<p>Inflammation critically influences mesenchymal stem cell (MSC) behavior, affecting their survival, plasticity, and immunomodulatory functions. MSC preconditioning has emerged as a promising strategy to enhance their therapeutic potential before clinical application. Among the factors capable of modulating MSC responses, selenium has gained attention due to its role in regulating inflammatory signaling. In this study, we investigated whether high-dose sodium selenite (Na₂SeO₃) preconditioning modulates MSC behavior under inflammatory stimulation and how these changes affect macrophage responses through paracrine mechanisms. MSCs were preconditioned with Na₂SeO₃ in the presence or absence of lipopolysaccharide (LPS). Cellular metabolism, apoptosis, cell-cycle progression, migration, intracellular signaling, reactive oxygen species (ROS) production, and soluble mediator release were evaluated. Macrophage responses were assessed using conditioned medium from treated MSCs. Na₂SeO₃ alone did not significantly affect MSC metabolic viability. However, under inflammatory conditions, Na₂SeO₃ enhanced apoptotic signaling and promoted cell-cycle accumulation in S/G2/M phases. In addition, Na₂SeO₃ preconditioning reduced intracellular ROS levels, attenuated NFκB and JNK activation, and altered the MSC secretory profile, increasing nitric oxide, transforming growth factor-β, and prostaglandin E2 production while decreasing IL-6 levels. Importantly, conditioned medium from preconditioned MSCs reprogrammed macrophage responses, reducing TNF-α and IL-1β production while increasing IL-1 receptor antagonist levels. These findings demonstrate that high-dose Na₂SeO₃ reprograms MSCs under inflammatory stress, enhancing their anti-inflammatory paracrine activity and capacity to modulate macrophage inflammatory responses. These results identify selenium as a key regulator of MSC-mediated immunomodulation and support sodium selenite preconditioning as a strategy to optimize MSC-based therapies for inflammatory diseases.</p>