BACKGROUND: Exercise therapy has been recognized as first line therapy of osteoarthritis (OA). The exercise related exosome involved in the interaction between fibroblast-like synoviocytes (FLSs) and chondrocytes could be a novel nanoparticle strategy for treating OA. METHODS: Single-cell transcriptome sequencing was used to investigate the exercise therapy-related gene. Cells were transfected by recombinant adenovirus carrying knockdown or overexpression sequences. Quantitative proteomics of FLS-derived exosomes via data-independent acquisition was used to investigate exosomal cargo. RESULTS: In this study, PPARγ was upregulated in FLSs under exercise stimulation. Our data revealed that overexpression (OE) PPARγ FLSs derived exosome could ameliorate the OA severity and activate autophagy, inhibit chondrocyte apoptosis, and reduced cartilage degeneration. But knockdown (KD) PPARγ FLSs derived exosome aggravate the OA. Moreover, we found PPARγ controls the endosomal sorting complex required for the transport (ESCRT)-dependent pathway to activate exosome biogenesis. Annexin A1 (ANXA1) was enriched in OE- PPARγ exosome. As a therapeutic cargo, FLSs exosomal ANXA1 was confirmed be internalization by chondrocyte via exosome labeled experiment and ANXA1 could inhibit the phosphorylation of ERK to activate the autophagy and decrease chondrocyte apoptosis. CONCLUSIONS: PPARγ controls ESCRT-dependent FLSs exosome biogenesis and alleviates chondrocyte osteoarthritis mediated by exosomal cargo ANXA1 protein. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: PPARγ/ESCRT - FLSs exosomal ANXA1 - ERK axis provides a deeper theoretical basis for exercise therapy of OA and a new idea for the clinical transformation of nanoparticles into OA therapy.