DESCRIPTION (provided by applicant): It is well established that adherent cells change their orientation in response to mechanical non- uniformities of the substrate, including substrate stretching and material anisotropy. While it is known that cells use their contractile machinery to mechanosense these nonuniformities, it is not known how mechanical signals from the substrate and intracellular contractile forces cooperate to govern cell orientation. The current paradigm holds that mechanical nonuniformities from the substrate are a precondition for a cascade of biochemical and remodeling events that govern cell to a new position. Our recent theoretical study suggests that reorientation and steering of the cell in response to substrate nonuniformities may be governed and even regulated by mechanical mechanisms. This led us to the following novel, mechanistic, and testable working hypothesis: In response to mechanical nonuniformities of the substrate, the cytoskeletal contractile stress dictates orientation of the cell via a simple physical mechanism - namely the contractile torque - that steers the cell towards a new orientation. The main goal of this study is therefore to investigate the relationship between the contractile torque and cell reorientation. To accomplish this, we need new experimental tools for measuring the contractile torque. Thus, we propose two aims: 1. To measure the contractile torque during cell reorientation in response to nonuniform, static and dynamic, substrate stretching using the cell mapping rheometry technique. 2. To design a substrate with alterable material isotropy in order to measure the contractile torque during cell reorientation in response to induced material anisotropy of the substrate. Although the proposed work is fundamental to any adherent cells, here we limit attention to the questions of airway smooth muscle (ASM) cell contractility, mechanosensing and positioning on the substrate. These questions are crucial to airway mechanics because, if appropriate, constriction of airways by ASM cells is the key to normal airway narrowing. If successful, this investigation will advance our understanding of the mechanisms by which the mechanical microenvironment dictates organization and contractility of ASM cells in their natural habitat. PUBLIC HEALTH RELEVANCE: A growing body of evidence indicates that the microenvironment of the airways may contribute to the development of pulmonary diseases, such as hypersensitivity to allergy and asthma, by altering structure and spatial organization of airway smooth muscle cells. The present study will investigate the physical mechanisms by which airway smooth muscle cells sense mechanical changes in their microenvironment and respond by altering their orientation and alignment. If successful, this study will establish physical principles that govern airway smooth muscle cell spatial organization and identify potential targets for therapeutic intervention in the future.

Project Narrative A growing body of evidence indicates that the microenvironment of the airways may contribute to the development of pulmonary diseases, such as hypersensitivity to allergy and asthma, by altering structure and spatial organization of airway smooth muscle cells. The present study will investigate the physical mechanisms by which airway smooth muscle cells sense mechanical changes in their microenvironment and respond by altering their orientation and alignment. If successful, this study will establish physical principles that govern airway smooth muscle cell spatial organization and identify potential targets for therapeutic intervention in the future.