Polypeptide N-acetylgalactosaminyl transferase 3 independently predicts high-grade tumours and poor prognosis in patients with renal cell carcinomas.

BACKGROUND: The polypeptide N-acetylgalactosaminyltransferases (GalNAc-Ts) family of enzymes regulates the initial steps of mucin-type O-glycosylation. N-acetylgalactosaminyltransferases might show novel patterns of GalNAc-T glycosylation on tumour-derived proteins, which could influence cancer biology, but its mechanisms are unclear. We investigated the association of GalNAc-T3 and -T6 expressions with clinicopathological features and prognoses of patients with renal cell carcinomas (RCCs). METHODS: Expressions of GalNAc-T3/6 and cell-adhesion molecules were analysed immunohistochemically in 254 paraffin-embedded tumour samples of patients with RCC. RESULTS: Of 138 GalNAc-T3+ cases, 46 revealed significant co-expression with GalNAc-T6. N-acetylgalactosaminyltransferases-3+ expression showed a close relationship to poor clinical performance and large tumour size, or pathologically high Fuhrman's grading, and presence of vascular invasion and necrosis. The GalNAc-T3-positivity potentially suppressed adhesive effects with a significantly low ß-catenin expression. Univariate and multivariate analyses showed the GalNAc-T3+ group, but not the GalNAc-T6+ group, to have significantly worse survival rates. CONCLUSION: N-acetylgalactosaminyltransferases-3 expression independently predicts high-grade tumour and poor prognosis in patients with RCC, and may offer a therapeutic target against RCC.

Polypeptide N-acetylgalactosaminyltransferase 6 expression in pancreatic cancer is an independent prognostic factor indicating better overall survival.

BACKGROUND: The family of polypeptide N-acetylgalactosaminyltransferases (GalNAc-Ts) is responsible for the altered glycosylation in cancer. The purpose of our study was to investigate the clinical significance of two isoforms, GalNAc-T6 and -T3, and their correlation with the prognosis of pancreatic cancer. METHODS: Immunohistochemistry was used to analyse GalNAc-T6 and -T3 expressions in 70 clinicopathologically characterised pancreatic cancer cases. RESULTS: Positive expressions of GalNAc-T6 and -T3 were immunohistochemically identified in 51% (36 of 70) and in 77% (54 of 70) of patients, respectively. A close relationship was noted between GalNAc-T6 positive expression and pathological well/moderate differentiated type (P=0.001), small tumour size (P=0.044), absence of vascular invasion (P=0.009), and low stage of the American Joint Committee on Cancer systems (P=0.043). The expression of GalNAc-T3 significantly correlated with good differentiation (P=0.001), but not with other clinicopathologic features. Furthermore, univariate and multivariate analyses revealed that GalNAc-T6 expression was an independent prognosis indicator for the disease, whereas GalNAc-T3 expression had no impact on clinical outcome, even though 33 of 36 GalNAc-T6-positive cases also had a positive expression of GalNAc-T3 (P=0.001, r=0.356). CONCLUSION: Both GalNAc-T6 and -T3 expressions correlated significantly with tumour differentiation, whereas only GalNAc-T6 expression predicted prognosis in pancreatic cancer.

Shortened microsatellite d(CA)21 sequence down-regulates promoter activity of matrix metalloproteinase 9 gene.

One characteristic elements in the promoter of the matrix metalloproteinase 9 (MMP-9) gene is the d(CA) repeat. To investigate whether this element regulates the transcription of the MMP-9 gene and its enzymatic activities, we sequenced the promoter region isolated from esophageal carcinoma cell lines. TE9 cells with low MMP-9 enzymatic activity had the number of d(CA) repeats shortened from 21 to 14 or 18. TE8, TE10 and TE11 cells with high MMP-9 activities had 21 or 23 d(CA) repeats. Luciferase assays using MMP-9 promoter containing 18, 14 or 0 d(CA) repeats showed transcriptional activities which were 50, 50 or 5%, respectively, of the level achieved with promoter containing 21 d(CA) repeats. Sequence analysis of the promoter of 223 Japanese subjects revealed that most had two alleles with 20, 21 or 22 d(CA) repeats, whereas six had one or two alleles with 14, 18 or 19 d(CA) repeats. We postulate that length alteration of the d(CA) repeat causes phenotypic differences among carcinoma cells and that microsatellite instability may contribute to the polymorphism of d(CA) repeat length.

Switch of histamine receptor expression from H2 to H1 during differentiation of monocytes into macrophages.

It is known that histamine suppresses gene expression and synthesis of tumor necrosis factor alpha (TNF-alpha) induced by lipopolysaccharide (LPS) in human peripheral blood mononuclear monocytes (HPM) or alveolar macrophages via histamine H2 receptors. We investigated the effect of histamine and differentiation in macrophages on the expression and secretion of TNF-alpha, TNF-alpha-converting enzyme (TACE), and histamine H1 and H2 receptors by use of a leukemia cell line, U937, and HPM. Differentiation of U937 and HPM cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) enhanced the H1 receptor expression and rather suppressed the H2 receptor, resulting in up-regulation of the histamine-induced expression and secretion of TNF-alpha, modulated via TACE. Therefore, histamine failed to inhibit up-regulated expression of TNF-alpha induced by LPS in macrophages. The switch from H2 to H1 receptors during differentiation in the monocyte/macrophage lineage could participate in the pathogenic processes of atherosclerosis and inflammatory reactions in the arterial wall.

A role for interleukin 4 in production of matrix metalloproteinase 1 by human aortic smooth muscle cells.

Effect of interleukin 4 (IL-4) on the production of matrix metalloproteinase 1 (MMP-1) by normal and immortalized human intimal smooth muscle cells (SMC) was investigated. The production of the precursors of MMP-1 by intimal SMC was enhanced in a dose-dependent manner by addition of IL-4 to the culture medium, whereas the cytokine also showed an inhibitory effect on DNA synthesis in the cells. In addition, mRNA of IL-4 was found in the atherosclerotic and nonatherosclerotic areas of the intima. Although the production of MMP-1 and the proliferation of SMC are thought to play an important role in reconstruction of the intima during atherogenesis, our results suggest a possible role of IL-4 induced MMP-1 in inhibiting tissue remodeling caused by a variety of arterial disorders including atherosclerosis.

Phenotypes of human esophageal squamous cell carcinoma based on matrix metalloproteinase production.

We investigated the effects of epidermal growth factor (EGF) and transforming growth factor-beta (TGF-beta) on cell growth and on regulation of matrix metalloproteinases (MMPs) in four cell lines of human esophageal squamous cell carcinomas (TE8, TE9, TE10, and TE11). EGF stimulated the production of proforms of gelatinase B (MMP-9) by three cell lines that could synthesize EGF by themselves, with TE9 being the exception. Particularly, both the production of MMP-9 and DNA synthesis in TE10 were stimulated significantly by EGF. TGF-beta slightly stimulated DNA synthesis in two cell lines, TE9 and TE11, and TGF-beta secretion by TE9 was detected. The production of proforms of gelatinases A (MMP-2) and MMP-9 was gradually induced by TGF-beta in a concentration-dependent manner in all the cell lines except for TE9. Flow cytometric analysis revealed that all the lines expressed both EGF- and TGF-beta-receptors. In conclusion, our present results indicate that at least there are possibly two distinct phenotypes in human esophageal squamous cell carcinoma: one (TE10) depends on autocrine EGF production that enhances DNA synthesis and MMP-9 production; and the other (TE9) on autocrine TGF-beta that stimulates DNA synthesis but not in relation to gelatinase production.

Extraskeletal myxoid chondrosarcoma: a clinicopathologic, immunohistochemical, and molecular analysis of 18 cases.

Extraskeletal myxoid chondrosarcoma (EMCS) is an uncommon clinicopathologically well-defined tumor, but its pathogenesis and biologic behavior are poorly understood. We reviewed 18 cases of EMCS to verify clinicopathologic features and immunohistochemical profiles together with molecular detection of the tumor-specific fusion genes. The tumors were located mainly in the proximal extremities and limb girdles (72%). Two tumors arose at unusual anatomic sites: the finger and the hip joint. Nine of the 17 followed-up patients were alive and disease free, 4 were alive with recurrences and/or metastases, and 4 died of the tumor. Fifteen tumors showed typical features of EMCS, and 3 had hypercellular areas in addition to conventional EMCS areas. The tumors were variably immunoreactive for S-100 protein (50%), NSE (89%), peripherin (60%), and synaptophysin (22%). Chromogranin A and some epithelial markers (AE1/AE3, CAM5.2, and epithelial membrane antigen) were entirely negative. Frequent expressions of the neural/neuroendocrine markers suggest possible neural/neuroendocrine differentiation in at least some EMCSs, in addition to chondroid differentiation. In a reverse-transcription polymerase chain reaction (RT-PCR) assay using paraffin-embedded specimens, EWS-CHN or TAF2N-CHN fusion gene transcripts characteristic of EMCS could be detected in 15 (83%) of the 18 cases: EWS-CHN type 1 in 11 cases, EWS-CHN type 2 in 1, and TAF2N-CHN in 3. Three fusion-negative cases included 2 conventional EMCSs and 1 considered a "cellular" variant of the tumor. None of 30 other soft tissue and bone tumors with myxoid or chondroid morphology that we examined contained these fusion genes. Thus, RT-PCR detection of EWS-CHN or TAF2N-CHN fusion gene using archival paraffin-embedded tissue is a feasible and useful ancillary technique for the diagnosis of EMCS.

Molecular detection of EWS-FLI1 chimeric transcripts in Ewing family tumors by nested reverse transcription-polymerase chain reaction: application to archival paraffin-embedded tumor tissues.

Chromosomal translocations generating unique chimeric genes are highly characteristic of specific sarcomas, and their use as diagnostic markers has been suggested. From a diagnostic pathologic point of view, detection of such cytogenetic or molecular aberrations applicable to routinely processed archival tissue specimens is considered a powerful tool for tumor diagnosis. To assess the feasibility and reliability of the molecular detection of the transcript originating from the chimeric gene in paraffin-embedded tumor specimens, we performed a nested reverse transcription-polymerase chain reaction (RT-PCR)-based assay to detect the EWS-FLI1 chimeric message in a series of Ewing family tumors. Of 24 paraffin-embedded tumor specimens from 23 cases analyzed, the chimeric message was detectable in 20 (83%) specimens from 20 cases (87%) by this nested RT-PCR assay, whereas none of 7 small round cell tumors not from this family (3 alveolar rhabdomyosarcomas, 2 neuroblastomas, 2 malignant lymphomas) showed detectable chimeric messages. In the sequence analysis of the PCR products, the amplified chimeric messages contained the junctions between exon 7 of the EWS gene and any one of exons 5, 6 and 8 of the FLI1 gene. The detection process was usually completed within 3 days, except for the subseqent sequence analysis. Our results endorse the use of this molecular assay as an ancillary technique in the diagnosis of Ewing family tumors using paraffin-embedded material.

Detection of COL1A1-PDGFB fusion transcripts in dermatofibrosarcoma protuberans by reverse transcription-polymerase chain reaction using archival formalin-fixed, paraffin-embedded tissues.

The reciprocal translocation t(17;22)(q22;q13) and a supernumerary ring chromosome, r(17;22), derived from the translocation, have been shown to be highly characteristic of dermatofibrosarcoma protuberans (DFSP). Its consequence is a fusion of two genes, a collagen type I alpha 1 gene (COL1A1) and platelet-derived growth factor B-chain gene (PDGFB). The COL1A1-PDGFB fusion gene, is expected to be a diagnostic molecular assay. However, previous studies on this subject were mostly based on frozen tissue specimens or cultured tumor cells. In this present study, the investigators conducted a reverse transcription (RT)-polymerase chain reaction (PCR) assay to detect the COL1A1-PDGFB fusion transcripts using archival formalin-fixed, paraffin-embedded tumor specimens from 12 patients with DFSP. To amplify the fusion transcripts, a specific COL1A1 forward and PDGFB reverse primers were designed for single step PCR. The COL1A1-PDGFB fusion transcripts could be detected in 10 of 12 paraffin-embedded DFSP tumor specimens (83%). Subsequent sequence analysis using the PCR products confirmed that the detected messages were derived from gene fusions composed of PDGFB exon 2 and different regions of the COL1A1 gene (exon 8, 10, 22, 24, 32, 38, 45 or 46). Two samples of Bednar tumor included in this series also contained the fusion transcripts. In sample of DFSP with fibrosarcomatous transformation, the COL1A1-PDGFB could not be detected in the fibrosarcoma areas of the third recurrence, though the chimeric transcripts were identified in the ordinary DFSP areas of the first recurrence. No fusion transcripts could be amplified in non-DFSP lesions, including 10 dermatofibromas and 9 malignant fibrous histiocytomas. These results indicate that this molecular assay could be applied to archival formalin-fixed, paraffin-embedded tumor tissues as a diagnostic aid for DFSP.

Detection of TLS/FUS-CHOP fusion transcripts in myxoid and round cell liposarcomas by nested reverse transcription-polymerase chain reaction using archival paraffin-embedded tissues.

The reciprocal translocation t(12;16)(q13;p11) has been shown to be highly characteristic of myxoid and round cell subtypes of liposarcoma, and the TLS/FUS-CHOP fusion gene that resulted from the translocation is expected to be a diagnostic molecular marker of these sarcomas. In this study, we conducted a nested reverse transcription-polymerase chain reaction (RT-PCR)-based assay to detect the TLS/FUS-CHOP fusion gene transcripts using archival formalin-fixed, paraffin-embedded tumor specimens. Of 18 paraffin-embedded specimens from 16 myxoid and round cell liposarcoma cases, the fusion transcripts could be identified in 16 (89%) specimens from 15 (94%) cases. A sequence analysis using the PCR products confirmed that the detected messages were derived from either type I or type II TLS/FUS-CHOP fusion gene, the latter of which was predominant (80%). The results were consistent in primary and recurrent lesions of the same patients and in paraffin-embedded and snap-frozen samples from the same tumors. In two negative specimens, transcripts of the beta-actin gene could not be detected by RT-PCR, and intact mRNA including the fusion messages might have been degraded. No fusion transcripts were detected in snap-frozen or paraffin-embedded material of other types of tumors with myxoid morphology (seven myxoid malignant fibrous histiocytomas and four lipomas with myxoid change). These results indicate that this molecular assay can be applied to formalin-fixed, paraffin-embedded tumor tissues as a diagnostic aid for these subtypes of liposarcoma.

Pathological study of splenomegaly associated with cadmium-induced anemia in rats.

Splenomegaly was observed both in male and female Sprague-Dawley rats after 1 week of exposure to CdCl2 (0.6 mg Cd/kg/day). Spleen weight reached about double that in controls by 8 weeks of Cd exposure. Histopathological examination of the enlarged spleen revealed that iron- and lipid-laden histiocytes were clustered in the periarterial lymphatic sheath, and the red pulp appeared to be expanded. It is noteworthy that electron microscopy revealed marked poikilocytosis and Heinz body formation in red blood cells (RBCs) in both the sinus and cord. Histiocytes were swollen by a granular substance in the cytoplasm and also many secondary lysosomes. These morphological findings indicate that degradation of damaged RBCs induced by exposure to Cd might be promoted in the spleen and possibly cause splenomegaly. This RBC damage-hemolysis-splenomegaly sequence is also considered to be associated with the etiology of Cd-induced anemia. In addition to the abnormal RBC degradation, nuclei of lymphocytes in the Cd-exposed spleen exhibited high electron density, consistent with a preapoptotic state suggesting the immunosuppressive effect of Cd.

[Cytofluorometric analysis of DNA content in proliferating cells of coronary arteriosclerotic lesions].

Cytofluorometric determination of DNA content was done on paraffin-embedded tissues of 19 cases of coronary arteriosclerotic lesions including fibrocellular intimal thickening lesions (FT) or atherosclerosis (AS). DNA distribution pattern of medial smooth muscle cells of coronary arteries with FT and AS was all diploid. The average proliferative index (PI) of both medial smooth muscle cells of coronary arteries with FT and AS was 4.8 +/- 0.6. DNA distribution patterns of intimal cells of coronary arteries with FT and AS were also diploid. The average PI of intimal cells of coronary arteries with FT and AS was 8.4 +/- 1.0 and 9.1 +/- 0.7, respectively. These results suggest that intimal cellular proliferation plays an important role in the development of atherosclerosis.

[Expression of PDGF-A and -B in human coronary atherosclerotic lesion: immunohistochemical and in situ hybridization study].

We studied the role of platelet-derived growth factor (PDGF) on the development of atherosclerosis of human coronary arteries of 63 autopsied cases. Smooth muscle cells in fibrocellular intimal thickening lesion showed no significant immunohistochemical reaction of antibodies for PDGF-A or PDGF-B or PDGF-receptor. In atherosclerotic lesions, foam cells derived from macrophages and smooth muscle cells showed intense immunohistochemical reaction with antibodies of PDGF-B and PDGF-receptor, but not with that of PDGF-A. By in situ hybridization, no significant signals of PDGF-A or PDGF-B or PDGF-receptor were demonstrated in proliferating intimal smooth muscle cells in fibrocellular intimal thickening lesions. Uncomplicated atherosclerotic lesions expressed significant amount of m-RNA of c-sis protooncogene and PDGF-B receptor in foam cells. However, no significant signals of PDGF-A were observed in uncomplicated atherosclerotic lesions. These results suggest that foam cells-producing PDGF-B play an important role for the progression of atherosclerotic lesions in human coronary arteries.

[An autopsy case of periarteritis nodosa associated with disseminated strongyloidiasis].

An autopsy case of periarteritis nodosa associated with disseminated strongyloidiasis in a 59-year-old woman is reported. The patient had unknown fever of 38 degrees C and marked impairment of renal function, for which pulse therapy of steroid was performed. Electromyogram showed myogenic pattern, and biopsy of the biceps presented necrotizing angitis of small arteries. Renal biopsy showed marked infiltration of lymphocytes in the stroma, and frequent hyalinosis and crescent formation of the glomeruli. Two months after admission, the patient died of respiratory failure associated with sepsis and disseminated intravascular coagulopathy. Postmortem examination disclosed strongyloides stercolaris and fungi in both lungs with extensive hemorrhage. Terminal ileum and ascending colon had multiple erosions, extensive hemorrhage, numerous strongyloides stercolaris, and frequent necrotizing angitis in the mucosa. Necrotizing angitis was also demonstrated in both kidneys.

Tumor consistency of pituitary macroadenomas: predictive analysis on the basis of imaging features with contrast-enhanced 3D FIESTA at 3T.

BACKGROUND AND PURPOSE: Preoperative evaluation of pituitary macroadenoma tumor consistency is important for neurosurgery. Thus, we aimed to retrospectively assess the role of contrast-enhanced FIESTA in predicting the tumor consistency of pituitary macroadenomas. MATERIALS AND METHODS: Twenty-nine patients with pituitary macroadenomas underwent conventional MR imaging sequences and contrast-enhanced FIESTA before surgery. Two neuroradiologists assessed the contrast-enhanced FIESTA, contrast-enhanced T1WI, and T2WI. On the basis of surgical findings, the macroadenomas were classified by the neurosurgeons as either soft or hard. Finally, Fisher exact probability tests and unpaired t tests were used to compare predictions on the basis of the MR imaging findings with the tumor consistency, collagen content, and postoperative tumor size. RESULTS: The 29 pituitary macroadenomas were classified as either solid or mosaic types. Solid type was characterized by a homogeneous pattern of tumor signal intensity without intratumoral hyperintense dots, whereas the mosaic type was characterized by many intratumoral hyperintense dots on each MR image. Statistical analyses revealed a significant correlation between tumor consistency and contrast-enhanced FIESTA findings. Sensitivity and specificity were higher for contrast-enhanced FIESTA (1.00 and 0.88-0.92, respectively) than for contrast-enhanced T1WI (0.80 and 0.25-0.33, respectively) and T2WI (0.60 and 0.38-0.54, respectively). Compared with mosaic-type adenomas, solid-type adenomas tended to have a hard tumor consistency as well as a significantly higher collagen content and lower postoperative tumor size. CONCLUSIONS: Contrast-enhanced FIESTA may provide preoperative information regarding the consistency of macroadenomas that appears to be related to the tumor collagen content.

Expression of Y-box-binding protein dbpC/contrin, a potentially new cancer/testis antigen.

Y-box-binding proteins are members of the human cold-shock domain protein superfamily, which includes dbpA, dbpB/YB-1, and dbpC/contrin. dbpC/contrin is a germ cell-specific Y-box-binding protein and is suggested to function as a nuclear transcription factor and RNA-binding protein in the cytoplasm. Whereas ubiquitous dbpB/YB-1 expression has been well studied in various types of human carcinomas as a prognostic or predictive marker, the dbpC/contrin expression in human tumour cells has not been reported. In this report, we provide the first evidence showing that dbpC was highly expressed in human testicular seminoma and ovarian dysgerminomas, and in carcinomas in other tissues and that its expression in normal tissues is nearly restricted to germ cells and placental trophoblasts. These results indicate that dbpC/contrin would be a potentially novel cancer/testis antigen.

Detection of SYT-SSX fusion transcripts in both epithelial and spindle cell areas of biphasic synovial sarcoma using laser capture microdissection.

To investigate the distribution of tumour cells expressing the SYT-SSX fusion gene in biphasic synovial sarcoma, modified reverse transcription polymerase chain reaction (RT-PCR) analysis was performed using microdissected specimens from haematoxylin and eosin stained sections of archival paraffin wax embedded tissues. This modified RT-PCR included a stage with degenerate oligonucleotide primed (DOP) PCR, which randomly amplified cDNA after reverse transcription. SYT-SSX fusion transcripts were detected in both epithelial and spindle cell areas of all three biphasic synovial sarcomas examined. Subsequent sequence analysis confirmed that the detected messages were derived from the SYT-SSX1 fusion gene in two cases and from SYT-SSX2 in one. These results indicate that SYT-SSX fusion transcripts are found in both epithelial and spindle cell areas of biphasic synovial sarcoma, and RT-DOP-PCR-PCR analysis is a useful method for detection of extremely small amounts of mRNA in microdissected samples from archival formalin fixed, paraffin wax embedded tumour tissues.

Complete remission of ovarian endometrioid adenocarcinoma associated with hyperamylasemia and liver metastasis treated by paclitaxel and carboplatin chemotherapy: a case report.

Complete remission in the case of a 45-year-old woman with ovarian endometrioid adenocarcinoma associated with hyperamylasemia and liver metastasis is described. Ultrasound examination and CT scan revealed several large solid and cystic intra-abdominal tumors and a metastatic liver tumor. The serum amylase was 600 micro/l (normal value: 60-200 micro/l), and electrophoresis identified isoamylases of the salivary type. A total abdominal hysterectomy with bilateral salpingo-oophorectomy was performed for invasive carcinoma of the left ovary. The histology of the left ovary showed endometrioid adenocarcinoma. Immunohistochemically, the adenocarcinoma cells were diffusely and strongly positive for amylase. The patient received six courses of paclitaxel and carboplatin combination chemotherapy. Two years later, the patient is alive and well, without evidence of disease. The prognosis of patients with ovarian cancer metastatic to the liver is uniformly poor. This represents the first report of complete remission of such a patient in the literature.