A platform to investigate the neuronal determinants of APOE genotype on neuron function

Abstract

Alzheimer’s disease (AD), the most prevalent form of dementia worldwide, affects a growing proportion of the elderly and poses a significant societal burden. Pathological hallmarks include the appearance of misfolded proteins in the brain. This is evidenced by the deposition of amyloid-β (Aβ) plaques and tau-containing neurofibrillary tangles leading to neuron loss.
Advanced age is the single most important factor that leads to the dementia associated with AD. Additionally, genetic factors underlie the severity and/or age of onset. APOE 𝜀4 is the strongest genetic risk factor for sporadic AD. Subtle changes in the gene lead to three common variants: APOE e4, APOE e3, and APOE e2. In terms of AD, e4 is associated with increased while e2 is associated with decreased incidence compared with 𝜀3. The protein from this gene, apolipoprotein E (ApoE), is involved in numerous functions in the brain including cholesterol homeostasis and clearance of Aβ. Functional complexity is increased as ApoE is primarily synthesized and secreted from astrocytes while neuronal expression has also been identified in the context of injury, damage, or stress. ApoE is a secreted molecule, which raises questions regarding which cell source and what fundamental functions of ApoE modulate AD susceptibility. Is there a neuron-intrinsic disease trigger and/or disproportionately high neuron-autonomous contribution to AD?
I compared isogenic cell lines in which the genome is engineered to carry homozygosity for each of the three APOE alleles listed above. I interrogated the impact of genotype on neuronal differentiation, morphology, and potential variation in vulnerability to stress. This latter aspect was modeled by exposing mature cultures to kainate-induced excitotoxic insult. These approaches revealed that APOE had little impact on neuron differentiation, except for subtly affecting connectivity-related gene expression in the e4/e4 versus neurons of the other two genotypes. Upon kainic acid exposure, e4/e4 neurons again showed a small difference; compared to e3/e3 and e2/e2 neurons, there was a delay in mounting an ATF4 response, a stress-protective
mechanism. No other differential response to excitotoxicity emerged. Although these neurons did show a detectable low level of APOE transcript, there was no upregulation of APOE upon kainate exposure. My established neuronal platform of APOE variants is now available for further studies to more deeply probe genotype-specific mechanistic differences based on my initial observations.
These subtle but clear changes in e4-related altered neurodevelopment and delayed stress response suggest that the e4 allele may have limited effect early in life. However, over multiple stress events that may occur over a lifetime, these changes could provoke neurodegeneration. Another implication is that because e2/e2 neurons did not simply show the opposite phenotype of 𝜀4 neurons, the neuroprotective mechanism of e2 does not overlap with the pathways responsible for 𝜀4’s detrimental effects.

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Identifiers

ORCID for Mercedes Erika Alexandra Beyna: orcid.org/0000-0003-0603-3981

ORCID for Katrin Deinhardt: orcid.org/0000-0002-6473-5298

ORCID for Vincent O'connor: orcid.org/0000-0003-3185-5709

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