"Rhabdoid" meningioma: an aggressive variant

It is has been suggested that rhabdoid morphology is associated with a poor prognosis, regardless of tumor histogenesis. We report a series of 15 meningiomas with rhabdoid features. Nine patients had undergone multiple resections. In six, the rhabdoid component was histologically apparent only in recurrences. Rhabdoid morphology was defined as sheets of loosely cohesive cells with eccentric nuclei and hyaline, paranuclear inclusions. Ultrastructurally, the latter consisted of whorls of intermediate filaments often entrapping lysosomes or other organelles. Meningothelial features included whorl formation and nuclear pseudoinclusions, immunohistochemical coexpression of vimentin and epithelial membrane antigen, and the ultrastructural finding of interdigitating cell membranes and intercellular junctions. At the histologic level, a conventional meningioma component was noted in most tumors; only four lesions were entirely rhabdoid. Histologic malignancy (brain invasion or anaplasia) was observed in nine cases, another two tumors being considered malignant on the basis of extracranial metastasis. In the majority, increased cell proliferation was evidenced by a high mitotic rate or MIB-1 LI. At last follow-up, 13 patients (87%) had experienced at least one recurrence and 8 (53%) were dead of disease. Median time to death was 5.8 years after initial surgery and 3.1 years after the first appearance of rhabdoid morphology. Our findings corroborate those from a smaller series recently reported by Kepes et al. on the same entity (Kepes JJ, Moral LA, Wilkinson SB, Abdullah A, Llena JF. Rhabdoid transformation of tumor cells in meningiomas: A histologic indication of increased proliferative activity. Report of four cases. Am J Surg Pathol 1998;22:231-8). They further suggest that rhabdoid meningiomas are highly aggressive tumors and that the rhabdoid phenotype represents a marker of malignant transformation in meningiomas.     

Secretory meningioma with severe perifocal edema--case report

An 82-year-old male presented with a small parasagittal meningioma associated with disproportionately severe perifocal edema. Histological examination including immunohistochemical staining and electron microscopy resulted in a diagnosis of secretory meningioma. In addition to tumor size, the edema could not be explained by location, growth rate, vascular involvement, or other factors. We conclude that secretory meningiomas may possess an innate ability to cause brain edema.     

Perceptual similarity of shapes generated from Fourier descriptors

BACKGROUND: Growth rates and tumor aggressiveness of meningiomas are thought to be closely related to brain edema development. However, histopathologic data alone are not consistently accurate predictors of the behavior and clinical course of a meningioma. METHODS: The authors examined 57 histologically proven intracranial meningiomas to identify factors, including growth fractions determined by MIB-1 immunostaining, that may influence the development of meningioma-associated peritumoral brain edema. There were 54 benign, 2 atypical, and 1 anaplastic meningiomas. The MIB-1 staining index (SI) percentage was defined as the number of MIB-1 positive cells divided by the total number of tumor cells in a 1.037-square millimeter area on the slide. The extent of peritumoral brain edema was determined using preoperative magnetic resonance imaging. The extent of edema was classified as Grade 0,1, or 2 (GR0, GR1, or GR2), in order of increasing severity. RESULTS: The MIB-1 SIs of the 57 cases ranged from 0.06-6.8% (median, 0.80%). There were 26 GR0, 20 GR1, and 11 GR2 edema cases. The MIB-1 SI rose in order of increasing edema severity. There was a statistically significant correlation between the MIB-1 SI and the extent of brain edema (P<0.0001), and also between the tumor size and the extent of brain edema (P=0.001). Meningothelial and atypical/anaplastic meningiomas were associated with peritumoral brain edema more often than any other subtype (P<0.005). CONCLUSIONS: Growth fractions, as determined by MIB-1 immunostaining, rise with increasing severity of peritumoral brain edema, indicating a close relationship between tumor aggressiveness and edema development.     

Cystadenolymphoma of the parotid gland an immunohistochemical study of the epithelial component of twenty cases

Carcinoembryonic antigen, epithelial membrane antigen, Keratin, Desmin, Vimentin, CD30, lysozyme, alpha 1-antitrypsin, alpha 1-antichymotrypsin, S-100 protein, somatostatin and glucagon were looked for using immunohistochemical methods in the epithelial component of 20 parotid gland cystadenolymphomas and 20 normal parotid glands. Carcino-embryonic antigen, ephithelial membrane antigen, S-100 protein, and somatostatin were found in the epithelial cells of most of the cystadenolymphomas. In normal parotid tissue, carcinoembryonic antigen, epithelial membrane antigen, Keratin, alpha 1-antitrypsin, alpha 1-antichymotrypsin, and S-100 protein were found in all three types of ductal cells, somatostatin only in intercalated and striated ductal cells, and lysozyme only in acinar and intercalated ductal cells. Desmin and CD30 were found in the epithelial component of seven of the 20 tumors versus none of the 20 normal parotid glands. Glucagon and Vimentin were negative both in tumor epithelial cells and in normal parotid ductal cells. Our results support the theory that cystadenolymphomas arise from epithelial cells. The presence of lysozyme in the epithelial tumor cells and in the intercalated ductal cells of normal parotid tissue suggest that cystadenolymphomas may arise from the intercalated ducts. The presence of S-100 and somatostatin may indicate that the tumor derives from neuroendocrine structures, but further studies are needed to clarify this point.     

Brain edema in meningiomas is associated with increased vascular endothelial growth factor expression

OBJECTIVE: Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF), an endothelial cell-specific cytokine, induces proliferation of endothelial cells and increases vascular permeability dramatically. All gliomas secrete significant amounts of VEGF, whereas meningiomas are variable in expression. Thus, we sought to determine whether the extent of VPF/VEGF expression in meningiomas correlated with differences in brain edema associated with these tumors. METHODS: Meningioma tissue samples from 37 patients (15 men, average age 65 +/- 13 yr; 22 women, average age 60 +/- 10 yr) who underwent surgery at or were referred to the University of Alabama Hospital were examined retrospectively for the extent of expression of immunoreactive VPF/VEGF. Additionally, peritumoral edema was assessed on a blinded basis radiographically from preoperative magnetic resonance imaging scans. Selected specimens were examined by in situ hybridization to document the source of VPF/VEGF. RESULTS: The predominant meningioma subclassifications were transitional (57%) or meningothelial (27%) subtypes. VPF/VEGF immunoreactivity ranged from 0 to 3.5, with a median value of 2 on a subjective 5-point scale; magnetic resonance imaging-assessed edema ranged in extent from 0 to 4 (subjective 5-point scale), with a median value of 2.5. The correlation of determination (R2) of magnetic resonance imaging-assessed tumor edema rating and VPF/VEGF staining intensity rating was 0.6087 (r = 0.78; P = 0.0001). In situ hybridization localized VPF/VEGF messenger ribonucleic acid in meningioma cells and not in normal parenchymal brain cells. CONCLUSION: These data suggest that meningioma-associated edema may be a result of the capacity of meningioma cells to produce VPF/VEGF locally, leading to increased tumor neovascularization and enhanced vascular permeability.     

Effect of glycerol on blood flow distribution in tumoral and peritumoral brain tissue

The effect of glycerol on blood flow in tumoral and peritumoral tissue was measured in 32 patients with brain tumor, 17 gliomas and 15 meningiomas. Blood flow before and after the administration of glycerol was measured by stable xenon-enhanced computed tomography. The tumor part of glioma was significantly hypoperfused. In contrast, the tumor part of meningioma was significantly hyperperfused. Peritumoral edema of both glioma and meningioma was hypoperfused. After the administration of glycerol, blood flow increased in all regions except for the tumor part of glioma. Vascular responses to glycerol may be different in these two tumor types. The steal phenomena of blood flow might occur in cases of glioma.     

The importance of pial blood supply to the development of peritumoral brain edema in meningiomas

In a retrospective analysis, the authors studied the pial and dural blood supplies in 74 intracranial meningiomas and quantified their associated peritumoral brain edema (PTBE). The extent and localization of pial blush in relation to the total tumor volume were determined angiographically. The amount of edema and tumor size were calculated using computerized tomography. The edema-tumor volume ratio was defined as Edema Index (EI). There were 49 meningiomas with PTBE; of those tumors, 46 were supplied by pial vessels, and three were supplied exclusively by dural vessels. Tumors without PTBE showed no pial blush. The mean EI in meningiomas with pial blush was significantly larger (EI = 3) than in meningiomas without pial supply (EI = 1.1; p < 0.0001). Meningiomas with a smaller pial supply than dural supply had a significantly smaller mean EI than tumors with a pial supply equal to or greater than the dural supply (EI = 2.9 vs. EI = 3.7; p < 0.015). In 69.9% of cases with pial blood supply, major portions of the edema were located adjacent to the tumor region supplied by pial vessels. Edema index differences among tumors of different subgroups, as defined by size or histology, were significantly related to the pial supply in each subset. Thus, pial blood supply may be associated with the development of PTBE in meningiomas.     

Mutational analysis of the p73 gene localized at chromosome 1p36.3 in colorectal carcinomas

A case of chordoid meningioma occurring in a 15-year-old girl is presented. The patient manifested seizures as the initial symptom and subsequently exhibited subclinical microcytic hypochromic anemia. The tumor, located in the falcotentorial region and associated with diffuse edema, was totally resected. On histological examination of the surgical specimen, the clustering pattern of partly vacuolated cells in the mucoid stroma mimicked chordoma; however, positive staining of individual cells for vimentin and epithelial membrane antigen led to a diagnosis of meningioma. Interestingly, the tumor cells were surrounded by a periodic acid-Schiff- and type IV collagen-positive substance. Electron microscopy demonstrated a strikingly dense and thick basal lamina. The patient's microcytic hypochromic anemia disappeared after the tumor was removed. Both the clinical and pathological features of this case resemble those of chordoid meningioma, a rare meningioma variant.     

The mechanism of positive scintigraphy with 99mTc-pertechnetate in adenolymphomas of the parotid grand

Inhibin is a peptide hormone produced by ovarian granulosa cells and testicular Sertoli cells. Ovarian granulosa cell and other sex cord-stromal tumors usually exhibit positive immunohistochemical staining with antiinhibin antibodies, and this may be valuable in differentiating these neoplasms from histologic mimics. In the present study, we investigated the immunohistochemical staining of testicular sex cord-stromal tumors using antiinhibin. Immunostaining with CAM5.2, vimentin, S-100 protein, desmin, epithelial membrane antigen (EMA), carcinoembryonic antigen (CEA), and placental alkaline phosphatase (PLAP) also was performed because few studies have investigated in detail the immunophenotype of testicular sex cord-stromal tumors. Fifteen of 16 Leydig cell tumors exhibited strong positive staining with antiinhibin. A proportion of Leydig cell tumors also stained positively with CAM5.2 (7 of 16), vimentin (14 of 16), S-100 protein (10 of 16), desmin (2 of 16) and epithelial membrane antigen (4 of 16). Four of six testicular sex cord-stromal tumors with varying degrees of Sertoli or granulosa cell differentiation were positive with antiinhibin, as were two of three sex cord-stromal tumors that were unclassified. Some of these tumors were positive with CAM 5.2, vimentin, S-100 protein, desmin, and epithelial membrane antigen. All tumors were negative with carcinoembryonic antigen and placental alkaline phosphatase. The immunohistochemical findings show that, analogous to their ovarian counterparts, most testicular sex cord-stromal tumors are immunoreactive with antiinhibin. Immunohistochemistry using this antibody as part of a panel may be valuable in confirming a diagnosis of testicular sex cord-stromal tumor and in differentiating these neoplasms from others that may mimic them.     

Cytology of primary pulmonary meningioma. Report of the first multiple case

BACKGROUND: Ectopic meningiomas arising in the lung are rare. We report here the first multiple primary case diagnosed by intraoperative imprint cytology. CASE: Asymptomatic pulmonary nodules, two in the left and three in the right lung, were found in a 61-year-old woman, and video-assisted thoracoscopic surgery was undertaken. Because the largest tumor was diagnosed as a meningioma by intraoperative imprint cytology using an excised biopsy specimen, further resection was not performed immediately. Histopathologically the tumor was characterized by whorled nests of cells accompanied by psammoma bodies intermingled with a fibrous pattern. The diagnosis was a transitional meningioma, positive for vimentin and epithelial membrane antigen and negative for keratin immunohistochemically. All the nodules were subsequently surgically resected and showed a similar cytohistologic appearance. Ultrastructurally the tumor cells demonstrated interdigitation of adjacent plasma membranes with numerous desmosomes and hemidesmosomes, typical of meningiomas. We failed to detect another primary tumor in the nervous system, and at this writing the patient was healthy three years after the operation. CONCLUSION: Because of the characteristic cytomorphologic features of primary pulmonary meningioma, the cytologic approach provides useful information for therapy.     

Estimation of different models of insulin therapy in noninsulin-dependent diabetes mellitus

A 21-year old woman underwent surgery in December 1996 for the removal of a presumed tuberculum sellae meningioma. However, some radio-clinical findings were proved somewhat intriguing:the patient's age, the presence of inflammatory and febrile syndromes together with the diagnosis of aseptic meningitis associated with perilesional edema intensity (an unusual feature in such cases) made us challenge the initial neuroradiological diagnosis evoked in connection with the tumoral location and dural attachment pattern. A right sub-fronto-temporal approach allowed complete tumor resection (confirmed with a postoperative MRI) and clinical recovery of the patient. But while pathological examination suggested a chordoma, the study of immunohistochemical stains revealed a meningioma. The final diagnosis was chordoid meningioma. Our review of the literature has shown that chordoid meningiomas display several areas of physaliferous cells which give the tumor a chordoma-like aspect. However, the results of immunohistochemical studies along with the location of the tumor were not consistent with the diagnosis of chordoma. Eight cases of chordoid meningiomas are reviewed in the literature. They are described as inducing systemic symptoms, particularly anemia. They could also be linked to Castleman's syndrome according to Kepes et al. After careful evaluation, we retained the hypothesis of a cause and effect relationship between the local and generalised inflammatory syndrome and chordoid meningioma.     

Reduced expression of schwannomin/merlin in human sporadic meningiomas

OBJECTIVE: The neurofibromatosis type 2 gene is frequently mutated in sporadic meningiomas. The protein product of the neurofibromatosis type 2 gene is called schwannomin or merlin. Its expression in leptomeningeal cells from which meningiomas are derived and the characteristics of mutated forms in meningiomas, to our knowledge, have not been previously studied. METHODS: Immunoblotting and immunoprecipitation experiments with two specific antibodies were used to determine the size and subcellular distribution of schwannomin/merlin in rabbit and human brain tissue and established human leptomeningeal LTAg2B cells. Immunoblotting was used to determine the expression level of schwannomin/merlin in 14 human sporadic meningiomas. RESULTS: Both antibodies detect a protein of approximately 66 kDa, which is predominantly expressed in the Triton X-100-insoluble fraction of the brain and LTAg2B cells. The levels of schwannomin/merlin were severely reduced in eight tumors (57%) when compared with the expression levels in the human brain, LTAg2B cells, and the remaining six meningiomas. All six tumors with the normal schwannomin/merlin expression were of meningotheliomatous type. In contrast, all other histological types and one meningotheliomatous tumor with psammoma bodies were deficient in the 66-kDa schwannomin/merlin. Although nonsense mutations leading to premature stop codons are common in the neurofibromatosis type 2 gene in meningiomas, we found no evidence of truncated schwannomin/merlin forms in the tumors analyzed. CONCLUSION: The absence of complete schwannomin/merlin in almost 60% of primary sporadic meningiomas seems to be an important factor in meningioma tumorigenesis. The development of meningotheliomatous meningiomas is probably linked to alterations in other oncogenes or tumor suppressor genes.     

Mammary ductal foam cells: macrophage immunophenotype

Mammary ductal foam cells are present in normal breast tissue as well as in a number of breast diseases. Such foam cells tend to be in particular abundance with fibrocystic changes of the breast. Foam cells may appear within duct lumens or plastered in cohesive masses along duct walls, simulating an epithelial structure. The nature and origin of these innocuous-appearing cells, based on morphologic studies, remain a controversy, for they appear to be of epithelial derivation. This study was undertaken to determine the nature of intraductal "foam" cells and their origin in the breast. Nine cases of adult fibrocystic disease were examined immunohistochemically with antibodies to cytokeratins (Mak-6, Cam 5.2), leukocyte common antigen, and the following macrophage antibodies: KP-1 (CD68), HAM 56, and MAC 387. The lysozyme and alpha-1-antitrypsin content of foam cells also was studied. The immunohistochemical data in this study confirm the macrophage character of these foam cells, which are positive for CD68, HAM 56, and MAC 387, lysozyme, and alpha-1-antitrypsin and negative for leukocyte-common antigen and cytokeratins.     

Initial experience using a catheterizable ileovesicostomy (Monti procedure) in children

OBJECT: The goal of this study was to evaluate gene delivery to a benign brain tumor. METHODS: A recombinant adenovirus vector bearing the Escherichia coli beta-galactosidase reporter gene was selectively injected into the vascular supply of a spontaneously occurring canine olfactory groove meningioma. The tumor and a small amount of peritumoral brain tissue were removed 5 days after viral injection and stained with X-Gal to assess gene delivery. The authors noted significant beta-galactosidase gene expression by the tumor, but not by surrounding brain tissue. No obvious viral-related cytotoxicity was noted. CONCLUSIONS: The authors found that meningiomas can be successfully transduced by adenovirus vectors by using endovascular techniques.     

Meningiomas with disomy of chromosome 22: study of 9 cases

BACKGROUND: Cytogenetic studies of meningiomas suggest that loss of (or parts of) chromosome 22 is a primary event in the development of these tumors; later on, other chromosomal changes would occur in the caryotypes. All these secondary changes are observed mainly in cases with high clinical aggressivity. However, in a few cases of meningiomas disomy 22 coexists, but with other chromosomic anomalies. We present clinical, histopathological and cytogenetic findings in a group of meningiomas with disomy of chromosome 22. PATIENTS AND METHODS: We collected 10 meningiomas from nine patients which ages ranged between 28-70 years. Fresh tumoral specimens were divided for histologic examination and cytogenetic study, performed after short-term culture. RESULTS: At microscopic examination 5 tumors were classified as benign meningiomas, four as atypical and one as malignant meningioma. Four cases were recurrent tumors. The cytogenetic studies showed that all tumors presented two chromosomes 22 and other chromosome abnormalities. Losses in chromosomes 4, 7, 10, 14, 16, 17 and 20 were frequent; cytogenetics rearrangements of chromosomes 1, 4, 5, 7, 14, 19 and 22 were frequently involved. CONCLUSIONS: In karyotypic evolution of meningiomas, secondary anomalies of chromosomes 1p, 10 and 14 are the most common and appear to be associated with a more aggressive clinical course. In this group of meningiomas with disomy 22, these anomalies were also frequently found, and were related in 50% of cases with atypical or malignant morphologies and of them with recurrent tumors in the 40%.     

Testicular Sertoli cell tumor with a heterologous sarcomatous component: immunohistochemical assessment of Sertoli cell differentiation

OBJECTIVE: Immunohistochemical staining is reported to be useful in distinguishing ovarian Sertoli-stromal cell tumors from carcinosarcomas. To assess Sertoli cell differentiation in a rare malignant biphasic testicular tumor, we compared the immunophenotypic profile of the tumor with that of Sertoli cell nodules and adenomas and mullerian carcinosarcomas. DESIGN: Immunohistochemical staining was performed on 6 testes (4 with hyperplastic Sertoli cell nodules, 2 with Sertoli cell adenomas) and 7 carcinosarcomas (6 involving the uterus, 1 involving the uterus and ovary) using primary monoclonal antibodies AE1/AE3, CAM 5.2, CA 19.9, and antibodies directed against epithelial membrane antigen, carcinoembryonic antigen (monoclonal and polyclonal), S100, placental alkaline phosphatase, and inhibin. These staining results were compared with those of the index case. RESULTS: All testes showed positive staining for inhibin and vimentin in the Sertoli cells of the nodules and adenomas. One Sertoli cell nodule showed focal staining with AE1/AE3 and CAM 5.2. Both adenomas showed focal positive staining for S100. All nodules and adenomas were negative for epithelial membrane antigen, monoclonal and polyclonal carcinoembryonic antigen, CA 19.9, and placental alkaline phosphatase. In contrast, the carcinomatous areas of the carcinosarcomas were all negative for inhibin but exhibited positive staining for AE1/AE3, CAM 5.2, and epithelial membrane antigen. The carcinosarcomas showed variable expression of vimentin, S100, carcinoembryonic antigen, CA 19.9, and placental alkaline phosphatase. The epithelial component of the tumor from the index case showed strong diffuse staining for inhibin and vimentin and only very faint focal staining with AE1/AE3 and CAM 5.2. The epithelial component was negative for epithelial membrane antigen, monoclonal and polyclonal carcinoembryonic antigen, S100, CA 19.9, and placental alkaline phosphatase. CONCLUSIONS: The immunohistochemical findings in the index case support the diagnosis of Sertoli cell tumor with a heterologous sarcomatous component over carcinosarcoma. Inhibin seems to be the best single marker for Sertoli cell differentiation. To our knowledge, only 1 other case of this rare testicular tumor has been reported in the literature.     

Exposure to high ambient temperature increases absorption and plasma concentrations of transdermal nicotine

Granulosa cell tumors with bizarre nuclei (GCT-BN) are rare lesions with a prognosis apparently similar to that of conventional granulosa cell tumors (GCT-NOS). The immunohistochemical features of GCT-BN have not been described, and the exact nature of the bizarre nuclei (BN) is unclear. Thirteen GCT-BN were studied with antibodies to cytokeratin, vimentin, epithelial membrane antigen, muscle-specific actin, alpha smooth muscle actin, desmin, and S-100 protein. Six cases were also examined by fluorescence in situ hybridization for trisomy 12, a nonrandom chromosomal aberration found in a proportion of ovarian sex-cord stromal tumors. Histologically, 12 tumors (86%) contained BN areas interspersed with large areas of GCT-NOS. The remaining tumor contained only microscopic foci of GCT-NOS. Immunohistochemically, the tumors stained for vimentin (13 tumors), S-100 protein (11 tumors), muscle-specific actin (10 tumors), cytokeratin (eight tumors), alpha smooth muscle actin (eight tumors), and desmin (one tumor), but none stained for epithelial membrane antigen. Immunostaining results for the BN and GCT-NOS areas were concordant in eight (73%) of the 11 tumors in which both areas could be independently assessed. The remaining three tumors (27%) showed discordant results for only one of the eight markers used. In five patients, trisomy 12 was detected by fluorescence in situ hybridization in areas of BN but not in areas of GCT-NOS present in the same tumor. Trisomy 12 was also present in another BN tumor in which the foci of GCT-NOS were too small to be evaluated. We conclude that within GCT-BN, areas with BN are immunohistochemically similar to areas of GCT-NOS present in the same tumor. The finding of trisomy 12 in areas with BN but not GCT-NOS in the same tumor, however, suggests that cells with BN represent a genetically distinct clone of tumor cells arising within GCT-NOS.     

Expression of a functional endothelin (ETA) receptor in human meningiomas

Endothelin (ET) receptor subtypes (ETA and ETB) in human meningiomas were characterized using quantitative receptor autoradiography. A single class of high-affinity 125I-ET-1 binding sites was localized in all meningioma tissue studied (dissociation constant: 2.4 +/- 0.3 nM, maximum binding capacity: 319 +/- 66 fmol/mg (mean +/- standard error of the mean for 13 tumors)). Unlabeled ET-1 showed a strong affinity for 125I-ET-1 binding to tissue sections of the tumors with a 50% inhibiting concentration (IC50) of 2.9 +/- 0.7 x 10(-9) M, whereas ET-3 showed a much lower affinity (IC50: 8.4 +/- 2.5 x 10(-6) M). Sarafotoxin S6c, a selective agonist for the ETB receptor, could not compete for 125I-ET-1 binding to meningiomas. Endothelin-1 significantly stimulated deoxyribonucleic acid (DNA) synthesis in a dose-dependent manner in cultured human meningioma cells. In contrast, no significant stimulation of DNA synthesis occurred with an S6c concentration up to 10(-7) M. Pretreatment of the meningioma cells with pertussis toxin, a bacterial toxin that adds adenosine 5'-diphosphate-ribose to the alpha subunit of guanine nucleotide binding (G) proteins such as Gi or G(o), induced a concentration-dependent reduction in ET-stimulated DNA synthesis in meningioma cells, but did not affect the epidermal growth factor-induced DNA synthesis. These observations suggest that the ETA receptor is predominantly expressed in human meningioma tissue and that ET may act as a growth factor on the meningioma cells by interacting with the ETA receptor and by pertussis toxin-sensitive mechanisms.     

No significant role of Epstein-Barr virus in the tumorigenesis of Warthin tumor

To determine the significance of Epstein-Barr virus (EBV) in the tumorigenesis of Warthin tumor, formaldehyde-fixed and paraffin-embedded specimens of 21 tumors from 18 patients were examined by polymerase chain reaction (PCR)-Southern hybridization, in situ hybridization (ISH), and immunohistochemistry. PCR-Southern hybridization revealed the presence of EBV DNA in 13 of the 21 tumors (61.9%). However, ISH for EBV RNA showed that nuclei of the neoplastic epithelial cells of all tumors were negative. Although ISH for EBV DNA revealed that the nuclei of the neoplastic epithelial cells were positive in 4 of the 21 tumors (19.0%), the positive cells were sparsely distributed and there was no evidence of monoclonal proliferation of EBV positive neoplastic epithelial cells. Moreover, immunohistochemical reactions with antibodies against latent membrane protein 1 (LMP1) and EBV nuclear antigen 2 (EBNA2), were negative in all cases. Judging from these findings, we conclude that EBV does not play a major role in the tumorigenesis of Warthin tumor.