Which isoforms of apolipoprotein E (apoE) we inherit determine our risk of developing late-onset Alzheimer’s Disease (AD), but the mechanism underlying this link is poorly understood. In particular, the relevance of direct interactions between apoE and amyloid-β (Aβ) remains controversial. Here, single-molecule imaging shows that all isoforms of apoE associate with Aβ in the early stages of aggregation and then fall away as fibrillation happens. ApoE-Aβ co-aggregates account for ~ 50% of the mass of soluble Aβ aggregates detected in the frontal cortices of homozygotes with the higher-risk APOE4 gene. Our results connect inherited APOE genotype with the risk of developing AD by demonstrating how, in an isoform- and lipidation-specific way, apoE modulates the aggregation, clearance and toxicity of Aβ. Selectively removing non-lipidated apoE4-Aβ co-aggregates enhances clearance of toxic Aβ by glial cells, and reduces inflammation and membrane damage, demonstrating a clear path to AD therapeutics.

Background: Astrocytes have multiple roles including providing neurons with metabolic substrates and maintaining neurotransmitter synaptic homeostasis. Astrocyte glucose metabolism plays a key role in learning and memory with astrocytic glycogen a key substrate supporting memory encoding. The neuronal support provided by astrocytes has a high metabolic demand. Deficits in astrocytic mitochondrial metabolic functioning and glycolysis could impair neuronal function. Changes to cellular metabolism are seen early in Alzheimers disease (AD). Understanding cellular metabolism changes in AD astrocytes could be exploited as a new biomarker or synergistic therapeutic agent when combined with anti-amyloid treatments in AD. Methods: In this project, we characterised mitochondrial and glycolytic function in astrocytes derived from patients with sporadic (n=6) and familial (PSEN1, n=3) forms of AD. Astrocytes were derived using direct reprogramming methods. Astrocyte metabolic outputs: ATP, and extracellular lactate levels were measured using luminescent and fluorescent protocols. Mitochondrial respiration and glycolytic function were measured using a Seahorse XF Analyzer. Hexokinase deficits identified where corrected by transfecting astrocytes with an adenovirus viral vector containing the hexokinase 1 gene. Results: There was a reduction of total cellular ATP of 20% (p=0.05 in sAD astrocytes) and of 48% (p<0.01) in fAD. A 44% reduction (p<0.05), and 80% reduction in mitochondrial spare capacity was seen in sAD and fAD astrocytes respectively. Reactive oxygen species (ROS) were increased in both AD astrocyte types (p=0.05). Mitochondrial complex I and II was significantly increased in sAD (p<0.05) but not in fAD. Astrocyte glycolytic reserve and extracellular lactate was significantly reduced when compared to controls in both sAD and fAD (p<0.05). We identified a deficit in the glycolytic pathway enzyme hexokinase, and correcting this deficit restored most of the metabolic phenotype in sAD but not fAD astrocytes. Conclusion: AD astrocytes have abnormalities in functional capacity of mitochondria and the process of glycolysis. These functional deficits can be improved by correcting hexokinase expression deficits with adenoviral vectors. This suggests that hexokinase 1 deficiency could potentially be exploited as a new therapeutic target for AD.