Goat Milk-derived Extracellular Vesicles modulate glial responses in vitro and attenuate neuroinflammation in vivo

Abstract

Milk-derived extracellular vesicles (MEV) represent a naturally derived, potentially scalable platform for immunomodulation, yet their effects on central nervous system (CNS) glial inflammation at defined particle-to-cell ratios remain insufficiently characterized. Here, we investigated the immunomodulatory effects of goat milk-derived MEV in cellular and murine models of neuroinflammation. MEV-enriched preparations were obtained by sequential filtration, ultrafiltration, and size-exclusion chromatography, and characterized by scanning electron microscopy, nanoparticle tracking analysis, and dynamic light scattering. In LPS-stimulated BV2 microglia-like cells, MEV at 250 and 2500 particles per cell significantly reduced CD40 expression. In primary rat mixed glial cultures, microglia, astrocytes, and oligodendrocytes internalized MEV. At 250 particles per cell, MEV significantly reduced CD40 and IL1{beta} gene expression at the mixed-culture level. In both cellular models, MEV also increased markers associated with an M2-like microglial phenotype under resting conditions. Following intrathecal administration in healthy mice, MEV did not induce significant changes in the inflammatory gene markers assessed at 8 h. In an LPS-induced mouse model of neuroinflammation, intrathecal MEV administration reduced CD40 protein expression towards control levels within 24 h and shifted the inflammatory marker profile towards a pro-resolutive state, as indicated by increased Arg1 levels and elevated CD206/CD86 and IL10/IL6 ratios. These findings identify goat milk-derived MEV-enriched preparations as potential modulators of CNS glial inflammation and support further investigation of their therapeutic potential.