The Cell with Many Faces: Lung Macrophage Plasticity and Function in Response to Environmental and Pathogenic Insults

Abstract

Alveolar macrophages (AM) are pivotal immune sentinels, essential for maintaining tissue homeostasis and mediating immune responses to inhaled particles and pathogens. They demonstrate remarkable plasticity by transitioning from pro-inflammatory (M1) and anti-inflammatory/reparative (M2) phenotypes in response to local signals. Upon exposure to environmental agents, such as particulate matter, atypical respiratory pathogens opportunistic gram-negative bacteria, or respiratory viruses they undergo dynamic activation that profoundly influences their functional repertoire. Acute or chronic environmental/biological insults disrupt normal AM activities such as phagocytosis, efferocytosis, cytokine production, inciting oxidative stress, inflammasome activation, and in some cases, forms of programmed cell death such as pyroptosis. Although these responses are indispensable for eliminating noxious particles and pathogens, such as Mycoplasma pneumoniae or Klebsiella pneumoniae, Influenza A, or SARS-CoV-2, they can also derail the resolution phase by perpetuating inflammation, driving tissue remodeling and fibrosis, and thereby fueling chronic lung disorders such as chronic obstructive pulmonary disease (COPD), pneumoconiosis, and post-COVID interstitial lung disease. Moreover, environmental and microbial exposures modify AM by altering receptor repertoires, intracellular phenotype by signaling cascades, and crosstalk with epithelial and mesenchymal cells that collectively determine the disease trajectory. Elucidating how diverse environmental agents, together with pathogens such as Mycoplasma pneumoniae, Klebsiella pneumoniae, Influenza A, and SARS-CoV-2, shape AM biology is therefore pivotal for understanding the pathogenesis of COPD, pneumoconiosis, and progressive fibrotic lung disease, and COVID-19 related pulmonary sequelae. This review brings together the current insights into exposure-driven modulation of AM functions, highlighting recent advances and identifying knowledge gaps relevant for therapeutic targeting of exposure-induced and pathogen-mediated lung pathology.

Graphical Abstract:

Graphical illustration of the acute, resolution, and chronic phases of inflammation orchestrated by AM in response to environmental toxicants and pathogens (Klebsiella pneumoniae, Mycoplasma, Influenza, SARS-CoV-2). (A) Acute phase: Environmental toxicants (e.g., cigarette smoke, PM silica, asbestos, or pathogens are phagocytosed and by AM. They may also damage the lung cells by generating reactive oxygen species (ROS). SP-A and SP-D, complement protein C1q, C3b, and/or immunoglobins, opsonize toxic particles and enhance phagocytosis by being recognized by mannose (MR)/TLR2, compliment CCR1/CCR3, and FcR receptors respectively expressed on AM. M1 macrophages secrete an array of inflammatory cytokines and chemokines, which leads to the recruitment of other immune cells including neutrophils and BMDMs to the lung and exacerbate the inflammatory condition. (B) Resolution: Effective clearance of apoptotic or dead cells by AM through efferocytosis as well as secretion of IL-4 and IL-13 by ILC2 cells polarize the macrophages towards M2 state. M2 macrophages secrete anti-inflammatory cytokines such as IL-10 and TGF-β and mark the resolution phase. (C) Chronic phase: Continuous inhalation of toxic particles (e.g., cigarette smoke, PM silica, asbestos, ozone, or mycoplasma) or defective efferocytosis leads to the secretion of various chemotactic factors by AM, which leads to accumulation of other inflammatory immune cells in the lung. CC-chemokine ligand 2 (CCL2) attracts monocytes via a number of receptors, which engage T helper (T_H) cells and cytotoxic CD8 T cells. These cells release proteases including MMP9 and elastase, which leads to the elastin degradation, damage of the alveolar wall, and emphysema. Additionally, AM and epithelial cells releases TGF-β which induces proliferation and activation of fibroblast and myofibroblast resulting in tissue remodeling and fibrosis. Figure created with a licensed version of Biorender.com

Introduction

Inhaled particles, airborne pathogens, and pollutants constantly bombard the lungs, with their large surface area and constant interaction with the external environment. Lung immunity is orchestrated by both innate and adaptive immune cells that reside in or are recruited to the airways, interstitial, and alveolar spaces. To maintain effective gas exchange while defending against inhaled threats, the lungs maintain a dynamic array of immune cells, which cooperate to maintain homeostasis. These cells are responsible for immune surveillance, rapid defense against pathogens, and tolerance to harmless antigens.(1, 2) The tight regulation of these cellular networks is crucial for cellular function, and their disruptions can lead to infections, allergies, or chronic inflammatory diseases.(1) Acute lung injury (ALI) and its severe form, acute respiratory distress syndrome (ARDS), involve complex interactions of various immune and structural cells. Stimuli from both infectious (e.g., bacterial or viral pneumonia) and non-infectious (e.g., chemical exposure, mechanical ventilation) (3) agents trigger robust immune responses that can be either protective or damaging.

Among the initial responders to lung injury are the highly adaptable macrophages, which are the most numerous immune cells in the lungs. Their ability to detect, react, and adjust to a wide range of harmful stimuli makes them key contributors not only to defend the lungs from external particulate matter and pathogens, but also to initiate the repair. The ability of these cells to shift between pro-inflammatory and anti-inflammatory roles is a key mediator in the aggravation and alleviation of lung injury.(4) Understanding their dual nature is crucial to unravel the complex pathogenesis of acute and chronic lung diseases.

Unlike traditional views that all macrophages derive from circulating monocytes, recent advances in fate-mapping and cellular lineage tracing have revealed that lung macrophages have diverse embryonic and postnatal origins, which influence their identity, maintenance, and function throughout life. In the healthy lung, there are two primary macrophage populations: alveolar macrophages (AM), which reside on the epithelial fluid layers, and interstitial macrophages (IM), located within the lung parenchyma.(5, 6) AM originate mainly from fetal monocytes that colonize the lungs during embryogenesis and then maintain their numbers through self-renewal, with little need for replenishment from bone marrow-derived cells. In steady-state conditions, these macrophages are the primary immune cells residing on the alveolar surface, where they perform specialized roles essential for maintaining lung homeostasis. In contrast, IM are embedded within the connective tissue of the lung parenchyma. They arise from a mixture of both embryonic precursors and adult blood monocytes. Nestled among structural and immune cells, IM guide tissue remodeling, temper immune reactions, and help preserve lung architecture. During inflammation or injury, circulating monocytes are rapidly recruited into the lungs by chemokine gradients, and differentiate into macrophages. These newly recruited macrophages augment the resident pool and support the immune response by aiding in pathogen elimination, tissue repair, and regulation of inflammation.(1, 7, 8) Recent advances have revealed a spectrum of macrophage subsets in the lungs, distinguished by their ontogeny and local niche. The lung, therefore, serves as a paradigm for understanding how tissue context endows macrophages with specialized identities.

The mammalian lungs are exposed to a million of particles and a plethora of reactive gases in daily life. Macrophages, strategically positioned at the air-tissue interface, clear debris, orchestrate immune responses, and maintain tolerance to innocuous antigens. Perturbation of macrophage homeostasis by environmental agents is increasingly recognized as a pivotal driver of chronic lung diseases such as COPD, asthma, pulmonary fibrosis, cancer, and others.(9–11) Understanding exposure-specific alterations in macrophages is crucial for gaining mechanistic insights, identifying biomarkers, and developing targeted therapies. Herein, we aim to discuss the origin, phenotypic diversity, and functional dynamics of AM, with a particular focus on how environmental/biological agents modulate their responses and contribute to pulmonary health and disease.

AM display remarkable phenotypic plasticity, allowing them to adjust rapidly to the dynamic lung microenvironment. They are the multitasking conductors of lung inflammation that exhibit extraordinary functional plasticity through complex networks of cytokines, pathogen recognition receptors (PRRs), and epigenetic modifications. Key functions include: (i) Particle recognition, (ii) Phagocytosis of opsonized pathogens and trafficking them into intracellular compartments for killing; (iii) Formation of extracellular traps (METs); (iv) Clearance of apoptotic cells by efferocytosis; (v) Release of reactive oxygen-nitrogen species (ROS/RNS), proteases, cytokines, and chemokines; (vi) Tissue repair and remodeling by phagocytosing pathogens, inhaled particles. Below, we discuss the main polarization pathways of AM, the key molecular signals that regulate these states, and the characteristic surface markers and functional traits that define each phenotype as well as metabolic shifts which play a crucial role in their maintenance of pro-inflammatory (M1) and tissue repair phenotypes (M2) (Table 1, Figure 1).

Table1.

Main categories of alveolar macrophages (AM) encountered in adult lungs, their origin and ontogeny, and their distinguishing molecular/functional traits.(8, 20, 80, 173, 659–668)

Alveolar-Macrophage Type / Phenotype	Typical Inducer or Ontogeny	Core Surface Markers (human)	Signature Transcription Factors	Key Effector Molecules which are upregulated	Dominant Functions	Resulting Lung Injury
Homeostatic Resident AM (quiescent)	Prenatal yolk-sac & fetal-liver origin; GM-CSF & surfactant cues	CD11c/HIGH (Itgax+), MARCO, CD169, Siglec-8 (Siglecf+), MerTK, CD11b/LOW, Pparg+, Car4+, TRPV4 (LOW/constitutive)	PPARγ, KLF4, BACH2	TGF-β (basal), GM-CSF, SP-A/D, cathepsins (low), C1qa/b/c+	Surfactant turnover, debris clearance, immune surveillance	Healthy lung; immune silence to innocuous inhaled particles
Inflammatory “M1-like” AM	Classical PRR ligation (LPS, TLR2/4 agonists, silica, cigarette smoke, Mycoplasma lipoproteins)	CD80, CD86, MHC-II-HIGH, CD11b↑, TLR2/4, MARCO, CCR7, C5ar1+, Ear1+, TRPV4 (↑/activated)	NF-κβ, AP-1, IRF1, HIF-1α	TNF-α, IL-1β, IL-6, CXCL8, iNOS/NO, NOX2-ROS, caspase-1, Krat79, Mki67+, Top2a+	ROS/RNS burst, pathogen killing, neutrophil recruitment	Acute infections (Mp, influenza), particulate exposure, early COPD exacerbation
Alternative “M2-like” AM	Th2 cytokines (IL-4, IL-13), IL-10, TGF-β, prolonged particle exposure, convalescent Mp infection	CD206 (MRC1), CD163, MerTK-HIGH, PD-L1, CCR2-LOW, Chil3+, Pf4+, TRPV4 (variable/linked to profibrotic signaling)	STAT6, STAT3, PPARγ, SMAD2/3	Arginase-1, IL-10, TGF-β, MMP-9/12, PDGF, VEGF, Chil3 (YM1)	Wound healing, collagen synthesis, phagocytosis of apoptotic cells	Allergic asthma, fibrosis (silicosis, asbestosis), late Mp pneumonia
Regulatory / IL-10 AM (Mreg)	Chronic low-grade bacterial ligands, apoptotic-cell uptake, glucocorticoids	CD11c, CD14, IL-10R, MerTK, low CD80/CD86, C1qa/b/c+, SinglecF+, TRPV(LOW/regulated)	STAT3, CREB	IL-10, IL-1Ra, PGE2	Resolution of inflammation, T-cell suppression	Post-viral or post-Mp convalescence; corticosteroid therapy
Monocyte-Derived AM (Mo-AM)	Recruitment of classical monocytes (CCR2+) during injury or infection	CD11b/HIGH, CD14, Lysozyme, CCR2+, Ly6c2+, Ly6C+, low Siglec-8 (Siglecf variable), Ccr2+, TRPV4 (HIGH/functional)	IRF8, RUNX1	Pro-IL-1β, TNF-α, cathepsin B/L, MMP-8, Ear1+, Pf4+, Mki67+, Top2a+	First-wave defense, debris removal; can replace resident AM niche	ARDS, severe pneumonia, smoke-induced lung injury
Senescent / Aged AM	Aging, chronic oxidative stress (smoke, PM2.5)	CD11cLOW, SA-β-gal+, reduced Siglec-8, SinglecF variable, TRPV4(dysregulated)	FOXO3, p53, decreased NRF2	SASP factors: IL-6, CCL2, MMP-3	Low phagocytosis, high inflammatory “noise”	Age-related decline, susceptibility to infection, emphysema
Foam-Cell / Lipid-laden AM	Surfactant overload, e-cigarette, alveolar proteinosis	CD11c, ADRP, TREM2, CD36, PLIN2, TRPV4(variable; may affect lipid handling)	LXR-α, SREBP-1c	Cholesteryl-esters, oxidized lipids, GM-CSFLOW	Lipid storage, impaired surfactant clearance	Exogenous lipoid pneumonia, smoke-associated lung injury
Trained-Immunity (Epigenetically Scared) AM	Prior smoke/PM exposure or Mp infection; persistent β-glucan	CD11c, MARCO, epigenetic markers (H3K4me3 at TNF/IL6), Krat79, TRPV4(sensitized)	RelA (accessible), loss of HDAC2	TNF-α, IL-6 (heightened on restimulation), Mki67+, Top2a+	Exaggerated secondary responses, steroid resistance	Former smokers, recurrent Mp-associated wheeze, rapid-decline COPD
Hemosiderin-Laden Macrophages	Alveolar hemorrhage, diffuse alveolar hemorrhage (e.g. Goodpasture’s)	CD68, CD163, Prussian blue+ iron deposits, TRPV4 (variable)	NRF2, HO-1	Ferritin, heme oxygenase-1 (HO-1), iron-binding proteins	RBC phagocytosis, iron recycling, antioxidant defense	Pulmonary hemorrhage syndromes, idiopathic pulmonary hemosiderosis
Multinucleated Giant Cells	Chronic granulomatous stimuli (TB, sarcoidosis, foreign bodies—silica/coal dust)	CD68+, CD11b, MHC-II, DC-SIGN, TRPV4(present in fused macrophages)	NFATc1, STAT6, NF-κβ	IFN-γ–inducible genes, TNF-α, MMP-9	Granuloma formation, walling-off pathogens/particles	Sarcoidosis, tuberculosis, silicosis, coal-workers’ pneumoconiosis

Figure 1: Graphical representation of molecular and cellular pathways involved in macrophage activation and polarization in response to environmental exposure.

(A) Homeostasis: tissue resident alveolar macrophages (TR-AM) in healthy lung interact with alveolar epithelial cells (AEC) (ATI and ATII) via CD200R-CD200 receptors. Secretion of granulocyte–macrophage colony-stimulating factor (GM-CSF) by AEC, transforming growth factor-β (TGF-β) by TR-AM and inhibitory signal via CD200R-CD200 orchestrate the immunosuppressive activity of the macrophages. AM maintain their population by self-renewal capabilities and expressing proliferation markers Mki67 and Top2a. During homeostasis TR-AM, rely on oxidative phosphorylation (OXPHOS) for their metabolic needs. (B) M1 polarization: Environmental toxicants generate reactive oxygen species (ROS) that triggers the release of damage associated molecular patterns (DAMPs) in the extracellular and intracellular environments. DAMPs activate pattern recognition receptors such as TLR (toll like receptors), NLRP3 inflammasome, and cGAS-STING pathways to stimulate macrophages and the release of pro-inflammatory mediators, which recruit neutrophils, and bone marrow derived macrophages to the lungs. Cytokines/chemokines further stimulate macrophages via interleukin and TNF receptors. Infiltrated macrophages also secrete pro-inflammatory mediators, thus establishing a feed forward loop of inflammation. M1 macrophages mainly utilize glycolysis for their energy demands. (C) M2 Polarization: Chronic exposure to environmental toxicants promotes increased levels of IL-13 and IL-4 secreted by T-helper type (Th2). These inhibitory cytokines activate lung macrophages via IL-13R1α and IL-4Rα receptors and skew them towards M2 polarization. M2 macrophages regulate inflammation by enhancing expression of anti-inflammatory molecules including CD200R and TREM2 and secretion of anti-inflammatory cytokines including IL-10 and TGF-β. αvβ6 integrin expressed on inflamed AEC type I cells bind to TGFβR expressed on AM and inhibit inflammation.(145, 676) Chronic M2 activation of macrophages and dysregulated TGF- β signaling causes proliferation and activation of fibroblasts and myofibroblasts, which results in fibrosis. Figure created with a licensed version of Biorender.com.

Stages of Alveolar Macrophage Activations

Classically Activated AM (M1 Macrophages)

In the absence of external stimuli, AM exist in an immune-quiescent, “tolerogenic” phenotype that protects the delicate gas-exchange surface from any undue inflammation. However, upon exposure to potent stimuli, such as bacterial lipopolysaccharide, viral components, and Th1 cytokines like interferon- γ (IFN- γ) and others, AM are rapidly reprogrammed into the classically activated or M1 state.(12, 13) This activation does not represent a fixed state but rather a signal-dependent activation program, that optimizes the cell for rapid, aggressive innate immune responses, aimed at pathogen clearance. The shift toward M1 is marked by changes in gene expression, surface receptor expression, and cellular behavior that together prioritize inflammation and antimicrobial activity over tissue repair.(8, 12, 14)

On a molecular level, classical activation is driven by signaling pathways initiated by cytokine receptors and pattern-recognition receptors. Exposure to a variety of external stimuli result in the release of Damage Associated Molecular Patterns (DAMPs), which activate pattern recognition receptors such as toll-like receptors (TLR), NLRP3 inflammasome, and cGAS-STING pathways to stimulate macrophages and release of pro-inflammatory mediators (cytokine/chemokines). Chemokines recruit other immune cells including neutrophils and bone marrow-derived macrophages. Cytokines further stimulate macrophages via interleukin receptors and TNF receptors.(15–19) (Figure 1)

Infiltrated macrophages also secrete proinflammatory mediators, thus establishing a feed forward loop of inflammation. This is further exacerbated by growth factors, notably granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF),(20) which engage their respective receptors and signal through JAK2 to enhance macrophage survival and differentiation. Converging signals from pattern-recognition (PRRs) and cytokine receptors activates three principal signaling cascades, JAK/STAT, NF-κβ, and IRF. All three cooperate at the chromatin level to drive a robust transcriptional program encoding iNOS, NADPH oxidase, and inflammatory chemokines and cytokines.(21) (Figure 2)

Figure 2: Generation of Superoxide (·O₂⁻), Nitric Oxide (NO·) and peroxynitrite (ONOO⁻) by AM:

(A) NADPH oxidase generates superoxide and nitric oxide synthase (iNOS) produces NO by metabolizing L-arginine. ·O₂⁻ is also generated through an electron transport chain in mitochondria. ·O₂⁻ and NO· combine to form the highly reactive ONOO⁻. ·O₂⁻ is converted into hydrogen peroxide, which reacts with chlorine (Cl−) to form hypochlorous acid. NO· diffuses through cell membranes while ·O₂⁻ and ONOO⁻ can cross through anion channels. NO· and ONOO⁻ inhibit Na⁺ (ENAC) and Cl⁻ (CFTR) channels on epithelial cells.(677–680) NO· and ONOO⁻ reduce the pathogen load ONOO⁻ causes lipid peroxidation and damage surfactant components in the alveolar space and also increases permeability of the blood-gas barrier.(434, 681–683) (B) Immunostaining of nitric oxide synthase (iNOS) in human AM from bronchoalveolar lavage fluid of patients with acute respiratory distress syndrome (ARDS) shows increased expression of iNOS at day 7 of ARDS pathogenesis. Panel B was reprinted from Sittipunt C, et al with permission.(684) Figure created with a licensed version of Biorender.com.

The JAK/STAT axis is initiated when binding of its heterodimeric receptor (IFN-γR1/IFN-γR2), bringing receptor-associated JAK1 and JAK2 into proximity and facilitating their trans-phosphorylation.(22, 23) Activated JAKs phosphorylate STAT1, prompting STAT1 dimerization and nuclear translocation. In the nucleus, STAT1 dimers bind to gamma-activated sequence (GAS) elements in promoters and enhancers of target genes such as Nos2 (iNOS), and TNF.(24, 25) STAT1 also induces the transcription factor IRF1, creating a feed-forward loop that maintains high STAT1 activity and further amplifies inflammatory gene expression.(26–29)

Simultaneously, engagement of TLR4 by Lipopolysaccharide (LPS), or TNFR1 and IL-1R1 signaling via autocrine TNF-α and IL-1β activates the MyD88/IRAK4/IRAK1–TAK1 cascade, culminating in IKKβ-mediated liberation of the NF-κβ p65/p50 heterodimer.(25, 30, 31) NF-κβ then drives expression of Il1b, Il6, TNF-α, Nos2, and NADPH-oxidase subunits, thereby fueling inflammatory gene transcription.

Activation of TLR3 and the TLR4–TRIF axis recruits TRAF3, activating TBK1 and IKKε, which phosphorylates IRF3 and IRF7, inducing their dimerization and nuclear translocation, initiating IFN-β transcription.(32–34) The secreted IFN-β then binds the type I interferon receptor (IFNAR1/2), triggering JAK1 and TYK2 to phosphorylate STAT1/STAT2, which together with IRF9 form the ISGF3 complex that binds interferon-stimulated response elements (ISREs) in promoters of Nos2 and NADPH oxidase. Meanwhile, IRF5, induced by both STAT1 and NF-κβ, binds to distinct promoter motifs in cytokine genes (e.g., Il12, Il6), thereby reinforcing the M1 phenotype. Cooperation among STAT1, NF-κβ, IRF1/5, and AP-1 (activated via MAPKs p38, JNK, ERK) assembles super-enhancers at key loci, recruit’s chromatin remodelers, and drives burst transcription of iNOS, and inflammatory cytokines.(33, 35–39)

In addition, mitochondrial or nuclear DNA released from stressed cells activates cyclic GMP-AMP synthase (cGAS), which generates cGAMP to engage the ER-resident adaptor STING.(40–42) STING couples to TBK1 for IRF3 phosphorylation and to IKK for additional NF-κβ activation, synergizing with surface TLR signals to solidify the M1 transcriptome and promote production of CXCL9/CXCL10.(43, 44) Simultaneously, extracellular ATP and UTP released from injured epithelial cells bind to the purinergic receptors, triggering potassium (K+) efflux, mitochondrial ROS generation (Figure 3), and lysosomal disruption, which are the classic cues for assembling the NLRP3 inflammasome. NF-κβ–mediated “priming” supplies the pro-IL-1β and NLRP3 protein, while the P2X7/ROS axis provides the “activation” signal; together they allow the NLRP3-ASC-caspase-1 complex to mature IL-1β and IL-18 and to cleave gasdermin D, driving pyroptosis.(45–49) The released cytokines feedback on AM and neighboring leukocytes, intensifying STAT1 and NF-κβ activity and locking the AM in the M1 state.

Figure 3: Reactive oxygen species (ROS) generation and their effect on AM activation during exposure of lungs to environmental toxicants and pathogens:

(A) Environmental toxicants such as particulate matter, silica, asbestos, and pathogens are phagocytosed by AM, leading to autophagolysosomal formation and generation of cytosolic ROS. Ozone and other environmental agents cause epithelial cell damage by oxidation of phospholipids (OxPL) and lipid peroxidation of the components of surfactants (LPP). These active byproducts cause the production of DAMPs, which lead to the activation of AM. (B) Cytosolic ROS also produced through NOX (NADPH oxidase) activate macrophages via NF-Kβ and HIF-1α pathways towards a pro-inflammatory phenotype and enhances glycolytic pathways. Cellular stress and altered activity of enzymes including electron transport chain (ETC) trigger the generation of mitochondrial ROS (mtROS). mtROS damages mitochondrial DNA which forms DAMPs that activate the NLRP3 inflammasome leading to the further activation of the macrophages.(685) Figure created with a licensed version of Biorender.com.

Metabolically, these signaling events trigger a shift toward aerobic glycolysis, which supplies rapid ATP and biosynthetic intermediates while generating lactate. This metabolic reprogramming also enhances production of ROS and reactive nitrogen species (RNS); NADPH derived from the oxidative state of the pentose phosphate pathway fuels the NOX2 complex to generate superoxide, while iNOS converts L-arginine to nitric oxide. Superoxide and NO combine to form peroxynitrite, a potent microbicidal molecule, and elevated ROS can oxidize phosphatase cysteines to prolong kinase signaling, creating feed-forward amplification loops.(14, 50)

Phenotypically, M1 polarized AM secrete pro-inflammatory cytokines (TNF-α, IL-1β, IL-6, IL-12, IL-23) and chemokines (CXCL8/IL-8, CXCL9-11) that recruit neutrophils, monocytes, lymphocytes, and eosinophils, which propel a Th1-dominated immune response.(51) M1 polarized AM are responsible for the rapid pathogen clearance during early lung infections (bacterial and viral), efficient antigen presentation, and orchestration of innate and adaptive immune responses. However, sustained or dysregulated M1 polarization often driven by chronic exposure to microbial products, or endogenous danger signals can lead to persistent inflammation and tissue injury.(12, 52–54) Thus, anti-inflammatory cytokines, inhibitory receptors,(55–58) and metabolic feedback help to switch/reprogram M1 AM to the M2 state.

Alternatively Activated AM (M2 Macrophages)

M2 AM represent an alternatively activated state dedicated to dampening inflammation and promoting tissue repair in the lungs. This phenotype is primarily induced by Th2-type cytokines, such as interleukin-4 (IL-4), interleukin-13 (IL-13), interleukin-10 (IL-10), and TGFβ, which trigger the STAT6 transcriptional pathway after binding to the shared IL-4Rα receptor(20, 59–62) (Figure 1). M2-AM contain a distinctive constellation of markers, including arginase-1 (Arg1), scavenger receptor-A, CCR2, and others which help them suppress the inflammatory responses and regulate the immune balance.(63)

When macrophages adopt the M2 phenotype, they switch from glycolysis to fatty-acid oxidation and oxidative phosphorylation to generate the energy and biosynthetic building blocks essential for tissue repair. (Figure 1) Additionally, anti-inflammatory signals such as IL-10, TGF-β, and glucocorticoids reinforce M2 by creating specialized functional subtypes (M2a, M2b, M2c and M2d) that help resolve distinct aspects of inflammation and promote tissue remodeling.(63–66) All four M2 subtypes share common hallmark features, such as high Arg1 activity, enhanced scavenger-receptor expression, and dampened production of classical pro-inflammatory cytokines. However, each M2 subtype is induced by a unique set of signals. IL-4 and IL-13 primarily induce M2a; immune complexes together with TLR ligation produce M2b; prolonged exposure to IL-10, TGF-β, glucocorticoids, or uptake of apoptotic cells gives rise to M2c; and adenosine, IL-6, hypoxia, or tumor-derived factors result in the formation of the M2d phenotype. These upstream signals sculpt distinct transcriptional profiles, surface-marker repertoires (e.g., CD206 high in M2a, CD163 high in M2c) and metabolic settings, giving each subtype a specialized toolkit for dealing with different stages of tissue injury.(59, 67, 68)

Functionally, M2a macrophages are the archetypal “wound-healing” cells. They secrete pro-fibrotic mediators such as fibronectin, insulin-like growth factor-1, and TGF-β, which up-regulate enzymes that remodel the lung extracellular matrix. M2b AM, by contrast, occupy a hybrid regulatory niche, releasing both IL-10 and moderate amounts of TNF-α and IL-6, which allow them to fine-tune lingering inflammation, while still providing antimicrobial support. M2c AM express high levels of MerTK and CD163, engulf apoptotic cells, and secrete IL-10 and TGF-β, which terminate effector T-cell responses.(12, 51, 59, 66, 67, 69–71) M2d cells express abundant vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), and matrix metalloproteinase-9 (MMP9), thereby fostering neovascularization that supplies oxygen to regenerating tissue (Figure 1).

Recognizing the specific signals that generate each subtype provides valuable therapeutic entry points: augmenting M2a and M2c responses accelerate wound healing or temper autoimmunity, whereas selectively inhibiting M2d polarization is a growing strategy for t cancer immunotherapies, which enhance T-cell responses.(12, 51, 59, 67, 69, 70, 72, 73) Metabolically, M2 AM rely mainly on fatty-acid oxidation and oxidative phosphorylation, a shift supported by PGC-1β-induced mitochondrial biogenesis.(74, 75) (Figure 4) This profile differs sharply from the glycolysis-driven, ROS-producing metabolic characteristic of classically activated M1 AM. M1 responses depend on the rapid and transient transcription regulated by NF-κβ and STAT1, whereas M2 AM responses develop more gradually but persist longer, an adaptation that fits with the lung’s need for ongoing repair and remodeling once pathogens have been eliminated. Therefore, achieving the proper timing and balance between M1 and M2 states is crucial for maintaining homeostasis. M2-polarized AM drive the resolution phase of inflammation and rebuild tissue architecture after injury or infection. Their proficiency in efferocytosis, swiftly removing apoptotic neutrophils and cellular debris, prevents the secondary release of harmful intracellular contents and supports orderly healing. (13, 76–79)

Figure 4: Metabolic rewiring of macrophage activation and function:

(A) AM in a homoeostatic state mainly depends upon oxidative phosphorylation (OXPHOS) for their metabolic needs. (B) Following exposure of AM to DAMPs or PAMPS, the majority of their pyruvate is converted into lactate. As a result, the TCA cycle is inhibited which leads to the accumulation of citrate and succinate. Citrate facilitates fatty acids synthesis (FAS) and succinate promotes stabilization of HIF-1α; both then enter the nucleus and stimulate the expression of glycolytic genes and proinflammatory mediators. This also leads to decreased OXPHOS and increased ROS generation, which drives AM toward the M1 pro-inflammatory phenotype. Enhanced pentose phosphate pathway (PPP) results into increased NADPH leading to the nitric oxide (NO·) production, which further inhibits OXPHOS. (C) Following stimulation by IL-4 and IL-13, AM pyruvate is converted into acetyl-CoA and enters the TCA cycle, which results in increased OXPHOS. Enhanced fatty acid oxidation (FAO) also upregulate OXPHOS leading to increased arginase1 (ARG1) and nuclear translocation of PPARγ/STAT6 which promotes tissue repair.(687, 688) Figure created with a licensed version of Biorender.com.

In the acute and resolving stages of pneumonia, ARDS, and asthma, M2 activation accelerates healing and restores effective gas exchange. Conversely, excessive or chronic M2 skewing may lead to idiopathic pulmonary fibrosis or post-viral interstitial lung disease, by stimulating excessive fibroblast activity and collagen deposition.(80) (Figure 5) Therefore, tight regulation of M2 AM polarization is critical for achieving a balance between effective repair and the prevention of maladaptive scarring in the lung. Under normal circumstances, once the initial injurious stimulus is cleared, a series of built-in “brakes” engage to prevent M2 cells from driving unchecked fibrosis. These include intracellular signaling inhibitors such as suppressor of cytokine signaling proteins (SOCS1 and SOCS3) and protein tyrosine phosphatases (e.g., SHP-1), which attenuate IL-4/IL-13 receptor-STAT6 activation.

Figure 5: Mechanism of macrophage activation and function in tissue repair and regeneration:

IL-4 and IL-13 released from innate lymphoid cells type II (ILC2) and helper T cells (Th1) activates both BMDM and resident AM into a regulatory and pro-resolving phenotype, which clear the apoptotic cells by efferocytosis. AM secrete AREG, PDGF, TGF-α, Plet1, and VEGF-α which promote repair and regeneration of epithelial and endothelial cells and reconstitution of the blood-gas barrier. In the later phase of repair, BMDM and resident AM acquire regulatory phenotype, self-proliferation, and maintain the normal tissue architecture and homeostasis. Chronic activation of M2-like macrophages causes activation and proliferation of fibroblasts and myofibroblast via TGF-β signaling leading to the deposition of an extracellular matrix (ECM) and fibrosis. TGF-β stimulates recruitment of the TRPV4 complex from the cytoplasm to the membrane of fibroblast which leads to the increased Ca²⁺ influx and promotes myofibroblast trans-differentiation.(668) Amphiregulin (AREG), Arginase-1 (Arg1), interferon regulatory factor 4 (IRF4), peroxisome proliferator-activated receptor gamma (PPARγ), Vascular Endothelial Growth Factor (VEGF), placenta-expressed transcript 1 (Plet1) Transient Receptor Potential Vanilloid 4 (TRPV4).(688) Figure created with a licensed version of Biorender.com.

Likewise, nuclear factors such as PPARγ and BACH1 compete with pro-fibrotic transcriptional programs, modulating excessive collagen and growth-factor production.(81–83) At the post-transcriptional level, specific microRNAs (e.g., miR-155, miR-21, and miR-34a) dampen M2-associated transcripts or promote the expression of negative regulators, effectively shortening the half-life of profibrotic mRNAs and resetting M2 cells toward homeostasis.(31, 84, 85) Metabolically, a switch away from fatty-acid oxidation, driven by AMPK activation or shifts in NAD+/NADH ratios, limit the bioenergetic support that M2 macrophages require for persistent matrix remodeling. Pro-resolving lipid mediators (such as resolvins, lipoxins, protectins), produced by neutrophils and platelets, restore phagocytic clearance functions and bind to specific G-protein coupled receptors on M2 macrophages, triggering apoptosis or re-polarization back toward a quiescent phenotype.(86–89)

Finally, crosstalk with neighboring cells provides another important layer of control. Alveolar type II (ATII) epithelial cells secrete GM-CSF and surfactant proteins SP-A and SP-D that favor balanced macrophage activation; regulatory T cells supply IL-10 or TGF-β to dampen but not abolish repair functions. Mechanical cues, such as increasing tissue stiffness during late-phase repair, feedback through integrin, and focal-adhesion kinase pathways further curtail M2-driven fibrosis. Together, these intersecting checkpoints ensure that once damage is mended, AM return to surveillance mode rather than perpetuating a pro-fibrotic cycle.(14, 67, 90–93)

M3 or Regulatory Alveolar Macrophages (Mres)

As stated above, macrophage polarization is traditionally recognized as consisting of M1, classically activated, pro-inflammatory, and M2, alternatively activated, reparative. However, accumulating evidence reveals an additional, functionally distinct population that emerges as inflammation subsides, commonly termed M3 or Mres.(94, 95) These AM represent a specialized activation state that emerges once the initial microbial or inflammatory insult are cleared. Metabolically, M3 macrophages favor high-efficiency fatty-acid oxidation and mitochondrial oxidative phosphorylation, sustained by AMPK-driven autophagy(96) and PGC-1β–mediated mitochondrial biogenesis,(97) generating ample NADPH for redox balance, without the ROS burst characteristic of M1 cells.

Functionally, M3 macrophages serve as the “off-switch” for inflammation and are the first architects of tissue restoration. However, they are molecularly distinct from conventional M2 AM. M3 AM contain Tim-4, MerTK, Stabilin-2, and CD36, four receptors that together endow them with superior efferocytosis, while they express little or no CD206 (mannose receptor), CD209, or CD301 that are present in M2a/b macrophages.(94, 98–101)Transcriptome analyses show the dominance of resolution-linked transcription factors, LXRα/β, NR4A1 (Nur77), ATF3, and KLF2; whereas the IL-4/IL-13-driven STAT6-IRF4 axis that defines M2 polarization is markedly muted.(102–107) This unique wiring drives a metabolic profile rich in cholesterol efflux genes (ABCA1, ABCG1) and oxidative phosphorylation enzymes, contrasting with the fatty-acid oxidation bias of classic M2 cells.(108–110)

These events translate into a specialized functional portfolio. By clustering Tim-4, MerTK, and Stabilin-2 at the membrane, M3 macrophages execute rapid, high-capacity efferocytosis, preventing secondary necrosis and the spill-over of DAMPs that would otherwise reignite inflammation.(69, 111, 112) M3 secrete anti-inflammatory cytokines (IL-10 and low-dose TGF-β) together with growth factors such as hepatocyte growth factor (HGF) and amphiregulin that favor epithelial and endothelial repair over fibroblast activation. M3 AM cells are also the dominant producers of specialized pro-resolving mediators, resolvin-D1, maresin-1, protectin-D1.(113, 114) However, whether pro-resolvins exist in significant concentrations in vivo to enhance repair is currently in dispute.(115) Pro-resolvins collectively inhibit further neutrophil recruitment, promote epithelial and endothelial repair, and helps restore normal surfactant turnover and fluid homeostasis.(87, 116, 117)

In clinical settings, the timely appearance of M3 macrophages following pneumonia, ARDS, or ventilator-induced injury is associated with faster recovery of gas exchange and a lower risk of maladaptive scarring. Failure to switch from M1/M2 intermediates to the M3 state leaves the lungs caught in a smoldering inflammatory loop, or, conversely, diverts repair into a rigid, fibrotic pathway dominated by unchecked M2-like activity. Thus, the combination of distinctive surface markers (Tim-4^high / CD206^low), transcription factors (LXRα/β-NR4A1 vs. STAT6-IRF4), and effectors (SPMs and amphiregulin) provides a clear molecular signature that separates M3 macrophages from the broader M2 family.(118–126)

Summary: An Integrated View of AM Polarization

Alveolar macrophage polarization spans a tightly regulated spectrum: pro-inflammatory, pathogen-clearing M1, tissue-remodeling and anti-inflammatory M2 subtypes, resolution-focused, efferocytosis-enhanced M3 AM. The timing, magnitude, and molecular character of each state are crucial for maintaining alveolar integrity and health. Premature or excessive M1/M2 activity can prolong injury or promote fibrosis, while failure to transition to M3 delays resolution and risks persistent dysfunction. Thus, it is vital to delineate the transcriptional, metabolic, and signaling mechanisms that regulate the AK states in order to discover new therapies that improve pathogen clearance, support repair, and promote healthy resolution while preserving lung structure,

Efferocytosis and LC3-Associated Phagocytosis (LAP) are Critical AM Functions

Efferocytosis, is essential for clearing apoptotic cells, thereby preventing secondary necrosis and promoting the resolution of inflammation.(127) (Figure 6) Alongside canonical phagocytosis and LAP, efferocytosis depends on proper phagosomal maturation and signaling. Apoptotic cells express phosphatidylserine (PtdSer) or “eat me signal” on their surface that are recognized by receptors such as MerTK, TIM4, CD36, and CD300b present on AM.(128–130) PtdSer is also recognized by GAS6 and facilitates the recognition by MerTK or AXL.(131, 132) Complement proteins such as C1q and C3b are also expressed on apoptotic cells and recognized by complement receptors, including CR1 and CR3, (expressed on AM), phosphatidylserine (PtdSer), T cell immunoglobulin, and mucin domain-containing molecule 4 (TIM4).(133, 134) Efficient efferocytosis keeps the airspaces free of cellular debris, prevents secondary necrosis and limits pro-inflammatory danger signals that would otherwise damage the blood-gas barrier and pulmonary surfactant.(135) Decreased efferocytosis is linked to unresolved inflammation, susceptibility to secondary infections and fibrotic remodeling.

Figure 6: Receptors and ligands mediated mechanism of apoptotic cell recognition and efferocytosis by AM:

Apoptotic cells express phosphatidylserine (PtdSer) or “eat me signal” on their surface, which are recognized by AM receptors such as MerTK. TIM4, CD36, and CD300b. Recognition of GAS6 enhances attachment to MerTK or AXL. Apoptotic cells also express the complement components C1q and C3b, which are recognized by the complement receptors CCR1 and CCR3 expressed on AM. Mucin domain-containing molecule 4 (TIM4) also recognizes PtdSer. Figure created with a licensed version of Biorender.com.

Recent work shows that efferocytosis in AM is often coupled to a specialized form of phagocytosis, named LC3-associated phagocytosis (LAP), that plays a critical role in the resolution of inflammation by facilitating the rapid, non-inflammatory clearance of bacteria, dead cells, and debris.(136, 137) (Figure 7) Once apoptotic cells are phagocytosed, NADPH oxidase (NOX2) generates ROS that recruit a Rubicon-Beclin-1-Vps34 lipid-kinase complex to the single-membrane phagosome. Autophagy-related proteins Atg5, Atg7, and Atg3 then lipidate LC3 onto the phagosomal membrane. LC3 decoration accelerates fusion with lysosomes, enhances acidification and speeds degradation of the apoptotic cargo.(138, 139)

Figure 7: LC3-mediated phagocytosis (LAP), Phagocytosis, and LC3-associated endocytosis (LANDO) of particles and pathogens by AM.

(A) AM initiate LAP by recognizing pathogens, opsonized particles, and dead cells via surface receptors TLR, Dectin-1, MR, and FcγR. Receptor engagement leads to a signaling cascade including recruitment of NADP oxidase (NOX2) to the phagosome, stabilized by Rubicon and PI3K complexes that includes Beclin1, VSP15, VSP34, and UVRAG. ROS produced by NOX2 and PI3K complex recruit downstream LC3 conjugation machinery including ATG12, ATG16, ATG 5 to the phagosome. Lapidated LC3-II decorated on single-membrane LAPosome, which fuses with lysosomes for the degradation of phagocytosed cargo. LAPosome-mediated phagocytosis enhances degradation and inhibits inflammation.(689, 690) (B) LC3-negative phagocytosis, which is devoid of LAPsome formation, reduces the efficiency of phagosome and lysosomal fusion and degradation of phagocytosed cargo thus promoting inflammatory responses. (C) LC3 associated endocytosis of protein aggregates including surfactant proteins A and D (SP-A, SP-D) is required for clearance and receptor recycling. This process restricts inflammation.(689) (D) Graphical illustration showing receptor engagement with various ligands present on opsonized particles. Figure created with a licensed version of Biorender.com.

In parallel, LAP alters the cytokine output of AM towards anti-inflammatory mediators (IL-10, TGF-β) and specialized pro-resolving lipid mediators, reinforcing a tissue-protective M2-like polarization program. Efficient LAP promotes antigen degradation and production of anti-inflammatory mediators, whereas defective LAP delays cellular clearance, prolongs NF-κβ and type I IFN signaling, and shifts AM toward an M1-like inflammatory state. In murine models this defect aggravates acute lung injury and remodeling, promotes autoantibody generation, and facilitates bacterial superinfection, whereas pharmacologic enhancement of LAP (e.g., PPARγ agonists, surfactant replacement) improves resolution.(137, 140–143)

Notably, the different polarization state of AM dictates the efferocytic process: anti-inflammatory M2-like AM are the most adept at both efferocytosis and LAP engagement, whereas M1- AM exhibit diminished efficiency, and the less-defined “M3” subset has not been shown to surpass M2 cells in these functions. Taken together, the LAP-efferocytosis axis is central to AM homeostasis and represents a therapeutic target to dampen injurious inflammation without compromising antimicrobial defense.

Metabolic Reprogramming

One of the defining features of AM is their ability to redistribute nutrient use and energy production in response to environmental or infectious cues. AM exist in a lipid-rich, relatively nutrient-poor milieu dominated by pulmonary surfactant. GM-CSF and TGF-β signaling induce transcription factors like PPARγ and LXR, which in turn switch on the genes that regulate fatty-acid uptake, β-oxidation, and cholesterol efflux.(144–146) Oxidative phosphorylation (OXPHOS) and mitochondrial fatty-acid oxidation (FAO) fuel the “resting” AM, and support longer, self-renewal and non-inflammatory clearance of surfactant lipids and apoptotic cells.(62, 147) (Figure 4) Any disruption in this lipid-catabolic program (as classically seen in GM-CSF or PPARγ-deficient mice), causes surfactant accumulation and pulmonary alveolar proteinosis, underscoring the essentiality of proper metabolism for AM homeostasis.(147, 148) Upon sensing pathogens or danger signals, AM undergo a rapid metabolic switch that parallels classical (M1) activation. PRRs, pro-inflammatory cytokines or hypoxia stabilize HIF-1α, suppress FAO, and drive a rapid increase in aerobic glycolysis.(149, 150)

Through glycolytic reprogramming cells generate Adenosine Triphosphate (ATP) independent of oxygen and divert glucose-6-phosphate into the pentose-phosphate pathway. This generates NADPH for NOX2-driven ROS, and accumulates the TCA intermediates, which further stabilizes HIF-1α and augments IL-1β production.(151–153) (Figure 4) Simultaneously, inducible nitric-oxide synthase (iNOS)-derived NO inhibits electron-transport chain (complex IV), forcing the cells to rely on glycolysis.(153) This M1-like metabolic shift enhances the ability of AM to kill pathogens; however, prolonged glycolysis can itself promotes tissue damage.

During the resolution of inflammation, AM undergo a second metabolic shift or a reversal of the inflammatory switch. STAT6 and PGC-1β promote mitochondrial biogenesis and activation of FAO enzymes. In contrast, arginase-1, collectively restores OXPHOS and promotes an M2-like, pro-resolving phenotype that clears debris and secretes tissue-repair mediators.(76, 154, 155) Thus, metabolic reprogramming in AM is not merely a consequence of activation but also a determinant of their functional spectrum that moves from immune surveillance to inflammation and repair.

Cross-Talk between Neutrophils and AM in Lung Inflammation and Chronic Diseases

Injurious agents cause an influx of neutrophils, monocytes, and other immune cells, together with high concentrations of cytokines and chemokines. Neutrophils arrive within 12-18 h and release potent effector molecules [e.g., ROS, neutrophil elastase, myeloperoxidase (MPO), and neutrophil extracellular traps (NETs)], that kill pathogens but also injure epithelial cells, components of surfactants, and the extracellular matrix.(156–158) ROS and proteases modify AM surface receptors and signaling pathways. For example, nitration or proteolytic cleavage of MerTK or CD36 impairs efferocytosis and phagolysosomal function and transform AM toward a more proinflammatory, glycolytic phenotype. Simultaneous exposure to IL-1β, TNF-α, and CXCL8 further amplifies the AM activation and chemokine production by creating a feed-forward loop that sustains neutrophil recruitment and tissue-damaging inflammation.(156, 159–163)

Direct interactions between neutrophils and AM influence both the onset and pathogenesis of chronic lung diseases. Impaired removal of apoptotic neutrophils, especially under high neutrophil burden or when apoptotic cells progress to secondary necrosis, results in sustained release of DAMPs that activate alveolar macrophage inflammasomes (e.g., NLRP3) and drive IL-1β production.(164–167) NETs and proteases generate matrix fragments that act as danger signals, driving macrophage secretion of matrix metalloproteinases (MMPs) and profibrotic mediators. Over time, this dysregulated cross-talk promotes protease–antiprotease imbalance, collagen degradation, and aberrant repair responses.(168–170) In COPD, recurrent neutrophil-mediated injury and AM dysfunction contribute to the emphysematous destruction and impaired host defense; in addition persistent neutrophil–macrophage signaling favors TGF-β activation, fibroblast recruitment, and deposition of the extracellular matrix leading to fibrosis.(93, 119, 171, 172)

Elucidating the specific cellular and molecular interactions between neutrophils and AM in inflamed lungs is crucial for therapeutic interventions. Targeting neutrophil-derived mediators or restoring AM function could decrease the vicious cycle of inflammation and tissue damage that results in the pathogenesis of chronic pulmonary disease.(173–176) Approaches that protect AM efferocytosis or regulate their metabolic programming toward resolution may reduce lung injury while maintaining the host defenses. Moreover, additional insight into how Neutrophil-AM interactions differ across different disease stages, could enhance the timing and specificity of treatments designed to stop or reverse pathological remodeling in COPD and fibrosis.(173, 177) Therefore, developing insights into the AM-neutrophil interplay is crucial for developing comprehensive strategies to manage and treat inflammatory lung diseases.

Impact of Environmental Agents and Food Particles on AM Polarization and Function

Upon encountering environmental agents, AM initiate an immune response by producing cytokines and chemokines, recruiting additional immune cells to the injury site, and increasing pathogen clearance. AM capacity to adapt and respond effectively is essential for preventing infections, limiting inflammation, and preserving normal lung function. Environmental agents include a wide range of substances that can either directly injure epithelial cells or activate AM, leading to inflammation and tissue remodeling. Some of these environmental agents, food particles, and their impact on macrophage phenotype and function (Table 2A and 2B) are discussed below.

Table 2A.

Phenotypic and molecular markers that characterize alveolar macrophages (AM) in three settings: (i) physiologic homeostasis, (ii) exposure to non-infectious environmental agents (e.g. Ozone, cigarette smoke, PM2.5, coal, silica, asbestos) and (iii) infection with Mycoplasma pneumoniae and Klebsiella pneumoniae. For the two exposure conditions, the table distinguishes the early “M1-like” pro-inflammatory phase from the later “M2-skewed” or reparative phase that often follows prolonged or repeated stimulation.(53, 146, 178, 195, 215, 254, 257, 297, 407, 566, 568, 583, 660, 669–672)

ALVEOLAR-MACROPHAGE Marker Matrix
	Non-exposed	Environmental Agents			Mycoplasma pneumoniae		Klebsiella pneumoniae
Category	Homeostatic / Quiescent	Early M1 burst	Chronic / Late M2 drift	Coal-Dust Exposure	Acute M1 burs	Late M2 / Regulatory	Acute M1 Burst	Late M2 / Regulatory
Surface / phenotypic markers	CD11c^high, Siglec–8, CD169, MerTK; low CD11b, MHC-II	↑CD80, CD86, MHC-II; ↑TLR2/4, MARCO; ↑CD11b, ↓Siglec–8	↑CD206, CD163, MerTKhigh, PD-L1; ↓CD11c	↑TLR2, MARCO, CD11b; ↑CD206, CD163, CD36	CD80, CD86, CD14, TLR2/6; NLRP3–ASC specks	CD206, CD163, MerTK, TIM-4, scavenger Rs	↑TLR4, CD14, MD-2, FcγR, CR3; ↑CD11b, CD80	↑CD206, CD163, MerTK, IL-10R; ↓TLR4
Signature cytokines / chemokines	GM-CSF, TGF-β, SP-A/D (basal)	TNF-α, IL-1β, IL-6, CXCL8/IL-8; HMGB1, S100A8/A9	IL-10, TGF-β, PDGF, VEGF, MMP-9/12	TNF-α, IL-1β, TGF-β, CCL18, PDGF	TNF-α, mature IL-1β, IL-6, CXCL8	IL-10, TGF-β, Arg1 metabolites, PDGF, MMP-9	IL-1β, IL-6, TNF-α, CXCL10	IL-10, TGF-β, PDGF, MMP-9
Enzymes / mediators	Balanced cathepsins; surfactant enzymes; basal HMOX1	iNOS (NOS2), NO, NOX2-ROS; active caspase-1	Arginase-1, MMP-9/12, chitotriosidase	NOX2, cathepsin B/L, MMP-9; heme oxygenase (HO-1)	High iNOS, ROS burst, caspase-1, GSDMD	Arginase-1, ABCA1/G1 efflux, CHI3L1	iNOS, NADPH-oxidase, caspase-1, GSDMD	Arginase-1, CHI3L1, MMP-9
Key signaling / TFs	PPARγ, KLF4, basal NRF2	NF-κβ (p65), AP-1, HIF-1α; NLRP3 inflammasome	STAT3, STAT6, SMAD2/3; re-engaged PPARγ	NF-κβ, AP-1, HIF-1α, STAT6	NF-κβ, p38-MAPK, MyD88, NLRP3	STAT3, STAT6, PPARγ; epigenetic gain at ARG1/MRC1	NF-κβ, IRF3, STAT1	STAT3, PPARγ, IRF4
Metabolic / redox profile	OXPHOS; FA β-oxidation; high GSH; low ROS	Aerobic glycolysis; NADPH-oxidase; GSH depletion; itaconate-low	FA oxidation; itaconate-high; partial antioxidant recovery	Chronic glycolysis + FAO; elevated ROS; iron handling	Glycolysis; PPP; NADPH for ROS	Arginine depletion; itaconate↑; FAO; glutamine anaplerosis	Aerobic glycolysis; succinate↑; ROS	FAO recovery; NADPH regeneration; itaconate↑
Functional outcome	Efficient phagocytosis; surfactant clearance	“Primed but paralyzed”; tissue injury; neutrophil influx	Fibrosis/emphysema; impaired clearance	Chronic inflammation; fibrogenic signaling; PM retention	Effective early killing; tissue injury risk	Resolution with risk of persistence/remodeling	Rapid bacterial killing; pyroptosis; inflammation	Suppressed inflammation; risk of chronic colonization
Clinical / pathology conditions	Healthy lung; immune silence	Acute COPD exacerbation; early particle toxicity	Late fibrosis (silicosis, asbestosis)	Coal-workers’ pneumoconiosis; PM fibrosis	Post-Mp organizing pneumonia; acute Mp pneumonia	Post-Mp convalescence; regulatory myeloid dominance	Acute Gram-negative pneumonia; necrotizing pneumonia	Post-Kp recovery; risk of bronchiectasis

^*Siglec-F is a mouse AM marker; the human analogue is Siglec-8; (↑) is increased; (↓) decreased; R = receptor.

Table 2B.

Phenotypic and molecular markers that characterize alveolar macrophages (AM) during infection with (i) infection with Influenza A (flu) and (ii) infection with SARS-CoV-2 (COVID-19). For the two exposure conditions, the table distinguishes the early “M1-like” proinflammatory phase from the later “M2-skewed” or reparative phase that often follows prolonged or repeated stimulation.(587, 610–614, 617–619, 654, 656, 661, 673–675)

	Influenza A (flu)		SARS-CoV-2 / COVID-19
Category	Early M1 burst	Chronic / Late M2 drift	Early M1 burst	Chronic / Late M2 drift
Surface / phenotypic markers	↑CD80, ↑CD86, ↑MHC-II, ↑TLR3, ↑TLR7, ↑CD11b, ↑CD38	↑CD206, ↑CD163, ↑MerTK, ↑PD-L1, ↑TIM-4, ↑SIRPα	↑CD80, ↑CD86, ↑MHC-II (variably), ↑TLR7/8 sensing, ↑CD11b; appearance of inflammatory monocyte-like macrophages (CD14+/CD16+/CCR2+)	↑CD206, ↑CD163, ↑MerTK, ↑PD-L1, ↑TIM-4, ↑SIRPα; expansion of tissue-resident reparative phenotype in convalescence
Signature cytokines / chemokines	IFN-α/β, TNF-α, IL-6, CXCL10, CCL2, GM-CSF	IL-10, TGF-β, PDGF, CCL22, IL-1Ra	IFN-I (often blunted), IL-6, TNF-α, IL-1β, CCL2, CCL5, CXCL10, GM-CSF	IL-10, TGF-β, CCL18, IL-1Ra, growth factors (VEGF/PDGF) during repair
Enzymes / mediators	iNOS, NOX2, cathepsin B, PKR, OAS1, ISG15	Arginase-1, MMP-9, CHI3L1, HO-1	iNOS (variable), ROS, inflammasome activation (NLRP3 → IL-1β/IL-18), high ferritin-associated responses	Arginase-1, MMPs (remodeling), HO-1, pro-resolving mediators (lipoxins, resolvins in recovery)
Key signaling / TFs	IRF3, IRF7, NF-κβ, AP-1, STAT1	STAT3, STAT6, PPARγ, IRF4	NF-κβ, IRF3/7 (dysregulated), NLRP3 inflammasome, STAT1, AP-1	STAT3, STAT6, PPARγ, IRF4; resolution-associated transcriptional programs
Metabolic / redox profile	Aerobic glycolysis↑, pentose phosphate pathway↑, ROS↑	FAO↑, glutathione↑, itaconate↑	Glycolysis↑ in inflammatory macrophages, mitochondrial stress, ROS↑, glycolytic rewiring in recruited monocyte-derived cells	FAO↑, mitochondrial recovery, increased antioxidant pathways, itaconate and pro-resolving metabolites ↑
Functional outcome	Early virus clearance; tissue injury; neutrophil influx	Resolution/remodeling; suppress viral inflammation	Excessive inflammation, endothelial activation, alveolar injury, microthrombosis; impaired efferocytosis and antigen presentation in severe cases	Tissue repair and fibrosis risk; suppression of inflammation but potential for persistent fibrotic remodeling and impaired host defense
Clinical / pathology conditions	Influenza A viral pneumonia, ARDS risk	Post-influenza organizing pneumonia; risk of secondary infection	COVID-19 pneumonia, hypoxemic respiratory failure, ARDS, thromboinflammation, multisystem ammatory features	Post-COVID organizing pneumonia, fibrosis, prolonged respiratory symptoms

Ozone

Ozone (O₃) is a potent oxidant gas that, upon inhalation, rapidly reacts with the thin aqueous film lining the airways and alveoli, generating a host of ROS/RNS as well as oxidized lipids and proteins. These oxidized molecules act as danger signals or DAMPs, engaging pattern-recognition receptors (TLRs, NLRP3) on AM, triggering a rapid shift towards a pro-inflammatory, M1-like phenotype.(2, 145, 172) Acute ozone exposure of AM up-regulates NF-κβ and AP-1 signaling pathways,(178) driving transcription of Tumor necrosis factor (TNF-α), Interleukin-1β (IL-1β), Interleukin-6 (IL-6), and chemokines (e.g., CCL2, CXCL1).(178–181) At the same time, iNOS is induced, fueling a surge of nitric oxide (NO) and superoxide production augmented by NADPH oxidase.(182–186) This oxidative burst helps clear oxidized debris but also damages the epithelial barrier, increases vascular permeability leading to edema, and accelerates neutrophil recruitment.

Paradoxically, macrophage phagocytic capacity is often reduced during this phase, as oxidative modifications of surfactant proteins and cytoskeletal elements impair phagocytosis.(178, 187–190) With repeated or chronic ozone challenge, AM undergo adaptive shifts that alter their phenotype and function. Persistent oxidative stress and ongoing tissue injury gradually induce antioxidant pathways, most notably Nrf2-driven expression of antioxidant enzymes, including heme oxygenase-1 (HO-1), glutathione synthesis enzymes, and superoxide dismutases to blunt the pro-inflammatory response.(181, 191, 192) Concurrently, their polarization begins to shift, and a partial M2 or pro-resolving phenotype (Mres) emerges. AM secrete higher levels of IL-10 and TGF-β, express scavenger receptors (CD206, MerTK), and shift their metabolic flux toward fatty acid oxidation and oxidative phosphorylation in place of glycolysis, which is a hallmark of M2 phenotype.(193)

Although this adaptive “resolution-oriented” reprogramming helps limit unchecked inflammation and promotes clearance of apoptotic cells, it can also foster airway remodeling, fibrosis, and stiffening of alveolar walls if TGF-β remains elevated.(194–196) Ozone-exposed AM may show reduced microbial killing and antigen presentation, which increases susceptibility to respiratory infections. Taken together, ozone exposure first drives AM into a hyperinflammatory state that clears oxidized damage at the cost of tissue integrity; over time, AM develop enhanced antioxidant defenses and adopt a partial reparative phenotype that, if unchecked, may lead to pathological remodeling and increased susceptibility to lung disease. Therefore, in ozone-exposed AM, the balance between acute defense driven by M1 and adaptive M2/pro-resolving programming is crucial in determining whether the lung injury resolves or progresses toward chronic airway disease.

Cigarette Smoke and Particulate Matter (PM)

Cigarette smoke, delivers a complex aerosol of chemicals including reactive aldehydes (acrolein, formaldehyde), polycyclic aromatic hydrocarbons, heavy metals, nitric oxide, and other oxygen-nitrogen intermediates and nicotine.(197, 198) Ambient or occupational particulate matter (PM) delivers to the lungs a heterogeneous mixture of carbon cores coated with transition metals, sulfates, nitrates, and organic compounds. Particles with aerodynamic diameter of ≤2.5 μm (PM_2.5) are readily deposited in the small airways and alveoli.(199–201) Normally, AM rapidly clear these inhaled particles by phagocytosis, subsequently undergoing apoptosis or removal by the mucociliary transport apparatus or efferocytosis by other macrophages.(145). However, when the particulate burden becomes chronic through repeated smoking, polluted work environments, or urban smog, AM experience sustained chemical and physical stress that rewires virtually every aspect of their biology.

The initial pivotal impact of these toxicants is an intense burst of oxidative and electrophilic stress that drives AM toward a classically activated M1 phenotype, characterized by the production of TNF-α, IL-1β, IL-6, CXCL8, iNOS, and ROS. Quinones and particulate matter-bound metals such as iron, nickel, and copper(199, 202–205) catalyze the generation of hydroxyl radicals and peroxynitrite.(206). Concurrently, reactive aldehydes form covalent adducts with thiol groups on proteins, deplete intracellular glutathione, and hamper critical antioxidant defenses. Initially activation of NRF2 results in a protective response by up-regulating heme oxygenase-1, NAD(P)H:quinone oxidoreductase-1, and glutamate–cysteine ligase. However, chronic exposure leads to the degradation of NRF2 and carbonyl modification of its co-activators.(207–214) The resulting redox imbalance damages lipids, proteins, and DNA (which impairs mitochondrial function), promotes AM toward a chronically pro-oxidant state, and perpetuates mitochondrial dysfunction.

Oxidative stress intersects with innate receptor signaling to generate a paradoxical “primed but paralyzed” AM phenotype. Engagement of TLRs (like TLR 2, TLR4) and scavenger receptors (MARCO) by smoke particles and PM activates NF-κβ, AP-1, and MAPK cascades, driving high-level secretion of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) and chemokines such as CXCL8/IL-8 that orchestrate neutrophil infiltration and perpetuate airway inflammation.(215–222) Furthermore, phagolysosomal rupture by engulfed particles allows cathepsin leakage into the cytosol, activating the NLRP3 inflammasome, which generates mature IL-1β and IL-18.(223) At the same time, carbonyls such as acrolein in smoke and soluble components of PM, inhibit MyD88-dependent signaling, reduce STAT1 phosphorylation, and down-regulate key phagocytic receptors (MARCO, SR-A, FcγRs)(224–228) resulting in AM that contribute to inflammation and lose their ability to phagocytose microbes and apoptotic-cell clearance (efferocytosis).(229). These maladaptive changes may contribute to the development of COPD and recurrent infections.

Repeated exposure to smoke and particulate matter drives a drift toward an immunosuppressive, tissue-remodeling M2-like state marked by arg-1, CD206, CD163, and secretion of IL-10, TGF-β, VEGF, and platelet-derived growth factors (PDGF). AM secrete matrix metalloproteinases (MMP-9, MMP-12) and cathepsins, while oxidative inactivation of α₁-antitrypsin and secretory leukocyte protease inhibitor removes the brakes on elastolytic activity.(230–234) Elastin and collagen fibers within the alveolar wall are progressively destroyed, expanding airspace and generating the hallmark emphysematous lesions of COPD. In parallel, TGF-β and PDGF released from stressed macrophages stimulate fibroblast proliferation and myofibroblast differentiation, linking particulate exposure not only to emphysema but also to small-airway fibrosis and remodeling.(235–237)

In addition to these functional alterations, repeated exposure to cigarette smoke and PM imprint long-lasting epigenetic and metabolic reprogramming on AM. DNA methylation changes at loci (e.g., aryl hydrocarbon receptor repressor (AHRR) and cytochrome P450 genes), loss of histone deacetylase-2 (HDAC2), and shifts in microRNA profiles (increased miR-21 and miR-155, decreased miR-146a) lock cells into a semi-activated, steroid-resistant state phenotype that persists even after smoking cessation. This is a form of maladaptive “trained immunity.” AM switch to glycolytic metabolisms which reinforces their pro-inflammatory states.(238–252)

Phenotypically, sustained oxidative stress, aryl hydrocarbon receptor signaling, excessive efferocytosis, and epigenetic rewiring promote this M1-M2 conversion, while a metabolic switch to fatty-acid oxidation dampens antimicrobial ROS production. This shift from an early injurious M1 surge to chronic, protease-rich yet immunosuppressive M2 dominance, underpins the clinical characterization of COPD: accelerated decline in lung-function, emphysema, small-airway fibrosis, frequent infectious exacerbations, and poor corticosteroid responsiveness.(253–261) Eliminating this maladaptive response requires elimination of exposure, such as smoking cessation, air-quality control, and re-calibrating macrophage biology (e.g., antioxidants that restore NRF2 activity, PPARγ or Arg-1 inhibitors that rebalance M1/M2 polarization, and agents that preserve HDAC2 function).(262–264)

Asbestos, Silica, and Coal Dust

Mineral dust, most notably crystalline silica, asbestos fibers, and coal mine dust, poses unique challenges that overwhelm the injury to AM degradative machinery. The interaction of these particles with AM is a key early step in the pathogenesis of pneumoconiosis, asbestosis, and progressive fibrosis. Upon encountering respirable crystalline silica, AM internalize the particles into phagolysosomes. Unlike organic material that is broken down by enzymes, silica crystals rupture and destabilize the phagolysosomal membrane, causing leakage of proteolytic enzymes and cathepsins into the cytosol. This breach activates the NLRP3 inflammasome, culminating in caspase-1-dependent cleavage of pro-IL-1β to mature IL-1β (a potent pyrogen and recruiter of neutrophils) and an explosive M1 response.(265–271) This event triggers macrophage apoptosis or an inflammatory cell death called pyroptosis. The dying macrophages then release their proinflammatory contents, including IL-1β and TNF-α, perpetuating a chronic inflammatory milieu in the lung parenchyma. In the sub-acute phase, many of the surviving macrophages phagocytose apoptotic debris and transition toward an M2/M2-like phenotype that expresses Arg-1, mannose receptor (CD206) and high levels of TGF-β.(272, 273) AM containing silica become paradoxically immunosuppressed with phagolysosomal acidification and IFN-γ signaling, thereby explaining the well-known susceptibility of silicotic lungs to Mycobacterium tuberculosis.(274–276)

Asbestos fibers present a different physical challenge to AM. Many amphibole and chrysotile fibers exceed the length macrophages can fully engulf, leading to “frustrated phagocytosis.”(Figure 8) AM adhere to protruding fiber ends while their NADPH oxidase remains chronically activated, producing a sustained burst of RO and RN species.(277–283) Early M1 polarization is e driven not only by classical cytokines but also by the oxidative stress pathways NF-κβ and AP-1. To sequester the foreign bodies, AM (and later adjacent type II pneumocytes) coat the fibers with an iron-protein shell, creating golden-brown “ferruginous bodies,” which become histologic hallmarks of exposure. In addition, AM switch toward an M2/M2-like profile.(284–290) The iron catalyzes Fenton reactions, generating hydroxyl radicals that impose additional oxidative stress on nearby epithelial and mesothelial cells. These activated macrophages secrete high levels of TGF-β, PDGF, VEGF, and matrix metalloproteinases, driving diffuse interstitial (asbestosis) and pleural fibrosis.(291–293) Simultaneously, asbestos-induced epigenetic reprogramming (histone acetylation, miR-21 up-regulation) locks the cells into a chronic, low-grade M2-dominant state. This dominant state supports fibroblast proliferation through ROS-mediated DNA damage, contributing to the initiation of bronchogenic carcinoma and malignant mesothelioma.(293–300)

Figure 8: Graphical illustration showing” frustrated phagocytosis” induced by a long asbestos fiber:

AM phagocytose and clear short asbestos fibers. However, asbestos fibers longer than 10μM present a challenge to AM engulfment, leading to frustrated phagocytosis. NADPH oxidase (NOX) produces ROS, which results in NF-kβ mediated expression of pro-inflammatory cytokines/chemokines. To sequester the uncleared fiber, AM coat the fiber with iron leading to generation of hydroxyl radicals. This chronic condition of oxidative stress results in the secretion of TGF-β and platelet-derived growth factor (PDGF) which may result in fibrosis. Figure created with a licensed version of Biorender.com.

In coal workers’ pneumoconiosis (“black lung”), inhaled coal dust often mixed with silica, is engulfed by AM but resists enzymatic degradation, due to its inert carbonaceous core. These pigment-laden “anthracotic macrophages,” which also contain silica, remain in a state of chronic low-level activation. Repeated lysosomal damage and ROS generation engage the NLRP3 inflammasome, yielding IL-1β, TNF-α, and further oxidative stress. Many macrophages undergo pyroptosis, releasing DAMPs that recruit additional inflammatory cells. Over time, surviving AM shift toward an M2-like, fibrogenic phenotype, up-regulating arg-1, CD206, and TGF-β to wall off dust in fibrotic nodules.(77, 170, 273, 301–304) Excessive extracellular matrix build-up and collagen cross-linking around these nodules drives progressive fibrosis, which stiffens lung tissue, disrupts ventilation-perfusion balance, and can culminate in respiratory failure.

Despite their distinct modes of injury, chemical lytic damage by silica, asbestos’s frustrated phagocytosis, iron-catalyzed oxidative stress, and inert pigment overload by coal dust, converge on a common pathophysiologic theme. This persistent activation converts a homeostatic sentinel into a chronic source of oxidants, proteases, and fibrogenic mediators. Over time, the resulting matrix deposition stiffens lung parenchyma, impairs pulmonary function, and, in the case of silica and coal dust, compromises antimicrobial activity, predisposing patients to tuberculosis. In the case of asbestos, the combination of iron-catalyzed free radicals and cytokine-driven cell proliferation underlies a marked increase in neoplastic risk.(305–307) Thus, the AM, designed to preserve pulmonary homeostasis, become the pivotal drivers of occupational lung diseases.

Food Particles--Gastric Aspiration

Pulmonary aspiration of acid and gastric contents is thought to cause acute lung injury.(308) The extent and severity of lung injury depends on the quantity and nature of the aspirate, the frequency of aspiration events, and the host’s response to the aspirated material.(309) Gastric acid aspiration has been attributed to as much as 15.6% of ARDS cases.(310–312) Clinically, pulmonary aspiration is characterized by bronchospasm, pulmonary edema, hypoxemia, and respiratory failure resulting from inflammation and airway hyperresponsiveness (AHR). Upon exposure to gastric contents or food particles, AM detect pathogen associated molecular patterns (PAMPs) and DAMPs via toll-like and NOD-like receptors, activating NF-κβ, inflammasomes, MAPK, and JAK-STAT pathways. This leads to the secretion of TNF-α, IL-1β, and IL-6, which recruit neutrophils and amplify local injury. (313–317) AM, thus, play a dual but diametrically opposite role: on the one hand, they clear aspirated material; on the other hand, they contribute to inflammation and tissue damage by generating injurious agents and attracting neutrophils to the lungs.(314, 315, 318)

Polarization into M1 or M2 phenotypes critically influences outcomes post-aspiration.(316) M1 macrophages, driven by IFN-γ and LPS engulf and phagocytose food particles. However, they may cause fibrosis if overly activated.(79, 319) M2 macrophages, triggered by IL-4/IL-13, support resolution, efferocytosis, and repair. Aspiration usually leads to early M1 dominance, followed by a transition to M2, helping to restore alveolar homeostasis.(79, 319) AM rapidly shift phenotypes in response to aspirated acid, food particles, or toxins, which contribute to chronic aspiration syndromes.(314, 320, 321) Acid aspiration suppresses phagocytosis and TNF-α release, whereas particulate matter intensifies inflammation. Activation states are further defined by metabolic reprogramming (glycolysis in M1, oxidative phosphorylation in M2, and epigenetic changes).(315, 322–324)

Interactions between AM, airway epithelium, and interstitium shape aspiration pathophysiology.(315, 325) Epithelial alarmins like IL-33 and HMGB1 activate AM by binding to ST2 and RAGE receptors, escalating inflammation.(326, 327) Macrophage-derived TGF-β and VEGF contribute to remodeling and fibrosis in recurrent aspiration.(328) Oxidative stress from epithelial injury promotes conversion of high molecular hyaluronan (HMW-HA), an anti-inflammatory agent, to pro-inflammatory low molecular hyaluronan (LMW-HA), which activates AM and recruits immune cells. This makes LMW-HA a potential biomarker for aspiration pneumonia.(329–332) Therapeutic strategies targeting polarization, enhancing M2 repair, or limiting M1 activation are under investigation. Agents like pioglitazone (PPAR-γ agonist) and NF-κβ inhibitors reduce cytokine release and neutrophil influx.(333, 334) Immunometabolic and epigenetic interventions, including HDAC inhibitors, show promise in promoting reparative macrophage phenotypes.(335). This makes AM an important therapeutic target in aspiration-related lung injury.

Another important component of aspiration is hydrochloric acid (HCl), which injures AM, reducing their adhesion to surfaces. This acid-induced injury can elicit a more robust inflammatory response in the lungs, with increased neutrophil recruitment and potential contribution to fibrosis. In vitro studies have shown that macrophage function can be negatively impacted by both the acid itself and by inflammatory factors released by other damaged cells, such as neutrophils.(336–338)

Summary: Alveolar-Macrophage Plasticity as a Unifying Determinant of Lung Outcome after Environmental Injury

AM are the lung’s gatekeepers. They phagocytose inhaled particles, patrol for microbes, and secrete growth factors that maintain the epithelial barrier. Virtually every inhaled or aspirated agent, whether ozone, cigarette smoke, mineral dust, or gastric contents, first encounters these resident phagocytes. The nature, intensity, and duration of that encounter dictate whether the lung fully recovers or proceeds toward chronic inflammation and fibrosis. A conserved “alarm” program characterizes the earliest macrophage response to all these insults. Pattern-recognition receptors (TLRs, NLRP3) on AM surfaces sense oxidized lipids, free radicals, crystalline edges of silica, and mtDNA from damaged mitochondria of epithelial cells. Within minutes NF-κβ and AP-1 are activated, iNOS synthase and NADPH oxidase generate a burst of ROS/RNS, and large amounts of TNF-α, IL-1β, IL-6, and neutrophil-recruiting chemokines are released. Although this classical M1-like response helps degrade or sequester harmful material, it also disrupts epithelial tight junctions, increases permeability, and transiently impairs phagocytosis.

Despite their distinct toxicants, all these events converge on several mechanistic themes: (i) Redox overload coupled with imperfect antioxidant recovery; (ii) Chronic NLRP3 inflammasome signaling; (iii) Epigenetic “locks” that maintain maladaptive phenotypes (HDAC2 loss, AHRR methylation, miR-21 up-regulation); and (iv) metabolic reprogramming acute glycolysis for M1 and fatty acid oxidation for M2. These shared pathways reveal therapeutic targets. Reactivating Nrf2, replenishing thiols, or supplying mitochondrial antioxidants can reset redox balance; PPAR-γ agonists or arginase-1 inhibitors may rebalance M1/M2 polarization; and preserving HDAC2 or blocking NLRP3/IL-1β may quell steroid-resistant inflammation and fibrosis.

Pulmonary Infections and their Impact on AM Polarization and Function

AM coordinate innate and adaptive responses during pulmonary infections. They detect and engulf pathogens, produce ROS and RNS, and secrete a spectrum of cytokines (e.g., TNF-α, IL-1β) and chemokines (e.g., CXCL8/IL-8) that attract neutrophils and monocytes. This recruitment fosters a multicellular antibacterial front, while antigen presentation by activated macrophages primes T-cell responses that tailor immunity to specific organisms.(8, 146, 339–341)

The outcome of pulmonary infections often hinges on the effectiveness and restraint of AM alveolar macrophage activity. If activation is insufficient, pathogens such as mycobacterium tuberculosis, streptococcus pneumoniae, mycoplasma pneumoniae, klebsiella pneumoniae, influenza virus, or SARS-CoV-2 can replicate unchecked, leading to pneumonia, acute lung injury, or disseminated disease. Conversely, excessive or dysregulated macrophage activation contributes to immunopathology, typified by the “cytokine storm” in severe viral pneumonias and by chronic inflammation and fibrosis in conditions such as hypersensitivity pneumonitis or ARDS.

Mycoplasma pneumoniae (M. pneumoniae)

AM along with SP-A and SP-D, which play crucial roles in innate immunity by opsonizing pathogens and modulating macrophage activation.(342–344) M. pneumoniae accounts for 20–30% of all pneumonias, leading to illnesses such as tracheobronchitis, bronchiolitis, pharyngitis, pneumonia, and possibly ARDS. In addition, M. pneumoniae may exacerbate other respiratory disorders such as COPD and asthma. Moreover, because of the widely diverse clinical manifestations and the need for specialized testing for identification of active infections, it is clear that M. pneumoniae diseases are underdiagnosed.(345–347) (Figure 9)

Figure 9: Phagocytosis of Mycoplasma pneumoniae (MP) by AM.

(A) Lipoproteins present on the surface of MP are recognized by TLR2/TLR6 heterodimers expressed on AM triggering activation of NF-κβ via the MyD88 dependent pathway. Additionally, MP secretes community-acquired respiratory distress syndrome (CARDS) toxin that activates NLRP3-mediated maturation of IL-1β. In the alveolar space, surfactants SP-A and SP-D help enhance phagocytosis of MP by opsonizing them. Altogether, these signals activate AM causing them to secrete pro-inflammatory cytokines, chemokines, and reactive nitrogen species (RNS). (B) MP activate AM into M1-phenotyple through MAPK-NF-kβ signaling and secrete various pro-inflammatory mediators including IL-23, IL-1, and IGF-1. IGF-1 inhibits endogenous inflammatory signals in alveolar epithelial cells.(686) AEC regulates inflammation by secreting inhibitory cytokines including IL-10, TGF-β, PGE2, and GMCSF which switch AM from the M1 to the M2 state. Figure created with a licensed version of Biorender.com.

When M. pneumoniae enter the alveolar space, their cholesterol-rich, Wall-less membrane and abundant surface lipoproteins are immediately sensed by AM pattern-recognition receptors, most prominently TLR2/TLR6 heterodimers.(348–353) Engagement of these receptors triggers MyD88-dependent signaling cascades culminating in NF-κβ and MAP kinase activation, which drives transcription of M1-associated genes.(349, 354, 355) Concurrently, M. pneumoniae secrete the community-acquired respiratory distress syndrome (CARDS) toxin, which activates the NLRP3 inflammasome, leading to caspase-1-mediated maturation and release of IL-1β(356) (Figure 8). Together, CARDS and IL-1β drive AM toward an M1 phenotype. The resulting pro-inflammatory response, regulated in part by interactions with surfactant protein A (SP-A), is essential for early containment of M. pneumoniae. AM phagocytose and kill mycoplasma by generating peroxynitrite. When infected intranasally with M. pulmonis C57BL/6 mice lacking SP-A or iNOS had significantly higher levels of mycoplasmas in their lungs as compared to wild type controls.(342, 357–359) Furthermore, depletion of AM by intranasal instillation of liposome-encapsulated dichloromethylene bisphosphonate (L-Cl2MBP) decreased the killing of M. Pulmonis in C57BL/6 mice.(360)

As the infection progresses, M. pneumoniae deploy several strategies to temper the M1 response and facilitate their existence. Engagement of TLR2 by lipoproteins, along with direct effects of CARDS toxin on host signaling pathways, gradually shift AM toward the M2-like or regulatory phenotype. The pathogens and infected epithelial cells secrete IL-10 and TGF-β, which activate STAT3 and STAT6 (361–364). These transcription factors induce expression of arg-1, which inhibit iNOS, and upregulate scavenger receptors and mannose receptors (CD206). SP-A and SP-D influence this phenotypic shift, as they can dampen the inflammation and further modulate macrophage responses. M. pneumoniae also interferes with phagolysosomal maturation and scavenges host antioxidants, allowing a fraction of organisms to persist intracellularly or remain attached to damaged epithelium.(356, 365, 366).

The shift between M1 and M2 polarization in Mycoplasma infection has important pathophysiologic consequences (367, 368). An exaggerated or prolonged M1 response can trigger excessive neutrophil recruitment, oxidative tissue damage, capillary leak, and clinical features such as cough, bronchospasm, and radiologic infiltrates(369). In contrast, an early or overly strong M2 bias may impair pathogen clearance and promote chronic or recurrent infection, airway remodeling, and persistent inflammation.(370) Surfactant proteins continually participate in these processes, either enhancing pathogen clearance or contributing to the resolution of inflammation. Thus, the timing and degree of macrophage polarization, shaped in part by surfactant proteins, critically influence whether the host experiences a self-limited pneumonia or develops complications such as bronchiolitis obliterans or post-infectious reactive airway disease.(371, 372)

Klebsiella pneumoniae (K. pneumoniae)

K. pneumoniae is a Gram-negative, heavily encapsulated bacterium that has become a leading cause of both hospital-acquired and community-acquired pneumonia. AM recognize K. pneumoniae primarily via TLR4 engagement of LPS, and to a lesser extent via TLR2 by recognition of outer-membrane lipoproteins. SP-A and SP-D bind to exposed surface carbohydrates or LPS motifs on Klebsiella, bridging the bacterium to AM scavenger receptors and enhancing uptake. Indeed, SP-A drives the killing of K. pneumoniae by human AM isolated from transplanted lungs in vitro through stimulation of phagocytosis and generation of ROS intermediates.(373)(Figure 10) Thus, the efficiency of early pathogen detection depends not only on classical PRRs but also on the availability and integrity of these surfactant proteins.(145, 374–378)

Figure 10: Interaction of AM with Klebsiella pneumoniae (KP):

(A) Surfactant proteins SP-A and SP-D opsonize KP, which facilitates their phagocytosis by AM, leading to ROS generation and NLRP3 inflammasome activation. KP also activates AM via surface TLRs to produce various inflammatory mediators, which results in the recruitment of various immune cells including neutrophils and Myeloid-Derived Suppressor Cells (MDSCs) to the lung alveolar space. (B) KP components, such as outer membrane protein A (OmpA), lipopolysaccharide (LPS), and capsular polysaccharides (CPS), are recognized by TLR2 and TLR4 and activate AM via the MAPK-NF-kβ pathway to secrete pro-inflammatory cytokines/chemokines and ROS. (C) Phagocytosed KP activates the type 6-secretion-system in response to generated ROS, which activates the NLRP3 inflammasome, which cleaves pro-IL-1β and pro-IL-18 into mature IL-1β and IL-18 and mounts an inflammatory response. (D) KP infected AM produces high levels of itaconate and α-ketoglutarate that promote fatty acid oxidation (FAO) and OXPHOS which favors M2 phenotype with upregulated arginase 1 (ARG1), CD206, and CD163. This activation leads to the accumulation of myeloid-derived suppressor cells (MDSCs) and M2-like macrophages in the lung.(627) Figure created with a licensed version of Biorender.com.

This initial encounter drives a rapid polarization of AM toward the M1 phenotype. TLR- and collectin-mediated signaling converges on NF-κβ and MAP-kinase pathways, inducing transcription of pro-inflammatory cytokines (TNF-α, IL-1β, IL-6) and chemokines (CXCL8/IL-8, CXCL1, CCL2). SP-A and SP-D amplify this response by boosting cytokine release and enhancing phagolysosomal fusion in the early hours of infection. M1-polarized AM increase glycolysis, up-regulate iNOS, and generate ROS, all of which are crucial for limiting bacterial replication and killing.(378–383)

K. pneumoniae counters these defenses through multiple evasion strategies. For example, it has been reported that K. pneumoniae ’s thick polysaccharide capsule, shields underlying antigens from SP-A and SP-D, allowing the pathogen to evade rapid clearance, to proliferate, and produce the dense, lobe-filling infiltrates and necrotic lung damage characteristic of severe K. pneumoniae.(373, 384–386)Its mucoid capsule contains SP-A/SP-D binding sites and blocks phagocytosis. In addition, ROS and RNS will nitrate and oxidize residues in the collectins carbohydrate recognition domain decreasing their abilities to opsonize pathogens.(387–389) Thus, K. pneumoniae may prolong their extracellular survival and sustain the M1-driven inflammatory milieu. The sustained influx of neutrophils and release of oxidants and proteases, although bactericidal, also injure the epithelial barrier, predisposing the lung to edema, hemorrhage, and, in severe cases, ARDS.(390–392)

As tissue damage accumulates, anti-inflammatory mediators such as IL-10, TGF-β become more prominent, driving AM toward the M2 phenotype. Furthermore, SP-A and SP-D promote this transition by engaging signal-regulatory protein-α (SIRPα) and other inhibitory receptors on the macrophage surface.(376, 393, 394, 394–397) A carefully timed M2 shift is essential for wound resolution; however, if it occurs prematurely or becomes dominant while viable bacteria are still present, it may result in the persistence and dissemination of Klebsiella.(76, 392, 398) Clinical studies have shown that patients with low levels or dysfunctional variants of SP-A or SP-D are more prone to chronic or relapsing Klebsiella infections, highlighting how deficiencies in surfactant protein driven regulation of macrophage phenotype can tip the balance toward bacterial persistence and maladaptive repair.(376, 382, 399)

Recognizing the dual role of surfactant proteins in regulating AM polarization has opened new therapeutic avenues. Recombinant SP-A/SP-D or small molecules that enhance their expression are being explored to improve opsonization and early M1 efficacy.(52, 400, 401) Alternatively, therapies that preserve collectin function while selectively decreasing NF-κβ signaling such as TLR4 antagonists and PPAR-γ agonists, are being considered.(402–407)

Influenza A Virus (IAV)

Influenza A virus (IAV), the principal agent of seasonal and pandemic “flu,” is a highly contagious respiratory pathogen capable of causing anything from mild, self-limited illness to fulminant viral pneumonia, ARDS and lethal secondary bacterial infections.(408, 409) While ciliated airway epithelium and type II pneumocytes are the primary sites of productive replication, IAV infection profoundly disturbs AM biology and function.(410–413) (Figure 11). Whether human AM support a productive IAV cycle is debated: several studies detect intracellular viral RNA and proteins, yet most report an abortive infection with little or no release of infectious progeny.(117, 414–416) This “shouldering” replication may still suffice to re-program AM and amplify inflammation while sparing neighboring epithelial cells from further seeding.

Figure 11: Detection of Influenza A virus by AM.

(A) Activation of the innate immune response: Dying influenza virus infected cells or influenza virus opsonized with surfactants proteins SP-A and SP-D are phagocytosed by AM via scavenger receptors and PRRs. Released viral RNA recognized by TLR3 and TLR7 either in endosomes or by RIG-1/MDA-5 in cytosols. This leads to the stimulated expression of NF-kβ and IRF-7-mediated pro-inflammatory cytokines/chemokines (e.g., TNF-α, IL-6, Il-1β, and IL-8) and type 1 interferon (IFN) via IRF3 transcription factors. Activated AM produce reactive oxygen and nitrogen intermediates (ROS/RNS) and rely on enhanced glycolysis for their metabolic needs. (B) Adaptive immune response initiation: Internalized viral components undergo antigen processing and present them to helper T cells and cytotoxic CD8 T cells via MHCII and MHCI respectively resulting in activation of the adaptive immune response. Activated T cells, particularly CD8 T cells, kill the virus and decreases the viral load. (C) AM anergy: Influenza virus downregulates the expression of scavenger and activation receptors as well as inhibits phagocytosis capability of AM. Influenza infection causes a drastic depletion of AM population (anergy) in the lung, which results in compromised innate and adaptive immune responses. This condition leads to the secondary infection caused by bacteria and fungi. (D) Recovery phase: Enhanced expression of PDL1 on AM and TGF-β signaling differentiates the naïve T cells to regulatory T cells (Treg) and release of IL-10. Inflammation is inhibited by increased expression of CD200R and scavenger receptors as well as activation of PPARγ/STAT6. Figure created with a licensed version of Biorender.com.

The scenario dramatically shifts when the influenza virus enters the airways. AM phagocytose flu virions and debris, acting as sentinels that sense viral RNA through endosomal TLR-7 and cytosolic RIG-I/MDA-5.(417, 418) Activation of MyD88-, TRIF-, and MAVS-dependent pathways drives AM away from resting homeostasis. Transcription factors such as NF-κβ, IRF3, and IRF7 move into the nucleus to induce interferon-stimulated genes and pro-inflammatory cytokines.(341, 419–421) Within just 4 hours, type I IFN genes dominate the transcriptome, and TNF-α peaks between 4 and 8 hours post-infection, confirming AM as the first-responder “cytokine sprinters.”(422) The AM metabolism shifts from oxidative phosphorylation to glycolysis to fuel an energetic burst needed for acute defense. Direct mitochondrial damage (accumulation of swollen, abnormal organelles and down-regulation of oxidative-phosphorylation genes) further enforces this glycolytic bias.(74, 423, 424)

At the same time, SP-A and SP-D, coat influenza virions promoting rapid phagocytosis and better detection by macrophage antiviral sensors.(8, 340, 425, 426) There is increased expression of antigen-presenting markers (MHC-II, CD80, CD86) and Fc receptors (CD64), while markers associated with the M2 phenotype (such as CD206) and arg-1 tend to decline.(427, 428) Concomitantly, type I IFN signaling suppresses PPAR-γ which correlates with delayed pneumonia resolution.(82) AM produce inflammatory cytokines (TNF-α, IL-1β, IL-6) and chemokines (CXCL10, CCL2, CCL5), which attract neutrophils, NK cells, and monocytes to the alveolar space.(420, 421, 429, 430) These recruited cells cooperate with AM to contain viral replication and clear infected epithelial cells. ROS and RNS generated by activated AM further destroy viral particles but can also inflict collateral injury on the delicate lung parenchyma and surfactant layer.(145, 431) Despite this early burst, AM still secrete fewer inflammatory mediators than blood-derived macrophages, revealing a built-in ceiling that limits by-stander damage under physiological conditions.(432)

If influenza infection is mild and short-lived, these changes are usually reversible. Many seasonal H1N1/H3N2 infections remain confined to the upper airway and may barely engage AM at all.(413) However, in more severe or repeated infections a more complex picture emerges. Persistent pattern-recognition signaling keeps AM fixed in an inflammatory M1-state, with high levels of iNOS, ROS, and cytokine output.(59, 145, 428) Moreover, chronic oxidative stress oxidizes surfactant phospholipids, undermining their biophysical role in maintaining alveolar stability and further impairing gas exchange.(433, 434) The resulting “immunoparalysis” can last for weeks, characterized by reduced phagocytosis and down-regulation of scavenger receptors such as CLEC7A and MSR1, thereby predisposing to secondary infections.(435)

In addition, Influenza viruses that entered AM activate the inflammasomes (notably NLRP3) that triggers pyroptosis, apoptosis, and necroptosis (PANoptosis).(436–439) The viral PB1-F2 protein, by targeting mitochondria and collapsing membrane potential, is a leading candidate for this cytotoxic effect.(440) In murine models, up to 60% of resident AM may be lost in the acute phase, depleting the local phagocyte pool just when robust clearance of debris and pathogens is needed.(441, 442) This “macrophage-disappearance reaction,” is absent in pigs and ferrets and not yet proven in humans, emphasizing species differences.(412, 443) Surviving AM often become functionally exhausted, showing diminished phagocytic capacity, reduced cytokine production upon secondary stimulation, and increased expression of inhibitory checkpoint molecules (PD-L1, Tim-3, CD200R).(13, 444, 445) This state, especially in the context of ongoing surfactant degradation, creates a vulnerability, which explains why convalescent patients are predisposed to secondary bacterial pneumonia caused by organisms such as Streptococcus pneumoniae.(446)

Over time, repeated or chronic infection can induce lasting epigenetic changes in AM. Histone modification patterns (such as H3K4me3 and H3K27ac) may produce ‘trained immunity,’ heightening responses to future pathogens.(447–451) Alternatively, chronic interferon exposure may drive endotoxin tolerance, leading to anergic macrophages that fail to respond to new infections. Both conditions interfere with the ability of AM to recycle surfactant, which may lead to alveolar proteinosis and compromised gas exchange.(452–454) Intriguingly, influenza-trained AM have recently been shown to confer long-term antitumor protection in the lung.(455)

During recovery, adaptive immunity, particularly neutralizing antibodies and cytotoxic T lymphocytes, reduce the viral load, and pro-resolving cytokines (IL-10, resolvins, lipoxins) help transition AM to an M2-like repair state.(456–460) Their metabolism shifts back to oxidative phosphorylation, scavenger receptors such as CD206 and CD163 reappear, and repair factors such as VEGF, PDGF and amphiregulin facilitate the restoration of the epithelial barrier and basement membrane.(125, 461–463, 463–467) Surfactant homeostasis is gradually reestablished as type II pneumocytes increase surfactant synthesis and AM ingest oxidized proteins and lipids. If recovery is unimpeded, lung function may return to normal; however, repeated influenza infections or unresolved inflammation can tip the balance toward chronic fibrosis, as AM secrete TGF-β and drive persistent myofibroblast activity and collagen deposition.(468–470) Here, surfactant turnover remains disrupted and alveolar walls stiffen, establishing a feed-forward loop of mechanical stress and inflammation.

In summary, influenza virus infection initiates a dynamic process of polarization that promotes the rapid switching of AM from M2 to inflammatory M. Depending on disease severity and chronicity, possibly culminating in exhaustion, tolerance, or fibrotic dysfunction. Each state reflects the interactions of viral signals, surfactants, immunometabolism, and epigenetic memory, emphasizing the double role of AM in preventing and inducing influenza-related lung pathology.

SARS-CoV-2 (COVID-19)

SARS-CoV-2, the causative agent of COVID-19, is a highly transmissible respiratory virus that may cause severe viral pneumonia, ARDS, multi-organ dysfunction with long-term sequelae. Beyond targeting the ACE2-expressing airway and alveolar cells, SARS-CoV-2 infections also perturb AM biology and function.(471–474) (Figure 12) When AM are infected with SARS-CoV-2, viral RNAs are recognized by endosomal and cytosolic pattern-recognition receptors of AM (TLR7/8 and RIG-I/MDA5), and their associated adaptor molecules (MyD88, TRIF, MAVS) that promote the translocation of key transcription factors (e.g., NF-κβ, IRF3, IRF7) into the nucleus.(475–482) This, induces the production of type I/III interferons, interferon-stimulated genes (ISGs), and proinflammatory cytokines (TNF-α, IL-1β, IL-6), that initiate an antiviral response and recruits additional innate effectors.(473, 483, 484) Unlike other respiratory viruses, the timing and magnitude of the interferon response to SARS-CoV-2 infection are often dysregulated, delayed, or blunted. Early responses of type I IFN have been associated with worse outcomes, whereas prolonged IFN signaling can contribute to tissue pathology in severe disease.(483, 485, 486)

Figure 12: Detection of SARS-CoV-2 by AM.

(A) Resident AM detect SARS-CoV-2 by endocytosis, ACE2 receptors, and the FCγR1-IgG immune complex pathway. Upon activation by viral components, AM releases various cytokines and chemokines which recruit other immune cells including bone marrow derived macrophages (BMDMS), neutrophils, NK cells, and T cells into the lungs. Macrophages process viral components and present them to T helper cells and cytotoxic CD8 T cells via MHCII and MHCI respectively. Activated T helper cells secrete IFN-γ, which further activates AM; CD8 T cells secrete granzyme B, which reduces viral load. During the acute phase of SARS-CoV-2 infection, the AM population is depleted (anergy) due to cell death pathways, including pyroptosis, apoptosis, and necroptosis. Infected alveolar epithelial cells (AEC) also engage in inflammation by secreting various cytokines and chemokines. (B) After internalization of the virus, released viral RNA is recognized by either TLR3 or TLR7 in the endosome or by RIG-1/MDA-5 in the cytosol. This leads to the activation of NF-kβ, IRF-7 and IRF-3 transcription factors that activate the AM to produce cytokines/chemokines (e.g., TNF-α, IL-6, Il-1β, CCL2, and IL-8) and type-1 interferon. Activated AM mount oxidative stress and rely on glycolysis for their metabolic requirement. ROS and RNS alter surfactants SP-A and SP-D function by oxidizing them or nitrating them, reducing their capacity to opsonize the virus. (C) Circulating viral components act as pathogen-associated molecular patterns (PAMPs) and stimulate monocytes, leading to expression of the tissue factor (coagulation factor III) on monocyte and endothelial surfaces.(531, 691) Tissue factors and neutrophil extracellular traps (NETs), released by neutrophils stimulate the coagulation pathway leads to accumulation of activated platelets, deposition of fibrin and collagen, and ultimately results into blood clotting. Angiotensin-converting enzyme two (ACE2); major histocompatibility complex (MHC); Fc gamma receptor type I (FCγR1). Figure created with a licensed version of Biorender.com.

Phagocytosis of SARS-CoV-2 by AM results in the upregulation of Fc receptors (CD64), antigen-presenting molecules (MHC-II, CD80/CD86), and results in the secretion of chemokines (CXCL10, CCL2, CCL5) that recruit monocytes, neutrophils, and NK cells to the alveolar space.(487–490) There is a metabolic shift of AM toward aerobic glycolysis that provides the energy needed for phagocytosis. However, when sustained, promotes robust M1-like features with high production of ROS that can damage alveolar epithelium and surfactant components. SP-A and SP-D that normally dampen inflammation may change their function in this milieu, altering opsonization and viral clearance dynamics.(491–493)

Although classical infection of SARS-CoV-2 in AM is limited due to the low expression of ACE2, viral uptake can still occur via phagocytosis of infected debris, Fc receptor–mediated pathways, or lectin interactions.(494, 495) Failed viral RNA sensing or infection can activate inflammasomes, most commonly NLRP3, leading to the release of IL-1β/IL-18 and triggering lytic cell death, which contributes to a PANoptotic milieu (pyroptosis, apoptosis, necroptosis).(496–498) These processes can reduce the resident AM pool during the acute phase, while the inflammatory byproducts amplify local tissue injury.

The chemokine-mediated influx of CCR2+monocytes differentiate into monocyte-derived macrophages within the alveolar space. These recruited cells adopt a highly proinflammatory transcriptional program distinct from that of homeostatic AM and can dominate the local cytokine milieu in severe infection.(499, 500) This shift in macrophage composition, loss of tissue-resident AM, and accumulation of inflammatory monocyte-derived macrophages, impairs surfactant recycling and amplifies the tissue-damaging responses.(341, 430, 501) AM-derived cytokines and chemokines further recruit neutrophils and prompts NETosis, endothelial activation, and cooperates with systemic inflammatory pathways to drive microvascular thrombosis and vascular leaks, the hallmarks of severe COVID-19 lung injury. ROS and proteases released from the AM degrade surfactant lipids and proteins and thereby increasing surface tension, predispose to alveolar collapse, and worsen gas exchange.(492, 502–504) The combined epithelial and endothelial injury disrupts alveolar fluid clearance and surfactant homeostasis, exacerbating hypoxemia.

Surviving AM and recruited monocyte-derived macrophages can become functionally exhausted during prolonged or severe illness, displaying impaired phagocytosis, increased cytokine responses to secondary stimuli, and upregulation of inhibitory receptors, such as PD-L1, TIM-3.(505–507) Persistent type I IFN exposure and chronic inflammatory signaling may also induce an immune tolerance in some macrophage populations.(508, 509) Along with, damaged surfactant components and impaired mucociliary clearance, this further increases the vulnerability to secondary bacterial pneumonias and delays recovery.

SARS-CoV-2 infection can leave long lasting epigenetic and metabolic marks on AM. Patterns of histone modifications and chromatin accessibility may confer a form of trained immunity, heightening responsiveness to later insults or, conversely, inducing sustained hypo-responsiveness.(510–512) In some individuals, prolonged macrophage-mediated profibrotic signaling (TGF-β production), together with epithelial injury, and altered surfactant turnover, promotes fibroblast activation and deposition of extracellular matrix, leading to post-COVID pulmonary fibrosis in some survivors.(513–516) By contrast, effective adaptive immunity, neutralizing antibodies, and virus-specific CD8+ T cells reduce viral load, while pro-resolving mediators (IL-10, specialized pro-resolving lipid mediators) and a shift toward reparative macrophage polarization helps restore homeostasis.(517–519) Post-infection or during recovery, the pool of AM are replenished, cellular metabolism reverts toward oxidative phosphorylation, reestablishing scavenger receptors (e.g., CD206, CD163), and surfactant-clearing functions, and most importantly type II pneumocytes resume surfactant synthesis.(493, 520–523) If resolution is timely, surfactant homeostasis and alveolar architecture can largely normalize; however, failure of resolution, leads to persistent inflammation, repeated injury, and maladaptive repair and chronic dysfunction. (524)

In summary, SARS-CoV-2 exposure triggers a dynamic, macrophage response that initially aims to control the virus but when dysregulated by viral factors, comorbidities, host genetics, or delayed interferon signaling, can drive inflammatory lung injury, surfactant dysfunction, and delayed repair. The balance between effective antiviral clearance and protracted immunopathology in AM largely determines disease severity and the risk of long-term pulmonary sequelae.

Differences and Similarities of Influenza A and SARS-CoV-2 Infections

Influenza A virus (IAV) and SARS-CoV- engage AM in distinct ways. IAV attaches to α2,3- or α2,6-linked sialic acids on the macrophage surface and enters the AM by endocytosis; some avian or highly pathogenic strains progress partway through the viral life-cycle, yet most human IAV strains achieve only abortive replication, producing little infectious progeny while still exposing the cell to viral RNA and proteins.(525–527) Conversely, SARS-CoV-2, depends upon the ACE2 receptor for canonical entry, so the virus usually reaches AM indirectly through phagocytosis of infected epithelial debris or via Fcγ-receptor uptake of antibody-opsonized virions.(497, 528, 529) Thus, SARS-CoV-2 is less efficient in completing a full replication cycle in AM as compared to IAV, which typically leaves behind a non-replicative viral genome that nonetheless alters macrophage physiology. On the other hand, IAV often triggers overt AM death (apoptosis or pyroptosis), whereas SARS-CoV-2 more commonly reprograms the surviving AM without causing extensive early depletion.(530–532)

These two viruses activate different inflammatory responses in AM. IAV drives an immediate, high-intensity M1-like activation, marked by a robust increase of type I interferons, TNF-α, IL-1β, IL-6, and neutrophil-attracting chemokines.(533–536) This sudden surge is further amplified by a metabolic shift towards aerobic glycolysis and succinate-driven HIF-1α signaling.(150, 536, 537) SARS-CoV-2 initially blunts type I interferon output and induces a more gradual, heterogeneous activation.(538–540) During mild COVID-19, AM adopt a mixed blend of regulatory (IL-10, TGF-β) and pro-inflammatory signals; whereas in severe cases, AM and the monocyte-derived macrophages that replace or augment them, produce high levels of IL-6, CXCL10, and CCL2, fueling the cytokine storm.(541–544) Accordingly, IAV provokes an early interferon-dominant surge that can be self-damaging, whereas SARS-CoV-2 often causes a delayed yet constant inflammatory state dominated by recruited myeloid cells.

These upstream differences result in distinct clinical outcomes. Following IAV infection, the loss of AM and their impaired phagocytic function hampers the surfactant clearance and bacterial killing, creating a favorable environment for classic post-influenza pneumonias caused by pathogens such as Streptococcus pneumoniae or Staphylococcus aureus.(117, 413, 545–547) In contrast, SARS-CoV-2 infection leaves a pool of metabolically and transcriptionally altered AM that, together with recruited monocyte-derived macrophages, sustain alveolar inflammation, promote endothelial injury and, in prolonged cases, contribute to pneumonia or fibrotic remodeling. Secondary bacterial infections after a SARS infections are less frequent.(541, 542, 548, 549) Therapeutically, these differences require different strategies: following IAV infection, restoring AM numbers and phagocytic capacity is critical to prevent superinfection, whereas in COVID-19 the priority is on suppressing dysregulated macrophage-driven inflammation while preserving their antiviral and reparative capacities.

Immunotherapeutic Interventions Targeting AM

A number of emerging therapies attempt to re-programming AM rather than merely suppressing downstream inflammation. Inhaled granulocyte-macrophage colony-stimulating factor (GM-CSF) replenishes depleted AM pools, restores phagocytic competence, and lowers post-influenza bacterial pneumonia.(550–552) Different metabolic modulators, such as PPAR-γ agonists and AMPK activators attempt to convert, highly glycolytic AM back toward oxidative phosphorylation while directly curbing viral replication.(553, 554) Epigenetic interventions that erase long-lasting “immunoparalysis” are being explored to re-establish balanced antimicrobial and reparative functions after acute lung injury.(555) Short, local blockade of inhibitory checkpoints and mitochondria-protective agents that neutralize viral factors offer additional routes to rescue AM bioenergetics without provoking a cytokine storm.(541) Presently, none of these interventions are FDA approved for the treatment of occupational or pathogen induced lung diseases. In addition, their long-term cardiotoxicity, well established for cancer chemotherapeutic agents and checkpoint inhibitors (556, 557) has not been investigated.

Insights from OMIC Studies

In recent years, OMIC technologies, including genomics, transcriptomics, epigenomics, proteomics, and metabolomics have transformed our understanding of how diverse inhaled insults have offered new insights into molecular rewiring that underlies dysfunctional AM states and unveiled novel targets for precision interventions.

Genomic and epigenomic studies have identified susceptible loci and long-lasting “Epigenetic scars” in AM exposed to pollutants, dust, M. Pneumonia, and K. Pneumoniae. Genome-wide association studies have linked variants in IL-1β, NLRP3, and MARCO to exaggerated inflammatory responses. Whole-genome bisulfite sequencing has uncovered DNA hypermethylation at regulatory regions of PPARγ and IL10, as well as altered chromatin accessibility at interferon and antioxidant response elements.(558–565) K. pneumoniae infected AM show pathogen-driven chromatin remodeling with increased accessibility at TLR4, IRF3, and STAT1 effectively priming these cells for heightened cytokine production upon secondary challenges.(566, 567)

Transcriptomic analysis, spanning both bulk and single-cell RNA sequencing (scRNA-seq), have identified heterogeneous AM populations following environmental or infectious conditions. For example, exposure to coal dust or particulate matter induces distinct AM subpopulations expressing high levels of inflammatory cytokine (NLRP3, IL1β), chemokines (CCL2, CXCL2), and pattern recognition receptors.(568–570) During M. Pneumoniae infection, RNA-seq analysis showed the presence of an early NF-κβ burst followed by a STAT1/IRF7-dominated interferon response; while, K. pneumoniae triggers a similar biphasic response, with an initial TLR4/NF-κβ activation followed by IRF3/IFN-β upregulation. scRNA-seq showed the presence of AM expressing high levels of CXCL10, IFNB1and IRG1, and SPP1 that coordinates downstream neutrophil and T-cell recruitment.(571–578). In K. pneumoniae, scRNA-seq identified an immunoregulatory subset expressing high levels of PD-L1 and a cathepsin-rich cluster generating high levels of ROS poised for pyroptotic death. These findings, highlight how distinct pathogens sculpt unique AM phenotypes.(394, 577, 579–583) AM of mice infected with influenza A virus exhibit distinct interferon-dominated AM and altered antigen presentation trajectories that shape antiviral responses and immunopathology.(584–592) Similarly, studies on SARS-CoV-2 infection reveal unique AM transcriptional landscapes characterized by delayed and dysregulated interferon signaling, enhanced inflammatory monocyte-derived macrophage recruitment, and the emergence of pro-fibrotic and thrombo-inflammatory gene signatures, which differentiate COVID-19 from other respiratory infections at the single-cell level.(542, 548, 549, 593) ScRNA-seq has also revealed a loss of homeostatic gene signatures and emergence of “trained immunity” profiles in AM following exposure to environmental and pathogenic agents. AM exhibit durable epigenetic and metabolic imprints, including sustained elevation of glycolytic enzymes and H3K4me3 marks at inflammatory loci, which heighten responsiveness to subsequent insults.(9, 548, 564, 565, 578, 594–597)

The integration of proteomics and metabolomics findings has further identified exposure and infection-driven transformations in AM. Proteomic studies of the AM after pollutants and M. pneumoniae challenge have identified elevated levels of oxidative enzymes (NOX2), inflammasome components (NLRP3, Caspase-1), proteases (cathepsins), remodeling proteins (MMPs), and phosphorylation of NF-κβ, MAPK cascades. Infection with M. Pneumoniae has shown increased IRG1, PKR, and antigen-presentation levels while depleting surfactant-handling proteins, indicating a metabolic diversion toward host defense.(194, 598–603) K. pneumoniae challenge enriched TLR4/MD-2 complexes and outer-membrane protein networks, signifying adaptation to Gram-negative LPS exposure.(566, 604–608) Influenza A virus infection likewise reshaped proteomic signatures toward antiviral effector and immune modulation, contributing to pathophysiology of influenza-driven lung disease.(609–613) Proteomic analysis of AM from COVID-19 patients demonstrates upregulation of pro-inflammatory cytokines, complement factors, coagulation mediators, and markers of cellular stress, reflecting the unique thrombo-inflammatory milieu and sustained tissue damage characteristic of severe SARS-CoV-2 infection.(542, 614–619) Metabolomic profiling of AM has revealed a significant decrease in itaconate and increase in succinate and lactate, supporting a shift towards glycolytic metabolism and hypoxia-inducible inflammation (HIF1α).(620–622)

In M. Pneumoniae-infected neutrophils or mixed lung-cell suspensions (rodents), LC–MS shows a significant rise in itaconate and a parallel accumulation of arachidonate-derived lipid mediators, underscoring immunometabolic rewiring that can either constrain or propagate inflammation.(603, 623–626) By contrast, AM following K. pneumoniae infection exhibit increased succinate, which promotes the release of IL-1β and a decrease in SDH–dependent itaconate synthesis.(150, 604, 627, 628) Similarly, Influenza A virus infection reprograms metabolic pathways in AM, balancing antiviral defense with inflammatory output and tissue injury, with notable shifts in glycolysis, fatty acid metabolism, and mitochondrial dynamics.(611, 612)

By integrating these multi-OMIC datasets, systems biology approaches identified central regulators (like NLRP3, NRF2, MARCO, and SLC7A11) and network “hubs” with the greatest influence over AM responses to environmental insults.(8, 629–632) These hubs provide strategic entry points for selective therapeutic targeting, ranging from NRF2 activators that temper pollutant-induced oxidative stress to metabolic modulators that curb excessive CXCL10 production during M. pneumoniae and K. pneumoniae infection.(366, 633)

Focusing specifically on M. Pneumoniae, complementary CRISPR-Cas9 dropout screens reinforce STING, PKR, IRG1, MARCO, and GBP2/5 as “Tier-1” host factors, while parallel screens in K. pneumoniae models highlight TLR4, MyD88, and Vav1 as essential for bacterial clearance. Epigenomic assays reveal a TLR2–MyD88–DNMT3B axis that hyper-methylates PPARγ/IL10 and builds H3K27ac/H3K4me3 super-enhancers at CXCL10, IFNB1, and IRG1.(349, 368, 571, 577, 634–640) BET inhibition by JQ1 or I-BET15 (a class of targeted, epigenetic drugs that bind to Bromodomain and Extra-Terminal proteins) eliminates these super-enhancers and selectively depletes the AM subset that expresses high levels of CXCL10.(641–644) In addition, DNMT3B knockdown restores IL-10. Proteo-phospho profiling has identified persistent phosphorylation of p65, p38-MAPK, TBK1, and STAT1. Small-molecule TBK1 or p38 blockade temper the interferon-dominated second wave of phosphorylation in M. pneumoniae-infected mice.(645–647) Metabolite-based interventions such as exogenous dimethyl-itaconate or 5-LOX/PGE₂-synthase inhibitors, dampen cytokine storms and airway hyper-reactivity in both M. pneumoniae and K. pneumoniae infection models.(623, 648–652) Emerging CRISPR and epigenetic studies in SARS-CoV-2 models similarly point to key host factors regulating interferon pathways, inflammasome activation, and metabolic reprogramming, highlighting potential therapeutic targets to modulate macrophage-driven inflammation in COVID-19.(653–658)

Altogether, OMIC approaches not only dissect the complex molecular rewiring of AM by environmental pollutants and M. pneumoniae, K. pneumoniae, influenza A, and SARS-CoV-2 (COVID-19), but also provide crucial insight into the development of precision therapies and biomarker-based diagnostics that may be used to monitor and mitigate combined pollutant- and infection-related lung diseases. High-Dimensional Single-Cell OMIC combined with spatial transcriptomics, proteomics, cell biological, and physiological measurements hold high promise in unraveling the heterogeneity and dynamic responses of AM subpopulations to a wide spectrum of environmental and pathological insults. Integration of these approaches with sophisticated in-vivo imaging and lineage-tracing models will illuminate how repeated or chronic exposures reshape macrophage phenotypes, their interactions with lung structural cells, and their roles in inflammation, tissue repair, and disease progression.

Unanswered Questions: Areas for Future Research

• Which signals propel AM towards pro-inflammatory, anti-inflammatory, reparative, and pro-resolving stages when exposed to simultaneous but conflicting signals (e.g., a viral infection in addition to environmental agents)?

• Which are the primary molecular triggers (DAMPs, oxidized lipids, crystalline edges, or lysosomal rupture) that activate the AM inflammasome and the Nrf2?

• Can we map exposure-specific AM phenotypes to distinct tissue remodeling outcomes (fibrosis vs. repair vs. emphysema) using spatial transcriptomics and single-cell proteomics?

• What are the mechanisms by which influenza A infection perturbs AM homeostasis and surfactant integrity, and how can AM-targeted strategies preserve antiviral defense while limiting tissue injury?

• How do co-exposures (environmental agent plus Mycoplasma pneumoniae or Klebsiella pneumoniae infection) reshape AM polarization, inflammasome activation, efferocytosis, and MET formation?

• Are there pathogen-specific AM subpopulations (e.g., PD-L1high, ROS^high, SPM^high) that emerge only under certain host–pathogen–exposure combinations, and can they be therapeutically targeted?

• Which single-cell and spatial-OMICS signatures best predict transition from acute exposure/infection to chronic lung remodeling, and how can these be translated to clinical biomarkers?

Conclusions

In conclusion, AM play a pivotal role in the lung’s defense, responding dynamically to an ever-widening array of environmental agents and pathogens. Repeated or co-exposures to coal dust and other respirable particles can skew AM toward pro-inflammatory or pro-fibrotic states, impairing surfactant recycling, efferocytosis and bacterial killing. Likewise, M. pneumoniae, K. pneumoniae, influenza A, and SARS-CoV-2 deploy toxins, lipoproteins, and LPS-rich outer-membrane vesicles to subvert phagolysosomal fusion, induce oxidative stress, and trigger inflammasome-driven pyroptosis. The net result is a dysregulated macrophage phenotype that fuels chronic inflammation, compromises barrier integrity, and propels diseases such as asthma exacerbation, COPD, organizing pneumonia, and pulmonary fibrosis.

Glossary with Description of Some Abbreviations

Abbreviation	Meaning	Description
ABCA1 / ABCG1	ATP-binding cassette transporter A1	It is involved in lipid export from macrophages.
AP-1	Activator Protein-1	A transcription factor, Stimulates inflammatory and stress responses.
AREG	Amphiregulin	It is an epidermal growth factor, which is crucial for cell growth, differentiation and tissue repair.
ASC specks	Apoptosis-associated speck-like protein	Apoptosis-associated speck-like protein containing a CARD; adaptor protein oligomers that scaffold active inflammasomes
ATG	Autophagy-related protein	It regulates formation and maturation of autophagosomes.
C1q a/b/c	Subunits of complement component C1q	Important in recognition and clearance of apoptotic cells and immune complexes.
Cathepsin B / L	Lysosomal proteases B and L	Implicated in phagolysosomal digestion and NLRP3 activation after lysosomal damage.
CCL22	C-C chemokine ligand 22	Chemokine often associated with regulatory/T-helper 2 recruitment
CCL5	C-C chemokine ligand 5	Also called RANTES, Chemokine recruiting T cells, monocytes and NK cells.
CD11b	Cluster of differentiation 11b	Integrin α M (ITGAM), marker of recruited/inflammatory myeloid cells and phagocytic activity.
CD11c	Cluster of differentiation 11c	Integrin alpha X (ITGAX), A myeloid integrin highly expressed on tissue macrophages and dendritic cells; associated with phagocytosis and adhesion.
CD14	Cluster of differentiation 14	Co-receptor for LPS (lipopolysaccharide).
CD16	Cluster of differentiation 16	marker distinguishing monocyte subsets (CD14+/CD16+)
CD163	Cluster of differentiation 200	Scavenger receptors expressed on AM and help in clearance of Hemoglobin-haptoglobin complexes.
CD169	Cluster of differentiation 169	Sialoadhesin (SIGLEC1), A macrophage lectin implicated in cell–cell interactions and antigen capture.
CD200	Cluster of differentiation 200	Type I transmembrane glycoprotein, acts as an inhibitory signal to AM.
CD200R	Cluster of differentiation 200 receptor	Inhibitory receptor expressed on alveolar epithelial cells that bind to CD200 to suppress macrophages activation.
CD206	Cluster of differentiation 200	Mannose Receptor (MRC1); function as endocytic receptor, mainly expressed on M2 macrophages and help in resolution of inflammation.
CD300b	Cluster of differentiation 300b	Receptors help in clearance of apoptotic cells.
CD36	Cluster of differentiation 36	Act as scavenger receptor for fatty acids, oxidized lipid and pathogens and help in platelets activation.
CD38	Cluster of differentiation 38	Ecto-enzyme and activation marker associated with inflammation and NAD+ metabolism; marks activated macrophages.
CD4+ T cells	Cluster of differentiation 4 expressing T cells	Helper T cells, crucial cells of immune system that shape adaptive immune responses by signaling B cells macrophages and CD8 T cells.
CD8+ T cell	Cluster of differentiation 8 expressing T cells	Cytotoxic T lymphocyte, crucial cells of adaptive immune system, which directly kill pathogens by identifying them through MHC, I molecules.
CD80	Cluster of Differentiation 80 B7-1	T-cell costimulatory ligand expressed on activated antigen-presenting cells; upregulated on pro-inflammatory macrophages to enhance T-cell activation.
CD86	Cluster of Differentiation 86 (B7-2)	Partner costimulatory molecule similar function to CD80.
CXCL8 (IL-8)	C-X-C motif chemokine ligand 8	Key neutrophil chemoattractant.
DAMPs	Damage-Associated Molecular Patterns	Endogenous molecules released by stressed or dying cells (e.g., mitochondrial DNA, ATP) that activate innate immune receptors and inflammasomes.
DC-SIGN	Dendritic Cell-Specific Intercellular adhesion molecule-3	Grabbing Non-integrin (CD209). A C-type lectin implicated in pathogen binding.
Dimethyl Itaconate	Dimethyl Itaconate	A synthetic derivative of itaconate used to modulate macrophage metabolism and inflammatory responses.
Ferruginous Bodies	Ferruginous Bodies	Iron-protein coated asbestos fibers found in lung tissues, indicative of asbestos exposure and associated with fibrosis.
FOXO3 / p53	Forkhead Box O3	Regulators of stress responses, senescence and apoptosis.
GSDMD	Gasdermin D	Components of pyroptotic cell-death pathway release IL-1β/IL-18 and amplify inflammation.
H3K4me3	Histone H3 Lysine 4 Trimethylation	Epigenetic activation mark
HDAC2	Histone Deacetylase 2	An epigenetic regulator whose loss contributes to chronic inflammation and steroid resistance in diseases like COPD.
HIF-1α	Hypoxia-Inducible Factor 1α	Coordinates glycolytic and inflammatory responses under hypoxia.
HMGB1	High mobility group box 1	A DAMP released by dying or stressed cells.
HO-1	Heme oxygenase-1	Antioxidant enzyme, key in heme/iron handling and antioxidant responses.
IL-10	Interleukin-10	Anti-inflammatory cytokine promoting resolution.
IL-1β	Interleukin-1 beta	Pro-inflammatory cytokine processed by caspase-1.
IL-6	Interleukin-6	Promotes acute-phase responses, systemic inflammation, and immune cell recruitment.
IRF3 / IRF4	Interferon regulatory factors 3/4	Involved in antiviral (IRF3) and alternative activation / tolerance (IRF4) programs.
IRF3 / IRF7	Interferon regulatory factors 3/7	Interferon regulatory factors driving type I IFN gene transcription after viral sensing (critical for IFN-α/β production).
ISG15	Interferon-stimulated gene 15	Ubiquitin-like modifier induced by type I IFNs with antiviral roles.
KRT79	Keratin 79	Epithelial marker; occasional detection may reflect epithelial–macrophage interactions or contamination in transcriptomes.
MAPK	Mitogen-Activated Protein Kinase	A signaling cascade involving p38, JNK, and ERK pathways that regulate inflammation and macrophage activation.
MARCO	Macrophage Receptor with Collagenous Structure	A scavenger receptor for bacteria and particulates.
MerTK	MER proto-oncogene, tyrosine kinase	It is highly expressed on macrophages and help in efferocytosis.
METs	Macrophage Extracellular Traps	Fibrous structures composed of DNA and antimicrobial proteins expelled by macrophages to trap pathogens.
MHC I	Major Histocompatibility Complex Class I	Important molecule of immune system, which expressed antigen presenting, cells and displays internal cell antigen to cytotoxic T lymphocyte (CD8) for the T cell activation.
MHCII	Major Histocompatibility Complex Class II	Expressed on antigen presenting cells such as macrophages and dendritic cells and present exogenous antigen to CD4 T cells.
MMP	Matrix metalloproteinases	Remodel extracellular matrix; contribute to tissue remodeling and pathology.
MyD88	Myeloid differentiation primary response 88	An adaptor for most TLRs driving NF-κβ activation.
NETosis	Neutrophil Extracellular Traps formation	The process by which neutrophils release extracellular traps to entrap pathogens.
NFATc1	Nuclear factor of activated T cells, c1	Implicated in macrophage fusion and giant cell formation.
NLRP3	Inflammasome	A multiprotein complex in macrophages that activates caspase-1 to process IL-1β and IL-18, inducing inflammatory responses and pyroptosis.
NLRP3	NOD-like receptor protein 3 Inflammasome	Multiprotein sensor complex that activates caspase-1 to process IL-1β/IL-18 and trigger pyroptosis.
NRF2	Nuclear Factor Erythroid 2–Related Factor 2	Master transcriptional regulator of antioxidant and detoxification genes.
OAS1	2′-5′-oligoadenylate synthetase 1	ISG that activates RNase L antiviral pathway. OH−: Hydroxyl radical
OmpA	Outer membrane protein A	Found on outer membrane of many gram-negative bacteria and act as a channel for small nutrients.
P2X7	Purinergic Receptors	Receptors activated by extracellular ATP that can trigger inflammasome activation and inflammatory responses.
PD-L1	Programmed Death Ligand-1	An inhibitory checkpoint ligand that dampens T-cell responses.
PGE2	Prostaglandin E2	Lipid mediator with immunomodulatory roles.
Plet1	Placenta-expressed transcript 1	It plays role in tissue repair and wound healing.
PLIN2	Perilipin-2	Lipid droplet coat protein
PPARγ	Peroxisome Proliferator-Activated Receptor gamma	Regulates lipid metabolism and ant-inflammatory programs in macrophages.
S100A8/A9	Calprotectin subunits (S100 proteins)	DAMPs and pro-inflammatory mediators released by myeloid cells.
SASP factors	Senescence-Associated Secretory Phenotype	Senescent cells secrete pro-inflammatory cytokines and proteases (IL-6, CCL2, MMPs).
Siglec-8	Sialic acid-binding Immunoglobulin-like Lectin 8	A lectin family receptor involved in recognition of sialylated glycans; Siglec-F is the murine analog.
SIRPα	Signal regulatory protein alpha	Binds CD47 and transmits “don’t eat me” signals; regulates phagocytosis.
STAT1 / STAT3 / STAT6	Signal transducer and activator of transcription 1/3/6	Key mediators of IFN, IL-10, and IL-4/IL-13 signaling, respectively.
TLR7 / TLR8	Toll-like receptors 7 and 8	Endosomal sensors for single-stranded RNA that trigger type I IFN and inflammatory signaling in myeloid cells.
TNF-α	Tumor necrosis factor α	Potent pro-inflammatory cytokine involved in cell activation, vascular effects, and recruitment.
VEGF	Vascular Endothelial Growth Factor	Pro-angiogenic factor.
VSP34	Vacuolar protein sorting 34	Class III phosphatidylinositol 3-kinase.

Clinical Highlights (Call-Out Box for Clinicians).

• Alveolar macrophages are central components of lung defense, repair, and remodeling. Their ability to transform from proinflammatory to pro-resolving states plays a vital role in defending the lungs from external environmental agents and pathogens and initiating tissue repair.

• AM clear apoptotic cells and debris which minimizes pro-fibrotic signaling and enhance repair

• Dysregulated AM responses contribute to COPD progression, pneumoconiosis/fibrosis, organizing pneumonia, decreased surfactant clearance and susceptibility to secondary infections due to impaired phagocytosis/efferocytosis or altered surfactant turnover.

• Surfactant proteins A and D work in unison with AM to phagocytose and pathogens

Glossary

ADRP

Adipocyte Differentiation-Related Protein

ALI

Acute Lung Injury

APCs

Antigen-Presenting Cells

ARG1

Arginase-1

BMDM

Bone marrow derive macrophage

Car4

Carbonic Anhydrase 4

Caspase-1

Cysteine-aspartic protease 1

CCR1

C-C chemokine receptor type 1

CCR3

C-C chemokine receptor type 3

CD68

Glycoprotein macrophage marker

cGAS

Cyclic GMP-AMP Synthase

CHIL3 (YM1)

Chitinase-Like Protein 3 (mouse Ym1)

CPS

Capsular polysaccharides

CREB

cAMP Response Element-Binding Protein

CXCL1

C-X-C motif chemokine ligand 1

CXCL9

C-X-C motif chemokine ligand 9

Ear1

Eosinophil-associated ribonuclease 1

ECM

Extracellular matrix

ETC

Electron transport chain

FAS

Fatty acids synthesis

FcγR

Fc gamma receptors

GAS6

Growth Arrest-Specific protein 6

GSH

Glutathione, Major intracellular antioxidant tripeptide

IFN-α/β

Type I interferons

ILC2

Innate lymphoid cells type II

IRF1

Interferon Regulatory Factor 1

ITGAX

Integrin alpha X gene

KP

Klebsiella pneumoniae

LAP

LC3-mediated phagocytosis

LMW-HA

Low Molecular Weight Hyaluronan

LPS

Lipopolysaccharide

Ly6c2/Ly6C

Lymphocyte Antigen 6 Complex, locus C2 (mouse monocyte marker)

MD-2

Myeloid differentiation factor 2

MDSCs

Myeloid-Derived Suppressor Cells

MKI67 (Mki67)

Marker of Proliferation Ki-67

MPO

Myeloperoxidase

mtROS

Mitochondrial reactive oxygen species

NETs

Neutrophil Extracellular Traps

NO·.

Nitric oxide

O₂⁻

Superoxide

ONOO⁻

Peroxynitrite

p53

Tumor Protein p53

PDGF

Platelet-derived growth factor. Mitogenic and pro-fibrotic mediator

PI3K

Phosphoinositide 3-Kinase

PM

Particulate matters

Pro-IL-1β

Precursor Interleukin-1 beta

Prussian blue+

Positive iron stain (ferric iron deposits)

RelA

NF-κβ p65

RNS

Reactive nitrogen species

RUNX1

Runt-Related Transcription Factor 1

SiglecF

Sialic acid-binding Immunoglobulin-like Lectin F (mouse)

SP-A and SP-D

Surfactant Protein-A / Surfactant Protein- D

STAT

Signal Transducer and Activator of Transcription

T6SS

Type 6-secretion-system

TGF-α

Transforming Growth Factor Alpha

TGFβR

Transforming growth factor-beta receptor

TIM-4

T cell immunoglobulin and mucin domain-containing protein-4

TLR2 / TLR4

Toll-like receptor 2 / Toll-like receptor 4

TOP2A

DNA Topoisomerase II Alpha

TREM2

Triggering Receptor Expressed on Myeloid Cells 2

UVRAG

UV radiation resistance-associated gene

VSP-15

Vacuolar protein sorting-15

AEC

Alveolar epithelial cell

AM

Alveolar Macrophages

ARDS

Acute Respiratory Distress Syndrome

BACH2

BTB and CNC Homology 1, Basic Leucine Zipper Transcription Factor 2

C5aR1 (C5ar1)

Complement Component 5a Receptor 1

CARDS

Community-Acquired Respiratory Distress Syndrome toxin

CCL2 (MCP-1)

Monocyte chemoattractant protein-1

CCR2 (Ccr2+)

C-C Chemokine Receptor 2

CCR7

C-C Chemokine Receptor 7

CFTR

Cystic Fibrosis Transmembrane Conductance Regulator

CHI3L1 (CHI3L1/YKL-40)

Chitinase-like protein implicated in remodeling and fibrosis

Cholesteryl esters

Stored cholesterol derivatives

CR3

Complement receptor 3

CS

Cigarette smoke

CXCL10

C-X-C motif chemokine ligand 10

DCs

Dendritic Cells

ELF

Epithelial lining fluid

ENAC

Epithelial Sodium Channel

EVALI

E-cigarette or vaping product use-associated lung injury

FAO

Fatty Acid Oxidation

Ferritin

Iron storage protein

GM-CSF

Granulocyte-Macrophage Colony-Stimulating Factor

IFN

Interferon

IGF-1

Insulin-like Growth Factor 1

iNOS (NOS2)

Inducible nitric oxide synthase

IRF8

Interferon Regulatory Factor 8

KLF4

Krüppel-Like Factor 4

LANDO

LC3-associated endocytosis

LC3

Microtubule-associated protein 1 light chain 3

LPP

Lipid peroxidation product

LXR-α

Liver X Receptor alpha (NR1H3)

Lysozyme

Antimicrobial muramidase enzyme

MDA-5

Melanoma Differentiation-Associated gene 5

OXPHOS

Oxidative phosphorylation

MP

Mycoplasma pneumoniae

MR

Mannose receptor

NADPH

Nicotinamide Adenine Dinucleotide Phosphate

NF-κβ

Nuclear factor kappa β

NOX2

NADPH Oxidase

OH⁻

Hydroxyl radical

OxPL

Oxidized phospholipid

PAMPs

Pathogen-Associated Molecular Patterns

PF4

Platelet Factor 4

PKR

Protein kinase R

PPP

Pentose phosphate pathway

PRRs

Pattern Recognition Receptors

PtdSer

Phosphatidylserine

RIG-1

Retinoic acid-inducible gene I

ROS

Reactive Oxygen Species

SA-β-gal+

Senescence-Associated beta-Galactosidase positive

SMAD2/3

Intracellular mediators of TGF-β signaling (pro-fibrotic programs)

SREBP-1c

Sterol Regulatory Element-Binding Protein-1c (SREBF1)

STING

Stimulator of interferon Genes

TCA

Tricarboxylic Acid Cycle

TGF-β

Transforming Growth Factor-beta

Th1

Helper T cells

TLR3

Toll-like receptor 3

TNF-α

Tumor necrosis factor α

Treg

Regulatory T cells

TRPV4

Transient Receptor Potential Vanilloid 4

VEGF- α

Vascular Endothelial Growth Factor A