Lipopolysaccharide-Neutralizing Peptide Modulates P2X7 Receptor-Mediated Interleukin-1β Release

Lipopolysaccharide (LPS)-neutralizing peptides are emerging as new potential therapeutic modalities to treat sepsis and skin infections. Purinergic ligand-gated ion channels (P2X receptors) play a critical role in various biological processes, including inflammation. Recent drug development efforts have significantly focused on the modulation of P2X receptors. Here, we investigated the effects of the synthetic LPS-neutralizing peptide Pep19-2.5 on human P2X receptors in cells of the innate immune system. Pep19-2.5 concentration-dependently triggered Ca²⁺ influx, interleukin (IL)-1β, and lactate dehydrogenase (LDH) release in Toll-like receptor-stimulated human macrophages and monocytes. Ca²⁺ influx was mediated at least partially by P2X7 receptors, and IL-1β and LDH release by P2X7 receptors, respectively. Confocal microscopy confirmed the colocalization of Pep19-2.5 with P2X7 receptors. Pep19-2.5-induced IL-1β release in primed cells was dependent on K⁺ efflux, caspase-1, and the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing protein 3 inflammasome. In the presence of the P2X7 receptor agonist 2'(3')-O-(4-benzoylbenzoyl)adenosine-5'-triphosphate, Pep19-2.5 reduced IL-1β and LDH release. In 1321N1, astrocytoma cells stably transfected with human P2X receptors, Pep19-2.5 potently modulated P2X7 and P2X4 receptors (IC₅₀ values of 0.346 and 0.146 μM, respectively) but showed less (P2X1, P2X3) or no activity (P2X2) at other P2X receptor subtypes. Our findings underline the potential of LPS-neutralizing peptides as modulators of P2X receptors, thus expanding their applicability beyond the treatment of sepsis to the treatment of inflammatory diseases.