TLR2 agonist prevents the progression of periapical lesions in mice by reducing osteoclast activity and regulating the frequency of Tregs.

AIM: To evaluate the role of regulatory T lymphocytes (Tregs) in the presence or absence of the synthetic ligand Pam3Cys during the progression of periapical lesion in wild-type (WT) and toll-like receptor 2 knockout (TLR2KO) mice. METHODOLOGY: A total of 130 C57BL/6 male WT and TLR2KO mice were allocated into control (n = 5) and experimental (periapical lesion induction) (n = 10) groups. In specific groups (WT+Pam3cys and TLR2KO+Pam3cys), the synthetic ligand Pam3cys was administered intraperitoneally every 7 days, according to the experimental period (14, 21 and 42 days). At the end of those periods, the animals were euthanized, and the mandible and the spleen were submitted to histotechnical processing. Mandible histological sections were analysed by haematoxylin and eosin, TRAP histoenzymology and immunohistochemistry (FOXP3, RANK, RANKL and OPG). Spleen sections were analysed by immunohistochemistry (FOXP3). RESULTS: The inflammatory infiltrate and bone resorption were more intense in the TLR2KO group compared to the WT group. The animals that received the Pam3cys had smaller periapical lesions when compared to the animals that did not receive the ligand (p < .05). TLR2KO animals showed a significant increase in the number of osteoclasts when compared to TLR2KO+Pam3cys group (p < .05). At 21 days, the WT+Pam3cys group had a lower number of osteoclasts when compared to the WT animals (p = .02). FOXP3 expression was more intense in the WT+Pam3cys groups when compared to the WT animals in the 42 days (p = .03). In the spleen analysis, the WT+Pam3cys group also had a higher expression of FOXP3 when compared to the WT animals at 14 and 42 days (p = .02). Concerning RANKL, there was a reduction in staining in the KOTLR2+Pam3cys groups at 21 and 42 days (p = .03) and a higher binding ratio between RANK/RANKL in animals that did not receive the ligand. CONCLUSION: Administration of the Pam3cys increased the proliferation of Tregs, showed by FOXP3 expression and prevented the progression of the periapical lesion in WT mice. On the other hand, in the TLR2KO animals, Treg expression was lower with larger areas of periapical lesions. Finally, systemic administration of the Pam3cys in KO animals was able to limit the deleterious effects of the absence of the TLR2 receptor.

The effects of NLRP3 inflammasome inhibition or knockout in experimental apical periodontitis induced in mice.

OBJECTIVE: To evaluate the effects of NLRP3 inflammasome inhibition or knockout in experimental apical periodontitis (AP) induced in mice. METHODS: The experimental AP was induced by pulpal exposure. To evaluate NLRP3-specific inhibitor medication (MCC950), WT mice received intraperitoneal injections, while the control received PBS (n = 10). In addition, to evaluate NLRP3 knockout, 35 wild-type (WT) and 35 NLRP3-/- mice were divided into a control group (without pulpal exposure, n = 5) and three experimental groups: after 2, 14 and 42 days after pulpal exposure (n = 10). Microscopic and molecular analyzes were carried out using a significance level of 5%. RESULTS: Exposure to MCC950 did not affect the periapical lesion size after 14 days (P = 0.584). However, exposed mice had a lower expression of IL-1ß, IL-18 and caspase-1 (P = 0.010, 0.016 and 0.002, respectively). Moreover, NLRP3-/- mice showed a smaller periapical lesion after 14 and 42 days (P = 0.023 and 0.031, respectively), as well as a lower expression of IL-1ß after 42 days (P < 0.001), of IL-18 and caspase-1 after 14 (P < 0.001 and 0.035, respectively) and 42 days (P = 0.002 and 0.002, respectively). NLRP3-/- mice also showed a lower mRNA for Il-1ß, Il-18 and Casp1 after 2 (P = 0.002, 0.036 and 0.001, respectively) and 14 days (P = 0.002, 0.002 and 0.001, respectively). CONCLUSIONS: NLRP3 inflammasome inhibition or knockout can attenuate the inflammatory events that result in the periapical lesion (AP) formation after pulpal exposure in mice. CLINICAL RELEVANCE: The NLRP3 inflammasome may be a therapeutic target for AP, and new approaches may verify the impact of its inhibition (through intracanal medications or filling materials) on the bone repair process and treatment success.

Root canal conicity determination of primary second molars using nanotomography: An in vitro study.

BACKGROUND: The conicity of the root canals of primary teeth is an important measure for endodontic therapies. However, determining this conicity depends on the methods employed, which requires further investigation. AIM: The aim of this study was to determine the conicity of the root canals of the upper and lower primary second molars using nanotomography (nCT). DESIGN: An in vitro study was performed using nine primary second molars, both upper and lower, subjected to nCT. Comparisons between the diameters of root canals were performed between the thirds (cervical-D0, middle-D5, and apical-D7). The conicity (%) was determined for each root canal from cervical to apical. Data were statistically analyzed with a significance level of 5%. RESULTS: The conicity ranged from 2% to 8% for the upper primary second molars. Significant differences in root canal diameter between the thirds (D0, D5, and D7 points) were observed in the mesio- and distobuccal roots (p < .05), but not in the palatal roots (p > .05). For the lower primary second molars, the conicity ranged from 2% to 17%, as well as significant differences in root canal diameter between the thirds (D0, D5, and D7 points) were observed in all roots (distal, mesiobuccal, and mesiolingual; p < .05). CONCLUSION: The conicity of the upper primary second molars was different from that of the lower ones, which showed a greater variability.

Analysis of the cytotoxicity and bioactivity of CeraSeal, BioRoot™ and AH Plus® sealers in pre-osteoblast lineage cells.

BACKGROUND: The objective of the present study was to evaluate in vitro the cytotoxicity and bioactivity of various endodontic sealers (CeraSeal, BioRoot™ and AH Plus®) in pre-osteoblast mouse cells (MC3T3 cells). METHODS: MC3T3 cells (ATCC CRL-2594) were plated in 1 × 104 cells/well in 96-well plates in contact with endodontic sealers at concentrations of 1:10 and 1:100. Cell viability was evaluated by MTT assay after 24 and 48 h. In addition, sealer bioactivity was measured by RT-PCR for mediator of inflammation (Tnf, Ptgs2) and mineralization (Runx2, Msx1, Ssp1 and Dmp1) after 24 h and by Alizarin Red S Assay of mineralization after 28 days. Data were analyzed using one-way ANOVA followed by the Tukey's post-test at a significance level of 5%. RESULTS: BioRoot™ presented 24-hour cytotoxicity (p < 0.05) at 1:10 concentration. In the period of 48 h, no endodontic cement was cytotoxic to the cells compared to the control (p > 0.05). TNF-α gene expression was induced by AH Plus® (p < 0.05), while Ptgs2 was induced by the CeraSeal and BioRoot™ (p < 0.05). The expression of Runx2 was stimulated by BioRoot™ and AH Plus® (p < 0.05). In contrast, the expression of Dmp-1 Dmp1 was higher for the CeraSeal and BioRoot™ (p < 0.05). Nonetheless, the sealers did not impact the formation of mineralization nodules (p > 0.05). CONCLUSION: CeraSeal, BioRoot™ and AH Plus® sealers were not cytotoxic to MC3T3 cells within 48 h, but differentially induced the expression of genes related to inflammation and mineralization without impacting biomineralization by the cells.

Genetic polymorphism in the tumour necrosis factor alpha gene (G-308A) is associated with persistent apical periodontitis in Brazilians.

AIM: To investigate if there was an association between genetic polymorphisms in tumour necrosis factor (TNF)-⍺ and its receptors TNFRSF1A and TNFRSF1B with persistent apical periodontitis (PAP) in Brazilian subjects. METHODOLOGY: Patients who had pulpal necrosis and apical periodontitis at the time of treatment, with at least 1-year of follow-up after non-surgical root canal treatment were recalled. Three hundred and seventy eight subjects were included, 150 subjects with signs/symptoms of PAP and 228 subjects with root canal-treated teeth exhibiting healthy perirradicular tissues (healed). Genomic DNA was extracted from saliva and used for TNF-⍺ (rs1800629), TNFRSF1A (rs1800693) and TNFRSF1B (rs1061622) genotyping by real-time PCR. Genotypes and alleles frequencies were evaluated by c2 or Fisher's exact tests and odds ratios were implemented (α = 5%). RESULTS: The genetic polymorphism in TNF-α (rs1800629) was associated as a protective factor for the development of PAP (p < .05), once subjects who presented at least one allele A (AA+AG X GG), had a higher chance to lesion repair (p < .05). The polymorphisms rs1800693 and rs1061622 in TNF receptors (TNFRSF1A and TNFRSF1B, respectively) were not associated with the development of PAP (p > .05). CONCLUSIONS: The observed results demonstrate that polymorphism in TNF-α but not in its receptors is associated with PAP.

Effect of chlorhexidine mouthwashes on periodontal parameters and extrinsic tooth staining in orthodontic patients.

INTRODUCTION: Periodontal health and biofilm control are primordial to success in orthodontic treatment. This study aimed to evaluate the effect of chlorhexidine (CHX) mouthwashes on periodontal status and extrinsic tooth staining in orthodontic patients. METHODS: Thirty-three patients of both sexes, aged 11-33 years, under orthodontic treatment with fixed appliances at <16 months, were randomly distributed into 2 groups. In the control group, patients received mechanical hygiene instruction, and in the experimental group, patients also used CHX wash twice a week for 60 days. The effectivity of the protocols was evaluated using the plaque, gingival, gingival bleeding, and discoloration indexes before the hygiene protocol was applied, 15, 30, and 60 days after protocol implementation. RESULTS: In the experimental group, there was a decrease in the plaque, gingival, and gingival bleeding indexes at the different evaluation periods (P <0.05). In addition, there was a significant difference in the discoloration index at 60 days compared with initial time points after implementing hygiene protocols in the experimental group (P <0.05). In contrast, there were no significant differences in plaque, gingival, gingival bleeding, and discoloration indexes in the control group at any time (P >0.05). CONCLUSIONS: CHX mouthwash administered 30 days, twice a week, significantly improved the periodontal status with mild brown staining. After this period, expressive extrinsic tooth staining was observed.

Assessment of microbial contamination in removable orthodontic appliances with and without the use of antimicrobial agents by checkerboard DNA-DNA hybridization analysis.

INTRODUCTION: This clinical, crossover, double-blind trial evaluated the microbial contamination of removable orthodontic appliances used by children and the efficacy of 0.12% chlorhexidine gluconate spray use for disinfection. METHODS: Twenty children aged 7-11 years were instructed to wear removable orthodontic appliances for 1 week. They were instructed to use a placebo solution (control) or 0.12% chlorhexidine gluconate (experimental) to clean the appliances on days 4 and 7 after installation. After this period, the microbial contamination on the surfaces of the appliance was analyzed using checkerboard DNA-DNA hybridization for 40 bacterial species. Data were analyzed by Fisher exact, t, and Wilcoxon tests (α = 0.05). RESULTS: Removable orthodontic appliances were heavily contaminated by the target microorganisms. Streptococcus sanguinis, Streptococcus oralis, Streptococcus gordonii, and Eikenella corrodens were found in 100% of the appliances. Among cariogenic microorganisms, Streptococcus mutans and Streptococcus sobrinus were more abundant than Lactobacillus acidophilus and Lactobacillus casei. Red complex pathogens were more abundant than orange complex species. Purple complex bacteria were the most prevalent among bacterial complexes not associated with specific pathologies, detected in 34% of the samples. After the use of chlorhexidine, the number of cariogenic microorganisms (S. mutans, S. sobrinus, and L. casei) decreased significantly (P <0.05), and the numbers of periodontal pathogenic species from the orange and red complex also decreased significantly (P <0.05). There was no reduction for Treponema socranskii. CONCLUSIONS: Removable orthodontic appliances were densely contaminated by several bacterial species. Twice-a-week application of chlorhexidine spray effectively reduced cariogenic and orange and red complex periodontal pathogens.

Systemic inhibition of 5-lipoxygenase by MK-886 exacerbates apical periodontitis bone loss in a mouse model.

BACKGROUND: To investigate if 5-LO selective inhibitor (MK-886) could be used for systemic treatment of experimentally induced apical periodontitis in a mouse model. METHODS: Twenty-four C57BL/6 mice were used. After coronal opening, a solution containing Escherichia coli LPS (1.0 µg/µL) was inoculated into the root canals of the lower and upper right first molars (n = 72 teeth). After 30 days apical periodontitis was established, and the animals were treated with MK-886 (5 mg/kg), a 5-LO inhibitor, for 7 and 14 days. The tissues were removed for histopathological and histometric analyses, evaluation of osteoclast number and gene expression for receptor activator of nuclear factor kappa-B (Tnfrsf11a), receptor activator of nuclear factor kappa-B ligand (Tnfsf11), osteoprotegerin (Tnfrsf11b), tartrate-resistant acid phosphatase (Acp5), matrix metalloproteinase-9 (Mmp9), cathepsin K (Ctsk) and calcitonin receptor (Calcr). Statistical data analysis was performed using Kruskal Wallis followed by Dunn's tests (α = 0.05). RESULTS: Administration of MK-886 for 7 days exerted no effect on apical periodontitis progression compared to LPS inoculation without treatment (p = 0.3549), while treatment for 14 days exacerbated bone loss (p < 0.0001). Administration of MK-886 enhanced osteoclastogenesis signaling and osteoclast formation within 7 days (p = 0.0005), but exerted no effect at 14 days (p > 0.9999). After 7 days of treatment, MK-886 induced mRNA expression for Acp5 (p = 0.0001), Calcr (p = 0.0003), Mmp9 (p = 0.0005) and Ctsk (p = 0.0008), however no effect in those gene expression was observed after 14 days (p > 0.05). CONCLUSION: Systemic treatment with MK-886 exacerbated LPS-induced apical periodontitis in a mouse model.

Interferon-gamma inducible protein 16 and type I interferon receptors expression in experimental apical periodontitis induced in wild-type mice.

AIM: The aim of this study was to evaluate the IFI16 and IFN-α/ß receptors expression during the genesis and development of experimental apical periodontitis (AP) in mice teeth. METHODOLOGY: Apical periodontitis was induced in the lower first molars of 40 C57BL/6 mice. They were divided according to the experimental periods 2, 7, 14, 21 and 42 days (n = 8 per group). Five animals were used as a control group (without AP). Specimens were submitted to histological processing for description of the inflammatory process, immunostaining for the presence/absence and localization of IFI16 and IFN-α/ß receptors (qualitative and semi-quantitative analysis) and tartrate-resistant acid phosphatase (TRAP) histoenzimology. RESULTS: The results showed a gradual development of AP over the experimental times. The expression of IFI16 was noticeably more exacerbated in the experimental early period (day 2) whilst the lowest expression was observed in the control group (p = .02). For IFN-α/ß receptors, a higher intensity staining was observed 42 days after AP induction, that was statistically different from the control group (p = .02). In addition, the number of TRAP-positive cells was higher on the later periods (days 21 and 42; p < .001). CONCLUSION: IFI16 protein expression was highest during the early periods after AP induction in mice teeth, whilst IFN-α/ß receptor expression was highest after AP became established.

Photoelastic, microhardness and roughness analysis of zirconia and stainless steel crowns for primary molars.

PURPOSE: To compare the stress distribution through photoelasticity, microhardness and roughness of intact crowns of primary molars (CC) and the preformed crowns of stainless steel (SSC) and zirconia (ZC) used in dental restorations in pediatric dentistry. METHODS: Six healthy primary molars were selected. For the photoelastic models, the teeth were fixed in photoelastic resin. A load of 100 N was applied, and the models were analyzed by transmission polariscope. The Tardy method was used to quantify the fringe order which calculates the maximum stress (T) value in each selected point. The teeth were prepared for cementation of the crowns. The photoelastic test was repeated for each experimental crown. Knoop microhardness was assessed on the buccal surfaces of the CCs, SSCs, and ZCs using a microhardness tester. Parameters were 50 gf for 5 seconds. Roughness was evaluated using a confocal 3-D laser scanning microscope/software at 216x magnification. Roughness average (Ra) values from each model (expressed in µm) were collected and group means were calculated. The stress distribution, microhardness, and roughness data were compared by using one-way ANOVA and the Tukey's test (α= 0.05). RESULTS: There was no difference in the stress distribution for the CCs, SSCs and ZCs. For the microhardness analysis, the ZCs obtained the highest values compared to the CCs and the SSCs (P< 0.001). The CCs were significantly higher than the SSCs (P= 0.027). There was no difference in roughness for the three models (P= 0.615). The SSCs and ZCs showed satisfactory mechanical behavior. CLINICAL SIGNIFICANCE: The use of preformed crowns, especially those made of esthetic materials, is currently increasing in the field of pediatric dentistry. The knowledge of mechanical properties of stainless steel- and zirconia-prefabricated crowns provides scientific foundation for safe clinical application, especially in primary teeth.

Self-ligating brackets exhibit accumulation of high levels of periodontopathogens in gingival crevicular fluid.

Different types of brackets seem to influence the disruption of the oral microbial environment. Therefore, the aim of this study was to evaluate the influence of self-ligating brackets on the gingival crevicular fluid levels of the putative periodontal pathogens Aggregatibacter actinomycetemcomitans sorotype a (Aaa), Tannerella forsythia, Fusobacterium nucleatum, and Porphyromonas gingivalis. Sixty samples of crevicular fluid of twenty patients (11 boys and 9 girls) were analysed at baseline (T0) and after 30 (T1) and 60 (T2) days of bonding of the self-ligating (In-Ovation®R, Dentsply, GAC or SmartClip™, 3 M Unitek, Monrovia, CA, USA) and of one conventional bracket (Gemini™, 3 M Unitek, Monrovia, CA, USA) used with elastomeric ligatures. Total DNA from samples was extracted using CTAB-DNA precipitation method and Real-time PCR was performed to analyse bacterial level. Non-parametric Friedman and Wilcoxon tests were used for data analysis (p value of < 0.05). F. nucleatum presented a different level among the different brackets at T1 (p = 0.025), the highest level in the Gemini™ bracket when compared to the SmartClip™ bracket (p = 0.043). P. ginigvalis levels increased in the In-Ovation®R (p = 0.028) at T1. The subgingival levels of bacterial species associated with periodontal disease P. ginigvalis increased in the self-ligating brackets In-Ovation®R.Clinical Relevance: Some kinds of brackets could provide more retentive sites than others, and it seems to modulate the subgingival microbiota, since, in this study, we could observe the increase of the species associated with periodontal disease. Preventive protocols should be adopted in the use of self-ligating brackets.

Use of the Probiotic Bifidobacterium animalis subsp. lactis HN019 in Oral Diseases.

The oral cavity is one of the environments on the human body with the highest concentrations of microorganisms that coexist harmoniously and maintain homeostasis related to oral health. Several local factors can shift the microbiome to a pathogenic state of dysbiosis. Existing treatments for infections caused by changes in the oral cavity aim to control biofilm dysbiosis and restore microbial balance. Studies have used probiotics as treatments for oral diseases, due to their ability to reduce the pathogenicity of the microbiota and immunoinflammatory changes. This review investigates the role of the probiotic Bifidobacterium animalis subsp. lactis (B. lactis) HN019 in oral health, and its mechanism of action in pre-clinical and clinical studies. This probiotic strain is a lactic acid bacterium that is safe for human consumption. It mediates bacterial co-aggregation with pathogens and modulates the immune response. Studies using B. lactis HN019 in periodontitis and peri-implant mucositis have shown it to be a potential adjuvant treatment with beneficial microbiological and immunological effects. Studies evaluating its oral effects and mechanism of action show that this probiotic strain has the potential to be used in several dental applications because of its benefit to the host.

Leukotriene B4 loaded in microspheres regulate the expression of genes related to odontoblastic differentiation and biomineralization by dental pulp stem cells.

BACKGROUND: Leukotriene B4 (LTB4) is a potent lipid mediator that stimulate the immune response. Because dental pulp inflammation and dentin repair are intrinsically related responses, the aim of this research was to investigate the potential of LTB4 in inducing differentiation of dental pulp stem cells. METHODS: Microspheres (MS) loaded with LTB4 were prepared using an oil emulsion solvent extraction evaporation process and sterility, characterization, efficiency of LTB4 encapsulation and in vitro LTB4 release assay were investigated. Mouse dental pulp stem cells (OD-21) were stimulated with soluble LTB4 or MS loaded with LTB4 (0.01 and 0.1 µM). Cytotoxicity and cell viability was determined by lactate dehydrogenase and methylthiazol tetrazolium assays. Gene expression were measured by quantitative reverse transcription polymerase chain reaction after 3, 6, 24, 48 and 72 h. Mineralized nodule formation was assessed after 28 days of OD-21 cell stimulation with LTB4 in mineralized media or not. Groups were compared using one-way ANOVA test followed by Dunnett's post-test (α = 0.05). RESULTS: Treatment with LTB4 or MS loaded with LTB4 (0.01 and 0.1 µm-µM) were not cytotoxic to OD-21 cells. Treatment with LTB4 modulated the expression of the Ibsp (integrin binding sialoprotein) and Runx2 (runt-related transcription factor 2) genes differently depending on the experimental period analyzed. Interestingly LTB4 loaded in microspheres (0.1 µM) allowed long term dental pulp cell differentiation and biomineralization. CONCLUSION: LTB4, soluble or loaded in MS, were not cytotoxic and modulated the expression of the Ibsp and Runx2 genes in cultured OD-21 cells. When LTB4 was incorporated into MS, odontoblast differentiation and mineralization was induced in long term culture.

Osteoclast formation, inflammation, and matrix metalloproteinase-9 are downregulated in bone repair following root canal treatment in dogs teeth.

OBJECTIVES: The aim of this study was to investigate the inflammatory infiltrate, osteoclast formation, and expression of MMP-9 during the healing phase following root canal treatment in teeth with apical periodontitis. MATERIALS AND METHODS: Apical periodontitis was induced in dogs teeth, and root canal treatment was performed in a single visit or using calcium hydroxide as intracanal medication. One hundred and eighty days following treatment the presence of inflammation was examined, and the tissues were stained to detect osteoclasts by means of a tartrate resistant alkaline phosphatase (TRAP) assay. Synthesis of MMP-9 was detected using Western blotting and immunohistochemistry. RESULTS: Teeth with apical periodontitis that had root canal therapy performed in a single visit presented a higher synthesis of MMP-9 compared with root canal treatment using calcium hydroxide. Treatment with calcium hydroxide resulted in a reduced amount of inflammatory cells and MMP-9 positive cells. Osteoclast formation, the number of MMP-9 positive osteoclasts and cementocytes, was reduced following root canal treatment, regardless of the root canal treatment protocol used. CONCLUSION: Root canal treatment reduced the amount of inflammatory cells and osteoclasts in periapical area. The use of calcium hydroxide as intracanal medication resulted in a lower synthesis of MMP-9, though the number of osteoclasts and MMP-9 positive osteoclasts were similar between the groups. CLINICAL RELEVANCE: Periapical bone repair following root canal treatment is impacted by therapy performed either in single visit or using calcium hydroxide dressing measured by inflammatory cell recruitment, osteoclast formation, and MMP-9 synthesis.

Alteration of the oral microbiota may be a responsible factor, along with estrogen deficiency, by the development of larger periapical lesions.

OBJECTIVES: To answer the questions: (1) Does reducing estrogen levels influence the microbial composition of the oral cavity? (2) Does the presence of periapical lesion (PL) cause changes in the oral microbiota? (3) Since estrogen deficiency alters the oral microbiota, can this be one of the factors that contribute to the increase of the PL? MATERIALS AND METHODS: Thirty-six rats were divided into four groups: sham (control), ovariectomy (OVX), control with PL (Sham + PL), and OVX + PL. After 9 weeks of OVX, the lower first molars were submitted to PL induction. After 21 days, the microbiological collection of the oral cavity was performed, and the animals were euthanized. The contents were evaluated by the checkerboard DNA-DNA hybridization method, to verify the prevalence of 40 bacterial species (divided into 7 microbial complexes). The blocks containing the lower first molars were submitted to histotechnical processing and staining with hematoxylin and eosin (HE), for the measurement of the periapical lesion area. The results were submitted to ANOVA and Kruskal-Wallis tests and Tukey and Dunn post-tests, with a significance level of 5%. RESULTS: In conditions of estrogen deficiency, there was alteration of the oral microbiota. The OVX groups had a higher amount of bacteria compared to the SHAM group in most of the microbial complexes (p < 0.001). The animals in the control group (with or without lesion) did not present a statistically significant difference (p > 0.001) in any of the microbial complexes. The PLs in OVX animals were significantly higher compared to SHAM animals (p < 0.001). CONCLUSIONS: Hypoestrogenicity conditions interfere in the oral microbiota by increasing the amount of bacteria in the saliva and influencing the progression of periapical lesions. CLINICAL RELEVANCE: This inedited study shows that deficiency of estrogen leads to alteration of the oral microbiota.

Effect of non-steroidal anti-inflammatory drugs on pulpal and periapical inflammation induced by lipopolysaccharide.

OBJECTIVES: The objective of this study was to evaluate the effect of non-steroidal anti-inflammatory drugs (NSAIDs) in controlling pulpal and periapical inflammation in vivo as a potential coadjutant systemic therapy for pulpitis. MATERIALS AND METHODS: A suspension containing E. coli lipopolysaccharide (LPS; 1.0 µg/µL) was inoculated into the pulp chamber of the first molars of C57BL/6 mice (n = 72), and the animals were treated daily with indomethacin or celecoxib throughout the experimental periods. After 7, 14, 21, and 28 days, the tissues were removed for histopathological, histoenzymology, histometric, and immunohistochemical evaluation. RESULTS: Inoculation of LPS into the pulp chamber induced the synthesis of the enzyme cyclooxygenase-2 (COX-2) in dental pulp and periapical region. Indomethacin and celecoxib treatment changed the profile of inflammatory cells recruited to dental pulp and to the periapex, which was characterized by a higher mononuclear cell infiltrate, compared to LPS inoculation alone which recruited a higher amount of polymorphonuclear neutrophils. Administration of indomethacin for 28 days resulted in the development of apical periodontitis and increased osteoclast recruitment, unlike celecoxib. CONCLUSIONS: NSAIDs indomethacin and celecoxib changed the recruitment of inflammatory cells to a mononuclear profile upon inoculation of LPS into the pup chamber, but indomethacin enhanced periapical bone loss whereas celecoxib did not. CLINICAL RELEVANCE: Celecoxib, a selective COX-2 inhibitor, can change the profile of inflammatory cells recruited to the dental pulp challenged with LPS and might a be potential systemic coadjutant for treatment of pulpitis.

Effect of root surface treatment with denusomab after delayed tooth replantation.

OBJECTIVES: To evaluate denosumab, a human monoclonal antibody that mimics the effects of osteoprotegerin in bone metabolism, as a topical treatment of root surface to be used prior to delayed tooth replantation. MATERIALS AND METHODS: Thirty-six rats' right incisors were used. Teeth were extracted and divided into: delayed replantation without root surface treatment (control); delayed replantation with root surface treatment with denosumab 60 mg/mL and 30 mg/mL, respectively, for 10 min both experimentals groups. After that, the root canals were filled with calcium hydroxide and replanted. After 15 and 60 days, the animals were euthanized, and the samples were collected and processed for microscopic analysis. Histological sections were performed, and stained with HE to describe the dental characteristics, measure ankylosis, replacement resorption, and dental resorption by conventional microscopy. Also, was performed Brown & Brenn staining and immunohistochemistry for RANKL, OPG, and periostin. RESULTS: Denosumab 60 mg/mL reducted ankylosis (p < 0.0001), replacement resorption (p < 0.0001), and tooth resorption, 60 days after replantation, compared to untreated replanted teeth (p < 0.005). Lower bacterial contamination in root surface in the denosumab treatment groups was found, regardless of the concentration used (p < 0.001). Also, denosumab treatment inhibited the expression of RANKL without modulating OPG. Periostin was observed in periodontal ligament of replanted tooth, although this labelling was absent in the ankylosis areas, in both experimental periods. CONCLUSION: Treatment of the root surface with denosumab at 60 mg/mL of rat teeth before delayed replantation reduced dental root resorption compared with the untreated teeth after 60 days. CLINICAL RELEVANCE: Survival of a replanted tooth has been a challenge in clinical practice. The use of a medication, such as denosumab, to limit dental root resorption represents an important therapeutical approach.

Vitamin D deficiency is a risk factor for delayed tooth eruption associated with persistent primary tooth.

OBJECTIVE: To verify the association between 25(OH)D level and polymorphisms in the vitamin D receptor gene (VDR) with the disturbance in the dental development and eruption. DESIGN: A total of 183 children from two datasets were evaluated. The first dataset was a case-control (15:15) designed to assess if persistent primary tooth (PPT) is associate with serum 25(OH)D level and with genetic polymorphisms in VDR. The second dataset of genomic DNA samples from 54 children with delayed tooth eruption (DTE) and 99 controls were analysed to verify if genetic polymorphisms in VDR (rs2228570 and rs739837) are associated with DTE. The 25(OH)D and the genotyping/allele distribution were analysed using the T-test and chi-square test, respectively. RESULTS: The level of 25(OH)D in the PPT group (24.9 ± 6.4 mg/mL) was significantly lower than the control (30.0 ± 7.0 mg/mL) (p=.047). Our data show that children with 25(OH)D deficiency are more likely to present PPT (OR = 2.36; 95%CI: 1.51, 3.70). The rs739837 and rs2228570 polymorphisms were not associated with DTE (OR = 1.44; 95%CI: 0.87, 2.39 and OR = 0.80; 95%CI: 0.45, 1.44, respectively). CONCLUSIONS: Vitamin D deficiency is a risk factor for PPT.

Periodontal ligament repair after active splinting of replanted dogs' teeth.

BACKGROUND/AIM: The high rate of root resorption resulting from tooth replantation represents a serious clinical problem. In order to prevent ankylosis and replacement resorption, the contemporary literature highlights the importance of using a flexible stabilization for traumatized teeth. For this purpose, orthodontic devices may be promising for obtaining a better prognosis and periodontal repair. The aim of this study was to evaluate the effect of an active splinting protocol with controlled force in dog's teeth following replantation. MATERIAL AND METHODS: Sixty premolar roots from three dogs were used. They were submitted to endodontic treatment, hemisected, atraumatically extracted and subsequently replanted. They were divided into four groups: Passive Stabilization (n = 20)-after 20 min in a dry medium; Active Stabilization (n = 20)-after 20 min in a dry medium; Negative control (n = 10)-immediate replantation and passive Stabilization; and Positive control (n = 10)-90 min of extra-alveolar time and passive Stabilization. The samples were collected and submitted to histologic processing. They were then evaluated for the count of inflammatory cells, expression of neurotrophin 4, osteoclasts, apoptotic cells and collagen fibres. The results were submitted to ANOVA or Kruskal-Wallis statistical tests followed by Tukey or Dunn post-tests (α = 5%). RESULTS: Passive Stabilization with orthodontic brackets without traction used after replantation had the highest number of inflammatory cells (p = .0122), osteoclasts (p = .0013) and percentage of collagen fibres in the periodontal ligament (p < .0001) when compared to Active Stabilization with orthodontic brackets applying amild tensile force. Neurotrophin 4 had no statistically significant difference (p = .05), regardless of the treatment. The apoptotic cells count revealed statistical differences (p < .0001) between Active Stabilization (189.70 ± 47.99) and Positive Control (198.90 ± 88.92) when compared to Passive Stabilization (21.19 ± 32.94). CONCLUSION: The active splinting protocol using orthodontic appliances generating a light and controlled force favoured periodontal ligament repair of replanted teeth.

Esthetic elastomeric ligatures: Quantification of bacterial endotoxin in vitro and in vivo.

INTRODUCTION: The objective of this study was to evaluate in vitro and in vivo bacterial endotoxin (LPS) adhesion in polyurethane and silicone esthetic elastomeric orthodontic ligatures. The null hypotheses tested were: (1) there is no LPS adhesion in esthetic elastomeric orthodontic ligatures; and (2) there is no difference in the LPS adhesion between different brands of these ligatures. METHODS: For the in vitro study, 4 types of esthetic elastomeric ligatures were used (Sani-Ties and Sili-Ties [Dentsply GAC, Islandia, NY;] and Mini Single Case Ligature Stick and Synergy low-friction ligatures [Rocky Mountain Orthodontics, Denver, Colo]), contaminated or not with endotoxin solution. Replicas of twisted wire and cast stainless steel ligatures were used as control. For the in vivo study, 10 male and 10 female patients, aged 15-30 years, received the same 4 types of ligatures, 1 of each inserted in the maxillary and mandibular canines, randomly. Twenty-one days later, the ligatures were removed, and endotoxin quantification was performed using the Limulus amebocyte lysate test. Data were analyzed (α = 0.05) using the Kruskal-Wallis test and Dunn's posttest or analysis of variance and Tukey's posttest. RESULTS: GAC silicone group had the lowest median contamination (1.15 endotoxin units/mL; P <0.0001) in vitro. In the in vivo study, the GAC silicone group had the lowest mean contamination (0.577 endotoxin units/mL; P <0.001). In both studies, the other groups did not present a significant difference when compared with each other (P >0.05). CONCLUSIONS: LPS exhibited an affinity for all the tested polyurethane and silicone elastomeric ligatures. GAC silicone ligatures presented with lower amounts of LPS attached to their surfaces. Thus, both null hypotheses were rejected.