Oral microbe-host interactions: influence of β-glucans on gene expression of inflammatory cytokines and metabolome profile

Silva et al. BMC Microbiology (2017) 17:53 DOI 10.1186/s12866-017-0946-1 RESEARCH ARTICLE Open Access Oral microbe-host interactions: influence of β-glucans on gene expression of inflammatory cytokines and metabolome profile Viviam de Oliveira Silva1,2, Luciano José Pereira3 and Ramiro Mendonça Murata4,5* Abstract Background: The aim of this study was to evaluate the effects of β-glucan on the expression of inflammatory mediators and metabolomic profile of oral cells [keratinocytes (OBA-9) and fibroblasts (HGF-1) in a dual-chamber model] infected by Aggregatibacter actinomycetemcomitans. The periodontopathogen was applied and allowed to cross the top layer of cells (OBA-9) to reach the bottom layer of cells (HGF-1) and induce the synthesis of immune factors and cytokines in the host cells. β-glucan (10 μg/mL or 20 μg/mL) were added, and the transcriptional factors and metabolites produced were quantified in the remaining cell layers and supernatant. Results: The relative expression of interleukin (IL)-1-α and IL-18 genes in HGF-1 decreased with 10 μg/mL or 20 μg/mL of β-glucan, where as the expression of PTGS-2 decreased only with 10 μg/mL. The expression of IL-1-α increased with 20 μg/mL and that of IL-18 increased with 10 μg/mL in OBA-9; the expression of BCL 2, EP 300, and PTGS-2 decreased with the higher dose of β-glucan. The production of the metabolite 4-aminobutyric acid presented lower concentrations under 20 μg/mL, whereas the concentrations of 2-deoxytetronic acid NIST and oxalic acid decreased at both concentrations used. Acetophenone, benzoic acid, and pinitol presented reduced concentrations only when treated with 10 μg/mL of β-glucan. Conclusions: Treatment with β-glucans positively modulated the immune response and production of metabolites. Keywords: Aggregatibacter actinomycetemcomitans, Periodontal disease, Host response, Keratinocyte, Fibroblast, Immunomodulation Background β-glucans from yeast have been used extensively as protective substances against infections with potent effects on the innate and adaptive immune responses. β-glucans are non-starch polysaccharides that make up structural cells of plants and microorganisms [1]. The cell wall of Saccharomyces cerevisiae is an important source of β-glucans and these represents about 50–60% of yeast [2]. The protective effect of these compounds has been demonstrated in * Correspondence: 4 School of Dental Medicine, Department Foundational Sciences, East Carolina University, 1851 MacGregor Downs Road, Greeville, NC 27834-4354, USA 5 Brody School of Medicine, Department of Microbiology and Immunology, East Carolina University, Greenville, NC, USA Full list of author information is available at the end of the article experimental infection [3]. Additionally, there are reports that these substances modulate allergy symptoms [4] and have anticancer properties [5, 6]. Many hypotheses have been put forward to explain the effects of β-glucans. Such compounds can act by inhibiting the adhesion of pathogens to epithelial tissues of the digestive tract by blocking carbohydrate-binding adhesins on bacteria; they stimulate the immunocompetent cells in Peyer’s patches and the consecutive activation of mechanisms of innate and adaptive immune defense; further, by adsorption of mycotoxins in food (when linked to the diet) β-glucans inhibit their toxic activity [2]. However, its effects on periodontal inflammation are still poorly studied. Periodontal disease is a highly prevalent disease in the adult population. It is characterized © The Author(s). 2017 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Silva et al. BMC Microbiology (2017) 17:53 by inflammation and progressive destruction of the periodontal tissues in response to specific microorganisms present in oral biofilm [7–10]. The pathogens associated with periodontal disease are frequently present in the human subgingival microbiota and are represented mainly by anaerobic gram-negative bacteria [11]. A. actinomycetemcomitans, Pasteurellaceae family, is a coccobacillus, fermentative, gram-negative, capnophilic, non-motile, and non-sporulating microorganism. This bacterium is considered the main etiological agent of localized aggressive periodontitis lesions, but is also associated with chronic periodontitis [12–18]. The progression of periodontal disease is associated with the virulence of the microorganism, together with the susceptibility of the host [19]. There are several virulence factors of A. actinomycetemcomitans that collaborate for its pathogenicity in periodontitis [20]. Leukotoxin, cytolethal distending toxins, bacteriocins, adhesins and lipopolysaccharide correspond to the variety of the microorganism virulence factors that may be associated with the pathogenesis of localized aggressive periodontitis [21]. These virulence factors attributed to A. actinomycetemcomitans are responsible for interacting with the host cells triggering an inflammatory response in the tissues supporting the teeth [22]. Fibroblasts and epithelial cells are the first cells to be activated in the oral cavity in response to exotoxic and endotoxic virulence factors of A. actinomycetemcomitans, performing an essential role in the production of cytokines involved in the inflammatory process. After this first local colonization, leukocytes (mainly monocytes and neutrophils) and dendritic cells are recruited to the site of infection giving sequence on inflammatory response [22, 23]. Recently, in vivo studies have demonstrated that β-glucans from S. cerevisiae present regulatory activity toward metabolism [24] and also modulate the expression of cycloxygenase-2 (COX-2), receptor activator of nuclear factor kappa-B ligand (RANK-L), and osteoprotegerin (OPG), decreasing alveolar bone loss caused by induced periodontal disease (ligature) in normal and diabetic animals [25]. However, knowledge of the molecular and biochemical mechanisms involved in β-glucan activity in periodontal disease is still not understood, demanding further research with advanced tissue culture techniques, examining the microbiota-host interaction. In that sense, the dual chamber model is an interesting in vitro model that mimics the human periodontum. It is constructed using a monolayer of epithelial keratinocytes and a subepithelial layer of fibroblasts on which the invasive periodontopathogen can be applied [26]. Thus, this study aims to evaluate the effects of β-gluc (...truncated)