First Description of Different Neutrophil Populations in the Mucosa of the Female Genital Tract Identified by Spatial Transcriptomics

Abstract

Women worldwide acquire the human immunodeficiency virus (HIV) mainly through sexual contact. The female genital tract (FGT) has a unique mucosal surface that combines reproductive function with host defense against incoming pathogens, with a complex array of local immune defenses coordinated to protect against viral infections, including HIV. The first line of defense is formed by the epithelial cell barrier, mucus and secreted immune mediators. In addition, neutrophil recruitment to the genital mucosa is essential to maintain homeostasis, reproductive function and, as first responders against an insult, fight pathogens. We hypothesize that different populations of neutrophils exist within the FGT mucosa to play distinct roles in homeostasis and defense. To gain insight into neutrophil distribution and function within FGT tissues, we performed immunofluorescent staining of human endocervical and ectocervical tissue sections using neutrophil elastase antibody. Neutrophils were found concentrated towards the mucosal surface, positioned close to or within the epithelium, and also scattered throughout the tissue. Using spatial transcriptomics, neutrophils were selected in regions of interest (ROIs) with a segmentation mask and RNA extracted for sequencing. Whole transcriptome analysis segregated the transcriptional profiles of subepithelial and tissue‐scattered neutrophils and detected differential gene expression based on tissue location. Interestingly, epithelium‐associated neutrophil transcripts were enriched for pathways characteristic of defense (i.e., “Antimicrobial peptides”, “Neutrophil degranulation”, “Chemokine receptors bind chemokines”, among others). In addition, epithelium‐associated neutrophils showed an upregulation of CD16 and CD62L measured by flow cytometry, which indicates an activated state and could be associated to microbiota‐neutrophil interactions and protection of the very first layer of mucosal tissue.For the first time, we describe different populations of neutrophils within the mucosal FGT and set foundation for mucosal immunity research, with potential for development of therapeutic interventions against sexually transmitted infections, including HIV.