The alpha 7 nicotinic receptor agonist PHA-543613 hydrochloride inhibits Porphyromonas gingivalis-induced expression of interleukin-8 by oral keratinocytes

Inflamm Res. 2014 Jul;63(7):557-68. doi: 10.1007/s00011-014-0725-5. Epub 2014 Mar 8.

Abstract

Objective: The alpha 7 nicotinic receptor (α7nAChR) is expressed by oral keratinocytes. α7nAChR activation mediates anti-inflammatory responses. The objective of this study was to determine if α7nAChR activation inhibited pathogen-induced interleukin-8 (IL-8) expression by oral keratinocytes.

Materials and methods: Periodontal tissue expression of α7nAChR was determined by real-time PCR. OKF6/TERT-2 oral keratinocytes were exposed to Porphyromonas gingivalis in the presence and absence of a α7nAChR agonist (PHA-543613 hydrochloride) alone or after pre-exposure to a specific α7nAChR antagonist (α-bungarotoxin). Interleukin-8 (IL-8) expression was measured by ELISA and real-time PCR. Phosphorylation of the NF-κB p65 subunit was determined using an NF-κB p65 profiler assay and STAT-3 activation by STAT-3 in-cell ELISA. The release of ACh from oral keratinocytes in response to P. gingivalis lipopolysaccharide was determined using a GeneBLAzer M3 CHO-K1-bla cell reporter assay.

Results: Expression of α7nAChR mRNA was elevated in diseased periodontal tissue. PHA-543613 hydrochloride inhibited P. gingivalis-induced expression of IL-8 at the transcriptional level. This effect was abolished when cells were pre-exposed to a specific α7nAChR antagonist, α-bungarotoxin. PHA-543613 hydrochloride downregulated NF-κB signalling through reduced phosphorylation of the NF-κB p65-subunit. In addition, PHA-543613 hydrochloride promoted STAT-3 signalling by maintenance of phosphorylation. Furthermore, oral keratinocytes upregulated ACh release in response to P. gingivalis lipopolysaccharide.

Conclusion: These data suggest that α7nAChR plays a role in regulating the innate immune responses of oral keratinocytes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetylcholine / immunology
  • Animals
  • Bridged Bicyclo Compounds, Heterocyclic / pharmacology
  • Bungarotoxins / pharmacology
  • CHO Cells
  • Cricetulus
  • Humans
  • Interleukin-8 / immunology*
  • Keratinocytes / drug effects
  • Keratinocytes / immunology*
  • Lipopolysaccharides
  • Mouth Mucosa / cytology
  • Periodontal Diseases / genetics
  • Periodontal Diseases / immunology
  • Porphyromonas gingivalis
  • Quinuclidines / pharmacology
  • RNA, Messenger / metabolism
  • STAT3 Transcription Factor / immunology
  • Transcription Factor RelA / immunology
  • alpha7 Nicotinic Acetylcholine Receptor / agonists
  • alpha7 Nicotinic Acetylcholine Receptor / antagonists & inhibitors
  • alpha7 Nicotinic Acetylcholine Receptor / genetics
  • alpha7 Nicotinic Acetylcholine Receptor / immunology*

Substances

  • Bridged Bicyclo Compounds, Heterocyclic
  • Bungarotoxins
  • CXCL8 protein, human
  • Interleukin-8
  • Lipopolysaccharides
  • N-(1-azabicyclo(2.2.2)oct-3-yl)furo(2,3-c)pyridine-5-carboxamide
  • Quinuclidines
  • RNA, Messenger
  • STAT3 Transcription Factor
  • STAT3 protein, human
  • Transcription Factor RelA
  • alpha7 Nicotinic Acetylcholine Receptor
  • Acetylcholine