BONE MORPHOGENETIC PROTEIN-2 INHIBITS THE SYNTHESIS OF INSULIN-LIKE GROWTH FACTOR-BINDING PROTEIN-5 IN BONE CELL-CULTURES

Previous work from our laboratory indicated that bone morphogenetic pr otein-2 (BMP-3) enhances the synthesis of insulin-like growth factor-I (IGF-I) and IGF-II by skeletal cells. The activity of EGF-I and -II i s regulated by six known IGF-binding proteins (IGFBPs). Although most IGFBPs inhibit the actions of IGF on bone growth, IGFBP-8 is stimulato ry, and its synthesis correlates with changes in osteoblast cell growt h. We tested the effects of BMP-2 on IGFBP-B expression in cultures of osteoblast-enriched cells from 22-day-old fetal rat calvariae (Ob cel ls). Treatment of Ob cells with BMP-2 caused a time- and dose-dependen t decrease in IGFBP-8 messenger RNA (mRNA) levels, as determined by No rthern blot analysis. The effect was maximal after 24 h of treatment a nd occurred at BMP-2 concentrations of 0.03-3.3 nM. Treatment with BMP -2 for 24 h also decreased IGFBP-5 polypeptide levels in the extracell ular matrix, as determined by Western blot analysis. The effects of BM P-2 on IGFBP-5 transcripts were independent of cell division, as they were observed in the presence and absence of hydroxyurea (1 mM). IGFBP -5 transcripts were barely detectable in the presence of the protein s ynthesis inhibitor cyclohexidmide at 3.6 mu M, and further suppressive effects of BMP-2 on IGFBP-5 mRNA could not be determined. BMP-2 did n ot modify the decay of IGFBP-5 mRNA in transcriptionally arrested Ob c ells. In addition, BMP-2 inhibited IGFBP-5 heterogeneous nuclear RNA, determined by reverse transcription polymerase chain reaction, after 2 -6 h of treatment, suggesting an inhibition of IGFBF-5 transcription o r processing. In conclusion, BMP-2 inhibits IGFBP-5 expression in Ob c ells through pathways that are independent of its mitogenic activity a nd through mechanisms that may involve decreased transcription or alte red RNA processing.