Extracellular vesicles derived from mesenchymal stem cells attenuate the immune response in Type 1 Diabetes by inducing regulatory dendritic cells
Introduction: Type 1 Diabetes (T1D) is a complex autoimmune disease characterized by T-cell driven destruction of pancreatic β-cells in the islets of Langerhans and resultant hyperglycemia1. The pathogenesis of T1D involves presentation of normally cryptic β-cell autoantigens and aberrant activation rather than suppression of immune cells1,2. The only treatment for T1D is exogenous insulin therapy, which does not target the etiology of the disease1. Extracellular vesicles derived from mesenchymal stem cells (MSC-EVs) hold therapeutic potential as they retain the immunomodulatory properties of MSCs while having the advantage of being cell-free and more predictable3,4. The mechanism by which MSC-EVs suppress T1D was investigated. Methods: MSCs and peripheral blood mononuclear cells (PBMCs) were obtained from healthy donors5. EVs were obtained from MSCs via ultracentrifugation5,6. Dendritic cells (DCs) were obtained from PBMCs via immunomagnetic isolation and matured by exposure to lipopolysaccharide (LPS) in the presence or absence of MSC-EVs and DC co-stimulatory markers were quantified via flow cytometry5. Supernatants from co-cultures of MSC-EVs with mature DCs (mDCs), as well as T cells with mDCs or mDCs + MSC-EVs, were analyzed via ELISA to measure cytokine concentration5. A one-way mixed lymphocyte reaction assay was performed using splenocytes from C57BL/6 and BALB/c mice, in the presence or absence of EVs, and flow cytometry was used to quantify DC co-stimulatory markers6. Splenocytes vs splenocytes + MSC-EVs were transfused from NOD to NOD/scid mice and the onset to T1D (two consecutive >250 mg/dL blood glucose tests) was measured6. Results: mDCs co-cultured with MSC-EVs showed significant reduction in co-stimulatory molecules CD80, CD83, and CD38 compared to culturing without MSC-EVs. Co-culture of mixed splenocytes with EVs showed dose-dependent reduction of co-stimulatory molecules CD80, CD40, CD86, and MHC II compared to co-culturing without MSC-EVs5. Supernatants derived from co-cultures of mDCs with MSC-EVs compared to without MSC-EVs showed reduced IL-6 and IL-12 and increased TGF-β6. Supernatants derived from co-cultures of T cells with MSC-EV-conditioned mDCs compared to non-conditioned mDCs showed reduced IFN-γ, IL-65. Infusion of NOD-derived splenocytes with, vs without NOD-derived MSC-EVs, into NOD/scid mice significantly delayed the onset of T1D6. Conclusions: MSC-EVs suppress the immune response in T1D by inducing a regulatory phenotype in DCs, evidenced by the reduction of co-stimulatory molecules and alterations in their secreted cytokine profiles. They have been shown to delay T1D onset in vivo. MSC-EVs are therefore a potential therapeutic agent for T1D.
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