Effects of actinomycin D on developing hamster molar tooth germs in vitro.

The aim of this study was to evaluate the toxic effects of actinomycin D on the developing hamster tooth germ in organ culture. Hamster tooth germs during early secretory amelogenesis were exposed in vitro for 24 h to 10(-9) M-5 x 10(-5) M actinomycin D. Actinomycin D dose-dependently (> or = 10(-7) M) decreased the tooth germ dry weight but mineralization was affected only by doses > or = 10(-5) M. However, the uptakes of TCA-insoluble 32P and [3H]thymidine were significantly reduced dose-dependently from > or = 10(-8) M actinomycin D, indicating that the drug inhibits the synthesis of phosphate-containing macromolecules as well as DNA synthesis. Histologically, 10(-8) M actinomycin D was the lowest dose which was not toxic to any cell type in the developing tooth germ. At 10(-7) M actinomycin D, the most sensitive cells were the proliferating pre-odontoblasts followed by pre-ameloblasts; the mature secretory ameloblasts and odontoblasts appeared unaffected. Higher doses resulted in increased cytotoxicity to the secretory cells and, eventually, total degeneration of most cells. The data suggest that children treated for cancer during tooth development using anti-chemotherapy cocktails containing actinomycin D (serum levels > 10(-7) M) may develop defects later on in the mature dentition as a direct consequence of the toxicity of the drug to the tooth organ.

Effects of vincristine on the developing hamster tooth germ in vitro.

Vincristine is one of the cytostatic drugs present in cocktails commonly used for the treatment of cancer in children. The aim of this study was to evaluate biochemically and histologically the toxic effects of this drug on the developing tooth in vitro using the organ culture model in order to be able to predict what damage the drug can induce in the developing teeth from children undergoing anti-neoplastic chemotherapy. The most profound effect of the drug (10(-8)M-10(-4)M vincristine) on the developing tooth germ was the induction of mitotic arrests at the cervical loop and in the inter-cuspal regions. The 10(-4)M-10(-6)M vincristine doses were cytotoxic to most cells in the developing tooth germ. The 10(-7)M vincristine dose apart from induction of mitotic arrests, did not appear to be cytotoxic to the mature differentiated secretory cells. However, this dose induced incomplete nuclear polarization of the differentiating ameloblasts and odontoblasts. At 10(-8)M vincristine, the only effect observed were mitotic arrests; the secretory cells did not appear to have been affected at all. On the other hand, mineralization (TCA-soluble 45Ca and 32P uptake) was dose-dependently decreased from 10(-7)M vincristine upwards. 10(-9)M vincristine, the lowest dose tested, did not induce any changes in the developing tooth germ. The organ culture data indicate that 10(-9)M vincristine is the highest (safe) dose which does not induce any toxic effects in the developing hamster tooth germ.(ABSTRACT TRUNCATED AT 250 WORDS)

Reversible and irreversible effects of temperature on amelogenesis of hamster tooth germs in vitro.

Hamster first hamster molar tooth germs in early secretory stage of amelogenesis were cultured for one day in vitro at 6 degrees C, 22 degrees C, 37 degrees C or 45 degrees C in the presence of 3H-proline, 45Ca and 32P-orthophosphate. Other explants were cultured without these labels and after culture examined by histology. The highest temperature tested was lethal to the explants, decreased total dry weight and rapidly increased total uptake of the radiolabelled mineral ions, probably merely due to physicochemical modification of the existing preculture minerals. Optimal synthesis and secretion of amelogenins were measured at physiological temperature (37 degrees C). Effects of exposure to both temperatures below the physiological value were virtually reversible when explants were grown at physiological temperature (37 degrees C) for another day. However, amelogenin secretion during this recovery period did not reach values as high as those found for the first day in explants initially grown at physiological temperature during the first day. We concluded from the four temperatures examined that the optimal temperature for enamel matrix deposition in vitro was 37 degrees C. At this temperature enamel biosynthesis and its secretion are high. Lowering the temperature slows down the metabolism without any apparent harmful effect. Normal development of the tooth explants in vitro resumes when the culture temperature is restored to physiological levels (37 degrees C). For temporary storage of tooth germ explants prior to any reimplantation, we therefore recommend a temperature of 6 degrees C.

Possible functions of alkaline phosphatase in dental mineralization: cadmium effects.

In mineralizing dental tissues the non-specific alkaline phosphatase, using paranitrophenylphosphate (p-NPP) as substrate, is also capable of splitting inorganic pyrophosphate (PPi). In contrast to the p-NPP-ase part of the enzyme, the PPi-ase part requires Zn2+ as a cofactor for its hydrolytic activity. The PPi-ase activity of the enzyme can be inhibited by cadmium ions (Cd2+), perhaps by replacing Zn2+ from the active site of the enzyme molecule. In addition to splitting PPi, the PPi-ase part of the enzyme may also be involved in the phosphorylation process of yet undetermined organic macromolecules. Cd2+ inhibits this phosphorylation process. Inhibition of the PPi-ase activity can also be accomplished by ascorbic acid known for its capacity to complex bivalent cations. Ascorbic acid may accordingly also remove Zn2+ from the active site of the PPi-ase. It is suggested that in developing dental tissues alkaline phosphatase is not only associated with the transport of phosphate ions towards the mineralization front, but is also involved in the phosphorylation of organic macromolecules, a process activated the PPi-ase part of the enzyme.

Correlation between some salivary tests, oral hygiene and caries in children.

gn 137 children whose ages varied from 2-14 years, salivary tests and oral hygiene levels were determined for boys and girls separately. Only the unstimulated flow (flow 1) differed significantly between boys and girls. For the initiation and further development of new caries lesions more precise definitions were used in which the results were corrected for number of teeth present and for the time interval between the observations. The caries scores for all children together were correlated with the several salivary test results as well as with oral hygiene levels. The initiation of caries was correlated with oral hygiene as well as with the Snyder test. The further development of caries correlated almost significantly with the inorganic composition of the saliva alone. No correlation was found between the initiation and the further development of caries.

Remineralization in human premolars of different posteruptive age.

In human premolars with posteruptive ages varying from 6 months to 4 years demineralized enamel was remineralized for 3 days at 37 degrees C in a recalcifying medium containing 2 mM Ca2+, 1.2 mM phosphate, 0.05 mMF- and 150 mM NaCl (pH = 7). The Ca, P and Ca/P molar ratio of the mineral gained were determined at various distances from the enamel surface. Whereas, particularly at 30 micrometers from the surface remineralization was higher and more extensive in young demineralized enamel, no difference could be found in Ca/P molar ratio, suggesting precipitation of an apatitic mineral in both young and old demineralized human enamel. The probable nature of the precipitate is that of a fluoridated hydroxyapatite.

Changes in calcium and phosphorus content in young human enamel during demineralization in vitro.

Premolars extracted about 6 months after eruption were divided into four parts by sectioning through the mesial- and distal midplanes. The enamel of each part was subjected to demineralization by hydroxy-ethyl-cellulose (H.E.C.) gel at pH 4 for periods varying between 2 and 24 hrs. The Ca and P contents of the gel as well as that of 10 microns slices prepared from the enamel after demineralization were determined. Then the Ca and P content of the HEC gel as well as that of 10 microns slices sequentially sectioned from the surface of the demineralized enamel inwards, were determined. The data indicate that initially, demineralization was predominant in the layers between 20 and 60 microns depth together with a reprecipitation in the surface 20 microns layer. After about 6 h of demineralization more mineral loss was found from layers deeper than 60 microns. Further, it was shown that the molar Ca/P ratio of mineral loss from these young teeth at depths of between 40 and 50 microns was extremely high (about 3) whereas in older teeth it was normal. This might be due either to preferential dissolution of carbonate containing mineral or to preferential precipitation of HPO4(2)--containing mineral or both especially at that depth.

Organ culture of tooth germs: relationship between alkaline phosphatase and mineralization in vitro.

In the 3rd day 2nd molar of the hamster, mineralization of the tooth germ has been studied in vitro starting from unmineralized predentine stage up to secretory amelogenesis. The explants were cultured in BGJb medium containing 15% foetal calf serum. The biochemical data on whole tooth germs were compared morphometrically with its main calcifying components. The alkaline phosphatase seems not to be related to cell-division but more to mineralization. There the alkaline phosphatase appeared to be more correlated with the phosphorus than with the calcium uptake. Its function in mineralization of the tooth germ seems also to be more expressed in dentinogenesis than in amelogenesis.