Project description:The full neutrophil heterogeneity and differentiation landscape remains incompletely characterized. Here we profiled more than 25,000 mouse differentiating and mature neutrophils using single-cell RNA sequencing to provide a comprehensive transcriptional landscape of neutrophil maturation, function, and fate decision in their steady state and during bacterial infection. Eight neutrophil populations (including the GMP population) were defined by distinct molecular signatures, including a new circulating mature neutrophil population highly expressing interferon-stimulated genes. The three mature peripheral blood neutrophil subsets arise from distinct maturing bone marrow neutrophil subsets, a novel mechanism that highlights the complex and precise regulation of neutrophil production. Neutrophil heterogeneity and differentiation are driven by both known and uncharacterized transcription factors. Neutrophils gradually acquire microbicidal capability as cells traverse the transcriptional landscape, representing an evolved mechanism for fine-tuned regulation of an effective but balanced neutrophil response. Bacterial infection reprograms the genetic architecture of neutrophil population, alters dynamic transition between each subpopulation, and primes neutrophils for augmented functionality without affecting overall heterogeneity. Bacterial infection-induced emergency granulopoiesis is mediated by augmented proliferation of early-stage neutrophil progenitors and accelerated post-mitotic maturation. In summary, single-cell transcriptomics enabled the reconstruction of neutrophil differentiation and maturation trajectories and uncovered neutrophil subpopulations, gene pathways, and regulators of neutrophil function and fate decisions. These data establish a reference model and general framework for studying neutrophil-related disease mechanisms, biomarkers, and therapeutic targets at single-cell resolution.
Project description:Purpose: The goals of this study are to compare bulk RNAseq profiles of tissue neutrophils. Methods: Bulk RNAseq of sorted neutrophils from bone marrow, spleen, blood, lung, peripheral blood, skin and intestine from wild-type (WT) mice, in triplicate, using Illumina. The sequence reads that passed quality filters were analyzed at the gene level with RSEM which mapped about 26 million sequence reads per sample to the mouse genome (build mm10/GRCm38) and identified 32328 genes.
Project description:Examination of open chromatin regions between clonal neutrophil progenitor populations. Conditionally immortalized CD45.1 naive paired granulocyte-monocyte progenitors (GMPs) and their mature neutrophil counterpart were profiled in an in vitro cell system. We identified 29,966 differentially accessible regions (DARs) between GMPs and neutrophils, from a total of 74,009 consensus peaks of chromatin accessibility. There is some correlation of open chromatin between GMP and mature neutrophil clones.
Project description:Purpose: The goals of this study are to compare bulk RNAseq profiles of tissue neutrophils in germ free mice. Methods: Bulk RNAseq of sorted neutrophils from spleen, blood, lung from spf and germ free mice, using Illumina. The sequence reads that passed quality filters were analyzed at the gene level with RSEM.
Project description:Sepsis is a life-threatening condition characterized by uncontrolled systemic inflammation and coagulation, leading to multi-organ failure. Therapeutic options to prevent sepsis-associated immunopathology remain scarce. Here, we established a mouse model of long-lasting disease tolerance during severe sepsis, manifested by diminished immunothrombosis and organ damage in spite of a high pathogen burden. We found that, both neutrophils and B cells emerged as key regulators of tissue integrity. Enduring changes in the transcriptional profile of neutrophils, included upregulated Cxcr4 expression in protected, tolerant hosts. Neutrophil Cxcr4 upregulation required the presence of B cells, suggesting that B cells promoted disease tolerance by improving tissue damage control via the suppression of neutrophils’ tissue damaging properties. Finally, therapeutic administration of a Cxcr4 agonist successfully promoted tissue damage control and prevented liver damage during sepsis. Our findings highlight the importance of a critical B-cell/neutrophil interaction during sepsis and establish neutrophil Cxcr4 activation as a potential means to promote disease tolerance during sepsis.
