Immunohistochemical detection of amelogenin and cytokeratin 19 in epithelial odontogenic tumors

OBJECTIVE: Epithelial odontogenic tumors exhibit considerable histological variation and are classified into several benign and malignant entities. Expression of amelogenin and cytokeratin 19 (CK19), that are potentially useful polypeptides for identification of odontogenic epithelial components, was evaluated in various types of epithelial odontogenic tumors. MATERIALS AND METHODS: Specimens of 33 ameloblastomas, three calcifying epithelial odontogenic tumors (CEOTs), two clear cell odontogenic tumors (CCOTs) and five malignant ameloblastomas were examined by immunohistochemistry using anti-amelogenin and anti-CK19 antibodies. RESULTS: Immunohistochemical reactivity for amelogenin was detected in many peripheral columnar or cuboidal cells and some central polyhedral cells in ameloblastomas, and histological variants showed various degrees of amelogenin expression. Expression of CK19 was diffusely present in neoplastic cells in ameloblastomas, and decreased expression was found in keratinizing cells of acanthomatous variants and some neoplastic cells of desmoplastic variants. In CEOTs, immunohistochemical reactivity for amelogenin was detected in neoplastic cells and intercellular amyloid-like materials, whereas CK19 was expressed in neoplastic cells. CCOTs showed positive reactivity for amelogenin and CK19 in neoplastic cells. Malignant ameloblastomas exhibited various degrees of amelogenin expression with constant CK19 expression in neoplastic cells. CONCLUSION: Diverse types of epithelial odontogenic tumors express amelogenin and CK19, suggesting that these tumors have ameloblastic differentiation or odontogenic epithelial properties.     

Clonal nature of odontogenic tumours

Background:  Although clonal origin is an essential step in the comprehension of neoplasias, there have been no studies to examine whether odontogenic tumours are derived from a single somatic progenitor cell. The purpose of this study was to investigate the clonal origin of odontogenic tumours.
Methods:  Fresh samples of seven ameloblastomas, two odontogenic mixomas, two adenomatoid odontogenic tumour, one calcifying odontogenic cyst, one calcifying epithelial odontogenic tumour (CEOT) and six odontogenic keratocyst (OKC) of female patients were included in this study. After DNA extraction, the HUMARA gene polymorphism assay was performed.
Results:  Most of the informative odontogenic lesions studied (12 out of 16) showed a monoclonal pattern. Among the polyclonal cases, two were OKC, one CEOT and one odontogenic mixoma.
Conclusions:  Our results suggest that most odontogenic tumours are monoclonal.     

Immunohistochemical detection of hepatocyte growth factor, transforming growth factor-β and their receptors in epithelial odontogenic tumors

BACKGROUND: Tumors derived from odontogenic epithelium exhibit considerable variation and are classified into several benign and malignant entities. To clarify the role of growth factors in oncogenesis, cytodifferentiation and progression of epithelial odontogenic tumors, expression of hepatocyte growth factor (HGF), transforming growth factor-beta (TGF-beta) and their receptors were analyzed in these tumors as well as in tooth germs. METHODS: Specimens of five tooth germs, 34 ameloblastomas, three calcifying epithelial odontogenic tumors (CEOTs), two clear cell odontogenic tumors (CCOTs), five adenomatoid odontogenic tumors (AOTs), six calcifying odontogenic cysts (COCs) and six malignant ameloblastomas were examined immunohistochemically with the use of antibodies against HGF, TGF-beta and their receptors. RESULTS: In tooth germs and epithelial odontogenic tumors, immunoreactivity for HGF and TGF-beta was detected in both epithelial and mesenchymal cells, while expression of their receptors was found only in epithelial cells. In tooth germs and main types of ameloblastomas, HGF and TGF-beta reactivity was marked in epithelial cells near the basement membrane, and their receptors were diffusely positive in most epithelial cells. In subtypes of ameloblastomas, reduced expression of HGF, c-Met and TGF-beta and increased reactivity for TGF-beta receptors were detected in keratinizing cells in acanthomatous ameloblastomas, and granular cells in granular cell ameloblastomas demonstrated little or no expression of HGF, TGF-beta or their receptors. As compared with main types of ameloblastomas, basal cell ameloblastomas showed high HGF reactivity, and desmoplastic ameloblastomas exhibited elevated reactivity for TGF-beta and its receptors. Neoplastic cells in CEOTs, AOTs and COCs showed reactivity for HGF, TGF-beta and their receptors. Elevated HGF and TGF-beta reactivity was found in pseudoglandular cells in AOTs, and high expression of their receptors was noted in ghost cells in COCs. Metastasizing ameloblastomas showed similar expression patterns of HGF, TGF-beta and their receptors to those of benign ameloblastomas, while CCOTs and ameloblastic carcinomas had increased HGF expression and low reactivity for TGF-beta and its receptors as compared with benign ameloblastomas. CONCLUSIONS: Immunohistochemical localization of HGF, TGF-beta and their receptors in tooth germs and epithelial odontogenic tumors supports the hypothesis that HGF and TGF-beta act on epithelial cells via paracrine and autocrine mechanisms. Altered expression of the agents in these epithelial odontogenic tumors, especially subtypes of ameloblastomas, AOTs and COCs, suggests that HGF and TGF-beta signaling might affect differentiation of neoplastic odontogenic epithelial cells. Activated HGF/c-Met pathway and reduced TGF-beta signaling in CCOTs and ameloblastic carcinomas may be associated with the malignant potential of these epithelial odontogenic tumors.     

Podoplanin expression in human tooth germ tissues and cystic odontogenic lesions: an immunohistochemical study

J Oral Pathol Med (2010) 39 : 115–120
Background:  Podoplanin expression was described in mouse tooth germ and apical bud cells. The aim of this study was to analyse the podoplanin expression of human tooth germ tissues, adult teeth and odontogenic lesions immunohistochemically.
Study Design:  Nine human tooth germ biopsies and seven healthy permanent teeth extracted for orthodontic reasons were examined. Anti-podoplanin (D2-40) reactivity was investigated immunohistochemically. Five well-defined cystic odontogenic lesions (10 radicular cysts, 10 follicular cysts, three keratocystic odontogenic tumours, five ameloblastomas, and two adenomatoid odontogenic tumours) were analysed simultaneously.
Results:  Podoplanin expression was detected in the majority of epithelial and ecto-mesenchymal cells of human tooth germ tissues, odontoblasts and superficial dental pulp fibroblasts of permanent teeth. Cystic odontogenic lesions revealed positive reactions predominantly at the invasion front edge within basal epithelial layers.
Conclusion:  Podoplanin appears to be involved in the orthologic and pathologic processes of the formation of elongated cell extensions and odontoblastic fibers, in the epithelial–mesenchymal transition and local invasion during tooth germ development as well as in both reactive and neoplastic odontogenic cystic lesions.     

Comparison of angiogenesis in keratocystic odontogenic tumours, dentigerous cysts and ameloblastomas

Objective:  The aim of the present study was to evaluate and compare angiogenesis in keratocystic odontogenic tumours, dentigerous cysts (DCs) and ameloblasomas using monoclonal antibody against CD34.
Materials and methods:  Microvessel density was assessed in a total of 53 cases including 20 keratocystic odontogenic tumours, 13 DCs and 20 ameloblastomas (14 solid and six unicystic variants). Microvessel density was expressed as the mean number of microvessels per high-power-field.
Results:  Statistically significant differences in mean microvessel density were observed between keratocystic odontogenic tumours, DCs and solid ameloblastomas ( P  < 0.001). Mean microvessel density was significantly higher in solid ameloblastomas compared with both keratocystic odontogenic tumours and DCs; and was also significantly higher in keratocystic odontogenic tumours than in DCs.
Conclusion:  Within the limitations of the present study, it can be suggested that angiogenesis may be one of the mechanisms possibly contributing to the different biological behaviours of keratocystic odontogenic tumours, DCs and solid ameloblastomas.     

Perlecan-rich epithelial linings as a background of proliferative potentials of keratocystic odontogenic tumor

Background:  The intraepithelial deposit of perlecan, a basement membrane-type heparan sulfate (HS) proteoglycan, has been demonstrated in neoplastic conditions such as salivary gland tumors, odontogenic tumors, and oral carcinoma in situ . Our aim was to determine whether perlecan turnover was enhanced in the lining cells of keratocystic odontogenic tumor (KCOT), which had been recently renamed from odontogenic keratocyst because of its accumulated evidence of neoplasm, as a possible background for neoplastic proliferation.
Methods:  Ten surgical specimens from each of KCOT, dentigerous cyst, and radicular cyst were examined for the expressions of perlecan core protein, HS chains, heparanase, and Ki-67 by immunohistochemistry and in situ hybridization.
Results:  In KCOT, perlecan core protein and HS chains were localized on the cell border from the parabasal to subkeratinized layers of the lining epithelium. Heparanase was localized in a similar fashion to those for perlecan and HS chains but was within the cytoplasm. mRNA signals for perlecan core protein and heparanase were mostly compatible with their protein signals. Ki-67-positive cells were localized mainly in the second basal cell layers with definitely higher labeling indices (approximately 31.3%, second layer). In contrast to KCOT, dentigerous cysts and radicular cysts had no perlecan, HS chains, and heparanase deposition in their linings with extremely lower Ki-67 indices (0.4–0.8%).
Conclusion:  The result suggests that the characteristic intra-lining-epithelial deposit of perlecan in KCOT, which has never been seen in other cystic jaw lesions, is a new evidence supporting the neoplastic nature of KCOT.     

Immunoexpression of MMPs-1, -2, and -9 in ameloblastoma and odontogenic adenomatoid tumor

Objective:  The aim of this study was to evaluate and compare the expression of metalloproteinases-1, -2, and -9 in solid ameloblastoma and adenomatoid odontogenic tumor.
Methods:  A total of 20 cases of solid ameloblastoma and 10 cases of adenomatoid odontogenic tumors were selected and immunohistochemically assessed. Metalloproteinases-1, -2, and -9 immunoexpression and their distribution pattern were noted and semiquantitatively scored. The scores obtained were statistically analyzed.
Results:  Matrix metalloproteinase (MMP)-1 showed a predominant expression in both tumors and was found in stroma and parenchyma. For MMP-2, there was a varied expression, with 80% and 60% of immunoreactive tumor cells in ameloblastoma and adenomatoid odontogenic tumor respectively. Regarding stromal cells, 65% of ameloblastomas and 80% of adenomatoid odontogenic tumors showed positivity. There was immunoexpression of the MMP-9 in parenchymal and stromal cells in all cases of both tumors analyzed. A statistically significant difference in the expression of MMP-1 in relation to the expression of MMP-2 and -9 in ameloblastomas ( P  < 0.001) was observed.
Conclusion:  The results suggest that these metalloproteinases are related to growth and progression of tumors analyzed, and particularly in ameloblastoma, its highest aggressiveness may be, in part, a result of the active participation of the stromal cells and their products, such as the MMPs studied.     

Cystic variant of calcifying epithelial odontogenic tumor

The calcifying epithelial odontogenic tumor is a rare benign odontogenic neoplasm of the jaw. Clinically, calcifying epithelial odontogenic tumor manifests as an intraosseous lesion (central type) in the majority of cases (95%). Extraosseous or peripheral lesions account for less than 5% of cases. Calcifying epithelial odontogenic tumor can be associated with an impacted tooth and give a radiographic simulation of dentigerous cyst. Most calcifying epithelial odontogenic tumors are solid in nature, histopathologically, and might have few cyst-like spaces within them. However, a true cystic calcifying epithelial odontogenic tumor is a rare possibility. We describe a case of a true cystic variant of calcifying epithelial odontogenic tumor in a 30-year-old male, which to our knowledge, is only the second reported case.     

Immunohistochemical demonstration of enamel proteins in odontogenic tumors

Immunohistochemical localization of two enamel proteins, amelogenin and enamelin, in comparison with that of keratin, was determined in odontogenic tumors and the allied lesions in order to verify functional differentiation of the tumor cells as ameloblasts. Amelogenin and enamelin were demonstrated in small mineralized foci and in the tumor cells surrounding them in adenomatoid odontogenic tumor (AOT), calcifying epithelial odontogenic tumor (CEOT), and calcifying odontogenic cyst (COC). Hyaline droplets in AOT showed positive staining for both enamel proteins. These mineralized and hyaline materials were not positive for keratin, although tumor cells were positive. On the other hand, no immunoreaction for enamel proteins was obtained in ameloblastoima and odontogenic epithelial cell nests within myxoma and epulis. The results suggest that tumor cells of AOT and CEOT and lining epithelial cells of COC show ameloblastic differentiation in part, but that ameloblastoma cells do not attain functional matauration as secretory phase ameloblasts.     

Immunohistochemical demonstration of bone morphogenetic protein in odontogenic tumors

The aim of the present study was to describe the expression and distribution of bone morphogenetic protein (BMP) in odontogenic tumors by immunohistochemistry using monoclonal antibody against bovine BMP (BMPMcAb). Eight types of odontogenic tumors (44 cases), including ameloblastoma (20 cases), cementifying fibroma (8 cases), benign cementoblastoma (5 cases), dentinoma (3 cases), compound odontoma (2 cases), adenomatoid odontogenic tumor (2 cases), calcifying epithelial odontogenic tumor (2 cases) and odontogenic fibroma (2 cases), were studied. The results showed that, according to the immunostaining pattern of BMPMcAb, tumors could be classified into two types: all cementifying fibro-mas, benign cementoblastomas. Dentinomas, odontogenic fibromas. and compound odontomas demonstrated a positive reaction, whereas all ameloblastomas, adenomatoid odontogenic tumors, and calcifying epithelial odontogenic tumors were negative. BMPMcAb-positive odontogenic tumors were those tumors with formation of enamel, dentin, cementum or bone. Therefore. BMP might play an important role in the formation of calcified dental tissues and the development of odontogenic tumors contaning such tissues.     

Immunostaining of involucrin in odontogenic epithelial tumors and cysts

An immunoperoxidase method was used to detect involucrin in 47 odontogenic tumors and 35 radicular cysts. Of a total of 40 ameloblastomas, 9 cases were positive for involucrin expression and those positive cases exhibited acanthomatous or follicular patterns. Squamous odontogenic tumors were strongly positive for involucrin, whereas adenomatoid odontogenic tumors gave a negative staining reaction. Involucrin expression in odontogenic tumors was divided into three categories: single cell positive, focally positive, and squamous metaplastic cell positive. Radicular cysts showed a very irregular distribution of involucrin; nonstratified epithelium was generally negative or showed only trace staining for involucrin, whereas suprabasilar stratified squamous epithelial cells were strongly positive. Cells positive for involucrin in odontogenic tumors and in cystic epithelium are probably direct signs of epithelial differentiation; such cells were squamoid in appearance.     

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牙源性肿瘤是颌骨最常见病损之一,临床上绝大多数牙源性肿瘤均发生在颌骨内。良性肿瘤多无自觉症状,直至肿瘤生长到一定程度出现颌面部组织膨隆方被发现,早期患者多是在做牙髓治疗或体检照片时无意中发现的,除非继发感染,患者一般没有疼痛等不适感觉。牙源性肿瘤的病理学分类很多,基于局部生物学行为的特点,WHO在2005年将原来的牙源性角化囊肿和牙源性钙化囊肿归列入牙源性肿瘤。牙源性肿瘤的临床表现多样,一般需要结合病理学检查确诊。手术处理是治疗牙源性肿瘤惟一有效的方法,但如何选择有效术式是临床医师需要慎重考虑的问题。本文就几种常见牙源性肿瘤的外科治疗作了简要概述。     

Frequency of odontogenic cysts and tumors: a systematic review

A systematic review of the literature from 1993 to 2011 was undertaken examining frequency data of the most common odontogenic cysts and tumors. Seven inclusion criteria were met for the paper to be incorporated. In the preliminary search 5231 papers were identified, of these 26 papers met the inclusion criteria. There were 18 297 odontogenic cysts reported. Of these there were 9982 (54.6%) radicular cysts, 3772 (20.6%) dentigerous cysts and 2145 (11.7%) keratocystic odontogenic tumors. With the reclassification of keratocystic odontogenic tumor in 2005 as an odontogenic tumor, there were 8129 odontogenic tumors reported with 3001 (36.9%) ameloblastomas, 1163 (14.3%) keratocystic odontogenic tumors, 533 (6.5%) odontogenic myxomas, 337 (4.1%) adenomatoid odontogenic tumors and 127 (1.6%) ameloblastic fibromas. This systematic review found that odontogenic cysts are 2.25 times more frequent than odontogenic tumors. The most frequent odontogenic cyst and tumor were the radicular cyst and ameloblastoma respectively.     

Expression of E-cadherin and α-catenin in epithelial odontogenic tumours: an immunohistochemical study

To clarify the possible role of cell adhesion in epithelial odontogenic tumors, expression of E-cadherin and alpha-catenin was examined by an immunohistochemical method. These molecules showed pericellular distribution in epithelial cells of the tooth germ and its derived tumors. In ameloblastomas, E-cadherin and alpha-catenin were expressed strongly in central polyhedral cells and slightly in peripheral columnar cells. These features resembled those of epithelial components in the tooth germ tissues, retaining cytodifferentiation of odontogenic epithelium. Expression of the molecules in the variants of ameloblastomas showed loss in the keratinizing areas and reduction in the granular cell clusters, suggesting terminal differentiation of the tumor cells. Calcifying epithelial odontogenic tumors and a clear cell odontogenic tumor preserved E-cadherin and alpha-catenin expression without a specific feature for histogenesis or cytodifferentiation. One case of two malignant ameloblastomas showed prominent reduction in expression of E-cadherin and alpha-catenin.     

Clear cell change in a calcifying odontogenic cyst

A case of calcifying odontogenic cyst exhibiting clear cell change is described. Histomorphologic similarities to the clear cell type of calcifying epithelial odontogenic tumor were noted and the importance of differential diagnosis from clear cell types of salivary gland neoplasms and metastatic hypernephroma are emphasized.     

International collaborative study on ghost cell odontogenic tumours: calcifying cystic odontogenic tumour, dentinogenic ghost cell tumour and ghost cell odontogenic carcinoma

Background:  Calcifying odontogenic cyst was described first by Gorlin et al. in 1962; since then several hundreds of cases had been reported. In 1981, Prætorius et al. proposed a widely used classification. Afterwards, several authors proposed different classifications and discussed its neoplastic potential. The 2005 WHO Classification of Odontogenic Tumours re-named this entity as calcifying cystic odontogenic tumour (CCOT) and defined the clinico-pathological features of the ghost cell odontogenic tumours, the CCOT, the dentinogenic ghost cell tumour (DGCT) and the ghost cell odontogenic carcinoma (GCOC).
Methods:  The aim of this paper was to review the clinical-pathological features of 122 CCOT, DGCT and GCOC cases retrieved from the files of the oral pathology laboratories from 14 institutions in Mexico, South Africa, Denmark, the USA, Brazil, Guatemala and Peru. It attempts to clarify and to group the clinico-pathological features of the analysed cases and to propose an objective, comprehensive and useful classification under the 2005 WHO classification guidelines.
Results:  CCOT cases were divided into four sub-types: (i) simple cystic; (ii) odontoma associated; (iii) ameloblastomatous proliferating; and (iv) CCOT associated with benign odontogenic tumours other than odontomas. DGCT was separated into a central aggressive DGCT and a peripheral non-aggressive counterpart. For GCOC, three variants were identified. The first reported cases of a recurrent peripheral CCOT and a multiple synchronous, CCOT are included.
Conclusions:  Our results suggest that ghost cell odontogenic tumours comprise a heterogeneous group of neoplasms which need further studies to define more precisely their biological behaviour.     

The immunoprofile of odontogenic keratocyst (keratocystic odontogenic tumor) that includes expression of PTCH, SMO, GLI-1 and bcl-2 is similar to ameloblastoma but different from odontogenic cysts

Background:  The aggressive biological behavior of odontogenic keratocysts (OKCs), unlike that of other odontogenic cysts, has argued for its recent re-classification as a neoplasm, 'keratocystic odontogenic tumor'. Identification of mutations in the PTCH gene in some of the OKCs that were expected to produce truncated proteins, resulting in loss of control of the cell cycle, provided additional support for OKCs having a neoplastic nature.
Methods:  We investigated the immunohistochemical expression of the sonic hedgehog (SHH) signaling pathway-related proteins, PTCH, smoothened (SMO) and GLI-1, and of the SHH–induced bcl-2 oncoprotein in a series of primary OKC (pOKC), recurrent OKC (rOKC) and nevoid basal cell carcinoma syndrome-associated OKCs (NBCCS-OKCs), and compared them to solid ameloblastomas (SAMs), unicystic ameloblastomas (UAMs), 'orthokeratinized' OKCs (oOKCs), dentigerous cysts (DCs) and radicular cysts (RCs).
Results:  All studied lesions expressed the SHH pathway-related proteins in a similar pattern. The expression of bcl-2 in OKCs (pOKCs and NBCCS-OKCs) and SAMs was significantly higher than in oOKCs, DCs and RCs ( P  <   0.001).
Conclusions:  The present results of the immunoprofile of OKCs (that includes the expression of the SHH-related proteins and the SHH-induced bcl-2 oncoprotein) further support the notion of OKC having a neoplastic nature. As OKCs vary considerably in their biologic behavior, it is suggested that the quality and quantity of interactions between the SHH and other cell cycle regulatory pathways are likely to work synergistically to define the individual phenotype and corresponding biological behavior of this lesion.     

Significance of podoplanin expression in keratocystic odontogenic tumor

J Oral Pathol Med (2010) 39 : 110–114
Background:  The most important clinical features of the keratocystic odontogenic tumor (KCOT) are its potential for locally destructive behavior, a tendency to recur, and its origin in the odontogenic epithelium. The clinical features of KCOT are similar to those of ameloblastoma (AM). Histologically, KCOT is distinguished from jaw cyst with keratinization (orthokeratinized odontogenic cyst; OOC). However, current scientifically based clinical parameters cannot predict any potential for neoplastic behavior, or aggressive and localized invasiveness, in patients with KCOT. We have shown that podoplanin, a lymphatic endothelial marker, is highly expressed in AM. The purpose of this study was to determine the usefulness of podoplanin for reclassification of the odontogenic keratocyst (OKC) from cyst to tumor status.
Methods:  Paraffin-embedded tissue specimens of 57 OKCs (46 KCOTs and 11 OOCs) and 15 dentigerous cysts (DCs) were immunohistochemically examined using antibody against podoplanin.
Results:  Immunohistochemical reactivity for podoplanin was detected in the cell membrane and cytoplasm of most of the basal and suprabasal layer, areas of budding basal cell proliferation, epithelial nests and peripheral cells of daughter cysts in the stromal connective tissue in KCOTs. In the case of OOC and DC, only cases associated with inflammation were positive for podoplanin.
Conclusion:  Podoplanin is strongly expressed in KCOTs in comparison with OOCs. The pattern of staining for podoplanin in KCOT could be related to its neoplastic nature, and suggests a role of the protein in tumor invasiveness.     

A comparative study of epithelial cell proliferation between the odontogenic keratocyst, orthokeratinized odontogenic cyst, dentigerous cyst, and ameloblastoma

OBJECTIVES: The aim of the present study was to compare the proliferation index of the epithelial cells between odontogenic keratocysts (OKC), orthokeratinized odontogenic cysts (OOC), dentigerous cysts (DC), and ameloblastomas. MATERIALS AND METHODS: The proliferation index, employing a novel cell proliferation marker IPO-38, was studied by the immunohistochemical technique in 10 OKC, seven OOC, eight DC and 10 ameloblastomas. RESULTS: The ameloblastoma had no higher labeling index (LI) of IPO-38 than the OKC (P = 0.910) but had higher LI than the OOC (P = 0.001) and DC (P = 0.000); the OKC had higher LI than the OOC (P = 0.002) and DC (P = 0.000); and the OOC had higher LI than the DC (P = 0.011). IPO-38-positive cells in the OKC and OOC were located principally in the suprabasal cell layers while the ameloblastoma were found in the peripheral portion in particularly, the follicular and plexiform types. CONCLUSION: These findings support previous studies that the proliferation indices are useful in predicting the different biological behavior of the odontogenic lesions and the OKC should be regarded as a benign tumor rather than simply an odontogenic cyst.