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The use of the stable isotope 44Ca in studies of calcium incorporation into dentin.

The incorporation into rat incisor dentin of two calcium isotopes, the stable 44Ca and the radioactive 45Ca, was studied using secondary ion mass spectrometry (SIMS) step-scanning and imaging, and autoradiography, respectively. The results demonstrated a time-dependent incorporation of the calcium isotopes into the mineral phase of dentin. With the SIMS step-scanning, detecting 44Ca, the ion yield was high in the odontoblasts 2 min after intravenous injection. After 10 min a marked increase in signal intensity was found at the dentin mineralization front. This result was consistent with those obtained by 45Ca autoradiography; a peak of incorporation occurred 10 min after injection of the isotope. Likewise, localization of 44Ca to the mineralization front could be demonstrated 10 min after injection by SIMS imaging. In images obtained at earlier intervals, no such increase in ion yield could be detected. The results show that the nonradioactive, stable isotope 44Ca can be used as a marker for biomineralization in a similar way to radioactive 45Ca.

Animals↗

The resistance of superficially sealed enamel to wear and carious attack in vitro.

The resistance to wear and carious attack has been investigated in vitro for enamel that has been subjected to several stages of the sealing procedure: etching, etching and remineralization in whole saliva and etching followed by sealing. After partial mechanical removal of the sealant, the remaining surface enamel proved to be resistant to carious attack as long as the resin tags were present. An estimation is made of the thickness of the affected layer. Pretreatment with monofluorophosphate has proved to influence the extent of the histological changes due to etching. A discussion on these phenomena is given.

Acid Etching, Dental↗

TGF-beta3 induces ectopic mineralization in fetal mouse dental pulp during tooth germ development.

Several members of the transforming growth factor (TGF)-beta superfamily are expressed in developing teeth from the initiation stage through adulthood. Of those, TGF-beta1 regulates odontoblast differentiation and dentin extracellular matrix synthesis. However, the molecular mechanism of TGF-beta3 in dental pulp cells is not clearly understood. In the present study, beads soaked with human recombinant TGF-beta3 induced ectopic mineralization in dental pulp from fetal mouse tooth germ samples, which increased in a dose-dependent manner. Further, TGF-beta3 promoted mRNA expression, and increased protein levels of osteocalcin (OCN) and type I collagen (COL I) in dental pulp cells. We also observed that the expression of dentin sialophosphoprotein and dentin matrix protein 1 was induced by TGF-beta3 in primary cultured dental pulp cells, however, not in calvaria osteoblasts, whereas OCN, osteopontin and osteonectin expression was increased after treatment with TGF-beta3 in both dental pulp cells and calvaria osteoblasts. Dentin sialoprotein was also partially detected in the vicinity of TGF-beta3 soaked beads in vivo. These results indicate for the first time that TGF-beta3 induces ectopic mineralization through upregulation of OCN and COL I expression in dental pulp cells, and may regulate the differentiation of dental pulp stem cells to odontoblasts.

Animals↗

Reconstructing impairment of secretory ameloblast function in porcine teeth by analysis of morphological alterations in dental enamel.

We studied the relationship between the macroscopic appearance of hypoplastic defects in the dental enamel of wild boar and domestic pigs, and microstructural enamel changes, at both the light and the scanning electron microscopic levels. Deviations from normal enamel microstructure were used to reconstruct the functional and related morphological changes of the secretory ameloblasts caused by the action of stress factors during amelogenesis. The deduced reaction pattern of the secretory ameloblasts can be grouped in a sequence of increasingly severe impairments of cell function. The reactions ranged from a slight enhancement of the periodicity of enamel matrix secretion, over a temporary reduction in the amount of secreted enamel matrix, with reduction of the distal portion of the Tomes' process, to either a temporary or a definite cessation of matrix formation. The results demonstrate that analysis of structural changes in dental enamel allows a detailed reconstruction of the reaction of secretory ameloblasts to stress events, enabling an assessment of duration and intensity of these events. Analysing the deviations from normal enamel microstructure provides a deeper insight into the cellular changes underlying the formation of hypoplastic enamel defects than can be achieved by mere inspection of tooth surface characteristics alone.

Ameloblasts↗

In vitro studies on periodontal ligament cells and enamel matrix derivative.

The recognition that periodontal regeneration can be achieved has resulted in increased efforts focused on understanding the mechanisms and factors required for restoring periodontal tissues so that clinical outcomes of such therapies are more predictable than those currently being used. In vitro models provide an excellent procedure for providing clues as to the mechanisms that may be required for regeneration of tissues. The investigations here were targeted at determining the ability of enamel matrix derivative (EMD) to influence specific properties of periodontal ligament cells in vitro. Properties of cells examined included migration, attachment, proliferation, biosynthetic activity and mineral nodule formation. Immunoassays were done to determine whether or not EMD retained known polypeptide factors. Results demonstrated that EMD under in vitro conditions formed protein aggregates, thereby providing a unique environment for cell-matrix interaction. Under these conditions, EMD: (a) enhanced proliferation of PDL cells, but not of epithelial cells; (b) increased total protein production by PDL cells; (c) promoted mineral nodule formation of PDL cells, as assayed by von Kossa staining; (d) had no significant effect on migration or attachment and spreading of cells within the limits of the assay systems used here. Next, EMD was screened for possible presence of specific molecules including: GM-CSF, calbindin D, EGF, fibronectin, bFGF, gamma-interferon, IL-1 beta, 2, 3, 6; IGF-1,2; NGF, PDGF, TNF, TGF beta. With immunoassays used, none of these molecules were identified in EMD. These in vitro studies support the concept that EMD can act as a positive matrix for cells at a regenerative site.

Calbindins↗

Effects of bicarbonate on remineralization of enamel.

Enamel surfaces were demineralized in acetic acid and remineralized in solutions that contained 1.5 mM calcium, 1.0 mM phosphate, and 0.0, 5, 15, and 25 mM NaHCO3, and 0.00 or 5.0 mM Naf at pH 7.0. NaF in CO2-free solutions increased the rates of calcium and phosphate uptake during remineralization. Bicarbonate in NaF-free solutions caused small increases of calcium and phosphate uptake. Bicarbonate with NaF in solutions synergistically increased calcium uptake, but did not affect phosphate uptake. Bicarbonate reduced fluoride uptake by about 50% during remineralization, and increased the Ca/P ratios of deposited minerals from about 1 (dicalcium phosphate) to approach and exceed the ratios of apatites.

Animals↗

The carbonate and fluoride in surfaces of remineralized enamel.

Analyses were made of samples of surface enamel collected before and after acid demineralization and after subsequent remineralization in solutions containing calcium and phosphate and various levels of bicarbonate and fluoride ions. Demineralization caused a preferential loss of carbonates. Remineralization was increased by both fluoride and bicarbonate ions. With no bicarbonate in the calcifying solution, 0.25 to 4.0 mM NaF increased mineral formation and thereby caused a relative reduction of its carbonate content. With 0.5 mM NaF (10 ppm F-) and 5, 15, and 25 mM HCO-3 in remineralizing solutions more carbonate was deposited than in the absence of NaF. Five and 15 mM HCO-3 increased, but 25 mM HCO-3 decreased, deposit of fluoride in enamel.

Acetates↗

Enamel remineralization as a factor in the pathogenesis of dental caries.

This paper described an investigation of local resistance acquired in response to cariogenic attacks and remineralization periods from environmental fluids. Surfaces from extracted human teeth with yellow and brown areas indicative of consolidated carious lesions were exposed to acid buffers. Subsurface lesions were developed on areas of sound enamel adjacent to, but not in areas of, consolidated lesions. The development of higher tooth resistance to acid through demineralization and remineralization was demonstrated experimentally on bovine enamel presoftened in acid, treated with fluoride, exposed to the oral environment, and finally exposed to acid buffers for development of subsurface lesions. These results are interpreted as an extension of theory of tissue adaptation to an injurious challenge, in that the tooth surface is primed by the challenge for possible remineralization and the development of higher resistance. This, cariogenic challenges can result in either local adaptation of the enamel to the challenge through lesion consolidation or to open cavitation. As in any biological adaptation, the determining factors for the final outcome are: (a) the frequency and duration of the challenge, (b) the frequency and duration of remineralizing conditions, and (c) the composition of the remineralizing fluid. The implications of the theory in the pathogenesis of caries are discussed in the light of clinical and laboratory investigations.

Animals↗

Remineralization of softened human enamel in mucin- or CMC-containing artificial salivas.

The rehardening properties of four saliva substitutes on artificially softened human enamel have been investigated by microhardness measurements. The saliva substitutes were all based on the same formula, containing calcium, phosphate and fluoride as the main electrolytes, and mucin or carboxymethylcellulose as the main macromolecules. It has been shown that a rehardening potential exists in the saliva substitutes when calcium and phosphate are present. Omitting F- greatly reduces the rehardening potential. The rehardening is better in case of the CMC-containing saliva than in that containing mucin.

Carboxymethylcellulose Sodium↗

Ultrastructure of altered rat enamel beneath fluoride-induced cysts.

The effect of a single injection of sodium fluoride (60 mg/kg) on the development of rat molar enamel beneath fluoride-induced subameloblastic cysts was studied by transmission electron microscopy using undecalcified sections. Three bands of altered enamel were identified and defined as the cyst surface band, the hypoplastic band, and the hypercalcified band. The irregular cyst surface band, not previously described, was found to have two components: electron-dense enamel globules and organic spherules. The electron-dense globules consisted of small, randomly arranged crystals (confirmed by selected area electron diffraction) occurring within a stippled organic matrix. The organic spherules have staining properties similar to stippled material and lack a crystalline component. They may be a form of organic material being extruded from the underlying developing enamel. The critical role of normal matrix production and ameloblast Tomes' process structure on the development of the crystal orientation and rod pattern is discussed.

Ameloblasts↗

Amelogenesis imperfecta: a scanning electron microscopic and microradiographic study.

The aim of the present study was to use scanning electron microscopy (SEM) to visualize the morphology of the enamel surface in 12 primary teeth from children with amelogenesis imperfecta (AI). The observations were correlated to genetic, clinical and microradiographic data from the same teeth and to non-affected control teeth. SEM showed similar disturbances in teeth with a clinical predominance of hypoplasias and in teeth with a predominance of hypomineralization. In the microradiographs the enamel of most teeth showed both hypoplasias and areas of hypomineralization, independently of the predominant clinical manifestation. In the one boy with an X-linked inheritance pattern, both SEM and microradiography showed the morphology of the enamel to be unique in the present study. In the other teeth, similar manifestations were found in cases with AI as an AD trait and in the sporadic cases.

Amelogenesis Imperfecta↗

Cyclosporin A-induced alterations of dentinogenesis in rat molars.

Cyclosporin A (CsA), a widely used immunosuppressive drug, induces gingival overgrowth and modifications of bone remodelling. The scope of this study was to investigate the possible effect of CsA on dentin. Thirty mg/kg/day of CsA were administered orally to male Sprague-Dawley rats for nineteen weeks. The same number of control rats received oil-based vehicle solution. Rats were anesthetized, and tissues were fixed by an intracardiac perfusion of fixative solution. Mandibles were dissected, demineralized, and processed for Epon embedding. Semi-thin sections of the first molars revealed alterations at the secondary dentin-pulp interface in four out of six experimental animals. The changes consisted of the formation of: 1) osteodentin spurs, in which the volume and interface with the secondary dentin varied from about 25,000 to 75,000 microns 3 and from 1400 to 3530 microns 2, respectively; 2) abnormally shaped and irregularly spaced incremental lines; and 3) numerous globular formations embedded in dentin or free in the pulp. These results indicate that CsA induces abnormal mineralized matrix formation in dentin and in the peripheral part of the pulp in rat molars.

Administration, Oral↗