Mesenchymal Stem Cell-Derived Extracellular Vesicles Mitigate Immune Cell Activation in an In Vitro Model of Post-Resuscitation Inflammation

ABSTRACT Background Systemic inflammation is a well-established component of post-cardiac arrest syndrome (PCAS), a condition responsible for significant morbidity and mortality in patients who are initially resuscitated from sudden cardiac arrest. Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) have emerged as promising immunomodulatory agents in various inflammatory conditions, including after ischemia-reperfusion injury (IRI). Here, we investigated the therapeutic potential of MSC-EVs in porcine peripheral blood mononuclear cells (PBMCs) stimulated with lipopolysaccharide (LPS) or mitochondrial DNA (mtDNA) to mimic immune cell activation in PCAS. Methods PBMCs were isolated from healthy pigs ( Sus scrofa ), cultured in vitro , stimulated with LPS or mtDNA, and treated with a range of MSC-EV concentrations. Flow cytometry, quantitative PCR, ELISA, and ROS/RNS measurements were performed to assess PBMC activation. Results MSC-EV treatment reduced LPS-induced inflammatory granulocyte activation and selectively modulated cytokine transcripts, including IFNα, IL-1β, and TNF-α, in a concentration-dependent manner. Similar immunosuppressive effects were observed in mtDNA-stimulated PBMCs, where MSC-EVs attenuated dendritic cell activation and inflammatory cytokine release. Furthermore, higher concentrations of MSC-EVs significantly decreased ROS/RNS production in both LPS- and mtDNA-challenged PBMCs. Conclusions MSC-EVs exhibit potent immunomodulatory properties against LPS- and mtDNA-induced activation of porcine PBMCs, highlighting their broad capacity to modulate immune responses and mitigate oxidative stress induced by pro-inflammatory stimuli that are relevant to PCAS. These findings provide further support for the administration of MSCs, or MSC-EVs themselves, as a potential therapeutic intervention to target immune activation in PCAS and other disorders characterized by an acute systemic inflammatory state.