In-vitro study of Zn uptake in developing rat enamel

Molar explants from 8-day-old rats were cultured for up to 12h. Addition of calf serum (10 per cent) to the defined medium (M199) markedly reduced ⁶⁵Zn uptake in enamel. Metabolic inhibition (DNP) or heat killing had no apparent effect on movement of tracer through the enamel organ to the enamel. Removal of the enamel organ prior to culture resulted in increased uptake of ⁶⁵Zn in the enamel.     

The in vitro uptake of fluoride by secretory and maturation stage bovine enamel

The objectives of this study were to determine the specific surface area of secretory-stage and of maturation-stage enamel, to compare the fluoride uptake by isolated enamel at these two stages on a surface-area basis, and to examine the effect of the organic matrix on the fluoride uptake by whole enamel. Fetal bovine secretory and maturation stage enamel samples were collected, and a portion of the enamel at each developmental stage was treated with hydrazine for removal of the organic matrix. The specific surface areas of the enamel mineral, as determined by the multi-point BET method, were 59.3 m2/g in the secretory stage and 37.9 m2/g in the maturation stage. Whole and deproteinated enamel samples were equilibrated in buffered solutions containing 10(-5) to 10(-3) mol/L fluoride, and the uptake was measured with a fluoride specific electrode. The results indicate that the in vitro fluoride uptake was controlled solely by the surface area of the apatitic mineral and that the organic matrix did not contribute to the fluoride uptake.     

In vivo and in vitro study of 48V uptake in developing rat molar enamel

The uptake of 48V in developing rat molar enamel was studied in vivo and in vitro using autoradiographic and quantitative methods. It was shown that the uptake of 48V in enamel is less than in bone and dentin, and that plasma protein binding of 48V is primarily responsible for the limited uptake in enamel.     

In-vitro study of the cellular control of 18F uptake in the enamel of developing rat molar teeth

Molar expiants from 6-day-old rats were cultured for 4 h in medium containing approx. 2μCi/ml¹⁸F. Control-cultured expiants were compared to their contralateral pairs which had been subjected to: (1) the enamel organ removed prior to incubation; (2) the expiants heat-treated (65 °C for 10 min); or (3) a metabolic inhibitor, 5mM dinitrophenol (DNP), added to the medium. The Ca concentration of the medium indicated that the expiants became mineralized during incubation. Both removal of enamel organ and heat-treatment enhanced uptake of ¹⁸F from the medium. DNP had no consistent effect. As the pattern of ¹⁸F uptake was similar to that for net Ca uptake by mineralizing enamel, the findings suggest that there is no direct cellular control of F^? flux through the enamel organ during the secretory phase of enamel formation.     

In-vitro study of calcium-45 and phosphorus-32 uptake in developing rat molar enamel using quantitative methods

In-vitro quantitative studies were conducted on rat molar tooth expiants to determine the effect of the enamel organ on enamel mineralization during the secretory phase of formation. Control explants were compared to contralateral pairs which had (1) the enamel organ removed, (2) heat-treatment, or (3) treatment with the metabolic inhibitor dinitrophenol (DNP). Removal of the enamel organ enhanced enamel mineralization and ⁴⁵Ca exchange with the enamel. Heat-treatment altered the enamel matrix to increase Ca uptake. DNP inhibited mineralization but increased ⁴⁵Ca exchange through the enamel organ. The findings suggest that the enamel organ controls Ca flux into the forming enamel during the secretory phase and slows the rate of mineralization.     

Fluoride uptake and retention at various stages of rat molar enamel development

Suckling rat pups were given intraperitoneal fluoride injections at selected ages so that we could study fluoride uptake in the enamel of the maxillary first molar at various stages of enamel development. Plasma fluoride levels in six-day-old and 11-day-old pups were monitored following the intraperitoneal injection of fluoride. The findings indicate that: (1) fluoride was more easily taken up and retained during the early stages of enamel formation, but fluoride uptake can occur during all stages of enamel formation; (2) when injections were started early in enamel formation, more fluoride was contained in the enamel of the maxillary first molar at 13 days of age; and (3) the same dose of fluoride per gram body weight resulted in greater exposure to elevated plasma fluoride levels in six-day-old pups than in 11-day-old pups.     

Appearance of freeze-fractured rat incisor enamel during the forming stage

On replicas of freeze-fractured fixed early forming enamel, differences were noted between newly-secreted areas and older ones. Fibres which were first organized as 30–50 nm aligned particles become more sophisticated structures constituted of 10 nm particles aggregated in 3 or 4 rows. This might reflect the degradation of the initially synthesized high molecular-weight species.     

Effect of turpentine-induced fever during the enamel formation of rat incisor

OBJECTIVE: Some epidemiological studies have indicated that diseases resulting in prolonged and sustained fever, such as exanthemata, respiratory infections and otitis media in infantile period or childhood are likely to have a marked deleterious effect on enamel formation, but the relationship between fever and enamel defects is unknown. The purpose of the present study was to induce a persistent high fever and examine the effects on the developing tooth enamel. METHODS: Twenty male Wistar rats weighting 140+/-10 g were used in this study. For the experimental group, a dose of 2.3 ml/kg steam-distilled turpentine was subcutaneously injected into both hind limbs five times at 12h intervals. Control rats received 2.3 ml/kg of sterile saline into the same injection site. The rectal temperatures of animals were measured at the febrile period. After constant periods, the animals were sacrificed, and the mandibular incisors were examined by contact microradiography (CMR) and histological observation. RESULTS: The febrile state lasted for 57 h and the average temperature rose 1.51 degrees C higher than that of the control group. The ground sections, semi-thin and ultra-thin sections of mandibular incisors were prepared and the enamel was observed. The microradiographs showed a radiolucent line along with the incremental line in the enamel. Moreover, microscopic examination indicated disorientation of enamel prism and crystal-free area within this radiolucent lesion. CONCLUSIONS: Persistent high fever pattern was established firmly by the turpentine injections and the process of enamel formation was influenced by the febrile period.     

Ultrastructure of fluoride-induced cysts in the rat molar enamel organ

Abstract — Earlier studies have shown that a single high dose of fluoride induces subameloblastic cysts separated by morphologically unaltered ameloblasts in the developing rat molar. In the present investigation, the ultrastnucture of both the ameloblasts forming the cystic wall, and the cells within the cystic lumina were studied by means of transmission electron microscopy. It was shown that 24 h after fluoride injection the ameloblasts of the cystic wall showed various degrees of cytopiasmic and nuclear alterations. Some cells displayed signs of necrosis as indicated by condensation of the chromatin. The cytopiasmic changes varied from altered organelle morphology to fragmentation and almost complete shedding of the whole cytoplasm. In the ameloblasts of the cystic wall secretory products accumulated intracellularly, in distended rough endoplasmatic reticulum, in vesicles of the Golgi region and extracellularly between ameioblasts as well as between cells in the stratum intermedium, indicating an altered matrix secretion. Electron lucent material, cell and cell fragments were found in the cystic lumina, the two latter apparently originating from the ameloblastic layer. The degenerative changes seemed to follow the normal pattern of cell degeneration.     

Ultrastructure of fluoride-induced cysts in the rat molar enamel organ

Earlier studies have shown that a single high dose of fluoride induces subameloblastic cysts separated by morphologically unaltered ameloblasts in the developing rat molar. In the present investigation, the ultrastructure of both the ameloblasts forming the cystic wall, and the cells within the cystic lumina were studied by means of transmission electron microscopy. It was shown that 24 h after fluoride injection the ameloblasts of the cystic wall showed various degrees of cytoplasmic and nuclear alterations. Some cells displayed signs of necrosis as indicated by condensation of the chromatin. The cytoplasmic changes varied from altered organelle morphology to fragmentation and almost complete shedding of the whole cytoplasm. In the ameloblasts of the cystic wall secretory products accumulated intracellularly, in distended rough endoplasmatic reticulum, in vesicles of the Golgi region and extracellularly between ameloblasts as well as between cells in the stratum intermedium, indicating an altered matrix secretion. Electron lucent material, cell and cell fragments were found in the cystic lumina, the two latter apparently originating from the ameloblastic layer. The degenerative changes seemed to follow the normal pattern of cell degeneration.     

Expression, localisation and synthesis of versican by the enamel organ of developing mouse molar tooth germ: An in vivo and in vitro study

Objective

Versican is a large, aggregating chondroitin sulphate proteoglycan. In dental tissue, versican expression occurs primarily in mesenchymal tissue but rarely in epithelial tissue. We investigated the expression, localisation and synthesis of versican in the enamel organ of the developing tooth germ.

Design

To elucidate versican localisation in vivo, in situ hybridisation and immunohistochemistry were conducted in foetal ICR mice at E11.5–E18.5. Epithelium and mesenchyme from the lower first molars at E16.0 were enzymatically separated and versican mRNA expression was investigated by semi-quantitative RT-PCR. Organ culture of the separated samples combined with metabolic labelling with [³⁵S], followed by gel filtration, was performed to analyse secreted proteoglycans.

Results

Versican mRNA was first expressed in the thickened dental epithelium at E12.0 and continued to be expressed in the enamel organ until the bell stage. Versican immunostaining was detected in the stellate reticulum areas from the bud stage to the apposition stage. The enamel organ at E16.0 expressed versican mRNA at a level comparable to that in dental mesenchyme. Furthermore, when compared to dental mesenchyme, about 1/2–3/4 of the [³⁵S]-labelled versican-like large proteoglycan was synthesised and released into tissue explants by the enamel organ.

Conclusions

The dental epithelium of developing tooth germ is able to synthesise significant amounts of versican.     

An autoradiographic study of calcium movement in the enamel organ of rat molar tooth germs

The distribution and movement of calcium through the enamel organ and into the forming enamel was studied in 6-day-old rats, intravenously injected with 45Ca. To prevent dislocation of radiocalcium in the specimens, the tooth germs were rapidly frozen/freeze-substituted and processed for 45Ca autoradiography under dry conditions. At 30 s after the 45Ca injection, there was a decrease in labelling intensity progressing from the overlying connective tissue to the enamel organ and, in the secretory ameloblasts, from the proximal to distal cytoplasm. The most intense labelling was in the enamel matrix, where it was restricted to the superficial layer extending approx. 15 microns below the surface. At later times the density of silver grains over the connective tissue decreased considerably, whereas secretory ameloblasts showed an increasing intensity in the distal portions. Enamel had the heaviest labelling: the width of the labelled enamel increased gradually to only 40 microns from the surface 60 min after the injection. The use of wet emulsion over similarly prepared sections caused a severe dislocation of radiocalcium in the specimens. These findings confirm the rapid penetration of systemically administered calcium to newly formed enamel, probably due to isotopic exchange. A relatively slow diffusion through the enamel organ and into the surface layer of enamel suggests that net transport of calcium through the enamel organ is transcellular.     

Fluoride-induced cystic changes in the enamel organ of the rat molar

Many substances have been found to cause enamel disturbances in toxic doses, and it has been postulated that these disturbances are linked to the formation of sub-ameloblastic cysts. In the present investigation, fluoride-induced sub-ameloblastic cysts in developing rat molars were related to fluoride dose, age of the animals, and the plasma fluoride level. The sub-ameloblastic cysts, which developed predominantly towards the end of the secretory stage of amelogenesis, appeared shortly after fluoride administration and regressed within 3 days. Hypoplasias and internal defects were found in the enamel under the disturbed ameloblast layer. The highest plasma fluoride levels were found in the youngest animals 24 h after injection. The frequency and size of the sub-ameloblastic cysts were clearly related to the fluoride-dose and age of the animal and, subsequently, to the plasma fluoride level.     

Fluoride-induced cystic changes in the enamel organ of the rat molar

Many substances have been found to cause enamel disturbances in toxic doses, and it has been postulated that these disturbances are linked to the formation of sub-ameloblastic cysts. In the present investigation, fluoride-induced sub-ameloblastic cysts in developing rat molars were related to fluoride dose, age of the animals, and the plasma fluoride level. The sub-ameloblastic cysts, which developed predominantly towards the end of the secretory stage of amelogenesis, appeared shortly after fluoride administration and regressed within 3 days. Hypoplasias and internal defects were found in the enamel under the disturbed ameloblast layer. The highest plasma fluoride levels were found in the youngest animals 24 h after injection. The frequency and size of the sub-ameloblastic cysts were clearly related to the fluoride-dose and age of the animal and, subsequently, to the plasma fluoride level.