Porphyromonas gingivalis lipopolysaccharide promotes T- helper 17 cell differentiation from human CD4+ naïve T cells via toll-like receptor-2 in vitro Publication date: November 2019 Source: Archives of Oral Biology, Volume 107 Author(s): Liping Zhang, Li Gao, Chenrong Xu, Xiting Li, Panpan Wang, Chi Zhang, Chuanjiang Zhao Abstract Objectives The persistence of T-helper 17 (Th17) cells has been shown to support chronic inflammation and mediate tissue destruction in periodontitis. However, little is known regarding the underlying mechanisms that regulate Th17 cell differentiation in the periodontal inflammatory context. The objective of this study was to explore the possible effect and mechanism of lipopolysaccharide (LPS) from Porphyromonas gingivalis on Th17 cell differentiation. Methods Activated human CD4+CD45RA+ naïve T cells were stimulated with different doses of LPS from virulent and avirulent P. gingivalis strains combined with Th17 driven cytokines in vitro. Flow cytometry was used to analyze the differentiation ratio of Th17 cells. IL-17A protein expression and IL-17, retinoid-related orphan receptor C (RORC) and toll-like receptor 2 (TLR2) mRNA transcription were analysed by ELISA and real-time qPCR, respectively. The role of TLR2 in Th17 cell differentiation was further confirmed by TLR2 blocking assay. Results LPS from P. gingivalis (Pg-LPS) up-regulated Th17 cell differentiation ratios, expression of IL-17 and RORC mRNA, and IL-17 concentration in culture supernatant, with 0.1 μg/mL LPS from the virulent P. gingivalis strain being the most effectively. Furthermore, Pg-LPS also up-regulated expression of TLR2 on T cells during Th17 differentiation, and the differentiation was attenuated by treatment with TLR2 antibody. Conclusions These results suggest that Pg-LPS promotes Th17 cell differentiation in vitro, and TLR2 signalling may be involved in this process. LPS from the virulent P. gingivalis strain up-regulated Th17 cell differentiation more effectively, which may be associated with the pathogenicity of different P. gingivalis strains.

Effects of full-length human amelogenin on the differentiation of dental epithelial cells and osteoblastic cells Publication date: November 2019 Source: Archives of Oral Biology, Volume 107 Author(s): Ayumi Takahashi, Takao Morita, Kaori Murata, Erika Minowa, Azmeree Jahan, Masato Saito, Akihiko Tanimura Abstract Background and objective Amelogenins are major components of extracellular matrix proteins in developing teeth, and regulate the growth of enamel crystals. They also function as signaling molecules in cell differentiation. This study aimed to determine the biological effects of amelogenins on the differentiation of HAT-7 dental epithelial cells and MC3T3-E1 pre-osteoblastic cells using full-length recombinant human amelogenin (rh-AMEL). Design rh-AMEL was expressed in a mammalian cell line (Expi293F™) and was purified by DDK agarose beads. Effects of rh-AMEL on differentiation were evaluated by Mineralization and Alkaline phosphatase (ALP) activity using Alizarin Red S staining and colorimetric substrate p-nitrophenol, respectively. Results Western blotting and silver staining confirmed the successful purification of rh-AMEL. Mineralization and ALP activity in HAT-7 cells were significantly higher after treatment with 4 μg/mL rh-AMEL, but not after treatment with Emdogain® (EMD). In MC3T3-E1 cells, on the other hand, rh-AMEL showed biphasic effects on differentiation. Treatment with low concentrations of rh-AMEL (0.001–0.1 μg/mL) and EMD (0.01–1 μg/mL) increased mineralization and ALP activity in MC3T3-E1 cells, whereas treatment with high concentrations of rh-AMEL (4 μg/mL) and EMD (100 μg/mL) had the opposite effect. Conclusion High concentrations of rh-AMEL and EMD decreased the differentiation of MC3T3-E1 cells. By contrast, a high concentration of rh-AMEL, but not that of EMD, promoted the differentiation of HAT-7 cells. This study demonstrates that the effects of rh-AMEL on cell differentiation differ between HAT-7 and MC3T3-E1 cells, and suggests that different regions on AMEL may induce the differentiation of these cell types.

Antimicrobial and biofilm anti-adhesion activities of silver nanoparticles and farnesol against endodontic microorganisms for possible application in root canal treatment Publication date: November 2019 Source: Archives of Oral Biology, Volume 107 Author(s): Gisselle Moraima Chávez-Andrade, Mário Tanomaru-Filho, Maria Inês Basso Bernardi, Renato de Toledo Leonardo, Gisele Faria, Juliane Maria Guerreiro-Tanomaru Abstract Objective This study aimed to evaluate the antimicrobial and biofilm anti-adhesion activities of poly(vinyl alcohol)-coated silver nanoparticles (AgNPs-PVA) and farnesol against Enterococcus faecalis, Candida albicans or Pseudomonas aeruginosa. Design Minimum inhibitory concentration (MIC) and minimum microbicidal concentration (MMC) of the solutions, as well as the effect on the biofilm biomass were evaluated. The biofilm anti-adhesion activity was evaluated using bovine root dentine treated with the solutions after 3 min of contact and analyzed by scanning electron microscopy (SEM) and by colony-forming units per milliliter (CFU mL−1) counting. Data were analyzed using ANOVA and Tukey's, the paired Student’s t-test or Kruskal-Wallis and Dunn’s tests (α = 0.05). Results The MIC and MMC values (MIC/MMC) of the AgNPs-PVA and farnesol against E. faecalis were 42.5/50 μM and 0.85/1.0%, respectively. For C. albicans, the values were 27.5/37.5 μM and 1.75/2.5%; and for P. aeruginosa, 32.5/32.5 μM and 2.5/2.75%, respectively. Both solutions showed reduced biofilm biomass (p < 0.05). SEM analysis showed that dentine blocks treated with both solutions had lower biofilm formation than the control (saline), except for C. albicans. In the CFU mL−1 counting, biofilm cells were viable in the all groups in comparison with control (p > 0.05). Conclusions AgNPs-PVA and farnesol showed antimicrobial and biofilm anti-adhesion activities, as well as potential for use as coadjuvant in endodontic treatment, and may be an option as auxiliary procedure for root canal disinfection or to inhibit biofilm formation.

Change of saliva composition with radiotherapy Publication date: October 2019 Source: Archives of Oral Biology, Volume 106 Author(s): Vera J. Müller, Georgios N. Belibasakis, Philipp P. Bosshard, Daniel B. Wiedemeier, Dominique Bichsel, Martin Rücker, Bernd Stadlinger Abstract Objective The aim of this study was to analyze the physiological and microbiological changes of saliva from patients with head and neck cancer during and after intensity-modulated radiotherapy (IMRT). Design In this prospective clinical trial saliva samples and oral candida swabs were collected from patients receiving IMRT due to head and neck cancer (examination group). The first measurement was scheduled before radiotherapy, the other measurements during and after radiotherapy up to a one year follow-up. Additionally samples from healthy controls were collected over six weeks. Salivary flow rate and pH were measured. Microbiological analysis of cariogenic and periodontopathogenic taxa was performed by fluorescence in situ hybridization and oral Candida spp occurrence was evaluated by swab tests. Results 11 patients and 19 controls were included. The salivary flow rate and the unstimulated pH of the examination group were significantly reduced during radiotherapy compared with the measurement before radiotherapy and to the control group. Total bacteria, streptococci and lactobacilli numbers slightly increased after radiotherapy, resuming baseline levels after one year. Mutans streptococci, Porphyromonas gingivalis and Treponema denticola were barely detectable, whereas Tannerella forsythia slightly increased following radiotherapy. No differences in Candida levels were observed in the study. Conclusions Salivary changes in quantitative, qualitative and microbial composition occur during and after radiotherapy, with resumption of the measurements towards baseline levels after one year. While low levels of cariogenic and periodontopathogenic species were detected, the lower pH and salivary flow combined with increased numbers of aciduric and acidogenic lactobacilli corroborates a higher risk for caries, necessitating prevention.

Effectiveness of poly-γ-glutamic acid in maintaining enamel integrity Publication date: October 2019 Source: Archives of Oral Biology, Volume 106 Author(s): Zeeshan Qamar, Zubaidah Binti Haji Abdul Rahim, Gan Seng Neon, Hooi Pin Chew, Tayyaba Zeeshan Abstract Objective The aim of the study was to determine demineralisation inhibition and remineralisation potential of poly-γ-glutamic acid with its possible mechanism of action on human dental enamel. Methodology Three sodium-fluoride(NaF) concentration(0.01%w/v,0.1%w/v and 0.5%w/v respectively)and two poly-γ-glutamic acid(PGGA)concentration(1%w/v and 2%w/v respectively)were prepared in 0.1 M acetic acid(pH4.0)and deionized distilled water.For de/re-mineralisation study, tooth samples (18 teeth varnished, leaving a 2 mm2 window on the mid-buccal surfaces) were immersed in respective acidified NaF and PGGA solutions. The Ca2+ release/uptake was monitored with ISE over 72-hr with increasing pH every 24-h from 4.0 to 6.0.These teeth were later subjected to cross-sectional microhardness to determine integrated mineral recovery of enamel on increasing pH of respective acidified solution.In order to determine mechanism of PGGA,two concentrations of PGGA in deionized-water-solutions were used for tooth samples immersion followed by overnight drying then later subjected to Fourier Transform Infra-Red(FT-IR) analysis.The FT-IR analysis was also carried out on PGGA powder.For control,the experiment was repeated using hydroxyapatite(HAp)pellets.The density of PGGA solutions(1%and2%)was also measured to determine their dynamic viscosities. Results The ISE and microhardness testing revealed statistically significant (ρ ≤ 0.05) dissolution inhibition and remineralisation potential for tooth sample treated with acidified 2%PGGA. From the FT-IR spectra, it was observed that the profiles of the enamel and HAp surfaces treated with 1%-and 2%-PGGA solutions were similar to those of PGGA powder.It was found that the viscosity of PGGA increases with increasing concentration. Conclusion The study implies that 2% PGGA is more effective than NaF as forms a coating layer to protect from demineralisation and promote remineralisation of the tooth surface.

Identification of candidate genes of nasopharyngeal carcinoma by bioinformatical analysis Publication date: October 2019 Source: Archives of Oral Biology, Volume 106 Author(s): Zhimin Ye, Fangzheng Wang, Fengqin Yan, Lei Wang, Bin Li, Tongxin Liu, Fujun Hu, Mingxiang Jiang, Zhenfu Fu Abstract Objective This study aimed to identify candidate genes as potential biomarkers in nasopharyngeal carcinoma (NPC) by bioinformatical analysis. Methods Three microarray datasets: GSE32906, GSE15170, GSE53819 were download from public database and analyzed to identify the differentially expressed genes (DEGs) between NPC and normal samples. Functional and pathway enrichment analysis of the DEGs were performed. Protein-protein interaction network and gene-transcription factor regulatory network of DEGs were constructed. And the expression of hub genes in NPC was also validated based on the public database. Results A total of 16 up-regulated and 27 down-regulated genes were screened out from the microarray datasets. Functional and pathway enrichment analysis showed that DEGs were mostly enriched in positive regulation of angiogenesis, mesenchymal cell proliferation, cell surface and DNA binding, ECM-receptor interaction pathway, PI3K-Akt signaling pathway, and pathways in cancer. Five hub genes JUN, VEGFA, FOXM1, MYB, and WNT5A were identified from the protein-protein interaction network. Subsequently, the hub gene-transcription factor regulatory network revealed that STAT3, MYC, SOX2, RUNX2 present key relations with hub genes. The expression of these five hub genes were also validated to be differentially expressed among NPC and normal samples. Conclusions The current study indicated that the hub DEGs JUN, VEGFA, FOXM1, MYB, and WNT5A we identified might be potential therapeutic biomarkers of NPC.

Bilateral intermittent nasal obstruction in adolescent rats leads to the growth defects of mandibular condyle Publication date: October 2019 Source: Archives of Oral Biology, Volume 106 Author(s): Xiaoling Wang, Huijun Sun, Yanfei Zhu, Yanmei Tang, Xiaochen Xue, Ping Nie, Min Zhu, Bing Wang Abstract Objective This study aimed to evaluate the effects of nasal obstruction on mandibular growth, especially condyle, in adolescent rats and explore the possible mechanism with a focus on mesenchymal stem cells (MSCs) from condylar tissues. Design 4-week-old male Sprague–Dawley rats were randomly divided into bilateral intermittent nasal obstruction (i.e. mouth-breathing, MB) and nasal-breathing (NB) groups. Self-made plugs were used to obstruct the nasal cavity in the MB group for 4 weeks, from 8:00 a.m. to 4:00 p.m. every day. The body weights were recorded. Three-dimensional computed tomography (3D-CT) scanning of the craniomaxillary region was performed after 2 and 4 weeks of nasal obstruction. Other rats were sacrificed, and MSCs were isolated from condylar tissues and cultured in vitro for examining the cell proliferation and expression of chondrogenic marker genes. Results Significant decreases in body weight were observed in the MB group compared with the NB group during 4 weeks of nasal obstruction. All mandibular parameters in the sagittal, vertical, and transverse dimensions (except bi-condylar width) measured via 3D-CT were significantly smaller in the MB group. No significant difference was found in the proliferative ability of cultured MSCs derived from condylar tissues between the two groups. However, the expression of chondrogenic marker genes Acan, Col2a1 and Sox9 was significantly lower in the MB group-derived MSCs, using Cell Counting Kit-8 and quantitative polymerase chain reaction. Conclusion The findings suggested that mouth breathing forced by nasal obstruction lead to developmental defects in the mandibular condyle, which might be related to the reduced cartilage differentiation of condylar MSCs.

Initial third molar development is delayed in jaws with short distal space: An early impaction sign? Publication date: October 2019 Source: Archives of Oral Biology, Volume 106 Author(s): D.F. Marchiori, G.V. Packota, J.C. Boughner Abstract Introduction The multifactorial aetiology of third molar (M3) impaction remains puzzling. While short retromolar jaw space is a well-established risk factor in young adults, space alone does not explain how impaction develops. Here we investigated in children and adolescents a potential new factor, delayed M3 development. We hypothesized that two important impaction correlates – lack of jaw space, and delayed M3 development – are also positively correlated with each other, and may together help suggest from early ages subsequent possible M3 impaction. Material and methods We studied 689 M3 regions on retrospective CBCT images of 179 Canadian patients aged 8–13 years, using multiple linear mixed-effects (LME) modeling to predict M3 development (using Demirjian’s stages) in both upper and lower jaws based upon available distal jaw space while also accounting for age, sex, jaw, and oral quadrant. Results Lack of distal jaw space was associated with less-developed M3s in upper and lower jaws (t = 3.209, p = .001). If space availability fell below a given threshold, absence of M3 development was predicted. Upper M3s developed more than lower M3s for each additional millimetre of retro-M1 space gained. Between-sex differences were not found. Conclusion Retromolar jaw space and M3 development are two positive correlates of M3 impaction that co-associate as early as childhood. Our results suggest that delayed initial M3 development merits investigation as a potential prognosticator for late M3 eruption relative to when jaw growth ceases – an outcome known to increase the likelihood of impaction.

The ATC/TTC haplotype in the Interleukin 8 gene in response to Gram-negative bacteria: a pilot study Publication date: Available online 24 July 2019 Source: Archives of Oral Biology Author(s): Suzane C. Pigossi, Giovana Anovazzi, Livia S. Finoti, Marcell C. de Medeiros, Tatiana Maria De Souza Moreira, Marcia P.A. Mayer, Cleslei Fernando Zanelli, Sandro Roberto Valentini, Carlos Rossa, Raquel M. Scarel-Caminaga ABSTRACT Objective The aim of this study was investigate the functionality of ATC/TTC (Hap-1) and ATT/TTC (Hap-2) Interleukin (IL) 8 gene haplotypes in the response of neutrophils to Gram-negative bacteria associated with periodontitis. Design Neutrophils were isolated by gradient centrifugation from whole peripheral blood of systemically healthy individuals presenting the two IL8 gene haplotypes. Neutrophils were stimulated with P.gingivalis, A.actinomycetemcomitans and PMA/ionomycin, and cytokine gene expression (RT-qPCR) and migration/chemotaxis (boyden chamber assay) were compared according to the presence of Hap-1 or Hap-2 haplotypes. Protein production was also evaluted in the multiplex assay using the mixed population of leukocytes present in the whole blood from the same individuals. The influence of these two haplotypes on the IL8 promoter activity was assessed in gene-reporter experiments. Results Hap-1 haplotype in neutrophils and leukocytes exacerbated the response to stimulation with Gram-negative bacteria, with higher levels of TNF-α (mRNA and protein), IL-1β, IL-2R and IFN-γ (protein) and with increased chemotaxis. Presence of the T allele at the rs4071 polymorphism (alias -251) was associated with increased activity of IL8 proximal promoter. Conclusions Neutrophils and leukocytes carrying the Hap-1 haplotype (ATC/TTC) in the IL8 gene present an enhanced response to stimulation with Gram-negative bacteria associated with periodontitis. Presence of the T allele (rs4073) in the IL8proximal promoter increases transcription activity.

A Novel Missense Mutation p.S305R of EDA Gene causes XLHED in a Chinese family Publication date: Available online 24 July 2019 Source: Archives of Oral Biology Author(s): Guannan Liu, Xin Wang, Man Qin, Lisha Sun, Junxia Zhu Abstract X-linked hypohidrotic ectodermal dysplasia (XLHED) can be characterized by hypohidrosis, sparse hair, hypodontia, and characteristic facial features and is usually caused by mutations of ectodysplasin A (EDA) gene located on the X chromosome. In this study, we examined a HED pedigree and studied the molecular genetics of the disease. A novel missense mutation was revealed by direct sequencing analysis in the EDA exon 7 (c.913 A > C, p.S305R). The function of the mutant EDA gene was subsequently confirmed by in vitro study in human embryonic kidney 293 T cells transfected with mutant or wild type forms of EDA. The mutant-type EDA1 protein showed impaired solubility comparing with wild-type EDA1. This novel missense EDA mutation was considered to be the cause of HED in the pedigree reported here. Our findings, combined with those reported elsewhere, provide an improved understanding of the pathogenic mechanism of HED as well as important information for a genetic diagnosis.