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Histopathological and Immunohistochemical Profile of Benign and Malignant Oral Vascular Lesions

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Histopathological and Immunohistochemical Profile of Benign and Malignant Oral Vascular Lesions Oral Med Pathol 11 (2006) 97 Histopathological and Immunohistochemical Profile of Benign and Malignant Oral Vascular Lesions. Rodelio L. Aguirre1, Mai Nishioka2, Renelson L. Esguerra3, Li-Hong Wang1,4, Naoya Arai1, Norihiko Okada2 and Teruo Amagasa1 1Maxillofacial Surgery, 2Diagnostic Oral Pathology, Tokyo Medical and Dental University, Tokyo, Japan 3College of Dentistry, Southwestern University, Cebu City, Philippines 4Department of Bioengineering, School of Sciences, Graduate School, Northeast University, Shenyang, P.R. China Aguirre RL, Nishioka M, Esguerra RL, Wang L, Arai N, Okada N and Amagasa T. Histo- pathological and immunohistochemical profile of benign and malignant oral vascular lesions. Oral Med Pathol 2006; 11: 97-103, ISSN 1342-0984 This study assessed the clinicopathological and immunohistochemical characteristics of benign and malignant vascular lesions using Factor VIII Related-Antigen (FVIIIRAg), CD31, CD34, VEGF, Ki-67, SMA (smooth-muscle actin), and D2-40. Eighty-one cases were collected: 15 VN (vascular neoplasms) (mean age of 43.34 ± 19.45 years old, 1.2:1 male:female ratio) and 66 VA (vascular anomalies) (mean age of 45.70 ± 19.66 years old, 1.37:1 male:female ratio). Most VN and VA were located on the lips, tongue, and buccal mucosa. VN showed increased endothelial cell (EC) proliferation and VA revealed dilated vascular channels lined by flat EC. Hemangiosarcoma demonstrated clusters of epithelioid and spindle cells and expressed immunoreactivity to panendothelial cell markers (CD31, CD34, and FVIIIRAg), Ki-67, VEGF, and D2-40. CD34 is the most consistent vascular EC marker, whereas, Ki-67 and VEGF are useful in delineating VN from VA. For differential diagnosis of hemangiosarcoma, panendothelial cell markers and D2- 40 is recommended. Key words: hemangioma; hemangiosarcoma; vascular anomaly; vascular malformation Correspondence: Rodelio L. Aguirre, Maxillofacial Surgery, Tokyo Medical and Dental University Graduate School 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8549, Japan Phone & Fax: +81-3-5803-5500, E-mail: [email protected] Introduction by the International Society of the Study of Vascular Oral vascular neoplasms (VN) and vascular anoma- Anomalies (ISSVA), distinguishing vascular neoplasm lies (VA) exhibit a broad spectrum of clinical and histo- from vascular anomaly (9, 10). According to this classifi- logical characteristics and the difficulty of distinguish- cation, VN are characterized by an active proliferation of ing these tumors coupled with disagreement on termi- endothelial cells (EC) and panendothelial cells, while VA nology and classification impedes proper treatment (1- are characterized by flat EC and categorized according 5). The term hemangioma is still being used in a generic to their vascular morphology: venous malformation (VM), sense to describe most vascular lesions without proper capillary malformation (CM), capillary-venous malforma- consideration of their underlying biologic characteristics tion (CMVM), arterio-venous malformation (AVM), or lym- and the use of modifiers such as “cavernous”, “strawberry” phatic malformation (LM). However, the phenomenon of and other hybrid terms to describe these lesions further capillary vessel overgrowth, plump endothelial cell lin- aggravates the confusion (2). Recent advances in vascu- ing the vascular vessels, and endothelial hyperplasia can lar research and medical procedures make accurate di- blur the distinction between VA and VN. Therefore, fur- agnosis paramount in proper clinical management of ther studies are needed. Whereas various studies have these lesions (6-8). investigated VA and VN of infancy and childhood, a clas- Among the existing frameworks, a classification sification framework to distinguish VA and VN in adults, based on the natural history and cellular kinetics of vas- particularly of the oral region is necessary. cular lesions of childhood and infancy is recommended In this study, we evaluated the clinopathological 98 Aguirre et al. IHC profile of oral vasclar lesions Table 1: Primary antibodies used in the study and immunohistochemical (IHC) profile of various VA and Details of primary antibodies (CD31, CD34, vWF/ VN of the oral cavity to provide a more accurate assess- FVIIIRAg, VEGF, Ki-67, SMA, D2-40), dilution, clonality, ment of these lesions. Also, to further characterize and and antigen retrieval methods are shown in Table 1. The comprehend the pathophysiology of these tumors, we uti- sections were deparaffinized with xylene and rehydrated lized specific markers for vascular endothelial cells (VEC) in series of graded ethanol. Heat-induced antigen retrieval and lymphatic endothelial cells (LEC). method was employed for vWF/FVIIIRAg (von Willerbrand factor/ Factor VIII related antigen), CD31, Materials and Methods VEGF (vascular endothelial growth factor), Ki-67, SMA Human samples and histopathological data (smooth muscle actin), and D2-40 (Anti-Lymphatic En- Vascular lesions diagnosed as VN (hemangioma and dothelium Marker), while proteinase digestion was per- hemangiosarcoma) and/or VA from 1992 to 2002, exclud- formed for CD34. Endogenous peroxidase activity was ing lymphangioma, were collected from the archives of blocked by immersing the slides in methanol containing Diagnostic Oral Pathology, Tokyo Medical and Dental 0.3% H2O2 and the slides were then incubated with the University (TMDU) for this research. The specimens, primary antibodies in a moist chamber and left in a re- which were fixed in 10% neutral buffered formalin, dehy- frigerator overnight. For color development, the slides drated in a series of graded concentration of ethanol and were immersed in 3,3’-diamino-benzidine (DAB) solution embedded in paraffin wax, were sectioned at 4μm thick- and counterstained with methyl green. Immunoreactiv- ness and mounted on silane coated slides. Details per- ity of the VEC (vascular endothelial cells) was evaluated taining to age, sex, and location were recorded, while his- using an Olympus B × 40 microscope (Olympus, Tokyo, tological diagnosis was performed by two pathologists Japan) at ×200 magnification. Microvessel density count without knowledge of previous diagnosis. While there are was performed for CD31, CD34, and vWF/FVIII RAg. As- various histologic and descriptive classification systems sessment of Ki-67 expression was determined by the pres- differentiating VA from VN, a classification based on en- ence of nuclear staining and cytoplasmic staining for dothelial characteristics (flat or plump vascular endot- VEGF. Only focal proliferation of EC or proliferation of helial cell lining, nuclear pleomorphism, EC proliferation, EC on the vessel lining was considered positive for VEGF thrombosis, and presence of fibroadipose tissues) was used and Ki-67 while intravascular papillary EC hyperplasia in this study. For the purpose of clarification, the term was not considered positive. The results were then orga- hemangioma was restricted to lesions exhibiting cellular nized into 4 categories based on the total percentage of proliferation, whereas vascular anomaly was used for le- cells staining positively as follows: (-) <20%, (+) = 20-40%, sions exhibiting endothelial mitotic activity within nor- (++) = 41-60%, (+++) > 60%. mal range. Results Immunohistochemistry (IHC) Clinicopathological findings Routine IHC technique has been employed using a 81 cases of VA and VN were collected; they con- conventional avidin-biotin complex (ABC) labeled with sisted of 66 VA (49 VEM, 5 CM, and 12 CMVM) and 15 peroxidase as a detection system (Nichirei, Tokyo, Japan). VN (13 hemangioma and 2 hemangiosarcoma). There Oral Med Pathol 11 (2006) 99 Table 2: Clinico-epidemiological characteristics of VA and VN M (male), F (female), BM (buccal mucosa), FM (floor of the mouth), T (tongue), N/F (no female), VM (venous malformation), CM (capillary malformation), CMVM (capillary-venous malformation), He (hemangioma), AS (hemangiosarcoma), *(palate, gingiva, vestibule) Table 3: The histological features of VA and hemangioma displaying the following characteristics a (focal or limited endothelial cell proliferation), b (frequent observation of endothelial cell proliferation with or without lumen formation), c (majority of vessels are dilated and lined with flat endothelial cell), d (proliferation of small vessels without endothelial cell hyperplasia), e (intravascular papillary endothelial hyperplasia), f (presence of thrombosis/phlebolith) Fig. 1: Dilated blood vessels as seen in venous malformation Fig. 2: Capillary malformation displaying plump endothelial cell (HE × 100). lining (HE × 200). were 48 males and 33 females, with a mean age of 46.43 (Table 2). Only 2 cases of hemangioma-of-infancy (HOI) ± 18.82. Most lesions developed on the lips, tongue, and were noted soon after birth and the remaining cases oc- buccal mucosa; increased frequency during the 5th and curred during late adolescence or adulthood. Precursor 6th decades was noted. The most common lesion was VM lesions were not noted in VA and the majority had a 3-5 (60.5%) followed by hemangioma (16%) and CM (14.8%), year duration characterized by swelling and discolora- whereas, hemangiosarcoma was the least common (2.5%) tion. Pain was not a common symptom. 100 Aguirre et al. IHC profile of oral vasclar lesions Table 4: IHC profile of VN and VA (N=81) Fig. 3: Epithelioid cells exhibiting marked nuclear pleo- Fig. 4: The lobular pattern of growth was a distinguishing morphism and increased mitosis in hemangiosarcoma characteristic of proliferative stage of
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