PROJECT SUMMARY Traumatic brain injury (TBI) is one of the leading causes of death and disability and represents the most well- established environmental risk factor for cognitive impairment, progression to clinical Alzheimer’s disease (AD) and AD related dementia (ADRD). Following injury, brain blood vessels undergo many degenerative changes ranging from, loss of pericytes (PC), remodeling of the vascular basement membrane (vBM), loss of blood-brain barrier (BBB) integrity and impaired angiogenesis, which ultimately cause neuronal damage, cognitive decline and ADRD/AD. BBB disruption in the human hippocampus predicts cognitive impairment in patients. In addition, the acute inflammatory phase following TBI correlates with post-TBI ADRD/AD trajectories, supporting the idea that the initial innate immune response, may influence the later adaptive immune response, resulting in unresolved inflammatory responses, and the risk of post-TBI cognitive impairments and ADRD/AD - progressive disorders that worsens over time. Yet, the molecular mechanisms that predispose TBI survivors to an increased risk of ADRD/AD are unknown and consequently there are no treatments. Our parent grant (R33HL159949), which focuses on understanding the effects of the blood components on NVU function following TBI, compared to control, is importantly related to the development of ADRD/AD. A major impediment to the development of therapeutic approaches to neurological disorders has been the lack of human in vitro NVU models that mirror the spatial organization and molecular characteristics of NVU cells in the human brain. To address this challenge, during the R61 phase, we established a 3D human blood-BBB interface microfluidic model, which allows for the integration of defined iPSC derived cell types forming the functional NVU, and sera from patient or controls. The focus of the R33 phase is to examine the effect of blood components (sera and immune cells) from donors with/without TBI on BBB/NVU function. This timely Supplement proposal requires research beyond our funded grant to test this hypothesis through two new aims in line with the original aims from our R33. The overall goal of this Supplement is to better understand how changes within the acute and chronic peripheral immune response following TBI, may influence BBB/NVU function over time, and predict progression of post-TBI associated cognitive impairments and ADRD/AD. In this proposal, we outline a plan to examine the effect of sera from Veterans (LIMBIC-CENC cohort) with chronic TBI whom develop post-TBI associated ADRD/AD, or do not, in the BBB/NVU. Specifically, we will examine the influence on BBB cell biology (structural and functional BBB properties) and NVU function (cell-cell interactions and -communication and functionality). We will use state-of- the-art single cell kinetic imaging (KIC), transcriptomics, and functional approaches to address these questions. The proposed work may facilitate discovery of pote...