Distinct osteoclast transcriptional states and heterotypic fusion arising from circulating pre-dendritic cells and monocytes.

Osteoclasts (OCs) are multinucleated bone-resorbing cells essential for bone remodelling and contribute to pathological bone loss in inflammatory and metabolic bone diseases. Although classically considered to arise from monocyte/macrophage precursors, circulating pre-dendritic cells (pre-DCs) also retain osteoclastogenic potential. However, whether OCs derived from these distinct precursors retain different transcriptional states, and how they interact during fusion, remain unclear. Here, we compared OCs generated from directly isolated circulating monocytes and pre-DCs and investigated how these populations contribute to OC formation and fusion within mixed cultures. RNA sequencing identified 41 genes significantly enriched in pre-DC-derived OCs and 9 in monocyte-derived OCs, while core osteoclastogenic and fusion-associated genes showed no significant differential expression. Gene set enrichment analysis (GSEA) identified distinct precursor-associated transcriptional programmes. Pre-DC-derived OCs were enriched for extracellular matrix organisation, skeletal development, canonical Wnt signalling and immune-regulatory pathways, whereas monocyte-derived OCs were enriched for cell-cycle, proteasomal, lysosomal and innate immune pathways. In mixed cultures, OC output increased with the proportion of pre-DCs, with the greatest OC formation observed in pre-DC-only cultures. Live-cell imaging demonstrated heterotypic fusion, with monocyte-derived precursors observed fusing with established pre-DC-derived multinucleated OCs, whereas exclusively monocyte-derived multinucleated OCs were infrequently observed. These findings indicate that both populations generate OCs with a conserved core transcriptional profile but distinct precursor-associated transcriptional states. Greater pre-DC abundance was associated with increased OC output, and live-cell imaging showed that monocyte-derived cells can be incorporated into pre-DC-derived multinucleated OCs. This heterogeneity may influence OC abundance, multinucleation, and functional properties in inflammatory and metabolic bone diseases.