Human apolipoprotein E (apoE) exists as three main isoforms called apoE2, apoE3, and apoE4, of which the E4 isoform is associated with increased Alzheimer’s disease (AD) risk. Brain glucose hypometabolism, linked to synaptic dysfunction, occurs years before symptom onset in AD, especially in APOEε4-carriers. An association between a higher ratio of plasma apoE4 to apoE3 levels and cerebral glucose hypometabolism was recently discovered in cognitively healthy APOEε3/ε4 subjects. A lower plasma apoE level, regardless of isoform, is linked to increased AD risk. How the plasma apoE level affects neurodegenerative processes in the brain is poorly understood, given that apoE doesn’t cross the blood-brain barrier (BBB). The main aim of this thesis was therefore to investigate a relationship between plasma apoE and features of AD pathophysiology. We explored plasma apoE levels and dimer/monomer formation in APOEε3 and APOEε4 homozygous controls, in patients with mild cognitive impairment (MCI) and AD. In APOEε4-carriers versus non-carriers, plasma apoE levels were lower and significantly correlated with AD biomarkers. ApoE3 homodimers were less in AD patients than in controls. We next examined potential links between plasma apoE, glucose, and insulin levels in the previously examined cognitively healthy APOEε3/ε4 subjects. Lower plasma apoE3 was associated with higher glucose levels in males and subjects with body max index above 25. Plasma glucose levels were negatively correlated with the cerebral metabolic rate of glucose and neuropsychological test scores. To explore the potential effects of a hepatic APOEε4 phenotype on the brain, we compared liver humanized mice with an APOEε4/ε4 versus an APOEε2/ε3 genotype. Mice with an APOEε4/ε4 liver exhibited reduced endogenous mouse apoE in the brain, accompanied by changes in markers of synaptic integrity and insulin signaling. Plasma apoE4 levels were negatively associated with some of the assessed markers. We further explored the effects of a high-fat diet (HFD) in mice with livers humanized with the AD risk-neutral APOEε3/ε3 genotype. Endogenous mouse apoE was increased in the hippocampus following an HFD, with concomitant effects on levels of synaptic markers. In the cortex, we found altered levels of insulin signaling and synaptic markers. Together, our findings indicate that alterations in apoE levels or distribution, hepatic APOEε4 phenotype, and HFD contribute to AD-related pathological processes.

Amyloidogenic processing of the amyloid precursor protein (APP) gives rise to Aβ peptides that assemble into the Aβ plaques found in AD. The binding of the adaptor protein Fe65, through its PTB2, to APP might enhance amyloidogenic APP processing. Fe65 is localized both in the cytoplasm and in the nucleus, with compartment-specific biological functions. Mechanisms affecting Fe65 subcellular localization are poorly understood. We explored the impact of the different Fe65 interaction domains WW and PTB2 and APP processing on Fe65 cellular localization. By transfecting Fe65-domain deletion constructs into neuroblastoma cell lines, we found that deleting the PTB2 domain almost abolished nuclear localization. Upon pharmacological inhibition of APP secretases, we found decreased Fe65 localization to the nucleus. To conclude, Fe65-APP interaction and APP processing may be important factors governing the Fe65 cellular localization.

The apolipoprotein E gene (APOE) is expressed as three major isoforms, APOE ε2, ε3 and ε4 that encode the apoE2, E3 and E4 proteins, respectively. APOE ε4 is associated with an increased risk for age-related neurodegenerative diseases including Alzheimer’s disease (AD) and dementia with Lewy bodies (DLB), and increases the risk of dementia in Parkinson’s disease (PD), however, the exact molecular mechanisms underpinning the associations between APOE ε4 and neurodegeneration are still unknown. Pathologically, α-synuclein (αSyn) is the primary component of Lewy neuropathology which is stereotypically found in PD, DLB and also in the majority of AD brains upon post mortem examination, and changes in the soluble levels of intra- and extracellular αSyn in the central nervous system have been found in AD patients compared to cognitively healthy individuals. With this in mind, we hypothesized that interactions, both direct and indirect, between apoE4 / APOE ε4 and αSyn might help explain their associations with the pathogenesis of multiple neurodegenerative diseases. Using mouse models with a humanized APOE ε4/ε4 or ε3/ε3 liver genotype we found that changing the liver genotype to APOE ε4/ε4 led to a re-compartmentalization of brain αSyn and significant changes in key pre- and postsynaptic protein levels We also found that in prodromal sporadic AD patients APOE ε4 carriers had significantly increased cerebrospinal fluid (CSF) αSyn levels, and in autosomal dominant AD APOE ε4 carriers higher CSF αSyn levels that were correlated with AD-onset and increasing brain amyloid-β plaque deposition. When next assessing any disease related changes in the molecular sizes of αSyn and apoE in AD and PD post mortem brain parenchyma we found that the amygdala had a differing distribution of αSyn and apoE4 species. Co-incubation of the recombinant apoE isoforms and αSyn led to high molecular weight αSyn species, increased high molecular weight apoE2 species and a stabilized pool of monomeric apoE3. Using, binding kinetics assays and human neuronal cell cultures we found that recombinant apoE isoforms bind with similar strength to αSyn and reduce αSyn cellular uptake, whereas astrocyte secreted apoE led to an apoE- isoform-dependent reduction in αSyn uptake (apoE4 > apoE3 > apoE2). Further, heparin sulfate proteoglycans were partially responsible for αSyn uptake mediated by apoE. Taken together, our results show that APOE / apoE is linked to changes in levels, species and compartmentalization of αSyn in multiple pathological contexts ranging from the central nervus system to the periphery in humans, mouse models and cell lines. Our data reinforce the growing body of literature linking APOE / apoE and αSyn to the pathogenesis of the most prevalent neurodegenerative diseases.