Local NO synthase inhibition produces histological and functional recovery in Achilles tendon of rats after tenotomy

Repair of injured tendon is a very slow process and involves the release of many molecules, including nitric oxide. We investigate the influence of local nitrergic inhibition in histological and functional recovery of injured Achilles tendon. A standard murine model of tendon injury by rupture was used. The animals were divided into three experimental groups: control, injury + vehicle (normal saline) and injury + Nω-nitro-L-arginine methyl ester (L-NAME). The products were injected into the paratendinous region every 2 days and body weight gain and Achilles functional index (AFI) were evaluated on days 0, 7, 14 and 21 after tendon injury. On day 21 post-injury, the animals were killed to evaluate nitric oxide production and tissue organization. We observed that tendon surgical division led to increased tissue nitrite levels, which were reduced in L-NAME-treated rats. The AFI revealed functional recovery of L-NAME-treated animals on day 21 post-injury, which was not observed in the saline-treated group. Microscopic analysis of hematoxylin-eosin staining and collagen autofluorescence showed that L-NAME-treated rats had more aligned areas of collagen fibers and that the diameter of newly organized collagen in this group was also greater than that in the vehicle-treated one. We demonstrate that local treatment with L-NAME significantly improves the functional parameters and accelerates histomorphological recovery.     

Crossing the threshold: an amphibian assemblage highly infected with Batrachochytrium dendrobatidis in the southern Brazilian Atlantic forest

The fungal infection caused by Batrachochytrium dendrobatidis (Bd) in amphibians is known to be lethal when infection intensity values exceed loads of 10,000 zoospores per individual. We investigated Bd infection intensity in 100 anurans of southern Brazil. Almost half of the individuals were infected and the intensity ranged from four to about 156,000 zoospore genomic equivalents. We found no clinical signs of chytridiomycosis and no evidence of mortality. However, we observed a reduction in the number of infected individuals with loads above 10,000 zoospores. This fact could be considered indirect evidence that individuals with high loads are removed from the population.     

Double-antigen sandwich ELISA based on chimeric antigens for detection of antibodies to Trypanosoma cruzi in human sera

BackgroundEnzyme-linked immunosorbent assays (ELISA) are generally the chosen test for Chagas disease (CD) diagnosis; however, its performance depends on the antigen preparation adsorbed to the solid phase, which may lead to false-positive results and cross-reactions. The use of chimeric recombinant antigens can overcome this limitation. Four chimeric antigens from Trypanosoma cruzi (IBMP-8.1, IBMP-8.2, IBMP-8.3 and IBMP-8.4) were developed and evaluated in phase I, II and III studies using indirect ELISA as diagnostic platform. However, peroxidase-labeled secondary anti-human IgG antibody, which is employed in indirect ELISAs, limits its use for the detection of species-specific and class-specific antibodies. To overcome this limitation, peroxidase-labeled antigens can be utilized, diagnosing both acute or chronic infection, in a species and immunoglobulin class-independent manner, through the use of a double-antigen sandwich ELISA (DAgS-ELISA). We aimed to evaluate and validate the diagnostic performance of the chimeric antigens IBMP-8.1, IBMP-8.2, IBMP-8.3 and IBMP-8.4 in the DAgS-ELISA platform.Methodology/Principal findingsDAgS-ELISA was optimized by checkerboard titration. In phase I study, 207 positive and 205 negative samples were evaluated. Cross-reactivity to other infections was also assessed using 68 samples. The selected conditions for the tests utilized 25 ng of antigen per well and the conjugate diluted at 1:2,000 for all molecules. In the phase I study, the areas under the curve of IBMP-8.1, IBMP-8.2, IBMP-8.3 and IBMP-8.4 were 98.7%, 99.5%, 98.6% and 98.8%, respectively. Among the positive samples, IBMP-8.1 antigen classified 53 (25.6%) as false negative, IBMP-8.2, 27 (13%), IBMP-8.3, 24 (11.6%) and IBMP-8.4, 43 (20.8%), giving sensitivities of 74.4%, 87%, 88.4% and 79.2%, respectively. The only antigen that did not reach 100% specificity was IBMP-8.3, with 96.6%. IBMP-8.3 was also the only molecule to show cross-reactivity with HTLV.Conclusions/SignificanceDAgS-ELISA is a promising tool for immunodiagnosis, and despite the high AUC values, the performance of this assay was different from the values obtained by our group when using these antigens in the indirect ELISA, for this reason, improvements are being considered to increase the sensitivity of the DAgS-ELISA.     

Detecting Multiple States of Trophic Connectivity Between Mangroves and Salt Marshes

The productivity gradient between adjacent habitats can fluctuate over time due to seasonal cycles and lead to both habitats being alternately subsidized. Although this process is well known for prey subsidies in stream-riparian forest ecotones, few studies are available for other systems or subsidy types. Moreover, the effects of transport intensity on this expected alternate subsidy exchange are still poorly understood. We assessed whether subsidy input and allochthonous carbon assimilation by resident benthic invertebrates alternated between adjacent mangroves and salt marshes during peaks of detritus productivity (summer and winter, respectively) in a subtropical estuary, by using detritus trapping techniques and stable isotope ratios. Sampling was performed simultaneously in the sheltered (inner sector) and exposed (outer sector) regions of the estuary to assess the influence of different physical conditions on the intensity of subsidy flow. Transport of mangrove litter into the salt marsh occurred mainly in the summer in both sectors; however, most of the litter remained trapped in the marsh boundary. The mixing model also showed that there was little influence of allochthonous carbon in the diet of salt marsh benthic invertebrates. Marsh litter supply to mangroves did not vary significantly between seasons but was significantly higher in the outer than in the inner sector. Likewise, the mixing model showed great contribution of salt marsh carbon to the diet of benthic invertebrates from the outer-sector mangroves, whereas autochthonous carbon predominated in those from the inner mangroves. Our findings reinforce the model that trophic connectivity relies on the relative proportion of allochthonous (subsidy) and autochthonous resources rather than only on asymmetric productivity between habitats. Differences in the proportion of resources result from interaction among productivity, permeability, and transport vectors that lead to many states of trophic connectivity.     

Apoplastic pH and inorganic ion levels in tomato fruit: A potential means for regulation of cell wall metabolism during ripening

Apoplastic pH and ionic conditions exert strong influence on cell wall metabolism of many plant tissues; however, the nature of the apoplastic environment of ripening fruit has been the subject of relatively few studies. In this report, a pressure-bomb technique was used to extract apoplastic fluid from tomato fruit ( Lycopersicon esculentum Mill.) pericarp at several developmental stages. pH and the levels of K⁺, Na⁺, Ca²⁺, Mg²⁺, Cl⁻ and P were determined and compared with the values for the bulk pericarp and locule tissues. The pH of the apoplastic fluid from pericarp tissue decreased from 6.7 in immature and mature-green fruits to 4.4 in fully-ripe fruit. During the same period, the K⁺ concentration increased from 13 to 37 m M . The levels of Na⁺ and divalent cations did not change, whereas the anions P and Cl⁻ increased in ripe fruit. Ca²⁺ levels remained relatively constant during ripening at 4–5 m M , concentrations that effectively limit pectin solubilization. The electrical conductivity of the apoplastic liquid increased 3-fold during ripening, whereas osmotically active solutes increased 2-fold. Pressure-treated fruit retained the capacity to ripen. The decline in apoplastic pH and increase in ionic strength during tomato fruit ripening may regulate the activity of cell wall hydrolases. The potential role of apoplastic changes in fruit ripening and softening is discussed.     

Cellular and molecular basis of decision‐making

People think they are in control of their own decisions: what to eat or drink, whom to marry or pick a fight with, where to live, what to buy. Behavioural economists and neurophysiologists have long studied decision‐making behaviours. However, these behaviours have only recently been studied through the light of molecular genetics. Here, we review recent research in mice, Drosophila melanogaster and Caenorhabditis elegans, that analyses the molecular and cellular mechanisms underlying decision‐making. These studies interrogate decision‐making about food, sexual behaviour, aggression or foraging strategies, and add molecular and cell biology understanding onto the consilience of brain and decision.     

A novel interleukin-17 receptor-like protein identified in human umbilical vein endothelial cells antagonizes basic fibroblast growth factor-induced signaling

We have previously utilized a combination of high throughput sequencing and genome-wide microarray profiling analyses to identify novel cell-surface proteins expressed in human umbilical vein endothelial cells. One gene identified by this approach encodes a type I transmembrane receptor that shares sequence homology with the intracellular domain of members of the interleukin-17 (IL-17) receptor family. Real-time quantitative PCR and Northern analyses revealed that this gene is highly expressed in human umbilical vein endothelial cells and in several highly vascularized tissues such as kidney, colon, skeletal muscle, heart, and small intestine. In addition, we also found that it is also highly expressed in the ductal epithelial cells of human salivary glands, seminal vesicles, and the collecting tubules of the kidney by in situ hybridization. This putative receptor, which we have termed human SEF (hSEF), is also expressed in a variety of breast cancer tissues. In co-immunoprecipitation assays, this receptor is capable of forming homomeric complexes and can interact with fibroblast growth factor (FGF) receptor 1. Overexpression of this receptor inhibits FGF induction of an FGF-responsive reporter gene in human 293T cells. This appears to occur as a result of specific inhibition of p42/p44 ERK in the absence of upstream MEK inhibition. This inhibitory effect is dependent upon a functional intracellular domain since deletion mutants missing the IL-17 receptor-like domain lack this inhibitory effect. These findings are consistent with the recent discovery of the zebrafish homologue, Sef (similar expression to fgf genes), which specifically antagonizes FGF signaling when ectopically expressed in zebrafish or Xenopus laevis embryos. Based on sequence and functional similarities, this novel IL-17 receptor homologue represents a potential human SEF and is likely to play critical roles in endothelial or epithelial functions such as proliferation, migration, and angiogenesis.     

Field evaluation of efficacy and persistence of an insect vacuum device against the tarnished plant bug (Hemiptera: Miridae) in a day-neutral strawberry field

The efficacy of a vacuuming device to reduce Lygus lineolaris (Palisot de Beauvois) populations was evaluated over 3 yr in a day-neutral strawberry field. Immediately after treatments, a significant reduction of population estimates (by tapping of flower clusters) was observed for adults (75% of the time) and nymphs (50% of the time). Under the same conditions, control (i.e., vacuum turbine off) treatments significantly reduced adult population estimates 25% of the time. Lygus lineolaris tapping samples taken from strawberry plants adjacent to the treated zone did not show significant variations over sampling time, suggesting that no escape behavior occurred. This was supported by adult catches on sticky traps located in zones adjacent to the treated one. Our results suggest that vacuuming had inconsistent effect on tarnished plant bug populations and that L. lineolaris mobility plays a marginal role in the use of this control method.     

Adhesion of flowing monocytes to hypoxia-reoxygenation-exposed endothelial cells: role of Rac1, ROS,and VCAM-1

Production ofreactive oxygen species (ROS) by ischemic tissue afterischemia-reperfusion (I/RP) is an important factor that contributes to tissue injury. The small GTPase Rac1 mediates the oxidative burst, and ROS act on signaling pathways involved in expression of inflammatory genes. Because there is evidence implicating monocytes in the pathogenesis of I/RP injury, our objective was todetermine the molecular mechanisms that regulate adhesive interactions between monocytes and hypoxia-reoxygenation (H/RO)-exposed cultured endothelial cells (ECs). When U937 cells were perfused over human umbilical vein ECs at 1 dyn/cm², H (1 h at 1%O₂)/RO (13 h) significantly increased the fluxes of rollingand stably adherent U937 cells. Either EC treatment with theantioxidant pyrrolidine dithiocarbamate (PDTC) or infection withAdRac1N17, which results in expression of the dominant-negative form ofRac1, abolished H/RO-induced ROS production, attenuated rolling, andabolished stable adhesion of U937 cells to H/RO-exposed ECs. Infectionwith AdRac1N17 also abolished H/RO-induced upregulation of vascularcell adhesion molecule (VCAM)-1. In turn, blocking VCAM-1 abolishedU937 cell stable adhesion and slightly increased rolling. We concludedthat the Rac1-dependent ROS partially regulate rolling and exclusivelyregulate stable adhesion of monocytic cells to ECs after H/RO and thatstable adhesion, but not rolling, is mediated by ROS-induced expressionof VCAM-1.