Macrophages in Microbial Pathogenesis: Commonalities of Defense Evasion Mechanisms

ABSTRACT

Macrophages are key arsenals of the immune system against invaders. After compartmental isolation of a pathogen in phagosomes, the host immune response attempts to neutralize the pathogen. However, pathogens possess the ability to subvert these assaults and can also convert macrophages into their replicative niche. The multiple host defense evasion mechanisms employed by these pathogens include phagosome maturation arrest, molecular mimicry through secretory antigens, interference with host signaling, active radical neutralization, inhibition of phagosome acidification, alteration of programmed cell death, and other mechanisms. Macrophage biology as a part of the host-pathogen interaction has expanded rapidly in the past decade. The present review aims to shed some light upon the macrophage defense evasion strategies employed by pathogens. We have also incorporated recent knowledge in the field of macrophage dynamics during infection and evolutionary perspectives of macrophage dynamics.

INTRODUCTION

Macrophages are among the first responders of the innate immune system during an infection. They show diverse anatomical appearances with phenotypic variability and plasticity. Depending on their location, macrophages go by many different names: osteoclasts in the bone, Kupffer cells in the liver, alveolar macrophages in the lungs, microglia in the central nervous system, and more.

More than a century ago, Elie Metchnikoff discovered that phagocytic cells could internalize particles and, specifically, that these particles were animate or inanimate irritants other than food entering the body (1). Now, macrophages are at the center stage in the study of innate immunity. Decades of research have revealed the roles of macrophages to involve detection-engulfment-killing of pathogens, secretion of cytokines and chemokines for recruitment of other immune cells, antigen presentation for invoking adaptive responses, and the cleaning and restoration of damaged tissue.

Macrophages recognize pathogen associated molecular patterns (PAMPs) using specific pattern recognition receptors (PRRs) present on their cell surface, inside vesicles, or in the cytoplasm. Following recognition, macrophages engulf the pathogen, leading to lysosomal fusion and phagosome maturation that subject the pathogen to multiple attacks that eventually result in microbial degradation. PRRs and associated mechanisms of detection of the pathogen are detailed in depth in a recent review by Gunter Weiss and Ulrich E. Schaible (2). Adaptive immunity is then activated when pathogen-loaded macrophages move to lymph nodes and activate T cells. However, activation is not exclusive to macrophages; dendritic cells are also involved in antigen presentation that activates adaptive immunity. In addition, both the humoral arm of the immune system and the complement system can recruit macrophages (2). Antibody opsonization of bacteria greatly facilitates their phagocytosis by macrophages. The classical complement activation assembles the membrane attack complex and eventually tags the bacterial surface with opsonins that allow for their recognition by macrophages and neutrophils.

Despite the molecular arsenal used by macrophages to defend against pathogens, macrophages are often exploited as a suitable replicative niche by intracellular pathogens. This is advantageous to the intracellular pathogen, as it offers protection from other immune challenges and sufficient nutrients for replication. The different molecular mechanisms employed by pathogens to neutralize macrophage defenses will be detailed in the present review. We will also discuss the evolutionary perspective of unicellular grazing and its analogy with macrophages and bacterial interaction (3).

MACROPHAGE-PATHOGEN INTERACTION: EVOLUTIONARY PERSPECTIVE AND WINNER’S CONUNDRUM

The primitive microscopic world has been locked in a constant battle for billions of years. This struggle between bacteria and bacteriophages or unicellular protozoa and bacteria is referred to as “unicellular grazing” (Fig. 1). From this struggle, lineages of bacteria and other microbes have evolved molecular tools to defend against predatory attacks, to become the predator, or to engage in a combination of both strategies. These tools could be multiples of secretory effectors, molecular mimics, and immune mediators. For example, a specific mode of defense against grazers is the secretion of antipredator molecules, such as antibacterial fatty acids like Chlorellin by Chlorella vulgaris or acrylate by the alga Emiliania huxleyi (4, 5). In addition, diatoms can secrete oxylipins that reduce the reproductive success of herbivorous grazers copepods (6). Another mode of defense against grazers is morphological adaptation, as shown by the planktonic microalgae Microcystis aeruginosa, which forms colonies when it senses the grazer Ochromonas sp., or the filamentous cyanobacteria Aphanizomenon, which form flake-like assemblages instead of single filaments when they sense the grazer Daphnia (7). Similarly, bacteria prevent bacteriophage attack by blocking DNA entry, utilizing restriction-modification systems, and undergoing programmed cell death after infection. Bacteria can also activate expression of phage-inducible chromosomal islands that inhibit phage reproduction or activate interference of the clustered regularly interspaced short palindromic repeats (CRISPR) (8, 9).

FIG 1.

The macrophage paradox: winner’s conundrum. A constant battle between pathogen and host exists, and both of them are developing their arsenals to defeat the other over evolutionary time scale.

The phagocytic mechanisms of single-celled eukaryotes are well conserved from amoeba to macrophages. In fact, several human pathogens that can replicate inside macrophages can also thrive and replicate within amoeba. For example, Shigella, Salmonella, Coxiella, Burkholderia, Francisella, Chlamydophila pneumoniae, Mycobacterium avium, Listeria monocytogenes, and Pseudomonas aeruginosa have all been shown to replicate within amoeba (3, 10). While hopping the host, these pathogens can parasitize the macrophages using similar molecular tools. Thus, they are successful against macrophages or other cell types. This ability to hop between hosts might have happened a myriad number of times during the coevolution of bacteria and humans. Thus, macrophages as a replicative niche may not be hostile for many microbes; instead, they could be familiar places for those that evolved the appropriate defense strategies to survive there (3).

Once infected, macrophages can either limit the access of pathogens or spread them systemically. Thus, either macrophages could be a winner by containing the pathogens or pathogens could be a winner by exploiting macrophages for their spread. We can further discuss the winner’s conundrum of this battle. For example, Mycobacterium tuberculosis can be confined within macrophages in a nonreplicative latent phase indefinitely, until, in some cases, the host’s immune system becomes compromised and M. tuberculosis reactivates. While the majority of M. tuberculosis cases can be viewed as macrophages having conquered the pathogen, one might object to that view since a large population of infected phagocytes may provide the pathogen a population-dependent advantage upon reactivation. In addition, M. tuberculosis dormancy can limit potential therapeutic interventions, making eradication of the pathogen from the population difficult, giving an edge to bacteria. Nevertheless, a large portion of the infected population remains disease- and symptom-free as M. tuberculosis remains dormant within macrophages. Because the lifetime risk of reactivation is 5 to 10% for a person with a recorded latent tuberculosis infection, it is hard to determine who the true “winner” is (11).

Despite many pathogens preferring macrophages as their replicative niche, the choice of cell type for replication is not always macrophages. Several viruses, protozoa, and fungi favor different host cell tropism (12, 13). In the case of viruses, cell surface receptors direct viral entry to only specific cells, e.g., Epstein-Barr virus (EBV) displaying tropism for B-cell gp42 gHgl receptors (14). Regulation of specific cell surface receptors or critical regulators drives a pathogen’s host cell tropism. For example, macrophages expressing deoxynucleoside triphosphate (dNTP) hydrolase SAMHD1 can resist retroviral infection by nucleotide starvation (15, 16), or phagocytosis of Streptococcus pneumoniae by lung macrophages is reduced after influenza A virus strain X31 infection due to dampening of CD36 receptor expression (12).

CELLULAR EVENTS OF MACROPHAGE DEFENSES

In the bone marrow, committed myeloid progenitor cells, generated from hematopoietic stem cells, give rise to macrophage precursors, known as monocytes. Monocytes migrate to different tissues, differentiate into specialized macrophage subpopulations, and participate in immune defenses via recognition, internalization, and presentation of antigens, along with regulation of inflammation and eventual pathogen clearance (17).

During macrophage-pathogen interactions, PRRs, like Toll-like receptors (TLRs), recognize conserved PAMPs like lipoproteins, lipopolysaccharides, bacterial flagellins, or nucleic acids (18). Recognition and internalization of the pathogen initiate activation of downstream signaling, such as MyD88/TRIF-mediated initiation of cytokine and chemokine signaling (19). Similar to TLRs, different PRRs may initiate different pathways, depending upon the nature of the external stimuli. Similarly, nucleotide-binding oligomerization domain 1 (NOD1), NOD2, and cyclic GMP-AMP are sensors for bacterial peptidoglycan and cytoplasmic DNA, respectively. The former activates proinflammation via NF-κB, and the latter proceeds with STING-mediated activation of type 1 interferons (20, 21).

Maturation of pathogen-loaded phagosomes within the macrophage leads to strong microbicidal effects on the pathogen mediated by multiple different strategies: production of reactive radicals, acidification of the phagosome environment, production of proteases, synthesis of antimicrobial peptides, nutrient starvation, and modulation of programmed cell death (2).

The antimicrobicidal effect of reactive radicals is generated through the actions of inducible nitric oxide synthase (NOS2) and NADPH oxidase 2 (NOX2). Activated macrophages generate reactive nitrogen intermediates, like nitric oxide (NO), through the activity of NOS2. NOS2 is a homodimeric enzyme that produces NO and citrulline using arginine and oxygen as substrates, with the exchange of electrons from one NOS2 subunit to the heme iron of the second NOS2 subunit facilitated by NADPH, FAD, and FMN (22, 23). Reactive oxygen radicals are generated in activated macrophages by NOX2 (24). The NOX2 complex is comprised of five catalytic subunits that generate superoxide anions, hydrogen peroxide, and hydroxyl radicals (2, 23). These reactive radicals exert antibacterial action by damaging DNA and other chemical moieties, such as Fe-S clusters, tyrosyl radicals, hemes, sulfhydryls, thioethers, and alkenes (23).

Phagosome acidification, as a part of phagosome maturation, depends on the vacuolar-type ATPase (V-ATPase) complex where hydrolysis of ATP by the complex’s cytoplasmic component leads to displacement of its membrane ring component, resulting in the transmembrane displacement of protons (25, 26). Acidification promotes phagolysosomal maturation and provides the optimal pH for proteases like cathepsins and the metal transporter NRAMP-1 (Slc11A1) (27, 28).

The nutrients and growth factors required by intracellular pathogens are heavily regulated by macrophages forming an important antimicrobial strategy. Macrophages infected with nonpathogenic Mycobacterium smegmatis deplete intracellular iron stores over time, but infection with pathogenic M. tuberculosis causes an increase in macrophage iron reserves (29). This suggests that pathogen-induced variations in microenvironments may exist within macrophages. Variations in the intracellular nutrient status of host cells affect the expression of several virulence-associated genes in inhabiting microbes. The presence of micronutrients is required for regulation of virulence factors, like magnesium for the PhoP/PhoQ two-component system in Salmonella, iron for Shiga toxin in Shigella, calcium for the Yop virulon in Yersinia, and iron for diphtheria toxin in Corynebacterium (30–33). Thus, unsuccessful microbes are restricted by both nutrient deprivation and phagolysosome toxicity, but successful pathogens are able to manage both. In a Legionella pneumophila mutant study, a tryptophan, but not a thymidine auxotroph, can proliferate within human monocytes, suggesting that the phagosome lacks only certain nutrients (34). Furthermore, activation of macrophages by cytokines can affect nutrient acquisition and availability within macrophages. Gamma interferon (IFN-γ)- and tumor necrosis factor alpha (TNF-α)-activated macrophages increase NRAMP1 (Slc11A1) expression, depleting iron, manganese, and other cations necessary for microbial growth (35, 36).

Coordination with antimicrobial peptides is also an important arm of macrophage defense. Macrophages, granulocytes, and other immune cells express antimicrobial peptides that create membrane pores that affect internal targets in the pathogen (37). Human cathelicidin class antimicrobial peptide LL-37 can reduce proinflammatory macrophage responses by decreasing reduced TNF-α and NO levels in vitro and in vivo (38). The macrophage-expressed antimicrobial peptide CRAMP, which is related to murine cathelicidin, impairs replication of phagocytosed Salmonella (39). The human neutrophil peptides 1 to 3 (HNP1–3), α-defensins, enhance macrophage phagocytosis and immunomodulatory action during a Staphylococcus aureus infection (40).

MACROPHAGES: IMMUNOLOGICAL CROSS-TALKS AND INFLAMMATORY CONFLICT

There is an inflammatory burst after infection that damages the host tissues which has to be subdued for repair. This inflammatory conflict is resolved by macrophages at the time of killing of pathogen or repair of damaged tissues. Earlier models explain the existence of the M1/M2 polarized macrophage phenotype dichotomy of activated macrophages and subsequent inflammatory responses.

Classically activated M1 polarized macrophages create a proinflammatory microenvironment mediated by interleukin 1β (IL-1β), IL-6, IL-12, IL-23, and TNF-α, whereas activated M2-polarized macrophages cause anti-inflammatory responses mediated by IL-10, transforming growth factor β (TGF-β), chemokine ligand 1 (CCL1), CCL2, CCL17, CCL18, and CCL22 (41). Cultured human macrophage cells are considered fully polarized and ready to use after being polarized to proinflammatory (M1) by IFN-γ (and often in the presence of lipopolysaccharide [LPS]) or polarized to anti-inflammatory (M2) by IL-4 and/or IL-13 (42, 43). This balance of polarization elicits either the T-helper-1 or T-helper-2 immune response, respectively.

In current views, the state of M1/M2 polarization of macrophages is flexible and dynamic. The phenotypic switching of macrophages greatly depends upon the microenvironment (44, 45). In addition, polarization states are not binary, and macrophages can exist in between the polarization extremities (45). Along with the canonical polarized population of M1/M2 macrophages, regulatory macrophages limit inflammatory responses (46).

Plasticity of macrophages allows for recalibration of their physiology in response to environmental cues (47). The hallmark of macrophage activation is expression of markers and secretion of cytokines and chemokines that recruit leucocytes and create or resolve inflammation. IFN-γ secreted by immune cells, such as natural killer (NK) cells, CD8⁺ T cells, or CD4⁺ Th1 cells, induces or upregulates many M1 markers in macrophages (45). An upregulation is achieved by stimulation of IL-12 and IL-18 in mice (48, 49). During an infection, M1 polarization is microbicidal and inflammatory in nature, in contrast to the M2 polarization, which is immunomodulatory and weakly microbicidal. In response to bacterial infection, expression of common host response genes in macrophages is strongly skewed to M1 polarization, expressing TNF-α, IL-6, IL-12, IL-1 β, and IL-23 and signaling multiple microbicidal mechanisms (17, 50). M. tuberculosis and Helicobacter pylori infections induce M1 polarization of macrophages, resulting in secretion of proinflammatory cytokines (41). Macrophages in M. tuberculosis lung granulomas and Salmonella enterica serovar Typhimurium splenic granulomas show heterogeneous polarization, anti-inflammatory at the periphery and proinflammatory at the inner region (51, 52). Bacterial effectors like S. Typhimurium SteE or M. tuberculosis ESAT-6 contribute to the polarization states of the macrophages, influencing the overall outcome of the infection (persistence versus clearance of intracellular bacteria) (52, 53). This is consistent with the developmental stages of a granuloma, assisted by macrophage M1 inflammatory response and time-dependent transition of response to M2 polarization (54). For further reading, post-M. tuberculosis infection tissue pathology in different study models has been presented by Sakamoto (55). Similarly, in Leishmania infections, M1 macrophage polarization is linked with inflammation and clearance of infection whereas M2 polarization is associated with susceptibility and disease progression (56). Aggravated inflammatory Th-1 response causes tissue damage and ulcer formation, but M2 polarization is permissive to infection.

M2 macrophages suppress the inflammatory responses and play a pivotal role in healing damaged tissues (17, 45). CD4⁺ T cells, basophils, eosinophils, and mast cells secrete IL-4 and IL-13 and in turn activate the M2 phenotype to support wound healing. M2 polarization is permissive in the Cryptococcus neoformans infection and fatal for monocytes (57). Inhibiting the anti-inflammatory microenvironment by blocking IL-4 and IL-13 improves the infection and promotes proinflammatory M1 polarization, further inducing fungicidal action (58–60). The anti-inflammatory phenotype in liver macrophages support hepatitis B-associated pathogenesis (61), while zika-infected macrophages show limited proinflammatory responses (62).

EVASION MECHANISMS OF MACROPHAGE DEFENSES

During host-pathogen interactions, multiple macrophage assaults are neutralized by invading pathogens at different phases of their interaction (Fig. 2). Evasion of macrophage defense starts with evasion of phagocytosis, followed by subversion of intraphagosomal assaults, disruption of phagosome maturation, disabling of nutrient sequestration, modulation of membrane repair processes, nonlytic expulsion, and finally, modulation of programmed cell death.

FIG 2.

Pictorial depiction of common modalities of macrophage defense evasion mechanisms. Surface pathogen recognition receptors (PRRs) recognize and internalize pathogens. Pathogens residing at phagosome secrete multiple effectors, neutralize multiple immune assault, target cytosolic PRR mediated signaling, inhibit vacuolar acidification, and arrest lysosomal fusion and phagosome maturation. Further, pathogens modulate vesicular trafficking, chromatin remodeling, and cytokine balance. Pathogens also use specialized secretion systems for iron and nutrient acquisition. They may escape to cytosol and target ubiquitination and lipid raft-dependent signaling network.

Evasion of recognition and engulfment.

Pathogens subvert macrophage engulfment by avoiding detection, impairing signaling, and disabling the phagocytic machinery. To limit recognition and phagocytosis, pathogens apply the strategies of encapsulation and inactivation of complement/antibody (63). Encapsulation also protects against opsonization-mediated mechanisms of detection. Attachment to the host surface without eventual internalization is achieved by interfering with host signaling and modulation of the host cytoskeleton (63).

Microbial immunogenic signatures essential for their recognition (PAMPs) are masked by encapsulation to evade host recognition. Neisseria meningitidis, Haemophilus influenzae, Pseudomonas aeruginosa, Streptococcus spp., and C. neoformans all utilize this strategy (64). Effector-mediated evasion of TLR signaling through LPS modification and flagella production are strategies used by Francisella novicida, S. Typhimurium, Porphyromonas gingivalis, H. pylori, L. pneumophila, Yersinia spp., Campylobacter jejuni, and Bartonella bacilliformis (65–69). Though TLR-5 is evolutionarily adapted for recognition of conserved sites in bacterial flagellin, this recognition can be easily evaded through alteration/mutation of these sites. Andersen-Nissen et al. have reported that Salmonella flagellin is strongly recognized by TLR-5; however, H. pylori flagellin, FlaA, contains mutations at residues 89 to 96, destroying TLR-5 recognition and, simultaneously, bacterial motility (70). Opsonization of pathogens facilitates their recognition and results in their clearance. Production of immunoglobin digestive proteases by Streptococcus pneumoniae, Neisseria spp., and Staphylococcus aureus (71–73) or sequestration of factor H by Neisseria meningitidis, S. aureus, and Borrelia burgdorferi (74–76) results in impairment of the complement pathway or scavenging of opsonizing antibody.

Fc-receptor-dependent phagocytosis requires phosphorylation of critical tyrosine residues by Src-family kinases. Activation of Rho GTPases, present downstream of these kinases, is required for completion of the signaling cascade for engulfment. The Yop H factor of Yersinia spp. is delivered to the host through its type-3 secretion system, and Yop H dephosphorylase activity is able to disrupt the phagocytic signaling cascade of macrophages (77). P. gingivalis secretory effectors, gingipans, impair macrophage migration and reduce the expression of surface CD14, impairing macrophage interaction with apoptotic cells and inducing inflammation. Poor bacterial clearance with high inflammation leads to periodontal pathology (78).

Of note, certain pathogens benefit from phagocytosis or transcytosis and use the processes to establish infections in the host. Phosphatidylinositol derivatives undergo kinase action, generating secondary messengers that mimic the signaling cascade or directly recruit pleckstrin homology domain-containing proteins that are part of actin polymerization signaling and microbial engulfment. Modulation of phosphatidylinositol kinases or phosphatases affects bacterial entry into the cell for pathogens such as S. enterica, Shigella flexneri, and Vibrio parahaemolyticus (79–82).

Evasion of phagosomal mechanisms.

During the phagosome maturation cascade, Rab5 GTPases in the early phagosome are replaced by Rab7 GTPases in the late phagosome, mediating fusion with lysosomes (83). Rab5, Rab22, Rab14, Rab20, and Rab21 are also a part of early phagosomes and participate in the maturation pathway (84). Pathogens target different Rab proteins to block lysosomal fusion, enhancing their intracellular survival. Rab32 is recruited by infected vacuoles containing M. tuberculosis, S. aureus, or Salmonella enterica serovar Typhi (85–87). Phagosomes loaded with M. tuberculosis modify Rab22, resulting in the accumulation of Rab14 that is inhibitory to lysosomal fusion (88). Mycobacterial secretory protein kinase G exclusively targets Rab7l1, blocking lysosomal fusion and promoting bacterial survival within the phagosome (89). It has been reported that Listeria-infected phagosomes exhibit reduced lysosomal fusion (90).

During phagosome maturation, V-type ATPases accumulate in the phagosome membrane and translocate protons into the phagosome at the expense of cytosolic ATP (25). The arrest of phagosome acidification helps in the survival of pathogens within the phagosome, and several pathogens specifically attenuate the activity of V-ATPases to prevent the acidification of vacuoles. For example, Yersinia pseudotuberculosis, M. avium, and M. tuberculosis block the action of V-H⁺ ATPases (91–93). The secretory M. tuberculosis PtpA (protein tyrosine phosphatase A) interacts with the H subunit of H⁺ V-ATPase, blocking complex assembly and thus preventing vacuolar acidification (93). M. tuberculosis can also degrade H⁺ V-ATPase from infected phagosomes by enhancing secretion of granulocyte-macrophage colony-stimulating factor (GM-CSF), which induces STAT5-mediated expression of cytokine-inducible SH2-containing protein (CISH) signaling, ubiquitinating and degrading the A subunit of V-ATPase (94).

Microbial growth within phagosomes requires a steady supply of essential nutrients, such as amino acids and iron. Siderophores produced by pathogens help subvert nutrient sequestration, as in the case of Aspergillus fumigatus and Histoplasma capsulatum (95, 96). Pathogens within the phagosome of macrophages utilize specialized secretion systems to release toxins and virulence factors into the host that facilitate nutrient acquisition. The M. tuberculosis ESX-3 secretion system is specialized for iron acquisition, allowing a continued supply of the essential metal despite the macrophages’ attempts to limit it (97). In a similar process, M. tuberculosis PE4-PPE5 proteins work as a siderophore for iron uptake, as well (98).

The NADPH oxidase complex is a key macrophage defensive response mechanism that delivers a regulated burst of ROS to phagosomal pathogens. This mechanism offers protection against Mycobacterium and Salmonella infection, among others (99, 100). M. tuberculosis effector protein CpsA inhibits reactive oxygen species (ROS) generation by NADPH oxidase and bypasses LC3-associated phagocytosis (101). Superoxide dismutase neutralizes the ROS by converting it to H₂O₂ in Pseudomonas aeruginosa and M. tuberculosis (102, 103). Similarly, sodA and sodM of S. aureus encode superoxide dismutases for ROS neutralization (104). KatG, a catalase-peroxidase, converts H₂O₂ into oxygen and water and is another defense mechanism against ROS utilized by M. tuberculosis and S. aureus (105, 106). Internalized microorganisms can also neutralize NO radicals generated by NOS2. The two-component system of S. aureus, SrrAB, recognizes nitrosative stress via histidine kinase sensors (107, 108).

Evasion of cytosolic defenses.

A nonlytic phagosome escape strategy of fungi, known as vomocytosis, allows them to escape without cell lysis of the host. This was first observed in C. neoformans and later on in Candida krusei and Candida albicans (109, 110). The mechanism of action is not yet clear, but what is known is that the absence of fungal phospholipase and urease, phagosome acidification, the formation of actin cages, and M2 macrophage polarization are inhibitory to vomocytosis (110). Several reports present a similar mechanism of bacterial escape (111–113).

Coronin proteins are actin-binding proteins in eukaryotes with a primary role in actin polymerization dynamics. Interesting, they are also important in pathogen-host interactions and are recruited to the vacuolar surface to help establish bacterial infections of M. tuberculosis, Mycobacterium leprae, and Helicobacter pylori and viral infection of lymphocytic choriomeningitis virus (LCMV) and vesicular stomatitis Indiana virus (VSV). Coronin-1 blocks lysosomal fusion by activating calcium–calcineurin signaling in infected phagosomes, which is facilitated by bacterial effectors like Lam, SapM, and PtpA (114, 115).

Evasion of membrane-related mechanisms.

Membrane microdomains enriched with cholesterol, sphingolipids, and glycosylphosphatidylinositol (GPI)-anchored proteins are known as lipid rafts. Lipid rafts participate in a variety of cellular processes and internalization of bacteria, viruses, and their toxins (116, 117). H. pylori glycosylates the host membrane cholesterol, reducing cholesterol from the host lipid rafts and promoting gastric pathology. Hp0421, a cholesterol-α-glucosyltransferase enzyme, modulates this lipid raft signaling and phagocytosis (118). Cholesterol glycosylation by H. pylori delays its internalization by subverting the inflammatory response, modulating autophagy, and blocking phagosome maturation, enhancing its intracellular survival (119–121).

Cell death in response to infection.

Infection manifests a multitude of responses in host physiology, like immune activation, inflammatory responses, and cell death. A pathogen-triggered cell death program displays morphological similarities to apoptosis, pyroptosis, and autophagic cell death. While this topic is too vast to be detailed at length in this review, a recent review by Labbé and Saleh provides excellent coverage on the subject (122). The choice of the specific pathway for cell death depends on many factors, such as the nature of pathogen, the severity of infection, and the site of infection. For example, macrophages and dendritic cells die primarily of pyroptosis (122).

Shigella flexneri-induced macrophage cell death is a classic example of pyroptosis, which is independent of effector caspases and instead initiated by inflammatory caspases. Contrastingly, in epithelial cells, the Shigella secretory inhibitor, OspC3, inhibits caspase-4 to delay epithelial cell death (123). This positions Shigella to easily infect the nearby epithelial cells as it escapes from the cytosol of dying macrophages. Similarly, caspase-1-induced death has been reported in Salmonella Typhimurium, Listeria monocytogenes, Pseudomonas aeruginosa, and Yersinia pseudotuberculosis (122). Following recognition of PAMPs by cytosolic receptors like NLRs, canonical caspase-1 inflammasomes or noncanonical caspase-11 inflammasomes are activated, eventually triggering pyroptosis.

ONTOGENY OF MACROPHAGES AND ITS IMPACT

The pathophysiology of several diseases that arise from pathogenic invasion and inflammatory imbalance is due to an inadequate macrophage response. A better understanding of tissue-specific macrophage heterogeneity, diversity of macrophages’ transcriptional signatures, and macrophages’ consequential impact on the disease outcome is required for the development of novel macrophage-targeted therapies.

For several decades, the inception of a “mononuclear phagocytic system” proposed by Van Furth and colleagues and strengthened by Fogg and others has established a hierarchy, where promonocytes and their precursor cells populate bone marrow, monocytes populate peripheral blood, and macrophages populate tissues (124–126). The lacuna in our understanding was knowledge of the extent to which tissue macrophages are replenished by bone marrow-derived (BM) circulatory macrophages. Technical advances from pattern of surface markers to single-cell transcriptome sequencing (RNA-seq) have begun to bridge the gap in our understanding of macrophage ontology (127–129). Ontogeny of macrophages and the contribution of differentially originated macrophage in tissue macrophage populations and embryonic hematopoiesis are detailed by Ginhoux (130). The current view of the subject accepts the embryonic ontogeny of tissue macrophages. Hematopoietic waves during embryonic development seed the still-unknown progenitors to develop tissue-resident macrophage populations. These are maintained independently of BM-derived macrophage populations even in adulthood (130).

The precise mechanism of embryonic hematopoiesis and the impact of the physical heterogeneity of macrophages in normal and pathological conditions are not known. In addition, there are several other unresolved questions related to macrophage ontogeny: identification of fetal progenitors and their maturation pathway, distribution of erythromyeloid precursor (EMP) versus hematopoietic stem cell (HSC) originated macrophages, and contribution of early versus late HSCs. EMP-derived arterial macrophages are predominant, with their population peaking at adulthood and then declining with age, whereas the population of BM-derived macrophages peaks at adolescence and remains stable for the rest of life (131). Age-related alteration in cellular metabolism, immune status, endoplasmic reticulum stress, and mitochondrial dysfunctions could reasonably account for the effect of aging on EMP-originated arterial macrophages and contribute to diseases related to arterial homeostasis and hypertension (132).

The comparison of transcriptional profiles using genotoxic irradiation models reveals that BM-derived lung and peritoneal macrophages overlap 90% with embryonic macrophages but less than 50% with Kupffer cells (133). M. tuberculosis infection in ontogenically distinct macrophage populations responds diversely. Alveolar and interstitial macrophages and embryogenic and monocyte ontogeny show distinct replication, transcriptome profiles, and bacterial burdens on macrophage depletion (134). In a study on mannose receptor-expressing dermal macrophages infected with Leishmania, embryonic-derived macrophages were shown to be more permissive to growth and more responsible for disease severity than bone marrow-derived macrophages (135).

Thus, ontogeny-dependent macrophage diversity translates into functionally distinct responses. However, macrophages’ precise functional consequence on the inflammatory balance, homeostasis, or repair in normal or infected conditions is not known. This understanding will help in the development of novel macrophage-targeted intervention strategies in pathological conditions.

CONCLUSION

The winner of pathogen-host competitive interactions for clinical manifestation of pathology versus pathogen clearance is still disputed (Fig. 1). Limiting the pathogen within the macrophage itself is a primary success of the immune system, as it seizes the opportunity to thrive and suppresses the clinical manifestation of the disease. Ineffective clearance of pathogenic bacteria, e.g., M. tuberculosis, from macrophages reflects that macrophages can be potential carriers and an infinite supply of new infection cycle upon reactivation, making it a successful situation for the pathogen. Further, screening the susceptible population for latent infections and improving the immune status of the risk group can dwindle the pathogen’s success. Future research in this field should be oriented to develop newer molecular tools to strengthen and support innate immune mechanisms and neutralize the pathogenic defenses. With the emergence of new infections and new drug resistance, improved understanding of host-pathogen interaction should be the need of the hour for developing novel therapeutic intervention strategies.

Biographies

Saurabh Pandey completed his bachelor’s degree in Applied Sciences. He then did his master’s degree and Ph.D. in Biochemistry from University of Hyderabad. His research work sprawls around infection biology with emphasis on tuberculosis. He moved to University of Pennsylvania for his postdoctoral work, where the major area of research was tumor virology and gastric cancer. In 2018, Dr. Pandey joined Jamia Hamdard as an Assistant Professor in the Department of Biochemistry. His group in Jamia Hamdard is investigating the role of chaperone proteins in virulence of Mycobacterium tuberculosis.

Sashi Kant earned his Bachelor of Science and Master of Science in Biotechnology from India. He received his Ph.D. at the University of Delhi, India. He was then trained as postdoctoral researcher at Ohio State University College of Medicine, USA. In his postdoctoral work, the major emphasis of his work was to understand the molecular basis of Gram-positive bacterial pathogenesis especially related to Streptococcus pyogenes, Streptococcus pneumoniae, and Staphylococcus aureus (including multidrug-resistant). In 2019, Dr. Kant moved to the University of Colorado-Anschutz School of Medicine, USA as Research Instructor. He is working on the long-standing question of molecular mechanisms by which Salmonella avoids ROS and NOS encounter and protects itself against oxidative and nitrosative stress. He is also involved in investigating the adaptations and defense systems of Salmonella in order to develop therapeutic approaches.

Deeksha Tripathi completed her bachelor’s and master’s degrees in Microbiology from University of Delhi, India. She moved to Jawaharlal Nehru University, New Delhi for her Ph.D. Her research revolves around the virulence attributes of tuberculosis pathogen, Mycobacterium tuberculosis. In her postdoctoral work at Indian Institute of Technology, Delhi, she investigated the role of peptidyl prolyl isomerases in immunomodulation and biofilm formation of the pathogen. She received AMI Young Scientist Award in 2015 for her contribution in the field of Medical Microbiology. She joined Central University of Rajasthan as Assistant Professor in 2016, where she is actively engaged in teaching and research. She has published her research work in journals of high repute.