Immunohistochemical characterization of neoplastic cells of breast origin

ABSTRACT: BACKGROUND: After skin cancer, breast cancer is the most common malignancy in women. Tumors of unknown origin account for 5-15% of malignant neoplasms, with 1.5% corresponding to breast cancer. We selected an immunohistochemical panel with conventional and newer markers, such as mammaglobin, for the detection of neoplastic cells of breast origin. The specific objectives are: 1) to determine the sensitivity and specificity of the panel, with an especial emphasis on the inclusion of the mammaglobin marker, and 2) to compare immunohistochemistry performed on whole tissue sections and on Tissue Micro-Array. Methods. Twenty-nine metastatic breast tumors were included and assumed as tumors of unknown origin. Other 48 biopsies of other tissues were selected and assumed as negative controls. Tissue Micro-Array was performed. Immunohistochemistry for mammaglobin, gross cystic disease fluid protein-15, estrogen receptor, progesterone receptor and cytokeratin 7 was done. Results. Mammaglobin positive staining was observed in 10/29 cases, in 13/29 cases for gross cystic disease fluid protein-15, in 20/29 cases for estrogen receptor, 9/29 cases for progesterone receptor, and in 25/29 cases for cytokeratin 7. Among the negative controls, mammaglobin was positive in 2/48, and gross cystic disease fluid protein-15 in 4/48. Conclusions. The inclusion of MAG antibody in the immunohistochemical panel for the detection of tumors of unknown origin increased the probability of detecting metastasis of breast cancer. For the diagnostic strategy with the highest positive predictive value (87.5%), hormone receptors and mammaglobin were considered in serial, non-parallel manner.     

Development and preliminary validation of an antibody filtration-assisted single-dilution chemiluminometric immunoassay for potency testing of Piscirickettsia salmonis vaccines

Challenge with live pathogens could be substituted by serology for many veterinary diseases, however little progress has been made in the development of alternative batch vaccine potency tests for fish. This study reports the development and preliminary validation of a single-dilution filtration-assisted chemiluminometric immunoassay (SD FAL-ELISA) applied to measure anti Piscirickettsia salmonis IgM in individual or pooled serum and mucus samples. The assay was set up to test a single-dilution of the sample. Serum SD FAL-ELISA yielded a sensitivity of 90% and a specificity of 96%. SD FAL-ELISA was applied to evaluate pooled and individual samples from P. salmonis challenge assessments. Relative-light units values (RLU) obtained by SD FAL-ELISA were proportional to antibody levels in serum. RLU values obtained from pooled and individual serum samples increased with the observed relative percent survival (RPS) values, indicating a correlation between protection and specific IgM levels. Results obtained for specific IgM in mucus samples was not related to the RPS, but discriminated the vaccine that yielded high RPS (86.4%) from the others (40.9 and 54.5%). This is the first report on the development of an indirect high-throughput serological assessment for P. salmonis vaccine potency testing using both pooled or individual serum and cutaneous mucus samples.     

Nutritional value and intake of aquatic ferns (Azolla filiculoides Lam. and Salvinia molesta Mitchell.) in sows

Aquatic ferns (AFs) such as Azolla filiculoides and Salvinia molesta are grown on swine lagoons in the tropics and used in diets for pigs. The present work is aimed at evaluating their potential as feed ingredients for sows. When presented with ad libitum AFs, gilts weighing 110 ± 14 kg (mean ± SD), were able to ingest 9.1–9.7 kg fresh AF per day (from 597 to 630 g dry matter (DM) per day) and from 1240 to 1428 g DM per day when presented in a dry, ground form. A digestibility study was conducted, using sows weighing 213 ± 9 kg (mean ± SD), which were fed diets containing maize, soybean meal and 0, 150 or 300 g AF kg⁻¹ diet. The presence of AFs had a negative impact on the faecal digestibility of the crude protein, NDF and energy content of the whole diet (P<0.001) and on the ileal protein digestibility, especially with 300 g AFs kg⁻¹ diet. The level of AFs in the diet had no effect on stomach weight (P>0.05) but increased the weight of the rest of the gastrointestinal tract (P<0.001). The rate of AF fibre fermentation in the pig large intestine was measured using an in vitro gas test. The rates were much lower than tropical tree foliage, which can also be used in pig diets in the tropics. This could partly explain the low apparent digestibility of AFs in pigs. In conclusion, the inclusion level of AFs in rations for sows should be limited to 150 g AFs kg⁻¹ diet due to the low digestibility and energy density, as well as the negative impact on the digestibility of the whole diet.     

Segregation of SE Pacific and SW Atlantic southern blue whiting stocks: integrating evidence from complementary otolith microchemistry and parasite assemblage approaches

Southern blue whiting Micromesistius australis support one of the largest industrial fisheries in South America. Two main spawning/nursery grounds are known: one in the SW Atlantic Ocean (SWA), southwest from the Falkland (Malvinas) Islands; and other in the SE Pacific Ocean (SEP), south from the Taitao Peninsula. Juveniles originating from both grounds are believed to mix during migration and/or in feeding areas in the Scotia Sea. Previous efforts to distinguish stocks in this area have yielded contradictory results between genetics and otolith microchemical analyses. In the present work we revisited the null hypothesis of a single stock occurring in the broader SWA-SEP region by comparing and integrating results from different approaches: trace metals (Ca, Sr, Ba, Mg, Mn) and stable isotopes (δ¹³C, δ¹⁸O) in otolith cores, and parasite assemblage compositions in adults from SWA and SEP spawning grounds. We found significant differences in Sr:Ca, δ¹³C and δ¹⁸O mean ratios between spawning grounds. The best trace element discriminant model classified 83% of the samples. Each stable isotope discriminated >90% of the samples, while combining them into a bivariate discriminant model led to 100% classification success. Higher δ¹⁸O levels in the SWA samples agreed with lower mean temperature and higher ambient δ¹⁸O levels in that area. Parasite assemblage compositions also showed significant differences between grounds regarding the prevalence of Chondracanthus, Contracaecum, Hepatoxylon and Grillotia and the abundance of Diclidophora, Anisakis, Contracaecum, Hysterothylacium and Hepatoxylon. Parasite-based discriminant models supported 90–100% correct assignment of samples to capture location. Although preliminary due to limited sampling coverage, our results support the existence of at least two ecologically distinct sub-populations of southern blue whiting in South America. The joint use of otolith microchemistry and parasitological techniques showed to be a promising way to test hypotheses concerning ecological stocks in marine fishes.     

Progesterone receptor isoforms differentially regulate the expression of tryptophan and tyrosine hydroxylase and glutamic acid decarboxylase in the rat hypothalamus

Progesterone exerts a variety of actions in the brain through the interaction with its receptors (PR) which have two isoforms with different function and regulation: PR-A and PR-B. Progesterone may modulate neurotransmission by regulating the expression of neurotransmitters synthesizing enzymes or their receptors in several brain regions. The role of PR isoforms in this modulation is unknown. We explored the role of PR isoforms in the regulation of tryptophan (TPH) and tyrosine (TH) hydroxylase, and glutamic acid decarboxylase (GAD) expression in the hypothalamus of ovariectomized rats. Two weeks after ovariectomy, animals were subcutaneously injected with 5 μg of estradiol benzoate (EB), and 40 h later, progesterone (P) was intracerebroventricularly (ICV) injected. Each animal received two ICV injections of 1 μg/μl (4 nmol) of PR-B and total PR (PR-A + PR-B) sense or antisense (As) oligonucleotides (ODNs). First injection was made immediately before sc EB injection, and 24 h later animals received the second one. Twenty-four hours after P administration, rats were euthanized and brains removed to measure the expression of PR-A and PR-B, TPH, TH and GAD by Western blot. We observed that sense ODNs modified neither PR isoforms nor enzymes expression in the hypothalamus, whereas PR A + B antisense (PR A + B As) clearly decreased the expression of both PR isoforms in this region. ICV administration of PR-B As only decreased PR-B isoform expression with no significant effects on PR-A expression. A differential protein expression of TPH, TH and GAD was observed after PR isoforms antisense administration. PR-B As administration decreased the expression of TPH (65% with respect to control). In contrast, PR A + B As and PR-B As administration increased (51.6% and 34.4%, respectively) TH expression. The administration of PR A + B As and PR-B As diminished GAD expression (33.4% and 41.6%, respectively). Our findings indicate that PR isoforms play a differential role in the regulation of the content of TPH, TH and GAD in the rat hypothalamus.     

Apical meristem exhaustion during determinate primary root growth in the moots koom 1 mutant of Arabidopsis thaliana

An indeterminate developmental program allows plant organs to grow continuously by maintaining functional meristems over time. The molecular mechanisms involved in the maintenance of the root apical meristem are not completely understood. We have identified a new Arabidopsis thaliana mutant named moots koom 1 (mko1) that showed complete root apical meristem exhaustion of the primary root by 9?days post-germination. MKO1 is essential for maintenance of root cell proliferation. In the mutant, cell division is uncoupled from cell growth in the region corresponding to the root apical meristem. We established the sequence of cellular events that lead to meristem exhaustion in this mutant. Interestingly, the SCR and WOX5 promoters were active in the mko1 quiescent center at all developmental stages. However, during meristem exhaustion, the mutant root tip showed defects in starch accumulation in the columella and changes in auxin response pattern. Therefore, contrary to many described mutants, the determinate growth in mko1 seedlings does not appear to be a consequence of incorrect establishment or affected maintenance of the quiescent center but rather of cell proliferation defects both in stem cell niche and in the rest of the apical meristem. Our results support a model whereby the MKO1 gene plays an important role in the maintenance of the root apical meristem proliferative capacity and indeterminate root growth, which apparently acts independently of the SCR/SHR and WOX5 regulatory pathways.     

Digestive tract morphology of the Neotropical piscivorous fish Cichla kelberi (Perciformes: Cichlidae) introduced into an oligotrophic Brazilian reservoir

Despite being one of the most well-known cichlid fish of importance to artisanal and sport fishing, and among the largest fishes in the Neotropics, data on digestive tract anatomy of peacock basses (Cichla spp.) are largely lacking, especially for non-native populations. In this paper, we describe for the first time the digestive tract morphology of Cichla kelberi, a voracious piscivore that was introduced in the 1950s into an oligotrophic and physically low-complex impoundment in Brazil. Peacock basses were collected between 1994 and 2002 in Lajes Reservoir, through gillnets (25 to 55mm mesh; 20-50 x 2m), seines (10 x 2.5m; 8.0mm mesh), cast nets and angling. All the fishes were kept on ice in the field and then transferred to the laboratory, where they were identified, measured, weighed and dissected for digestive tract analyses. The Index of Relative Importance-IRI was calculated for diet characterization while linear and non-linear regressions were performed to assess growth patterns of four morphological characters related to feeding (e.g. mouth width, mouth height, stomach length and intestine length) and the number of gill rakers during the C. kelberi ontogeny. Most digestive tract structures were directly related to the piscivorous diet of C. kelberi, indicating that peacock bass is a diurnal, bathypelagic and gape-size limited predator that feeds largely on shallow-water prey species within the littoral zone. Mouth width and height grew allometrically (b > 1) with the size of peacock bass, broadening the size range in which prey can be eaten, but especially for predators smaller than -400mm of total length. Differently, stomach and intestine lengths increased isometrically (b = 1), which could constrain prey consumption for adult C. kelberi, especially those at advanced stages of gonadal maturation. The presence of longer-drawn, sharp and furcated gill rakers in C. kelberi may be related to increased prey retention in the resource-limited Lajes Reservoir, but further studies are necessary whether such features are randomly triggered by genetic or phenotypic anomalies, or effectively bring ecological advantages to the predator. In addition to contribute to improve the current biological knowledge on peacock basses, our results can be also useful to further comparisons on whether those morphological features related to feeding will change with transitions on prey consumption by C. kelberi and/or with the particular conditions of the invaded ecosystem.