Heidelberg University (Germany)/Harvard Medical School Cambridge, MA, United States
Nicolaj hackert1, Felix Radtke1, Tarik Exner1, Carsten Müller-Tidow1, Hanns-Martin Lorenz1, Peter Nigrovic2, Guido Wabnitz1 and Ricardo Grieshaber-Bouyer1, 1Heidelberg University Hospital, Heidelberg, Germany, 2Boston Children's Hospital, Boston, MA
Background/Purpose: Neutrophils mediate a range of homeostatic and inflammatory processes and display substantial phenotypic and functional heterogeneity. While animal models enable important mechanistic discoveries, differences between organisms can pose roadblocks for translation of findings. Research increasingly benefits from seamless transitions between the murine and human system, necessitating knowledge about conserved biology and methods for integrative analyses of cellular systems across species.
Methods: We performed an integrative analysis of gene expression in resting and inflamed leukocytes from humans and mice. Raw data from 247 samples from 21 different studies was obtained from GEO and integrated by focusing on protein coding, one-to-one orthologous genes with high confidence according to ENSEMBL. Transcription factor enrichment was assessed using ChEA3. We used DESeq2 to identify differentially expressed genes in each study individually compared to healthy control cells and then used a Fisher's combined test to compute the significant overlap. Validation studies were performed using flow cytometry on stimulated human and murine neutrophils.
Results: Gene expression data from human and murine immune cells could be analyzed together, and lineage-specific gene expression was well conserved across species. In resting blood neutrophils, we observed high concordance in overall gene expression, transcription factor expression and lineage-specific gene expression across humans and mice. In inflammation, neutrophils displayed varied transcriptional responses to different contexts. Within these, we identified a core inflammation program conserved across a broad range of conditions and between humans and mice. This core inflammation program includes genes encoding IL-1 family members, CD14, IL-4R, CD69 and PD-L1. Neutrotime scores trended higher in inflammation, suggesting more advanced maturation in activated neutrophils. Transcription factor enrichment analysis nominated NF-κB family members and AP-1 complex members as key drivers, consistent with strong enrichment for gene sets associated with NFκB activation and cytokine response in inflamed cells. In vitro perturbations confirmed the predicted surface protein changes characteristic for inflamed neutrophils in both species. Activated neutrophils from all tissues displayed the core inflammation program. Yet, differences between blood, bone marrow and spleen highlight that maturation stage, tissue and other factors act in concert to drive variable neutrophil phenotypes.
Conclusion: Here, we demonstrate that integrative analyses of gene expression in human and murine immune cells can be performed by mapping protein coding, one-to-one orthologous genes. Resting blood neutrophils demonstrate substantial transcriptional conservation across species. In inflammation, we predict and experimentally validate a core inflammation program in neutrophils conserved across a range of conditions and across species. This approach of focusing on targets with conserved biology can be transferred to other cell types and enable more accurate translational studies in the immune system.
Disclosures: N. hackert, None; F. Radtke, None; T. Exner, None; C. Müller-Tidow, None; H. Lorenz, None; P. Nigrovic, None; G. Wabnitz, None; R. Grieshaber-Bouyer, None.