A Sustained-Release AMD3100-Hyaluronic Acid Complex Ameliorates Osteoarthritis by Promoting FHL1-Mediated Negative Feedback on SDF-1 Signaling

Background Osteoarthritis (OA) is a prevalent degenerative joint disease with limited therapeutic options. The Stromal Cell-Derived Factor-1 (SDF-1)/CXCR4 signaling pathway is a key driver of OA catabolism and inflammation. However, its downstream regulatory mechanisms are incompletely understood, and strategies for sustained intra-articular drug delivery are needed to effectively target this pathway. Methods We developed and comprehensively characterized a sustained-release hydrogel complex of the CXCR4 antagonist AMD3100 and hyaluronic acid (HA-AMD3100). Its therapeutic efficacy was evaluated in a surgically-induced rabbit OA model. To investigate the underlying molecular mechanisms, we employed RNA-sequencing, gene knockdown, co-immunoprecipitation (Co-IP), site-directed mutagenesis, and molecular docking in SDF-1-stimulated chondrocytes. Results The HA-AMD3100 hydrogel exhibited excellent biocompatibility, biodegradability, and a three-day sustained-release profile. In vivo, intra-articular injection of the hydrogel significantly attenuated cartilage erosion, proteoglycan loss, and suppressed inflammatory cytokine expression, demonstrating superior chondroprotective effects compared to HA alone. Mechanistically, RNA-sequencing identified Four and a half LIM domains 1 (FHL1) as a key protective factor downregulated in OA chondrocytes. FHL1 knockdown exacerbated chondrocyte apoptosis, inflammation, and matrix degradation. Critically, we discovered for the first time that FHL1 acts as a negative feedback regulator of SDF-1 signaling by directly interacting with SDF-1. This physical interaction, mapped to the GLU241 residue of FHL1, was essential for suppressing the downstream pro-inflammatory AKT and ERK signaling pathways. Conclusion Our findings demonstrate that the sustained-release HA-AMD3100 hydrogel effectively alleviates OA progression. We unveil a novel chondroprotective mechanism where the therapeutic effect is mediated by enhancing a newly identified FHL1-mediated negative feedback loop on SDF-1 signaling. This FHL1/SDF-1 interaction is crucial for restraining catabolic and inflammatory responses in chondrocytes, establishing FHL1 as a promising therapeutic target for OA.