Seminiferous epithelium cycle of a hantavirus reservoir, the long-tailed mouse Oligoryzomys flavescens (Rodentia-Cricetidae).

Oligoryzomys flavescens, a common rodent of the temperate regions of South America, has been identified as a Hantavirus reservoir. There is still little information concerning its reproductive biology, which is essential to devise effective control measurement of natural populations. This rodent is a seasonal breeder and adult males exhibit a short period of testicular regression during winter months (June-August). In the present study we provided a histological and ultrastructural analysis of the composition of the testis of sexually mature O. flavescens during the breeding season. Over 95% of the testicular parenchyma was occupied by the seminiferous tubules and less than 5% by the interstitial tissue. The mean tubular diameter and epithelium height were 147.2 and 57.8 microm, respectively. The spermatogenic wave was characterized and eight spermatogenesis stages were identified according to the tubular morphology method. Their length, estimated as their relative frequencies, were (I-VIII) 8.8, 14.9, 4.0, 5.0, 10.4, 5.8, 27.0, and 23.9. Ultrastructural features of spermiogenesis are shown for the first time in a sigmodontine rodent.

Sex differentiation pattern in the annual fish Austrolebias charrua (Cyprinodontiformes: Rivulidae).

Sex differentiation process, determination of sexual strategy, and gametogenesis of the annual fish Austrolebias charrua are established. Evidence of histological sex differentiation in an antero-posterior gradient was observed in pre-hatching stages. Sexual strategy corresponds to the "differentiated gonochoric" pattern. Histological analyses of adult gonads showed an asynchronous spawning mode for females and continuous spawning for males. Mature oocytes presented fluid yolk. Testis organization corresponded to a restricted spermatogonial model. Herein, we report the ultrastructural organization of the vitelline envelope and the main features of the sperm of A. charrua. Taking together these results also contribute to phylogenetic studies and provide base line data to propose A. charrua as a biomonitor of contamination in a protected area.

Molecular detection of cancer cells in bone marrow and peripheral blood of patients with operable breast cancer. Comparison of CK19, MUC1 and CEA using RT-PCR.

We have compared three different RT-PCR procedures to measure cytokeratin 19 (CK19), carcinoembryonic antigen (CEA) and mucin MUC1 gene expression in order to determine their diagnostic value in detecting tumour cells in bone marrow aspirates of patients with operable breast cancer. In an experimental model, the best sensitivity was observed for CK19 and MUC1 RT-PCR assays, although only the CEA and CK19 assays showed good specificity. The study of 42 patients showed that a 'CK19 positive/CEA positive' RT-PCR assay in bone marrow correlated positively with a positive axillary lymph node status (N(0) versus N(1-3), P<0.05). Both assays were also positive in 17% of node negative patients. RT-PCR assays were more sensitive in bone marrow than in peripheral blood. Our results suggest that CK19 and CEA RT-PCR assays are powerful methods for detecting disseminated breast cancer cells. A larger study with long-term follow-up is required in order to clarify their clinical usefulness.

Detection of rare human breast cancer cells. Comparison of an immunomagnetic separation method with immunocytochemistry and RT-PCR.

BACKGROUND: The detection of occult carcinoma cells in patients with breast cancer may aid determination of prognosis and the development of new therapeutic approaches. In this study, we report a new method to detect rare human breast cancer cells, which combines an immunomagnetic separation (IMS) procedure with cytokeratin 19 (CK 19) immunostaining. MATERIALS AND METHODS: Four monoclonal antibodies (MAb) previously characterized against cell surface antigens (1BE12, ED8, 7B10 and 83D4), were evaluated for IMS optimization. Immunoseparated epithelial cells were identified using a MAb against CK 19. We compared the IMS procedure with the immunocytochemistry (ICC) and the RT-PCR for CK 19 on an "in vitro" experimental model. RESULTS: The best results in IMS procedures were obtained using MAbs 1BE12 (directed against Lewis y antigen) and ED8 (directed against MUC 1). In reconstitution experiments, using several ratios of T47D cells mixed with peripheral-blood mononuclear (PBMN) cells, the IMS procedure reliably detects one mammary carcinoma cell in 5 x 10(5) PBMN cells, whereas the ICC detects up to one T47D cell per 10(5) PBMN cells. The best sensitivity was observed with the RT-PCR (up to one T47D cell per 10(6) PBMN cells). We found the same high specificity with the three methods evaluated. CONCLUSIONS: The IMS procedure using MAbs 1BE12 or ED8 associated with CK 19 immunostaining is a specific, sensitive, and feasible method for the detection of rare human breast cancer cells. This method proved to be better than the ICC staining but its sensitivity was lower than that of RT-PCR for CK 19.

[Histology of the ovary of Macrodon ancylodon (Bloch and Schneider, 1801) Teleostei: Sciaenidae). Oogenesis. Post-ovulatory follicles. Atresia]. / Histología del ovario de Macrodon ancylodon (Bloch y Schneider, 1801) Teleostei: Sciaenidae) ovogénesis. Folículos post-ovulatorios. Atresia.

The reproductive cycle of Teleostean fishes may be studied from different points of view. One of them is to examine the histological changes that take place in the gonad. A histological description of the gonad is to be done first. In this work we have studied the oogenesis, atresia and post-ovulatory follicles of Macrodon ancylodon. Specimens were collected from April 1982 to May 1983 on the coast of Río de la Plata (Montevideo, Uruguay). Material used was preserved in neutralized 10% formalin inbedded in paraffin and paraffin-celloidin, sectioned at 5 - 10 microns and stained with haematoxilin-eosin. Pas-haematoxilin, and specific techniques for lipid detection were used. Six oogenetic stages were recognized: oogonias, and basophilic, lipid yolk vesicles, protein yolk vesicles, matures and ripe hydrated oocytes. Theca and granulosa are negative to lipid and cholesterol histochemic reaction techniques. Post-ovulatory follicles show structural degenerative changes. Two types of atresia are described: hypertrophic and nonhypertrophic, which apparently would not have an endocrine function. It is important to recognize post-ovulatory follicles to establish the spawning pattern and also to recognize atretic follicles due to their incidence in the fecundity of a species.

Tn antigen is a pre-cancerous biomarker in breast tissue and serum in n-nitrosomethylurea-induced rat mammary carcinogenesis.

The Tn determinant (GalNAcalpha-O-Ser/Thr), normally a cryptic structure in mucin-type O-glycans, is a tumor-associated marker which has attracted particular interest in cancer biology. We herein report the characterization of N-nitrosomethylurea (NMU)-induced breast cancer in rats as a new model for the study of aberrant O-glycosylation products. Tn-antigen expression is detectable not only in mammary carcinoma induced by NMU but also in carcinogen-initiated mammary epithelium, indicating that Tn could be a pre-cancerous biomarker in rats treated with NMU. Serum Tn levels were followed up longitudinally in 30 rats from the time of the first injection of NMU to the development of advanced breast cancer. Tn antigen increased in serum several weeks before tumor development, and became highly positive after 56 days of carcinogenesis (prior to breast-cancer occurrence), and the levels correlated with Tn expression in mammary tissues. However, during the follow-up after detection of mammary cancer, all animals displayed a significant decrease of serum Tn antigen, and low levels were observed in animals with advanced breast cancer. We have shown that the humoral immune response to cancer, with the production of anti-Tn antibodies, could hamper the detection of Tn antigen in animals with advanced breast cancer. These results suggest that NMU-induced rat mammary carcinogenesis is a useful experimental model to study the regulation of O-glycosylation at the cellular level during malignant transformation.

Analysis of a heterogeneous group of human breast carcinoma associated glycoproteins bearing the Tn determinant.

The Tn determinant (GalNAc alpha-O-Ser/Thr) is expressed by about 90% of human carcinomas, but is cryptic in most normal human tissues. A murine monoclonal antibody (MAb) 83D4, developed following immunization with human breast carcinoma cells, reacts with a Tn-related epitope. In the present study we characterized the glycoprotein antigen identified by 83D4 in the human breast carcinoma cell line MCF-7. We further showed that the 83D4 antigenic determinant is masked in human milk fat globule membranes (HMFGM), and can be exposed upon mild m-periodate treatment after desialylation. Western-blot analysis resolved the 83D4 antigen from MCF-7 into two main components of 120-190 kD and > 500 kD respectively. Non equilibrium pH gradient electrophoresis/SDS PAGE revealed the acidic nature of the reactive glycoproteins (pI 4.43-4.70). 83D4 antigenic activity resolved by CsCl gradient ultracentrifugation layered on a wide range of densities (1.30-1.46 g/ml) including typical densities of mucin-like glycoproteins but also lower densities. The amino acid composition of the antigen, relatively rich in serine but poor in threonine and proline, confirmed the divergence from other mucin-like carcinoma-associated glycoproteins. Dicarboxylic amino acids were abundant, accounting in part for the acidic nature of the molecules. ELISA and Western-blot analysis of the subcellular fractions from MCF-7 cells revealed that the 83D4 antigen is mainly contained in plasma membranes (85%) from which it may be resolved into two broad bands (slow and fast migrating components). These results provide information on a group of breast carcinoma associated glycoproteins related to but different from typical mucins, and provide data on alteration of O-glycosylation in tumor cells.