Edésio José Tenório de Melo

PvD1 defensin, a plant antimicrobial peptide with inhibitory activity against Leishmania amazonensis.

PvD1 was able to inhibit the proliferation of Leishmania amazonensis promastigotes; PvD1 caused cell membrane permeabilization and alterations in the cytoplasmic contents of these cells; PvD1 was internalized in these cells, what suggests a possible intracellular target.

Anti-Parasite effects of new Thiosemicarbazones and their ProductsThiazolidinone including Cellular aspects of Intracellular Elimination ofTrypanosoma Cruzi in Vitro

Trypanosoma cruzi, agent of Chagas disease in humans, invades and replicates within a wide variety of nucleated mammalian cells until the lysis of the host cells. This study was undertaken to evaluate the cellular features of effect of new compounds of thiosemicarbazone and their thiazolidinone derivate on the multiplication of extra- and intracellular T. cruzi. Most of the compounds interrupted epimastigote proliferation at 1 mM, and above 5 mM led to drastic cytoplasm reduction, nuclear condensation and death. Ultrastructural assays showed that epimastigotes treated with 1 mM of the compounds retained main parasite organelles such as the Golgi complex and the mitochondria, but underwent drastic reduction in cytoplasm volume and led to the formation of blebs on the plasma membrane, suggesting a cell death process by apoptosis. Infected cultures treated for 24 h with the compounds showed similar effects, with drastic decrease in infection and the elimination of intracellular parasite at 1 mM without toxic effects on the host cells. Intracellular amastigotes showed progressive disorganization leading to the rupture and elimination of the parasite. The results strongly suggest that in presence of thiosemicarbazones and their derivate, intracellular amastigotes arrested the proliferation, leading to irreversible ultrastructural disorganization, death, and then elimination. When the intracellular T. cruzi elimination process was analysed, it showed that autophagy was present, in intriguing and new features to futures studies. In addition, our results in vitro also suggest that these compounds are promising molecules against intracellular T. cruzi when compared to the anti-proliferative drugs Hydroxyurea and Benznidazole.

Activity of recombinant and natural defensins from Vigna unguiculata seeds against Leishmania amazonensis.

Antimicrobial peptides (AMPs), which are differentiated from other antibiotic peptides, such as gramicidins and polymyxins, because they are synthesized by large enzymatic complex and bear modified amino acids including d-amino acids, are short polymers of l-amino acids synthesized by ribosomes upon which all living organisms rely to defend themselves from invaders or competitor microorganisms. AMPs have received a great deal of attention from the scientific community as potential new drugs for neglected diseases such as Leishmaniasis. In plants, they include several families of compounds, including the plant defensins. The aim of the present study was to improve the expression of recombinant defensin from Vigna unguiculata seeds (Vu-Defr) and to test its activity against Leishmania amazonensis promatigotes. Recombinant expression was performed in LB and TB media and under different conditions. The purification of Vu-Defr was achieved by immobilized metal ion affinity and reversed-phase chromatography. The purified Vu-Defr was analyzed by circular dichroism (CD), and its biological activity was tested against L. amazonenis promastigotes. To demonstrate that the recombinant production of Vu-Defr did not interfere with its fold and biological activity, the results of all experiments were compared with the results from the natural defensin (Vu-Def). The CD spectra of both peptides presented good superimposition indicating that both peptides present very similar secondary structure and that the Vu-Defr was correctly folded. L. amazonensis treated with Vu-Defr led to the elimination of 54.3% and 46.9% of the parasites at 24 and 48h of incubation time, respectively. Vu-Def eliminated 50% and 54.8% of the parasites at 24 and 48 h, respectively. Both were used at a concentration of 100 μg/mL. These results suggested the potential for plant defensins to be used as new antiparasitic substances.

Étude de l’effet des thiosemicarbazones sur Toxoplasma gondii

Toxoplasmosis is a neglected disease, with an estimated occurrence of one-third of the population worldwide. Research in medicinal chemistry has for some years been pursuing the development of new drugs against toxoplasmosis, because current treatments cause serious side effects in the patient. The use of thiosemicarbazones as an alternative option for the treatment of various diseases has been published in recent years, due to their, among others, anticancer, antimalarial, antitrypanosomal, antibacterial, and antitoxoplasmosis activities, the latter being the subject of this study, which is based upon biological analyses and tests of the response of Toxoplasma gondii in the presence of thiosemicarbazones.

The toxic effect of Vu-Defr, a defensin from Vigna unguiculata seeds, on Leishmania amazonensis is associated with reactive oxygen species production, mitochondrial dysfunction, and plasma membrane perturbation

Plant defensins are plant antimicrobial peptides that present diverse biological activities in vitro, including the elimination of Leishmania amazonensis. Plant defensins are considered promising candidates for the development of new drugs. This protozoan genus has great epidemiological importance and the mechanism behind the protozoan death by defensins is unknown, thus, we chose L. amazonensis for this study. The aim of the work was to analyze the possible toxic mechanisms of Vu-Defr against L. amazonensis. For analyses, the antimicrobial assay was repeated as previously described, and after 24 h, an aliquot of the culture was tested for viability, membrane perturbation, mitochondrial membrane potential, reactive oxygen species (ROS) and nitric oxide (NO) inductions. The results of these analyses indicated that after interaction with L. amazonensis, the Vu-Defr causes elimination of promastigotes from culture, membrane perturbation, mitochondrial membrane collapse, and ROS induction. Our analysis demonstrated that NO is not produced after Vu-Defr and L. amazonensis interaction. In conclusion, our work strives to help to fill the gap relating to effects caused by plant defensins on protozoan and thus better understand the mechanism of action of this peptide against L. amazonensis.

In Vitro Anti-Toxoplasma gondii and Antimicrobial Activity of Amides Derived from Cinnamic Acid

Most cinnamic acids, their esters, amides, aldehydes, and alcohols present several therapeutic actions through anti-inflammatory, antitumor, and inhibitory activity against a great variety of microorganisms. In this work, eight amines derived from cinnamic acid were synthesized and tested against host cells infected with Toxoplasma gondii and the bacteria Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, and three strains of Staphylococcus aureus. Compounds 3 and 4 showed the best result against intracellular T. gondii, presenting antiparasitic activity at low concentrations (0.38 and 0.77 mM). The antibacterial activity of these compounds was also evaluated by the agar microdilution method, and amides 2 and 5 had a minimum inhibitory concentration of 250 µg mL−1 against two strains of S. aureus (ATCC 25923 and bovine strain LSA 88). These also showed synergistic action along with a variety of antibiotics, demonstrating that amines derived from cinnamic acid have potential as pharmacological agents.