Permissive lung neutrophils facilitate tuberculosis immunopathogenesis in male phagocyte NADPH oxidase-deficient mice

Abstract

NADPH oxidase 2 (NOX2) is an enzyme responsible for generating reactive oxygen species, primarily found in phagocytes. Chronic Granulomatous Disease (CGD), along with bacterial infections such as Mycobacterium tuberculosis (Mtb), is a representative NOX2-deficient X-linked disease characterized by uncontrolled inflammation. However, the precise roles of host-derived factors that induce infection-mediated hyperinflammation in NOX2-deficient condition remain incompletely understood. To address this, we compared Mtb-induced pathogenesis in Nox2-/- and wild type (WT) mice in a sex-dependent manner. Among age- and sex-matched mice subjected to Mtb infection, male Nox2-/- mice exhibited a notable increase in bacterial burden and lung inflammation. This was characterized by significantly elevated pro-inflammatory cytokines such as G-CSF, TNF-alpha, IL-1 alpha, IL-1 beta, and IL-6, excessive neutrophil infiltration, and reduced pulmonary lymphocyte levels as tuberculosis (TB) progressed. Notably, lungs of male Nox2-/- mice were predominantly populated with CD11bintLy6GintCXCR2loCD62Llo immature neutrophils which featured mycobacterial permissiveness. By diminishing total lung neutrophils or reducing immature neutrophils, TB immunopathogenesis was notably abrogated in male Nox2-/- mice. Ultimately, we identified G-CSF as the pivotal trigger that exacerbates the generation of immature permissive neutrophils, leading to TB immunopathogenesis in male Nox2-/- mice. In contrast, neutralizing IL-1 alpha and IL-1 beta, which are previously known factors responsible for TB pathogenesis in Nox2-/- mice, aggravated TB immunopathogenesis. Our study revealed that G-CSF-driven immature and permissive pulmonary neutrophils are the primary cause of TB immunopathogenesis and lung hyperinflammation in male Nox2-/- mice. This highlights the importance of quantitative and qualitative control of pulmonary neutrophils to alleviate TB progression in a phagocyte oxidase-deficient condition. While tuberculosis (TB) remains a global threat to public health, the immunologic factors contributing to TB susceptibility are not yet fully defined. To enhance our understanding of TB immunopathogenesis, we utilized TB-susceptible male Nox2-/- mice to dissect the immunologic factors accelerating TB pathogenesis. In this study, we observed that male Nox2-/- mice infected with Mycobacterium tuberculosis (Mtb) exhibited increased lung hyperinflammation and mycobacterial burden compared to female Nox2-/- mice and WT mice. Exacerbated TB immunopathogenesis in male Nox2-/- mice was strongly correlated with an extreme infiltration of pulmonary neutrophils and the loss of pulmonary lymphocytes. The pulmonary neutrophils in male Nox2-/- mice displayed immature phenotypes and mycobacterial permissiveness. The depletion of total neutrophils or the AM80-induced maturation of neutrophils ameliorated TB pathogenesis in Nox2-/- mice, reducing pulmonary neutrophil counts, lung hyperinflammation, and mycobacterial load. Furthermore, neutralization of G-CSF mitigated TB pathogenesis in male Nox2-/- mice by decreasing immature neutrophil counts. Thus, we identified that G-CSF-mediated generation of permissive immature neutrophils contributes to TB susceptibility in male Nox2-/- mice.