Advisor: Ahmet F. Coskun, Ph.D. (BME, Georgia Institute of Technology)                 

Committee:

Peng Qiu, Ph.D. (BME, Georgia Institute of Technology)

Saurabh Sinha (BME, Georgia Institute of Technology)

Rabindra Tirouvanziam, Ph.D. (BME, Emory University)

Marcus Cicerone, Ph.D. (Chem, Georgia Institute of Technology)  

Timeseries Spatial Omics Of Immune Cell Signaling In Inflammation  

Cystic fibrosis (CF) affects 100,000 people worldwide. Although FDA-approved therapies have remarkably succeeded, chronic inflammation and bacterial infections persist, causing tissue damage and worsening quality of life, despite a large immune cell presence. We investigate the spatiotemporal dynamics of these immune cells, specifically the Nuclear Factor kappa B (NFκB) pathway, during inflammation. We combine emerging spatial omics techniques with a time-series fixation method involving static stimulation and programmable formaldehyde perfusion to map pseudo-Signaling with Omics signatures (pSigOmics) of single-cell data from 100K+ cells. Examining NFκB in mouse fibroblasts, we discovered a novel asynchronous pseudotime regulation (APR) between p65 RNA and protein in the quintessential NFκB p65 protein using single-molecule spatial imaging. The observed p65 translational APR is evident in both statically sampled timepoints and dynamic response gradients from perfused fixation. This translational regulation is vital in CF, where resident macrophages and transmigrating neutrophils persist in high-inflammation lung areas despite successful treatment. The spatiotemporal activity of immune cells – fibroblasts, macrophages, neutrophils – poses significant clinical challenges because they fail to clear infection and engender tissue damage with prolonged activation. Examining the immune landscape spatiotemporal patterns is critical to identifying opportune windows for treatment and investigating why bactericidal capabilities are malfunctioning.