Endometrial metaplasia: correlation of histological and cytological specimens obtained from 103 cases undergoing hysterectomy for endometrial carcinoma.

OBJECTIVE: To assess the frequency of endometrial metaplasia in histological and cytological specimens from the same cases, and to determine the relationship between various types of metaplasia and clinicopathological findings. METHODS: We reviewed 103 histological specimens diagnosed as endometrioid adenocarcinoma, in which endometrial smears had been obtained before surgery. We examined the correlation between the frequency of endometrial metaplasia occurring in association with carcinoma in both histological and cytological specimens. The categories of metaplasia were eosinophilic metaplasia, squamous metaplasia, mucinous metaplasia, ciliated cell metaplasia and others. We compared the incidence of endometrial metaplasia with the clinicopathological findings for each case. RESULTS: Endometrial metaplasia was recognized in 90 (87.4%) of the histological and 80 (77.7%) of the cytological specimens of 103 specimens, with the respective frequency of subtypes as follows: eosinophilic metaplasia (36.0% and 43.7%), squamous metaplasia (70.9% and 68.0%), mucinous metaplasia (38.8% and 19.4%), ciliated cell metaplasia (22.3% and 2.9%) and others (11.7% and 0%). Mixed subtypes were seen in 58.3% and 41.7% of histological and cytological specimens, respectively. In histology, mucinous metaplasia was significantly more frequent in G1-G2 than G3 carcinomas (P = 0.0089). Ciliated cell metaplasia was significantly related to endometrial hyperplasia (P = 0.0068). In cytology, eosinophilic and mucinous metaplasia were significantly associated with G1-G2 cases (P = 0.0061 and P = 0.0385). CONCLUSIONS: Endometrial metaplasia was seen in 87.4% of the histological and 77.7% of the cytological specimens. Where routine endometrial cytopathology is practiced, it is important to understand the detailed histological and cytological features of these changes.

Increasing methylation of the CDKN2A gene is associated with the progression of adult T-cell leukemia.

In this study, we examined the methylation status of the CDKN2A gene in patients with different forms of adult T-cell leukemia (ATL) using Southern blot analysis, methylation-specific PCR (MSPCR), and nucleotide sequencing. We found that the CDKN2A gene was more frequently methylated in fresh tumor cells isolated from patients with acute ATL (47%) or lymphoma-type ATL (73%) than in those with less malignant chronic (17%) and smoldering (17%) ATL. In addition, deletions of the CDKN2A gene were found in 24% of acute ATL patients; thus, abnormalities of the CDKN2A gene totaled 71% in acute ATL patients. In contrast, no CDKN2A gene methylation was found in asymptomatic carriers or uninfected individuals. Methylation of the p15 gene was not found in any samples from 36 ATL patients. Direct sequencing of the CDKN2A gene after sodium bisulfite treatment of genomic DNA revealed that the methylation of CpG sites had occurred in 24 of 32 ATL cases (75%) including chronic and smoldering ATL, even when MSPCR and the Southern blot had failed to detect CDKN2A gene methylation. Among fresh ATL samples with methylation, methylation was detected in the promoter region and exon in 17 of 24 cases, and methylation in the exon without promoter region was detected in 7 of 24 cases. In one case, the pattern of methylation proved to be different between peripheral blood cells and lymph node cells, suggesting the presence of multiple subclones with regard to methylation patterns, despite the same HTLV-I integration site. Quantitative PCR showed a marked decrease in CDKN2A mRNA expression in the cells with a methylated CDKN2A gene, especially if the promoter region was methylated. These findings suggest that CpG methylation decreases CDKN2A expression and represents a critical factor in the disease progression of ATL.

Persistent clonal proliferation of human T-lymphotropic virus type I-infected cells in vivo.

Clonal proliferation of human T-lymphotropic virus type I (HTLV-I)-infected cells has been detected by Southern blot analysis and inverse PCR in patients with adult T-cell leukemia, patients with HTLV-I-associated diseases, and even in asymptomatic carriers. Combining inverse PCR with long PCR, we amplified the genomic DNA regions flanking the integration sites of the HTLV-I provirus to detect clones of infected cells. Inverse long PCR revealed that increased virus load was associated with an increase of both the number of cells in each clone and the number of clones. Clonal proliferations were found in both CD4- and CD8-positive cells in a carrier and a patient with HTLV-I-associated neuropathy/tropical spastic paraparesis. These HTLV-I-infected clones persisted over several years in the same carriers, and, moreover, most of the persistent clones were CD4 positive in a HTLV-I carrier. These findings indicate that HTLV-I infection plays an important role in the clonal expansion of lymphocytes and the prolonged survival of CD4-positive cells in vivo. Surviving T-lymphocytes may be susceptible to genetic changes, leading to the onset of leukemia.

HTLV-I provirus in the clinical subtypes of ATL.

Adult T cell leukemia (ATL) is an aggressive neoplasm of mature helper T cell, which is etiologically linked with human T-lymphotropic virus type 1 (HTLV-I). We studied HTLV-I provirus in 61 cases of ATL with Southern blot analyses and long PCR. These methods detected defective virus in 34 cases (56%). Furthermore, it found two types of defective virus. The first type (type 1) defective virus had both LTRs, but lacked internal sequences, such as gag and pol. Type 1 defective virus was seen in 50% of all defective virus. The second form (type 2) of defective virus had only one LTR, and 5'-LTR was preferentially deleted. This type of defective virus could be more frequently detected in aggressive types of ATL (16/44 cases) than chronic type (1/17 cases). This defective virus might be associated with clinical subtype.

Adult T cell leukemia following HTLV-I-associated myelopathy/tropical spastic paraparesis: case reports and implication to the natural course of ATL.

Adult T cell leukemia (ATL) is the T cell malignancy caused by human T lymphotropic virus type I (HTLV-I), and HTLV-I is also the causative agent of HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Although HTLV-I causes both diseases, concomitant occurrence is reported to be rare. This paper describes two cases of HAM/TSP that developed into lymphoma-type ATL after the onset of HAM/TSP. In one case, the same HTLV-I infected clone could be detected by polymerase chain reaction in peripheral blood obtained when the patient was diagnosed as HAM/TSP. This finding showed that the HTLV-I clone already existed at the stage of HAM/TSP. Since frequent detection of clonal proliferation of HTLV-I infected cells has been reported previously in patients with HAM/TSP, careful follow-up is needed for patients with HAM/TSP.

Pleomorphic leiomyosarcoma: clinicopathologic and immunohistochemical study with special emphasis on its distinction from ordinary leiomyosarcoma and malignant fibrous histiocytoma.

Pleomorphic leiomyosarcoma (PLMS) was recently described as a morphologic variant of leiomyosarcoma; however, its diagnostic criteria, as shown by morphologic features and biologic behavior, remain controversial. We describe 28 cases of pleomorphic sarcoma with pleomorphic areas in more than two thirds of the tumor and an ordinary leiomyosarcomatous fascicular area covering less than one third as PLMS. PLMS comprised 8.6% of all the leiomyosarcomas (322 cases) registered in our institute. Patients ranged in age from 31 to 89 years (average, 57.9 years). Seventeen patients (60.7%) were male and 11 were female. Tumor location was as follows: the extremities in 17 cases, the retroperitoneum or abdominal cavity in 7 cases, the chest/abdominal wall in 3 cases, and the scalp in 1 case. Histologically, all cases showed at least small foci of fascicles consisting of smooth muscle tumor cells, in addition to pleomorphic areas mimicking storiform-pleomorphic malignant fibrous histiocytoma. The border between pleomorphic and leiomyosarcomatous fascicular areas was sharp in 3 cases, gradual in 2 cases, and blending in 23 cases. Sixteen cases (57.1%) showed a typical storiform pattern, 6 cases revealed extensive stromal hyalinization, 6 cases showed a chronic inflammatory infiltrate, 2 cases had the foci of foamy xanthomatous cells, and 7 cases contained myxoid malignant fibrous histiocytoma-like areas covering less than 50% of the tumor. The tumors had a tendency to be of a morphologically higher grade (10 tumors were French Federation of Cancer Centers grade 2, 18 were grade 3). Five of 28 cases (18%) showed rhabdoid features. Immunohistochemically, all of the 28 tumors examined showed a positive reactivity for at least one smooth muscle marker (desmin, muscle-specific actin, and alpha-smooth muscle actin) in the leiomyosarcomatous fascicular areas. In the pleomorphic areas the expression of smooth muscle markers (desmin 10 of 28, muscle-specific actin 13 of 28, and alpha-smooth muscle actin 14 of 28) was significantly reduced, compared with that in leiomyosarcomatous fascicular area (desmin 18 of 28, muscle-specific actin 26 of 28, and alpha-smooth muscle actin 24 of 28). No significant difference was observed between the MIB-1 labeling index in the leiomyosarcomatous fascicular areas (26.10 on average) and that in the pleomorphic areas (26.17 on average). However, the MIB-1 labeling index in PLMS was significantly higher than that in ordinary leiomyosarcoma (n = 20, 12.86 on average) or storiform-pleomorphic malignant fibrous histiocytoma (n = 16, 16.63 on average). In 23 patients follow-up data were available with a duration of 1-239 months. Eleven patients developed metastases, and lung accounted for the most common site of metastasis (9 cases). Fifteen of 23 patients (65.2%) died of disease. Our results indicate that PLMS should be differentiated from ordinary leiomyosarcoma because of its high proliferative activities and rather aggressive biologic behavior.

Hepatocyte growth factor function and c-Met expression in human lens epithelial cells.

PURPOSE: Hepatocyte growth factor (HGF) and its receptor c-met perform a multitude of functions. However, despite the significant degree of study of HGF and c-met in numerous tissues and cell types, relatively few investigations have been performed on the lens. In the current study, therefore, the role of HGF and the receptor c-met in human lens epithelial cells was investigated. METHODS: Anterior epithelium and capsular bags were prepared from human donor eyes and maintained in Eagle's minimum essential medium (EMEM) in a 5% CO(2) atmosphere at 35 degrees C. In addition, the human lens cell line FHL124, was routinely cultured and seeded onto glass coverslips (c-met immunodetection), 12-well plates (DNA and protein synthesis), and tissue culture dishes (migration). c-Met was detected by immunocytochemistry and fluorescence-activated cell scanning (FACS). HGF was measured using enzyme-linked immunosorbent assay (ELISA) techniques. Proliferation and protein synthesis were determined by [(3)H]thymidine and (35)S-methionine incorporation into DNA and proteins, respectively. Migration was assessed using a scratch-wound assay and time-lapse video microscopy. RESULTS: HGF was detected at all stages of culture of capsular bags in protein-free medium. Moreover, c-met was present on the native epithelium and after mechanical trauma was seen to be upregulated. Immunolocalization and FACS analysis demonstrated c-met expression on FHL124 cells throughout the whole population. Furthermore, FACS analysis showed that serum-maintained cells sustained a higher level of receptor expression relative to serum-deprived cells. Additionally, HGF was found to stimulate proliferation, protein synthesis, and migratory responses. CONCLUSIONS: c-Met receptors are expressed in native epithelium, capsular bag cultures, and FHL124 cells. Receptor is distributed across the entire cell population; however, this expression is environmentally and mechanically sensitive. HGF is also present in capsular bags at all stages of culture. In addition, HGF can stimulate migration, proliferation, and protein synthesis. It therefore appears that a multifunctional autocrine loop involving HGF and c-met is in place and could be important in the development of posterior capsule opacification.

FGF: an autocrine regulator of human lens cell growth independent of added stimuli.

PURPOSE: Posterior capsule opacification (PCO) arises because of a persistent growth of lens epithelial cells. Cultured human lens cells residing on their native collagen capsule and maintained in serum-free medium actively grow and thus show an intrinsic capacity for regulation. In the present study, the authors investigated the role of the putative FGF autocrine system in human capsular bags. METHODS: Capsular bags were prepared from human donor eyes and maintained in a 5% CO(2) atmosphere at 35 degrees C. On-going observations were by phase-contrast microscopy. Cellular architecture was examined by fluorescence cytochemistry. De novo protein synthesis was determined by the incorporation of 35S-methionine. Basic fibroblast growth factor (FGF) and FGF receptor (R)-1 were detected using enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR) techniques. FGFR-1 inhibition was achieved using the specific antagonist SU5402. RESULTS: Human lens epithelial cells can maintain metabolic activity for more than 1 year in a protein-free medium. Basic FGF was shown to be present in capsular bags throughout culture and also in capsular bags removed from donor eyes that had previously undergone cataract surgery. Furthermore, FGFR-1 was identified. Inhibition of FGFR-1 caused a significant retardation of growth on the posterior capsule. On no occasion did any treated bag reach confluence, whereas all match-paired control samples did. CONCLUSIONS: The results provide evidence that FGF plays an integral role in the long-term survival and growth of human lens epithelial cells, independent of external stimuli. Inhibition of FGFR-1 by specific synthetic molecules, such as SU5402, could provide a potential therapeutic approach to resolving PCO.

Relative quantitation of p53 and MDM2 gene expression in leiomyosarcoma; real-time semi-quantitative reverse transcription-polymerase chain reaction.

To investigate the etiology of leiomyosarcoma, we examined abnormalities of p53 and its regulation in 13 cases of leiomyosarcoma using fresh tumor specimens. We estimated p53 and MDM2 mRNA level and MDM2 gene amplification using a real-time semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) based on the TaqMan fluorescence method. We also used immunohistochemistry (IHC) for p53 and MDM2 protein overexpression, polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) and direct sequencing to detect p53 mutation. Eight of the 13 cases (62%) showed an overexpression of p53 protein on IHC and eight of 13 cases (62%) had p53 gene point mutations. Five of the 13 cases (38%) showed positive staining for MDM2 protein and only one case (7.7%) demonstrated MDM2 gene amplification. The relative p53 mRNA level of the tumors compared with normal tissue ranged from 1.14 to 12.19 arbitrary units (AU), and the MDM2 mRNA level ranged from 1.06 to 17.17 AU. The mRNA level in the p53-positive cases was higher than in the negative cases (positive: 7.70 AU on average; negative: 3.38 AU on average; P=0.0344). However, there was no significant correlation between the MDM2 mRNA level and other factors, such as p53 IHC, p53 mutation status, p53 mRNA level and MDM2 IHC. Our results indicate that p53 abnormalities are major events and that an increasing level of p53 mRNA is associated with an overexpression of p53 protein in leiomyosarcoma and they may play an important role in the tumorigenesis in this tumor.

Immunoexpression of neurofibromin, S-100 protein, and leu-7 and mutation analysis of the NF1 gene at codon 1423 in osteofibrous dysplasia.

The NF1 (neurofibromatosis type 1, or von Recklinghausen disease) gene, is a tumor-suppressor gene, and its product, neurofibromin, down-regulates ras protein by its guanosine triphosphatase-activating protein (GAP)-related domain. Osteofibrous dysplasia (OFD) is characterized by fibroblast-like spindle cells and osseous tissue and is generally seen in the tibia or fibula during childhood. The precise nature of OFD remains controversial. Cosegregations of OFD and NF1 have been reported, and it has been surmised that OFD is associated with the NF1 gene. We studied the expressions of NF1 gene product (neurofibromin) and so-called Schwann cell markers (S-100 protein, Leu-7) in 17 cases of OFD immunohistochemically. Ten cases of fibrous dysplasia (FD) were also used for the purpose of comparison. Five OFD and 7 FD cases were analyzed for NF1 gene mutation at codon 1423, which is a GAP-related domain, by single-strand conformation polymorphism. Fibroblast-like cells of OFD showed the expression of neurofibromin (5 of 17), S-100 protein (9 of 17), and Leu-7 (5 of 17), and those of FD did not show these expressions, with the exception of 1 case that showed Leu-7 expression. Regarding the OFD cases, significant correspondence was found between cases showing expression of neurofibromin and S-100 protein, between cases showing expression of neurofibromin and Leu-7, and between cases showing expression of S-100 protein and Leu-7 (P < .01). NF1 gene mutation at codon 1423 was not detected in either the OFD (0 of 5) or FD (0 of 7) cases. These results seem to suggest the possible involvement of neurofibromin in the development of OFD, which is associated with the expression of Schwann cell markers (S-100 protein and Leu-7). Furthermore, NF1 gene mutation at codon 1423 did not seem to be related to OFD.

H-, K-, and N-ras gene mutation in atypical fibroxanthoma and malignant fibrous histiocytoma.

Atypical fibroxanthoma (AFX) which is histologically similar to malignant fibrous histiocytoma (MFH), occurs in the sun-exposed skin. The presence of mutations at codons 12 and 13 of the H- and K-ras genes and in exons 1 and 2, which include codons 12, 13, and 61, of the N-ras gene was studied in 8 cases of AFX and 8 cases of storiform-pleomorphic-type MFH using polymerase chain reaction (PCR)-restriction fragment length polymorphism and PCR-single-conformation polymorphism. Two of the 8 cases of MFH showed ras mutations in the H-ras gene at codon 12 (GGC-AGC) and in the K-ras gene at codon 13 (GGC-GAC). H- and K-ras gene mutations were not seen in any of the cases of AFX (0 of 8). N-ras gene mutation was not detected in either the AFX (0 of 8) or MFH (0 of 8) cases. In conclusion, although the number of cases in this study was small, H- and K-ras genes were present in some of the MFH cases and accordingly may play an important role in the pathogenesis of MFH. In addition, the finding that H-, K-, and N-ras gene mutations are not present in AFX may indicate why AFX has a more favorable behavior than MFH.

Evaluation of Epstein-Barr virus infection in sinonasal small round cell tumors.

Sinonasal undifferentiated carcinoma, olfactory neuroblastoma and malignant melanoma of the sinonasal regions are included within the category of small round cell tumors of the sinonasal region. It is difficult to diagnose these tumors on the basis of light-microscopic features alone, but, in some instances, immunohistochemical staining evaluating cytokeratin and S-100 protein, for example, is of value. On the other hand, the sinonasal region is a significant site for Epstein-Barr-virus (EBV)-related tumors, including sinonasal undifferentiated carcinoma or malignant lymphoma. Twenty-three sinonasal small round cell tumors (SSRCT) comprising 5 sinonasal undifferentiated carcinomas, 9 olfactory neuroblastomas and 9 malignant melanomas were evaluated for the presence of EBV infection by in situ hybridization for EBV-encoded RNA, combined with immunostaining for EBV-related proteins (LMP-1 and EBNA2). Furthermore, 55 SSRCT comprising 37 sinonasal undifferentiated carcinomas, 9 olfactory neuroblastomas, and 9 malignant melanomas were examined for the presence of cytokeratins (AE1/ AE3 and CAM5.2), S-100 protein and p53 protein using immunohistochemical staining. According to in situ hybridization for detecting EBV-encoded RNA 1 (EBER1), all of the sinonasal undifferentiated carcinomas showed clear, intense hybridization signals localized over the nuclei of the tumor cells and, in 3 out of 9 (33.3%) malignant melanomas, hybridization signals were also recognized. However, none of the olfactory neuroblastomas revealed hybridization signals. Immunohistochemically, 4 out of 5 (80%) sinonasal undifferentiated carcinomas were positive for LMP-1, whereas only 2 out 9 (22.2%) malignant melanomas and no olfactory neuroblastomas were positive. With regard to EBNA2, sinonasal undifferentiated carcinomas, malignant melanomas and olfactory neuroblastomas were all negative. Out of 37 sinonasal undifferentiated carcinomas 35 (94.6%) showed a diffuse positive immunoreaction for AE1/AE3, whereas neither olfactory neuroblastoma nor malignant melanoma revealed a positive reaction. All 9 malignant melanomas and 6 out of 9 olfactory neuroblastomas (75%) were positive for S-100 protein, whereas only 6 cases of sinonasal undifferentiated carcinomas (19.4%) were positive. As for p53 protein, 16 of 37 sinonasal undifferentiated carcinomas (43.2%) were positive, whereas neither olfactory neuroblastoma nor malignant melanoma revealed any positive reaction. The above results suggest that EBV infection is closely associated with sinonasal undifferentiated carcinomas, and that some malignant melanomas may also have a relationship with its infection. For the differential diagnosis of SSRCT, it is important to evaluate EBV infection along with immunohistochemical staining for cytokeratins and S-100 protein. The overexpression of p53 protein was found to be related to the oncogenesis of sinonasal undifferentiated carcinoma; however, there was no association between its overexpression and malignant melanoma or olfactory neuroblastoma.

Mutations of the p53 gene in malignant rhabdoid tumors of soft tissue and the kidney: immunohistochemical and DNA direct sequencing analysis.

Malignant rhabdoid tumor (MRT) is characterized by the presence of intracytoplasmic eosinophilic inclusions composed of whorls of intermediate filaments. This tumor was originally described as an entity of the abortive type of Wilms' tumor in childhood. Recently, it has been proved that these rhabdoid cells can be observed in various types of malignant tumors, including soft tissue sarcoma or carcinoma. To investigate the oncogenesis of this tumor, we examined the p53 gene alteration by means of immunohistochemical analysis and DNA direct sequencing in three cases of malignant rhabdoid tumor (MRT) of the soft tissue and three cases of MRT of the kidney. All the cases of MRT of the soft tissue and two of the cases of MRT of the kidney showed immunopositivity for p53 protein. Among them, one of the cases of MRT of the soft tissue and two of the cases of MRT of the kidney showed missense mutations of the p53 gene. These results strongly suggest that p53 gene alterations may have an important role to play in the aggressive biological behavior and poor prognosis of this tumor.

H-ras oncogene mutation in dedifferentiated liposarcoma. Polymerase chain reaction-restriction fragment length polymorphism analysis.

Point mutations of the ras gene family (K-ras, H-ras, and N-ras) are thought to be involved in the development of a variety of human tumors. Dedifferentiated liposarcoma is characterized by the coexistence of well-differentiated (WD) and high-grade anaplastic (HG) components. The presence of point mutations at codons 12 and 13 of the H-ras gene was studied in 34 liposarcomas, comprising 15 well-differentiated liposarcomas and 19 dedifferentiated liposarcomas, and in 8 storiform-pleomorphic type malignant fibrous histiocytomas (MFHs) using polymerase chain reaction-restriction fragment length polymorphism and direct sequencing analysis. The 2 components of dedifferentiated liposarcoma were analyzed independently. H-ras mutations were seen only in dedifferentiated liposarcomas (4/19 [21%]), 1 in WD components and 3 in HG components. The mutation was not seen in any of 15 cases of well-differentiated liposarcoma. MFHs showed an H-ras mutation in 1 (12%) of 8 cases. Our results seem to suggest that the H-ras mutation is a relatively uncommon event in dedifferentiated liposarcoma, which may demonstrate an epiphenomenon of dedifferentiation in dedifferentiated liposarcoma.

Malignant peripheral nerve sheath tumour arising within neurofibroma. An immunohistochemical analysis in the comparison between benign and malignant components.

AIMS: To compare the expression of immunohistochemical variables between benign and malignant components of malignant peripheral nerve sheath tumour (MPNST) arising within neurofibroma. METHODS: Eight cases of MPNST arising within a neurofibroma, associated with neurofibromatosis type 1 (NF1), were studied. The areas of MPNST and neurofibroma were compared immunohistochemically with regard to the expression of proliferative activity (MIB-1), growth factors, p53, bcl-2, neural cell adhesion molecule (N-CAM), and CD34. RESULTS: The expression of transforming growth factor beta 1 (TGF-beta 1), TGF-beta receptor type II, hepatocyte growth factor alpha (HGF-alpha), c-met, p53, and N-CAM was higher in the areas of MPNST than in the neurofibromatous areas in four, five, five, eight, five, and three of the eight cases, respectively. CD34 expression was lower in the areas of MPNST than in the neurofibroma areas in three of the eight cases. CONCLUSIONS: On the basis of these findings, TGF-beta 1, HGF-alpha, and p53 might be involved in the malignant transformation of neurofibroma to MPNST.

Calponin and h-caldesmon expression in atypical fibroxanthoma and superficial leiomyosarcoma.

To evaluate smooth muscle differentiation, myogenic markers [desmin, alpha-smooth muscle actin (SMA), and muscle-specific actin (HHF35)] have been widely used. Calponin and h-caldesmon, which are cytoskeleton-associated actin-binding proteins, have been reported to be more specific myogenic markers, especially since myofibroblasts express a small amount of h-caldesmon. Atypical fibroxanthoma (AFX) occurs in the sun-exposed skin of the elderly and follows a benign clinical course. Histologically, AFX, which is a pleomorphic spindle cell tumor and considered to be a superficial variant of malignant fibrous histiocytoma, also mimics leiomyosarcoma. AFX has been thought to differentiate along pathways with fibrohistiocytic and myofibroblastic phenotypes. AFX ( n=10), superficial leiomyosarcoma (S-LMS) ( n=17) and benign fibrous histiocytoma (BFH) ( n=17) were analyzed for myofibroblastic and smooth muscle differentiation immunohistochemically from the viewpoint of comparison. AFX and BFH showed immunoreactivities respectively for calponin (3/10, 11/17), desmin (3/10, 1/17), SMA (3/10, 13/17), and HHF35 (1/10, 5/17), but failed to express h-caldesmon (0/10, 0/17). S-LMS had a high immunoreactive rate of calponin (17/17), desmin (13/17), SMA (16/17), and HHF35 (16/17), while also expressing caldesmon (11/17). The results reveal that AFX and BFH have immunoreactivities for several myogenic markers, with myofibroblastic differentiation (calponin: +/-, h-caldesmon: -), but without the smooth muscle differentiation seen in S-LMS (calponin:+, h-caldesmon: +/-). In addition, calponin and h-caldesmon are considered to be useful markers for distinguishing AFX from S-LMS.

Establishment and characterization of malignant rhabdoid tumor of the kidney.

Malignant rhabdoid tumor of the kidney (MRTK) is a highly aggressive tumor which occurs in childhood and which is histologically characterized by the existence of eosinophilic intracytoplasmic inclusions. We established and characterized a cell line from this tumor with histological, immunohistochemical and cytogenetical analysis. Histologically, the tumor cells demonstrate typical eosinophilic inclusions, while immunohistochemically the cells demonstrate common mesenchymal and epithelial differentiation. Although the conventional karyotyping of this tumor lacked the abnormalities of 22q chromosome, Southern blot analysis and microsatellite analysis verified abnormalities of the BCR gene and of the hSNF5/INI1 gene. Despite the variety of locations, these common genetic abnormalities appear to contribute to distinguish rhabdoid tumor from such other small round cell tumors as primitive neuroectodermal tumor, rhabdomyosarcoma, poorly differentiated synovial sarcoma and desmoplastic small round cell tumor.

H-ras-1 point mutation in malignant peripheral nerve sheath tumors: polymerase chain reaction restriction fragment length polymorphism analysis and direct sequencing from paraffin-embedded tissues.

It has been shown that the NF1 (neurofibromatosis type 1) gene encodes a tumor suppressor which inactivates ras proteins. Among malignant mesenchymal tumors, H-ras-1 mutations have been found in malignant fibrous histiocytoma, leiomyosarcoma and embryonal rhabdomyosarcoma. However, studies on H-ras-1 mutation of many cases of malignant peripheral nerve sheath tumors (MPNST) have not been documented. Therefore, we investigated H-ras-1 mutations of MPNST. In 45 cases of MPNSTs of our files, DNA was extracted from the formalin-fixed paraffin-embedded tissue, and the mutations of the H-ras-1 gene were detected by using PCR-RFLP (polymerase chain reaction- restriction fragment length polymorphisms) method and direct sequencing. We found two cases with H-ras-1 point mutation in MPNST for the first time. Both cases showed the same mutation in codon 13.1 [GGT(Gly) to AGT(Ser) transition]. Interestingly, both cases were associated with NF1. It is possibile that the mutation of the H-ras-1 gene occurred after the mutation of the NF1 gene in the MPNST.

Immunoreactivity of p53, mdm2, and p21WAF1 in dedifferentiated liposarcoma: special emphasis on the distinct immunophenotype of the well-differentiated component.

Alteration of the p53/mdm2 pathway has been reported in the well-differentiated liposarcoma (WDLS)/dedifferentiated liposarcoma (DDLS) group. We investigated the immunoreactivity of p53, mdm2, and p21WAF1, along with the MIB-1-labeling index (MIB-1-LI) in 21 WDLS and 21 DDLS cases, to clarify the association of these markers with the morphologic changes and the biological factors responsible for the aggressiveness of DDLS. Within DDLS, p53 and p21WAF1 expression and mdm2 overexpression were significantly more prevalent in the dedifferentiated (DD) components than in the well-differentiated (WD) components. The mdm2 overexpression and p21WAF1 expression was significantly associated with sclerosing liposarcomas in both WDLS and the WD components of DDLS. There was no significant difference in the immunoreactivity of p53, mdm2, or p21WAF1 or MIB-1-LI between WDLS and the WD components of DDLS. An association was found between p53 expression and mdm2 overexpression in the WD group (comprising WDLS and WD components of DDLS) and in the DD group, significantly so in the WD group. Notably, this correlation was found in the subtype of sclerosing liposarcoma but not in that of lipoma-like liposarcoma. Within DDLS, the clinical outcome of the nonaccessible soft tissue (non-AST: comprising retroperitoneum and mediastinum) group was significantly worse than that of the accessible soft tissue (AST: comprising extremities, buttocks, axilla, and scrotum) group; however, the immunophenotypes of p53, mdm2, and p21WAF1 and the MIB-1-LI showed no correlation with survival in the AST group alone, in the non-AST group alone, or in the 2 together. This study suggests that the immunoreactivity of p53, mdm2, and p21WAF1 is associated with the morphologic changes, but not with the biological factors responsible for the aggressiveness of DDLS.

Cytokeratin subunits of inclusion bodies in rhabdoid cells: immunohistochemical and clinicopathological study of malignant rhabdoid tumor and epithelioid sarcoma.

Extrarenal malignant rhabdoid tumor (MRT), which is recognized as being histologically similar to renal MRT, is characterized by the presence of "rhabdoid cell" (RC) and a highly aggressive biological behavior. Recently it has been proposed that "proximal variant" of epithelioid sarcoma (ES), whose morphology is similar to that of MRT, actually has a more aggressive clinical course than classical type ES. Detailed immunohistochemical analysis of cytokeratin (CK) subunits was performed in 3 cases of extrarenal MRT, 3 cases of renal MRT, and 11 cases of ES comprising 2 "proximal variants" and 9 classical types. Renal and extrarenal MRTs showed positive immunoreactivity for both CK8 and CK18. Classical type ESs were diffusely positive, not only for CK8 and CK18, but also for other cytokeratin subunits including CK4, 6, 10, 13, 16, 17, and "high-molecular-weight" CKs (CK1, 5, 10, and 14). On the other hand, proximal ES revealed limited immunohistochemical reactivity for cytokeratins, compared with classical ES. In conclusion, the inclusion bodies of RCs show immunoreactivity confined to CK8, CK18, and vimentin. Furthermore, ES has additional CK expressions, while proximal ES possesses characteristics intermediate between those of classical ES and those of external MRT.