Mycobacterium tuberculosis Promotes Apoptosis in Human Neutrophils by Activating Caspase-3 and Altering Expression of Bax/Bcl-xL Via an Oxygen-Dependent Pathway

Abstract

In addition to direct bactericidal activities, such as phagocytosis and generation of reactive oxygen species (ROS), neutrophils can regulate the inflammatory response by undergoing apoptosis. We found that infection of human neutrophils with Mycobacterium tuberculosis (Mtb) induced rapid cell death displaying the characteristic features of apoptosis such as morphologic changes, phosphatidylserine exposure, and DNA fragmentation. Both a virulent (H37Rv) and an attenuated (H37Ra) strain of Mtb were equally effective in inducing apoptosis. Pretreatment of neutrophils with antioxidants or an inhibitor of NADPH oxidase markedly blocked Mtb-induced apoptosis but did not affect spontaneous apoptosis. Activation of caspase-3 was evident in neutrophils undergoing spontaneous apoptosis, but it was markedly augmented and accelerated during Mtb-induced apoptosis. The Mtb-induced apoptosis was associated with a speedy and transient increase in expression of Bax protein, a proapoptotic member of the Bcl-2 family, and a more prominent reduction in expression of the antiapoptotic protein Bcl-xL. Pretreatment with an inhibitor of NADPH oxidase distinctly suppressed the Mtb-stimulated activation of caspase-3 and alteration of Bax/Bcl-xL expression in neutrophils. These results indicate that infection with Mtb causes ROS-dependent alteration of Bax/Bcl-xL expression and activation of caspase-3, and thereby induces apoptosis in human neutrophils. Moreover, we found that phagocytosis of Mtb-induced apoptotic neutrophils markedly increased the production of proinflammatory cytokine TNF-α by human macrophages. Therefore, the ROS-dependent apoptosis in Mtb-stimulated neutrophils may represent an important host defense mechanism aimed at selective removal of infected cells at the inflamed site, which in turn aids the functional activities of local macrophages.

Induction of apoptosis in human neutrophils by Mycobacterium tuberculosis is dependent on mature bacterial lipoproteins

“…The involvement of p38 is consistent with findings reported by Aleman et al (2004), who showed that p38 MAPK was activated in circulating PMNs from patients with tuberculosis, and that these PMNs showed accelerated apoptosis compared to uninfected control cells [31]. However, it is unlikely that circulating PMNs have come in contact Apoptosis elicited by opsonized Mtb is dependent on ROS production by the NADPH oxidase [11,17]. The present experiments show that apoptosis mediated by unopsonized Mtb also requires intracellular ROS, as indicated by the findings that Mtb-induced apoptosis was abrogated by DPI, and that the extent and kinetics of the ROS production were similar for opsonized and unopsonized bacteria.…”

“…Our results showed that, even though PMNs that phagocytose Mtb are left with large numbers of unfused primary granules [14], the moderate ROS production induced no lysosomal damage. However, the mitochondria were central in the apoptosis induction and Δ m was significantly reduced, supporting our previous observation that expression of antagonizing Bcl-2 proteins is altered early during Mtb-induced apoptosis [17]. Low levels of oxidative stress induce apoptosis in several cell types, and ROS or lipid peroxidation products may do so through the redox sensitive mitochondrial permeabilization pore [39,40].…”

“…Unopsonized and opsonized Mtb induced apoptosis to a level of 26 and 35% respectively compared to spontaneously apoptotic control cells (8%) (Fig. 3B), confirming previous findings [7,11,17]. Human PMNs infected with virulent Mtb displayed an Mtb or from Mtb-induced apoptotic PMNs.…”

Murine Macrophages Produce Secretory Leukocyte Protease Inhibitor During Clearance of Apoptotic Cells: Implications for Resolution of the Inflammatory Response

“…It is not known at present why apoptotic CTLL-2 and apoptotic thymocytes have different potencies in inducing TNF-␣ from macrophages. A recent report by Perskvist et al (45) showed that phagocytosis of apoptotic neutrophils markedly increased TNF-␣ production by human macrophages. Thus, apoptotic cells with different origins may differ in their ability to induce inflammatory cytokines during their clearance by macrophages, perhaps due to their distinct surface presentations.…”

Delayed apoptosis of tumor associated neutrophils in the absence of endogenous IFN‐β

“…20 For neutrophils, the expression of the proapoptotic Bcl-2 protein Bax and its antiapoptotic counterpart BCL-xL is of special importance. 21,22 To address whether IFN-b regulates the expression of such proteins and thus influences longevity of neutrophils, we sorted CD11b 1 Gr1 1 cells from For mice on C57BL/6 background B16F10 melanoma cells were injected s.c. into the flank. After 14 days, mice were sacrificed, total cell suspensions of blood and tumor were prepared and neutrophils were isolated using FACS Aria for cell sorting.…”