Moiz, BilalBrain microvascular endothelial cells (BMECs) form the blood-brain barrier, which protects the brain from neurotoxic elements and simultaneously transports glucose and other vital nutrients into the brain. Neurovascular dysfunction is implicated in pathogenesis of neurodegenerative diseases such as Alzheimer’s disease and Parkinson’s disease; however, little is known about how neurovascular changes contribute to rare inherited neurogenetic disorders such as Niemann-Pick Disease (NP-C). NP-C is caused by mutations in the intracellular cholesterol trafficking proteins NPC1 and NPC2, which leads to endolysosomal cholesterol accumulation and membrane cholesterol depletion. Clinical manifestations vary by age and genetic factors but include neurological symptoms such as developmental delay, cognitive impairment, ataxia, and seizures. Current clinical management strategies are challenged by diagnostic difficulties and poor therapeutic efficacy. Hydroxypropyl-beta-cyclodextrin (HPβCD), an agent believed to release accumulated cholesterol, has shown promising clinical results; however, its efficacy is limited due to poor brain penetration. The major objective of this thesis was to determine how NPC1-deficiency impacts BMEC barrier function, metabolism, and nanoparticle uptake. I found that NPC1 deficiency diminishes barrier integrity in BMECs by disrupting claudin-5 and occludin morphology. Using isotope labeling, mass spectrometry, and computational flux analysis, I also observed that NPC1 inhibition leads to systemic metabolic changes, including increased glycolytic flux, elevated activity in peripheral glycolytic pathways, and reduced mitochondrial respiration. HPβCD treatment attenuated barrier function changes and partially restored BMEC metabolic phenotype. Finally, I found that isoproprylacrylamide (NIPAA-m) nanogels loaded with HPβCD were transported across NPC1-deficient BMECs, suggesting their potential for HPβCD delivery to the brain. This thesis demonstrates a unique, integrated computational-translational approach that unveils the role of BMEC in NP-C pathology, possibly leading to improved therapeutic strategies. In addition, this thesis improves our understanding of how variants in cholesterol metabolism and trafficking, as well as in proteins such as NPC1, which has been implicated in Alzheimer’s, diabetes, obesity, and atherosclerosis, contribute to brain endothelial dysfunction.enDissertationBioengineering