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1.
Pharmaceuticals (Basel) ; 17(3)2024 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-38543056

RESUMO

The bottleneck in drug discovery for central nervous system diseases is the absence of effective systemic drug delivery technology for delivering therapeutic drugs into the brain. Despite the advances in the technology used in drug discovery, such as Adeno-Associated Virus (AAV) vectors, the development of drugs for central nervous system diseases remains challenging. In this manuscript, we describe, for the first time, the development of a workflow to generate a novel brain-targeted drug delivery system that involves the generation of genetically engineered exosomes by first selecting various functional AAV capsid-specific peptides (collectively called CAPs) known to be involved in brain-targeted high-expression gene delivery, and then expressing the CAPs in frame with lysosome-associated membrane glycoprotein (Lamp2b) followed by expressing CAP-Lamp2b fusion protein on the surface of mesenchymal stem cell-derived exosomes, thus generating CAP-exosomes. Intravenous injection of green fluorescent protein (GFP) gene-loaded CAP-exosomes in mice transferred the GFP gene throughout the CNS as measured by monitoring brain sections for GFP expression with confocal microscopy. GFP gene transfer efficiency was at least 20-fold greater than that of the control Lamp2b-exosomes, and GFP gene transduction to mouse liver was low.

2.
J Neuroimmune Pharmacol ; 17(1-2): 228-241, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-34028667

RESUMO

Alzheimer's disease (AD) and other neurodegenerative diseases are characterized by chronic neuroinflammation and a reduction in brain energy metabolism. An important role has emerged for small, non-coding RNA molecules known as microRNAs (miRNAs) in the pathophysiology of many neurodegenerative disorders. As epigenetic regulators, miRNAs possess the capacity to regulate and fine tune protein production by inhibiting translation. Several miRNAs, which include miR-146a, are elevated in the brain, CSF, and plasma of AD patients. miR-146a participates in pathways that regulate immune activation and has several mRNA targets which encode for proteins involved in cellular energy metabolism. An additional role for extracellular vesicles (EVs) has also emerged in the progression AD, as EVs can transfer functionally active proteins and RNAs from diseased to healthy cells. In the current study, we exposed various cell types present within the CNS to immunomodulatory molecules and observed significant upregulation of miR-146a expression, both within cells and within their secreted EVs. Further, we assessed the effects of miR-146a overexpression on bioenergetic function in primary rat glial cells and found significant reductions in oxidative phosphorylation and glycolysis. Lastly, we correlated miR-146a expression levels within various regions of the AD brain to disease staging and found significant, positive correlations. These novel results demonstrate that the modulation of miR-146a in response to neuroinflammatory stimuli may mediate the loss of mitochondrial integrity and function in cells, thereby contributing to the progression of beta-amyloid and tau pathology in the AD brain. Multiple inflammatory stimuli can upregulate miRNA-146a expression within neurons, mixed glial cells, and brain endothelial cells, which is either retained within these cells or released from them as extracellular vesicle cargo. The upregulation of miR-146a disrupts cellular bioenergetics in mixed glial cells. This mechanism may play a critical role in the neuroinflammatory response observed during Alzheimer's disease.


Assuntos
Doença de Alzheimer , MicroRNAs , Animais , Ratos , Doença de Alzheimer/genética , Células Endoteliais , Imunomodulação , Metabolismo Energético , MicroRNAs/genética
3.
Neurobiol Aging ; 105: 115-128, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34062487

RESUMO

Heart disease and vascular disease positively correlate with the incidence of Alzheimer's disease (AD). Although there is ostensible involvement of dysfunctional cerebrovasculature in AD pathophysiology, the characterization of the specific changes and development of vascular injury during AD remains unclear. In the present study, we established a time-course for the structural changes and degeneration of the angioarchitecture in AD. We used cerebrovascular corrosion cast and µCT imaging to evaluate the geometry, topology, and complexity of the angioarchitecture in the brain of wild type and 3xTg AD mice. We hypothesized that changes to the microvasculature occur early during the disease, and these early identifiable aberrations would be more prominent in the brain subregions implicated in the cognitive decline of AD. Whole-brain analysis of the angioarchitecture indicated early morphological abnormalities and degeneration of microvascular networks in 3xTg AD mice. Our analysis of the hippocampus and cortical subregions revealed microvascular degeneration with onset and progression that was subregion dependent.


Assuntos
Envelhecimento/metabolismo , Envelhecimento/patologia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Encéfalo/irrigação sanguínea , Microvasos/patologia , Placa Amiloide/metabolismo , Doença de Alzheimer/diagnóstico por imagem , Doença de Alzheimer/psicologia , Animais , Encéfalo/diagnóstico por imagem , Cognição , Modelos Animais de Doenças , Progressão da Doença , Masculino , Camundongos Mutantes , Camundongos Transgênicos , Microvasos/diagnóstico por imagem , Microtomografia por Raio-X
4.
Front Aging Neurosci ; 12: 92, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32317959

RESUMO

Despite the extensive use of hormonal methods as either contraception or menopausal hormone therapy (HT), there is very little known about the potential effects of these compounds on the cellular processes of the brain. Medroxyprogesterone Acetate (MPA) is a progestogen used globally in the hormonal contraceptive, Depo Provera, by women in their reproductive prime and is a major compound found in HT formulations used by menopausal women. MPA promotes changes in the circulating levels of matrix metalloproteinases (MMPs), such as MMP-9, in the endometrium, yet limited literature studying the effects of MPA on neurons and astroglia cells has been conducted. Additionally, the dysregulation of MMPs has been implicated in the pathology of Alzheimer's disease (AD), where inhibiting the secretion of MMP-9 from astroglia reduces the proteolytic degradation of amyloid-beta. Thus, we hypothesize that exposure to MPA disrupts proteolytic degradation of amyloid-beta through the downregulation of MMP-9 expression and subsequent secretion. To assess the effect of progestins on MMP-9 and amyloid-beta, in vitro, C6 rat glial cells were exposed to MPA for 48 h and then the enzymatic, secretory, and amyloid-beta degrading capacity of MMP-9 was assessed from the conditioned culture medium. We found that MPA treatment inhibited transcription of MMP-9, which resulted in a subsequent decrease in the production and secretion of MMP-9 protein, in part through the glucocorticoid receptor. Additionally, we investigated the consequences of amyloid beta-degrading activity and found that MPA treatment decreased proteolytic degradation of amyloid-beta. Our results suggest MPA suppresses amyloid-beta degradation in an MMP-9-dependent manner, in vitro, and potentially compromises the clearance of amyloid-beta in vivo.

5.
J Alzheimers Dis ; 75(1): 119-138, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32250296

RESUMO

Cerebrovascular pathology is pervasive in Alzheimer's disease (AD), yet it is unknown whether cerebrovascular dysfunction contributes to the progression or etiology of AD. In human subjects and in animal models of AD, cerebral hypoperfusion and hypometabolism are reported to manifest during the early stages of the disease and persist for its duration. Amyloid-ß is known to cause cellular injury in both neurons and endothelial cells by inducing the production of reactive oxygen species and disrupting intracellular Ca2+ homeostasis. We present a mechanism for mitochondrial degeneration caused by the production of mitochondrial superoxide, which is driven by increased mitochondrial Ca2+ uptake. We found that persistent superoxide production injures mitochondria and disrupts electron transport in cerebrovascular endothelial cells. These observations provide a mechanism for the mitochondrial deficits that contribute to cerebrovascular dysfunction in patients with AD.


Assuntos
Peptídeos beta-Amiloides/farmacologia , Cálcio/metabolismo , Células Endoteliais/metabolismo , Mitocôndrias/metabolismo , Fragmentos de Peptídeos/farmacologia , Superóxidos/metabolismo , Regulação para Cima/efeitos dos fármacos , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Linhagem Celular , Células Endoteliais/efeitos dos fármacos , Camundongos , Mitocôndrias/efeitos dos fármacos , Fosforilação Oxidativa , Espécies Reativas de Oxigênio/metabolismo
6.
Sci Rep ; 10(1): 3233, 2020 02 24.
Artigo em Inglês | MEDLINE | ID: mdl-32094435

RESUMO

Blood-brain barrier (BBB) dysfunction occurs in cerebrovascular diseases and neurodegenerative disorders such as stroke. Opening of the BBB during a stroke has a negative impact on acute outcomes. We have recently demonstrated that miR-34a regulates the BBB by targeting cytochrome c (CYC) in vitro. To investigate the role of miR-34a in a stroke, we purified primary cerebrovascular endothelial cells (pCECs) from mouse brains following 1 h transient middle cerebral artery occlusion (tMCAO) and measured real-time PCR to detect miR-34a levels. We demonstrate that the miR-34a levels are elevated in pCECs from tMCAO mice at the time point of BBB opening following 1 h tMCAO and reperfusion. Interestingly, knockout of miR-34a significantly reduces BBB permeability, alleviates disruption of tight junctions, and improves stroke outcomes compared to wild-type (WT) controls. CYC is decreased in the ischemic hemispheres and pCECs from WT but not in miR-34a-/- mice following stroke reperfusion. We further confirmed CYC is a target of miR-34a by a dural luciferase reporter gene assay in vitro. Our study provides the first description of miR-34a affecting stroke outcomes and may lead to discovery of new mechanisms and treatments for cerebrovascular and neurodegenerative diseases such as stroke.


Assuntos
Citocromos c/metabolismo , MicroRNAs/metabolismo , Acidente Vascular Cerebral/genética , Animais , Barreira Hematoencefálica/patologia , Isquemia Encefálica/genética , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Junções Íntimas/metabolismo , Resultado do Tratamento
7.
J Alzheimers Dis ; 70(1): 139-151, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31177221

RESUMO

Mitochondrial dysfunction is often found in Alzheimer's disease (AD) patients and animal models. Clinical severity of AD is linked to early deficiencies in cognitive function and brain metabolism, indicating that pathological changes may begin early in life. Previous studies showed decreased mitochondrial function in primary hippocampal neurons from triple-transgenic Alzheimer's disease (3xTg-AD) mice and mitochondrial movement and structure deficits in primary neurons exposed to amyloid-ß oligomers. The present study characterized mitochondrial movement, number, and structure in 3xTg-AD primary cortical neurons and non-transgenic (nonTg) controls. We found a significant reduction in mitochondrial number and movement in 3xTg-AD primary cortical neurons with modest structural changes. Additionally, application of the sigma-1 receptor agonist, (+)SKF-10,047, markedly increased mitochondrial movement in both 3xTg-AD and nonTg primary cortical cultures after one hour of treatment. (+)SKF-10,047 also led to a trend of increased mitochondrial number in 3xTg-AD cultures. Embryonic mitochondrial movement and number deficits could be among the key steps in the early pathogenesis of AD that compromise cognitive or metabolic reserve, and amelioration of these deficits could be a promising area for further preclinical and clinical study.


Assuntos
Doença de Alzheimer/metabolismo , Córtex Cerebral/metabolismo , Mitocôndrias/metabolismo , Neurônios/metabolismo , Doença de Alzheimer/patologia , Animais , Córtex Cerebral/patologia , Modelos Animais de Doenças , Camundongos , Camundongos Transgênicos , Mitocôndrias/patologia , Dinâmica Mitocondrial/fisiologia , Neurônios/patologia
8.
Aging Dis ; 10(2): 329-352, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31011481

RESUMO

Aging is a complex and integrated gradual deterioration of cellular activities in specific organs of the body, which is associated with increased mortality. This deterioration is the primary risk factor for major human pathologies, including cancer, diabetes, cardiovascular disorders, neurovascular disorders, and neurodegenerative diseases. There are nine tentative hallmarks of aging. In addition, several of these hallmarks are increasingly being associated with acute brain injury conditions. In this review, we consider the genes and their functional pathways involved in brain aging as a means of developing new strategies for therapies targeted to the neuropathological processes themselves, but also as targets for many age-related brain diseases. A single microRNA (miR), which is a short, non-coding RNA species, has the potential for targeting many genes simultaneously and, like practically all other cellular processes, genes associated with many features of brain aging and injury are regulated by miRs. We highlight how certain miRs can mediate deregulation of genes involved in neuroinflammation, acute neuronal injury and chronic neurodegenerative diseases. Finally, we review the recent progress in the development of effective strategies to block specific miR functions and discuss future approaches with the prediction that anti-miR drugs may soon be used in the clinic.

9.
Front Cell Neurosci ; 13: 51, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30837842

RESUMO

Extracellular vesicles (EVs) are small, membrane-bound nanoparticles released from most, if not all cells, and can carry functionally active cargo (proteins, nucleic acids) which can be taken up by neighboring cells and mediate physiologically relevant effects. In this capacity, EVs are being regarded as novel cell-to-cell communicators, which may play important roles in the progression of neurodegenerative diseases, like Alzheimer's disease (AD). Aside from the canonical physical hallmarks of this disease [amyloid ß (Aß) plaques, neurofibrillary tangles, and widespread cell death], AD is characterized by chronic neuroinflammation and mitochondrial dysfunction. In the current study, we sought to better understand the role of tumor necrosis factor-alpha (TNF-α), known to be involved in inflammation, in mediating alterations in mitochondrial function and EV secretion. Using an immortalized hippocampal cell line, we observed significant reductions in several parameters of mitochondrial oxygen consumption after a 24-h exposure period to TNF-α. In addition, after TNF-α exposure we also observed significant upregulation of two microRNAs (miRNAs; miR-34a and miR-146a) associated with mitochondrial dysfunction in secreted EVs. Despite this, when naïve cells are exposed to EVs isolated from TNF-α treated cells, mitochondrial respiration, proton leak, and reactive oxygen species (ROS) production are all significantly increased. Collectively these data indicate that a potent proinflammatory cytokine, TNF-α, induces significant mitochondrial dysfunction in a neuronal cell type, in part via the secretion of EVs, which significantly alter mitochondrial activity in recipient cells.

10.
Neurochem Int ; 127: 73-79, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30365981

RESUMO

Aging of the nervous system, and the occurrence of age-related brain diseases such as stroke, are associated with changes to a variety of cellular processes controlled by many distinct genes. MicroRNAs (miRNAs), short non-coding functional RNAs that can induce translational repression or site-specific cleavage of numerous target mRNAs, have recently emerged as important regulators of cellular senescence, aging, and the response to neurological insult. Here, we focused on the assessment of the role of miR-34a in stroke. We noted increases in miR-34a expression in the blood of stroke patients as well as in blood and brain of mice subjected to experimental stroke. Our methodical genetic manipulation of miR-34a expression substantially impacted stroke-associated preclinical outcomes and we have in vitro evidence that these changes may be driven at least in part by disruptions to blood brain barrier integrity and mitochondrial oxidative phosphorylation in endothelial cells. Finally, aging, independent of brain injury, appears to be associated with shifts in circulating miRNA profiles. Taken together, these data support a role for miRNAs, and specifically miR-34a, in brain aging and the physiological response to age-related neurological insult, and lay the groundwork for future investigation of this novel therapeutic target.


Assuntos
Isquemia Encefálica/genética , Infarto Cerebral/genética , MicroRNAs/genética , Acidente Vascular Cerebral/genética , Envelhecimento/fisiologia , Animais , Barreira Hematoencefálica/metabolismo , Encéfalo/metabolismo , Isquemia Encefálica/metabolismo , Senescência Celular/genética , Infarto Cerebral/metabolismo , Células Endoteliais/metabolismo , Mitocôndrias/genética , Mitocôndrias/metabolismo , Fatores de Risco , Acidente Vascular Cerebral/metabolismo
11.
Mitochondrion ; 47: 244-255, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30594729

RESUMO

Astrocytes serve to maintain proper neuronal function and support neuronal viability, but remain largely understudied in research of cerebral ischemia. Astrocytic mitochondria are core participants in the metabolic activity of astrocytes. The objective of this study is to assess astrocyte mitochondrial competence during hypoxia and post-hypoxia reoxygenation and to determine cellular adaptive and pathological changes in the mitochondrial network. We hypothesize that during metabolic distress in astrocytes; mitochondrial networks undergo a shift in fission-fusion dynamics that results in a change in the morphometric state of the entire mitochondrial network. This mitochondrial network shift may be protective during metabolic distress by priming mitochondrial size and facilitating mitophagy. We demonstrated that hypoxia and post-hypoxia reoxygenation of rat primary astrocytes results in a redistribution of mitochondria to smaller sizes evoked by increased mitochondrial fission. Excessive mitochondrial fission corresponded to Drp-1 dephosphorylation at Ser 637, which preceded mitophagy of relatively small mitochondria. Reoxygenation of astrocytes marked the initiation of elevated mitophagic activity primarily reserved to the perinuclear region where a large number of the smallest mitochondria occurred. Although, during hypoxia astrocytic ATP content was severely reduced, after reoxygenation ATP content returned to near normoxic values and these changes mirrored mitochondrial superoxide production. Concomitant with these changes in astrocytic mitochondria, the number of astrocytic extensions declined only after 10-hours post-hypoxic reoxygenation. Overall, we posit a drastic mitochondrial network change that is triggered by a metabolic crisis during hypoxia; these changes are followed by mitochondrial degradation and retraction of astrocytic extensions during reoxygenation.


Assuntos
Astrócitos/metabolismo , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Mitofagia , Oxigênio/metabolismo , Animais , Astrócitos/patologia , Hipóxia Celular , Células Cultivadas , Dinaminas/metabolismo , Mitocôndrias/patologia , Proteínas Mitocondriais/metabolismo , Oxigênio/farmacologia , Ratos
12.
J Cereb Blood Flow Metab ; 36(2): 387-92, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26661155

RESUMO

The blood-brain barrier is composed of cerebrovascular endothelial cells and tight junctions, and maintaining its integrity is crucial for the homeostasis of the neuronal environment. Recently, we discovered that mitochondria play a critical role in maintaining blood-brain barrier integrity. We report for the first time a novel mechanism underlying blood-brain barrier integrity: miR-34a mediated regulation of blood-brain barrier through a mitochondrial mechanism. Bioinformatics analysis suggests miR-34a targets several mitochondria-associated gene candidates. We demonstrated that miR-34a triggers the breakdown of blood-brain barrier in cerebrovascular endothelial cell monolayer in vitro, paralleled by reduction of mitochondrial oxidative phosphorylation and adenosine triphosphate production, and decreased cytochrome c levels.


Assuntos
Barreira Hematoencefálica/fisiologia , Citocromos c/genética , Citocromos c/fisiologia , MicroRNAs/genética , Mitocôndrias/genética , Mitocôndrias/fisiologia , Difosfato de Adenosina/biossíntese , Animais , Circulação Cerebrovascular/genética , Circulação Cerebrovascular/fisiologia , Biologia Computacional , Células Endoteliais , Endotélio Vascular/citologia , Cinética , Camundongos , Fosforilação Oxidativa , Consumo de Oxigênio/fisiologia , Permeabilidade , Junções Íntimas
13.
J Clin Cell Immunol ; 7(6)2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28652929

RESUMO

This short communication describes our research which demonstrates that TNF-α causes a rapid decline in mitochondrial function, leading to neuronal cell death. As such, this neurotoxic proinflammatory cytokine may play a role in brain damage from stroke and neurodegeneration in chronic conditions such as Alzheimer's disease (AD) and Parkinson's disease. We have extended this initial observation by demonstrating that TNF-α stimulates a microRNA (miR-34a) which we have shown reduces five key proteins in the mitochondrial electron transport chain through base-pair complementarity. miR-34a is increased in affected brain regions of Alzheimer's patients and transgenic AD mouse models. We have further shown that oligomeric amyloid beta 42 (oAß42) stimulates miR-34a. Collectively, these data suggest that TNF-α, oAß42, and miR-34a participate in a vicious cycle, resulting in mitochondrial dysfunction, which is critical to the neuropathology of AD.

14.
Brain Res ; 1616: 101-11, 2015 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-25964165

RESUMO

Perturbations in dynamic properties of mitochondria including fission, fusion, and movement lead to disruption of energy supply to synapses contributing to neuropathology and cognitive dysfunction in Alzheimer׳s disease (AD). The molecular mechanisms underlying these defects are still unclear. Previously, we have shown that ERß is localized in the mitochondria and ERß knock down disrupts mitochondrial functions. Because a selective ERß modulator (DPN) can activate PKA, and localized PKA signaling in the mitochondrial membrane regulates mitochondrial structure and functions, we reasoned that ERß signaling in the mitochondrial membrane rescues many of the mitochondrial defects caused by soluble Aß oligomer. We now report that DPN treatment in primary hippocampal neurons attenuates soluble Aß-oligomer induced dendritic mitochondrial fission and reduced mobility. Additionally, Aß treatment reduced the respiratory reserve capacity of hippocampal neuron and inhibited phosphorylation of Drp1 at its PKA site, which induces excessive mitochondrial fission, and DPN treatment ameliorates these inhibitions. Finally, we discovered a direct interaction of ERß with a mitochondrial resident protein AKAP1, which induces the PKA-mediated local signaling pathway involved in increased oxidative phosphorylation and inhibition of mitochondrial fission. Taken together, our findings highlight the possibility that ERß signaling pathway may be a useful mitochondria-directed therapeutic target for AD.


Assuntos
Peptídeos beta-Amiloides/farmacologia , Estrogênios/farmacologia , Mitocôndrias/efeitos dos fármacos , Proteínas Mitocondriais/metabolismo , Neurônios/ultraestrutura , Fragmentos de Peptídeos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Proteínas de Ancoragem à Quinase A/genética , Proteínas de Ancoragem à Quinase A/metabolismo , Animais , Encéfalo/citologia , Células Cultivadas , Dendritos/efeitos dos fármacos , Dendritos/ultraestrutura , Dinaminas/metabolismo , Embrião de Mamíferos , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Humanos , Ratos , Transfecção
15.
Brain Res ; 1383: 1-12, 2011 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-21262203

RESUMO

Neuromodulation of synaptic plasticity by 17ß-estradiol (E2) is thought to influence information processing and storage in the cortex and hippocampus. Because E2 rapidly affects cortical memory and synaptic plasticity, we examined its effects on phosphorylation of calcium/calmodulin-dependent protein kinase II (CaMKII), extracellular signal-regulated kinase (ERK), and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) [AMPA-type glutamate receptor subunit 1 (GluR1 subunit)], all of which are important for the induction and maintenance of synaptic plasticity and memory. Acute E2 treatment resulted in an increased temporal and spatial phosphorylation pattern of CaMKII, ERK, and AMPAR (GluR1 subunit). By using inhibitors, we were able to attribute GluR1 phosphorylation to CaMKII at serine 831, and we also found that E2 treatment increased GluR1 insertion into the surface membrane. Because soluble amyloid-beta (Aß) oligomers inhibit CaMKII and ERK activation, which is necessary for synaptic plasticity, we also tested E2's ability to ameliorate Aß-induced dysfunction of synaptic plasticity. We found that estrogen treatment in neuronal culture, slice culture, and in vivo, ameliorated Aß oligomer-induced inhibition of CaMKII, ERK, and AMPAR phosphorylation, and also ameliorated the Aß oligomer-induced reduction of dendritic spine density in a CaMKII-dependent manner. These phosphorylation events are correlated with the early stage of inhibitory avoidance learning, and our data show that E2 improved inhibitory avoidance memory deficits in animals treated with soluble Aß oligomers. This study identifies E2-induced signaling that attenuates soluble Aß peptide-mediated dysfunction of pathways in synaptic plasticity.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Estradiol/farmacologia , Estrogênios/farmacologia , Plasticidade Neuronal/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Feminino , Fosforilação , Ratos
16.
J Biol Chem ; 284(14): 9540-8, 2009 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-19189968

RESUMO

We recently demonstrated mitochondrial localization of estrogen receptor beta (ERbeta). We herein confirm the mitochondrial localization of ERbeta by the loss of mitochondrial ERbeta immunoreactivity in ERbeta knockdown cells. A phenotype change characterized as an increase in resistance to oxidative stressors is associated with ERbeta knockdown. ERbeta knockdown results in a lower resting mitochondrial membrane potential (Deltapsim) and increase in resistance to hydrogen peroxide-induced Deltapsim depolarization in both immortal hippocampal cells and primary hippocampal neurons. ERbeta knockdown cells maintained ATP concentrations despite insults that compromise ATP production and produce less mitochondrial superoxide under oxidative stress. Furthermore, similar mitochondrial phenotype changes were identified in primary hippocampal neurons derived from ERbeta knock-out mice. These data demonstrate that ERbeta is expressed in mitochondria and function as a mitochondrial vulnerability factor involved in Deltapsim maintenance, potentially through a mitochondrial transcription dependent mechanism.


Assuntos
Receptor beta de Estrogênio/metabolismo , Mitocôndrias/metabolismo , Animais , Células Cultivadas , Receptor beta de Estrogênio/deficiência , Receptor beta de Estrogênio/genética , Humanos , Potencial da Membrana Mitocondrial , Camundongos , Camundongos Knockout , Interferência de RNA , Ratos , Técnicas de Cultura de Tecidos
17.
Proc Natl Acad Sci U S A ; 105(39): 15148-53, 2008 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-18815371

RESUMO

L-type voltage-gated Ca(2+)channels (VGCC) play an important role in dendritic development, neuronal survival, and synaptic plasticity. Recent studies have demonstrated that the gonadal steroid estrogen rapidly induces Ca(2+) influx in hippocampal neurons, which is required for neuroprotection and potentiation of LTP. The mechanism by which estrogen rapidly induces this Ca(2+) influx is not clearly understood. We show by electrophysiological studies that extremely low concentrations of estrogens acutely potentiate VGCC in hippocampal neurons, hippocampal slices, and HEK-293 cells transfected with neuronal L-type VGCC, in a manner that was estrogen receptor (ER)-independent. Equilibrium, competitive, and whole-cell binding assays indicate that estrogen directly interacts with the VGCC. Furthermore, a L-type VGCC antagonist to the dihydropyridine site displaced estrogen binding to neuronal membranes, and the effects of estrogen were markedly attenuated in a mutant, dihydropyridine-insensitive L-type VGCC, demonstrating a direct interaction of estrogens with L-type VGCC. Thus, estrogen-induced potentiation of calcium influx via L-type VGCC may link electrical events with rapid intracellular signaling seen with estrogen exposure leading to modulation of synaptic plasticity, neuroprotection, and memory formation.


Assuntos
Canais de Cálcio Tipo L/metabolismo , Estrogênios/metabolismo , Neurônios/metabolismo , Animais , Canais de Cálcio Tipo L/genética , Linhagem Celular , Estradiol/metabolismo , Estradiol/farmacologia , Estrogênios/farmacologia , Humanos , Mutação , Neurônios/efeitos dos fármacos , Ratos , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
18.
Mol Cell Endocrinol ; 290(1-2): 51-9, 2008 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-18571833

RESUMO

Estrogens are potent neuroprotective hormones and mitochondria are the site of cellular life-death decisions. As such, it is not surprising that we and others have shown that estrogens have remarkable effects on mitochondrial function. Herein we provide evidence for a primary effect of estrogens on mitochondrial function, achieved in part by the import of estrogen receptor beta (ERbeta) into the mitochondria where it mediates a number of estrogen actions on this vital organelle. ERbeta is imported into the mitochondria, through tethering to cytosolic chaperone protein and/or through direct interaction with mitochondrial import proteins. In the mitochondria, ERbeta can affect transcription of critical mitochondrial genes through the interaction with estrogen response elements (ERE) or through protein-protein interactions with mitochondrially imported transcription factors. The potent effects of estrogens on mitochondrial function, particularly during mitochondrial stress, argues for a role of estrogens in the treatment of mitochondrial defects in chronic neurodegenerative diseases like Alzheimer's disease (AD) and Parkinson's disease (PD) and more acute conditions of mitochondrial compromise, like cerebral ischemia and traumatic brain injury.


Assuntos
Estrogênios/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Animais , Citoproteção , Receptor beta de Estrogênio/metabolismo , Humanos , Doenças Neurodegenerativas/metabolismo , Transporte Proteico
19.
Neuroreport ; 15(9): 1515-8, 2004 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-15194886

RESUMO

Estrogens exert neuroprotective activity in both in vivo and in vitro model systems. Herein, we report that both 17beta-estradiol and low concentrations of nitric oxide (NO) attenuate hydrogen peroxide (H2O2) induced toxicity in SK-N-SH cells, which express the neuronal nitric oxide synthase (nNOS). 17beta-estradiol rapidly induced an increase in NO levels. A nNOS inhibitor was able to block the neuroprotection of 17beta-estradiol. Cyclic guanylyl mono-phosphate (cGMP) also protected against H2O2 induced toxicity, while NO's protection was attenuated by ODQ, a soluble guanylyl cyclase (sGC) inhibitor. In SK-N-SH cells, the major estrogen receptor isoforms is estrogen receptor beta. Our current study suggests that increased activity of nNOS may be involved in the neuroprotection conferred by 17beta-estradiol.


Assuntos
Estradiol/farmacologia , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Fármacos Neuroprotetores/farmacologia , Óxido Nítrico Sintase/metabolismo , Linhagem Celular Tumoral , GMP Cíclico/metabolismo , Humanos , Peróxido de Hidrogênio/toxicidade , Neuroblastoma , Óxido Nítrico/metabolismo , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico Sintase Tipo I , Compostos Nitrosos/farmacologia , Oxidantes/toxicidade
20.
J Neurochem ; 87(2): 333-43, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-14511111

RESUMO

Presenilin (PS) in association with nicastrin (NICA) forms a gamma-secretase complex that plays a crucial role in facilitating intramembranous processing of Notch, a signaling receptor that is essential for neuronal fate specification and differentiation. Loss of function studies have implicated a role for PS1 in regulating neuronal differentiation in association with the down-regulation of Notch signaling during neurogenesis. By using a system for stable, as well as tetracycline-inducible expression of interfering RNAs (RNAi), we studied the functions of PS1 during neuronal differentiation in the murine pluripotent p19 embryonic carcinoma cell line. After retinoic acid (RA) treatment and in the absence of doxycycline, neuronal progenitor cells in the p19 clone were found to extend their processes towards the neighboring colony to form network-like connections, as revealed by neuron-specific microtubule-associated protein 2 staining and laser scanning confocal microscopy. However, doxycycline-induced expression of PS1 small interfering RNA (siRNA) in the p19 clone resulted in a severe defect in the formation of network-like connections. Expression of the NICA and Notch down-stream effector genes Hes1 and Hes5 was unaffected in p19 cells expressing doxycycline-induced PS1 siRNA. In contrast to PS1, constitutive inactivation of NICA by siRNA in p19 cells resulted in premature and partial differentiation without RA treatment. In these NICA siRNA-expressing p19 cells the expression of the Notch1 down-stream effector Hes1 gene was substantially reduced. After RA treatment the NICA siRNA clone failed to differentiate completely into networks of neurons. These results taken together provide direct evidence that PS1 and NICA may participate in neuronal differentiation during neurogenesis in vitro.


Assuntos
Diferenciação Celular/fisiologia , Glicoproteínas de Membrana/metabolismo , Proteínas de Membrana/metabolismo , Neurônios/metabolismo , Receptores de Superfície Celular , Fatores de Transcrição , Secretases da Proteína Precursora do Amiloide , Animais , Diferenciação Celular/efeitos dos fármacos , Doxiciclina/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Glicoproteínas de Membrana/antagonistas & inibidores , Glicoproteínas de Membrana/genética , Proteínas de Membrana/genética , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , Células-Tronco Neoplásicas/citologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Neurônios/citologia , Presenilina-1 , RNA de Cadeia Dupla/biossíntese , RNA Interferente Pequeno/biossíntese , RNA Interferente Pequeno/farmacologia , Receptor Notch1 , Tretinoína/farmacologia
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