New Evidence on the Antiproliferative Activity of Campomanesia Adamantium (Cambess.) O. Berg Extracts in Melanoma Lung Metastasis.

Given the importance of discovering plant species from the Brazilian Cerrado biome with anticancer potential, this study evaluated the antitumor activity of two extracts of Campomanesi adamantium fruits in in vitro and in vivo models of melanoma lung metastasis. Pulp and peel extracts (DEGPU and DEGPE, respectively) were extracted from fresh fruit using dichloromethane as a solvent. As cytotoxicity parameter, concentration values that inhibited 50% cell growth (GI50), total growth inhibition (TGI), and selectivity index (SI) were established. The melanoma lung metastasis model was obtained by injecting 5 × 105/50 µL B16-F10 cells via the tail vein of mice, which received treatment on the 15th day. Metastatic lungs were collected for fluorescence analysis with the IR-780 marker and also macro- and microscopic assessment. In vitro analyses showed that DEGPU was active in K562 (GI50 32.99; TGI 47.93) and U-251 (GI50 32.10; TGI 249.92), whereas DEGPE showed better cytotoxicity results for all tumor cell lines, but was more efficient in K562 (GI50 27.42; TGI 40.20) and U-251 (GI50 4.89; TGI 12.77). Both showed a cytocidal effect on B16F10 at the highest concentration tested, with approximately 25% (DEGPU) and 88% (DEGPE) of cell death. In vivo analyzes showed that both extracts showed significant activity in metastatic lung. Fluorescence images showed differences in intensity between groups owing to greater tumor involvement. Macro- and microscopic images showed that treatments with extracts limited tumor growth and prevented proliferation. The extracts tested have promising activity, thus requiring further research on their active compounds.

Prevention of hospital pathogen biofilm formation by antimicrobial peptide KWI18.

This study investigated the antimicrobial and antibiofilm activity of KWI18, a new synthetic peptide. KWI18 was tested against planktonic cells and Pseudomonas aeruginosa and Candida parapsilosis biofilms. Time-kill and synergism assays were performed. Sorbitol, ergosterol, lipid peroxidation, and protein oxidation assays were used to gain insight into the mechanism of action of the peptide. Toxicity was evaluated against erythrocytes and Galleria mellonella. KWI18 showed antimicrobial activity, with minimum inhibitory concentration (MIC) values ranging from 0.5 to 10 µM. KWI18 at 10 × MIC reduced P. aeruginosa and C. parapsilosis biofilm formation and cell viability. Time-kill assays revealed that KWI18 inhibited the growth of P. aeruginosa in 4 h and that of C. parapsilosis in 6 h. The mechanism of action was related to ergosterol as well as induction of oxidative damage in cells and biofilms. Furthermore, KWI18 demonstrated low toxicity to erythrocytes and G. mellonella. KWI18 proved to be an effective antibiofilm agent, opening opportunities for the development of new antimicrobials.

Rational design of mimetic peptides based on the interaction between Inga laurina inhibitor and trypsins for Spodoptera cosmioides pest control.

The interaction of Inga laurina Kunitz inhibitor with insect trypsins is an example of protein-protein interaction with potential application for the pest control. However, the crop field application of proteins as inhibitors is limited due to high production cost, the large molecular size and low environmental stability. The use of mimetic peptides that have molecular features associated with the protein inhibitor can result in a product with lower cost and higher efficiency for the agricultural application. Here, we designed mimetic peptides deriving from globular domains of ILTI that are predicted to interact with trypsin enzymes of Lepidoptera pest. Two linear peptides were identified and synthetized from the interface of interaction between trypsin-ILTI complexes. These peptides were derived due to its high-energy contribution for the biding affinity between the enzyme-protein inhibitor. The peptides showed structural stability, propensity to adopt the bound conformation also without the context of the protein, inhibitory activity of digestive trypsins and toxic effects on the S. cosmioides, indicating that they can be used as potential inhibitor for pest control.

Nonalcoholic Fatty Liver Disease Induced by High-Fat Diet in C57bl/6 Models.

Researchers have a range of animal models in which to study Nonalcoholic fatty liver disease (NAFLD). Induction of NAFLD by a high-fat diet in the C57BL/6 strain is the most widely used among mice. In this study, we review works that performed NAFLD induction by a high-fat diet using the C57BL/6 strain, focusing on experiments on the effects of lipid ingestion. Studies are initially distinguished into researches in which mice received lipids by oral gavage and studies in which lipid was added to the diet, and each of these designs has peculiarities that must be considered. Oral gavage can be stressful for animals and needs trained handlers but allows accurate control of the dose administered. The addition of oils to the diet can prevent stress caused to mice by gavage, but possible changes in the consistency, taste, and smell of the diet should be considered. Regarding the experimental design, some variables, such as animal sex, treatment time, and diet-related variables, appear to have a definite pattern. However, no pattern was found regarding the number of animals per group, age at the beginning of the experiment, time of adaptation, the substance used as a vehicle, and substance used as a control.

Novel Peptidase Kunitz Inhibitor from Platypodium elegans Seeds Is Active against Spodoptera frugiperda Larvae.

A novel Kunitz-type inhibitor from Platypodium elegans seeds (PeTI) was purified and characterized. The mass spectrometry analyses of PeTI indicated an intact mass of 19 701 Da and a partial sequence homologous to Kunitz inhibitors. PeTI was purified by ion exchange and affinity chromatographies. A complex with a 1:1 ratio was obtained only for bovine trypsin, showing a Ki = 0.16 nM. Stability studies showed that PeTI was stable over a wide range of temperature (37-80 °C) and pH (2-10). The inhibitory activity of PeTI was affected by dithiothreitol (DTT). Bioassays of PeTI on Spodoptera frugiperda showed negative effects on larval development and weight gain, besides extending the insect life cycle. The activities of digestive enzymes, trypsin and chymotrypsin, were reduced by feeding larvae with 0.2% PeTI in an artificial diet. In summary, we describe a novel Kunitz inhibitor with promising biotechnological potential for pest control.

Effect of Powdered Shells of the Snail Megalobulimus lopesi on Secondary-Intention Wound Healing in an Animal Model.

Topical administration of powdered shells of the land snail Megalobulimus lopesi was evaluated in Wistar rats for their healing activity in an excision wound model. The animals were distributed into three groups-G1 (control): no therapeutic intervention; G2 (vehicle controls): Lanette cream once daily; G3 (experimental animals): treated with powdered shells. Variables investigated were: wound area contraction, angiogenic activity, morphometric data, leukocytic inflammatory infiltrate, and total leukocyte count in peripheral blood. Thermogravimetric analysis and quantification and characterization of powdered shell proteins were also performed. Wound area on days 3, 7, and 14 was smaller in G3, besides presenting wound closure on day 21 for all these animals. Topical administration of the powdered shells also led to an increased number of vessels at the wound site, higher leukocyte counts in peripheral blood, and increased leukocytic inflammatory infiltrate. The results lend support to the southern Brazilian folk use of M. lopesi powdered shells, as shown by the enhanced secondary-intention healing achieved with their topical administration to wounds in rats. Topical administration caused inflammatory response modulation, crucial to accelerating the healing process, the chronification of which increases the risks of wound contamination by opportunistic pathogens.

Morphological and growth alterations in Vero cells transformed by cisplatin.

Cisplatin is an antineoplastic agent used to treat solid tumours, such as ovarian, testicular and bladder tumours. However, studies in vitro and in vivo have shown that cisplatin is mutagenic, genotoxic and tumorigenic in other tissues and organs. In this work, we examined the effect of cisplatin on Vero cells, a fibroblast-like cell line. The morphological characteristics were investigated using phase contrast microscopy, scanning electron microscopy and the actin cytoskeleton was labelled with fluorescein isothiocyanate-phalloidin. Cell proliferation was assessed based on the growth curve. Cultured Vero cells treated with cisplatin showed behavioural and morphological alterations associated with cellular transformation. The transformed cells grew in multilayers and formed cellular aggregates. The proliferation and morphological characteristics of the transformed cells were very different from those of control ones. Since transformed Vero cells showed several characteristics related to neoplastic growth, these cells could be a useful model for studying tumour cells in vitro.

A trypsin inhibitor from Peltophorum dubium seeds active against pest proteases and its effect on the survival of Anagasta kuehniella (Lepidoptera: Pyralidae).

A novel trypsin inhibitor was purified from the seeds of Peltophorum dubium (Spreng.). SDS-PAGE under reducing conditions showed that the inhibitor consisted of a single polypeptide chain (ca. 20 kDa). The dissociation constants of 4 x 10(-10) and 1.6 x 10(-10) M were obtained with bovine and porcine trypsin, respectively. This constant was lower (2.6 x 10(-7) M) for chymotrypsin. The inhibitory activity was stable over a wide range of temperature and pH and in the presence of DTT. The N-terminal sequence of the P. dubium inhibitor showed a high degree of homology with other Kunitz-type inhibitors. When fed to the insect Anagasta kuehniella, in an artificial diet (inhibitor concentration 1.6%), the inhibitor produced approximately 56% and delayed the development of this lepidopteran. The concentration of inhibitor in the diet necessary to cause a 50% reduction in the weight (ED50) of fourth instar larvae was approximately 1%. The action of the P. dubium trypsin inhibitor (PDTI) on A. kuehniella may involve inhibition of the trypsin-like activity present in the larval midgut, resistance of the inhibitor to digestion by midgut enzymes and bovine trypsin, and association of the inhibitor with a chitin column and chitinous structures in the peritrophic membrane and/or midgut of the insect.