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Project Topic
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Alzheimer’s disease (AD) is the most common form of dementia. Although traditional animal models of
AD have been extremely helpful in advancing our understanding of the cellular and molecular pathways
leading to neurodegeneration and cognitive decline, pharmaceutical or non-pharmaceutical options that
have a proven and significant impact on the incidence and progression of AD are still very scarce. One
reason for this translational roadblock is that most animal models typically do not adequately reflect the
complex and multifactorial pathophysiology of sporadic (late-onset) AD.
One particularly strong and common inherited risk factor for late-onset AD is the apolipoprotein E
(ApoE) ε4 genotype. In addition, many environmental and acquired risk factors – such as traumatic
brain injury, sleep disorders, systemic inflammation, dietary factors and vascular pathology – also
strongly amplify the risk for AD, but how they interact with ApoE and contribute to AD pathogenesis has
remained incompletely understood. In previous publications, we have found that astroglial derived ApoE
is critically involved in the microglial clearance of amyloid-β (Aβ), and that many pathways attributed to
ApoE function are also strongly modified by acquired risk factors – for example, that sleep, aging and
traumatic brain injury all strongly affect Aβ clearance from the brain; that chronic brain inflammation as
well as systemic inflammation both greatly modulate disease progression and Aβ phagocytosis; and that
chronic inflammation contributes to network dysfunction between neurons and astrocytes. Moreover, in
preliminary experiments, we have found that ApoE isoforms are differentially distributed and transported
through the brain's glymphatic clearance system.
The goal of this proposal is to provide a comprehensive analysis of "nature and nurture" based on
analysis of a novel AD mouse model that combines the most common genetic risk factor (ApoE-ε4) as
well as many of the most prevalent acquired risk factors. To this end, we have generated mouse lines
that express the human ApoE-ε4 or ApoE-ε2 isoforms in an APP/PS1 mouse model of AD. We
hypothesize that hallmarks of AD pathology are differentially modified by the presence of the diseaseaggravating
ε4 isoform or the disease-attenuating ε2 isoform compared to APP/PS1 mice with wildtype
mouse ApoE. Secondly, we will expose these mouse lines to environmental and acquired risk factors,
aiming to create models that more realistically reflect ‘real-world’ scenarios leading to AD. We
hypothesize that this combination of genetic and acquired risk factors strongly aggravates the initiation
and progression of pathological changes.
All partners of this proposal have a long-standing expertise and outstanding track records in a highly
complementary set of research questions and techniques related to AD pathophysiology including its
genetic and acquired risk factors. Therefore, in comparison to most other consortia, our approach will
allow us to investigate pathological changes in our models and their interactions with risk factors in a
shorter time frame, at a much deeper level and from many more angles. At the same time, our project will
be unified by use of the same mouse models and specific pre-defined common study endpoints.
In Work Package (WP) 1 of the proposal, we will investigate how expression of ApoE-ε4 or -ε2 isoforms
influence disease initiation and progression in the APP/PS1 model at different age points. Aβ metabolism
and pathology will be assessed using biochemical/electrochemiluminescence assays and
immunohistochemisty. Cerebral and systemic inflammation will be assessed by microglial phenotyping,
cytokine assays of brain, CSF and plasma, and immunohistochemisty. We will also investigate local Aβ
clearance through microglia (using FACS-based phagocytosis assays) and through the blood via the
glymphatic system (by radiolabeled tracer studies). Blood-brain barrier integrity and vessel density will be
studied using two-photon microscopy and ultrahigh-field MRI. Pericyte function will be assessed using
single-cell RNA-seq and novel reporter lines. The development and degree of cellular hypo- and
hyperactivity will be studied using genetic calcium indicators with in vivo two-photon microscopy in awake
mice. Finally, we will investigate behavioral changes using a battery of tests (open field, novel object
recognition, Morris water maze, elevated plus maze), and we will also study electrophysiological
correlates of memory by in vitro hippocampal recordings. This comprehensive characterization will serve
as a reference point for future studies using this model, and will be the foundation of the second part of
our proposal.
In WP 2, we will expose these lines to established acquired risk factors to determine if and through which
pathways neurodegeneration and cognitive decline are modified following a combined impact of genetic
and environmental risk factors. To this end, our mouse lines will be subjected to: high-fat or 'western'
diets; traumatic brain injury, using open and closed skull models of cortical injury; systemic inflammation,
using lipopolysaccharide treatment; vascular pathology, using a carotid artery stenosis model of chronic
hypoxia and a model of embolic cerebral microinfarcts; and perturbed sleep homoeostasis. All mice
subjected to these environmental risk factors will be studied using the methods described above.
In summary, by introducing the strongest known genetic risk factor for sporadic AD into an AD mouse
model and investigating interactions with environmental risk factors, we will create an animal model that
better reflects the multifactorial and highly prevalent, yet currently understudied, interplay between
inherited and acquired risk factors in the pathophysiology of late-onset AD. This model may therefore be
more predictive of possible translatability into clinical studies, and may potentially lead to the
development of new avenues of primary prevention or treatment.
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