A THREE-DIMENSIONAL RECONSTRUCTION OF THE VASCULAR SYSTEM TO THE LODICULES,ANDROECIUM, AND GYNOECIUM OF A FERTILE FLORET OF PANICUM DICHOTOMIFLORUM (GRAMINEAE)

A three-dimensional reconstruction of a fertile floret stele of Panicum dichotomiflorum approaching anthesis was made by a new technique using superimposition of tracings of 80, 1-μm thick serial sections, cleared tracing film, and mounting adhesive. From a collateral bundle, which also served as the median trace to the fertile lemma, most of the vascular tissue branched adaxially and horizontally to become the sole vascular supply to the two lodicules, three stamens, and pistil. The xylem branched at a low level to form a broad and long vessel plexus. The phloem branched at a higher level to overlay the vessel plexus on the right and left with an arc of horizontal sieve tubes in a phloem plexus. Those sieve tubes and vessels which rose after branching from the horizontal plexi assumed a more vertical course in the floret stele. Traces to the right and left lodicules arose from the lower abaxial portions of the flanks of the floret stele. Vessels ascended vertically from the xylem plexus and passed through the phloem plexi and joined with the sieve tubes there to exit at the same level and become the right and left lodicule traces. The vascular tissues to the three filament traces arose from different higher levels of the stele. The sieve tubes for the median filament trace arose vertically from the abaxial side between but above the lodicule traces. At higher levels the sieve tubes for the lateral filaments rose from the horizontal arcs of the flanks of the stele and departed it tangentially. The vessels destined to the filament traces arose in the center of the floret stele from adaxial portions of the horizontal plexus, ascended between the arcs of phloem, exited the stele simultaneously above the phloem of the traces, and followed the courses of their respective sieve tubes. The adaxially displaced apex of the floret stele then contained the vascular tissue related to the pistil. All the sieve tubes and vessels of the floret stele were embedded in a matrix of intermediary cells. The peripheral intermediary cells associated with the vessel plexus were xylem transfer cells with pronounced wall ingrowths. At higher levels in the floret stele, intermediary cells in scattered locations near sieve tubes or vessels had less conspicuous wall ingrowths. No preferred orientation of transfer cells with any particular trace was noted.     

An inhibitory mechanism of action of coiled‐coil peptides against type three secretion system from enteropathogenic Escherichia coli

Human pathogenic gram‐negative bacteria, such as enteropathogenic Escherichia coli (EPEC), rely on type III secretion systems (T3SS) to translocate virulence factors directly into host cells. The coiled‐coil domains present in the structural proteins of T3SS are conformed by amphipathic alpha‐helical structures that play an important role in the protein‐protein interaction and are essential for the assembly of the translocation complex. To investigate the inhibitory capacity of these domains on the T3SS of EPEC, we synthesized peptides between 7 and 34 amino acids based on the coiled‐coil domains of proteins that make up this secretion system. This analysis was performed through in vitro hemolysis assays by assessing the reduction of T3SS‐dependent red blood cell lysis in the presence of the synthesized peptides. After confirming its inhibitory capacity, we performed molecular modeling assays using combined techniques, docking‐molecular dynamic simulations, and quantum‐mechanic calculations of the various peptide‐protein complexes, to improve the affinity of the peptides to the target proteins selected from T3SS. These techniques allowed us to demonstrate that the peptides with greater inhibitory activity, directed against the coiled‐coil domain of the C‐terminal region of EspA, present favorable hydrophobic and hydrogen bond molecular interactions. Particularly, the hydrogen bond component is responsible for the stabilization of the peptide‐protein complex. This study demonstrates that compounds targeting T3SS from pathogenic bacteria can indeed inhibit bacterial infection by presenting a higher specificity than broad‐spectrum antibiotics. In turn, these peptides could be taken as initial structures to design and synthesize new compounds that mimic their inhibitory pharmacophoric pattern.     

Serous cutaneous glands in new world hylid frogs: an ultrastructural study on skin poisons confirms phylogenetic relationships between Osteopilus septentrionalis and Phrynohyas venulosa

Transmission electron microscope investigations of the serous (poison) skin glands in the New World tree frogs Osteopilus septentrionalis and Phrynohyas venulosa revealed that they produce granules with closely similar substructures, namely, a dense cortex and pale medulla. In both species these features, that contrast the complex, sometimes repeating patterns described in other hylid frogs, derive from similar secretory and maturational processes starting from the Golgi phase of poison biosynthesis. Observations on secretory discharge showed that the two species share common release mechanisms, based on bulk discharge (holocrine) processes. Our data provide novel evidence of the extensive ultrastructural polymorphism of serous skin products in Hylidae and agree with phylogenies that regard this family as polyphyletic in origin. Assuming that ultrastructural features of cutaneous poison biosynthesis and maturation are adequate clues for tracking anuran phylogeny, the present findings also support a close relationship between Osteopilus and Phrynohyas taxa as previously suggested by osteological evidence.     

Molecular profile of androgen-independent prostate cancer xenograft LuCaP 23.1

After castration or therapeutic hormone deprivation, most cancer of the prostate (CaP) cells develop androgen-independent (AI) growth. In this work, we studied the effect of androgen depletion (castration) on the growth of experimental model LuCaP 23.1 xenograft. A total of 101 nude mice were implanted and analysed for their growth profile before experimental period 1 (11 weeks) and after castration experimental period 2 (15 weeks). For specific periods, tumors were harvested and assessed for molecular marker expression specific for CaP. Taking into account tumor dynamic growth, prior to castration we found 37 fast growing (FG) tumors (948.9 ± 76.9 mm³) and 63 slow growing (SG) tumors (229.6 ± 18.4 mm³). Real-time quantitative RT-PCR showed that in comparison to SGs, FGs contained elevated expression of epidermal growth factor receptor type 1 (HER1), urokinase plasminogen activator (uPA), thymidine phosphorylase (TP) and thymidilate synthase (TS) mRNAs expression and low levels of 5-reductase 2 (5-R2) mRNA. After castration all FG tumors progressed rapidly (by 5 weeks) to AI growth (FG-P). In SG castrated tumors, 66% of tumors showed retarded progression (by 12 weeks) to AI (SG-P), whereas 34% responded to castration (SG-R). Molecular analysis demonstrated distinct molecular profiles integrating different pathways associated with AI progression. The progressive tumors FG-P, and some tumors of SG-P subgroup, presented significantly high levels of HER1, epidermal growth factor receptor type 2 (HER2), TS, uPA, TP, tumor necrosis factor superfamily member 6 (FAS) and peptidylglycine -amidating mono-oxygenase (PAM) mRNA all of which correlated with androgen receptor (AR) mRNA. The second subgroup of SG-P tumors showed a high expression of the anti-apoptotic gene Bcl-2. A third subgroup of SG-P tumors showed significant expression of hypoxia-related genes such as adrenomedullin (AM) after castration. LuCaP 23.1 xenograft represent a useful dynamic model to study pre-clinically new therapeutic molecules and evaluate non-randomized therapeutics protocols combining different target inhibition specific to each AI pathways.     

Vulnerability of microcrustaceans to predation by the invasive filter-feeding mussel Limnoperna fortunei (Dunker)

Limnoperna fortunei is an Asian mussel introduced to South America around 1990. One of the most important impacts of this invader is probably its grazing on the plankton. In this study we evaluate the vulnerability of several planktonic microcrustaceans from the Paraná River floodplain to predation by adult L. fortunei. We conducted 2-h laboratory feeding experiments where the bivalves were offered microcrustaceans differing in overall body shape, size, and locomotive abilities. Ingestion and clearance rates for each taxon were estimated. Results suggest that, in addition to detritus and phytoplankton, microcrustaceans may be a very important food item for this invasive mollusc. Limnoperna fortunei can prey on larger organisms (up to 1100?µm) than Dreissena polymorpha, the European and North American invasive mussel.     

Analysis of vitelline envelope synthesis and composition during early oocyte development in gilthead seabream (Sparus aurata)

The oocyte vitelline envelope (VE) of gilthead seabream is composed of four known zona pellucida (ZP) proteins, ZPBa, ZPBb, ZPC, and ZPX. We have previously shown that the gilthead seabream ZP proteins are differentially transcribed in liver and ovary, with the expression in liver being under estrogenic control. However, although mRNA was found in both liver and ovary, only low ZPBa protein levels were detected in liver and plasma. Using isoform-specific ZP antibodies we show that ZPBa and ZPX translation products are present in the cytosol of stage I and II oocytes. In addition, the zpBa and zpX mRNAs were detected in early developing oocytes. During oocyte growth (vitellogenesis), the VE increased in thickness (>10 microm), and we show that the four ZP isoforms are present in different regions of the VE. ZPX was detected closest to the oocyte plasma membrane while the intermediate region was composed of ZPBa, ZPBb, and ZPC. At the outer layer, only ZPC was detected. When oocytes reach the fully grown stage they resume meiosis and hydration. As the oocyte expands, thinning to 4 microm, the VE acquire a striped and compact appearance at the electron microscopy level. This study provides further evidence for the oocyte origin of some ZP proteins in the gilthead seabream and suggests that the ZP proteins are differentially distributed within the VE.     

DNA diversification in two Sinorhizobium species

The comparative analysis of genomic characteristics and single-nucleotide polymorphism patterns from large fragments borne on different replicons of Sinorhizobium spp. genomes clearly demonstrate that DNA recombination among closely related bacteria is a major event in the diversification of this genome, especially in pSymA, resulting in mosaic structure.