The Potential Role of Matrix Metalloproteinases 8 and 9 and Myeloperoxidase in Predicting Outcomes of Bacterial Meningitis of Childhood.

BACKGROUND: Matrix metalloproteinases (MMPs) and myeloperoxidase (MPO) contribute to the inflammatory cascade in the cerebrospinal fluid (CSF) during bacterial meningitis. We determined levels of MPO, MMP-8, MMP-9, and tissue inhibitor of metalloproteinase- (TIMP-) 1 in the CSF of children with bacterial meningitis and investigated how these inflammatory mediators relate to each other and to the disease outcomes. METHODS: Clinical data and the diagnostic CSF samples from 245 children (median age eight months) with bacterial meningitis were obtained from a clinical trial in Latin America in 1996-2003. MMP-9 levels in the CSF were assessed by zymography, while MMP-8, MPO, and TIMP-1 concentrations were determined with immunofluorometric and enzyme-linked immunosorbent assays. RESULTS: MPO correlated positively with MMP-8 (rho 0.496, P < 0.001) and MMP-9 (rho 0.153, P = 0.02) but negatively with TIMP-1 (rho -0.361, P < 0.001). MMP-8 emerged as the best predictor of disease outcomes: a CSF MMP-8 concentration above the median increased the odds of death 4.9-fold (95% confidence interval 1.8-12.9). CONCLUSIONS: CSF MMP-8 presented as an attractive prognostic marker in children with bacterial meningitis.

Comparison of gender identification using exfoliated cells obtained from toothbrush and miswak: A longitudinal study.

Gender identification plays a pivotal role in forensic medicine. Among the various methods used for gender identification, deoxyribose nucleic acid (DNA) based methods are considered accurate. Exfoliated oral mucosal cells that are harvested from oral hygiene aids can be potentially used for gender identification using real-time polymerase chain rection (PCR). The aim of the present longitudinal study is to assess and compare the efficacy of toothbrush and miswak as potential tools to harvest exfoliated cells for gender identification. Forty healthy volunteers were recruited and asked to clean their teeth using new toothbrush and fresh miswak each day for 4 days. Toothbrush and miswak used by the participants were subjected to DNA analysis immediately, 1st, 2nd and 6th month. The absorbance of DNA samples were quantified and gender identification was done by amplification of sex determining gene-Sex determining region Y gene (SRY) and ALT1 genes using real-time PCR. The number of correct and positive identification for samples at various time points were tabulated and subjected to statistical analysis. Post hoc power analysis showed that the study had a power of 93%. Correct and positive gender identification was observed for the samples (100%) obtained using miswak, for tooth brush it reduced to 95%, 80%, and 35% at the end of 1st, 2nd, and 6th month. The differences seen at the end of 2nd month and 6th month were statistically significant. Miswak is a better tool to harvest exfoliated cells for gender identification when compared to a toothbrush. Hence, miswak can serve as a potential tool in forensic medicine for DNA extraction and subsequently victim identification.

Active MMP-8 point-of-care (PoC)/chairside enzyme-test as an adjunctive tool for early and real-time diagnosis of peri-implantitis.

OBJECTIVE: The aim of this study was to investigate the utility of the active matrix metalloproteinase (aMMP-8)-point-of-care (PoC) test as a quantitative real-time chair-side diagnostic tool for peri-implant diagnosis, as well as assess the potentially developing and ongoing risk relative to the traditional clinical methods. BACKGROUND: Current peri-implant and periodontal disease diagnoses rely on clinical and radiological examinations. This case-control study investigated the applicability of aMMP-8-PoC immunotest for quantitative real-time diagnosis and monitoring of dental implants in health and disease. METHODS: Sixty-eight patients visiting a specialist clinic for maintenance following dental implant placement underwent assessment of their peri-implant health. aMMP-8-PoC peri-implant sulcular fluid (PISF) lateral-flow immunotests were performed using ImplantSafe® technology quantitated by ORALyzer®. In addition, the PISF samples were analyzed for total MMP-8, calprotectin, and interleukin (IL)-6 by enzyme-linked immunosorbent assays (ELISA), aMMP-8 by western immunoblot, and MMP-2 and MMP-9 by gelatin zymography. RESULTS: The aMMP-8-PoC test promptly recorded and reflected peri-implant disease, differentiating it clearly from health. X-ray findings (bone loss > 2 mm), peri-implant pocket depth ≥ 3 mm, and bleeding on probing were significantly more prevalent among implants positive for the aMMP-8-PoC test. aMMP-8/ORALyzer analysis was more precise in recording disease than total MMP-8, calprotectin, IL-6, MMP-2, and MMP-9. CONCLUSIONS: The aMMP-8-PoC test can be conveniently implemented to alert for and detect active collagenolysis affecting peri-implant tissues, both in the early and advanced stages of the disease. Active and fragmented MMP-8 exhibits a strong and significant association with peri-implantitis as compared to total MMP-8 and other biomarkers and can be utilized as the POC/chairside biomarker of choice in the new classification of peri-implantitis.

Evaluation of active matrix metalloproteinase-8 (aMMP-8) chair-side test as a diagnostic biomarker in the staging of periodontal diseases.

OBJECTIVE: There is a need for a reliable complementary diagnostic tool that ideally helps to screen, differentiate sites, activities of and predict future periodontal tissue destruction. The purpose of this cross-sectional study was to investigate the screening and prevention potential of the chair-side/point-of-care (PoC) diagnostic test of salivary active matrix metalloproteinase-8 (aMMP-8) levels at different stages of periodontal disease and periodontal health. MATERIAL & METHODS: 80 individuals were included in this study; 18 with periodontitis stage 3 (P-Stage III), 19 with periodontitis stage-4 (P-Stage IV), 21 with gingivitis, and 22 with clinically healthy periodontium (H). The aMMP-8 levels in GCF and saliva were analyzed by chairside point-of-care aMMP-8 lateral flow immunotest and also by a time-resolved immunofluorescence assay (IFMA). RESULTS: The sensitivity of the chair-side/PoC test was 83.9 % while specificity was 79.2 %. The aMMP-8 IFMA levels in GCF were significantly higher in P-Stage IV group than P-Stage III, gingivitis and healthy groups (p = 0.01, p = 0.001, p = 0.00, respectively). Moreover, P-Stage III and gingivitis groups had significantly higher aMMP-8 IFMA levels than the healthy group (p < 0.05). CONCLUSION: The aMMP-8 chair-side test showed promising results in its ability to recognize and predict the inflammatory status even at the very initial/early stages. aMMP-8 chair-side test could be a valuable adjunctive diagnostic and preventive tool to conventional clinical methods in detecting periodontal disease.

Active MMP-8 (aMMP-8) as a Grading and Staging Biomarker in the Periodontitis Classification.

The aim of this study was to investigate the utility of incorporating active matrix metalloproteinase-8 (aMMP-8) as a biomarker into the new periodontitis classification system (stage/grade) presented in 2018. This study included 150 Greek adults aged 25-78, of whom 74 were men and 76 women. Participants were tested with an aMMP-8 point-of-care mouthrinse test, after which a full-mouth clinical examination was performed to assess their periodontal and oral health. The aMMP-8 levels in mouthrinse were significantly lower among healthy patients compared with patients in more severe periodontitis stages and grades (Kruskal-Wallis test and Dunn-Bonferroni test for pairwise post-hoc comparisons; p < 0.01 and p < 0.05, respectively). Furthermore, aMMP-8 levels were less correlated with plaque levels than bleeding on probing (BOP) (Spearman's rho = 0.269, p < 0.001; Spearman's rho = 0.586, p < 0.001); respectively). Thus, aMMP-8 was more robust to the confounding effects of oral hygiene than traditional periodontal parameter bleeding on probing. The aMMP-8 point-of-care mouthrinse test can be utilized as an adjunctive and preventive diagnostic tool to identify periodontal disease, classified by stage and grade, and ongoing periodontal breakdown chairside in clinical practice in only 5 min. Overall, integrating aMMP-8 into the new periodontitis classification system seems beneficial.

Label-Free Quantitative Proteomics versus Antibody-Based Assays to Measure Neutrophil-Derived Enzymes in Saliva.

PURPOSE: This study aims to validate label-free quantitative proteomics (LFQ) against antibody-based methods for quantifying established periodontal disease biomarkers in saliva. EXPERIMENTAL DESIGN: In an experimental gingivitis model, healthy volunteers (n = 10) provide saliva at baseline (d0), during the induction (d7, d14, d21) and resolution (d35) of gingival inflammation (total n = 50). Biomarker levels are analyzed by LFQ and time-resolved immunofluorometric assay (IFMA) or enzyme-linked immunosorbent assay (ELISA). Molecular matrix metalloproteinase (MMP)-8 forms are assessed by Western blot (WB) analysis. RESULTS: LFQ detects significantly (p < 0.05) elevated MMP-8 (d21vsd7, d35vsd7) and tissue inhibitor of matrix metalloproteinases (TIMP)-1 (d35vsd7). Latent MMP-8 (70-80 kDa) is present (d0-d35), but not active MMP-8 (50-60 kDa). LFQ and immunoassay data significantly correlate for MMP-8 (r = 0.36), myeloperoxidase (r = 0.39), polymorphonuclear leukocyte elastase (r = 0.33), and TIMP-1 (r = -0.24). CONCLUSION AND CLINICAL RELEVANCE: LFQ can quantify enzyme levels in saliva, however lacks the ability to measure enzymatic activity. WB analysis reveals that MMP-8 may not be activated during induction of gingival inflammation. Significant but weak correlations between IFMA or ELISA and LFQ suggest a limited capacity of available antibodies to reliably quantify salivary biomarkers for periodontal diseases. Novel "anti-peptide" antibodies designed by newer targeted mass spectrometry-based approaches can help to overcome these drawbacks.

Adjunctive Effects of a Sub-Antimicrobial Dose of Doxycycline on Clinical Parameters and Potential Biomarkers of Periodontal Tissue Catabolism.

OBJECTIVES: The aim of the present randomized, double-blind, placebo-controlled, parallel-arm study was to examine the effectiveness of a sub-antimicrobial dose of doxycycline (SDD) in combination with nonsurgical periodontal therapy, compared to nonsurgical periodontal therapy alone, on potential gingival crevicular fluid (GCF) biomarkers of periodontal tissue catabolism related to the clinical outcomes over a 12-month period. Materials and Methods: GCF was collected and clinical parameters were recorded from 30 periodontitis patients randomized either to an SDD or placebo group. The SDD group received SDD (20 mg) b.i.d for 3 months plus scaling and root planing (SRP), while the placebo group was given placebo capsules b.i.d for 3 months plus SRP. The patients were evaluated every 3 months during the 12-month study period. At each visit, clinical parameters and GCF sampling were repeated. Matrix metalloproteinase (MMP)-8, MMP-9, MMP-13, myeloperoxidase (MPO), osteoprotegerin (OPG), and tartrate-resistant acid phosphatase-5 (TRAP-5) were determined by IFMA and ELISA. Results: Significant improvements were observed in all clinical parameters in both groups over 12 months (p < 0.0125) while the SDD group showed significantly better reduction in gingival index (GI) and pocket depth and a gain in clinical attachment compared to the placebo group (p < 0.05). GCF MMP-8 and OPG levels significantly reduced in the SDD group compared to baseline (p < 0.05). GCF MMP-9 significantly decreased in both groups compared to baseline (p < 0.05). GCF MPO significantly decreased at 3 and 9 months in the SDD group, while it significantly decreased at 6 months in the placebo group (p < 0.05). TRAP and MMP-13 could be detected in none of the samples. Conclusions: The present results indicate that three months of adjunctive usage of SDD to nonsurgical periodontal therapy compared to nonsurgical periodontal therapy alone in periodontitis patients results in further improvement of clinical periodontal parameters and GCF markers of periodontal tissue breakdown over a 12-month period. Beneficial effects of adjunctive SDD therapy is likely to be related to the reduced levels of two major periodontitis-associated MMPs, MMP-8 and -9, and their potential oxidative activator MPO.

Matrix Metalloproteinase-8 as an Inflammatory and Prevention Biomarker in Periodontal and Peri-Implant Diseases.

Levels of and especially the degree of activation of matrix metalloproteinase (MMP-8) in oral fluids (i.e., saliva, mouth rinse, gingival crevicular fluid (GCF) and peri-implantitis sulcular fluid (PISF)) increase to pathologically elevated levels in the periodontal and peri-implant diseases. This study aimed at collecting and collating data from previously published studies and determining whether active MMP-8 (aMMP-8) could serve as a biomarker for the diagnosis and prevention of periodontal and peri-implant diseases. The literature search identified a total of 284 articles. Out of 284 articles, 61 articles were found to be relevant. Data obtained from the selected studies were combined, and it indicated that aMMP-8 in oral fluids exerts the strong potential to serve as a useful adjunctive diagnostic and preventive biotechnological tool in periodontal and peri-implant diseases. aMMP-8 can be used alone or in combination with other proinflammatory and/or microbiological biomarkers.

The Ability of Quantitative, Specific, and Sensitive Point-of-Care/Chair-Side Oral Fluid Immunotests for aMMP-8 to Detect Periodontal and Peri-Implant Diseases.

The analysis of the disease-specific oral and systemic biomarkers in saliva and oral fluids (i.e., mouth rinse, gingival crevicular fluid (GCF), and peri-implantitis fluid (PISF)) is demanding. Several hosts and microbial factors may influence their expression, release, and levels. The type of saliva/oral fluids utilized for the diagnostics affects the analysis. High sensitivity and specificities together with sophisticated methods and techniques are essential for valuable outcome. We describe here recently developed practical, convenient, inexpensive, noninvasive, and quantitative mouth rinse and PISF/GCF/chair-side/point-of-care (PoC) lateral-flow aMMP-8 immunoassays (PerioSafe and ImplantSafe/ORALyser) to detect, predict, and monitor successfully the course, treatment, and prevention of periodontitis and peri-implantitis, respectively. The tests have been independently and successfully validated to differentiate periodontal and peri-implant health and disease in Finland, Germany, Netherland, Sweden, Turkey, Nigeria, Malawi, and USA. The clinical use of salivary/oral fluid biomarkers to identify oral and systemic conditions requires additional studies utilizing these noninvasive screening, diagnostic, and preventive aMMP-8 PoC/chair-side technologies.