Concentration-dependent Way Research Articles

Comparison Antioxidant Properties Extract of Touyserkani Variety of Walnut Leave with two Extraction Methods (Traditional Method and Microwave Assisted Extraction)

The aim of this study is extraction of phenolic compounds from walnut leaves of touyserkani variety with two extraction methods (traditional method and with microwave assisted extraction (MAE)) and evaluation of their antioxidant activity. Total Phenolic contents and antioxidant properties of aqueous, methanol (80%) and ethanol (50%) extracts in different times was determined. At the next stage, effect of methanol extract on retarding of oxidation of soy oil was assessed. All of extracts showed antioxidant activity in a concentration-dependent way. In MAE, methanol extract had highest total phenol whose content is 89.15 ± 0.25 mg/g. the ethanol extract presented highest activity in radical-scavenging activity (EC50 = 27.90 µg/ml), reducing power assay (EC50 = 93.26 µg/ml) and antioxidant activity assay (EC50=68.23 µg/ml). Also extract at 1000 ppm could control oxidation of soy oil and this extract was effective than all of synthesis antioxidant in retarding oil oxidation Hence we can present walnut leaves as a potential source of phenolic compounds and natural antioxidant.

Identification of genes involved in the ACC-mediated control of root cell elongation in Arabidopsis thaliana

BackgroundAlong the root axis of Arabidopsis thaliana, cells pass through different developmental stages. In the apical meristem repeated cycles of division increase the numbers of cells. Upon leaving the meristem, these cells pass the transition zone where they are physiologically and mechanically prepared to undergo subsequent rapid elongation. During the process of elongation epidermal cells increase their length by 300% in a couple of hours. When elongation ceases, the cells acquire their final size, shape and functions (in the differentiation zone). Ethylene administered as its precursor 1-aminocyclopropane-1-carboxylic acid (ACC) is capable of inhibiting elongation in a concentration-dependent way. Using a microarray analysis, genes and/or processes involved in this elongation arrest are identified.ResultsUsing a CATMA-microarray analysis performed on control and 3h ACC-treated roots, 240 differentially expressed genes were identified. Quantitative Real-Time RT-PCR analysis of the 10 most up and down regulated genes combined with literature search confirmed the accurateness of the analysis. This revealed that inhibition of cell elongation is, at least partly, caused by restricting the events that under normal growth conditions initiate elongation and by increasing the processes that normally stop cellular elongation at the end of the elongation/onset of differentiation zone.ConclusionsACC interferes with cell elongation in the Arabidopsis thaliana roots by inhibiting cells from entering the elongation process and by immediately stimulating the formation of cross-links in cell wall components, diminishing the remaining elongation capacity. From the analysis of the differentially expressed genes, it becomes clear that many genes identified in this response, are also involved in several other kind of stress responses. This suggests that many responses originate from individual elicitors, but that somewhere in the downstream signaling cascade, these are converged to a ’common pathway’. Furthermore, several potential keyplayers, such as transcription factors and auxin-responsive genes, were identified by the microarray analysis. They await further analysis to reveal their exact role in the control of cell elongation.

Nicotine induces the expression of C-reactive protein via MAPK-dependent signal pathway in U937 macrophages.

Atherosclerosis is an inflammatory disease in the vessel wall. Nicotine, a major component of cigarette smoke, is an independent risk factor for cardiovascular diseases including atherosclerosis. As an inflammatory molecule, C- reactive protein (CRP) participates in atherogenesis. Although it has been confirmed that CRP level in smoking patient is significantly higher than non-smokers and cigarette withdrawal, it is unknown whether nicotine induces CRP expression in macrophages. The present study was to observe effect of nicotine on CRP production and the related signal pathway in U937 macrophages. The results showed that nicotine significantly increased mRNA and protein expression of CRP in U937 macrophages in time- and concentration-dependent ways. Nicotinic acetylcholine receptor (nAChR) blocker hexamethonium, MEK1/2 inhibitor PD98059, p38 MAPK inhibitor SB203580 and NF-κB inhibitor PDTC almost completely abolished nicotineinduced CRP expression in mRNA and protein levels in U937 macrophages. The further study indicated that hexamethonium, PD98059, and SB203580 significantly inhibited ERK1/2 and p38 MAPK phosphorylation. These demonstrate that nicotine has ability to induce CRP expression in macrophages through nAChR-ERK1/2/p38 MAPK-NF-κB signal pathway, which contributes to better understanding of the pro-inflammatory and pro-atherosclerotic effects of nicotine in cigarette smokers.

Alterations of fibrin network structure mediated by dermatan sulfate

Dermatan sulfate (DS) is well-known for its anticoagulant activity through binding to heparin cofactor II (HCII) to enhance thrombin inhibition. It has also been reported that DS has a profibrinolytic effect. We have evaluated the effects of DS solutions (4-20 μg/mL) on the formation (by kinetic studies), structure (by electron microscopy and compaction assays) and lysis (with urokinase-type plasminogen activator) of plasma fibrin networks. The results showed that DS significantly prolonged the lag phase and decreased the fibrin formation rate and the optical density of the final networks versus control, in a concentration dependent way. DS-associated networks presented a minor network percentage compared with control, composed of lower number of fibers per field, which resulted significantly thinner and longer. Moreover, DS rendered gels more sensible to rupture by centrifugal force and more susceptible to lysis. When fibrin formation kinetic assays were performed with purified fibrinogen instead of plasma, in the absence of HCII, the optical density of final DS-associated networks was statistically lower than control. Therefore, a direct effect of DS on the thickness of fibers was observed. Since in all in vitro assays low DS concentrations were used, it could be postulated that the fibrin features described above are plausible to be found in in vivo thrombi and therefore, DS would contribute to the formation of less thrombogenic clots.

In Vivo Antifungal Activities of the Methanol Extracts of Invasive Plant Species Against Plant Pathogenic Fungi

Plants are the promising reservoirs for natural products with their diverse secondary metabolites. Many invasive plants have been introduced in Korea, which adversely affect on the native ecosystem but holds difficulty removing them due to their proliferation. In this study, we evaluated disease control efficacy of methanol extracts from four invasive plant species against 7 representative crop pathogens. Methanol extract of Phytolacca americana effectively suppressed rice blast, tomato gray mold, and tomato late blight in a dose dependent manner. The methanol extract of Amorpha fruticosa also exhibited potent antifungal activity against pepper anthracnose in a concentration dependent way. These data suggest that the extracts of P. americana and A. fruticosa can be developed as plant disease protection agents against rice blast, tomato gray mold, tomato late blight, and pepper anthracnose. Furthermore, more extensive research will be required to identify and isolate active compounds from problematic invasive plant species to develop valuable agrochemicals.

Neuroprotective effect of α-mangostin and curcumin against iodoacetate-induced cell death

Curcumin is a phenolic yellow curry pigment with anti-inflammatory and antioxidant activities and α-mangostin is a xanthone isolated from mangosteen fruit with antioxidant properties. Iodoacetate (IAA) is an inhibitor of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase that induces a model of metabolic inhibition in neurons where reactive oxygen species (ROS) production is a significant mechanism. Furthermore, it has been shown that the induction of heme oxygenase-1 (HO-1) protects against IAA-induced neuronal death.ObjectivesTo study the effects of α-mangostin and curcumin against the IAA-induced cell death and on HO-1 expression in primary cultures of cerebellar granule neurons (CGNs).MethodsCGNs were treated with curcumin or α-mangostin before the addition of IAA. Cell viability and ROS production were measured 24 and 4 hours after IAA addition, respectively. HO-1 expression was measured by western blot.ResultsBoth α-mangostin and curcumin pretreatment ameliorated the neuronal death induced by IAA in a concentration-dependent way, which was associated with an amelioration of IAA-induced ROS formation. In addition, it was found that α-mangostin and curcumin induced HO-1 expression.DiscussionTreatment with α-mangostin and curcumin provided a neuroprotective effect against IAA in primary cultures of CGNs, an effect associated with an amelioration of the IAA-induced ROS production. HO-1 induced by these antioxidants may also be involved in the neuroprotective effect. Future work will be required to determine whether α-mangostin may cross the blood–brain barrier and achieve enough bioavailability to elicit a protective response in the brain being an effective nutraceutical compound for preventive therapy of neurodegenerative diseases.

Synergistic neurotoxicity of oxygen-glucose deprivation and tetrabromobisphenol A in vitro: role of oxidative stress

BackgroundTetrabromobisphenol A (TBBPA) is a toxic brominated flame retardant. Previous studies have demonstrated that exposure of primary cultures of rat cerebellar granule cells (CGC) to ≥10μM TBBPA induces toxicity and excitotoxicity, and the underlying mechanism may involve calcium imbalance and oxidative stress. Here we examined whether the application of TBBPA at subtoxic concentrations may exacerbate acute damage of CGC challenged with oxygen-glucose deprivation (OGD), and evaluated with fluorescent indicators the involvement of calcium imbalance, mitochondrial depolarization and oxidative stress. MethodsSurvival of CGC was assessed 24h after OGD/TBBPA using fluorescent dyes. An OGD challenge lasting for 45, 60 or 75min induced a duration-dependent injury to the neurons. ResultsApplication of 2.5, 5 or 7.5μM TBBPA for 45min to normoxic and glucose-containing incubation medium did not reduce the viability of cultured CGC, but this compound exacerbated the toxic effects of OGD in a concentration-dependent way. Moreover, TBBPA had a slight effect on calcium homeostasis and mitochondrial membrane potential, but significantly activated the production of reactive oxygen species in CGC. The application of H2O2 at 5, 10 and 25μM mimicked the effects of TBBPA on OGD toxicity, while 0.1mM ascorbic acid or 1mM glutathione ameliorated this toxicity. ConclusionThese results suggest the involvement of oxidative stress in the synergistic neurotoxic effects of TBBPA and OGD.

Behavior of Platelet Activating Factor in Membrane-Mimicking Environment. Langmuir Monolayer Study Complemented with Grazing Incidence X-ray Diffraction and Brewster Angle Microscopy

1-O-octadecyl-2-acetyl-sn-glycero-3-phosphocholine (PAF) belonging to the class of single-chained ether phospholipids is widely known from its essential biological activities. There is a growing body of evidence that some significant aspects of PAF actions are connected with its capability to direct intercalation into biomembranes' environment. Although this mechanism is of great importance in the perspective of understanding PAF implications in various physiological processes, in the literature, there is a lack of studies devoted to this subject. It is still unknown which is the exact influence of membrane composition, molecular organization, and its other properties on the PAF impact on cells and tissues. Unfortunately, the biological studies carried out on cell cultures do not provide satisfactory results, mainly because of the complexity of natural systems. In order to obtain insight into the behavior of PAF in a lipid environment at the molecular level, the application of appropriate model systems is required. Among them, Langmuir monolayers are very often applied as a simple but very efficient platform for studies of the interactions between membrane lipids. In the present paper, special attention is focused on the issue concerning the interactions between PAF and two representatives of membrane components occurring mainly in the outer leaflet of natural bilayers, namely, cholesterol and DPPC. The application of Langmuir monolayers enabled us to construct the effective model mimicking the exogenous incorporation of PAF into membrane environment. On the basis of the obtained results, a thorough discussion was carried out and the conclusions derived from the traditional thermodynamic analysis were confronted with microscopic analysis of surface domains and the GIXD results. The selection of experimental techniques enables us to obtain information regarding the miscibility and interactions in the binary mixed films as well as the molecular organization of film-forming molecules on water surface. The experiments revealed that the addition of the investigated single-chained ether phospholipid into both cholesterol and DPPC monolayers causes a considerable decrease of monolayer condensation. On the basis of thermodynamic analysis, it was found that PAF mixes and consequently interacts strongly with cholesterol, whereas its interactions with DPPC are thermodynamically unfavorable. Differences between the PAF influence on cholesterol and DPPC monolayer found its corroboration in the results obtained with the GIXD technique. Namely, the monolayer of DPPC can incorporate more PAF than the model membrane containing cholesterol. The obtained results indicate that short chained sn-2 ether phospholipid is able to modify model membrane properties in a concentration-dependent way.

Alterations observed in the endothelial HUVEC cell line exposed to pure Cylindrospermopsin

The cyanobacterial toxin Cylindrospermopsin (CYN) is receiving great interest due to its increasing presence in waterbodies, which has lead to recognize it as a potential threat to drinking water safety. CYN is a potent inhibitor of protein and glutathione synthesis. The present work studies for the first time the effects of CYN in endothelial cells. The basal cytotoxicity endpoints studied at 24 and 48h were total protein content (PC), neutral red (NR) uptake and the tretazolium salt, MTS, reduction. Moreover, the effect of subcytotoxic concentrations of CYN on the generation of intracellular reactive oxygen species (ROS), the activity of γ-glutamylcysteine synthetase (GCS) and glutathione (GSH) content have been investigated. In addition, morphological alterations of HUVEC cells subsequent to CYN exposure were recorded.The cytotoxicity endpoints revealed a decrease in the cellular viability in a time and concentration-dependent way. The most sensitive cytotoxicity endpoint was NR uptake assay, with reductions in cell viability of 95% at 48h of exposure to 40μgmL−1 CYN. Intracellular ROS production was increased only at the lowest concentration assayed, while GCS activity and GSH content underwent concentration-dependent enhancements. The most remarkable morphological alterations observed were: nucleolar segregation with altered nuclei, degenerated Golgi apparatus, increases in the presence of granules and apoptosis.

Relaxant Effect of a Water Soluble Carbon Monoxide-Releasing Molecule (CORM-3) on Spontaneously Hypertensive Rat Aortas

Both carbon monoxide (CO) and nitric oxide (NO) are two gaseous molecules performing relevant functions in mammals. In order to better understand their actions in the cardiovascular system, we have investigated the effects of CORM-3, (tricarbonylchloro(glycinato)ruthenium(II), a water soluble CO-releasing molecule and SNAP (S-nitroso-N-acetyl-DL-penicillamine, a well known NO-releasing molecule) on aortas of normotensive Wistar Kyoto (WKY) and spontaneously hypertensive rats (SHR). The isometric contraction of angiotensin II (AT-II) and endothelin-1 (ET-1) was evaluated in endothelium-denuded aortic strips. In control conditions, AT-II induced a similar concentration-dependent contraction in both WKY and SHR, while ET-1 was more effective in SHR aortic strips. CORM-3 or SNAP (10(-7)-3 × 10(-4) M) reduced the contraction induced by AT-II or ET-1 in a concentration-dependent way. Whereas the median inhibitory concentration of SNAP was significantly lower in WKY than in SHR, CORM-3 had a similar effect in both strains. The scaffold compound iCORM-3 was ineffective. Pretreatment with an inhibitor of soluble guanylyl cyclase (ODQ, 3 × 10(-6) M) marginally reduced CORM-3 relaxation in both strains, whereas it reduced relaxation induced by SNAP in WKY and, to a lesser extent, in SHR. The benzylindazole derivative YC-1 (10(-6) M), a sensitizer of soluble guanylate cyclase to the action of NO, significantly increased the relaxant effect of SNAP in AT-II precontracted aortic strips. The blocker of calcium-activated potassium channels, charybdotoxin (10(-8) M), reduced the relaxation induced by CORM-3 in both strains. Different mechanisms seem to be implicated in CO- and NO-mediated vascular relaxation. Since the relaxant properties of CO are conserved in SHR aortas, CORM-3 could be a new potential agent for the treatment of hypertension, when NO donors show sub-optimal or absent responses.

EphA3 Expressed in the Chicken Tectum Stimulates Nasal Retinal Ganglion Cell Axon Growth and Is Required for Retinotectal Topographic Map Formation

BackgroundRetinotopic projection onto the tectum/colliculus constitutes the most studied model of topographic mapping and Eph receptors and their ligands, the ephrins, are the best characterized molecular system involved in this process. Ephrin-As, expressed in an increasing rostro-caudal gradient in the tectum/colliculus, repel temporal retinal ganglion cell (RGC) axons from the caudal tectum and inhibit their branching posterior to their termination zones. However, there are conflicting data regarding the nature of the second force that guides nasal axons to invade and branch only in the caudal tectum/colliculus. The predominant model postulates that this second force is produced by a decreasing rostro-caudal gradient of EphA7 which repels nasal optic fibers and prevents their branching in the rostral tectum/colliculus. However, as optic fibers invade the tectum/colliculus growing throughout this gradient, this model cannot explain how the axons grow throughout this repellent molecule.Methodology/Principal FindingsBy using chicken retinal cultures we showed that EphA3 ectodomain stimulates nasal RGC axon growth in a concentration dependent way. Moreover, we showed that nasal axons choose growing on EphA3-expressing cells and that EphA3 diminishes the density of interstitial filopodia in nasal RGC axons. Accordingly, in vivo EphA3 ectodomain misexpression directs nasal optic fibers toward the caudal tectum preventing their branching in the rostral tectum.ConclusionsWe demonstrated in vitro and in vivo that EphA3 ectodomain (which is expressed in a decreasing rostro-caudal gradient in the tectum) is necessary for topographic mapping by stimulating the nasal axon growth toward the caudal tectum and inhibiting their branching in the rostral tectum. Furthermore, the ability of EphA3 of stimulating axon growth allows understanding how optic fibers invade the tectum growing throughout this molecular gradient. Therefore, opposing tectal gradients of repellent ephrin-As and of axon growth stimulating EphA3 complement each other to map optic fibers along the rostro-caudal tectal axis.

Oxidatively modified high density lipoprotein promotes inflammatory response in human monocytes–macrophages by enhanced production of ROS, TNF-α, MMP-9, and MMP-2

It has been proposed that high-density lipoprotein (HDL) loses its cardioprotective ability through oxidative modifications by reactive oxygen species (ROS) and promote atherogenesis. However, the pro-atherogenic pathways undergone by oxidized HDL remain poorly understood. Since monocytes play a crucial role in atherogenesis, this study was aimed to investigate the influence of both native and oxidized HDL (oxHDL) on monocytes-macrophages functions relevant to atherogenesis. HDL particles were isolated from human blood samples by ultracentrifugation and subjected to in vitro oxidation with CuSO(4). The extent of oxidation was quantitated by measurement of lipid peroxides. Human peripheral blood mononuclear cells were isolated and cultured under standard conditions. Cells were treated with native and oxHDL at varying concentrations for different time intervals and used for several analyses. Intracellular ROS production was assessed based on ROS-mediated DCFH fluorescence of the cells. The release of TNF-α and matrix metalloproteinases (MMPs) was quantitated using ELISA kit and gelatine zymography, respectively. Treatment of cells with oxidized HDL enhanced the production of ROS in a concentration-dependent way, while native HDL had no such effect. Further, the release of TNF-α, MMP-9, and MMP-2 was found to be remarkably higher in cells incubated with oxHDL than that of native HDL. Results demonstrate that oxidative modification of HDL induces pro-inflammatory response and oxidative stress in human monocytes-macrophages.

Comparison of the tolerance on the venous relaxation induced by nitroglycerin and by the new nitrite donor cis‐[Ru(bpy)2(py)(NO2)](PF6)

Organic nitrates such as nitroglycerin (GNT, glycerol trinitrate), are commonly used in clinical cardiovascular medicine in the acute treatment of angina and severe arterial hypertension. The development of tolerance due to the continuous treatment with high doses leads to a decrease of the magnitude of the vasodilatation induced by GNT. It is a major limitation for the chronic clinical use of GNT. The present study aimed to verify if the new nitrite donor cis‐[Ru(bpy)2(py)(NO2)](PF6) (Rubpy) induces in vitro tolerance in the rat cava vein and to compare this effect with GTN. We have analyzed the maximum relaxing effect (ME) and the potency (EC50) induced by GNT and Rubpy in concentration‐effect curves constructed in vein rings pre‐contracted with endothelin‐1. The experiments were performed on intact endothelial rings following pre‐incubation for 30 min with GTN (EC100: 0.1mmol/L or EC50: 4 μmol/L) or Rubpy (EC100: 0.1μmol/L or EC50: 2 μmol/L). After pre‐incubation with GTN (EC100), the ME was reduced (from 57.5±2.5%, n=4 to 28.1±0.8%, n=4, p<0.001). Pre‐incubation with the EC50 of GTN had no effect. Rubpy induced relaxation in a concentration‐dependent way, that has not been changed by pre‐incubation for 30 min with RuBPY in the EC50 or EC100 (78.6±2.8%, n=4). In conclusion, prior exposure with GTN reduces the relaxation induced by GTN whereas it does not occur with Rubpy. Supported by FAPESP/CNPq.

Involvement of DNA in biofilm formation II: From bacterial adhesion to biofilm formation

The effect of extracellular DNA (eDNA) on bacterial attachment was characterized using Escherichia coli, DNase I, and several different kinds of DNA. Here, we showed that eDNA enhanced bacterial attachment to solid surface in a concentration-dependent way. Either plasmid DNA or chromosome DNA, even or eukaryotic DNA fragments showed the promotion effect, suggesting that the effect of DNA on bacterial attachment is nonspecific. This promotion effect of eDNA is separable from that of conjunctive pili. In a static culture system, biofilm can form even with the presence of active DNase I. DNase I impaired bacterium-to-bacterium adhesion and microcolony formation efficiently but had little effect on bacterial attachment to a solid surface and mature biofilm. Consequently, this study provides a rational description for the role of DNA in bacterial biofilm formation in natural environments.

Restoring Nitric Oxide Cytosolic Calcium Regulation by Cyclic Guanosine Monophosphate Protein Kinase I Alpha Transfection in Coronary Endothelial Cells of Spontaneously Hypertensive Rats

In microcoronary endothelial cells (RCEs) from spontaneously hypertensive rats (SHR), the nitric oxide (NO)/cyclic guanosine monophosphate (GMP)-dependent proteinkinase I (cGKI) pathway cannot regulate the cytosolic calcium ([Ca²⁺]_i) dynamic as in RCEs from Wistar Kyoto rats (WKY). We investigated the altered downstream NO target in SHR cells and, since cGKI expression was low, whether the re-expression of cGKIα in SHR RCEs could restore NO calcium responsiveness. We measured [Ca²⁺]_i dynamic by fura-2 imaging analysis and the cGKI level by RT-PCR and Western blot in SHR and WKY RCEs. Plasmids encoding for enhanced green fluorescence protein or cGKIα-enhanced green fluorescence protein were transiently transfected in SHR RCEs, and [Ca²⁺]_i was evaluated. Angiotensin-II (AT-II) increased [Ca²⁺]_i in a concentration-dependent way in both strains. Whereas in WKY, endogenously produced NO and cyclic GMP analog decreased the AT-II-induced [Ca²⁺]_i transient, they were ineffective in SHR RCEs. The cGKI level was low in SHR cells. However, after cGKIα re-expression, endogenous NO decreased the AT-II-induced [Ca²⁺]_i transient, while endothelial NO synthase and cGKI inhibition prevented it. The low expression of cGKI in SHR accounts for the absent regulation of the agonist-induced [Ca²⁺]_i transient by the NO/cyclic GMP pathway. Studies on cGKI in humans could contribute to a better understanding of cardiovascular pathologies.

A new methyl-thiochroman-4-ketone derivative induces apoptosis in MCF-7 human breast cancer cells <italic>in vitro</italic>

To investigate effects of ( Z )-3-chloromethylene-5-methyl-thiochroman-4-ketone (CMTK) on apoptosis in human breast cancer MCF-7 cells in vitro , cultured MCF-7 cells were treated with different concentrations of CMTK. The MTT method was used to study inhibition rates; the trypan blue method for cell viability and to construct growth curves; flow cytometry for cell cycles; the TUNEL assay and flow cytometry to assess proportion of apoptotic cells; Ac-IETD-pNA caspase-8/Ac-LEHD-pNA caspase 9 Colorimetric Assay Kit for caspase-8 and caspase-9 activities; and ELISA assays to detect DR3, TNFR1, TNF-α and caspase-3 proteins. Results showed that CMTK had dramatic anti-tumor activities at low concentrations (IC 50 : 14.24 ± 0.12 μmol L -1 ). Importantly, the IC 50 of CMTK on MCF-7 cells was much higher than that of cisplatin. Growth curves show CMTK can inhibit cell proliferation in time- and dose-dependent manners. In cell-cycle analysis, proportions of MCF-7 cells in G 0 /G 1 phase after 12-h and 24-h treatment with CMTK were significantly different from the control group. In contrast, the proportion of cells in S phase tended to decrease, but not significantly. Use of TUNEL and flow cytometry indicated that CMTK induces apoptosis. In MCF-7 cells, CMTK induces TNFR1 production, activates caspase-8 and caspase-9, and enhances caspase-3 levels in a concentration-dependent way; however, CMTK does not affect production of DR3 and TNF-α. These results suggest that the anti-tumor mechanism of CMTK involves inducing apoptosis via caspase-8 activation by death receptor, and caspase-9 activation by the mitochondrial-dependent pathway. Our results provide preliminary data for further investigation of the apoptotic mechanism.

Concentration-Response Analysis of Differential Gene Expression in the Zebrafish Embryotoxicity Test Following Flusilazole Exposure

The zebrafish embryotoxicity test (ZET) is considered a promising alternative model in predictive toxicology. Currently, morphological assessment of the embryo is the main readout for this assay. However, implementation of transcriptomics may help to detect more subtle effects, which may increase the sensitivity and predictability of the test. In this study, we tested a concentration response of flusilazole in the ZET. After exposure for 24 h postfertilization, microarray analysis revealed a number of processes to be regulated in a concentration-dependent way. We identified development related processes, retinol metabolism and transcription, as well as processes corresponding to the antifungal mechanism of action, steroid biosynthesis, and fatty acid metabolism, to be differentially regulated. Retinol metabolism and transcription were already significantly altered at concentrations that were not inducing morphological effects. Differential expression of genes related to steroid biosynthesis and fatty acid metabolism showed a concentration response similar to morphological response. An increase in concentration was also positively associated with an increase in magnitude of expression for individual genes within functional processes. Our study shows that transcriptomics analysis in the ZET is a more sensitive readout of compound-induced effects than morphological assessment. However, the interpretation of differential gene expression in terms of predicting morphological effects is not straightforward and requires further study.

Effects of rare earth elements La and Yb on the morphological and functional development of zebrafish embryos

AbstractIn recent years, with the wide applications and mineral exploitation of rare earth elements, their potential environmental and health effects have caused increasing public concern. Effect of rare earth elements La and Yb on the morphological and functional development of zebrafish embryos were studied. The embryos were exposed to La3+ or Yb3+ at 0, 0.01, 0.1, 0.3, 0.5 and 1.0 mmol/L, respectively. Early life stage parameters such as egg and embryo mortality, gastrula development, tail detachment, eyes, somite formation, circulatory system, pigmentation, malformations, hatching rate, length of larvae and mortality were investigated. The results showed La3+ and Yb3+ delayed zebrafish embryo and larval development, decreased survival and hatching rates, and caused tail malformation in a concentration-dependent way. Moreover, heavy rare-earth ytterbium led to more severe acute toxicity of zebrafish embryo than light rare-earth lanthanum.

Avaliação das atividades antioxidante, anti e pró-hemolítica do extrato etanólico das folhas de Pterogyne nitens Tul. (Fabaceae-Caesalpinioideae)

A pesquisa de produtos naturais permite a descoberta de novos princípios ativos, ou ainda, a descoberta de novas atividades para extratos de plantas (amplamente utilizados pela população brasileira) e princípios ativos naturais já conhecidos. Pterogyne nitens é uma planta cuja descrição das atividades é relativamente recente e, portanto, tem no extrato bruto boa fonte para pesquisas na área de produtos naturais. Desta forma, o objetivo deste trabalho foi estudar o perfil antioxidante do extrato bruto etanólico das folhas de P. nitens e possível interferência sobre a hemólise provocada pelo radical AAPH•. No estudo da ação antioxidante das espécies estudadas, ABTS•+, DPPH•, H2O2 e HOCl, encontrou-se os valores de IC50 de 5,0 µg mL-1, 17 µg mL-1, sem ação e 3,9 µg mL-1, respectivamente, valores relativamente baixos e que indicam bom potencial antioxidante. Foram encontradas atividades pró-hemolítica e anti-hemolítica para o extrato de forma concentração-dependente. O extrato estudado mostro boa fonte de moléculas naturais com potencial de ação biológica.

Sigma-1 receptors do not regulate calcium influx through voltage-dependent calcium channels in mouse brain synaptosomes

Several lines of evidence suggest that σ1 receptors regulate intracellular calcium concentration [Ca2+]i. However, no previous studies have demonstrated a consistent role for these receptors in the modulation of extracellular calcium entry through plasmalemmal voltage-dependent calcium channels (VDCCs). To search for evidence of such a role we compared [Ca2+]i under basal conditions and after depolarization with KCl in fura-2-loaded synaptosomes from wild-type and σ1 receptor knockout (σ1R-KO) mice. We also tested the effects of the selective σ1 receptor agonists PRE-084 and (+)-pentazocine and antagonists BD-1047 and NE-100 on the increase in [Ca2+]i induced by depolarization with 60mM KCl. Mibefradil, a nonselective blocker of VDCCs, was used as a positive control. Basal [Ca2+]i and the increase in [Ca2+]i caused by KCl-induced depolarization were similar in brain synaptosomes from both wild-type and σ1R-KO mice. Mibefradil (1–30μM) and all σ1 receptor ligands studied (3–100μM) inhibited the KCl-induced increase in [Ca2+]i in a concentration-dependent way. The order of maximum inhibition for the ligands compared here was NE-100>BD-1047=PRE 084>(+)-pentazocine. There were no appreciable differences in their effects between wild-type and σ1R-KO mice. These findings indicate that σ1 receptors are not involved in calcium influx through VDCCs or in the inhibitory effects of these σ1 ligands on Ca2+ channels.