Investigating Regulation of Neutrophil-mediated Inflammation in Mycoplasma pneumoniae Infection

No vaccine exists for Mycoplasma pneumoniae (Mp) infections, largely due to the gap in understanding in the virulence and immunopathogenesis of this bacterium. We have recently demonstrated that neutrophils are maladaptive in the context of Mp infection, and drive disease severity while also impairing bacterial clearance. Furthermore, we observed that B cells play a regulatory role in limiting disease severity by modulating neutrophilic inflammation. We investigated this phenomenon further to elucidate the mechanisms by which B cells regulate neutrophilic inflammation. We utilized a monoclonal antibody to deplete B cells during Mp infection and found that B-cell-depleted mice had elevated neutrophil numbers in the airways compared to B-cell-sufficient controls, indicating exacerbated suppurative pneumonia. We phenotyped airway inflammation via flow cytometry and found an accumulation of Fas+ neutrophils in the airways of B- cell-depleted mice; a population absent in controls. We observed no differences in other assessed death receptors (TNF-R1, TRAIL-R, and PD-1). These data indicate that B cells may regulate neutrophilia via the Fas/FasL pathway. Indeed, using a flow-cytometric based Annexin V assay, we found that greater than 70% of Fas- expressing neutrophils in Mp infected mice are apoptotic – and a small population of B-cells appear to provide the FasL ligand. Collectively, our data indicate that B- cells ameliorate Mp-induced suppurative pneumonia by inducing neutrophil apoptosis via the Fas/FasL pathway. We are currently further interrogating this mechanism via RNAscope staining to identify spatial patterns of expression of Fas and FasL in the lung parenchyma. Understanding this regulatory mechanism can advance immunological research by further characterizing the function of regulatory B cells in disease and may provide insight into regulation of neutrophils during infection. By contributing to basic and applied immunology and microbiology, our work has broad implications for both fields.