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

Abstract

ObjectiveThe 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 methodsPeriodontal 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.ResultsExpression 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.ConclusionThese data suggest that α7nAChR plays a role in regulating the innate immune responses of oral keratinocytes.

Clinical associations between acetylcholine levels and cholinesterase activity in saliva and gingival crevicular fluid and periodontal diseases

“…The role of ACh as an anti‐inflammatory neuroendocrine mediator has received a great deal of attention since the discovery of the cholinergic anti‐inflammatory pathway (Borovikova et al., ). It is now well established that non‐neuronal ACh, acting via the α7nAChR, can also negatively regulate proinflammatory mediator expression by both professional and non‐professional immune cells (de Jonge et al., ; Macpherson et al., ; Yoshikawa et al., ). However, evidence from in vivo and in vitro infection models suggests that it is too simplistic to state that within complex mucosal tissues ACh acts purely in an anti‐inflammatory capacity (Giebelen, Leendertse, Florquin, & van der Poll, ; Giebelen et al., ; Rajendran et al., ).…”

“…ChAT + B cell populations in the intestinal mucosa have also been found to release ACh upon antigen stimulation which in turn modulates neutrophil recruitment (Reardon et al., ). Furthermore, oral keratinocytes have been found to release ACh in response to stimulation by Porphyromonas gingivalis and the α7nAChR on oral keratinocytes has been demonstrated to downregulate P. gingivalis induced expression of IL‐8 (CXCL8) (Macpherson et al., ). The evidence therefore suggests that ACh can modulate innate immune responses within the oral mucosa and subsequently the pathogenesis of periodontal diseases (Zoheir et al., ).…”

“…The oral mucosa can be defined as having a non-neuronal cholinergic system (Arredondo et al, 2003;Macpherson et al, 2014;Nguyen et al, 2000;Zoheir, Lappin, & Nile, 2012) due to the fact it expresses ChAT, acetylcholine receptors and cholinesterases (Rana et al, 2010;Wessler & Kirkpatrick, 2008). Roles for non-neuronal ACh within mucosal tissues include; cell proliferation and differentiation, cytoskeletal organization, cell-cell contact, maintenance of effective barrier functions, controlling ion and water movements and modulation of immune responses Wessler & Kirkpatrick, 2008).…”

Acetylcholine Protects against Candida albicans Infection by Inhibiting Biofilm Formation and Promoting Hemocyte Function in a Galleria mellonella Infection Model

“…In addition, the efferent vagus nerve interacts with the splenic nerve to activate a unique ACh-producing memory phenotype T-cell population that can propagate ACh-mediated immunoregulation throughout the body (6). Furthermore, ACh is produced by numerous cells outside of neural networks, and nonneuronal ACh can also play a vital role in immune regulation through its cytotransmitter capabilities (7,8).…”

Nicotine inhibits osteogenic differentiation of human periodontal ligament cells under cyclic tensile stress through canonical Wnt pathway and α7 nicotinic acetylcholine receptor

“…Diverse subtypes consist of 17 subunits including α1‐α10, β1‐β4, δ and γ . α7 nAChR, a specific receptor that binds to nicotine, is not only expressed in the central nervous system but is also found in the periodontium, including oral epithelial cells, oral keratinocytes and fibroblasts, thereby regulating the negative effect of nicotine on the osteogenic differentiation of hPDLCs . However, α‐BTX can specifically bind to the α7 nAChR, thereby impacting downstream signal transmission .…”