RESUMEN
This study evaluated the effects of acute exposure of Aedes aegypti third instar (L3 ) larvae to the saline extract of Opuntia ficus-indica cladodes on the biological cycle and fertility of the emerging adults. For this, larvae were treated for 24 h with the extract at » LC50 (lethal concentration to kill 50% of larvae), ½ LC50 or LC50 ; the development and reproduction of the emerged adults were evaluated after a recovery period of 9 days. The resistance of proteins in the extract to hydrolysis by L3 digestive enzymes and histomorphological alterations in the larval midgut were also investigated. The extract contained lectin, flavonoids, cinnamic derivatives, terpenes, steroids, and reducing sugars. It showed a LC50 of 3.71% for 48 h. The data indicated mean survival times similar in control and extract treatments. It was observed development delay in extract-treated groups, with a lower number of adults than in control. However, the females that emerged laid similar number of eggs in control and treatments. Histological evaluation revealed absence of bacterial and fungal microorganisms in the food content in midguts from larvae treated with cladode extract. Electrophoresis revealed that three polypeptides in the extract resisted to hydrolysis by L3 digestive proteases for 90 min. The lectin activity was not altered even after 24-h incubation with the enzymes. In conclusion, the extract from O. ficus-indica can delay the development of Ae. aegypti larvae, which may be linked to induction of an axenic environment at larval midgut and permanence of lectin activity even after proteolysis.
Asunto(s)
Aedes , Insecticidas , Opuntia , Femenino , Animales , Lectinas/química , Larva , Extractos Vegetales/farmacología , Extractos Vegetales/química , Insecticidas/farmacologíaRESUMEN
Angiogenesis (budding of new blood vessels) is involved in several processes, including the development of embryos and growth of tumors. Schinus terebinthifolia leaves express an antitumor lectin (SteLL). This work hypothesized that SteLL can interfere with the formation of a vascular network from preexisting vessels. To test this hypothesis, the effect of SteLL on the angiogenesis process was assessed using an in vivo model of yolk sac membrane of Coturnix japonica embryos. SteLL was isolated with purification factor of 46.6. As expected, polyacrylamide gel electrophoresis (PAGE) for native basic proteins confirmed the homogeneity and PAGE in presence of dodecyl sodium sulphate revealed a single 14-kDa polypeptide band. The fractal analysis by box counting and information dimension measurements indicated that SteLL at 1.35 mg/mL significantly decreased by ca. 12% the angiogenesis within the C. japonica yolk sac membrane regarding the control. The inhibition of the vascular network formation in the yolk sac membrane resulted in decreased blood supply to the embryos. Consequently, the area of embryos was significantly reduced by 9.2% regarding the control, which corroborated with the antiangiogenic activity of SteLL. The findings implicate SteLL as an antiangiogenic agent and add to the panel of biological activities of this lectin.
Asunto(s)
Anacardiaceae , Coturnix , Inhibidores de la Angiogénesis/farmacología , Animales , Lectinas/farmacología , Hojas de la PlantaRESUMEN
Haemonchus contortus is a hematophagous parasite causing damage to the production of ruminant animals throughout the world. This study evaluated the in vitro effect of proteins from Moringa oleifera (WSMoL - Water Soluble M. oleifera Lectin and cMoL - coagulant M. oleifera Lectin) on the motility of infective larvae and adult male and female worms of H. contortus. The specific activity of total proteases and the morphology of the worms exposed to the lectins were observed. Both lectins inhibited motility of all parasite stages tested. WSMoL and cMoL at 500 µg mL-1 interfered in the motility of larvae. Values of 11.1% and 8.1% were the lowest motility indices of larvae with sheath, and 30.6% and 16.4% were the lowest motility indices of exsheathed larvae treated with WSMoL and cMoL, respectively. In 1 mg mL-1 solutions of WSMoL and of cMoL, the motility index of adult male worms was 23.3% (p < 0.001) and 20% (p < 0.001), while the motility index of adult female worms was 63.3% (p > 0.05) and 26.6% (p < 0.001), respectively. Greater proteolytic activity was detected in extracts obtained from adult worms, male and female, after incubation with the lectins. Morphological changes caused by the lectins were revealed by changes in the crests of the cuticle, in the longitudinal striations and at the vulva.
Asunto(s)
Haemonchus/efectos de los fármacos , Moringa oleifera/química , Lectinas de Plantas/farmacología , Semillas/química , Animales , Femenino , Haemonchus/enzimología , Haemonchus/fisiología , Haemonchus/ultraestructura , Larva/efectos de los fármacos , Masculino , Microscopía Electrónica de Rastreo , Movimiento/efectos de los fármacos , Péptido Hidrolasas/metabolismo , Lectinas de Plantas/administración & dosificación , Lectinas de Plantas/aislamiento & purificaciónRESUMEN
Lactones are organic cyclic esters that have been described as larvicides against Aedes aegypti and as components of oviposition pheromone of Culex quinquefasciatus. This work describes the effect of six α,ß-unsaturated lactones (5a-5f) on survival of A. aegypti fourth instar larvae (L4). It is also reported the effects of the lactones on L4 gut trypsin activity and oviposition behavior of A. aegypti females. Five lactones were able to kill L4 being the lactones 5a (LC50 of 39.05 ppm), 5e (LC50 of 36.30 ppm) and 5f (LC50 of 40.46 ppm) the most promising larvicides. Only the lactone 5a inhibited L4 gut trypsin activity, with an IC50 of 115.15 µg/mL. Lactones 5a, 5c, 5d and 5e did not exert deterrent or stimulatory effects on oviposition, whereas lactone 5b exhibited a strong deterrent oviposition activity. In conclusion, this work introduces new α,ß-unsaturated lactones as promising alternatives to control A. aegypti dissemination. The larvicidal mechanism of the lactone 5a can involve the disruption of proteolysis at larval gut.
Asunto(s)
Aedes/efectos de los fármacos , Insectos Vectores/efectos de los fármacos , Insecticidas/farmacología , Lactonas/farmacología , Oviposición/efectos de los fármacos , Tripsina/efectos de los fármacos , Aedes/fisiología , Animales , Femenino , Concentración 50 Inhibidora , Insectos Vectores/fisiología , Insecticidas/química , Lactonas/química , Larva/efectos de los fármacos , Tripsina/metabolismo , Inhibidores de Tripsina/farmacologíaRESUMEN
Arterial thrombosis is an important complication of diabetes and cancer, being an important target for therapeutic intervention. Crataeva tapia bark lectin (CrataBL) has been previously shown to have hypoglycemiant effect and also to induce cancer cell apoptosis. It also showed inhibitory activity against Factor Xa (Kiapp=8.6 µm). In the present study, we evaluated the anti-thrombotic properties of CrataBL in arterial thrombosis model. CrataBL prolongs the activated partial thromboplastin time on human and mouse plasma, and it impairs the heparin-induced potentiation of antithrombin III and heparin-induced platelet activation in the presence of low-dose ADP. It is likely that the dense track of positive charge on CrataBL surface competes with the heparin ability to bind to antithrombin III and to stimulate platelets. In the photochemically induced thrombosis model in mice, in the groups treated with 1.25, 5.0, or 10 mg/kg CrataBL, prior to the thrombus induction, the time of total artery occlusion was prolonged by 33.38%, 65%, and 66.11%, respectively, relative to the time of the control group. In contrast to heparin, the bleeding time in CrataBL-treated mice was no longer than in the control. In conclusion, CrataBL was effective in blocking coagulation and arterial thrombus formation, without increasing bleeding time.
Asunto(s)
Coagulación Sanguínea/efectos de los fármacos , Inhibidores del Factor Xa/farmacología , Lectinas de Plantas/farmacología , Trombosis/patología , Animales , Capparaceae/química , Arterias Carótidas/efectos de los fármacos , Cromatografía de Afinidad , Modelos Animales de Enfermedad , Humanos , Hidrólisis/efectos de los fármacos , Ratones Endogámicos C57BL , Óxido Nítrico/metabolismo , Tiempo de Tromboplastina Parcial , Agregación Plaquetaria/efectos de los fármacos , Tiempo de Protrombina , Flujo Sanguíneo Regional/efectos de los fármacos , Sefarosa/análogos & derivados , Sefarosa/química , Especificidad por Sustrato/efectos de los fármacosRESUMEN
Although numerous reports are available concerning the larvicidal potential of essential oils, very few investigations have focused on their mechanisms of action. In the present study, we have investigated the chemical composition of the leaf oil of Croton rhamnifolioides during storage and its effects on oviposition and survival of larvae of the dengue fever mosquito Aedes aegypti. In addition, we have established a possible mechanism of action for the larvicidal activity of the essential oil. GC-MS analyses revealed marked differences in the composition of oil that had been freshly isolated and that of a sample that had been stored in a sealed amber-glass vial under refrigeration for three years. However, both fresh and stored oil exhibited substantial larvicidal activities with LC50 values of 122.35 and 89.03 ppm, respectively, and oviposition deterrent effects against gravid females at concentrations of 50 and 100 µg·mL-1. These results demonstrate that the larvicidal effect of the essential oil was unchanged during three years of storage even though its chemical composition altered. Hence, the essential oil could be used in the preparation of commercial products. In addition, we observed that the trypsin-like activity of mosquito larvae was inhibited in vitro by the essential oil of C. rhamnifolioides, suggesting that the larvicidal effect may be associated with inhibition of this enzyme.
Asunto(s)
Aedes/anatomía & histología , Croton/química , Aceites Volátiles/farmacología , Oviposición/efectos de los fármacos , Tripsina/metabolismo , Aedes/efectos de los fármacos , Animales , Aceite de Crotón/farmacología , Femenino , Cromatografía de Gases y Espectrometría de Masas , Larva/efectos de los fármacos , Hojas de la Planta/químicaRESUMEN
CrataBL, a protein isolated from Crataeva tapia bark, which is both a serine protease inhibitor and a lectin, has been previously shown to exhibit a number of interesting biological properties, including anti-inflammatory, analgesic, antitumor, and insecticidal activities. Using a glycan array, we have now shown that only sulfated carbohydrates are effectively bound by CrataBL. Because this protein was recently shown to delay clot formation by impairing the intrinsic pathway of the coagulation cascade, we considered that its natural ligand might be heparin. Heparin is a glycosaminoglycan (GAG) that interacts with a number of proteins, including thrombin and antithrombin III, which have a critical, essential pharmacological role in regulating blood coagulation. We have thus employed surface plasmon resonance to improve our understanding of the binding interaction between the heparin polysaccharide and CrataBL. Kinetic analysis shows that CrataBL displays strong heparin binding affinity (KD = 49 nM). Competition studies using different size heparin-derived oligosaccharides showed that the binding of CrataBL to heparin is chain length-dependent. Full chain heparin with 40 saccharides or large oligosaccharides, having 16-18 saccharide residues, show strong binding affinity for CrataBL. Heparin-derived disaccharides through tetradecasaccharides show considerably lower binding affinity. Other highly sulfated GAGs, including chondroitin sulfate E and dermatan 4,6-disulfate, showed CrataBL binding affinity comparable to that of heparin. Less highly sulfated GAGs, heparan sulfate, chondroitin sulfate A and C, and dermatan sulfate displayed modest binding affinity as did chondroitin sulfate D. Studies using chemically modified heparin show that N-sulfo and 6-O-sulfo groups on heparin are essential for CrataBL-heparin interaction.
Asunto(s)
Capparaceae/metabolismo , Glicosaminoglicanos/metabolismo , Heparina/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Unión Competitiva , Capparaceae/genética , Glicosaminoglicanos/química , Heparina/química , Cinética , Modelos Moleculares , Datos de Secuencia Molecular , Proteínas de Plantas/genética , Unión Proteica , Conformación Proteica , Resonancia por Plasmón de SuperficieRESUMEN
BACKGROUND: The resistance of a Culex quinquefasciatus strain to the binary (Bin) larvicidal toxin from Lysinibacillus sphaericus is due to the lack of expression of the toxin's receptors, the membrane-bound Cqm1 α-glucosidases. A previous transcriptomic profile of the resistant larvae showed differentially expressed genes coding Cqm1, lipases, proteases and other genes involved in lipid and carbohydrate metabolism. This study aimed to investigate the metabolic features of Bin-resistant individuals by comparing the activity of some enzymes, energy reserves, fertility and fecundity to a susceptible strain. METHODS: The activity of specific enzymes was recorded in midgut samples from resistant and susceptible larvae. The amount of lipids and reducing sugars was determined for larvae and adults from both strains. Additionally, the fecundity and fertility parameters of these strains under control and stress conditions were examined. RESULTS: Enzyme assays showed that the esterase activities in the midgut of resistant larvae were significantly lower than susceptible ones using acetyl-, butyryl- and heptanoyl-methylumbelliferyl esthers as substrates. The α-glucosidase activity was also reduced in resistant larvae using sucrose and a synthetic substrate. No difference in protease activities as trypsins, chymotrypsins and aminopeptidases was detected between resistant and susceptible larvae. In larval and adult stages, the resistant strain showed an altered profile of energy reserves characterized by significantly reduced levels of lipids and a greater amount of reducing sugars. The fertility and fecundity of females were similar for both strains, indicating that those changes in energy reserves did not affect these reproductive parameters. CONCLUSIONS: Our dataset showed that Bin-resistant insects display differential metabolic features co-selected with the phenotype of resistance that can potentially have effects on mosquito fitness, in particular, due to the reduced lipid accumulation.
Asunto(s)
Bacillus , Toxinas Bacterianas , Culex , Animales , Femenino , Toxinas Bacterianas/metabolismo , Culex/metabolismo , Lípidos , Larva/genéticaRESUMEN
Allied to the problem of gastrointestinal parasites, especially Haemochus contortus, the use of lectins of plant origin has contributed to the research of alternative anthelmintics. The nematicidal effect of a water soluble Moringa oleifera lectin (WSMoL) was investigated in an experimental model with H. contortus infected Wistar rodents. Three concentrations were tested orally: 5 mg/kg, 2.5 mg/kg and 1 mg/kg. The reduction in the number of larvae recovered in the experimental groups was analyzed, as well as biochemical, hematological and histological parameters. Treatments with 5, 2.5 and 1 mg/kg of WSMoL reduced the number of larvae recovered of animals by 74.7 %, 72.8 % and 66 %, respectively. Untreated infected animals had anemia, moderate mononuclear multifocal hepatitis, vascular congestion in the liver and kidneys, white pulp hyperplasia in the spleen, and presence of eosinophils in the intestine. Infected animals treated with 5 mg/kg of WSMoL showed liver with moderate bleeding, kidney with vascular congestion, spleen with white pulp hyperplasia and intestine with moderate presence of mononuclear cells. An increase in the serum level of glutamic pyruvic transaminase and a reduction in the level of hemoglobin (p < 0.001) were also observed in this group when compared to the uninfected group. However, the administered concentrations of 2.5 and 1 mg/kg of WSMoL were both satisfactory in terms of reducing the number of recovered larvae and not promoting negative changes in the biochemical, hematological and histological parameters evaluated. These results indicate an in vivo nematicidal effect of WSMoL on the H. contortus parasite.
Asunto(s)
Antihelmínticos , Haemonchus , Moringa oleifera , Animales , Lectinas , Roedores , Agua , Hiperplasia , Antihelmínticos/farmacología , Antihelmínticos/uso terapéutico , Antinematodos , LarvaRESUMEN
Serine proteases play crucial biological roles and have their activity controlled by inhibitors, such as the EcTI, a serine protease inhibitor purified from Enterolobium contortisiliquum seeds, which has anticancer activity. This study aimed to conjugate EcTI with quantum dots (QDs), fluorophores with outstanding optical properties, and investigate the interaction of QDs-EcTI nanoprobe with cancer cells. The conjugation was evaluated by fluorescence correlation spectroscopy (FCS) and fluorescence microplate assay (FMA). EcTI inhibitory activity after interaction with QDs was also analyzed. From FCS, the conjugate presented a hydrodynamic diameter about 4× greater than bare QDs, suggesting a successful conjugation. This was supported by FMA, which showed a relative fluorescence intensity of ca. 3815% for the nanosystem, concerning bare QDs or EcTI alone. The EcTI inhibitory activity remained intact after its interaction with QDs. From flow cytometry analyses, approximately 62% of MDA-MB-231 and 90% of HeLa cells were labeled with the QD-EcTI conjugate, suggesting that their membranes have different protease levels to which EcTI exhibits an affinity. Concluding, the QD-EcTI represents a valuable nanotool to study the interaction of this inhibitor with cancer cells using fluorescence-based techniques with the potential to unravel the intricate dynamics of interplays between proteases and inhibitors in cancer biology.
Asunto(s)
Fabaceae , Neoplasias , Puntos Cuánticos , Humanos , Inhibidores de Tripsina/farmacología , Células HeLa , Fabaceae/química , Serina Proteasas , ColorantesRESUMEN
Moringa oleifera is an important source of antioxidants, tools in nutritional biochemistry that could be beneficial for human health; the leaves and flowers are used by the population with great nutritional importance. This work investigates the antioxidant activity of M. oleifera ethanolic (E1) and saline (E2) extracts from flowers (a), inflorescence rachis (b), seeds (c), leaf tissue (d), leaf rachis (e) and fundamental tissues of stem (f). The radical scavenging capacity (RSC) of extracts was determined using dot-blots on thin layer chromatography stained with a 0.4 mM 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) solution; spectrophotometric assays were recorded (515 nm). Antioxidant components were detected in all E1 and E2 from a, b and d. The best RSC was obtained with E1d; the antioxidants present in E2 reacted very slowly with DPPH. The chromatogram revealed by diphenylborinate-2-ethylamine methanolic solution showed that the ethanolic extract from the flowers, inflorescence rachis, fundamental tissue of stem and leaf tissue contained at least three flavonoids; the saline extract from the flowers and leaf tissue revealed at least two flavonoids. In conclusion, M. oleifera ethanolic and saline extracts contain antioxidants that support the use of the plant tissues as food sources.
Asunto(s)
Antioxidantes/química , Moringa oleifera/química , Extractos Vegetales/química , Etanol , Flavonoides/química , Depuradores de Radicales Libres/química , Extractos Vegetales/análisis , Hojas de la Planta/química , Tallos de la Planta/química , Semillas/química , Cloruro de SodioRESUMEN
This work aimed to characterize the coagulant properties of protein preparations from Moringa oleifera seeds in the removal of humic acids from water. Three distinct preparations were assayed, namely extract (seeds homogenized with 0.15 M NaCl), fraction (extract precipitated with 60% w/v ammonium sulphate) and cMoL (protein purified with guar gel column chromatography). The extract showed the highest coagulant activity in a protein concentration between 1 mg/L and 180 mg/L at pH 7.0. The zeta potential of the extract (-10 mV to -15 mV) was less negative than that of the humic acid (-41 mV to -42 mV) in a pH range between 5.0 and 8.0; thus, the mechanism that might be involved in this coagulation activity is adsorption and neutralization of charges. Reduction of total organic carbon (TOC) and dissolved organic carbon (DOC) was observed in water samples containing 9 mg/L carbon as humic acid when treated with 1 mg/L of the extract. A decrease in colour and in the aromatic content of the treated water was also observed. These results suggested that the extract from M. oleifera seeds in a low concentration (1 mg/L) can be an interesting natural alternative for removing humic acid from water in developing countries. The extract dose determined in the present study does not impart odour or colour to the treated water.
Asunto(s)
Sustancias Húmicas/análisis , Moringa oleifera/química , Contaminantes Químicos del Agua/aislamiento & purificación , Purificación del Agua/métodos , Proteínas de Plantas/química , Semillas/químicaRESUMEN
Lectins, proteins which selectively recognize carbohydrates, have been used in histochemistry for the evaluation of changes in glycosylation in processes of cellular differentiation and/or dedifferentiation. Cratylia mollis seed lectins (Cramoll 1,4 and Cramoll 3), conjugated to horseradish peroxidase, were used as histochemical probes in human prostate tissues: normal (NP), hyperplasia (BPH), and prostate carcinoma (PCa). The staining pattern of Con-A and Cramoll 1,4 in BPH was more intense than in NP. These lectins also showed staining differences between BPH and PCa; the latter showing decreased staining intensity with an increased degree of malignancy. PNA and Cramoll 3 stained epithelial cells similarly in all diagnoses although they did present intense staining of PCa glands lumen. Corpora amylacea were not differentially recognized by any of the lectins. Cramoll 1,4 and Cramoll 3 seed lectins present themselves as candidates for histochemical probes for prostate pathologies when compared to commercial lectins such as Con-A and PNA.
Asunto(s)
Histocitoquímica/métodos , Lectinas de Plantas/química , Próstata/metabolismo , Hiperplasia Prostática/metabolismo , Neoplasias de la Próstata/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Concanavalina A/química , Concanavalina A/metabolismo , Fabaceae , Glicosilación , Humanos , Masculino , Persona de Mediana Edad , Aglutinina de Mani/química , Aglutinina de Mani/metabolismo , Lectinas de Plantas/metabolismo , Semillas/química , Estadísticas no ParamétricasRESUMEN
Glycoconjugates found on cell walls of Candida species are fundamental for their pathogenicity. Laborious techniques have been employed to investigate the sugar composition of these microorganisms. Herein, we prepared a nanotool, based on the fluorescence of quantum dots (QDs) combined with the specificity of Cramoll lectin, to evaluate glucose/mannose profiles on three Candida species. The QDs-Cramoll conjugates presented specificity and bright fluorescence emission. The lectin preserved its biological activity after the conjugation process mediated by adsorption interactions. The labeling of Candida species was analyzed by fluorescence microscopy and quantified by flow cytometry. Morphological analyses of yeasts labeled with QDs-Cramoll conjugates indicated that C. glabrata (2.7 µm) was smaller when compared to C. albicans (4.0 µm) and C. parapsilosis sensu stricto (3.8 µm). Also, C. parapsilosis population was heterogeneous, presenting rod-shaped blastoconidia. More than 90% of cells of the three species were labeled by conjugates. Inhibition and saturation assays indicated that C. parapsilosis had a higher content of exposed glucose/mannose than the other two species. Therefore, QDs-Cramoll conjugates demonstrated to be effective fluorescent nanoprobes for evaluation of glucose/mannose constitution on the cell walls of fungal species frequently involved in candidiasis.
Asunto(s)
Candida/química , Colorantes Fluorescentes/química , Glucosa/análisis , Lectinas/química , Manosa/análisis , Microscopía Fluorescente/métodos , Candida/crecimiento & desarrollo , Candida/aislamiento & purificación , Candida/metabolismo , Candidiasis/diagnóstico , Candidiasis/microbiología , Pared Celular/química , Pared Celular/metabolismo , Glucosa/metabolismo , Humanos , Manosa/metabolismo , Microscopía Fluorescente/instrumentación , Nanopartículas/química , Puntos Cuánticos/químicaRESUMEN
Mosquito-borne arboviruses compromise human health worldwide. Due to resistance to chemical insecticides, natural compounds have been studied to combat mosquitoes. Previous works have demonstrated a larvicidal activity of the water-soluble Moringa oleifera lectin (WSMoL) against Aedes aegypti, suggesting a mechanism of action based on the interaction between lectin and chitin present in the larvae's peritrophic matrix. In this work, it was investigated the WSMoL activity against Aedes aegypti larvae, by using luminescent bioconjugates of WSMoL conjugated to l-glutathione capped CdTe quantum dots. The conjugation was confirmed by ITC experiments, presenting high enthalpy associated to hydrogen bond interactions between nanoparticles and lectins. The bioconjugate luminescence stability was evaluated by the quantum yield (QY) at different pHs, ionic strengths and heat treatment time. The best parameters reached were pH 7.0, absence of electrolytes and heat treatment, giving QYâ¯=â¯4.4 %. The larvae were exposed to the bioconjugates and analyzed by confocal and fluorescence microscopy. CdTe-WSMol were detected along the entire midgut tract, suggesting a strong interaction with peritrophic matrix and lumen of the Aedes aegypti.
Asunto(s)
Compuestos de Cadmio/química , Glutatión/química , Larva/efectos de los fármacos , Lectinas/química , Moringa oleifera/química , Puntos Cuánticos/química , Telurio/química , Aedes/efectos de los fármacos , Animales , Biomarcadores/análisis , Calor , Enlace de Hidrógeno , Concentración de Iones de Hidrógeno , Luminiscencia , Microscopía Electrónica de Transmisión , Microscopía Fluorescente , Puntos Cuánticos/ultraestructura , Difracción de Rayos XRESUMEN
Carbohydrates perform important physiological functions in eukaryotic and prokaryotic cells. Indeed, alterations in glycan patterns may be associated with disorders. The analysis of these sugars can be reached using nanoprobes composed by lectins associated with fluorescent nanoparticles. This study reports the conjugation of a galactose-binding lectin (BmoLL) isolated from Bauhinia monandra leaves with quantum dots (QDs) by adsorption. QDs-BmoLL conjugates showed bright fluorescence and the hemagglutination assay revealed that the lectin preserved its carbohydrate-binding ability after the conjugation. To evaluate the efficiency/specificity of the bioconjugate, ABO human red blood cells (RBCs) were used as biological models and the labeling was analyzed by flow cytometry. Among ABO blood groups, higher labeling (71.7 ± 5.9%) was detected for B-type RBCs, whose antigens have galactose in their structure. The specificity of labeling was confirmed since A- and O-types RBCs incubated with QDs-BmoLL, as well as B-type cells incubated with previously galactose-inhibited conjugates, were labeled below 6%. In AB-type RBCs, which simultaneously have B and A (N-acetylgalactosamine) antigens on their membrane, the labeling was ca. 14.1 ± 4.8%. Therefore, a successful conjugation was reached and QDs-BmoLL conjugates can be considered promising fluorescent nanoprobes for biological investigations.
Asunto(s)
Bauhinia/química , Eritrocitos/química , Nanopartículas/química , Hojas de la Planta/química , Puntos Cuánticos/química , HumanosRESUMEN
A simple and inexpensive procedure used magnetite and levan to synthesize a composite recovered by a magnetic field. Lectins from Canavalia ensiformis (Con A) and Cratylia mollis (Cramoll 1 and Cramoll 1, 4) did bind specifically to composite. The magnetic property of derivative favored washing out contaminating proteins and recovery of pure lectins with glucose elution. Cramoll 1 was purified by this affinity binding procedure in two steps instead of a previous three-step protocol with ammonium sulfate fractionation, affinity chromatography on Sephadex G-75, and ion exchange chromatography through a CM-cellulose column.
Asunto(s)
Óxido Ferrosoférrico/química , Fructanos/química , Lectinas de Plantas/aislamiento & purificación , Canavalia/química , Cromatografía Liquida , Concanavalina A/aislamiento & purificación , Fabaceae/química , Fructanos/metabolismo , Pruebas de Inhibición de Hemaglutinación , Resonancia Magnética Nuclear Biomolecular , Lectinas de Plantas/metabolismo , Zymomonas/químicaRESUMEN
This work aimed to evaluate the effects of CasuL on growth and viability of 15 mastitis isolates from cows and goats, to determine the synergistic potential between CasuL and antibiotics, and to investigate the effects on bacterial ultrastructure and antibiofilm activity. The lectin inhibited the growth of Staphylococcus isolates from either bovine (Ssp6PD and Sa) or caprine (Ssp5D and Ssp01) mastitis. The minimal inhibitory concentrations were ranged from 3.75 to 15 µg/ml. Synergistic effect was observed for CasuL-tetracycline against Sa and Ssp6PD and CasuL-ampicillin against Ssp01. No structural damage was observed under the scanning electron microscope in CasuL treatments. Flow cytometry analysis using thiazol orange and propidium iodide demonstrated that CasuL was unable to reduce the cell viability of the isolates tested. At sub-inhibitory concentrations, CasuL reduced biofilm formation by the isolates Sa and Ssp5D. However, CasuL-tetracycline and CasuL-ampicillin combinations inhibited biofilm formation by Ssp6PD and Ssp01, respectively. In conclusion, CasuL is a bacteriostatic and antibiofilm agent against some mastitis isolates and displayed a synergistic potential when used in combination with either ampicillin (against one isolate) or tetracycline (against two isolates). The results stimulate the evaluation of CasuL for the treatment of mastitis, particularly when used in conjunction with antibiotics.
Asunto(s)
Antibacterianos/farmacología , Enfermedades de los Bovinos/microbiología , Fabaceae/química , Enfermedades de las Cabras/microbiología , Lectinas/farmacología , Mastitis/veterinaria , Staphylococcus/efectos de los fármacos , Animales , Antibacterianos/aislamiento & purificación , Biopelículas/efectos de los fármacos , Biopelículas/crecimiento & desarrollo , Bovinos , Sinergismo Farmacológico , Cabras , Lectinas/aislamiento & purificación , Mastitis/microbiología , Pruebas de Sensibilidad Microbiana , Viabilidad Microbiana/efectos de los fármacos , Extractos Vegetales/aislamiento & purificación , Extractos Vegetales/farmacología , Infecciones Estafilocócicas/microbiología , Infecciones Estafilocócicas/veterinaria , Staphylococcus/crecimiento & desarrollo , Staphylococcus/aislamiento & purificaciónRESUMEN
BACKGROUND: Moringa oleifera is used in traditional medicine as well as in food, cosmetic, and pharmaceutical industries. Water-soluble M. oleifera lectin (WSMoL) is an anionic protein isolated from the seeds of this tree. Until now, immune responses promoted by this lectin in human PBMC have not been investigated. OBJECTIVE: The main objective of this study was to investigate the immunomodulatory effects of WSMoL on human PBMC through measurement of lymphocytes subsets, cytokine and nitric oxide levels. METHODS: Human peripheral blood mononuclear cells (PBMC) were isolated through Ficoll technique, were incubated with WSMoL (10 µg/mL) for 24, 48 and 72 hours, and was performed immunophenotyping assay of lymphocytes and monocytes. Culture supernatants were used to determined cytokine and nitric oxide levels. Assays with cells subsets and cytokine production were performed through cytometry. Nitric oxide release assay was determinate by spectrophotometry. RESULTS: WSMoL induced the release of the cytokines TNF-α, IL-2, IL-6, IL-10 as well as nitric oxide. Incubation of PBMC with this lectin also led to activation of CD8+ T lymphocytes. CONCLUSION: WSMoL promotes immunomodulation in human PBMC inducing a potential wound healing profile and, in future in vivo assays, can be evaluated as adjuvant in immunosuppressive diseases and wound repair.
Asunto(s)
Factores Inmunológicos/farmacología , Leucocitos Mononucleares/efectos de los fármacos , Moringa oleifera/química , Lectinas de Plantas/farmacología , Semillas/química , Adulto , Supervivencia Celular , Citocinas/metabolismo , Humanos , Factores Inmunológicos/aislamiento & purificación , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/inmunología , Óxido Nítrico/metabolismo , Extractos Vegetales/aislamiento & purificación , Lectinas de Plantas/aislamiento & purificación , Solubilidad , Agua , Adulto JovenRESUMEN
In this study, Moringa oleifera flower extract and a trypsin inhibitor (MoFTI) isolated from it were evaluated for anti-protozoal activity against Trypanosoma cruzi and cytotoxicity to mammalian cells. The presence of flavonoids was remarkable in the HPLC fingerprints of the extract at 254 and 360 nm. Amino acid sequences of peptides derived from in-gel digestion of MoFTI were determined. Both the extract and MoFTI caused lysis of T. cruzi trypomastigotes with LC50/24 h of 54.18 ± 6.62 and 41.20 ± 4.28 µg/mL, respectively. High selectivity indices (7.9 to >12) for T. cruzi cells over murine peritoneal macrophages and Vero cells were found for the extract and MoFTI. The results show that MoFTI is a trypanocidal principle of the flower extract.