The role of the alpha7 nicotinic receptor in regulating the Pophyromonas gingivalis-induced expression of interleukin-8 by oral keratinocytes

There is increasing evidence for a role of acetylcholine in modulating the inflammatory response. This has been demonstrated to occur predominantly via the ‘cholinergic anti-inflammatory pathway’ and mediated by the α7 nicotinic acetylcholine receptor (α7nAChR). Therefore, there is now an abundance of in vitro and in vivo evidence that suggest pharmacologically target the α7nAChR is a potential anti-inflammatory therapy to treat chronic inflammatory diseases. The role of acetylcholine and the α7nAChR in modulating the periodontal immune response is at present unknown. However, as nu-merous cells within the periodontium express the α7nAChR it is interesting to speculate that targeting this receptor may modulate the periodontal immune response. Therefore, his study aims to investigate the effects of the α7nAChR on the periodontal pathogen (P. gingivalis) induced expression of Interleukin-8 (IL-8) by oral keratinocytes. Expression of the α7nAChR mRNA was demonstrated to be upregulated in diseased periodontal tissue. In line with previous studies, oral keratinocytes were found to ex-press the α7nAChR using PCR. The acetylcholine mimic; Carbachol when used in high concentrations was found to inhibit dead P. gingivalis induced IL-8 expression by OKF6-TERT2 cells in vitro. The expression of IL-8 was investigated at both the protein level by ELISA and the transcriptional level by real time PCR. In contrast, the specific α7nAChR agonist (PHA 543613 hydrochloride) when used at pharmacological concen-trations potently inhibited expression of IL-8 by OKF6-TERT2 cells cultured with a live P. gingivalis biofilm. This was again determined using ELISA and real time PCR. A membrane integrity assay confirmed that the agonist exhibited no toxic effects on the cells. The inhibition of IL-8 expression was found to be mediated at the transcriptional level. Indeed, using a Fast Activated Cell-based ELISA (FACE) NF-κB p65 Profiler Kit activation of the α7nAChR inhibited the phosphorylation of the NF-κB p65 subunit at Serine 468 and serine 536. Therefore, it can be hypothesised that targeting the α7nAChR leads to inhibition of NF-κB p65 subunit translocation into the nucleus and thus downregulated IL-8 transcription. These data suggest that the α7nAChR plays a role in regulating the pathogen induced immune response at epithelial surfaces. As dysregulated immunity is a hallmark of peri-odontal disease it is interesting to speculate that pharmacologically targeting the α7nAChR may offer a novel immune-modulatory therapy

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

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.&lt;p&gt;&lt;/p&gt; Materials and methods: Periodontal tissue expression of α7nAChR was determined by real-time PCR. OKF6/TERT-2 oral keratinocytes were exposed to &lt;i&gt;Porphyromonas gingivalis&lt;/i&gt; 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 &lt;i&gt;P. gingivalis&lt;/i&gt; lipopolysaccharide was determined using a GeneBLAzer M3 CHO-K1-blacell reporter assay.&lt;p&gt;&lt;/p&gt; Results: Expression of α7nAChR mRNA was elevated in diseased periodontal tissue. PHA-543613 hydrochloride inhibited &lt;i&gt;P. Gingivalis&lt;/i&gt;-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 &lt;i&gt;P. Gingivalis&lt;/i&gt; lipopolysaccharide.&lt;p&gt;&lt;/p&gt; Conclusion: These data suggest that α7nAChR plays a role in regulating the innate immune responses of oral keratinocytes.&lt;p&gt;&lt;/p&gt

Periodontal infectogenomics:a systematic review update of associations between host genetic variants and subgingival microbial detection

OBJECTIVE: The aim of this study was to systematically update the evidence for associations between host genetic variants and subgingival microbial detection and counts. MATERIALS AND METHODS: Following a previous systematic review (Nibali et al. J Clin Periodontol 43(11): 889-900, 15), an update of a systematic search of the literature was conducted in Ovid Medline, Embase, LILACS, and Cochrane Library for studies reporting data on host genetic variants and detection of microbes subgingivally published in the last 6 years. RESULTS: A total of 19 studies were included in the review, from an initial search of 2797 titles. Studies consisted mainly of candidate gene studies and of one genome-wide analysis. A total of 62 studies were considered for summary findings, including 43 identified in the previous systematic review of studies published up to 2015. Meta-analyses were done when appropriate including both papers in the original review and in the update. Meta-analyses revealed lack of associations between IL1 composite genotype and subgingival detection of Aggregatibacter acinomycetemcomitans, Poprhyromonas gingivalis, Tannerella forsythia, Treponema denticola, and Prevotella intermedia. Promising evidence is emerging from other genetic variants and from sub-analyses of data from genome-association studies. Among other studies with candidate-gene, target SNPs were mainly within the IL10, IL6, IL4, IL8, IL17A, and VDR gene. CONCLUSIONS: IL1 composite genotype does not seem to be associated with subgingival microbial detection. Promising associations should be pursued by future studies, including studies employing -OMICS technologies. CLINICAL RELEVANCE: A better knowledge of which host genetic variant predispose to subgingival microbial colonization and to the development of progression of periodontal disease could potentially help to better understand periodontal disease pathogenesis and help with its management. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00784-021-04233-8

Acetylcholine and the alpha 7 nicotinic receptor: a potential therapeutic target for the treatment of periodontal disease?

&lt;p&gt;Objectives: The aim of this review is to examine the evidence for a functional cholinergic system operating within the periodontium and determine the evidence for its role in periodontal immunity.&lt;/p&gt; &lt;p&gt;Introduction: Acetylcholine can influence the immune system via the ‘cholinergic anti-inflammatory pathway’. This pathway is mediated by the vagus nerve which releases acetylcholine to interact with the α7 subunit of the nicotinic acetylcholine receptor (α7nAChR) on proximate immuno-regulatory cells. Activation of the α7nAChR on these cells leads to down-regulated expression of pro-inflammatory mediators and thus regulates localised inflammatory responses. The role of the vagus nerve in periodontal pathophysiology is currently unknown. However, non-neuronal cells can also release acetylcholine and express the α7nAChR; these include keratinocytes, fibroblasts, T cells, B cells and macrophages. Therefore, by both autocrine and paracrine methods non-neuronal acetylcholine can also be hypothesised to modulate the localised immune response.&lt;/p&gt; &lt;p&gt;Methods: A Pubmed database search was performed for studies providing evidence for a functional cholinergic system operating in the periodontium. In addition, literature on the role of the ‘cholinergic anti-inflammatory pathway’ in modulating the immune response was extrapolated to hypothesise that similar mechanisms of immune regulation occur within the periodontium.&lt;/p&gt; &lt;p&gt;Conclusion: The evidence suggests a functional non-neuronal ‘cholinergic anti-inflammatory pathway’ may operate in the periodontium and that this may be targeted therapeutically to treat periodontal disease.&lt;/p&gt