No abstract

Tamoxifen inhibits bone resorption by disrupting calmodulin-dependent processes. Since tamoxifen inhibits protein kinase C in other cells, we compared the effects of tamoxifen and the PKC inhibitor, bis indolylmaleimide II (bIM), on bone resorption and acid transport activity in isolated membrane vesicles. Bis indolylmaleimide inhibited bone resorption 50% with an IC50 approximately 3 microM, as well as acid transport activity in a concentration -dependent manner with an IC50 of approximately 0.4 IM. The IC50 of bIM for inhibiting acid transport activity was similar to that of calmodulin antagonists. The potassium ionophore, valinomycin, failed to restore bIM or tamoxifen-dependent inhibition of acid transport, suggesting that bIM and tamoxifen both inhibit H(+)-ATPase activity. Half maximal inhibitory concentrations of tamoxifen and bIM were not additive in acid transport assays, suggesting different sites of action. Furthermore, exogenous calmodulin blocked tamoxifen, but not bIM, -dependent inhibition of acid transport. We also compared the effects of tamoxifen and bIM on phosphorylation of proteins in isolated membrane fractions as determined by 32P incorporation and autoradiography. Tamoxifen had no effect on protein phosphorylation in contrast to bIM, which inhibited phosphorylation of eight proteins with different apparent kinetics. The data suggest that, while tamoxifen and bIM both affect H(+)-ATPase activity, the mechanisms of action are different.

Tamoxifen inhibits bone resorption by disrupting calmodulin-dependent processes. Since tamoxifen inhibits protein kinase C in other cells, we compared the effects of tamoxifen and the phorbol ester, phorbol myristate acetate, on osteoclast activity. Phorbol esters stimulate bone resorption and calmodulin levels four-fold (k0.5 = 0.1-0.3 microM). In contrast, tamoxifen inhibited osteoclast activity approximately 60% with an IC50 of 1.5 microM, had no apparent effect on protein kinase C activity in whole-cell lysates, and reduced protein kinase C alpha recovered by immunoprecipitation 75%. Phorbol esters stimulated resorption in a time-dependent manner that was closely correlated with a similar-fold increase in calmodulin. Protein kinase C alpha, beta, delta, epsilon, and zeta were all down-regulated in response to phorbol ester treatment. Tamoxifen and trifluoperazine inhibited PMA-dependent increases in bone resorption and calmodulin by 85 +/- 10%. Down-regulation of protein kinase C isoforms by phorbol esters suggests that the observed increases in bone resorption and calmodulin levels are most likely due to a mechanism independent of protein kinase C and dependent on calmodulin. In conclusion, the data suggest that protein kinase C negatively regulates calmodulin expression and support the hypothesis that the effects of both phorbol esters and tamoxifen on osteoclast activity is mediated by calmodulin.

One phase treatment for Class II malocclusion with high-pull headgear followed by fixed orthodontic appliances resulted in correction to Class I molar through restriction of horizontal maxillary growth with continued horizontal mandibular growth and vertical skeletal changes unaffected. The anteroposterior molar correction and skeletal effects of this treatment were stable long term.

Cyclosporine results in rapid and profound bone loss in transplant patients, an effect ascribed to osteoclasts. Cyclosporine, complexed with the appropriate immunophilin, inhibits calcineurin (the calcium/calmodulin dependent serine/threonine phosphatase) activity. We tested the hypothesis that cyclosporine inhibits calcineurin activity in osteoclasts, resulting in stimulation of osteoclast activity. We compared the effects of cyclosporine A and the calmodulin antagonist, tamoxifen, on bone resorption by avian osteoclasts. Tamoxifen inhibits bone resorption ϳ60%, whereas cyclosporine A only inhibited bone resorption 12%. One-hour treatment with 100 nM cyclosporine inhibited osteoclast calcineurin activity 70% in whole cell lysates, whereas 10 M tamoxifen only inhibited calcineurin activity 25%. We compared the effects of cyclosporine A and tamoxifen on acid transport activity in isolated membrane vesicles and in isolated membrane vesicles obtained from osteoclasts treated with cyclosporine A or tamoxifen under conditions that inhibit calcineurin activity. Direct addition of cyclosporine A in the acid transport assay, or pretreatment of cells with cyclosporine A followed by membrane isolation, had no effect on acid transport activity in membrane vesicles. In contrast, direct addition of tamoxifen to membranes inhibits acid transport activity, an effect that can be prevented by addition of exogenous calmodulin. Furthermore, acid transport activity was also inhibited in membrane vesicles isolated from cells treated with tamoxifen. In conclusion, cyclosporine A inhibits osteoclast calcineurin activity; however, calcineurin inhibition does not correspond to a significant effect on acid transport activity in isolated membrane vesicles or bone resorption by osteoclasts. (J Bone Miner Res 2003;18:451-457)