Using iPSCs to model the combined effects of genetics and aging on the human blood-brain barrier - Project Summary In addition to the ethical considerations of using animals for biomedical research, the high costs, low throughput, and limited translation of mice motivates their reduction and replacement with human tissue-on-chip (hTOC) for disease modeling, drug development, and discovery. One frontier in hTOC development is the modeling of aging as a confounding factor in disease. Our focus is on the blood-brain barrier (BBB), which is known to become less robust at regulating the passage of cells and molecules from blood as people age. Age- related changes in the BBB include cellular and transport changes that make the brain more vulnerable to injurious circulating factors and neurodegeneration. This includes impairment of lipoprotein receptor related protein-1(LRP-1), which leads to amyloid beta (Ab) accumulation in the brain, a hallmark of Alzheimer’s disease (AD). In addition, the presence of the Apoe4 allele, a risk factor for AD, accelerates most of the aging related changes in the BBB. Traditional animal models of AD and the BBB lack human relevance. Importantly, the mouse does not naturally develop human neurodegenerative diseases, including AD, and must be genetically engineered to approximate the condition. In animals that do naturally develop AD or cognitive decline such as non-human- primates or canines, the age-of-onset is too long for practical progress. For these reasons we will combine induced-pluripotent stem cell (iPSC) and microphysiolocial systems technologies with small molecule ‘aging’ cocktails to develop the first hTOC aged model of the blood brain barrier: the µSiM-aBBB (microphysiological system enabled by a silicon membrane-aged BBB). We will model geriatric vulnerabilities to genetic predisposition of the Apoe4 allele in the BBB by mirroring the cellular changes that occur during aging having shown preliminary induction of senescence, shifts in protein expression and increases in barrier permeability in our aged model. Thus, I hypothesize that consistent with the onset of AD occurring in older patients homozygous for Apoe4, aged cellular phenotypes combined with the Apoe4 mutation produce a BBB that is intrinsically compromised compared to Apoe4 or aging alone. To test this hypothesis, I will pursue two aims. Aim 1 will establish and characterize our model, the µSiM-aBBB using colorimetric assays, immunohistochemistry, permeability assays and ELISAs. Aim 2 will determine how the presence of the genetic risk factor, Apoe4, combined with aging affects barrier integrity and function in the BBB using RNA sequencing as well as the methods mentioned in aim 1. The proposed experiments will create a novel tool that can be used to determine ways to prevent barrier dysfunction under compromised conditions. This project will demonstrate the value of hTOC models to emulate human health and disease and expand our knowledge on the combined effect of genetics and aging on the human BBB.
