Crystal ribcage: a platform for probing real-time lung function at cellular resolution in health and disease

Abstract

Real-time, cellular resolution imaging is essential for probing the highly dynamic functions of the lung at the interface of physics, biology, and immunology. CT and MRI modalities have low spatial resolution, and histological approaches provide only snapshots of fixed lungs with little temporal information. Existing intravital imaging approaches cannot include and manipulate the physical and spatiotemporal changes involved in respiratory function. Here, we describe the development of a platform, termed "LungEx", to visualize and mechanistically probe the dynamics of a functioning lung at optical resolutions. LungEx is equipped with a novel transparent ribcage, termed "crystal" ribcage, that provides physiological conditions for a functioning lung and allows high-resolution and real-time optical imaging of nearly the entire lung surface. This imaging capability is obtained while LungEx preserves the complex 3-D architecture, cellular diversity, and integrative function of the ex vivo ventilated and perfused lung at near in vivo conditions. Utilizing LungEx in health and key lung diseases such as metastasis, pneumonia, and fibrosis, we probed a wide range of lung dynamic functions and remodeling at multiple spatial scales including alveolar deformation and elasticity, circulation-respiration coupling at the capillary level, cellular deformation, immune cell motility, and vascular transport. By modulating the biophysical environment of LungEx, we discovered that intravascular and interstitial, but not intra-alveolar neutrophils are highly and reversibly responsive to vascular pressure. Thus, by allowing high spatiotemporal imaging coupled with controlled biophysical interventions, LungEx, provides a powerful platform to mechanistically probe dynamic and multiscale lung functions in health and disease.

Particulate matter composition drives differential molecular and morphological responses in lung epithelial cells

“…With the development of single-cell technologies, in both transcriptional and morphological profiling, and the advances in RNA fluorescence in situ hybridization techniques, further studies could be performed to more directly link the expression of different transcripts with morphological features of individual cells. Additionally, the use of primary cells in a more realistic ECM environment or the use of ex vivo lung model technologies ( 62 , 63 ), and the testing of additional freshly collected PM mixtures containing volatile organic components could further improve the biological context of future work. Finally, the use of live-cell imaging to monitor cellular changes over time could lead to a better understanding of cluster stability among morphological patterns and would further enhance our ability to determine susceptibility to PM mixtures and other bioactive agents.…”