RESUMO
Alzheimer's disease (AD) is the most common progressive neurodegenerative disorder causing dementia. Massive deposition of amyloid ß peptide (Aß) as senile plaques in the brain is the pathological hallmark of AD, but oligomeric, soluble forms of Aß have been implicated as the synaptotoxic component. The apolipoprotein E ε 4 (apoE ε4) allele is known to be a genetic risk factor for developing AD. However, it is still unknown how apoE impacts the process of Aß oligomerization. Here, we found that the level of Aß oligomers in APOE ε4/ε4 AD patient brains is 2.7 times higher than those in APOE ε3/ε3 AD patient brains, matched for total plaque burden, suggesting that apoE4 impacts the metabolism of Aß oligomers. To test this hypothesis, we examined the effect of apoE on Aß oligomer formation. Using both synthetic Aß and a split-luciferase method for monitoring Aß oligomers, we observed that apoE increased the level of Aß oligomers in an isoform-dependent manner (E2 < E3 < E4). This effect appears to be dependent on the ApoE C-terminal domain. Moreover, these results were confirmed using endogenous apoE isolated from the TBS-soluble fraction of human brain, which increased the formation of Aß oligomers. Together, these data show that lipidated apoE, especially apoE4, increases Aß oligomers in the brain. Higher levels of Aß oligomers in the brains of APOE ε4/ε4 carriers compared with APOE ε3/ε3 carriers may increase the loss of dendritic spines and accelerate memory impairments, leading to earlier cognitive decline in AD.
Assuntos
Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Apolipoproteína E4/genética , Apolipoproteína E4/metabolismo , Encéfalo/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/genética , Peptídeos beta-Amiloides/farmacologia , Análise de Variância , Apolipoproteína E2/genética , Apolipoproteína E2/metabolismo , Apolipoproteína E3/genética , Apolipoproteína E3/metabolismo , Astrócitos/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Proteínas de Fluorescência Verde/genética , Células HEK293/efeitos dos fármacos , Células HEK293/metabolismo , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/genética , Masculino , Morfolinos/farmacologia , Fragmentos de Peptídeos/farmacologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , TransfecçãoRESUMO
The apolipoprotein E ε4 gene is the most important genetic risk factor for sporadic Alzheimer's disease, but the link between this gene and neurodegeneration remains unclear. Using array tomography, we analysed >50000 synapses in brains of 11 patients with Alzheimer's disease and five non-demented control subjects and found that synapse loss around senile plaques in Alzheimer's disease correlates with the burden of oligomeric amyloid-ß in the neuropil and that this synaptotoxic oligomerized peptide is present at a subset of synapses. Further analysis reveals apolipoprotein E ε4 patients with Alzheimer's disease have significantly higher oligomeric amyloid-ß burden and exacerbated synapse loss around plaques compared with apolipoprotein E ε3 patients. Apolipoprotein E4 protein colocalizes with oligomeric amyloid-ß and enhances synaptic localization of oligomeric amyloid-ß by >5-fold. Biochemical characterization shows that the amyloid-ß enriched at synapses by apolipoprotein E4 includes sodium dodecyl sulphate-stable dimers and trimers. In mouse primary neuronal culture, lipidated apolipoprotein E4 enhances oligomeric amyloid-ß association with synapses via a mechanism involving apolipoprotein E receptors. Together, these data suggest that apolipoprotein E4 is a co-factor that enhances the toxicity of oligomeric amyloid-ß both by increasing its levels and directing it to synapses, providing a link between apolipoprotein E ε4 genotype and synapse loss, a major correlate of cognitive decline in Alzheimer's disease.
Assuntos
Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/fisiologia , Apolipoproteína E4/metabolismo , Degeneração Neural/metabolismo , Sinapses/patologia , Doença de Alzheimer/genética , Peptídeos beta-Amiloides/metabolismo , Apolipoproteína E3/genética , Apolipoproteína E4/genética , Estudos de Casos e Controles , Genótipo , Humanos , Neurônios/metabolismo , Placa Amiloide/metabolismo , Cultura Primária de Células , Transporte Proteico , Sinapses/metabolismoRESUMO
The paradoxical appearance of aggregated α-synuclein (αsyn) in naive transplanted embryonic stem cells in Parkinson's disease (PD) brains has recently been reported, highlighting the possibility of neuron to neuron transmission of αsyn in PD. Here, we demonstrate in a cellular model the presence of αsyn oligomers in the extracellular space, their uptake by neurons, retrograde axonal transport to cell soma, and detrimental effects on neighboring cells. Moreover, we demonstrate that Hsp70 chaperones αsyn in the extracellular space and reduces extracellular αsyn oligomer formation and related toxicity. These novel findings provide evidence that extracellular αsyn oligomers may represent a crucial player in the propagation of pathology in PD, with their modulation by Hsp70 representing a potential new target for therapeutic interventions.
Assuntos
Proteínas de Choque Térmico HSP70/metabolismo , Neurônios/fisiologia , Transmissão Sináptica/fisiologia , alfa-Sinucleína/metabolismo , Animais , Axônios/metabolismo , Benzoquinonas/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Meios de Cultivo Condicionados/farmacologia , Dependovirus/genética , Espaço Extracelular/metabolismo , Vetores Genéticos/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Proteínas de Choque Térmico HSP70/antagonistas & inibidores , Proteínas de Choque Térmico HSP70/genética , Humanos , Lactamas Macrocíclicas/farmacologia , Luciferases/genética , Luciferases/metabolismo , Camundongos , Neurônios/citologia , Neurônios/metabolismo , Multimerização Proteica , Transfecção , alfa-Sinucleína/química , alfa-Sinucleína/genéticaRESUMO
Tau protein is present in six different splice forms in the human brain and interacts with microtubules via either 3 or 4 microtubule binding repeats. An increased ratio of 3 repeat to 4 repeat isoforms is associated with neurodegeneration in inherited forms of frontotemporal dementia. Tau over-expression diminishes axonal transport in several systems, but differential effects of 3 repeat and 4 repeat isoforms have not been studied. We examined the effects of tau on mitochondrial transport and found that both 3 repeat and 4 repeat tau change normal mitochondrial distribution within the cell body and reduce mitochondrial localization to axons; 4 repeat tau has a greater effect than 3 repeat tau. Further, we observed that the 3 repeat and 4 repeat tau cause different alterations in retrograde and anterograde transport dynamics with 3 repeat tau having a slightly stronger effect on axon transport dynamics. Our results indicate that tau-induced changes in axonal transport may be an underlying theme in neurodegenerative diseases associated with isoform specific changes in tau's interaction with microtubules.
Assuntos
Transporte Axonal/fisiologia , Mitocôndrias/fisiologia , Neurônios/fisiologia , Tauopatias/fisiopatologia , Proteínas tau/genética , Proteínas tau/metabolismo , Animais , Linhagem Celular Tumoral , Córtex Cerebral/citologia , Glioma , Proteínas de Fluorescência Verde/genética , Humanos , Camundongos , Técnicas Analíticas Microfluídicas , Neurônios/citologia , Transporte Proteico/fisiologia , Sequências Repetitivas de Ácido Nucleico/fisiologia , Tauopatias/genética , Tauopatias/metabolismo , TransfecçãoRESUMO
The ability of the low density lipoprotein receptor-related protein (LRP) to form homo-dimers was studied in mouse neuroblastoma and human neuroglioma cells as well as in primary cortical cultures from adult mouse brain. Homo-dimerization of LRP light chain (LC) was shown by several methods including co-immunoprecipitation, fluorescence lifetime imaging microscopy, and bimolecular fluorescence complementation assay. The requirement of intact NPXY motifs of LRP LC for homo-dimerization was ruled out by co-immunoprecipitation assay.