INHIBITION OF BMP RECEPTOR SYNTHESIS BY ANTISENSE OLIGONUCLEOTIDES ATTENUATES OP-1 ACTION IN PRIMARY CULTURES OF FETAL-RAT CALVARIA CELLS

Osteogenic protein-1 (OP-1 or bone morphogenetic protein-7 [BMP-7]) st imulates osteoblast differentiation in vitro and induces bone formatio n in vivo, BMPs exert their effects through complex formation with a h eterodimeric receptor composed of a type I and a type II polypeptide, In the present study, mRNAs for three BMP subtype I receptors (ActR-I, BMPR-IA, and BMPR-IB) and one BMPR-II receptor were detected by North ern analysis in two human osteosarcoma cell lines (SaOS-2 and TE85) an d in the primary cultures of fetal rat calvaria (FRC) cells, OP-1 affe cted the steady-state mRNA levels of these receptors differently among these cell types, To study the role of each receptor type in OF-l act ion in FRC cells, receptor synthesis was inhibited by antisense oligon ucleotides. Inhibition of receptor synthesis was confirmed by immunopr ecipitation of radiolabeled cellular proteins with specific antibodies . The osteogenic action of OP-1 was measured by alkaline phosphatase ( ALP) activity and mineralized bone nodule formation in FRC cells. Resu lts showed that inhibition of synthesis of a single subtype I receptor alone did not affect significantly the OP-1-stimulated ALP activity. Inhibition of BMPR-II synthesis reduced the OP-1-stimulated ALB activi ty by about 50%, Inhibition of synthesis of any one of the type I rece ptor plus the BMPR-II receptor did not reduce the OP-1-stimulated ALB activity significantly beyond that observed by inhibition of BMPR-II a lone. Under these conditions, nodule formation was affected similarly, thus supporting the observations made with the ALP measurements. The present results suggest that the ActR-I, BMPR-IA, and BMPR-IB receptor s and the BMPR-II receptor are expressed and functional for OP-1 in FR C cells and that regulation of synthesis of these receptors may be a m echanism by which a specific cell type responds to OP-1. The turnover rate of these receptor proteins might be relatively long and another t ype II: receptor(s) for OP-1. might be functional in FRC cells.