Wednesday, January 15, 2025
1/15/2025
PROJECT SUMMARY Senescent cells (SnCs) accumulate in tissues with increased organismal age. SnCs have been associated with multiple age-associated chronic diseases and implicated as contributing pathological factors in Alzheimer’s disease (AD). Brain vasculature aging contributes to AD progression and vascular dysfunction is observed early on in the disease. Recent studies suggest that endothelial senescence impairs the barrier function of the brain endothelium. However, SnCs within the brain vasculature have not been studied in the context of AD. I examined multiple human and mouse single-cell RNA-seq datasets from organisms of different ages to create a resource that identifies SnCs in different cell types and tissues. My preliminary data indicate that the brain vasculature has an increased SnC burden in AD and that the blood-brain barrier expresses receptors for SnC signaling. However, the phenotype and molecular signatures of AD-associated SnCs have not been resolved. I hypothesize that senescent brain vascular cells have distinct and targetable molecular patterns and that their disruptive effect on healthy vascular cells can be lessened by interrupting specific SnC signaling pathways. Moreover, I have shown that transposable elements and human endogenous viral elements are upregulated in endothelial inflammation, cellular senescence, and AD. Transposable elements differentially regulated in endothelial inflammation were co-expressed with important inflammatory and senescence regulators, such as NFKB and CDKN2A. I hypothesize that transposable elements expressed during inflammation and senescence contribute to sterile inflammation and loss of blood-brain barrier integrity. I will transcriptionally characterize SnCs within the brain vasculature of human AD patients and mouse models by examining existing single-cell AD datasets. I will find unique patterns of mRNA expression, regulatory mechanisms, and cell-cell interactions specific to AD- associated SnCs. I will train a semi-supervised convolutional neural network to identify senescent cells within the AD brain vasculature. I will test how SnC signaling contributes to AD by ablating receptors required in SnC signaling in brain microvascular endothelial cells with CRISPR-Cas9 then test the effect on barrier integrity in the presence of SASP molecules. Next, I will find transposable elements uniquely expressed in senescent brain endothelial cells and bystander cells during AD. I will uncover regulatory mechanisms within the cells that contribute to the aberrant activation of transposable elements and examine the pathways these elements activate that contribute to barrier dysregulation. I will test how shRNA knockdown of expressed transposable element RNA affects the inflammatory and barrier phenotypes in brain endothelial cells. The proposed studies will teach me multiple computational approaches, such as machine learning, as well as advanced genetic manipulation in cells and animal models.
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