Heterologous production of α-chain of Dioclea sclerocarpa lectin: Enhancing the biological effects of a wild-type lectin.

Lectins from Diocleinae subtribe species (family Leguminosae) are of special interest since they present a wide spectrum of biological activities, despite their high structural similarity. During their synthesis in plant cells, these proteins undergo post-translational processing resulting in the formation of three chains (α, ß, γ), which constitute the lectins' subunits. Furthermore, such wild-type proteins are presented as isolectins or with different combinations of these chains, which undermine their biotechnological potential. Thus, the present study aimed to produce a recombinant form of the lectin from Dioclea sclerocarpa seeds (DSL), exclusively constituted by α-chain. The recombinant DSL (rDSL) was successfully expressed in E. coli BL21 (DE3) and purified by affinity chromatography (Sephadex G-50), showing a final yield of 74 mg of protein per liter of culture medium and specificity for D-mannose, α-methyl-mannoside and melibiose, unlike the wild-type protein. rDSL presented an effective vasorelaxant effect in rat aortas up to 100% and also interacted with glioma cells C6 and U87. Our results demonstrated an efficient recombinant production of rDSL in a bacterial system that retained some biochemical properties of the wild-type protein, showing wider versatility in sugar specificities and better efficacy in its activity in the biological models evaluated in this work.

Detection, purification and characterization of a lectin from freshwater green algae Spirogyra spp.

Freshwater algae are rich sources of structurally biologically active metabolites, such as fatty acids, steroids, carotenoids and polysaccharides. Among these metabolites, lectins stand out. Lectins are proteins or glycoproteins of non-immune origin which bind to carbohydrates or glycoconjugates, without changing ligand structure. Many studies have reported on the use of Spirogyra spp. as effective bioindicators of heavy metals; however, reports on Spirogyra molecular bioprospecting are quite limited. Therefore, this study aimed to detect, isolate, purify and characterize a lectin present in the freshwater green algae Spirogyra. Presence of the lectin protein in the extract was detected by hemagglutination assays. Subsequently, the protein extract was subjected to a sugar inhibition assay to identify the lectin-specific carbohydrate. Following this, the extract was applied to a guar gum column to afford the pure lectin. The lectin was inhibited by N-acetyl-glucosamine and N-acetyl-beta-D-mannose, but more strongly by D-galactose. The apparent molecular mass of the purified lectin was evaluated by Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE). Electrophoretic analysis revealed a single protein band with an apparent molecular mass of 56 kDa. Thus, it could be concluded that a lectin was purified from Spirogyra spp.

Detection, purification and characterization of a lectin from freshwater green algae Spirogyra spp

ABSTRACT Freshwater algae are rich sources of structurally biologically active metabolites, such as fatty acids, steroids, carotenoids and polysaccharides. Among these metabolites, lectins stand out. Lectins are proteins or glycoproteins of non-immune origin which bind to carbohydrates or glycoconjugates, without changing ligand structure. Many studies have reported on the use of Spirogyra spp. as effective bioindicators of heavy metals; however, reports on Spirogyra molecular bioprospecting are quite limited. Therefore, this study aimed to detect, isolate, purify and characterize a lectin present in the freshwater green algae Spirogyra. Presence of the lectin protein in the extract was detected by hemagglutination assays. Subsequently, the protein extract was subjected to a sugar inhibition assay to identify the lectin-specific carbohydrate. Following this, the extract was applied to a guar gum column to afford the pure lectin. The lectin was inhibited by N-acetyl-glucosamine and N-acetyl-beta-D-mannose, but more strongly by D-galactose. The apparent molecular mass of the purified lectin was evaluated by Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE). Electrophoretic analysis revealed a single protein band with an apparent molecular mass of 56 kDa. Thus, it could be concluded that a lectin was purified from Spirogyra spp.

Can Transcranial Direct Current Stimulation Improve the Resistance Strength and Decrease the Rating Perceived Scale in Recreational Weight-Training Experience?

Lattari, E, Andrade, ML, Filho, AS, Moura, AM, Neto, GM, Silva, JG, Rocha, NB, Yuan, T-F, Arias-Carrión, O, and Machado, S. Can transcranial direct current stimulation improve the resistance strength and decrease the rating perceived scale in recreational weight-training experience? J Strength Cond Res 30(12): 3381-3387, 2016-The goal of this study was to evaluate the acute efficacy of anodic transcranial direct current stimulation on the total volume of repetitions and perceived exertion in recreationally trained individuals in strength. The sample consisted of 10 participants trained in exercise against resistance for at least 3 months. Participants underwent elbow flexion exercise at barbell with a specific load of 10 repetition maximum (10RM), responded immediately after the OMNI-RES scale, and were stimulated for 20 minutes with a tDSC protocol (2 mA), depending on randomization. After applying the tDSC, subjects were again subjected to perform elbow flexion with 10RM load and, soon after, again responded to OMNI-RES scale. All subjects underwent the 3 experimental conditions of the study, c-tDSC, a-tDSC, and sham-tDSC, which were randomized. A range of 48-72 hours was allowed between each assessment visit. An interaction to condition and time (F = 52.395; p ≤ 0.001) has shown that repetitions completed after anodic condition were higher compared with the other conditions in the postsession. In relation to perceived exertion, verified by OMNI-RES scale, 2-way analysis of variance for repeated measures showed an interaction between condition and time (F = 28.445; p ≤ 0.001), where the perceived exertion was decreased after the a-tDSC condition and increased after the c-tDSC condition. In strict terms of performance, it seems to be beneficial to attend a session of 20 minutes a-tDSC, when strength training practitioners can no longer support high-volume training and have increased responses in the perceived exertion.

Applied investigation on the interaction of hazardous elements binding on ultrafine and nanoparticles in Chinese anthracite-derived fly ash.

A multifaceted instrumental approach was employed to determine the chemistry and mineralogy of pulverized-coal-combustion fly ashes from two Chinese power plants. Techniques included traditional optical microscopy, X-ray diffraction, and chemical analysis along with a variety of electron beam methods. The aim is to demonstrate and bring together the wide variety of procedures dealing with F as the key element of concern, and determining its location in the mineral nanoparticles. The Hg content of the Anwen (Songzao coalfield) fly ashes is higher than that of the Diandong (East Yunnan) fly ashes, possibly owing to the greater C and Cl in the Anwen fly ashes. Both fly ash sources contain a variety of amorphous and nano-crystalline trace-element-bearing particles, both associated with multi-walled carbon nanotubes and as particles independent of carbons.

The occurrence of hazardous volatile elements and nanoparticles in Bulgarian coal fly ashes and the effect on human health exposure.

Low-rank, high-mineral matter Bulgarian coals were studied using a variety of chemical, optical, and electron beam methods. The larger fly ash carbon phases include charred carbons in contrast to coked carbons present in the fly ashes of bituminous-coal-derived fly ashes. Nanoscale carbons include multi-walled carbon nanotubes (MWCNTs) encapsulating Hg, Se, and As, among other elements. In addition to the glass which dominates the fly ash, relatively coarse 'rock fragments', consisting of an unmelted to partially melted core surrounded by a glassy rim, are present in the fly ash. Nano-scale minerals can contain hazardous elements and, along with metal-bearing multiwalled nanotubes, can be a path for the entry of hazardous particles into the lungs and other organs.

Influence of n-3 polyunsaturated fatty acids on soluble cellular adhesion molecules as biomarkers of cardiovascular risk in young healthy subjects.

BACKGROUND AND AIM: Serum levels of soluble cellular adhesion molecules (CAMs) and blood lipid parameters have been used as markers of inflammatory processes associated with cardiovascular disease (CVD) events. The present study evaluated the effects of the intake of n-3 polyunsaturated fatty acids (PUFAs) in fish and fish oil within energy-restricted diets, on soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular adhesion molecule-1 (sVCAM-1). METHODS AND RESULTS: Two hundred and seventy-five healthy European subjects aged between 20 and 40 years, were randomized to one of four hypocaloric dietary groups: control (sunflower oil capsules, no seafood), lean fish (3 x 150 g portions of cod/week), fatty fish (3 x 150 g portions of salmon/week), fish oil ((docosahexaenoic acid (DHA)+eicosapentaenoic acid (EPA) capsules, no seafood)). Diets rich in lean fish significantly decreased ICAM-1 levels, around 5% from baseline to endpoint (p<0.05), and had no effect on VCAM-1 levels. No significant differences were observed in sICAM-1 levels after the intervention with fatty fish or fish oils. On the other hand, these two seafood based diets were responsible for a significant increase of VCAM-1 levels [fatty fish; 16.1% and fish oil; 21.9%] respectively (p<0.05). CONCLUSIONS: CAMs as inflammatory biomarkers in young and healthy subjects are not conclusive for the evaluation of CVD risk. Hypocaloric fish diets had a different effect on CAMs, being lean fish responsible for the highest decrease in ICAM-1. On the other hand, VCAM-1 results allow speculation that a low dose of n-3 PUFA may be anti-inflammatory contrarily to a high dose which can have a pro-inflammatory effect. CAMs mechanism is complex and affected by multiple factors such as lifestyle, gender, and n-3 dose and source.

Production of transgenic goat (Capra hircus) with human Granulocyte Colony Stimulating Factor (hG-CSF) gene in Brazil.

In order to produce transgenic goats with hG-CSF, a total of 24 adult Saanen and 48 adult undefined breed goats were used as donors and recipients, respectively. Donors were estrus-synchronized with vaginal sponges and superovulated by a treatment with 200 mg FSH given twice daily in decreasing doses over 3 days starting 48 h before sponge removal. Ovulation was induced by injecting 100 microg GnRH 36 h after sponge removal. The recipients also received an estrus synchronization treatment. Donors were mated with fertile Saanen bucks and, approximately 72 h after sponge removal, zygotes were recovered surgically by flushing oviducts. The recovered zygotes were briefly centrifuged to a reliable visualization of the pronuclei. The DNA construct containing hG-CSF gene flanked by goat and bovine alphas1-casein sequences was injected into pronuclei of 129 zygotes. The microinjected embryos (3-6 per female) were transferred to 27 recipients. Ten recipients became pregnant and 12 kids were born. One transgenic male founder was identified in the group of kids. This is the first report of a birth of a transgenic goat in Latin America.

Production of transgenic goat (Capra hircus) with human Granulocyte Colony Stimulating Factor (hG-CSF) gene in Brazil

In order to produce transgenic goats with hG-CSF, a total of 24 adult Saanen and 48 adult undefined breed goats were used as donors and recipients, respectively. Donors were estrus-synchronized with vaginal sponges and superovulated by a treatment with 200 mg FSH given twice daily in decreasing doses over 3 days starting 48 h before sponge removal. Ovulation was induced by injecting 100µg GnRH 36 h after sponge removal. The recipients also received an estrus synchronization treatment. Donors were mated with fertile Saanen bucks and, approximately 72 h after sponge removal, zygotes were recovered surgically by flushing oviducts. The recovered zygotes were briefly centrifuged to a reliable visualization of the pronuclei. The DNA construct containing hG-CSF gene flanked by goat and bovine alphas1-casein sequences was injected into pronuclei of 129 zygotes. The microinjected embryos (3-6 per female) were transferred to 27 recipients. Ten recipients became pregnant and 12 kids were born. One transgenic male founder was identified in the group of kids. This is the first report of a birth of a transgenic goat in Latin America.

A fim de produzir caprinos transgênicos para o hG-CSF, utilizou-se 24 cabras Saanen adultas e 48 cabras sem raça definida adultas como doadoras e receptoras, respectivamente. As doadoras tiveram o estro sincronizado por esponjas vaginais e foram superovuladas com 200 mg de FSH em doses decrescentes, duas vezes ao dia e iniciando 48 h antes da retirada da esponja. A ovulação foi induzida pela injeção de 100 µg de GnRH às 36 h após a retirada da esponja. As receptoras também receberam um tratamento de sincronização do estro. As doadoras foram cobertas por bodes Saanen férteis e, aproximadamente 72 h após a retirada da esponja, os zigotos foram colhidos cirurgicamente por lavagem dos ovidutos. Os zigotos colhidos foram rapidamente centrifugados para uma melhor visualização dos pró-núcleos. A construção de DNA, contendo o gene do hG-CSF flanqueado pelos genes caprino e bovino da alfas1-caseína, foi injetada em 129 embriões. Os embriões microinjetados (3 a 6 por receptora) foram transferidos para 27 receptoras que responderam ao tratamento. Dez receptoras ficaram gestantes e 12 crias foram produzidas. Um macho transgênico fundador foi identificado no grupo de crias nascidas. Este é o primeiro relato do nascimento de um caprino transgênico na América Latina.