Intraperitoneal clodronate liposomes remodel the local macrophage niche and potentiate biomaterial-induced osteoinduction

Background Osteoinductive biomaterials hold promise for repairing critical-size bone defects, yet the mechanisms underlying materials-instructed osteoinduction (MIOI) remain unclear. Macrophages are central, and local liposomal clodronate (LipClod) typically suppresses osteogenesis. Whether systemic LipClod modulates MIOI is unknown. We investigated the effect of intraperitoneal (i.p.) LipClod on ectopic osteoinduction. Methods Male FVB mice received subcutaneous β-tricalcium phosphate (TCPs) scaffolds and a single i.p. LipClod dose 1 day after implantation; vehicle liposomes served as controls (n = 6 per group; 10 µL/g). Recombinant macrophage colony-stimulating factor (M-CSF; 0.5 or 2 µg/mL; 100 µL per pocket) was injected locally, with contralateral pockets receiving PBS. Outcomes were assessed by histology, immunohistochemistry, RT-qPCR, flow cytometry, and Transwell assays of macrophage-conditioned media on bone marrow mesenchymal stem cell (BMSC) migration and osteogenic genes. Results i.p. LipClod enhanced ectopic bone formation at 8 weeks, supported by histology and increased bone sialoprotein transcripts. Rather than depleting macrophages, it expanded monocyte–macrophage at the implant site and increased BMSC recruitment. Macrophage-conditioned media promoted BMSC migration in vitro. Local M-CSF further boosted macrophage infiltration, favored reparative polarization, and augmented osteogenesis. Conclusions Systemic LipClod augments MIOI by eliciting compensatory expansion of local macrophages that enhances early BMSC trafficking. Carefully calibrated systemic macrophage-targeted interventions may improve osteoinductive biomaterial performance.