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APOE4-carrying human astrocytes oversupply cholesterol to promote neuronal lipid raft expansion and Aβ generation.

ABSTRACT: The ε4 allele of APOE-encoding apolipoprotein (ApoE) is one of the strongest genetic risk factors for Alzheimer's disease (AD). One of the overarching questions is whether and how this astrocyte-enriched risk factor initiates AD-associated pathology in neurons such as amyloid-β (Aβ) accumulation. Here, we generate neurons and astrocytes from isogenic human induced pluripotent stem cells (hiPSCs) carrying either APOE ε3 or APOE ε4 allele and investigate the effect of astrocytic ApoE4 on neuronal Aβ production. Secretory factors in conditioned media from ApoE4 astrocytes significantly increased amyloid precursor protein (APP) levels and Aβ secretion in neurons. We further found that increased cholesterol secretion from ApoE4 astrocytes was necessary and sufficient to induce the formation of lipid rafts that potentially provide a physical platform for APP localization and facilitate its processing. Our study reveals the contribution of ApoE4 astrocytes to amyloidosis in neurons by expanding lipid rafts and facilitating Aβ production through an oversupply of cholesterol.

SUBMITTER: Lee SI

PROVIDER: S-EPMC8452535 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

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APOE4-carrying human astrocytes oversupply cholesterol to promote neuronal lipid raft expansion and Aβ generation.

Lee Se-In SI Jeong Woojin W Lim Heejin H Cho Sukhee S Lee Hyein H Jang Yonghee Y Cho Joonho J Bae Simsung S Lin Yuan-Ta YT Tsai Li-Huei LH Moon Dae Won DW Seo Jinsoo J

Stem cell reports 20210826 9

The ε4 allele of APOE-encoding apolipoprotein (ApoE) is one of the strongest genetic risk factors for Alzheimer's disease (AD). One of the overarching questions is whether and how this astrocyte-enriched risk factor initiates AD-associated pathology in neurons such as amyloid-β (Aβ) accumulation. Here, we generate neurons and astrocytes from isogenic human induced pluripotent stem cells (hiPSCs) carrying either APOE ε3 or APOE ε4 allele and investigate the effect of astrocytic ApoE4 on neuronal ...[more]

PMID: 34450034

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Project description:Astrocytes are important glia cell type in the central nervous system. These cells can undergo transformation to a reactive state upon injury such as focal ischemic stroke (FIS). Reactive astrocytes are distinct from normal or homeostatic astrocytes in morphology, protein profiles and metabolic functions. Glial cell-derived neurotrophic factor (GDNF) was discovered as a potent survival neurotrophic factor for multiple subtypes of neurons and can be released from reactive astrocytes. In our previous study, we found that GDNF expression was upregulated in reactive astrocytes following ischemic stroke. Specific knock out of GDNF in reactive astrocytes exacerbated brain damage and motor deficits after ischemic stroke. Here, using in vitro and in vivo ischemia models, we investigated the effects of GDNF overexpression in astrocytes on neuronal survival and brain recovery after ischemia. We observed that astrocyte specific GDNF overexpression by viral transduction could decrease brain infarction and promote motor function recovery after photothrombosis (PT)-induced FIS. In addition, GDNF overexpression in astrocytes could increase the proliferation of reactive astrocytes and reduce oxidative stress after PT. Using the oxygen-glucose deprivation (OGD) model of cultured astrocytes, we confirmed that this ischemic insult could upregulate GDNF expression and increase its release to extracellular space. Transfection of GDNF DNA plasmid could further increase GDNF release after OGD. To further study the effects of reactive astrocytes-derived extracellular GDNF on neuronal survival after ischemia, cultured neurons subjected to OGD were exposed to astrocyte conditioned medium (ACM). The ACM collected from OGD subjected astrocyte culture could significantly reduce neuronal death, while neutralizing antibodies against GDNF and its receptors including GFRα1, RET and p-RET could suppress this beneficial effect. We also found that reactive astrocytes-derived GDNF could trigger the activation of RET receptors in cultured neurons and suppress neuronal mitochondrial fission and caspase-dependent cell apoptosis after OGD. Overall, our results indicate that reactive astrocytes-derived GDNF could play an important role in neuronal survival and functional recovery and underscore the non-cell autonomy underlying astrocyte-neuron interactions in brain repair after ischemic stroke.

Project description:BackgroundRecent findings suggest that the pathological effects of apoE4, the most prevalent genetic risk factor for Alzheimer's disease (AD), start many years before the onset of the disease and are already detectable at a young age. In the present study we investigated the extent to which such pathological and cognitive impairments also occur in young apoE4 mice.ResultsThis study revealed that the levels of the presynaptic glutamatergic vesicular transporter, VGlut, in the CA3, CA1, and DG hippocampal subfields were lower in hippocampal neurons of young (4-month-old) apoE4-targeted replacement mice than in those of the apoE3 mice. In contrast, the corresponding inhibitory GABAergic nerve terminals and perikarya were not affected by apoE4.This synaptic effect was associated with hyperphosphorylation of tau in these neurons. In addition, apoE4 increased the accumulation of neuronal Aβ42 and induced mitochondrial changes, both of which were specifically pronounced in CA3 neurons. Spatial navigation behavioral studies revealed that these hippocampal pathological effects of apoE4 are associated with corresponding behavioral impairments. Time-course studies revealed that the effects of apoE4 on tau hyperphosphorylation and the mitochondria were already apparent at the age of 1 month and that the apoE4-driven accumulation of neuronal Aβ and reduced VGlut levels evolve later and are apparent at the age of 2-4 months. Furthermore, the levels of tau phosphorylation decrease in apoE3 mice and increase in apoE4 mice between 1 and 4 months, whereas the levels of Aβ42 decrease in apoE3 mice and are not affected in apoE4 mice over the same time period.ConclusionsThese findings show that apoE4 stimulates the accumulation of Aβ42 and hyperphosphorylated tau and reduces the levels of VGlut in hippocampal neurons of young apoE4-targeted replacement mice and that these neurochemical effects are associated with cognitive impairments. This model is not associated with hypothesis-driven mechanistic manipulations and is thus most suitable for unbiased studies of the mechanisms underlying the pathological effects of apoE4.

Project description:Background: The beta-amyloid peptide (Aβ) involved in Alzheimer's disease (AD) has been described to associate/aggregate on the cell surface disrupting the membrane through pore formation and breakage. However, molecular determinants involved for this interaction (e.g., some physicochemical properties of the cell membrane) are largely unknown. Since cholesterol is an important molecule for membrane structure and fluidity, we examined the effect of varying cholesterol content with the association and membrane perforation by Aβ in cultured hippocampal neurons. Methods: To decrease or increase the levels of cholesterol in the membrane we used methyl-β-cyclodextrin (MβCD) and MβCD/cholesterol, respectively. We analyzed if membrane fluidity was affected using generalized polarization (GP) imaging and the fluorescent dye di-4-ANEPPDHQ. Additionally membrane association and perforation was assessed using immunocytochemistry and electrophysiological techniques, respectively. Results: The results showed that cholesterol removal decreased the macroscopic association of Aβ to neuronal membranes (fluorescent-puncta/20 μm: control = 18 ± 2 vs. MβCD = 10 ± 1, p < 0.05) and induced a facilitation of the membrane perforation by Aβ with respect to control cells (half-time for maximal charge transferred: control = 7.2 vs. MβCD = 4.4). Under this condition, we found an increase in membrane fluidity (46 ± 3.3% decrease in GP value, p < 0.001). On the contrary, increasing cholesterol levels incremented membrane rigidity (38 ± 2.7% increase in GP value, p < 0.001) and enhanced the association and clustering of Aβ (fluorescent-puncta/20 μm: control = 18 ± 2 vs. MβCD = 10 ± 1, p < 0.01), but inhibited membrane disruption. Conclusion: Our results strongly support the significance of plasma membrane organization in the toxic effects of Aβ in hippocampal neurons, since fluidity can regulate distribution and insertion of the Aβ peptide in the neuronal membrane.

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APOE4-carrying human astrocytes oversupply cholesterol to promote neuronal lipid raft expansion and Aβ generation.

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APOE4-carrying human astrocytes oversupply cholesterol to promote neuronal lipid raft expansion and Aβ generation.

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