Impact of neutrophil heterogeneity on pathogenesis of C. difficile infection

Cincinnati Va Medical Center Research, Cincinnati OH

Investigators

Linked publications, trials & patents

Abstract

Clostridium difficile is a leading nosocomial infection in the U.S., and a major concern for VA acute and long- term care facilities. Although host factors are a key predictor of disease outcomes after C. difficile infection (CDI), almost all current therapies are focused on controlling the pathogen. Our lab focuses on studying key facets of the host responses during CDI. The overall goal is to identify deleterious and beneficial host factors after CDI, so that we can develop novel therapies that target the host instead of pathogen alone. Neutrophil-mediated inflammation is an important determinant of CDI outcomes, and excessive neutrophilia is associated with severe disease. In the previous grant period, we explored role of neutrophils and neutrophil- derived molecules in regulating CDI pathogenesis. Through meticulous investigations, we unveiled an intricate interplay between neutrophil-mediated response and intestinal epithelial barrier integrity after CDI. Utilizing cutting-edge single cell transcriptomics approaches, we next established the first-ever transcriptomics atlas of bone marrow, blood and colonic neutrophils after CDI; our manuscript (MS ID# BIORXIV/2023/553751) is under consideration at Science Immunology. Specifically, we identified a pathogenic neutrophil subset in CDI: neutrophils that express a particular glycoprotein, Olfactomedin-4 (OLFM4), exacerbate the epithelial damage caused by C. difficile and its toxins. Both humans and mice with CDI had higher concentration of circulating OLFM4 than uninfected controls; and in mice OLFM4 deficiency resulted in expedited recovery and better survival after CDI without any effects on the pathogen. Further, we show that mice with a common single nucleotide polymorphism (SNP) in the gene for leptin receptor (Q223R; change of Glutamine [Q] to Arginine [R] in LEPR) that is associated with more severe CDI, also have more OLFM4+ neutrophils and higher serum OLFM4, compared to controls. In this renewal grant, we now want to explore the mechanisms that underlie the pathogenic nature of OLFM4+ neutrophils. Our central hypothesis is that: LEPR Q to R substitution amplifies neutrophil OLFM4 expression and its release leading to exaggerated CDI-induced IEC damage. We will address the following questions: 1) What is the effect of a LEPR SNP on CDI-induced pathways of Olfm4 transcription and protein release? 2) What are the mechanisms by which OLFM4+ neutrophils exacerbate CDI-induced epithelial damage? 3) Can the proportion of OLFM4+ neutrophils in circulation and/or serum OLFM4 concentration be used to predict CDI severity? We expect that our studies will identify the effects of LEPR Q223R SNP on OLFM4 biology and neutrophil heterogeneity, provide a greater understanding of the drivers of excessive IEC damage in CDI, and determine the utility of OLFM4+ neutrophils and/or serum OLFM4 levels in predicting CDI severity. Data generated here can form the basis of future host-targeted therapies that can deplete OLFM4+ neutrophils or block OLFM4 as a way to ameliorate CDI severity.