Alveolar macrophages in pulmonary host defence the unrecognized role of apoptosis as a mechanism of intracellular bacterial killing

Abstract

Alveolar macrophages play an essential role in clearing bacteria from the lower airway, as the resident phagocyte alveolar macrophages must both phagocytose and kill bacteria, and if unable to do this completely must co-ordinate an inflammatory response. The decision to escalate the inflammatory response represents the transition between subclinical infection and the development of pneumonia. Alveolar macrophages are well equipped to phagocytose bacteria and have a large phagolysosomal capacity in which ingested bacteria are killed. The rate-limiting step in control of extracellular bacteria, such as Streptococcus pneumoniae, is the capacity of alveolar macrophages to kill ingested bacteria. Therefore, alveolar macrophages complement canonical microbicidal strategies with an additional level of apoptosis-associated killing to help kill ingested bacteria.

Keywords: Streptococcus pneumoniae; bacteria; killing; macrophage.

Fig. 1

Macrophages play critical roles in host defence against Streptococcus pneumoniae. S. pneumoniae (Spn) are ingested and killed by alveolar macrophages (AM) in the distal airway. When the bacterial inoculum increases above the level at which AM can contain bacteria without recruitment of further phagocytes cross-talk between AM, epithelial cells (EC) and T cells results in the release of chemokines and recruitment of further inflammatory cells, in particular polymorphonuclear leucocytes (PMN). PMN become critical effectors of bacterial clearance. Ultimately, PMN undergo apoptosis. Monocytes may play an important role in regulating inflammatory cells and may induce apoptosis in activated T cells. Ingestion of apoptotic cells (apoPMN and apoT cells) as well as apoptotic monocytes/macrophages by macrophages, a process termed efferocytosis, helps to down-regulate the inflammatory response and results in the generation of anti-inflammatory cytokines and other factors that help to down-regulate the inflammatory response.

Fig. 2

Streptococcus pneumoniae resists anti-microbial killing. Streptococcus pneumoniae (Spn) express several genes, which help them resist reactive oxygen species, generated within phagocytes. Spn express several enzymes that generate hydrogen peroxide (H₂O₂); SpxB encodes a pyruvate oxidase, Lox a lactate oxidase and CarB a carbamoyl phosphate synthase. Pneumococcal proteins which help resist H₂O₂ include high temperature requirement A (HtrA), a heat shock-stimulated serine protease, the alkylhydroperoxidase D (AhpD), the caseinolytic peptidase P (ClpP), another heat shock-stimulated serine protease, the thioredoxin-like protein A (TlpA), the pneumococcal surface antigen A (PsaA) a manganese permease, PsaD a putative glutathione peroxidase and an alcohol dehydrogenase (AdhC) that generates the reduced glutathione required by PsaD. Resistance to superoxide (O₂^•−) involves the superoxide dismutase A (SodA) and the transcriptional regulator Rgg. Nox is a nicotinamide adenine dinucleotide (NADH) oxidase that converts O₂ to H₂O, limiting the availability of O₂ to form O₂^•−. Spx allows acetyl phosphate (Acetyl P) to provide a source of adenosine triphosphate (ATP) during oxidative stress. In addition, pneumococcal surface protein C (PspC) helps Spn to withstand nitric oxide (NO) and ClpP-reactive nitrogen species (RNS), produced by macrophages.

Fig. 3

Induction of apoptosis-associated killing complements bacterial killing in macrophages. Streptococcus pneumoniae Spn is internalized following phagocytosis into a phagolysosome, where it can resist killing by reactive oxygen species (ROS). With time, the phagolysosome becomes permeabilized and activation of the lysosomal protease cathepsin D (cD) results in down-regulation of the anti-apoptotic protein myeloid cell leukaemia sequence 1 (Mcl-1). Mcl-1 down-regulation involves ubiquitination mediated by interaction with the Mcl-1 ubiquitin ligase E3 (Mule) and proteasomal degradation. Following Mcl-1 down-regulation, pro-apoptotic Bcl-2 family members result in the ultimate activation of Bcl-2-associated X protein (Bax) and Bcl-2 homologous antagonist/killer (Bak), which interact with the mitochondrion (aBax/aBak) to induce mitochondrial outer membrane permeabilization and cytochrome c (Cytc) release. This results in caspase activation (aCasp) and downstream features of apoptosis, illustrated by nuclear fragmentation. Apoptosis results in macrophage cell death but also killing of the residual viable Spn contained in the phagolysosome.