to explore relationships among serum adipokines, vitamin D, clinical and microbial parameters of chronic periodontitis before and after treatment.
weight, height and smoking status were recorded for 56 patients with chronic periodontitis. Plaque, gingivitis, bleeding on probing (BOP), suppuration, pocket depth (PD) and attachment level (AL) were measured at all teeth present. Subgingival biofilm samples from each tooth were analyzed for levels of 40 bacterial species using checkerboard DNA-DNA hybridization. Serum levels of interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), adiponectin, leptin, resistin and vitamin D were measured at baseline. Sample collection was then performed in a subset of the population 6 months post-therapy (n=17). Serum samples were analyzed using ELISA and immunoassays. Differences in clinical, microbial and serum factors among groups were sought using the Mann-Whitney test. Correlations among factors were evaluated using regression analysis. Effects of therapy were sought using the Wilcoxon signed ranks test
There were positive correlations between adiponectin/vitamin D and between IL-6/leptin; negative correlations between IL-6/vitamin D, and leptin/vitamin D, but no associations between serum analytes and clinical or microbial parameters. Gender and BMI were associated with levels of adipokines. Periodontal therapy improved clinical and microbiological parameters, but did not influence the levels of serum analytes.
Adipokines and IL-6 levels were affected by gender and BMI. Serum analytes were not influenced by periodontal therapy.
adipokines; cytokines; calcitriol; periodontitis; subgingival scaling; biofilm; microbiota
Vitamin D has anti-inflammatory and anti-microbial properties that, together with its influence on bone health, may confer periodontal benefit.
We investigated cross-sectional associations (1997–2000) between plasma 25-hydroxyvitamin D concentrations [25(OH)D] and periodontal measure among 920 postmenopausal women. Chronic measures of disease were defined based on: 1) alveolar crestal height (ACH) measures from intraoral radiographs and tooth loss, and the 2) Center for Disease Control and Prevention (CDC)/American Academy of Periodontology (AAP) criteria using measures of clinical attachment level (CAL) and probing pocket depth (PD). Acute oral inflammation was assessed by the % of gingival sites that bled upon assessment with a probe. Logistic regression was used to estimate the odds ratios (OR) and 95% confidence intervals (CIs) for periodontal disease among participants with adequate ([25(OH)D]≥50 nmol/L) compared to deficient/inadequate ([25(OH)D]<50 nmol/L) vitamin D status adjusted for age, dental visit frequency, and body mass index.
No association was observed between vitamin D status and periodontal disease defined by ACH and tooth loss (adjusted OR=0.96, 95% CI: 0.68–1.35). In contrast, women with adequate compared to deficient/inadequate vitamin D status had a 33% lower odds (95% CI: 5%–53%) of periodontal disease defined using the CDC/AAP definition and a 42% lower odds (95% CI: 21%-58%) of having ≥50% of gingival sites that bled.
Vitamin D status was inversely associated with gingival bleeding, an acute measure of oral health and inflammation and inversely associated with clinical categories of chronic periodontal disease that incorporated PD, an indicator of oral inflammation. However, vitamin D was not associated with chronic periodontal disease based on measures of ACH in combination with tooth loss.
vitamin D; 25-hydroxyvitamin D; periodontal diseases; postmenopausal period; epidemiology; women
Low dietary intakes of vitamin D and calcium hasten bone loss and osteoporosis. Because vitamin D metabolites may also alter the inflammatory response and have anti-microbial effects, we studied whether use of vitamin D and calcium supplements affects periodontal disease status.
A cohort of 51 subjects receiving periodontal maintenance therapy was recruited from 2 dental clinics. Of these, 23 were taking vitamin D (≥400 international units/day) and calcium (≥1000mg/day) supplementation, and 28 were not taking such supplementation. All subjects had ≥2 interproximal sites with ≥3 mm clinical attachment loss. Daily calcium and vitamin D intakes (from food and supplements) were estimated by nutritional analysis. The following clinical parameters of periodontal disease were recorded for the mandibular posterior teeth: gingival index, probing depth, cementoenamel junction-gingival margin distance (attachment loss), bleeding upon probing, and furcation involvement. Posterior photostimulable-phosphor bitewing radiographs were taken to determine cementoenamel-junction-alveolar-crest distances (alveolar crest height loss). Data were analyzed with a repeated-measures, multivariate analysis of variance.
Relative to subjects who did not take vitamin D and calcium supplementation, supplement takers had shallower probing depths, fewer bleeding sites, lower gingival index values, fewer furcation involvements, less attachment loss and less alveolar crest height loss. The repeated-measures analysis indicated that collectively these differences for clinical parameters were borderline significant (P=0.08).
In these subjects receiving periodontal maintenance therapy, there was a trend for better periodontal health with intake of vitamin D and calcium supplementation. More expanded longitudinal studies are required to determine the potential of this relationship.
vitamin D; calcium; chronic periodontitis; alveolar bone
One sentence summary
Women who indicate DMPA use have a significantly increased risk of prevalence of periodontal conditions as compared to women who have never used DMPA.
Maternal periodontal disease diagnosed by a detailed oral health examination is associated with preeclampsia. Our objective was to measure the association between maternal self-report of oral symptoms/problems, oral hygiene practices, and/or dental service utilization prior to or during pregnancy and severe preeclampsia.
A written questionnaire was administered to pregnant women at the time of prenatal ultrasound, and outcomes ascertained by chart abstraction. Chi square test compared maternal oral symptoms/problems, hygiene practices, and dental service utilization between women with severe preeclampsia versus normotensive women. Multivariable logistic regression was used to calculate adjusted odds ratios (aOR) and 95% confidence intervals (CI) for severe preeclampsia. Results: 48 (10%) of 470 women reported ≥ 2 oral symptoms/problems in the 6 months prior to pregnancy and 77 (16%) since pregnancy. 51(11%) reported prior periodontal treatment. 28 (6%) of 470 developed severe preeclampsia. Women with a history of periodontal treatment were more likely to develop severe preeclampsia (aOR, 95%CI: 3.71, 1.40-9.83) than women without a prior history of periodontal treatment. Self-reported oral health symptoms/problems, oral hygiene practices, or dental service utilization prior to or during pregnancy were not associated with severe preeclampsia when considered in the context of other maternal risk factors. Conclusion: Maternal self report of previous periodontal treatment prior to pregnancy is associated with severe preeclampsia.
Pregnancy complications; periodontitis; epidemiology
Periodontitis can ultimately result in tooth loss. Many natural and synthetic materials have been tried to achieve periodontal regeneration, but the results remain variable and unpredictable. We hypothesized that exogenous treatment with dentin matrix protein 1 (DMP1) activates specific genes and results in phenotypic and functional changes in human periodontal ligament stem cells (hPDLSCs).
hPDLSCs were isolated from extracted teeth and cultured in the presence or absence of DMP1. Quantitative polymerase chain reactions were performed to analyze the expression of several genes involved in periodontal regeneration. hPDLSCs were also processed for immunocytochemical and Western blot analysis using phosphorylated extracellular signal-regulated kinase (pERK) and ERK antibodies. Alkaline phosphatase and von Kossa staining were performed to characterize the differentiation of hPDLSCs into osteoblasts. Field emission scanning electron microscopic analysis of the treated and control cell cultures were also performed.
Treatment with DMP1 resulted in the upregulation of genes, such as matrix metalloproteinase-2, alkaline phosphatase, and transforming growth factor β1. Activation of ERK mitogen-activated protein kinase signaling pathway and translocation of pERK from the cytoplasm to the nucleus was observed. Overall, DMP1-treated cells showed increased expression of alkaline phosphatase, increased matrix, and mineralized nodule formation when compared with untreated controls.
DMP1 can orchestrate a coordinated expression of genes and phenotypic changes in hPDLSCs by activation of the ERK signaling pathway, which may provide a valuable strategy for tissue engineering approaches in periodontal regeneration.
Bone; bone regeneration; guided tissue regeneration; periodontal; mitogen-activated protein kinases
This pilot study examined whether a novel diabetes screening approach using gingival crevicular blood (GCB) could be used to test for hemoglobin A1c (HbA1c) during the periodontal visit.
At a large periodontics clinic, finger stick blood (FSB) samples from 120 patients as well as GCB samples from those patients with adequate bleeding on probing were collected on special blood collection cards and were analyzed for HbA1c levels in a laboratory. The Pearson correlation coefficient was used to measure correlation between FSB and GCB HbA1c values for 75 paired FSB and GCB samples. A Receiver Operator Characteristic Curve (ROC) analysis was performed to determine an optimal GCB HbA1c criterion value for a positive diabetes screen.
For the 75 paired samples, the Pearson correlation coefficient was .842. The ROC analysis identified a criterion value of 6.3% for the GCB HbA1c test with high sensitivity (.933) and high specificity (.900) corresponding to FSB HbA1c values of 6.5% or greater (in the diabetes range). Using this GCB HbA1c criterion value for 27 additional paired samples in which there was an unidentified component observed to co-elute within the elution window of GCB HbA1c in the laboratory, there was agreement between FSB and GCB values for 24 of the pairs according to whether they were both within, or both outside of the diabetes range.
Using a criterion value of 6.3%, GCB samples are acceptable for HbA1c testing to screen for diabetes in most persons with bleeding on probing at the GCB collection site.
diabetes mellitus; periodontitis; public health dentistry
This study compared the subgingival microbiota of subjects with refractory periodontitis (RP) to those in subjects with treatable periodontitis (GR) or periodontal health (PH) using the Human Oral Microbe Identification Microarray (HOMIM).
At baseline, subgingival plaque samples were taken from 47 periodontitis and 20 PH individuals, and analyzed for the presence of 300 species by HOMIM. The periodontitis subjects were classified as RP (n=17) based on mean attachment loss (AL) and/or >3 sites with AL ≥2.5 mm after SRP, surgery and systemically administered amoxicillin and metronidazole or as GR (n=30) based on mean attachment gain and no sites with AL ≥2.5 mm after treatment. Significant differences in taxa among groups were sought using the Kruskal Wallis and Chi-square tests.
More species were detected in diseased patients (GR or RP) than those without disease (PH). RP subjects were distinguished from GR and PH by a significantly high frequency of putative periodontal pathogens such as, Parvimonas micra, Campylobacter gracilis, Eubacterium nodatum, Selenomonas noxia, Tannerella forsythia, Porphyromonas gingivalis, Prevotella spp., Treponema spp., Eikenella corrodens, as well as “unusual” species (Pseudoramibacter alactolyticus, TM7 spp. oral taxon (OT) 346/356, Bacteroidetes spp. OT 272/274, Solobacterium moorei, Desulfobulbus sp. OT 041, Brevundimonas diminuta, Sphaerocytophaga sp. OT 337, Shuttleworthia satelles, Filifactor alocis, Dialister invisus/pneumosintes, Granulicatella adiacens, Mogibacterium tidmidum, Veillonella atypica, Mycoplasma salivarium, Synergistes sp. cluster II, Acidaminococcaceae [G-1] sp. OT 132/150/155/148/135) [p<0.05]. Species that were more prevalent in PH than in periodontitis patients included Actinomyces sp. OT 170, Actinomyces spp. cluster I, Capnocytophaga sputigena, Cardiobacterium hominis, Haemophilus parainfluenzae, Lautropia mirabilis, Propionibacterium propionicum, Rothia dentocariosa/mucilagenosa, Streptococcus sanguinis (p<0.05).
RP patients present a distinct microbial profile compared to patients in the GR and PH groups as determined by HOMIM.
Refractory periodontitis; subgingival microbiota; periodontal pathogen; HOMIM; periodontal therapy
Most studies comparing prevalence of periodontal disease and risk factors by using partial protocols were performed in adult populations, with several studies being conducted in clinical settings. The aim of this study is to assess the accuracy of partial protocols in estimating the prevalence of periodontal outcomes in adolescents and young adults from two population-based birth cohorts from Pelotas, Brazil, and to assess differences in the estimation and strength of the effect measures when partial protocols are adopted compared to full-mouth examination.
Gingival bleeding at probing among adolescents (n = 339) and young adults (n = 720) and dental calculus and periodontal probing depth among young adults were assessed using full-mouth examinations and four partial protocols: Ramfjord teeth (RT), community periodontal index (CPI), and two random diagonal quadrants (1 and 3, 2 and 4). Socioeconomic, demographic, and periodontal health-related variables were also collected. Sensitivity, absolute and relative bias, and inflation factors were calculated. Prevalence ratio for each periodontal outcome for the risk factors was estimated.
Two diagonal quadrants showed better accuracy; RT had the worst, whereas CPI presented an intermediate pattern when compared to full-mouth examination. For bleeding assessment in adolescence, RT and CPI underestimated by 18.4% and 16.2%, respectively, the true outcome prevalence, whereas among young adults, all partial protocols underestimated the prevalence. All partial protocols presented similar magnitude of association measures for all investigated periodontal potential risk factors.
Two diagonal quadrants protocol may be effective in identifying the risk factors for the most relevant periodontal outcomes in adolescence and in young adulthood.
Data collection; epidemiologic studies; periodontal index
Periodontal diseases are inflammatory diseases resulting in the destruction of tissues of the periodontium. Although bacteria must be present for periodontal disease to occur, a susceptible host is also required, which is determined by genetic, environmental, and acquired factors. One such factor, autoimmunity, may play a role in the tissue destruction. Data indicate that some antibodies that occur in the gingival lesion are directed to host tissue components, such as type I collagen, although investigations of other periodontal autoimmune targets are limited.
Histologic sections and extracts from periodontally healthy teeth and the associated soft tissues were probed with serum from localized aggressive periodontitis (LAgP), chronic periodontitis (CP), and periodontally healthy subjects to determine autoreactivity to components of the periodontium. Any autoreactivity observed was characterized further by mass spectrometry and enzyme-linked immunosorbent assay.
Autoreactivity to components of the periodontium was observed in CP and LAgP. Known autoimmune targets, such as collagen and heat shock protein, were identified along with multiple potential autoimmune targets, including members of the extracellular matrix, such as vimentin, spectrin, filamin, actin, lamin, keratin, and tubulin. Finally, it was determined that the autoreactivity observed in LAgP was more severe and diverse than that observed in CP.
These data demonstrated that autoimmune reactivity can play a role in the tissue destruction of periodontal disease but that the nature of the autoreactivity may differ based on the type and/or stage of periodontal disease.
Antibody; autoimmunity; collagen; periodontal diseases
Reports from studies of twins, disease aggregation in families, animal models for periodontal disease, and various genetic-analysis studies have determined that genetics plays a role in the susceptibility to periodontal disease. The purpose of this pilot study was to evaluate the effect of genetics on periodontal disease by evaluating the heritability of alveolar bone loss in a captive baboon population.
A collection of baboon skulls from a pedigreed colony (for which scientists and veterinarians maintain complete genealogical and veterinary records) were obtained from the Southwest National Primate Research Center and used in this pilot study. Measurements of alveolar bone loss were performed on 390 dry baboon skulls. A periodontal probe was used to measure alveolar bone loss. Maximum likelihood methods (designed to handle complex genealogies) were used to determine the heritability of alveolar bone loss. This software utilized known pedigrees in the captive baboon sample and tested the relationship between pairwise kinship and alveolar bone loss data to determine the heritability of alveolar bone loss from periodontal disease.
Genetic data were available for 347 of the 390 specimens. Using age and sex as covariates, genetic analysis indicated a heritability of 35% (standard error=20%, p=0.01). While sex was not a significant factor in periodontal disease (p=0.96), age was highly significantly associated with periodontal disease (p<0.0001).
In this pilot study, analysis of alveolar bone loss measurements from captive baboons indicates that bone loss increases with age and that a portion of periodontal disease risk may be due to genetic variance. These findings provide evidence that periodontal disease is heritable in captive baboons and indicate that a larger, more-detailed study is warranted.
periodontitis; alveolar bone; genetics; baboons
Current scientific evidence addressing the relation between periodontitis and hypertension is limited to a few studies producing inconsistent results.
All participants of an on-going representative cohort of Puerto Rican elderly who were 70 years and older and residing in San Juan metropolitan area were invited to this cross-sectional study. Periodontal probing depth (PD) and attachment loss (AL) were summarized using CDC-AAP definition for severe periodontitis (≥2 teeth with AL ≥6mm and ≥1 tooth with PD ≥5mm). We averaged three repeated blood pressure (BP) measurements taken using a standardized auscultatory method. Information on hypertension history, use of anti-hypertensive medications and potential confounders (age, gender, smoking, heavy and binge drinking, diabetes, utilization of preventive dental services, flossing, body mass index, fruit and vegetable, whole wheat bread and high-fiber cereal consumption) was collected during in-person interviews. High BP was defined as average systolic BP ≥140 mmHg or diastolic ≥90 mmHg. Multivariate logistic regression models were used to study the relation between severe periodontitis, hypertension history and high BP.
The study population comprised 182 adults. In multivariate analysis, there was no association between severe periodontitis and hypertension history (OR=0.99, 95% CI: 0.40–2.48). Severe periodontitis was associated with high BP, with OR of 2.93 (95% CI: 1.25–6.84), after adjusting for age, gender, smoking, and binge drinking. This association was stronger when restricted to those with hypertension or taking anti-hypertensive medications: OR=4.20 (95% CI: 1.28–13.80).
Our results suggest that periodontitis may contribute to poor blood pressure control among older adults.
Periodontal diseases; periodontitis; hypertension; blood pressure
The periodontal pathogen Actinobacillus actinomycetemcomitans produces a leukotoxin that is considered a primary virulence factor in localized juvenile Periodontitis (LJP). Select strains of the bacterium contain a 530-bp deletion in the promoter region of the leukotoxin gene operon which results in enhanced transcription of the leukotoxin. DNA hybridization and polymerase chain reaction (PCR) were used to examine genetic variants of A. actinomycetemcomitans in 24 LJP-susceptible children from 21 families having a history of the disease and 34 control children from non-LJP families. A significant association was found between the detection of variants that had a deletion in the leukotoxin promoter region, indicative of a high level expression leukotoxin genotype, and conversion from a healthy periodontal status to disease. Subjects harboring A. actinomycetemcomitans of this genotype were more likely to convert to LJP than those subjects who had variants containing the full length leukotoxin promoter region (odds ratio = 22.50, 95% C.I.). These findings support the concept that highly virulent strains or clonal types of periodontal pathogens play a major role in the initiation of periodontal disease in susceptible hosts.
Periodontitis; juvenile/epidemiology; Actinobacillus actinomycetemcomitans; leukotoxin; epidemiology; polymerase chain reaction; polymorphism; restriction fragment length
Biofilm-induced inflammatory osteolytic oral infections, such as periodontitis and peri-implantitis, have complex etiology and pathogenesis. A significant obstacle to research has been the lack of appropriate animal models where the inflammatory response to biofilms can be investigated. The aim of this study is to develop a novel animal model to study the host response to Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans)–biofilm colonizing titanium implants.
Titanium implants were inoculated in vitro with A. actinomycetemcomitans, establishing a biofilm for 1 to 3 days. Biofilm-inoculated and control implants were transmucosally placed into rat hard palate or alveolar ridge. Analysis included documentation of clinical inflammation, polymerase chain reaction, and culture detection of A. actinomycetemcomitans and microcomputed tomography quantitation of peri-implant bone volume.
Viable A. actinomycetemcomitans biofilm was successfully established on titanium implants in vitro, detected by confocal laser scanning microscopy. An inflammatory response characterized by clinical inflammation, bleeding, ulceration, hyperplasia, and necrosis was observed around biofilm-inoculated implants. A. actinomycetemcomitans was detected by polymerase chain reaction and culture analysis on 100%of biofilm-inoculated implants for up to 3 weeks and 25%for up to 6 weeks. Microcomputed tomography analysis demonstrated significantly lower bone volume (P <0.05) around biofilm-inoculated implants (29.6% ± 7.6%) compared to non-inoculated implants (50.5% ± 9.6%) after 6 weeks.
These results describe a novel animal model where A. actinomycetemcomitans biofilm was established in vitro on titanium implants before placement in rat oral cavity, leading to an inflammatory response, osteolysis, and tissue destruction. This model may have potential use for investigation of host responses to biofilm pathogens and antibiofilm therapy.
Aggregatibacter actinomycetemcomitans; biofilms; dental implants; infection; models; animal; rats
Dental biofilms contain a protein that inhibits mammalian cell growth, possibly lysine decarboxylase from Eikenella corrodens. This enzyme decarboxylates lysine, an essential amino acid for dentally attached cell turnover in gingival sulci. Lysine depletion may stop this turnover, impairing the barrier to bacterial compounds. The aims of this study were to determine biofilm lysine and cadaverine contents before oral hygiene restriction (OHR), and their association with plaque index (PI) and gingival crevicular fluid (GCF) after OHR for a week.
Laser-induced fluorescence after capillary electrophoresis was used to determine lysine and cadaverine contents in dental biofilm, tongue biofilm and saliva before OHR and in dental biofilm after OHR.
Before OHR, lysine and cadaverine contents of dental biofilm were similar and 10-fold greater than in saliva or tongue biofilm. After a week of OHR, the biofilm content of cadaverine increased and that of lysine decreased, consistent with greater biofilm lysine decarboxylase activity. Regression indicated that PI and GCF exudation were positively related to biofilm lysine post-OHR, unless biofilm lysine exceeded the minimal blood plasma content in which case PI was further increased but GCF exudation was reduced.
After OHR, lysine decarboxylase activity seems to determine biofilm lysine content and biofilm accumulation. When biofilm lysine exceeds minimal blood plasma content after OHR, less GCF appeared despite more biofilm. Lysine appears important for biofilm accumulation and the epithelial barrier to bacterial proinflammatory agents.
Inhibiting lysine decarboxylase may retard the increased GCF exudation required for microbial development and gingivitis.
Gingival crevicular fluid; Gingivitis; Oral hygiene; Pathogenesis of periodontal disease(s); Microbiology
We previously reported in a cross-sectional study that patients who were in periodontal maintenance programs and were taking vitamin D and calcium supplementation had a trend for better periodontal health compared with patients not taking supplementation. The objective of the present study was to determine, for the same group of subjects, whether there was a difference in periodontal health over a one–year period.
Fifty-one patients enrolled in maintenance programs from two dental clinics were recruited. Twenty-three were taking vitamin D (≥400 international units/day) and calcium (≥1000mg/day) supplementation, and twenty-eight were not taking supplementation. All subjects had ≥2 interproximal sites with ≥3 mm clinical attachment loss. For mandibular-posterior teeth, these clinical parameters were recorded: gingival index, plaque index, probing depth, attachment loss, bleeding upon probing, calculus index and furcation involvement. Photostimulable-phosphor, posterior bitewing radiographs were taken to assess alveolar bone. Daily vitamin D and calcium intakes were estimated by nutritional analysis. Data were collected at baseline, 6 months, and 12 months.
Clinical parameters improved with time in both groups (p<0.01). When clinical measures were considered collectively, the results were borderline significant at baseline (p=0.061), significant at 6 months (p=0.049) but not significant at 12 months (p=0.114). After adjusting for covariates, the effect of supplements was significant at baseline (p=0.037), borderline at 6 months (p=0.058) and not significant at 12 months (p=0.142)
Calcium and vitamin D supplementation has a modest positive effect on periodontal health, and consistent dental care improves clinical parameters of periodontal disease regardless of such supplements. Calcium and vitamin D supplementation has a modest positive effect on periodontal health, and consistent dental care improves clinical parameters of periodontal disease regardless of such supplements. Our findings raise the possibility that vitamin D, perhaps at higher doses, may positively impact on periodontal disease severity.
vitamin D; calcium; chronic periodontitis; alveolar bone
Inflammatory periodontal diseases are a leading cause of tooth loss and are linked to multiple systemic conditions, such as cardiovascular disease and stroke. Reconstruction of the support and function of affected tooth-supporting tissues represents an important therapeutic endpoint for periodontal regenerative medicine. An improved understanding of periodontal biology coupled with current advances in scaffolding matrices has introduced novel treatments that use cell and gene therapy to enhance periodontal tissue reconstruction and its biomechanical integration. Cell and gene delivery technologies have the potential to overcome limitations associated with existing periodontal therapies, and may provide a new direction in sustainable inflammation control and more predictable tissue regeneration of supporting alveolar bone, periodontal ligament, and cementum. This review provides clinicians with the current status of these early-stage and emerging cell- and gene-based therapeutics in periodontal regenerative medicine, and introduces their future application in clinical periodontal treatment. The paper concludes with prospects on the application of cell and gene tissue engineering technologies for reconstructive periodontology.
Macrolide antibiotics yield high concentrations in inflamed tissue, suggesting that their levels in gingival crevicular fluid (GCF) could be increased at gingivitis sites. However, the increased volume of GCF associated with gingivitis could potentially dilute macrolides. To determine whether these assumptions are correct, the bioavailability of systemically-administered azithromycin was compared in GCF from healthy and gingivitis sites.
Materials and methods
Experimental gingivitis was induced in one maxillary posterior sextant in nine healthy subjects. Contralateral healthy sextants served as controls. Subjects ingested 500 mg of azithromycin followed by a 250 mg dose 24 hours later. Four hours after the second dose, plaque was removed from experimental sites. GCF was collected from 8 surfaces in both the experimental and control sextants and pooled separately. GCF samples were subsequently collected on the 2nd, 3rd, 8th and 15th days and azithromycin content was determined by agar diffusion bioassay.
On days 2 and 3, the pooled GCF volume at experimental sites was significantly higher than at control sites (P <0.01) and the total azithromycin mass in 30 second GCF samples pooled from experimental sites was significantly higher than at control sites (P < 0.02). However, there were no significant differences in azithromycin concentration between the experimental and control pools at any point. Concentrations exceeded 7.3 μg/ml on day 2 and 2.5 μg/ml on day 15.
Azithromycin concentrations are similar in GCF from gingivitis sites and healthy sites, suggesting that the processes that regulate GCF azithromycin concentration can compensate for local inflammatory changes.
Anti-infective agents; inflammation; pharmacokinetics
Understanding the molecular features of bone repair and osseointegration may aid in the development of therapeutics to improve implant outcomes. The purpose of this investigation is to determine the gene expression dynamics during alveolar bone repair and implant osseointegration.
An implant osseointegration preclinical animal model was used whereby maxillary defects were created at the time of oral implant placement, while a tooth extraction socket healing model was established on the contralateral side of each animal. The surrounding tissues in the zone of the healing defects were harvested during regeneration for temporal evaluation using histology, immunohistochemistry, laser capture microdissection, and quantitative reverse transcription–polymerase chain reaction for the identification of a panel of 17 putative genes associated with wound repair.
In both models, three distinct expression patterns were displayed: 1) genes that are slowly increased during the healing process, such as bone morphogenetic protein 4, runt-related transcription factor 2, and osteocalcin; 2) genes that are upregulated at the early stage of healing and then downregulated at later stages, such as interleukin and chemokine (C-X-C motif) ligands 2 and 5; and 3) genes that are constitutively expressed over time, such as scleraxis. Although some similarities between osseointegration and tooth extraction socket were seen, distinct features developed and triggered a characteristic coordinated expression and orchestration of transcription factors, growth factors, extracellular matrix molecules, and chemokines.
Characterization of these events contributes to a better understanding of cooperative molecular dynamics in alveolar bone healing, and highlights potential pathways that could be further explored for the enhancement of osseous regenerative strategies.
Analysis of samplings from periodontal pockets is important in diagnosis and therapy control of periodontitis. In this study, three different sampling techniques were compared to determine if one method can yield samples suitable for reproducible and simultaneous determination of bacterial load, cytokines, neutrophil elastase, and Arg-specific gingipains. R-gingipains are an important virulence factor of Porphyromonas gingivalis, the exact concentration of which in gingival crevicular fluid (GCF) has not yet been quantified.
GCF was sampled from four sites per patient (each two sites one method) in 36 chronic periodontitis patients. One week later, the procedure was repeated with alternative methods. The variables that had been determined were: loads of Aggregatibacter actinomycetemcomitans and P. gingivalis, levels of interleukin-6 and interleukin-8, activity of neutrophil elastase and level of R-gingipains.
The detected cytokine levels were higher using paper strips compared to paper points. Bacteria were found in similar loads from the paper strips and paper points. R-gingipains were detectable in high quantities only by washing of the periodontal pocket. The level of R-gingipains correlated with the load of P. gingivalis.
The use of paper strips is suitable for simultaneous determination of microbial and immunological parameters. Obtaining GCF by washing can be useful for special purposes. Gingipain concentration in periodontal pockets was directly determined to be up to 1.5 μM. This value indicates that most of so far identified substrates of these proteases by in vitro assays can be easily degraded in P. gingivalis infected sites.
gingival crevicular fluid; sampling techniques; Porphyromonas gingivalis; Arg-gingipain; cytokines
Fluoxetine, a selective serotonin reuptake inhibitor, has recently been found to possess anti-inflammatory properties. The present study investigated the effects of fluoxetine on inflammatory tissue destruction in a rat model of ligature-induced periodontitis (PD).
Male Wistar rats were randomly assigned into three groups (n=10 animals/group): 1) Control rats (without ligature); 2) rats with ligature + placebo (saline; oral gavage); 3) rats with ligature + fluoxetine (20 mg/kg/day in saline; oral gavage). Histological analyses were performed on the furcation region and mesial of mandibular first molars of rats sacrificed at 15 days after ligature-induced PD. Reverse transcriptase-polymerase chain reaction (RT-PCR) and zymography were performed to analyze the mRNA expression of interleukin (IL)-1β, cyclooxygenase (COX)-2, matrix metalloproteinase (MMP)-9 and inducible nitric oxide synthase (iNOS), and the MMP-9 activity, respectively, in gingival tissues samples.
Compared to the ligature + placebo group, alveolar bone loss was reduced in the fluoxetine group (P < 0.05), and the integrity of collagen fibers in the gingival tissue was maintained. Moreover, in gingival tissue sampled 3 days after ligature attachment, fluoxetine administration reduced IL-1β and COX-2 mRNA expression. Fluoxetine down-regulated MMP-9 activity, without affecting MMP-9 mRNA expression induced by ligature, compared to the ligature + placebo group (P < 0.05). These data suggested that fluoxetine suppressed proinflammatory responses, as well as proteolytic enzyme activity, induced by ligature.
In the present study, fluoxetine suppressed the inflammatory response and protected against periodontal bone resorption and destruction of collagen fibers, suggesting that fluoxetine can constitute a promising therapeutic approach for periodontal diseases.
Fluoxetine; inflammation; periodontitis; bone resorption; collagen
The proinflammatory chemokine interleukin-8 (IL8) is important in the regulation of the inflammatory response. Single nucleotide polymorphism (SNP) rs4073 have shown that the A allele up regulates the IL8 levels after stimulation with lipopolysaccharides. We investigated the association of the SNP rs4073 with chronic periodontitis.
Genotyping was performed by standard polymerase chain reaction-restriction fragment length polymorphism assay in 289 genomic DNA samples of healthy control and chronic periodontitis patients; analyses were adjusted by the multivariate logistic regression modeling. Real-time PCR performance was used to detect the levels of the IL8 mRNA.
Analysis points to a statistical significant association of chronic periodontitis with the heterozygous TA genotype (p = 0.001); the results showed an increase in the frequency of the A allele in the diseased group (36% in the control × 48% in the periodontitis group). The higher levels of the IL8 mRNA were found in the periodontitis group, mainly in individuals who presented the TA genotype (p=0.03).
The SNP rs4073 is associated with chronic periodontitis, in non-smoker Brazilian subjects, since the frequency of A allele is higher in the disease than control group and the TA genotype was associated with increased levels of IL8 mRNA transcripts.
periodontitis; polymorphism; single nucleotide; Interleukin-8; genotype; alleles
Inflammatory stimuli activate inducible nitric oxide synthase (iNOS) in a variety of cell types, including osteoclasts (OC) and osteoblasts, resulting in sustained NO production. In this study, we evaluate the alveolar bone loss in rats with periodontitis under long-term iNOS inhibition, and the differentiation and activity of OC from iNOS-knockout (KO) mice in vitro.
Oral aminoguanidine (an iNOS inhibitor) or water treatment was started 2 weeks before induction of periodontitis. Rats were sacrificed 3, 7, or 14 days after ligature placement, and alveolar bone loss was evaluated. In vitro OC culture experiments were also performed to study the differentiation of freshly isolated bone marrow cells from both iNOS KO and wild-type C57BL/6 mice. OC were counted 6 days later after tartrate-resistant acid phosphatase staining (a marker of osteoclast identity), and bone resorption activity was assessed by counting the number of resorption pits on dentin disks.
Rats with ligature showed progressive and significant alveolar bone loss compared to sham animals, and aminoguanidine treatment significantly inhibited ligature-induced bone loss at 7 and 14 days after the induction. In comparison to bone marrow cells from wild-type mice, cells from iNOS KO mice showed decreased OC growth and the resulting OC covered a smaller culture dish area and generated fewer resorption pit counts.
Our results demonstrate that iNOS inhibition prevents alveolar bone loss in a rat model of ligature-induced periodontitis, thus confirming that iNOS-derived NO plays a crucial role in the pathogenesis of periodontitis, probably by stimulating OC differentiation and activity.
Animal experimental use; bone resorption; iNOS enzyme; nitric oxide; osteoclasts; periodontitis
A major cause of chronic inflammatory periodontal diseases is Porphyromonas gingivalis (P. gingivalis), a non-motile, gram-negative, rod-shaped, anaerobic bacterium. Within gingival tissue, both macrophages and fibroblasts participate in the immune response to foreign entities by releasing cytokines and expressing molecules to recruit and activate lymphocytes. However, the contribution of gingival macrophages and fibroblasts to the immune response to P. gingivalis infection is not fully known.
The AMJ2-C8 cell line (AM cells), a mouse alveolar macrophage cell line, and ESK-1 cells, a mouse gingival fibroblast cell line made in our laboratory, were treated with lipopolysaccharide (LPS) from either P. gingivalis or Escherichia coli (E. coli). The expression of immune response molecules was quantified by real-time PCR and enzyme-linked immunoassay.
AM and ESK-1 cells responded differently to P. gingivalis and E. coli LPS stimulation. The ESK-1 gingival fibroblast cell line was more responsive to E. coli LPS stimulation as seen by elevated levels of interleukin (IL)-6, inducible nitric oxide (iNOS), and monocyte chemotactic protein-1 (MCP-1) expression relative to stimulation by P. gingivalis LPS. Conversely, the AM macrophage cell line was more responsive to P. gingivalis LPS stimulation, particularly for interleukin IL-1β, IL-6, and MCP-1, relative to stimulation by E. coli LPS.
These findings demonstrate that E. coli LPS induces a stronger cytokine/chemokine response in gingival fibroblasts, while P. gingivalis LPS induces a stronger response in macrophages.
cytokines; fibroblasts; host response; immunology; inflammation and innate immunity
This report is a further analysis of a study designed to determine clinical and microbial risk indicators for progressing periodontitis.
MATERIAL AND METHODS
One hundred and ninety subjects periodontally healthy or adults with early signs of periodontitis (20–40 years) were monitored clinically at 6-month intervals followed by supragingival cleaning. At each visit, GCF and blood were collected for determination of IL-1β content (GCF) and IL-1 genotype (blood). Inter-proximal sites with >1.5 mm increase in clinical attachment over 18 months were considered disease active. Characteristics were compared between active and inactive subjects.
IL-1β levels in GCF increased with severity of disease and correlated well with clinical signs of incipient disease. However, IL-1 genotype did not show any significant associations with disease or extent of disease.
Indicators of inflammation may be important clinical determinants of future periodontal disease progression, but IL-1 genotype was not a risk indictor for early (slight) periodontitis as defined in this subject population.
genotype; gingival crevicular fluid; health; Periodontitis; race/ethnicity smoking