Ulvi K. Gursoy

Salivary MMP‐8, TIMP‐1, and ICTP as markers of advanced periodontitis

AIM Salivary matrix metalloproteinase (MMP)-8 and -14, tissue inhibitor of matrix metalloproteinase (TIMP)-1, and pyridinoline cross-linked carboxyterminal telopeptide of type I collagen (ICTP) were analysed aiming to detect potential markers of advanced periodontitis in saliva. In addition, we compared two MMP-8 detection methods, a time-resolved immunofluorometric assay (IFMA) and an enzyme-linked immunoassay (ELISA), to differentiate periodontitis subjects from controls. MATERIAL AND METHODS Concentrations of MMP-8, MMP-14, TIMP-1, and ICTP were analysed from salivary specimens of 165 subjects, including 84 subjects having at least 14 teeth with periodontal pocket (pocket depth > or =4 mm) and 81 subjects without pocket depth as their controls. RESULTS Salivary MMP-8 detection by IFMA differentiated periodontitis subjects from controls more strongly than by ELISA. Salivary MMP-8, TIMP-1, and ICTP concentrations were higher in periodontitis subjects than those in controls. When only smokers were included in the analysis these differences were lost. The MMP-8/TIMP-1 ratio and the combination of MMP-8 and ICTP differentiated periodontitis and control groups even in smoker subjects. CONCLUSION Salivary MMP-8, TIMP-1, ICTP, and especially their ratios and combinations are potential candidates in the detection of advanced periodontitis. Differentiating periodontitis and control subjects with salivary MMP-8 detection is dependent on the selected techniques.

Salivary MMP-8, TIMP-1, and ICTP as markers of advanced periodontitis

AIM Salivary matrix metalloproteinase (MMP)-8 and -14, tissue inhibitor of matrix metalloproteinase (TIMP)-1, and pyridinoline cross-linked carboxyterminal telopeptide of type I collagen (ICTP) were analysed aiming to detect potential markers of advanced periodontitis in saliva. In addition, we compared two MMP-8 detection methods, a time-resolved immunofluorometric assay (IFMA) and an enzyme-linked immunoassay (ELISA), to differentiate periodontitis subjects from controls. MATERIAL AND METHODS Concentrations of MMP-8, MMP-14, TIMP-1, and ICTP were analysed from salivary specimens of 165 subjects, including 84 subjects having at least 14 teeth with periodontal pocket (pocket depth > or =4 mm) and 81 subjects without pocket depth as their controls. RESULTS Salivary MMP-8 detection by IFMA differentiated periodontitis subjects from controls more strongly than by ELISA. Salivary MMP-8, TIMP-1, and ICTP concentrations were higher in periodontitis subjects than those in controls. When only smokers were included in the analysis these differences were lost. The MMP-8/TIMP-1 ratio and the combination of MMP-8 and ICTP differentiated periodontitis and control groups even in smoker subjects. CONCLUSION Salivary MMP-8, TIMP-1, ICTP, and especially their ratios and combinations are potential candidates in the detection of advanced periodontitis. Differentiating periodontitis and control subjects with salivary MMP-8 detection is dependent on the selected techniques.

Analysis of matrix metalloproteinases, especially MMP-8, in gingival creviclular fluid, mouthrinse and saliva for monitoring periodontal diseases.

Matrix metalloproteinase-8 is a promising candidate biomarker for oral fluid (gingival crevicular fluid, peri-implant sulcular fluid and saliva) and mouthrinse chair-side/point-of-care diagnostics to predict, diagnose and determine the progressive phases of episodic periodontitis and peri-implantitis, as well as to monitor the treatments and medications. Matrix metalloproteinase-8 can be used alone or together with interleukin-1beta and Porphyromonas gingivalis to calculate cumulative risk score at the subject level as a successful diagnostic tool, especially in large-scale public health surveys, in which a thorough periodontal examination is not feasible.

Salivary biomarkers of bacterial burden, inflammatory response, and tissue destruction in periodontitis

AIM Chronic periodontitis has an episodic and multifactorial character, with fluctuations in bacterial burden, inflammatory response, and tissue destruction. We investigated the association of selected salivary biomarkers with periodontal parameters and validated the use of a novel salivary diagnostic approach, the cumulative risk score (CRS), in detection of periodontitis in subjects with angiographically verified coronary artery disease diagnosis. MATERIALS AND METHODS The concentrations of matrix metalloproteinase (MMP)-8, interleukin (IL)-1β, and Porphyromonas gingivalis were analysed from saliva of 493 subjects. The subjects participated in a detailed clinical and radiographic oral examination. The CRS index, combining the three salivary biomarkers, was calculated for each subject. RESULTS High salivary concentrations of MMP-8, IL-1β, and P. gingivalis were associated with deepened periodontal pockets and alveolar bone loss, and MMP-8 and IL-1β with bleeding on probing. The CRS index had a stronger association with moderate to severe periodontitis (OR 6.13; 95% CI 3.11-12.09) than any of the markers alone. CONCLUSIONS Salivary concentrations of MMP-8, IL-1β, and P. gingivalis are associated with various clinical and radiographic measures of periodontitis. The CRS index, combining the three salivary biomarkers, is associated with periodontitis more strongly than any of the markers alone regardless of the coronary artery disease status of the patients.

Use of host- and bacteria-derived salivary markers in detection of periodontitis: a cumulative approach.

In the present study, we propose a novel diagnostic approach, using 3 different salivary markers, representing periodontal pathogen burden, inflammation, and tissue degradation, for detecting periodontitis. The salivary concentrations of Porphyromonas gingivalis, interleukin-1β, and matrix metalloproteinase-8, available from salivary specimens of 165 subjects (84 subjects with advanced periodontitis and 81 controls), were calculated together to obtain a cumulative risk score (CRS). In the calculation of CRS, the concentrations of each marker were divided into tertiles, and cumulative sub-score per each subject were calculated by the multiplication of the tertile values. Three CRS groups, indicating the lowest, medium, or highest risk, were formed with the cumulative sub-scores. Logistic regression analysis and ROC curves were performed to study the association of CRS with periodontitis. The results indicate that CRS, calculated from the 3 salivary biomarkers, is associated with advanced periodontitis more strongly than any of the markers individually. CRS offers a novel, non-invasive model for advanced periodontitis risk categorization that is especially useful in large population surveys where a periodontal examination is not feasible.

Periodontal pathogen carriage, rather than periodontitis, determines the serum antibody levels

AIM We investigated in a nationally representative sample, how periodontitis modifies the association between the carriage of periodontal pathogens and serology. MATERIALS AND METHODS The population comprised 1586 dentate subjects who participated in an interview, clinical and radiological oral health examination, and saliva collection. Serum immunoglobulin A (IgA)- and IgG-class antibody levels against Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis and their salivary occurrence were determined in the whole population. The quantity of the pathogens was measured in a subpopulation. RESULTS In the univariate analyses, the corresponding antibody levels were higher in the pathogen carriers compared with the non-carriers, and clearly higher in the carriers with periodontal pockets compared with the carriers without. In the multi-variate analyses, however, all antibody levels associated strongly with age (p<0.001) and the carriage of the corresponding pathogen (p<0.001), but only weakly with the presence or number of teeth with periodontal pockets. In the subpopulation, the antibody levels and the numbers of corresponding bacteria in saliva had a positive association, which was not affected by the disease. CONCLUSIONS The carriage of A. actinomycetemcomitans and P. gingivalis is the strongest determinant of the systemic antibody response to these pathogens, and the extent of periodontitis has at most a modest modifying effect.

Biofilm Formation Enhances the Oxygen Tolerance and Invasiveness of Fusobacterium nucleatum in an Oral Mucosa Culture Model

BACKGROUND The present study evaluates the survival capability of Fusobacterium nucleatum strains in an aerobic environment and compares the invasive capability of F. nucleatum in biofilm and planktonic forms in an organotypic cell culture (OCC) model. METHODS Biofilms of F. nucleatum American Type Culture Collection (ATCC) 25586 or Anaerobe Helsinki Negative (AHN) 9508 were produced by culturing on semipermeable membranes on brucella agar plates. The oxygen tolerance of the F. nucleatum strains was examined by incubating 3-day-old anaerobically grown biofilms in an aerobic environment (CO(2) [5% in air] incubator) for an additional 48 hours. The OCC model was constructed by seeding keratinocytes on a fibroblast-containing collagen gel. In invasion assays, a 3-day-old anaerobically grown biofilm (and planktonic bacteria in solution as the control) was placed upside down on the top of OCC and incubated under 5% CO(2) for 24 hours. Invasion of the bacteria and morphologic changes in OCC were assessed using hematoxylin and eosin, Ki-67, and periodic acid-Schiff stainings. RESULTS In biofilms, both F. nucleatum strains continuously increased their cell numbers in an aerobic environment for 48 hours. After incubating the bacterial biofilm in contact with the OCC model, F. nucleatum AHN 9508 was able to pass through the epithelial/basement membrane barrier and invade the collagen matrix. The invasiveness of biofilm F. nucleatum ATCC 25586 was limited to the epithelium. Cytotoxic effects and invasiveness of F. nucleatum on the OCC were much stronger when the bacteria were in biofilms than in the planktonic form. CONCLUSION Biofilm formation regulates the survival and invasiveness of F. nucleatum in an aerobic environment.

Understanding the roles of gingival beta-defensins.

Abstract Gingival epithelium produces β-defensins, small cationic peptides, as part of its contribution to the innate host defense against the bacterial challenge that is constantly present in the oral cavity. Besides their functions in healthy gingival tissues, β-defensins are involved in the initiation and progression, as well as restriction of periodontal tissue destruction, by acting as antimicrobial, chemotactic, and anti-inflammatory agents. In this article, we review the common knowledge about β-defensins, coming from in vivo and in vitro monolayer studies, and present new aspects, based on the experience on three-dimensional organotypic culture models, to the important role of gingival β-defensins in homeostasis of the periodontium.

A Novel Organotypic Dento-Epithelial Culture Model: Effect of Fusobacterium nucleatum Biofilm on B-Defensin-2,-3, and LL-37 Expression

BACKGROUND In the present study, the expression and localization of three epithelial peptides (human β-defensin [hBD]-2 and -3, and cathelicidin [LL-37]) are studied in an organotypic dento-epithelial (OD-E) model exposed to Fusobacterium nucleatum (Fn) biofilm. METHODS Biofilm of Fn ATCC 25586 or AHN 9508 were produced by culturing each strain on semipermeable membranes. The OD-E model was constructed by seeding keratinocytes on fibroblast-containing collagen gels and by placing dentin pieces on the top. A 3-day-old biofilm was placed on the top of the OD-E and the coculture was incubated for 5 hours or 24 hours. Production of epithelial antimicrobial peptides was determined immunohistochemically. RESULTS After 5 hours of incubation, the biofilm of each Fn strain stimulated expression of hBD-2 and -3. hBD-2 was localized on superficial layers and hBD-3 on basal cell layers of the epithelium and dento-epithelial junctions, whereas LL-37 was only weakly expressed. After 24 hours, hBD-2 expression was extended toward basal cell layers of the epithelium. In contrast, hBD-3 expression extended toward superficial layers of the epithelium. In the case of Fn AHN 9508 biofilm, LL-37 was localized in the cell layers of the dento-epithelial junction. CONCLUSION In our OD-E model, epithelial antimicrobial peptide responses to Fn biofilms have distinct regulation and localization characteristics, resembling those known to occur in the gingival epithelium in vivo.

High Salivary Estrogen and Risk of Developing Pregnancy Gingivitis

BACKGROUND Estrogen regulates the cellular functions of several tissues that may disturb the host response against bacteria. The present aim is to evaluate the contribution of estrogen to the severity of gingival inflammation during pregnancy. METHODS Salivary estrogen levels from 30 pregnant and 24 non-pregnant females were related to their periodontal health parameters, including visible plaque index (VPI) and bleeding on probing (BOP) from six sites per tooth. The pregnant group was examined three times during pregnancy and twice during postpartum, and the non-pregnant group was examined three times, once per subsequent month. RESULTS Salivary estrogen levels increased significantly during the second (P <0.01) and third (P <0.05) trimesters. In both participant groups, BOP scores correlated significantly with VPI scores (r = 0.498 to 0.870) but not with estrogen levels. In all trimesters and postpartum, the individuals with both high estrogen and high VPI levels had the highest frequency of pregnancy gingivitis. During the second and third trimesters, simultaneously enhanced estrogen levels and VPI scores brought an additional risk of developing gingivitis compared with a high VPI score alone. CONCLUSION The present findings suggest that, during pregnancy, the estrogen level determines the magnitude of gingival inflammation developed against microbial plaque at the gingival margin.