[Show abstract][Hide abstract] ABSTRACT: Periodontal infection (Periodontitis) is a chronic inflammatory disease, which results in the breakdown of the supporting tissues of the teeth. Previous epidemiological studies have suggested that resistance to chronic periodontitis is controlled to some extent by genetic factors of the host. The aim of this study was to determine the phenotypic response of inbred and Collaborative Cross (CC) mouse populations to periodontal bacterial challenge, using an experimental periodontitis model. In this model, mice are co-infected with Porphyromonas gingivalis and Fusobacterium nucleatum, bacterial strains associated with human periodontal disease. Six weeks following the infection, the maxillary jaws were harvested and analyzed for alveolar bone loss relative to uninfected controls, using computerized microtomography (microCT). Initially, four commercial inbred mouse strains were examined to calibrate the procedure and test for gender effects. Subsequently, we applied the same protocol to 23 lines (at inbreeding generations 10--18) from the newly developed mouse genetic reference population, the Collaborative Cross (CC) to determine heritability and genetic variation of control bone volume prior to infection (CBV, naive bone volume around the teeth of uninfected mice), and residual bone volume (RBV, bone volume after infection) and loss of bone volume (LBV, the difference between CBV and RBV) following infection RESULTS: BALB/CJ mice were highly susceptible (P<0.05) whereas DBA/2J, C57BL/6J and A/J mice were resistant. Six lines of the tested CC population were susceptible, whereas the remaining lines were resistant to alveolar bone loss. Gender effects on bone volume were tested across the four inbred and 23 CC lines, and found not to be significant. Based on ANOVA analyses, broad-sense heritabilities were statistically significant and equal to 0.4 for CBV and 0.2 for LBV.
The moderate heritability values indicate that the variation in host susceptibility to the disease is controlled to an appreciable extent by genetic factors. These results strongly support the possibility of using the Collaborative Cross, as well as developing dedicated F2 (resistant x susceptible inbred strains) resource populations, for future dissection of genetic factors in periodontitis.
[Show abstract][Hide abstract] ABSTRACT: To investigate the in vivo role of gingipains in Porphyromonas gingivalis' virulence, and suggest a possible host mechanisms through which the bacteria cause alveolar bone loss.
Mice were orally infected with P. gingivalis wild type, or the gingipains mutants (RgpA(-) , Kgp(-) , RgpA(-) /Kgp(-) ). Mice were analysed for alveolar bone loss using micro-computed tomography. The molecular effects of the proteases were evaluated using the subcutaneous chamber model. Mice were infected with P. gingivalis wild type or mutants. Exudates were analysed for cytokine and leukocytes levels, in vivo phagocytosis, P. gingivalis survival and serum anti-P. gingivalis IgG titres.
Only RgpA-expressing bacteria induced significantly alveolar bone loss, and suppressed phagocytosis resulting in increased survival of P. gingivalis in the chamber exudates. In addition, RgpA-expressing bacteria induced higher levels of leukocytes and cytokines 2 h post-infection, and reduced levels of serum anti-P. gingivalis IgG titres 7 days post-infection.
Our findings showed that elimination of RgpA from P. gingivalis diminished inflammation, but augmented phagocytosis and antibody titres, coincidental with reduced alveolar bone loss. These findings support the hypothesis that RgpA is a critical virulence factor in the pathogenesis of experimental periodontitis in mice.
Journal Of Clinical Periodontology 07/2013; · 3.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: T cells, particularly CD4(+) T cells, play a central role in both progression and control of periodontal disease, whereas the contribution of the various CD4(+) T helper subsets to periodontal destruction remains controversial, the activation, and regulation of these cells is orchestrated by dendritic cells. As sentinels of the oral mucosa, dendritic cells encounter and capture oral microbes, then migrate to the lymph node where they regulate the differentiation of CD4(+) T cells. It is thus clear that dendritic cells are of major importance in the course of periodontitis, as they hold the immunological cues delivered by the pathogen and the surrounding environment, allowing them to induce destructive immunity. In recent years, advanced immunological techniques and new mouse models have facilitated in vivo studies that have provided new insights into the developmental and functional aspects of dendritic cells. This progress has also benefited the characterization of oral dendritic cells, as well as to their function in periodontitis. Here, we provide an overview of the various gingival dendritic cell subsets and their distribution, while focusing on their role in periodontal bone loss.
[Show abstract][Hide abstract] ABSTRACT: We have developed a technique to precisely isolate and process murine gingival tissue for flow cytometry and molecular studies. The gingiva is a unique and important tissue to study immune mechanisms because it is involved in host immune response against oral biofilm that might cause periodontal diseases. Furthermore, the close proximity of the gingiva to alveolar bone tissue enables also studying bone remodeling under inflammatory conditions. Our method yields large amount of immune cells that allows analysis of even rare cell populations such as Langerhans cells and T regulatory cells as we demonstrated previously (1). Employing mice to study local immune responses involved in alveolar bone loss during periodontal diseases is advantageous because of the availability of various immunological and experimental tools. Nevertheless, due to their small size and the relatively inconvenient access to the murine gingiva, many studies avoided examination of this critical tissue. The method described in this work could facilitate gingival analysis, which hopefully will increase our understating on the oral immune system and its role during periodontal diseases.
[Show abstract][Hide abstract] ABSTRACT: Excessive bone resorption is frequently associated with chronic infections and inflammatory diseases. Whereas T cells were demonstrated to facilitate osteoclastogenesis in such diseases, the role of dendritic cells, the most potent activators of naive T cells, remains unclear. Using a model involving inflammation-driven alveolar bone loss attributable to infection, we showed that in vivo ablation of Langerhans cells (LCs) resulted in enhanced bone loss. An increased infiltration of B and T lymphocytes into the tissue surrounding the bone was observed in LC-ablated mice, including receptor activator of NF-κB ligand (RANKL)-expressing CD4(+) T cells with known capabilities of altering bone homeostasis. In addition, the absence of LCs significantly reduced the numbers of CD4(+)Foxp3(+) T-regulatory cells in the tissue. Further investigation revealed that LCs were not directly involved in presenting antigens to T cells. Nevertheless, despite their low numbers in the tissue, the absence of LCs resulted in an elevated activation of CD4(+) but not CD8(+) T cells. This activation involved elevated production of IFN-γ but not IL-17 or IL-10 cytokines. Our data, thus, reveal a protective immunoregulatory role for LCs in inflammation-induced alveolar bone resorption, by inhibiting IFN-γ secretion and excessive activation of RANKL(+)CD4(+) T cells with a capability of promoting osteoclastogenesis.
Proceedings of the National Academy of Sciences 04/2012; 109(18):7043-8. · 9.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Periodontitis is a common human chronic inflammatory disease that results in the destruction of the tooth attachment apparatus and tooth loss. Although infections with periopathogenic bacteria such as Porphyromonas gingivalis (P. gingivalis) and Fusobacterium nucleatum (F. nucleatum) are essential for inducing periodontitis, the nature and magnitude of the disease is determined by the host's immune response. Here, we investigate the role played by the NK killer receptor NKp46 (NCR1 in mice), in the pathogenesis of periodontitis. Using an oral infection periodontitis model we demonstrate that following F. nucleatum infection no alveolar bone loss is observed in mice deficient for NCR1 expression, whereas around 20% bone loss is observed in wild type mice and in mice infected with P. gingivalis. By using subcutaneous chambers inoculated with F. nucleatum we demonstrate that immune cells, including NK cells, rapidly accumulate in the chambers and that this leads to a fast and transient, NCR1-dependant TNF-α secretion. We further show that both the mouse NCR1 and the human NKp46 bind directly to F. nucleatum and we demonstrate that this binding is sensitive to heat, to proteinase K and to pronase treatments. Finally, we show in vitro that the interaction of NK cells with F. nucleatum leads to an NCR1-dependent secretion of TNF-α. Thus, the present study provides the first evidence that NCR1 and NKp46 directly recognize a periodontal pathogen and that this interaction influences the outcome of F. nucleatum-mediated periodontitis.
[Show abstract][Hide abstract] ABSTRACT: Background: Implantoplasty is one of the options in treating peri-implantitis. The efficacy of the dental bur used can reduce the time needed for the procedure and, as a consequence, minimize the risk of overheating that can negatively affect the remaining bone surrounding the implant. Purpose: The aim of this study was to evaluate the efficacy of three dental burs in removing implant substance (titanium) and to determine the amount of heat generated by each bur. Materials and Methods: Four burs with different surface properties (diamond, diamond - Premium Line, carbide, and smooth bur - control [Strauss Co., Raanana, Israel]) were attached to a high-speed handpiece and applied to a titanium implant for a total of 60 seconds after cooling by water spray. Variations in temperature were recorded every 5 seconds, and the amount of implant substance removed (reduction in weight of the implant) was evaluated. Results: The diamond Premium Line bur removed 59.24 mg; carbide, 29.39 mg; diamond, 11.35 mg; and smooth bur (control) 0.19 mg, statistically significant. Only minimum thermal changes (∼1.5°C) were recorded for all four burs. Conclusions: There are considerable differences in efficiency of different burs working on titanium. Selecting the proper bur can reduce working time. Under proper cooling conditions, implantoplasty does not generate excess temperature increases that can damage soft tissue or bone surrounding the treated implant.
Clinical Implant Dentistry and Related Research 07/2011; · 3.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Objectives: To examine whether oral lichen planus (OLP) affects the success rate of dental implants and if the manifestations of OLP are altered by implant-borne prostheses. Materials and Methods: OLP patients, treated in the oral medicine department, with (the study group) and without (control group) dental implants were included. Pocket depth, mobility, bleeding on probing, erythema, pain and radiolucency around the implants, as well as clinical findings and OLP symptoms were recorded. Follow-up ranged from 12-24 months. Ordinal variables and visual analog scale score were compared using the Mann-Whitney test. The significance of the trend within each of the groups was assed using the Friedman test. Categorical variables were compared using Pearson chi-squared test and Fisher's exact test. Results: Fourteen patients in the study group with 1-15 implants per patient and 15 in the control group were included. No implant failures were recorded. Comparison between the clinical manifestations of OLP in both groups did not reveal any significant differences. Conclusions: Success of implant rehabilitation among treated OLP patients does not seem to be different from the success rate in the general population. Nor does implant placement influence the disease manifestations.
Clinical Implant Dentistry and Related Research 05/2011; · 3.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Although oral dendritic cells (DCs) were shown to induce cell-mediated immunity, the identity and function of the various oral DC subsets involved in this process is unclear. In this study, we examined the mechanisms used by DCs of the buccal mucosa and of the lining mucosa to elicit immunity. After plasmid DNA immunization, buccally immunized mice generated robust local and systemic CD8(+) T cell responses, whereas lower responses were seen by lining immunization. A delayed Ag presentation was monitored in vivo in both groups; yet, a more efficient presentation was mediated by buccal-derived DCs. Restricting transgene expression to CD11c(+) cells resulted in diminished CD8(+) T cell responses in both oral tissues, suggesting that immune induction is mediated mainly by cross-presentation. We then identified, in addition to the previously characterized Langerhans cells (LCs) and interstitial dendritic cells (iDCs), a third DC subset expressing the CD103(+) molecule, which represents an uncharacterized subset of oral iDCs expressing the langerin receptor (Ln(+)iDCs). Using Langerin-DTR mice, we demonstrated that whereas LCs and Ln(+)iDCs were dispensable for T cell induction in lining-immunized mice, LCs were essential for optimal CD8(+) T cell priming in the buccal mucosa. Buccal LCs, however, failed to directly present Ag to CD8(+) T cells, an activity that was mediated by buccal iDCs and Ln(+)iDCs. Taken together, our findings suggest that the mechanisms engaged by oral DCs to prime T cells vary between oral mucosal tissues, thus emphasizing the complexity of the oral immune network. Furthermore, we found a novel regulatory role for buccal LCs in potentiating CD8(+) T cell responses.
The Journal of Immunology 01/2011; 186(2):891-900. · 5.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To assess the potential of using vaccination with Porphyromonas gingivalis or Fusobacterium nucleatum, in modulating local subcutaneous inflammatory response and alveolar bone loss following coinfection with both bacteria.
Mice were immunized against either P. gingivalis or F. nucleatum. The cytokine response to mixed infection with P. gingivalis and F. nucleatum was evaluated using the subcutaneous chamber model. The alveolar bone loss induced by oral mixed infection was evaluated by micro-CT using the experimental periodontitis model. Serum levels of specific antibodies were determined by ELISA.
Vaccination with either bacterium produced a specific humoral response before infection. Animals immunized against either bacteria following a mixed infection with P. gingivalis and F. nucleatum, showed decreased TNFalpha (but not IL-1beta) levels as compared with non-immunized animals. However, the vaccination did not change the level of mixed infection-induced alveolar bone loss when compared with non-immunized animals. Six weeks following the oral mixed infection, specific antibody titres remained high. Furthermore, specific antibodies against the non-immunized bacterium were present at high levels.
While vaccination produced specific antibodies and suppressed the inflammatory response, it failed to prevent or reduce the progression of experimental periodontitis induced by mixed infection with P. gingivalis and F. nucleatum.
Journal Of Clinical Periodontology 09/2010; 37(9):812-7. · 3.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To evaluate the effect of oral infection with three Porphyromonas gingivalis strains on alveolar bone loss (ABL) and its correlation with the mouse immune response.
Mice were orally infected with P. gingivalis strains 381, 33277 and 53977. After 42 days, maxillae were analysed for ABL using micro-computed tomography and the serum for anti-P.gingivalis IgG1 and IgG2a levels. The cytokine response to P. gingivalis was tested using the subcutaneous chamber model.
The P. gingivalis 53977-infected group showed the highest ABL, which was significantly different from all other groups (p<0.001). In addition, the humoral response to P. gingivalis 53977 was significantly lower than the response to P. gingivalis 381 and 33277 (p< or =0.01). The IgG2a/IgG1 ratio was higher in the P. gingivalis 33277-infected group (1.6) compared with the P. gingivalis 381-infected group (0.51). Four days post-infection, interleukin (IL)-1beta levels remained significantly higher in the P. gingivalis 53977-infected group only (1198.2+/-260.0, p<0.05), while IL-4 levels remained significantly higher in the P. gingivalis 381-infected group (265.8+/-131.6, p<0.05).
The high levels of ABL induced by P. gingivalis 53977 were inversely correlated with the humoral response to this bacterium. In addition, ABL was correlated with an elevated pro-inflammatory response.
Journal Of Clinical Periodontology 09/2009; 36(11):915-21. · 3.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The chemistry of titanium is a key factor in determining implant-tissue interactions. Reports that a vanadium-based titanium alloy (Ti-6Al-4V) exhibits some cytotoxicity led to a search for an alternative.
The aim of the present study was to investigate the behavior of human osteoblast-like cells (Saos-2) cultured on Ti-6Al-4V or Ti-6Al-7Nb disks with a rough or a machined surface.
In all four groups, the cells proliferated rapidly between days 1 and 3, and then plateaued. On day 1 of culture, the highest proliferation rate was of cells cultured on disks containing Nb with a machined surface. On day 7, there was no significant difference in cell density on all the tested surfaces. Alkaline phosphatase (ALP) activity was lower on the machined surfaces, regardless of the material used, suggesting that cells on rough surfaces exhibit a more mature phenotype. On day 3, cells cultured on rough disks made of Ti-6Al-7Nb showed the highest ALP activity; the lowest activity was observed on the machined Ti-6Al-4V surface. The highest level of osteocalcin (day 7) was found in the cells cultured on rough Ti-6Al-7Nb disks. Also, higher levels of transforming growth factor (TGFbeta) were noted for cells cultured on the rough Ti-6Al-7Nb disks, suggesting that the Nb-containing alloy supports more rapid maturation of the osteoblast.
The results of the present study suggest that according to our cell culture preclinical model, Ti-6Al-7Nb may replace the Ti-6Al-4V alloy as an implant material.
Clinical Oral Implants Research 07/2009; 20(6):578-82. · 3.43 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To compare the effect of oral infection with Porphyromonas gingivalis or Fusobacterium nucleatum versus infection with both bacteria on mouse periodontal tissues, and to characterize the inflammatory response.
Mice were orally infected with P. gingivalis, F. nucleatum or both. At 42 days post-infection, alveolar bone loss was quantified using micro-computerized tomography. Tumour necrosis factor-alpha (TNF-alpha) and interleukin (IL)-1beta levels induced by the infection were quantified using the subcutaneous chamber model.
Mice orally infected with F. nucleatum/P. gingivalis exhibited significantly more bone loss compared with that of mono-infected and sham-infected mice. F. nucleatum/P. gingivalis infection also increased the levels of TNF-alpha and IL1beta compared with the levels found in the mono-infected groups.
Polymicrobial infection with P. gingivalis/F. nucleatum aggravates alveolar bone loss and induces a stronger inflammatory response compared with that observed upon infection with either bacterium alone. The results suggest that oral infection of mice with a mixture of P. gingivalis and F. nucleatum may be superior to mono-infection models of experimental periodontitis.
Journal Of Clinical Periodontology 06/2009; 36(5):406-10. · 3.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Distraction osteogenesis is routinely used for reconstruction of bone. Conversely, it was hypothesized that mechanical traction of the periosteum would induce bone formation, and hence the use of periosteal distraction for induction of osteogenesis has been proposed. Further, it was postulated that intracallus administration of vascular endothelial growth factor (VEGF) would facilitate osteogenesis. To investigate this hypothesis, formation of newly synthesized bone was evaluated using micro-computerized tomography (microCT) and histomorphometry.
Periosteal distractors were placed subperiosteally in one side of the mandible of rabbits, whereas the contralateral served as control. One group of animals received VEGF into the forming callus. Formation of bone was measured using microCT and histological analysis.
The results demonstrate formation of new bone following periosteal distraction. Addition of VEGF to the distraction site increased bone synthesis.
microCT and histological analysis validate the hypothesis that mechanical distraction of the periosteum induces osteogenesis and that VEGF has a positive effect on osteogenesis. Periosteal distraction is emerging as a reliable technique for bone regeneration.
Tissue Engineering Part A 03/2008; 14(2):247-53. · 4.07 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Animal models are routinely used for the study of the pathogenesis of periodontal disease. However, some of the methods used for evaluating alveolar bone loss are limited to linear or two dimension measurements, while other methods, such as histology, are labor consuming. The present study was designed to evaluate a novel method for assessing the volume of alveolar bone loss in mice and to compare it to the traditional morphometric (linear) technique.
Seven- to 8-week-old BALB/c mice were divided into three equal groups of six each; two groups were infected orally with Porphyromonas gingivalis (P. gingivalis) 381 or 53,977, while the third group was used as non-infected control. Forty-two days following infection, serum samples were obtained and maxillae were harvested. Bone loss was evaluated by micro-computed tomography (micro-CT) and by the morphometric technique.
Significant decrease in residual supportive bone volumes (RSBV) was observed in both P. gingivalis-infected groups compared to the control group (P<0.0001). The P. gingivalis 53,977-infected group showed less residual supportive bone volume compared to the P. gingivalis 381-infected group, but there were no significant differences between these two groups. Using the morphometric technique, the differences between the three groups were not found to be statistically significant (P>0.05).
The present study describes a novel micro-CT technique for volumetric evaluation of alveolar bone loss in mice. This technique is relatively simple and accurate, and due to its high sensitivity, enables the investigator to reduce the number of animals needed in each experimental group.
Journal of Periodontology 08/2005; 76(8):1282-6. · 2.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To review the association between genetic variability and the inflammatory response induced by periodontal infection.
A search of MEDLINE-PubMed was performed from January 2000 up to and including March 2005. The search included all types of publications, published in English without other limitations. The following search terms were used: "cytokine polymorphism", "gene polymorphism", "periodontitis", "gingivitis", "inflammation" and "host-response". The papers resulting from the above search were used as an additional source for relevant articles.
Genetic variability was examined for the correlation to clinical indicators of inflammation such as bleeding on probing (BOP), gingival inflammation, cytokine in gingival crevicular fluid (GCF) and cytokine production by inflammatory cells. According to the current literature, most of the studies found no association between genetic variability and BOP, gingival inflammation or cytokine concentrations in the GCF. These studies were hampered by inappropriate study designs and the use of inflammatory parameters as secondary rather than primary outcome variables. The data suggest that the production of inflammatory mediators by inflammatory cells may be affected by different genetic traits but further studies are needed in order to establish this association.
To date, there is no clear correlation between any of the gene polymorphisms and clinical indicators of inflammation. The powering of studies to reveal associations between single or multiple nucleotide polymorphisms and inflammatory parameters will need to involve a much larger number of subjects than were used in the past. The available data (including the interleukin-1 composite genotype) do not currently support the utility of such tests in the diagnosis and prognostic assessments of periodontal diseases.