RESUMO
BACKGROUND: Protease inhibitors have been isolated from plants and present several biological activities, including immunomodulatory action. OBJECTIVE: This work aimed to evaluate a Moringa oleifera flower trypsin inhibitor (MoFTI) for acute toxicity in mice, hemolytic activity on mice erythrocytes and immunomodulatory effects on mice splenocytes. METHODS: The acute toxicity was evaluated using Swiss female mice that received a single dose of the vehicle control or MoFTI (300 mg/kg, i.p.). Behavioral alterations were observed 15-240 min after administration, and survival, weight gain, and water and food consumption were analyzed daily. Organ weights and hematological parameters were analyzed after 14 days. Hemolytic activity of MoFTI was tested using Swiss female mice erythrocytes. Splenocytes obtained from BALB/c mice were cultured in the absence or presence of MoFTI for the evaluation of cell viability and proliferation. Mitochondrial membrane potential (Δψm) and reactive oxygen species (ROS) levels were also determined. Furthermore, the culture supernatants were analyzed for the presence of cytokines and nitric oxide (NO). RESULTS: MoFTI did not cause death or any adverse effects on the mice except for abdominal contortions at 15-30 min after administration. MoFTI did not exhibit a significant hemolytic effect. In addition, MoFTI did not induce apoptosis or necrosis in splenocytes and had no effect on cell proliferation. Increases in cytosolic and mitochondrial ROS release, as well as Δψm reduction, were observed in MoFTI-treated cells. MoFTI was observed to induce TNF-α, IFN-γ, IL-6, IL-10, and NO release. CONCLUSION: These results contribute to the ongoing evaluation of the antitumor potential of MoFTI and its effects on other immunological targets.
Assuntos
Moringa oleifera/enzimologia , Proteínas de Plantas , Inibidores da Tripsina , Animais , Comportamento Animal/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Feminino , Flores/química , Hemólise/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Proteínas de Plantas/farmacologia , Proteínas de Plantas/toxicidade , Baço/citologia , Testes de Toxicidade Aguda , Inibidores da Tripsina/química , Inibidores da Tripsina/metabolismo , Inibidores da Tripsina/farmacologia , Inibidores da Tripsina/toxicidadeRESUMO
The biological activities and the structural arrangement of adevonin, a novel antimicrobial peptide, were investigated. The trypsin inhibitor ApTI, isolated from Adenanthera pavonina seeds, was used as a template for screening 18-amino acid peptides with predicted antimicrobial activity. Adevonin presented antimicrobial activity and minimum inhibitory concentrations (MIC) ranging from 1.86 to 7.35 µM against both Gram-positive and - negative bacterial strains. Moreover, adevonin exerted time-kill effects within 10 min and both susceptible and drug-resistant bacterial strains were affected by the peptide. In vitro and in vivo assays showed that, at MIC concentration, adevonin did not affect human fibroblasts (MRC-5) viability or Galleria mellonella survival, respectively. Hemolytic activity was observed only at high peptide concentrations. Additionally, nucleic acid efflux assays, gentian violet uptake and time-kill kinetics indicate that the antimicrobial activity of adevonin may be mediated by bacterial membrane damage. Furthermore, molecular dynamic simulation in the presence of SDS micelles and anionic membrane bilayers showed that adevonin acquired a stable α-helix secondary structure. Further studies are encouraged to better understand the mechanism of action of adevonin, as well as to investigate the anti-infective activity of this peptide.
Assuntos
Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/genética , Peptídeos Catiônicos Antimicrobianos/farmacologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacologia , Inibidores da Tripsina/farmacologia , Animais , Anti-Infecciosos/toxicidade , Peptídeos Catiônicos Antimicrobianos/toxicidade , Bioensaio , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Fabaceae/enzimologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Hemólise , Humanos , Lepidópteros/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Proteínas Recombinantes/toxicidade , Análise de Sobrevida , Inibidores da Tripsina/toxicidadeRESUMO
Soybean world production has been increasing at a rate of 5.2% per year (average yield is around 1,400 kg/ha). This production has been solely used for oil extraction and the protein meal obtained for animal rations, but lately it is being used for human consumption. Brazil, the third largest producer, has had a yearly rate of production increase of 32% in the last years. Average yields in Brazil are still low (around 1,500 kg/ha), but in experimental results, yields over 3,000 kg/ha have been obtained. Some problems needstill to be solved, such as obtention of adapted varieties, soil fertility, adequate agronomic practices, damage by insects and diseases. Protein and oil contents are highly negative correlated, they are genetically controlled and can also be influenced by environmental conditions and agronomic practices. To breed for high protein (above 48%) enhances a decrease in oil and yield, but new varieties containing 43% protein and with a good yielding capacity have been developed lately. Methionine content varies from 1.0 to 1.6% g/16g N; there is a correlation of 0.56 to 0.58 between methionine in the protein and protein in the seed. Particular attention has been given to toxic factors such as trypsin inhibitors, whose action is related to the availability or utilization of methionine; this effect, however, can be eliminated by heat.