Morphological and immunocytochemical analyses on the effects of diet-induced hypocalcemia on enamel maturation in the rat incisor

During the maturation stage of amelogenesis, the loss of matrix proteins co mbined with an accentuated but regulated influx of calcium and phosphate io ns into the enamel layer results in the "hardest" tissue of the body. The a im of the present investigation was to examine the effects of chronic hypoc alcemia on the maturation of enamel. Twenty-one-day old male Wistar rats we re given a calcium-free diet and deionized water for 28 days, while control animals received a normal chow. The rats were perfused with aldehyde and t he mandibular incisors were processed for histochemical and ultrastructural analyses and for postembedding colloidal gold immunolabeling with antibodi es to amelogenin, ameloblastin, and albumin. The maturation stage enamel or gan in hypocalcemic rats exhibited areas with an apparent increase in cell number and the presence of cyst-like structures. In both cases the cells ex pressed signals for ameloblastin and amelogenin. The content of the cysts w as periodic acid-Schiff- and periodic acid-silver nitrate-methanamine-posit ive and immunolabeled for amelogenin, ameloblastin, and albumin. Masses of a similar material were also found at the enamel surface in depressions of the ameloblast layer. In addition, there were accumulations of glycoprotein aceous matrix at the interface between ameloblasts and enamel. In decalcifi ed specimens, the superficial portion of the enamel matrix sometimes exhibi ted the presence of tubular crystal "ghosts." The basal lamina, normally se parating ameloblasts and enamel during the maturation stage, was missing in some areas. Enamel crystals extended within membrane invaginations at the apical surface of ameloblasts in these areas. Immunolabeling for amelogenin , ameloblastin, and albumin over enamel was variable and showed a heterogen eous distribution. In contrast, enamel in control rats exhibited a homogene ous labeling for amelogenin, a concentration of ameloblastin at the surface , and weak reactivity for albumin. These results suggest that diet-induced chronic hypocalcemia interferes with both cellular and extracellular events during enamel maturation.