Cytokine preconditioning enhances the therapeutic potential of MSCs in ARDS

<bold>Rationale:</bold> Mesenchymal stem cell (MSC) therapy for acute respiratory distress syndrome (ARDS) has shown safety in clinical trials but efficacy remains unproven. Thus, MSC preconditioning has been proposed as a strategy to increase MSC efficacy. This project aims to explore the therapeutic enhancement of MSCs through preconditioning with a cytokine cocktail. <bold>Methods:</bold> Bone marrow-derived MSCs (BM-MSCs) were preconditioned with a cytokine cocktail (IL-1β, TNF-α and IFN-γ, 50ng/mL, “Cytomix”). Apoptosis and activation of the BM-MSCs were analysed by flow cytometry. Conditioned media (CM) was collected to quantify cytokine secretion by ELISA and to analyse the inhibitory effect of the MSC secretome on bacterial proliferation. <bold>Results:</bold> Compared to naïve BM-MSCs, the early and late apoptosis and necrosis induced by serum-free media decreased significantly in the Cytomix preconditioned group. The levels of surface markers CD54 and CD274 were significantly higher in Cytomix preconditioned BM-MSCs. In Cytomix preconditioned CM, higher levels of IL-6, IL-8 and MCP-1 were detected. Moreover, Cytomix preconditioning enhanced BM-MSC inhibitory effects on <italic>Klebsiella pneumoniae</italic> and <italic>Staphylococcus aureus</italic>. <bold>Conclusion:</bold> Cytomix preconditioning can enhance BM-MSC function, inducing resistance to starvation-induced apoptosis and necrosis, and regulates the phenotype of BM-MSCs to increase immunomodulatory properties. Besides this, cytomix preconditioning can augment the anti-microbial effects of BM-MSCs, which makes these preconditioned BM-MSCs a promising treatment for bacterial-induced ARDS.