THE INTEGRIN LIGAND ECHISTATIN PREVENTS BONE LOSS IN OVARIECTOMIZED MICE AND RATS

Integrins that bind RGD (arginine-glycine-aspartic acid) containing pe ptides, especially the vitronectin receptor alpha(v) beta(3), have bee n implicated in the regulation of osteoclast function. Echistatin, an RGD-containing snake venom peptide with high affinity for beta(3) inte grins, as well as nonpeptide RGD mimetics, were shown to inhibit osteo clastic bone resorption in vitro and in vivo. To evaluate the role of RGD-binding integrins in bone metabolism, we examined by several metho ds the effects of echistatin on ovariectomy (OVX)-induced bone loss in mice and rats. First, we confirmed that echistatin binds in vitro wit h high affinity (K-d, 0.5 nM) to alpha(v) beta(3) integrin purified fr om human placenta and established a competitive binding assay to measu re echistatin concentrations in serum. We find that echistatin infused for 2 or 4 weeks at 0.36 mu g/h.g body weight (similar to 50 nmol/day .mouse) completely prevents OVX-induced cancellous bone loss in the di stal femora of ovariectomized mice. Echistatin has no effect on uterin e weight, body weight, and femoral length changes induced by OVX, nor does it cause any apparent changes in major organs other than bone. In OVX rats, echistatin infusion at 0.26 mu g/hg for 4 weeks effectively prevents bone loss, evaluated by dual energy x-ray absorptiometry of the femur, by femoral ash weight, and by bone histomorphometry of the proximal tibia. At effective serum concentrations of 20-30 nM, measure d at the end of the infusion period, echistatin maintains histomorphom etric indices of bone turnover at control levels but does not decrease osteoclast surface. In conclusion, these results provide in vivo evid ence, at the level of bone histology, that RGD-binding integrins, prob ably alpha(v) beta(3), play a rate-limiting role in osteoclastic bone resorption and suggest a therapeutic potential for integrin ligands in the suppression of bone loss.