Neutrophil heterogeneity and plasticity in wound healing

PROJECT SUMMARY/ABSTRACT The long-term goal of my research program is to understand neutrophil heterogeneity and functions in diverse tissues and biomedical contexts. We have combined high-dimensional and single-cell profiling with integrated bioinformatics approaches to defining context-specific neutrophil landscape. Neutrophils, the most abundant human immune cell type, play crucial, first-line roles in regulating swift responses against infections and pathological responses. Emerging evidence has shown that neutrophil response is systematic and context- specific with distinct phenotypes and functions. Burn injuries are among the most common traumatic injuries worldwide. In severe cases, patients can die of infection, shock, and/or organ failure, requiring expeditious clinical responses. Proper wound healing requires a coordinated time-dependent balance of pro- and anti- inflammatory immune pathways to prevent irreversible systemic damage. Despite being the first to mobilize to sites of injury, there remains little known about neutrophil heterogeneity, their fates, and which factors control the plasticity of neutrophils in wound healing. We hypothesize that neutrophils' cellular and molecular landscape change over time during the wound healing process, and that blood neutrophils could be a marker to predict complications and outcomes in severe burn injury. To this end, we will focus on two important discoveries in neutrophil biology over the past decade: (i) neutrophil heterogeneity and adaption to specific tissue environments revealed by high-dimensional flow cytometry and single-cell transcriptome data and (ii) neutrophil reverse migration phenotypes, shown in model organisms demonstrating that neutrophils migrate back to the vasculature in response to inflammation. In this MIRA application, I plan to 1) identify transcriptional and spatial landscapes of neutrophils and their interactions with other immune cells in human burns using multimodal single-cell profiling approaches, 2) define reverse migration neutrophil phenotypes in human burns by performing cross-species single-cell transcriptome analyses of neutrophil heterogeneity in wound healing using clinical samples and pre- clinical zebrafish burn models, and 3) develop analysis pipelines and methods to predict neutrophil behaviors defined by imaging using single-cell genomic data in humans and publicly available data from zebrafish and mouse models. In the next five years, I will establish a research program to study neutrophil heterogeneity and plasticity in wound healing using multidisciplinary approaches. Our approaches are innovative because we will employ: (i) state-of-the-art single-cell technologies, (ii) novel integrated bioinformatics method development and analysis, and (iii) profiling data cross-tissues in humans with in vivo validation models using model organisms. These studies will generate comprehensive data and provide cellular and molecular landscapes of neutrophil heterogeneity to the community studying wound repair and neutrophil biology.