Mechanisms of lung macrophage programming by MUC5B during health and disease

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PROJECT SUMMARY Mucus and macrophages protect the lungs in health, but they can also contribute to disease following lung injury. How their protective vs. pathological functions are calibrated is poorly understood. We seek to determine mechanisms that control their interactions during responses to lung injury. For mucus-mediated defense, the mucin glycoprotein MUC5B is essential. In mice, absence of Muc5b causes particles and bacteria to accumulate in the lungs, ultimately resulting in early mortality. Despite its requirement for health, MUC5B is an important risk factor in human pulmonary fibrosis, where it is misexpressed in bronchiolar club cells and type 2 alveolar epithelia. Overexpression of Muc5b in these cell types in mice potentiates fibrosis following bleomycin-induced lung injury. These data suggest that the levels and locations of MUC5B/Muc5b- expression are significant factors in the pathogenesis of pulmonary fibrosis. Nonetheless, we do not yet fully understand cellular and molecular mechanisms that explain how MUC5B/Muc5b promotes lung fibrosis. Our recent work suggests that defensive and pathologic effects of airway mucus are regulated by interactions between Muc5b and airspace macrophages (AMs). Resident AMs are present constitutively and are required for non-inflammatory defense. In response to injury, AM pools increase through recruitment of blood monocytes that mature into macrophages. These recruited AMs are more inflammatory than resident AMs, but they are also short-lived, resulting in transient expansion and then contraction of the AM pools. Mechanisms of acute and resolving inflammation that distinguish resident and recruited AM types also impinge on fibrotic repair. The presence of both MUC5B/Muc5b and AMs in distal airspaces, along with our prior observation of AM dysfunction in Muc5b knockout mice, implicate a connection between MUC5B/Muc5b and AM functions. We identified a potential mechanism mediated by mucin glycans and AM glycan receptors. MUC5B/Muc5b is heavily coated with sialic acid (SA) that is attached to galactose via an α2,3-linkage. It is also a ligand for sialic acid binding immunoglobulin like lectin-F (Siglec-F), an inhibitory signaling molecule found almost exclusively on AMs in healthy lungs. We found that a Muc5b-SA-Siglec-F axis is critical for resolving inflammation, as shown by prolonged recruited AM accumulation in mice lacking each component. We now also show that bleomycin-induced lung fibrosis is suppressed in Muc5b-SA-Siglec-F axis disrupted mice. Thus, while protective in response to bacterial inflammation, this mechanism appears to be detrimental in a pro-fibrotic injury setting. We hypothesize that fibrotic repair of lung tissues is mediated by a Muc5b-SA-Siglec-F dependent AM programming mechanism. This will be tested in three aims that test whether 1) promotion of lung fibrosis by Muc5b requires α2,3-sialylation; 2) ligation of Siglec-F by sialylated Muc5b mediates fibrosis; and 3) the Muc5b-SA-Siglec-F axis regulates fibrotic programming of resident vs. recruited AMs.