Bone morphogenetic protein-2 inhibits the synthesis of insulin-like growth factor-binding protein-5 in bone cell...

Previous work from our laboratory indicated that bone morphogenetic protein-2 (BMP-2) enhances the synthesis of insulin-like growth factor-I (IGF-I) and IGF-II by skeletal cells. The activity of IGF-I and -II is regulated by six known IGF-binding proteins (IGFBPs). Although most IGFBP's inhibit the actions of IGF on bone growth, IGFBP-5 is stimulatory, and its synthesis correlates with changes in osteoblast cell growth. We tested the effects of BMP-2 on IGFBP-5 expression in cultures of osteoblast-enriched cells from 22-day-old fetal rat calvariae (Ob cells). Treatment of Ob cells with BMP-2 caused a time- and dose-dependent decrease in IGFBP-5 messenger RNA (mRNA) levels, as determined by Northern blot analysis. The effect was maximal after 24 h of treatment and 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 extracellular matrix, as determined by Western blot analysis. The effects of BMP-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 synthesis inhibitor cycloheximide at 3.6 microM, and further suppressive effects of BMP-2 on IGFBP-5 mRNA could not be determined. BMP-2 did not modify the decay of IGFBP-5 mRNA in transcriptionally arrested Ob cells. 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 IGFBP-5 transcription or processing. In conclusion, BMP-2 inhibits IGFBP-5 expression in Ob cells through pathways that are independent of its mitogenic activity and through mechanisms that may involve decreased transcription or altered RNA processing.