Herança da Resistência de Lolium multiflorum ao Iodosulfuron-Methyl Sodium

Populações de azevém resistente aos inibidores da enzima ALS têm aumentado rapidamente nos campos cultivados. Para o manejo da resistência, são necessários estudos de herança da resistência, os quais permitem entender a evolução da resistência, a estrutura genética da população e a dinâmica de adaptação dos biótipos resistentes. Este trabalho teve como objetivo identificar o tipo de herança, o número de genes envolvidos e o grau de resistência dos biótipos de azevém, homozigotos e heterozigotos, resistentes ao iodosulfuron. A partir da seleção dos biótipos homozigoto resistente (R) e homozigoto suscetível (S), foram realizados cruzamentos (R x S) para obtenção de plantas F1, e estas, cruzadas para obtenção da F2, e realizaram-se retrocruzamentos entre plantas F1 e os respectivos genitores masculinos e femininos resistentes (RCr) e sensíveis (RCs). As sementes F1, F2, RCr, RCs e dos genitores foram semeadas em bandejas e avaliadas, com aplicação do iodosulfuron, quanto à sua suscetibilidade ou resistência. Plantas F1 e dos genitores foram tratadas com doses crescentes do herbicida. A avaliação de controle dessas plantas pelo iodosulfuron foi feito por meio de notas (0 a 100), referentes aos sintomas de intoxicação e pela massa da matéria seca da parte aérea acumulada. Os genitores masculino ou feminino transmitiram a característica para a prole, sendo esta 100% resistente, indicando gene de resistência dominante. A geração F2 apresentou segregação 3:1 resistente/suscetível, confirmando a característica de dominância. O teste de dominância das plantas F1 evidenciou que as plantas homozigotas resistentes e as heterozigotas apresentam grau de resistência semelhante. Conclui-se que a resistência do azevém ao iodosulfuron é codificada por gene dominante nuclear com dominância completa.

Large-scale production and purification of recombinant protein from an insect cell/baculovirus system in Erlenmeyer flasks: application to the chicken poly(ADP-ribose) polymerase catalytic domain

A simple and inexpensive shaker/Erlenmeyer flask system for large-scale cultivation of insect cells is described and compared to a commercial spinner system. On the basis of maximum cell density, average population doubling time and overproduction of recombinant protein, a better result was obtained with a simpler and less expensive bioreactor consisting of Erlenmeyer flasks and an ordinary shaker waterbath. Routinely, about 90 mg of pure poly(ADP-ribose) polymerase catalytic domain was obtained for a total of 3 x 109 infected cells in three liters of culture

Poly-DL-lactide-co-glycolide microspheres as a controlled release antigen delivery system

Successful vaccine application means maximum protection with minimal number of administrations. A rational development of vaccines involves studies of the nature of the antigen as well as of the adjuvant to be used to improve the immune responses. This has provided the impetus for studies to design the degradable devices and for different approaches to antigen delivery by different routes of administration. The development of controlled release systems based on polymeric devices that permit a sustained or pulsed release of encapsulated antigens has attracted much interest. Polymeric delivery systems consist of polymers that release their content continuously in a controlled manner over a period of time. The development of a biocompatible delivery system for parenteral administration offers several advantages in terms of immunoadjuvanticity over other compounds. It was found that, in contrast to other carriers, microspheres are more stable, thus permitting administration by the oral or parenteral route. In the present study, we describe the main characteristics and potentialities of this new immunoadjuvant for oral and parenteral administration.

Adhesion of the human pathogen Sporothrix schenckii to several extracellular matrix proteins

The pathogenic fungus Sporothrix schenckii is the causative agent of sporotrichosis. This subcutaneous mycosis may disseminate in immunocompromised individuals and also affect several internal organs and tissues, most commonly the bone, joints and lung. Since adhesion is the first step involved with the dissemination of pathogens in the host, we have studied the interaction between S. schenckii and several extracellular matrix (ECM) proteins. The binding of two morphological phases of S. schenckii, yeast cells and conidia, to immobilized type II collagen, laminin, fibronectin, fibrinogen and thrombospondin was investigated. Poly (2-hydroxyethyl methacrylate) (poly-HEMA) was used as the negative control. Cell adhesion was assessed by ELISA with a rabbit anti-S. schenckii antiserum. The results indicate that both morphological phases of this fungus can bind significantly to type II collagen, fibronectin and laminin in comparison to the binding observed with BSA (used as blocking agent). The adhesion rate observed with the ECM proteins (type II collagen, fibronectin and laminin) was statistically significant (P<0.05) when compared to the adhesion obtained with BSA. No significant binding of conidia was observed to either fibrinogen or thrombospondin, but yeast cells did bind to the fibrinogen. Our results indicate that S. schenckii can bind to fibronectin, laminin and type II collagen and also show differences in binding capacity according to the morphological form of the fungus.

Electrophysiological evidence for the presence of NR2C subunits of N-methyl-D-aspartate receptors in rat neurons of the nucleus tractus solitarius

The nucleus tractus solitarius (NTS) plays an important role in the control of autonomic reflex functions. Glutamate, acting on N-methyl-D-aspartate (NMDA) and non-NMDA ionotropic receptors, is the major neurotransmitter in this nucleus, and the relative contribution of each receptor to signal transmission is unclear. We have examined NMDA excitatory postsynaptic currents (NMDA-EPSCs) in the subpostremal NTS using the whole cell patch clamp technique on a transverse brainstem slice preparation. The NMDA-EPSCs were evoked by stimulation of the solitary tract over a range of membrane potentials. The NMDA-EPSCs, isolated pharmacologically, presented the characteristic outward rectification and were completely blocked by 50 µM DL-2-amino-5-phosphonopentanoic acid. The I-V relationship of the NMDA response shows that current, with a mean (± SEM) amplitude of -41.2 ± 5.5 pA, is present even at a holding potential of -60 mV, suggesting that the NMDA receptors are weakly blocked by extracellular Mg2+ at near resting membrane potentials. This weak block can also be inferred from the value of 0.67 ± 0.17 for parameter delta obtained from a fit of the Woodhull equation to the I-V relationship. The maximal inward current measured on the I-V relationship was at -38.7 ± 4.2 mV. The decay phase of the NMDA currents was fitted with one exponential function with a decay time constant of 239 ± 51 and 418 ± 80 ms at a holding potential of -60 and +50 mV, respectively, which became slower with depolarization (e-fold per 145 mV). The biophysical properties of the NMDA receptors observed in the present study suggest that these receptors in the NTS contain NR2C subunits and may contribute to the synaptic signal integration.

Use of blends of bioabsorbable poly(L-lactic acid)/poly(hydroxybutyrate- co-hydroxyvalerate) as surfaces for Vero cell culture

Vero cells, a cell line established from the kidney of the African green monkey (Cercopithecus aethiops), were cultured in F-10 Ham medium supplemented with 10% fetal calf serum at 37°C on membranes of poly(L-lactic acid) (PLLA), poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) and their blends in different proportions (100/0, 60/40, 50/50, 40/60, and 0/100). The present study evaluated morphology of cells grown on different polymeric substrates after 24 h of culture by scanning electron microscopy. Cell adhesion was also analyzed after 2 h of inoculation. For cell growth evaluation, the cells were maintained in culture for 48, 120, 240, and 360 h. For cytochemical study, the cells were cultured for 120 or 240 h, fixed, processed for histological analysis, and stained with Toluidine blue, pH 4.0, and Xylidine ponceau, pH 2.5. Our results showed that cell adhesion was better when 60/40 and 50/50 blends were used although cells were able to grow and proliferate on all blends tested. When using PLLA/PHBV (50/50) slightly flattened cells were observed on porous and smooth areas. PLLA/PHBV (40/60) blends presented flattened cells on smooth areas. PLLA/PHBV (0/100), which presented no pores, also supported spreading cells interconnected by thin filaments. Histological sections showed that cells grew as a confluent monolayer on different substrates. Cytochemical analysis showed basophilic cells, indicating a large amount of RNA and proteins. Hence, we detected changes in cell morphology induced by alterations in blend proportions. This suggests that the cells changed their differentiation pattern when on various PLLA/PHBV blend surfaces.

Prazosin blocks the glutamatergic effects of N-methyl-D-aspartic acid on lordosis behavior and luteinizing hormone secretion in the estrogen-primed female rat

We have observed that intracerebroventricular (icv) injection of selective N-methyl-D-aspartic acid (NMDA)-type glutamatergic receptor antagonists inhibits lordosis in ovariectomized (OVX), estrogen-primed rats receiving progesterone or luteinizing hormone-releasing hormone (LHRH). When NMDA was injected into OVX estrogen-primed rats, it induced a significant increase in lordosis. The interaction between LHRH and glutamate was previously explored by us and another groups. The noradrenergic systems have a functional role in the regulation of LHRH release. The purpose of the present study was to explore the interaction between glutamatergic and noradrenergic transmission. The action of prazosin, an alpha1- and alpha2b-noradrenergic antagonist, was studied here by injecting it icv (1.75 and 3.5 µg/6 µL) prior to NMDA administration (1 µg/2 µL) in OVX estrogen-primed Sprague-Dawley rats (240-270 g). Rats manually restrained were injected over a period of 2 min, and tested 1.5 h later. The enhancing effect induced by NMDA on the lordosis/mount ratio at high doses (67.06 ± 3.28, N = 28) when compared to saline controls (6 and 2 µL, 16.59 ± 3.20, N = 27) was abolished by prazosin administration (17.04 ± 5.52, N = 17, and 9.33 ± 3.21, N = 20, P < 0.001 for both doses). Plasma LH levels decreased significantly only with the higher dose of prazosin (1.99 ± 0.24 ng/mL, N = 18, compared to saline-NMDA effect, 5.96 ± 2.01 ng/mL, N = 13, P < 0.05). Behavioral effects seem to be more sensitive to the alpha-blockade than hormonal effects. These findings strongly suggest that the facilitatory effects of NMDA on both lordosis and LH secretion in this model are mediated by alpha-noradrenergic transmission.

Glutamate-N-methyl-D-aspartate receptor modulation and minocycline for the treatment of patients with schizophrenia: an update

Growing consistent evidence indicates that hypofunction of N-methyl-D-aspartate (NMDA) transmission plays a pivotal role in the neuropathophysiology of schizophrenia. Hence, drugs which modulate NMDA neurotransmission are promising approaches to the treatment of schizophrenia. The aim of this article is to review clinical trials with novel compounds acting on the NMDA receptor (NMDA-R). This review also includes a discussion and translation of neuroscience into schizophrenia therapeutics. Although the precise mechanism of action of minocycline in the brain remains unclear, there is evidence that it blocks the neurotoxicity of NMDA antagonists and may exert a differential effect on NMDA signaling pathways. We, therefore, hypothesize that the effects of minocycline on the brain may be partially modulated by the NMDA-R or related mechanisms. Thus, we have included a review of minocycline neuroscience. The search was performed in the PubMed, Web of Science, SciELO, and Lilacs databases. The results of glycine and D-cycloserine trials were conflicting regarding effectiveness on the negative and cognitive symptoms of schizophrenia. D-serine and D-alanine showed a potential effect on negative symptoms and on cognitive deficits. Sarcosine data indicated a considerable improvement as adjunctive therapy. Finally, minocycline add-on treatment appears to be effective on a broad range of psychopathology in patients with schizophrenia. The differential modulation of NMDA-R neurosystems, in particular synaptic versus extrasynaptic NMDA-R activation and specific subtypes of NMDA-R, may be the key mediators of neurogenesis and neuroprotection. Thus, psychotropics modulating NMDA-R neurotransmission may represent future monotherapy or add-on treatment strategies in the treatment of schizophrenia.

The methyl ester of rosuvastatin elicited an endothelium-independent and 3-hydroxy-3-methylglutaryl coenzyme A reductase-independent relaxant effect in rat aorta

The relaxant effect of the methyl ester of rosuvastatin was evaluated on aortic rings from male Wistar rats (250-300 g, 6 rats for each experimental group) with and without endothelium precontracted with 1.0 µM phenylephrine. The methyl ester presented a slightly greater potency than rosuvastatin in relaxing aortic rings, with log IC50 values of -6.88 and -6.07 M, respectively. Unlike rosuvastatin, the effect of its methyl ester was endothelium-independent. Pretreatment with 10 µM indomethacin did not inhibit, and pretreatment with 1 mM mevalonate only modestly inhibited the relaxant effect of the methyl ester. Nω-nitro-L-arginine methyl ester (L-NAME, 10 µM), the selective nitric oxide-2 (NO-2) inhibitor 1400 W (10 µM), tetraethylammonium (TEA, 10 mM), and cycloheximide (10 µM) partially inhibited the relaxant effect of the methyl ester on endothelium-denuded aortic rings. However, the combination of TEA plus either L-NAME or cycloheximide completely inhibited the relaxant effect. Inducible NO synthase (NOS-2) was only present in endothelium-denuded aortic rings, as demonstrated by immunoblot with methyl ester-treated rings. In conclusion, whereas rosuvastatin was associated with a relaxant effect dependent on endothelium and hydroxymethylglutaryl coenzyme A reductase in rat aorta, the methyl ester of rosuvastatin exhibited an endothelium-independent and only slightly hydroxymethylglutaryl coenzyme A reductase-dependent relaxant effect. Both NO produced by NOS-2 and K+ channels are involved in the relaxant effect of the methyl ester of rosuvastatin.

Poly(3-hydroxybutyrate-co-3-hydroxyhexanoate)-based scaffolds for tissue engineering

Development and selection of an ideal scaffold is of importance for tissue engineering. Poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) is a biocompatible bioresorbable copolymer that belongs to the polyhydroxyalkanoate family. Because of its good biocompatibility, PHBHHx has been widely used as a cell scaffold for tissue engineering. This review focuses on the utilization of PHBHHx-based scaffolds in tissue engineering. Advances in the preparation, modification, and application of PHBHHx scaffolds are discussed.

Phase transitions in biodegradable films based on blends of gelatin and poly (vinyl alcohol)

The aim of this work was to study the effect of the hydrolysis degree (HD) and the concentration (C PVA) of two types of poly (vinyl alcohol) (PVA) and the effect of the type and the concentration of plasticizers on the phase properties of biodegradable films based on blends of gelatin and PVA, using a response-surface methodology. The films were made by casting and the studied properties were their glass (Tg) and melting (Tm) transition temperatures, which were determined by diferential scanning calorimetry (DSC). For the data obtained on the first scan, the fitting of the linear model was statistically significant and predictive only for the second melting temperature. In this case, the most important effect on the second Tm of the first scan was due to the HD of the PVA. In relation to the second scan, the linear model could be fit to Tg data with only two statistically significant parameters. Both the PVA and plasticizer concentrations had an important effect on Tg. Concerning the second Tm of the second scan, the linear model was fit to data with two statistically significant parameters, namely the HD and the plasticizer concentration. But, the most important effect was provoked by the HD of the PVA.