Extracellular Vesicles Derived from H 2 O 2-Stimulated Adipose-Derived Stem Cells Alleviate Senescence in Diabetic Bone Marrow Mesenchymal Stem Cells and Restore Their Osteogenic Capacity

Introduction: Autologous stem cell transplantation has emerged as a promising strategy for bone repair. However, the osteogenic potential of mesenchymal stem cells derived from diabetic patients is compromised, possibly due to hyperglycemia -induced senescence. The objective of this study was to assess the preconditioning effects of extracellular vesicles derived from H 2 O 2 - stimulated adipose -derived stem cells (ADSCs) and non -modified ADSCs on the osteogenic potential of diabetic bone marrow mesenchymal stem cells (BMSCs). Methods: Sprague-Dawley (SD) rats were experimentally induced into a diabetic state through a high -fat diet followed by an injection of streptozotocin, and diabetic BMSCs were collected from the bone marrow of these rats. Extracellular vesicles (EVs) were isolated from the conditioned media of ADSCs, with or without hydrogen peroxide (H 2 O 2 ) preconditioning, using density gradient centrifugation. The effects of H 2 O 2 preconditioning on the morphology, marker expression, and particle size of the EVs were analyzed. Furthermore, the impact of EV-pretreatment on the viability, survivability, migration ability, osteogenesis, cellular senescence, and oxidative stress of diabetic BMSCs was examined. Moreover, the expression of the Nrf2/HO-1 pathway was also assessed to explore the underlying mechanism. Additionally, we transplanted EV-pretreated BMSCs into calvarial defects in diabetic rats to assess their in vivo bone formation and anti -senescence capabilities. Results: Our study demonstrated that pretreatment with EVs from ADSCs significantly improved the viability, senescence, and osteogenic differentiation potential of diabetic BMSCs. Moreover, in -vitro experiments revealed that diabetic BMSCs treated with H 2 O 2 -activated EVs exhibited increased viability, reduced senescence, and enhanced osteogenic differentiation compared to those treated with non -modified EVs. Furthermore, when transplanted into rat bone defects, diabetic BMSCs treated with H 2 O 2 -activated EVs showed improved bone regeneration potential and enhanced anti -senescence function t compared to those treated with non -modified EVs. Both H 2 O 2 -activated EVs and non -modified EVs upregulated the expression of the Nrf2/HO-1 pathway in diabetic BMSCs, however, the promoting effect of H 2 O 2 -activated EVs was more pronounced than that of nonmodified EVs. Conclusion: Extracellular vesicles derived from H 2 O 2 -preconditioned ADSCs mitigated senescence in diabetic BMSCs and enhanced their bone regenerative functions via the activation of the Nrf2/HO-1 pathway.