17ß-estradiol delays 6-OHDA-induced apoptosis by acting on Nur77 translocation from the nucleus to the cytoplasm.

Nuclear receptors (Nurs) represent a large family of gene expression regulating proteins. Gathering evidence indicates an important role for Nurs as transcription factors in dopamine neurotransmission. Nur77, a member of the Nur superfamily, plays a role in mediating the effects of antiparkinsonian and neuroleptic drugs. Besides, Nur77 survival and apoptotic roles depend largely on its subcellular localization. Estrogens are known for their neuroprotective properties, as demonstrated in animal and clinical studies. However, their action on Nur77 translocation pertaining to neuroprotection has not been investigated yet. The aim of our study was to perform a kinetic study on the effect of neurotoxic 6-hydroxydopamine (6-OHDA) and 17ß-estradiol (E2) on the subcellular localization of Nur77 with reference to the modulation of apoptosis in PC12 cells. Our results demonstrate that E2 administration alone does not affect Nur77 cytoplasmic/nuclear ratio, mRNA levels, or apoptosis in PC12 cells. The neurotoxin 6-OHDA significantly enhances cytoplasmic localization of Nur77 after merely 3 h, while precipitating apoptosis. 6-OHDA also increases Nur77 transcription, which could partly explain the rise in cytoplasmic localization of the protein. Finally, treatment with both E2 and 6-OHDA delays Nur77 accumulation in the cytoplasm and delays cell death for a few hours in our cellular paradigm. Pre-treatment with E2 does not alter the increase in levels of Nur77 mRNA produced by 6-OHDA, suggesting that a raise in nuclear translocation is likely responsible for the stabilization of the cytoplasmic/nuclear ratio until 6 h. These results suggest an intriguing cooperation between E2 and Nur77 toward cellular fate guidance.

Resveratrol protects DAergic PC12 cells from high glucose-induced oxidative stress and apoptosis: effect on p53 and GRP75 localization.

Resveratrol (RESV), a polyphenolic natural compound, has long been acknowledged to have cardioprotective and antiinflammatory actions. Evidence suggests that RESV has antioxidant properties that reduce the formation of reactive oxygen species leading to oxidative stress and apoptotic death of dopaminergic (DAergic) neurons in Parkinson's disease (PD). Recent literature has recognized hyperglycemia as a cause of oxidative stress reported to be harmful for the nervous system. In this context, our study aimed (a) to evaluate the effect of RESV against high glucose (HG)-induced oxidative stress in DAergic neurons, (b) to study the antiapoptotic properties of RESV in HG condition, and c) to analyze RESV's ability to modulate p53 and GRP75, a p53 inactivator found to be under expressed in postmortem PD brains. Our results suggest that RESV protects DAergic neurons against HG-induced oxidative stress by diminishing cellular levels of superoxide anion. Moreover, RESV significantly reduces HG-induced apoptosis in DAergic cells by modulating DNA fragmentation and the expression of several genes implicated in the apoptotic cascade, such as Bax, Bcl-2, cleaved caspase-3, and cleaved PARP-1. RESV also prevents the pro-apoptotic increase of p53 in the nucleus induced by HG. Such data strengthens the correlation between hyperglycemia and neurodegeneration, while providing new insight on the high occurrence of PD in patients with diabetes. This study enlightens potent neuroprotective roles for RESV that should be considered as a nutritional recommendation for preventive and/or complementary therapies in controlling neurodegenerative complications in diabetes.

Cucurbitacin E has neuroprotective properties and autophagic modulating activities on dopaminergic neurons.

Natural molecules are under intensive study for their potential as preventive and/or adjuvant therapies for neurodegenerative disorders such as Parkinson's disease (PD). We evaluated the neuroprotective potential of cucurbitacin E (CuE), a tetracyclic triterpenoid phytosterol extracted from the Ecballium elaterium (Cucurbitaceae), using a known cellular model of PD, NGF-differentiated PC12. In our postmitotic experimental paradigm, neuronal cells were treated with the parkinsonian toxin 1-methyl-4-phenylpyridinium (MPP(+)) to provoke significant cellular damage and apoptosis or with the potent N,N-diethyldithiocarbamate (DDC) to induce superoxide (O2(•-)) production, and CuE was administered prior to and during the neurotoxic treatment. We measured cellular death and reactive oxygen species to evaluate the antioxidant and antiapoptotic properties of CuE. In addition, we analyzed cellular macroautophagy, a bulk degradation process involving the lysosomal pathway. CuE showed neuroprotective effects on MPP(+)-induced cell death. However, CuE failed to rescue neuronal cells from oxidative stress induced by MPP(+) or DDC. Microscopy and western blot data show an intriguing involvement of CuE in maintaining lysosomal distribution and decreasing autophagy flux. Altogether, these data indicate that CuE decreases neuronal death and autophagic flux in a postmitotic cellular model of PD.

Quercetin and sesamin protect dopaminergic cells from MPP+-induced neuroinflammation in a microglial (N9)-neuronal (PC12) coculture system.

A growing body of evidence indicates that the majority of Parkinson's disease (PD) cases are associated with microglia activation with resultant elevation of various inflammatory mediators and neuroinflammation. In this study, we investigated the effects of 2 natural molecules, quercetin and sesamin, on neuroinflammation induced by the Parkinsonian toxin 1-methyl-4-phenylpyridinium (MPP(+)) in a glial-neuronal system. We first established that quercetin and sesamin defend microglial cells against MPP(+)-induced increases in the mRNA or protein levels of 3 pro-inflammatory cytokines (interleukin-6, IL-1ß and tumor necrosis factor-alpha), as revealed by real time-quantitative polymerase chain reaction and enzyme-linked immunoabsorbent assay, respectively. Quercetin and sesamin also decrease MPP(+)-induced oxidative stress in microglial cells by reducing inducible nitric oxide synthase protein expression as well as mitochondrial superoxide radicals. We then measured neuronal cell death and apoptosis after MPP(+) activation of microglia, in a microglial (N9)-neuronal (PC12) coculture system. Our results revealed that quercetin and sesamin rescued neuronal PC12 cells from apoptotic death induced by MPP(+) activation of microglial cells. Altogether, our data demonstrate that the phytoestrogen quercetin and the lignan sesamin diminish MPP(+)-evoked microglial activation and suggest that both these molecules may be regarded as potent, natural, anti-inflammatory compounds.

Quercetin and sesamin protect neuronal PC12 cells from high-glucose-induced oxidation, nitrosative stress, and apoptosis.

Complications of diabetes are now well-known to affect sensory, motor, and autonomic nerves. Diabetes is also thought to be involved in neurodegenerative processes characteristic of several neurodegenerative diseases. Indeed, it has been acknowledged recently that hyperglycemia-induced oxidative stress contributes to numerous cellular reactions typical of central nervous system deterioration. The goal of the present study was to evaluate the effects of the polyphenol quercetin and the lignan sesamin on high-glucose (HG)-induced oxidative damage in an in vitro model of dopaminergic neurons, neuronal PC12 cells. When incubated with HG (13.5 mg/mL), neuronal PC12 cells showed a significant increase of cellular death. Our results revealed that quercetin and sesamin defend neuronal PC12 cells from HG-induced cellular demise. An elevated level of reactive oxygen and nitrogen species is a consequence of improved oxidative stress after HG administration, and we demonstrated that this production diminishes with quercetin and sesamin treatment. We also found that quercetin and sesamin elicited an increment of superoxide dismutase activity. DNA fragmentation, Bax/Bcl-2 ratio, nuclear translocation of apoptosis-inducing factor, as well as poly(adenosine diphosphate [ADP]-ribose) polymerase cleavage were significantly reduced by quercetin and sesamin administration, affirming their antiapoptotic features. Also, HG treatment impacted caspase-3 cleavage, supporting caspase-3-dependent pathways as mechanisms of apoptotic death. Our results indicate a powerful role for these natural dietary compounds and emphasize preventive or complementary nutritional strategies for diabetes control.

Protective effects of resveratrol and quercetin against MPP+ -induced oxidative stress act by modulating markers of apoptotic death in dopaminergic neurons.

Reactive oxygen species produced by oxidative stress may participate in the apoptotic death of dopamine neurons distinctive of Parkinson's disease. Resveratrol, a red wine extract, and quercetin, found mainly in green tea, are two natural polyphenols, presenting antioxidant properties in a variety of cellular paradigms. The aim of this study was to evaluate the effect of resveratrol and quercetin on the apoptotic cascade induced by the administration of 1-methyl-4-phenylpyridinium ion (MPP(+)), a Parkinsonian toxin, provoking the selective degeneration of dopaminergic neurons. Our results show that a pre-treatment for 3 h with resveratrol or quercetin before MPP(+) administration could greatly reduce apoptotic neuronal PC12 death induced by MPP(+). We also demonstrated that resveratrol or quercetin modulates mRNA levels and protein expression of Bax, a pro-apoptotic gene, and Bcl-2, an anti-apoptotic gene. We then evaluated the release of cytochrome c and the nuclear translocation of the apoptosis-inducing factor (AIF). Altogether, our results indicate that resveratrol and quercetin diminish apoptotic neuronal cell death by acting on the expression of pro- and anti-apoptotic genes. These findings support the role of these natural polyphenols in preventive and/or complementary therapies for several human neurodegenerative diseases caused by oxidative stress and apoptosis.

Sesamin modulates tyrosine hydroxylase, superoxide dismutase, catalase, inducible NO synthase and interleukin-6 expression in dopaminergic cells under MPP+-induced oxidative stress.

Oxidative stress is regarded as a mediator of nerve cell death in several neurodegenerative disorders, such as Parkinson's disease. Sesamin, a lignan mainly found in sesame oil, is currently under study for its anti-oxidative and possible neuroprotective properties. We used 1-methyl-4-phenyl-pyridine (MPP(+)) ion, the active metabolite of the potent parkinsonism-causing toxin 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine, to produce oxidative stress and neurodegeneration in neuronal PC12 cells, which express dopamine, as well as neurofilaments. Our results show that picomolar doses of sesamin protected neuronal PC12 cells from MPP(+)-induced cellular death, as revealed by colorimetric measurements and production of reactive oxygen species. We also demonstrated that sesamin acted by rescuing tyrosine hydroxylase levels from MPP(+)-induced depletion. Sesamin, however, did not modulate dopamine transporter levels, and estrogen receptor-alpha and -beta protein expression. By examining several parameters of cell distress, we found that sesamin also elicited a strong increase in superoxide dismutase activity as well as protein expression and decreased catalase activity and the MPP(+) stimulated inducible nitric oxide synthase protein expression, in neuronal PC12 cells. Finally, sesamin possessed significant anti-inflammatory properties, as disclosed by its potential to reduce MPP(+)-induced interleukin-6 mRNA levels in microglia. From these studies, we determined the importance of the lignan sesamin as a neuroprotective molecule and its possible role in complementary and/or preventive therapies of neurodegenerative diseases.

Oxidative stress and 17-alpha- and 17-beta-estradiol modulate neurofilaments differently.

Oxidative stress plays an important role in the pathogenesis of neurodegenerative diseases such as Parkinson's disease (PD). Neuronal death in the substantia nigra of PD patients is partly caused by exacerbated oxidative damage. Our previous studies demonstrated that oxidative stress can alter the structure and stability of neurofilament (NF) proteins and that 17-alpha- and 17-beta-estradiol are potent neuroprotective agents. The aim of this study was to investigate the cytoskeletal target of neuroprotection by estrogens in neuronal PC12 cells. We induced oxidative stress by MPP+ administration for 24 h, and 17-alpha- and 17-beta-estradiol were used as neuroprotective drugs. We measured gene expression and protein expression of each NF subunit, NFL, NFM, and NFH, by semiquantitative RT-PCR, Western blot, and immunofluorescence. Our results demonstrate that NFL mRNA and protein levels are not modulated by MPP+ or estradiol isomers, whereas NFM gene expression, as well as protein expression, are strongly influenced by MPP+, 17-alpha-, and 17-beta-estradiol after a 24-h treatment. Finally, mRNA levels of the most phosphorylated subunits, NFH, are not changed by MPP+ or treatment with both estradiol isomers, whereas NFH protein expression is decreased by the same treatments. These results suggest that oxidative stress affects neuronal cytoskeleton, maybe though proteolysis and/or abnormal structural changes in NFs. Then, 17-alpha- and 17-beta-estradiol might help the neuronal cell in recovering after oxidative stress by inducing protein expression of NFM and NFH subunits.