Despite significant efforts, no cure exists for the major respiratory diseases such as asthma, emphysema and pulmonary fibrosis due to a lack of proper preclinical methods. Drug development is often carried out in cultured cells. While amenable to imaging and manipulation, this approach lacks the native tissue environment and neglects the effects of mechanical forces due to breathing which influence many signaling pathways. Alternatively, animal experiments are low throughput and performed using knock-out or transgenic mice which is also limited in its ability to target, image and pharmacologically manipulate individual cells. The Precision Cut Lung Slice (PCLS) is a preparation at an intermediate scale that permits visualization of cells and the extracellular matrix in the native lung tissue. However, current methods 1) do not deliver periodic stretch that mimics breathing; 2) do not determine parenchymal or airway wall stiffness, or airway contractile force, functional manifestations of lung disease; and 3) are not suitable for high throughput drug screening. We propose to design, build and test a universal high throughput mechanobiological tool, called the AccuStretch, which can stretch up to 20 PCLS samples with breathing-like waveforms for extended periods of time (>21 days) while simultaneously allowing measurement of stiffness and airway contractile force as well as cellular imaging and pharmacological manipulation.
Our specific aims are as follows:
Aim 1 : Design, build and test a versatile multi-well mechanobiology tool, the AccuStretch. (6 months). We will attach gels of known stiffness to the wells and verify the stiffness measurements in the AccuStretch. Using computational finite element analysis, we will fully validate the method.
Aim 2 : Stretch rat PCLS samples including an airway with breathing-like waveforms and determine parenchymal stiffness and airway contractile force in response to methacholine challenge. (6 months) Rat PCLS samples will be prepared and stretched in the AccuStretch for up to 21 days to verify tissue viability and to determine stiffness and airway contractile force. For comparison, parenchymal stiffness and airway contractile force will be independently measured in tissue strips from the PCLS in an organ bath system. In this Phase I application, we will combine the complementary expertise at Mechanobiologix LLC and Boston University to produce a prototype AccuStretch system. To achieve these goals, scientists at Mechanobiologix LLC will organize the studies, design the AccuStretch, carry out computational simulations and create various test membranes. System building and testing as well experimental procedures using PCLS will be contracted to Boston University. In Phase II, we plan to move on toward commercialization by prototype refinement and disease modeling using human PCLS. The end result will be a versatile device that will find countless applications in respiratory as well as other laboratories suitable for basic research as well as drug discovery with an estimated total addressable market of $120M worldwide.
The major respiratory diseases - asthma, pulmonary fibrosis and emphysema - contribute significantly to mortality and morbidity worldwide. Despite significant efforts into basic science and drug development, no cure exists for these diseases due in large part to a lack of proper preclinical methods. This proposal seeks to design, build and test a novel screening device that is expected to revitalize respiratory basic research and drug discovery.
