Inhaled Microplastics Inhibit Tissue Maintenance Functions of Pulmonary Macrophages

Abstract Rationale: Microplastics, particles smaller than 5 millimeters which slough off plastic as it degrades, are ubiquitous pollutants which have been observed in human lung tissue, sputum, and bronchoalveolar lavage. Yet, the effects of microplastics on the lung, and the pulmonary immune system in particular, remain undefined. Microplastics in the lung often cluster in macrophages suggesting that they are not readily broken down and cleared. Notably, studies have identified plastic laden macrophages within plaques removed after carotid endarterectomy which correlated with a higher risk for myocardial infarction, stroke, and all cause death. We hypothesized that inhaled microplastics cause both long-lasting pulmonary macrophage dysfunction as well as extra-pulmonary deposition with systemic effects. Methods: Macrophage cell lines (Raw264.7, THP-1, U937) and human monocyte-derived macrophages were cultured in vitro with fluorescent polystyrene microplastics of sizes 0.02µm, 0.1µm, 1.0µm, 5.0µm, and 10µm at various ratios and phagolysosomal processing was determined by immunofluorescence microscopy. Macrophage zymosan and E.coli uptake and digestion, migration, and cytokine production post-exposure were measured in the presence or absence of the AMP kinase activator AICAR (Acadesine). Female and male FVB/N mice were intranasally exposed to combined microplastics (0.02-10µm), with each size particle marked by a unique fluorescent signal, and pulmonary macrophage phenotype and function were determined ex vivo. Extrapulmonary dissemination of sized microplastics in tissues and association with macrophages was detected by immunohistochemistry. Results: Exposure to any size microplastic reduced macrophage phagocytosis at 24hrs. Larger microplastics (10µm) inhibited lysosomal processing of E.coli by 24hrs whereas smaller plastics (1µm) reducing processing at 3 days post-exposure. Phagocytosed microplastics found to amass in the late (Rab7+) phagosome but were largely absent from either the early (Rab5a+) phagosome or the lysosome (LAMP-1+). To restore function, microplastic exposed macrophages were co-cultured with increasing concentrations of the Acadesine which significantly improved macrophage phagocytosis and lysosomal processing at 100µM. Microplastics were detected in the lung, brain, liver, kidney, heart, and colon for up to 7-days after respiratory exposure, with smaller particles demonstrating wider extrapulmonary dissemination but larger particles present for longer periods of time. Conclusions: These findings indicate that microplastics can directly inhibit the ability of macrophages to clear particulate matter, a process which may be reversed pharmacologically. Notably, microplastic-induced pulmonary macrophage dysfunction may lead to increase susceptibility to infection, chronic tissue damage, and ultimately lung cancer.