Fine structure of Henneguya hemiodopsis sp. n. (Myxozoa), a parasite of the gills of the Brazilian teleostean fish Hemiodopsis microlepes (Hemiodontidae)

A fish-infecting myxosporean, Henneguya hemiodopsis sp. n., found infecting the gills of Hemiodopsis microlepis and collected from the Poty River near the city of Teresina, Brazil, was described based on ultrastructural studies. The parasite occurred within large whitish polysporic plasmodia (up to 200 μm in diameter) containing asynchronous developmental sporogonic stages, mainly mature spores. The spores measured 19.7 ± 0.9 μm in total length (n = 30) and the ellipsoidal spore body was 10.8 ± 0.5 μm long, 3.3 ± 0.4 μm wide and 2.5 ± 0.5 μm thick. The spores were composed of two equal shell valves adhering together along the straight suture line, with each valve having equal-sized caudal tapering tails measuring 8.7 ± 0.6 μm in length. The spores were surrounded by a thin anastomosed network of microfibrils, more evident on the tails. There were two symmetric elongated bottle-like polar capsules 3.5 ± 0.3 μm long and 1.0 ± 0.2 μm wide, each with a polar filament with five to six coils. Given the morphological and ultrastructural differences from previously described parasites and the specificity of the host species, we propose a new species, named H. hemiodopsis sp. n.

Light and electron microscopy of Myxobolus sciades n. sp. (Myxozoa), a parasite of the gills of the Brazilian fish Sciades herzbergii (Block, 1794) (Teleostei: Ariidae)

A myxosporean parasite in the gill lamellae of the freshwater teleost fish, Sciades herzbergii (Ariidae) (Block, 1794), from the Poti River (Northeast of Brazil) was described by light and electron microscopy studies. Polysporic histozoic cyst-like plasmodia containing several life-cycle stages, including mature spores, were observed. The spores were pyriform and uninucleate, measuring 9.15 ± 0.39 μm (n = 50) long, 4.36 ± 0.23 μm (n = 25) wide and 2.61 ± 0.31 μm (n = 25) thick. Elongated pyriform polar capsules (PC) were of equal size (4.44 ± 0.41 μm long and 1.41 ± 0.42 μm in diameter) and each contained a polar filament with 9-10 coils obliquely arranged in relation to the axis of PC. The PC wall was composed of two layers of different electron densities. Histological analysis revealed the close contact of the cyst-like plasmodia with the basal portion of the epithelial gill layer, which exhibited some alterations in the capillary vessels. Based on the morphological and ultrastructural differences, the similarity of the spore features to those of the genus Myxobolus and the specificity of this host to previously described species, we describe a new species named Myxobolus sciades n. sp. in this study.

2D-immunoblotting analysis of Sporothrix schenckii cell wall

We utilized two-dimensional gel electrophoresis and immunoblotting (2D-immunoblotting) with anti-Sporothrix schenckii antibodies to identify antigenic proteins in cell wall preparations obtained from the mycelial and yeast-like morphologies of the fungus. Results showed that a 70-kDa glycoprotein (Gp70) was the major antigen detected in the cell wall of both morphologies and that a 60-kDa glycoprotein was present only in yeast-like cells. In addition to the Gp70, the wall from filament cells showed four proteins with molecular weights of 48, 55, 66 and 67 kDa, some of which exhibited several isoforms. To our knowledge, this is the first 2D-immunoblotting analysis of the S. schenckii cell wall.

Pauciconfibula patagonensis sp. nov. (Monogenea: Microcotylidae) parasitizing the horsefish, Congiopodus peruvianus (Pisces: Congiopodidae), from the Patagonian Shelf, Argentina

Pauciconfibula patagonensis sp. nov. (Monogenea: Microcotylidae), parasite of gill filaments of the horsefish, Congiopodus peruvianus (Congiopodidae) collected in the Patagonian Shelf, Argentina, is described and illustrated. The new species is characterized by having intestinal caeca not confluent and entering into the haptor, vitelline follicles extending from the genital pore to near the posterior portion of haptor, two parallel rows each comprised of 16-20 microcotylid clamps in the haptor, 25-43 testes and a fusiform egg with one very long tangled polar filament. P. patagonensis is the only member of the genus known to parasitize a scorpaeniform host and represents the first record of a representative of this genus in the southern Atlantic Ocean.

Lack of evidence for KRAS oncogenic mutations in triple-negative breast cancer.

BACKGROUND Mutational analysis of the KRAS gene has recently been established as a complementary in vitro diagnostic tool for the identification of patients with colorectal cancer who will not benefit from anti-epidermal growth factor receptor (EGFR) therapies. Assessment of the mutation status of KRAS might also be of potential relevance in other EGFR-overexpressing tumors, such as those occurring in breast cancer. Although KRAS is mutated in only a minor fraction of breast tumors (5%), about 60% of the basal-like subtype express EGFR and, therefore could be targeted by EGFR inhibitors. We aimed to study the mutation frequency of KRAS in that subtype of breast tumors to provide a molecular basis for the evaluation of anti-EGFR therapies. METHODS Total, genomic DNA was obtained from a group of 35 formalin-fixed paraffin-embedded, triple-negative breast tumor samples. Among these, 77.1% (27/35) were defined as basal-like by immunostaining specific for the established surrogate markers cytokeratin (CK) 5/6 and/or EGFR. KRAS mutational status was determined in the purified DNA samples by Real Time (RT)-PCR using primers specific for the detection of wild-type KRAS or the following seven oncogenic somatic mutations: Gly12Ala, Gly12Asp, Gly12Arg, Gly12Cys, Gly12Ser, Gly12Val and Gly13Asp. RESULTS We found no evidence of KRAS oncogenic mutations in all analyzed tumors. CONCLUSIONS This study indicates that KRAS mutations are very infrequent in triple-negative breast tumors and that EGFR inhibitors may be of potential benefit in the treatment of basal-like breast tumors, which overexpress EGFR in about 60% of all cases.

Myxidium volitans sp. nov., a parasite of the gallbladder of the fish, Dactylopterus volitans (Teleostei: Triglidae) from the Brazilian Atlantic coast: morphology and pathology

Myxidium volitans sp. nov. (Myxozoa: Myxidiidae) parasitizing the hypertrophied green-brownish gallbladder of the teleost Dactylopterus volitans, collected in the Atlantic coast near Niterói, Brazil was described based on ultrastructural studies. The spores were fusiform, sometimes slightly crescent-shaped on average 21.7 ± 0.3 µm (mean ± standard deviation) (n = 50) long and 5.6 ± 0.4 µm (n = 30) wide. The spore wall was thin and smooth, comprising two equally-sized valves joined by a hardly visible sutural ridge. Spores containing two pyriform polar capsules (PC) (5.0 ± 0.4 × 2.3 ± 0.3 µm) (n = 30) are situated in each extremity of the spore. The PC wall was composed of hyaline layer (0.20-0.29 µm thick) and by a thin external granular layer. Each PC contains a polar filament (PF) with irregular arrangements that was projected from its apical region to the bases of PC and coiled laterally from bases to the tip of PC. Some regular striations and S-like structures in the periphery of the PFs with four-five irregular sections were observed. Based on the spore morphology, ultrastructural differences and the specificity of the host we describe this parasite as a new myxosporidian, named M. volitans sp. nov.

Prokaryotic cells: structural organisation of the cytoskeleton and organelles

For many years, prokaryotic cells were distinguished from eukaryotic cells based on the simplicity of their cytoplasm, in which the presence of organelles and cytoskeletal structures had not been discovered. Based on current knowledge, this review describes the complex components of the prokaryotic cell cytoskeleton, including (i) tubulin homologues composed of FtsZ, BtuA, BtuB and several associated proteins, which play a fundamental role in cell division, (ii) actin-like homologues, such as MreB and Mb1, which are involved in controlling cell width and cell length, and (iii) intermediate filament homologues, including crescentin and CfpA, which localise on the concave side of a bacterium and along its inner curvature and associate with its membrane. Some prokaryotes exhibit specialised membrane-bound organelles in the cytoplasm, such as magnetosomes and acidocalcisomes, as well as protein complexes, such as carboxysomes. This review also examines recent data on the presence of nanotubes, which are structures that are well characterised in mammalian cells that allow direct contact and communication between cells.

Ultrastructural description of Ceratomyxa microlepis sp. nov. (Phylum Myxozoa): a parasite infecting the gall bladder of Hemiodus microlepis, a freshwater teleost from the Amazon River

A new ceratomyxid parasite was examined for taxonomic identification, upon being found infecting the gall bladder of Hemiodus microlepis (Teleostei: Hemiodontidae), a freshwater teleost collected from the Amazon River, Brazil. Light and transmission electron microscopy revealed elongated crescent-shaped spores constituted by two asymmetrical shell valves united along a straight sutural line, each possessing a lateral projection. The spores body measured 5.2 ± 0.4 µm (n = 25) in length and 35.5 ± 0.9 µm (n = 25) in total thickness. The lateral projections were asymmetric, one measuring 18.1 ± 0.5 µm (n = 25) in thickness and the other measuring 17.5 ± 0.5 µm (n = 25) in thickness. Two equal-sized subspherical polar capsules measuring 2.2 ± 0.3 µm in diameter were located at the same level, each possessing a polar filament with 5-6 coils. The sporoplasm was binucleate. Considering the morphometric data analyzed from the microscopic observations, as well as the host species and its geographical location, this paper describes a new myxosporean species, herein named Ceratomyxa microlepis sp. nov.; therefore representing the first description of a freshwater ceratomyxid from the South American region.

PARP-1 regulates metastatic melanoma through modulation of vimentin-induced malignant transformation.

PARP inhibition can induce anti-neoplastic effects when used as monotherapy or in combination with chemo- or radiotherapy in various tumor settings; however, the basis for the anti-metastasic activities resulting from PARP inhibition remains unknown. PARP inhibitors may also act as modulators of tumor angiogenesis. Proteomic analysis of endothelial cells revealed that vimentin, an intermediary filament involved in angiogenesis and a specific hallmark of EndoMT (endothelial to mesenchymal transition) transformation, was down-regulated following loss of PARP-1 function in endothelial cells. VE-cadherin, an endothelial marker of vascular normalization, was up-regulated in HUVEC treated with PARP inhibitors or following PARP-1 silencing; vimentin over-expression was sufficient to drive to an EndoMT phenotype. In melanoma cells, PARP inhibition reduced pro-metastatic markers, including vasculogenic mimicry. We also demonstrated that vimentin expression was sufficient to induce increased mesenchymal/pro-metastasic phenotypic changes in melanoma cells, including ILK/GSK3-β-dependent E-cadherin down-regulation, Snail1 activation and increased cell motility and migration. In a murine model of metastatic melanoma, PARP inhibition counteracted the ability of melanoma cells to metastasize to the lung. These results suggest that inhibition of PARP interferes with key metastasis-promoting processes, leading to suppression of invasion and colonization of distal organs by aggressive metastatic cells.

Pattern of recurrence of early breast cancer is different according to intrinsic subtype and proliferation index.

INTRODUCTION Recurrence risk in breast cancer varies throughout the follow-up time. We examined if these changes are related to the level of expression of the proliferation pathway and intrinsic subtypes. METHODS Expression of estrogen and progesterone receptor, Ki-67, human epidermal growth factor receptor 2 (HER2), epidermal growth factor receptor (EGFR) and cytokeratin 5/6 (CK 5/6) was performed on tissue-microarrays constructed from a large and uniformly managed series of early breast cancer patients (N = 1,249). Subtype definitions by four biomarkers were as follows: luminal A (ER + and/or PR+, HER2-, Ki-67 <14), luminal B (ER + and/or PR+, HER2-, Ki-67 ≥14), HER2-enriched (any ER, any PR, HER2+, any Ki-67), triple-negative (ER-, PR-, HER2-, any Ki-67). Subtype definitions by six biomarkers were as follows: luminal A (ER + and/or PR+, HER2-, Ki-67 <14, any CK 5/6, any EGFR), luminal B (ER + and/or PR+, HER2-, Ki-67 ≥14, any CK 5/6, any EGFR), HER2-enriched (ER-, PR-, HER2+, any Ki-67, any CK 5/6, any EGFR), Luminal-HER2 (ER + and/or PR+, HER2+, any Ki-67, any CK 5/6, any EGFR), Basal-like (ER-, PR-, HER2-, any Ki-67, CK5/6+ and/or EGFR+), triple-negative nonbasal (ER-, PR-, HER2-, any Ki-67, CK 5/6-, EGFR-). Each four- or six-marker defined intrinsic subtype was divided in two groups, with Ki-67 <14% or with Ki-67 ≥14%. Recurrence hazard rate function was determined for each intrinsic subtype as a whole and according to Ki-67 value. RESULTS Luminal A displayed a slow risk increase, reaching its maximum after three years and then remained steady. Luminal B presented most of its relapses during the first five years. HER2-enriched tumors show a peak of recurrence nearly twenty months post-surgery, with a greater risk in Ki-67 ≥14%. However a second peak occurred at 72 months but the risk magnitude was greater in Ki-67 <14%. Triple negative tumors with low proliferation rate display a smooth risk curve, but with Ki-67 ≥14% show sharp peak at nearly 18 months. CONCLUSIONS Each intrinsic subtype has a particular pattern of relapses over time which change depending on the level of activation of the proliferation pathway assessed by Ki-67. These findings could have clinical implications both on adjuvant treatment trial design and on the recommendations concerning the surveillance of patients.

Chronic nitric oxide synthase inhibition and carotid artery distensibility in renal hypertensive rats.

The goal of the present study was to examine the viscoelastic properties of the carotid artery in genetically identical rats exposed to similar levels of blood pressure sustained by different mechanisms. Eight-week old male Wistar rats were examined 2 weeks after renal artery clipping (two-kidney, one clip [2K1C] Goldblatt rats, n = 53) or sham operation (n = 49). One half of the 2K1C and sham rats received the nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME, 1.48 mmol/L) in their drinking water for 2 weeks after the surgical procedure. Mean blood pressure increased significantly in the 2K1C-water (182 mm Hg), 2K1C-L-NAME (197 mm Hg), and sham-L-NAME (170 mm Hg) rats compared with the sham-water rats (127 mm Hg). Plasma renin activity was not altered by L-NAME but significantly enhanced after renal artery clipping. A significant and similar increase in the cross-sectional area of the carotid artery was observed in L-NAME and vehicle-treated 2K1C rats. L-NAME per se did not modify cross-sectional area in the sham rats. There was a significant upward shift of the distensibility-pressure curve in the L-NAME- and vehicle-treated 2K1C rats compared with the sham-L-NAME rats. L-NAME treatment did not alter the distensibility-pressure curve in the 2K1C rats. These results demonstrate that the mechanisms responsible for artery wall hypertrophy in renovascular hypertension are accompanied by an increase in arterial distensibility that is not dependent on the synthesis of nitric oxide.

EMT and EGFR in CTCs cytokeratin negative non-metastatic breast cancer.

Circulating tumor cells (CTCs) are frequently associated with epithelial-mesenchymal transition (EMT).The objective of this study was to detect EMT phenotype through Vimentin (VIM) and Slug expression in cytokeratin (CK)-negative CTCs in non-metastatic breast cancer patients and to determine the importance of EGFR in the EMT phenomenon. In CK-negative CTCs samples, both VIM and Slug markers were co-expressed in the most of patients. Among patients EGFR+, half of them were positive for these EMT markers. Furthermore, after a systemic treatment 68% of patients switched from CK- to CK+ CTCs. In our experimental model we found that activation of EGFR signaling by its ligand on MCF-7 cells is sufficient to increase EMT phenotypes, to inhibit apoptotic events and to induce the loss of CK expression. The simultaneous detection of both EGFR and EMT markers in CTCs may improve prognostic or predictive information in patients with operable breast cancer.

Estimation of inelastic seismic material properties of a surgical sea-bed from multi-component marine seismic data

To date, state-of-the-art seismic material parameter estimates from multi-component sea-bed seismic data are based on the assumption that the sea-bed consists of a fully elastic half-space. In reality, however, the shallow sea-bed generally consists of soft, unconsolidated sediments that are characterized by strong to very strong seismic attenuation. To explore the potential implications, we apply a state-of-the-art elastic decomposition algorithm to synthetic data for a range of canonical sea-bed models consisting of a viscoelastic half-space of varying attenuation. We find that in the presence of strong seismic attenuation, as quantified by Q-values of 10 or less, significant errors arise in the conventional elastic estimation of seismic properties. Tests on synthetic data indicate that these errors can be largely avoided by accounting for the inherent attenuation of the seafloor when estimating the seismic parameters. This can be achieved by replacing the real-valued expressions for the elastic moduli in the governing equations in the parameter estimation by their complex-valued viscoelastic equivalents. The practical application of our parameter procedure yields realistic estimates of the elastic seismic material properties of the shallow sea-bed, while the corresponding Q-estimates seem to be biased towards too low values, particularly for S-waves. Given that the estimation of inelastic material parameters is notoriously difficult, particularly in the immediate vicinity of the sea-bed, this is expected to be of interest and importance for civil and ocean engineering purposes.

A regulatory network for coordinated flower maturation.

For self-pollinating plants to reproduce, male and female organ development must be coordinated as flowers mature. The Arabidopsis transcription factors AUXIN RESPONSE FACTOR 6 (ARF6) and ARF8 regulate this complex process by promoting petal expansion, stamen filament elongation, anther dehiscence, and gynoecium maturation, thereby ensuring that pollen released from the anthers is deposited on the stigma of a receptive gynoecium. ARF6 and ARF8 induce jasmonate production, which in turn triggers expression of MYB21 and MYB24, encoding R2R3 MYB transcription factors that promote petal and stamen growth. To understand the dynamics of this flower maturation regulatory network, we have characterized morphological, chemical, and global gene expression phenotypes of arf, myb, and jasmonate pathway mutant flowers. We found that MYB21 and MYB24 promoted not only petal and stamen development but also gynoecium growth. As well as regulating reproductive competence, both the ARF and MYB factors promoted nectary development or function and volatile sesquiterpene production, which may attract insect pollinators and/or repel pathogens. Mutants lacking jasmonate synthesis or response had decreased MYB21 expression and stamen and petal growth at the stage when flowers normally open, but had increased MYB21 expression in petals of older flowers, resulting in renewed and persistent petal expansion at later stages. Both auxin response and jasmonate synthesis promoted positive feedbacks that may ensure rapid petal and stamen growth as flowers open. MYB21 also fed back negatively on expression of jasmonate biosynthesis pathway genes to decrease flower jasmonate level, which correlated with termination of growth after flowers have opened. These dynamic feedbacks may promote timely, coordinated, and transient growth of flower organs.

Selective changes in the neurofilament and microtubule cytoskeleton of NF-H/LacZ mice.

This study focused mainly on changes in the microtubule cytoskeleton in a transgenic mouse where beta-galactosidase fused to a truncated neurofilament subunit led to a decrease in neurofilament triplet protein expression and a loss in neurofilament assembly and abolished transport into neuronal processes in spinal cord and brain. Although all neurofilament subunits accumulated in neuronal cell bodies, our data suggest an increased solubility of all three subunits, rather than increased precipitation, and point to a perturbed filament assembly. In addition, reduced neurofilament phosphorylation may favor an increased filament degradation. The function of microtubules seemed largely unaffected, in that tubulin and microtubule-associated proteins (MAP) expression and their distribution were largely unchanged in transgenic animals. MAP1A was the only MAP with a reduced signal in spinal cord tissue, and differences in immunostaining in various brain regions corroborate a relationship between MAP1A and neurofilaments.