Transcriptomic Differences in Endothelial Cells Across Five Brain Regions in Normal Aging and Alzheimer's Disease

Background Altered cerebral vasculature is recognized as a key feature of Alzheimer’s disease (AD) neuropathology, with current evidence suggesting a diverse collection of changes including aberrant angiogenesis, vascular pruning, inflammation, senescence, and other remodeling events. Previous studies examining the endothelial cell transcriptome in AD have been limited to 1‐2 brain regions and hampered by low numbers of isolated endothelial cell nuclei, potentially missing relevant regional heterogeneity. We hypothesized that such regional heterogeneity in endothelial cell gene expression in the normal aging brain may contribute to the varied vascular responses to AD neuropathology. Method Nuclei were isolated from five brain areas in subjects with increasing AD pathology (n=32 donors). The five brain areas chosen represent the hierarchical accumulation of tau pathology in AD: entorhinal cortex > inferior temporal gyrus > prefrontal cortex > secondary visual cortex > primary visual cortex. All subjects had detailed p‐tau, in vitro tau seeding, and amyloid‐beta plaque measurements available. Nuclei were FACS sorted to exclude NeuN+ and Olig2+ cells, and the resulting neuron‐ and oligodendrocyte‐depleted fraction was subject to single‐nucleus RNAseq. Endothelial cell nuclei were identified based on high von Willebrand factor expression and filtered to exclude contaminating glia, yielding a dataset comprised of 51,586 total nuclei. Result Transcriptomic analysis confirmed that endothelial cells share a core set of commonly upregulated genes across the five brain regions compared to other cell types. However, there is also regional heterogeneity with 193‐311 differentially upregulated genes expressed in endothelial cells within each brain area relative to all other areas. Compared to normal aging brain, endothelial cells from AD donors downregulated 962 genes and upregulated 936 genes that are enriched in vasculogenesis, blood‐brain barrier maintenance, senescence, and leukocyte interactions. Further analysis indicates regional differences in upregulation of these pathways and highlights the relationship to disease burden. Conclusion This dataset demonstrates a previously unrecognized regional heterogeneity in endothelial cell transcriptome in the normal aging human brain. These regional differences may impact the endothelial cell response to the local AD pathology. This dataset will inform ongoing research efforts to unravel the molecular underpinnings of AD‐