Differentiation of myoepithelial cells in the developing rat sublingual gland

Sublingual glands of rats were prepared for light and electron microscopy and for the histochemical demonstration of myofibrils and alkaline phosphatase (AkPase) activity. Through 17 days in utero, the epithelial cells of the glandular rudiment are relatively undifferentiated. At 18 days, the inner cells of the terminal buds begin to assemble around a lumen and accumulate secretory granules, while the outer cells flatten and form long processes. At 19 days, many of the outer cells have dilated cisternae of rough endoplasmic reticulum engorged with finely granular material. At 20 days, some of the outer cells have thin bands of microfilaments in their processes, suggesting that they are differentiating into myoepithelial cells (MEC). Though the secretory cells are almost mature at birth, only a few of the MEC have myofibrils detected with an actomyosin reaction, and AkPase activity is very weak. Progressive increases in AkPase activity and in myofibril size and number continue until the acini and intercalated ducts are fully invested with mature MEC at about 14 days after birth. Thus, the MEC and secretory cells begin to differentiate at the same time, but the MEC subsequently differentiate asynchronously with the secretory cells and with each other. Although the sublingual MEC are only partly differentiated in the newborn rat, their overall development occurs somewhat more rapidly than in the adjacent submandibular gland.     

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.     

Chronology of peroxidase activity in the developing rat parotid gland

The course of development of salivary peroxidase, an enzyme that has an important role in oral defense mechanisms, has been well documented in rat submandibular glands. However, the only report on salivary peroxidase activity in the other major salivary glands of the rat has been a cytochemical study of the adult parotid gland. In the present investigation, the accumulation of salivary peroxidase activity in developing parotid glands of rats was followed both biochemically and cytochemically. Specific activity (units per mg protein) attributable to salivary peroxidase began at 1 day after birth, then rose rapidly but unevenly, with peaks at 21 and 70 days, and no difference between the sexes at any age. Activity per gland increased progressively to 42 days in both sexes and was significantly higher in males at 70 days. The cytochemical observations on peroxidase activity localized to the rough endoplasmic reticulum and secretory granules of the developing acini were well correlated with the biochemical findings. Peroxidase-negative cells occurred in immature acini at 1 and 7 days, but only in the intercalated ducts thereafter. This observation suggests that the acini are a source of some of the ductal cells, at least during early postnatal development. The developmental pattern of specific activity differed from those of other rat parotid secretory enzymes, indicating that control of their synthesis during development is noncoordinate. The patterns of specific activity of the parotid and submandibular glands were complementary, suggesting that their combined secretions may supply biologically significant peroxidase activity to the oral cavities of rats throughout postnatal development. © 1993 Wiley-Liss, Inc.     

Cytopathological features of an epithelial-myoepithelial carcinoma with predominant clear myoepithelial cells in the parotid gland

Epithelial-myoepithelial carcinoma (EMC) is a rare low-grade carcinoma occurring most frequently in the parotid gland. Most EMCs consist of two cell types that typically form double-layered ductal structures. However, occasionally EMC presents predominantly clear myoepithelial cells. A 34-year-old man visited in August 1993 and was diagnosed as having clear-cell carcinoma. The tumor was curatively resected. However, in the following 5 years, recurrence developed a total of five times. The imprint cytological feature of the recurrence at the third time showed monophasic clear cells in sheet clusters with overlapping. Most of the clear tumor cells presented an expression to alpha-smooth muscle actin (SMA). The imprint cytological feature of the recurrence at the fifth time showed increase of nuclear atypia with coarse chromatin patterns and large nucleoli. In addition to cytological findings, the cytological diagnosis of EMC with predominant clear myoepithelial cells requires a definite expression to SMA.     

Proliferative activity by cell type in the developing rat parotid gland

Background: In contrast to the considerable amount of research that has been done on the proliferative activity of the several types of parenchymal cells in the developing submandibular glands of rodents, systematic studies of cellular proliferation in the developing parotid gland have been confined to the acinar cells. The purpose of the present study was to attempt to fill this knowledge gap. Methods: Tritiated thymidine was parenterally administered to Sprague-Dawley rats at ages representative of the pre- and postnatal development of the parotid gland, and glands were harvested for autoradiography 90 min after injection. Mitotic activity among all cell types was verified by electron microscopy. Results: At all ages, the % labeled cells was much greater among the acini than any other cell type, including well-differentiated cells at 25 and 40 days. However, there were only small alterations in the proportions of cells comprised by the major cell types. Conclusions: Current theories on the histogenesis of salivary glands and their neoplasms are based on the renewing population model, in which both normal differentiated cells and neoplastic cells arise from undifferentiated stem cells in the ducts. However, these results suggest that most of the migration and redifferentiation in the developing rat parotid gland must be in the opposite direction, i.e., the acinar cells redifferentiate into ductal cells. They also indicate that until there are precise data on the rates of cell death among the several cell types, it remains more appropriate for salivary glands to be categorized as an expanding, rather than renewing, population. © 1995 Wiley-Liss, Inc.     

Dedifferentiated acinic cell carcinoma of the parotid gland with myoepithelial features

Dedifferentiated acinic cell carcinoma of the salivary gland is an uncommon variant of acinic cell carcinoma, characterized by the coexistence of both an usual low-grade acinic cell carcinoma and a high-grade dedifferentiated component, as well as by an accelerated clinical course. We describe a case of acinic cell carcinoma of the parotid gland in a 67-year-old woman, which recurred 4 times after surgery and radiotherapy. The recurrences consisted of residual foci of acinic cell carcinoma intermingled with a high-grade epithelial proliferation; the latter was focally constituted by cells with morphologic and immunohistochemical features of myoepithelium.     

Spindle cell myoepithelial tumours of the parotid gland with extensive lipomatous metaplasia

We report four cases of parotid gland tumours composed predominantly of spindle-shaped myoepithelial cells and mature adipocytes. The central portion of one tumour showed extensive adipose differentiation, whereas in the peripheral parts there were small foci of ductal epithelium arranged in cords and tubules within an abundant myxoid stroma. The other cases were adipose spindle cell myoepitheliomas without an obvious glandular component. Under high-power examination, a transition between modified spindle-shaped myoepithelial cells and adipocytes was observed, and this was confirmed with immunohistochemistry. Ultrastructurally, the modified myoepithelial cells showed intracytoplasmic tonofilaments, bundles of actin microfilaments and lipid droplets. A possible pathogenesis is proposed of true metaplastic transformation of myoepithelial cells to adipocytes. This lesion is important to identify correctly, as inadequate surgery can lead to recurrence.     

Differentiation of chromaffin cells in the developing adrenal gland of Testudo hermanni

The aim of this study was to investigate the development and differentiation of chromaffin cells in the adrenal gland of the turtle Testudo hermanni during ontogenesis using histological, immunocytochemical and ultrastructural methods. The 26 developmental stages were divided into three periods: in the early period (stages 1-18, up to 20 days of incubation at 37 degrees Celsius and 85% humidity), the chromaffin cells were observed from stage 12. They followed a ventro-lateral migration pathway with respect to the notochord and dorsal aorta, forming groups embedded in undifferentiated mesenchymal tissue. They reached the kidney surface only at the end of this period. Under the EM the chromaffin cells showed typical embryonic characters, such as rounded shape, high nucleus/plasmatic ratio, cell membrane with elongated processes; the cytoplasm contained a large number of free ribosomes, Golgi complexes, RER and a few chromaffin granules distributed in small sets. The granules were small and displayed a high electrondensity. Numerous unmyelinated fibres ran close to the chromaffin cells. At the end of this period both nervous elements and chromaffin cells were positive to the antigen for DbetaH. The intermediate period (stages 19-22, incubation days 21-35) was characterized by the first occurrence of steroidogenic cells on the ventro-medial kidney surface. Some chromaffin cells were still found in the same position, whereas other cells were still migrating, maintaining their embryonic character. It was possible to divide the secretory granules into two types according to their shape and electrondensity: the more numerous N-type granules had a dark content, whereas the small number of A-type granules (consistent with the scarce PNMT reaction) displayed a light content. They occurred for the first time in this period. In the advanced period (stages 23-26, from incubation day 36 to hatching) the adrenal gland reached its definitive shape, although remaining immature; groups of variously sized chromaffin cells intermingled with steroidogenic cells, both lying on the kidney surface. Chromaffin granules were more numerous and larger than in the previous stages, frequently mingling in the same cell. A migration pathway of the chromaffin cells along the nerve fibres can be hypothesized on the basis of their common origin and closeness. The polymorphic shape of chromaffin cells with long cytoplasmic processes also accounts for their migrating fitness. We can assume that steroidogenic differentiation from the mesodermic blastema begins after the first chromaffin cells have completed their migration.     

Regeneration of the parotid gland in the rat

Summary Parotid gland regeneration was studied in rats in which all of the left gland and part (50–60%) of the right had been removed. In young rats weighing 60–100 g, regeneration occurred in 19% of cases, as judged by the restoration of the weight of the organ. The weight of the glands did not always reach the initial level, however, and averaged 80% of the weight in control animals.The number of cases with gland regeneration increased in adult rats when the capsule was left open after the operation. Histological investigation showed that regeneration does not occur by growth starting at the injured surface, but rather by the proliferation of the secretory epithelium (regenerative hypertrophy) of the acini over the whole organ.(Presented by Active Member AMN SSSR V. V. Parin) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 50, No. 10, pp. 113–118, October, 1960     

Changes in the secretory acinar cells of the rat parotid gland during aging

The secretory acinar cells of parotid glands from rats of varying ages have been examined by electron microscopy to determine what age-related changes occur in these cells. The most prominent change noted in these cells is the progressive increase in the amount of lipofuscin granules with age. Lipofuscin granules are membrane-bound structures consisting of lipids, other subcomponents, and a matrix. In addition, these cells contain lipid droplets that are not associated with any other components and tend to accumulate at the base of the cells in older rats. Also, many acinar cells in the glands of old rats contain altered secretory granules which appear to be in the process of degeneration. The accumulation of lipid and degenerating secretory granules appears to be related to the reduced level of cellular secretory activity in the glands of older rats. It is possible that these two types of inclusions contribute to the formation of lipofuscin granules. Lipofuscin and degenerating secretory granules are associated with acid phosphatase, which is demonstrated cytochemically, indicating that these granules are lysosomal structures.     

Changing myoepithelial cell distribution during regeneration of rat parotid glands

The distribution of the myoepithelial cells during regeneration of the rat parotid gland after atrophy induced by one week of parotid duct ligation was investigated by immunohistochemistry for actin and transmission electron microscopy (TEM). Immunohistochemically, residual ducts were surrounded by actin-positive cells when clips were removed from the duct. Three days later, most of the newly formed acini originating from the residual ducts were also embraced by actin-positive cells. After 10 days, actin-positivity tended to be seen as dots around acini that decreased in number day by day. On day 21 actin-positive cells mainly surrounded intercalated ducts with only a few positive reactions identified at the acinar periphery. Electron microscopically, residual ducts and newly formed acini were peripherally embraced by myoepithelial cells before day 5. After day 7, shift of myoepithelial cells from the periphery of acini to the duct-acinar junctional region was identified. Then few myoepithelial cells were identified at the periphery of acini. These observations indicate that myoepithelial cells migrate from the acinar periphery to the duct-acinar junctional region during rat parotid regeneration, and that such behaviour is closely related to that seen during rat parotid development.     

Intercalated duct cells in the rat parotid gland may behave as tissue stem cells

This study was conducted to determine whether intercalated duct cells in the rat parotid gland have the properties of tissue stem cells. After induction of cellular proliferation by repeated administration of isoproterenol (IPR), a β-adrenergic agonist, proliferation activity in acinar, intralobular, and intercalated ductal cells was quantified using Ki-67 immunohistochemistry. The total number of each type of component cell in a gland was also estimated in the course of IPR treatment. IPR was found to induce proliferation of acinar and intercalated duct cells, but not intralobular duct cells. The total number of acinar cells in a gland on day 5 of IPR treatment was 1.6 times of that at day 0 (baseline). In contrast, the total numbers of intercalated and intralobular duct cells did not change from baseline, indicating a high possibility that the proliferated intercalated duct cells differentiated into acinar cells. On days 2 to 3 of IPR treatment, intercalated duct cells with amylase-positive secretory granules were recognized in a region very close to the acini, and were suspected of being transitional cells from intercalated duct to acinar cells. This quantitative study indicates that intercalated duct cells may have the properties of tissue stem cells upon IPR stimulation.     

Dissociation of rat parotid gland.

Rat parotid gland was dissociated by sequential collagenase and hyaluronidase digestions, chelation with ethylenediaminetetraacetic acid, and mild shearing force to yield predominantly single cells. The isolated acinar cells retained their morphologic characteristics and their amylase activity. The functional integrity of the isolated cells was assessed by measuring their secretory response to isoproterenol, epinephrine, and carbamylcholine and by their ability to incorporate radioactively labeled leucine and thymidine. The discharge of amylase from the dissociated cells was not effected by isoproterenol or norepinephrine and the response to carbamylcholine was minimal. The data indicate a destruction or perturbation of hormone receptors during the dissociation procedure. The maintenance of the cells in culture for up to 18 hours failed to restore the responsiveness of the isolated parotid gland acinar cells to isoproterenol. The isolated cells incorporated 14C-leucine into proteins at a linear rate between 30 and 180 minutes. Chromatographic and electrophoretic profiles of newly synthesized proteins indicated that all major proteins synthesized in vivo were also synthesized by the isolated cells. The isolated cells incorporated tritiated thymidine into DNA. Furthermore, stimulation of DNA synthesis by isoproterenol in vivo was reflected by a higher rate of thymidine incorporation by the isolated cells as compared with controls. The dissociated parotid gland cells offer a convenient system for studying various cellular processes, particularly the synthesis of macromolecules with high specific activity. However, some functions, notably the response to beta- adrenergic agonists, are lost during the dissociation procedure.     

A scanning electron microscope study of myoepithelial cells in the intercalated ducts of rat parotid and exorbital lacrimal glands

Myoepithelial cells in the intercalated ducts of rat parotid and exorbital lacrimal glands were examined by scanning electron microscopy. The basal surface of the intercalated ducts revealed myoepithelial cells running parallel with its long axis. These myoepithelial cells were linked with one another, forming a well-developed network, and numerous wrinkles running transversely were observed on the surface of the myoepithelial cells. Also, some myoepithelial cells in the terminal portion linked with those in the intercalated duct. Based on these findings, it is suggested that myoepithelial cells in the intercalated duct may function as a protective wall against constriction of the narrow lumen of the intercalated duct when it is subjected to pressure by the surrounding tissues.     

Melatonin release by exocytosis in the rat parotid gland

Several beneficial effects on oral health are ascribed to melatonin. Due to its lipophilic nature, non‐protein‐bound circulating melatonin is usually thought to enter the saliva by passive diffusion through salivary acinar gland cells. Recently, however, using transmission electron microscopy (TEM), melatonin was found in acinar secretory granules of human salivary glands. To test the hypothesis that granular located melatonin is actively discharged into the saliva by exocytosis, i.e. contrary to the general belief, the β‐adrenergic receptor agonist isoprenaline, which causes the degranulation of acinar parotid serous cells, was administered to anaesthetised rats. Sixty minutes after an intravenous bolus injection of isoprenaline (5 mg kg^?1), the right parotid gland was removed; pre‐administration, the left control gland had been removed. Samples were processed to demonstrate melatonin reactivity using the immunogold staining method. Morphometric assessment was made using TEM. Gold particles labelling melatonin appeared to be preferentially associated with secretory granules, occurring in their matrix and at membrane level but, notably, it was also associated with vesicles, mitochondria and nuclei. Twenty‐six per cent of the total granular population (per 100 μm² per cell area) displayed melatonin labelling in the matrix; three‐quarters of this fraction disappeared (P < 0.01) in response to isoprenaline, and melatonin reactivity appeared in dilated lumina. Thus, evidence is provided of an alternative route for melatonin to reach the gland lumen and the oral cavity by active release through exocytosis, a process which is under the influence of parasympathetic and sympathetic nervous activity and is the final event along the so‐called regulated secretory pathway. During its stay in granules, anti‐oxidant melatonin may protect their protein/peptide constituents from damage.     

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.     

Distribution of carbonic anhydrase isozyme VI in the developing bovine parotid gland

The distribution of bovine carbonic anhydrase isozyme VI (CA-VI), purified from bovine saliva, was studied immunohistochemically using antiserum against bovine CA-VI in bovine parotid glands during fetal and postnatal development. A weak expression of CA-VI in undifferentiated epithelial cells and ductal cells was observed in a 4- to 5-month-old fetus with a 26-cm crown-rump length. The reaction in both acinar and ductal cells subsequently persisted during late gestation and birth. Although anti-CA-VI reactivity was still seen in both regions immediately following birth, the reactivity had almost completely disappeared from most duct segments by 1 month following birth. Changes in the localization and time-dependent expression of the isozyme in parotid glands may reflect changes in the biological function of structurally closely related isozymes.     

Characteristics of stimulated parotid gland secretion in the aging rat

The production of pilocarpine-stimulated parotid saliva was evaluated in young adult and aged male and female rats. Parotid salivary flow rate was about 50% lower in aged animals of both sexes. Saliva of aged animals had the same Na+ concentration as that of young rats but contained about 40% more protein. Salivary K+ concentration was similar in young and aged males but not females.