RESUMO
The molecular mechanism by which the lipido-sterolic extract of Serenoa repens (LSESr, Permixon) affects prostate cells remains to be fully elucidated. In androgen-independent PC3 prostate cancer cells, the LSESr-induced effects on proliferation and apoptosis were evaluated by counting cells and using a FACScan cytofluorimeter. PC3 cells were stained with JC-1 dye to detect mitochondrial membrane potential. Cell membrane lipid composition was evaluated by thin layer chromatography and gas chromatographic analysis. Akt phosphorylation was analyzed by Western blotting and cellular ultrastructure through electron microscopy. LSESr (12.5 and 25 microg/ml) administration exerted a biphasic action by both inhibiting proliferation and stimulating apoptosis. After 1 h, it caused a marked reduction in the mitochondrial potential, decreased cholesterol content and modified phospholipid composition. A decrease in phosphatidylinositol-4,5-bisphosphate (PIP2) level was coupled with reduced Akt phosphorylation. After 24 h, all of these effects were restored to pre-treatment conditions; however, the saturated (SFA)/unsaturated fatty acid (UFA) ratio increased, mainly due to a significant decrease in omega 6 content. The reduction in cholesterol content could be responsible for both membrane raft disruption and redistribution of signaling complexes, allowing for a decrease of PIP2 levels, reduction of Akt phosphorylation and apoptosis induction. The decrease in omega 6 content appears to be responsible for the prolonged and more consistent increase in the apoptosis rate and inhibition of proliferation observed after 2-3 days of LSESr treatment. In conclusion, LSESr administration results in complex changes in cell membrane organization and fluidity of prostate cancer cells that have progressed to hormone-independent status.
Assuntos
Antagonistas de Androgênios/farmacologia , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Preparações de Plantas/farmacologia , Neoplasias da Próstata , Serenoa/química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral/citologia , Linhagem Celular Tumoral/efeitos dos fármacos , Membrana Celular/química , Proliferação de Células/efeitos dos fármacos , Humanos , Masculino , Lipídeos de Membrana/química , Lipídeos de Membrana/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Potencial da Membrana Mitocondrial/fisiologia , Fitoterapia , Preparações de Plantas/química , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Oxidative stress-mediated LDL modification has a key role in initiation of the atherosclerotic process. Platelets produce reactive oxidant species (ROS) upon stimulation with agonist, but it is uncertain whether they are able to oxidatively modify LDL. Human platelets taken from healthy subjects were incubated with LDL, then stimulated with collagen. Compared with unstimulated platelets, collagen-stimulated platelets induced LDL modification as shown by enhanced conjugated dienes and lysophosphatidylcholine formation, electrophoretic mobility, Apo B-100 degradation, and monocyte LDL uptake. Activated platelets also induced a marked reduction of vitamin E contained in LDL. A significant inhibition of LDL oxidation was observed in platelets treated with arachidonyl trifluomethyl ketone (AACOCF3), an inhibitor of phospholipase A2. The experiments reported above were also conducted in patients with hereditary deficiency of gp91phox, the central core of NADPH oxidase, and in patients with hypercholesterolemia. Platelets from gp91 phox-deficient patients produced a small amount of ROS and weakly modified LDL. Conversely, platelets from hypercholesterolemic patients showed enhanced ROS formation and oxidized LDL more than platelets from healthy subjects. This study provides evidence that platelets modify LDL via NADPH oxidase-mediated oxidative stress, a phenomenon that could be dependent on arachidonic acid activation. This finding suggests a role for platelets in favoring LDL accumulation within atherosclerotic plaque.
Assuntos
Plaquetas/metabolismo , Lipoproteínas LDL/metabolismo , Glicoproteínas de Membrana/metabolismo , Monócitos/metabolismo , NADPH Oxidases/metabolismo , Eletroforese em Gel de Poliacrilamida , Humanos , Hipercolesterolemia/metabolismo , Hipercolesterolemia/patologia , Lisofosfatidilcolinas/isolamento & purificação , NADPH Oxidase 2 , Oxirredução , Espécies Reativas de Oxigênio/metabolismo , Vitamina E/metabolismoAssuntos
Artérias/fisiopatologia , Isquemia/fisiopatologia , Glicoproteínas de Membrana/metabolismo , NADPH Oxidases/metabolismo , Estresse Oxidativo , Vasodilatação , Adulto , Humanos , Isquemia/metabolismo , Masculino , Glicoproteínas de Membrana/deficiência , Erros Inatos do Metabolismo/genética , Pessoa de Meia-Idade , NADPH Oxidase 2 , NADPH Oxidases/deficiência , Projetos PilotoRESUMO
Several studies demonstrated an inverse association between polyphenol intake and cardiovascular events. Platelet recruitment is an important phase of platelet activation at the site of vascular injury, but it has never been investigated whether polyphenols influence platelet recruitment. The aim of the study was to analyze in vitro whether two polyphenols, quercetin and catechin, were able to affect platelet recruitment. Platelet recruitment was reduced by NO donors and by NADPH oxidase inhibitors and was enhanced by L-NAME, an inhibitor of NO synthase. Quercetin and catechin, but not single polyphenol, significantly inhibited platelet recruitment in a concentration-dependent fashion. The formation of superoxide anion was significantly inhibited in platelets incubated with quercetin and catechin but was unaffected by a single polyphenol. Incubation of platelets with quercetin and catechin resulted in inhibition of PKC and NADPH oxidase activation. Treatment of platelets with quercetin and catechin resulted in an increase of NO and also down-regulated the expression of GpIIb/IIIa glycoprotein. This study shows that the polyphenols quercetin and catechin synergistically act in reducing platelet recruitment via inhibition of PKC-dependent NADPH oxidase activation. This effect, resulting in NO-mediated platelet glycoprotein GpIIb/IIIa down-regulation, could provide a novel mechanism through which polyphenols reduce cardiovascular disease.
Assuntos
Antioxidantes/farmacologia , Plaquetas/efeitos dos fármacos , Flavonoides/farmacologia , NADPH Oxidases/antagonistas & inibidores , Óxido Nítrico/metabolismo , Fenóis/farmacologia , Agregação Plaquetária/efeitos dos fármacos , Adulto , Plaquetas/enzimologia , Catequina/farmacologia , Ativação Enzimática , Inibidores Enzimáticos/farmacologia , Feminino , Humanos , Masculino , NADPH Oxidases/metabolismo , NG-Nitroarginina Metil Éster/farmacologia , Doadores de Óxido Nítrico/farmacologia , Estresse Oxidativo , Polifenóis , Proteína Quinase C/antagonistas & inibidores , Quercetina/farmacologia , Superóxidos/metabolismoRESUMO
Aim of this study was to analyse the relationship between the plasma levels of polyphenols and the antioxidant activity of red and white wine. Twenty healthy subjects (HS) were randomly allocated to drink 300 ml of red (n = 10) or white n = 10 wine for 15 days. Ten HS who refrained from any alcohol beverage for 15 days were used as control. Urinary PGF-2alpha-III, a marker of oxidative stress and plasma levels of polyphenols were measured. Urinary PGF-2alpha-III significantly fell in subjects taking wine with a higher percentage decrease in subjects given red wine (-38.5 +/- 6%, p < 0.001) than in those given white wine (-23.1 +/- 6%). Subjects taking red wine had higher plasma polyphenols than those taking white wine (1.9 +/- 0.6 microM versus 1.5 +/- 0.33 microM, p < 0.001). Plasma polyphenols were inversely correlated with urinary PGF2alpha (r = 0.77, p < 0.001). No changes of urinary isoprostanes were observed in subjects who refrained from wine intake. In vitro study demonstrated that only a mixture of polyphenols, all in a range corresponding to that found in human circulation, inhibited LDL oxidation and PKC-mediated NADPH oxidase activation. Such inhibitory effects were more marked using the concentrations of polyphenols detected in human circulation after red wine intake. This study shows that red wine is more antioxidant than white wine in virtue of its higher content of polyphenols, an effect that may be dependent upon a synergism among polyphenols.