In vivo imaging reveals PKA regulation of ERK activity during neutrophil recruitment to inflamed intestines

J Exp Med. 2014 Jun 2;211(6):1123-36. doi: 10.1084/jem.20132112. Epub 2014 May 19.

Abstract

Many chemical mediators regulate neutrophil recruitment to inflammatory sites. Although the actions of each chemical mediator have been demonstrated with neutrophils in vitro, how such chemical mediators act cooperatively or counteractively in vivo remains largely unknown. Here, by in vivo two-photon excitation microscopy with transgenic mice expressing biosensors based on Förster resonance energy transfer, we time-lapse-imaged the activities of extracellular signal-regulated kinase (ERK) and protein kinase A (PKA) in neutrophils in inflamed intestinal tissue. ERK activity in neutrophils rapidly increased during spreading on the endothelial cells and showed positive correlation with the migration velocity on endothelial cells or in interstitial tissue. Meanwhile, in the neutrophils migrating in the interstitial tissue, high PKA activity correlated negatively with migration velocity. In contradiction to previous in vitro studies that showed ERK activation by prostaglandin E2 (PGE2) engagement with prostaglandin receptor EP4, intravenous administration of EP4 agonist activated PKA, inhibited ERK, and suppressed migration of neutrophils. The opposite results were obtained using nonsteroidal antiinflammatory drugs (NSAIDs). Therefore, NSAID-induced enteritis may be caused at least partially by the inhibition of EP4 receptor signaling of neutrophils. Our results demonstrate that ERK positively regulates the neutrophil recruitment cascade by promoting adhesion and migration steps.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Anti-Inflammatory Agents, Non-Steroidal / pharmacology
  • Benzamides / pharmacology
  • Cell Adhesion / drug effects
  • Cell Movement / drug effects
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • Diphenylamine / analogs & derivatives
  • Diphenylamine / pharmacology
  • Endothelial Cells / metabolism
  • Enteritis / metabolism*
  • Extracellular Signal-Regulated MAP Kinases / antagonists & inhibitors
  • Extracellular Signal-Regulated MAP Kinases / metabolism*
  • Female
  • Fluorescence Resonance Energy Transfer
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Intestine, Small / metabolism
  • Intestine, Small / pathology
  • Male
  • Methyl Ethers / pharmacology
  • Mice
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Microscopy, Fluorescence, Multiphoton
  • Naphthalenes / pharmacology
  • Neutrophil Infiltration*
  • Neutrophils / drug effects
  • Neutrophils / metabolism
  • Neutrophils / pathology
  • Phenylbutyrates / pharmacology
  • Receptors, Prostaglandin E, EP4 Subtype / agonists
  • Receptors, Prostaglandin E, EP4 Subtype / antagonists & inhibitors
  • Receptors, Prostaglandin E, EP4 Subtype / metabolism
  • Time-Lapse Imaging / methods

Substances

  • 4-(4-cyano-2-(2-(4-fluoronaphthalen-1-yl)propionylamino)phenyl)butyric acid
  • Anti-Inflammatory Agents, Non-Steroidal
  • Benzamides
  • Cyan Fluorescent Protein
  • Methyl Ethers
  • Naphthalenes
  • ONO-AE1-329
  • Phenylbutyrates
  • Ptger4 protein, mouse
  • Receptors, Prostaglandin E, EP4 Subtype
  • Green Fluorescent Proteins
  • mirdametinib
  • Diphenylamine
  • Cyclic AMP-Dependent Protein Kinases
  • Extracellular Signal-Regulated MAP Kinases