Journal of Dental Research

The oral environment contains diverse communities of micro-organisms including bacteria, fungi, protozoa, and viruses. Studies of oral ecology have led to an appreciation of the complexity of the interactions that oral micro-organisms have with the host in both health and disease. Despite this, diseases such as dental caries and periodontal diseases are still worldwide human ailments, resulting in a high level of morbidity and an economic burden to society. Proteomics offers a new approach to the understanding of holistic changes occurring as oral micro-organisms adapt to environmental change within their habitats in the mouth.

Expression of human β-defensins is correlated with differentiation in the oral epithelium, consistent with their function as part of the epithelial antimicrobial barrier. Because calcium is a known regulator of epithelial differentiation, we tested the hypothesis that calcium concentration mediates β-defensin expression. Gingival epithelial cells were cultured in medium containing low calcium concentration (0.03 mM), then either changed to high extracellular calcium concentrations or stimulated with thapsigargin to release intracellular calcium stores in the presence or absence of BAPTA-AM, a calcium chelator. Human β-defensin-2 (hBD-2) mRNA expression was rapidly induced by thapsigargin, and more slowly induced by high extracellular calcium. Induction of hBD-2 peptide was confirmed by immunofluorescence. BAPTA-AM inhibited hBD-2 induction by both thapsigargin and calcium in a dose-dependent fashion. In addition, BAPTA-AM inhibited hBD-2 induction by a bacterial stimulant. Collectively, these findings demonstrate that intracellular calcium is a critical mediator of hBD-2 expression. Abbreviations used in this study are: BAPTA-AM, 1,2-bis(2-aminophenoxy)-ethane-N,N,N′,N′-tetra-acetic acid tetrakis (acetoxymethyl ester); DMSO, dimethylsulfoxide; F. nucleatum, Fusobacterium nucleatum; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HBDs, human β-defensins; HGECs, human gingival epithelial cells; MAP, mitogen-activated protein; and RT-PCR, reverse-transcriptase/polymerase chain-reaction.

The membrane-bound mucin MUC1 is expressed ubiquitously on epithelial surfaces and is thought to provide protection from bacterial and chemical injury. The present study was undertaken to determine whether MUC1 was expressed in cultured oral epithelial cells and whether expression is modulated by pro-inflammatory mediators released as part of the host response to infection by oral pathogens. Northern and Western blotting experiments showed that KB cells express MUC1 mRNA and protein. When cells were treated with interleukins (IL-1β, IL-6), tumor necrosis factor-alpha (TNF-α), or interferon-gamma (IFN-γ), or combinations of these, real-time PCR demonstrated that MUC1 mRNA increased 1.4- to 3.2-fold. Interestingly, a significant increase in levels of MUC1 protein was also observed. While no effect was observed when KB cells were incubated with LPS from Porphyromonas gingivalis, infection of KB monolayers with this oral pathogen caused a 2.85-fold increase in MUC1 transcript levels. These results suggest that increased MUC1 synthesis may be a key element in the host response to infection with oral pathogens.

Adenosine has various biological effects on human gingival fibroblasts (HGF) and epithelial cells closely associated with inflammation, such as cytokine production and cell adhesion. However, the mechanism of adenosine formation in periodontal tissues is not yet defined. In this study, we examined the involvement of CD73 (ecto-5′-nucleotidase) in adenosine generation by HGF. CD73 was detected on in vitro-maintained HGF by immunocytochemistry and flow cytometric analysis. Adenosine production was observed following the addition of 5′-AMP, the substrate of CD73-associated ecto-5′-nucleotidase. Moreover, the addition of 5′-AMP to cultured HGF resulted in the elevation of cyclic adenosine monophosphate (cAMP). The 5′-AMP-induced increase in intracellular cAMP level was inhibited markedly by xanthine amine congener, an adenosine receptor antagonist, and partially by α,β-methylene adenosine 5′-diphosphate, an ecto-5′-nucleotidase inhibitor. These results suggest that CD73 on HGF is a critical enzyme responsible for the generation of adenosine, an immunomodulator that activates adenosine receptors.

Lymphocytes in peripheral blood do not express CD13 (aminopeptidase N), a membrane alanyl metallopeptidase. However, it has been demonstrated that locally infiltrated lymphocytes in chronic inflammatory sites can be CD13-positive, and possible involvement of stromal cell adherence in the induction of CD13 has been suggested. In this study, we examined whether T-lymphocyte/gingival-fibroblast interaction can activate T-lymphocytes to express CD13. CD13 expression was induced on PMA-activated T-lymphocytes only when they adhered directly to human gingival fibroblasts (HGF) at 2 hrs after the co-culture began, while an increase in the enzyme activity of CD13 was also confirmed in activated T-lymphocytes that had been co-cultured with HGF. Furthermore, CD13-positive T-lymphocytes were detected in inflamed gingival tissues in vivo. Analysis of these results indicates that direct interaction with HGF is essential for the induction of CD13 expression on T-lymphocytes that was also observed in periodontitis lesions.

The submandibular gland is a cell-rich encapsulated organ with high transport of fluid through the interstitial space during salivation. We hypothesized that the gland is a low-compliant tissue, i.e., that a modest increase in fluid volume will produce a rise in interstitial fluid pressure (IFP) counteracting fluid filtration into the interstitium. To test this hypothesis, we measured IFP with micropipettes and glandular blood flow (GBF) with a laser-Doppler flowmeter during changes in perfusion. Clamping of the carotid artery or the jugular vein, or electrical stimulation of the sympathetic or parasympathetic nerve to the gland, induced changes in perfusion. Baseline IFP averaged 3.5 ± 0.5 mm Hg. Clamping of the artery reduced IFP and GBF (−56.5 ± 8.4% and −53.1 ± 6.4%, respectively), whereas clamping of the vein decreased GBF (−21.6 ± 14.3%) and increased IFP (141.2 ± 27.4%). Sympathetic nerve stimulation reduced both parameters (−86.9 ± 16.5% and −74.4 ± 7.0%, respectively). In contrast, stimulation of the parasympathetic nerve elicited an increase in GBF (133.2 ± 5.9%) and in IFP (173.3 ± 41.4%). Thus, changes in vascular volume led to concomitant changes in IFP consistent with low tissue compliance, a phenomenon of importance for fluid volume regulation.

It is widely assumed that coupling the degradation rate of polymers used as cell transplantation carriers to the growth rate of the developing tissue will improve its quantity or quality. To test this hypothesis, we developed alginate hydrogels with a range of degradation rates by gamma-irradiating high-molecular-weight alginate to yield polymers of various molecular weights and structures. Decreasing the size of the polymer chains increased the degradation rate in vivo, as measured by implant retrieval rates, masses, and elastic moduli. Rapidly and slowly degrading alginates, covalently modified with RGD-containing peptides to control cell behavior, were then used to investigate the effect of biodegradation rate on bone tissue development in vivo. The more rapidly degrading gels led to dramatic increases in the extent and quality of bone formation. These results indicate that biomaterial degradability is a critical design criterion for achieving optimal tissue regeneration with cell transplantation.

Despite fluoride’s (F) well-documented ability to prevent caries, the effects of F concentrations on enamel and dentin apatite crystals are unknown. The present study examined the hypothesis that tooth F concentration and tooth crystallite size correlate. One hundred human unerupted third molars were studied—53 from Fortaleza-Brazil (F water 0.7 ppm), 23 from Toronto (1.0 ppm), and 24 from Montreal (0.2 ppm). F concentration was analyzed by Neutron Activation Analysis and apatite crystal size by powder x-ray diffraction. A positive correlation between dentin F concentration and enamel crystallite length and width was found. Enamel crystallite length was significantly greater in teeth from Fortaleza than in teeth from Toronto (p = 0.011) and Montreal (p = 0.003). Enamel crystallite widths were significantly greater in Fortaleza teeth compared with those from Toronto (p = 0.020) and Montreal (p < 0.001). No difference in the dentin crystallite size was seen in the 3 regions. Thus, tooth F concentration and crystallite size correlate.

Casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) nanocomplexes have been shown to prevent demineralization and promote remineralization of enamel subsurface lesions in animal and in situ caries models. The aim of this study was to determine the effect of incorporating CPP-ACP into a self-cured glass-ionomer cement (GIC). Incorporation of 1.56% w/w CPP-ACP into the GIC significantly increased microtensile bond strength (33%) and compressive strength (23%) and significantly enhanced the release of calcium, phosphate, and fluoride ions at neutral and acidic pH. MALDI mass spectrometry also showed casein phosphopeptides from the CPP-ACP nanocomplexes to be released. The release of CPP-ACP and fluoride from the CPP-ACP-containing GIC was associated with enhanced protection of the adjacent dentin during acid challenge in vitro.

Antiphospholipid antibodies are commonly found in patients with systemic lupus erythematosus or the antiphospholipid syndrome, and a subset of such antibodies is associated with prothrombotic events such as stroke and with adverse pregnancy outcomes and fetal loss. We examined sera from 411 patients who were clinically characterized as to their periodontal disease status for serum levels of β2-glycoprotein I-dependent anti-cardiolipin autoantibodies (anti-CL). The prevalence of patients with chronic periodontitis (CP) and generalized aggressive periodontitis (GAgP) positive for anti-CL (16.2% and 19.3%, respectively) was greater than that in healthy controls (NP) and localized aggressive periodontitis (LAgP) patients (6.8% and 3.2%). Patients with these autoantibodies demonstrated increased pocket depth and attachment loss compared with patients lacking the antibodies. Analysis of the data indicates that patients with generalized periodontitis have elevated levels of autoantibodies reactive with phospholipids. These antibodies could be involved in elevated risk for stroke, atherosclerosis, or pre-term birth in periodontitis patients.

Survivin is a recently described apoptosis inhibitor selectively over-expressed in most tumors. Immunohistochemistry was used to investigate a potential role of survivin as an early predictor of malignant transformation in precancerous and cancerous lesions of the oral cavity. Survivin was present in 10/30 cases (33%) of oral precancerous lesions without malignant progression, and in 15/16 cases (94%) of oral precancerous lesions evolved into full-blown squamous cell carcinoma. Tumors that progressed from these precancerous lesions retained widespread survivin positivity (100%). Variations among group means were highly statistically significant (p < 0.001). No significant correlation was found between survivin expression and the degree of dysplasia. High expression of cytoplasmic/nuclear survivin is an early event during oral carcinogenesis and may provide a useful tool for the identification of precancerous lesions at higher risk of progression into invasive carcinoma.

Recent research shows that what people hear can influence what they feel. We investigated whether the perception of an electric toothbrush might also be affected by the sound that it makes. Participants were required to make stereotypical brushing movements with a standard electric toothbrush while they rated either the pleasantness or the roughness of the vibrotactile stimulation they felt on their teeth. The results demonstrate that the perception of the sensations experienced during toothbrush use were systematically altered by variations in the auditory feedback elicited by the brushing action. Participants reported that the toothbrush felt more pleasant and less rough when either the overall sound level was reduced, or when just the high-frequency sounds were attenuated. These results highlight the significant role that auditory cues can play in modulating the perception and evaluation of everyday products in use, and provide a paradigm for future study in this area.