Fine structure of transforming-type granules in mucous cells in the early postnatal rat parotid gland when processed by rapid freezing followed by freeze-substitution fixation

The present study was designed to clarify the more precise ultrastructural feature of granules, especially mucous granules in the early postnatal rat parotid gland by using rapid freezing followed by freeze-substitution fixation. The parotid gland of Wistar rats (aged 0-10 days) was removed under anesthesia and immediately underwent cryofixation followed by substitution with osmium tetroxide. After fixation, the samples were embedded in Epon-Araldite, cut into ultrathin section, and then examined by transmission electron microscopy. Electron microscopy showed that the mucous granules of samples treated by freeze-substitution fixation had low electron density and were almost spherical in shape with a clear limiting membrane. By Day 5, granules that were a little more electron dense than mucous granules, granules with a more electron dense portion at their periphery, and heterogeneous granules with an internal highly electron dense portion were found. Moreover, cells containing both homogeneous granules with a high electron density similar to that of mature serous granules and heterogeneous granules were observed. These findings demonstrated that the morphology of the transforming-type mucous granules by chemical fixation in the previous study was an artifact and, as a result, strongly suggested that on the sequential morphological changes of transitional mucous/serous granules by rapid freezing method in the present study, the mucous cells in the early postnatal rat parotid gland undergo transformation to serous cells.     

High‐grade epithelial‐myoepithelial carcinoma of the parotid gland with mucous cell differentiation

Epithelial‐myoepithelial carcinoma (EMC) is a rare salivary gland tumor with a low‐grade malignancy, and EMC with high‐grade histopathological features is exceedingly rare. Furthermore, EMC with intracellular mucin is also extremely rare. We report an uncommon case of a high‐grade EMC of the parotid gland with mucous cell differentiation in a 66‐year old Japanese woman who noticed a right palpable parotid mass increasing in size within a one‐year period. The cytological specimen showed a focally biphasic structure and included isolated or discohesive piled‐up clusters with hyaline globules surrounded by neoplastic cells with nuclear atypia. The gross examination revealed a relatively well‐demarcated, multinodular gray‐whitish and solid mass. Histologically, the tumor consisted of variably sized solid nests or trabeculae with central necrosis and increased mitotic activity, and invaded into adjacent skeletal muscles. Immunohistochemically, the biphasic ductal and myoepithelial differentiation of this tumor confirmed the diagnosis of high‐grade EMC. Furthermore, numerous small nests with d‐PAS and alcian blue‐positive mucous cells predominated in about 5% of the whole tumor, and these mucous cells were encompassed by neoplastic myoepithelial cells. We should recognize this variant of EMC because we can't rule out the possibility of EMC even in the presence of mucous cells.     

Morphological and histochemical changes in the secretory granules of mucous cells in the early postnatal mouse parotid gland

It has previously been known that the developing parotid glands in humans and rats contain mucous cells in their terminal clusters and acini, but these cells disappear within a short period of time. Using rat parotid glands, IKEDA and AIYAMA (1997, 1999) suggested that the mucous cells might change into serous cells in the early postnatal period, but it is uncertain whether mucous cells appear only in the developing parotid gland of a few species such as humans and rats, or whether the cell transformation actually occurs. To clarify these points, the present study investigated the developing mouse parotid glands. Light microscopy showed cells with secretory granules that stained extensively with PAS and alcian blue in the terminal clusters of a 1-day-old mouse parotid gland. Mucous cell numbers in the terminal clusters and the acini reached a peak on day 5 and decreased on day 7. By day 10, the mucous cells had disappeared altogether. Thus, the presence of mucous cells in the developing mouse parotid gland was confirmed. Electron microscopy showed granules of low-electron-density and bipartite granules in the mucous cells. Bipartite granules and highly electron-dense granules sometimes co-existed in a single cell. Immuno-electron microscopy revealed a positive reaction for amylase to the low-electron-density granules and the low-electron-density portions of the bipartite granules, in addition to the highly electron-dense granules and the electrondense cores of the bipartite granules. No mucous cells with nuclei displaying characteristics of apoptosis were recognizable. Lectin histochemistry both at the light and electron microscopic levels showed that the secretory granules in the mouse parotid gland mucous cells had sugar residues similar to those of the mature serous granules. These findings demonstrate that mucous cells appear in the early postnatal mouse parotid gland, and that almost all of these cells may be converted into serous cells.     

Enzyme histochemical localization of Na+,K+‐ATPase and NADH‐DE in the developing rat parotid gland

Information on ductal differentiation in the developing rat parotid gland is sparse. Striated and excretory ducts are rich in a number of enzymes related to ion movement. The objective of this investigation was to delineate histochemically the chronology of two of these, ouabain‐sensitive Na⁺,K⁺‐ATPase and NADH‐DE, in the developing rat parotid gland. Parotid glands were excised from rats at representative ages from 20 days in utero to 42 days. Enzyme histochemistry was performed on air‐dried frozen sections. For Na⁺,K⁺‐ATPase, some sections also were fixed in phosphate‐buffered formalin. Ouabain blocked Na⁺,K⁺‐ATPase activity, and neither enzyme reacted without substrate. Weak Na⁺,K⁺‐ATPase reactions were initially seen in unfixed sections at 1 day, and increased steadily to the adult pattern of strong (concentrated basolaterally) in striated ducts and excretory ducts, respectively, and weak to modest (diffuse) in acini and intercalated ducts at 28 days. In fixed sections, localization was sharper but the reaction was somewhat reduced. NADH‐DE was modest in terminal buds and ducts before birth, then progressively changed to the adult pattern of weak in acini and intercalated ducts and strong (concentrated basally and luminally) in striated and excretory ducts at 28 days. As demonstrated by enzyme histochemistry of Na⁺,K⁺‐ATPase and NADH‐DE, differentiation of rat parotid striated ducts and excretory ducts occurs mainly between birth and 28 days. Anat Rec 256:72–77, 1999. Published 1999 Wiley‐Liss, Inc.     

Going beyond “Basaloid neoplasm”: Fine needle aspiration cytology of epithelial‐myoepithelial carcinoma of the parotid gland

Epithelial‐myoepithelial carcinoma (EMC) is a rare salivary gland malignancy with variable cytologic findings. Its rarity, variable morphologic findings, and similarities with more common salivary gland entities make it a difficult cytologic diagnosis. As the name signifies, the key feature of this tumor is presence of an epithelial and myoepithelial component. However, when one of these two components is scant on the fine needle aspiration (FNA) smears, it may be overlooked. We present a case from a 62 year‐old female who presented to the clinic with a parotid nodule and episodes of sharp, throbbing pain. A fine needle aspiration was performed which revealed a highly cellular specimen comprised primarily of aggregates of cells with small, round nuclei and scant to absent cytoplasm. Abundant hyaline stromal material was also noted. The case was signed out as basaloid neoplasm with a recommendation for surgical resection. The subsequent resection specimen revealed EMC. By reviewing the FNA specimen following the surgical resection of the tumor, we were able to utilize the benefit of hindsight to more clearly identify the subtle, biphasic components of the tumor. Diagn. Cytopathol. 2016;44:422–425. © 2016 Wiley Periodicals, Inc.     

Temporary accumulation of glycogen in the epithelial cells of the developing mouse submandibular gland

Temporary accumulation of glycogen in the epithelial cells of the developing mouse submandibular gland was examined under light microscopic histochemistry and electron microscopy. To avoid loss of water-soluble glycogen during histological tissue preparation, fixation with ethanol and embedding in hydrophilic glycol methacrylate resin was used for light microscopy, and high-pressure freezing/freeze substitution for electron microscopy. Glycogen was detected on periodic acid-Schiff stain, periodic acid-thiosemicarbazide-silver proteinate reaction, and the digestion test with alpha-amylase. On embryonic day 14, glycogen began to accumulate in the proximal portions of the developing epithelial cords. On embryonic day 17, marked glycogen particles were seen at the basal portion of the ductal epithelial cells and an abrupt increase of glycogen accumulation occurred in the secretory cells in the terminal bulbs. Ultrastructural observation indicated large clumps of glycogen particles localized in the basal portion of the terminal bulb cells. The initiation of glycogen accumulation preceded the formation of lumens in the ducts and terminal bulbs. Furthermore, proliferation analysis by bromodeoxyuridine labeling showed that this glycogen accumulation followed the cessation of the epithelial cell proliferation. Postnatally, glycogen accumulation in the terminal bulbs became gradually inconspicuous and completely disappeared by postnatal day 3, but that in the ducts was retained until around postnatal day 12. Temporary glycogen accumulation after the cell proliferation and before/during the lumen formation and secretory granule formation suggests significant involvement of the carbohydrate metabolism in the organogenesis of the submandibular gland.     

Effects of β-adrenergic agonists in the parotid gland of the rat–an electrophysiological study

The effects of some β-adrenergic agonists were studied in the parotid gland of the rat by electrophysiological techniques. In the unoperated gland, isoprenaline caused depolarizations which were slowly developing, long-lasting and of low amplitude. The same response was seen when noradrenaline was combined with α-adrenoceptor blocking drugs. A greater number of cells responded to this combination than to isoprenaline. After either parasympathetic or sympathetic denervation 1–3 weeks in advance, to induce supersensitivity, the number of cells responding to β-adrenoceptor stimulating drugs was significantly increased. In the latter case the threshold dose required to evoke a response was also significantly lowered. Atropine did not have any effect on the isoprenaline-evoked response. The combined parasympathetic and sympathetic denervation did not further increase the responsiveness. It is concluded that β-adrenoceptor stimulation in the parotid gland of the rat may cause membrane depolarizations. The response is mediated by β₁- adrenoceptors. The responsiveness is increased in the denervated gland. Secretory studies have demonstrated a supersensitivity to β-adrenergic agonists as a result of denervation. On the other hand, β-adrenoceptor stimulation is believed mainly to activate the adenylate cyclase/cyclic AMP system independent of membrane potential changes. It is thus not known if the present ‘supersensitivity’ is correlated to the increased secretory response earlier demonstrated in this gland.     

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. Parotid glands were obtained from male or female Wistar rats (aged 0-14 days and adults). Fixed tissue sections were reacted with soybean agglutinin (SBA) and wheat germ agglutinin (WGA) to detect glycoconjugates, or were stained using an anti-neonatal submandibular gland protein B1 (SMG-B1) antibody to identify serous acinar cells. The sections were observed by transmission electron microscopy. Electron microscopy revealed that cells with characteristics intermediate between those of mucous and serous cells (transitional cells) appeared around day 8 and that the nuclei of these cells did not show chromatin condensation, a characteristic of apoptotic cells. Lectin histochemistry showed that the mucous cells had the same sugar residues as the serous cells, which appeared after day 10. Immunohistochemistry with an anti-SMG-B1 antibody gave a positive reaction not only in the cells with highly electron-dense granules but also in the electron-dense cores of bipartite or tripartite granules in the transitional cells. Cells with morphological characteristics intermediate between those of mucous and serous cells (transitional cells) appearing in the early postnatal rat parotid gland begin to produce B1-immunoreactive protein common to serous acinar cells during development of the gland.     

Long‐term dexamethasone treatment alters the histomorphology of acinar cells in rat parotid and submandibular glands

Glucocorticoids (GCs) induce insulin resistance (IR), a condition known to alter oral homeostasis. This study investigated the effects of long‐term dexamethasone administration on morphofunctional aspects of salivary glands. Male Wistar rats received daily injections of dexamethasone [0.1 mg/kg body weight (b.w.), intraperitoneally] for 10 days (DEX), whereas control rats received saline. Subsequently, glycaemia, insulinaemia, insulin secretion and salivary flow were analysed. The parotid and submandibular glands were collected for histomorphometric evaluation and Western blot experiments. The DEX rats were found to be normoglycaemic, hyperinsulinaemic, insulin resistant and glucose intolerant (P < 0.05). DEX rat islets secreted more insulin in response to glucose (P < 0.05). DEX rats had significant reductions in the masses of the parotid (29%) and submandibular (16%) glands (P < 0.05) that was associated with reduced salivary flux rate. The hypotrophy in both glands observed in the DEX group was associated with marked reduction in the volume of the acinar cells in these glands of 50% and 26% respectively (P < 0.05). The total number of acinar cells was increased in the submandibular glands of the DEX rats (P < 0.05) but not in the parotid glands. The levels of proteins related to insulin and survival signalling in both glands did not differ between the groups. In conclusion, the long‐term administration of dexamethasone caused IR, which was associated with significant reductions in both mass and flux rate of the salivary glands. The parotid and submandibular glands exhibited reduced acinar cell volume; however, the submandibular glands displayed acinar hyperplasia, indicating a gland‐specific response to GCs. Our data emphasize that GC‐based therapies and insulin‐resistant states have a negative impact on salivary gland homeostasis.     

Multinucleated giant cells in HIV‐associated benign lymphoepithelial cyst‐like lesions of the parotid gland on FNAC

Benign lymphoepithelial cyst‐like lesion (BLEL), a previously rare lesion of the parotid gland consisting of marked lymphoid hyperplasia with accompanying squamous‐lined cysts, has been described in patients with the acquired immune deficiency syndrome (AIDS) or AIDS risk factors. Histologically, these cysts are lined by a squamous or cuboidal epithelium. The lumen contains a pale homogenous material with foamy macrophages and lymphocytes with the cyst wall having germinal centers and a dense infiltrate of lymphoid cells. On FNAC, the aspirates are mostly cystic with the presence of reactive lymphoid tissue, numerous histiocytes, and metaplastic cell clusters. Multinucleated giant cells (MGCs) are also rarely seen in such lesions. We report a case of HIV‐associated BLEL with numerous large sized multinucleated giant cells. Diagn. Cytopathol. 2009. © 2009 Wiley‐Liss, Inc.     

Giant cell‐rich osteosarcoma of the parotid gland: An exceptionally rare entity at an unusual site

Giant cell‐rich osteosarcoma is a rare histologic variant of conventional osteosarcoma that affects mainly the extremities. Extraskeletal giant cell‐rich osteosarcoma is therefore exceedingly rare. Here, we report the first case of this uncommon tumor involving the parotid gland in a 62‐year‐old male who presented with initial right jaw swelling. Radiologic work‐up revealed a 6.2 cm mass involving the right parotid gland. Fine‐needle aspiration cytology showed numerous multinucleated giant cells in a background of dyshesive epithelioid cells and rare clusters of spindle stromal cells, suspicious for malignancy. The subsequent excisional biopsy showed histopathologic features diagnostic for giant cell‐rich osteosarcoma. Diagn. Cytopathol. 2016;44:1107–1111. © 2016 Wiley Periodicals, Inc.     

Transient occurrence of 27 kDa heat‐shock protein in the terminal tubule cells during postnatal development of the rat submandibular gland

It has been suggested that the 27 kDa heat‐shock protein (Hsp27) plays a role at crucial cellular checkpoints for proliferation, apoptosis, and differentiation. We examined the immunolocalization of Hsp27 in the rat submandibular gland during postnatal development, wherein acinar cells proliferate and differentiate at earlier postnatal periods. At 2 weeks of age, weak Hsp27 immunoreactivity was distributed diffusely over all gland components. At 3 weeks, Hsp27 immunoreactivity disappeared in most parts of the acini and ducts, but was intensely accumulated in a small cell population located in the acinar center. This population was composed mostly of terminal tubule (TT) type I cells. At 4 weeks, the Hsp27‐immunopositive cell population in the acinar center was composed primarily of immature (type II) acinar cells, partly of immature (granulated) intercalated duct (ID) cells, and occasionally of apoptotic cells. After 5 weeks, all acinar components became mature and were no longer immunoreactive for Hsp27. When acinar cell differentiation was accelerated by administration of isoproterenol to 3‐week‐old rats for 7 days, the number of Hsp27‐positive cells was significantly lower than in the control gland at 4 weeks, confirming that Hsp27 expression is downregulated in mature acinar cells. These results suggest that at around 3–4 weeks in postnatal development, the centroacinar TT cells stop proliferating and begin to differentiate into acinar and ID cells, and occasionally undergo apoptosis. Hsp27 is transiently expressed in the centroacinar TT cells during this critical period, and thus may play a role in their differentiation into the immediate descendants. Anat Rec 264:358–366, 2001. © 2001 Wiley‐Liss, Inc.     

Necrotizing sialometaplasia of the parotid gland associated with angiocentric T‐cell lymphoma: A case report and review of the Literature

A very rare case of necrotizing sialometaplasia of the parotid gland associated with angiocentric T‐cell lymphoma was described. A 66‐year‐old male had left neck and pharyngeal masses and biopsy specimen showed a monotonous proliferation of atypical lymphoid cells with massive necrosis in the parotid gland. Angiocentric pattern or vascular invasion by the lymphoid cells was observed and the involved parotid gland exhibited squamous metaplasia of the ducts and acini; necrotizing sialometaplasia. Immunohistochemical analysis revealed a cytotoxic T‐cell phenotype of the lymphoid cells (CD3+, CD4‐, CD5+, CD8+, CD56‐, Granzyme B+, TIA‐1+, Perforin‐) but in situ hybridization showed no relation to Epstein‐Barr virus. Although necrotizing sialometaplasia is relatively rare in the parotid gland, angiocentric T‐cell lymphoma should be considered for a causative condition of necrotizing sialometaplasia.     

The fine structure of testicular interstitial cells in the mouse. A comparison after conventional double fixation versus rapid freezing followed by freeze substitution

The interstitial cells in the testis of the mouse have been studied both after conventional double fixation with glutaraldehyde followed by osmium tetroxide and after rapid freezing and cryosubstitution with osmium tetroxide-acetone by light, scanning and transmission electron microscopy. Differences have been found especially concerning the preservation of lipids and the appearance of mitochondria. Varying proportions and different aspects of rough and smooth endoplasmic reticulum were observed corresponding after both preparation methods thus confirming their real existence. Numerous specific contacts are found between macrophages and Leydig cells, indicating a functional interdependence between these two cell types. Though structural preservation is superior after freeze substitution in the outermost zone, artifacts caused by the initial impact, by ice crystal formation, and during substitution must be considered.