Lung Interstitial Macrophages Redefined: It Is Not That Simple Anymore

Tissue resident macrophages (TRMΦs) are required for embryonic development and postnatal tissue homeostasis, as well as for reinstating steady-state functionality after tissue injury (1). TRMΦs function by secreting various growth factors and cytokines that promote cell regeneration and survival. In addition, TRMΦs clear invading pathogens, surround dead cells (through a process known as efferocytosis), and clear extraneous cellular debris and fluids (2). Dysregulation of TRMΦ function has been associated with various acute and chronic diseases, including inflammatory lung injury (2, 3). Thus, it is crucial to understand the complexity of TRMΦ function to devise novel strategies for suppressing inflammation while accelerating tissue regeneration.

In the lung, two major populations of TRMΦs were previously thought to exist: alveolar (AMΦs) and interstitial (IMΦs) macrophages. AMΦs reside within the alveolus, whereas IMΦs localize to the interstitial space of the alveolar septum or alveolar corners (4) (Figure 1). Although AMΦs have been the principal focus of study because of their salient localization and the feasibility of tracking their behavior during lung injury and repair, the identity of other subsets of TRMΦs has remained largely elusive. AMΦs act as lung sentinels by sensing pathogenic insults and rapidly triggering leukocyte recruitment to mount host defense (5). Recently, subsets of nonsessile and sessile AMΦs have been described (6). Nonsessile AMΦs patrol the alveolar space and initiate an inflammatory response after sensing inhaled pathogens or toxicants, whereas sessile AMΦs remain attached to the alveolar epithelium and communicate with those cells to dampen the inflammatory response (6). In contrast, IMΦs appear to be functionally and phenotypically distinct from AMΦs (7, 8). IMΦs are proportionally less abundant (∼2–3-fold), have low phagocytic potential, and respond to pathogenic or inflammatory stimuli differently than do AMΦs (7–9). IMΦs have been shown to play an important role in blunting unwarranted immune reactions elicited by dendritic cells in steady-state and in response to pathologic stimuli (7). Strategies for depleting IMΦs or AMΦs using clodronate or F4/80 antibody (a pan monocytic/macrophage marker), respectively, have produced varied results. For example, depletion of IMΦs, but not AMΦs, exaggerated endotoxin/antigen-induced airway allergic responses (7), whereas depletion of sessile AMΦs augmented LPS-induced lung injury (6). Thus, a more thorough molecular characterization of IMΦs and AMΦs and a better characterization of their exact functions is required for understanding the complexity of MΦ populations in the lung under normal conditions and in disease states.

Figure 1.

Graphic representation of resident macrophages present in the lung. In the alveolus, sessile (attached) and nonsessile (unattached) alveolar macrophages (AMΦs) are shown. Alveolar interstitial macrophages (AIMΦs) and bronchial IMs (BIMΦs) represent MΦs residing in alveolar and bronchial interstitium, respectively. Phenotypic cell surface markers used to identify and characterize the specific MΦs are shown. Illustration by Jacqueline Schaffer.

In the July issue of the Journal, Gibbings and colleagues (pp. 66–76) describe studies using complementary in vivo and ex vivo approaches to provide convincing evidence detailing the lack of IMΦs in the alveolar–capillary lung interstitium during normal healthy steady-state conditions (10). These authors demonstrate that IMΦs in fact reside within the bronchial interstitium in close proximity to lymphatic vessels, cells we herein refer to as bronchial IMΦs (BIMΦs). The authors classified them as BIMΦ1 (CD11c^lo+MHCII^lo), BIMΦ2 (CD11c^lo+MHCII^hi), and BIMΦ3 (CD11c^hi+MHCII^hi) subsets. The BIMΦs have phagocytic potential similar to AMΦs both in vivo and ex vivo, but express distinct yet overlapping MΦ-specific cell surface phenotype markers compared with AMΦs. Gene expression analysis of these BIMΦs clearly revealed the presence of monocyte-specific markers (e.g., CD14, CD163, and Csfr1) and the absence of AMΦ-specific expression markers (e.g., Marco, Siglec-F, and Csfr2), supporting the notion that BIMΦs probably originate from circulating monocytes. Consistent with this view, increased expression of inflammatory mediators and receptors are noted in BIMΦs compared with AMΦs under steady-state. Using BrdU labeling and parabiotic mice, the authors show that within the three BIMΦ subsets, BIMΦ3 turnover rate is faster and they exhibit less phagocytic potential in vivo than do other two subtypes. They also report that three BIMΦs subpopulations exist in other organs such as the skin, gut, and heart and express similar gene expression patterns, suggesting a ubiquitous nature. The existence of macrophages in the bronchial interstitium was noted previously in the lungs of the rhesus monkey (7, 8), further supporting the authors’ findings and implicating their potential roles in human lung disease or diseases.

Previous studies have shown that MΦs exist in the interstitium of alveoli and capillary. For example, immuno-colocalization studies in lung tissue sections with anti-CD11c and anti-F4/F80 or CD206 antibodies revealed coexistence of IMΦs and AMΦs in the alveoli (7, 8), which we refer to as alveolar IMΦs (AIMΦs). However, in the current study, the investigators used c-mer proto-oncogene tyrosine kinase (MerTK)-positive staining as the bona fide marker for detecting MΦs (11) in the alveolar region of CX₃CR1^GFP+ reporter mice. It is noteworthy that the authors detected CX₃CR1^GFP+ cells within and around alveoli that were negative for MerTK immunostaining. It is possible that these cells either expressed low-to-undetectable levels of MerTK or represent another subset or subsets of AIMΦs. Also, the authors detected MerTK⁺GFP^- cells in the bronchial interstitium, but their identity remains unclear.

The findings of Gibbings and colleagues (10) no doubt represent an important step toward defining the role of BIMΦs in maintaining bronchial homeostasis. Further, their findings suggest that IMΦs have a role in modulating immune and angiogenic mechanisms not only in the alveolar region but also in the bronchiolar region in healthy as well as injured/diseased lungs. Nevertheless, the study raises several intriguing questions: Are these BIMΦs sessile or nonsessile, and do they communicate with the AMΦs to mitigate infection or facilitate repair after lung injury? Do these BIMΦ subsets acquire phenotypic transition by themselves, or do they acquire characteristics of AMΦs and/or AIMΦs as part of the tissue repair process? To address these important issues, specific tools such as subset-specific lineage tracing of BIMΦ populations will be needed to unravel the complexity of “BIMΦ subsets,” as well as their potential relationships with AMΦs and/or AIMΦs during the process of acute lung injury and repair, and in chronic airway diseases.