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Objective: One of the most important families of proteases associated with periodontal disease is the family of the matrix metalloproteinases (MMPs). Their activity is regulated by tissue inhibitors of metalloproteinases (TIMPs), and an imbalance between MMP activity and regulation by TIMPs has been associated with the progression of periodontal disease. This strong interaction between TIMPs and MMPs might be an indication that TIMPs can be used as a biomarker to monitor periodontal disease progression in oral fluids. In particular, TIMP-1 is a frequently studied biomarker for periodontal diseases. Therefore, the aim of this systematic review was to evaluate the scientific literature regarding TIMP-1 concentrations in oral fluids of patients suffering from periodontitis or gingivitis in comparison to healthy individuals., Material and Methods: PubMed/ MedLine and Web of Science databases were searched electronically. Studies that met the inclusion criteria were systematically evaluated and assessed for eligibility and risk of bias. Meta-analysis was performed through the random effects model to assess the association between periodontitis/gingivitis and TIMP-1 concentration in stimulated saliva, unstimulated saliva, and gingival crevicular fluid (GCF)., Results: The search strategy provided a total of 322 studies of which 10 studies met all inclusion criteria. Two studies investigated TIMP-1 concentrations in GCF, three studies in unstimulated saliva, and five studies investigated TIMP-1 concentrations in stimulated saliva. Three studies revealed that TIMP-1 levels in oral fluids were significantly decreased in periodontal disease. Meta-analysis revealed that there is no statistically significant difference between TIMP-1 concentration in oral fluids of periodontitis/gingivitis patients in comparison to healthy individuals., Conclusions: This systematic review with meta-analysis shows that periodontal diseases are not associated with a statistically significant change in TIMP-1 concentration in oral fluids., (© 2021 The Authors. Journal of Periodontal Research published by John Wiley & Sons Ltd.)

Aim: To explore the feasibility of screening for periodontitis by measuring biomarkers, namely total proteolytic activity (TPA), matrix metalloproteinase (MMP)-8, chitinase, lysozyme or their combination, in saliva, oral rinse and gingival crevicular fluid (GCF)., Material and Methods: Subjects were recruited among healthy/gingivitis individuals and untreated periodontitis patients in Academic Centre for Dentistry Amsterdam (ACTA). All participants donated samples of unstimulated whole saliva, oral rinse and GCF. The protein concentrations and MMP-8 levels were determined by ELISA. Enzymatic activities were measured using appropriate fluorogenic substrates., Results: In oral rinse samples, periodontitis patients (n = 19) exhibited significantly higher concentrations of MMP-8 and TPA than controls (n = 20). MMP-8 in combination with chitinase explained 88% of the variance and assigned a subject to control or periodontitis group, with best accuracy (87.2%) in oral rinse., Conclusions: The combination of MMP-8 and chitinase in the current oral rinse procedure has the potential to discriminate periodontitis from periodontal health/gingivitis., (© 2021 The Authors. Journal of Clinical Periodontology published by John Wiley & Sons Ltd.)

The black pigmentation-related genes in Porphyromonas gingivalis are primarily involved in regulating gingipain functions. In this study, we identified a pigmentation-related gene, designated as pgn_0361. To characterize the role of pgn_0361 in regulating P. gingivalis-mediated epithelial cell detachment and inhibition of wound closure, PgΔ0361, an isogenic pgn_0361-defective mutant strain, and PgΔ0361C, a complementation strain, were constructed using P. gingivalis ATCC 33277. The gingipain and hemagglutination activities, as well as biofilm formation, were examined in all three strains. The effect of P. gingivalis strains on epithelial cell detachment was investigated using the HO-1-N-1 and Ca9-22 epithelial cell lines. The inhibition of wound closure by heat-killed P. gingivalis cells and culture supernatant was analyzed using an in vitro wound closure assay. Compared to the wild-type strain, the PgΔ0361 strain did not exhibit gingipain or hemagglutination activity but exhibited enhanced biofilm formation. Additionally, the PgΔ0361 strain exhibited attenuated ability to detach the epithelial cells and to inhibit wound closure in vitro. Contrastingly, the culture supernatant of PgΔ0361 exhibited high gingipain activity and strong inhibition of wound closure. The characteristics of PgΔ0361C and wild-type strains were comparable. In conclusion, the pgn_0361 gene is involved in regulating gingipains. The PGN_0361-defective strain exhibited reduced virulence in terms of epithelial cell detachment and inhibition of wound closure. The culture supernatant of the mutant strain highly inhibited wound closure, which may be due to high gingipain activity., (© 2020 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

Aim: To profile gingival tissue levels of human beta-defensin (hBD)-2 and hBD-3 in relation to gingival inflammation, Th17-related cytokine concentrations, Porphyromonas gingivalis counts, and gingipain and total protease activities., Materials and Methods: Gingival tissue and subgingival plaque samples were collected from 21 periodontitis patients including 48 periodontal pocket sites with marginal, mild, or moderate to severe inflammation. hBD levels were determined by immunodetection, P. gingivalis counts with real-time polymerase chain reaction, protease activities with fluorogenic substrates, and cytokine concentrations with Luminex technique. Data were statistically analysed using Kruskal-Wallis and Mann-Whitney U tests and Spearman correlation coefficients., Results: Subgingival plaque counts of P. gingivalis (p = .001) and gingipain activity (p < .001), as well as interleukin (IL)-1β (p = .012), IL-10 (p = .024), IL-17A (p = .002), IL-17F (p = .006), and IL-23 (p = .036) concentrations were elevated in severely inflamed sites, whereas no change was observed in hBD-2 and hBD-3 levels. Negative correlations were found between protease activity and hBD-2 (p = .033) and hBD-3(p = .003) levels., Conclusions: Shift in gingival inflammation from marginal to mild stage is related to elevations in subgingival plaque P. gingivalis counts and gingipain activity, but not to tissue hBD levels. Negative correlations between hBDs and total protease activity suggest the degradation of these antimicrobial peptides in progressed inflammation., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

A selection of commercially available products containing stannous fluoride (SnF2 )/sodium fluoride (NaF), SnF2 /amorphous calcium phosphate (ACP), SnF2 /NaF/ACP, tin (Sn)/fluorine (F)/chitosan were compared with phytosphingosine (PHS) with respect to their anti-erosive properties in vitro. One-hundred and twenty bovine enamel specimens were immersed in the respective product slurries for 2 min, twice daily. The formulations were diluted with either remineralization solution or artificial saliva. After each treatment, an erosive challenge was performed for 10 min, twice daily, using citric acid, pH 3.4. The specimens were stored in remineralization solution or artificial saliva until the next treatment-erosion challenge. After 10 d, tissue loss was determined using profilometry. Enamel softening was determined through surface microhardness measurements. Tissue-loss values (measured in μm and expressed as mean ± SD) for PHS, SnF2 /NaF, SnF2 /ACP, SnF2 /ACP/NaF, and Sn/F/chitosan treatment groups and for the negative-control group, were, respectively, 35.6 ± 2.8, 15.8 ± 1.8, 22.1 ± 2.0, 22.9 ± 1.8, 16.2 ± 1.2, and 51.2 ± 4.4 in the presence of remineralization solution and 31.7 ± 3.3, 15.6 ± 2.9, 16.5 ± 2.7, 16.8 ± 2.1, 13.1 ± 3.0, and 50.7 ± 2.8 in the presence of artificial saliva. There were no significant differences in surface microhardness measurements between the treatment groups. In conclusion, PHS resulted in a significant reduction of tissue loss compared with the negative control, but in comparison, the toothpastes containing Sn(2+) and F(-) ions were significantly more effective compared with PHS., (© 2016 Eur J Oral Sci.)

This study evaluated the anti-biofilm activity of sphingosine, phytosphingosine (PHS), and sphinganine for: (i) anti-adherence activity on hydroxyapatite (HA) surfaces; and (ii) bactericidal activity on different Streptococcus mutans phenotypes (i.e. planktonic cells and cells from a disrupted biofilm). For this, HA discs treated with sphingolipids were incubated with S. mutans and the number of adherent cells was evaluated by both culture and confocal microscopy. Sphinganine strongly inhibited bacterial adherence by 1000-fold compared with an untreated surface. Phytosphingosine and sphingosine inhibited bacterial adherence by eight- and five-fold, respectively, compared with an untreated surface. On saliva-coated HA, sphinganine and PHS inhibited bacterial adherence by 10-fold. Bactericidal activity of sphingolipids was evaluated by culture. For biofilms, the strongest bactericidal activity was exhibited by sphingosine compared with PHS and sphinganine. At a concentration of 12.5 μg ml(-1) , PHS and sphingosine were profoundly effective against planktonic and disrupted biofilms; and sphinganine reduced the number of cells in planktonic form by 100-fold and those derived from a disrupted biofilm by 1000-fold. Atomic force microscopy studies suggested that mechanical stability does not appear to be a factor relevant for anti-fouling activity. The results suggest that sphingolipids may be used to control oral biofilms, especially those loaded with S. mutans., (© 2015 Eur J Oral Sci.)

The salivary agglutinin glycoprotein (SAG) is present in saliva but is also part of the salivary pellicle, playing a seemingly paradoxical role with regard to bacterial homeostasis. On the one hand, SAG aggregates bacteria in solution, thereby preventing bacterial colonization. On the other hand, when bound to the tooth surface, SAG facilitates bacterial colonization and microbial growth. The protein part of SAG is predominantly composed of conserved scavenger receptor cysteine-rich (SRCR) domains. Previously it was found that bacterial binding and aggregation is mediated via a single peptide loop, designated SRCRP2 (P2), within the SRCR domains of SAG. The current data suggest that the SRCR domains also harbour a hydroxyapatite (HA)-binding moiety, SRCRP3 (P3). The observation that P2 and P3 individually play unique roles in the function of SAGs contributes to our understanding of the dual role of SAGs in bacterial binding. Inspired by the bacterial-modulating capacity of SAGs, we created a P3-polyethylene glycol (PEG) conjugate. It was found that a P3 coating resulted in an increased antifouling activity of 20% compared with the uncoated surface in vitro. An additional PEG moiety resulted in an antifouling activity of up to 40% and 30% for Streptococcus mutans and Staphylococcus epidermidis, respectively., (© 2013 Academic Centre for Dentistry Amsterdam, University of Amsterdam and VU University Amsterdam.)

Background and Objective: The aim of the study was to compare the detection of Porphyromonas gingivalis using a fluorescence resonance energy transfer (FRET) technology with commonly used diagnostic methods in salivary and subgingival plaque samples from subjects with dental implants. P. gingivalis was considered as a marker for a pathogenic microbiota., Material and Methods: Ninety-seven adult subjects were recruited, including periodontally healthy controls with no dental implants, implant controls with no peri-implant disease and patients with peri-implant disease. Saliva and subgingival/submucosal plaque samples were collected from all subjects and were analyzed using culture, real-time PCR and FRET technology employing P. gingivalis-specific substrates., Results: It was found that the P. gingivalis-specific substrates were highly suitable for detecting the presence of P. gingivalis in saliva and in subgingival plaque samples, showing comparable specificity to culture and real-time PCR., Conclusion: We applied the FRET technology to detect P. gingivalis in implant patients with or without an implant condition and in controls without implants. The technique seems suitable for detection of P. gingivalis in both plaque and saliva samples. However, with all three techniques, P. gingivalis was not very specific for peri-implantitis cases. Future work includes fine-tuning the FRET technology and also includes the development of a chair-side application., (© 2012 John Wiley & Sons A/S.)

As part of a research program to develop novel methods for diagnosis of sulfur mustard exposure in the human skin the suitability of phage display was explored. Phage display is a relative new method that enables researchers to quickly evaluate a huge range of potentially useful antibodies, thereby bypassing the more costly and time-consuming hybridoma technique. The Tomlinson I and J phage libraries were used to select phage antibodies exhibiting affinity for sulfur mustard adducts on keratins, isolated from human callus. Two kinds of phage antibodies were obtained: antibodies recognizing keratin and antibodies recognizing keratin which was exposed to sulfur mustard. These phage antibodies retained activity after repeated culturing and culturing in larger volumes. For the first time antibody phage display was successfully applied for immunodiagnostics of a chemical warfare agent.

Drosocin is a 19-mer, cationic antimicrobial peptide from Drosophila melanogaster. The aim of the study was to examine the antibacterial spectrum of unglycosylated drosocin analogues. Furthermore, the amino acid sequence of DnaK, drosocin's intracellular target, from susceptible species was aligned and studied for sequence homology. From this a panel of 31 bacterial strains, including Salmonella strains with truncated lipopolysaccharide structures, was tested for susceptibility towards three drosocin analogues. Available bacterial DnaK amino acid sequences were retrieved from the ExPASy proteomics server of the Swiss Institute of Bioinformatics studied for sequence homology. Seventeen of the 31 strains tested were susceptible for the drosocin analogues. Minimal inhibitory concentration values against mainly Gram-negative bacteria ranged from 3.1 to 100 microm. With the exception of Micrococcus luteus and Xanthomonas campestris all drosocin analogue-susceptible strains were Enterobacteriaceae showing a high DnaK amino acid sequence homology.