Abstract: Objective To investigate the therapeutic efficacy of adoptively transferred sEV in mice with collagen-induced arthritis (CIA), we isolated small extracellular vesicles (sEV) from the culture supernatant of human peripheral blood-derived induced CD8⁺Treg cells. Methods sEV were isolated via differential ultracentrifugation and characterized by nanoparticle tracking analysis, transmission electron microscopy and western blotting. CIA mice were randomly assigned into control and sEV-treated groups. After intervention, clinical arthritis scoring and histopathological examination of joint tissues were examined. Plasma levels of proinflammatory cytokines and anti-type II collagen antibodies were detected, and CD4⁺ T cell subsets in lymph nodes were quantified by flow cytometry. Results Purified sEV exhibited a typical cup-shaped morphology with an average diameter of 106.8±3.3 nm and expression of CD63 and TSG101. Compared with controls, the sEV-treated group displayed delayed disease onset, alleviated articular inflammation and reduced cartilage erosion. Consistently, plasma TNF-α, IL-6 and anti-type Ⅱ collagen antibodies were markedly reduced, while the frequencies of lymph node CD4⁺IFN-γ⁺ and CD4⁺IL-4⁺ T cells were elevated. Conclusion CD8⁺Treg-derived sEV alleviate joint inflammation in CIA mice by suppressing inflammatory cytokines and modulating CD4⁺ T cell differentiation.

Key words: CD4⁺ cell subsets, Inflammatory cytokine, CD8⁺ regulatory T cells, Extracellular vesicles, Collagen-induced arthritis