Date of Award
Spring 2024
Document Type
Dissertation
Degree Name
Doctor of Philosophy (PhD)
Department
Biomedical Engineering (ENAS)
First Advisor
Fan, Rong
Abstract
With the introduction of next-generation high-throughput DNA sequencing, “Omics” has become staple in biological and biomedical research. Currently, single-cell omics has enabled a wide range of biomolecular assays at single cell level including proteomics, transcriptomics, epigenomics, metabolomics, and more. These techniques have the power to interrogate the different dimensional states of single cells and in parallel with one another. Extensive and rich datasets have been produced with these methods ranging from normal to developing to diseased cells, especially informative in delineating heterogeneous populations. However, the trade-off for high-throughput single-cell Omics is the loss of cellular phenotypic and positional information. When discussing heterogeneous cell populations, or even relatively uniform ones, not only are their specific location in the tissue crucial for deeply understanding their nature, but also their neighboring cells. This need led to the recent development and rise in spatially-resolved Omics which aimed to retain the identities lost in next-generation sequencing and to study cells and tissues in their native context as well as how they interact between one another. This type of information is especially critical in studying the human brain which is composed of diverse cell types that are organized in highly complex and intricate networks with each anatomical region responsible for specific functions. My PhD dissertation project aims to adapt and (1) apply multiplexed fluorescence image-based spatial proteomics for high resolution characterization of cell types and interactions within the human hippocampus, (2) apply the spatial platform Deterministic Barcoding in Tissue for spatial Omics sequencing (DBiT-seq) to target brain tissue and specifically map the human hippocampal region in a multilayered facet with transcriptomics and epigenomics, and (3) expand these tools for spatial mapping of major depressive disorder (MDD) patient hippocampi. First, to characterize the spatial organization of cell types and cell states in the human hippocampus, we incorporated proteomics in the form of the platform called Co-detection by indexing, or CODEX. Several key features from this platform that made it a highly informative modality were the single-cell resolution, the capability for a multiplexed panel of protein markers to be targeted, and the ability to capture a large region up to the entire hippocampus cross-section. This work allowed us to precisely describe the major cell-types identified in each sublayer of the hippocampus in 9 neurotypical patient samples. Second, we utilized the microfluidics spatial sequencing platform DBiT-seq on brain tissue and mapped the transcriptomic and chromatin accessibility landscape of the human hippocampus dentate gyrus region. By inserting a set of dual barcodes introduced by separate orthogonal parallel channels, a matrix of barcodes referencing the location can be appended to the transcriptome or genome and retrieved after sequencing. Parameters affecting crucial steps such as reverse transcription and in situ barcode ligation were optimized to improve the efficiency of the platform for brain-related tissues. Here, we describe the pipeline for targeting spatial mRNA-sequencing and spatial chromatin accessibility in brain sections and the subsequent work in developing a mapped omics atlas based on 6 neurotypical postmortem human hippocampi. Third, by leveraging the multi-Omics atlas of the human hippocampus we further examined the molecular landscape within a clinical cohort of major depressive disorder (MDD) patient brains. Additional spatial proteomics of 8 MDD hippocampi, transcriptomics of 6, and epigenomics of 5 were carried out to describe cellular and function changes that may play pivotal roles in the pathophysiology of the disease. In summary, my PhD project led to the implementation of the spatial Omics platform DBiT-seq on brain-related tissue and the development of a spatial atlas for the human hippocampus in both healthy and MDD patient brains. These studies provide a large resource for not only furthering our understanding of the human hippocampus and dentate gyrus at a molecular level, but also the potential for new avenues in studying many different biological questions such as the mechanisms behind MDD.
Recommended Citation
Su, Graham, "Spatial Multi-Omics Mapping of the Human Brain Hippocampus" (2024). Yale Graduate School of Arts and Sciences Dissertations. 1299.
https://elischolar.library.yale.edu/gsas_dissertations/1299
