Date of Award
2026
Document Type
Thesis
Degree Name
Doctor of Philosophy (PhD)
Thesis Advisor
Junyue Cao
Abstract
Mammalian aging is a complex and multifaceted biological process, and its mechanism remains poorly understood. Several decades of research suggest several “hallmarks” (López-Otín et al. 2023) that are universally observed in the aging process, and “pillars” (Kennedy et al. 2014) that are thought to play causative roles. However, much of our understanding of aging is based on bulk studies of tissue or on studies of nonmammalian model organisms that lack critical mammalian cell types. Many aging-associated diseases are driven by rare cell populations: the key cell type lost Parkinson’s disease–dopaminergic neurons in the substantia nigra–represents only 0.00064% of the neurons in the brain. (Pakkenberg et al. 1991; Azevedo et al. 2009) Modifying the aging process to prevent these diseases and extend human lifespan and healthspan will require systematic ways to screen the effects of aging and longevity interventions on many mammalian cell populations. We can thereby identify critical subpopulations that drive the aging process or modulate the effects of longevity interventions. Highly-scalable single-cell genomics methods such as EasySci-RNA make it possible to simultaneously profile the effect of any biological perturbation on many cell populations in the body. Our lab has applied EasySci-RNA to study aging in the mouse brain (Sziraki et al. 2023) and in 14 other tissues/organs, (Zhang et al. 2025) revealing key cell population proportion changes that occur during the aging process, including early loss of stem and progenitor cells and late expansion of inflammatory cells. In the first part of this study, I expand on this previous work to determine how a key longevity intervention, caloric restriction, modulates the aging process in the mouse brain. I employ EasySci-RNA to profile >500,000 brain nuclei from mice at three different ages in the presence or absence of caloric restriction. To determine the spatial effects of caloric restriction, I also employ the optics-free spatial transcriptomics method IRISeq (Abdulraouf et al. 2024) to profile brain sections from aged ad libitum and caloric restriction mice. I thereby show that caloric restriction slows several aging-associated changes in cell populations and gene expression, providing key evidence that metabolism and dietary regulation act upstream of key aging hallmarks such as inflammation and senescence. This work also establishes a template for future studies of additional longevity interventions. In the second part of this study, I develop JointSci-RNA, a modification of EasySci-RNA to capture single-cell genotype information alongside the transcriptome. I initially validate JointSci-RNA in a mixture of three cell culture lines with a variety of known homozygous and heterozygous variants. I distinguish cell lines by clustering, and show that the correct genotypes are assigned to each cell line. I then apply JointSci-RNA to profile a human glioma tumor with four putative variants of interest, successfully employing genotyping data to identify the cluster of malignant cells and the distribution and relevance of each variant. This work provides a proof of concept that JointSci-RNA can be employed in human tissue, and paves the way for future studies of the role of clonal expansions in aging.
License and Reuse Information
This work is licensed under a Creative Commons Attribution-NonCommercial-Share Alike 4.0 International License.
Recommended Citation
Epstein, Alexander, "Pushing the Limits of Single-Cell Genomics to Study Aging and Cancer" (2026). Student Theses and Dissertations. 857.
https://digitalcommons.rockefeller.edu/student_theses_and_dissertations/857
Comments
A Thesis Presented to the Faculty of The Rockefeller University in Partial Fulfillment of the Requirements for the degree of Doctor of Philosophy