Modulation of bone morphogenetic protein-2 and bone morphogenetic protein-4 gene expression in osteoblastic cell lines.

Bone morphogenetic proteins (BMPs) belong to the transforming growth factor-beta (TGF-beta) superfamily and are crucial factors in the process of bone formation. Despite knowledge on their wide distribution and expression, however, there is very little information on the biological factors that affect gene transcription of these osteoinductive agents. To investigate this aspect of BMP gene regulation we have studied the effect of a number of factors known to affect osteogenic cells. Northern analysis showed modulation of the expression of BMP-2 and BMP-4 mRNAs in two human osteosarcoma cell lines, MG63 and Saos-2, by prostaglandin E2 (PGE2), interleukin-1beta (IL-1beta), interleukin-6 (IL-6), interferon-alpha (IFN-alpha), retinoic acid and 1,25(OH)2 vitamin D3. mRNA expressions of the normally used "housekeeping genes", glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and beta-actin, were found to be susceptible to influence by some of the factors used. Hence, an oligo(dT)15-18 probe was used to reliably estimate the relative quantities of mRNA present for normalization of data. In general, all factors down-regulated mRNA expressions of BMP-2 and BMP-4 in MG63 cells. IL-6 completely abolished detectable expression of BMP-2 mRNA, which was also greatly reduced by IL-1beta, retinoic acid and 1,25(OH)2 vitamin D3. PGE2 had similar influences on BMP-2 and BMP-4 expressions, showing reductions to approximately 60% of normal. In Saos-2 cells only 1,25(OH)2 vitamin D3 had any great effect on BMP-2 expression, which was down-regulated to approximately 60% of control values. BMP-4 was down-regulated by IFN-alpha (approximately 60%) and IL-1beta (approximately 20%). We conclude that BMPs are subject to regulation by a variety of factors and that this is dependent on the stage of the cell in the osteogenic lineage. Furthermore, the use of GAPDH and beta-actin genes as "housekeeping genes" in expression-modulation studies must be treated with care.