Identifying molecular signatures of parapneumonic effusion reveals HMGB2 as the disease progression biomarker

Background: Parapneumonic effusion (PPE) exacerbation is associated with high morbidity and mortality in pneumonia patients. However, the molecular signatures underlying the progression and severity of PPE remain poorly understood. We therefore employed whole-transcriptome profiling to comprehensively characterize the molecular landscape associated with PPE exacerbation and its clinical manifestations. Methods: In the discovery cohort, 26 PPE patients underwent capture-based targeted RNA sequencing and were stratified by disease severity into non-severe PPE (UPPE, n = 9) and severe PPE (n = 17), comprising CPPE (n = 12) and empyema (n = 5). Bioinformatic functional annotation was performed to delineate gene expression signatures associated with disease progression from non-severe to severe PPE. Integrated analyses of the transcriptome and secretome of pleural effusions identified high-mobility group box 2 (HMGB2) as a novel biomarker for severe PPE. In the validation cohort, 220 patients with different pleural effusion etiologies were enrolled for enzyme-linked immunosorbent assay (ELISA) analysis to evaluate HMGB2 levels. The functional relevance of HMGB2 in PPE progression was characterized in neutrophils and macrophages. Results: In pneumonia patients, severe PPE was associated with higher expression of 232 genes and lower expression of 212 genes compared with non-severe PPE. Upregulated gene signatures associated with disease progression in severe PPE were enriched in neutrophil degranulation, glycolysis/gluconeogenesis, HIF-1 signaling, and inflammatory response pathways. Elevated HMGB2 levels in severe PPE enabled accurate discrimination from other pleural effusion cases, with an AUC of 0.93. In vitro infection with Streptococcus pneumoniae significantly increased HMGB2, IL1β, and IL8 expression in neutrophils and macrophages. Moreover, HMGB2 overexpression enhanced NF-κB activation as well as IL-1β and IL-8 levels in PGN-stimulated macrophages. These results suggest that HMGB2 regulates IL-1β and IL-8 via the NF-κB signaling pathway. Conclusions: Our study delineates molecular signatures driving PPE exacerbation and highlights HMGB2 as a novel biomarker and therapeutic target with potential to prevent severe PPE.