A Soluble PrPC Derivative and Membrane-Anchored PrPC in Extracellular Vesicles Attenuate Innate Immunity by Engaging the NMDA-R/LRP1 Receptor Complex

: Nonpathogenic cellular prion protein (PrPC) demonstrates anti-inflammatory activity; however, the responsible mechanisms are incompletely defined. PrPC exists as a GPI-anchored membrane protein in diverse cells; however, PrPC may be released from cells by ADAM proteases or when packaged into extracellular vesicles (EVs). In this study, we show that a soluble derivative of PrPC (S-PrP) counteracts inflammatory responses triggered by pattern recognition receptors in macrophages, including TLR2, TLR4, TLR7, TLR9, NOD1, and NOD2. S-PrP also significantly attenuates the toxicity of LPS in mice. The response of macrophages to S-PrP is mediated by a receptor assembly that includes the N-methyl-d-aspartate receptor (NMDA-R) and low-density lipoprotein receptor-related protein-1 (LRP1). PrPC was identified in EVs isolated from human plasma. These EVs replicated the activity of S-PrP, inhibiting cytokine expression and IκBα phosphorylation in LPS-treated macrophages. The effects of plasma EVs on LPS-treated macrophages were blocked by PrPC-specific Ab, by antagonists of LRP1 and the NMDA-R, by deleting Lrp1 in macrophages, and by inhibiting Src family kinases. Phosphatidylinositol-specific phospholipase C dissociated the LPS-regulatory activity from EVs, rendering the EVs inactive as LPS inhibitors. The LPS-regulatory activity that was lost from phosphatidylinositol-specific phospholipase C-treated EVs was recovered in solution. Collectively, these results demonstrate that GPI-anchored PrPC is the essential EV component required for the observed immune regulatory activity of human plasma EVs. S-PrP and EV-associated PrPC regulate innate immunity by engaging the NMDA-R/LRP1 receptor system in macrophages. The scope of pattern recognition receptors antagonized by S-PrP suggests that released forms of PrPC may have broad anti-inflammatory activity.