The Role of Tension Anisotropy in Fibroblast Activation

Project Summary Fibroblasts play a crucial role in tissue function by generating and modulating contractile forces in response to environmental cues. Upon activation, they upregulate actin and myosin to enhance contractile force generation, a process vital for wound closure and tissue repair. However, prolonged fibroblast activation can lead to pathological outcomes, including fibrocontractile diseases. Traditional mechanobiology has identified four primary mechanical factors influencing fibroblast activation: ECM stiffness, ECM microstructure, ECM viscoelasticity, and mechanical stress magnitude. Our recent work introduces a fifth factor, stress anisotropy, which refers to directional stress differences (Nature Materials, 2024). Our findings show that stress anisotropy significantly impacts fibroblast activation and can be modulated. Furthermore, we demonstrate that stress anisotropy can induce cellular mechanical memory, allowing fibroblasts to retain an activated state long after the initial stress is removed (PNAS, 2024). Building on these discoveries, we will investigate the effects of stress anisotropy at the cellular, tissue, and clinical scales: Specific Aim 1 will explore strategies for controlling long-term fibroblast activation by reducing stress anisotropy. Using our advanced micropatterning technology, we will independently manipulate ECM stiffness, cell shape, cell area, and the magnitude and direction of stretching to test whether reducing stress field anisotropy can prevent prolonged fibroblast activation in clinical settings such as skin grafting. Specific Aim 2 will investigate the interplay of extracellular and intracellular factors in fibroblast responses to anisotropic stress fields. We hypothesize that changing ECM properties — due to aging or diseases like fibrosis — affects mechanical signal transmission, impacting fibroblast activation. Our in vitro model of cell-collagen tissues, which generates both isotropic and anisotropic stress fields within a single tissue specimen, will enable us to dissect these responses. Specific Aim 3 will focus on ex vivo studies to optimize skin grafting techniques by controlling stress anisotropy through meshing parameters. Traditional grafts are meshed with parallel slits and stretched to cover larger wounds. Our preliminary data suggest that modifying slit length and orientation can optimize tension anisotropy control. We will also investigate kirigami-inspired meshing, which may provide superior control over anisotropy, in skin grafting. Success in these aims will yield validated tools for stress measurement and control, refined surgical techniques, and clear markers of cellular response, advancing therapeutic strategies for skin grafting.