Antimicrobial and Antibiofilm Activity of Copper-Based Metallic Compounds Against Bacteria Related with Healthcare-Associated Infections.

Health care-associated infections (HAIs) contribute to a significant rate of morbidity, mortality, and financial burden on health systems. These infections are caused by multidrug-resistant bacteria that produce biofilm as the main virulence factor. This study aimed to evaluate the effect of the copper-based metallic compounds [Cu(phen)(pz)NO2]Cl (I), [Cu(bpy)(pz)(NO2)]Cl (II), and [Cu(phen)(INA)NO2]Cl (III), where phen = phenanthroline, bpy = bipyridine, pz = pyrazinamide, and INA = isonicotinic acid, against planktonic cells and biofilms formation of Staphylococcus aureus, Staphylococcus epidermidis, and Escherichia coli. The susceptibility of the microorganisms was evaluated by minimum inhibitory concentration (MIC), minimum bacterial concentration (MBC), and time-kill curve assay on planktonic cells. The biofilm formation was evaluated by biomass quantification through staining with crystal violet (CV), colony-forming units (CFUs) quantification, and biofilm metabolic activity determination by XTT assay. The compounds showed bacteriostatic and bactericidal activity on all microorganisms analyzed. Regarding the antibiofilm activity, all metallic compounds were able to reduce significantly the biofilm biomass, colony-forming units, and the metabolic activity of remaining cells, varying the efficient concentration according to the strain analyzed. Interestingly, compounds (I), (II) and (III) did not exhibit DNA degradation activity even with up to 100 µM of these metal complexes. On the other hand, complexes (I) and (III) showed a remarkable capacity to cleave DNA upon addition of glutathione, a reducing agent (CuII/CuI) that leads to reactive oxygen species (ROS) formation. The results presented in this study showed promising antimicrobial and antibiofilm effects.

New lectin isolated from the tropical sponge Haliclona (Reniera) implexiformis (Hechtel, 1965) shows antibiofilm effect.

A lectin from the marine sponge Haliclona (Reniera) implexiformis (HiL) was isolated by affinity chromatography on Sepharose™ matrix. HiL showed specificity for galactose and its derivatives. The glycoproteins porcine stomach mucin (PSM) and bovine stomach mucin (BSM) were potent inhibitors. Hemagglutinating activity of the lectin was maximal between pH 5.0 and 9.0. The lectin remained active until 60°C. The presence of CaCl2 and EDTA did not affect the hemagglutinating activity. In SDS-PAGE, HiL showed a single band of 20 kDa under reduced conditions, whereas in the non-reducing conditions, it showed a band of 20 kDa and one additional band of 36 kDa. The average molecular mass determined by Electrospray Ionization Mass Spectrometry (ESI-MS) was 35.874 ± 2 Da in native and non-reducing conditions, whereas carboxyamidomethylated-lectin showed 18,111 Da. These data indicated that HiL consists in a dimer formed by identical subunits linked by disulfide bonds. Partial amino acid sequence of HiL was determined by mass spectrometry, and revealed that it is a new type of lectin, which showed no similarity with any protein. Secondary structure consisted of 6% α-helice, 31% ß-sheet, 18% ß-turn and 45% random coil. HiL showed significant reduction in the number of viable cells of Staphylococcus biofilms.

Purification, biochemical characterization of a lectin from marine sponge Ircinia strobilina and its effect on the inhibition of bacterial biofilms.

A new lectin from marine sponge Ircinia strobilina, denominated IsL, was isolated by combination of affinity chromatography in Guar gum matrix followed by size exclusion chromatography. IsL was able to agglutinate native and enzymatically treated rabbit erythrocytes, being inhibited by galactosides, such as α-methyl-D-galactopyranoside, ß-methyl-D-galactopyranoside and α-lactose. IsL hemagglutinating activity was stable at neutral to alkaline pH, however the lectin loses its activity at 40° C. The molecular mass determinated by mass spectrometry was 13.655 ± 5 Da. Approximately 40% of the primary structure of IsL was determined by mass spectrometry, but no similarity was observed with any protein. The secondary structure of IsL consists of 28% α-helix, 26% ß-sheet, and 46% random region, as determined by dichroism circular. IsL was a calcium-dependent lectin, but no significant variations were observed by circular dichroism when IsL was incubated in presence of calcium and EDTA. IsL was not toxic against Artemia nauplii and did not have antimicrobial activity against bacterial cells. However, the IsL was able to significantly inhibit the biofilm formation of Staphylococcus aureus and Staphylococcus epidermidis.

An unusual bidentate methionine ruthenium(II) complex: photo-uncaging and antimicrobial activity.

The cis-[Ru(bpy)2(Met)](PF6)2 complex, where Met = L-methionine and bpy = 2,2'-bipyridine, was prepared and fully characterized. This complex was subjected to blue and green light photolysis (453 and 505 nm, respectively) in aqueous solution, leading to the release of methionine and formation of the cis-[Ru(bpy)2(H2O)2]2+ ion. This latter photoproduct was shown to subsequently interact with DNA, while DNA photocleavage was noticed. In agreement with these reactivities, this compound exhibited an exciting antibacterial action, particularly against Gram-positive bacteria Staphylococcus aureus and Staphylococcus epidermidis, which was enhanced upon blue light irradiation. Altogether, these results showed that our strategy was successful in producing light-triggered DNA-binding agents with pharmacological potential and a likely blocking reagent for efficient peptide chemistry formation.

Evaluation of the antimicrobial and antioxidant activity of 7-hydroxy-4', 6-dimethoxy-isoflavone and essential oil from Myroxylon peruiferum L.f.

This study evaluated the antibacterial, antifungal, and antioxidant effect of 7-hydroxy-4',6-dimethoxy-isoflavone and essential oil of Myroxylon peruiferum. The compound was isolated and its structure elucidated by NMR. The chemical composition of essential oil determined by GC-MS analysis. To evaluation of antimicrobial activity, the Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC) and Minimum Fungicidal Concentration (MFC) were performed. In addition to analysis of antioxidant activity, DPPH radical scavenging tests, iron chelating assay (FIC), antioxidant reducing power assay (FRAP) and ß-carotene bleaching assay (BCB) were performed. For the essential oil were identified 24 organized compounds having as main constituents; Germacrene D (17.2%), α-pinene (14.8%) and E-caryophyllene (10.8%). The results showed that isoflavone (2000 to 156 µg/mL) and essential oil (5.0 to 1.25%) present antibacterial and antifungal activity against Gram-positive bacteria and filamentous fungi. The isoflavone and the essential oil also presented antioxidant activity in all the tests, mainly on inhibition of the oxidation of ß-carotene test concentrations ranging from 60 to 100%. In conclusion, isoflavone and essential oil from M. peruiferum present an antimicrobial alternative against Gram-positive bacteria, especially of the genus Staphylococcus and dermatophyte fungi of the genus Trichophyton, as well as a natural compound antioxidant.

Aryl-Substituted Ruthenium(II) Complexes: A Strategy for Enhanced Photocleavage and Efficient DNA Binding.

Ruthenium polypyridine complexes have shown promise as agents for photodynamic therapy (PDT) and tools for molecular biology (chromophore-assisted light inactivation). To accomplish these tasks, it is important to have at least target selectivity and great reactive oxygen species (ROS) photogeneration: two properties that are not easily found in the same molecule. To prepare such new agents, we synthesized two new ruthenium complexes that combine an efficient DNA binding moiety (dppz ligand) together with naphthyl-modified (1) and anthracenyl-modified (2) bipyridine as a strong ROS generator bound to a ruthenium complex. The compounds were fully characterized and their photophysical and photochemical properties investigated. Compound 2 showed one of the highest quantum yields for singlet oxygen production ever reported (ΦΔ= 0.96), along with very high DNA binding (log Kb = 6.78). Such photochemical behavior could be ascribed to the lower triplet state involving the anthracenyl-modified bipyridine, which is associated with easier oxygen quenching. In addition, the compounds exhibited moderate selectivity toward G-quadruplex DNA and binding to the minor groove of DNA, most likely driven by the pendant ligands. Interestingly, they also showed DNA photocleavage activity even upon exposure to a yellow light-emitting diode (LED). Regarding their biological activity, the compounds exhibited an exciting antibacterial action, particularly against Gram-positive bacteria, which was enhanced upon blue LED irradiation. Altogether, these results showed that our strategy succeeded in producing light-triggered DNA binding agents with pharmacological and biotechnological potential.

ConBr, a lectin from Canavalia brasiliensis seeds, modulates signaling pathways and increases BDNF expression probably via a glycosylated target.

In the central nervous system, many receptors, ion channels and neurotransmitter transporters are glycoproteins, where the glycan chains are modulator elements. Lectins are proteins, which recognize and bind carbohydrate complexes. We have previously shown that ConBr, a lectin purified from Canavalia brasiliensis seeds, produced antidepressant-like effect and blocked hippocampal neurotoxicity induced by quinolinic acid and glutamate. Noteworthy, all these effects occurred in a dependence of its carbohydrate recognition domain. Therefore, the present study was undertaken in order to elucidate intracellular signaling pathways regulated by ConBr that may be potentially associated with the antidepressant and neuroprotective effects previously reported to be dependent on carbohydrate interaction. ConBr (10 µg/site) was injected into the ventricle (i.c.v.) of mice, and the hippocampi were removed 0.5, 1, 3, 6, 8, 12, 18, and 24 h after treatment. Our results showed that in the period of 0.5-3 h, ConBr induced activation of the protein kinases Akt, ERK1, and PKA. Furthermore, the phosphorylation of CREB-Ser133 was stimulated by ConBr (1-6 h), while brain-derived neurotrophic factor (BDNF) mRNA was increased at 12 h and BDNF protein at 18-24 h. Our data suggest that an early activation of protein kinases may trigger CREB-dependent BDNF transcription, resulting in a subsequent increase of BDNF protein in response to ConBr. Later, increment of Akt phosphorylation was observed 24 h after ConBr administration, possibly due to BDNF/TrkB-dependent activation of Akt. Our findings indicate that ConBr is a multifunctional molecule capable to activate signaling pathways involved in neuroplasticity and neuroprotection.

Glucose-Binding Dioclea bicolor Lectin (DBL): Purification, Characterization, Structural Analysis, and Antibacterial Properties.

In this study, we purified a lectin isolated from the seeds of Dioclea bicolor (DBL) via affinity purification. Electrophoresis analysis revealed that DBL had three bands, α, ß, and γ chains, with molecular masses of approximately 29, 14, and 12 kDa, respectively. Gel filtration chromatography revealed that the native form of DBL had a molecular mass of approximately 100 kDa, indicating that it is a tetramer. Interestingly, DBL-induced hemagglutination was inhibited by several glucosides, mannosides, ampicillin, and tetracycline with minimum inhibitory concentration (MIC) values of 1.56-50 mM. Analysis of the complete amino acid sequence of DBL revealed the presence of 237 amino acids with high similarity to other Diocleinae lectins. Circular dichroism showed the prominent ß-sheet secondary structure of DBL. Furthermore, DBL structure prediction revealed a Discrete Optimized Protein Energy (DOPE) score of -26,642.69141/Normalized DOPE score of -1.84041. The DBL monomer was found to consist a ß-sandwich based on its 3D structure. Molecular docking showed the interactions between DBL and α-D-glucose, N-acetyl-D-glucosamine, α-D-mannose, α-methyl-D-mannoside, ampicillin, and tetracycline. In addition, DBL showed antimicrobial activity with an MIC of 125 µg/mL and exerted synergistic effects in combination with ampicillin and tetracycline (fractional inhibitory concentration index ≤ 0.5). Additionally, DBL significantly inhibited biofilm formation and showed no toxicity in murine fibroblasts (p < 0.05). These results suggest that DBL exhibits antimicrobial activity and works synergistically with antibiotics.

Purification and primary structure of a mannose/glucose-binding lectin from Parkia biglobosa Jacq. seeds with antinociceptive and anti-inflammatory properties.

Parkia biglobosa (subfamily Mimosoideae), a typical tree from African savannas, possess a seed lectin that was purified by combination of ammonium sulfate precipitation and affinity chromatography on a Sephadex G-100 column. The P. biglobosa lectin (PBL) strongly agglutinated rabbit erythrocytes, an effect that was inhibited by d-mannose and d-glucose-derived sugars, especially α-methyl-d-mannopyranoside and N-acetyl-d-glucosamine. The hemagglutinating activity of PBL was maintained after incubation at a wide range of temperature and pH and also was independent of divalent cations. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, PBL exhibited an electrophoretic profile consisting of a single band with apparent molecular mass of 45 kDa. An analysis using electrospray ionization-mass spectrometry indicated that purified lectin possesses a molecular average mass of 47 562 ± 4 Da, and the analysis by gel filtration showed that PBL is a dimer in solution. The complete amino acid sequence of PBL, as determined using tandem mass spectrometry, consists of 443 amino acid residues. PBL is composed of a single non-glycosylated polypeptide chain of three tandemly arranged jacalin-related domains. Sequence heterogeneity was found in six positions, indicating that the PBL preparations contain highly homologous isolectins. PBL showed important antinociceptive activity associated to the inhibition of inflammatory process.

Purification and partial characterization of a new mannose/glucose-specific lectin from Dialium guineense Willd seeds that exhibits toxic effect.

A new mannose/glucose-specific lectin, named DigL, was purified from seeds of Dialium guineense by a single step using a Sepharose 4b-Mannose affinity chromatography column. DigL strongly agglutinated rabbit erythrocytes and was inhibited by d-mannose, d-glucose, and derived sugars, especially α-methyl-d-mannopyranoside and N-acetyl-d-glucosamine. DigL has been shown to be a stable protein, maintaining its hemagglutinating activity after incubation at a wide range of temperature and pH values and after incubation with EDTA. DigL is a glycoprotein composite by approximately 2.9% of carbohydrates by weight. By sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis, the purified DigL exhibited an electrophoretic profile consisting of a broad band of 28-30 kDa. Analysis using electrospray ionization mass spectrometry indicated that purified DigL possesses a molecular average mass of 28 452 ± 2 Da and shows the presence of possible glycoforms. In addition, DigL exhibited an intermediary toxic effect on Artemia sp. nauplii, and this effect was both dependent on native structure and mediated by a carbohydrate-binding site.

ConBr, a lectin from Canavalia brasiliensis seeds, protects against quinolinic acid-induced seizures in mice.

Lectins are proteins capable of reversible binding to carbohydrates or glycoconjugates. In the central nervous system of mammals, lectins with affinity for mannose/glucose or galactose can modulate cellular communication. ConBr, a lectin isolated from the seeds of Canavalia brasiliensis, previously showed antidepressant effect in the forced swimming test in mice, with involvement of the monoaminergic system. In this study, we investigated the neuroprotective effects of ConBr against quinolinic acid (QA), a well-known NMDA agonist that produces severe neurotoxicity when administered in vivo. ConBr (10 µg/site) administered via intracerebroventricular (i.c.v.) showed a neuroprotective activity against seizures induced by QA (36.8 nmol/site; i.c.v.) when administered 15 min prior to QA, with a percentage of protection around 50%. ConBr was also able to significantly decrease the severity of the seizures but without changes in the latency of the first convulsion or the duration of the seizures. This effect was dependent on the structural integrity of the ConBr protein and its binding capacity to oligosaccharides residues. ConA, a lectin with high similarity to ConBr, did not reverse the QA-induced seizures. Moreover, ConBr was able to protect against hippocampal cell death caused by QA, which was measured by propidium iodide incorporation. QA caused activation of JNK2 and improved the phosphorylation of Ser831 and 845 on the AMPA receptor GluR1 subunit, and both of these effects were counteracted by ConBr. Our data suggest that the lectin ConBr may exert a modulatory action on NMDA receptors, which inhibits its activity in response to QA.

Expression, purification and structural analysis of recombinant rBdh-2His6, a spermadhesin from buck (Capra hircus) seminal plasma.

Spermadhesins, a family of secretory proteins from the male genital tract of ungulate species, belong to the group of animal lectins. Spermadhesins have a prominent role in different aspects of fertilisation, such as spermatozoid capacitation, acrosomal stabilisation, sperm-oviduct interaction and during sperm-oocyte fusion. Proteins (spermadhesins) in buck seminal plasma were described. In the present study, bodhesin Bdh-2 cDNA present in buck seminal plasma was subcloned with the expression plasmid pTrcHis TOPO used to transform Escherichia coli Top10 One shot cells. The recombinant clones were selected by growth in 50 µg mL⁻¹ ampicillin-containing LB broth and polymerase chain reaction amplification. Recombinant rBdh-2His6 synthesis was monitored by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and followed by immunoblotting using monoclonal anti-His antibody. Production of rBdh-2 using low temperatures was not satisfactory. Greater production of rBdh-2 occurred with 1.5mM isopropyl ßd-thiogalactoside after 2h of induction. The method used to purify rBdh-2 was affinity chromatography on a His-Trap column following ion-exchange chromatography on a DEAE-Sephacel column. The secondary structure of the rBdh-2His6 was evaluated by spectral profile circular dichroism (CD). The prevalence of secondary structures like ß-sheets, with fewer unfolded structures and α-helices, was confirmed. The structure of rBdh-2His6 remained stable up to 35°C. However, significant structural changes were observed at temperatures higher than 40 °C related to a distortion of the CD spectrum.

Pharmacological potential of the triterpene 3ß,6ß,16ß-trihidroxilup-20 (29)-ene isolated from Combretum leprosum: A literature review.

Globally, plant-derived medicines have been playing an increasing and relevant role in the treatment of several diseases, thus fostering the search for new bioactive substances. Among the various families of plants studied, those of the Combretum genus can be highlighted since they are widely used in folk medicine for the treatment of hepatitis, malaria, respiratory infections, cancer, skin hemorrhage, and anxiety. Phytochemical studies carried out on species of the Combretum genus demonstrated the presence of several classes of bioactive chemical compounds, including the triterpene 3ß,6ß,16ß-trihydroxilup-20(29)-ene (CLF-1). In this perspective, the objective of this review was to gather all pharmacological activities attributed to the CLF-1 triterpene, highlighting its importance for the pharmaceutical industry. The research was performed in scientific databases such as PubMed, SciELO, LILACS, SciFinder and Science Direct. The literature indicates a great pharmacological potential of CLF-1, evidencing its antioxidant, anti-inflammatory, antiviral, antiparasitic, antinociceptive, healing, and antibacterial action, antinociceptive and antitumor effect. Therefore, based on the different research above, it is plausible to consider CLF-1, obtained from different parts of the C. leprosum plant, as a molecule with biotechnological potential that may contribute to the development of new drugs and, consequently, in the treatment of various human pathologies.

Antiproliferative activity on Trypanosoma cruzi (Y strain) of the triterpene 3ß,6ß,16ß-trihidroxilup-20 (29)-ene isolated from Combretum leprosum.

Chagas disease infects approximately seven million people worldwide. Benznidazole is effective only in the acute phase of the disease, with an average cure rate of 80% between acute and recent cases. Therefore, there is an urgent need to find new bioactive substances that can be effective against parasites without causing so many complications to the host. In this study, the triterpene 3ß-6ß-16ß-trihydroxilup-20 (29)-ene (CLF-1) was isolated from Combretum leprosum, and its molecular structure was determined by NMR and infrared spectroscopy. The CLF-1 was also evaluated in vitro and in silico as potential trypanocidal agent against epimastigote and trypomastigote forms of Trypanosoma cruzi (Y strain). The CLF-1 demonstrated good results highlighted by lower IC50 (76.0 ± 8.72 µM, 75.1 ± 11.0 µM, and 70.3 ± 45.4 µM) for epimastigotes at 24, 48 and 72 h, and LC50 (71.6 ± 11.6 µM) for trypomastigotes forms. The molecular docking study shows that the CLF-1 was able to interact with important TcGAPDH residues, suggesting that this natural compound may preferentially exert its effect by compromising the glycolytic pathway in T. cruzi. The ADMET study together with the MTT results indicated that the CLF-1 is well-absorbed in the intestine and has low toxicity. Thus, this work adds new evidence that CLF-1 can potentially be used as a candidate for the development of new options for the treatment of Chagas disease.Communicated by Ramaswamy H. Sarma.

Antinociceptive Effect of the Essential Oil of Schinus terebinthifolius (female) Leaves on Adult Zebrafish (Danio rerio).

Schinus terebinthifolius Raddi (Anacardiaceae) is popularly known in Brazil as aroeira-da-praia and has pharmacological use as an astringent, antidiarrheal, anti-inflammatory, depurative, diuretic, and antifebrile agent. Although the neuropathic antinociceptive potential of S. terebinthifolius fruits has already been investigated, this study is the first one to analyze the acute antinociceptive effect of the essential oil of S. terebinthifolius (female) leaves (EOFSt) on adult zebrafish. EOFSt was submitted to antioxidant activity evaluation by two methods (ferrous ion-chelating capacity [FIC] and ß-carotene). The animals (n = 6/group) were treated orally (20 µL) with EOFSt (0.1, 0.5, or 1.0 mg/mL) or vehicle (0.9% sodium chloride [NaCl]; 20 µL), and submitted to nociception (formalin, cinnamaldehyde, capsaicin, glutamate, acidic saline, and hypertonic saline). Possible neuromodulation mechanisms, as well motor alterations and toxicity were also evaluated. In the FIC assay, EOFSt showed ferrous ion-chelating capacity in ∼40% to 90%. Regarding the ß-carotene bleaching assay, EOFSt showed inhibition in a 58% to 80% range. Oral administration of EOFSt showed no acute toxicity and did not alter the locomotor system of aZF, and reduced the nociceptive behavior in all tested models. These effects of EOFSt were significantly similar to those of morphine, used as a positive control. The antinociceptive effect of EOFSt was inhibited by naloxone, L-NAME, ketamine, camphor, ruthenium red, and amiloride. The antinociceptive effect of the EOFSt cornea was inhibited by capsazepine. EOFSt has the pharmacological potential for acute pain treatment and this effect is modulated by the opioid system, NMDA receptors, and transient receptor potential ankyrin 1 (TRPA1), transient receptor potential vanilloid 1 (TRPV1), and acid-sensing ion channels. The EOFSt also has the pharmacological potential for corneal pain treatment and this effect is modulated by the TRPV1 channel.

Halilectin-3, a Lectin from the Marine Sponge Haliclona caerulea, Induces Apoptosis and Autophagy in Human Breast Cancer MCF7 Cells Through Caspase-9 Pathway and LC3-II Protein Expression.

BACKGROUND: An ideal strategy for cancer treatment is the specific induction of tumor cell death, sparing normal cells. Marine sponges are rich biological reservoirs of biomolecules, especially lectins, which have attracted considerable attention due to potential biological effect on human cells. Lectins are proteins that bind specific carbohydrate signatures and some gained further interest for their capacity to bind tumor associated carbohydrates antigens and induce tumor cell apoptosis. OBJECTIVE: This study aimed to evaluate the antitumor potential of H3, a lectin, recently reported from marine sponge Haliclona caerulea on the human breast cancer cell line MCF7. RESULTS: H3 reduced MCF7 cell viability with an IC50 of 100 µg/ml, without a significant effect on normal cells. At 24 h, H3 induced a significant arrest in the G1 cell cycle phase. Consistently, almost 50% of the cells were in early apoptosis and showed remarkable increased expression of caspase-9 (CASP 9). H3 impaired dramatically the adhesiveness of MCF7 cells in culture. Assays conducted with Lysotracker Red probe showed increased organelle acidity, suggesting autophagic cell death, which was further supported by increased expression of microtubuleassociated protein light chain 3 (LC3) and observable conversion of LC3-I in LC3-II by western blot. CONCLUSION: The apoptotic effect of H3 may be related to a balance between apoptotic and autophagic cell death, mediated by increased expression of CASP 9 and LC3-II. To the best of our knowledge this is the first report about a sponge lectin triggering both apoptosis and autophagy in MCF7 cell.

Antidepressant-like effect of lectin from Canavalia brasiliensis (ConBr) administered centrally in mice.

This study investigates the action of the central administration of the lectins isolated from Canavalia brasiliensis seeds (ConBr) and from Canavalia ensiformes seeds, (Concanavalin A, ConA) in the forced swimming test (FST) in mice. ConBr (1-10 micro g/site, i.c.v.), but not ConA, produced a decrease in the immobility time in the FST (observed at the time points 15, 30, 60 and 120 min after the injection), without changing the locomotor activity in the open-field test. The effect of ConBr in the FST was dependent on its protein structure integrity. ConBr (0.1 micro g/site, i.c.v.) caused a potentiation of the action of fluoxetine, a selective 5-HT reuptake inhibitor. The anti-immobility effect elicited by ConBr (10 micro g/site, i.c.v.) in the FST was prevented by the pretreatment of mice with pindolol (32 mg/kg, a 5-HT(1A/1B) receptor/beta-adrenoceptor antagonist), NAN-190 (0.5 mg/kg, a 5-HT(1A) receptor antagonist), ketanserin (5 mg/kg, a 5-HT(2A/2C) receptor antagonist), sulpiride (50 mg/kg, a D(2) receptor antagonist) or yohimbine (1 mg/kg, an alpha(2)-adrenoceptor antagonist), but not with SCH 23390 (0.05 mg/kg, a D(1) receptor antagonist) or prazosin (1 mg/kg, an alpha(1)-adrenoceptor antagonist). These results indicate that the antidepressant-like effect of ConBr in the FST is dependent on its interaction with the serotoninergic (via 5-HT(1A) and 5-HT(2)), noradrenergic (via alpha(2)-adrenoceptors) and dopaminergic (via D(2) receptors) systems. Considering the presence of lectins in the brain and based on the results, it will be important to determine a possible role of endogenous lectins in the modulation of the central nervous system function.

Renal effects induced by the lectin from Vatairea macrocarpa seeds.

Lectins are glycoproteins that interact reversibly and specifically with carbohydrates. The renal effects of the galactose-binding lectin from the seeds of Vatairea macrocarpa were investigated. Isolated kidneys from Wistar rats (240-280 g) were perfused with Krebs-Henseleit solution containing 6% bovine serum albumin. The V. macrocarpa lectin (10 microg mL(-1)) increased the perfusion pressure, renal vascular resistance, urinary flow and glomerular filtration rate. However, V. macrocarpa lectin did not change the percentage sodium, potassium or chloride tubular transport. Pre-treatment with lectin-galactose complex significantly blocked the increase in perfusion pressure, renal vascular resistance, urinary flow and glomerular filtration rate. The control group showed a small amount of a proteinaceous material in the urinary space, although no alteration in the renal tubules was detected. The administration of galactose alone did not modify the functional parameters of the kidney. Kidneys perfused with V. macrocarpa lectin showed moderate deposits of a proteinaceous material in the tubules and urinary space. Those pre-treated with lectin-galactose complex had only small amount of a proteinaceous material in the urinary space. No abnormalities were seen in renal tubules. The results suggest that lectin from V. macrocarpa seeds has important effects on the carbohydrate-binding sites of the renal system, given the reversal of renal effects with the use of that specific inhibitor.

Renal alterations promoted by the lectins from Canavalia ensiformis (ConA) and Dioclea guianensis (DguiL) seeds.

The lectin from the seeds of Canavalia ensiformis (ConA) and Dioclea guianensis (DguiL) was tested upon its renal effects using the isolated perfusion rat kidney method. Both lectins (10 microg/ml) affected perfusion pressure and renal vascular resistance, but DguiL showed a much greater action than ConA. However, ConA, but not DguiL, affected potassium tubular transport.

Expression and purification of the recombinant ConBr (Canavalia brasiliensis lectin) produced in Escherichia coli cells.

ConBr, a D-glucose/D-mannose-specific lectin from Canavalia brasiliensis seeds, was produced in Escherichia coli from a (c)DNA clone subcloned to pET15b expression vector. The recombinant lectin (rConBr) was purified by one-step immobilized metal-affinity chromatography using an amino-terminal hexahistidine tag. By SDS-PAGE and Western blot, rConBr was highly pure with an apparent molecular mass of 37 kDa. N-terminal sequence analysis revealed a single sequence, confirming the identity of the expressed protein as the pre-pro-ConBr.