Cytoplast^CXCR4: An enucleated self-eliminating and mitochondria delivering strategy for targeted cartilage defect therapy.

Abstract

Joint cartilage regeneration presents a significant challenge due to its limited self-repair capacity, hindered by local inflammation and metabolic dysregulation. Mitochondrial quality control (MQC) is crucial for maintaining chondrocyte differentiation and metabolism; however, strategies targeting MQC remain underexplored. Here, we generated an acellular delivery vehicle, cytoplast^CXCR4, which was designed to facilitate mitochondria transport and restore MQC, from enucleated CXCR4 transfected mesenchymal stem cells (MSCs) through ultracentrifugation. CXCR4 was overexpressed to enhance migration toward inflamed cartilage via its interaction with SDF-1α, which is a chemokine elevated at the site of cartilage damage. The CXCR4 gene-transfected, nuclear-depleted cytoplast^CXCR4 exhibited enhanced homing ability, via CXCR4-SDF-1α axis. In vitro results showed that cytoplast^CXCR4 treatment improved mitochondrial quality-structure, oxygen consumption, and ATP synthesis-in osteoarthritic chondrocytes, thus re-establishing metabolic balance. Compared to exosomes, cytoplasts retain cellular bioactivity, including functional organelles and paracrine signaling. Notably, we observed mitochondrial transfer via cytoplast-derived vesicles, which contributed to the restoration of morphology and function in damaged mitochondria within injured chondrocytes. Furthermore, in vivo results demonstrated that cytoplasts^CXCR4 had a longer retention time, and could precisely target the damaged cartilage and significantly facilitate cartilage repair compared with exosomes in a rat knee cartilage defect model. This study presents an MQC-targeted and mitochondria-preserving approach, offering an improved strategy for joint cartilage regeneration.