Background/Purpose: Macrophages and monocytes are increasingly implicated in the pathogenesis of autoimmune induced pulmonary fibrosis. In addition to the well-recognized classes of tissue resident macrophages alveolar macrophages (AMs) and interstitial macrophages (IMs), recent studies have identified a novel class lung tissue resident macrophage termed CD169+ Nerve and Airway-associated Macrophages (NAMs) whose role in autoimmune pulmonary pathology is unclear. Further, recent studies have demonstrated monocytes playing a critical role in the progression of pulmonary fibrosis. Interestingly, monocytes can be further subdivided into classical, non-classical, intermediate and fibrocytes, that are thought to play a key role in fibrosis/wound healing, but whose exact roles in autoimmune induced pulmonary fibrosis are unknown.

Methods: We utilized the bleomycin-induced murine model of pulmonary fibrosis in C57BL/6 wild-type (WT) mice in combination with transgenic murine models that selectively deplete AMs, NAMs, or both AMs/NAMs upon Intraperitoneal injection of diphtheria toxin (DT). DT injection (1000ng) were given 16 hours prior to bleomycin treatment (2.5u/kg). Mice were sacrificed on days 4, 11, 18 and 42. Lungs were fixed, sectioned and stained for IF or disassociated into a single cell suspension, stained, and analyzed by flow cytometry for both surface and intracellular targets, including a-Smooth Actin (aSMA)/Collagen-1a (Col1a).

Results: Bleomycin-treated mice whose NAMs were depleted were sicker than WT littermates, with greater weight loss (34% vs. 24%), reduced survival (35% vs 65%) and increased lung dry weight (67.7mg vs 55.9mg) Interestingly, NAM depletion resulted in increased monocytic infiltration compared to WT bleomycin-treated littermates. We further characterized the infiltrating monocytic compartment, via flow cytometry, as either ‘true’ monocytes (CD45+,CD11b+,CD140a-,SMA-,Col1a-), putative fibrocytes (CD45+,CD11b+,CD140a+,aSMA+, Col1a+) or a transitioning population (CD45+,CD11b+,CD140a+,aSMA+, Col1a-). Our analyses demonstrates that depletion of NAMs alters the infiltrating monocytic compartment in two key ways. First, we observed a 39% increase in the numbers of infiltrating monocytes that are (CD45+,CD11b+,CD140a-) (p=0.0195). However, these monocytes were noted to have more fibrocytic phenotype with higher levels of aSMA or Col1a expression, 23% (aSMA+/Col1a-) (p=0.0426), and 16% (aSMA+/Col1a+) (p=0.0292).

Conclusion: Depleting tissue-resident macrophages led to an increase in infiltrating monocytes skewed towards a more pro-fibrotic phenotype and resulting in a poorer clinical outcome. Taken together, these findings potentially provide insight into the mechanisms of inflammatory pulmonary fibrosis in autoimmune diseases.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/nerve-and-airway-associated-tissue-resident-pulmonary-macrophages-limit-infiltration-and-alter-phenotype-of-infiltrating-monocytes-and-fibrocytes-to-reduce-pulmonary-fibrosis/