Journal of Dental Research

Monomers are released from dental resin materials, and thus cause adverse biological effects in mammalian cells. Cytotoxicity and genotoxicity of some of these methacrylates have been identified in a vast number of investigations during the last decade. It has been well-established that the co-monomer triethylene glycol dimethacrylate (TEGDMA) causes gene mutations in vitro. The formation of micronuclei is indicative of chromosomal damage and the induction of DNA strand breaks detected with monomers like TEGDMA and 2-hydroxyethyl methacrylate (HEMA). As a consequence of DNA damage, the mammalian cell cycle was delayed in both G1 and G2/M phases, depending on the concentrations of the monomers. Yet, the mechanisms underlying the genetic and cellular toxicology of resin monomers have remained obscure until recently. New findings indicate that increased oxidative stress results in an impairment of the cellular pro- and anti-oxidant redox balance caused by monomers. It has been demonstrated that monomers reduced the levels of the natural radical scavenger glutathione (GSH), which protects cell structures from damage caused by reactive oxygen species (ROS). Depletion of the intracellular GSH pool may then significantly contribute to cytotoxicity, because a related increase in ROS levels can activate pathways leading to apoptosis. Complementary, cytotoxic, and genotoxic effects of TEGDMA and HEMA are inhibited in the presence of ROS scavengers like N-acetylcysteine (NAC), ascorbate, and Trolox (vitamin E). Elevated intracellular levels of ROS can also activate a complex network of redox-responsive macromolecules, including redox-sensitive transcription factors like nuclear factor kappaB (NF-κB). It has been shown that NF-κB is activated probably to counteract HEMA-induced apoptosis. The induction of apoptosis by TEGDMA in human pulp cells has been associated with an inhibition of the phosphatidylinositol 3-kinase (PI3-K) cell-survival signaling pathway. Although the details of the mechanisms leading to cell death, genotoxicity, and cell-cycle delay are not completely understood, resin monomers may be able to alter the functions of the cells of the oral cavity. Pathways regulating cellular homeostasis, dentinogenesis, or tissue repair may be modified by monomers at concentrations well below those which cause acute cytotoxicity.

Dental caries is a biofilm-dependent oral disease, and fermentable dietary carbohydrates are the key environmental factors involved in its initiation and development. However, among the carbohydrates, sucrose is considered the most cariogenic, because, in addition to being fermented by oral bacteria, it is a substrate for the synthesis of extracellular (EPS) and intracellular (IPS) polysaccharides. Therefore, while the low pH environment triggers the shift of the resident plaque microflora to a more cariogenic one, EPS promote changes in the composition of the biofilms’ matrix. Furthermore, it has recently been shown that the biofilm formed in the presence of sucrose presents low concentrations of Ca, P_i, and F, which are critical ions involved in de- and remineralization of enamel and dentin in the oral environment. Thus, the aim of this review is to explore the broad role of sucrose in the cariogenicity of biofilms, and to present a new insight into its influence on the pathogenesis of dental caries.

The paired-related homeobox genes, Prx1 and Prx2, encode transcription factors critical for orofacial development. Prx1 ^−/−/Prx2 ^−/− neonates have mandibular hypoplasia and malformed mandibular incisors. Although the mandibular incisor phenotype has been briefly described (ten Berge et al., 1998, 2001; Lu et al., 1999), very little is known about the role of Prx proteins during tooth morphogenesis. Since the posterior mandibular region was relatively normal, we examined molar tooth development in Prx1 ^−/−/Prx2 ^−/− embryos to determine whether the tooth malformation is primary to the loss of Prx protein or secondary to defects in surrounding tissues. Three-dimensional (3D) morphological reconstructions demonstrated that Prx1 ^−/−/Prx2 ^−/− embryos had molar malformations, including cuspal changes and ectopic epithelial projections. Although we demonstrate that Prx1 protein is expressed only mesenchymally, 3D reconstructions showed important morphological defects in epithelial tissues at the cap and bell stages. Analysis of these data suggests that the Prx homeoproteins are critical for mesenchymal-epithelial signaling during tooth morphogenesis.

Amelogenin RNA transcripts undergo extensive alternative splicing, and MMP-20 processes the isoforms following their secretion. Since amelogenins have been ascribed cell-signaling activities, we asked if a lack of proteolytic processing by MMP-20 affects amelogenin signaling and consequently alters amelogenin splice site selection. RT-PCR analyses of amelogenin mRNA between control and Mmp20 ^−/−mice revealed no differences in the splicing pattern. We characterized 3 previously unidentified amelogenin alternatively spliced transcripts and demonstrated that exon-8-encoded amelogenin isoforms are processed by MMP-20. Transcripts with exon 8 were expressed approximately five-fold less than those with exon 7. Analyses of the mouse and rat amelogenin gene structures confirmed that exon 8 arose in a duplication of exons 4 through 5, with translocation of the copy downstream of exon 7. No downstream genomic sequences homologous to exons 4–5 were present in the bovine or human amelogenin genes, suggesting that this translocation occurred only in rodents.

Although eugenol is widely used in dentistry, little is known about the molecular mechanisms responsible for its anesthetic properties. In addition to calcium channels, recently demonstrated by our group, there could be another molecular target for eugenol. Using a whole-cell patch-clamp technique, we investigated the effect of eugenol on voltage-gated sodium channel currents (I _Na ) in rat dental primary afferent neurons identified by retrograde labeling with a fluorescent dye in maxillary molars. Eugenol inhibited action potentials and I _Na in both capsaicin-sensitive and capsaicin-insensitive neurons. The pre-treatment with capsazepine, a competitive antagonist of transient receptor potential vanilloid 1 (TRPV1), failed to block the inhibitory effect of eugenol on I _Na , suggesting no involvement of TRPV1. Two types of I _Na , tetrodotoxin (TTX)-resistant and TTX-sensitive I _Na , were inhibited by eugenol. Our results demonstrated that eugenol inhibits I _Na in a TRPV1-independent manner. We suggest that I _Na inhibition by eugenol contributes to its analgesic effect.

Bone tissue can adapt to orthodontic load. Mechanosensing in bone is primarily a task for the osteocytes, which translate the canalicular flow resulting from bone loading into osteoclast and osteoblast recruiting signals. Apoptotic osteocytes attract osteoclasts, and inhibition of osteocyte apoptosis can therefore affect bone remodeling. Since TNF-α is a pro-inflammatory cytokine with apoptotic potency, and elevated levels are found in the gingival sulcus during orthodontic tooth movement, we investigated if mechanical loading by pulsating fluid flow affects TNF-α-induced apoptosis in chicken osteocytes, osteoblasts, and periosteal fibroblasts. During fluid stasis, TNF-α increased apoptosis by more than two-fold in both osteocytes and osteoblasts, but not in periosteal fibroblasts. One-hour pulsating fluid flow (0.70 ± 0.30 Pa, 5 Hz) inhibited (−25%) TNF-α-induced apoptosis in osteocytes, but not in osteoblasts or periosteal fibroblasts, suggesting a key regulatory role for osteocyte apoptosis in bone remodeling after the application of an orthodontic load.

Although the contribution of the oral microbiota to oral malodor is well-documented, the potential role of Gram-positive micro-organisms is unclear. In the current study, we tested the hypothesis that Gram-positive micro-organisms contribute to malodor production by deglycosylating oral glycoproteins, rendering them susceptible to subsequent proteolysis. To this end, we examined the effect of Streptococcus salivarius on Porphyromonas gingivalis-mediated putrefaction of a model glycoprotein (pig gastric mucin). Malodor was scored by two odor judges, and volatile sulfides were determined with the use of a sulfide monitor. Mucin degradation was followed by electrophoresis on SDS-PAGE. Results showed that the addition of S. salivarius or β-galactosidase promoted mucin degradation and concomitant malodor production. Addition of glycosidic inhibitors (p-APTG and glucose) inhibited this process. These results suggest that Gram-positive micro-organisms such as S. salivarius contribute to oral malodor production by deglycosylating salivary glycoproteins, thus exposing their protein core to further degradation by Gram-negative micro-organisms.

We have previously reported that a targeted anti-caries DNA vaccine, pGJA-P, induced accelerated and increased antibody responses compared with a non-targeted anti-caries DNA vaccine. Recently, pGJA-P/VAX, a new targeted anti-caries DNA vaccine for human trials, was constructed by replacing the pCI vector used in the construction of pGJA-P with pVAX1, the only vector authorized by the US Food and Drug Administration in clinical trials. Here, we report on our exploration of the kinetics of the antibody responses generated following pGJA-P/VAX immunization and the persistence of pGJA-P/VAX at both the inoculation site and the draining lymph nodes. Intranasal vaccination of mice with pGJA-P/VAX induced strong antibody responses that lasted for more than 6 months. Furthermore, pGJA-P/VAX could still be detected at both the inoculation site and the draining cervical lymph nodes 6 months after immunization. Thus, the persistent immune responses are likely due to the DNA depot in the host, which acts as a booster immunization.

Although the individual cariostatic effects of laser and fluoride have been shown, the combined effect of CO₂ laser and fluoride on root demineralization remains uncertain and was the main aim of this study. By using a pH-cycling system and Polarized Light Microscopy, we demonstrated the synergistic effect of fluoride combined with CO₂ laser treatment on reducing root demineralization. The mean lesion depths (in μm) for each group were 160 ± 14 (Control), 113 ± 8 (Laser treatment alone), 111 ± 6 (Fluoride treatment alone), and 25 ± 7 (Fluoride followed by laser treatment). A significant laser-enhanced fluoride uptake, characterized by the ToF-SIMS analysis, was revealed by the 37% and 400% increments in loosely and firmly bound fluorides (both p < 0.002) in laser-irradiated areas, compared with the non-irradiated controls. In conclusion, there is a significant synergistic effect of combined CO₂ laser and fluoride treatment on the inhibition of root demineralization, possibly due to laser-enhanced fluoride uptake in the root.

Dental screening of children in schools is undertaken in many countries. There is no evidence that this activity is effective. The objective of our study was to determine if school dental screening of children reduces untreated disease or improves attendance at the population level. A four-arm cluster-randomized controlled trial was undertaken in the northwest of England. In total, 16,864 children aged 6–9 years in 168 schools were randomly allocated to 3 test groups, which received screening according to different models, and a control, which received no intervention. There were no significant differences in caries increment in the primary and secondary dentitions or in the proportions of children attending a dentist after screening between the control group and the 3 intervention arms. School dental screening delivered according to 3 different models was not effective at reducing levels of active caries and increasing attendance in the population under study.

Successful use of atraumatic restorative treatment (ART) in children has been reported, but little information is available regarding its use in older adults. The hypothesis of this study was that survival rates of root restorations placed by both ART and the conventional technique were similar. Root-surface caries lesions in 103 institutionalized elders in Hong Kong were treated randomly by either: (1) the conventional approach—caries removed by dental burs, and the cavity filled with light-cured resin-modified glass ionomer; or (2) the ART approach—caries removed by hand instruments, and the cavity filled with chemically cured high-strength glass ionomer. In total, 84 conventional and 78 ART restorations were placed. After 12 months, 63 conventional and 59 ART restorations were reviewed, and the respective 12-month survival rates were 91.7% and 87.0% (p > 0.05). It is concluded that the survival rates of both types of root restorations were high and similar.

Antimicrobial peptides play an important role in the innate immune response. Deficiency in salivary LL-37 antimicrobial peptide has been implicated in periodontitis in patients with morbus Kostman syndrome. Down syndrome is associated with periodontitis, diminished salivary flow, and salivary immunoglobulin deficiency. In the present study, levels of LL-37 and its hCAP18 precursor were measured in saliva samples from young individuals with Down syndrome and compared with levels in those from age-matched healthy controls. LL-37 and human cathelicidin antimicrobial protein (hCAP18) were detected in whole but not in parotid saliva. hCAP18 was more abundant than LL-37. The concentrations of salivary hCAP18 and LL-37 were found to be higher in individuals with Down syndrome than in healthy controls, but their secretion rates were similar. We concluded that, while the adaptive immunity of individuals with Down syndrome is impaired at the oral mucosa, the secretion rate of the LL-37 component of the innate immune system is normal.

Van der Woude syndrome (VWS) (OMIM 119300) is a dominantly inherited, developmental disorder that is characterized by pits and/or sinuses of the lower lip and a cleft lip and/or cleft palate. Mutations in the interferon regulatory factor 6 gene (IRF6) have been recently identified in patients with VWS, with more than 60 mutations reported. However, the VWS phenotype, IRF6 mutation genotypes, and their interrelationships in Chinese VWS patients have not been studied. Here, we report 11 Chinese families with variable clinical phenotypes of VWS and identified mutations in all patients. Of the 11 mutations, 8 appeared to be novel: CC5.6GT, T342A, 566delA, C748T, C756A, C989A, C1209G, and 1316delT. Seven mutations caused a change or loss of the IRF6 domain. The marked phenotypic variation may be caused by the action of certain modifier genes on IRF6 function. Abbreviations: VWS, Van der Woude syndrome; IRF6, interferon regulatory factor 6; CL/P, cleft lip and/or cleft palate; DBD, DNA-binding domain; SMIR, Smad-interferon regulatory factor-binding domain; Kb, kilobase; PCR, polymerase chain-reaction.

Among functional monomers used in contemporary dental adhesives, 10-methacryloyloxydecyl dihydrogen phosphate (MDP) has been found to interact chemically with hydroxyapatite (HAp) most intensively and stably. This effect was thought to be the basis of the superior bonding effectiveness of MDP-based self-etch adhesives to enamel/dentin. To elucidate fully the chemical interaction and reactivity of MDP with HAp, we used ³¹P CP-MAS NMR spectroscopy and powder x-ray diffraction. In an aqueous ethanol solution, Ca ions were leached from HAp to form, at short term, a MDP-calcium salt (CaMHP₂) layered structure on the HAp surface. When MDP was allowed to interact for longer time (< 24 hrs), CaHPO₄·2H₂O precipitated on top of this MDP-calcium salt layered structure. In conclusion, the intense chemical interaction of MDP with HAp must be ascribed to superficial dissolution of HAp induced by the MDP adsorption and subsequent deposition of MDP-calcium salt with a solubility lower than that of CaHPO₄·2H₂O.

During polymerization, dental composites develop residual stresses that may compromise the marginal integrity and properties of the restorative. The objective of this study was to use the thin-walled ring-slitting method to measure and compare residual stresses. The hypotheses to be tested were that composites would generate different levels of residual stress based on their specific formulations and slitting times. Rings made from composites (Z100, Herculite, and Heliomolar) were cut at different times (10 min, 1 and 24 hrs) after being light-cured, and stress was measured. Residual stress was higher at the earlier cutting times, except for Heliomolar (α < 0.05). For the 10-minute and one-hour cutting groups, stress followed this order: Z100 > Herculite > Heliomolar. Early slitting was better to capture residual stress, and the thin-walled rings showed higher values than thick-walled rings and were better able to discriminate residual stress in composites.

Chlorhexidine salts are available in various formulations for dental applications. This study tested the hypothesis that the release of chlorhexidine from a urethane dimethacrylate and triethylene glycol dimethacrylate resin system can be effectively controlled by the chlorhexidine diacetate content and pH. The filler concentrations were 9.1, 23.1, or 33.3 wt%, and the filled resins were exposed to pH 4 and pH 6 acetate buffers. The results showed that Fickian diffusion was the dominant release mechanism. The rates of release were significantly higher in pH 4 buffer, which was attributed to the increase of chlorhexidine diacetate solubility at lower pH. The higher level of filler loading reduced the degree of polymerization, leading to a greater loss of organic components and higher chlorhexidine release rates.

Light propagation in human dentin exhibits a strong directional dependence featuring the long-known optical magnification property. We hypothesized that this anisotropic effect is caused by multiple scattering at the dentin tubules, and not by fiberoptic effects, as had been previously assumed. We performed measurements of the transmitted intensity from dentin disks and compared them with Monte Carlo simulations of light propagation in dentin, considering the scattering by the tissue’s microstructure. We found that the optical anisotropy of dentin can be fully explained with this model. We concluded that the magnification property of dentin is due to multiple scattering by the dental microstructure.