Browsing by Author "Vasconcelos, Andreanne G."
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- In Silico, In Vitro and In Vivo Toxicological Assessment of BPP-BrachyNH2, A Vasoactive Proline-Rich Oligopeptide from Brachycephalus ephippiumPublication . Arcanjo, Daniel D. R.; Mafud, Ana Carolina; Vasconcelos, Andreanne G.; Silva-Filho, José Couras da; Amaral, Maurício P. M.; Brito, Lucas M.; Bemquerer, Marcelo P.; Kückelhaus, Selma A. S.; Plácido, Alexandra; Delerue-Matos, Cristina; Vale, Nuno; Mascarenhas, Yvonne P.; Carvalho, Fernando Aécio A.; Oliveira, Aldeidia P.; Leite, José Roberto Souza AlmeidaBPP-BrachyNH2 is a proline-rich oligopeptide (PRO) firstly identified in skin secretion of the frog Brachycephalus ephippium, which possess in vitro inhibitory activity of angiotensin-I converting enzyme (ACE) and endothelium-dependent vasorelaxant activity. Considering its potential application in the treatment of cardiovascular diseases, the present work assessed the toxicological profile of the BPP-BrachyNH2. The in silico toxicity prediction was performed from the best model obtained through the optimization of the FASTA query peptide. This prediction study revealed that BPP-BrachyNH2 induced high predicted LD50 values for both humans and rats, and then is well-tolerated in the recommended range. The MTT assay was applied for the in vitro cytotoxic evaluation in murine macrophages. In this assay, a decrease of cell viability was not observed. The in vivo acute toxicological study was performed after the intraperitoneal administration of BPP-BrachyNH2 at doses of 5 and 50 mg/kg. After intraperitoneal administration, no death, alterations in behavioral parameters or weight gain curve was observed, as well as none in the serum biochemical parameters, and gross pathological and histopathological analyses. These observations demonstrates an acceptable safety profile for BPP-BrachyNH2, leading towards further studies focused on investigation of pharmacological and therapeutical applications for this peptide.
- Layer-by-layer films containing peptides of the Cry1Ab16 toxin from Bacillus thuringiensis for potential biotechnological applicationsPublication . Plácido, Alexandra; Farias, Emanuel Airton de Oliveira; Marani, Mariela M.; Vasconcelos, Andreanne G.; Mafud, Ana C.; Mascarenhas, Yvonne P.; Eiras, Carla; Leite, José R.S.A.; Delerue-Matos, CristinaCry1Ab16 is a toxin of crystalline insecticidal proteins that has been widely used in genetically modified organisms (GMOs) to gain resistance to pests. For the first time, in this study, peptides derived fromthe immunogenic Cry1Ab16 toxin (from Bacillus thuringiensis) were immobilized as layer-by-layer (LbL) films. Given the concern about food and environmental safety, a peptide with immunogenic potential, PcL342–354C, was selected for characterization of the electrochemical, optical, and morphological properties. The results obtained by cyclic voltammetry (CV) showed that the peptide have an irreversible oxidation process in electrolyte of 0.1 mol·L−1 potassiumphosphate buffer (PBS) at pH7.2. Itwas also observed that the electrochemical response of the peptide is governed mainly by charge transfer. In an attempt to maximize the electrochemical signal of peptide, it was intercalated with natural (agar, alginate and chitosan) or synthetic polymers (polyethylenimine (PEI) and poly(sodium 4-styrenesulfonate (PSS)). The presence of synthetic polymers on the film increased the electrochemical signal of PcL342–354C up to 100 times. Images by Atomic Force Microscopy (AFM) showed that the immobilized PcL342–354C formed self-assembled nanofibers with diameters ranging from 100 to 200 nm on the polymeric film. By UV–Visible spectroscopy (UV–Vis) it was observed that the ITO/PE/PSS/PcL342–354C film grows linearly up to the fifth layer, thereafter tending to saturation. X-ray diffraction confirmed the presence on the films of crystalline ITO and amorphous polypeptide phases. In general, the ITO/PEI/PSS/PcL342–354C film characterization proved that this systemis an excellent candidate for applications in electrochemical sensors and other biotechnological applications for GMOs and environmental indicators.
- Lycopene-rich extract from red guava ( Psidium guajava L.) displays cytotoxic effect against human breast adenocarcinoma cell line MCF-7 via an apoptotic-like pathwayPublication . Santos, Raimunda C. dos; Ombredane, Alicia S.; Souza, Jéssica Maria T.; Vasconcelos, Andreanne G.; Plácido, Alexandra; Amorim, Adriany das G.N.; Barbosa, Eder Alves; Lima, Filipe C.D.A.; Ropke, Cristina D.; Alves, Michel M.M.; Arcanjo, Daniel D.R.; Carvalho, Fernando A.A.; Delerue-Matos, Cristina; Joanitti, Graziella A.; Leite, José Roberto de S.A.This study investigated a lycopene-rich extract from red guava (LEG) for its chemical composition using spectrophotometry, mass spectrometry, attenuated total reflectance-fourier transform infrared spectroscopy (ATR-FTIR), and computational studies. The cytotoxic activity of LEG and the underlying mechanism was studied in human breast adenocarcinoma cells (MCF-7), murine fibroblast cells (NIH-3T3), BALB/c murine peritoneal macrophages, and sheep blood erythrocytes by evaluating the cell viability with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method and flow cytometry. Spectrophotometry analysis showed that LEG contained 20% of lycopene per extract dry weight. Experimental and theoretical ATR-FTIR suggests the presence of lycopene, whereas MS/MS spectra obtained after fragmentation of the molecular ion [M]+• of 536.4364 show fragment ions at m/z 269.2259, 375.3034, 444.3788, and 467.3658, corroborating the presence of lycopene mostly related to all-trans configuration. Treatment with LEG (1600 to 6.25μg/mL) for 24 and 72h significantly affected the viability of MCF-7 cells (mean half maximal inhibitory concentration [IC50]=29.85 and 5.964μg/mL, respectively) but not NIH-3T3 cells (IC50=1579 and 911.5μg/mL, respectively). Furthermore LEG at concentrations from 800 to 6.25μg/mL presented low cytotoxicity against BALB/c peritoneal macrophages (IC50≥800μg/mL) and no hemolytic activity. LEG (400 and 800μg/mL) caused reduction in the cell proliferation and induced cell cycle arrest, DNA fragmentation, modifications in the mitochondrial membrane potential, and morphologic changes related to granularity and size in MCF-7 cells; however, it failed to cause any significant damage to the cell membrane or display necrosis or traditional apoptosis. In conclusion, LEG was able to induce cytostatic and cytotoxic effects on breast cancer cells probably via induction of an apoptotic-like pathway.
- Structure-function studies of BPP-BrachyNH 2 and synthetic analogues thereof with Angiotensin I-Converting EnzymePublication . Arcanjo, Daniel D.R.; Vasconcelos, Andreanne G.; Nascimento, Lucas A.; Mafud, Ana Carolina; Plácido, Alexandra; Alves, Michel M.M.; Delerue-Matos, Cristina; Bemquerer, Marcelo P.; Vale, Nuno; Gomes, Paula; Oliveira, Eduardo B.; Lima, Francisco C.A.; Mascarenhas, Yvonne P.; Carvalho, Fernando Aécio A.; Simonsen, Ulf; Ramos, Ricardo M.; Leite, José Roberto S.A.The vasoactive proline-rich oligopeptide termed BPP-BrachyNH2 (H-WPPPKVSP-NH2) induces in vitro inhibitory activity of angiotensin I-converting enzyme (ACE) in rat blood serum. In the present study, the removal of N-terminal tryptophan or C-terminal proline from BPP-BrachyNH2 was investigated in order to predict which structural components are important or required for interaction with ACE. Furthermore, the toxicological profile was assessed by in silico prediction and in vitro MTT assay. Two BPP-BrachyNH2 analogues (des-Trp1-BPP-BrachyNH2 and des-Pro8-BPP-BrachyNH2) were synthesized, and in vitro and in silico ACE inhibitory activity and toxicological profile were assessed. The des-Trp1-BPP-BrachyNH2 and des-Pro8-BPP-BrachyNH2 were respectively 3.2- and 29.5-fold less active than the BPP-BrachyNH2-induced ACE inhibitory activity. Molecular Dynamic and Molecular Mechanics Poisson-Boltzmann Surface Area simulations (MM-PBSA) demonstrated that the ACE/BBP-BrachyNH2 complex showed lower binding and van der Wall energies than the ACE/des-Pro8-BPP-BrachyNH2 complex, therefore having better stability. The removal of the N-terminal tryptophan increased the in silico predicted toxicological effects and cytotoxicity when compared with BPP-BrachyNH2 or des-Pro8-BPP-BrachyNH2. Otherwise, des-Pro8-BPP-BrachyNH2 was 190-fold less cytotoxic than BPP-BrachyNH2. Thus, the removal of C-terminal proline residue was able to markedly decrease both the BPP-BrachyNH2-induced ACE inhibitory and cytotoxic effects assessed by in vitro and in silico approaches. In conclusion, the aminoacid sequence of BPP-BrachyNH2 is essential for its ACE inhibitory activity and associated with an acceptable toxicological profile. The perspective of the interactions of BPP-BrachyNH2 with ACE found in the present study can be used for development of drugs with differential therapeutic profile than current ACE inhibitors.