(I-351) Design and Validation of a Biomimetic Alveoli-on-a-Chip System

Introduction:: In the human respiratory system, the site for air exchange is the alveoli. Alveoli are highly dynamic organs, and they are constantly being inflated and deflated to allow the occurrence of air exchange. Inside the alveoli walls are the alveolar epithelial cells (AECs), which consist of Type I and Type II cells. In traditional in vitro studies, AECs are cultured on flat (2D) surfaces, which lack physiological alveolar architecture. Thus, the need exists for a platform with relevant alveolar architecture enabling the recapitulation of both physiological and aberrant microenvironments to evaluate physiological and pathological cellular changes in human alveoli. Herein, we propose a biomimetic alveoli-on-chip system composed of a controllable stretchable substrate.


Materials and Methods:: The constructed alveoli-on-a-chip system uses a programmable pressure controller (PreciGenome® Pressure Control Lite Version) that connects with an external vacuum source and a computer that has real-time control on the controller. The controller is connected an opening of a commercially available microfluidic Ibidi® μ-slide. A stainless-steel disc with holes (350μm in diameter) is placed on top of the other opening, and then sealed with a thin Gel-Pak® PDMS diaphragm (38.1μm in thickness). To measure deflection of the PDMS diaphragm, the collagen coating was stained via immunofluorescence and subsequently imaged using confocal microscopy. Lung epithelial cells (A549 cells) were seeded on the diaphragm, and viability was confirmed with a Live/Dead Staining Kit. YAP and F-actin staining were performed to evaluate the morphological response of cells to static and dynamic stretch.


Results, Conclusions, and Discussions:: We demonstrate that PDMS diaphragms in the system can withstand static stretch between -2.5 PSI to -7.5 PSI. This pressure difference can induce strain beyond the physiological range in alveoli. For proof-of-concept and to validate the system, we characterized the response of AEC cells, under physiologic stretch conditions. Since the physiologic range of strain in alveoli is 5-15%, we selected our physiologic pressure range to be -3.5 PSI to -4.5 PSI, with a maximum strain of 13.8%. Confocal imaging showed that the range of deflection achieved for physiological pressure change was from 112.7±1.7 mm to 140.0±0.8 mm. Viability analysis over 24 hours of static and dynamic physiological stretch showed that A549 cells adhered and grew well on PDMS diaphragms. We observed a higher density of cells in the peripheral region of the dimples where there was higher level of tensile forces. YAP quantification showed the decrease in cytoplasmic YAP in static and cyclic stretch culture compared to that of flat culture. F-actin staining showed that cells aligned perpendicular to the direction of radial stretch in dynamically stretched groups, with significantly higher expression compared to that found in cells under static stretch conditions.
We present here a novel, biomimetic culture system for lung alveolar epithelial cells with physiologically relevant architecture. We also showed that our system is capable of recapitulating both the physiological and aberrant biophysical microenvironment of human alveoli. Our future plan to evaluate lung epithelial cell performance under pathological cyclic stretch, as well as examining the effect of cyclic stretch on primary human Alveolar Type II (ATII) cells and pluripotent-derived human alveolar epithelial cells.


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