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Histomorphology and Histogenesis of Pleomorphic Adenoma

Z. Tepavcevic1,*, J. Sopta2 and S. Sankovi-Babi3
1 2 3

Faculty of Dentistry, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia Faculty of Medicine, University of Belgrade, 11000 Belgrade, Serbia ORL Clinic, KBC Zvezdara, Belgrade, Serbia
Abstract: The recent histogenetical concepts, the diagnostic criteria and the classification of the benign salivary gland tumours especially pleomorphic adenoma were pointed out in this paper, as well as nowadays assumptions of histogenesis and morphogenesis, the classification and pathological diagnosis of benign salivary gland tumours. The general insight in the studies that analysed advantages and controversies in the use of tissue markers, in histogenesis, and growth factors was performed. The immunohistochemical analysis, hybridisation in situ, the cell and tissue cultures and the chromosomal analyses can contribute to accurate pathological diagnosis of the salivary gland tumours.

Keywords: Immunohistochemistry, salivary gland tumours, pleomorphic adenoma. INTRODUCTION The primary epithelial salivary gland tumours are infrequent and make about 2% of the neoplasms of the head and the neck. These tumours have been showing a wide spectrum of the tumour histomorphology. As a dynamic and evolving field, the tumours of salivary glands were studied extensively and their classification was frequently changed [1, 2]. The histopathologic characteristics are varying in different parts of the same tumour, and different tumour subtypes are showing the similar or almost identical histomorphologic findings, which makes difficulties in differential diagnosis. This lead to the controversies in their nomenclature and diagnostic criteria. The recent progress in accurate definition of these modalities has been done by application of immune chemistry, hybridisation in situ, the cell and tissue cultures and the chromosomal analyses. In that way the controversies in histogeneses, clinicopathological fetures and the prognosis of salivary gland tumours could be clarified [2, 3]. The myoepithelial cells in normal salivary glands are of the unique structures and are the only cells in the normal tissue containing cytokeratin and the isoforms of actin and myosin. Associated with the muscle cells they announced the combined epithelial and mygenic differentiation. They develop by the early modification and differentiation out of the pluripotential ductal salivary cell about the tenth week of the intrauterine development. It has been assumed that the precursor of the myoepithelial cell has been a bright cell located in the terminal and striped channels. Their most important characteristic has been the presence of cytoplasmic filaments on the basal side. They consist of actin myofilaments, tropomyosin and myosin having the distribution similar to that in non striped muscle cells. The genesis of myoepithelial cells during the embryonic development had begun before the ductal differentiation. In the normal gland the myoepithelium is intimately connected to the acini, the intercalated and striped channels within the basal lamina [4, 6, 7, 9]. In the normal salivary gland the myoepithelial and basal cells form a continuous layer under the basal membrane outside the acini and ductal luminal cells. The subpopulation of ductal cells located on the non luminal parts of excretory and striped channels with much more heterogeneous and functionally more complex characteristics had been identified by the ultrastructure and immune histochemical studies. These cells had been called the ductal basal cells, but their nature and function had not been completely explained. They share numerous common markers with the myoepithelial ones and some of them, if not all, serve as the progenitory reserve cells for the neoplasia of salivary glands [6, 7, 11, 12]. The myoepithelial cells in the normal parotid gland could express glial fibrillar acidic protein (GFAP), an intermediary filament which normally has been expressed by the glial cells and the Schwanns cells of peripheral nerves [13-15]. Vimentin is a protein, an intermediary filament of mesenchymal cells and its expression by myoepithelial cells has still been the object of disputation among the authors [3, 5, 13]. The myoepithelial cells take part in the formation of the basal lamina. This is of importance in some hyperplastic and neoplastic alterations, where myoepithelial cells produced the fibrokinetins, laminin

*Address correspondence to this author at the Departement of Pathology, Faculty of Dentistry, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia; Tel: +381 63 7738 588; E-mail: tzvezdana@yahoo.com

2013 Pharma Professional Services

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and type III collagen. All these proteins are the constituent parts of basal lamina. The myoepithelial cells are also involved in the production of tenascin, of extracellular matrix, of glycoproteins. The altered myoepithelial cell may in the neoplastic proliferation show one or both of these characteristics. These cells have been assumed to be the key factors in the morphology of salivary gland tumours and in the morphologic variability. The role of myoepithelial cells in histogenesis of pleomorphic adenoma had been outstandingly studied. Nowadays it has been accepted that myoepithelial cells played the most significant role in the neoplastic process by expressing both the epithelial and the mesechymal structures in the majority of pleomorphic adenoma [5, 8, 10,16, 15]. The main morphologic types of modified myoepithelial cells are: 1 The star - like or myxoid cells present in the chondromyxoid fields of pleomorphic adenoma. 2 The fusiform and myoid cells that could be found in pleomorphic adenoma and in some types of myoepithelioma. 3 The hyalin or plasmocytoid cells that could be seen in pleomorphic adenoma and could be present in myoepithelioma. These cells show the cytoplasmic filaments in abundance creating the hyaline. 4 The bright or epithelial cells that could be found in many salivary tumours on the outer surface of ducts or of the duct like structures. They are characteristic for the myoepithelial carcinoma. The myoepithelial cells with mesenchymal characteristics secrete the mesenchymal mucin such as acid glycosamino glycane, elastin and tenascin. Vimentin is especially intensely positive in fusiform cells. The S-100 protein being a common marker for myoepithelium has been positive in chondroid, myxoid and star like cells, especially if they are associated with the myxoid stroma. The plasmocytoid myoepithelial cells have often been negative to the muscle markers. The epithelial differentiation could have the form of bright or squamous cells containing both the vimentin and keratin filaments [7, 8, 15]. The myoepithelial cells have been the central element in the histologic formation and organization of different salivary gland tumours. The cytomorphologic characteristics and the variability of extracellular products do explain the morphologic heterogeneity of these lesions [8, 15]. THE TUMOUR HISTOGENESIS AND MORPHOGENESIS The histopathologic diagnosis of salivary gland tumours has been based on the formula of the cell

differentiation. The architectonic orientations of the neoplastic tumour cell types and their similarities with the normal gland architecture are evident. The salivary neoplasms have shown the complex formula of the cell differentiation and organization. Nevertheless, the histogenetical concept had mainly been based on the belief that all salivary gland neoplasms had originated from the spare stem cells because of the presence of the spare stem cells in the normal salivary gland and of the semi-pluripotential concept for the tumour induction postulate [4, 9, 15]. It has been assumed that completely differentiated cell types in the mature gland have not been capable of proliferation and neoplastic transformation and that the spare cells have been either the non engaged stem cells or the only cells having a role in supplementation and reparation of the glandular tissue [4]. The detailed characteristics of intercalated and excretory ductal segments for confirmation of the semipotential bicellular concept and the role of specific cells in the geneses of salivary neoplasms had shown that a solid proof had been lacking to be able to describe the roles of individual stem cells and their differentiation pathways. The cellular organization of the salivary gland ducts, besides the presence or absence of myoepithelial cells, had an important role in the histogenetic classification of salivary tumours. Nevertheless, a smaller role of these cells might be contained in the diagnostics of numerous salivary gland tumours [13, 15, 18]. Numerous cell kinetics studies had shown the specific cell types such as the basal cells having been postulated in the semi-pluripotential bicellular concept as well as in their pathways of differentiation as the unique mechanism in the normal gland, glandular hyperplasia and regeneration. The studies had shown all cell types in the salivary glands: the acinus cells, the cells at all levels of ductal segments and the myoepithelial cells which, if not at all levels, could proliferate under different physiological and pathological conditions [3-5]. In human subjects, on the basis of the findings in the irradiated submandibular glands, the acinus cells, that had been excluded from the histogenetical theories of tumours, have shown a great proliferation potential. [3-5, 15, 16]. In in vitro findings in the human parotid and submandibular glands, the acinus cells had shown to possess the potential for the cellular proliferation. The basic cellular architecture in the tumour support or inclination to represent their normal duplication during the tumour genesis in the salivary gland tumours has reflected the basal cellular organisation of the normal

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tissue. That is why the new trends in tumour classification have been based mostly on histological and cytological similarities on the immune histochemical and ultrastructural levels between the mother tissue and the tumour. This might have the implications to the specific tumour type or types in their histogeneses. The differentiation of acinus cells into the ductlike cells and the squamous metaplasia and the potential of laminal cells to differentiate into the squamous cells as well as the metaplasia of the goblet shaped cells into the ciliar cells have been supported in the geneses of very different morphologies of salivary gland tumours [15, 16]. IMMUNOHISTOCHEMISTRY The immunohistochemical and ultrastructural studies had been widely applied to explain the cellular characteristics of salivary gland tumours. The complex and various profiles of the expression of cytokeratin in luminal cells at different levels of the ductal system and the heterogeneous potentials of differentiation in human parotid and submandibular glands equally with the participation of ductal/basal/myoepithelial cells may present a very heterogeneous formula of differentiation in normal and neoplastic glands. This concept of the ductal-acinus unity as the basic architectonic structure has further been under investigation. An extended version of the concept has proposed that the differentiation formula of salivary tumour cells from the ductal or acinus cell type and myoepithelial or basal cell type could appear individually, to get overlapped or to appear in various combinations [3, 5-7, 12, 16, 19]. Pleomorphic adenoma had been the most studied histopathologically, ultrastructurally and immune chemically. The main criteria for the diagnosis was based on the recognition of the unique mixture of evidently not linked together epithelial tumour cell elements in the tubular or duct like structures or of the solid fields and neoplastic myoepithelial cells with luminal and non luminal components in hyaline and myxochondroid regions. At the ultrastructural level, the luminal and non luminal cells may have a co-ordinated proliferation, that may be one of the factors in the production of the characteristic morphology. The dominant histomorphology in the tumour may be influenced by comparatively non co-ordinated proliferation even in different regions of the same tumours and the different differentiation of luminal and non luminal neoplastic myoepithelial cells and the potential for secondary extracellular products such as glycosaminoglycans from neoplastic myoepithelial cells which could influence the dominant histomorphology.

The intensive immune responsiveness to cytokeratin, moderate to epithelial membraneous antigen (EMA) and carcinoma embryonic antigen (CEA) has usually been present in luminal tumour cells of tubulo-ductal structures [5, 3, 7, 21]. The non luminal or neoplastic myoepithelial cells are morphologically heterogeneous and may be showing a positive response to cytokeratin, vimentin, S-100 protein, the neuron-specific enolase (NSE) and the glial fibrillar acid protein (GFAP) [3, 12, 15, 20]. The immunohistochemical and ultrastructural studies have shown that the fibronectin, laminin, collagen IV, tenascin and the collagen fibres and the glycosaminoglycans have been the main excretory products, and that they have been the possible factors in the production of the tumour stroma [2, 5]. The ductal epithelium of the normal salivary gland does not contain the muscle specific intermediary filament desmin or neurofilaments. The normal myoepithelial cells do contain the specific muscle actin (SMA). In pleomorphic adenoma the neoplastic myoepithelial cells do not always contain the specific muscle actin (SMA). These cells may express desmin, although in rare cases the expression has been independent on the muscular cell skeleton [3, 8, 13, 16, 20]. The heterogeneous expression of Ca binding S-100 proteins (S 100A1, S 100A2,S 100A4, S 100A6 and S 100B) has significantly been differing from the normal gland and had been found in the luminal and nonluminal tumour cell components of the pleomorphic adenoma. mRNA for S-100, obtained in situ by hybridisation in tumour cells of pleomorphic adenoma has been in correlation with immune histochemically detected S-100 protein.. The presence of not stained tumour cells may point to the heterogeneous population of the cells in-between luminal and nonluminal cells in relation to the S-100 protein expression. The immune fluorescence and the electron microscopy had shown that the S-100 protein had been absent in normal myoepithelium and that the S-100 immune responsive structures similar to the myoepithrlium had been in fact the abundant plexuses of non-myelinized nerves which the salivary acinus cells had been richly endowed with [3, 5, 19, 20]. On the level of the light microscopy two types of modified myoepithelial cells: -the fusiform or myofibrillar and the plasmocytoid-have been described. Nevertheless, the lack of myogenic differentiation in plasmocytoid cells points out that they have been the subtypes of the modified ones, although the careful ultrastructural examination may discover the myofilaments in the plasmocytoid myoepitheliomas [1,

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2]. These cells have shown a heterogeneous features by showing the epithelial, mesenchymal end neural differentiation on the immune histochemical and ultrastructural levels [3, 5, 9, 13]. The explanation of the cellular characteristics of non-luminal or the neoplastic myoepithelial cells in pleomorphic adenoma and myoepitheliomas, the lack of detected myofilaments by immune histochemistry or by the routine ultrastructure studies of the specific muscle actin (SMA), of the marker specific to the normal myoepithelial cells have not excluded the diagnosis of pleomorphic adenoma, or myoepithelioma when the growth indices and the cytology have pointed out to such a diagnosis. The immune histochemical microscopy may show the specific muscle actin (SMA) that could be detected when the myofilaments of muscle actin seem to be absent or at the minimum on the routine preparations. When the markers of the normal salivary gland epithelium have been used to determine the range of their expression in the neoplastic myoepithelial cells of pleomprphic adenoma and myoepithelioma it has been shown that in nonluminal cells there has been incomplete expression or the absence of myoepithelial/basal cell markers the specific muscle actin (SMA), cytokeratin 14 (CK 14) and that there has been a general expression of vimentin and glial fibrillar acidic protein (GFAP). The myoepithelial cells have been responsible for the production and accumulation of extracellular materials, often in an excessive quantity. The extracellular regions in pleomorphic adenoma have been no true stroma, but they have been a secondary alteration produced by neoplastic myoepithelial cells. The basal lamina and the regions containing glycosaminoglycan, and having been located in -between, have been surrounded by the modified myoepithelial cells and their cytoplasmatic extensions may be the initial phase in the development of myxoid alterations. The next one during the synthesis of the basal lamina, of collagen, flycosaminoglycan and glycoprotein has gradually been separating the modified myoepithelial cells resulting in the separation of individual tumour cells or their small groups in the fusion with the adjacent intercellular spaces leading to the formation of the myxoid or hyaline stroma [3, 5, 9]. The histogenesis and differentiation of myoepithelium-like cells called neoplastic cells or modified myoepithelial cells have frequently formed the dominant histopathologic characteristic, especially in pleomorphic adenoma and in numerous salivary neoplasms and have incompletely determined the

criterion for discrimination of the neoplastic product from the normal myoepithelial cell and have still remained unexplained. The presence of myofilaments at the level of ultrastructure and the expression of the specific muscle actin (SMA) have been the criteria for identification of the normal myoepithelium. The differentiation of these cells out of the normal myoepithelium or the common stem cells has presented the aberrant differentiation of the tumour cell component has not been sufficiently explained. The incomplete expression or the absence of the myoepithelial/basal cell markers, the specific muscle actin (SMA) and cytokeratin (CK) and the expression of vimentin and glial fibrillar acidic protein (GFAP) may be common to both tumour types. For the reasons of description the myoepithelial cells may be classified into those being positive and those being negative to the specific muscle actin (SMA) and/or the myofilaments. The ductal cells in the early stage of development may be expressing the specific muscle actin (SMA) [3-5, 21]. The immune responsiveness of the osseal morphogenetic protein (BMP) producing the chondroosseal tissue in the osseal system by activation and proliferation of non-differentiated mesenchymal cells has been found in modified myoepithelial cells of pleomorphic adenoma, producing chondroid alterations [5]. These findings have been of help to conclude that the modified myoepithelial cells have been responsible for the different histopathologic morphology of pleomorphic adenoma with myxoid, hyaline and chondroid fields. The excessive accumulation of these materials had been studied in adenoid cystic carcinoma, carcinoma in the pleomorphic adenoma, myoepitheliomas, epithelial-myoepithelial carcinomas and in numerous salivary adenomas [3-5]. Therefore the surplus of secretion products from tumour cells, especially from the neoplastic myoepithelial cells has been determining various histologic pictures in many salivary gland tumours altering the cell to-cell and cellto-extracellular matrix interactions. MOLECULAR GENETICS Until the beginning of 1980 the belief that the benign tumours, except the meningioma had normal chromosomal constitution had been dominant. In 1980 a specific chromosomal aberration in benign tumours of salivary glands of a certain histological type Adenoma polymorphae had been described. These tumours have shown specific chromosomal translocations including the regions 3p21, 8q12 and

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12q13-15 [23]. The most frequent aberration has been t (3,8) (p21;q12) representing at the same time the first tumour specific translocation noticed in the benign forms. The cytogenetical analyses of pleomorphic adenomas have shown the presence of clonal chromosomal aberrations on the base of which the classification in 3 cytogenetic subgroups had been made: 1) Pleomorphic adenomas with structural chromosomal aberrations of the chromosome 8 in the region 8q12. 2) Pleomorphic adenomas with structural aberrations of the chromosome 12 in the region 12q1315. 3) Pleomorphic adenomas with normal karyotypes possessing also the cell lines with the trisomy 8 or the loss of the chromosome Y as mosaicism [22]. THE PROLIFERATION POTENTIAL In a tumour the invasion to the capsule may appear in spite of that a benign tumour was in question. The pleomorphic adenoma may also become malignantly altered into carcinoma in. a certain percent of cases. Numerous authors had established a significant correlation between the cellular proliferation activity and the tumour prognosis. The pleomorphic adenoma has shown a low proliferating nuclear cell antigen (PCNA) index. Ki-67 has shown a greater affinity to the solid-trabecular and ductal regions while, according to the findings of the authors it has shown a nearly negative response in mesenchymal regions [12, 14, 19, 23-25]. DIFFERENTIAL DIAGNOSIS Basal Cell Adenoma The cellular variant of pleomorphic adenoma with few or no myxodhondroid stroma may be difficult to be differentiated from the tubular or trabecular basal cell adenoma. In the pleomorphic adenoma the tumour cells usually neither are well organised into trabecular structures nor are they separated by narrow stripes of stroma. The sharp difference between the fibrous stroma and the outer basal/myopithelial cells in basal cell adenoma and the clear separation of myoepithelial cells from the myxoid stroma in pleomorphic adenoma have made the essential difference. Myoepithelioma The pleomorphic adenoma may have an outstanding component of neoplastic myoepithelial cell, what might present a diagnostic problem. The myoepitheliomas have however been devoid of ductal cells or have less than 5 percent of these structures,

while the ductal and glandular formations have been a constituent part of pleomorphic adenoma. Mucoepidermoid Carcinoma This neoplasm may be confused with the pleomorphic adenoma because the intermediary cell population of carcinoma has been very similar to the basal/myoepithelial cells of pleomorphic adenoma. The intermediary cells, in spite of their potential for the production of extracellular material cannot create the myxochondroid stroma. The squamous differentiation, when present in pleomorphic adenoma, is generally well developed and may show keratinisation, the characteristic little evident in carcinoma. The goblet like cells are infrequently present in pleomorphic adenoma. The plasmocytoid cells had not been described in carcinoma, so that their presence makes a marker of pleomorphic adenoma [25]. Adenoid Cystic Carcinoma This neoplasm has been, in contrast to the pleomorphic adenoma, of the infiltrative growth and it often has been associated with the perineural invasion. The cribriform growth may also appear in pleomorphic adenoma. It usually has been bound to the basal/myoepithelial cells which have a wide leaf like growth. This is in contrast with the sharp circumscriptions of tumour nests in carcinoma within which there is the extracellular material. The tubular structures being the general characteristic of pleomorphic adenoma, in carcinoma have been small and infrequent, especially in solid and cribriform carcinoma forms. Carcinoma in the Pleomorphic Adenoma At the great magnification it has been noticed the cellular atypism and the high grade mitotic activity both in luminal and in non-luminal cells. The cellular pleomorphism may be present focally in pleomorphic adenoma, while in carcinoma it is widespread. In carcinoma it appears the infiltration of the capsule [25]. When the pleomorphic adenoma is of the outstanding myxoid type, in differential diagnosis there could be involved schwannoma, myxoma or even the malignant fibrous histiocytoma. The diagnosis of the pleomorphic adenoma could be made on the base of the local presence of epithelial elements on the borderlines as the presence of minimum tubular structures. Sometimes it has been necessary to do the immune histochemical staining.

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Received on 14-11-2013

Accepted on 21-11-2013

Published on 30-11-2013

2013 Tepavcevic et al.; Licensee Pharma Professional Services. This is an open access article licensed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted, non-commercial use, distribution and reproduction in any medium, provided the work is properly cited.