Yasuko Nagai

Apoptosis in mouse taste buds after denervation.

Abstract.Apoptotic cells in the taste buds of mouse circumvallate papillae after the sectioning of bilateral glossopharyngeal nerves were examined by the method of DNA nick-end labeling (TUNEL), together with standard electron microscopy. The taste buds decreased in number and size 3–11 days after denervation and disappeared at 11 days. The TUNEL method revealed only a few positively stained nuclei in normal taste buds but, in those of mice 1–5 days after denervation, the number of positive nuclei had increased to 3–5 times that of taste buds from normal mice. Electron-microscopic observation after denervation demonstrated taste bud cells containing condensed and fragmentary nuclei in a cytoplasm with increased density. The results show that taste bud cells under normal conditions die by apoptosis at the end of their life span, and that gustatory nerve sectioning causes apoptosis of taste bud cells with taste buds decreasing in number and ultimately disappearing.

Expression of E- and P-cadherin during tooth morphogenesis and cytodifferentiation of ameloblasts

Abstract Cell-cell adhesion is fundamental in morphogenesis and is known to be mediated by several groups of cell adhesion molecules. Cadherins are a group of such molecules involved in the Ca2+-dependent cell-cell adhesion mechanism and are found in most kinds of tissue. In this study using indirect immunofluorescence microscopy, we analyzed the distribution of two kinds of cadherins, E- and P-cadherin, in developing tooth germs. In the molar tooth germs at the early bud stage, marginal cells of the epithelial tooth bud expressed both E- and P-cadherin, whereas central cells expressed only E-cadherin. At the cap stage, in addition to the cells of the inner and outer enamel epithelium, which outline the enamal organ, cells of the enamel knot, which is thought to control tooth morphogenesis, strongly expressed P-cadherin. The expression of P-cadherin was prominent in the inner enamel epithelium during the early to mid bell stage, and was also evident in the non-dividing cell masses at future cusp tips, which are the so-called secondary enamel knots. In the tooth germ at the late bell stage when the cells of the inner enamel epithelium began to polarize to differentiate into ameloblasts, the polarizing ameloblasts lost P-cadherin and strongly expressed E-cadherin. However, E-cadherin was also lost from polarized ameloblasts at later stages. The stratum intermedium and the stellate reticulum were E-cadherin positive from the bell stage onward even at the stages when the ameloblasts became E-cadherin negative again. These results suggest that the differential expression of E- and P-cadherin during morphogenetic stages plays a role in the regulation of tooth morphogenesis, whereas alteration of E-cadherin expression during later stages of tooth development is related to differentiation and function of the ameloblasts and other cells supporting amelogenesis.

Expression of neural cell-adhesion molecule mRNA during mouse molar tooth development.

This study employed in situ hybridisation using a probe recognising all isoforms of the molecule. Expression of the molecule in tooth germs started at embryonic day 13, when they were at the bud stage. Both inner cells of the epithelial bud and peripheral cells of the dental mesenchyme were positive. At the cap stage, positive cells were found in the inner part of the enamel organ but only in a limited area near the outer enamel epithelium. In the mesenchyme at the cap stage, expression was weak in the dental papilla and strong in the follicle. From the bell stage onward, epithelial cells in the enamel organ were negative except for the cells of the stratum intermedium, which were transiently positive at early and late bell stages. In the dental papilla, expression had mostly ceased during and after the bell stage, although transient expression was found in cuspal areas at the early bell stage. The dental follicle strongly expressed neural cell-adhesion molecule (NCAM) to the end of the experimental period, at post-natal day 4. In contrast to the first molar at its earliest stage of appearance, in which both the thickened epithelium and surrounding mesenchyme were negative for the expression of the molecule, the second molar appeared as a combination of extending epithelial thickenings and mesenchymal cells strongly positive for its expression. This study newly identifies the dental papilla and the stratum intermedium as NCAM-expressing sites.

Induction of apoptosis by colchicine in taste bud and epithelial cells of the mouse circumvallate papillae.

Abstract. Apoptotic cells in the taste buds and epithelia of mouse circumvallate papillae after colchicine treatment were examined by the methods of in situ DNA nick-end labeling, immunocytochemistry, and electron microscopy. After colchicine treatment, numerous positive cells appeared in the taste buds by DNA nick-end labeling, and some epithelial cells in the basal and suprabasal layers in and around the circumvallate papillae also revealed positive staining. Condensed and fragmented nuclei with a high density were occasionally found in the taste bud cells and in the basal and suprabasal layer epithelial cells by electron-microscopic observation. An immunocytochemical reaction for tubulin revealed weak staining in taste bud cells, because of the depolymerization of microtubules, and a decrease of the microtubules in the taste bud cells was observed by electron microscopy. These results indicate that colchicine treatment of mice induces the apoptosis of taste bud and epithelial cells in the circumvallate papillae and dorsal epithelial cells around the circumvallate papillae.

Immunofluorescence detection of cadherins in mouse tooth germs during root development

The distribution of two cell-adhesion molecules, E- and P-cadherin, was studied in relation to morphological changes in Hertwigs epithelial root sheath Before root dentinogenesis had started, the root sheath expressed both cadherins. As dentinogenesis proceeded, the sheath fragmented and lost P-cadherin rapidly and E-cadherin slowly, whereas the intact sheath at the apical end continued to express both. These results suggest that the two cadherins play a part in root as well as in crown development, and indicate that the decrease in the amount of these molecules and the fragmentation of the epithelial root sheath are interrelated.

A comparative study of an olfactory epithelial area lacking olfactory neurons and a nearby presence of TGF-α-like immunoreactive olfactory neurons

Our previous study has shown that ddY mice have special patches of nasal epithelium in the posterior roof of the nasal cavity that exclusively consists of olfactory supporting cells and horizontal basal cells. Here, we extend this finding to Balb/c and DBA/2 mice, Wistar and Sprague-Dawley rats, hamsters, and guinea pigs. In the mice, rats, and hamsters studied, the patches lacked olfactory cells and their precursor, globose basal cells. In rats and hamsters, the supporting cells were arranged in a single layer, in mice as three or four layers. Horizontal basal cells were located in a single layer in these species. In the guinea pigs, the specialized roof structure was less clear and could be seen at the level of ultrastructure as an olfactory neuron-lacking area. Distinct populations of transforming growth factor (TGF)-α-like immunoreactive olfactory cells occupied an area close to the epithelial patches. In this region, the TGF-α-like immunoreactive neurons were negative for the usual olfactory markers, either OMP or protein gene product (PGP) 9.5 or β-tubulin. These cells are suggested to project to the so-called ’necklace glomeruli’ and use a different cGMP-driven, transduction pathway. Three-dimensional analysis of double- labeled (TGF-α, PGP9.5) serial sections revealed a unique relation among the epithelial patches, TGF-α-like immunoreactive neurons and olfactory epithelium.

Lamellar Bodies of Mouse Taste Buds

It has been reported that fixation with tannic acid preceding osmication preserves saturated phospholipids. Tannic acid reacts with the choline base of phosphatidylcholine and sphingomyelin to form a complex, which is then stabilized by treatment with OsO4 and survives during embedding procedures. Thus, the ordered lamellar structures of phospholipids can be demonstrated under an electron microscope [1].