Apoptotic vesicles derived from human adipose-derived mesenchymal stem cells facilitate the transition of netosis to neutrophil apoptosis

Background　Mesenchymal stem cell-derived apoptotic vesicles (MSCs-apoVs) have demonstrated the capacity to promote apoptosis in polymorphonuclear neutrophils (PMNs) and reduce their neutrophilic inflammatory necrosis (NETosis). Consequently, MSCs-apoVs have emerged as a novel therapeutic strategy for inhibiting NET formation by suppressing NETosis in PMNs and enhancing their apoptotic processes.Objective　This study aimed to investigate the effects of adipose-derived stem cell derived apoptotic vesicles (ADSCs-apoVs) on promoting PMNs apoptosis and inhibiting NETosis.Methods　Primary mouse bone marrow-derived polymorphonuclear neutrophils (PMNs) were isolated via differential centrifugation combined with sterile fluorescence-activated cell sorting (FACS). PMNs were co-cultured with adipose-derived stem cell apoptotic vesicles (ADSCs apoVs) at varying concentrations (1, 5, and 10 μg/mL) for 8 hours. NETs formation, NETosis, and apoptosis were quantified using immunofluorescence and flow cytometry. Results　ADSCs-apoVs regulated PMNs fate through dual pathways. NETosis suppression showed that at 5 μg/mL, significant reductions occurred in H3cit production (48.20 ± 0.96 vs 75.30 ± 1.00, P < 0.001), NETosis incidence (36.07% ± 3.07% vs control 57.03% ± 5.35%, P=0.002); At 10 μg/mL, synergistic inhibition was observed for H3cit levels (24.73 ± 0.90 vs 75.30 ± 1.00, P < 0.001),NETosis incidence (16.48% ± 8.09% vs 57.03% ± 5.35%, P < 0.001). Apoptosis threshold effect showed that 10 μg/mL elevated total apoptosis (50.70% ± 10.86% vs 10.30% ± 1.55%, P=0.001),early apoptosis (14.30% ± 1.97% vs 0.25% ± 0.21%, P 0.999), 1μg/mL (9.14% ± 4.82 vs 10.30% ± 1.55, P=0.997).Conclusion　ADSCs-apoVs regulate PMNs fate through dual pathways by inhibiting histone H3 citrullination to suppress NETs formation, thereby preventing NETosis, and concurrently activating apoptosis pathways to induce extensive PMNs apoptosis. 5 μg/mL ADSCs-apoVs defines a therapeutic window for NETosis-selective inhibition, while 10 μg/mL represents an effective clearance dose for synergistic NETosis suppression and apoptosis activation.