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Projeto de investigação
Valorisation of Molecules Isolated from Aquatic Portuguese Amphibians
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Novel ocellatin peptides mitigate LPS-induced ROS formation and NF-kB activation in microglia and hippocampal neurons
Publication . Sousa, Nayara A.; Oliveira, Guilherme A. L.; de Oliveira, Ana Patrícia; Lopes, André Luís F.; Iles, Bruno; Nogueira, Kerolayne M.; Araújo, Thiago S. L.; Souza, Luan K. M.; Araújo, Alyne R.; Ramos-Jesus, Joilson; Plácido, Alexandra; Amaral, Constança Pais Do; Campelo, Yuri D. M.; Barbosa, Eder Alves; Portugal, Camila C.; Socodato, Renato; Lobo, Andrea; Relvas, Joao; Bemquerer, Marcelo; Eaton, Peter; Leite, José Roberto S. A.; Medeiros, Jand Venes R.
Cutaneous secretions of amphibians have bioactive compounds, such as peptides, with potential for biotechnological applications. Therefore, this study aimed to determine the primary structure and investigate peptides obtained from the cutaneous secretions of the amphibian, Leptodactylus vastus, as a source of bioactive molecules. The peptides obtained possessed the amino acid sequences, GVVDILKGAAKDLAGH and GVVDILKGAAKDLAGHLASKV, with monoisotopic masses of [M + H]± = 1563.8 Da and [M + H]± = 2062.4 Da, respectively. The molecules were characterized as peptides of the class of ocellatins and were named as Ocellatin-K1(1-16) and Ocellatin-K1(1-21). Functional analysis revealed that Ocellatin-K1(1-16) and Ocellatin-K1(1-21) showed weak antibacterial activity. However, treatment of mice with these ocellatins reduced the nitrite and malondialdehyde content. Moreover, superoxide dismutase enzymatic activity and glutathione concentration were increased in the hippocampus of mice. In addition, Ocellatin-K1(1-16) and Ocellatin-K1(1-21) were effective in impairing lipopolysaccharide (LPS)-induced reactive oxygen species (ROS) formation and NF-kB activation in living microglia. We incubated hippocampal neurons with microglial conditioned media treated with LPS and LPS in the presence of Ocellatin-K1(1-16) and Ocellatin-K1(1-21) and observed that both peptides reduced the oxidative stress in hippocampal neurons. Furthermore, these ocellatins demonstrated low cytotoxicity towards erythrocytes. These functional properties suggest possible to neuromodulatory therapeutic applications.
Neuroprotective effects on microglia and insights into the structure–activity relationship of an antioxidant peptide isolated from Pelophylax perezi
Publication . Plácido, Alexandra; Amaral, Constança Pais Do; Teixeira, Cátia; Nogueira, Ariane; Brango‐Vanegas, José; Alves Barbosa, Eder; C. Moreira, Daniel; Silva‐Carvalho, Amandda E.; da Silva, Maria da Gloria; do Nascimento Dias, Jhones; Albuquerque, Patrícia; Saldanha‐Araújo, Felipe; C. D. A. Lima, Filipe; Batagin‐Neto, Augusto; Kuckelhaus, Selma; Bessa, Lucinda J.; Freitas, Jaime; Dotto Brand, Guilherme; Santos, Nuno C.; B. Relvas, João; Gomes, Paula; S. A. Leite, José Roberto; Eaton, Peter
Tryptophyllins constitute a heterogeneous group of peptides that are one of the first classes of peptides identified from amphibian's skin secretions. Here, we report the structural characterization and antioxidant properties of a novel tryptophyllin-like peptide, named PpT-2, isolated from the Iberian green frog Pelophylax perezi. The skin secretion of P. perezi was obtained by electrical stimulation and fractionated using RP-HPLC. De novo peptide sequencing was conducted using MALDI MS/MS. The primary structure of PpT-2 (FPWLLS-NH2 ) was confirmed by Edman degradation and subsequently investigated using in silico tools. PpT-2 shared physicochemical properties with other well-known antioxidants. To test PpT-2 for antioxidant activity in vitro, the peptide was synthesized by solid phase and assessed in the chemical-based ABTS and DPPH scavenging assays. Then, a flow cytometry experiment was conducted to assess PpT-2 antioxidant activity in oxidatively challenged murine microglial cells. As predicted by the in silico analyses, PpT-2 scavenged free radicals in vitro and suppressed the generation of reactive species in PMA-stimulated BV-2 microglia cells. We further explored possible bioactivities of PpT-2 against prostate cancer cells and bacteria, against which the peptide exerted a moderate antiproliferative effect and negligible antimicrobial activity. The biocompatibility of PpT-2 was evaluated in cytotoxicity assays and in vivo toxicity with Galleria mellonella. No toxicity was detected in cells treated with up to 512 µg/ml and in G. mellonella treated with up to 40 mg/kg PpT-2. This novel peptide, PpT-2, stands as a promising peptide with potential therapeutic and biotechnological applications, mainly for the treatment/prevention of neurodegenerative disorders.
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Entidade financiadora
Fundação para a Ciência e a Tecnologia
Programa de financiamento
3599-PPCDT
Número da atribuição
PTDC/BII-BIO/31158/2017