Investigating the role of lipopolysaccharide-stimulated monocytes in the development of immune tolerance in community-acquired pneumonia patients

Abstract

Introduction: Pneumonia remains a leading cause of morbidity and mortality worldwide. Beyond antibiotic treatment, disease severity hinges on how monocytes balance immune defence and tolerance, a process which may paradoxically worsen outcomes. Background: Emerging evidence links monocyte “tolerance” in CAP to DNA methylation and metabolic shifts, yet why patients differ in cytokine output remains unclear. We hypothesized that transcriptional changes in monocytes underlie this heterogeneity. Methodology: The study included samples from a previously executed prospective observational investigation of CAP patients and control subjects (ELDERBIOME; NCT02928367). Specifically, RNA-seq data of monocytes purified from 75 patients, stimulated with LPS- or not. In addition, TNF-α levels in supernatants were used to stratify samples as LPS responders or non-responders. Read libraries were prepared using the KAPA RNA HyperPrep Kit with RiboErase. Sequencing was performed on the Illumina HiSeq 4000 platform. Bioinformatics included standard quality control metrics, graph-based read alignment, and subsequent DESeq2 modelling, and pathway/network enrichment. Results and Discussion: At padj ≤ 0.01, LPS reshaped expression of 7,033 genes (3,878 upregulated; 3,155 downregulated). Enrichment pinpointed heightened cytokine and interferon pathways, with type I interferons (IFNB1) strongly induced. TNF-α stratification revealed two distinct monocyte states: high responders amplified interferon signalling and HLA class II expression; low responders favoured antioxidant, ECM, and solute transport programmes. Conclusion: These findings uncover transcriptional blueprints explaining patientspecific monocyte behaviour in CAP. Understanding this immune polarity could guide strategies to rebalance hyperinflammation without compromising pathogen clearance.