Gingival Epithelial Cells Research Articles

Cytotoxic effects of different detergent containing children's toothpastes on human gingival epithelial cells

BackgroundThis study aimed to evaluate possible cytotoxic effects to gingival epithelial cells exposed to children toothpastes containing different detergent.MethodsTissues required for the isolation of human gingival epithelial cells were obtained by biopsy during the extraction of the impacted third molar tooth. Toothpaste solutions of different concentrations were prepared from five different children’s toothpastes with different detergent contents. Isolated gingival epithelial cells were stimulated with experimental groups consisting of toothpaste solutions (Colgate, Sensodyne, Splat, Nenedent, Perlodent) at different concentrations and a control group consisting of complete Dulbecco’s modified eagle medium. After the experiments, cell viability was evaluated using flow cytometry. 2 Way ANOVA was used to see the interaction effect of the main effects of toothpaste solution and concentration factors. Pairwise comparisons were made by Tukey post hoc tests. In the study, the significance level was taken as 0.05.ResultsAs a result of the analysis, it was seen that the toothpaste solution and concentration factors and the interactions of these 2 factors were effective on the viable, early apoptotic, late apoptotic and necrotic cell rates. The statistically highest live cell ratios were detected in Splat’s toothpaste solutions (90.14% at 0.4% concentration) after the control group (90.82%) and the group with the lowest viability values was determined in Colgate group (75.74% at 0.4% concentration) (p < 0.05).ConclusionsAccording to the results of the study, it was observed that toothpastes containing SLS affected the viability of cells more negatively than toothpastes with other detergent contents.

The effects of mouthwashes in human gingiva epithelial progenitor (HGEPp) cells

ObjectivesThe gingiva epithelium accounts for a significant proportion of the surface around the tooth. An inflammatory reaction occurs in the presence of bacterial biofilm, adhesion is reduced, and the depth of the sulcus gingivalis increases. The most common antiseptic agents in oral rinses are chlorhexidine digluconate (CHX) and cetylpyridinium chloride. We examined long-lasting effects of residual concentrations of eight commercially available rinses. Our main goals were (i) to analyze the effect of different chemical compositions on cell proliferation, (ii) to examine apoptosis, and (iii) cell morphology on human epithelial progenitor cell line (HGEPp).Materials and methodsCell proliferation was measured in a real-time system (0–48 h) by impedimetry (xCELLigence). Apoptosis was measured with labeled Annexin-V (BD-FACScalibur).ResultsChanges in proliferation were measured at certain concentrations: (i) H2O2 proved to be cytotoxic at almost all concentrations; (ii) low concentrations of CHX (0.0001%; 0.0003%) were proliferation inducers, while higher concentrations were cytotoxic; (iii) for ClO2, advantageous proliferative effect was observed over a broad concentration range (0.06–6 ppm). In mouthwashes, additives in the formulation (e.g., allantoin) appeared to influence cellular responses positively. Apoptosis marker assay results suggested a low-level activation by the tested agents.ConclusionsMouthwashes and their reference compounds proved to have concentration-dependent cytotoxic effects on human gingival epithelial cells.Clinical relevanceA better understanding of the effects of mouthwashes and their reference compounds is particularly important. These concentration-dependent effects (cytotoxic or proliferation inducing) interfere with human cells physiology while being used in the fight against the pathogenic flora.

Macrophage migration inhibitory factor regulates specific innate immune sensor responses in gingival epithelial cells.

The gingival epithelium protects periodontal tissues and the alveolar bone by maintaining a steady state of regulated inflammatory surveillance, also known as healthy homeostasis. Accordingly, the repertoire of receptors present within the gingival epithelium showcases its ability to recognize microbial colonization and contribute to bacterial sensing. Macrophage migration inhibitory factor (MIF) is one of many cytokines that are expressed in this protective state and is involved in neutrophil regulation. However, its role in the maintenance of healthy gingival tissue has not been described. Gingival tissues from wild-type (WT) and Mif knock-out (KO) mice were stained for neutrophils and three key neutrophil chemoattractants: MIF, Gro-α/CXCL1, and Gro-β/CXCL2 in the junctional epithelium (JE). In addition, gene silencing studies were performed using gingival epithelial cells (GECs) to examine the role of MIF on transcription of key bacterial recognition receptors Toll-like receptors (TLR)-1, -2, -4, -6, -9 and interleukin-1 receptors (IL-1R1 and IL-1R2) in response to oral bacterial stimulation. WT murine gingival tissues demonstrated high expression of MIF in the JE. In Mif KO mice, despite the significant reduction of Gro-α/CXCL1 and Gro-β/CXCL2, there was a slight increase in neutrophils. Gene silencing experiments showed that MIF down-regulated the mRNA expression of TLR4, IL-1R1, and IL-1R2 in GEC, in addition to decreasing secreted IL-8/CXCL8 in response to bacteria. MIF regulates the expression of TLR4, IL-1Rs, and IL-8/CXCL8, components that are all involved in maintaining oral health. Our data demonstrate that MIF is a significant contributor to the maintenance of healthy oral homeostasis.

Periodontal Pathogen Adhesion, Cytotoxicity, and Surface Free Energy of Different Materials for an Implant Prosthesis Screw Access Hole.

Background and Objectives: Oral implant restorations are an excellent treatment option for edentulous patients; however, periodontopathogenic bacteria have been found in the microgaps between implant−abutment junctions. Implant designs to limit the microgaps have been extensively studied. However, studies have shown microgaps continue to exist, allowing for the leakage of bacteria into the implant system. Screw access hole materials are used to fill the access hole void. The use of materials with beneficial properties could provide bacterial leakage prevention. The aim of this study was to examine the surface free energy, cytotoxicity, and bacterial adhesion of selected screw access hole materials such as cotton, polytetrafluoroethylene (PTFE) tape, paraffin wax−polyolefin thermoplastic (PF), paraffin wax (Wax), gutta-percha (GP), and caviton EX (CE). Materials and Methods: A sessile drop test was performed to observe the contact angle and calculate the surface free energy of each material in order to determine the level of hydrophobicity. Cytotoxicity was examined in a mouse gingival epithelial cell line for day 1 and day 3. Bacterial adhesion was tested with Porphyromonas gingivalis, Fusobacterium nucleatum, and Aggregatibacter actinomycetemcomitans. Results: PTFE, PF, and wax presented low surface free energies of 19.34, 23.041, and 24.883 mN.m-1, respectively. No cytotoxicity was observed, except for GP and CE. Concurrently, the bacterial adhesion was also the lowest in PTFE and PF. Conclusions: Within the limits of this study, PTFE and PF showed an excellent biocompatibility with few bacterial adhesions. These materials could be potential screw access hole materials in clinical settings.

Interleukin-1β regulates odontogenic ameloblast-associated protein gene transcription in human gingival epithelial cells.

Junction epithelium (JE) is located apical to the bottom of the gingival sulcus and binds enamel to hemidesmosomes to protect the periodontal tissue from bacterial infection. Function of odontogenic ameloblast-associated protein (ODAM) is suggested by its expression sites (JE and maturation stage ameloblasts) to be involved in the adhesion between the JE and enamel, and odontogenesis. To analyze the changes in ODAM gene and protein expressions in inflamed gingiva, Ca9-22 gingival epithelial cells were stimulated with 1ng/ml interleukin-1β (IL-1β) for 3-24h, and ODAM mRNA and protein levels were analyzed by real-time PCR and Western blotting. Luciferase (LUC) constructs were made ligating various lengths of human ODAM gene promoters and performed LUC analyses in Ca9-22 cells. Gel shift and chromatin immunoprecipitation (ChIP) assays were performed. IL-1β induced ODAM mRNA and protein levels at 6-24h. IL-1β increased LUC activities of the ODAM gene promoter constructs from - 85 to - 950. These activities were blocked by protein kinase A, tyrosine kinase, mitogen-activated protein (MAP) kinase kinase and phosphoinositide 3-kinase inhibitors. Gel shift and ChIP assays showed that IL-1β induced CCAAT/enhancer-binding protein (C/EBP) β and Yin Yang1 (YY1) binding to C/EBP1, 2, 3, and YY1 elements. These data indicate that IL-1β stimulates ODAM gene transcription mediated through C/EBP1, C/EBP2, C/EBP3, and YY1 elements in the human ODAM gene promoter.

Antibacterial and anti-biofilm activities of probiotic Lactobacillus curvatus BSF206 and Pediococcus pentosaceus AC1-2 against Streptococcus mutans

Streptococcus mutans (S. mutans) is the most important oral pathogenic bacterial cause of dental caries. Here we investigated the abilities of probiotic lactobacillus strains of Lactobacillus curvatus (L. curvatus) BSF206 and Pediococcus pentosaceus (P. pentosaceus) AC1-2 to control S. mutans. Both probiotic strains are acid and bile salt tolerant and are resistant to hydrogen peroxide and lysozyme to promote their survival within the oral environment. In addition, both strains are highly hydrophobic and are also capable of engaging in electrostatic interactions. These properties enhance abilities of both strains to adhere to gingival epithelial cells and HT-29 for improved colonization of oral tissues, while also enabling these probiotics auto-aggregate and to form aggregates with S. mutans that both may prevent S. mutans from colonizing oral tissues and facilitate the clearance of the cariogenic bacteria from the mouth during swallowing of food and saliva. Furthermore, results presented herein revealed that L. curvatus BSF206 and P. pentosaceus AC1-2 effectively inhibited S. mutans activities (biofilm formation, secretion of extracellular matrix components, synthesis of water-insoluble glucans) and led to downregulation of expression of key S. mutans genes involved in biofilm production (gtfA, gtfB, ftf, brpA). Taken together, these results indicate that L. curvatus BSF206 and P. pentosaceus AC1-2 can inhibit S. mutans biofilm formation as a new strategy for preventing dental caries.

TNF-α regulates the composition of the basal lamina and cell-matrix adhesions in gingival epithelial cells

ABSTRACT Laminin 5, type 4 collagen, and α6β4 integrin contribute to the formation of hemidesmosomes in the epithelia of periodontal tissues, which is critical for the development and maintenance of the dentogingival junction. As it is not known whether TNF-α alters the composition of the epithelial pericellular matrix, human gingival epithelial cells were cultured in the presence or absence of TNF-α. Treatment with TNF-α accelerated epithelial cell migration and closure of in vitro wounds. These data indicate unexpectedly, that TNF-α promotes the formation of the pericellular matrix around epithelial cells and enhances adhesion of epithelial cells to the underlying matrix, properties which are important for cell migration and the integrity of the dentogingival junction.

Tyrosine Kinases and Phosphatases: Enablers of the Porphyromonas gingivalis Lifestyle.

Tyrosine phosphorylation modifies the functionality of bacterial proteins and forms the basis of a versatile and tunable signal transduction system. The integrated action of tyrosine kinases and phosphatases controls bacterial processes important for metabolism and virulence. Porphyromonas gingivalis, a keystone pathogen in periodontal disease, possesses an extensive phosphotyrosine signaling network. The phosphorylation reaction is catalyzed by a bacterial tyrosine (BY) kinase, Ptk1, and a Ubiquitous bacterial Kinase UbK1. Dephosphorylation is mediated by a low-molecular-weight phosphatase, Ltp1 and a polymerase and histidinol phosphatase, Php1. Phosphotyrosine signaling controls exopolysaccharide production, gingipain activity, oxidative stress responses and synergistic community development with Streptococcus gordonii. Additionally, Ltp1 is secreted extracellularly and can be delivered inside gingival epithelial cells where it can override host cell signaling and readjust cellular physiology. The landscape of coordinated tyrosine kinase and phosphatase activity thus underlies the adaptive responses of P. gingivalis to both the polymicrobial environment of bacterial communities and the intracellular environment of gingival epithelial cells.

Aggregatibacter actinomycetemcomitans Outer Membrane Proteins 29 and 29 Paralogue Induce Evasion of Immune Response.

Aggregatibacter actinomycetemcomitans (Aa) is abundant within the microbial dysbiotic community of some patients with periodontitis. Aa outer membrane protein 29 (OMP29), a member of the OMPA family, mediates the invasion of Aa to gingival epithelial cells (GECs). This study evaluated the effect of OMP29 and its paralogue OMP29par on the response of GECs to Aa. The omp29 or/and omp29par deletion mutants AaΔ29, AaΔ29P, and AaΔ29Δ29P were constructed, and recombinant Aa OMP29His was obtained. Microarray analysis and the evaluation of cxcl-8 gene expression were performed to examine the response of GECs line OBA-09 to Aa and its mutants. The expression of cxcl-8 and its product CXCL-8 was examined in LPS-stimulated OBA-09 cells with Aa OMP29His. Proteomics analysis showed that the deletion of omp29 led to overexpression of both OMP29par and another membrane protein OMP39, the expression of which was further increased in AaΔ29Δ29P. OBA-09 cells challenged with AaΔ29Δ29P exhibited a higher expression of cxcl-8 in comparison to wildtype Aa strain AaD7S or single-deletion mutants AaΔ29 or AaΔ29P. LPS-stimulated OBA-09 cells challenged with Aa OMP29His showed reduced expressions of cxcl-8 and its product CXCL-8. OBA-09 cells challenged with AaΔ29Δ29P in comparison to Aa strain AaD7S resulted in higher expressions of genes involved in apoptosis and inflammatory response such as bcl2, birc3, casp3, c3, ep300, fas, fosb, grb2, il-1α, il-1β, il-6, cxcl-8, nr3c1, prkcq, socs3, and tnfrsf1β and reduced expressions of cd74, crp, faslg, tlr1, and vcam1. The results suggested a novel strategy of Aa, mediated by OMP29 and OMP29par, to evade host immune response by inhibiting CXCL-8 expression and modulating the genes involved in apoptosis and inflammatory response in GECs. Pending further confirmation, the strategy might interfere with the recruitment of neutrophils and dampen the host inflammatory response, leading to a more permissive subgingival niche for bacterial growth.

The neutrophil elastase-upregulated placenta growth factor promotes the pathogenesis and progression of periodontal disease.

Periodontal disease is a chronic inflammatory disease. Given its high prevalence, especially in aging population, the detailed mechanisms about pathogenesis of periodontal disease are important issues for study. Neutrophil firstly infiltrates to periodontal disease-associated pathogen loci and amplifies the inflammatory response for host defense. However, excessive neutrophil-secreted neutrophil elastase (NE) damages the affected gingival. In lung and esophageal epithelium, NE had been proved to upregulate several growth factors including placenta growth factor (PGF). PGF is an angiogenic factor with proinflammatory properties, which mediates the progression of inflammatory disease. Therefore, we hypothesize excessive NE upregulates PGF and participates in the pathogenesis and progression of periodontal disease. In gingival epithelial cells (GEC), growth factors array demonstrated NE-increased growth factors and further be corroborated by Western blot assay and ELISA. The GEC inflammation was evaluated by ELISA. In mice, the immunohistochemistry results demonstrated ligature implantation-induced neutrophil infiltration and growth factor upregulation. By multiplex assay, the ligature-induced proinflammatory cytokines level in gingival crevicular fluid (GCF) were evaluated. Finally, alveolar bone absorption was analyzed by micro-CT images and H &amp; E staining. NE upregulated PGF expression and secretion in GEC. PGF promoted GEC to secret IL-1β, IL-6, and TNF-α in GCF In periodontal disease animal model, ligature implantation triggered NE infiltration and PGF expression. Blockade of PGF attenuated the ligature implantation-induced IL-1β, IL-6, TNF-α and MIP-2 secretion and ameliorated the alveolar bone loss in mice. In conclusion, the NE-induced PGF triggers gingival epithelium inflammation and promotes the pathogenesis and progression of periodontal disease.

Effects of grape seed proanthocyanidins on the expression of inflammatory mediators in gingival epithelial cells.

This study aims to determine the effect of grape seed proanthocyanidin (GSP) pretreatment on lipopolysaccharide (LPS)-induced inflammation of human gingival epithelial cells (HGECs). HGECs were cultivated with different concentrations of GSPs (0, 1, 5, 10, 20, 40, 60, 80, 100 μg·mL-1) for 6, 12, 24, and 48 h. CCK-8 was used to detect the proliferation activity of HGECs. HGECs were treated with different concentrations of GSPs (0, 10, 20, and 40 μg·mL-1) for 24 h and then cultured with 1.0 μg·mL-1 LPS. Enzyme-linked immunosorbent assay (ELISA) was used to detect the expression levels of pro-inflammatory cytokine tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6 and anti-inflammatory cytokines IL-4, IL-10, and transforming growth factor-β (TGF-β). Quantitative real-time polymerase chain reaction (QRT-PCR) was used to detect the mRNA expression levels of TNF-α, IL-1β, IL-6, IL-4, IL-10, and TGF-β. When the GSP concentration was 0-40 μg·mL-1, the cell proliferation had no significant difference. When the action time reached 24 h, the cell proliferation was the highest. The results of ELISA and QRT-PCR showed that 10, 20, and 40 μg·mL-1 GSPS decreased the expression levels of TNF-α, IL-1β, and IL-6 (P<0.05) and increased the expression levels of IL-4, IL-10, and TGF-β compared with 0 μg·mL-1 GSPS (P<0.05). GSPS (0-40 μg·mL-1) has no significant effect on the proliferation activity of HGECs. Pretreatment with GSPS can inhibit the expression of pro-inflammatory factors and enhance the expression of anti-inflammatory factors. Hence, GSPS has a certain preventive effect on the resistance of HGECs to the stimulation of endotoxin.

Porphyromonas gingivalis upregulates calbindin 1 and thus promotes the proliferation of gingival epithelial cells.

This study aims to investigate the effect of calbindin 1 on the proliferation and apoptosis of gingival epithelial cells affected by Porphyromonas gingivalis (P. gingivalis)invitro. A model of P. gingivalis infecting CA9-22 was established in vitro. At 24 h after infection, the expression of calbindin 1 (CALB1) was detected by real-time fluorescent quantitative polymerase chain reaction, Western blot, and immunofluorescence analyses. The expression of CALB1 was further inhibited by RNA interference. Cell proliferation was detected by BrdU analysis, and cell apoptosis was detected by caspase 3 activity. The expression of MDM2 and p53 was detected by Western blot analysis. P. gingivalis infection upregulated the expression of CALB1 in CA9-22 cells with multiplicity-dependent manner. CALB1 promoted the proliferation of CA9-22 cells, increased the expression of MDM2, and inhibited the expression of p53. Inhibiting CALB1 expression did not affect the inhibitory effect of P. gingivalis infection on CA9-22 apoptosis. P. gingivalis infection can promote the proliferation of CA9-22 cells by increasing CALB1 expression. The related mechanism may be associated with MDM2-p53.

A glutathione-responsive silica-based nanosystem capped with in-situ polymerized cell-penetrating poly(disulfide)s for precisely modulating immuno-inflammatory responses

HypothesisPrecise modulation of immuno-inflammatory response is crucial to control periodontal diseases and related systemic comorbidities. The present nanosystem with the controlled-release and cell-penetrating manner enhances the inflammation modulation effects of baicalein in human gingival epithelial cells (hGECs) for better oral healthcare. ExperimentsWe constructed a red-emissive mesoporous silica nanoparticle-based nanosystem with cell-penetrating poly(disulfide) (CPD) capping, through a facile in-situ polymerization approach. It was featured with a glutathione-responsive manner and instant cellular internalization capacity for precisely delivering baicalein intracellularly. Laboratory experiments assessed whether and how the nanosystem per se with the delivered baicalein could modulate immuno-inflammatory responses in hGECs. FindingsThe in-situ polymerized CPD layer capped the nanoparticles and yet controlled the release of baicalein in a glutathione-responsive manner. The CPD coating could facilitate cellular internalization of the nanosystem via endocytosis and thiol-mediated approaches. Notably, the intracellularly released baicalein effectively downregulated the expression of pro-inflammatory cytokines through inhibiting the NF-κB signaling pathway. The nanosystem per se could modulate immuno-inflammatory responses by passivating the cellular response to interlukin-1β. This study highlights that the as-synthesized nanosystem may serve as a novel multi-functional vehicle to modulate innate host response via targeting the NF-κB pathway for precision healthcare.

In vitro and in vivo study of the pathogenic role of PPARα in experimental periodontitis.

Objective The purpose of this study is to investigate the pathogenic role of PPARα in periodontal antigen treated gingival cells in vitro and in experimental periodontitis in vivo .Methodology Gingival fibroblasts, gingival epithelial cells and splenocytes were isolated from C57BL/6J wild type (WT) mice and treated with fixed P. gingivalis at for 48 hours. The mRNA levels of PPARs, TNFα, IL-1β and IL-10 were detected by Real-time quantitative PCR. Silk ligatures after being soaked in the P.gingivalis suspension were tied around both maxillary second molars of WT mice or PPARα knock-out (KO) mice for two weeks. PPARα agonist fenofibrate and vehicle control were injected into the different side of the palatal gingiva on days 3, 6, and 9. At day 14, bone resorption and gingival mRNA expression levels of PPARs, TNFα, IL-1β and IL-10 were measured by micro-computed tomography and RT-qPCR respectively.ResultsP. gingivalis treatment downregulated the expression of PPARα, but not PPARβ or PPARγ, and increased the expression of TNF-α and IL-1β in Gingival fibroblasts, gingival epithelial cells and splenocytes from WT mice. Gingival mRNA levels of PPARα were significantly decreased in experimental periodontitis in WT mice. The bone loss of PPARα KO mice in experimental periodontitis was significantly higher than WT mice and was not reduced by fenofibrate treatment. Gingival TNFα protein expressions were significantly increased by P. gingivalis associated ligation and decreased by fenofibrate treatment in WT mice but not in PPARα KO mice.Conclusion This study suggests that PPARα plays an essential role in periodontitis.

Inflammasome Activation in Gingival Epithelial Cells.

Inflammasomes are multiprotein complexes that assemble in host cells upon recognition of infection or danger via pattern recognition receptors and/or danger recognition receptors. The assembly of inflammasomes results in the activation of caspase-1 and is followed by the enzymatic maturation and secretion of inflammatory cytokines like interleukin 1β (IL-1β) and IL-18.In the oral cavity, gingival epithelial cells (GECs) line the mucosa and have a barrier role for invading pathogens. In these cells, the NLRP3 inflammasome is the best studied and has been shown to play a role in the inflammatory immune response against a variety of oral pathogens. As such, in order to study gingivitis and other oral pathologic inflammatory conditions, studying the activation of inflammasomes is important. The simplest way to detect inflammasome activation is to detect the activated caspase-1, as well as the secretion of mature IL-1β and IL-18, via ELISA, Western blot, and immunofluorescence techniques.Here we describe the detection of NLRP3 inflammasome activation by the oral pathogen Porphyromonas gingivalis in human GECs via these three methods and mention other methods and techniques that we have successfully used together with these in our quest to understand the host-pathogen interaction.

In vitro biocompatibility testing of different base materials used for elevation of proximal subgingival margins using human gingival epithelial cells.

To analyze the biological effects of four base materials used for elevation of proximal subgingival margins on gingival epithelial cells. Twenty-eight specimens for each of the four base materials (total 112 specimens) were used: resin-modified glass ionomer (RMGI), glass hybrid (HV-GIC), flowable bulk fill resin composite (Bulk Flow) and bioactive ionic resin (Activa). Proximal enamel and root dentin were used as controls. Gingival epithelial cell viability was calculated after direct incubation on all four types of material for either 24 h or 72 h using both the methyl tetrazolium and trypan blue dye exclusion assays. Data were analyzed statistically using one-way analysis of variance, Tukey post hoc test and independent sample t-test (P < 0.05). Cell viability values in both assays showed significant differences among the study groups. Bulk Flow showed the highest values, followed in order by Activa and the control groups. Both HV-GIC and RMGI had the lowest values. Cell viability in all of the study groups was higher after incubation for 72 h than after 24 h. In terms of biocompatibility with epithelial tissues, bulk fill resin composite appears to be most suitable, followed by bioactive composite, for subgingival placement than glass ionomer-based materials, especially that containing 2-hydroxy-ethyl methacrylate.

Comparative assessment of exfoliated gingival cells in healthy individuals and chronic periodontitis patients with and without tobacco chewing habit: A cytomorphometric study.

Background:Cytomorphometry is the analysis of the morphometry of exfoliated cells. Studies on exfoliated gingival epithelial cells in chronic periodontitis were not evident on the literature search. In the backdrop of this information, we were inspired to conduct a cytomorphometric study.Aim:The aim of the study is to correlate the nuclear area (NA), cytoplasm area (CA), and the nuclear: cytoplasm ratio (N/C) in the gingival cells of healthy individuals, patients with chronic periodontitis, and chronic periodontitis patients who are tobacco chewers.Materials and Methods:Ninety subjects, aged 35–45 years, were categorized into three equal groups of 30 each. Group I had individuals with healthy gingiva, Group II patients with chronic periodontitis without any deleterious habits, while Group III patients with chronic periodontitis and tobacco chewing habit. Exfoliated gingival cells were stained with Papanicolaou stain. Analysis of cells was done using binocular light microscope under ×40 and i-progress® image analyzer software.Results:There was statistically significant increase in CA in all the groups, but NA and N/C ratio did not show any statistical significant difference. When Group II was compared with Group III, we found significant correlation of CA with pocket probing depth and clinical attachment loss (CAL) in Group II. Similarly, we also found a correlation of CA and N/C with pocket depth and CA with CAL in Group III.Conclusion:The results suggest that molecular level changes occur in the cells. Cytomorphometry can be used to analyze the cellular changes in chronic periodontitis patients even before clinical signs manifest. Hence, it can be used as an investigative tool.

Cannabinoid control of gingival immune activation in chronically SIV-infected rhesus macaques involves modulation of the indoleamine-2,3-dioxygenase-1 pathway and salivary microbiome.

SummaryBackgroundHIV/SIV-associated periodontal disease (gingivitis/periodontitis) (PD) represents a major comorbidity affecting people living with HIV (PLWH) on combination anti-retroviral therapy (cART). PD is characterized by chronic inflammation and dysbiosis. Nevertheless, the molecular mechanisms and use of feasible therapeutic strategies to reduce/reverse inflammation and dysbiosis remain understudied and unaddressed.MethodsEmploying a systems biology approach, we report molecular, metabolome and microbiome changes underlying PD and its modulation by phytocannabinoids [delta-9-tetrahydrocannabinol (Δ9-THC)] in uninfected and SIV-infected rhesus macaques (RMs) untreated (VEH-untreated/SIV) or treated with vehicle (VEH/SIV) or Δ9-THC (THC/SIV).FindingsVEH- untreated/SIV but not THC/SIV RMs showed significant enrichment of genes linked to anti-viral defense, interferon-β, NFκB, RIG-1, and JAK-STAT signaling. We focused on the anti-microbial DUOX1 and immune activation marker IDO1 that were reciprocally regulated in the gingiva of VEH-untreated/SIV RMs. Both proteins localized to the gingival epithelium and CD163+ macrophages, and showed differential expression in the gingiva of THC/SIV and VEH/SIV RMs. Additionally, inflammation-associated miR-21, miR-142–3p, miR-223, and miR-125a-5p showed significantly higher expression in the gingiva of VEH/SIV RMs. In human primary gingival epithelial cells, miR-125a-5p post-transcriptionally downregulated DUOX1 and THC inhibited IDO1 protein expression through a cannabinoid receptor-2 mediated mechanism. Interestingly, THC/SIV RMs showed relatively reduced plasma levels of kynurenine, kynurenate, and the neurotoxic quinolinate compared to VEH/SIV RMs at 5 months post SIV infection (MPI). Most importantly, THC blocked HIV/SIV-induced depletion of Firmicutes and Bacteroidetes, and reduced Gammaproteobacteria abundance in saliva. Reduced IDO1 protein expression was associated with significantly (p<0.05) higher abundance of Prevotella, Lactobacillus (L. salivarius, L. buchneri, L. fermentum, L. paracasei, L. rhamnosus, L. johnsonii) and Bifidobacteria and reduced abundance of the pathogenic Porphyromonas cangingivalis and Porphyromonas macacae at 5MPI.InterpretationThe data provides deeper insights into the molecular mechanisms underlying HIV/SIV-induced PD and more importantly, the anti-inflammatory and anti-dysbiotic properties of THC in the oral cavity. Overall, these translational findings suggest that phytocannabinoids may help reduce gingival/systemic inflammation, salivary dysbiosis and potentially metabolic disease/syndrome in PLWH on cART and those with no access to cART or do not suppress the virus under cART.FundingResearch reported in this publication was supported by the National Institutes of Health Award Numbers R01DA052845 (MM and SNB), R01DA050169 (MM and CO), R01DA042524 and R56DE026930 (MM), and P51OD011104 and P51OD011133. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.

Influence of Anodized Titanium Surfaces on the Behavior of Gingival Cells in Contact with: A Systematic Review of In Vitro Studies

Electrochemically anodized (EA) surfaces promise enhanced biological properties and may be a solution to ensure a seal between peri-implant soft tissues and dental transmucosal components. However, the interaction between the modified nano-structured surface and the gingival cells needs further investigation. The aim of this systematic review is to analyze the biological response of gingival cells to EA titanium surfaces in in vitro studies with a score-based reliability assessment. A protocol aimed at answering the following focused question was developed: “How does the surface integrity (e.g., topography and chemistry) of EA titanium influence gingival cell response in in vitro studies?”. A search in three computer databases was performed using keywords. A quality assessment of the studies selected was performed using the SciRAP method. A total of 14 articles were selected from the 216 eligible papers. The mean reporting and the mean methodologic quality SciRAP scores were 87.7 ± 7.7/100 and 77.8 ± 7.8/100, respectively. Within the limitation of this review based on in vitro studies, it can be safely speculated that EA surfaces with optimal chemical and morphological characteristics enhance gingival fibroblast response compared to conventional titanium surfaces. When EA is combined with functionalization, it also positively influences gingival epithelial cell behavior.

Anti-bacterial Efficacy of Probiotics used in Provisional Polymethyl Methacrylate Crowns against Porphyromonos gingivalis – an In vitro Study

Aim: To evaluate the effect of Probiotic used in PMMA Temporary restorations on gingival inflammatory response by its action on the marginal gingival epithelial cells.&#x0D; Materials and Methods: This is an in vitro Interventional pilot study using discs of PMMA ( Group A ) and PMMA incorporated with Probiotics (Group B . The effect of probiotics incorporated PMMA on strains of Porphyromonos gingivalis grown on culture plates will be assessed using zone of inhibition test and minimum inhibitory concentration was assessed . Data was recorded, tabulated and statistically evaluated using SPSS software.&#x0D; Results: The zone of Inhibition in the Group B was (Mean = 16.30mm) is comparatively higher than that of Group A with mean value 12.92mm. Results states that among the various concentration of Probiotic Lozenges (2.5, 5, 10, 15 and 20 µg/ml) used to determine the antibacterial activity , Highest mean zone of inhibition was observed with 20 µg/ml with 16 mm, followed by 15 µg/ml with 14mm , 10 µg/ml with 9mm, 5 µg/ml with 11 mm &amp; with the lowest inhibition observed in 2.5 µg/ml with 8mm. There is significant correlation between concentration of probiotics and its antibacterial efficacy. The increase in concentration of probiotics in the PMMA discs is directly proportional to its antibacterial efficacy against P. gingivalis.&#x0D; Conclusion: Thus the study proved a significant correlation between Probiotic used in PMMA Temporary crowns and reduction in gingival Inflammation at the margins of the restoration which will be beneficial when used in crowns used in patients undergoing Orthodontic treatment or Immediate Implant Loading with acrylic crowns.