Pathogens and host immunity in the ancient human oral cavity.

Calcified dental plaque (dental calculus) preserves for millennia and entraps biomolecules from all domains of life and viruses. We report the first, to our knowledge, high-resolution taxonomic and protein functional characterization of the ancient oral microbiome and demonstrate that the oral cavity has long served as a reservoir for bacteria implicated in both local and systemic disease. We characterize (i) the ancient oral microbiome in a diseased state, (ii) 40 opportunistic pathogens, (iii) ancient human-associated putative antibiotic resistance genes, (iv) a genome reconstruction of the periodontal pathogen Tannerella forsythia, (v) 239 bacterial and 43 human proteins, allowing confirmation of a long-term association between host immune factors, 'red complex' pathogens and periodontal disease, and (vi) DNA sequences matching dietary sources. Directly datable and nearly ubiquitous, dental calculus permits the simultaneous investigation of pathogen activity, host immunity and diet, thereby extending direct investigation of common diseases into the human evolutionary past.

Root malformation associated with a cervical mineralized diaphragm--a distinct form of tooth abnormality?

OBJECTIVE: Deviations in length and shape of tooth roots result from hard tissue resorption or occur as a developmental disorder. The purpose of this report is to present a type of root malformation which seems to have gone unreported so far. STUDY DESIGN: Two patients showing severely dysplastic roots of all permanent first molars were evaluated using radiography, histology as well as scanning and transmission electron microscopy. RESULTS: Medical histories of the patients revealed significant, but diverse events in the first year after birth. Radiographically the pulp cavity floors of the affected molars in large part were occupied by ectopic mineralized plates. Microscopically these plates consisted of hard tissue, densely calcified globules, and a network of canals which contained large blood vessels and were lined by cementum and periodontal ligament. CONCLUSIONS: We propose that the ectopic mineralized plate was derived from the dental follicle, had developed during crown formation around the vascular plexus at the base of the dental papilla, and represented a mechanical obstacle interfering with normal root development.

Resorption pattern and radiographic diagnosis of invasive cervical resorption. A correlative microCT, scanning electron and light microscopic evaluation of a case series.

The aim of this study was to evaluate, whether and how the histologic structure of invasive cervical resorption (external granulomas) affect their clinical radiographic diagnosis. For this purpose, nine more or less intact extracted teeth, of which intraoral radiographs were available, were processed for examination in the scanning electron (SEM) and light microscope. From some of the specimens non-decalcified ground sections were prepared and some were decalcified for preparation of histologic sections. Five teeth were scanned in an X-ray microtomography (microCT) device and digitally reconstructed in three dimensions. Three histologic characteristics, namely (1) the location and extension of the resorptive defect, (2) the size of the communication with the periodontium, and (3) the amount of mineralized substitution tissue formed in the lesion affected the radiographic diagnosis. At early and advanced stages, invasive cervical resorption had encircled the pulp cavities, however, without destroying the innermost dentin. Even if the dentinal wall adjacent to the pulp was histologically intact and thick, this important diagnostic feature frequently could not be recognized in conventional radiographs. Also, the communication between the resorptive lesion and the periodontium often was not visible radiographically, although it could always be identified in the microscopic evaluation, particularly at early stages when it proved to be very small. Invasive cervical resorption lesions containing large amounts of mineralized substitution tissue were difficult to recognize and, therefore, could easily be overseen. Thus, three features which are deemed essential for the differential diagnosis of invasive cervical resorption were not readily apparent in conventional radiographs. From these three features, the dentinal wall against the pulp cavity and the communication to the peri-odontium were, however, clearly visible in the microCT reconstructions.

E-cadherin regulates the behavior and fate of epithelial stem cells and their progeny in the mouse incisor.

Stem cells are essential for the regeneration and homeostasis of many organs, such as tooth, hair, skin, and intestine. Although human tooth regeneration is limited, a number of animals have evolved continuously growing teeth that provide models of stem cell-based organ renewal. A well-studied model is the mouse incisor, which contains dental epithelial stem cells in structures known as cervical loops. These stem cells produce progeny that proliferate and migrate along the proximo-distal axis of the incisor and differentiate into enamel-forming ameloblasts. Here, we studied the role of E-cadherin in behavior of the stem cells and their progeny. Levels of E-cadherin are highly dynamic in the incisor, such that E-cadherin is expressed in the stem cells, downregulated in the transit-amplifying cells, re-expressed in the pre-ameloblasts and then downregulated again in the ameloblasts. Conditional inactivation of E-cadherin in the cervical loop led to decreased numbers of label-retaining stem cells, increased proliferation, and decreased cell migration in the mouse incisor. Using both genetic and pharmacological approaches, we showed that Fibroblast Growth Factors regulate E-cadherin expression, cell proliferation and migration in the incisor. Together, our data indicate that E-cadherin is an important regulator of stem cells and their progeny during growth of the mouse incisor.

MRI of the temporo-mandibular joint: which sequence is best suited to assess the cortical bone of the mandibular condyle? A cadaveric study using micro-CT as the standard of reference.

OBJECTIVE: To determine the best suited sagittal MRI sequence out of a standard temporo-mandibular joint (TMJ) imaging protocol for the assessment of the cortical bone of the mandibular condyles of cadaveric specimens using micro-CT as the standard of reference. METHODS: Sixteen TMJs in 8 human cadaveric heads (mean age, 81 years) were examined by MRI. Upon all sagittal sequences, two observers measured the cortical bone thickness (CBT) of the anterior, superior and posterior portions of the mandibular condyles (i.e. objective analysis), and assessed for the presence of cortical bone thinning, erosions or surface irregularities as well as subcortical bone cysts and anterior osteophytes (i.e. subjective analysis). Micro-CT of the condyles was performed to serve as the standard of reference for statistical analysis. RESULTS: Inter-observer agreements for objective (r = 0.83-0.99, P < 0.01) and subjective (κ = 0.67-0.88) analyses were very good. Mean CBT measurements were most accurate, and cortical bone thinning, erosions, surface irregularities and subcortical bone cysts were best depicted on the 3D fast spoiled gradient echo recalled sequence (3D FSPGR). CONCLUSION: The most reliable MRI sequence to assess the cortical bone of the mandibular condyles on sagittal imaging planes is the 3D FSPGR sequence. KEY POINTS: MRI may be used to assess the cortical bone of the TMJ. • Depiction of cortical bone is best on 3D FSPGR sequences. • MRI can assess treatment response in patients with TMJ abnormalities.

Discoloration of teeth from tetracyclines--even today?

The aim of this study was to examine whether brownish crown and root discoloration of wisdom teeth was related to treatment of acne with tetracyclines. For this purpose, 17 discolored third molars from nine patients were embedded without being decalcified, ground along the tooth axis, and examined using fluorescence microscopy. A thorough medical history served to determine the start and duration of any administration of tetracyclines. This confirmed the use of drugs against acne containing minocycline in all cases except one. The microscopic analyses of all teeth revealed intensely fluorescent bands in the dentin, which corresponded to the mineralization front at the time of tetracycline intake. More or less uniform discoloration of the entire crown was seen in association with treatment against acne prior to the completion of crown formation at the age of about 15 years. This uniform staining can be attributed to incorporation of minerals during ongoing maturation of the occlusal enamel, which is concomitant with the formation of the cervical crown regions. When acne was treated between 15 and 22 years of age, only the roots of the third molars displayed annular discolorations, which seemed to result from the incorporation of tetracyclines into dentin, while fine fluorescent incremental lines in root cementum were too thin to be apparent clinically. Three accidentally removed interradicular bony septa revealed that tetracyclines incorporated into alveolar bone remained there for about 2 years, but thereafter disappeared as a result of physiological remodelling.

Rare dental peculiarities associated with the hypomyelinating leukoencephalopathy 4H syndrome/ADDH.

PURPOSE: 4H syndrome/ADDH, a disease of the cerebral white matter, seems to be associated with delayed tooth eruption and other dental abnormalities, which so far could not be assessed conclusively-mainly because patients were too young. The aim of this study was to characterize these abnormalities in a sample of patients old enough for a reliable assessment. METHODS: Three children, all diagnosed with 4H syndrome/ADDH, were followed from approximately 4 to 10 years of age and examined clinically and radiographically. In one case, a histopathological analysis supplemented these records. RESULTS: All 3 patients showed a generalized delay in eruption of the primary and permanent teeth, which culminated in complete retention of all primary maxillary central incisors. Permanent mandibular second premolars were missing in all children and permanent maxillary central incisors of 2 individuals exhibited a concave labial surface, while agenesis of the permanent maxillary lateral incisors and natal or neonatal teeth were observed in one patient. CONCLUSION: 4H syndrome/ADDH seems to be associated with a delay in primary tooth eruption, complete retention of the primary maxillary central incisors, and shape abnormalities of the permanent maxillary central incisors, which otherwise are very rare. Therefore, a neurological examination would appear warranted when these peculiarities are encountered.

Misfit of suprastructures on implants processed by electrical discharge machining or the Cresco method.

OBJECTIVE: To assess the accuracy of fit of frameworks on implants processed with electrical discharge machining (EDM) or the Cresco technique (Astra Tech). METHOD AND MATERIALS: On 12 identical master casts with implants at positions 9(21), 11(23), and 13(25), high-gold alloy frameworks were produced by standard casting procedure. Six frameworks were used for the Cresco technique (group CRE) by employing specific fixed partial denture supports. The remaining 6 frameworks were cast with prefabricated gold copings and served as control. The finished frameworks were screwed onto implant 25 of the corresponding master cast. Dimensions of the marginal gaps were measured at 4 locations on each implant under the scanning electron microscope, applying the replica technique. Subsequently, the control group was processed by EDM (SAE EDM 2000) (group EDM) and analyzed alike. Statistical analysis of the results was performed with Kruskal-Wallis and Mann-Whitney U tests. RESULTS: The mean marginal gaps were measured as follows (CRE/EDM/control): position 25: 0.0 microm/1.0 +/- 1.6 microm/1.5 +/- 2.1 microm; position 23: 5.2 +/- 5.6 microm/18.7 +/- 29.3 microm/23.6 +/- 30.7 microm; and position 21: 36.0 +/- 21.6 microm/40.7 +/- 31.0 microm/46.0 +/- 41.1 microm. The only statistically significant difference was found at location 23 between group CRE on one side and both group EDM and control on the other side. The strong increase of misfit for group CRE from location 23 to location 21 indicates that laser welding is the crucial parameter in this technique. CONCLUSION: The Cresco technique has a potential to reduce the marginal gap between implants and suprastructures.

Enamel-free teeth: Tbx1 deletion affects amelogenesis in rodent incisors.

TBX1 is a principal candidate gene for DiGeorge syndrome, a developmental anomaly that affects the heart, thymus, parathyroid, face, and teeth. A mouse model carrying a deletion in a functional region of the Tbx1 gene has been extensively used to study anomalies related to this syndrome. We have used the Tbx1 null mouse to understand the tooth phenotype reported in patients afflicted by DiGeorge syndrome. Because of the early lethality of the Tbx1-/- mice, we used long-term culture techniques that allow the unharmed growth of incisors until their full maturity. All cultured incisors of Tbx1-/- mice were hypoplastic and lacked enamel, while thorough histological examinations demonstrated the complete absence of ameloblasts. The absence of enamel is preceded by a decrease in proliferation of the ameloblast precursor cells and a reduction in amelogenin gene expression. The cervical loop area of the incisor, which contains the niche for the epithelial stem cells, was either severely reduced or completely missing in mutant incisors. In contrast, ectopic expression of Tbx1 was observed in incisors from mice with upregulated Fibroblast Growth Factor signalling and was closely linked to ectopic enamel formation and deposition in these incisors. These results demonstrate that Tbx1 is essential for the maintenance of ameloblast progenitor cells in rodent incisors and that its deletion results in the absence of enamel formation.

Deletion of BMP7 affects the development of bones, teeth, and other ectodermal appendages of the orofacial complex.

Sequential and reciprocal epithelial-mesenchymal interactions govern the development of most tissues and organs of the craniofacial region. Bone morphogenetic proteins (BMPs) are members of the transforming growth factor-beta family of secreted signaling molecules that have long been implied to have a significant contribution in this process. However, evidence for such a role during craniofacial development is largely missing. Using a lacZ reporter mouse we mapped the spatiotemporal expression of BMP7 in the developing craniofacial region. The observed pattern suggested a potential involvement of BMP7 in epithelial-mesenchymal interactions and thus a direct role for this molecule in the development of ectodermal appendages (teeth, hair follicle, lachrymal and sweat glands, taste buds) and, furthermore, palatal formation. To correlate the expression to function we analyzed germline deleted conditional BMP7-deficient embryos for malformations. We found developmental defects in many craniofacial structures such as teeth, eyes, whiskers, hair follicles, salivary glands, and palate. These findings place BMP7 as a central mediator of epithelial-mesenchymal interactions that are necessary for the correct development of structures belonging to the orofacial complex.

Distribution of insulin-like growth factors in condylar hyperplasia.

Condylar hyperplasia (CH) is a local overgrowth of the condylar process of the temporomandibular joint (TMJ) of unknown etiology. Probably, growth factors like the insulin-like growth factors (IGFs) are involved in its pathogenesis. Specimens from 12 patients were investigated histologically and immunohistochemically to obtain the distribution of the IGFs-I and -II and the IGF1 receptor. The results revealed juvenile and adult subtypes. While generally IGF-II could only be detected weakly, in the juvenile cases strong immunostaining for IGF-I in cartilage and bone supposes an influence on pathological growth processes.

Biocompatibility of Beta-tricalcium phosphate root replicas in porcine tooth extraction sockets - a correlative histological, ultrastructural, and x-ray microanalytical pilot study.

This investigation studies porcine tissue response in tooth extraction sockets treated with root replicas made out of Beta-tricalcium phosphate (Beta-TCP; Beta-Ca(3)(PO(4))(2)) granules, molded and held together by thermal fusion of a thin film of polyglycolic-polylactic acid copolymer. Six left mandibular third incisors (n (1)/4 6) of experimental pigs are treated with the root replicas and four contralateral incisors are used as nontreated controls (n (1)/4 4). Two animals each were killed at 20, 40, and 60 weeks of observation periods. The mandibular jaw segments were prepared in toto for light microscopy by resin embedding and serial ground sectioning. Additionally, one Beta-TCP-treated socket at 60 weeks was thoroughly investigated by correlative light, electron microscopic and electron probe X-ray microanalysis to assess the bio-absorbability and host removal of the replica material from the implant site. The extraction wounds of the animals healed satisfactorily with very little histologically observable differences in the healing pattern of the test and control sites. The Beta-TCP was completely removed from extracellular sites, but at 60 weeks, remnants of it were found in the cytoplasm of multinucleated giant cells. The root replicas made out of Beta-TCP were biocompatible and bioabsorbable. Osseous healing occurred both in the test and control sockets, but the healing process was delayed due to the presence of Beta-TCP particles.

Performance of visual inspection, electrical conductance and laser fluorescence in detecting occlusal caries in vitro.

The aim of this study was to compare visual inspection (VI) and electrical conductance (EC) and laser fluorescence (LF) measurements in detecting occlusal caries. VI was based on fissure discoloration and performed with the naked eye. EC was measured with the ECM device (Lode Diagnostic, Groningen, The Netherlands), and LF was assessed with the DIAGNOdent apparatus (KaVo, Biberach, Germany). In extracted human premolars and molars, clinically sound sites (D0-/D1-lesions), enamel caries (D2-lesions), and dentinal caries (D3-/D4-lesions) were identified using recommended cut-off values. Thereafter, the teeth were cut longitudinally and analyzed by scanning electron microscopy for verification of caries depth. Reproducibility of VI was good, that of EC and LF excellent. In identifying caries at both the enamel and dentin level, the sensitivities of VI and LF were significantly (p < 0.05) higher than that of EC, while EC was significantly (p < 0.05) more specific. The positive predictive values, however, did not exceed 43%. Improved diagnoses at the dentinal level were obtained, when EC and LF were used as an adjunct to VI and when cut-off values were raised. Thus, visual inspection relying exclusively on fissure discoloration seems to allow only proper identification of sound occlusal surfaces. In cases of discolored fissures, the appliance-based methods help to avoid false positive identification of dentinal caries. However, attainable reliabilities of diagnoses do not seem to exceed about 50% to 60%.

Pulpal response to Er:YAG laser drilling of dentine in healthy human third molars.

BACKGROUND AND OBJECTIVES: Maintenance of pulpal health is a critical prerequisite for successful application of light amplification by stimulated emission of radiations (lasers) in the hard tissue management of vital teeth. The purpose of this study was to investigate the short- and long-term pulpal effects to cavity-preparations in healthy human teeth using erbium-doped:yttrium, aluminum, and garnet (Er:YAG) laser. MATERIALS AND METHODS: A total of seven healthy third molars that were to be removed due to space-problem were used. Following the laser excavation, the cavities in dentine were closed temporarily and the teeth were extracted after 7 days (n = 5) and 3 months (n = 2) post-operation. The specimens were fixed, decalcified, subdivided, and processed for light and transmission electron microscopy. RESULTS: In the short-term group, four of the five laser-drilled teeth did not reveal any pathological changes in the pulp-dentine complex. One tooth showed mild disruption of odontoblasts (OB) and vascular dilatation subjacent to the deepest point of the cavity-preparation with a remaining dentine thickness (RDT) of less than 80 microm. The two teeth under long-term observation revealed distinct apposition of tertiary dentine (TD), lined predominantly with cuboidal cells on its pulpal aspect. CONCLUSIONS: These results would allow a conclusion to be drawn that the Er:YAG laser under investigation is a pulp preserving hard-tissue drilling tool when used with the specific energy settings and emitting radiation at a wavelength of 2.94 microm.

Ciliated epithelium-lined radicular cysts.

OBJECTIVE: This report describes 3 cases of ciliated epithelium-lined radicular cysts among 256 apical periodontitis lesions and also illustrates the occurrence of an Actinomyces-infected periapical cyst. STUDY DESIGN: Serial and step serial sections of 256 plastic-embedded root apices with attached apical periodontitis lesions that were prepared for a previous investigation were reviewed for the presence of ciliated epithelium-lined radicular cysts. The lesions that were found to have such epithelial lining were examined in a transmission electron microscope to elaborate the fine structure of the ciliated cells. RESULTS: A total of 3 ciliated columnar epithelium-lined cysts was found among the 256 apical periodontitis lesions examined. Two of the lesions also contained stratified squamous epithelium. All 3 lesions affected maxillary premolars. One of the lesions was a true cyst, and the other 2 were periapical pocket cysts. The lumen of 1 of the latter revealed the presence of typical "ray-fungus" actinomycotic colonies. CONCLUSION: Although the stratified squamous component of the epithelia that lined the radicular cysts reported here may be derived from the cell rests of Malassez, the ciliated epithelial cells may be of sinus origin. Microbial agents from diseased root canals can advance into radicular cysts, particularly in pocket cysts, with the possible threat of such infection in upper posterior teeth spreading into the maxillary sinus.

Factors affecting degeneration in human temporomandibular joints as assessed histologically.

The influence of sex, age, tooth loss, and articular disc position on temporomandibular joint (TMJ) degeneration was evaluated in specimens collected at autopsy from 15 women and 38 men ranging in age from 15 to 92 yr. The position of the articular discs was classified as normal or abnormal, tooth loss was both counted and categorized. Degenerative changes of the articular tissues were assessed histologically and quantified, taking into account both the severity of structural alterations and their extension along the articular surface. This was recorded separately in the condyle, disc, and temporal component, three latero-medial joint regions, and putative load-bearing and non-load-bearing surfaces. Analysis of covariance with repeated measures served for testing contributing factors. It showed that the effects of sex and the number of missing teeth were insignificant, whereas age up to about 55-60 yr and reduction of dental arch length proved to be the most important factors. Load-bearing seemed to play a significant role mainly at younger ages, and the effect of disc position was significant, when internal derangement was combined with reduction of dental arch length. Thus, rising severity of TMJ degenerative changes appears to be associated primarily with increasing age. In addition, it may also depend on mechanical factors, in particular loss of molar support and, to a minor degree, abnormal disc position.