Journal of Dental Research recent issues

Pre-clinical Models for Oral and Periodontal Reconstructive Therapies

Pellegrini, G., Seol, Y.J., Gruber, R., Giannobile, W.V. — Fri, 20 Nov 2009 15:47:48 PST

The development of new medical formulations (NMF) for reconstructive therapies has considerably improved the available treatment options for individuals requiring periodontal repair or oral implant rehabilitation. Progress in tissue engineering and regenerative medicine modalities strongly depends on validated pre-clinical research. Pre-clinical testing has contributed to the recent approval of NMF such as GEM 21S^® and INFUSE^® bone grafts for periodontal and oral regenerative therapies. However, the selection of a suitable pre-clinical model for evaluation of the safety and efficacy of a NMF remains a challenge. This review is designed to serve as a primer to choose the appropriate pre-clinical models for the evaluation of NMF in situations requiring periodontal or oral reconstruction. Here, we summarize commonly used pre-clinical models and provide examples of screening and functional studies of NMF that can be translated into clinical use.

Engineering Craniofacial Structures: Facing the Challenge

Zaky, S.H., Cancedda, R. — Fri, 20 Nov 2009 15:47:49 PST

The human innate regenerative ability is known to be limited by the intensity of the insult together with the availability of progenitor cells, which may cause certain irreparable damage. It is only recently that the paradigm of tissue engineering found its way to the treatment of irreversibly affected body structures with the challenge of reconstructing the lost part. In the current review, we underline recent trials that target engineering of human craniofacial structures, mainly bone, cartilage, and teeth. We analyze the applied engineering strategies relative to the selection of cell types to lay down a specific targeted tissue, together with their association with an escorting scaffold for a particular engineered site, and discuss their necessity to be sustained by growth factors. Challenges and expectations for facial skeletal engineering are discussed in the context of future treatment.

Walter Loesche--a Maverick in Translational Research in Dentistry

Banas, J.A. — Fri, 20 Nov 2009 15:47:49 PST

Effects of Chemical Cross-linkers on Caries-affected Dentin Bonding

Macedo, G.V., Yamauchi, M., Bedran-Russo, A.K. — Fri, 20 Nov 2009 15:47:49 PST

The achievement of a strong and stable bond between composite resin and dentin remains a challenge in restorative dentistry. Over the past two decades, dental materials have been substantially improved, with better handling and bonding characteristics. However, little attention has been paid to the contribution of collagen structure/stability to bond strength. We hypothesized that the induction of cross-linking in dentin collagen improves dentin collagen stability and bond strength. This study investigated the effects of glutaraldehyde-and grape seed extract-induced cross-linking on the dentin bond strengths of sound and caries-affected dentin, and on the stability of dentin collagen. Our results demonstrated that the application of chemical cross-linking agents to etched dentin prior to bonding procedures significantly enhanced the dentin bond strengths of caries-affected and sound dentin. Glutaraldehyde and grape seed extract significantly increased dentin collagen stability in sound and caries-affected dentin, likely via distinct mechanisms.

Inhibition of Enzymatic Degradation of Adhesive-Dentin Interfaces

Fri, 20 Nov 2009 15:47:49 PST

Adhesive procedures activate dentin-associated matrix metalloproteinases (MMPs), and so iatrogenically initiate bond degradation. We hypothesized that adding MMP inhibitors to adhesive primers may prevent this endogenous enzymatic degradation, thereby improving bond durability. A non-specific MMP inhibitor (chlorhexidine) and a MMP-2/9-specific inhibitor (SB-3CT) were admixed to the primers of an etch & rinse and a self-etch adhesive, both considered as gold-standard adhesives within their respective categories. For dentin powder exposed to the adhesives under clinical application conditions, gelatin zymography revealed the release of MMP-2 (not of MMP-9) by the etch & rinse adhesive, while no release of enzymes could be detected for the mild self-etch adhesive, most likely because of its limited dentin demineralization effect. The built-in MMP inhibitors appeared effective in reducing bond degradation only for the etch & rinse adhesive, and not for the self-etch adhesive. Water sorption of adhesive interfaces most likely remains the principal mechanism of bond degradation, while endogenous enzymes appear to contribute to bond degradation of only etch & rinse adhesives.

Appositional Bone Formation by OCP-Collagen Composite

Fri, 20 Nov 2009 15:47:49 PST

Synthetic octacalcium phosphate (OCP) has been shown to enhance bone formation and to biodegrade if implanted into bone defects. Here, we hypothesized that an OCP-atelocollagen complex (OCP/Col) is biodegradable and can induce bone formation in a thickness-dependent manner when implanted into the calvaria. OCP/Col disks (diameter, 9 mm; thickness, 1 or 3 mm) were implanted into a subperiosteal pocket in the calvaria of 12-week-old Wistar rats for 4, 8, and 12 weeks and subsequent bone formation was monitored. X-ray diffraction analysis and Fourier transform infrared spectroscopy showed that OCP in the OCP/Col implants was converted into a carbonate-rich apatite after 4 weeks. Although thinner disks tended to be replaced by new bone, thicker disks were progressively resorbed by osteoclast-like cells until 12 weeks, possibly via the increased mechanical load in the subperiosteal pocket. Therefore, OCP/Col can increase appositional intra-membranous bone formation if the appropriate size of the implant is applied.

Pharmacological Retention of Oral Mucosa Progenitor/Stem Cells

Izumi, K., Inoki, K., Fujimori, Y., Marcelo, C.L., Feinberg, S.E. — Fri, 20 Nov 2009 15:47:49 PST

Oral mucosa progenitor/stem cells reside as a small-sized cell population that eventually differentiates concurrently with an increase in cell size. Activation of the mammalian target of rapamycin (mTOR) leads to an increase in cell size. We hypothesized that rapamycin, a specific inhibitor of mTOR, will maintain primary human oral keratinocytes as a small-sized, undifferentiated cell population capable of retaining their proliferative capacity. Primary, rapamycin-treated (2 nM, 20 nM) oral keratinocytes showed a diminished cell size that correlated with a higher clonogenicity, a longer-term proliferative potential, and a slower cycling cell population concurrent with decreased expression of a differentiation marker when compared with untreated cells. Only the 2-nM rapamycin-treated oral keratinocytes maintained their ability to regenerate oral mucosa in vitro after 15 weeks of culture. Rapamycin, a Food and Drug Administration-approved drug, may have applicability for use in creating a highly proliferative cell population for use in regenerative medicine.

PDK1 Regulates Chemotaxis in Human Neutrophils

Fri, 20 Nov 2009 15:47:49 PST

Phosphoinositide-dependent kinase (PDK1) plays a central role in signal transduction mediated by phosphatidylinositol 3-kinases (PI3K) and regulates cellular functions in neutrophils. Neutrophils from individuals diagnosed with localized aggressive periodontitis (LAP) present an in vivo phenotype with depressed chemotaxis. The aim of this study was to test the hypothesis that PDK1 regulates chemotaxis in neutrophils and is responsible for the abnormal neutrophil chemotaxis LAP. Neutrophil chemotaxis was significantly suppressed by the PDK1 inhibitor staurosporine. When cells were transfected with PDK1 siRNA, there was a significant reduction in chemotaxis, while superoxide generation was not significantly affected. In primary neutrophils from persons with LAP, PDK1 expression and activation levels were significantly reduced, and this reduction was associated with the reduced phosphorylation of Akt (Thr308) and chemotaxis. Analysis of these data demonstrates that PDK1 is essential for the chemotactic migration of neutrophils, and in the absence of PDK1, neutrophil chemotaxis is impaired.

MAP Kinase Phosphatase-1 Protects against Inflammatory Bone Loss

Sartori, R., Li, F., Kirkwood, K.L. — Fri, 20 Nov 2009 15:47:49 PST

The mitogen-activated protein (MAP) kinase phosphatase (MKP) family plays an important function in regulating the pro-inflammatory cytokines by deactivating MAP kinases. MKP-1 is essential for the dephosphorylation of p38 MAP kinase that regulates expression of IL-6, TNF-, and IL-1β. We hypothesized that MKP-1 regulates inflammatory bone loss in experimental periodontitis. Wild-type and Mkp-1^–/– mice received A. actinomycetemcomitans LPS injection in the palatal region or PBS control 3 times/wk for 30 days. Mice were killed, and maxillae were assessed by microcomputed tomography, histological analysis, and TRAP staining for measurement of bone loss, extent of inflammation, and degree of osteoclastogenesis. Results indicated that, in LPS-injected Mkp-1^–/– mice, significantly greater bone loss occurred with more inflammatory infiltrate and a significant increase in osteoclastogenesis compared with Mkp-1^–/– control sites or either wild-type group. Analysis of these data indicates that MKP-1 plays a key role in the regulation of inflammatory bone loss.

Cyclosporin A and Phenytoin Modulate Inflammatory Responses

Suzuki, A.M.M., Yoshimura, A., Ozaki, Y., Kaneko, T., Hara, Y. — Fri, 20 Nov 2009 15:47:49 PST

Gingival overgrowth is a common side-effect of administration of the immunosuppressant cyclosporin A and the anti-epileptic drug phenytoin. While cyclosporin-induced gingival overgrowth is often accompanied by gingival inflammation, phenytoin-induced gingival overgrowth usually forms fibrotic lesions. To determine whether these drugs alter the inflammatory responses of gingival fibroblasts, we investigated the effects of cyclosporin and phenytoin on Toll-like receptor (TLR)-mediated responses to microbial components. In Chinese hamster ovary reporter cell lines, cyclosporin alone triggered signaling, whereas phenytoin down-regulated signaling induced by the TLR2 or TLR4 ligand. In human gingival fibroblasts, cyclosporin alone did not induce evident inflammatory responses, but augmented the expression of CD54 and the production of interleukin (IL)-6 and IL-8 induced by TLR ligands, whereas phenytoin attenuated those responses. Cyclosporin also augmented CD54 expression in gingiva of mice injected with lipopolysaccharide. These results indicated that cyclosporin positively and phenytoin negatively modulated inflammatory responses of human gingival fibroblasts.

Genetic Risk Factors for Periodontitis in a Japanese Population

Fri, 20 Nov 2009 15:47:49 PST

Genetic variants at multiple loci have been shown to be associated with susceptibility to periodontitis. To better assess the genetic risk factors for periodontitis, we performed a case-control study in 319 Japanese individuals with periodontitis (172 aggressive and 147 chronic disease) and 303 race-matched healthy control individuals. Thirty-five functional gene polymorphisms that had been previously associated with immune responses were genotyped. For all gene polymorphisms tested, no significant differences were observed in the allele frequencies of persons with aggressive, chronic, and combined (aggressive and chronic) periodontitis, compared with control individuals. Multiple logistic regression analysis revealed a significant association of the vitamin D receptor +1056 T/C polymorphism with susceptibility to chronic periodontitis, after adjustment for age, gender, and smoking status (P = 0.002). These results suggest that none of the polymorphisms tested was strongly associated with periodontitis in a Japanese population. However, the vitamin D receptor +1056 polymorphism may be related to chronic periodontitis.

Immunomodulatory Activity of IL-27 in Human Periapical Lesions

Fri, 20 Nov 2009 15:47:49 PST

IL-27, a cytokine with pro-inflammatory and anti-inflammatory properties, is a new member of the IL-6/IL-12 family, whose function in periapical lesions is unknown. We hypothesized that the production of IL-27 and its effect depend upon the type of immune/inflammatory response and clinical presentation of periapical lesions. We tested this hypothesis by studying the expression and function of IL-27 in human periapical lesions, both in situ and in culture. Immunohistochemistry demonstrated the strongest expression of IL-27 by endothelial cells and mononuclear phagocytes. Its production by periapical lesion mononuclear cells (PL-MNC), especially in symptomatic lesions, was significantly higher compared with that in peripheral blood MNC and correlated with the frequency of CD14⁺ and CD3⁺ cells. Exogenous IL-27 stimulated Th1 and down-regulated Th17 cytokine production by PL-MNC from symptomatic lesions, but down-regulated Th1 and Th2 responses in asymptomatic lesions. These findings suggest that IL-27 is an immunomodulatory cytokine in periapical lesions, with complex biological effects.

Regression of Post-orthodontic Lesions by a Remineralizing Cream

Fri, 20 Nov 2009 15:47:49 PST

Orthodontic patients have an increased risk of white-spot lesion formation. A clinical trial was conducted to test whether, in a post-orthodontic population using fluoride toothpastes and receiving supervised fluoride mouthrinses, more lesions would regress in participants using a remineralizing cream containing casein phosphopeptide- amorphous calcium phosphate compared with a placebo. Forty-five participants (aged 12–18 yrs) with 408 white-spot lesions were recruited, with 23 participants randomized to the remineralizing cream and 22 to the placebo. Product was applied twice daily after fluoride toothpaste use for 12 weeks. Clinical assessments were performed according to ICDAS II criteria. Transitions between examinations were coded as progressing, regressing, or stable. Ninety-two percent of lesions were assessed as code 2 or 3. For these lesions, 31% more had regressed with the remineralizing cream than with the placebo (OR = 2.3, P = 0.04) at 12 weeks. Significantly more post-orthodontic white-spot lesions regressed with the remineralizing cream compared with a placebo over 12 weeks.

Diversity of Endodontic Microbiota Revisited

Siqueira, J.F., Rocas, I.N. — Wed, 14 Oct 2009 16:10:11 PDT

Although fungi, archaea, and viruses contribute to the microbial diversity in endodontic infections, bacteria are the most common micro-organisms occurring in these infections. Datasets from culture and molecular studies, integrated here for the first time, showed that over 460 unique bacterial taxa belonging to 100 genera and 9 phyla have been identified in different types of endodontic infections. The phyla with the highest species richness were Firmicutes, Bacteroidetes, Actinobacteria, and Proteobacteria. Diversity varies significantly according to the type of infection. Overall, more taxa have been disclosed by molecular studies than by culture. Many cultivable and as-yet-uncultivated phylotypes have emerged as candidate pathogens based on detection in several studies and/or high prevalence. Now that a comprehensive inventory of the endodontic microbial taxa has been established, future research should focus on the association with different disease conditions, functional roles in the community, and susceptibility to antimicrobial treatment procedures.

Bacterial Interactions in Dental Biofilm Development

Hojo, K., Nagaoka, S., Ohshima, T., Maeda, N. — Wed, 14 Oct 2009 16:10:11 PDT

Recent analyses with ribosomal RNA-based technologies have revealed the diversity of bacterial populations within dental biofilms, and have highlighted their important contributions to oral health and disease. Dental biofilms are exceedingly complex and multispecies ecosystems, where oral bacteria interact cooperatively or competitively with other members. Bacterial interactions that influence dental biofilm communities include various different mechanisms. During the early stage of biofilm formation, it is known that planktonic bacterial cells directly attach to surfaces of the oral cavity or indirectly bind to other bacterial cells that have already colonized. Adherence through co-aggregation may be critical for the temporary retention of bacteria on dental surfaces, and may facilitate eventual bacterial colonization. It is likely that metabolic communication, genetic exchange, production of inhibitory factors (e.g., bacteriocins, hydrogen peroxide, etc.), and quorum-sensing are pivotal regulatory factors that determine the bacterial composition and/or metabolism. Since each bacterium can easily access a neighboring bacterial cell and its metabolites, genetic exchanges and metabolic communication may occur frequently in dental biofilms. Quorum-sensing is defined as gene regulation in response to cell density, which influences various functions, e.g., virulence and bacteriocin production. In this review, we discuss these important interactions among oral bacteria within the dental biofilm communities.

Fam83h is Associated with Intracellular Vesicles and ADHCAI

Wed, 14 Oct 2009 16:10:11 PDT

Defects in FAM83H on human chromosome 8q24.3 cause autosomal-dominant hypocalcified amelogenesis imperfecta (ADHCAI). FAM83H does not encode a recognizable signal peptide, so we predicted that the Fam83h protein functions within the cell. We tested this hypothesis by constitutively expressing mouse Fam83h with green fluorescent protein (GFP) fused to its C-terminus in HEK293 and HeLa cell lines. Green fluorescent signal from the Fam83h-GFP fusion protein was associated with perinuclear vesicles, usually in the vicinity of the Golgi apparatus. No signal was observed within the nucleus. In addition, we identified FAM83H nonsense mutations in Hispanic (C1330C>T; p.Q444X) and Caucasian (c.1192C>T; p.Q398X) families with ADHCAI. We conclude that Fam83h localizes in the intracellular environment, is associated with vesicles, and plays an important role in dental enamel formation. FAM83H is the first gene involved in the etiology of amelogenesis imperfecta (AI) that does not encode a secreted protein.

Regulatory T-cells in Periapical Lesions

Wed, 14 Oct 2009 16:10:11 PDT

CD4⁺CD25^hiFoxp3⁺ regulatory T-cells (Tregs) are of crucial importance in regulating the immune response, including the control of any defense against infection. Their presence in periapical lesions has not been demonstrated, as yet. We hypothesized that Tregs infiltrate periapical lesions, where they inhibit T-cell proliferation. The aim of this study was to characterize Tregs in periapical lesions by confocal microscopy, flow cytometry, and functional assays. We showed that CD4⁺CD25^hiFoxp3⁺ cells in periapical lesions expressed IL-10 and TGF-β. Their frequency was significantly higher than in peripheral blood and correlated with the levels of TGF-β and IL-10 in culture supernatants of periapical lesion mononuclear cells. Tregs inhibited the proliferation of responder T-cells in vitro, at least in part, by stimulating the production of IL-10. These findings suggest that CD4⁺CD25^hiFoxp3⁺ cells in periapical lesions may play regulatory roles in controlling local immune/inflammatory processes.

Osterix Enhances BMSC-associated Osseointegration of Implants

Wed, 14 Oct 2009 16:10:12 PDT

Cellular and molecular events in osseointegration at the dental implant surface remain largely unknown. We hypothesized that bone marrow stromal cells (BMSCs) participate in this process, and that osterix (Osx) promotes implant osseointegration. To prove this hypothesis, we tracked double-labeled BMSCs in implantation sites created in nude mice transplanted with these cells. We also inserted implants into the femurs of our established transgenic mice after local administration of viruses encoding Osx, to determine the osteogenic effects of Osx. Immunohistochemical results demonstrated that BMSCs can recruit from peripheral circulation and participate in wound healing and osseointegration after implantation. Microcomputed tomography (microCT) analysis revealed an increased bone density at the bone-to-implant interface in the Osx group, and histomorphometric analysis indicated an elevated level of bone-to-implant contact in the Osx group. We concluded that exogenous BMSCs participate in the osseointegration after implantation, and that Osx overexpression accelerates osseointegration.

Periodontal Gene Transfer by Ultrasound and Nano/Microbubbles

Chen, R., Chiba, M., Mori, S., Fukumoto, M., Kodama, T. — Wed, 14 Oct 2009 16:10:12 PDT

A non-viral gene delivery approach with nano/microbubbles and ultrasound offers opportunities for targeting soft tissues for gene therapy. The periodontium is a complex structure comprised of hard (cementum, alveolar bone) and soft tissues (periodontal ligament, gingivae). We hypothesized that our established gene delivery method would allow the periodontal tissue to be targeted for transfection for gene therapy. Expression kinetics and sites of transfection sites with this approach were investigated in rat periodontal tissue. Bioluminescence imaging revealed that transient gene expression was induced at day 1 posttransfection, while confocal microscopy showed that gene expression was localized in the muscle cells of gingival tissues. These findings indicate that regular transfection with this approach results in high gene expression, facilitating gene therapy for periodontal disease involving alveolar bone resorption.

Odontoblast TRP Channels and Thermo/Mechanical Transmission

Son, A.R., Yang, Y.M., Hong, J.H., Lee, S.I., Shibukawa, Y., Shin, D.M. — Wed, 14 Oct 2009 16:10:12 PDT

Odontoblasts function as mechanosensory receptors because of the expression of mechanosensitive channels in these cells. However, it is unclear if odontoblasts direct the signal transmission evoked by heat/cold or osmotic changes. This study investigated the effects of heat/cold or osmotic changes on calcium signaling and the functional expression of the thermo/mechanosensitive transient receptor potential (TRP) channels in primary cultured mouse odontoblastic cells, with the use of RT-PCR, fluorometric calcium imaging, and electrophysiology. TRPV1, TRPV2, TRPV3, TRPV4, and TRPM3 mRNA was expressed, but TRPM8 and TRPA1 mRNA was not. The receptor-specific stimulation of TRPV1-3 (heat-sensing receptors) and TRPV4/ TRPM3 (mechanic receptors) caused increases in the intracellular calcium concentration. Moreover, the channel activities of TRPV1-4 and TRPM3 were confirmed by a whole-cell patch-clamp technique. These results suggest that primary cultured mouse odontoblasts express heat/mechanosensitive TRP channels and play a role in the underlying mechanisms of thermo/mechanosensitive sensory transmission.

Mineralized Tissue Formation by BMP2-transfected Pulp Stem Cells

Yang, X., van der Kraan, P.M., Bian, Z., Fan, M., Walboomers, X.F., Jansen, J.A. — Wed, 14 Oct 2009 16:10:12 PDT

Previously, in vitro differentiation of odontoblasts was shown for dental pulp stem cells (DPSCs) transfected with bone morphogenetic protein-2 (Bmp2). For this study, we hypothesized that such cells also show potential for mineralized tissue formation in vivo. DPSCs were transfected with Bmp2 and seeded onto a ceramic scaffold. These complexes were cultured in medium without dexamethasone, and thereafter placed subcutaneously in nude mice for 1, 4, and 12 weeks. Samples were evaluated by histology and real-time PCR for osteocalcin, bone sialoprotein, dentin sialophosphoprotein, and dentin matrix protein 1. Results indicated that only the transfected DPSCs showed obvious mineralized tissue generation, and 12 weeks of implantation gave the highest percentage of mineralized tissue formation (33 ± 7.3% of implant pore area). Real-time PCR confirmed these results. In conclusion, Bmp2-transfected DPSCs effectively show mineralized tissue formation upon ectopic implantation.

Microbial Interactions Influence Inflammatory Host Cell Responses

Wed, 14 Oct 2009 16:10:12 PDT

The inflammatory response plays an important role in the tissue destruction associated with periodontitis. Bacterial species can regulate the inflammatory responses of host cells, triggered by pathogens. It was hypothesized that, in the field of oral microbiology/immunology, such effects of bacterial interactions on inflammatory host cell responses might also be present. In this study, the effects of beneficial, commensal, and pathogenic species on Aggregatibacter actinomycetemcomitans-induced interleukin-8 (IL-8) production by human cells were investigated. The beneficial species, Streptococcus mitis, Streptococcus salivarius, and Streptococcus sanguinis, were able to lower the IL-8 production triggered by A. actinomycetemcomitans. The inhibitory effect was also achieved by the application of streptococcal supernatants. In contrast, the commensal Streptococcus gordonii caused no reduction, and the pathogen Fusobacterium nucleatum increased IL-8 production by the host cells. These results show that bacterial species can influence the inflammatory responses of host cells triggered by infection with A. actinomycetemcomitans.

Involvement of Cytosolic Phospholipase A2{alpha} in MMP-9 Up-regulation

Montreekachon, P., Chotjumlong, P., Reutrakul, V., Krisanaprakornkit, S. — Wed, 14 Oct 2009 16:10:12 PDT

Matrix metalloproteinase-9 (MMP-9) is important in the pathogenesis of periodontitis. Cytosolic phospholipase A₂ (cPLA₂) is involved in MMP-9 up-regulation in human monocytes. We tested the hypothesis that cPLA₂ also regulates MMP-9 induction by Fusobacterium nucleatum and by phorbol 12-myristate-13-acetate (PMA) in gingival epithelial cells. While PMA induced MMP-9 expression considerably, F. nucleatum did so moderately. This time-course study demonstrated that MMP-9 mRNA up-regulation occurred at 3 hours, whereas MMP-9 secretion and activity in cell-free supernatants occurred at 12 hours. cPLA₂ mRNA was constitutively expressed in gingival epithelial cells. Transient activation of cPLA₂ by Ser505 phosphorylation was observed in the nuclei upon stimulation, suggesting its role as a transcription factor, while cPLA₂ protein expression remained unchanged. Induction of MMP-9 expression and activity was significantly inhibited by 1 µM of the specific cPLA₂ inhibitor (P < 0.01). These findings demonstrate the involvement of cPLA₂ in MMP-9 up-regulation.

CCR5 Down-regulates Osteoclast Function in Orthodontic Tooth Movement

Wed, 14 Oct 2009 16:10:12 PDT

During orthodontic tooth movement, there is local production of chemokines and an influx of leukocytes into the periodontium. CCL5 plays an important role in osteoclast recruitment and activation. This study aimed to investigate whether the CCR5-receptor influences these events and, consequently, orthodontic tooth movement. An orthodontic appliance was placed in wild-type mice (WT) and CCR5-deficient mice (CCR5^–/–). The expression of mediators involved in bone remodeling was evaluated in periodontal tissues by Real-time PCR. The number of TRAP-positive osteoclasts and the expression of cathepsin K, RANKL, and MMP13 were significantly higher in CCR5^–/–. Meanwhile, the expression of two osteoblastic differentiation markers, RUNX2 and osteocalcin, and that of bone resorption regulators, IL-10 and OPG, were lower in CCR5^–/–. Analysis of the data also showed that CCR5^–/– exhibited a greater amount of tooth movement after 7 days of mechanical loading. The results suggested that CCR5 might be a down-regulator of alveolar bone resorption during orthodontic movement.

Diclofenac Sodium Inhibits NF{kappa}B Transcription in Osteoclasts

Wed, 14 Oct 2009 16:10:12 PDT

A non-steroidal anti-inflammatory drug, diclofenac, acts efficiently against inflammation; however, down-regulation of diclofenac on bone remodeling has raised concerns. The inhibitory mechanisms of diclofenac are poorly understood. We hypothesized that diclofenac down-regulates osteoclast differentiation and activation via inhibition of the translocation of phosphorylated nuclear factor kappa B (NFB). When osteoclasts prepared from mouse hematopoietic stem cells were treated with diclofenac, tartrateresistant acid phosphatase-positive multinucleated cells decreased in a concentration-dependent manner. Pit formation assay revealed the abolition of osteoclastic bone resorption; levels of cathepsin K transcripts, an osteoclastic resorption marker, were down-regulated time-dependently. Diclofenac induced the accumulation of the inhibitor of kappa B in cytosol, which led to suppression of the nuclear translocation of NFB and phosphorylated NFB. These results suggest that the novel mechanism of diclofenac for bone remodeling is associated with phosphorylated NFB reduction, which regulates osteoclast differentiation and activation.

Regulation of Trigeminal Respiratory Motor Activity in the Brainstem

Wed, 14 Oct 2009 16:10:12 PDT

The trigeminal motor system participates in the control of respiration as well as suckling and mastication. However, the central mechanism underlying respiratory activity in trigeminal motoneurons is not well-understood. In this study, we aimed to elucidate brainstem circuitry for rhythm generation and signal transmission of trigeminal respiratory activity in in vitro neonatal rat brainstem-spinal cord preparations. We further examined the role of midline-crossing trigeminal interneurons in the bilateral synchronization of respiratory and suckling activity in trigeminal motor nerves. The results of brainstem-sectioning experiments indicated that respiratory rhythms were generated in the medulla and ipsilaterally transmitted to trigeminal motoneurons in the pons. We conclude that the trigeminal motor system, as well as the hypoglossal and phrenic motor system, is regulated by medullary respiratory networks, and that pontine interactions between bilateral trigeminal interneurons are not critical for the generation or synchronization of trigeminal respiratory activity, but are crucial for trigeminal suckling activity.

Influence of Genetic Background on Fluoride Metabolism in Mice

Wed, 14 Oct 2009 16:10:12 PDT

A/J and 129P3/J mouse strains have different susceptibilities to dental fluorosis, due to their genetic backgrounds. This study tested whether these differences are due to variations in water intake and/or F metabolism. A/J (susceptible to dental fluorosis) and 129P3/J mice (resistant) received drinking water containing 0, 10, or 50 ppm F. Weekly F intake, excretion and retention, and terminal plasma and femur F levels were determined. Dental fluorosis was evaluated clinically and by quantitative fluorescence (QF). Data were tested by two-way ANOVA. Although F intakes by the strains were similar, excretion by A/J mice was significantly higher due to greater urinary F excretion, which resulted in lower plasma and femur F levels. Compared with 129P3/J mice given 50 ppm F, significantly higher QF scores were recorded for A/J mice. In conclusion, these strains differ with respect to several features of F metabolism, and amelogenesis in the 129P3/J strain seems to be unaffected by high F exposure.

ERRATUM

Wed, 14 Oct 2009 16:10:12 PDT

Hutchinson-Gilford Progeria Syndrome: Its Presentation in F. Scott Fitzgerald's Short Story 'The Curious Case of Benjamin Button' and Its Oral Manifestations

Maloney, W.J. — Fri, 25 Sep 2009 15:02:33 PDT

Hutchinson-Gilford Progeria Syndrome (HGPS) was first documented in the medical literature in 1886. A HGPS patient has the physical characteristics and appearances of an elderly individual. In 1921, F. Scott Fitzgerald published a short story entitled ‘The Curious Case of Benjamin Button’. The main character of Fitzgerald’s fictional work is born with a very rare condition in which he looks like an elderly person. The main difference between the fictional individual and individuals with HGPS is that Fitzgerald’s character becomes younger as the years go by.

This paper serves three purposes. The first purpose is to scientifically present the possibility that Fitzgerald consciously based his character, Benjamin Button, upon individuals with HGPS. The second purpose is to describe the rare condition of HGPS, along with its many manifestations in the head and neck region. The third purpose is to postulate that HGPS individuals might not only have the appearance of an aged person, but also might actually undergo true physical aging, which would enable researchers to gain valuable information into the treatment of ailments commonly associated with the natural process of aging.

The Impact of Fluoride on Ameloblasts and the Mechanisms of Enamel Fluorosis

Bronckers, A.L.J.J., Lyaruu, D.M., DenBesten, P.K. — Fri, 25 Sep 2009 15:02:33 PDT

Intake of excess amounts of fluoride during tooth development cause enamel fluorosis, a developmental disturbance that makes enamel more porous. In mild fluorosis, there are white opaque striations across the enamel surface, whereas in more severe cases, the porous regions increase in size, with enamel pitting, and secondary discoloration of the enamel surface. The effects of fluoride on enamel formation suggest that fluoride affects the enamel-forming cells, the ameloblasts. Studies investigating the effects of fluoride on ameloblasts and the mechanisms of fluorosis are based on in vitro cultures as well as animal models. The use of these model systems requires a biologically relevant fluoride dose, and must be carefully interpreted in relation to human tooth formation. Based on these studies, we propose that fluoride can directly affect the ameloblasts, particularly at high fluoride levels, while at lower fluoride levels, the ameloblasts may respond to local effects of fluoride on the mineralizing matrix. A new working model is presented, focused on the assumption that fluoride increases the rate of mineral formation, resulting in a greater release of protons into the forming enamel matrix.

Sensitivity of Salivary Glands to Radiation: from Animal Models to Therapies

Grundmann, O., Mitchell, G.C., Limesand, K.H. — Fri, 25 Sep 2009 15:02:33 PDT

Radiation therapy for head and neck cancer causes significant secondary side-effects in normal salivary glands, resulting in diminished quality of life for these individuals. Salivary glands are exquisitely sensitive to radiation and display acute and chronic responses to radiotherapy. This review will discuss clinical implications of radiosensitivity in normal salivary glands, compare animal models used to investigate radiation-induced salivary gland damage, address therapeutic advances, and project future directions in the field.

Runx2, Osx, and Dspp in Tooth Development

Fri, 25 Sep 2009 15:02:33 PDT

The transcription factors Runx2 and Osx are necessary for osteoblast and odontoblast differentiation, while Dspp is important for odontoblast differentiation. The relationship among Runx2, Osx, and Dspp during tooth and craniofacial bone development remains unknown. In this study, we hypothesized that the roles of Runx2 and Osx in the regulation of osteoblast and odontoblast lineages may be independent of one another. The results showed that Runx2 expression overlapped with Osx in dental and osteogenic mesenchyme from E12 to E16. At the later stages, from E18 to PN14, Runx2 and Osx expressions remained intense in alveolar bone osteoblasts. However, Runx2 expression was down-regulated, whereas Osx expression was clearly seen in odontoblasts. At later stages, Dspp transcription was weakly present in osteoblasts, but strong in odontoblasts where Osx was highly expressed. In mouse odontoblast-like cells, Osx overexpression increased Dspp transcription. Analysis of these data suggests differential biological functions of Runx2, Osx, and Dspp during odontogenesis and osteogenesis.

Primary Cilia of Odontoblasts: Possible Role in Molar Morphogenesis

Fri, 25 Sep 2009 15:02:33 PDT

A primary cilium, a sensory organelle present in almost every vertebrate cell, is regularly described in odontoblasts, projecting from the surfaces of the cells. Based on the hypothesis that the primary cilium is crucial both for dentin formation and possibly in tooth pain transmission, we have investigated the expression and localization of the main cilium components and involvement of the OFD1 gene in tooth morphogenesis. Odontoblasts in vitro express tubulin, inversin, rootletin, OFD1, BBS4, BBS6, ALMS1, KIF3A, PC1, and PC2. In vivo, cilia are aligned parallel to the dentin walls, with the top part oriented toward the pulp core. Close relationships between cilium and nerve fibers are evidenced. Calcium channels are concentrated in the vicinity of the basal body. Analysis of these data suggests a putative role of cilia in sensing the microenvironment, probably related to dentin secretion. This hypothesis is enhanced by the huge defects observed on molars from Ofd1 knockout mice, showing undifferentiated dentin-forming cells.

Increased Oral Fibroblast Lifespan Is Telomerase-independent

Fri, 25 Sep 2009 15:02:33 PDT

Oral mucosal wound-healing is characterized by rapid re-epithelialization and remodeling, with minimal scar formation. This may be attributed to the distinct phenotypic characteristics of the resident fibroblasts. To test this hypothesis, we investigated patient-matched oral mucosal and skin fibroblasts. Compared with skin fibroblasts, oral mucosal fibroblasts had longer proliferative lifespans, underwent more population doublings, and experienced senescence later, which was directly related to longer telomere lengths within oral mucosal fibroblasts. The presence of these longer telomeres was independent of telomerase expression, since both oral oral mucosal fibroblasts and skin fibroblasts were negative for active telomerase, as assessed according to the Telomeric Repeat Amplification Protocol. This study has demonstrated that, compared with skin fibroblasts, oral mucosal fibroblasts are ‘younger’, with a more embryonic/fetal-like phenotype that may provide a notable advantage for their ability to repair wounds in a scarless fashion.

Impact of Hydrodynamics on Oral Biofilm Strength

Paramonova, E., Kalmykowa, O.J., van der Mei, H.C., Busscher, H.J., Sharma, P.K. — Fri, 25 Sep 2009 15:02:33 PDT

Mechanical removal of oral biofilms is ubiquitously accepted as the best way to prevent caries and periodontal diseases. Removal effectiveness strongly depends on biofilm strength. To investigate the influence of hydrodynamics on oral biofilm strength, we grew single- and multi-species biofilms of Streptococcus oralis J22, Actinomyces naeslundii TV14-J1, and full dental plaque at shear rates ranging from 0.1 to 50 1/sec and measured their compressive strength. Subsequently, biofilm architecture was evaluated by confocal laser scanning microscopy. Multi-species biofilms were stronger than single-species biofilms, with strength values ranging from 6 to 51 Pa and from 5 to 17 Pa, respectively. In response to increased hydrodynamic shear, biofilm strength decreased, and architecture changed from uniform carpet-like to more "fluffy" with higher thickness. S. oralis biofilms grown under variable shear of 7 and 50 1/sec possessed properties intermediate of those measured at the respective single shears.

Role of Purinergic Receptor in Alpha Fodrin Degradation in Par C5 Cells

Fri, 25 Sep 2009 15:02:33 PDT

Autoantibodies specific for alpha-fodrin fragments are found in the tissues of persons afflicted with Sjögren’s syndrome (SS). However, the mechanism for alpha-fodrin degradation remains elusive. The following experiments utilized Par C5 cells to examine the role of P2X7 receptor (P2X7R) in apoptosis, particularly in the cleavage and release of alpha-fodrin, an apparent SS autoantigen. Five mM ATP stimulation induced apoptotic cell death with a sustained Ca²⁺ influx, which was mimicked in HEK cells transfected with P2X7R. ATP also induced cleavage of alpha-fodrin mediated by caspase-3 and calpain, releasing alpha-fodrin fragments through membrane blebs. However, both apoptotic cell death and alpha-fodrin cleavage were inhibited in the presence of 300 µM oxidized-ATP (ox-ATP), an irreversible blocker of P2X7R, or in Ca²⁺-free solution. We concluded that P2X7R plays an important role in apoptosis and alpha-fodrin degradation in salivary epithelial cells, providing an important clue elucidating the presence of alpha-fodrin fragments in SS tissues.

How to Influence Patient Oral Hygiene Behavior Effectively

Clarkson, J.E., Young, L., Ramsay, C.R., Bonner, B.C., Bonetti, D. — Fri, 25 Sep 2009 15:02:33 PDT

Considerable resources are expended in dealing with dental disease easily prevented with better oral hygiene. The study hypothesis was that an evidence-based intervention, framed with psychological theory, would improve patients’ oral hygiene behavior. The impact of trial methodology on trial outcomes was also explored by the conducting of two independent trials, one randomized by patient and one by dentist. The study included 87 dental practices and 778 patients (Patient RCT = 37 dentists/300 patients; Cluster RCT = 50 dentists/478 patients). Controlled for baseline differences, pooled results showed that patients who experienced the intervention had better behavioral (timing, duration, method), cognitive (confidence, planning), and clinical (plaque, gingival bleeding) outcomes. However, clinical outcomes were significantly better only in the Cluster RCT, suggesting that the impact of trial design on results needs to be further explored.

Lifecourse Socio-economic Mobility and Oral Health in Middle Age

Pearce, M.S., Thomson, W.M., Walls, A.W.G., Steele, J.G. — Fri, 25 Sep 2009 15:02:33 PDT

Socio-economic variations in health exist for a wide range of health outcomes, including oral health and oral-health-related quality of life (OHRQoL). Less is known regarding how socio-economic trajectories may influence oral health and OHRQoL. This study examined whether social mobility is related to the number of teeth retained by age 50 years and OHRQoL measured at the same time, using data from the Newcastle Thousand Families Study, a birth cohort established in 1947. Women remaining in the non-manual class had the greatest tooth retention. While promotion of a healthier lifestyle and continued improvements in oral hygiene throughout life appear to be the public health interventions most likely to improve oral health into middle age, there may be sub-groups of the population on which different approaches in terms of public health interventions need to be focused.

Missing Posterior Teeth and Risk of Temporomandibular Disorders

Wang, M.Q., Xue, F., He, J.J., Chen, J.H., Chen, C.S., Raustia, A. — Fri, 25 Sep 2009 15:02:33 PDT

There is disagreement about the association between missing posterior teeth and the presence of temporomandibular disorders (TMD). Here, the purpose was to investigate whether the number of missing posterior teeth, their distribution, age, and gender are associated with TMD. Seven hundred and forty-one individuals, aged 21–60 years, with missing posterior teeth, 386 with and 355 without TMD, were included. Four variables—gender, age, the number of missing posterior teeth, and the number of dental quadrants with missing posterior teeth—were analyzed with a logistic regression model. All four variables—gender (OR = 1.59, men = 1, women = 2), age (OR = 0.98), the number of missing posterior teeth (OR = 0.51), and the number of dental quadrants with missing posterior teeth (OR = 7.71)—were entered into the logistic model (P < 0.01). The results indicate that individuals who lose posterior teeth, with fewer missing posterior teeth but in more quadrants, have a higher prevalence of TMD, especially young women.

Periodontal Therapy Improves Gastric Helicobacter pylori Eradication

Fri, 25 Sep 2009 15:02:33 PDT

The oral cavity has been proposed as a reservoir for H. pylori that could be responsible for the refractoriness of gastric infection to triple therapy (antibiotics, antimicrobials, and proton pump inhibitors). The aim of this study was to evaluate the efficiency of combined periodontal and triple therapy vs. triple therapy alone, in gastric H. pylori eradication in persons with H. pylori in the subgingival biofilm. Individuals positive for H. pylori in their gastric and oral samples, as determined by nested PCR, were treated either with periodontal and triple therapy or with triple therapy alone. Our results indicate that 77.3% of those treated with the combined therapy exhibited successful eradication of gastric H. pylori, compared with 47.6% who underwent only triple therapy. Analysis of these data suggests that periodontal treatment in combination with systemic therapy could be a promising approach to increasing the therapy’s efficacy and decreasing the risk of infection recurrence.

Influence of Luting Material Filler Content on Post Cementation

Fri, 25 Sep 2009 15:02:33 PDT

Luting of fiber posts to intra-radicular dentin represents the worst-case scenario in terms of control of polymerization shrinkage. This study tested the hypothesis that filler content of resin cements does not influence luting of fiber posts to intra-radicular dentin, by assaying polymerization stress, push-out bond strength, and nanoleakage expression. The polymerization stress of experimental cements containing 10%, 30%, 50%, or 70% in filler content was investigated. Post spaces were prepared in endodontically treated teeth, and fiber posts were cemented with the experimental cements. A push-out test was performed, and interfacial nanoleakage expression was analyzed. Results showed that luting cements with higher filler content were related to increased polymerization stress (p < 0.05), decreased push-out bond strength (p < 0.05), and increased interfacial nanoleakage expression (p < 0.05). Conversely, lower-stress luting materials increased bonding of fiber posts to intra-radicular dentin. Further in vivo studies are needed to investigate the long-term clinical performance of these materials.

Micro-organism and Cell Viability on Antimicrobially Modified Titanium

Omori, S., Shibata, Y., Arimoto, T., Igarashi, T., Baba, K., Miyazaki, T. — Fri, 25 Sep 2009 15:02:33 PDT

When titanium is anodized by discharge in NaCl solution, both antimicrobial activity and osteoconductivity are conferred. The viability of adherent micro-organisms and cells on antimicrobial titanium remains uncertain. We hypothesized that a thin peroxidation barrier would efficiently destroy adherent bacteria, whereas adherent osteoblastic cells would be viable, since these cells adhere to the surface indirectly though serum proteins. The efficacy of antimicrobial titanium appears to be based on peroxidation, since peroxidation products were detected in parallel with the destruction of bacterial cell-surface structures. The peroxidation effect of antimicrobial titanium was confined to the surface within narrow limits. The viability of osteoblastic cells on the surface was strongly dependent on the presence of serum protein, whereas that of adherent Streptococcus mutans was not affected by the presence of serum proteins. Therefore, differences in the adherent systems used by bacteria and osteoblastic cells are important determinants of their viability on antimicrobial titanium.

Eruption of Primary Dentition--a Grave Health Problem According to Spanish Doctors of the XVI-XVIII Centuries

Romero-Maroto, M., Saez-Gomez, J.M. — Fri, 18 Sep 2009 15:59:46 PDT

Neural Crest Lineage Segregation: a Blueprint for Periodontal Regeneration

Fri, 18 Sep 2009 15:59:46 PDT

During the recent decade, the periodontal attachment apparatus has become one of the premier areas of the body for the development of novel tissue-engineering strategies. In the present review, we describe a developmental biology approach to characterize current concepts in periodontal regeneration and to discuss strategies for future applications in periodontal therapies. To decipher the developmental make-up of the periodontal region, we have followed the path of the migratory neural crest, since it gives rise to periodontal progenitor tissues, which in turn are subjected to the influence of diverse craniofacial extracellular matrices and peptide growth factors. Based on this developmental perspective, we have conducted a systematic analysis of the factors, progenitor cells, and matrices used in current periodontal tissue-engineering approaches. We propose that the developmental history of a tissue is a highly instructive design template for the discovery of novel bioengineering tools and approaches.

Mesenchymal Stem Cells Derived from Dental Tissues vs. Those from Other Sources: Their Biology and Role in Regenerative Medicine

Huang, G.T.-J., Gronthos, S., Shi, S. — Fri, 18 Sep 2009 15:59:46 PDT

To date, 5 different human dental stem/progenitor cells have been isolated and characterized: dental pulp stem cells (DPSCs), stem cells from exfoliated deciduous teeth (SHED), periodontal ligament stem cells (PDLSCs), stem cells from apical papilla (SCAP), and dental follicle progenitor cells (DFPCs). These postnatal populations have mesenchymal-stem-cell-like (MSC) qualities, including the capacity for self-renewal and multilineage differentiation potential. MSCs derived from bone marrow (BMMSCs) are capable of giving rise to various lineages of cells, such as osteogenic, chondrogenic, adipogenic, myogenic, and neurogenic cells. The dental-tissue-derived stem cells are isolated from specialized tissue with potent capacities to differentiate into odontogenic cells. However, they also have the ability to give rise to other cell lineages similar to, but different in potency from, that of BMMSCs. This article will review the isolation and characterization of the properties of different dental MSC-like populations in comparison with those of other MSCs, such as BMMSCs. Important issues in stem cell biology, such as stem cell niche, homing, and immunoregulation, will also be discussed.

Mechanical Properties of Tannic-acid-treated Dentin Matrix

Bedran-Russo, A.K.B., Yoo, K.J., Ema, K.C., Pashley, D.H. — Fri, 18 Sep 2009 15:59:46 PDT

Dentin collagen is a major component of the hybrid layer, and its stability may have a great impact on the properties of adhesive interfaces. We tested the hypothesis that the use of tannic acid (TA), a collagen cross-linking agent, may affect the mechanical properties and stability of the dentin matrix. The present study evaluated the effects of different concentrations of TA on the modulus of elasticity and enzymatic degradation of dentin matrix. Hence, the effect of TA pre-treatment on resin-dentin bond strength was assessed with the use of two bonding systems. Sound human molars were used and prepared according to each experimental design. The use of TA affected the properties of demineralized dentin by increasing its stiffness. TA treatment inhibited the effect of collagenase digestion on dentin matrix, particularly for 10%TA and 20%TA. The TA-dentin matrix complex resulted in improved bond strength for both adhesive systems.

Osteoblast Mechanoresponses on Ti with Different Surface Topographies

Sato, N., Kubo, K., Yamada, M., Hori, N., Suzuki, T., Maeda, H., Ogawa, T. — Fri, 18 Sep 2009 15:59:46 PDT

During implant healing, mechanical force is transmitted to osteogenic cells via implant surfaces with various topographies. This study tested a hypothesis that osteoblasts respond to mechanical stimulation differently on titanium with different surface topographies. Rat bone-marrow-derived osteoblastic cells were cultured on titanium disks with machined or acid-etched surfaces. A loading session consisted of a 3-minute application of a 10- or 20-µm-amplitude vibration. Alkaline phosphatase activity and gene expression increased only when the cells were loaded in 3 sessions/day on machined surfaces, regardless of the vibration amplitude, whereas they were increased with 1 loading session/day on the acid-etched surface. The loading did not affect the osteoblast proliferation on either surface, but selectively enhanced the cell spreading on the machined surface. Analysis of the data suggests that osteoblastic differentiation is promoted by mechanical stimulation on titanium, and that the promotion is disproportionate, depending on the titanium surface topography. The frequency of mechanical stimulation, rather than its amplitude, seemed to have a key role.

Surface Conditioning Influences Zirconia Ceramic Bonding

Kern, M., Barloi, A., Yang, B. — Fri, 18 Sep 2009 15:59:46 PDT

Air-abrasion seems to be mandatory for durable resin bonding to zirconia ceramic. Air-abrasion might compromise the ceramic strength by creating surface defects. Therefore, omitting air-abrasion or using reduced air-pressure seems desirable. We tested the null hypotheses that omitting air-abrasion or using reduced air-pressure does not affect zirconia ceramic bonding independent of using primers. Three mechanical surface conditions (polished, air-abraded at 0.05 or at 0.25 MPa) and 4 priming conditions were tested. After different surface conditioning, zirconia ceramic specimens were bonded, and tensile bond strengths were evaluated after water storage for 3 days or for 150 days with additional 37,500 thermal cyclings for artificial aging. Omitting air-abrasion resulted in debonding during artificial aging independent of using primers. The combination of air-abrasion and priming improved long-term resin bonding to zirconia ceramic significantly. With low-pressure air-abrasion, surface roughness was reduced without affecting long-term bond strength, provided that adequate adhesive primers were applied.

Mmp-20 and Klk4 Cleavage Site Preferences for Amelogenin Sequences

Fri, 18 Sep 2009 15:59:46 PDT

Mmp-20 and Klk4 are the two key enamel proteases. Can both enzymes process amelogenin to generate the major cleavage products that accumulate during the secretory stage of amelogenesis? We isolated Mmp-20 and Klk4 from developing pig teeth and used them to digest the tyrosine-rich amelogenin polypeptide (TRAP), the leucine-rich amelogenin protein (LRAP), and 5 fluorescence peptides. We characterized the digestion products by LC-MSMS, SDS-PAGE, and C18 RP-HPLC monitored with fluorescence and UV detectors. Mmp-20 cleaves amelogenin sequences after Pro¹⁶², Ser¹⁴⁸, His⁶², Ala⁶³, and Trp⁴⁵. These cleavages generate all of the major cleavage products that accumulate in porcine secretory-stage enamel: the 23-kDa, 20-kDa, 13-kDa, 11-kDa, and 6-kDa (TRAP) amelogenins. Mmp-20 cleaves LRAP after Pro⁴⁵ and Pro⁴⁰, producing the two LRAP products previously identified in tooth extracts. Among these key cleavage sites, Klk4 was able to cleave only after His⁶². We propose that Mmp-20 alone processes amelogenin during the secretory stage.

Eph/ephrinB Mediate Dental Pulp Stem Cell Mobilization and Function

Arthur, A., Koblar, S., Shi, S., Gronthos, S. — Fri, 18 Sep 2009 15:59:46 PDT

Damage to the dentin matrix instigates the proliferation and mobilization of dental progenitor cells to the injury site, the mechanisms of which are not defined. EphB receptors and ephrin-B ligands expressed within the perivascular niche of dental pulp have been implicated following tooth injury. We propose that elevated levels of ephrin-B1 following injury may prevent the proliferation and migration of dental pulp stem cell (DPSC), while EphB/ephrin-B interaction facilitates odontoblastic differentiation. The migration, proliferation, and differentiation of DPSC in response to Eph/ephrin-B molecules was assessed in an established ex vivo tooth injury model and by in vitro assays for the assessment of colony formation and differentiation. Analysis of our data demonstrated that EphB forward signaling promoted DPSC proliferation, while inhibiting migration. Conversely, reverse signaling enhanced DPSC mineral production. These observations suggest that EphB/ephrin-B molecules are important for perivascular DPSC migration toward the dentin surfaces and differentiation into functional odontoblasts, following damage to the dentin matrix.

Angiogenic Signaling Triggered by Cariogenic Bacteria in Pulp Cells

Soden, R.I., Botero, T.M., Hanks, C.T., Nor, J.E. — Fri, 18 Sep 2009 15:59:46 PDT

The inflammation observed in the dental pulp of teeth with deep caries lesions is characterized by a significant increase in blood vessel density. It is known that lipoteichoic acid (LTA) from Gram-positive cariogenic bacteria induces expression of vascular endothelial growth factor (VEGF) in dental pulp cells. The hypothesis underlying this study was that LTA induces VEGF expression in dental pulp cells through TLR2 and PI3k/Akt signaling. Odontoblast-like cells (MDPC-23) and undifferentiated pulp cells (OD-21) were exposed to LTA from Streptococcus sanguis, and the role of TLR2, PI3K/Akt, and IKK signaling in LTA-induced VEGF expression was evaluated. These studies demonstrated that TLR2 signaling through the PI3K-Akt pathway is necessary for LTA-induced VEGF expression in pulp cells. In contrast, inhibition of IKK signaling did not prevent VEGF up-regulation in response to LTA. Understanding signaling pathways triggered by cariogenic bacteria may reveal novel therapeutic targets for the clinical management of pulpitis.

Poisson Analysis of Streptococcal Bond-strengthening on Saliva-coated Enamel

Mei, L., Ren, Y., Busscher, H.J., Chen, Y., van der Mei, H.C. — Fri, 18 Sep 2009 15:59:46 PDT

The forces responsible for bond-strengthening in initial oral bacterial adhesion are unknown. Since Lifshitz-Van der Waals and electrostatic forces work instantaneously upon approach, it is hypothesized that bond-strengthening is governed by hydrogen bonding. Poisson analysis of adhesion forces observed during the retraction of bacterial probes from surfaces in atomic force microscopy can be used to analyze the nature of the adhesion forces. Streptococcal adhesion forces increased from about –0.7 to –10.3 nN when the contact time between cell surfaces and salivary films on enamel was increased from 0 to 120 sec. Initial and final adhesion forces were stronger for initial colonizers of tooth surfaces (S. mitis, S. sanguinis) than for later, more cariogenic, strains (S. sobrinus, S. mutans). Retraction curves after increased contact times showed minor peaks, representative of hydrogen bonds, and Poisson analyses indicated repulsive non-specific forces of around +0.3 nN and slightly more attractive hydrogen-bonding forces (–1.0 nN) for initial than for late colonizers (–0.8 nN).

Histatins Enhance Wound Closure with Oral and Non-oral Cells

Fri, 18 Sep 2009 15:59:46 PDT

The role of human saliva in oral wound-healing has never been fully elucidated. We previously demonstrated that parotid-salivary histatins enhance in vitro wound closure. The question remains whether other salivary-gland secretions enhance wound closure, and also the effects of histatins on primary and non-oral cells. Since the presence of histatins is not limited to parotid saliva, we expected to observe wound-closure activity of other salivary-gland secretions. However, here we show that non-parotid saliva does not stimulate wound closure, most probably due to the presence of mucins, since the addition of MUC5B to parotid saliva abolished its effect. Furthermore, we found that histatins stimulated wound closure of (primary) cells of both oral and non-oral origin. This suggests that the cellular receptor of histatins is widely expressed and not confined to cells derived from the oral cavity. These findings encourage the future therapeutic application of histatins in the treatment of all kinds of wounds.

Acceleration of Purine Degradation by Periodontal Diseases

Fri, 18 Sep 2009 15:59:46 PDT

Periodontal diseases, such as gingivitis and periodontitis, are characterized by bacterial plaque accumulation around the gingival crevice and the subsequent inflammation and destruction of host tissues. To test the hypothesis that cellular metabolism is altered as a result of host-bacteria interaction, we performed an unbiased metabolomic profiling of gingival crevicular fluid (GCF) collected from healthy, gingivitis, and periodontitis sites in humans, by liquid and gas chromatography mass spectrometry. The purine degradation pathway, a major biochemical source for reactive oxygen species (ROS) production, was significantly accelerated at the disease sites. This suggests that periodontal-disease-induced oxidative stress and inflammation are mediated through this pathway. The complex host-bacterial interaction was further highlighted by depletion of anti-oxidants, degradation of host cellular components, and accumulation of bacterial products in GCF. These findings provide new mechanistic insights and a panel of comprehensive biomarkers for periodontal disease progression.

Stability of the JP2 Clone of Aggregatibacter actinomycetemcomitans

Haubek, D., Ennibi, O.-K., Vaeth, M., Poulsen, S., Poulsen, K. — Fri, 18 Sep 2009 15:59:46 PDT

The JP2 clone of Aggregatibacter actinomycetemcomitans is strongly associated with aggressive periodontitis. To obtain information about colonization dynamics of the JP2 clone, we used PCR to examine its presence in 365 Moroccan juveniles from whom periodontal plaque samples were collected at baseline and after one and two years. Periodontal attachment loss was measured at baseline and at the two-year follow-up. At baseline, 43 (12%) carriers of the JP2 clone were found. Nearly half (44 %) of these were persistently colonized with the clone. The relative risk for the development of aggressive periodontitis, adjusted for the concomitant presence of other genotypes of A. actinomycetemcomitans, was highest for individuals continuously infected by the JP2 clone (RR = 13.9; 95% CI, 9.0 to 21.4), indicating a relationship between infectious dose and disease, which further substantiates the evidence for the JP2 clone as a causal factor in aggressive periodontitis.

Novel RUNX2 Mutations in Chinese Individuals with Cleidocranial Dysplasia

Zhang, C.Y., Zheng, S.G., Wang, Y.X., Zhu, J.X., Zhu, X., Zhao, Y.M., Ge, L.H. — Fri, 18 Sep 2009 15:59:47 PDT

Cleidocranial dysplasia (CCD) is an inherited autosomal-dominant skeletal disease caused by heterozygous mutations in the osteoblast-specific transcription factor, RUNX2. We performed mutation analysis of RUNX2 on four unrelated Chinese individuals with CCD. Three novel distinct mutations were detected in the coding region of RUNX2: two missense and one frameshift. These mutations were exclusively clustered within the Runt domain. One missense mutation converts threonine to isoleucine at codon 200 (T200I). The other one substitutes leucine for arginine at codon 225 (R225L), which affects many family members. The frame-shift mutation (214fs) in exon3 leads to the introduction of a translational stop codon at codon 221, resulting in a truncated RUNX2 protein. The reporter gene assays revealed that all the mutants exhibited significantly reduced transactivation activities on the osteocalcin promoter. Our results provide new genetic evidence that mutations involved in RUNX2 contribute to CCD. Abbreviations: AML3, gene encoding acute myeloid leukemia protein 3; bp, base pair; CBFA1, gene encoding core-binding factor 1; CBFβ, gene encoding core-binding factor β; CCD, cleidocranial dysplasia; NLS, nuclear localization signal; OSE2, osteoblast-specific cis-acting element 2; PEBP2A, gene encoding polyoma enhancer binding protein 2A; PST, proline/serine/ threonine-rich domain; Q/A, glutamine-alanine repeat domain; Runt, Runt Homology Domain; RUNX2, the mammalian runt-related genes 2; RUNX2, Runt-related protein 2.