**5. Physiological roles of alveolar macrophage efferocytosis in acute LRTIs**

#### **5.1 Roles of alveolar macrophages in viral infection**

Alveolar macrophages can directly or indirectly endocytose viruses via pinocytosis or opsonization, respectively. In the case of SARS-CoV-2, alveolar macrophages also recognize viral components such as envelop protein [73], spike protein [74–77], and single-stranded RNA [78, 79] using TLR2, TLR4, and TLR3/7, respectively, which trigger pro-inflammatory responses. However, the phagocytic and pro-inflammatory responses of alveolar macrophages against viruses appear to be dispensable for protecting the host from viral infection. Indeed, the absence of mature alveolar macrophages in GM-CSF-deficient mice resulted in severe respiratory failure and increased mortality after pulmonary infection with a non-lethal dose of influenza A virus, and these conditions were improved by neonatal transplantation of alveolar macrophage progenitor cells from wild-type mice [11]; however, alveolar macrophage-depleted mice exhibited severe manifestations, with viral clearance not being largely impaired and the functions of antibody-producing B lymphocytes and cytotoxic CD8-positive T-lymphocytes being normally activated [11]. Similarly, critically ill patients with COVID-19 have been characterized by a depletion of alveolar macrophages and a remarkably increased proportion of recruited pro-inflammatory monocyte-derived macrophages in bronchoalveolar lavage fluid [12]. These suggest that alveolar macrophages contribute to host survival by suppressing excessive pulmonary inflammation, which is caused by removing endogenous apoptotic cells rather than by phagocytosing the exogenous virus itself during infection.

#### **5.2 Regulation and roles of efferocytosis in alveolar macrophages**

Notably, clearance of apoptotic cells, termed efferocytosis, is an essential process for maintaining tissue homeostasis under both healthy and diseased conditions. Efferocytosis differs morphologically and mechanistically from the classical form of phagocytosis against pathogens and requires the expression of *Physiological Role of Alveolar Macrophage in Acute Lower Respiratory Tract Infection… DOI: http://dx.doi.org/10.5772/intechopen.110509*

receptors that recognize "eat me" signatures such as phosphatidylserine (Ptd-L-Ser) exposed on the membrane surface of apoptotic cells [80]. Macrophages perform efferocytosis primarily using tyrosine receptor kinases as Ptd-L-Ser receptors, including Tyro 3, Axl, and proto-oncogene c-mer tyrosine kinase (MerTK) (collectively abbreviated as TAM) [81]. In a recent study, transcriptome and flow-cytometric analyses revealed that murine alveolar macrophages highly express Axl and MerTK, but little or no expression was found in lung-mobilized monocytes after the LPS challenge [82]. Moreover, human alveolar macrophages predominantly express Axl, and peripheral monocytes do not express either Axl or MerTK [83]. Although Axl-knockout mice did not manifest inflammatory disorders under healthy conditions, they exhibited exaggerated severity during pulmonary infection with influenza A virus, accompanied by increased accumulation of apoptotic cells, elevated infiltration of neutrophils and T-lymphocytes, and increased secretion of pro-inflammatory cytokines and chemokines, without compromising virus clearance [10]. In addition, during acute lung injury after LPS challenge in mice, alveolar macrophages engulfed Pst-L-Ser-exposed microparticles but not lung-mobilized monocytes, and deletion of MerTK abrogated efferocytosis activity in both in vivo and in vitro experiments [82]. Therefore, alveolar macrophages prevent excessive pulmonary inflammation via efferocytosis using Axl and MerTK in lung injuries caused by viruses and bacteria; notably, lung-mobilized pro-inflammatory monocytes do not contribute to efferocytosis, at least at the early stage of infection.

#### **5.3 Anti-inflammatory properties of efferocytosis in alveolar macrophages**

Notably, TAM receptor-mediated recognition of Ptd-L-Ser requires soluble cross-linking molecules in the serum (growth arrest-specific gene 6 or protein S) [84]. Similar to pathogen recognition by phagocytic receptors, ligation of TAM receptors results in the activation of Rac1, leading to membrane ruffling to engulf apoptotic bodies [85, 86]. Phagocytic receptors are linked to pro-inflammatory responses [4, 87], whereas TAM receptors activate anti-inflammatory responses in macrophages. For example, TAM receptor ligation activates type I IFN receptor signaling to upregulate the expression of suppressors of cytokine signaling 1 and 3. This induces negative feedback to suppress type I IFN receptor signaling and both MyD88- and TRIF-dependent TLR signaling [88]. Moreover, the detailed molecular mechanisms underlying the promotion of anti-inflammatory IL-10 and TGF-β production during efferocytosis in macrophages have also been elucidated. The coenzyme NAD<sup>+</sup> , generated by mitochondrial β-oxidation of apoptotic cellderived fatty acids, activates sirtuin-1 and downstream transcription factor PBX homeobox 1, producing IL-10 in macrophages [89]. Higher expression of cholesterol 25-hydroxylase, characteristically found in alveolar macrophages, contributes to the biosynthesis of 25-hydroxycholesterol, which stimulates the nuclear receptor liver X receptor to increase transcriptional activity during efferocytosis, leading to the escalation of TGF-β production [90]. Thus, alveolar macrophages have advanced efferocytosis activity, enabling them to promptly and effectively eliminate the apoptotic bodies that prominently appear during viral infection. Furthermore, this property is indispensable for preventing excessive pulmonary inflammation owing to the massive production of viruses and damage-associated molecular patterns (DAMPs) from apoptotic bodies that lose cell membrane integrity.
