Structural characterization of a galectin from the marine sponge Aplysina lactuca (ALL) with synergistic effects when associated with antibiotics against bacteria.

Lectins presents the ability to interact with glycans and trigger varied responses, including the inhibition of the development of various pathogens. Structural studies of these proteins are essential to better understand their functions. In marine sponges, so far only a few lectins have their primary structures completely determined. Thus, the objective of this work was to structurally characterize and evaluate antibacterial potential, in association with different antibiotics, of the lectin isolated from the marine sponge Aplysina lactuta (ALL). ALL is a homotetramer of 60 kDa formed by four 15 kDa-subunits. The lectin showed affinity only for the glycoproteins fetuin, asialofetuin, mucin type III, and bovine submaxillary mucin type I. The complete amino acid sequences of two isoforms of ALL, named ALL-a and ALL-b, were determined by a combination of Edman degradation and overlapped peptides sequenced by tandem mass spectrometry. ALL-a and ALL-b have 144 amino acids with molecular masses of 15,736 Da and 15,985 Da, respectively. Both structures contain conserved residues typical of the galectin family. ALL is a protein with antibacterial potential, when in association with ampicillin and oxacillin the lectin potentiates its antibiotic effect, included Methicillin-resistant Staphylococcus strains. Thus, ALL shows to be a molecule with potential for the development of new antibacterial drugs.

Chemical composition determination and evaluation of the antibacterial activity of essential oils from Ruellia asperula (Mart. Ex Ness) Lindau and Ruellia paniculata L. against oral streptococci.

This study investigated the chemical composition and evaluated the antibacterial and antibiofilm activities of essential oils (EOs) extracted from Ruellia asperula (EORA) and Ruellia paniculata (EORP) against oral streptococci. The EO constituents were analyzed by gas chromatography/mass spectrometry. The antimicrobial potential of EOs was evaluated using the minimum inhibitory concentration, minimum bactericidal concentration, and time-kill determination. Furthermore, the quantification of total biomass and the number of viable cells in the biofilms were evaluated. The major constituents of EORA were cariophylla-4(12)-8-(13)-dien-5ß-ol (14.1%), (ß)-caryophyllene (22.7%), and caryophyllene oxide (29.4%). For EORP, the major constituents were (ß)-caryophyllene (11.0%), spathulenol (13.1%), and δ-amorphene (14.9%). The tested EOs exhibited antibacterial activity against planktonic growth and biofilm formation. Thus, the EOs from R. asperula and R. paniculata prove to be promising alternatives for bacterial growth control and biofilm formation prevention of oral streptococci.

Wound healing activity of lectin isolated from seeds of Centrolobium microchaete Mart. ex Benth. on cutaneous wounds in mice.

This study aims to evaluate the wound healing potential of lectin isolated from the seeds of Centrolobium microchaete (Mart. ex Benth) (CML) on cutaneous wounds in mice. CML did not show cytotoxicity on murine dermal fibroblasts (L929 cell line). The wounds treated with CML (200 µg/mL) showed a decrease in area within 12 days post-operative (P.O.) when compared to control. On 3rd and 7th day P.O., the CML-treated group exhibited fibroblast proliferation and neovascularization. On 12th day P.O., complete restructuring of the epithelial layer and connective tissue was observed in the CML-treated group, whereas control groups exhibited incomplete reepithelialization. CML treatment enhanced the wound closure via the wound contraction process, resulting in the restructuring of the skin layers on 12th day P.O. In conclusion, CML induced a fast and efficient wound healing, suggesting that it can be used as a promising therapeutic tool to heal acute wounds.

Antihyperglycemic and antioxidant activities of a lectin from the marine red algae, Bryothamnion seaforthii, in rats with streptozotocin-induced diabetes.

The aim of the study was to assess the antihyperglycemic, antilipidemic, and antioxidant effects of a lectin isolated from Bryothamnion seaforthii (BSL), on rats with streptozotocin (STZ)-induced diabetes. The disease model was induced by low-dose injections of STZ. Diabetic rats were treated with NaCl 150 mM, metformin, and BSL at different concentrations. Blood collection was carried out at 0, 30, 60, 90, and 120 days after hyperglycemia confirmation via the assessment of seric glucose, total cholesterol, and triglycerides, assessment of the enzymatic levels of glutathione peroxidase (GPx), catalase (CAT), and superoxide dismutase (SOD), and the determination of insulin resistance by a homeostasis model of assessment-insulin resistance (HOMA-IR) as well as a homeostasis model of assessment of ß-cells resistance (HOMA-ß). The BSL-treated animals at all three concentrations showed a significant reduction in levels of glucose, cholesterol, total cholesterol, and triglycerides. Moreover, BSL increased the enzymatic activity of GPx and SOD. Index assessments of HOMA-IR and HOMA-ß confirmed that BSL treatment significantly decreased insulin resistance and ß-cell hypersecretion, respectively. In conclusion, BSL treatment might exert hypoglycemic and hypolipidemic effects, diminish insulin resistance, and ameliorate pancreatic ß-cell function along with enzymatic activities toward oxidative stress caused by diabetes mellitus type 2 (T2DM).

Mycorrhization and saline stress response in Hyptis suaveolens / Micorrização e resposta ao estresse salino em Hyptis suaveolens

ABSTRACT: The objective of this study was to evaluate the physiological and biochemical mechanisms of adaptation of Hyptis suaveolens under osmotic stress due to salinity inoculations with a mycorrhizal fungal species. H. suaveolens seeds were germinated in polyethylene pots containing a substrate associated with or without arbuscular mycorrhizal fungi (AMF). After plant formation, treatments were treated with different salt concentrations (0.0, 35, 70, and 105 mM) and fungi (control and two types of AMF), totaling 12 treatments with three replicates. The experimental design used randomized blocks in a 4 x 3 factorial scheme, totaling 12 treatments with three replicates each. Salinity affected all measured physiological and biochemical variables, and the stress reduced dry matter content. Plants associated with AMF had increased dry matter compared to non-associated plants, and there were increased biochemical and physiological responses of AMF-colonized plants in the 35 mM NaCl treatment. However, saline stress negatively affected the development of H. suaveolens; and therefore, no attenuation of fungi was observed.

RESUMO: Este estudo teve como objetivo avaliar mecanismos fisiológicos e bioquímicos de adaptação de Hyptis suaveolens submetidas ao estresse osmótico decorrente da salinidade inoculadas com espécies de fungos micorrízicos. Sementes de H. suaveolens foram colocadas para germinar em vasos de polietileno contendo substrato associado, ou não, a FMA. Após o estabelecimento das plantas, foram iniciados os tratamentos com diferentes concentrações de sais (0,0; 35; 70 e 105 mM) e fungos (controle e dois tipos de FMA), perfazendo 12 tratamentos com três repetições. O delineamento experimental foi em blocos casualizados em esquema fatorial 4 x 3, perfazendo 12 tratamentos com três repetições cada. Observou-se, através de variáveis fisiológicas e bioquímicas, que a salinidade afetou todas as variáveis analisadas e o estresse promoveu redução no conteúdo de matéria seca. Verificou-se que plantas associadas com FMA apresentaram ganho de matéria seca em relação as não associadas e incremento de respostas bioquímicas e fisiológicas das plantas colonizadas no tratamento 35mM de NaCl. No entanto, o estresse salino afeta negativamente o desenvolvimento de H. suaveolens não sendo observado, portanto, atenuação dos fungos.

Chemical composition, antioxidant, antimicrobial and antibiofilm activities of Vitex gardneriana schauer leaves's essential oil.

This study aimed to determinate the chemical composition and evaluate the antimicrobial and antioxidant activity of the essential oil obtained from leaves of V. gardneriana. The Vitex gardneriana leaves's were hydrodistilled to obtain the essential oil and the chemical composition determined by GC/MS analysis. The antimicrobial activities were determined by microdilution method. The activity of essential oil on biofilm was evaluated by quantification of total biomass and enumeration of biofilm-entrapped viable cells. The antioxidant activity was assessed by DPPH free radical assay, ferrous ion chelating assay, ferric-reducing antioxidant power and ß-carotene bleaching assay. Furthermore, the essential oil was tested on viability of health human, animal cells and the microcrustacean Artemia sp. The essential oil showed high content of sesquiterpenes and very low content of monoterpenes. Regarding activity on planktonic cells, the essential oil reduced the growth of the all species tested but showed MIC values only to S. aureus (0.31%). In general, the essential oil reduced significantly the biofilm biomass and the number of viable cells of bacteria and yeasts, mainly on biofilm formation. The essential oil showed a potential antioxidant activity, mainly on ß-carotene oxidation. Moreover, the essential oil reduced the cell viability of murine fibroblasts but not show viability reduction of human keratinocytes. Furthermore, the oil not show toxicity against the microcrustacean. Thus, the essential oil from V. gardneriana leaves may be considered as an important alternative against biofilms formed by bacteria and yeasts related to infections, as well as a natural antioxidant and non-toxic substance on human cells.

A new mucin-binding lectin from the marine sponge Aplysina fulva (AFL) exhibits antibiofilm effects.

A new mucin-binding lectin (AFL) was isolated from the marine sponge Aplysina fulva. AFL was purified by affinity chromatography on Sepharose™ matrix. Its hemagglutinating activity was independent of divalent ions, and it was weakly inhibited by simple sugars. However, porcine stomach mucin was a powerful inhibitor. In SDS PAGE, piridylethylated AFL showed one band of approximately 16 kDa, whereas in the non-reducing conditions, AFL showed at least two bands of 30 and 70 kDa. Mass spectrometry MALDI-ToF analysis showed one major ion of 31,652 ±â€¯5 Da, which corresponded to a dimer formed by subunits linked by disulfide bonds. The first fifteen amino acids of AFL were determined, and no sequence similarity was observed with any known protein. Internal sequences were obtained by mass spectrometry analysis of tryptic digestion of AFL spots. These peptides showed similarity with a lectin from marine sponge Aplysina lactuca. Secondary structure of AFL was predominantly formed by ß-conformations, which were stable at variations of pH and temperature. AFL did not inhibit planktonic growth of Gram-positive and Gram-negative bacteria tested. However, the lectin did significantly reduce the biomass biofilm of the bacteria Staphylococcus aureus, S. epidermidis, and Escherichia coli.

Antibacterial activity of a new lectin isolated from the marine sponge Chondrilla caribensis.

A new lectin from the marine sponge Chondrilla caribensis (CCL) was isolated by affinity chromatography in Sepharose 6B media. CCL is a homotetrameric protein formed by subunits of 15,445 ±2Da. The lectin showed affinity for disaccharides containing galactose and mucin. Mass spectrometric analysis revealed about 50% of amino acid sequence of CCL, which showed similarity with a lectin isolated from Aplysina lactuca. Secondary structure consisted of 10% α-helix, 74% ß-sheet/ß-turn and 16% coil, and this profile was unaltered in a broad range of pH and temperatures. CCL agglutinated Staphylococcus aureus, S epidermidis and Escherichia coli, and it was able to reduce biofilm biomass, but showed no inhibition of planktonic growth of these bacteria. CCL activity was inhibited by α-lactose, indicating that Carbohydrate Recognition Domain (CRD) of the lectin was involved in antibiofilm activity.

Purification and primary structure of a novel mannose-specific lectin from Centrolobium microchaete Mart seeds.

This study aimed to purify and characterize a novel mannose-binding lectin from the seeds of Centrolobium microchaete. Centrolobium microchaete lectin (CML) was purified by affinity chromatography in mannose-Sepharose-4B column. CML agglutinated rabbit erythrocytes and was inhibited by D-mannose, α-methyl-D-mannoside, D-glucose, N-Acetyl-D-glucosamine and sucrose. The lectin was stable at pH 7.0 and 8.0 and temperatures up to 60°C. The monomeric form of CML showed approximately 28kDa, and its native form is probably a homodimer, as determined by gel filtration chromatography. The primary structure of CML was determined by tandem mass spectrometry that showed CML as a protein with two distinct forms (isolectins CML-1 and CML-2) with 246 and 247 residues, respectively. CML-2 possesses one residue of Asn more than CML-1 in C-terminal. The primary structure of CML agrees with the molecular weights found by electrospray ionization mass spectrometry: 27,224 and 27,338Da for CML-1 and CML-2, respectively. CML is a metal-dependent glycoprotein. Moreover, the glycan composition of CML and its structure were predicted.

Effect of the lectin of Bauhinia variegata and its recombinant isoform on surgically induced skin wounds in a murine model.

Lectins are a structurally heterogeneous group of highly specific carbohydrate-binding proteins. Due to their great biotechnological potential, lectins are widely used in biomedical research. The purpose of the present study was to evaluate the healing potential of the lectin of Bauhinia variegata (nBVL) and its recombinant isoform (rBVL-1). Following surgical creation of dorsal skin wounds, seven groups of mice were submitted to topical treatment for 12 days with lectin, D-galactose, BSA and saline. The animals were anesthetized and euthanized on POD 2, 7 and 12 in order to evaluate the healing potential of each treatment. The parameters considered included wound size, contraction rate, epithelialization rate and histopathological findings. Wound closure was fastest in animals treated with rBVL-1 (POD 7). nBVL was more effective than the controls. All skin layers were reconstructed and keratin deposition increased. Our findings indicate that the lectin of Bauhinia variegata possesses pro-healing properties and may be employed in the treatment of acute skin wounds.