[Effect of molar ligation and local Porphyromonas gingivalis inoculation on alveolar bone loss in the mouse].

OBJECTIVE: To compare the extent and time course of alveolar bone loss and osteoclast activation in two murine models of periodontal disease: molar ligation and Porphyromonas gingivalis (P. gingivalis) oral inoculation. METHODS: A split-mouth design was applied to two groups of mice (C57BL6, 6-8 weeks old, n=24 in both groups), resulting in four treatment groups: (1) Control group: unligated upper right 2nd molars receiving CMC only, (2)Ligature group: ligation of a 9-0 suture around the upper left 2nd molar, (3) P. gingivalis group: unligated upper right 2nd molar receiving P. gingivalis challenge only, (4)Ligature+P.gingivalis group: ligation of the upper left 2nd molar in combination with oral inoculation with 109 colony-forming units(CFU) P. gingivalis. Alveolar bone loss was measured as the cementoenamel junction and alveolar bone crest (CEJ-ABC) distance. In the study, 48 C57BL6 mice were designed and treated as described above, and osteoclasts were counted on histological sections following tartrate-resistant acid phosphatase (TRAP) staining and counts were normalized to alveolar bone surface distance. Then 36 C57BL6 mice were investigated, of which 30 were ligated a 9-0 silk ligature around the 2nd molar in the left maxillary quadrant and 6 were not ligated. After ligation for 1 week, the ligatures in 12 mice were taken off for either 1 week or 2 weeks. The CEJ-ABC distance of the 6 mice without ligation was baseline. The CEJ-ABC distances were measured and analyzed. The data were analyzed with one-way ANOVA. RESULTS: Molar ligation induced marked alveolar bone loss after 3, 6, 9 and 12 weeks [(0.16±0.04) mm, (0.16±0.02) mm, (0.18±0.03) mm, (0.17±0.02) mm], vs. corresponding controls [(0.09±0.03)mm,(0.10±0.01)mm,(0.12±0.04)mm,(0.12±0.01)mm] and P. gingivalis group [(0.09±0.03)mm, (0.12±0.01)mm,(0.12±0.02)mm,(0.10±0.01)mm], P<0.05. Combined treatment with molar ligation and P. gingivalis did not further increase the CEJ-ABC distance. Evidence for osteoclast activation was found one day after molar ligation, and TRAP-positive cell numbers peaked on day 3 (12±4 vs. control 2±2, P<0.01). After taking off ligature following ligation for 2 weeks, it showed significantly regrowth of alveolar bone compared with that before removal of the ligature on day 7 [(0.07±0.02)mm vs. (0.13±0.01)mm, P<0.01]. CONCLUSION: Molar ligation is a rapid and effective way to induce periodontal bone loss in mice. Osteoclast activation occurs within 24 hours of ligature placement, and the extent of bone loss well exceeds that of the P.gingivalis-induced bone loss. Removing ligature after periodontal disease might help bone regeneration by regrowth of the alveolar bone.

Connective tissue graft plus resin-modified glass ionomer restoration for the treatment of gingival recession associated with non-carious cervical lesions: microbiological and immunological results.

OBJECTIVES: It was previously reported the clinical results of placing subgingival resin-modified glass ionomer restoration for treatment of gingival recession associated with non-carious cervical lesions. The aim of this study was to evaluate the influence of this treatment on the subgingival biofilm and gingival crevicular fluid (GCF) inflammatory markers. MATERIALS AND METHODS: Thirty-four patients presenting the combined defect were selected. The defects were treated with either connective tissue graft plus modified glass ionomer restoration (CTG+R) or with connective tissue graft only (CTG). Evaluation included bleeding on probing and probing depth, 5 different bacteria targets in the subgingival plaque assessed at baseline, 45, and 180 days post treatments, and 9 inflammatory mediators were also assessed in the GCF. RESULTS: The levels of each target bacterium were similar during the entire period of evaluation (p > 0.05), both within and between groups. The highest levels among the studied species were observed for the bacterium associated with periodontal health. Additionally, the levels of all cyto/chemokines analyzed were not statistically different between groups (p > 0.05). CONCLUSION: Within the limits of the present study, it can be concluded that the presence of subgingival restoration may not interfere with the subgingival microflora and with GCF inflammatory markers analyzed. CLINICAL RELEVANCE: This approach usually leads to the placement of a subgingival restoration. There is a lack of information about the microbiological and immunological effects of this procedure. The results suggest that this combined approach may be considered as a treatment option for the lesion included in this study.

[Exogenous tooth discoloration in children: black stains]. / Les colorations dentaires noires exogènes chez l'enfant : Black-stains.

Black-stains are a coloring frequently met in pediatric dentistry. They can be medically diagnosed as 1-mm borders or unfinished lines formed by a dark exogenous substance which follows the gingival festoon of bet coronary (in cervical third of the crown) temporary teeth and permanent, or they can appear in like points or dark spots. They are caused by bacteria anaerobic chromogenous. The dominant responsible species are actinomyces. Blacks-stains are ferrous depots, formed following a chemical interaction on the surface of the tooth between sulphide of hydrogen (under the effect of the anaerobic bacteria which are producing hydrogen) and the iron contained in the saliva (by a healthy diet) or that released by red blood corpuscles (in case of bloody gums). Black-stains are a shape of characteristic dental plaque by its flora with trend to calcify. It contains an insoluble iron salt with a content raised in calcium and in inorganic phosphor. The coloring Black-stain is a mild pathology and has no incidence on the vitality of the tooth. Certainly these spots are unsightly. The dental surgeon in current practice can deprive them. The pediatrician plays a leading role in the diagnosis and advice to parents and patients affected by these stains.

The efficacy of a cervical barrier in preventing microleakage of Enterococcus faecalis in endodontically treated teeth.

The clinical failure of coronal restorations can compromise the healthy periapical status of endodontically treated teeth. The purpose of the present ex vivo study was to assess the effectiveness of the cervical barrier in preventing bacterial microleakage in the eventual loss of the coronal restoration. Following removal of the smear layer and obturation to the cementoenamel junction using gutta-percha, 70 single-rooted mandibular premolars were randomly divided into five different groups: Group 1 received no additional treatment; Groups 2 and 3 had 2 mm and 3 mm of the obturation removed, respectively, followed by placement of a cervical barrier that was the same thickness as the obturation. In Group 4 (positive control), the root canals were filled only with gutta-percha, while in Group 5 (negative control), the root canals were completely impermeabilized following obturation. The filled root canals were incorporated into a split-chamber model system using Enterococcus faecalis as a microbial marker. The apices were suspended in the lower chambers. Leakage was determined daily for 60 days and evaluated for turbidity. According to Fisher's exact test, the cervical barrier in Groups 2 and 3 prevented the microleakage of E. faecalis into the root canals when compared with Groups 1 and 4. This result was similar to that for the completely sealed samples in Group 5 (p = 0.001).

The CEJ: a biofilm and calculus trap.

There is a direct correlation between increasing periodontal probing depth and increasing presence of residual biofilms and calculus. One of the more common areas to find postscaling and root-planing residual biofilm and calculus is the cemento-enamel junction (CEJ)--an area that in most cases is easily accessible. Yet few studies have addressed the question of why the CEJ is a potential biofilm and calculus trap. This article discusses the various anatomical relationships of enamel, dentin, and cementum at the CEJ, the role of CEJ anatomy in the retention of biofilm (with SEM photographs as supporting evidence), and the biological and clinical implications of subgingival residual biofilm and calculus.

Chemical, mechanical and morphological properties of hypomineralized enamel of permanent first molars.

OBJECTIVE: The microstructure of hypomineralized enamel in permanent teeth has been described in several studies as less distinct prism sheaths and disorganized enamel with lack of organization of the enamel crystals. The mechanical properties, hardness and modulus of elasticity of the hypomineralized enamel have lower values compared with normal. The aim of this study was to examine normal and hypomineralized enamel using scanning electron microscopy (SEM), hardness measurements and X-ray microanalysis (XRMA). MATERIAL AND METHODS: Four extracted hypomineralized permanent first molars, sectioned and cut in half, were analyzed with SEM, XRMA and hardness measurements. RESULTS: An inverse relation was found between the micro hardness and the Ca:C ratio in hypomineralized and normal enamel. The acid-etched hypomineralized enamel appeared on SEM to be covered with a structureless layer and the prisms appeared disorganized, with thick prism sheaths and loosely packed crystallites. Furthermore, bacteria were found deep in porous hypomineralized enamel close to the enamel-dentin junction. CONCLUSIONS: Teeth diagnosed with molar incisor hypomineralization have significantly lower hardness values in the hypomineralized enamel compared with normal enamel. The hardness values vary according to the morphological and chemical properties.

Biofilm removal with a dental water jet.

OBJECTIVE: The objective of this study was to evaluate the effect of a dental water jet on plaque biofilm removal using scanning electron microscopy (SEM). METHODOLOGY: Eight teeth with advanced aggressive periodontal disease were extracted. Ten thin slices were cut from four teeth. Two slices were used as the control. Eight were inoculated with saliva and incubated for 4 days. Four slices were treated using a standard jet tip, and four slices were treated using an orthodontic jet tip. The remaining four teeth were treated with the orthodontic jet tip but were not inoculated with saliva to grow new plaque biofilm. All experimental teeth were treated using a dental water jet for 3 seconds on medium pressure. RESULTS: The standard jet tip removed 99.99% of the salivary (ex vivo) biofilm, and the orthodontic jet tip removed 99.84% of the salivary biofilm. Observation of the remaining four teeth by the naked eye indicated that the orthodontic jet tip removed significant amounts of calcified (in vivo) plaque biofilm. This was confirmed by SEM evaluations. CONCLUSION: The Waterpik dental water jet (Water Pik, Inc, Fort Collins, CO) can remove both ex vivo and in vivo plaque biofilm significantly.

Detection of Streptococcus mutans and Streptococcus sobrinus in dental plaque samples from Brazilian preschool children by polymerase chain reaction.

The purposes of this study were to detect S. mutans and S. sobrinus by polymerase chain reaction (PCR) amplification, and to relate their presence to the incidence of dental caries in 42 Brazilian preschool children. Dental plaque samples were collected from the cervical margin of all erupted teeth of 5-6 years old children with primary dentition, using a sterile explorer. Examination of the dmft (decayed, missing, filled teeth) index, performed following the World Health Organization (WHO) caries diagnostic criteria, showed a 2.71 score. Prevalence of S. mutans and S. sobrinus was respectively, of 85.7% and 14.3%; no dental plaque sample was either positive or negative for both bacterial species. Children harboring either S. mutans or S. sobrinus presented the same caries prevalence. PCR showed good discriminative ability for differentiation between these species, and suggested that it is a technique suitable for epidemiological studies on mutans streptococci.

Detection of streptococcus mutans and streptococcus sobrinus in dental plaque samples from Brazilian preschool children by polymerase chain reaction

The purposes of this study were to detect S. mutans and S. sobrinus by polymerase chain reaction (PCR) amplification, and to relate their presence to the incidence of dental caries in 42 Brazilian preschool children. Dental plaque samples were collected from the cervical margin of all erupted teeth of 5-6 years old children with primary dentition, using a sterile explorer. Examination of the dmft (decayed, missing, filled teeth) index, performed following the World Health Organization (WHO) caries diagnostic criteria, showed a 2.71 score. Prevalence of S. mutans and S. sobrinus was respectively, of 85.7 percent and 14.3 percent; no dental plaque sample was either positive or negative for both bacterial species. Children harboring either S. mutans or S. sobrinus presented the same caries prevalence. PCR showed good discriminative ability for differentiation between these species, and suggested that it is a technique suitable for epidemiological studies on mutans streptococci.

A presença de Streptococcus mutans e S. sobrinus, detectadas através da técnica da reação em cadeia da polimerase (PCR) foi comparada com a incidência de cárie dental em 42 crianças pré-escolares brasileiras. As amostras da placa dental foram obtidas utilizando-se um explorador estéril, o qual foi passado na margem cervical de todos os dentes erupcionados nas crianças de 5-6 anos de idade, as quais possuíam a dentição primária. Foi realizado um exame dental para a avaliação do índice ceod (índice de dentes decíduos cariados, extraídos por cárie ou obturados) de acordo com os critérios de diagnóstico de cárie preconizados pela Organização Mundial de Saúde (OMS). O índice ceod foi de 2,71. A presença de S. mutans e S. sobrinus foi detectada em 85,7 por cento e 14,3 por cento das crianças analisadas respectivamente, nenhuma amostra da placa dental foi positiva ou negativa para a presença das duas bactérias ao mesmo tempo. As crianças que apresentavam S. mutans ou S. sobrinus isoladamente apresentaram o mesmo índice de cárie. A técnica de PCR apresentou um grande poder discriminatório entre as duas espécies o que sugere que ela é adequada para estudos epidemiológicos no grupo mutans.

Black stains in the mixed dentition: a PCR microbiological study of the etiopathogenic bacteria.

The aim of this work is to emphasize that particular stains on the third cervical of the buccal and lingual surfaces in mixed dentition, called "black stain." Previous research showed the microbiological etiology of this discoloration by chromogen bacterias. Our study shows bacteria spp involved in stains by means of PCR process and electrophoresis gel on the agarose medium. Sample was formed by 100 subject with black stain and 100 control subjects stain-free. A statistical analysis (SPSS 10.0) using X2 was performed in this study. Porphyromonas gingivalis and Prevotella melaninogenica, were not involved in both in black stain subjects and in the control. On the contrary, Actinomyces could be involved in the pigmentation process.

Morphological analysis of subgingival biofilm formation on synthetic carbonate apatite inserted into human periodontal pockets.

BACKGROUND: Details of the development of human subgingival biofilm are unknown due to the difficulties in conducting experiments and especially in obtaining undisturbed materials. METHODS: This study was performed using deposits on carbonate apatite that had been inserted into human periodontal pockets for up to three weeks. Scanning electron microscopy using the vertically sectioned method and transmission electron microscopy using the freeze-substitution method were adopted. RESULTS: The development of subgingival biofilm occurred in five sequential phases: pellicle formation, microbial adherence, initial colonization, microbial organization, and establishment. Certain species in each of the initial, secondary and tertiary colonizers were considered to have a predilection for biofilm formation. Gram-positive, bacillary initial colonizers and gram-negative, filamentous secondary colonizers organized one stable structure that served as the framework for biofilm formation, and gram-negative, rod-shaped tertiary colonizers with cell-surface vesicles showed multigeneric coaggregation. The microbiota in the tertiary colonizers underwent repeated microflora alteration. CONCLUSIONS: Subgingival biofilm is constituted by initial, secondary and tertiary colonizers. Microflora alteration which is suggested to be related to periodontal disease, frequently occurred in the tertiary colonizers.

Relating visual and radiographic ranked scoring systems for occlusal caries detection to histological and microbiological evidence.

This study compared a visual ranked scoring system and a radiographic ranked scoring system for occlusal caries detection with the level of infection of dentin. Seventy-five third-molars, designated for extraction, were professionally cleaned. Caries was scored according to a visual ranked scoring system at a selected site in the groove-fossa system. Radiographs of the teeth were available and caries was recorded along a five-point ranked scoring system. Each tooth was extracted and hemi-sectioned through the investigation site under aseptic conditions. A burful of dentin was removed from the EDJ of one of the section faces and these samples were processed to establish the level of dentin infection. The depth of the lesion was assessed on the other section face using a five-point ranked histological scoring system. A strong relationship was observed between the histological lesion depth and visual score (r(s)=0.93) while a moderate relationship was seen between lesion depth and radiographic scores (r(s)=0.77). The dentin from teeth with cavities exposing dentin was heavily infected. The dentin from teeth with microcavities or grey discoloration of the dentin was less infected than the lesions with frank cavitation (score 4) (p<0.05, t-test), but more infected than the initial lesions (p<0.05, t-test). The latter lesions showed minimal infection. A similar tendency was seen with respect to increasing radiographic scores and the level of infection of the dentin.

Effect of sonic and mechanical toothbrushes on subgingival microbial flora: a comparative in vivo scanning electron microscopy study of 8 subjects.

OBJECTIVES: The purpose of this initial study was to evaluate the effects of both a sonic and a mechanical toothbrush versus the effects of no treatment on depth of subgingival penetration of epithelial and tooth-associated bacteria. METHOD AND MATERIALS: Eight adult subjects exhibiting advanced chronic periodontitis with at least 3 single-rooted teeth that were in separate sextants with facial pockets > or = 4 mm and < or = 8 mm and that required extraction constituted the experimental sample. Teeth were either subjected to 15 seconds of brushing with a mechanical toothbrush or a sonic toothbrush or left untreated. The test tooth and the associated soft tissue wall of the periodontal pocket were removed as a single unit. Samples were processed and coded for blind examination by scanning electron microscopy. Distributional and morphologic characteristics of dominant bacteria with specific emphasis on spirochetes were evaluated for both epithelial- and tooth-associated plaque. RESULTS: No differences were found in morphotypes or distributional and aggregational characteristics of epithelial-associated microbes in the 1- to 3-mm subgingival zone between the mechanical and sonic toothbrush-treated groups and the control group. Both toothbrush groups featured disruption of microbes that extended up to 1 mm subgingivally. Root surfaces on the sonic-treated samples appeared plaque-free at low magnification; however, at 4,700x, a thin layer of mixed morphotypes and intact spirochetes was found subgingivally and slightly subgingivally. In comparison, mechanical brush samples featured incompletely removed plaque, both subgingivally and subgingivally, with intact spirochetes present on subgingival root surfaces. CONCLUSION: Results suggest similar effects for both sonic and mechanical toothbrushes on epithelial- and tooth-associated bacterial plaque in periodontal pockets and adjacent root surfaces that extend up to 1 mm subgingivally. Further, the presence of intact subgingival spirochetes suggests limited exposure to acoustical or mechanical energy from the toothbrushes evaluated.

Bonding of a self-etching primer to non-carious cervical sclerotic dentin: interfacial ultrastructure and microtensile bond strength evaluation.

PURPOSE: The objectives of this study were 1) to examine the ultrastructural features of the resin-sclerotic dentin interface following the application of Clearfil Liner Bond II sigma to natural cervical wedge-shaped lesions, and 2) to evaluate the regional tensile bond strength of this self-etching primer at different locations on natural and artificially-created cervical lesions. MATERIALS AND METHODS: Deep cervical natural lesions were bonded using the self-etching primer. Micromorphology of the bonded interface at different locations within the lesions were examined using scanning electron microscopy (SEM), transmission electron microscopy (TEM) and scanning transmission electron microscopy/energy dispersive x-ray analysis (STEM/EDX). Ultrastructural features were further compared with the use of the same self-etching primer on artificial lesions created in sound cervical dentin. A nontrimming technique was used to evaluate the regional tensile bond strength from the occlusal, gingival, and the deepest central part of both natural and artificial cervical lesions. Beams with a mean area of 0.46 +/- 0.03 mm2 were prepared and were pulled to failure using a Bencor Multi-T testing device attached to an Instron universal tester. Bond strength results were evaluated using a two-way ANOVA design. RESULTS: A hypermineralized layer devoid of intact, banded collagen was invariably present on the surface of the natural lesions. Depending upon its thickness at different locations of the lesion, the action of a self-etching primer may be limited to this surface layer alone, producing a hybridized hypermineralized surface layer. Penetration of the self-etching primer into the underlying sclerotic dentin produced a hybridized complex containing a hybridized hypermineralized surface layer as well as a subsurface layer of hybridized intertubular dentin. Bacterial colonization of the lesion surface resulted in the formation of an additional zone of hybridized intermicrobial matrix over the surface of the lesions. Dentinal tubules remained blocked with sclerotic casts, and resin tags were rarely observed. Regional tensile bond strength results showed that the overall bond strength to natural sclerotic dentin was about 20% lower than sound cervical dentin, but was independent of the different locations within the lesions from which bond strength was evaluated. CONCLUSION: There were four factors that may have influenced the overall decrease in bond strength in natural cervical sclerotic lesions: a) the presence of a hybridized intermicrobial matrix together with entrapped bacteria may have weakened the bonds, b) inability of a self-etching primer to etch through a thick, hypermineralized surface layer, c) presence of a layer of possibly remineralized, denatured collagen at the base of the hypermineralized surface layer, and d) retention of acid-resistant sclerotic casts that obliterate the tubular lumina and prevent effective resin tag formation.

Root surface characteristics of primary teeth from children with prepubertal periodontitis.

This study describes the histologic characteristics of root surfaces of primary teeth from children with prepubertal periodontitis (PP). Fifteen primary teeth from 4 children with PP, and 2 control primary teeth from 2 healthy children were examined. Light microscopy revealed normal root surfaces in the control teeth. In contrast, the PP specimens revealed bacteria inside dentin tubules or covering cementum, a cuticle, or resorbed dentin; normal, wider than normal, or hypoplastic cementum; resorption lacunae with various depths; aplastic root resorption; alternate resorption and repair; and active repair. No cementoclasts were found in the resorption lacunae. Scanning electron microscopy revealed intrabony and suprabony root areas, and a "plaque free zone" (PFZ). Colonies of filaments were evident at the cemento-enamel junction (CEJ). The suprabony root surfaces had resorption lacunae, isolated short rods, calculus, colonies of filaments, or colonies composed by an heterogeneous bacterial population. The coronal boundary of the PFZ was the border of a sheet-like structure, which included isolated rods or filaments. At the PFZ, isolated filaments and rods, and a fibril matrix were evident. The apical boundary of the PFZ consisted of bundles of soft tissue remnants or the insertion of the periodontal fibers. The intrabony surfaces were mostly covered by soft tissue, which included isolated filaments and short rods. Resorption lacunae with or without soft tissue were also evident in this area. Crystals of calcium oxalate dihydrate and erythrocytes in distinct forms were found at various root areas. The present findings are different from those previously reported for hypophosphatasia specimens.

Pathohistology of undecalcified primary teeth in vitamin D-resistant rickets: review and report of two cases.

The basic dental defects in vitamin D-resistant rickets seem to be manifested in dentin. Enamel is usually reported to be normal. This histologic examination showed the penetration of microorganisms through the calcified structures of the enamel layer without visible caries. The microorganisms passed through the dentinoenamel junction and invaded dentin, which was characterized by calcospherites and large amounts of interglobular dentin. Furthermore, microorganisms could be detected in dentinal tubules, which were exposed to the oral cavity when enamel was removed. However, large areas of tertiary dentin extended between such tubules and the pulp. These light microscopic results suggest that clinical manifestations, such as, pulp recrosis and periapical lesions (without carious defects) may be caused by the penetration of microorganisms through microclefts of the enamel layer as well as pathologically altered enamel microstructures of affected teeth.