RESUMO
Antioxidants and diets supplemented with foods high in oxygen radical absorbance capacity (ORAC) reverse age-related decreases in cerebellar beta-adrenergic receptor function. We examined whether this effect was related to the antioxidant capacity of the food supplement and whether an antioxidant-rich diet reduced the levels of proinflammatory cytokines in the cerebellum. Aged male Fischer 344 rats were given apple (5 mg dry weight), spirulina (5 mg), or cucumber (5 mg) either in 0.5 ml water by oral gavage or supplied in the rat chow daily for 14 d. Electrophysiologic techniques revealed a significant decrease in beta-adrenergic receptor function in aged control rats. Spirulina reversed this effect. Apple (a food with intermediate ORAC) had an intermediate effect on cerebellar beta-adrenergic receptor physiology, and cucumber (low ORAC) had no effect, indicating that the reversal of beta-adrenergic receptor function decreases might be related to the ORAC dose. The mRNA of the proinflammatory cytokines tumor necrosis factor-alpha (TNFalpha) and TNFbeta was also examined. RNase protection assays revealed increased levels of these cytokines in the aged cerebellum. Spirulina and apple significantly downregulated this age-related increase in proinflammatory cytokines, whereas cucumber had no effect, suggesting that one mechanism by which these diets work is by modulation of an age-related increase in inflammatory responses. Malondialdehyde (MDA) was measured as a marker of oxidative damage. Apple and spirulina but not cucumber decreased MDA levels in the aged rats. In summary, the improved beta-adrenergic receptor function in aged rats induced by diets rich in antioxidants is related to the ORAC dose, and these diets reduce proinflammatory cytokine levels.
Assuntos
Envelhecimento/efeitos dos fármacos , Antioxidantes/farmacologia , Cerebelo/efeitos dos fármacos , Citocinas/metabolismo , Suplementos Nutricionais , Receptores Adrenérgicos beta/metabolismo , Envelhecimento/sangue , Envelhecimento/fisiologia , Animais , Proteínas de Bactérias , Cerebelo/metabolismo , Cucumis sativus , Citocinas/genética , Eletrofisiologia , Inflamação/dietoterapia , Inflamação/metabolismo , Iontoforese , Masculino , Malondialdeído/sangue , Malus , Modelos Animais , Inibição Neural/efeitos dos fármacos , Norepinefrina/sangue , Fitoterapia , Células de Purkinje/efeitos dos fármacos , Células de Purkinje/fisiologia , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos F344 , Receptores Adrenérgicos beta/efeitos dos fármacos , Spirulina , Ácido gama-Aminobutírico/administração & dosagemRESUMO
We have previously reported that Bcl-2 is up-regulated in the CNS of aged F344 rats as a consequence of oxidative stress. In addition to increased levels of expression, we now report that there is a subcellular redistribution of Bcl-2 in the CNS of aged F344 rats. Using western blotting, we found Bcl-2 predominantly located in the cytosol of young rats. However, in aged rats Bcl-2 was found primarily in the nucleus. This distribution, in the hippocampus and cerebellum, was reversed by treatment with the nitrone spin trap N-tert-butyl-alpha-phenylnitrone (PBN). Paradoxically, PBN treatment in young rats had the opposite effect, changing Bcl-2 from predominantly cytosolic to nuclear. We also detected an increase in Bax in aged hippocampal samples (both nuclear and cytosolic), which was reversed by treatment with PBN. The distribution of Bcl-2 and Bax in the cytosol of aged rats dramatically decreased the Bcl-2/Bax ratio, a probable indicator of neuronal vulnerability, which was restored upon treatment with PBN. In order to assess the effect of nuclear association of Bcl-2 we used PC12 cells stably transfected with a Bcl-2 construct to which we added the nuclear localization sequence of the SV40 large T antigen to the N-terminus which resulted in nuclear targeting of Bcl-2. Measurement of cell death using lactate dehydrogenase assays showed that, contrary to wild-type Bcl-2, Bcl-2 localized to the nucleus was not effective in protecting cells from treatment with 250 microm H2O2. These results suggest that nuclear localization of Bcl-2 observed in the aged CNS may not reflect a protective mechanism against oxidative stress, a major component of age-associated CNS impairments.