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1.
FASEB J ; 29(9): 3702-12, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25985800

RESUMO

Epidermal growth factor receptor (EGFR) plays pivotal roles in cell proliferation, differentiation, and tissue development, while EGFs protect neurons from toxic insults by binding EGFR and stimulating survival signaling. Furthermore, recent evidence implicates this receptor in neurometabolic disorders like Alzheimer disease and aging. Here we show that absence of presenilin 1 (PS1) results in dramatic decrease (>95%) of neuronal EGFR and that PS1-null (PS1(-/-)) brains have reduced amounts of this receptor. PS1(-/-) cortical neurons contain little EGFR and show no epidermal growth factor-induced survival signaling or protection against excitotoxicity, but exogenous EGFR rescues both functions even in absence of PS1. EGFR mRNA is greatly reduced (>95%) in PS1(-/-) neurons, and PS1(-/-) brains contain decreased amounts of this mRNA, although PS1 affects the stability of neither EGFR nor its mRNA. Exogenous PS1 increases neuronal EGFR mRNA, while down-regulation of PS1 decreases this mRNA. These effects are neuron specific, as PS1 affects the EGFR of neither glial nor fibroblast cells. In addition, PS1 controls EGFR through novel mechanisms shared with neither γ-secretase nor PS2. Our data reveal that PS1 functions as a positive transcriptional regulator of neuronal EGFR controlling its expression in a cell-specific manner. Severe downregulation of EGFR may contribute to developmental abnormalities and lethal phenotype found in PS1, but not PS2, null mice. Furthermore, PS1 may affect neuroprotection and Alzheimer disease by controlling survival signaling of neuronal EGFR.


Assuntos
Doença de Alzheimer/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Receptores ErbB/biossíntese , Regulação da Expressão Gênica , Neurônios/metabolismo , Presenilina-1/metabolismo , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Secretases da Proteína Precursora do Amiloide/genética , Animais , Receptores ErbB/genética , Camundongos , Camundongos Knockout , Neuroglia/metabolismo , Neuroglia/patologia , Neurônios/patologia , Presenilina-1/genética , Transcrição Gênica
2.
J Biol Chem ; 288(42): 30495-30501, 2013 Oct 18.
Artigo em Inglês | MEDLINE | ID: mdl-24025330

RESUMO

Abnormally high concentrations of extracellular glutamate in the brain may cause neuronal damage via excitotoxicity. Thus, tight regulation of glutamate release is critical to neuronal function and survival. Excitotoxicity is caused mainly by overactivation of the extrasynaptic NMDA receptor (NMDAR) and results in specific cellular changes, including calcium-induced activation of calpain proteases. Here, we report that presenilin-1 (PS1) null mouse cortical neuronal cultures have increased amounts of calpain-dependent spectrin breakdown products (SBDPs) compared with WT cultures. NMDAR antagonists blocked accumulation of SBDPs, suggesting abnormal activation of this receptor in PS1 null cultures. Importantly, an increase in SBDPs was detected in cultures of at least 7 days in vitro but not in younger cultures. Conditioned medium from PS1 null neuronal cultures at 8 days in vitro contained higher levels of glutamate than medium from WT cultures and stimulated production of SBDPs when added to WT cultures. Use of glutamate reuptake inhibitors indicated that accumulation of this neurotransmitter in the media of PS1 null cultures was due to increased rates of release. PS1 null neurons showed decreased cell surface expression and phosphorylation of the GluN2B subunit of NMDAR, indicating decreased amounts of extrasynaptic NMDAR in the absence of PS1. Inhibition of γ-secretase activity in WT neurons caused changes similar to those observed in PS1 null neurons. Together, these data indicate that the PS1/γ-secretase system regulates release of glutamate, tyrosine phosphorylation, and surface expression of GluN2B-containing NMDARs.


Assuntos
Secretases da Proteína Precursora do Amiloide/metabolismo , Córtex Cerebral/metabolismo , Regulação da Expressão Gênica/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Presenilina-1/metabolismo , Receptores de N-Metil-D-Aspartato/biossíntese , Secretases da Proteína Precursora do Amiloide/genética , Animais , Células Cultivadas , Córtex Cerebral/citologia , Camundongos , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Neurônios/citologia , Fosforilação/fisiologia , Presenilina-1/genética , Receptores de N-Metil-D-Aspartato/genética , Espectrina/genética , Espectrina/metabolismo , Fatores de Tempo , Tirosina/genética , Tirosina/metabolismo
3.
Neurobiol Aging ; 34(2): 499-510, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22475621

RESUMO

Activation of EphB receptors by ephrinB (efnB) ligands on neuronal cell surface regulates important functions, including neurite outgrowth, axonal guidance, and synaptic plasticity. Here, we show that efnB rescues primary cortical neuronal cultures from necrotic cell death induced by glutamate excitotoxicity and that this function depends on EphB receptors. Importantly, the neuroprotective function of the efnB/EphB system depends on presenilin 1 (PS1), a protein that plays crucial roles in Alzheimer's disease (AD) neurodegeneration. Furthermore, absence of one PS1 allele results in significantly decreased neuroprotection, indicating that both PS1 alleles are necessary for full expression of the neuroprotective activity of the efnB/EphB system. We also show that the ability of brain-derived neurotrophic factor (BDNF) to protect neuronal cultures from glutamate-induced cell death depends on PS1. Neuroprotective functions of both efnB and BDNF, however, were independent of γ-secretase activity. Absence of PS1 decreases cell surface expression of neuronal TrkB and EphB2 without affecting total cellular levels of the receptors. Furthermore, PS1-knockout neurons show defective ligand-dependent internalization and decreased ligand-induced degradation of TrkB and Eph receptors. Our data show that PS1 mediates the neuroprotective activities of efnB and BDNF against excitotoxicity and regulates surface expression and ligand-induced metabolism of their cognate receptors. Together, our observations indicate that PS1 promotes neuronal survival by regulating neuroprotective functions of ligand-receptor systems.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/farmacologia , Córtex Cerebral/metabolismo , Efrina-B2/farmacologia , Neurônios/metabolismo , Presenilina-1/metabolismo , Receptor EphB2/metabolismo , Receptor trkB/metabolismo , Animais , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Córtex Cerebral/citologia , Córtex Cerebral/efeitos dos fármacos , Camundongos , Camundongos Knockout , Neurônios/citologia , Neurônios/efeitos dos fármacos , Presenilina-1/genética , Ratos , Receptor EphB2/genética , Receptor trkB/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
4.
Aging Cell ; 11(4): 683-93, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22577879

RESUMO

In normal retinas, amyloid-ß (Aß) accumulates in the subretinal space, at the interface of the retinal pigment epithelium, and the photoreceptor outer segments. However, the molecular and cellular effects of subretinal Aß remain inadequately elucidated. We previously showed that subretinal injection of Aß(1-42) induces retinal inflammation, followed by photoreceptor cell death. The retinal Müller glial (RMG) cells, which are the principal retinal glial cells, are metabolically coupled to photoreceptors. Their role in the maintenance of retinal water/potassium and glutamate homeostasis makes them important players in photoreceptor survival. This study investigated the effects of subretinal Aß(1-42) on RMG cells and of Aß(1-42)-induced inflammation on retinal homeostasis. RMG cell gliosis (upregulation of GFAP, vimentin, and nestin) on day 1 postinjection and a proinflammatory phenotype were the first signs of retinal alteration induced by Aß(1-42). On day 3, we detected modifications in the protein expression patterns of cyclooxygenase 2 (COX-2), glutamine synthetase (GS), Kir4.1 [the inwardly rectifying potassium (Kir) channel], and aquaporin (AQP)-4 water channels in RMG cells and of the photoreceptor-associated AQP-1. The integrity of the blood-retina barrier was compromised and retinal edema developed. Aß(1-42) induced endoplasmic reticulum stress associated with sustained upregulation of the proapoptotic factors of the unfolded protein response and persistent photoreceptor apoptosis. Indomethacin treatment decreased inflammation and reversed the Aß(1-42)-induced gliosis and modifications in the expression patterns of COX-2, Kir4.1, and AQP-1, but not of AQP-4 or GS. Nor did it improve edema. Our study pinpoints the adaptive response to Aß of specific RMG cell functions.


Assuntos
Peptídeos beta-Amiloides/administração & dosagem , Gliose/patologia , Inflamação/patologia , Fragmentos de Peptídeos/administração & dosagem , Degeneração Retiniana/patologia , Peptídeos beta-Amiloides/toxicidade , Animais , Apoptose/efeitos dos fármacos , Barreira Hematorretiniana/efeitos dos fármacos , Barreira Hematorretiniana/patologia , Barreira Hematorretiniana/fisiopatologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Fragmentos de Peptídeos/toxicidade , Células Fotorreceptoras de Vertebrados/efeitos dos fármacos , Células Fotorreceptoras de Vertebrados/patologia , Células Fotorreceptoras de Vertebrados/fisiologia , Retina/efeitos dos fármacos , Retina/patologia , Retina/fisiopatologia , Degeneração Retiniana/genética , Degeneração Retiniana/fisiopatologia
6.
Neurobiol Aging ; 32(12): 2326.e5-16, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21820214

RESUMO

To reduce damage from toxic insults such as glutamate excitotoxicity and oxidative stresses, neurons may deploy an array of neuroprotective mechanisms. Recent reports show that progranulin (PGRN) gene null or missense mutations leading to inactive protein, are linked to frontotemporal lobar degeneration (FTLD), suggesting that survival of certain neuronal populations needs full expression of functional PGRN. Here we show that extracellular PGRN stimulates phosphorylation/activation of the neuronal MEK/extracellular regulated kinase (ERK)/p90 ribosomal S6 kinase (p90RSK) and phosphatidylinositol-3 kinase (PI3K)/Akt cell survival pathways and rescues cortical neurons from cell death induced by glutamate or oxidative stress. Pharmacological inhibition of MEK/ERK/p90RSK signaling blocks the PGRN-induced phosphorylation and neuroprotection against glutamate toxicity while inhibition of either MEK/ERK/p90RSK or PI3K/Akt blocks PGRN protection against neurotoxin MPP(+). Inhibition of both pathways had synergistic effects on PGRN-dependent neuroprotection against MPP(+) toxicity suggesting both pathways contribute to the neuroprotective activities of PGRN. Extracellular PGRN is remarkably stable in neuronal cultures indicating neuroprotective activities are associated with full-length protein. Together, our data show that extracellular PGRN acts as a neuroprotective factor and support the hypothesis that in FTLD reduction of functional brain PGRN results in reduced survival signaling and decreased neuronal protection against excitotoxicity and oxidative stress leading to accelerated neuronal cell death. That extracellular PGRN has neuroprotective functions against toxic insults suggests that in vitro preparations of this protein may be used therapeutically.


Assuntos
Sobrevivência Celular/fisiologia , Córtex Cerebral/metabolismo , Líquido Extracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Neurônios/metabolismo , Fármacos Neuroprotetores/farmacologia , Transdução de Sinais/fisiologia , 1-Metil-4-fenilpiridínio/toxicidade , Animais , Células Cultivadas , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/fisiologia , Líquido Extracelular/efeitos dos fármacos , Espaço Extracelular/efeitos dos fármacos , Espaço Extracelular/fisiologia , Ácido Glutâmico/toxicidade , Células HEK293 , Humanos , Progranulinas , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos
7.
Neurobiol Dis ; 42(1): 55-72, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21220018

RESUMO

Age-related macular degeneration is characterized by the formation of drusen containing amyloid-ß (Aß) and the degeneration of photoreceptors. To explore the largely unknown role of Aß in the retina, we investigated the effects on photoreceptors of the oligomeric form of Aß(1-42). Subretinal injection of the Aß peptide induced misplaced expression of recoverin and synaptophysin in the photoreceptors, oxidative stress in their inner and outer segments, and finally apoptosis. Aß did not induce cell death in purified photoreceptor cell cultures, but did so in retinal cell cultures, thereby suggesting that the cellular environment plays a role in Aß-induced photoreceptor apoptosis. Subretinal injection of Aß was followed by activation and migration of microglial cells and then by photoreceptor apoptosis. Microglial cells phagocytosed rhodopsin-containing debris and Aß in the subretinal space. Quantitative RT-PCR allowed us to identify a specific gene expression profile associated with the Aß-induced progression of retinal degeneration and consistent with oxidative stress, inflammation, and an apoptotic program. The gene most highly upregulated in Aß-injected retinas was that for the chemokine CCL2, and its absence or that of its cognate receptor CCR2 greatly reduced migration of activated microglial cells to the site of retinal injury and profoundly worsened photoreceptor degeneration and disorganization of the retinal pigment epithelium in Aß-injected retinas. Our study pinpoints the roles of Aß and of CCL2/CCR2 axis-dependent inflammation in photoreceptor apoptosis.


Assuntos
Peptídeos beta-Amiloides/toxicidade , Apoptose/fisiologia , Quimiocina CCL2/genética , Citoproteção , Inflamação/metabolismo , Fragmentos de Peptídeos/toxicidade , Células Fotorreceptoras de Vertebrados/metabolismo , Células Fotorreceptoras de Vertebrados/patologia , Receptores CCR2/genética , Animais , Quimiocina CCL2/deficiência , Citoproteção/genética , Humanos , Inflamação/genética , Inflamação/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores CCR2/deficiência
8.
Acta Neuropathol ; 121(3): 351-63, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20978902

RESUMO

Very few studies have examined expression and function of amyloid precursor protein (APP) in the retina. We showed that APP mRNA and protein are expressed according to the different waves of retinal differentiation. Depletion of App led to an absence of amacrine cells, a 50% increase in the number of horizontal cells and alteration of the synapses. The retinas of adult APP(-/-) mice showed only half as many glycinergic amacrine cells as wild-type retinas. We identified Ptf1a, which plays a role in controlling both amacrine and horizontal cell fates, as a downstream effector of APP. The observation of a similar phenotype in sorLA knockout mice, a major regulator of APP processing, suggests that regulation of APP functions via sorLA controls the determination of amacrine and horizontal cell fate. These findings provide novel insights that indicate that APP plays an important role in retinal differentiation.


Assuntos
Precursor de Proteína beta-Amiloide/fisiologia , Diferenciação Celular/fisiologia , Retina/embriologia , Retina/crescimento & desenvolvimento , Envelhecimento/fisiologia , Células Amácrinas/citologia , Células Amácrinas/fisiologia , Animais , Proliferação de Células , Camundongos , Camundongos Knockout , Modelos Animais , Retina/citologia , Células Horizontais da Retina/citologia , Células Horizontais da Retina/fisiologia , Sinapses/fisiologia , Fatores de Transcrição/fisiologia
9.
Aging Cell ; 8(2): 162-77, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19239420

RESUMO

Age-related macular degeneration (AMD) is characterized by the formation of drusen, extracellular deposits associated with atrophy of the retinal pigmented epithelium (RPE), disturbance of the transepithelial barrier and photoreceptor death. Amyloid-beta (Abeta) is present in drusen but its role during AMD remains unknown. This study investigated the in vitro and in vivo effects of the oligomeric form of Abeta(1-42) - OAbeta(1-42) - on RPE and found that it reduced mitochondrial redox potential and increased the production of reactive oxygen species, but did not induce apoptosis in RPE cell cultures. It also disorganized the actin cytoskeleton and halved occludin expression, markedly decreasing attachment capacity and abolishing the selectivity of RPE cell transepithelial permeability. Antioxidant pretreatment partially reversed the effects of OAbeta(1-42) on mitochondrial redox potential and transepithelial permeability. Subretinally injected OAbeta(1-42) induced pigmentation loss and RPE hypertrophy but not RPE cell apoptosis in C57BL/6 J mice. Rapid OAbeta(1-42)-induced disorganization of cytoskeletal actin filaments was accompanied by decreased RPE expression of the tight junction proteins occludin and zonula occludens-1 and of the visual cycle proteins cellular retinaldehyde-binding protein and RPE65. The number of photoreceptors decreased by half within a few days. Our study pinpoints the role of Abeta in RPE alterations and dysfunctions leading to retinal degeneration and suggests that targeting Abeta may help develop selective methods for treating diseases involving retinal degeneration, such as AMD.


Assuntos
Peptídeos beta-Amiloides/toxicidade , Degeneração Macular/fisiopatologia , Estresse Oxidativo/efeitos dos fármacos , Fragmentos de Peptídeos/toxicidade , Epitélio Pigmentado da Retina/efeitos dos fármacos , Epitélio Pigmentado da Retina/patologia , Envelhecimento/metabolismo , Envelhecimento/patologia , Peptídeos beta-Amiloides/metabolismo , Animais , Proteínas de Transporte/efeitos dos fármacos , Proteínas de Transporte/metabolismo , Linhagem Celular , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Citoesqueleto/patologia , Proteínas do Olho/efeitos dos fármacos , Proteínas do Olho/metabolismo , Humanos , Hipertrofia/induzido quimicamente , Hipertrofia/metabolismo , Hipertrofia/fisiopatologia , Degeneração Macular/induzido quimicamente , Degeneração Macular/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Potencial da Membrana Mitocondrial/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Estresse Oxidativo/fisiologia , Fragmentos de Peptídeos/metabolismo , Células Fotorreceptoras de Vertebrados/efeitos dos fármacos , Células Fotorreceptoras de Vertebrados/metabolismo , Células Fotorreceptoras de Vertebrados/patologia , Espécies Reativas de Oxigênio/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismo , Junções Íntimas/patologia , cis-trans-Isomerases
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