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Clinico-Pathological Evaluation of Oral Melanotic Macule, Oral Pigmented Nevus and Oral Mucosal Melanoma

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Clinico-Pathological Evaluation of Oral Melanotic Macule, Oral Pigmented Nevus and Oral Mucosal Melanoma Rosario Rivera Buery et al.: Evaluation of Oral Melanotic Lesions Journal of Hard Tissue Biology 19[1] (2010) p57-64 © 2010 The Hard Tissue Biology Network Association Printed in Japan, All rights reserved. CODEN-JHTBFF, ISSN 1341-7649 Online ISSIN 1888-828X Original Clinico-pathological Evaluation of Oral Melanotic Macule, Oral Pigmented Nevus and Oral Mucosal Melanoma Rosario Rivera Buery1) ,Chong Huat Siar2), Naoki Katase3), Masae Fujii 3), Han Liu3), Midori Kubota3), Ryo Tamamura3), Hidetsugu Tsujigiwa3) and Hitoshi Nagatsuka3) 1)College of Dentistry, University of the East, Manila, Philippines. 2)Department of Oral Pathology, Oral Medicine and Periodontology, Faculty of Dentistry, University of Malaya, Malaysia. 3)Department of Oral Pathology and Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama City, Japan. (Accepted for publication, February 8, 2010) Abstract: Focal pigmented melanocytic lesions rarely occur in the oral cavity but should not be taken for granted for they may represent markers or risks for oral mucosal melanoma (OMM). The study investigated the clinical and pathological features of focal pigmented lesions of melanocytic origin focusing on oral melanotic macule (oral MM), oral pigmented nevus (OPN) and OMM. Immunohistochemistry was employed with S100, HMB-45, Melan A, c-kit and Ki-67. Oral MM mostly occurred on the gingiva and buccal mucosa while OPN mostly occurred on the buccal mucosa. OMM occurred on the palate and gingiva. A female gender predilection was observed in oral MM. Most of the benign lesions were less than 6 mm in diameter while OMM had greater than 10 mm in diameter. Benign lesions occurred in almost the same location as that of OMM. S100 and c-kit were detected in most benign cases while HMB-45 and Melan A were focally detected in some cases. S100, HMB-45 and Melan A expressions were detected in all cases of OMM. Ki-67 was only detected at the epithelial basal layer in oral MM and was completely negative in nevus cells. In OMM, Ki-67 was more than 80%. In conclusion, oral MM and OPN may not be markers of risk for OMM but excisional biopsy is highly recommended for clinically undefined lesions greater than 6 mm in diameter. The combination of S100, c-kit, HMB-45 and Melan A can be utilized to support the diagnosis of OMM. Keywords: Oral melanotic macule, Oral pigmented nevus, Oral mucosal melanoma, Immunohistochemistry, Melanoma markers Introduction tumor so-called oral malignant melanoma (OMM). This justifies Pigmented lesions in the oral cavity may be classified as that melanocytic lesions should not be ignored and a definite melanotic and non-melanotic lesions 1). One factor causing oral diagnosis must be established. pigmentation is over production of melanin pigments. Melanocytes In contrast to cutaneous melanoma, predisposing factors and may be found in the oral mucosa but may not be noticeable because precursor lesions have not been thoroughly clarified in OMM. of their low level of pigment production. However, when they are Precise distinction of a pigmented melanocytic lesion is essential active in pigment production or proliferation, they may be as it may represent a benign stimulant, a potential precursor or as responsible for several oral pigmentations ranging from focal to a marker of risk for OMM. Consequently, focal pigmentations diffuse and physiologic to malignant neoplasm 1). Diffuse such as oral melanotic macule (oral MM), oral pigmented nevus pigmentations are mostly systemic-related, drug-induced or caused (OPN) and OMM were investigated. Oral MM is a benign by exogenous pigments while most focal pigmentations are pigmented lesion caused by increased melanin production without brought about by over production of melanin. Although most increased melanocyte proliferation. They are most commonly seen pigmented lesions are benign, it is necessary to distinguish the on the gingiva, buccal mucosa and palate. Those occurring on the origin of pigmentation for melanocytic lesion may be a malignant lower lip are simply called melanotic macule 2, 3). OPN is a very rare occasion in the oral cavity characterized by the accumulation Correspondence to: Hitoshi Nagatsuka, DDS, PhD, Department of Oral of nevus cells at the junctional epithelium or in the submucosa or Pathology and Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University 2-5-1 Shikata-cho, a combination of both 4). The most common of which are found in Okayama City, Japan 700-8525, Tel No: +81-86-235-6650, Fax No: +81- the buccal mucosa with nevus cells accumulating in the submucosa 86-235-6654, Email: [email protected] 57 J.Hard Tissue Biology Vol. 19(1):57-64, 2010 5). OMM is a malignant pigmented melanocytic tumor of the oral Table 1. Oral melanotic macule cavity characterized by tremendous proliferation of tumor cells in the epithelium or in the connective tissues or a combination of Case Age/gender Location Size (mm) both. OMM has been regarded as a separate entity and different 1 50/F BM Not specified from its cutaneous counterpart in many aspects and for reason 2 14/F Lower lip 3* that it is not related to ultra-violet exposure 6 - 9). 3 30/F Gingiva Not specified Melanocytic lesions are renowned as the most difficult 4 67/F BM Not specified diagnostic fields in histopathology 10). For instance, melanoma 5 32/F Gingiva Not specified may have varying morphological features. Particularly, non- 6 70/F Gingiva 5x15 cutaneous primary melanoma as well as metastatic melanomas 7 51/F Soft palate Not specified may resemble other neoplasms ranging from carcinoma, sarcoma, 8 64/M Gingiva 5x5 plasmacytoma, lymphoma and germ cell tumors 11). Another 9 62/M BM Not specified challenging feature might be a case of amelanotic melanoma 10 74/M BM Not specified having epithelioid shape cells. Moreover, atypical melanocytic 11 47/F Lower lip 4* proliferation may occur in the oral mucosa, having equivocal 12 46/F Palate 5x7 features but not sufficient to categorize as melanoma in situ 6). 13 63/F Gingiva 3* Nevertheless, atypical melanocytic proliferation may suggest the 14 63/F BM Not specified earliest stage of OMM progression 12, 13). 15 43/M Gingiva 7* Although H and E remains the basis of diagnosis, the advent of *Greatest diameter immunohistochemistry(IHC) has greatly expanded the histological analysis. Melanoma differentiation markers are used to distinguish pigmented nodular type is an amelanotic lesion without radial poorly differentiated melanomas from non-melanocytic tumors. growth phase. The histological classification of OMM had been Among the panel of markers, S100, HMB-45, Melan A are mainly previously described 12). used to establish the diagnosis of melanoma 14 - 19). Nevertheless, variations in the expressions of these molecules in different types Immunohistochemistry of melanomas have been reported 9). This study aimed to determine New sections of 3µ were cut for IHC. For comparative purposes, the clinical and pathological features of focal pigmented sections of neuroma, neurofibroma and schwannoma were also melanocytic lesions occurring in the oral cavity as well as the included in the study. Sections were deparaffinized in series of expressions of different melanocytic markers in these lesions. No xylene and dehydrated in series of alcohol. Sections were blocked sign of malignant feature both clinically or histologically, was for endogenous peroxidase activity using 3% hydrogen peroxide observed in oral MM and OPN, nonetheless, it is recommended in methanol for 30 min and washed with Tris-buffered saline that focal pigmentation with a diameter of greater than 6 mm on solution. IHC protocols were optimized for each antibody. Antigen preferred sites for OMM be subjected to biopsy. The combination retrieval for S100 and HMB-45 was done by immersing the slides of S100, HMB-45 and Melan A is a reliable aid in the diagnosis in pre-warmed (37oC) 0.1% trypsin for 15 min. For Melan, A, c- of OMM. kit and Ki-67, slides were immersed in citrate buffer, pH 6.0 (heat pressure for 5 min at 121°C). The slides were covered with serum- Materials and methods free protein block (DakoCytomation, Carpinteria, USA) for 15 Case selection from paraffin-embedded tissues min, followed by primary antibodies with the following dilutions: Cases were obtained from the archive of the Department of Oral HMB-45, c-kit and Ki-67 with 1:100, Melan A with 1:50 and Pathology and Medicine, Okayama University conforming to the S100 is ready to use. Slides were incubated overnight at 4°C. ethics imposed by the university hospital. H and E sections were Immunoreaction was carried out with Envision™ Detection thoroughly reviewed, representative paraffin blocks were chosen Reagent Peroxidase (DakoCytomation) and detected with 3- and new sections for H and E staining were made. Cases selected amino-ethylcarbazole substrate chromogen (DakoCytomation). for the study were limited to those diagnosed as oral MM, OPN and OMM. Clinical findings of OMM were classified based on Results Tanaka et al’s report 20). Briefly, pigmented macular type is a flat, Oral MM pigmented lesion with radial growth phase, pigmented nodular All fifteen cases were observed in patients over 30 years of age type is a pigmented tumor with vertical growth phase and rarely except for one case, which was observed in a 14-year old patient. radial growth phase, pigmented mixed type is characterized by There seemed to be a female gender predilection observed in 11/ the presence of radial growth phase surrounding the tumor, non- 15 cases (Table 1). Most cases were located on the gingiva (6/15) 58 Rosario Rivera Buery et al.: Evaluation of Oral Melanotic Lesions Table 2. Oral pigmented nevus Case Age/gender Location Size (mm) Histological type 1 69/M BM Not specified Subepithelial 2 30/F Palate 2x4 Subepithelial 3 36/M BM 4* Subepithelial 4 38/M BM 4* Subepithelial 5 22/F BM 2-3* Subepithelial 6 32/F BM 5* Subepithelial *Greatest diameter Table 3.
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