Abstract

Senescent BMSCs-derived exosome NR4A2 inhibits osteogenic differentiation of ST2 cells. • NR4A2 inhibits osteogenic differentiation of ST2 cells by transcriptionally inhibiting NELL2. • NELL2 promotes osteogenic differentiation of ST2 cells by promoting the binding of Fn1 to integrin β1. As a transcription factor, nuclear receptor subfamily 4 group A member 2 (NR4A2) regulates target gene transcription by directly binding to response elements within gene promoters. However, the exact role and molecular mechanism of NR4A2 in fracture healing have not been fully elucidated. Here, we identified NR4A2 as a key functional molecular cargo enriched in exosomes secreted by aged bone marrow stromal cells (BMSCs). Upon uptake by recipient cells, exosomal NR4A2 functioned as a transcriptional repressor that specifically bound to the promoter region of the Neural EGFL-like 2 (NELL2) gene, thereby suppressing its transcription. We further demonstrated that NELL2 acted as a critical adaptor protein that enhanced the binding affinity between fibronectin 1 (Fn1) and integrin β1, thereby licensing FAK/AKT cascade activation essential for osteogenic differentiation. Mechanistically, NR4A2-mediated suppression of NELL2 impaired the assembly of the NELL2/Fn1/integrin β1 complex and attenuated FAK/AKT phosphorylation, thereby inhibiting osteogenesis and culminating in delayed fracture healing. Collectively, our findings delineate a novel exosome-dependent regulatory axis, through which aged BMSCs deteriorated the bone repair microenvironment via the NR4A2/NELL2/FAK-AKT signaling cascade. This study reveals a previously unrecognized molecular mechanism underlying age-associated fracture healing impairment.