Sclerostin blockade inhibits bone resorption through PDGF receptor signaling in osteoblast lineage cells
Cyril Thouverey, Pierre Apostolides, Julia Brun, Joseph Caverzasio, Serge Ferrari
Cyril Thouverey, Pierre Apostolides, Julia Brun, Joseph Caverzasio, Serge Ferrari
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Research Article Bone biology

Sclerostin blockade inhibits bone resorption through PDGF receptor signaling in osteoblast lineage cells

Abstract

While sclerostin-neutralizing antibodies (Scl-Abs) transiently stimulate bone formation by activating Wnt signaling in osteoblast lineage cells, they exert sustained inhibition of bone resorption, suggesting an alternate signaling pathway by which Scl-Abs control osteoclast activity. Since sclerostin can activate platelet-derived growth factor receptors (PDGFRs) in osteoblast lineage cells in vitro and PDGFR signaling in these cells induces bone resorption through M-CSF secretion, we hypothesized that the prolonged anticatabolic effect of Scl-Abs could result from PDGFR inhibition. We show here that inhibition of PDGFR signaling in osteoblast lineage cells is sufficient and necessary to mediate prolonged Scl-Ab effects on M-CSF secretion and osteoclast activity in mice. Indeed, sclerostin coactivates PDGFRs independently of Wnt/β-catenin signaling inhibition, by forming a ternary complex with LRP6 and PDGFRs in preosteoblasts. In turn, Scl-Ab prevents sclerostin-mediated coactivation of PDGFR signaling and consequent M-CSF upregulation in preosteoblast cultures, thereby inhibiting osteoclast activity in preosteoblast/osteoclast coculture assays. These results provide a potential mechanism explaining the dissociation between anabolic and antiresorptive effects of long-term Scl-Ab.

Authors

Cyril Thouverey, Pierre Apostolides, Julia Brun, Joseph Caverzasio, Serge Ferrari

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Figure 5

Scl-Ab blocked sclerostin-mediated PDGFR coactivation and M-CSF secretion in preosteoblast cultures, and calcitriol-induced osteoclast formation and activity in cocultures.

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Scl-Ab blocked sclerostin-mediated PDGFR coactivation and M-CSF secretio...
(A) Pdgfrfl/fl (control) and Pdgfr-cKO (without PDGFRs) preosteoblasts were cocultured with nonadherent bone marrow cells isolated from WT mice in the presence of Veh, 10–8 M 1,25-dihydroxyvitamin D3 (calcitriol, Vit.D3), and/or 250 ng/mL recombinant sclerostin (SOST), with or without 1.25 μg/mL Scl-Ab or 500 ng/mL anti–M-CSF antibody (MCSF-Ab) for 8 days before quantification of TRAP-positive multinucleated cells. (B) The same cocultures performed in synthetic matrix–coated multiwell plates for 15 days before quantification of demineralized areas. (C) Pdgfrfl/fl and Pdgfr-cKO preosteoblasts were pretreated with Veh, 250 ng/mL SOST with and without 1.25 μg/mL Scl-Ab for 1 hour, and treated with Veh or 25 ng/mL PDGF-BB for 24 hours before measurement of Csf1 expression by quantitative RT-PCR. Data in A–C were analyzed by 2-way ANOVA followed by Tukey’s post hoc test. (D) WT preosteoblasts were pretreated with Veh, 250 ng/mL SOST with and without 1.25 μg/mL Scl-Ab for 1 hour, and treated with Veh or 25 ng/mL PDGF-BB for 48 hours before quantification of M-CSF in culture media by ELISA. Data in D were analyzed by 1-way ANOVA followed by Tukey’s post hoc test. (E) WT preosteoblasts were pretreated with Veh or 500 ng/mL SOST with and without 2.5 μg/mL Scl-Ab for 1 hour, and then treated with 15 ng/mL PDGF-BB for the indicated time periods. PDGFR signaling and M-CSF protein level were determined by Western blot analyses. (F and G) WT preosteoblasts were pretreated with Veh or 500 ng/mL SOST for 2 hours, and then treated with 25 ng/mL PDGF-BB for 15 minutes. (F) Proteins were immunoprecipitated by anti-LRP6 antibody and detected by Western blot analyses. (G) PDGFR signaling was assessed in the remaining cell lysates.