A Case of Doxycycline-induced Melanin in the Gingiva Tissue: Case Report.

BACKGROUND: Gingival pigmentation is a discoloration of the gingiva due to a variety of lesions and conditions associated with several endogenous and exogenous etiologic features. OBJECTIVE: The purpose of this study is to describe a report of gingival pigmentation in a patient who used doxycycline. CASE REPORT: A 21-year-old Caucasian female was under dermatological treatment and antibiotic therapy with doxycycline 100 mg (one time a day) for 90 days. She presented brown pigmentation at the gingival margin on the facial surfaces of the upper and lower anterior incisors and premolars. The patient was evaluated by immunohistochemical (S-100, Melan-A, and HMB-45) and histopathologic analyses, and clinical history. Blood levels of malondialdehyde (MDA), glutathione (GSH), and superoxide dismutase (SOD) were analyzed by UV/Vis spectroscopy. The adrenaline, noradrenaline, and dopamine in blood were analyzed by high-performance liquid chromatography (HPLC); dehidroepiandrosterone (DHEA) in serum by radioimmunoassay; and luteinizing hormone (LH) and 25-Hydroxyvitamin D by chemiluminescence. Hematoxylin-eosin stained sections revealed keratinocytes with pigment compatible with melanin. The Fontana-Masson staining was positive in melanophages and in some basal keratinocytes. S-100, Melan A and HMB-45 were confirmed as positive markers of melanocytic differentiation in gingival tissue. We observed a significant increase in malondialdehyde (p˂0.05) and a decrease in superoxide dismutase levels (p˂0.05). The dopamine value was found to be 15 pg/ml (reference value ≤ 10 pg/ml). CONCLUSION: The use of doxycycline is associated with an increase in oxidative stress and of dopamine with melanin pigments in the gingival tissue. This case report showed a cause-effect relationship between exposure to doxycycline and pigmentation of the marginal gingiva.

Physicochemical and in vitro digestibility properties on complexes of fermented wheat starches with konjac gum.

In this study, the wheat starch with natural fermentation for 72 h was combined with konjac gum (KGM) at different concentrations (0, 0.1, 0.3, 0.5%, w/w), and the changes in physicochemical and digestible characteristics of the complexes were investigated. The results showed that KGM clumped fermented starch (FS) granules together and caused the FS gels to form a close network structure. The addition of KGM significantly decreased the amylose content and swelling power, and reduced peak viscosity, final viscosity, and setback value (SB), which indicated that FS-KGM complexes possessed soft gel structure and could resist the short-term retrogradation. KGM impeded the increase of relative crystallinity, retrogradation enthalpy and gel firmness of FS during storage, suggesting the long-term retrogradation of FS was retarded by KGM. All starch pastes had a weak gel-like structure, and higher storage modulus (G') and loss tangent (tan δ) values obtained after the addition of KGM. In vitro digestion results showed that KGM could slow the hydrolysis of FS, resulting in the increase of slowly digested starch (SDS) and resistant starch (RS). In particularly, the FS-0.3KGM showed the ideal structure, the best anti-retrogradation effected, and slowest the hydrolysis.

Knockdown of MALAT1 Inhibits the Progression of Chronic Periodontitis via Targeting miR-769-5p/HIF3A Axis.

PURPOSE: Chronic periodontitis (CP) is a long-lasting inflammatory disease that seriously affects oral health. This study is aimed at investigating the regulatory mechanism of metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in CP. METHODS: Primary human periodontal ligament cells (PDLCs) were treated with P. gingivalis lipopolysaccharide (LPS) to establish a CP model. Quantitative real-time PCR (qRT-PCR) was used to measure the expression of MALAT1 and miR-769-5p in gingival tissues of patients with CP and LPS-treated PDLCs. Cell viability was detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of inflammatory cytokines. The protein levels of caspase-3, Bax, Bcl-2, and hypoxia-inducible factor (HIF) 3A were determined by western blot assay. Dual-luciferase reporter (DLR) assay was applied to validate the target relationships between miR-769-5p and MALAT1/HIF3A. RESULTS: The expression of MALAT1 and HIF3A was enhanced, and the expression of miR-769-5p was reduced in gingival tissues of patients with CP and LPS-treated PDLCs. MALAT1 knockdown promoted cell viability and inhibited inflammation and cell apoptosis in LPS-treated PDLCs. MALAT1 targeted miR-769-5p and negatively regulated miR-769-5p expression. miR-769-5p overexpression promoted cell viability and inhibited inflammation and cell apoptosis in LPS-treated PDLCs. Besides, miR-769-5p targeted HIF3A and negatively modulated HIF3A expression. Both miR-769-5p inhibition and HIF3A overexpression reversed the inhibitory effects of MALAT1 silencing on LPS-induced PDLC injury in vitro. CONCLUSION: MALAT1 knockdown attenuated LPS-induced PDLC injury via regulating the miR-769-5p/HIF3A axis, which may supply a new target for CP treatment.

Mandibular biomechanical behavior of rats submitted to chronic intermittent or continuous hypoxia and periodontitis.

BACKGROUND: The aim of this study was to investigate the effects of exposure to continuous (CH) and intermittent (IH) hypoxia on biomechanical properties of the mandible and periodontal tissue of animals submitted to experimental periodontitis (EP) when applying loads in a hypoxic environment. METHODS: Adult female Wistar rats were exposed during 90 days to IH or CH (simulated high altitude of 4200 m above sea level). Fourteen days prior to the euthanasia, EP was induced to half of the animals of each group. RESULTS: Only in the rats with EP, IH decreased the maximum capacity of the mandible to withstand load and the limit of elastic load. Indicators of intrinsic properties of the bone material were significantly reduced by both types of hypoxia in rats with EP. Hypoxia enhanced the alveolar bone loss induced by EP in the buccal side of the mandible, without showing additional effects in lingual or interradicular bone. Hypoxia increased prostaglandin E2 content in gingival tissue of healthy animals and further elevated the E2 levels increased by EP. CONCLUSIONS: When periodontitis is present, hypoxic stress induces a decrease in mineral properties that ultimately affects the ability of the mandible to resist load, mainly during intermittent exposure to hypoxia. These effects on bone may be related to the higher levels of prostaglandin E2 reached in the surrounding gingival tissue. The findings of this study may stimulate strategies to prevent unwanted effects of hypoxia on periodontal tissues.

Evaluation of the Sealing Ability of Direct versus Direct-Indirect Veneer Techniques: An In Vitro Study.

BACKGROUNDS: Marginal discoloration, microleakage, wear, and marginal fractures are all prevalent problems with composite veneers, and this scenario leads the esthetic outcome to deteriorate with time, resulting in patient discontent. Aim of the Study. The study's goal was to determine the marginal sealing ability of composite laminate veneers when employing two types of veneer techniques: direct and direct-indirect veneers, as well as two types of composite resin: nanohybrid and microfilled composite resin restorations, using dye penetration method. MATERIALS AND METHODS: In this study, forty extracted human teeth were utilized. Following a standardized veneer preparation on the labial surface of the teeth, they were separated into two groups of 20 teeth each, using the following composite application techniques: group A: direct veneers and group B: direct-indirect veneers. Following that, each major group was separated into two subgroups of ten teeth each, based on the type of composite employed: subgroup 1 used nanohybrid composite resin, while subgroup 2 used microfilled composite resin. All of the samples were kept in distilled water, thermocycled, and soaked in 2% basic fuchsine dye. These specimens were sectioned and examined under a stereomicroscope for dye penetration at the gingival margin. The data was analyzed using independent T-tests using SPSS 22. RESULT: Using direct-indirect veneer technique with nanohybrid composite resin material resulted in the most negligible dye penetration at the gingival margin, while using direct veneer technique with microfilled composite resin material resulted in the maximum dye penetration. For both composite materials, gingival microleakage was lower when using the direct-indirect veneer technique than when using the direct technique, and the difference was statistically significant (P < 0.05). In both techniques, gingival microleakage was lower with nanohybrid composite than with microfilled composite, and the difference was statistically highly significant (P = 0.001). CONCLUSION: The sealing ability of the gingival margin of tooth/composite interface is better when applying direct-indirect veneer technique with nanohybrid composite resin than that of direct veneer technique with microfilled composite resin material.

Hydrogels based on low-methoxyl amidated citrus pectin and flaxseed gum formulated with tripeptide glycyl-l-histidyl-l-lysine improve the healing of experimental cutting wounds in rats.

Hydrogels based on natural and modified polysaccharides represent growing group of suitable matrices for the construction of effective wound healing materials. Bioactive tripeptide glycyl-l-histidyl-l-lysine and amino acid α-l-arginine are known to accelerate wound healing and skin repair. In this study, hydrogels based on low-methoxyl amidated citrus pectin or flaxseed gum were prepared and used for the transport of these healing agents to the experimental cutting wounds affected by extensive skin damage. Fourier-transform infrared spectroscopy, rheology, differential scanning calorimetry, scanning electron microscopy, swelling and release tests confirmed that these hydrogels differed in structure and physical properties. The cationic tripeptide was found to bind to carboxylic groups in LMA pectin, and the C3OH hydroxyl and ring oxygen O5 are involved in this interaction. The pectin hydrogel showed high viscosity and strong elastic properties, while the flaxseed gum hydrogel was characterised as a viscoelastic system of much lower viscosity. The former hydrogel released the drugs very slowly, while the latter hydrogel demonstrated zero order releasing kinetics optimal for drug delivery. In the in vivo wound healing testing on rats, both polysaccharide hydrogels improved the healing process mediated by the mentioned biomolecules. The tripeptide applied in the hydrogels showed significantly higher healing degree and lower healing time than in the control animals without treatment and when it was applied in an aqueous solution. Despite the absence of a synergistic effect, the mixture of the tripeptide and α-l-arginine in the hydrogels was also quite effective in wound healing. According to histological analysis, complete healing was achieved only when using the tripeptide in the flaxseed gum hydrogel. These observations might have an important prospect in clinical application of polysaccharide hydrogels.

A therapeutic oxygen carrier isolated from Arenicola marina decreased P. gingivalis induced inflammation and tissue destruction.

The control of inflammation and infection is crucial for periodontal wound healing and regeneration. M101, an oxygen carrier derived from Arenicola marina, was tested for its anti-inflammatory and anti-infectious potential based on its anti-oxidative and tissue oxygenation properties. In vitro, no cytotoxicity was observed in oral epithelial cells (EC) treated with M101. M101 (1 g/L) reduced significantly the gene expression of pro-inflammatory markers such as TNF-α, NF-κΒ and RANKL in P. gingivalis-LPS stimulated and P. gingivalis-infected EC. The proteome array revealed significant down-regulation of pro-inflammatory cytokines (IL-1ß and IL-8) and chemokine ligands (RANTES and IP-10), and upregulation of pro-healing mediators (PDGF-BB, TGF-ß1, IL-10, IL-2, IL-4, IL-11 and IL-15) and, extracellular and immune modulators (TIMP-2, M-CSF and ICAM-1). M101 significantly increased the gene expression of Resolvin-E1 receptor. Furthermore, M101 treatment reduced P. gingivalis biofilm growth over glass surface, observed with live/dead analysis and by decreased P. gingivalis 16 s rRNA expression (51.7%) (p < 0.05). In mice, M101 reduced the clinical abscess size (50.2%) in P. gingivalis-induced calvarial lesion concomitant with a decreased inflammatory score evaluated through histomorphometric analysis, thus, improving soft tissue and bone healing response. Therefore, M101 may be a novel therapeutic agent that could be beneficial in the management of P. gingivalis associated diseases.

Effects of cashew gum and nanoparticles on cooled stallion semen.

BACKGROUND: Cryopreservation of stallion spermatozoa tends to cause plasma membrane damage due to the low ratio of cholesterol to phospholipids. Gums have been suggested as an alternative cryoprotectant to glycerol for stallion spermatozoa. Therefore, the present experiment was designed to verify whether the effect of addition of cashew gum (CG), or nanoparticles (NP) containing CG, to the extender before cooling on sperm quality in stallion semen. Ejaculates from 6 stallions were extended and split between six treatment groups (control, a-tocopherol [TOC], CG1, CG0.5, NP1 and NP0.5), stored in cryotubes at 4 °C. RESULTS: Aliquots were analysed by computer-assisted sperm motility analysis on the day of collection, and after 24 h and 48 h of cold storage. After 48 h, the total motility with NP1 (78.53 + 6.31%) was similar to control 85.79 + 6.31% at 0 h. The same pattern was observed for progressive motility. Membrane integrity assessed by flow cytometer was similar between control, TOC and G1 at all storage times. The DNA fragmentation in the control group increased at all time points, whereas chromatin integrity was maintained after 24 h in TOC and NP0.5 compared to 0 h. There was no increase in the proportion of live spermatozoa producing hydrogen peroxide, but there was a tendency for an increased proportion of spermatozoa in the live superoxide category in CG1 after 24 h cooled storage. CONCLUSIONS: The addition of CG or CG-derived NP to extender for stallion semen was not harmful to the sperm cells.

Kaolinite/cashew gum bionanocomposite for doxazosin incorporation and its release.

Incorporation of drugs in clay minerals has been widely proposed for the controlled-release or increased solubility of drugs. In this context, a bionanocomposite based on kaolinite and cashew gum (Kln/Gum) was synthesized and characterized by X-ray diffraction (XRD), thermal analysis (TG/DTA), and Fourier transform infrared spectroscopy (FTIR). The bionanocomposite was applied to the incorporation and further release of doxazosin mesylate (DB). The influence of solution pH (1-3), adsorbent dose (20-50 mg), initial drug concentration (20.0-70.0 mg L-1), contact time (15-300 min), and temperature (25, 35, and 45 °C) were systematically evaluated. Equilibrium was reached around 60 min, with a maximum adsorption capacity of 31.5 ± 2.0 mg g-1 at a pH of 3.0 and 25 °C. Hydrogen bonding contributed to DB incorporation on the Kln/Gum. In addition, DB maximum amounts of 16.80 ± 0.58 and 77.00 ± 2.46% were released at pH values of 1.2 and 7.4, respectively. These results indicated that the Kln/Gum bionanocomposite is an effective and promising material for the incorporation/release of drugs with similar structures to DB.

Crude Methanol Extract of Rosin Gum Exhibits Specific Cytotoxicity against Human Breast Cancer Cells via Apoptosis Induction.

BACKGROUND: Rosin (Colophony) is a natural resin derived from species of the pine family Pinaceae. It has wide industrial applications including printing inks, photocopying paper, adhesives and varnishes, soap and soda. Rosin and its derivatives are employed as ingredients in various pharmaceutical products such as ointments and plasters. Rosin-based products contain allergens that may exert some occupational health problems such as asthma and contact dermatitis. OBJECTIVE: Our knowledge of the pharmaceutical and medicinal properties of rosin is limited. The current study aims at investigating the cytotoxic potential of Rosin-Derived Crude Methanolic Extract (RD-CME) and elucidation of its mode-of-action against breast cancer cells (MCF-7 and MDA-MB231). METHODS: Crude methanol extract was prepared from rosin. Its phenolic contents were analyzed by Reversed- Phase High-Performance Liquid Chromatography (RP-HPLC). Antioxidant activity was evaluated by DPPH radical-scavenging assay. Antiproliferation activity against MCF-7 and MDA-MB231 cancerous cells was investigated by MTT assay; its potency compared with doxorubicin as positive control and specificity were assessed compared to two non-cancerous cell lines (BJ-1 and MCF-12F). Selected apoptosis protein markers were assayed by western blotting. Cell cycle analysis was performed by Annexin V-FITC/PI FACS assay. RESULTS: RD-CME exhibited significant and selective cytotoxicity against the two tested breast cancer cells (MCF-7 and MDA-MB231) compared to normal cells as revealed by MTT assay. ELISA and western blotting indicated that the observed antiproliferative activity of RD-CME is mediated via the engagement of an intrinsic apoptosis signaling pathway, as judged by enhanced expression of key pro-apoptotic protein markers (p53, Bax and Casp 3) relative to vehicle solvent-treated MCF-7 control cells. CONCLUSION: To our knowledge, this is the first report to investigate the medicinal anticancer and antioxidant potential of crude methanolic extract derived from colophony rosin. We provided evidence that RD-CME exhibits strong antioxidant and anticancer effects. The observed cytotoxic activity against MCF-7 is proposed to take place via G2/M cell cycle arrest and apoptosis. Colophony resin has a great potential to join the arsenal of plantderived natural anticancer drugs. Further thorough investigation of the potential cytotoxicity of RD-CME against various cancerous cell lines is required to assess the spectrum and potency of its novel activity.

Fabrication of a multifunctional hydrogel with a robust interface bioinspired by the structure of the dentogingival junction.

The dentogingival junction with a perforating fiber structure is an anatomical and functional interface between the gingiva and tooth structure. Inspired by this structure, this study involves the development of a handy method to fabricate a multifunctional hydrogel with a robust interface, which is also universal for preparing diverse materials that require a Janus structure.

Graft and crosslinked polymerization of polysaccharide gum to form hydrogel wound dressings for drug delivery applications.

The present article discusses the exploration of the potential of moringa gum (MOG) polysaccharides for developing a hydrogel dressing as a slow drug carrier to the enhanced wound healing. The polymer films were formed by grafting of carbomer onto MOG by radiation induced crosslinking technique. The polymers were characterized by cryo-SEM, AFM, FTIR, 13C NMR spectroscopy, and swelling studies. Drug delivery and biomedical properties of the dressings were also determined. Polymer dressing absorbed 4.20 ± 0.09 g/g simulated wound fluid. The release of levofloxacin was observed without burst effect and followed non-Fickian diffusion mechanism and best fitted in Higuchi kinetic model. The hydrogel films were permeable to O2 and H2O vapour and impermeable to microbes and showed antioxidant activity.

The physico-chemistry of adhesions of protein resistant and weak polyelectrolyte brushes to cells and tissues.

The non-specific adhesion of polymers and soft tissues is of great interest to the field of biomedical engineering, as it will shed light on some of the processes that regulate interactions between scaffolds, implants and nanoparticles with surrounding tissues after implantation or delivery. In order to promote adhesion to soft tissues, a greater understanding of the relationship between polymer chemistry and nanoscale adhesion mechanisms is required. In this work, we grew poly(dimethylaminoethyl methacrylate) (PDMAEMA), poly(acrylic acid) (PAA) and poly(oligoethylene glycol methacrylate) (POEGMA) brushes from the surface of silica beads, and investigated their adhesion to a variety of substrates via colloidal probe-based atomic force microscopy (AFM). We first characterised adhesion to a range of substrates with defined surface chemistry (self-assembled monolayers (SAMs) with a range of hydrophilicities, charge and hydrogen bonding), before studying the adhesion of brushes to epithelial cell monolayers (primary keratinocytes and HaCaT cells) and soft tissues (porcine epicardium and keratinized gingiva). Adhesion assays to SAMs reveal the complex balance of interactions (electrostatic, van der Waals interactions and hydrogen bonding) regulating the adhesion of weak polyelectrolyte brushes. This resulted in particularly strong adhesion of PAA brushes to a wide range of surface chemistries. In turn, colloidal probe microscopy on cell monolayers highlighted the importance of the glycocalyx in regulating non-specific adhesions. This was also reflected by the adhesive properties of soft tissues, in combination with their mechanical properties. Overall, this work clearly demonstrates the complex nature of interactions between polymeric biomaterials and biological samples and highlights the need for relatively elaborate models to predict these interactions.

Cellular and collagen reference values of gingival and periodontal ligament tissues in rats: a pilot study.

Reference data are lacking on the periodontal ligament and the gingival tissue of the rat model, which would be useful for studies of new medical or biomaterial periodontal treatments. The objective of the current study was to propose cellular and collagen reference values of gingival and periodontal ligament tissues in rat, using a simple and reliable quantitative method after decalcification. Mandibular samples of ten adult Sprague-Dawley rats were used. Mild decalcification was carried out using ethylenediaminetetraacetic acid (EDTA) to preserve the morphology of tissues. Half of the samples were decalcified and the other half were not. The gingiva and the periodontal ligament were analyzed. Descriptive histology and computer-assisted image analysis were performed. The data showed that qualitatively, cellular and extracellular matrix morphologies were well preserved compared to non-decalcified periodontal soft tissue biopsies. Histomorphometrically, constitutive cellularity and the total amount of native collagen, collagen directionality and collagen anisotropy in both experimental conditions did not significantly differ. Taken together, these results suggested that EDTA decalcification did not negatively affect the studied endpoints. Moreover, this mild decalcification method allowed in situ maintenance of the periodontal soft and hard tissue integrity. The structural and compositional computerized assessment performed in the healthy periodontal soft tissue could provide reference values that will be required for future assessment on the effects of pathological, reparative and regenerative processes in rat periodontal soft tissues.

Gliclazide reduced oxidative stress, inflammation, and bone loss in an experimental periodontal disease model.

OBJECTIVE: The aim of this study was to evaluate the effects of gliclazide on oxidative stress, inflammation, and bone loss in an experimental periodontal disease model. MATERIAL AND METHODS: Male albino Wistar rats were divided into no ligature, ligature, and ligature with 1, 5, and 10 mg/kg gliclazide groups. Maxillae were fixed and scanned using micro-computed tomography to quantify linear and bone volume/tissue volume (BV/TV) and volumetric bone loss. Histopathological, immunohistochemical and immunofluorescence analyses were conducted to examine matrix metalloproteinase-2 (MMP-2), cyclooxygenase 2 (COX-2), cathepsin K, members of the receptor activator of the nuclear factor kappa-Β ligand (RANKL), receptor activator of nuclear factor kappa-Β (RANK), osteoprotegerin (OPG) pathway, macrophage migration inhibitory factor (MIF), superoxide dismutase-1 (SOD-1), glutathione peroxidase-1 (GPx-1), NFKB p 50 (Cytoplasm), NFKB p50 NLS (nuclear localization signal), PI3 kinase and AKT staining. Myeloperoxidase activity, malondialdehyde and glutathione levels, while interleukin-1 beta (IL-1ß) and tumor necrosis factor-alpha (TNF-α) levels were evaluated by spectroscopic ultraviolet-visible analysis. A quantitative reverse transcription polymerase chain reaction was used to quantify the gene expression of the nuclear factor kappa B p50 subunit (NF-κB p50), phosphoinositide 3-kinase (PI3k), protein kinase B (AKT), and F4/80. RESULTS: Micro-computed tomography showed that the 1 mg/kg gliclazide treatment reduced linear bone loss compared to the ligature, 5 mg/kg gliclazide, and 10 mg/kg gliclazide treatments. All concentrations of gliclazide increased bone volume/tissue volume (BV/TV) compared to the ligature group. Treatment with 1 mg/kg gliclazide reduced myeloperoxidase activity, malondialdehyde, IL-1ß, and TNF-α levels (p≤0.05), and resulted in weak staining for COX-2, cathepsin k, MMP-2, RANK, RANKL, SOD-1, GPx-1,MIF and PI3k. In addition, down-regulation of NF-κB p50, PI3k, AKT, and F4/80 were observed, and OPG staining was strong after the 1 mg/kg gliclazide treatment. CONCLUSIONS: This treatment decreased neutrophil and macrophage migration, decreased the inflammatory response, and decreased bone loss in rats with ligature-induced periodontitis.

In vitro evaluation of Eugenia dysenterica in primary culture of human gingival fibroblast cells

Abstract: Eugenia dysenterica is a Brazilian tree investigated for its properties and bioactive compounds, which are believed to have both pharmacological and phytochemical therapeutic effects. The leaves of this tree contain tannins, flavonoids, terpenes, and saponins, with reportedly beneficial effects to the human body. Despite these therapeutic applications, its effects have never been tested on oral tissues. Therefore, the aim of the present study was to evaluate the cytotoxic and antioxidant effects and the anti-inflammatory and repair properties of the acetone fraction of E. dysenterica on primary culture of human gingival fibroblasts and on the immortalized murine macrophage cell line (RAW 264.7). For this purpose, a metabolic activity assay, a wound healing assay, a nitric oxide assay, and RT-qPCR were performed. The assays revealed a cytoprotective effect of this plant, suggested by the increase in the expression of SOD1 and NRF2. An antioxidant potential effect was observed in the DPPH• assay. However, the fraction of E. dysenterica did not show anti-inflammatory activity. In conclusion, Eugenia dysenterica may promote cytoprotection when associated with chlorhexidine digluconate because of its antioxidant effect. However, additional studies are necessary on other human dental tissues using other parts of the plant in order to develop a possible mouthwash to assist patients with oral disorders.

Gliclazide reduced oxidative stress, inflammation, and bone loss in an experimental periodontal disease model

Abstract Objective The aim of this study was to evaluate the effects of gliclazide on oxidative stress, inflammation, and bone loss in an experimental periodontal disease model. Material and Methods Male albino Wistar rats were divided into no ligature, ligature, and ligature with 1, 5, and 10 mg/kg gliclazide groups. Maxillae were fixed and scanned using micro-computed tomography to quantify linear and bone volume/tissue volume (BV/TV) and volumetric bone loss. Histopathological, immunohistochemical and immunofluorescence analyses were conducted to examine matrix metalloproteinase-2 (MMP-2), cyclooxygenase 2 (COX-2), cathepsin K, members of the receptor activator of the nuclear factor kappa-Β ligand (RANKL), receptor activator of nuclear factor kappa-Β (RANK), osteoprotegerin (OPG) pathway, macrophage migration inhibitory factor (MIF), superoxide dismutase-1 (SOD-1), glutathione peroxidase-1 (GPx-1), NFKB p 50 (Cytoplasm), NFKB p50 NLS (nuclear localization signal), PI3 kinase and AKT staining. Myeloperoxidase activity, malondialdehyde and glutathione levels, while interleukin-1 beta (IL-1β) and tumor necrosis factor-alpha (TNF-α) levels were evaluated by spectroscopic ultraviolet-visible analysis. A quantitative reverse transcription polymerase chain reaction was used to quantify the gene expression of the nuclear factor kappa B p50 subunit (NF-κB p50), phosphoinositide 3-kinase (PI3k), protein kinase B (AKT), and F4/80. Results Micro-computed tomography showed that the 1 mg/kg gliclazide treatment reduced linear bone loss compared to the ligature, 5 mg/kg gliclazide, and 10 mg/kg gliclazide treatments. All concentrations of gliclazide increased bone volume/tissue volume (BV/TV) compared to the ligature group. Treatment with 1 mg/kg gliclazide reduced myeloperoxidase activity, malondialdehyde, IL-1β, and TNF-α levels (p≤0.05), and resulted in weak staining for COX-2, cathepsin k, MMP-2, RANK, RANKL, SOD-1, GPx-1,MIF and PI3k. In addition, down-regulation of NF-κB p50, PI3k, AKT, and F4/80 were observed, and OPG staining was strong after the 1 mg/kg gliclazide treatment. Conclusions This treatment decreased neutrophil and macrophage migration, decreased the inflammatory response, and decreased bone loss in rats with ligature-induced periodontitis.

A Secreted Bacterial Peptidylarginine Deiminase Can Neutralize Human Innate Immune Defenses.

The keystone oral pathogen Porphyromonas gingivalis is associated with severe periodontitis. Intriguingly, this bacterium is known to secrete large amounts of an enzyme that converts peptidylarginine into citrulline residues. The present study was aimed at identifying possible functions of this citrullinating enzyme, named Porphyromonas peptidylarginine deiminase (PPAD), in the periodontal environment. The results show that PPAD is detectable in the gingiva of patients with periodontitis, and that it literally neutralizes human innate immune defenses at three distinct levels, namely bacterial phagocytosis, capture in neutrophil extracellular traps (NETs), and killing by the lysozyme-derived cationic antimicrobial peptide LP9. As shown by mass spectrometry, exposure of neutrophils to PPAD-proficient bacteria reduces the levels of neutrophil proteins involved in phagocytosis and the bactericidal histone H2. Further, PPAD is shown to citrullinate the histone H3, thereby facilitating the bacterial escape from NETs. Last, PPAD is shown to citrullinate LP9, thereby restricting its antimicrobial activity. The importance of PPAD for immune evasion is corroborated in the infection model Galleria mellonella, which only possesses an innate immune system. Together, the present observations show that PPAD-catalyzed protein citrullination defuses innate immune responses in the oral cavity, and that the citrullinating enzyme of P. gingivalis represents a new type of bacterial immune evasion factor.IMPORTANCE Bacterial pathogens do not only succeed in breaking the barriers that protect humans from infection, but they also manage to evade insults from the human immune system. The importance of the present study resides in the fact that protein citrullination is shown to represent a new bacterial mechanism for immune evasion. In particular, the oral pathogen P. gingivalis employs this mechanism to defuse innate immune responses by secreting a protein-citrullinating enzyme. Of note, this finding impacts not only the global health problem of periodontitis, but it also extends to the prevalent autoimmune disease rheumatoid arthritis, which has been strongly associated with periodontitis, PPAD activity, and loss of tolerance against citrullinated proteins, such as the histone H3.

Evaluation of the intensity of gingival melanin pigmentation at different age groups in the Indian population: An observational study.

BACKGROUND: The brownish-black pigmentation of the gingiva has been reported in several countries. However, the available literature pertaining to the Asian population suggests that this pigmentation is more predominant in individuals with dark complexion. Although there is available literature on the intensity of gingival pigmentation with different age groups in Japanese and Israel population, no such literature pertaining to age and intensity of melanin pigmentation is available in the Indian population. AIM: This study aims to observe the intensity of melanin pigmentation in different age groups in the Indian population. METHODS AND MATERIALS: Intensity of melanin pigmentation was observed by the gingival melanin pigmentation index as given by Ponnaiyan et al. in 250 individuals who were grouped under the age group of 0-5 years, 6-9 years, 10-12 years, 13-15 years, and 16-20 years. Each group comprised 25 individuals. RESULTS AND CONCLUSION: The results were subjected to statistical analysis so as to observe the intensity of melanin pigmentation at different age groups in the Indian population. It was concluded that with age, the intensity of gingival melanin pigmentation, as well as its distribution to the posterior gingiva was increased. As age increased, gingival pigmentation was more in the attached and interdental papilla, while in the younger age groups, it was confined either only to the interdental, marginal or attached gingival, or both depending on the skin color.

Impact of obesity and aging on crestal alveolar bone height in mice.

Obesity and aging are associated with periodontitis, which represents a chronic inflammatory disease of the tooth-supporting tissues, i.e. the periodontium. However, if both risk factors also have a negative impact on crestal alveolar bone in a clinically healthy periodontium, has yet to be elucidated and was analyzed in this in-vivo study. Eight C57BL/6 mice were fed a normal diet during the entire study. Half of these mice were sacrificed at week 19 (group 1: younger lean mice), whereas the other half of the animals were sacrificed at week 25 (group 2: older lean mice). In addition, four mice were fed a high-fat diet until their sacrifice at week 19 (group 3: younger obese mice). Mandibles and maxillae were scanned by micro-computed tomography and, subsequently, the distance between the cementoenamel junction and alveolar bone crest (CEJ-ABC) at all molars was determined. Levels of interleukin-6, cyclooxygenase-2, visfatin and adiponectin in gingival samples were quantified by real-time PCR. For statistical analyses, the Mann-Whitney-U test was applied (p<0.05). As compared to lean mice, obese animals presented a significantly increased CEJ-ABC distance, i.e. reduced alveolar bone crest height, at week 19. The alveolar bone loss was mainly found at the first molars of the mandibles. In animals fed a normal diet, the alveolar bone crest height in the mandibles and maxillae was significantly lower in the older mice as compared to the younger animals. Furthermore, gingival cyclooxygenase-2 and visfatin expressions were higher in the obese versus lean mice and in the older versus younger mice. This in-vivo investigation shows that obesity and older age can result in reduced alveolar bone crest height and suggests that they represent risk factors even in a clinically healthy periodontium.