Elucidating mechanisms of blood-brain barrier dysfunction in Alzheimer's disease using a molecularly and cellularly tunable in vitro model

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PROJECT SUMMARY Dysfunction of the blood-brain barrier (BBB) begins during early stages of Alzheimer’s Disease (AD) and contributes to AD pathogenesis in a manner both independent of and dependent on the accumulation of amyloid beta. The ε4 allele of the APOE gene is the greatest genetic risk factor for late-onset AD, but the molecular mechanisms underlying the connection between the APOE-ε4 allele and BBB dysfunction and are not well understood. In addition, the extent to which non-genetic risk factors for AD act alone and synergistically with the APOE-ε4 allele to disrupt BBB integrity and function is not known. The objective of this work is to gain mechanistic insight into how the APOE-ε4 allele and modifiable risk factors for AD influence BBB functions that are known to be disrupted in AD patients, with the goal of utilizing this information to identify novel therapeutic targets and/or interventive measures to prevent cognitive decline. Toward this end, human induced pluripotent stem cells (hiPSCs) will be used to generate isogenic models of the BBB with tunable molecular and cellular components. These models will be used to test our central hypothesis that expression of ApoE4, encoded by APOE-ε4, inherently alters the dynamics of key BBB functions and that molecular features of modifiable risk factors for AD compound these effects. The proposed research consists of two aims. (1) Determine the dominant molecular mechanisms by which ApoE4 effectuates BBB dysfunction for the prioritization of drug targets in early stage AD. Brain microvascular endothelial cells, pericytes, and astrocytes will be differentiated from hiPSCs containing either zero, one or two copies of the APOE-ε4 allele for quantitative analysis of how ApoE4 impacts dysregulated BBB transport, neuroimmune, and cell maintenance functions associated with AD pathogenesis. (2) Determine how molecular features common among modifiable risk factors for AD act in combination to drive BBB dysfunction and whether BBB susceptibility to these factors is ApoE-isoform dependent. In vitro BBB models of differing APOE status will be subjected to conditions that mimic modifiable risk factors for AD at the cellular level, including elevated concentrations of glucose, and inflammatory cytokines. This will be done using a fractional factorial design approach to systematically and efficiently evaluate combinations of genetic and non- genetic factors. Analysis of downstream signaling nodes will elucidate mechanistic connections between input cues and output responses, identifying druggable targets for therapeutic intervention. This approach will establish a detailed understanding of how the APOE-ε4 allele and modifiable risk factors act in concert to convey risk for AD through BBB dysfunction, providing preventative strategies and novel routes for therapeutic intervention in the early stages of AD, particularly in individuals with the APOE-ε4 allele.