Inflammatory and bone remodelling related biomarkers following periodontal transplantation of the tissue engineered biocomplex.

OBJECTIVES: To assess gingival crevicular fluid (GCF) levels of inflammatory and bone remodelling related biomarkers following transplantation of a tissue-engineered biocomplex into intrabony defects at several time-points over 12-months. MATERIALS AND METHODS: Group-A (n = 9) received the Minimal Access Flap (MAF) surgical technique combined with a biocomplex of autologous clinical-grade alveolar bone-marrow mesenchymal stem cells in collagen scaffolds enriched with an autologous fibrin/platelet lysate (aFPL). Group-B (n = 10) received the MAF surgery, with collagen scaffolds enriched with aFPL and Group-C (n = 8) received the MAF surgery alone. GCF was collected from the osseous defects of subjects via paper strips/30 sec at baseline, 6-weeks, 3-, 6-, 9-, 12-months post-surgery. Levels of inflammatory and bone remodelling-related biomarkers in GCF were determined by ELISA. RESULTS: Group-A demonstrated significantly higher GCF levels of BMP-7 at 6-9 months than baseline, with gradually decreasing levels of pro-inflammatory and pro-osteoclastogenic markers (TNF-α, RANKL) over the study-period; and an overall decrease in the RANKL/OPG ratio at 9-12 months than baseline (all p < 0.001). In comparison, only modest interim changes were observed in Groups-B and -C. CONCLUSIONS: At the protein level, the approach of MAF and biocomplex transplantation provided greater tissue regeneration potential as cell-based therapy appeared to modulate inflammation and bone remodelling in residual periodontal defects. CLINICAL RELEVANCE: Transplantation of a tissue engineered construct into periodontal intrabony defects demonstrated a biochemical pattern for inflammatory control and tissue regeneration over 12-months compared to the control treatments. Understanding the biological healing events of stem cell transplantation may facilitate the design of novel treatment strategies. CLINICAL DATABASE REGISTRATION: ClinicalTrials.gov ID: NCT02449005.

Limitations of using 16S rRNA microbiome sequencing to predict oral squamous cell carcinoma.

A new era of next-generation sequencing has changed our perception of the oral microbiome in health and disease, and with this there is a growing understanding that the oral microbiome is a contributing factor to oral squamous cell carcinoma (OSCC), a malignancy of the oral cavity. This study aimed to analyse the trends and relevant literature based on the 16S rRNA oral microbiome in head and neck cancer using next-generation sequencing technologies, and to conduct a meta-analysis of the studies with OSCC cases and healthy controls. A literature search using the databases Web of Science and PubMed was conducted in a scoping-like review to collect information based on the study design, and plots were generated using RStudio. We selected case-control studies using 16S rRNA oral microbiome sequencing analysis in OSCC cases versus healthy controls for re-analysis. Statistical analyses were conducted using R. Out of 916 original articles, we filtered and selected 58 studies for review, and 11 studies for meta-analysis. Differences between sampling type, DNA extraction methods, next-generation sequencing technology and region of the 16S rRNA were identified. No significant differences in the α- and ß-diversity between health and oral squamous cell carcinoma were observed (p < 0.05). Random Forest classification marginally improved predictability of four studies (training set) when split 80/20. We found an increase in Selenomonas, Leptotrichia and Prevotella species to be indicative of disease. A number of technological advances have been accomplished to study oral microbial dysbiosis in oral squamous cell carcinoma. There is a clear need for standardization of study design and methodology to ensure 16S rRNA outputs are comparable across the discipline in the hope of identifying 'biomarker' organisms for designing screening or diagnostic tools.

Acetylcholine Receptors in Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) are well known for their regenerative potential. Even though the ability of MSCs to proliferate and differentiate has been studied extensively, there remains much to learn about the signaling mechanisms and pathways that control proliferation and influence the differentiation phenotype. In recent years, there has been growing evidence for the utility of non-neuronal cholinergic signaling systems and that acetylcholine (ACh) plays an important ubiquitous role in cell-to-cell communication. Indeed, cholinergic signaling is hypothesized to occur in stem cells and ACh synthesis, as well as in ACh receptor (AChR) expression, has been identified in several stem cell populations, including MSCs. Furthermore, AChRs have been found to influence MSC regenerative potential. In humans, there are two major classes of AChRs, muscarinic AChRs and nicotinic AChRs, with each class possessing several subtypes or subunits. In this review, the expression and function of AChRs in different types of MSC are summarized with the aim of highlighting how AChRs play a pivotal role in regulating MSC regenerative function.

Expression of Toll-like receptor and cytokine mRNAs in feline odontoclastic resorptive lesion (FORL) and feline oral health.

Feline odontoclastic resorptive lesion (FORL) is a common chronic inflammatory condition whose aetiopathogenesis remains unclear. FORL affects 20-75% of cats and causes excruciating pain and tooth loss. The purpose of this study was to evaluate chronic inflammation in FORL by assessing differences in Toll-like receptor (TLR) and cytokine transcripts in gingival tissues between diseased and healthy cats. Gingival tissue samples were collected from 14 healthy cats with no known clinical signs of oral disease and 41 cats with FORL. Levels of mRNA encoding TLR2, TLR3, TLR4, TLR7, TLR9 and the cytokines interleukin-1ß (IL-1ß), IL-4, IL-6, IL-10, IL-12, interferon-γ (IFN-γ) and tumour necrosis factor-α (TNF-α) was evaluated using quantitative real-time PCR. Statistical significance of the results was assessed using non-parametric tests. Levels of TLR and cytokine transcripts were upregulated in gingival tissue from cats with FORL as compared with healthy gingival tissue: TLR2, TLR3 and TLR9, p ≤ 0.001; TLR4 and TLR7, p ≤ 0.01; IFN-γ, IL-4, IL-6, IL-10, IL-12, IL-1ß and TNF-α, p ≤ 0.001). In conclusion, expression of TLR and both pro- and anti-inflammatory cytokines were significantly increased, confirming an ongoing chronic inflammatory response to the microbiome in FORL. It is likely that dysbiosis of the oral microbiota in cats with FORL activates the innate immune response, leading to active inflammation that results in tooth resorption.

Effects of smoking on non-surgical periodontal therapy in patients with periodontitis Stage III or IV, and Grade C.

BACKGROUND: To evaluate possible effects of smoking on clinical, biochemical, and microbiological outcomes of non-surgical periodontal treatment in patients with periodontitis Stage III or IV and Grade C. METHODS: Conventional quadrant-wise non-surgical periodontal treatment was performed and whole-mouth periodontal measurements were recorded at baseline, 1, 3, and 6 months after completion of treatment. Saliva, gingival crevicular fluid, subgingival plaque, and blood samples were obtained at the same time points. Inflammatory cytokine levels, presence, and quantities of 11 different bacterial species were determined. Smoking status was validated by cotinine assay. RESULTS: Fourteen smoker and 13 non-smoker patients completed the study protocol and revealed similar clinical findings except for the higher plaque scores in the non-smokers at 6 months (P <0.01). Significant differences were found between the study groups in biofluid cytokine levels at 1 and 3 months (P <0.01). Gram-negative bacteria were more abundant in the smokers at baseline and so were Gram-positive bacteria in the non-smokers (P <0.01). Gram-negative bacteria repopulated in the smokers faster than in the non-smokers (P <0.01). CONCLUSIONS: The present findings suggest that smoker patients with periodontitis Stage III and IV, Grade C respond well to the non-surgical periodontal treatment during the 6-month follow-up. However, smokers exhibit faster repopulation of Gram-negative bacteria.

Plaque Accumulation and Inflammation Adjacent to Restorations of Amorphous Calcium Phosphate-containing Composite in Early Childhood Caries.

PURPOSE: To evaluate the clinical, biochemical, and microbiological reactions to nanocomposite containing amorphous calcium phosphate (ACP) in comparison to a traditional composite restorative material in early childhood caries. MATERIALS AND METHODS: Eighteen teeth were restored with the test material (ACP-containing resin) and 18 teeth were restored with the control material (traditional composite, TC) in fourteen paediatric patients using a split-mouth design. One caries- and restoration-free intact tooth in each patient was selected as the healthy control. Gingival crevicular fluid (GCF) and supragingival plaque samples were collected at baseline before the treatment and also on days 1, 7, 14 and 30 after treatment. Unstimulated whole saliva samples were obtained from each patient at baseline, and 1 and 6 months after restoration. GCF and saliva samples were assayed for IL-17A, IL-17F IL-17A/F, IL-17E, OPG and RANKL levels by ELISA, and plaque composition was assessed using RT-PCR. RESULTS: Clinical evaluation indicated no statistically significant differences between the two restorative materials according to the FDI criteria surface lustre, material retention and marginal adaptation properties. Pro-inflammatory IL-17 levels decreased statistically significantly at 6 months compared to baseline and 1-month values (p < 0.05). The baseline pro-inflammatory IL-17 cytokine levels in GCF samples around the carious teeth were higher than those obtained around the healthy teeth (p < 0.05), but similar in GCF from the ACP-test and TC teeth. Microbiological findings were similar in the ACP and T groups. CONCLUSION: It may be suggested that both ACP-containing and traditional resin composites show similar antimicrobial and biochemical effects in early childhood caries.

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

AIM: The oral mucosa possesses a non-neuronal cholinergic system. This study aimed to determine clinical evidence for a role of cholinergic mechanisms in the pathogenesis of periodontal diseases. MATERIALS AND METHODS: Fifty healthy participants, 52 patients with gingivitis and 49 with periodontitis were recruited. Full periodontal parameters were recorded and saliva and gingival crevicular fluid (GCF) collected. Levels of acetylcholine and inflammatory mediators were quantified using commercially available assay kits. Acetylcholinesterase and butyrylcholinesterase activities were measured using a published biochemical assay. RESULTS: Acetylcholine levels are significantly elevated in saliva and GCF, whereas GCF levels of butyrylcholinesterase activity are significantly decreased, in patients with periodontal diseases. Acetylcholine levels in saliva and GCF correlated positively with clinical markers of disease severity and with increased levels of IL-17A and IL-17F. In contrast, butyrylcholinesterase activity levels in GCF showed significant negative correlations with clinical markers of disease severity and IL-17A and IL-17F levels. None of the findings were due to smoking. CONCLUSIONS: Elevated acetylcholine levels and reduced butyrylcholinesterase activity are clinically associated with periodontal diseases and elevated levels of IL-17A and IL-17F. Therefore, non-neuronal cholinergic mechanisms may influence IL-17 biology and the aetiopathogenesis of periodontal diseases and therefore are possible therapeutic targets.

The Anti-Adhesive Effect of Curcumin on Candida albicans Biofilms on Denture Materials.

The use of natural compounds as an alternative source of antimicrobials has become a necessity given the growing concern over global antimicrobial resistance. Polyphenols, found in various edible plants, offers one potential solution to this. We aimed to investigate the possibility of using curcumin within the context of oral health as a way of inhibiting and preventing the harmful development of Candida albicans biofilms. We undertook a series of adsorption experiments with varying concentrations of curcumin, showing that 50 µg/ml could prevent adhesion. This effect could be further synergized by the curcumin pre-treatment of yeast cells to obtain significantly greater inhibition (>90%, p < 0.001). Investigation of the biological impact of curcumin showed that it preferentially affected immature morphological forms (yeast and germlings), and actively promoted aggregation of the cells. Transcriptional analyses showed that key adhesins were down-regulated (ALS1 and ALS3), whereas aggregation related genes (ALS5 and AAF1) were up-regulated. Collectively, these data demonstrated that curcumin elicits anti-adhesive effects and that induces transcription of genes integrally involved in the processes related to biofilm formation. Curcumin and associated polyphenols therefore have the capacity to be developed for use in oral healthcare to augment existing preventative strategies for candidal biofilms on the denture surface.

Biomarkers and Bacteria Around Implants and Natural Teeth in the Same Individuals.

BACKGROUND: This cross-sectional study assesses cytokine levels in peri-implant crevicular fluid (PICF)/gingival crevicular fluid (GCF) and a selection of subgingival/submucosal plaque bacteria from clinically healthy or diseased sites in the same individuals. METHODS: Samples from 97 implants/teeth (58 implants [19 healthy, 20 mucositis, 19 peri-implantitis] and 39 natural teeth [19 healthy, 12 gingivitis, eight periodontitis] in 15 systemically healthy patients were investigated by immunoassay and real-time polymerase chain reaction. Samples were obtained first, with probing depth, clinical attachment level, bleeding on probing, plaque index scores, and keratinized tissue width then recorded. Data were analyzed by Wilcoxon, Mann-Whitney U, and permutation tests on dependent, independent, and mixed dependent and independent samples and Spearman correlation. RESULTS: Interleukin (IL)-1ß levels were significantly higher in PICF samples of healthy implants than in GCF samples of healthy teeth (P = 0.003), and soluble receptor activator of nuclear factor-κB ligand (sRANKL) concentrations were significantly higher in the gingivitis than the mucositis group (P = 0.004). Biomarker levels were similar in peri-implantitis and periodontitis groups (P >0.05). Actinomyces naeslundi and Streptococcus oralis levels were significantly higher in the healthy implant group than in healthy teeth (P <0.05). Prevotella intermedia and Treponema denticola (Td) levels were lower in the mucositis group than the gingivitis group (P <0.05). Prevotella oralis and S. oralis levels were significantly higher in the periodontitis group (P <0.05), and Td levels were significantly higher in the peri-implantitis group (P <0.05). CONCLUSION: There were many similarities but, crucially, some differences in biomarker levels (IL-1ß and sRANKL) and bacterial species between peri-implant and periodontal sites in the same individuals, suggesting similar pathogenic mechanisms.

New strategic insights into managing fungal biofilms.

Fungal infections have dramatically increased in the last decades in parallel with an increase of populations with impaired immunity, resulting from medical conditions such as cancer, transplantation, or other chronic diseases. Such opportunistic infections result from a complex relationship between fungi and host, and can range from self-limiting to chronic or life-threatening infections. Modern medicine, characterized by a wide use of biomedical devices, offers new niches for fungi to colonize and form biofilm communities. The capability of fungi to form biofilms is well documented and associated with increased drug tolerance and resistance. In addition, biofilm formation facilitates persistence in the host promoting a persistent inflammatory condition. With a limited availability of antifungals within our arsenal, new therapeutic approaches able to address both host and pathogenic factors that promote fungal disease progression, i.e., chronic inflammation and biofilm formation, could represent an advantage in the clinical setting. In this paper we discuss the antifungal properties of myriocin, fulvic acid, and acetylcholine in light of their already known anti-inflammatory activity and as candidate dual action therapeutics to treat opportunistic fungal infections.

The Influence of Glycated Hemoglobin on the Cross Susceptibility Between Type 1 Diabetes Mellitus and Periodontal Disease.

BACKGROUND: Periodontal disease is a major complication of type 1 diabetes mellitus (T1DM). The aim of the present study is to investigate the relationship between glycated hemoglobin and circulating levels of interleukin (IL)-6, IL-8, and C-X-C motif chemokine ligand 5 (CXCL5) in non-smoking patients suffering from T1DM, with and without periodontitis. In addition, to determine the effect of advanced glycation end products (AGE) in the presence and absence of Porphyromonas gingivalis lipopolysaccharide (LPS) on IL-6, IL-8, and CXCL5 expression by THP-1 monocytes and OKF6/TERT-2 cells. METHODS: There were 104 participants in the study: 19 healthy volunteers, 23 patients with periodontitis, 28 patients with T1DM, and 34 patients with T1DM and periodontitis. Levels of blood glucose/glycated hemoglobin (International Federation of Clinical Chemistry [IFCC]) were determined by high-performance liquid chromatography. Levels of IL-6, IL-8, and CXCL5 in plasma were determined by enzyme-linked immunosorbent assay (ELISA). In vitro stimulation of OKF6/TERT-2 cells and THP-1 monocytes was performed with combinations of AGE and P. gingivalis LPS. Changes in expression of IL-6, IL-8, and CXCL5 were monitored by ELISA and real-time polymerase chain reaction. RESULTS: Patients with diabetes and periodontitis had higher plasma levels of IL-8 than patients with periodontitis alone. Plasma levels of IL-8 correlated significantly with IFCC units, clinical probing depth, and attachment loss. AGE and LPS, alone or in combination, stimulated IL-6, IL-8, and CXCL5 expression in both OKF6/TERT-2 cells and THP-1 monocytes. CONCLUSIONS: Elevated plasma levels of IL-8 potentially contribute to the cross-susceptibility between periodontitis and T1DM. P. gingivalis LPS and AGE in combination caused significantly greater expression of IL-6, IL-8, and CXCL5 from THP-1 monocytes and OKF6/TERT-2 cells than LPS alone.

The alpha 7 nicotinic receptor agonist PHA-543613 hydrochloride inhibits Porphyromonas gingivalis-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. 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.

Leptin enhances the secretion of interleukin (IL)-18, but not IL-1ß, from human monocytes via activation of caspase-1.

Circulating levels of leptin are elevated in type-2 diabetes mellitus (T2DM) and leptin plays a role in immune responses. Elevated circulating IL-18 levels are associated with clinical complications of T2DM. IL-18 regulates cytokine secretion and the function of a number of immune cells including T-cells, neutrophils and macrophages and as such has a key role in immunity and inflammation. Pro-inflammatory monocytes exhibiting elevated cytokine secretion are closely associated with inflammation in T2DM, however, little is known about the role of leptin in modifying monocyte IL-18 secretion. We therefore aimed to investigate the effect of leptin on IL-18 secretion by monocytes. We report herein that leptin increases IL-18 secretion in THP-1 and primary human monocytes but has no effect on IL-18mRNA. Leptin and LPS signalling in monocytes occurs by overlapping but distinct pathways. Thus, in contrast to a strong stimulation by LPS, leptin has no effect on IL-1ßmRNA levels or IL-1ß secretion. In addition, LPS stimulates the secretion of IL-6 but leptin did not whereas both treatments up regulate IL-8 secretion from the same cells. Although leptin (and LPS) has a synergistic effect with exogenous ATP on IL-18 secretion in both THP-1 and primary monocytes, experiments involving ATP assays and pharmacological inhibition of ATP signalling failed to provide any evidence that endogenous ATP secreted by leptin-stimulated monocytes was responsible for enhancement of monocyte IL-18 secretion by leptin. Analysis of the action of caspase-1 revealed that leptin up regulates caspase-1 activity and the effect of leptin on IL-18 release is prevented by caspase-1 inhibitor (Ac-YVAD-cmk). These data suggest that leptin activates IL-18 processing rather than IL-18 transcription. In conclusion, leptin enhances IL-18 secretion via modulation of the caspase-1 inflammasome function and acts synergistically with ATP in this regard. This process may contribute to aberrant immune responses in T2DM and other conditions of hyperleptinemia.

Investigating the biological properties of carbohydrate derived fulvic acid (CHD-FA) as a potential novel therapy for the management of oral biofilm infections.

BACKGROUND: A number of oral diseases, including periodontitis, derive from microbial biofilms and are associated with increased antimicrobial resistance. Despite the widespread use of mouthwashes being used as adjunctive measures to control these biofilms, their prolonged use is not recommended due to various side effects. Therefore, alternative broad-spectrum antimicrobials that minimise these effects are highly sought after. Carbohydrate derived fulvic acid (CHD-FA) is an organic acid which has previously demonstrated to be microbiocidal against Candida albicans biofilms, therefore, the aims of this study were to evaluate the antibacterial activity of CHD-FA against orally derived biofilms and to investigate adjunctive biological effects. METHODS: Minimum inhibitory concentrations were evaluated for CHD-FA and chlorhexidine (CHX) against a range of oral bacteria using standardised microdilution testing for planktonic and sessile. Scanning electron microscopy was also employed to visualise changes in oral biofilms after antimicrobial treatment. Cytotoxicity of these compounds was assessed against oral epithelial cells, and the effect of CHD-FA on host inflammatory markers was assessed by measuring mRNA and protein expression. RESULTS: CHD-FA was highly active against all of the oral bacteria tested, including Porphyromonas gingivalis, with a sessile minimum inhibitory concentration of 0.5%. This concentration was shown to kill multi-species biofilms by approximately 90%, levels comparable to that of chlorhexidine (CHX). In a mammalian cell culture model, pretreatment of epithelial cells with buffered CHD-FA was shown to significantly down-regulate key inflammatory mediators, including interleukin-8 (IL-8), after stimulation with a multi-species biofilm. CONCLUSIONS: Overall, CHD-FA was shown to possess broad-spectrum antibacterial activity, with a supplementary function of being able to down-regulate inflammation. These properties offer an attractive spectrum of function from a naturally derived compound, which could be used as an alternative topical treatment strategy for oral biofilm diseases. Further studies in vitro and in vivo are required to determine the precise mechanism by which CHD-FA modulates the host immune response.

Comparison of circulating tumour necrosis factor superfamily cytokines in periodontitis patients undergoing supportive therapy: a case-controlled cross-sectional study comparing smokers and non-smokers in health and disease.

BACKGROUND: B-cells are prominent immune cells in established periodontitis lesions. Tumour necrosis factor superfamily (TNFSF) cytokines play roles in supporting B-cell function as well as bone re-modelling. The influence of smoking on factors that support B-cell function in periodontitis remains unclear. AIM: To investigate plasma concentrations of TNF (TNSF1A), soluble receptor activator of nuclear-factor Kappa-B ligand (sRANKL/TNFSF11), a proliferation-inducing ligand (APRIL/TNFSF13), B-cell activating factor (BAFF/TNFSF13B) and Osteoprotegerin (OPG/TNFRSF11B) in smokers and non-smokers with and without chronic periodontitis MATERIALS & METHODS: Plasma concentrations of TNFSF and OPG were evaluated in 200 systemically healthy subjects divided into four groups: non-smokers with periodontitis (n = 101), smokers with periodontitis (n = 55), healthy non-smokers (n = 27) and healthy smokers (n = 17). RESULTS: Periodontitis patients had significantly higher plasma sRANKL, TNF, APRIL and BAFF and lower OPG than healthy subjects (p < 0.01). TNF and sRANKL were significantly greater in smokers with periodontitis (p = 0.011, p = 0.001) and OPG concentrations significantly lower (p = 0.001), whereas APRIL or BAFF were little changed. Plasma APRIL, BAFF, sRANKL and TNF correlated with probing depth and clinical attachment loss. CONCLUSION: TNFSF cytokines correlate with periodontitis disease severity. However, only TNF, sRANKL and OPG levels were altered by cigarette smoking. APRIL and BAFF appear as good indicators of disease severity.

Gingival crevicular fluid, serum levels of receptor activator of nuclear factor-κB ligand, osteoprotegerin, and interleukin-17 in patients with rheumatoid arthritis and osteoporosis and with periodontal disease.

BACKGROUND: This study is performed to evaluate gingival crevicular fluid (GCF) and serum levels of soluble receptor activator of nuclear factor-κB ligand (sRANKL), interleukin (IL)-17A, IL-17E, IL-17F, IL-17A/F, and osteoprotegerin (OPG) in women with rheumatoid arthritis (RA), osteoporosis (OPR), and those who are systemically healthy (SH), all with periodontal disease. METHODS: GCF and serum samples were obtained before any periodontal intervention from 17 women with RA, 19 with OPR, and 13 who were SH with periodontitis. Full-mouth clinical periodontal measurements were recorded. sRANKL, OPG, and IL-17 levels were determined by enzyme-linked immunosorbent assay. RESULTS: Clinical periodontal measurements were similar in the three study groups. Although the total amounts of GCF albumin, OPG, IL-17A, and IL-17A/F were similar in the study groups, there were statistically significant differences in GCF concentrations of sRANKL, OPG, IL-17A, IL-17E, IL-17F, and IL-17A/F. The sRANKL/OPG ratios were significantly higher in the RA group than in the OPR and SH groups (P <0.05). Serum sRANKL, sRANKL/OPG, and IL-17A/IL-17E ratios were significantly higher, whereas OPG concentrations were significantly lower in the RA group compared to other groups (P <0.05). Serum IL-17A concentrations were significantly higher in the RA and OPR groups than in the SH group (P <0.05). CONCLUSION: Increased inflammatory mediator levels in patients with RA, despite the long-term use of various anti-inflammatory drugs, suggest that these patients may have a propensity to overproduce these inflammatory mediators.

Is interleukin-17 involved in the interaction between polycystic ovary syndrome and gingival inflammation?

BACKGROUND: Polycystic ovarian syndrome (PCOS) is a hormonal disorder of females of reproductive age that impacts their oral and systemic health. The aim of this study is to evaluate interleukin-17A (IL-17A), IL-17F, IL-17A/F, and IL-17E (IL-25) levels in gingival crevicular fluid (GCF), saliva, and serum of non-obese females with PCOS and with either a clinically healthy periodontium or gingivitis. METHODS: Thirty-one females with PCOS, 30 females with PCOS and gingivitis, and 12 systemically and periodontally healthy females participated in the study. Clinical periodontal measurements, body mass index, and Ferriman-Gallwey score (FGS) (a measure of hirsutism in females) were recorded. Circulating levels of sex hormones, cortisol, and insulin were also determined. Levels of IL-17 cytokines were measured by enzyme-linked immunosorbent assay. RESULTS: The general linear model multivariate analysis, adjusting for age or plaque index, showed that the two groups with PCOS had higher concentrations of IL-17A, IL-17F, and IL-17A/F in serum and higher levels of IL-17A and IL-17F in GCF and saliva but lower serum IL-17E than systemically healthy females. Levels of IL-17E were lowest in females with PCOS and gingivitis who also had the highest FGS. Serum IL-17A and IL-17F levels correlated positively with FGS and periodontal probing depth (all ρ >0.33; P <0.005). Serum IL-17E showed the reverse relationship and also correlated negatively with IL-17A (ρ >-0.28; P <0.05). CONCLUSIONS: IL-17 levels are altered in non-obese females with PCOS and may influence gingival inflammation. Additional studies are warranted to clarify the relationship between PCOS and gingivitis.

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

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. 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. 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. 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.

Expression and regulation of interleukin-33 in human monocytes.

Interleukin-33 (IL-33) is an IL-1 family cytokine that has a role in regulating T helper type 2 cytokines and mast cell development. Expression of IL-33 is also associated with chronic inflammatory conditions such as rheumatoid arthritis. However, there is little information regarding IL-33 in myeloid cell immune responses, which are important in immunity and inflammation. We therefore investigated the expression, intracellular location and regulation of myeloid cell IL-33 by lipopolysaccharide (LPS) from Escherichia coli and the periodontal pathogen Porphyromonas gingivalis. We detected IL-33 messenger RNA in the human promonocytic cell line THP-1, in monocytes derived from these cells and in primary human monocytes. However, IL-33 was not expressed in primary monocyte-derived dendritic cells. Stimulation of monocytes with E. coli LPS (Toll-like receptor 4 agonist) and LPS from P. gingivalis (Toll-like receptor 2 agonist) up-regulated IL-33 at both the messenger RNA and protein levels but IL-1beta and tumour necrosis factor-alpha had no effect. The IL-33 protein was mainly found in the cytoplasm of monocytes with no evidence of nuclear translocation in stimulated cells. Furthermore, no IL-33 secretion was detected after stimulation with LPS and/or ATP. These data indicate that the function, if any, of IL-33 in activated monocytes is primarily intracellular. Interestingly, immunofluorescence analysis indicated that IL-33 was sequestered in the nucleus of monocytes undergoing apoptosis but released into the extracellular milieu by LPS-stimulated cells in which necrosis had been induced by freeze-thawing. Therefore, this endorses the view that IL-33 may function as an 'alarmin' and have a role in signalling cellular damage and inflammatory disease pathogenesis through release from damaged or necrotic cells.

Expression and functional analyses of liver expressed antimicrobial peptide-2 (LEAP-2) variant forms in human tissues.

The antimicrobial peptide Liver Expressed Antimicrobial Peptide-2 (LEAP-2) is proposed to function as part of the vertebrate innate immune system. However, the highly conserved nature of the LEAP-2 peptide primary structure among vertebrates suggests more fundamental physiological roles. RT-PCR analyses confirmed expression of LEAP-2 mRNA variants in human gastro-intestinal (GI) epithelial tissues and THP-1 monocytes. Three cDNA products indicative of at least three different spliced transcripts were observed. Translation of the cDNA sequences supported synthesis of transcripts encoding the secreted LEAP-2 peptide and two variants lacking signal sequences suggesting intracellular localisation. The synthesis and cytoplasmic localisation of LEAP-2 peptides in epithelia was supported by immunohistochemical analyses. Functional data suggested that LEAP-2 is not involved in the physiological response of GI epithelia to iron, nor is it mitogenic for epithelial cells or chemotactic for THP-1 monocytes. However, changes in the LEAP-2 transcript patterns associated with the challenge of THP-1 monocytes with lipopolysaccharide (100ng/ml) were supportive of the peptides having multiple roles in the innate immune response.