The APOE 4 allele is the strongest genetic risk factor for late-onset Alzheimer's disease (AD). ApoE protein aggregation plays a central role in AD pathology, including the accumulation of ␤-amyloid (A␤). Lipid-poor ApoE4 protein is prone to aggregate and lipidating ApoE4 protects it from aggregation. The mechanisms regulating ApoE4 aggregation in vivo are surprisingly not known. ApoE lipidation is controlled by the activity of the ATP binding cassette A1 (ABCA1). ABCA1 recycling and degradation is regulated by ADP-ribosylation factor 6 (ARF6). We found that ApoE4 promoted greater expression of ARF6 compared with ApoE3, trapping ABCA1 in late-endosomes and impairing its recycling to the cell membrane. This was associated with lower ABCA1-mediated cholesterol efflux activity, a greater percentage of lipid-free ApoE particles, and lower A␤ degradation capacity. Human CSF from APOE 4/4 carriers showed a lower ability to induce ABCA1-mediated cholesterol efflux activity and greater percentage of aggregated ApoE protein compared with CSF from APOE 3/3 carriers. Enhancing ABCA1 activity rescued impaired A␤ degradation in ApoE4-treated cells and reduced both ApoE and ABCA1 aggregation in the hippocampus of male ApoE4-targeted replacement mice. Together, our data demonstrate that aggregated and lipidpoor ApoE4 increases ABCA1 aggregation and decreases ABCA1 cell membrane recycling. Enhancing ABCA1 activity to reduce ApoE and ABCA1 aggregation is a potential therapeutic strategy for the prevention of ApoE4 aggregation-driven pathology.

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.

The allele ɛ4 of apolipoprotein E (apoE4) is the most prevalent genetic risk factor for Alzheimer's disease (AD) and is therefore a promising therapeutic target. Human and animal model studies suggest that apoE4 is hypolipidated; accordingly, we have previously shown that the retinoid X receptor (RXR) agonist bexarotene upregulates ABCA1, the main apoE-lipidating protein, resulting in increased lipidation of apoE4, and the subsequent reversal of the pathological effects of apoE4, namely: accumulation of Aβ42 and hyperphosphorylated tau, as well as reduction in the levels of synaptic markers and cognitive deficits. Since the RXR system has numerous other targets, it is important to devise the means of activating ABCA1 selectively. We presently utilized CS-6253, a peptide shown to directly activate ABCA1 in vitro, and examined the extent to which it can affect the degree of lipidation of apoE4 in vivo and counteract the associated brain and behavioral pathologies. This revealed that treatment of young apoE4-targeted replacement mice with CS-6253 increases the lipidation of apoE4. This was associated with a reversal of the apoE4-driven Aβ42 accumulation and tau hyperphosphorylation in hippocampal neurons, as well as of the synaptic impairments and cognitive deficits. These findings suggest that the pathological effects of apoE4 in vivo are associated with decreased activation of ABCA1 and impaired lipidation of apoE4 and that the downstream brain-related pathology and cognitive deficits can be counteracted by treatment with the ABCA1 agonist CS-6253. These findings have important clinical ramifications and put forward ABCA1 as a promising target for apoE4-related treatment of AD.

We presently investigated the effects of apolipoprotein E4 (apoE4), the most prevalent genetic risk factor for Alzheimer's disease, on the cognitive performance of young targeted replacement apoE4 mice. We revealed that these mice were impaired in the object recognition and Morris water maze tests, both of which are associated with hippocampal learning and memory, relative to that of the apoE3 mice. These results are consistent with previous histological and biochemical findings that hippocampal neurons are specifically affected by apoE4. The suggestion that the behavioral impairments of the apoE4 mice are related to the hippocampal neuropathology of these mice is further supported by the fear conditioning test. This test revealed that the performance of the apoE4 mice in the contextual component, which is hippocampus related, was impaired, whereas their cued test response, which is amygdala driven, was not. The stress levels of the apoE4 and apoE3 mice, as unraveled by the light/dark anxiety test, were similar, suggesting that the observed cognitive impairments of the apoE4 mice are not related to differences in the basal anxiety levels of these mice. In conclusion, the present study shows that young apoE4 targeted replacement mice are impaired in numerous hippocampus-related learning and memory tasks.

This study investigated the possible role of the ApoE receptors Lrp1 and Apoer2 in mediating the pathological effects of ApoE4 in ApoE-targeted-replacement mice expressing either the human ApoE3 or ApoE4 allele. In this study we show that activation of the amyloid cascade by inhibition of the Aβ-degrading enzyme neprilysin results in up-regulation of the ApoE receptor Lrp1 in the CA1 hippocampal neurons of 4-month-old ApoE4 mice, but not in the corresponding ApoE3 or ApoE-deficient (KO) mice. These results are in accordance with the previous findings that activation of the amyloid cascade induces Aβ accumulation in the CA1 neurons of ApoE4 mice, but not in ApoE3 or ApoE-KO mice. This suggests that the apoE4-driven elevation of Lrp1 is mediated via a gain of function mechanism and may play a role in mediating the effects of ApoE4 on Aβ. In contrast, no changes were observed in the levels of the corresponding Apoer2 receptor following the neprilysin inhibition. The ApoE receptors of naive ApoE4 mice were also affected differentially and isoform specifically by ApoE4. However, under these conditions, the effect was an ApoE4-driven reduction in the levels of Apoer2 in CA1 and CA3 pyramidal neurons, whereas the levels of Lrp1 were not affected. RT-PCR measurements revealed that the levels of Apoer2 and Lrp1 mRNA in the hippocampus of naïve and neprilysin-inhibited mice were not affected by ApoE4, suggesting that the observed effects of ApoE4 on the levels of these receptors is post-transcriptional. In conclusion, this study shows that the levels of hippocampal ApoE receptors Lrp1 and Apoer2 in vivo are affected isoform specifically by ApoE4 and that the type of receptor affected is context dependent.