2025 Theses Doctoral

Integrative systems analysis of the influence of tissue niches on human immune cells

Caron, Daniel Phillip

A preponderance of the body’s immune cells resides and functions within tissue, rather than in circulation. Murine models suggest the tissue microenvironment drives tissue-specific molecular profiles in immune cells; however, the restricted availability of healthy human tissue samples for research has limited our understanding of how various lineages and subsets of human immune cells are impacted by their tissue contexts. Here, we employ a systems approach to profile RNA, surface protein and immune receptor expression of >1.5 million immune cells isolated from human organ donor blood, lymphoid tissues, and mucosal barriers. Using these data, we define transcriptomic signatures of tissue-residency, identify tissue- and subset-specific trends with age, and probe hypotheses regarding how tissue-resident populations are maintained throughout life.

While transcriptomic data has proven invaluable for relatively unbiased profiling of heterogeneous cellular states, transcriptomic profiles cannot always delineate the immunological subsets defined by decades of surface proteome profiling. Multimodal sequencing technology, such as Cellular Indexing of Transcriptomes and Epitopes (CITE)-seq enables simultaneous profiling of single-cell transcriptomes and surface proteomes, offering potential to greatly improve cell type annotation accuracy. Here, we propose and evaluate algorithmic advancements in batch-correction (landmark registration) and joint-classification (Multi-Modal Classifier Hierarchy; MMoCHi) of these data, together facilitating accurate, granular annotation of immune subsets, such as discriminating CD4+ and CD8+ memory T cell subsets, various populations of cytotoxic lymphocytes, and rare subsets of dendritic cells.

We next explored molecular signatures associated with tissue-specific populations to understand the impacts of tissue on human immune cells. This high-quality annotation allowed us to distinguish between molecular programs underlying tissue-adaptation in immune cells and tissue-differences brought about by shifts in immune subset composition. We identify signatures of tissue-residency affecting expression of adhesion molecules, metabolism pathways, as well as soluble mediators and receptors of immune signaling. By relating these tissue-specific signatures across these diverse immune lineages (T cells, NK cells, ILCs, B cells, macrophages, etc.), or specific subsets (e.g., memory T cell subsets, niche-specific populations of macrophages) we identify broadly shared molecular programs suggestive of conserved regulatory mechanisms driving tissue-specific processes. As a consequence of this tissue-residency, we identified that age-associated effects were manifested in specific lineages and sites as revealed by macrophages in mucosal sites, B cells in lymphoid organs, and specific T and NK cell subsets across blood and tissues.

Finally, in addition to evaluating these molecular signatures, we also leveraged molecular profiling and immune receptor sequencing to understand maintenance of two major tissue-resident immune lineages, macrophages and T cells. Examining macrophage heterogeneity in tissues, we identify evidence for a dynamic equilibrium of monocyte-replenishment and self-renewal at homeostasis. T cell compartmentalization across tissues indicated that localized expansion and in situ differentiation of resident memory T cell subsets occur upon antigen encounter in barrier tissues.

Together, our results reveal tissue-specific signatures of immune homeostasis throughout the body, providing a molecular basis for immune variation from which to better understand immune pathologies across the human lifespan.