Fabrication of a Lung-on-chip Microfluidic Device Suited for Modeling the Alveolar-capillary Interface in a Healthy Lung and in a Lung with Adult Respiratory Distress Syndrome

Abstract

With each breath, the mammalian lung must undergo a sequence of expansion and compression, bringing fresh air into the lungs and exhaling waste carbon dioxide. As that air is bought into the lungs it flows through structures of increasingly small dimensions, maximizing the surface area available, until reaching the alveoli, the cellular structures responsible for gas exchange. With healthy lungs we are able to breathe with an ease that belies the complexity of the lung surfactant (LS) at work, enabling us to do so. At the cellular level a monolayer of lung surfactant, a lipid-protein mixture, coats the alveolar surface and minimizes the work required to inhale. LS has evolved to meet the demands of this expansion-compression cycle. Among these are spreading rapidly during inhalation, minimizing the surface tension but resisting monolayer collapse during exhalation, and acting as a barrier to the outside world. To meet these requirements, therefore, LS must consist of many components acting in concert. With such an important task, it comes as no surprise that the lack of LS or its dysfunction can result in disease states. For instance, a condition known as neonatal respiratory distress syndrome (NRDS) occurs in infants born prematurely, because in these babies the alveolar cells have not yet formed or secreted enough LS. These infants cannot overcome the additional work required to breathe and must undergo replacement LS therapy. In adult patients, existing surfactant can be displaced or inactivated by fluid buildup in the alveoli due to injury or illness in a condition collectively known as adult respiratory syndrome (ARDS). Treatment of these conditions using LS therapies could be improved or discovered with increased knowledge of the ways in which LS components interact at a molecular level to result in desired surfactant properties. The evaluation of lipid-protein interactions is a primary focus of our lab. Work of this nature is typically done using a Langmuir Trough coupled with Wilhelmy plate surface pressure measurements and fluorescent imaging. A microrheology system using magnetic nanorods has also enabled detection of small changes in interfacial viscosity that could not be detected by commercial rheometers. However, in order to truly evaluate LS properties, it is necessary to match the microscale environment of the alveoli as closely as possible. To this end, this thesis discusses the construction of a lung-on-chip device with the goal of creating an in vitro model of lung function that could be used in combination with the LS evaluation methods currently used in our lab. The lung model discussed here consists of a three layer microfluidic device constructed entirely of polydimethylsiloxane (PDMS) polymer. This device was constructed using microfabrication techniques capable of creating a microenvironment mimicking that of the alveolar-capillary interface. The model consists of top and bottom center channels separated laterally by a porous membrane (PM) and flanked by functional side channels. The top and bottom center channels create the alveolar and capillary sides of the interface, respectively. The flanking side channels serve to simulate breathing by subjecting the PM of the center channel to expansion and compression caused by cyclic application of negative pressure on the side channels. The PM serves as the scaffold on which alveolar cells and endothelial cells can be grown to create the in vitro model and can withstand the stretching associated with simulated breathing. Methods of microfabricating an intact porous membrane with through-holes were evaluated. Additionally, two methods of opening the top center channel of the device, CO2 laser ablation and precision milling via CNC, in order to create a true open air-liquid interface were evaluated