C20-01 Mosaic Pattern: Lung Functional Heterogeneity at the Alveolus Level

Abstract Rationale Inhaled particles carrying pathogens, pollutants (e.g., microplastics, smoke), therapeutics, and diagnostics are increasingly relevant to public health. However, real-time transport of aerosols in functional alveoli is poorly understood as clinical imaging methods (CT, MRI) lack sufficient spatiotemporal resolution for probing alveoli, and histology provides only static endpoints. Subsequently, factors governing aerosol deposition dynamics in alveoli have remained unexplored. We developed a transparent “crystal” ribcage that enables optical access to intact, ventilated lungs, allowing real-time visualization of single aerosol droplets during physiological breathing. Using this platform, we observed the formation of a deterministic, heterogeneous “mosaic pattern” of aerosol distribution, in which only specific alveolar clusters received particles. Methods Fluorescent aerosols (1-10 µm) were delivered under spontaneous breathing in vivo or during ex vivo mechanical ventilation in mouse lungs. We utilized the crystal ribcage (Banerji & Grifno, Nat. Methods, 2023) to probe the dynamics of aerosol transport at high spatiotemporal resolution. Results Inhaled aerosols deposited in a spatially heterogeneous, deterministic “mosaic pattern,” consisting of high-deposition regions (“tiles”) separated by alveoli with negligible deposition (“bands”) (Fig. 1a-b). Sequential delivery of aerosols confirmed the reproducibility of this pattern in vivo and ex vivo, indicating that deterministic structural features, not stochastic events, govern deposition (Fig. 1c). The mosaic pattern consistently formed regardless of aerosol composition (small molecules, nanobodies, nanoparticles, microplastics, and pathogens) and was independent of ventilation modality (spontaneous breathing, positive- and negative-pressure ventilation) (Fig. 1d-e). Similar heterogeneity was observed in porcine and human lungs, and the mosaic pattern evolved with age in mice, where the size and distribution of tiles increased from birth to adulthood (Fig. 1f-h). The post-deposition stability of the pattern was age and molecular weight-dependent, lasting from a few minutes for small molecular-weight particles to multiple days for cell-binding particles (Fig. 1i-j). Mouse models of emphysema, fibrosis, and asthma did not disrupt the overall architecture of the mosaic pattern at the lobe level, but remodeled the shape and size of individual tiles. Conclusion We identified a previously unrecognized, spatially heterogeneous “mosaic” pattern of aerosol deposition that evolves from birth through aging in mice with indication in larger animals and humans. This finding challenges the assumption of uniform alveolar exposure, revealing deterministic, structured aerosol distribution that produces local variability in epithelial and immune exposure. The mosaic organization may reflect intrinsic biological compartmentalization, providing new insights into lung development and disease origins such as pneumonia, COPD, asthma, fibrosis, and lung cancer. This abstract is funded by: DP2HL168562; Beckman Young Investigator Award; NSF CAREER Award; Hevolution/AFAR New Investigator Award