Increased Receptor for Advanced Glycation Endproducts Immunocontent in the Cerebral Cortex of Vitamin A-Treated Rats

Vitamin A, beyond its biological role, is an alternative choice in treating some life threatening pathologies, for instance leukemia and immunodeficiency. On the other hand, vitamin A therapy at moderate to high doses has caused concern among public health researchers due to the toxicological aspect resulting from such habit. It has been described hepatotoxicity, cognitive disturbances and increased mortality rates among subjects ingesting increased levels of vitamin A daily. Then, based on the previously reported data, we investigated here receptor for advanced glycation endproducts (RAGE) immunocontent and oxidative damage levels in cerebral cortex of vitamin A-treated rats at clinical doses (1,000–9,000 IU/kg day⁻¹). RAGE immunocontent, as well as oxidative damage levels, were observed increased in cerebral cortex of vitamin A-treated rats. Whether increased RAGE levels exert negative effects during vitamin A supplementation it remains to be investigated, but it is very likely that deleterious consequences may arise from such alteration.     

SHORT‐TERM MEASUREMENT OF CARBON STABLE ISOTOPE DISCRIMINATION IN PHOTOSYNTHESIS AND RESPIRATION BY AQUATIC MACROPHYTES,WITH MARINE MACROALGAL EXAMPLES1

Progress in the study of stable isotope discrimination in carbon assimilation by aquatic macrophytes has been slower than for other groups of primary producers, such as phytoplankton and terrestrial plants. A probable reason has been the methodologies employed for such a study: field collections or long‐term incubations, both relying on the observation of changes in carbon isotope composition of plant tissue. Here, we present a short‐term incubation method based on the change in carbon stable isotope composition in water. Its fundamental advantage over the other approaches is that the change in stable isotope composition in water in a closed system is much faster than in the plant tissue. We applied the method to investigate the relationship between carbon assimilation intensity and isotope discrimination. The results included a relatively small discrimination in respiration, a significant influence of carbon assimilation rate on discrimination, and the suggestion of HCO₃^? or CO₂ uptake in photosynthesis. The information gathered using this method would be difficult to obtain in other ways, and so we believe that it should contribute to a better understanding of the physiology and ecology of aquatic macrophytes.     

Inferring a weighted elastic network from partial unfolding with coarse‐grained simulations

A number of studies have demonstrated that simple elastic network models can reproduce experimental B‐factors, providing insights into the structure–function properties of proteins. Here, we report a study on how to improve an elastic network model and explore its performance by predicting the experimental B‐factors. Elastic network models are built on the experimental coordinates, and they only take the pairs of atoms within a given cutoff distance r_c into account. These models describe the interactions by elastic springs with the same force constant. We have developed a method based on numerical simulations with a simple coarse‐grained force field, to attribute weights to these spring constants. This method considers the time that two atoms remain connected in the network during partial unfolding, establishing a means of measuring the strength of each link. We examined two different coarse‐grained force fields and explored the computation of these weights by unfolding the native structures. Proteins 2014; 82:119–129. © 2013 Wiley Periodicals, Inc.     

In Vitro Trypanocidal Activity of Macela (Achyrocline satureioides) Extracts against Trypanosoma evansi

The aim of this study was to verify the trypanocidal effectiveness of aqueous, methanolic, and ethanolic extracts of Achyrocline satureioides against Trypanosoma evansi in vitro. A. satureioides extracts, known as macela, were used on trypomastigotes at different concentrations (1, 5, 10, 50, 100, 500, and 1,000 µg/ml) and exposure times (0, 1, 3, 6, and 9 hr). A dose-dependent effect was observed when the 3 extracts were tested. The concentrations of 1, 5, and 10 µg/ml were not able to kill trypomastigotes until 3 hr after exposure, and the highest concentrations (500 and 1,000 µg/ml) were able to kill all trypomastigotes after 1 hr. When the time of exposure was increased up to 9 hr, the concentrations at 50 and 100 µg/ml were 100% effective to 3 extracts. The chemical analysis of the extracts revealed the presence of flavonoids, a trypanocidal compound already described. Based on the results, we can conclude that the A. satureioides extracts exhibit trypanocidal effects.     

Methotrexate-Related Response on Human Peripheral Blood Mononuclear Cells May Be Modulated by the Ala16Val-SOD2 Gene Polymorphism

Methotrexate (MTX) is a folic acid antagonist used in high doses as an anti-cancer treatment and in low doses for the treatment of some autoimmune diseases. MTX use has been linked to oxidative imbalance, which may cause multi-organ toxicities that can be attenuated by antioxidant supplementation. Despite the oxidative effect of MTX, the influence of antioxidant gene polymorphisms on MTX toxicity is not well studied. Therefore, we analyzed here whether a genetic imbalance of the manganese-dependent superoxide dismutase (SOD2) gene could have some impact on the MTX cytotoxic response. An in vitro study using human peripheral blood mononuclear cells (PBMCs) obtained from carriers with different Ala16Val-SOD2 genotypes (AA, VV and AV) was carried out, and the effect on cell viability and proliferation was analyzed, as well as the effect on oxidative, inflammatory and apoptotic markers. AA-PBMCs that present higher SOD2 efficiencies were more resistance to high MTX doses (10 and 100 µM) than were the VV and AV genotypes. Both lipoperoxidation and ROS levels increased significantly in PBMCs exposed to MTX independent of Ala16Val-SOD2 genotypes, whereas increased protein carbonylation was observed only in PBMCs from V allele carriers. The AA-PBMCs exposed to MTX showed decreasing SOD2 activity, but a concomitant up regulation of the SOD2 gene was observed. A significant increase in glutathione peroxidase (GPX) levels was observed in all PBMCs exposed to MTX. However, this effect was more intense in AA-PBMCs. Caspase-8 and -3 levels were increased in cells exposed to MTX, but the modulation of these genes, as well as that of the Bax and Bcl-2 genes involved in the apoptosis pathway, presented a modulation that was dependent on the SOD2 genotype. MTX at a concentration of 10 µM also increased inflammatory cytokines (IL-1β, IL-6, TNFα and Igγ) and decreased the level of IL-10 anti-inflammatory cytokine, independent of SOD2 genetic background. The results suggest that potential pharmacogenetic effect on the cytotoxic response to MTX due differential redox status of cells carriers different SOD2 genotypes.     

An efficient Agrobacterium tumefaciens-mediated transformation method for Simplicillium subtropicum (Hypocreales: Cordycipitaceae)

Filamentous fungi are the organisms of choice for most industrial biotechnology. Some species can produce a variety of secondary metabolites and enzymes of commercial interest, and the production of valuable molecules has been enhanced through different molecular tools. Methods for genetic manipulation and transformation have been essential for the optimization of these organisms. The genus Simplicillium has attracted increased attention given several potential biotechnological applications. The Simplicillium genus harbors several entomopathogenic species and some isolates have been explored for bioremediation of heavy metal contaminants. Furthermore, the myriad of secondary metabolites isolated from Simplicillium spp. render these organisms as ideal targets for deep exploration and further biotechnological mining possibilities. However, the lack of molecular tools hampered the exploration of this genus. Thus, an Agrobacterium tumefaciens-mediated transformation method was established for Simplicillium subtropicum, employing the far-red fluorescent protein TURBOFP635/Katushka, as a visual marker, and the selection marker SUR gene, that confers resistance to chlorimuron ethyl. Notably, one round of transformation using the established method yielded almost 400 chlorimuron resistant isolates. Furthermore, these transformants displayed mitotic stability for, at least, five generations. We anticipate that this method can be useful for deep molecular exploration and improvement of strains in the Simplicillium genus.     

Co-stimulation of PAFR and CD36 is required for oxLDL-induced human macrophages activation

The oxidative process of LDL particles generates molecules which are structurally similar to platelet-activating factor (PAF), and some effects of oxidized LDL (oxLDL) have been shown to be dependent on PAF receptor (PAFR) activation. In a previous study, we showed that PAFR is required for upregulation of CD36 and oxLDL uptake. In the present study we analyzed the molecular mechanisms activated by oxLDL in human macrophages and the contribution of PAFR to this response. Human adherent monocytes/macrophages were stimulated with oxLDL. Uptake of oxLDL and CD36 expression were determined by flow cytometry; MAP kinases and Akt phosphorylation by Western blot; IL-8 and MCP-1 concentration by ELISA and mRNA expression by real-time PCR. To investigate the participation of the PI3K/Akt pathway, Gαi-coupled protein or PAFR, macrophages were treated with LY294002, pertussis toxin or with the PAFR antagonists WEB2170 and CV3988, respectively before addition of oxLDL. It was found that the addition of oxLDL to human monocytes/macrophages activates the PI3K/Akt pathway which in turn activates the MAPK (p38 and JNK). Phosphorylation of Akt requires the engagement of PAFR and a Gαi-coupled protein. The upregulation of CD36 protein and the uptake of oxLDL as well as the IL-8 production are dependent on PI3K/Akt pathway activation. The increased CD36 protein expression is dependent on PAFR and Gαi-coupled protein. Transfection studies using HEK 293t cells showed that oxLDL uptake occurs with either PAFR or CD36, but IL-8 production requires the co-transfection of both PAFR and CD36. These findings show that PAFR has a pivotal role in macrophages response to oxLDL and suggest that pharmacological intervention at the level of PAFR activation might be beneficial in atherosclerosis.     

Nutritional Status of the Cauliflower Cultivar ‘Verona’ Grown with Omission of out Added Macronutrients

Knowledge of plant nutritional status allows an understanding of the physiological responses of plants to crop fertilization. A hydroponic experiment evaluated the symptoms of macronutrient deficiency in cauliflower ‘Verona’ and determined: a) the macronutrient contents of foliar tissues when visual symptoms were observed, b) macronutrients content of foliar and inflorescence tissues at harvest. The effect of nutrient deficiency on inflorescence mass was also evaluated. Nitrogen deficiency caused chlorosis followed by purple color in the old leaves, while P deficiency caused only chlorosis in old leaves. Chlorosis at the edge of old leaves progressing to the center of the leaves was observed with the omission of K, and after was observed necrosis in the chlorotic areas. Ca deficiency caused tip burn in new leaves, while Mg deficiency caused internerval chlorosis in old leaves. The omission of each macronutrient reduced inflorescence dry matter. This deleterious effect was larger for N, P, and K deficiencies, reducing inflorescence dry matter by 87, 49, and 42%, respectively. When the nutrient solutions without N, P, K, Ca, or Mg were supplied to cauliflower plants, the macronutrient contents at harvest were 8.8, 0.6, 3.5, 13.0, and 0.8 g kg^-1 in the foliar tissues and 27.3, 2.2, 21.6, 1.1, and 0.7 g kg^-1 in the inflorescence tissues, respectively.     

Meiotic irregularities in diploid and tetraploid plants of <Emphasis Type="Italic">Lolium multiflorum</Emphasis> Lam.

The genus Lolium comprises several species of economical importance in temperate countries, mostly due to their high nutritional value and adaptability to cold environments, including southern regions of Brazil. Recently, several diploid cultivars and populations, as well as synthetic tetraploid cultivars have been explored. In order to viabilize or to direct crossings, it is important that the accessions present regular meiosis, thus, producing viable pollen grains. In this way, this study aimed at investigating the meiosis of nine accessions of Lolium multiflorum used in breeding programs, seeking to evaluate its viability in future crossings. The meiosis of diploid plants (2n?=?2×?=?14) is more regular than the artificially tetraploidized genotypes (2n?=?4×?=?28). In the tetraploids, univalent, bivalent, and multivalent configurations were observed. The irregularities were associated to mixoploidy, irregular segregation of chromosomes, spindle disorders, asynchrony, micronuclei, and cellular fusion and formation of syncyte. The abnormalities affected the meiotic index of tetraploid cultivars, characterizing them as unstable. Nevertheless, both diploid and tetraploid genotypes are considered able to participate in crossings because, although there are abnormalities, they do not occur at levels that could affect the production of viable pollen grains.