Age-related inhibitory activity of rat bone marrow supernatant on osteoblast proliferation

Because histomorphometric indices of bone formation (osteoblastic index, tetracyclin-labeled perimeter) are deeply depressed in aged rats, while in vitro proliferation of trabecular bone cells was found increased, we hypothesized that a signal to proliferate, correctly induced by increased strains on scarce bone, could be opposed in vivo by an inhibitor present in the bone marrow extracellular medium. Thus, we tested the effect of bone marrow extracellular fluid (BM supernatant) of rat femoral diaphysis on cultures of primary osteoblasts and osteoblastic cell lines and found that it inhibited bone cell proliferation. In a group of 69 female rats aged 4, 12, and 15/21 months, there was a stepwise increase in the inhibitory activity of the BM supernatant. The double reciprocal plots relating inhibition power of the medium to BM supernatant dilution suggest that we deal with a simple system and that the kinetics of the phenomenon are the same in older and younger animals. Moreover, proliferation inhibition by BM supernatant and trabecular bone surface measured by histomorphometry in the distal femoral metaphysis were inversely correlated. Because the extracellular fluid of bone marrow is also the medium surrounding the osteoblasts and their precursor cells, our results suggest that the bone marrow negatively regulates osteogenic cells and that this inhibition could contribute to the inability of older animals to supply osteoblasts to bone in proportion to the demand. Preliminary biochemical characterization of the inhibitor suggests it to be a protein of 30-40 kDa with an isoelectric point (pI) of about 6.5.