A two phase regulation of bone regeneration: IL-17F mediates osteoblastogenesis via C/EBP-β in vitro

T lymphocytes and pro-inflammatory cytokines, specifically interleukin-17F (IL-17F) have been identified as important regulators in bone regeneration during fracture repair. To better understand the molecular mechanisms of IL-17F-mediated osteoblastogenesis, a mouse pre-osteoblast cell line (MC3T3-E1) was utilized to characterize the intracellular signal transduction of IL-17F. Comparisons to the established canonical Wnt signaling pathway were made using Wnt3a ligand. Our results demonstrated greater bone marker gene expression in IL-17F-treated cells, compared to cells treated with Wnt3a. Western blot analysis confirmed degradation of β-catenin and up-regulation of two key proteins in osteoblast differentiation, Runx2 and C/EBP-β, in response to IL-17F treatment. RNA silencing of IL-17F receptors, IL-17Ra and IL-17Rc via siRNA transfection resulted in decreased expression of Act2, Runx2, and C/EBP-β, demonstrating the direct ligand-receptor interaction between IL-17F and IL-17Ra/c as an activator of osteoblastogenesis. Our findings suggest that IL-17F promotes osteoblast differentiation independent of the canonical Wnt pathway and β-catenin signaling, presenting new insights on modulating the adaptive immune response in the inflammatory phase, temporally distinct from the reparative and remodeling phases of fracture healing.This work was funded by Canadian Institutes of Health Research (CIHR) Institute of Musculoskeletal Health and Arthritis (IMHA) and Academic Enrichment Fund at Sunnybrook Health Science Centre, Division of Orthopaedic Surgery