LPS-neutralizing peptides reduce outer membrane vesicle-induced inflammatory responses

Biochim Biophys Acta Mol Cell Biol Lipids. 2019 Oct;1864(10):1503-1513. doi: 10.1016/j.bbalip.2019.05.018. Epub 2019 Jun 1.

Abstract

Outer membrane vesicles (OMVs) are secreted by Gram-negative bacteria and induce a stronger inflammatory response than pure LPS. After endocytosis of OMVs by macrophages, lipopolysaccharide (LPS) is released from early endosomes to activate its intracellular receptors followed by non-canonical inflammasome activation and pyroptosis, which are critically involved in sepsis development. Previously, we could show that the synthetic anti-endotoxin peptide Pep19-2.5 neutralizes inflammatory responses induced by intracellular LPS. Here, we aimed to investigate whether Pep19-2.5 is able to suppress cytoplasmic LPS-induced inflammation under more physiological conditions by using OMVs which naturally transfer LPS to the cytosol. Isothermal titration calorimetry revealed an exothermic reaction between Pep19-2.5 and Escherichia coli OMVs and the Limulus Amebocyte Lysate assay indicated a strong endotoxin blocking activity. In THP-1 macrophages and primary human macrophages Pep19-2.5 and polymyxin B reduced interleukin (IL)-1β and tumor necrosis factor (TNF) release as well as pyroptosis induced by OMVs, while the Toll-like receptor 4 signaling inhibitor TAK-242 suppressed OMV-induced TNF and IL-1β secretion, but not pyroptosis. Internalization of Pep19-2.5 was at least partially mediated by the P2X7 receptor in macrophages but not in monocytes. Additionally, a cell-dependent difference in the neutralization efficiency of Pep19-2.5 became evident in macrophages and monocytes, indicating a critical role for peptide-mediated IL-1β secretion via the P2X7 receptor. In conclusion, we provide evidence that LPS-neutralizing peptides inhibit OMV-induced activation of the inflammasome/IL-1 axis and give new insights into the mechanism of peptide-mediated neutralization of cytoplasmic LPS suggesting an essential and cell-type specific role for the P2X7 receptor.

Keywords: Cytoplasmic lipopolysaccharide; Interleukin-1beta; Outer membrane vesicles; P2X7 receptor; Pyroptosis; Synthetic anti-endotoxin peptides.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Inflammatory Agents / pharmacology*
  • Bacterial Outer Membrane / drug effects*
  • Bacterial Outer Membrane / immunology
  • Cell Line
  • Escherichia coli / drug effects*
  • Escherichia coli / immunology
  • Escherichia coli Infections / immunology
  • Escherichia coli Infections / microbiology
  • Humans
  • Inflammation / drug therapy
  • Inflammation / immunology
  • Inflammation / microbiology
  • Lipopolysaccharides / antagonists & inhibitors*
  • Lipopolysaccharides / immunology
  • Macrophages / drug effects*
  • Macrophages / immunology
  • Peptides / pharmacology*
  • Pyroptosis / drug effects

Substances

  • Anti-Inflammatory Agents
  • Lipopolysaccharides
  • Peptides