Background/Purpose: Gout is the most prevalent form of inflammatory arthritis, with rising global incidence. It results from the deposition of monosodium urate (MSU) crystals in joints, leading to severe inflammation and pain in individuals with hyperuricemia. Dental pulp stem cells (DPSCs), a type of mesenchymal stem cell (MSC), release extracellular vesicles (EVs) with intrinsic anti-inflammatory properties, which are further enhanced following inflammatory stimulation. This study aimed to evaluate the anti-inflammatory effects of EVs from both untreated (naïve) and MSU-preconditioned DPSCs on MSU-activated macrophages and to identify EV-associated microRNAs (miRNAs) mediating these effects.

Methods: DPSCs were obtained from Lonza. EVs were isolated from DPSCs cultured under normal conditions or after preconditioning with MSU, and characterized by immunoblotting and nanoparticle tracking analysis (NTA). The anti-inflammatory activity of EVs was assessed in vitro using human peripheral blood mononuclear cells (PBMCs) stimulated with 100 µg/mL MSU for 24 hours, with or without treatment with naïve or MSU-preconditioned DPSC EVs. Inflammatory gene and protein expression were analyzed by qRT-PCR and ELISA. Phagocytic activity was measured using the Vybrant™ Phagocytosis Assay Kit. miRNA profiles of naïve and preconditioned EVs were determined by sequencing and bioinformatics analysis. Statistical significance was assessed using one-way ANOVA followed by Tukey’s post hoc test (P< 0.05).

Results: EVs from both naïve and MSU-preconditioned DPSCs displayed similar physical and molecular characteristics. Both EV types significantly reduced inflammatory responses in MSU-stimulated PBMCs. Notably, MSU-preconditioned DPSC EVs showed enhanced suppression of pro-inflammatory markers (IL-1β, TNF-α, iNOS) and increased expression of anti-inflammatory markers (IL-10, ARG1, TGF-β). Phagocytosis was also significantly reduced by preconditioned EVs. miRNA sequencing revealed upregulation of anti-inflammatory miRNAs in EVs from MSU-preconditioned DPSCs compared to naïve controls.

Conclusion: MSU-preconditioned DPSC EVs exhibit enhanced anti-inflammatory activity in MSU-activated PBMCs, including suppression of phagocytosis. These effects are likely mediated by specific EV-resident miRNAs, offering a promising platform for miRNA-based EV engineering in the treatment of inflammatory diseases like gout.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/modulation-of-inflammatory-responses-by-dental-pulp-stem-cell-extracellular-vesicles-in-monosodium-urate-stimulated-macrophages/