Quorum Sensing in Oral Biofilms: Influence on Host Cells.

Quorum sensing molecules (QSMs) in the oral cavity regulate biofilm formation, the acquisition of iron, stress responses, and the expression of virulence factors. However, knowledge of the direct QSM-host interactions in the oral environment is limited, although their understanding could provide greater insight into the cross-kingdom communication occurring during oral disease development. This review aims to explore the literature on oral QSM-host interactions and to highlight areas of advancement in this field. The studies included in this review encompass an array of cell types and oral QSMs, with particular emphasis on immune cells and their relationship to periodontal diseases. It can be inferred from the current literature that QSMs are utilised by host cells to detect bacterial presence and, in the majority of cases, elicit an immune response towards the environmental QSMs. This may provide a base to target QSMs as a novel treatment of oral diseases. However, N-acyl homoserine lactone (AHL) detection methods remain an area for development, through which a greater understanding of the influence of oral QSMs on host cells could be achieved.

The role of the host-Neutrophil biology.

Neutrophilic polymorphonuclear leukocytes (neutrophils) are myeloid cells packed with lysosomal granules (hence also called granulocytes) that contain a formidable antimicrobial arsenal. They are terminally differentiated cells that play a critical role in acute and chronic inflammation, as well as in the resolution of inflammation and wound healing. Neutrophils express a dense array of surface receptors for multiple ligands, ranging from integrins to support their egress from bone marrow into the circulation and from the circulation into tissues, to cytokine/chemokine receptors that drive their navigation to the site of infection or tissue damage and also prime them for a second stimulus, to pattern recognition receptors and immunoglobulin receptors to facilitate the destruction and removal of infective agents or debridement of damaged tissues. When afferent neutrophil signals are proportionate and coordinated they will phagocytose opsonized and unopsonized bacteria, activating the nicotinamide adenine dinucleotide phosphate oxidase (respiratory burst) to generate reactive oxygen species, which augment the proteolytic destruction of microbes secured within the phagosome. A highly orchestrated process of apoptosis follows, forming membrane-bound substructures that are removed by macrophages. Neutrophils are capable of various other forms of programmed cell death, such as NETosis and pyroptotic cell death, as well as nonprogrammed cell death by necrosis. In recent years, research has revealed that neutrophils are capable of far more subtle cell-cell interactions than previously thought possible. This includes synthesis of various inflammatory mediators and also myeloid cell training within bone marrow, where epigenetic and metabolic signals associated with returning neutrophils that undergo reverse egress from tissues into the vasculature and back to bone marrow program a hyperreactive subset of neutrophils during myelopoiesis that are capable of hypersensitive reactions to microbial aggressors. These characteristics are evident in various neutrophil subsets/subpopulations, creating broad heterogeneity in the behavior and biological repertoire of these seemingly schizophrenic immune cells. Moreover, neutrophils are critical effector cells of adaptive and innate immunity, binding to opsonized bacteria and destroying them by extracellular and intracellular methods. The former creates substantial collateral host tissue damage, as they are less specific than T-cytotoxic cell-killing mechanisms, and in conditions such as peri-implantitis, where plasma cells and neutrophils dominate the immune infiltrate, bone and tissue destruction are rapid and appear relentless. Finally, the role of neutrophils as conduits for periodontal-systemic disease connections and for oxidative damage to act as a causal link between the two has only recently been realized. In this chapter, we attempt to expand on these issues, emphasizing the contributions of European scientists throughout a detailed appraisal of the benefits and side effects of neutrophilic inflammation and immune function.

Characterization, Quantification, and Visualization of Neutrophil Extracellular Traps.

Following the discovery of neutrophil extracellular traps (NETs) in 2004 by Brinkmann and colleagues, there has been extensive research into the role of NETs in a number of inflammatory diseases, including periodontitis. This chapter describes the current methods for the isolation of peripheral blood neutrophils as well as of oral neutrophils for subsequent NET experiments, including approaches to quantify and visualize NET production, the ability of NETs to entrap and kill bacteria, and the removal of NETs by nuclease-containing plasma.

Multiple external root resorptions in a patient with Stage IV, Grade C periodontitis and autoimmune diseases: A case report.

BACKGROUND: Pathological root resorption affects permanent teeth and is usually triggered by infectious-inflammatory stimuli and/or dental trauma. Periodontitis and traumatic occlusion have been reported as possible causative factors of root resorptions, whilst the impact of systemic diseases is less well understood. This case highlights the need for consideration of multiple risk factors, especially when presenting in combination. METHODS AND RESULTS: A 62-year-old South Asian female presented with unstable Stage IV Grade C periodontitis, poor oral hygiene and multiple autoimmune conditions including oral lichen planus. Clinical and radiographic examination revealed multiple advanced apical and external root resorptions of the patient's molars associated with periapical bone loss, despite of a minimally restored dental status. CONCLUSION: A likely etiology of this patient's multiple root resorptions is the presence of unstable periodontitis with periodontal-endodontic lesions, exacerbated by a dysbalanced immune response to infectious agents. Appropriate monitoring and managing of such patients can prevent or limit the pathological process of inflammatory root resorption. KEY POINTS: Why is this case new information? This is the first report documenting advanced multiple external inflammatory root resorptions in a periodontitis patient with oral and systemic co-morbidities. What are the keys to successful management of this case? Early diagnosis, prevention and intervention to limit periodontal inflammation, endodontic infection and occlusal trauma. What are the primary limitations to success in this case? Late diagnosis of multiple root resorptions, palliative periodontal treatment due to poor oral hygiene compliance, and poorly controlled systemic inflammation favoring the persistence of a dysregulated immune response to the oral microbiota.

Fusobacterium nucleatum Subspecies Differ in Biofilm Forming Ability in vitro.

Development of dysbiosis in complex multispecies bacterial biofilms forming on teeth, known as dental plaque, is one of the factors causing periodontitis. Fusobacterium nucleatum (F. nucleatum) is recognised as a key microorganism in subgingival dental plaque, and is linked to periodontitis as well as colorectal cancer and systemic diseases. Five subspecies of F. nucleatum have been identified: animalis, fusiforme, nucleatum, polymorphum, and vincentii. Differential integration of subspecies into multispecies biofilm models has been reported, however, biofilm forming ability of individual F. nucleatum subspecies is largely unknown. The aim of this study was to determine the single-subspecies biofilm forming abilities of F. nucleatum ATCC type strains. Static single subspecies F. nucleatum biofilms were grown anaerobically for 3 days on untreated or surface-modified (sandblasting, artificial saliva, fibronectin, gelatin, or poly-L-lysine coating) plastic and glass coverslips. Biofilm mass was quantified using crystal violet (CV) staining. Biofilm architecture and thickness were analysed by scanning electron microscopy and confocal laser scanning microscopy. Bioinformatic analysis was performed to identify orthologues of known adhesion proteins in F. nucleatum subspecies. Surface type and treatment significantly influenced single-subspecies biofilm formation. Biofilm formation was overall highest on poly-L-lysine coated surfaces and sandblasted glass surfaces. Biofilm thickness and stability, as well as architecture, varied amongst the subspecies. Interestingly, F. nucleatum ssp. polymorphum did not form a detectable, continuous layer of biofilm on any of the tested substrates. Consistent with limited biofilm forming ability in vitro, F. nucleatum ssp. polymorphum showed the least conservation of the adhesion proteins CmpA and Fap2 in silico. Here, we show that biofilm formation by F. nucleatum in vitro is subspecies- and substrate-specific. Additionally, F. nucleatum ssp. polymorphum does not appear to form stable single-subspecies continuous layers of biofilm in vitro. Understanding the differences in F. nucleatum single-subspecies biofilm formation may shed light on multi-species biofilm formation mechanisms and may reveal new virulence factors as novel therapeutic targets for prevention and treatment of F. nucleatum-mediated infections and diseases.

Aggregatibacter actinomycetemcomitans Leukotoxin Activates the NLRP3 Inflammasome and Cell-to-Cell Communication.

Carriers of highly leukotoxic genotypes of Aggregatibacter actinomycetemcomitans are at high risk for rapid degradation of tooth-supporting tissues. The leukotoxin (LtxA) expressed by this bacterium induces a rapid pro-inflammatory response in leukocytes that results in cell death. The aim of the present study was to increase the understanding of LtxA-induced leukocyte activation mechanisms and of possible associated osteoclast differentiation. The effect of LtxA on activation of the inflammasome complex was studied in THP-1 wild type and in NLRP3- and ASC knockout cells. Cell-to-cell communication was assessed by fluorescent parachute assays, and THP-1 differentiation into osteoclast-like cells was investigated microscopically. The results showed that LtxA induced inflammatory cell death, which involved activation of the NLRP3 inflammasome and gap junction cell-to-cell communication. THP-1 cells treated with lipopolysaccharide (LPS) and LtxA together differentiated into an osteoclast-like phenotype. Here, LPS prevented LtxA-mediated cell death but failed to induce osteoclast differentiation on its own. However, pit formation was not significantly enhanced by LtxA. We conclude that A. actinomycetemcomitans leukotoxicity mediates activation of the NLRP3 inflammasome and cell-to-cell communication in the induced pro-inflammatory cell death. In addition, LtxA stimulated differentiation towards osteoclasts-like cells in LPS-treated THP-1 cells.

Periodontal health, neutrophil activity and cardiovascular health in captive chimpanzees.

OBJECTIVE: To investigate the association of dental and cardiac disease in a cohort of captive chimpanzees DESIGN: 12 captive chimpanzees underwent periodontal and cardiac examinations under anaesthesia during a relocation to a new enclosure. Blood samples were taken for analysis of circulating markers of cardiac health, nutritional status and isolation of neutrophils for functional assays. They were then observed for three years for signs of heart disease. RESULTS: Although the chimpanzees displayed large quantities of supragingival plaque, they had low bleeding scores. Peripheral blood neutrophils responded to innate and adaptive immune stimuli. In the follow up period two animals died and post mortem confirmed heart disease. Levels of NT-proBNP were found to be high in chimpanzees that died from heart disease. CONCLUSIONS: Whilst there appeared to be a correlation between probing depth and age, there appeared to be no correlation between dental data and heart data in this cohort.

Regenerative surgical treatment of furcation defects: A systematic review and Bayesian network meta-analysis of randomized clinical trials.

AIMS: To investigate the clinical performance of regenerative periodontal surgery in the treatment of furcation defects versus open flap debridement (OFD) and to compare different regenerative modalities. MATERIAL AND METHODS: A systematic search was conducted to identify RCTs evaluating regenerative surgical treatment of furcations with a minimum of 12-month follow-up. Three authors independently reviewed, selected and extracted data from the search conducted and assessed risk of bias. Primary outcomes were tooth loss, furcation improvement (closure/conversion) (FImp), gain of horizontal bone level (HBL) and attachment level (HCAL). Secondary outcomes were gain in vertical attachment level (VCAL), probing pocket depth (PPD) reduction, PROMs and adverse events. Data were summarized into Bayesian standard and network meta-analysis in order to estimate direct and indirect treatment effects and to establish a ranking of treatments. RESULTS: The search identified 19 articles, reporting on 20 RCTs (19 on class II, 1 on class III furcations) with a total of 575 patients/787 defects. Tooth loss was not reported. Furcation closure ranged between 0% and 60% (10 trials), and class I conversion from 29% to 100% (six trials). Regenerative techniques were superior to OFD for FImp (OR = 20.9; 90% CrI = 5.81, 69.41), HCAL gain (1.6 mm), VCAL gain (1.3 mm) and PPD reduction (1.3 mm). Bone replacement grafts (BRG) resulted in the highest probability (Pr = 61%) of being the best treatment for HBL gain. Non-resorbable membranes + BRG ranked as the best treatment for VCAL gain (Pr = 75%) and PPD reduction (Pr = 56%). CONCLUSIONS: Regenerative surgery of class II furcations is superior to OFD. FImp (furcation closure or class I conversion) can be expected for the majority of defects. Treatment modalities involving BRG are associated with higher performance.

Systemic disease or periodontal disease? Distinguishing causes of gingival inflammation: a guide for dental practitioners. Part 2: cancer related, infective, and other causes of gingival pathology.

Periodontitis and gingivitis are highly prevalent inflammatory diseases of the oral cavity, and typically are characterised by the presence of dental plaque. However, other causes of oral inflammation exist, which can resemble plaque-induced gingivitis and periodontitis, and may thus first be seen by a dental practitioner. This paper aims to provide dentists with an understanding of the manifestations of systemic diseases to the periodontium and highlights anamnestic and clinical clues important for distinguishing between plaque-induced and non plaque-induced lesions. In the first part of this series immune-mediated and hereditary conditions as causes of gingival lesions were discussed; this second part highlights cancer-related gingival lesions as well as those caused by specific pathogens, medication or malnutrition. A clear clinical, epidemiological and visual overview of the different conditions is provided. Early diagnosis of non plaque-related causes of gingival lesions can be vital for affected patients. Therefore, dental practitioners should be aware of the various manifestations of systemic diseases to the periodontium.

Systemic disease or periodontal disease? Distinguishing causes of gingival inflammation: a guide for dental practitioners. Part 1: immune-mediated, autoinflammatory, and hereditary lesions.

Periodontitis and gingivitis remain two of the most common diseases that affect the oral cavity. As they are caused by plaque, effective oral hygiene, elimination of plaque-retentive factors and successful periodontal treatment will result in resolution of gingival and periodontal inflammation. Certain systemic diseases can have a clinical appearance similar to periodontal diseases or exacerbate existing periodontitis/gingivitis and vice versa. This paper aims to provide the dental practitioner with an understanding of the manifestations of systemic diseases to the periodontium and highlights elements in the clinical assessment, which will aid in establishing a correct diagnosis. Additional anamnestic and clinical clues are important for distinguishing between plaque-induced and non-plaque-induced lesions. The first part of this compendium covers immune-mediated and hereditary conditions as causes of gingival lesions, which can resemble those caused by dental plaque. The different conditions are presented concisely and exemplified by clinical photographs. Dental practitioners should be aware of the various manifestations of systemic diseases to the periodontium in order to offer appropriate diagnosis and treatment, which can reduce both patient morbidity and mortality.

Interest in orthodontic tooth alignment in adult patients affected by periodontitis: A questionnaire-based cross-sectional pilot study.

BACKGROUND: Orthodontic treatment can successfully align pathologically migrated teeth and lead to improvement of periodontal stability in patients with periodontitis. Periodontic-orthodontic approaches have gained increasing attention in the past years. Here, we investigated the interest of adults affected by chronic periodontitis in undergoing orthodontic treatment as well as patient-related and tooth-related influence factors. METHODS: Periodontal and orthodontic measurements/indices were taken from 115 adult patients with moderate-to-severe periodontitis. The study participants answered a questionnaire investigating patient demographics, quality of life aspects, and their interest in undergoing orthodontic treatment. Correlations between clinical data, questionnaire responses, and this interest were analyzed by means of an age- and sex-adjusted multiple regression model. RESULTS: Two-thirds of the participants were interested in orthodontic therapy and indicated long-term healthy and esthetically appealing teeth as their main motives. A significant correlation was found between subjectively felt impaired dental esthetics and an interest in orthodontic treatment. However, there were no correlations with the severity of periodontitis, tooth alignment, or patient demographics, including sex. Older patients were significantly more often interested in orthodontic treatment. Most participants had never been provided with information about orthodontic treatment options for adults. CONCLUSIONS: A considerable number of adult patients with periodontitis were interested in orthodontics to improve tooth alignment. However, severity of periodontitis and tooth misalignment or demographic factors may not be indicative thereof. Therefore, dental practitioners need to be aware of patients wishing to align their teeth and to provide them with the relevant information and, if appropriate, enable interdisciplinary treatment planning.

Oral Neutrophils Characterized: Chemotactic, Phagocytic, and Neutrophil Extracellular Trap (NET) Formation Properties.

Maintenance of oral health is in part managed by the immune-surveillance and antimicrobial functions of polymorphonuclear leukocytes (PMNs), which migrate from the circulatory system through the oral mucosal tissues as oral PMNs (oPMNs). In any microorganism-rich ecosystem, such as the oral cavity, PMNs migrate toward various exogenous chemoattractants, phagocytose bacteria, and produce neutrophil extracellular traps (NETs) to immobilize and eliminate pathogens. PMNs obtained from the circulation through venipuncture (hereafter called cPMNs) have been widely studied using various functional assays. We aimed to study the potential of oPMNs in maintaining oral health and therefore compared their chemotactic and antimicrobial functions with cPMNs. To establish chemotactic, phagocytic, and NET forming capacities, oPMNs and cPMNs were isolated from healthy subjects without obvious oral inflammation. Directional chemotaxis toward the chemoattractant fMLP was analyzed using an Insall chamber and video microscopy. fMLP expression was assessed by flow cytometry. Phagocytosis was analyzed by flow cytometry, following PMN incubation with heat-inactivated FITC-labeled micro-organisms. Furthermore, agar plate-based killing assays were performed with Escherichia coli (Ec). NET formation by oPMNs and cPMNs was quantified fluorimetrically using SYTOX™ Green, following stimulation with either PMA or RPMI medium (unstimulated control). In contrast to cPMNs, the chemotactic responses of oPMNs to fMLP did not differ from controls (mean velocity ± SEM of cPMNs: 0.79 ± 0.24; of oPMNs; 0.10 ± 0.07 micrometer/min). The impaired directional movement toward fMLP by oPMNs was explained by significantly lower fMLP receptor expression. Increased adhesion and internalization of various micro-organisms by oPMNs was observed. oPMNs formed 13 times more NETs than stimulated cPMNs, in both unstimulated and stimulated conditions. Compared to cPMNs, oPMNs showed a limited ability for intracellular killing of Ec. In conclusion, oPMNs showed exhausted capacity for efficient chemotaxis toward fMLP which may be the result of migration through the oral tissues into the oral cavity, being a highly "hostile" ecosystem. Overall, oPMNs' behavior is consistent with hyperactivity and frustrated killing. Nevertheless, oPMNs most likely contribute to maintaining a balanced oral ecosystem, as their ability to internalize microbes in conjunction with their abundant NET production remains after entering the oral cavity.

The guardians of the periodontium-sequential and differential expression of antimicrobial peptides during gingival inflammation. Results from in vivo and in vitro studies.

AIM: To evaluate the sequential and differential expression of a variety of antimicrobial peptides (AMPs) during the development of an experimentally induced gingivitis in humans. MATERIAL AND METHODS: In twenty healthy volunteers, gingival inflammation was induced by abstention from oral hygiene at 6 teeth. Bleeding on probing (BOP) and plaque index (PI) were assessed, and gingival biopsies and gingival crevicular fluid (GCF) were collected at 8 different time points (t0-t35). Gingival epithelial cells (GECs) were stimulated with various receptor agonists. In biopsies and GECs, mRNA expression of human beta-defensins (hBD-2, hBD-3), CC-chemokine ligand 20 (CCL20), S100A7/psoriasin (S100A7), and calgranulin A/B (S100A8, S100A9) was evaluated using real-time PCR, and protein profiles were measured by ELISA. Statistical analysis was performed using non-parametric tests. RESULTS: The clinical parameters BOP, PI and GCF increased over time (p < 0.0001). Tissue AMP mRNA expression was elevated, but at different and AMP-specific time points (p < 0.05). Protein analysis revealed a similar expression pattern for hBD-2 and CCL20 in GCF (p < 0.05). In GECs, multiple receptor stimulation was required to induce AMP gene expression (p < 0.0001). CONCLUSIONS: For the first time, this study showed the sequential and differential expression of AMPs during a developing inflammation in vivo providing further evidence for their role as guardians of a healthy periodontium.

Neutrophil Subsets in Periodontal Health and Disease: A Mini Review.

Neutrophils are amongst the most abundant immune cells within the periodontal tissues and oral cavity. As innate immune cells, they are first line defenders at the tooth-mucosa interface, and can perform an array of different functions. With regard to these, it has been observed over many years that neutrophils are highly heterogeneous in their behavior. Therefore, it has been speculated that neutrophils, similarly to other leukocytes, exist in distinct subsets. Several studies have investigated different markers of neutrophils in oral health and disease in recent years in order to define potential cell subsets and their specific tasks. This research was inspired by recent advancements in other fields of medicine in this field. The aim of this review is to give an overview of the current evidence regarding the existence and presence of neutrophil subsets and their possible functions, specifically in the context of periodontitis, gingivitis, and periodontal health.

Consequences of cathepsin C inactivation for membrane exposure of proteinase 3, the target antigen in autoimmune vasculitis.

Membrane-bound proteinase 3 (PR3m) is the main target antigen of anti-neutrophil cytoplasmic autoantibodies (ANCA) in granulomatosis with polyangiitis, a systemic small-vessel vasculitis. Binding of ANCA to PR3m triggers neutrophil activation with the secretion of enzymatically active PR3 and related neutrophil serine proteases, thereby contributing to vascular damage. PR3 and related proteases are activated from pro-forms by the lysosomal cysteine protease cathepsin C (CatC) during neutrophil maturation. We hypothesized that pharmacological inhibition of CatC provides an effective measure to reduce PR3m and therefore has implications as a novel therapeutic approach in granulomatosis with polyangiitis. We first studied neutrophilic PR3 from 24 patients with Papillon-Lefèvre syndrome (PLS), a genetic form of CatC deficiency. PLS neutrophil lysates showed a largely reduced but still detectable (0.5-4%) PR3 activity when compared with healthy control cells. Despite extremely low levels of cellular PR3, the amount of constitutive PR3m expressed on the surface of quiescent neutrophils and the typical bimodal membrane distribution pattern were similar to what was observed in healthy neutrophils. However, following cell activation, there was no significant increase in the total amount of PR3m on PLS neutrophils, whereas the total amount of PR3m on healthy neutrophils was significantly increased. We then explored the effect of pharmacological CatC inhibition on PR3 stability in normal neutrophils using a potent cell-permeable CatC inhibitor and a CD34+ hematopoietic stem cell model. Human CD34+ hematopoietic stem cells were treated with the inhibitor during neutrophil differentiation over 10 days. We observed strong reductions in PR3m, cellular PR3 protein, and proteolytic PR3 activity, whereas neutrophil differentiation was not compromised.

Therapeutic targeting of cathepsin C: from pathophysiology to treatment.

Cathepsin C (CatC) is a highly conserved tetrameric lysosomal cysteine dipeptidyl aminopeptidase. The best characterized physiological function of CatC is the activation of pro-inflammatory granule-associated serine proteases. These proteases are synthesized as inactive zymogens containing an N-terminal pro-dipeptide, which maintains the zymogen in its inactive conformation and prevents premature activation, which is potentially toxic to the cell. The activation of serine protease zymogens occurs through cleavage of the N-terminal dipeptide by CatC during cell maturation in the bone marrow. In vivo data suggest that pharmacological inhibition of pro-inflammatory serine proteases would suppress or attenuate deleterious effects mediated by these proteases in inflammatory/auto-immune disorders. The pathological deficiency in CatC is associated with Papillon-Lefèvre syndrome (PLS). The patients however do not present marked immunodeficiency despite the absence of active serine proteases in immune defense cells. Hence, the transitory pharmacological blockade of CatC activity in the precursor cells of the bone marrow may represent an attractive therapeutic strategy to regulate activity of serine proteases in inflammatory and immunologic conditions. A variety of CatC inhibitors have been developed both by pharmaceutical companies and academic investigators, some of which are currently being employed and evaluated in preclinical/clinical trials.

Modulation of Neutrophil Extracellular Trap and Reactive Oxygen Species Release by Periodontal Bacteria.

Oral bacteria are the main trigger for the development of periodontitis, and some species are known to modulate neutrophil function. This study aimed to explore the release of neutrophil extracellular traps (NETs), associated antimicrobial proteins, and reactive oxygen species (ROS) in response to periodontal bacteria, as well as the underlying pathways. Isolated peripheral blood neutrophils were stimulated with 19 periodontal bacteria. NET and ROS release, as well as the expression of NET-bound antimicrobial proteins, elastase, myeloperoxidase, and cathepsin G, in response to these species was measured using fluorescence-based assays. NET and ROS release was monitored after the addition of NADP (NADPH) oxidase pathway modulators and inhibitors of Toll-like receptors (TLRs). Moreover, bacterial entrapment by NETs was visualized microscopically, and bacterial killing was assessed by bacterial culture. Certain microorganisms, e.g., Veillonella parvula and Streptococcus gordonii, stimulated higher levels of ROS and NET release than others. NETs were found to entrap, but not kill, all periodontal bacteria tested. NADPH oxidase pathway modulators decreased ROS production but not NET production in response to the bacteria. Interestingly, TLR inhibitors did not impact ROS and NET release. These data suggest that the variability in the neutrophil response toward different bacteria may contribute to the pathogenesis of periodontal diseases by mechanisms such as bacterial avoidance of host responses and activation of neutrophils. Moreover, our results indicate that bacterium-stimulated NET release may arise in part via NADPH oxidase-independent mechanisms. The role of TLR signaling in bacterium-induced ROS and NET release needs to be further elucidated.

Assessment of the involvement of the macrophage migration inhibitory factor-glucocorticoid regulatory dyad in the expression of matrix metalloproteinase-2 during periodontitis.

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine and counter-regulator of endogenous glucocorticoids (GCs). It is implicated in acute and chronic inflammatory diseases. This study investigated the role of the MIF-GC regulatory dyad in the expression and release of matrix metalloproteinase-2 (MMP-2) during periodontitis, in vivo and in vitro. In a Mif-knockout (KO) mouse model of ligature-induced periodontitis, gingival tissues and blood were collected and analysed for levels of interleukin-6 (IL-6), MIF, MMP-2, and corticosterone. In addition, human gingival fibroblasts (HGFs) were tested for production of IL-6 and MMP-2 after stimulation with hydrocortisone (HC), MIF, tumour necrosis factor-alpha (TNF-α), or Fusobacterium nucleatum, a pathogen known to elicit immune responses during periodontitis. Wild-type (WT) mice showed a local and systemic increase of MIF levels during inflammation, which was confirmed by increased local IL-6 concentrations. Systemic GC levels were reduced in WT and Mif-KO mice during inflammation, with overall lower concentrations in Mif-KO mice. In vivo and in vitro, MMP-2 production was not dependent on MIF or inflammatory stimuli, but was inhibited by HC. Therefore, MIF does not appear to stimulate expression of MMP-2 in the gingival tissues, whereas GC upregulates MIF and downregulates MMP-2. Our findings further suggest that MIF may regulate systemic GC levels.

Long-term release of antibiotics by carbon nanotube-coated titanium alloy surfaces diminish biofilm formation by Staphylococcus epidermidis.

Bacterial biofilms cause a considerable amount of prosthetic joint infections every year, resulting in morbidity and expensive revision surgery. To address this problem, surface modifications of implant materials such as carbon nanotube (CNT) coatings have been investigated in the past years. CNTs are biologically compatible and can be utilized as drug delivery systems. In this study, multi-walled carbon nanotube (MWCNT) coated TiAl6V4 titanium alloy discs were fabricated and impregnated with Rifampicin, and tested for their ability to prevent biofilm formation over a period of ten days. Agar plate-based assays were employed to assess the antimicrobial activity of these surfaces against Staphylococcus epidermidis. It was shown that vertically aligned MWCNTs were more stable against attrition on rough surfaces than on polished TiAl6V4 surfaces. Discs with coated surfaces caused a significant inhibition of biofilm formation for up to five days. Therefore, MWCNT-modified surfaces may be effective against pathogenic biofilm formation on endoprostheses.