction of type X collag en mRNA. Treatment of the cultures with PTHrP (10(-7) M) at the time of pla ting completely abolished BMP-6 and type X collagen mRNA expression. Remova l of PTHrP after 6 days was followed by the rapid (within 24 h) expression of BMP-6 and type X collagen mRNA, with BMP-6 again preceding type X collag en expression. The addition of exogenous BMP-6 (100 ng/ml) to the cultures accelerated the maturation process both in the presence and absence of asco rbate and resulted in the highest levels of type X collagen. When exogenous BMP-6 was added to PTHrP containing cultures, maturation occurred with the expression of high levels of type X collagen, despite the presence of PTHr P in the cultures. Furthermore, BMP-6 did not stimulate expression of its o wn mRNA in the PTHrP treated cultures, but it did stimulate the expression of Indian hedgehog (Ihh) mRNA. These latter findings suggest that while PTH rP directly inhibits BMP-6, it indirectly regulates Ihh expression through BMP-6. Other phenotypic changes associated with chondrocyte differentiation were also stimulated by BMP-6, including increased alkaline phosphatase ac tivity and decreased proliferation. The results suggest that BMP-6 is an au tocrine factor that initiates chondrocyte maturation and that PTHrP may pre vent maturation by inhibiting the expression of BMP-6.