ABSTRACT
Seminal studies stated that bean proteins are efficient neuronal tracers with affinity for brain tissue. A low molecular weight peptide fraction (<3kDa) from Phaseolus vulgaris (PV3) was previously reported to be antioxidant, non-cytotoxic, and capable of reducing reactive oxygen species and increasing nitric oxide in cells. We evaluated the effects of PV3 (5, 50, 100, 500, and 5000 µg/kg) on behavior and the molecular routes potentially involved. Acute and chronic PV3 treatments were performed before testing Wistar rats: i) in the elevated plus-maze (EPM) to assess the anxiolytic-like effect; ii) in the open field (OF) to evaluate locomotion and exploration; and iii) for depression-like behavior in forced swimming (FS). Catecholaminergic involvement was tested using the tyrosine hydroxylases (TH) enzyme inhibitor, α-methyl-DL-tyrosine (AMPT). Brain areas of chronically treated groups were dissected to assess: i) lipid peroxidation (LPO); ii) carbonylated proteins (CP); iii) superoxide dismutase (SOD) and catalase (CAT) enzymatic activities. Neuronal nitric oxide synthases (nNOS) and argininosuccinate synthase (ASS) protein expression was evaluated by western blotting. Acute treatment with PV3 increased the frequency and time spent in the EPM open arms, suggesting anxiolysis. PV3 increased crossing episodes in the OF. These PV3 effects on anxiety and locomotion were absent in the chronically treated group. Acute and chronic PV3 treatments reduced the immobility time in the FS test, suggesting an antidepressant effect. TH inhibition by AMPT reverted acute PV3 effects. PV3 decreased LPO and CP levels and SOD and CAT activities, whereas nNOS and ASS were reduced in few brain areas. In conclusion, PV3 displayed central antioxidant actions that are concomitant to catecholaminergic-dependent anxiolytic and antidepressant effects.
ABSTRACT
About 3000 tons of beans are not used in human food due to hardening. Several studies on bean-derived bioactive peptides have shown potential to treat some diseases, including those relying on oxidative dysfunctions. We assessed the effects of peptides extracted from hardened bean Phaseolus vulgaris (PV) on reactive oxygen species (ROS) and nitric oxide (NO) production, cytotoxic and cytoprotective effects in endothelial cells, and oxidonitrergic-dependent vasodilating effects. Extract was composed by peptide fraction <3 kDa (PV3) from hardened common bean residue. PV3 sequences were obtained and analyzed with bioinformatics. Human umbilical vein endothelial cells were treated with 10, 20, 30, and 250 µg/mL PV3. Oxidative stress was provoked by 3% H2O2. Cytotoxicity and cytoprotective effects were evaluated by MTT assay, whereas, ROS and NO were quantified using DHE and DAF-FM fluorescent probes by confocal microscopy. NO- and endothelium-dependent vasodilating effects of PV3 were assessed in isolated aortic rings. We found 35 peptides with an average mass of 1.14 kDa. There were no cell deaths with 10 and 20 μg/mL PV3. PV3 at 30 μg/mL increased cell viability, while cytotoxicity was observed only with 250 μg/mL PV3. PV3 at 10 μg/mL was able to protect cells from oxidative stress. PV3 also increased NO release without causing cell death. It also reduced relative ROS production induced by H2O2. PV3 vasodilating effects relied on endothelium-dependent NO release. PV3 obtained from low-commercial-value bean displays little cytotoxicity and exerts antioxidant effects, whereas it increases endothelial NO release.
Subject(s)
Humans , Phaseolus , Peptides/pharmacology , Endothelium , Hydrogen Peroxide , Molecular Weight , Antioxidants/pharmacologyABSTRACT
Uma preparaçäo de proteínas de membrana plasmática de células tumorais obtidas por processo original de vesículaçäo de membrana celular é usada como antígeno específico do tumor. Em sistema singênico protegeu 80% dos camundongos contra inóculo tumoral. Inóculo tumoral jé estabelecido, em início de crescimento, é curado em 70 a 80% com inoculaçäo de antígeno de membrana com afjuvante, em tumores experimentais de camundongo. A imunoterapia adotiva específica, isto é, a transferência de linfócitos pré-sensibilizados, näo protegeu camundongos contra tumor já desenvolvido. A imunoquimioterapia adotiva, transferência de linfócitos de baço de animal imunizados pelo tumor + 1 dose de ciclofosfamida foi eficiente, curando 80% dos animais com tumor singênico já estabelecido, sendo que a ciclofosfamida apenas retarda temporariamente o crescimento tumoral. Este projeto näo tem aplicaçäo clínica, pois para o doente näo se dispöe de linfócitos isogênicos sensibilizados especialmente contra seu câncer. Está sendo tentada a substituiçäo de linfócitos T sensibilizados por "Interleukin-2" + ciclofosfamida. A "Interkeukin-2" é obtida in vitro por açäo de macrófago + linfócito T helper + concanavalina A. Experiência preliminares em tumores experimentais deram nítido retardamento do crecimento dos mesmos. Este produto está sendo purificado e concentrado e produzido especificamente com antígeno do próprio tumor