Dexamethasone stimulation of osteoprogenitor differentiation in adult rat bone cell populations is mediated in part through an increased response to insulin-like growth factors

I have tested the hypothesis that dexamethasone (Dex)-stimulated osteoprogenitor differentiation in rat bone cell populations is mediated, in part, through the insulin-like growth factor (IGF) system. Since osteoprogenitors and adipocyte progenitors are believed to originate from the same precursors and their differentiation has been shown to be reciprocally regulated, the effects of Dex and IGFs on adipocyte formation were also evaluated. Vertebral bone cells from 3-month-old female rats were cultured for up to 20 days with or without Dex. IGF stimulation of osteoblastic colony formation was Dex-independent, whereas IGF stimulation of adipocyte formation was Dex-dependent. Dex alone did not induce adipocyte formation. An anti-type I IGF receptor (IGF-1R) antibody blocked Dex-induced osteoprogenitor differentiation, but did not affect adipocyte formation, suggesting that Dex stimulation of osteoprogenitor differentiation is mediated through the IGF-IGF-1R axis and that although an increased IGF response is involved in Dex stimulation of adipocyte formation, the signaling is not via IGF-1R. Northern and RT-PCR analyses at various time points showed that mRNA levels of virtually all the components of the IGF system were changed after Dex treatment. Interestingly, both IGF-I and IGF-1R mRNA levels were lower in Dex-treated cultures. To explain the discordance between these findings and the increased IGF responses in Dex-treated cultures, I examined the possibility that the increased IGF responses result from reduced inhibition of IGF action by IGF binding protein-4 (IGFBP-4). Protein levels of IGFBP-4 were lower in Dex-treated cultures than in controls at all 4 time points tested as shown by Western blotting, although IGFBP-4 mRNA levels were lower at day 14 but higher at day 20 in Dex-treated cultures than in controls. Proteolysis analysis showed that conditioned medium (CM) from Dex-treated cultures was more effective than CM from controls in degrading exogenous IGFBP-4, and that the proteolysis could be blocked by pre-incubating CM with an anti-pregnancy associated plasma protein-A (PAPP-A) antibody. These findings suggest that reduction of IGFBP-4 protein levels via increased proteolysis by PAPP-A appears to be the main mechanism whereby Dex increases IGF responses.