Decoding Spatially Resolved Single Cell Metabolic Trajectory of Tonsil Tissues and Organoids

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Germinal centers (GCs) are the microstructural sites in secondary lymphoid organs that control B cell clonal expansion to produce high-affinity antibodies for achieving humoral immunity. GC structures are impaired in obstructive sleep apnea (OSA) patients, leading the deficiencies in immune responses in infected individuals. The role of B cell development through GC reactions has been well established. B-cell immunometabolism is crucial to meet the energy needs of rapid proliferation. OSA patients exhibited associations with metabolic syndrome and vulnerability to flu. However, the coordination of metabolic trajectories in B cells of OSA+ and OSA- patients is still not clearly understood. There is a critical need to decipher the B cell immunometabolism at the single cell level in GCs for identifying the metabolic defects in the immune system of OSA patients making them prone to life-threatening infections. Thus, this project will leverage the recently developed spatially resolved metabolic profiling framework (3D-SMF) to map B cell subsets and their metabolism in the tonsil tissues and organoids. Our long-term goal is to generate single cell metabolic insights of B cell development in GCs of OSA+ and OSA- patients in response to influenza. The goal of this project is to define spatially resolved B cell immunometabolism pixel-by-pixel in fixed human tissues and living tissues. We hypothesize that metabolic trajectories and spatial distributions of B-cell subsets will be defective in OSA+ compared to OSA- tissues and influenza response in OSA- tonsil-derived organoids will be more competent than OSA+ tonsil-derived organoids. The rationale for this hypothesis is based on the 3D-SMF data showing the depletion and enrichment of fatty acids in GCs located in native tonsil tissues and the recent evidence on OSA patients’ vulnerability to flu infections associated with metabolic syndrome. The central hypothesis will be tested by pursuing two specific Aims. Aim 1 will provide an integral understanding of the lipid-associated immunometabolism in B-cell subsets in human OSA+ and OSA- tonsil tissues (n=10 each) and engineered tonsil organoids. Aim 2 will define how the metabolic trajectory modeling of B cell subtypes differs in tonsil organoids exposed to influenza antigens in OSA+ and OSA- donors. To accomplish these Aims, 3D-SMF and multiplexed cytokine gene expression profiling will be used to analyze B cell immunometabolism through a pseudotime B cell development modeling and longitudinal metabolic trajectory comparisons of B cell subsets in biomaterial-based tonsil organoids. This project builds an interdisciplinary team integrating experts from spatial omics, biomaterials, pediatric OSA, and bioinformatics. The proposed application is innovative because it uses cutting-edge technology to define spatial metabolomics and proteomics of tonsil organoids and shifts from the traditional focus on T cell and B cell co-cultures, toward differences of B cell metabolism in physiologically relevant and dynamic OSA+ and OSA- tonsil organoids ex vivo. This research is significant because it defines B cell immunometabolism to understand why OSA patients are prone to flu infections.