Ironing Out the Roles of Macrophages in Idiopathic Pulmonary Fibrosis.

Airway macrophages (AMs) act to remove inhaled particles, debris, allergens, and microbes, making them crucial to host defense and epithelial homeostasis (1). Cross-talk between AMs, dendritic cells, alveolar epithelial cells, and T cells regulates how the immune system responds to environmental lung stimuli. Thus, AMs are pivotal in the response to alveolar injury and the subsequent biological pathways regulating resolution or persistence of alveolar inflammation (2). Given this keystone role, AMs have been studied in both human disease and animal models of pulmonary fibrosis (3–5), where the prevailing notion is that AMs derived from circulating monocytes worsen disease (5, 6). However, the mechanism(s) responsible for the ability of AMs to promote pulmonary fibrogenesis are unclear.

In this issue of the Journal (pp. 209–219), Allden and colleagues advance knowledge about macrophages and idiopathic pulmonary fibrosis (IPF) (7). They used BAL acquired from two independent IPF patient cohorts from observational clinical studies and interrogated specific lung leukocyte phenotypes by complimentary techniques, including multiparametric flow cytometry matched with immunohistochemistry. They show increased proportions of AMs lacking surface CD71 (the transferrin receptor) in human patients with IPF. The authors carefully interrogated the phenotype of these CD71− AMs. Remarkably, CD71− AMs demonstrated impaired function with defective phagocytosis, reduced markers of macrophage maturity, and profibrotic gene activation. In a further provocative turn, the authors demonstrated an association between reduced survival and higher proportions of CD71− AMs in the BAL using a cohort of 97 patients with IPF. They conclude that CD71− AM percentages may serve as a novel biomarker of IPF disease progression and that the CD71 pathway may represent a target for therapeutic manipulation.

CD71 is an integral membrane protein that binds diferric transferrin complexes to mediate uptake into the cell via receptor-mediated endocytosis. The majority of iron in circulation in the steady state is bound to transferrin. Transferrin–iron complexes bind CD71, which serves as a cellular receptor but also serves to limit the ability of iron to catalyze the formation of free radicals from reactive oxygen species, resulting in iron toxicity (8). Furthermore, control of free iron is an important host defense function because it limits availability of iron to potentially pathogenic bacteria in vivo.

Given these associations between macrophages, iron, bacteria, and fibrosis, it is interesting to speculate about how and why loss of CD71 on IPF macrophages may be important for disease pathogenesis. Previous studies have shown elevated levels of iron and altered iron metabolism in IPF lungs (9–11). Allden and colleagues also demonstrated higher levels of transferrin, although they could not measure free iron, in the BAL of patients with IPF compared with control subjects.

Although there is an intriguing correlation among the patients with IPF themselves which shows the highest levels of BAL transferrin in patients with the lowest percentages of CD71+ AMs, the fact that overall there were still more CD71+ AMs in patients with IPF than in control subjects (in whom BAL transferrin levels are low) suggests that mechanisms in addition to CD71+ AM clearance (e.g., vascular leak and synthesis by other cells) may regulate the levels of BAL iron/transferrin.

How might the abundance of transferrin in the BAL of patients with IPF promote disease, and how is this related to CD71 expression by AMs? One intriguing possibility is that transferrin-bound iron in lungs of patients with IPF may promote lung microbiome alterations or dysbiosis.

Interestingly, transferrin-bound iron is a growth factor for Staphylococcus aureus (12). In IPF, studies have shown that increased lung bacterial load (16S ribosomal DNA gene copies) predicts patients who will have rapidly progressive disease (13, 14). Furthermore, Streptococcus and Staphylococcus species were identified as key taxa found in patients with IPF exhibiting progressive disease (15). Thus, it is tempting to speculate that low levels of AM CD71 may promote pathobiont growth of S. aureus via increased availability of iron–transferrin complexes in the airspaces.

Another equally interesting hypothesis is that CD71− AMs showed reduced phagocytosis. Certainly, a defect in bacterial clearance by these AMs may promote increased bacterial load noted in IPF progression. How CD71 and/or iron–transferrin complexes might regulate phagocytosis is unknown. This could be via direct signaling in the AMs or via regulation of cellular homeostasis and metabolism. Additional studies are needed to determine whether loss of CD71 on AMs is playing a pathogenic role via regulation of phagocytosis or microbiota directly or whether loss of CD71 is merely a passive biomarker of the progressive disease environment. Although the authors have validated the loss of CD71 on AMs in two different IPF patient cohorts, this measurement requires BAL sampling, which limits implementation of this as a biomarker for clinical use. Recent guideline recommendations for diagnosis of IPF may reduce the number of BALs performed for diagnostic purposes in patients with IPF.

A final possibility is that loss of CD71 identifies a profibrotic macrophage phenotype. Of note, the CD71-deficient AMs expressed higher levels of MMP2 (matrix metalloproteinase 2), MMP8, vascular endothelial growth factor, and plasminogen activation inhibitor 1, which might lead to increased activation of TGFβ (transforming growth factor-β), increased angiogenesis, decreased matrix degradation, and increased deposition of extracellular matrix. Again, how CD71 signaling may mediate these effects is unclear. Expression of CD71 has been linked with tumorigenic proliferation in esophageal squamous cell carcinoma (16); thus, it may have proliferative effects in AMs also.

Potentially, CD71+ AMs may expand and have a regulatory role in some patients. Certainly, recent single-cell transcriptomic analyses of human IPF lungs have revealed macrophage heterogeneity that is incompletely understood (17). It is also possible that low levels of CD71 characterize immature, recently recruited, monocyte-derived AMs, consistent with murine studies suggesting that monocyte-derived AMs persist and promote lung fibrosis (6).

In conclusion, the study by Allden and colleagues provides another intriguing analysis of the role of macrophages in IPF. This study highlights correlations between disease progression, impaired innate immune function, and profibrotic mediator production and hints that these features may reside in inflammatory or recruited lung AMs. Thus, what’s old is new again, and the field is rediscovering the likely importance of inflammatory/recruited AMs in the pathogenesis of IPF. Perhaps the finding that CD71 loss marks a potentially pathogenic subset of AMs will help to eventually “iron” out the role of this cell type in disease progression.