Formation of intermediate cementum. II: a scanning electron microscopic study of the epithelial root sheath of Hertwig in monkey

The intermediate cementum is a layer of calcified tissue between the dentin and the cementum at the periphery of dental roots. The mineralization pattern of the intermediate cementum and the innermost layer of aprismatic enamel in the crowns has been shown to be very similar. Since the formation of these tissues is incompletely known and there have been diverging opinions whether they are of epithelial or mesenchymal origin, the present study aimed at investigating the surface morphology of the root sheath with scanning electron microscopy and correlate it to the mineralization of the intermediate cementum. The advancing mineralization front of the intermediate cementum was covered by the root sheath. Numerous microvilli facing the root were found on this part of the root sheath. The corresponding surface facing the dental follicle was covered with bulb-type junctions. Microvilli and bulb-type junctions have also been demonstrated on the surface of the presecretory ameloblast and associated with formation of aprismatic enamel. Thus, the epithelial root sheath seems to actively take part in the formation of intermediate cementum.     

Detecting demineralization of enamel and cementum after gamma irradiation using radiographic densitometry

The purpose of this study was to evaluate the ability of radiographic densitometry in detecting the early demineralization of human enamel and cementum in irradiated and non-irradiated teeth. Sixty extracted teeth were divided into two groups: irradiated group and non-irradiated group. After irradiation, the groups were subjected to demineralization–remineralization (PH) cycling. Radiographic densitometric measurements (gray values) of a selected area of interest in the enamel and in the cementum of each tooth were performed at baseline and after PH cycling. After PH cycling, there was a significant reduction in gray values for both groups. The difference between “baseline” and “after PH cycling” values represents the reduction in the mineral content of the hard tissue, i.e., the demineralization. Results show that the demineralization of irradiated tooth enamel and cementum was significantly higher compared to that of non-irradiated tissues as determined by gray-level values. It is concluded that densitometric measurements by means of digital radiographs allow for the detection of demineralization of enamel and cementum, and can be used successfully for diagnosis of the early carious lesions in patients who received head and neck radiotherapy. This will allow implementation of remineralizing therapy and avoid the risk of progression of radiation caries. Furthermore, it is concluded that gamma irradiation with typical therapeutic doses for head and neck carcinoma is a direct cause of demineralization of tooth enamel and cementum.     

Functional remineralization of dentin lesions using polymer-induced liquid-precursor process

It was hypothesized that applying the polymer-induced liquid-precursor (PILP) system to artificial lesions would result in time-dependent functional remineralization of carious dentin lesions that restores the mechanical properties of demineralized dentin matrix. 140 μm deep artificial caries lesions were remineralized via the PILP process for 7-28 days at 37°C to determine temporal remineralization characteristics. Poly-L-aspartic acid (27 KDa) was used as the polymeric process-directing agent and was added to the remineralization solution at a calcium-to-phosphate ratio of 2.14 (mol/mol). Nanomechanical properties of hydrated artificial lesions had a low reduced elastic modulus (E(R) = 0.2 GPa) region extending about 70 μm into the lesion, with a sloped region to about 140 μm where values reached normal dentin (18-20 GPa). After 7 days specimens recovered mechanical properties in the sloped region by 51% compared to the artificial lesion. Between 7-14 days, recovery of the outer portion of the lesion continued to a level of about 10 GPa with 74% improvement. 28 days of PILP mineralization resulted in 91% improvement of E(R) compared to the artificial lesion. These differences were statistically significant as determined from change-point diagrams. Mineral profiles determined by micro x-ray computed tomography were shallower than those determined by nanoindentation, and showed similar changes over time, but full mineral recovery occurred after 14 days in both the outer and sloped portions of the lesion. Scanning electron microscopy and energy dispersive x-ray analysis showed similar morphologies that were distinct from normal dentin with a clear line of demarcation between the outer and sloped portions of the lesion. Transmission electron microscopy and selected area electron diffraction showed that the starting lesions contained some residual mineral in the outer portions, which exhibited poor crystallinity. During remineralization, intrafibrillar mineral increased and crystallinity improved with intrafibrillar mineral exhibiting the orientation found in normal dentin or bone.     

Effects of Fluoridated Milk on Root Dentin Remineralization

Background

The prevalence of root caries is increasing with greater life expectancy and number of retained teeth. Therefore, new preventive strategies should be developed to reduce the prevalence of root caries. The aim of this study was to investigate the effects of fluoridated milk on the remineralization of root dentin and to compare these effects to those of sodium fluoride (NaF) application without milk.

Methods

Thirty extracted human molars were divided into 6 groups, and the root cementum was removed from each tooth. The dentin surface was demineralized and then incubated with one of the following six solutions: Sodium chloride NaCl, artificial saliva, milk, milk+2.5 ppm fluoride, milk+10 ppm fluoride and artificial saliva+10 ppm fluoride. Serial sections were cut through the lesions and investigated with polarized light microscopy and quantitative morphometry, scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS). The data were statistically evaluated using a one-way ANOVA for multiple comparisons.

Results

The depth of the lesion decreased with increasing fluoride concentration and was the smallest after incubation with artificial saliva+10 ppm fluoride. SEM analysis revealed a clearly demarcated superficial remineralized zone after incubation with milk+2.5 ppm fluoride, milk+10 ppm fluoride and artificial saliva+10 ppm fluoride. Ca content in this zone increased with increasing fluoride content and was highest after artificial saliva+10 ppm fluoride incubation. In the artificial saliva+10 ppm fluoride group, an additional crystalline layer was present on top of the lesion that contained elevated levels of F and Ca.

Conclusion

Incubation of root dentin with fluoridated milk showed a clear effect on root dentin remineralization, and incubation with NaF dissolved in artificial saliva demonstrated a stronger effect.     

Effects of 1-hydroxyethylidene-1, 1-bisphosphonate (HEBP) on the formation of dentin and the periodontal attachment apparatus in the mouse

The effects of the bisphosphonate HEBP on dentin formation and the structural relationship between the dentin and the developing periodontal attachment apparatus have been studied in the continuously growing mouse incisor. It was observed that HEBP (in doses greater than or equal to 8 mg P/kg b.w/day) not only inhibited the deposition of mineral crystallites in newly formed dentin matrix, but also entirely prevented the formation of a layer of acellular root cementum. It was further noticed that the drug interfered with the deposition of 3H-serine-containing substances at the predentin-dentin border. This was not always accompanied by an inhibition of dentin mineralization, thereby suggesting that 3H-serine-containing proteins (presumably phosphoproteins) do not play a critical role in the deposition of mineral layers onto previously formed ones. The absence of a cementum layer did not prevent the developing periodontal ligament from establishing matrix-to-matrix connections with the root-analogue dentin. Collagen fibrils of the ligament intermingled with those of the mantle dentin, which in contrast to teeth not exposed to the drug were clearly visible and not masked by electron-dense matrix components. Finally, it was found that the drug had distinct effects on the formation of root-analogue and crown-analogue dentin. Whereas during the course of the experiment the odontoblasts along the crown-analogue aspect of the tooth continued to produce circumpulpal dentin matrix, those along the root-analogue aspect of the tooth did so only when the mantle dentin layer had been mineralized prior to HEBP administration. This phenomenon is interpreted as being indicative of fundamental differences between the formation of crown and root dentin.     

The developmental biology of cementum.

In conclusion, we have reviewed an extensive literature on early cementogenesis and performed a detailed morphological and molecular analysis to illustrate and verify key issues in the current debate about epithelial and mesenchymal contributions to root cementum. We have demonstrated that prior to cementogenesis, Hertwig's epithelial root sheath disintegrates and dental follicle cells penetrate the epithelial layer to invade the root surface. Our studies confirmed that HERS became disrupted or disintegrated prior to cementum deposition. We visualized how mesenchymal cells from the dental follicle penetrated the HERS bilayer and deposited initial cementum, while immediately adjacent epithelial cells were separated from the root surface by a basal lamina and did not secrete any cementum. Human specimen from the Gottlieb collection indicated that HERS was removed from the root surface prior to cementum deposition. Our in situ hybridization and immolocalization data revealed that both amelogenin mRNAs and enamel proteins were restricted to the crown enamel and were absent from the root surface and from the cervical-most ameloblasts adjacent to the root margin. On Western blots, cementum protein extracts did not cross-react with amelogenin antibodies. Our studies in conjunction with our literature review together confirmed the classical theory of cementum as a dental follicle derived connective tissue that forms subsequent to HERS disintegration.     

Restoration of Carious Lesions of the Root1

This paper summarizes and evaluates the indications and limitations of various dental materials and techniques for the treatment and restoration of root surface dental caries. Development of a severity index for classifying and sorting lesions is described. Historical materials such as silver amalgam and gold foil are briefly discussed as are contemporary materials such as composite resins and dentin bonding primers. Emphasis is placed on materials and techniques requiring minimal invasion or preparation of the root surface. Eradication of root lesions by recontouring and smoothing the root surface is discussed at length. A new dental material, glass ionomer cement, which contains fluoride and forms a physicochemical bond with enamel and dentin. and requires no preparation of the root surface other than lesion debridement, is described in detail. Guidelines for managing arrested lesions are offered. In addition, the importance of topical fluoride in the treatment and management of root caries is discussed. Lastly, some suggestions are offered for future directions of research in this area.     

Human dentin production in vitro

The main hard tissues of teeth are composed of dentin and enamel, synthesized by the mesenchyme-derived odontoblasts and the epithelial-derived ameloblasts, respectively. Odontoblasts are highly differentiated post-mitotic cells secreting the organic matrix of dentin throughout the life of the animal. Pathological conditions such as carious lesions and dental injuries are often lethal to the odontoblasts, which are then replaced by other pulp cells. These cells are able to differentiate into odontoblast-like cells and produce a reparative dentin. In this study we reproduced this physiological event in an in vitro culture system using pulps of human third molars. Pulp cells cultured in presence of beta-glycerophosphate formed mineralization nodules, which grew all over the culture period. The immunohistochemical study revealed that, as odontoblasts, pulp cells contributing to the nodule formation express type I collagen, osteonectin, and nestin. By the exception of nestin, these proteins are also detected in the nodules. The composition of the nodules was also analyzed by Fourier transform infrared microspectroscopy. The spectra obtained showed that both the organic and the mineral composition of the nodules have the characteristics of the human dentin and differ from those of enamel and bone. Taken together, these results show that both the molecular and the mineral characteristics of the human dentin matrix are respected in the in vitro culture conditions.     

两种脱矿体系制备人工龋损的病理学比较

目的评价两种不同脱矿体系在人牙釉质与牛牙釉质形成人工龋样损害的情况,确定模拟早期龋实验研究的脱矿系统。方法两种脱矿体系：部分饱和酸缓冲溶液和酸性凝胶溶液,分别在人牙釉质与牛牙釉质制备人工早期釉质龋损,采用偏振光显微镜和扫描电子显微镜观察实验人工龋的组织病理学变化。结果部分饱和的酸缓冲体系制备的人工龋损都具有完整的表层和表层下脱矿,牛牙釉质除了脱矿深度大于人牙釉质外,其它病理学特点都类似于人牙釉质,与早期自然龋损相似。而凝胶液脱矿体系形成的人工龋损不管是人牙还是牛牙都没有明显的表层结构,实验牙表面多出现被腐蚀或溶解。结论部分饱和的酸缓冲液脱矿系统形成的人工龋损病理学变化更接近于自然龋;与人牙具有相似化学组成的牛牙很适合代替人牙来检测脱矿和再矿化的效果。     

The SCPP gene repertoire in bony vertebrates and graded differences in mineralized tissues

The vertebrate tooth is covered with enamel in most sarcopterygians or enameloid in chondrichthyans and actinopterygians. The evolutionary relationship among these two tissues, the hardest tissue in the body, and other mineralized tissues has long been controversial. We have recently reported that specific combinations of secretory calcium-binding phosphoprotein (SCPP) genes are involved in the mineralization of bone, dentin, enameloid, and enamel. Thus, the early repertoire of SCPP genes would elucidate the evolutionary relationship across these tissues. However, the diversity of SCPP genes in teleosts and tetrapods and the roles of these genes in distinct tissues have remained unclear, mainly because many SCPP genes are lineage-specific. In this study, I show that the repertoire of SCPP genes in the zebrafish, frog, and humans includes many lineage-specific genes and some widely conserved genes that originated in stem osteichthyans or earlier. Expression analysis demonstrates that some frog and zebrafish SCPP genes are used primarily in bone, but also in dentin, while the reverse is true of other genes, similar to some mammalian SCPP genes. Dentin and enameloid initially use shared genes in the matrix, but enameloid is subsequently hypermineralized. Notably, enameloid and enamel use an orthologous SCPP gene in the hypermineralization process. Thus, the hypermineralization machinery ancestral to both enameloid and enamel arose before the actinopterygian–sarcopterygian divergence. However, enamel employs specialized SCPPs as structuring proteins, not used in enameloid, reflecting the divergence of enamel from enameloid. These results show graded differences in mineralized dental tissues and reinforce the hypothesis that bone–dentin–enameloid–enamel constitutes an evolutionary continuum. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Mechanical recovery of dentin following remineralization in vitro--an indentation study

This study sought to gain insights into the steps leading to remineralization and mechanical recovery of hydrated dentin. Mechanical recovery in water was hypothesized to result from effective mineral matrix binding and to occur from the innermost regions outwards due to an increase in the number of nucleation sites. Partially demineralized (0.05 M acetate, pH=5.0, 8h) dentin was remineralized using calcium and phosphate solutions of 10.1 or 9.8 degree of saturation (DS) for hydroxyapatite (pH=7.4) for 4, 8 or 24h. Remineralization used a constant solution composition approach, which allowed for a continuous mineral growth with relatively constant thermodynamic driving forces. Crystal growth rates (R) were calculated using concentrations of calcium and phosphate. Before and after de- and re-mineralization, specimens had their surface and cross-section elastic moduli measured using AFM-nanoindentation in water. DS=10.1 provided higher R and higher mechanical recovery at the surface (p<0.0001). Cross-sectional measurements showed that subsurface mechanical recovery occurred from the innermost demineralized areas gradually outwards for both groups with no statistical differences at different DS, thus suggesting that remineralization is driven by mineral growth within nucleation sites with preserved collagen fibrils. Further, mechanical recovery appeared to initially obey a heterogeneous pattern, which vanished with time. This study provides evidence of mechanical recovery of hydrated dentin after remineralization and novel insights into the steps leading to mechanical recovery of carious dentin.     

Activity assessment of root caries lesions with thermal and near‐IR imaging methods

The purpose of this study was to evaluate thermal and near‐infrared (NIR) reflectance imaging methods for the assessment of the activity of root caries lesions. In addition, changes in the lesion structure were monitored with polarization sensitive optical coherence tomography (PS‐OCT). Artificial bovine and natural root caries lesions were imaged with PS‐OCT, and their dehydration rate was measured with thermal and NIR cameras. The lesion activity of the natural root caries samples was also assessed by two clinicians by conventional means according to ICDAS II guidelines. The thickness of the highly mineralized transparent surface layer measured using PS‐OCT increased and the area enclosed by the time‐temperature curve, ΔQ, measured with thermal imaging decreased significantly with longer periods of remineralization in simulated dentin lesions, but the NIR reflectance intensity differences, ΔI, failed to show any significant relationship with the degree of remineralization. The PS‐OCT algorithm for the automated assessment of remineralization successfully detected the highly mineralized surface layer on both natural and simulated lesions. Thermal imaging provided the most accurate diagnosis of root caries lesion activity. These results demonstrate that thermal imaging and PS‐OCT may be ideally suited for the nondestructive root caries lesion activity during a clinical examination.

     

Cementum is key to periodontal tissue regeneration: A review on apatite microstructures for creation of novel cementum-based dental implants

The cementum is the outermost layer of hard tissue covering the dentin within the root portion of the teeth. It is the only hard tissue with a specialized structure and function that forms a part of both the teeth and periodontal tissue. As such, cementum is believed to be critical for periodontal tissue regeneration. In this review, we discuss the function and histological structure of the cementum to promote crystal engineering with a biochemical approach in cementum regenerative medicine. We review the microstructure of enamel and bone while discussing the mechanism underlying apatite crystal formation to infer the morphology of cementum apatite crystals and their complex structure with collagen fibers. Finally, the limitations of the current dental implant treatments in clinical practice are explored from the perspective of periodontal tissue regeneration. We anticipate the possibility of advancing periodontal tissue regenerative medicine via cementum regeneration using a combination of material science and biochemical methods.     

Effects of gamma radiation on hard dental tissues of albino rats: investigation by light microscopy

The present work aims at studying the effect of gamma radiation on the hard dental tissues. Eighty adult male albino rats with weights of about 250 g were used. The rats were irradiated at 0.2, 0.5, 1.0, 2.0, 4.0 and 6.0 Gy whole-body gamma doses. The effects on hard dental tissue samples were investigated after 48 h in histological and ground sections using light microscopy. Areas of acid phosphatase activity were detected using tartrate-resistant acid phosphatase (TRAP) stains. Observation of histological sections revealed disturbance in predentin thickness and odontoblastic layer as the irradiation dose increased. In cementum, widened cementocytes lacunae were occasionally detected even with low irradiated doses. On the other hand, relatively homogenous enamel was detected with darkened areas in enamel surface at doses over than 0.5 Gy. TRAP-positive cells were detected on the surface of the dentin of irradiated groups as well as cementum surface. Minimal detectable changes were observed in ground sections.     

Nuclear analytical methods for trace element studies in calcified tissues

Various nuclear analytical methods have been developed and applied to determine the elemental composition of calcified tissues (teeth and bones). Fluorine was determined by prompt gamma activation analysis through the¹⁹F(p, α ψ)¹⁶O reaction. Carbon was measured by activation analysis with He-3 ions, and the technique of Proton-Induced X-ray Emission (PIXE) was applied to simultaneously determine Ca, P, and trace elements in well-documented teeth. Dental hard tissues: enamel, dentine, cementum, and their junctions, as well as different parts of the same tissue, were examined separately. Furthermore, using a Proton Microprobe, we measured the surface distribution of F and other elements on and around carious lesions on the enamel. The depth profiles of F, and other elements, were also measured right up to the amelodentin junction.

     

Mechanical vulnerability of lower second premolar utilising visco-elastic dynamic stress analysis

Stress analysis determines vulnerability of dental tissues to external loads. Stress values depend on loading conditions, mechanical properties and constrains of structural components. The critical stress levels lead to tissue damage. The aim of this study is to analyse dynamic stress distribution of lower second premolar due to physiological cyclic loading, and dependency of pulsatile stress characteristics to visco-elastic property of dental components by finite element modelling. Results show that visco-elastic property markedly influences stress determinants in major anatomical sites including dentin, cementum–enamel and dentin–enamel junctions. Reduction of visco-elastic parameter leads to mechanical vulnerability through elevation of stress pulse amplitude, maximum stress value; and reduction of stress phase shift as a determinant of stress wave propagation. The results may be applied in situations in which visco-elasticity is reduced such as root canal therapy and post and core restoration in which teeth are more vulnerable to fracture.     

Aluminum concentrations in human deciduous enamel and dentin related to dental caries

The aluminum (Al) concentrations in the enamel and dentin of 314 human deciduous teeth were determined in order to examine the relationship between Al and dental caries. The sample teeth were divided into three groups: the sound tooth group, carious tooth group and filled tooth group. The teeth of the carious tooth group were further classified into three groups depending on the stage of caries. The Al content was determined using graphite furnace atomic absorption spectrometry. In both the enamel and dentin, the Al concentrations were unaffected by sex, but did depend on tooth type. In enamel, the Al concentration was significantly higher in the sound tooth group (42.8 +/- 37.3 microg/g) than in the three carious groups (20.7 +/- 17.1-24.9 +/- 22.0 microg/g) and the filled tooth group (27.3 +/- 25.5 microg/g). As for dentin, the Al concentration was also significantly higher in the sound tooth group (36.2 +/- 35.1 microg/g) than in the three carious groups (15.1 +/- 13.3-24.5 +/- 23.4 microg/g) and the filled tooth group (17.2 +/- 20.6 microg/g). Even when analyzing incisors alone, the Al concentrations were significantly higher in the sound tooth group than in the other groups, for both enamel and dentin. Furthermore, the Al levels in carious enamel and dentin did not decrease with the advance of caries. These findings indicated that the deciduous teeth containing higher Al concentrations on average had less caries than the teeth with lower Al concentrations, and suggest that Al acts as a possible cariostatic agent by itself.     

FAM20C Plays an Essential Role in the Formation of Murine Teeth

FAM20C is highly expressed in bone and tooth. Previously, we showed that Fam20C conditional knock-out (KO) mice manifest hypophosphatemic rickets, which highlights the crucial roles of this molecule in promoting bone formation and mediating phosphate homeostasis. In this study, we characterized the dentin, enamel, and cementum of Sox2-Cre-mediated Fam20C KO mice. The KO mice exhibited small malformed teeth, severe enamel defects, very thin dentin, less cementum than normal, and overall hypomineralization in the dental mineralized tissues. In situ hybridization and immunohistochemistry analyses revealed remarkable down-regulation of dentin matrix protein 1 (DMP1) and dentin sialophosphoprotein in odontoblasts, along with a sharply reduced expression of ameloblastin and amelotin in ameloblasts. Collectively, these data indicate that FAM20C is essential to the differentiation and mineralization of dental tissues through the regulation of molecules critical to the differentiation of tooth-formative cells.     

Determination of two different types of cellular cementogenesis in rat molars: a histological and immunohistochemical study.

To elucidate the attachment mechanism of dentin and cellular cementum, developing and developed cellular cementum of rat molars was examined by light microscopy. Routine histological staining, immunohistochemical staining for bone sialoprotein (BSP) and osteopontin (OPN), and digestion tests with trypsin were conducted. Two different types of cellular cementogenesis were established, one on the mesial (type I cementogenesis) and one on the distal sides (type II cementogenesis) of the examined roots. In the type I cementogenesis a thin initial cementum layer, which was fibril-poor, hematoxylin-stained, and immunopositive for BSP and OPN, appeared on the mineralized dentin. With cellular cementogenesis, the layer became the cemento-dentinal junction. The cementum mineralization did not precede the dentin mineralization. After trypsin treatment the cemento-dentinal junction lost immunoreactivity for BSP and OPN and the cementum was detached from the dentin. In the type II cementogenesis the cellular cementum formed directly on the predentin without the initial cementum layer and the cementum mineralization preceded the dentin mineralization. Cemental and predentinal fibrils appeared to intermingle, as the cemento-dentinal junction was indiscernible by any staining. Trypsin treatment did not cause cementum detachment. The findings of the present study suggest that: (1) The type I cementogenesis requires the intervening initial cementum to bind cementum and dentin and to induce the cementum mineralization. (2) In the type II cementogenesis the cemento-dentinal attachment depends on fibril intermingling and the cementum mineralization advances apically and very rapidly, probably producing mineralization foci. (3) The formation of the initial cementum depends on the speed of the cementogenesis in the apical direction.