Shigeo Aiyama

Developmental changes of sugar residues and secretory protein in mucous cells of the early postnatal rat parotid gland

Mucous cells have been identified in the terminal portions of the early postnatal parotid gland in human and rat, although mature parotid gland acini are composed of serous cells or seromucous cells. Previously, Ikeda et al. demonstrated that mucous cells are present in the rat parotid gland on days 1 to 8 after birth and that the secretory granules within these mucous cells share some histochemical characteristics with mature serous cells. However, it is still not clear whether the mucous cells change into serous cells as the gland develops. The purpose of this study was to determine whether the mucous cells that appear in the early postnatal rat parotid gland change into serous cells.

Changes in the number and distribution of myoepithelial cells in the rat parotid gland during postnatal development

The mature rat parotid gland shows hardly any cell bodies of myoepithelial cells around the acini, only a few cell processes being visible. However, in the early postnatal period, the rat parotid gland shows many myoepithelial cell bodies around the acini, including the intercalated ducts. In order to clarify the reason for the disappearance of myoepithelial cells from the area around the acinus during postnatal development, changes in the number and distribution of myoepithelial cells in the rat parotid gland were examined histochemically and chronologically, with particular reference to cell proliferation and cell death. From day 7 to day 14, many myoepithelial cells showing a positive reaction with anti-actin antiserum were found around the acini and intercalated ducts, but thereafter the number of such cells decreased gradually, particularly around the acini, and had almost disappeared after day 35. BrdU/PCNA-positive myoepithelial cells surrounding the acini were easily detected on day 14, but disappeared by day 21, whereas BrdU/PCNA-positive acinar cells remained numerous even after day 21. TUNEL/ISEL staining showed no positive myoepithelial cells throughout the observation period. Transmission electron microscopy also demonstrated no myoepithelial cells with chromatin condensation characteristic of apoptosis through the observation period. These findings suggest that the main reason for the disappearance of myoepithelial cells from the area around the acinus during postnatal development is the large difference between the number of myoepithelial cells and that of acinar cells, because the acinar cells retain their proliferative activity even after myoepithelial cells have become quiescent.

Effect of Glucocorticoid on the Differentiation and Development of Terminal Tubules in the Fetal Rat Submandibular Gland

Glucocorticoids (CORT) are known to promote branching of the epithelial cords during the development of the rat submandibular gland. The aim of this study was to examine the effect of CORT (triamcinolone) on the differentiation of cells forming the terminal tubules in the developing fetal rat submandibular gland and the properties of the secretory granules. Light and electron microscopy showed that the terminal tubules of the glands in the experimental group contained more type III cells, which have been identified as proacinar cells, than those in the control group, whereas the relative number of type I cells, which have been identified as terminal tubule cells, was reduced. Immunoelectron microscopy using an antibody against neonatal submandibular gland secretory protein B (SMGB) revealed the presence of more gold particles over type III cell granules in the experimental group than in the control group. Lectin histochemistry demonstrated more wheat germ agglutinin (WGA)-labeled gold particles over type III cell granules in the experimental group than in the control group. These findings suggest that CORT promote the differentiation of type III cells, and moreover stimulate the production of secretory granules reactive for SMGB and WGA by acting on the terminal tubules of the developing rat submandibular gland.

Effects of Exogenous Thyroid Hormone on the Postnatal Morphogenesis of the Rat Parotid Gland

Administration of thyroid hormone has been shown to accelerate the early postnatal development of the rat parotid gland, but these studies have dwelt almost entirely on biochemical changes. The objective of this study was to describe the effects of exogenous thyroid hormone on morphologic aspects of the developing parotid gland, in particular the transient appearance of scattered mucous cells in this otherwise serous gland. Pups were given a daily subcutaneous injection of thyroxine (T4) of 0.1, 0.5, or 5.0 μg/g body weight, vehicle only (injection control), or no injection (normal control) beginning at 4 days, and killed for the collection of blood and parotid glands at intervals through 15 days. The serum was analyzed for T4 and the glands were examined by light and electron microscopy. The results indicated that both serum T4 and the pace of gland development were proportional to the dose of T4. In particular, T4 accelerated decreases in acinar size and gland area occupied by stroma and translocation of a subset of cells with small secretory granules, deeply stained with periodic acid–Schiff, from acini to intercalated ducts. However, the chronology of mucous cell disappearance was indifferent to treatment. In addition, signs of toxicity, including slower gain in body weight and greatly increased apoptosis and vacuoles in the glands, occurred with the higher doses of T4. Anat Rec, 291:94–104, 2007.

Effects of gadolinium and tetrodotoxin on the response of slowly adapting type I mechanoreceptors to mechanical stimulation in frog dorsal skin

Abstract  To elucidate the excitatory mechanism of mechanoreceptors innervating the frog skin, we examined the effects of gadolinium (Gd3+) and tetrodotoxin (TTX) on the response of single‐unit activity of slowly adapting type I mechanoreceptors to mechanical stimulation topically applied to the receptive field (RF). Recordings were made from 46 fibers responding to mechanical stimulation with von Frey hairs, which caused an irregular firing pattern with slow adaptation. Application of a mechanically gated channel blocker, Gd3+ (30 µM), and a Na+ channel blocker, TTX (3 µM), caused the suppression of discharge rates, which was characterized by the conversion of a slowly adapting to a rapidly adapting discharge pattern. The administration of a high‐voltage‐activated (HVA) Ca2+ channel blocker, Cd2+ (100 µm), inhibited the unit discharge and caused the conversion of a slowly adapting to a rapidly adapting discharge pattern. Tonic discharges evoked by anodal electrical stimulation were inhibited by the application of Gd3+ or TTX. Electron microscopic examination showed that the cytoplasm of Merkel cells seen in the RF contained numerous Merkel granules. These results suggest that the excitatory mechanism of frog cutaneous mechanoreceptors may be mediated by the activation of Gd3+‐sensitive stretch‐activated channels in the Merkel cell–neurite complex, which are related to the Na+ influx via voltage‐gated Na+ channels and/or the Ca2+ influx through HVA Ca2+ channels.

Exogenous thyroid hormone affects myoepithelium and proliferation in the developing rat parotid gland.

Abstract In the mature rat parotid gland, myoepithelial cells (MEC) invest intercalated ducts, but not acini. During postnatal development, however, these cells differentiate around both intercalated ducts and acini, then translocate to only intercalated ducts during weaning. Previously, we found that thyroxine (T4) accelerates translocation of cells with small secretory granules from acini into intercalated ducts and the number of apoptotic cells increased tremendously with high doses. We present here additional analysis of the effects of T4 on developing rat parotid gland, namely, the distribution of MEC and the proliferation of parenchymal cells. Beginning at age four days, pups were given daily subcutaneous injections of low, medium, and high doses of T4 or vehicle or no injection. At ages 4, 7, 10, and 15 days, glands were excised and processed for light microscopy. Sections were double-immunostained with antibodies against proliferating cell nuclear antigen (PCNA) and actin, and counterstained with hematoxylin. Proliferative activity was assessed via PCNA histochemistry and MEC were identified using actin histochemistry. MEC in the T4 groups invested mostly acini at 15 days in vehicle/normal glands and mostly intercalated ducts after 10 days in the T4 groups. The proliferative activity of acinar cells and MEC in vehicle/normal glands declined progressively with age and T4 increased the rate of this decline in the MEC in a dose-dependent manner. We conclude that T4 accelerates the translocation of MEC from acini to intercalated ducts and that an important mechanism is the more rapid decline in the proliferative activity of MEC than in acinar cells in the T4 groups. Some of the decline in the proliferative activity of all cells in the high and medium dose T4 groups after seven days may have been due to dose-related thyroxine toxicity.

Strain-Specific and Endocrine Control of Granular Convoluted Tubule Cells and Epidermal Growth Factor Expression in the Mouse Submandibular Gland

Submandibular glands (SMGs) of 11‐week‐old mice from four strains, ICR, C57BL/6J, BALB/c, and C3H/HeN were examined by immunohistochemistry for epidermal growth factor (EGF). In addition to sex‐related differences in granular convoluted tubules (GCTs), the GCT cells were significantly larger in ICR mice than in other three strains. In males from each of the strains, almost all the GCT cells were strongly positive for EGF. The EGF‐positive cells in the females, however, were markedly fewer in number, and were stained weaker in C57BL/6J, BALB/c, and C3H/HeN mice than in ICR mice. The GCT cells and their EGF expression in the F1 progeny from ICR and C3H/HeN strains were approximately intermediate between those of the parent strains of the same sex. T3 and/or dihydrotestosterone (DHT) enhanced the GCT phenotype in the C3H/HeN mice, and remarkably increased the EGF‐positive cells in females. Electron microscopy revealed that gold‐labeling of EGF was confined to the secretory granules, and that the GCT cells in females, given T3 + DHT, had a well‐developed Golgi apparatus and net‐like RER but few basal infoldings, whereas the equivalent cells in the untreated females had poor RER and prominent basal infoldings. These results suggest that the EGF concentration in SMGs is genetically high in ICR mice and low in other strain mice and that, considering the same response of GCT cells to T3 and/or DHT between the high and low EGF strains, the low EGF concentrations might be partly caused by a lesser sensitivity of the GCT cells to thyroid hormones. Anat Rec, 291:105–113, 2007.

Uptake of cationized ferritin by the epithelium of the main excretory duct of the rat submandibular gland

Previous studies demonstrated that the main excretory duct (MED) of the rat submandibular gland can internalize exogenous protein in addition to reabsorbing and secreting electrolytes. However, more precise studies have not been conducted. The aim of this study was to elucidate the cell types responsible for endocytosis of an exogenous protein (ferritin) and to follow the movements of the endocytosed protein in the ductal epithelial cells.

Morphological Changes in the Rat Sublingual Gland Parenchyma with Aging

Background: The characteristics of mucous cells in the aging rat sublingual gland were investigated in this study. Particular attention was paid to accumulated amyloid protein and changes of the properties of the secretory granules at the histochemical and ultrastructural level. Objective: This study was designed to examine age-related morphological changes in the sublingual gland of male Wistar rats from 12 to 27 months. Methods: For light microscopy, the sublingual glands were fixed with 10% neutral-buffered formalin, embedded in paraffin, and processed for Alcian blue, Congo red, and TUNEL staining. For transmission electron microscopy, some of the samples were fixed with Karnovsky solution, postfixed with 2% osmium tetroxide, and embedded in epoxy resin for pronase treatment. Results: The sublingual gland showed slight shrinkage after 21 months. After 24 months, Congo red staining showed positive reaction to the intralobular connective tissue surrounding the terminal portions and to the interlobular connective tissue around the blood vessels and the excretory ducts. At 27 months, some of the granules in the serous demilunes had difficulty in digesting with pronase treatment. The appearance rate of TUNEL-positive cells was low in both mucous and serous portions during the observation period, though the positive cell number was higher in the serous than in the mucous portion. Conclusions: These findings indicate that the rat sublingual gland accumulates amyloid protein in the parenchyma and changes the properties of secretory granules of the acinar cells in the serous demilune with aging, though apoptosis of the parenchymal cells and the decrease of the gland weight are slight.

Histological changes in the mouse submandibular gland subjected to parasympathetic nerve block or ischemia: Comparison between chorda tympani resection and trophic vessel transection

The purpose of this study was to determine the effect of disorders of parasympathetic innervation or trophic vessels on the parenchymal tissue of the submandibular gland. ICR mice underwent resection of the chorda tympani nerve or transection of the trophic vessels. After treatment, the submandibular glands were removed at intervals and processed for examination using light and electron microscopy.1.From 1 week after nerve resection, the submandibular gland parenchymal tissue showed expanded blood vessels, infiltration of wandering cells and growth of connective tissue fibers, etc. Degeneration of the acinar cells extended from the periphery of the gland proper toward the inside, and by 3–4 weeks degeneration was evident over a wide area of the gland. Striated duct and granular convoluted tubule (GCT) cells showed slight damage. At 8 weeks, GCT cells and at 10 weeks acinar and intercalated duct-like cells had begun to increase in number. At 20 weeks, the glandular tissue showed almost normal parenchyma.2.From 3 weeks after nerve resection, the submandibular gland proper showed a marked decrease in weight, but returned to approximately 70% of the original weight by 20 weeks.3.At 1 hour after trophic vessel transection, acinar, striated duct and GCT cells showed vacuolation of the cytoplasm in addition to nuclear atrophy. At 9 hours, growth of connective tissue in the capsule and phagocytosis of collapsed cells by wandering cells were observed. At 18 hours, almost all parenchymal cells inside the gland proper had degenerated, and at 72 hours the gland proper finally disappeared.4.Electron microscopy and TUNEL staining showed that critical injury of the parasympathetic nerve induced necrocytosis in the acinar and GCT cells, whereas serious damage to the trophic vessels caused an apoptosis-like cell death in these cells.