CD105 Expression on CD34-Negative Spindle-Shaped Stromal Cells of Primary Tumor Is an Unfavorable Prognostic Marker in Early Breast Cancer Patients

Several studies have confirmed that the breast tumor microenvironment drives cancer progression and metastatic development. The aim of our research was to investigate the prognostic significance of the breast tumor microenvironment in untreated early breast cancer patients. Therefore, we analyzed the association of the expression of α-SMA, FSP, CD105 and CD146 in CD34-negative spindle-shaped stromal cells, not associated with the vasculature, in primary breast tumors with classical prognostic marker levels, metastatic recurrence, local relapse, disease-free survival, metastasis-free survival and the overall survival of patients. In the same way, we evaluated the association of the amount of intra-tumor stroma, fibroblasts, collagen deposition, lymphocytic infiltration and myxoid changes in these samples with the clinical-pathological data previously described. This study is the first to demonstrate the high CD105 expression in this stromal cell type as a possible independent marker of unfavorable prognosis in early breast cancer patients. Our study suggests that this new finding can be useful prognostic marker in the clinical-pathological routine.     

Intrapericardial Administration of Mesenchymal Stem Cells in a Large Animal Model: A Bio-Distribution Analysis

The appropriate administration route for cardiovascular cell therapy is essential to ensure the viability, proliferative potential, homing capacity and implantation of transferred cells. At the present, the intrapericardial administration of pharmacological agents is considered an efficient method for the treatment of cardiovascular diseases. However, only a few reports have addressed the question whether the intrapericardial delivery of Mesenchymal Stem Cells (MSCs) could be an optimal administration route. This work firstly aimed to analyze the pericardial fluid as a cell-delivery vehicle. Moreover, the in vivo biodistribution pattern of intrapericardially administered MSCs was evaluated in a clinically relevant large animal model. Our in vitro results firstly showed that, MSCs viability, proliferative behavior and phenotypic profile were unaffected by exposure to pericardial fluid. Secondly, in vivo cell tracking by magnetic resonance imaging, histological examination and Y-chromosome amplification clearly demonstrated the presence of MSCs in pericardium, ventricles (left and right) and atrium (left and right) when MSCs were administered into the pericardial space. In conclusion, here we demonstrate that pericardial fluid is a suitable vehicle for MSCs and intrapericardial route provides an optimal retention and implantation of MSCs.     

A system for the directed evolution of the insecticidal protein from Bacillus thuringiensis

Theoretically, the activity of AB-type toxin molecules such as the insecticidal toxin (Cry toxin) from B. thuringiensis, which have one active site and two binding site, is improved in parallel with the binding affinity to its receptor. In this experiment, we tried to devise a method for the directed evolution of Cry toxins to increase the binding affinity to the insect receptor. Using a commercial T7 phage-display system, we expressed Cry1Aa toxin on the phage surface as fusions with the capsid protein 10B. These recombinant phages bound to a cadherin-like protein that is one of the Cry1Aa toxin receptors in the model target insect Bombyx mori. The apparent affinity of Cry1Aa-expressing phage for the receptor was higher than that of Cry1Ab-expressing phage. Phages expressing Cry1Aa were isolated from a mixed suspension of phages expressing Cry1Ab and concentrated by up to 130,000-fold. Finally, random mutations were made in amino acid residues 369–375 in domain 2 of Cry1Aa toxin, the mutant toxins were expressed on phages, and the resulting phage library was screened with cadherin-like protein-coated beads. As a result, phages expressing abnormal or low-affinity mutant toxins were excluded, and phages with high-affinity mutant toxins were selected. These results indicate that a method combining T7 phage display with selection using cadherin-like protein-coated magnetic beads can be used to increase the activity of easily obtained, low-activity Cry toxins from bacteria.     

Trace metal contents in water and sediments in Samborombón Bay wetland,Argentina

Samborombón Bay is the most extensive myxohaline wetland of Argentina. This Ramsar site is considered a priority area for the conservation of biodiversity in the country. Numerous rivers and channels that drain the Pampean plain, such as the Salado River and the channels Lower Salado, 15, 9, A, 1, and 2, cross this wetland and flow into Samborombón Bay. Trace metal concentration (As, Cd, Cr, Cu, Mn, Ni, Pb, and Zn) in water and sediments were determined in low and high water periods. In the first case, contents were higher than the Guide Levels for the Protection of the Aquatic Biota (GLPAB) in most of the sampling stations. In high water period, only As and Zn were detected. Relevant physico-chemical parameters were analyzed by a principal component analysis (PCA). Trace metal concentrations in sediments suggest, as a whole, that this wetland has been so far exposed to low to moderate levels of anthropic influence. It must be emphasized however, that the presence of metals in both the water and sediments of this bay might have negative effects on the biota and on the regional trophic web.     

Phosphorous amount in floating and rooted macrophytes growing in wetlands from the Middle Paraná River floodplain (Argentina)

Biomass, P concentration, P amount and chlorophyll in three floating and three rooted macrophytes growing in wetlands of the Middle Paraná River floodplain measured in winter and summer were compared. Macrophytes were sampled three times in summer and twice in winter, in the period 2002/2004. Although Pistia stratiotes was the species with the highest P concentration in leaves and roots, Typha domingensis, Eichhornia crassipes and Pontederia cordata were the most efficient species in P retention in natural wetlands because of their higher biomass. Total P amount in rooted species did not show seasonal variations. However, T. domingensis accumulated a greater P amount in its aerial part in summer, whereas in winter it did so in its below-ground parts, indicating an important P dynamic regarding translocation within the plant. In summer, floating species were able to accumulate great quantities of P in a short period due to their high growth rate. In order to optimize and maintain the efficiency of constructed wetlands for P removal throughout the year, a selection of floating and rooted species should be used.     

Biochemical and pharmacological characterization of toxins obtained from the fire coral Millepora complanata

Millepora complanata is a normal resident of coral reefs in the Mexican Caribbean. In this study, we describe for the first time the vasoconstrictor, phospholipase A2 (PLA2), and hemolytic activities elicited by a crude extract obtained from M. complanata. This extract caused a concentration-dependent contraction of isolated rat aortic rings (EC50=22.4+/-1.1 microg protein/mL). This effect was endothelium independent and significantly reduced in the absence of extracellular Ca2+ and when the intracellular Ca2+ stores were depleted. In addition, the crude extract obtained from M. complanata showed PLA2 activity (7.231+/-0.092 mmol min(-1) mg(-1)) and hemolysis of rat erythrocytes (HU50=1.64+/-1.04 mug protein/mL). The hemolysis increased in the presence of Ca2+ and decreased in the presence of cholesterol. Furthermore, this hemolysis was significantly reduced after incubation with an inhibitor of PLA2 enzymes. The hemolytic and vasoconstrictor effects were abolished after incubating the extract under denaturing conditions. Reverse phase chromatography of the M. complanata extract afforded 19 fractions (F1 to F19). F4 induced hemolysis and contained mainly a protein of 30 kDa, probably a PLA2 enzyme, while F8 and F11, containing mainly proteins of 15 and 20 kDa respectively, produced vasoconstrictor effects mediated by different mechanisms of action.     

Unfolding pathway of the dimeric and tetrameric forms of phosphofructokinase-2 from Escherichia coli

Escherichia coli phosphofructokinase-2 (Pfk-2) is an oligomeric enzyme characterized by two kinds of interfaces: a monomer-monomer interface, critical for enzymatic activity, and a dimer-dimer interface formed upon tetramerization due to allosteric binding of MgATP. In this work, Pfk-2 was denatured by guanidine hydrochloride (GdnHCl) and the impact of ligand binding on the unfolding pathway of the dimeric and the tertrameric forms of the enzyme was examined. The unligated dimeric form unfolds and dissociates from 0.15 to 0.8 M GdnHCl without the accumulation of native monomers, as indicated by circular dichroism and size exclusion chromatography measurements. However, a monomeric intermediate with an expanded volume and residual secondary structure accumulates above 0.8 M GdnHCl. The dimeric fructose-6-P-enzyme complex shows a shift in the simultaneous dissociation and unfolding process to elevated GdnHCl concentrations (from 0.8 to 1.4 M) together with the expulsion of the ligand detected by intrinsic fluorescence measurements. The unfolding pathway of the tetrameric MgATP-enzyme complex shows the accumulation of a tetrameric intermediate with altered fluorescence properties at about 0.4 M GdnHCl. Above this concentration a sharp transition from tetramers to monomers, without the accumulation of either compact dimers or monomers, was detected by light scattering measurements. Indeed, the most populated species was a partially unfolded monomer about 0.7 M GdnHCl. On the basis of these results, we suggest that the subunit contacts are critical for the maintenance of the overall structure of Pfk-2 and for the binding of ligands, explaining the reported importance of the dimeric state for enzymatic activity.     

Genomic evolution of the class Acidithiobacillia: deep-branching Proteobacteria living in extreme acidic conditions

Members of the genus Acidithiobacillus, now ranked within the class Acidithiobacillia, are model bacteria for the study of chemolithotrophic energy conversion under extreme conditions. Knowledge of the genomic and taxonomic diversity of Acidithiobacillia is still limited. Here, we present a systematic analysis of nearly 100 genomes from the class sampled from a wide range of habitats. Some of these genomes are new and others have been reclassified on the basis of advanced genomic analysis, thus defining 19 Acidithiobacillia lineages ranking at different taxonomic levels. This work provides the most comprehensive classification and pangenomic analysis of this deep-branching class of Proteobacteria to date. The phylogenomic framework obtained illuminates not only the evolutionary past of this lineage, but also the molecular evolution of relevant aerobic respiratory proteins, namely the cytochrome bo₃ ubiquinol oxidases.Subject terms: Bacterial evolution, Bacterial genomics, Phylogenetics     

Detección molecular de Toxoplasma gondii en carnes para consumo humano en Ibagué, Colombia

Introduction:

Toxoplasma gondii is a parasite with great zoonotic potential. It can infect a broad range of warm-blooded hosts (including livestock) and causes significant losses in the industry. In humans, it has been described as a pathogen in immunosuppressed people, it affects the fetus development in congenital infections, and is associated with various behavioral disorders in healthy people. Humans can acquire T. gondii by consuming undercooked, contaminated meat.

Objective:

To determine T. gondii positivity (currently unknown) in meat for human consumption (i.e., beef, chicken, and pork) in the city of Ibague, Colombia.

Materials and methods:

We used conventional nested PCR and the T. gondii B1 gene sequence as amplification target. We collected samples of meat (N=186) sold in the urban area of Ibagué (62 beef, 62 chicken, and 62 pork samples) and determined the T. gondii positivity percentage for each type of meat.

Results:

The study found an average of 18.8% positivity for all meat samples, pork having the highest percentage (22.5%; 14/62), followed by beef (19.3%; 12/62) and chicken (14.5%; 9/62). The best-amplified products were sequenced by macrogen and aligned with the B1 gene sequences in GenBank, thereby confirming their identity.

Conclusions:

This is the first study of T. gondii prevalence in meat for human consumption carried out in the city of Ibagué and the department of Tolima. All three types of meat sampled represent a risk for local human infection.