Vanadate prevents glucocorticoid-induced apoptosis of osteoblasts in vitro and osteocytes in vivo

Skeletal mass is maintained by a balance between formation and resorption, cell proliferation and apoptosis. In vitro, glucocorticoids (GCs) decrease extracellular signal-regulated kinases (ERK) activation by mitogens, thus inhibiting osteoblast proliferation. Both ERK activity and proliferation are restored by co-treatinent with the protein tyrosine phosphatase inhibitor, vanadate. Since ERK signalling may also be anti-apoptotic, we explored the effects of vanadate on GC-induced apoptosis in vitro and in vivo. Apoptosis in MBA-15.4 pre-osteoblasts increased from 6 h and remained up to eightfold higher through 6 days of 10(-6) M dexamethasone (Dex) treatment. Co-incubation with 10(-7) M vanadate markedly reduced apoptosis at all time points. Vanadate also prevented GC-induced poly-ADP-ribose polymerase cleavage. We assessed the transcriptional profiles of seven antiapoptotic proteins (Bcl-2, Bcl-X-L, inhibitors of apoptosis protein-1 (IAP-1), IAP-2, X-linked IAP (XIAP), Fas-associated death-domain-like IL-1 beta-converting enzyme-inhibitory protein (FLIPLong) and FLIPShort) in osteoblasts subjected to various stimuli using real-time quantitative PCR. Although these anti-apoptotic genes responded to different mitogenic conditions, Dex failed to repress their expression, IAP-2 and and in fact significantly up-regulated Bcl-X-L, XIAP. Dex may therefore induce apoptosis by up-regulating pro-apoptotic gene expression. We have previously demon-strated that rats treated with GC develop low formation osteoporosis (bone histoinorphometry and DEXA) and skeletal fragility (breaking strength) that were largely prevented by co-treatment with vanadate. We report here that vertebrae from rats treated with 3-5 mg/kg per day methylprednisolone for 9 weeks showed increased incidence of terminal deoxynucleotidyl transferase-mediated biotindUTP nick end-labelling-positive apoptotic osteocytes, which was reduced by vanadate co-treatment. We conclude that vanadate prevents GC-induced apoptosis of pre-osteoblasts in vitro and osteocytes in vivo, and this may contribute to its bone-sparing effects in vivo.