Lung necrosis and neutrophils reflect common pathways of susceptibility to Mycobacterium tuberculosis in genetically diverse, immune competent mice

Abstract

Pulmonary tuberculosis (TB) is caused by Mycobacterium tuberculosis in susceptible humans. Here, we infected Diversity Outbred (DO) mice with ∼100 bacilli by aerosol to model responses in a highly heterogeneous population. Following infection, ‘supersusceptible’, ‘susceptible’ and ‘resistant’ phenotypes emerged. TB disease (reduced survival, weight loss, high bacterial load) correlated strongly with neutrophils, neutrophil chemokines, tumor necrosis factor (TNF) and cell death. By contrast, immune cytokines were weak correlates of disease. We next applied statistical and machine learning approaches to our dataset of cytokines and chemokines from lungs and blood. Six molecules from the lung: TNF, CXCL1, CXCL2, CXCL5, interferon-γ (IFN-γ), interleukin 12 (IL-12); and two molecules from blood – IL-2 and TNF – were identified as being important by applying both statistical and machine learning methods. Using molecular features to generate tree classifiers, CXCL1, CXCL2 and CXCL5 distinguished four classes (supersusceptible, susceptible, resistant and non-infected) from each other with approximately 77% accuracy using completely independent experimental data. By contrast, models based on other molecules were less accurate. Low to no IFN-γ, IL-12, IL-2 and IL-10 successfully discriminated non-infected mice from infected mice but failed to discriminate disease status amongst supersusceptible, susceptible and resistant M.-tuberculosis-infected DO mice. Additional analyses identified CXCL1 as a promising peripheral biomarker of disease and of CXCL1 production in the lungs. From these results, we conclude that: (1) DO mice respond variably to M. tuberculosis infection and will be useful to identify pathways involving necrosis and neutrophils; (2) data from DO mice is suited for machine learning methods to build, validate and test models with independent data based solely on molecular biomarkers; (3) low levels of immunological cytokines best indicate a lack of exposure to M. tuberculosis but cannot distinguish infection from disease.

Relationship Between HIV Coinfection, Interleukin 10 Production, andMycobacterium tuberculosisin Human Lymph Node Granulomas

“…Neutrophil presence within granulomas increases inflammation and is associated with active tuberculosis [35, 36]. We hypothesize that this influx of neutrophils (a proinflammatory response) in LNs of patients with <50 peripheral CD4 + T cells/μL of blood ultimately led to the increased production of IL-10 (an antiinflammatory response), since their presence was positively correlated within caseous granulomas.…”

Spatial and temporal localization of immune transcripts defines hallmarks and diversity in the tuberculosis granuloma

“…In addition to the encapsulated granuloma with a caseous necrotic center, TB granulomas can be non-necrotizing, neutrophilrich, mineralized, fibrotic or cavitary 4,5 . TB granulomas in commonly used inbred mouse strains such as C57BL/6 do not develop necrotizing lesions, while encapsulated necrotizing granulomas are found in other strains such as the C3HeB/FeJ [6][7][8] . While the histological features of granulomas have been well characterized, the immune and inflammatory mechanisms that underlie variable granuloma dynamics and clinical outcomes of TB infection remain to be further elucidated.…”

Addressing diversity in tuberculosis using multidimensional approaches

“…The most widely used animal model for TB is inbred mice exposed to aerosols of virulent M. tuberculosis ; however, whilst this model likely captures some aspects of TB, the spectrum of disease (early clearance/latent TB/active TB) is not reflected. Instead, the use of outbred animals, including nonhuman primates , guinea pigs and mice generates results that have more validity for TB, as they come closer to human TB disease patterns. For example, a recent study performed by Beamer and colleagues shows that M. tuberculosis infection in ‘diversity outbred’ mice is reflected in heterogeneity, ranging from highly susceptible through susceptible and resistant to noninfected, as observed in human TB .…”

“…Instead, the use of outbred animals, including nonhuman primates , guinea pigs and mice generates results that have more validity for TB, as they come closer to human TB disease patterns. For example, a recent study performed by Beamer and colleagues shows that M. tuberculosis infection in ‘diversity outbred’ mice is reflected in heterogeneity, ranging from highly susceptible through susceptible and resistant to noninfected, as observed in human TB . Furthermore, those mice that developed disease displayed a human‐like TB pathology, including granulomas and neutrophil‐mediated lung necrosis .…”

“…For example, a recent study performed by Beamer and colleagues shows that M. tuberculosis infection in 'diversity outbred' mice is reflected in heterogeneity, ranging from highly susceptible through susceptible and resistant to noninfected, as observed in human TB [64]. Furthermore, those mice that developed disease displayed a human-like TB pathology, including granulomas and neutrophil-mediated lung necrosis [64]. In another study, based on outbred guinea pigs that were 'naturally' exposed to M. tuberculosis though out-air from wards with TB patients, the guinea pigs also displayed a heterogeneous disease outcome pattern ranging from tuberculin skin test reversion (self-healing) to severe pathology.…”