Identification of Differentially Expressed Genes and Signaling Pathways in Neutrophils During Sepsis-Induced Immunosuppression via Bioinformatics Analysis
Sepsis is a life-threatening condition characterized by organ dysfunction due to a dysregulated host response to infection. It profoundly disrupts immune homeostasis, impairing both innate and adaptive immunity by altering the lifespan, production, and function of effector cells responsible for maintaining homeostasis. One of the critical phases of sepsis is sepsis-induced immunosuppression, which is associated with altered neutrophil chemotaxis, oxidative burst, lactoferrin content, and an increased number of circulating immature granulocytes. These changes are linked to a higher risk of mortality following septic shock. Despite these findings, the key genes and signaling pathways involved in neutrophils during sepsis-induced immunosuppression remain poorly understood. This study employs bioinformatics analysis to identify differentially expressed genes (DEGs) and signaling pathways in neutrophils during this phase, aiming to elucidate their role in sepsis-induced immunosuppression.
The study utilized the GSE64457 gene expression dataset, which includes gene expression profiles from 15 sepsis-induced immunosuppression patients and eight healthy volunteers. This dataset was downloaded from the Gene Expression Omnibus (GEO) database. Differential gene expression analysis was performed using the limma package within the Bioconductor package, implemented in R software. DEGs were identified based on a log2 fold change (FC) greater than 1 and a corrected P value less than 0.05, adjusted for false discovery. The Database for Annotation, Visualization and Integrated Discovery (DAVID) was used for Gene Ontology (GO) annotations and enrichment analysis, focusing on biological processes (BP), molecular functions (MF), and cellular components (CC). Additionally, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was conducted using the KOBAS 3.0 database. Protein-protein interaction (PPI) networks were constructed using the STRING database and visualized using CYTOSCAPE software to identify interactions and pathway relationships among proteins encoded by DEGs in neutrophils during sepsis-induced immunosuppression.
Batch normalization of the GSE64457 dataset revealed 407 DEGs, comprising 227 upregulated and 180 downregulated genes. GO enrichment analysis indicated that these DEGs were significantly enriched in inflammatory response, canonical glycolysis, and positive regulation of nitric oxide biosynthetic process in the BP category. In the MF category, the genes were enriched in phospholipase inhibitor activity, phospholipase A2 inhibitor activity, and major histocompatibility complex class II protein complex binding. In the CC category, the genes were primarily enriched in extracellular exosome, basolateral plasma membrane, and cytosol. These findings suggest that the DEGs in neutrophils during sepsis-induced immunosuppression are predominantly involved in inflammatory response, respiratory burst, metabolic processes, and extracellular exosome functions.
KEGG pathway enrichment analysis identified 97 significantly enriched pathways, with the most notable being the tumor necrosis factor signaling pathway, transcriptional misregulation in cancer, metabolic pathways, and mitogen-activated protein kinase signaling pathway. These pathways are critical in understanding the molecular mechanisms underlying sepsis-induced immunosuppression.
The PPI network analysis, conducted using the STRING database and visualized with CYTOSCAPE, identified the top ten hub genes associated with neutrophils in sepsis-induced immunosuppression. These genes include MMP9, GAPDH, AKT1, JUN, CSF1R, FCGR2B, TLR5, KIT, ANXA5, and ARG1. Among these, MMP9, GAPDH, JUN, TLR5, ANXA5, and ARG1 were upregulated, while AKT1, CSF1R, FCGR2B, and KIT were downregulated. GO analysis revealed that these hub genes are involved in critical biological processes, molecular functions, and cellular components, and are implicated in multiple pathways identified by KEGG analysis.
The upregulation of genes such as JUN and MMP9 suggests their role in promoting neutrophil proliferation and inflammatory responses. Conversely, the downregulation of CSF1R may contribute to reduced neutrophil development and maturation, leading to an increased number of immature granulocytes in the circulation. These findings align with previous studies indicating that even during sepsis-induced immunosuppression, neutrophils retain some functional capacity, contributing to persistent inflammation and impaired immune responses.
The shift from oxidative phosphorylation (OXPHOS) to glycolysis in activated leukocytes, as indicated by the enrichment of canonical glycolysis pathways, highlights the increased energy demands of immune cells during sepsis. This metabolic shift enables rapid ATP production, supporting the specialized activities of immune cells during an inflammatory response.
In conclusion, this study identifies key genes and pathways involved in the neutrophil phenotype during sepsis-induced immunosuppression. The findings are consistent with previous cell, animal, and human studies, providing novel insights into the molecular mechanisms underlying this condition. The identified hub genes, including GAPDH, AKT1, JUN, MMP9, CSF1R, FCGR2B, TLR5, KIT, ANXA5, and ARG1, may serve as potential targets for the diagnosis and treatment of sepsis-induced immunosuppression. Future research should focus on validating these findings and exploring therapeutic interventions targeting these genes and pathways to improve outcomes in sepsis patients.
doi.org/10.1097/CM9.0000000000001878
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