Synergistic function of C5aR1 and C5aR2 complement receptors in defending against bacterial pneumonia 4441

Description Streptococcus pneumoniae (Spn) is the leading cause of pneumonia-associated mortality in adults. Complement activation is critical for encapsulated Spn clearance but also augments ARDS pathogenesis. C5a binding to its two receptors, C5aR1 and C5aR2, activates inflammatory cells. While C5a neutralization is protective in sepsis and inflammation models, its role in Spn infection remains unclear. To address this gap, we generated a novel C5aR1/2-/- double-knockout mouse strain using CRISPR/Cas9. Alveolar leukocytes were profiled at the single-cell level using CITE-sequencing containing a ∼200 antibody panel. Despite the lack of C5a-induced neutrophil influx, C5aR1/2-/- mice exhibited paradoxical exacerbation of alveolar inflammation after Spn infection, with increased neutrophil infiltration, elevated cytokine/chemokine release, and enhanced lung vascular permeability. The recruited neutrophils in C5aR1/2-/- mice showed greater NETosis but impaired bactericidal function evidenced by higher CFUs in bronchoalveolar lavage fluid. CITE-seq revealed distinct subclusters of alveolar macrophages and neutrophils after Spn infection, with heterogeneous C5aR1 and C5aR2 expression in WT mice. C5aR1/2-/- neutrophils showed increased apoptosis and signs of exhaustion (elevated PD-L1/ICAM-1 and reduced CXCR2/CD62L). In conclusion, C5aR1/2-/- neutrophils fail to control bacterial growth, fostering a potent chemotactic local environment for sustained inflammation, thereby promoting lung damage.