Inhibition of estrogen-responsive gene activation by the retinoid X receptor beta: evidence for multiple inhibitory pathways.

The retinoid X receptor beta (RXR beta; H-2RIIBP) forms heterodimers with various nuclear hormone receptors and binds multiple hormone response elements, including the estrogen response element (ERE). In this report, we show that endogenous RXR beta contributes to ERE binding activity in nuclear extracts of the human breast cancer cell line MCF-7. To define a possible regulatory role of RXR beta regarding estrogen-responsive transcription in breast cancer cells, RXR beta and a reporter gene driven by the vitellogenin A2 ERE were transfected into estrogen-treated MCF-7 cells. RXR beta inhibited ERE-driven reporter activity in a dose-dependent and element-specific fashion. This inhibition occurred in the absence of the RXR ligand 9-cis retinoic acid. The RXR beta-induced inhibition was specific for estrogen receptor (ER)-mediated ERE activation because inhibition was observed in ER-negative MDA-MB-231 cells only following transfection of the estrogen-activated ER. No inhibition of the basal reporter activity was observed. The inhibition was not caused by simple competition of RXR beta with the ER for ERE binding, since deletion mutants retaining DNA binding activity but lacking the N-terminal or C-terminal domain failed to inhibit reporter activity. In addition, cross-linking studies indicated the presence of an auxiliary nuclear factor present in MCF-7 cells that contributed to RXR beta binding of the ERE. Studies using known heterodimerization partners of RXR beta confirmed that RXR beta/triiodothyronine receptor alpha heterodimers avidly bind the ERE but revealed the existence of another triiodothyronine-independent pathway of ERE inhibition. These results indicate that estrogen-responsive genes may be negatively regulated by RXR beta through two distinct pathways.

Radiotherapy suppresses angiogenesis in mice through TGF-betaRI/ALK5-dependent inhibition of endothelial cell sprouting.

BACKGROUND: Radiotherapy is widely used to treat cancer. While rapidly dividing cancer cells are naturally considered the main target of radiotherapy, emerging evidence indicates that radiotherapy also affects endothelial cell functions, and possibly also their angiogenic capacity. In spite of its clinical relevance, such putative anti-angiogenic effect of radiotherapy has not been thoroughly characterized. We have investigated the effect of ionizing radiation on angiogenesis using in vivo, ex vivo and in vitro experimental models in combination with genetic and pharmacological interventions. PRINCIPAL FINDINGS: Here we show that high doses ionizing radiation locally suppressed VEGF- and FGF-2-induced Matrigel plug angiogenesis in mice in vivo and prevented endothelial cell sprouting from mouse aortic rings following in vivo or ex vivo irradiation. Quiescent human endothelial cells exposed to ionizing radiation in vitro resisted apoptosis, demonstrated reduced sprouting, migration and proliferation capacities, showed enhanced adhesion to matrix proteins, and underwent premature senescence. Irradiation induced the expression of P53 and P21 proteins in endothelial cells, but p53 or p21 deficiency and P21 silencing did not prevent radiation-induced inhibition of sprouting or proliferation. Radiation induced Smad-2 phosphorylation in skin in vivo and in endothelial cells in vitro. Inhibition of the TGF-beta type I receptor ALK5 rescued deficient endothelial cell sprouting and migration but not proliferation in vitro and restored defective Matrigel plug angiogenesis in irradiated mice in vivo. ALK5 inhibition, however, did not rescue deficient proliferation. Notch signaling, known to hinder angiogenesis, was activated by radiation but its inhibition, alone or in combination with ALK5 inhibition, did not rescue suppressed proliferation. CONCLUSIONS: These results demonstrate that irradiation of quiescent endothelial cells suppresses subsequent angiogenesis and that ALK5 is a critical mediator of this suppression. These results extend our understanding of radiotherapy-induced endothelial dysfunctions, relevant to both therapeutic and unwanted effects of radiotherapy.

Properties of nitrogen doped silicon films deposited by low-pressure chemical vapor deposition from silane and ammonia

Nitrogen doped silicon (NIDOS) films have been deposited by low-pressure chemical vapor deposition from silane SiH4 and ammonia NH3 at high temperature (750°C) and the influences of the NH3/SiH4 gas ratio on the films deposition rate, refractive index, stoichiometry, microstructure, electrical conductivity, and thermomechanical stress are studied. The chemical species derived from silylene SiH2 into the gaseous phase are shown to be responsible for the deposition of NIDOS and/or (silicon rich) silicon nitride. The competition between these two deposition phenomena leads finally to very high deposition rates (100 nm/min) for low NH3/SiH4 gas ratio (R¿0.1). Moreover, complex variations of NIDOS film properties are evidenced and related to the dual behavior of the nitrogen atom into silicon, either n-type substitutional impurity or insulative intersticial impurity, according to the Si¿N atomic bound. Finally, the use of NIDOS deposition for the realization of microelectromechanical systems is investigated.

Ammonia volatilization of urea in the out-of-season corn

The aim of this study was to evaluate the N losses due to volatilization at different rates of common urea, polymer coated urea and urease inhibitor-treated urea in the out-of-season corn, using semi-open static collectors. The treatments consisted of N levels on side-dressing fertilization with urea in different treatments: (a) control (without N), (b) urea 40 kg ha-1 N, (c) urea 80 kg ha-1 N, (d) polymer coated urea 40 kg ha-1 N, (e) polymer coated urea 80 kg ha-1 N and (f) urea with the urease inhibitor (UI) N 80 kg ha-1 N. The results showed that the treatments with polymer coated urea and with urease inhibitor-treated urea reduced the volatilization of N around 50 % compared to common urea, either in the first and the second N side-dressing fertilizations. Thus, they demonstrate that the polymer coat and the urease inhibitors were effective in reducing the volatilization of urea N applied in coverage, which resulted in higher productivity. There was also increasing urease activity in the treatments with application of common urea.

Dexamethasone stimulates the biochemical differentiation of fetal forebrain cells in reaggregating cultures.

The influence of dexamethasone on the development of neurons and oligodendrocytes was studied in serum-free, aggregating rat brain cell cultures. Synaptogenesis and myelination occur in this culture system. The concentration of myelin basic protein and the activity of 2',3'-cyclic nucleotide 3'-phosphodiesterase were used as oligodendroglia and myelin markers. Choline acetyltransferase and acetylcholinesterase served as neuronal markers, glutamine synthetase reflected astrocyte differentiation, while ornithine decarboxylase served as a general marker for cell growth and maturation. This study showed that dexamethasone stimulated the differentiation of cholinergic neurons and astrocytes. The effect of dexamethasone on oligodendroglial differentiation and myelination depended on the stage of development: during the early phase of myelination dexamethasone had a stimulatory effect, whereas at a later stage it showed a significant inhibition.

Hepatitis C virus-mediated mitochondrial dysfunctions are prevented and rescued by cyclophilin inhibition

Alisporivir (Debio-025) is an analogue of cyclosporine A andrepresents the prototype of a new class of non-immunosuppressivecyclophilin inhibitors. In vitro and in vivo studies have shownthat alisporivir inhibits hepatitis C virus (HCV) replication andongoing clinical trials are exploring its therapeutic potential inpatients with chronic hepatitis C. Recent data suggest that theantiviral effect is mediated by inhibition of cyclophilin A whichis an essential host factor in the HCV life cycle. However, alisporiviralso inhibits mitochondrial permeability transition by bindingto cyclophilin D. As HCV is known to affect mitochondrialfunction, we explored the effect of alisporivir on HCV proteinmediatedmitochondrial dysfunction. By the use of inducible celllines, which allow to investigate the effects of HCV polyproteinexpression independent from viral RNA replication and whichrecapitulate the major alterations of mitochondrial bioenergeticsobserved in infectious cell systems, we show that alisporivir preventsHCV protein-mediated cytochrome c redistribution,decrease of cell respiration, collapse of mitochondrial membranepotential, overproduction of reactive oxygen species and mitochondrialcalcium overload. Strikingly, some of the HCV-mediatedmitochondrial dysfunctions could even be rescued byalisporivir. These observations provide new insights into thepathogenesis of HCV-related liver disease and reveal an additionalmechanism of action of alisporivir that is likely beneficialin the treatment of chronic hepatitis C.

Ammonia volatilization in no-till system in the south-central region of the State of Paraná, Brazil

Ammonia (NH3) volatilization can reduce the efficiency of urea applied to the surface of no-till (NT) soils. Thus, the objectives of this study were to evaluate the magnitude of NH3 losses from surface-applied urea and to determine if this loss justifies the urea incorporation in soil or its substitution for other N sources under the subtropical climatic conditions of South-Central region of Paraná State, Brazil. The experiment, performed over four harvesting seasons in a clayey Hapludox followed a randomized block design with four replicates. A single dose of N (150 kg ha-1) to V5 growth stage of corn cultivated under NT system was applied and seven treatments were evaluated, including surface-applied urea, ammonium sulfate, ammonium nitrate, urea with urease inhibitor, controlled-release N source, a liquid N source, incorporated urea, and a control treatment with no N application. Ammonia volatilization was evaluated for 20 days after N application using a semi-open static system. The average cumulative NH3 loss due to the superficial application of urea was low (12.5 % of the applied N) compared to the losses observed in warmer regions of Southeastern Brazil (greater than 50 %). The greatest NH3 losses were observed in dry years (up to 25.4 % of the applied N), and losses decreased exponentially as the amount of rainfall after N application increased. Incorporated urea and alternative N sources, with the exception of controlled-release N source, decreased NH3 volatilization in comparison with surface-applied urea. Urea incorporation is advantageous for the reduction of NH3 volatilization; however, other aspects as its low operating efficiency should be considered before this practice is adopted. In the South-Central region of Paraná, the low NH3 losses from the surface-applied urea in NT system due to wet springs and mild temperatures do not justify its replacement for other N sources.

Ammonia losses estimated by an open collector from urea applied to sugarcane straw

The quantification of ammonia (NH3) losses from sugarcane straw fertilized with urea can be performed with collectors that recover the NH3 in acid-treated absorbers. Thus, the use of an open NH3 collector with a polytetrafluoroethylene (PTFE)-wrapped absorber is an interesting option since its cost is low, handling easy and microclimatic conditions irrelevant. The aim of this study was to evaluate the efficiency of an open collector for quantifying NH3-N volatilized from urea applied over the sugarcane straw. The experiment was carried out in a sugarcane field located near Piracicaba, São Paulo, Brazil. The NH3-N losses were estimated using a semi-open static collector calibrated with 15N (reference method) and an open collector with an absorber wrapped in PTFE film. Urea was applied to the soil surface in treatments corresponding to rates of 50, 100, 150 and 200 kg ha-1 N. Applying urea-N fertilizer on sugarcane straw resulted in losses NH3-N up to 24 % of the applied rate. The amount of volatile NH3-N measured in the open and the semi-open static collector did not differ. The effectiveness of the collection system varied non-linearly, with an average value of 58.4 % for the range of 100 to 200 kg ha-1 of urea-N. The open collector showed significant potential for use; however, further research is needed to verify the suitability of the proposed method.

Inhibition of cell migration and invasion mediated by the TAT-RasGAP317-326 peptide requires the DLC1 tumor suppressor.

TAT-RasGAP317-326, a peptide corresponding to the 317-326 sequence of p120 RasGAP coupled with a cell-permeable TAT-derived peptide, sensitizes the death response of various tumor cells to several anticancer treatments. We now report that this peptide is also able to increase cell adherence, prevent cell migration and inhibit matrix invasion. This is accompanied by a marked modification of the actin cytoskeleton and focal adhesion redistribution. Interestingly, integrins and the small Rho GTP-binding protein, which are well-characterized proteins modulating actin fibers, adhesion and migration, do not appear to be required for the pro-adhesive properties of TAT-RasGAP317-326. In contrast, deleted in liver cancer-1, a tumor suppressor protein, the expression of which is often deregulated in cancer cells, was found to be required for TAT-RasGAP317-326 to promote cell adherence and inhibit migration. These results show that TAT-RasGAP317-326, besides its ability to favor tumor cell death, hampers cell migration and invasion.

Ligand-dependent inhibition of CD1d-restricted NKT cell development in mice transgenic for the activating receptor Ly49D.

In addition to their CD1d-restricted T cell receptor (TCR), natural killer T (NKT) cells express various receptors normally associated with NK cells thought to act, in part, as modulators of TCR signaling. Immunoreceptor-tyrosine activation (ITAM) and inhibition (ITIM) motifs associated with NK receptors may augment or attenuate perceived TCR signals respectively, potentially influencing NKT cell development and function. ITIM-containing Ly49 family receptors expressed by NKT cells are proposed to play a role in their development and function. We have produced mice transgenic for the ITAM-associated Ly49D and ITIM-containing Ly49A receptors and their common ligand H2-Dd to determine the importance of these signaling interplays in NKT cell development. Ly49D/H2-Dd transgenic mice had selectively and severely reduced numbers of thymic and peripheral NKT cells, whereas both ligand and Ly49D transgenics had normal numbers of NKT cells. CD1d tetramer staining revealed a blockade of NKT cell development at an early precursor stage. Coexpression of a Ly49A transgene partially rescued NKT cell development in Ly49D/H2-Dd transgenics, presumably due to attenuation of ITAM signaling. Thus, Ly49D-induced ITAM signaling is incompatible with the early development of cells expressing semi-invariant CD1d-restricted TCRs and appropriately harmonized ITIM-ITAM signaling is likely to play an important role in the developmental program of NKT cells.

Urea coated with oxidized charcoal reduces ammonia volatilization

Urea is the most consumed nitrogen fertilizer in the world. However, its agronomic and economic efficiency is reduced by the volatilization of NH3, which can reach 78 % of the applied nitrogen. The coating of urea granules with acidic compounds obtained by charcoal oxidation has the potential to reduce the volatilization, due to the acidic character, the high buffering capacity and CEC. This work aimed to evaluate the effect of HNO3-oxidized carbon on the control of NH3 volatilization. These compounds were obtained by oxidation of Eucalyptus grandis charcoal, produced at charring temperatures of 350 and 450 ºC, with 4.5 mol L-1 HNO3. The charcoal was oxidized by solubilization in acidic or alkaline medium, similar to the procedure of soil organic matter fractionation (CHox350 and CHox450). CHox was characterized by C, H, O, N contents and their respective atomic relations, by the ratio E4 (absorbance 465 nm) by E6 (absorbance 665 nm), and by active acidity and total acidity (CEC). The inhibitory effect of CHox on the urease activity of Canavalia ensiformis was assessed in vitro. The NH3 volatilization from urea was evaluated with and without coating of oxidized charcoal (U-CHox350 or U-CHox450) in a closed system with continuous air flow. The pH of both CHox was near 2.0, but the total acidity of CHox350 was higher, 72 % of which was attributed to carboxylic groups. The variation in the ionization constants of CHox350 was also greater. The low E4/E6 ratios characterize the high stability of the compounds in CHox. CHox did not inhibit the urease activity in vitro, although the maximum volatilization peak from U-CHox450 and U-CHox350 occurred 24 h after that observed for uncoated urea. The lowest volatilization rate was observed for U-CHox350 as well as a 43 % lower total amount of NH3 volatilized than from uncoated urea.

Loss of ammonia from nitrogen fertilizers applied to maize and soybean straw

In Brazilian agriculture, urea is the most commonly used nitrogen (N) source, in spite of having the disadvantage of losing considerable amounts of N by ammonia-N volatilization. The objectives of this study were to evaluate: N lossby ammonia volatilization from: [urea coated with copper sulfate and boric acid], [urea coated with zeolite], [urea+ammonium sulfate], [urea coated with copper sulfate and boric acid+ammonium sulfate], [common urea] and [ammonium nitrate]; and the effect of these N source son the maize yield in terms of amount and quality. The treatments were applied to the surface of a soil under no-tillage maize, in two growing seasons. The first season (2009/2010) was after a maize crop (maize straw left on the soil surface) and the second cycle (2012/2011) after a soybean crop. Due to the weather conditions during the experiments, the volatilization of ammonia-N was highest in the first four days after application of the N sources. Of all urea sources, under volatilization-favorable conditions, the loss of ammonia from urea coated with copper sulfate and boric acid was lowest, while under high rainfall, the losses from the different urea sources was similar, i.e., an adequate rainfall was favorablet o reduce volatilization. The ammonia volatilization losses were greatest in the first four days after application. Maize grain yield differed due to N application and in the treatments, but this was only observed with cultivation of maize crop residues in 2009/2010. The combination of ammonium+urea coated with copper sulfate and boric acid optimized grain yield compared to the other urea treatments. The crude protein concentration in maize was not influenced by the technologies of urea coating.

Ammonia volatilization from coated urea forms

Nitrogen fertilization is a major component of the cost of agricultural production, due to the high cost and low efficiency of fertilizers. In the case of urea, the low efficiency is mainly due to losses by volatilization, which are more pronounced in cultivation systems in which plant residues are left on the soil. The objective of this work was to compare the influence of urea coated with sulfur or boric acid and copper sulfate with conventional N fertilizers on N volatilization losses in sugar cane harvested after stubble burning. The sources urea, sulfur-coated urea, urea coated with boric acid and copper sulfate, as well as nitrate and ammonium sulfate, were tested at amounts containing N rates of 120 kg ha-1 N. The integration of new technologies in urea fertilization can reduce N losses by volatilization. These losses were most reduced when using nitrate and ammonium sulfate. The application of a readily acidified substance (boric acid) to urea was more efficient in reducing volatilization losses and nutrient removal by sugar cane than that of a substance with gradual acidification (elemental sulfur).

Evaluation of ammonia volatilization losses by adjusted parameters of a logistic function

The dynamics of N losses in fertilizer by ammonia volatilization is affected by several factors, making investigation of these dynamics more complex. Moreover, some features of the behavior of the variable can lead to deviation from normal distribution, making the main commonly adopted statistical strategies inadequate for data analysis. Thus, the purpose of this study was to evaluate the patterns of cumulative N losses from urea through ammonia volatilization in order to find a more adequate and detailed way of assessing the behavior of the variable. For that reason, changes in patterns of ammonia volatilization losses as a result of applying different combinations of two soil classes [Planossolo and Chernossolo (Typic Albaqualf and Vertic Argiaquolls)] and different rates of urea (50, 100 and 150 kg ha-1 N), in the presence or absence of a urease inhibitor, were evaluated, adopting a 2 × 3 × 2 factorial design with four replications. Univariate and multivariate analysis of variance were performed using the adjusted parameter values of a logistic function as a response variable. The results obtained from multivariate analysis indicated a prominent effect of the soil class factor on the set of parameters, indicating greater relevance of soil adsorption potential on ammonia volatilization losses. Univariate analysis showed that the parameters related to total N losses and rate of volatilization were more affected by soil class and the rate of urea applied. The urease inhibitor affected only the rate and inflection point parameters, decreasing the rate of losses and delaying the beginning of the process, but had no effect on total ammonia losses. Patterns of ammonia volatilization losses provide details on behavior of the variable, details which can be used to develop and adopt more accurate techniques for more efficient use of urea.