Tuberculosis (TB) is a chronic inflammatory disease resulting from a dysregulated immune response to Mycobacterium tuberculosis (Mtb) infection. Bacterial transmission is mediated by cavitary lung destruction; a consequence of caseating granulomas at the centre of which are necrotic neutrophils. TB patient neutrophils overexpress interferon-gamma (IFN-g)-inducible genes, suggesting that this pro-inflammatory cytokine may be a driver of immune dysregulation in TB patients and highlights the importance of neutrophils in TB disease.
Neutrophil extracellular traps (NETs) are extrusions of decondensed chromatin coated in neutrophil granule derived antimicrobial agents and extracellular matrix degrading enzymes. Released by neutrophils during NETosis this primary effector function aims to trap and kill infecting pathogens. Once thought to be host-protective, accumulating evidence suggests that excessive NET extrusion, as a result of immune dysregulation, culminates in pathological tissue and organ damage.
First, we aimed to address the effect of IFN-g on neutrophil NET production during Mtb infection. Second, to identify the proteins involved in human NETosis that serve as druggable targets to prevent excessive NET production.
Mtb infection, in the presence of IFN-g, enhanced NET release that was profoundly reduced by inhibiting caspase-1 and -4. Concurrently, IFN-g promoted neutrophil viability by inhibiting nuclear decondensation during Mtb infection. Mtb infection also resulted in the formation of processed caspase-1, -4 and -5 capable of activating gasdermin-D, the executioner of NETosis and pyroptosis. Gasdermin-D was found to be post-translationally modified during Mtb infection in multiple ways, in human neutrophils and macrophages. Long gasdermin-D smears manifested concurrently with gasdermin-D p30 subunits in human neutrophils and macrophages. Additionally, smaller than 10kDa gasdermin-D subunits were present as early as 1 hour post-infection in human neutrophils.
Collectively, these findings reveal that IFN- g modulates human neutrophil response during Mtb infection and requires caspase-1 and -4 for enhanced NET production. Furthermore, Mtb interferes with classical gasdermin-D processing, the consequence of which is under further investigation.