Cytokine activation leads to acidification and increases maturation of Mycobacterium avium-containing phagosomes in murine macrophages

Department of Molecular Microbiology, Washington University, School of Medicine, St. Louis, MO 63110, USA.
The Journal of Immunology (Impact Factor: 5.36). 03/1998; 160(3):1290-6.
Source: PubMed

ABSTRACT Mycobacterium avium (MAC) organisms multiply in phagosomes that have restricted fusigenicity with lysosomes, do not acidify due to a paucity of vacuolar proton-ATPases, yet remain accessible to recycling endosomes. During the course of mycobacterial infections, IFN-gamma-mediated activation of host and bystander macrophages is a key mechanism in the regulation of bacterial growth. Here we demonstrate that in keeping with earlier studies, cytokine activation of host macrophages leads to a decrease in MAC viability, demonstrable by bacterial esterase staining with fluorescein diacetate as well as colony-forming unit counts from infected cells. Analysis of the pH of MAC phagosomes demonstrated that the vacuoles in activated macrophages equilibrate to pH 5.2, in contrast to pH 6.3 in resting phagocytes. Biochemical analysis of MAC phagosomes from both resting and activated macrophages confirmed that the lower intraphagosomal pH correlated with an increased accumulation of proton-ATPases. Furthermore, the lower pH is reflected in the transition of MAC phagosomes to a point no longer accessible to transferrin, a marker of the recycling endosomal system. These alterations parallel the coalescence of bacterial vacuoles from individual bacilli in single vacuoles to communal vacuoles with multiple bacilli. These data demonstrate that bacteriostatic and bactericidal activities of activated macrophages are concomitant with alterations in the physiology of the mycobacterial phagosome.

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    • "IFN-γ and TNF are key cytokines that lead to cell activation and enhanced phagosome–lysosome fusion in mycobacteria-infected macrophages (Schaible et al., 1998; Via et al., 1998). Phagolysosomes are formed upon fusion of phagosomes with intracellular organelles, in particular lysosomes. "
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    ABSTRACT: Infection of humans with Mycobacterium tuberculosis remains frequent and may still lead to death. After primary infection, the immune system is often able to control M. tuberculosis infection over a prolonged latency period, but a decrease in immune function (from HIV to immunosenescence) leads to active disease. Available vaccines against tuberculosis are restricted to BCG, a live vaccine with an attenuated strain of M. bovis. Immunodeficiency may not only be associated with an increased risk of tuberculosis, but also with local or disseminated BCG infection. Genetic deficiency in the reactive oxygen species (ROS)-producing phagocyte NADPH oxidase NOX2 is called chronic granulomatous disease (CGD). CGD is among the most common primary immune deficiencies. Here we review our knowledge on the importance of NOX2-derived ROS in mycobacterial infection. A literature review suggests that human CGD patient frequently have an increased susceptibility to BCG and to M. tuberculosis. In vitro studies and experiments with CGD mice are incomplete and yielded - at least in part - contradictory results. Thus, although observations in human CGD patients leave little doubt about the role of NOX2 in the control of mycobacteria, further studies will be necessary to unequivocally define and understand the role of ROS.
    Cellular Microbiology 06/2014; 16(8). DOI:10.1111/cmi.12322 · 4.82 Impact Factor
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    • "MtbDwhiB4 grew at a level similar to wt Mtb inside resting macrophages (Fig. S9). Since macrophages activated by IFN-g and LPS generate a more hostile environment by inducing vacuole acidification , nutrient depletion, and enhanced ROI/RNI production to constrain bacterial growth (James et al., 1995; Schaible et al., 1998), we analysed the influence of the activation status of the infected macrophages on the survival of MtbDwhiB4. In activated macrophages, we observed an approximately 50-fold enhanced survival of MtbDwhiB4 as compared with wt Mtb at day 2 post infection (Fig. 7A). "
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    ABSTRACT: Host-generated oxidative stress is considered one of the main mechanisms constraining Mycobacterium tuberculosis (Mtb) growth. The redox-sensing mechanisms in Mtb are not completely understood. Here we show that WhiB4 responds to oxygen (O(2) ) and nitric oxide (NO) via its 4Fe-4S cluster and controls the oxidative stress response in Mtb. The WhiB4 mutant (MtbΔwhiB4) displayed an altered redox balance and a reduced membrane potential. Microarray analysis demonstrated that MtbΔwhiB4 overexpresses the antioxidant systems including alkyl hydroperoxidase (ahpC-ahpD) and rubredoxins (rubA-rubB). DNA binding assays showed that WhiB4 [4Fe-4S] cluster is dispensable for DNA binding. However, oxidation of the apo-WhiB4 Cys thiols induced disulphide-linked oligomerization, DNA binding and transcriptional repression, whereas reduction reversed the effect. Furthermore, WhiB4 binds DNA with a preference for GC-rich sequences. Expression analysis showed that oxidative stress repressed whiB4 and induced antioxidants in Mtb, while their hyper-induction was observed in MtbΔwhiB4. MtbΔwhiB4 showed increased resistance to oxidative stress in vitro and enhanced survival inside the macrophages. Lastly, MtbΔwhiB4 displayed hypervirulence in the lungs of guinea pigs, but showed a defect in dissemination to their spleen. These findings suggest that WhiB4 systematically calibrates the activation of oxidative stress response in Mtb to maintain redox balance, and to modulate virulence.
    Molecular Microbiology 07/2012; 85(6):1148-1165. DOI:10.1111/j.1365-2958.2012.08165.x · 5.03 Impact Factor
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    • "Bacilli which survive proliferate logarithmically within alveolar macrophages and DCs and induce the production of immune mediators such as TNF-α, IL-6, IL-12p80, IL-1α, and IL-1β that activate macrophages to induce early bacterial killing [5] [16]. IFN-γ is a proinflammatory cytokine produced by CD4+ and CD8+ T cells as well as by activated NK cells in response to IL-12 and IL-18 produced by alveolar macrophages and DCs [15, 17– 19]. "
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    ABSTRACT: Mycobacterium tuberculosis (Mtb) infection is a major international public health problem. One-third of the world's population is thought to have latent tuberculosis, a condition where individuals are infected by the intracellular bacteria without active disease but are at risk for reactivation, if their immune system fails. Here, we discuss the role of nonspecific inflammatory responses mediated by cytokines and chemokines induced by interaction of innate receptors expressed in macrophages and dendritic cells (DCs). We also review current information regarding the importance of several cytokines including IL-17/IL-23 in the development of protective cellular and antibody-mediated protective responses against Mtb and their influence in containment of the infection. Finally, in this paper, emphasis is placed on the mechanisms of failure of Mtb control, including the immune dysregulation induced by the treatment with biological drugs in different autoimmune diseases. Further functional studies, focused on the mechanisms involved in the early host-Mtb interactions and the interplay between host innate and acquired immunity against Mtb, may be helpful to improve the understanding of protective responses in the lung and in the development of novel therapeutic and prophylactic tools in TB.
    Clinical and Developmental Immunology 05/2012; 2012:193923. DOI:10.1155/2012/193923 · 2.93 Impact Factor
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