Pleural fluid proteomics from patients with pleural infection shows signatures of diverse neutrophilic responses: The Oxford Pleural Infection Endotyping Study (TORPIDS-2)

Background Pleural infection is a complex disease with poor clinical outcomes and increasing incidence worldwide, yet its biological endotypes remain unknown. Methods We analysed 80 pleural fluid samples from the PILOT study, a prospective study on pleural infection, using unlabelled mass spectrometry. A total of 449 proteins were retained after filtering. Unsupervised hierarchical clustering and UMAP analyses were used to cluster samples and pathway analysis was performed to identify the biological processes. Protein signatures as identified by the pathway analysis were compared to microbiology as defined by 16S rRNA next generation sequencing. Spearman and exact Fischer's methods were used for correlation assessment. Results Higher neutrophil degranulation was correlated with increased glycolysis (OR=281, p<2.2E−16) and pentose phosphate activation (OR=371.45, p<2.2E−16). Samples dominated by Streptococcus pneumoniae exhibited higher neutrophil degranulation (OR=12.08, p=0.005), glycolysis (OR=11.4, p=0.006), and pentose phosphate activity (OR=12.82, p=0.004). On the other hand, samples dominated by anaerobes and Gram-negative bacteria exhibited lower neutrophil degranulation (OR=0.15, p=0.01, glycolysis (OR=0.14, p=0.01), and pentose phosphate activity (OR=0.07, p=0.001). Increased activity of the liver and retinoid X receptors (LXR-RXR) pathway was associated with lower risk of one-year mortality (OR=0.24, p=0.04). Conclusions These findings suggest that pleural infection patients exhibit diverse responses of neutrophil mediated immunity, glycolysis, and pentose phosphate activation which are associated with microbiology. Therapeutic targeting of the LXR-RXR pathway with agonists is a possible treatment approach.