Overexpression of E2F-5 correlates with a pathological basal phenotype and a worse clinical outcome.

The purpose of the present study is to identify genes that contribute to cell proliferation or differentiation of breast cancers independent of signalling through the oestrogen receptor (ER) or human epidermal growth factor receptor 2 (HER2). An oligonucleotide microarray assayed 40 tumour samples from ER(+)/HER2(-), ER(+)/HER2(+), ER(-)/HER2(+), and ER(-)/HER2(-) breast cancer tissues. Quantitative reverse transcriptase PCR detected overexpression of a cell cycle-related transcription factor, E2F-5, in ER-negative breast cancers, and fluorescence in situ hybridisation detected gene amplification of E2F-5 in 5 out of 57 (8.8%) breast cancer samples. No point mutations were found in the DNA-binding or DNA-dimerisation domain of E2F-5. Immunohistochemically, E2F-5-positive cancers correlated with a higher Ki-67 labelling index (59.5%, P=0.001) and higher histological grades (P=0.049). E2F-5-positive cancers were found more frequently in ER(-)/progesterone receptor (PgR)(-)/HER2(-) cancer samples (51.9%, P=0.0049) and in breast cancer samples exhibiting a basal phenotype (56.0%, P=0.0012). Disease-free survival in node-negative patients with E2F-5-positive cancers was shorter than for patients with E2F-5-negative cancers. In conclusion, we identify, for the first time, a population of breast cancer cells that overexpress the cell cycle-related transcription factor, E2F-5. This E2F-5-positive breast cancer subtype was associated with an ER(-)/PgR(-)/HER2(-) status, a basal phenotype, and a worse clinical outcome.

Demonstration of nuclear 3,5,3'-triiodothyronine receptor proteins in gonadotrophs and corticotrophs in rat anterior pituitary by double immunohistochemical staining.

Previously we raised an antiserum (4BII) against nuclear T3 receptor (NT3R), which recognizes c-erb A/NT3R alpha 1 and beta, but not alpha 2. An immunohistochemical study using 4BII revealed that among the tissues examined, the cerebral cortex, anterior pituitary, and thyroid gland exhibited strong immunostaining in the nuclei. In the present study we examined the localization of NT3R proteins in individual cell types of rat anterior pituitary, using the technique of double immunostaining with 4BII and antibodies against pituitary hormones. The cryostat sections and paraffin sections of rat pituitary were incubated with 4BII at 4 C overnight. The NT3R proteins were visualized as brown in the nuclei by avidin-biotin peroxidase staining. The control sections incubated with an antiserum which had previously absorbed with c-erb A peptide or an inactive antiserum showed very poor nuclear staining under the same condition. After being washed with 0.1 M glycine-HCl buffer (pH 2.2) for 2 h to remove 4BII, the sections were incubated with antiserum against individual anterior pituitary hormones at 4 C overnight. Using the method of alkali phosphatase/naphthol/Fast blue staining, the peptide hormones in the cytosol were stained blue. Double staining with 4BII and anti-FSH beta or anti-LH beta anti-serum clearly demonstrated that gonadotrophs contained the NT3R proteins. Similarly, the NT3R proteins in corticotrophs were demonstrated by the immunostaining with 4BII and anti-ACTH antiserum. As expected, the NT3R proteins were present in somatotrophs, thyrotrophs, and lactotrophs. On the other hand, folliculo-stellate cells, which are nonhormone secreting cells and are identified by their S-100 protein immunoreactivity, were stained by 4BII only faintly. The present study demonstrated the existence of NT3R proteins not only in somatotrophs, thyrotrophs and lactotrophs but also gonadotrophs and corticotrophs, suggesting some action of thyroid hormone through the receptor in these cells.

Distribution and characterization of immunoreactive corticostatin in the hypothalamic-pituitary-adrenal axis.

Using an antiserum against synthetic rabbit corticostatin-1 (CS-1), we established a specific RIA for CS-1 and examined its distribution in various tissues, including the hypothalamic-pituitary-adrenal axis. Among the tissues examined, the highest levels of CS-1-like immunoreactivity (-LI) were found in the lung and spleen. CS-1-LI was also detected at relatively high levels in the pituitary, adrenal medulla, and small intestine, while it was barely detectable in the hypothalamus. Immunocytochemical studies revealed the widespread distribution of CS-1 in these tissues. Plasma CS-1 levels averaged 7.8 ng/ml and increased to 185.4 ng/ml in the presence of infection. CS-1-LI in the adrenal gland, small intestine, and hypothalamus also increased in rabbits with active inflammation. These data suggest that CS-1 may modify the hypothalamic-pituitary-adrenal axis in an endocrine or paracrine manner in response to infection.

Production and secretion of endothelin by hepatocellular carcinoma.

To clarify whether hepatocellular carcinoma (HCC) may produce and secrete endothelin (ET), we measured plasma levels of ET-1 and big ET-1, a precursor form of ET-1, in 30 patients with HCC. When compared to normal subjects, a substantial number of patients had elevated plasma ET-1 and big ET-1 levels, determined by specific enzyme immunoassays. The mean (+/- SD) plasma concentrations of ET-1 (1.7 +/- 0.9 pmol/L) and big ET-1 (6.1 +/- 4.8 pmol/L) in patients' group were significantly (P < 0.01) higher than those (1.0 +/- 0.3 and 2.0 +/- 0.8 pmol/L, respectively) in control group. There was a significant positive correlation between plasma big ET-1 and alpha-fetoprotein (r = 0.77, P < 0.01). Some of the 29 patients with liver cirrhosis also had modestly elevated plasma big ET-1 levels. The mean (+/- SD) plasma big ET-1 concentration (3.1 +/- 0.9 pmol/L) in patients with liver cirrhosis was significantly (P < 0.01) higher than that in control group, although there was no significant difference between the mean plasma ET-1 levels of both groups. Raised plasma big ET-1 and, less markedly, ET-1 levels in patients with HCC decreased after successful transcatheter arterial embolization concomitantly with a reduction in tumor sizes and a decrease in plasma alpha-fetoprotein levels. In six patients, an arteriovenous difference in ET-1 and big ET-1 levels across the tumor bed with a higher concentration in the venous circulation was found. Reverse-phase high performance liquid chromatography revealed that major portions of immunoreactive ET-1 and big ET-1 in hepatic venous plasma coeluted with synthetic ET-1 and big ET-1, respectively. Immunohistochemistry of HCC tissues from two patients demonstrated HCC cells positive for ET-1 and big ET-1, whereas no ET immunoreactivity was found in adjacent nontumorous hepatocytes. We conclude from these results that ET is produced by and released from a substantial number of HCC, which may stimulate proliferation of carcinoma cells as an autocrine or paracrine growth factor.

Characterization of AL amyloid protein identified by immunoelectron microscopy: a simple method using the protein A-gold technique.

The classification of amyloidosis depends on the chemical nature of the specific amyloid protein involved. Because AL amyloid protein consists mainly of variable regions of light chain (LC), immunohistochemical staining with conventional anti-LC antisera cannot identify its protein. We were able to classify three cases of AL amyloidosis, including one case of AL-kappa LC and two cases of AL-lambda LC, using post-embedding protein A-gold immunoelectron microscopy on autopsy-derived tissues. We describe here our procedure in which a protein A-gold staining apparatus was used. The main advantage of this method is that many sections can be stained and washed simultaneously under the same conditions. These results suggest that the post-embedding protein A-gold technique using conventional kappa or lambda LC may be useful in diagnosing AL amyloidosis.

Expression of prolactin mRNA in rat mammary gland during pregnancy and lactation.

We studied the expression of prolactin (PRL) mRNA in the mammary gland of resting, pregnant, lactating, and weanling rats using in situ and solution reverse transcriptase-polymerase chain reaction (RT-PCR). In mid- to late pregnancy and throughout lactation, PRL mRNA was detected in both in situ and solution RT-PCR. These PRL mRNA signals were clearly identified in the cytoplasm of alveolar and ductal mammary epithelial cells by the in situ RT-PCR method. In mid- to late pregnancy, such as at the initiating point of PRL mRNA expression, we confirmed in some cases a lack of PRL mRNA by solution RT-PCR. In addition, in the early weaning phase, no signals were detected by solution RT-PCR. However, slight focal signals were detected in some poorly vacuolated cytoplasm of regressing acinar cells by in situ RT-PCR. These findings suggest that PRL mRNA in rat mammary gland begins in mid- to late pregnancy in parallel with the development of the mammary gland, continues throughout lactation, and declines in the early phase of weaning, with regression of mammary epithelial cells.

Interleukin 10 expression is correlated with thrombospondin expression and decreased vascular involvement in colon cancer.

Interleukin 10 (IL-10) is an immuno-suppressive cytokine produced by T-lymphocytes, and a regulatory molecule for angiogenesis in various cancers. We examined IL-10 gene expression in 53 colon cancer patients who underwent surgical resection. IL-10 gene expression was correlated with TSP1 and TSP2 gene expression (P=0.0049, P=0.0285). Colon cancer with IL-10 gene expression (19/53) showed significantly decreased venous involvement (P=0.0433). The mean vessel counts in the colon cancers with IL-10 gene expression were significantly lower than those without IL-10 gene expression (P<0.001). These results suggested that IL-10 stimulates angiostatic factor gene expression, and results in suppression of venous involvement.

Gene expression of angiogenesis related factors in glioma.

Angiogenesis plays an important role in growth and proliferation of cancer. Various angiogenic and angiostatic factors regulate angiogenesis. In this study, we examined gene expression of the angiopoietin family including angiopoietin 1 (Ang1) and angiopoietin 2 (Ang2) in 39 gliomas and 5 glioma-xenografts by RT-PCR. Ang1 and Ang2 genes were expressed in 54%, and 77% of gliomas, respectively. The expression of Ang1 was significantly correlated with the expression of Ang2. Both Ang1 and Ang2 were shown to be expressed in the glioma cells. Ang2 gene expression was correlated with VEGF gene expression. Angiopoietin molecules may synergistically cooperate in growth and vascularization in glioma.

Catamenial pneumothorax associated with endometriosis of the diaphragm.

A case of catamenial pneumothorax is documented. At thoracotomy, there was a small black mass measuring 10 x 6 mm associated with surrounding petechiae on the top of the dome of the right diaphragm. Microscopic examination revealed the presence of endometrial tissue. We could not find any diaphragmatic defect in this case.

Expression of cadherins and their undercoat proteins (alpha-, beta-, and gamma-catenins and p120) and accumulation of beta-catenin with no gene mutations in synovial sarcoma.

E-cadherin, the major intercellular adhesion molecule of epithelial cells, is important in determining the architecture of sarcomas, especially those showing epithelioid features. In addition to its role in cell adhesion, beta-catenin, a cadherin undercoat protein, has been shown to function as a downstream transcriptional activator of the Wnt/Wingless signaling pathway. In order to evaluate the significance of the cadherin cell adhesion system and the Wnt/Wingless signaling pathway in the morphogenesis and/or tumorigenesis of synovial sarcoma (a major type of sarcoma with epithelioid features), immunoreactivity for pan-cadherin, E-cadherin, and their undercoat proteins (alpha-, beta-,and gamma-catenins and p120) was evaluated in 15 synovial sarcomas. Immunoreactivity for pan-cadherin, E-cadherin, alpha-catenin, beta-catenin, and p120 was observed in all 15 specimens. Immunoreactivity for pan-cadherin was stronger than that for E-cadherin. Expression of gamma-catenin was detected in ten specimens. Although beta-catenin was observed only at the cell-cell boundaries in four specimens, it was present in the nucleus and cytoplasm and at the cell-cell boundaries in the other 11, suggesting constitutional activation of the Wnt/Wingless signaling pathway in synovial sarcoma. Direct sequencing for exon 3 of the beta-catenin gene, however, revealed no mutations in any of the 15 specimens. In conclusion, other types of cadherin besides E-cadherin, together with cadherin undercoat proteins, may play a role in cell adhesion in synovial sarcoma. Furthermore, mechanisms other than mutation of exon 3 of the beta-catenin gene may activate the Wnt/Wingless signaling pathway in this type of tumor.

Surgical resection of a second primary lung carcinoma in a survivor of small cell carcinoma.

Resection of a second primary lung carcinoma in a long-term survivor of small cell lung carcinoma is reported. The patient had adenocarcinoma as the second primary tumor and underwent lobectomy with mediastinal node dissection. He was alive 99 months after the initial diagnosis of small cell carcinoma and 54 months after curative pulmonary resection for his second primary carcinoma.

Tracheoesophageal anastomosis for intractable aspiration pneumonia.

Permanent tracheostomy and tracheoesophageal anastomosis were performed as a means of surgical intervention for the treatment of intractable aspiration pneumonia. Conventional methods of tracheoesophageal anastomosis have entailed various problems. The improved method devised by us uses the special histological features and enables safe and reliable anastomosis. By this method, the tracheal perichondrium is retained and the strength of the anastomosed portion of the trachea is maintained, the failure of sutures due to tension on the anastomotic site being prevented. Also, reanastomosis can be performed after the cure of primary disease.

Lymphocytic hypophysitis: case report.

We report a rare case of lymphocytic hypophysitis in a 52-year-old man who presented with a combination of hypopituitarism and diabetes insipidus. Magnetic resonance imaging with a contrast medium revealed an expanding sellar mass and thickening of the pituitary stalk with homogeneous enhancement. These findings may be useful in differentiating lymphocytic hypophysitis from pituitary adenoma. The unique clinical and radiological features of this case are discussed.

Acromegaly, diabetes insipidus, and visual loss caused by metastatic growth hormone-releasing hormone-producing malignant pancreatic endocrine tumor in the pituitary gland. Case report.

The case of a 52-year-old woman with acromegaly, diabetes insipidus, and visual impairment caused by a metastatic growth hormone-releasing hormone (GRH)-produced pancreatic tumor is reported. Serum growth hormone (GH) and somatomedin C levels were elevated to 14 ng/ml (normal < 5 ng/ml), and 3.20 U/ml (normal < 1.88 U/ml), respectively. Paradoxical increases were observed in GH levels after glucose tolerance and thyrotropin-releasing hormone-stimulation tests. Biopsy of a pituitary tumor observed on computerized tomography scans and magnetic resonance studies revealed a metastatic cancer. When circulating GRH levels were measured, a marked increase in plasma GRH (1145 pg/ml; normal < 4-1 pg/ml) was observed. The patient died of cachexia due to metastases. Postmortem examination revealed that a primary tumor, a malignant endocrine lesion, was present in the pancreas, with metastatic tumors in the pituitary, lung, liver, and adrenal glands. Synthesis and production of GRH by the tumor was demonstrated by Northern blotting and immunohistochemical analysis. The pituitary gland showed hyperplastic, but not adenomatous changes. The authors stress the importance of both exploration for an ectopic source of GRH and the search for a GH-producing pituitary adenoma when unusual signs and symptoms are seen in patients with acromegaly.

Small mucosal carcinoma of the stomach with para-aortic lymph node metastasis: a case report and review of the literature.

A 38-year-old woman presented with a mucosal gastric carcinoma measuring 0.7 x 0.5 cm and para-aortic lymph node metastasis. Radiographic and endoscopic studies showed a small depressed lesion on the anterior border of the gastric angle, which was classified as a type II c + III lesion. Histological examination of the biopsy specimen revealed a signet-ring cell carcinoma. Distal gastrectomy with wide lymph node excision was performed. Detailed study of the resected specimen revealed that the tumour was limited to the mucosa, but metastasized to both the perigastric and para-aortic lymph nodes. The patient received adjuvant immunochemotherapy postoperatively. However, multiple bone metastases developed at 3 years and she died 4 years after the operation.

Parvovirus B19 infection induces apoptosis of erythroid cells in vitro and in vivo.

OBJECTIVE: intrauterine parvovirus B19 infection is related to non-immune hydrops fetalis, the pathogenesis of which is based on the strict tropism of B19 for erythroid precursor cells and the massive destruction of the infected erythroid cells, although the mechanism of beta19-induced cytotoxicity has not been studied in detail. The purpose of this study is to provide empirical evidence that beta19 induces apoptosis of erythroid cells both in vitro and ill vivo. METHODS: we analysed culture cells infected in vitro by B19 and tissues of nine cases of hydrops fetalis caused by B19 intrauterine infection by histological and biological methods. RESULTS: cells infected iil vitro by B19 showed nuclear changes characteristic of apoptosis by light microscopic examination and DNA extracted from the infected cells was fragmented. Electron microscopic examination showed the nuclei of infected cells contained crescent-shaped clumps of heterochromatin with increased density and double staining with anti-B1 9 antibody and terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate-digoxigenin nick-end labeling (TUNEL) confirmed apoptosis of individual cells. Tissues of cases of hydrops fetalis caused by B19 contained erythroid cells with nuclear inclusions and characteristic nuclear changes of apoptosis by light microscopy. The double-staining confirmed apoptosis of erythroid cells in the tissues. Immunohistochemical analysis with antibodies against cellular factors involved in apoptosis showed that caspase3, p53 and p21 were positive in infected cells.

Histochemical studies of metaplastic lesions in the human gallbladder.

We studied goblet cell metaplasia and pseudopyloric gland metaplasia in 25 surgically removed, paraffin-embedded gallbladder specimens using mucin histochemistry, silver methods for endocrine cells, and the indirect immunoperoxidase method for 12 peptide hormones, secretory component, and lysozyme. Goblet cell metaplasia was closely related to the occurrence of endocrine cells that showed argentaffinity, argyrophilia, or immunoreactive gastrin, somatostatin, pancreatic polypeptide, or motilin. Mucosal areas without goblet cell metaplasia were devoid of such endocrine cells. Metaplastic pseudopyloric glands showing lysozyme immunoreactivity were positive for class III mucin with paradoxical concanavalin A staining. Specimens with florid metaplastic lesions revealed a low tendency to form Rokitansky-Aschoff sinuses whose cells never showed a metaplastic nature. We compared the pathophysiological significance of metaplastic lesions in the gallbladder with intestinal metaplasia of the stomach.

Checkup interval and cancers in automated multiphasic health testing and services.

The purpose of this study was to disclose which types of cancer and how many persons with cancer were detected among the AMHTS examinees of our AMHTS center by using the hospital information retrieval system, and to study the relationship between cancer and the number of examinees, checkup intervals, and frequency in AMHTS. The examinees who had checkups more than twice were divided into three groups based on their checkup intervals: within one year, one to two years, and over two years. The relationship between cancer ratios and checkup intervals was evaluated in each group of examinees. In those having checkups within one year and from one to two years the cancer rate was 2.9 patients per 1,000 persons. However, in those having checkups after a two-year period or longer, the cancer rate was 4.3, clearly greater than the rate of the other two groups.

Acute gastric mucosal lesions associated with cytomegalovirus infection in a non-immunocompromised host.

A 75-year-old woman with epigastric pain and tarry stool was admitted to our hospital, where upper gastrointestinal endoscopic study revealed multiple gastric ulcers. The endoscopic biopsy specimens obtained on the seventh hospital day disclosed a few typical intranuclear cytomegalovirus inclusions. Cytomegalovirus-DNA was detected using polymerase chain reaction in a biopsy specimen. No immunologic abnormalities were demonstrated by any laboratory tests. While only a few cases of cytomegalovirus-associated gastric ulcer in non-immunocompromised hosts have been reported, this entity may be more frequently detected when careful histological examination is performed in the active stage rather than postponed until after healing of the ulcer.

Cyclic Cushing's disease in long-term remission with a daily low dose of bromocriptine.

A 56-year-old male patient with cyclic Cushing's disease remained in a state of remission for more than one year with a relatively low dose of bromocriptine (2.5-3.75 mg/day). It has been reported that bromocriptine treatment for cyclic Cushing's disease induces only a transient remission; in the most effective cases, a relatively high dose (40 mg/day) was necessary. In the hypercortisolemic state, plasma adrenocorticotropic hormone (ACTH) and serum cortisol were not suppressed by dexamethasone and did not respond to corticotropin-releasing factor (CRF). An antehypophysectomy was not effective, even though the resected tissue contained ACTH-positive microadenomas. The present observations thus indicate the effectiveness of bromocriptine for some patients with this rare disorder.