ULTRASTRUCTURAL CYTOCHEMICAL DEMONSTRATION OF ELASTIN IN THE MATRIX OF SALIVARY GLAND TUMORS

Following light microscopic survey of the incidence of elastic tissue in 80 salivary gland tumors, tissue samples from 14 pleomorphic adenomas, three myoepitheliomas, and eight adenoid cystic carcinomas were processed for cytochemical demonstration of elastin with the tannic add stain for ultrathin sections. For comparative study, some other tumor types devoid of elastic tissue at the light microscopic level and non-neoplastic submandibular glands were also investigated. Elastic deposits of varying amounts were clearly revealed on the basal-lamina-like material and/or masses of microfibrils in the matrix close to the neoplastic myoepithelium and, to a lesser degree, immediately beneath the non-neoplastic myoepithelium. None of the other tumor types without myoepithelial differentiation contained elastic deposits closely associated with the neoplastic cells. Intimate topographical relationship of such an immature elastic fiber or developing elastic tissue to the neoplastic myoepithelial cells strongly indicated the primary origin of elastic components from these cells. It is postulated that the potential of salivary tumor cells to produce elastin is regarded as an indicator of their myoepithelial nature or differentiation.     

Tyrosine-rich crystalloids in neoplasms and tissues of the head and neck

Tyrosine-rich crystalloids were identified in three benign mixed tumors of the parotid gland, one terminal duct adenocarcinoma of a minor salivary gland, and the fibrous connective tissue of two laryngectomy specimens. Light and electron microscopic studies showed the crystalloids to be composed of irregular deposits of amorphous electron-dense material. In the salivary gland tumors this material was commonly associated with interstitial collagen and was found in greatest abundance near myoepithelial cells. This proximity suggests that the tyrosine-rich crystalloids result from the precipitation on stromal collagen of products secreted by neoplastic myoepithelial cells.     

Myoepithelial differentiation markers in salivary gland neoplasia]

Salivary gland tumors frequently present myoepithelial cell differentiation that is not always easily identified on routinely stained sections. Recently novel markers of myoepithelium have been studied, such as calponin (CALP), caldesmon (CALD), and smooth muscle myosin heavy chain. These markers, together with smooth muscle actin may be useful tools for identifying myoepithelial cells. We immunohistochemically studied a series of 23 benign and malignant salivary gland tumors using antibodies to these four markers. The tumors were classified as follows: pleomorphic adenoma (n = 8), basal cell adenoma (n = 3), myoepithelioma with plasmacytoid cells (n = 2), epithelial-myoepithelial cell carcinoma (n = 6) and adenoid cystic carcinoma (n = 4). All tumors were positive for at least one of the four markers. CALP and smooth muscle actin were the markers more frequently expressed. Positivity was mostly located in the myoepithelial cells that constitute the external layer of the glandular or tubular neoplastic structures. In poorly differentiated epithelial myoepithelial carcinomas, composed of solid sheets of neoplastic cells and sometimes of clear cells, immunohistochemical staining for myoepithelial markers evidenced rudimentary glandular structures. CALP and smooth muscle actin were positive in the two cases of myoepithelioma with plasmacytoid cells. In conclusion, the combined staining with four markers helps to disclose myoepithelial cell differentiation and can be a useful tool for the correct histopathological diagnosis of salivary gland tumors. Among the four markers studied, CALP and smooth muscle actin were the most useful to identify myoepithelial cell differentiation.     

Histogenesis of acinic cell carcinoma of the major and minor salivary glands. An ultrastructural study

This study describes the light microscopic, histochemical and electron-microscopic findings of 10 acinic cell carcinomas from the major and minor salivary glands. Ultrastructurally, four cell types were identified: secretory acinar cells, intercalated duct-like cells, pluripotential reserve/stem cells and myoepithelial cells. This cellular composition suggests that the tumours are derived from neoplastic proliferation, cytodifferentiation and functional maturation of pluripotential reserve/stem cells which normally reside at the acinar-intercalated duct junctions and/or in the intercalated ducts proper of adult salivary glands. This study further supports the concept that different salivary gland tumours recapitulate various developmental stages in the normal embryogenesis of the salivary glands.     

Immunohistochemical identification of actomyosin-containing (myoepithelial) cells in non-neoplastic and neoplastic tissues.

Actomyosin-containing cells in both non-neoplastic and neoplastic tissues of the salivary gland, lung, breast and some other organs were studied by immunofluorescent microscopy using antiactomyosin rabbit serum. In the breast, myoepithelial-like cells with positive immunofluorescence in the cytoplasm were observed not only in sclerosing adenosis and fibroadenoma but also in scirrhous and medullary-tubular duct carcinomas. No positive cells were observed in medullary carcinomas with lymphoid infiltration. The actomyosin positive cells were also seen at the outer layer of tubules of "mixed tumors" and of cell nests in adenoid cystic carcinoma and in myoepithelioma of the salivary gland, but not in the metaplastic squamous cells or in the cells of myxomatous and chondroid areas of "mixed tumor". In carcinoma of the lung, actomyosin-positive cells were observed in adenoid cystic carcinomas and adenocarcinoma of the bronchial gland type, but they were not seen in squamous cell carcinomas or papillary adenocarcinomas. It was concluded that the actomysoin-containing cells with structural appearances of myoepithelial cells in a variety of tumors were neoplastic myoepithelial cells.     

Pleomorphic adenoma, I: Ultrastructural organization of "epithelial" regions

Twenty-four major and minor salivary gland pleomorphic adenomas were studied ultrastructurally to determine the growth patterns, organization, and cytologic modifications of the proliferating neoplastic cells. In compact and highly cellular regions, two cell types--luminal epithelial and myoepithelial--could often be identified; their organization mimicked that of the normal salivary gland duct or acinar unit. Results of the study indicate that the principal proliferating tumor cell is a structurally modified myoepithelial cell that frequently shows squamous differentiation. At the immediate margins of cellular regions of many tumor cells, gradual dedifferentiation of modified myoepithelial cells with a loss of squamous features occurs, although in some cells the squamous features are retained to varying degrees. Within cellular regions, the earliest development of matrix occurs in relation to small, basal lamina-lined extracellular spaces between myoepithelial-like cells. Modifications of such intercellular spaces are helpful in tracing the development of myxoid zones and the evolution of cell types in this unique region. The authors postulate that salivary gland pleomorphic adenomas result from the neoplastic transformation of the complete ductal-acinar unit rather than from one particular ductal "reserve" cell.     

IMMUNOHISTOCHEMICAL IDENTIFICATION OF ACTOMYOSIN-CONTAINING (MYOEPITHELIAL) CELLS IN NON-NEOPLASTIC AND NEOPLASTIC TISSUES

Actomyosin-containing cells in both non-neoplastic and neoplastic tissues of the salivary gland, lung, breast and some other organs were studied by immunofluorescent microscopy using antiactomyosin rabbit serum. In the breast, myoepithelial-like cells with positive immunofluorescence in the cytoplasm were observed not only in sclerosing adenosis and fibroadenoma but also in scirrhousgand medullary-tubular duct carcinomas. No positive cells were observed in medullary carcinomas with lymphoid infiltration. The actomyosin positive cells were also seen at the outer layer of tubules of "mixed tumors" and of cell nests in adenoid cystic carcinoma and in myoepithelioma of the salivary gland, but not in the metaplastic squamous cells or in the cells of myxomatous and chondroid areas of "mixed tumor". In carcinoma of the lung, actomyosin-positive cells were observed in adenoid cystic carcinomas and adenocarcinoma of the bronchial gland type, but they were not seen in squamous cell carcinomas or papillary adenocarcinomas.
It was concluded that the actomysoin-containing cells with structural appearances of myoepithelial cells in a variety of tumors were neoplastic myoepithelial cells.     

Oncocytic adenoma of the parotid gland with psammoma bodies

An unusual slow-growing tumor was found in the superficial lobe of the right parotid gland. It was multilobulated and encapsulated and consisted of sheets of epithelial oncocytes and minor foci of myoepithelium and ducts. Psammoma bodies were abundant. An antibody directed against keratin protein was localized in all tumor cells and in ductal but not acinar elements of adjacent parotid tissue. Ultrastructurally, the neoplastic cells proved to be ductal epithelial and myoepithelial oncocytes.     

Findings of myoepithelial cells in human breast cancer. Ultrastructural and immunohistochemical study by means of anti-myosin antibody

Detailed light and electron microscopic, and immunohistochemical observations were made on the distribution and morphological characteristics of myoepithelial cells in the 53 cases of breast cancer. In non-invasive carcinoma, myoepithelial cells in the normal duct were found to be remaining at the outer margin of the cancer nests, but neoplastic myoepithelial cells were not detected in the carcinoma tissue. In invasive carcinoma, a small number of fluorescence-weakly-positive cells could be observed in more than 50% of medullary-tubular carcinoma, in all cases of papillary-tubular carcinoma, and two of three cases of invasive lobular carcinoma. Almost all of these cells were ultrastructurally intermediate cells which have the morphological characteristics of both epithelial cell and myoepithelial cell. Fluorescence-positive cells were observed in all cases of scirrhous carcinoma. Moreover, these cells showed a stronger fluorescence than that of other types of carcinoma and were ultrastructurally more similar to normal myoepithelial cell. The tumor cells having myoepithelial characteristics in invasive carcinoma showed a stronger tendency for arranging at the margin of carcinoma nests in contact with the stroma. The results of the present study indicate that in invasive carcinoma of the breast, neoplastic myoepithelial cells could be demonstrated together with ductal epithelial cells and as to its histogenesis, there is a possibility that breast cancer develops from common stem cells which have the ability of differentiating into both epithelial and myoepithelial cell because of the presence of intermediate cells.     

Adenoid cystic carcinoma: a comparative pathologic study of tumors in salivary gland, breast, lung, and cervix

Histologic, histochemical, and ultrastructural features of eight adenoid cystic carcinomas arising at diverse sites were compared in order to determine diagnostic values and to investigate histogenetic mechanisms. These tumors originated in the salivary glands, breast, uterine cervix, and tracheobronchial tree. By light microscopy each tumor was seen to have morphologic features of adenoid cystic carcinoma, yet only five of the eight cases showed differential staining for the two mucin types, stromal and epithelial, which are reportedly present in these tumors. In contrast, every case showed a set of fine structural features which, in aggregate, are specific for adenoid cystic carcinoma. These features include pseudocysts, intercellular spaces, basal lamina, and true glandular lumens. The most prominent feature is the pseudocyst, which mimics a glandular lumen by light microscopy but is actually a rounded extracellular space containing basal lamina. Ultrastructurally, the variation in composition of the extracellular compartments, including pseudocysts and true lumens, appears to explain the lack of uniformity in the histochemical staining. The tumors also contained cytoplasmic microfilaments in parallel bundles, consistent with myofilaments. The presence of these filaments combined with basal lamina suggests myoepithelial differentiation, yet it is not known whether these tumors truly originate from myoepithelium or show differentiation toward myoepithelium as a part of the neoplastic process. Regardless of their histogenesis, this study shows that true adenoid cystic carcinomas do arise in different organs. Knowledge of the specific ultrastructural features of adenoid cystic carcinomas can be useful in classifying these tumors in some cases.     

Findings Of Myoepithelial Cells In Human Breast Cancer Ultrastructural And Immunohistochemical Study By Means Of Anti-Myosin Antibody

Detailed light and electron microscopic, and immunohistochemical observations were made on the distribution and morphological characteristics of myoepithelial cells in the 53 cases of breast cancer. In non-invasive carcinoma, myoepithelial cells in the normal duct were found to be remaining at the outer margin of the cancer nests, but neoplastic myoepithelial cells were not detected in the carcinoma tissue. In invasive carcinoma, a small number of fluorescence-weakly-positive cells could be observed in more than 50% of medullary-tubular carcinoma, in all cases of papillary-tubular carcinoma, and two of three cases of invasive lobular carcinoma. Almost all of these cells were ultrastructurally intermediate cells which have the morphological characteristics of both epithelial cell and myoepithelial cell. Fluorescence-positive cells were observed in all cases of scirrhous carcinoma. Moreover, these cells showed a stronger fluorescence than that of other types of carcinoma and were ultrastructurally more similar to normal myoepithelial cell. The tumor cells having myoepithelial characteristics in invasive carcinoma showed a stronger tendency for arranging at the margin of carcinoma nests in contact with the stroma. The results of the present study indicate that in invasive carcinoma of the breast, neoplastic myoepithelial cells could be demonstrated together with ductal epithelial cells and as to its histogenesis, there is a possibility that breast cancer develops from common stem cells which have the ability of differentiating into both epthelial and myoepithelial cell because of the presence of intermediate cells. ACTA PATHOL. JPN. 34: 537–552, 1984.     

Basal cell adenocarcinoma arising from the minor salivary gland in the soft palate: a case report

Basal cell adenocarcinomas (BCACs) of the oral minor salivary gland are very rare neoplasms. We report on an 86-year-old woman with BCAC arising from the minor salivary gland in the soft palate. Histologically, the tumor was located in the submucosa and showed microinvasion into the adjacent soft tissue without encapsulation. It contained tiny tumor islands with solid and tubular patterns, as well as myxoid stroma. The neoplastic cells were basaloid cells and were composed of large pale cells and small dark cells. They were positive for alpha-smooth muscle actin, cytokeratin 14, and vimentin in the periphery of the tumor island, showing a myoepithelial differentiation. The myxoid stroma was positive for alcian blue and colloidal iron. Apical membranes of the neoplastic cells were positive for MUC1 and CEA. The present case is the 14th documented case of oral BCAC (the fifth case of palatal BCAC).     

DOG1: a novel marker of salivary acinar and intercalated duct differentiation

Anoctamin-1 (ANO1) (DOG1, TMEM16a) is a calcium-activated chloride channel initially described in gastrointestinal stromal tumors, but now known to be expressed in a variety of normal and tumor tissues including salivary tissue in murine models. We herein perform a comprehensive survey of DOG1 expression in 156 cases containing non-neoplastic human salivary tissues and tumors. ANO1 mRNA levels were significantly higher (8-fold increase, P<0.0001) in normal parotid tissue (n=6) as compared with squamous mucosa (n=15). By immunohistochemistry, DOG1 showed a diffuse moderate (2+) apical membranous staining pattern in normal serous acini, 1+ apical membranous pattern in mucous acini, and variable 1-2+ apical staining of distal intercalated ducts. Myoepithelial cells, striated and excretory ducts were invariably negative. All acinic cell carcinomas (n=28) were DOG1 positive demonstrating a complex mixture of intense (3+) apical membranous, cytoplasmic and complete membranous staining. Most ductal tumor types were negative or only showed a subset of positive cases. Within the biphasic tumor category, adenoid cystic carcinomas (18/24 cases) and epithelial-myoepithelial carcinomas (8/15 cases) were frequently positive, often showing a distinctive combined apical ductal and membranous/cytoplasmic myoepithelial staining profile. Thus, DOG1 staining is a marker of salivary acinar and to a lesser extent intercalated duct differentiation. Strong staining can be used to support the diagnosis of acinic cell carcinoma. DOG1 may also be a marker of a 'transformed' myoepithelial phenotype in a subset of biphasic salivary gland malignancies.     

Low-grade mucoepidermoid carcinoma of salivary glands: characteristic immunohistochemical profile and evidence of striated duct differentiation

The purpose of the present study is to determine the presence and distribution of epithelial and myoepithelial cells in mucoepidermoid carcinoma (MEC) of salivary glands and to compare them with normal salivary gland tissue and other primary carcinomas. This is in order to establish novel diagnostic criteria and to better understand MEC histogenesis. Formalin-fixed paraffin-embedded tissues from ten well-differentiated MECs, three adenoid cystic carcinomas (ACC), four acinic cell carcinomas (AC), and three epithelial-myoepithelial carcinomas (EMCC) of salivary glands were studied with immunohistochemistry using antibodies that recognise antigens indicative of epithelial and myoepithelial cell differentiation. An anti-mitochondrial antibody was also employed. Normal salivary tissue was present for comparative study in non-tumorous areas of the same section from 12 cases. MEC contained numerous keratin-positive cells. Anti mitochondrial antibody was diffusely positive in all ten of these tumours. Smooth muscle actin, h-caldesmon, and smooth muscle heavy chain myosin, which are indicative of myoepithelial cell differentiation, were negative. Rare cells in only one case were stained by calponin. Cytokeratin 14 (CK14) and anti mitochondrial antibody stained cells located mainly at the periphery of neoplastic nests and cystic spaces, while CK7 was mainly present in cells bordering gland lumina (zoning pattern). The immunohistochemical cell profile was similar to that seen in striated normal ducts. All others tumours studied showed a different immunohistochemical pattern, mostly consisting of a lack of mitochondrion-rich cells and the presence of myoepithelial cells in ACC and EMCC. Immunoreactivity in MEC for CK7, CK14 and mitochondrial antibodies appears as a peculiar pattern of staining, different from that of other salivary gland tumors; this seems helpful for diagnostic purposes. In addition, a differentiation of the "striated duct phenotype" is suggested.     

Salivary gland monomorphic adenoma. Ultrastructural,immunoperoxidase, and histogenetic aspects.

Monomorphic adenoma of basal cell type is a salivary gland tumor believed to result from a proliferation of a single type of cell. However, ultrastructural and immunocytochemical investigations of 6 monomorphic adenomas (5 from parotid and 1 from intraoral minor salivary gland) indicate that there are two classes of these lesions, one composed of two types of tumor cells and the other wholly or predominantly made up of one type of cell (isomorphic). In the former group, the organization of the tumor cells closely mimicked that of normal and hyperplastic salivary gland intercalated ducts. Aggregates of tumor cells were arranged as an inner layer of luminal epithelial cells which were surrounded by an outer layer of cells that, in some cases, had ultrastructural and immunohistochemical features indicating myoepithelial cell differentiation. In some adenomas formed by two types of tumor cells, basal-lamina-lined extracellular spaces were identified ultrastructurally in relation to modified myoepithelial cells; such spaces had the same fine-structural features as those reported in pleomorphic adenoma and adenoid cystic carcinoma. Predominantly isomorphic adenomas were composed exclusively of luminal epithelial cells. These results indicate that despite the varied histologic patterns in the numerous subtypes of monomorphic adenoma, there is a central theme of differentiation and organization in this type of neoplasm which recapitulates the ductoacinar unit of normal salivary gland parenchyma.     

Nestin is a wide‐spectrum abluminal cell marker of salivary gland tumors

Nestin is an intermediate filament that was first identified in neural progenitor cells. It is expressed in various cell types in the nervous system as well as in other systems. In the present study, we investigated nestin expression in non‐neoplastic salivary gland tissue and in salivary gland tumors. In non‐neoplastic salivary glands, nestin expression was observed in only a few abluminal cells. In contrast, diffuse nestin staining was observed in the abluminal cells of pleomorphic adenoma (11 of 11 cases), basal cell adenoma (7 of 7 cases), and epithelial‐myoepithelial carcinoma (2 of 2 cases). The stromal cells in basal cell adenoma also expressed nestin. In adenoid cystic carcinoma (6 of 7 cases) and polymorphous low‐grade adenocarcinoma (3 of 3 cases), nestin positive cells were observed focally. Nestin was not detected in Warthin tumor (6 cases), classical acinic cell carcinoma (2 cases), mucoepidermoid carcinoma (5 cases), or salivary duct carcinoma (4 cases). Because the nestin expression pattern in each histological salivary gland tumor type is unique, nestin could be a very useful abluminal cell marker for the diagnosis of salivary gland tumors.     

Elastofibroma

Summary Four cases of elastofibroma located in the subscapular region of 3 men aged 66, 74, and 83 years, and a woman 70 years old are reported. A correlated light and electron microscopic study including ultrastructural examination of Verhoeff's iron-hematoxylin (VIH)-stained sections was performed. Light microscopically, the elastofibromas were characterized by connective tissue built up by collagen fibers and sclerotic masses mingled with numerous fibers and globules of elastin material. In one micron thick Epon sections these elastin fibers often revealed a central axis surrounded by a mantle composed of periodic segments giving them a necklace-like appearance. The ultrastructural findings of these elastin structures, stained with VIH, and the appearance of the stroma cells and their relation to the elastin indicate that elastofibroma is a non-neoplastic reactive lesion in which elastin material is synthesized by the stromal cells and predominantly laid down around preexisting elastic fibers.     

Benign apocrine papillary lesions of the breast lacking or virtually lacking myoepithelial cells-potential pitfalls in diagnosing malignancy

Two cases of apocrine papillary lesions are presented, in which myoepithelial cells were not visualized on hematoxylin and eosin-stained slides. A complex sclerosing papillary lesion did not exhibit myoepithelial cells on p63 and CD10 immunostaining, although a thin rim of myoepithelial cell cytoplasm was revealed by smooth muscle actin, calponin, and S100 immunostains. In the other lesion, without sclerosis, myoepithelium was not detected in the central papillary cores or at the periphery of the cystic wall on hematoxylin and eosin-stained sections or with any of the myoepithelial markers tested, although two small wall areas did display the presence of myoepithelium. These two cases strengthen the view that the lack of myoepithelium alone in apocrine lesions cannot be equated with malignancy. As recent literature reports have demonstrated that myoepithelial cells may be missing from benign apocrine changes and proliferations of different types, a reconsideration of the diagnostic criteria of the malignancy of such lesions appears essential.     

A case report of basal cell adenoma showing elastic fiber (elastin-basement membrane complex) formation of the submandibular gland

A biopsy case of basal cell adenoma of the submandibular gland was reported in a 15-year-old boy. The tumor was pigeon's egg-sized, spherical in shape and encapsulated by fibrous tissue, and its cut-surface was grayish white. Histologic feature of this neoplasm was a trabecular or tubular monomorphic adenoma with well-developed elastic fibers (elastin-basement membrane complex) in the interstitial tissue. Electron microscopy disclosed 4 kinds of tumor cells, that is, secreting cells, squamous cell-like cells with tonofibrils and tonofilaments, clear cells with a few filaments, and myoepithelial cells. The interstitium contained elastin, collagen fibrils and basement membrane-like substance. Possible production of elastic fibers by the myoepithelial cells of this adenoma was discussed.     

Immunohistochemical and electron microscopical study of the myoepithelium in mastopathy and carcinoma of the mammary gland

Biopsies obtained from 45 cases of invasive ductal carcinoma and 41 cases of cystic mastopathy were immunohistochemically investigated, with electron microscopy being used on some of them, for the purpose of clearing up presence, distribution, and ultrastructural peculiarities of the myoepithelial cells. Focal or diffuse myoepithelial proliferations were observed in all cases of cystic mastopathy, with peripheral layers of myoepithelium at the basal membrane being widely preserved. Myoepithelial cells had lost their normal position and cellular orientation in the regions with intraductal carcinoma. Actin-positive, high-microfilament myoepithelial cells were frequent findings recorded from parenchyma in cases of invasive ductal carcinoma without specific differentiation, though cells of that kind were rarely detectable from parenchyma in cases of medullary carcinoma. Particular attention should be given to the presence of tumor cells differing from each other for their morphological and functional differentiation in cases of carcinoma of the mammary gland.