Histochemical studies on the distribution of calcium in the cells related to the formation of dental and periodontal hard tissues.

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The distribution of calcium in the enamel organ, pulp and periodontal tissues was examined by means of histochemical, electron microscopic and X-ray microanalytical methods. All specimens were rapidly frozen/freeze-substituted and embedded in epoxy resin for the preservation of morphology and cell calcium. This method allowed a histochemical demonstration of cell calcium as diffuse or discrete red Ca-GBHA precipitates under the light microscope, and an electron microscopic examination of adjacent ultrathin sections.Our study revealed intense calcium reactions in the cytoplasm of both secretory and maturation ameloblasts, and its absence in other enamel organ cells. Electron microscopy and X-ray microanalytical studies confirmed the presence of calcium-containing granules and crystalline figures in the tubulovesicular structures of secretory ameloblasts, implicating the contribution of such organelles in transcellular translocation of calcium. In the pulp, Ca-GBHA reactins were shown in differentiated odontoblasts but not in other pulp cells. In molar tooth germs at the stage of root formation, Ca-GBHA reactions appeared in cementoblasts and precementoblasts located at disrupted portions of unreactive epithelial root sheath. Osteoblasts and osteoclasts on the forming and absorbing surfaces of the alveolar bone also showed intense Ca-GBHA reactions in the cytoplasm.In this project, we thus established a tissue preparation method whereby both cell morphology and unstable cell calcium is preserved. A histochemical calcium staining method was also established. With these methods, data suggesting the exclusive localization of high concentrations of calcium in cells directly related to formation or absorption of enamel, dentin or bone were presented. The elucidation of the physiological significance of present findings awaits further studies.