Archives of Oral Biology

Editorial Board

2010-08-01

Collagenolytic activity in sonic extracts of Tannerella forsythia

Norio Kawase, Jun-ichi Kishi, Hiroshi Nakamura, Taro Hayakawa — 2010-08-01

Abstract: Objective: The purpose of the present study was to characterize the collagenolytic activity in a sonicated extract of Tannerella forsythia and to investigate the activation of proMMMP-2 and -9 by the T. forsythia extract.Methods: The T. forsythia extract was incubated with type I collagen. The cleaved products were then analyzed by SDS-PAGE using the method of Laemmli. We studied the effects of cysteine, DTT, CaCl2, and various proteinase inhibitors on collagenolytic activity. A HT1080 cell culture supernatant containing proMMP-2 and -9 was incubated with the T. forsythia extract and analyzed for the activation of proMMP-2 and -9 by gelatin zymography.Results: The T. forsythia extract degraded type I collagen. Cysteine increased the collagenolytic activity of the extract, and 5mM CaCl2 was required for this activity. The collagenolytic activity of T. forsythia was inhibited by N-ethylmaleimide, iodoacetamide, iodoacetic acid, EDTA, and leupeptin, but not by PMSF, E-64, TLCK, or TPCK. When proMMP-2 and -9 were incubated with the T. forsythia extract, gelatinases with the relative molecular masses of MMP-2 and -9 were produced.Conclusion: The present study suggests that T. forsythia extract is able to degrade type I collagen and activate proMMP-2 and -9.

Association between biofilm-forming isolates of mutans streptococci and caries experience in adults

Rodrigo A. Giacaman, Enrique Araneda, Carlos Padilla — 2010-08-01

Abstract: Objectives: Although it is still controversial, mutans streptococci (MS) have been typically considered the primary etiological agents of dental caries. Besides the acidogenic and aciduric properties, extracellular polysaccharide synthesis leading to biofilm formation from sugar constitutes one of the key virulence factors of MS. The aim of this study was to investigate whether biofilm formation by MS was associated to an increased experience of caries in young adults.Methods: A cross sectional study with a total of 96 randomly selected patients aged 15–27 years old was carried out. DMFT was determined by clinical examination and bite-wing radiographs. A sample of stimulated saliva was obtained and seeded on agar plates to culture MS. Colonies with and without biofilm formation were identified and quantified.Results: When the total MS count was considered, levels of MS were not associated with higher caries experience. 50% of the patients showed at least one biofilm-forming colony. Patients with biofilm-forming colonies showed significantly higher DMFT (p<0.001) than individuals whose plates did not reveal the structure surrounding the colony, but only at the low and moderate MS count.Conclusion: Biofilm formation in MS appears to be associated with higher caries experience in individuals with low counts of the cariogenic microorganism.

Antimicrobial activity of n-6, n-7 and n-9 fatty acids and their esters for oral microorganisms

Chifu B. Huang, Brian George, Jeffery L. Ebersole — 2010-08-01

Abstract: Objective: This study is to assess the antibacterial activity of omega-6, -7, -9 (n-6, n-7, n-9) fatty acids against various oral microorganisms.Methods: The n-6, n-7, n-9 fatty acids, such as γ-linoleic acid (GLA), linoleic acid (LA), arachidonic acid (ARA), palmitoleic acid (PA), and oleic acid (OA), their fatty acid ethyl esters, GLA-EE, LA-EE, ARA-EE, PA-EE, OA-EE, and their fatty acid methyl esters, GLA-ME, LA-ME, ARA-ME, PA-ME, OA-ME, were investigated for antimicrobial activity against oral pathogens Streptococcus mutans, Candida albicans, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, and Porphyromonas gingivalis. Various concentrations of the fatty acids, their methyl and ethyl esters were tested against various oral pathogens in 96-well plates and blood-agar plate. The plates were incubated anaerobically or aerobically at 37°C for 48h, and the colony forming units (CFU) were determined.Results: The data demonstrated that select n-6, n-7, n-9 fatty acids and their esters exhibited strong antimicrobial activity against these oral microorganisms, demonstrating some specificity for individual microbial species.Conclusion: The potential use or the combinations of the n-6, n-7, n-9 fatty acids and/or their esters, provided in a local delivery vehicle to infected sites in the oral cavity, could be considered as an additional therapeutic approach to improving oral health.

Distribution pattern of cholinesterase enzymes in human tooth germs

T.L. Nandasena, C.K. Jayawardena, W.M. Tilakaratne, C.D. Nanayakkara — 2010-08-01

Abstract: The two distinct molecular forms of cholinesterase (ChE) are acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). Our previous studies have reported that ChE is involved in tooth development. However, further experiments are needed to understand the precise action of ChE in tooth development. This study aimed to localise types of ChE in human tooth germs, and identify their distribution pattern.ChE were localised in frozen sections of jaws which were prepared from dead fetuses, neonates and stillborns who were free from visible abnormalities by Karnovsky and Root method.AChE was identified in the inner and outer enamel epithelia including the cervical loop region, stratum intermedium and preameloblasts of tooth germs at bell stage. Secretory ameloblasts were free from staining. The bud and cap stages of permanent tooth germs showed AChE activity on the lingual aspect and top surface of the epithelial ingrowths, respectively. BuChE activity was localised in the degenerating dental lamina.Our study reported the first evidence of localisation of ChE in human tooth development and identified the possible molecular form of ChE in tooth germs as AChE. Also, our results have provided strong evidence to speculate the action of AChE is on the cells of enamel organ during tooth development.

Cell proliferation and apoptosis in the primary enamel knot measured by flow cytometry of laser microdissected samples

2010-08-01

Abstract: Laser capture microdissection (LCM) uniquely allows the selection of specific cell populations from histological sections. These selected cells are then catapulted into a test tube without any contamination from surrounding tissues. During the last ten years, many significant results have been achieved, particularly at the level of DNA and RNA where amplification techniques are available. However, where amplification procedures are difficult, the benefits of LCM diminish.To overcome such difficulties, a novel approach, combining laser capture microdissection and flow cytometry, has been tested here for detection of apoptosis and proliferation in tissue bound cell populations without any amplification steps. The mouse cap stage molar tooth germ was used as a model. At the centre of the inner enamel epithelium, the primary enamel knot is a clearly defined apoptotic population with minimal proliferation, flanked by the highly proliferative cervical loops on each side. Thus within the tooth germ epithelium at this stage, two distinct populations of cells are found side by side. These populations were selected by laser capture microdissection and then analysed by flow cytometry for apoptosis and proliferation. Flow cytometric results correlated well with immunohistochemical findings, demonstrating the success and sensitivity of this combined procedure.

The case of an insular molarless black rat: Effects on lifestyle and mandible morphology

Pere Bover, Josep A. Alcover, Jacques Michaux, Sabrina Renaud — 2010-08-01

Abstract: We report a specimen of an insular black rat (Rattus rattus) from Illa den Colom (Menorca, Western Mediterranean) displaying a singular dental characteristic. It has no molar teeth but displays regular incisors. Its mere occurrence as a regular adult rat is puzzling and we attempted to evaluate what diet and morphological changes in jaw shape were promoted by the total lack of molars, and allowed the successful survival of this specimen. Two approaches were performed: first, bone tissue was analysed to obtain δ15N and δ13C values in order to estimate dietary preferences of the rat. Second, the shape of the jaw was analysed through elliptic Fourier analysis, using outlines as markers of diet. The values for C and N fractionation (−19.89‰ and 10.06‰, respectively) suggest that the molarless rat included animal food in the diet and not exclusively plant material as observed in other mainland rat populations. The morphometric analysis in which the shape of the molarless mandible falls into the range of omnivorous groups leads to a similar conclusion. The adult age of the specimen suggests that it fed efficiently enough with its incisors to allow a normal growth. Although displaying a lack of molar teeth, no deep changes in remodelling jaw morphology can be observed and its shape falls into the variation of regular murines. The molarless rat exemplifies that special ecological features on small islands allow the survival of aberrant morphotypes.

Participation of the endocannabinoid system in lipopolysaccharide-induced inhibition of salivary secretion

2010-08-01

Abstract: Objective: The aim of the present paper was to assess whether lipopolysaccharide (LPS)-induced inhibition of salivary secretion involves the activation of the endocannabinoid system and the participation of tumor necrosis factor (TNF)α in the submandibular gland.Design: Pharmacological approaches were performed by using CB1 and/or CB2 cannabinoid receptor antagonists, AM251 and AM630, respectively, injected into the submandibular gland, to study the participation of the endocannabinoid system in LPS inhibitory effects on metacholine-induced salivary secretion. To assess the participation of TNFα on LPS inhibitory effects, salivary secretion was studied in LPS treated rats after the intraglandular injection of etanercept, a soluble form of TNF receptor which blocks TNFα action. Finally, to evaluate the possible interplay between endocannabinoids and TNFα on the submandibular gland function reduced during LPS challenge, the salivary secretion was studied after the intraglandular injection of this cytokine alone or concomitantly with AM251 and AM630.Results: AM251 and AM630, injected separately or concomitantly, partially prevented LPS-induced inhibition of salivation. Also, anandamide synthase activity was increased in submandibular glands extracted from rats 3h after LPS injection, suggesting that the endocannabinoid system was activated in response to this challenge. On the other hand, etanercept, prevented the inhibitory effect of LPS on salivary secretion and moreover, TNFα injected intraglandularly inhibited salivary secretion, being this effect prevented by AM251 and AM630 injected concomitantly.Conclusion: The present results demonstrate the participation of the endocannabinoid system and TNFα on salivary responses during systemic inflammation induced by LPS.

The expression of estrogen receptors in rat genioglossus muscle-derived satellite cells and its relationship to intracellular Ca2+ mobilization

Tao Guo, Xiao-Ling Yu, Yin Ding — 2010-08-01

Abstract: Genioglossus (GG) is the most important pharyngeal dilator muscle in maintaining upper airway (UA) patency in human; therefore, its dysfunction plays an important role in pathogenesis of sleep-related breathing disorder. Recently, the expression of estrogen receptors (ERs) on mRNA and protein level has been evidenced in GG muscle; however, the cellular localization of two subtypes of ER in GG myoblasts remains unclear. The present study was designed to clarify the expression and cellular distribution of ERs in rat GG muscle-derived satellite cells (MDSCs) and further probe the effect of ERs expression on regulation of intracellular Ca2+. The immunocytochemistry revealed positive staining for both ERα and ERβ in nuclei and cytoplasm of GG MDSCs. Noticeably, positive signals for ERα and ERβ were comparable in cytoplasm, whereas the positive staining of ERα in nuclear was obviously strong than that of ERβ. More intriguingly, by using Fluo 4-AM as a fluorescent Ca2+ indicator and 17β-estradiol (E2) as a stimulant, we observed that the level of intracellular Ca2+ was not affected by E2 application, which implied that Ca2+ signaling may not be involved in ER-mediated estrogenic effects on GG MDSCs. Taken together, the present study clearly indicates the differential cellular localization of ERs in rat GG MDSCs; moreover, ER-mediated estrogenic effect in rat GG MDSCs bears no relationship to intracellular Ca2+ mobilization. In addition, the GG MDSCs express both ERα and ERβ and therefore, provide a suitable and convenient in vitro cell model for investigating the molecular mechanisms of estrogenic effects on rat GG muscle.

Estrogen deficiency increases variability of tissue mineral density of alveolar bone surrounding teeth

2010-08-01

Abstract: Objective: Estrogen deficiency increases bone remodeling leading to increased variability of tissue mineral density (TMD). Due to the functional demands of mastication, alveolar bone around teeth is inherently a highly remodeled region of bone tissue with a highly variable distribution of TMD. This study investigated the effect of estrogen deficiency on the TMD distribution of alveolar bone.Design: Using three-dimensional micro-computed tomography images of sham surgery (Sham) and ovariectomized (OVX) rat mandible sections, alveolar bone region (AB) and control bone region (CB) of interest were isolated. Based on histograms of gray levels equivalent to TMD values, mean (Mean), standard deviation (SD) and coefficient of variation (COV=SD/Mean) were computed. Fifth and 95th percentile gray level values were also obtained (Low5 and High5, respectively). Absolute value of percentage (%) differences of the gray level parameters between AB and CB regions were computed.Results: Both SD and COV were significantly higher in AB region than those in CB region for all specimens of both Sham and OVX groups (p<0.001). The mean values of % differences for SD were moderately higher (p<0.073) and those for COV and Low5 were significantly higher for the OVX group than for the Sham group (p<0.04).Conclusions: Higher variability of mineralization observed in AB of OVX group indicates that estrogen deficiency amplifies the active bone remodeling of AB already present due to the mastication. These findings provide an insight that the increased variability of TMD induced by estrogen deficiency may compromise the mechanical stability of the tooth-bearing alveolar bone.