Center of Biomedical Research Excellence in Matrix Biology Phase II

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1. PROJECT SUMMARY - Role of Cellular Mechanotransduction of Low Intensity Vibrations in Regulating Extracellular Matrix Synthesis 1.1. Summarize the goal of the parent award: The long-term goal of the Center of Biomedical Research Excellence (COBRE) in Matrix Biology is to establish, enhance, and actively advance a multidisciplinary research center focusing on improving our understanding of the role of the extracellular matrix in development, health, and disease, and contributing to the prevention, treatment, and cure for diseases of high priority. The specific aims of the COBRE Matrix Biology Parent award are: 1) enhance and grow upon the critical mass of investigators established around the thematic multidisciplinary focus of matrix biology, 2) enhance biomedical research core capabilities, 3) grow research collaborations with existing programs, and 4) enhance research training opportunities. This project will supplement the existing COBRE Matrix Biology award to form a new team of investigators that bring together three investigators from IDeA states with different perspectives and expertise to address complex basic, behavioral, clinical and/or translational research questions with complementary approaches. The research question does not duplicate those currently being pursued by the parent award and clearly benefits from the collective efforts of the collaboration. 1.2 Research question to be addressed by the supplement award: Engineering biophysical signals promises non-pharmacologic interventions to direct tissue regeneration in conditions that devastate bone such as osteoporosis, aging, injury, bedrest, or microgravity. Externally applied Low-Intensity Vibrations (LIV), a mechanical signal similar to muscle activity, offers a readily usable technology to stimulate Mesenchymal Stem Cell (MSC) anabolism for both tissue engineering and clinical approaches. LIV does not generate significant matrix deformations in vivo, thus excluding most mechano-transduction mechanisms previously proposed for high-magnitude and low-frequency mechanical signals (e.g., exercise). This presents a significant gap knowledge about bone mechanobiology and prevents utilization of LIV as an effective treatment for bone loss. MSC’s ability to replace and rejuvenate bone cell populations are informed by both dynamic mechanical forces generated during daily activities (e.g. muscle activity) and by the quality of the Extracellular Matrix (ECM). Yes1 Associated Protein (YAP) is a transcriptional co-activator that can activate the expression of genes in response to mechanical force, including ECM molecules such as Connective Tissue Growth Factor (CTGF) to regulate collagen production in cells. For tissue engineering and clinical approaches to ultimately succeed, causative information on how high-frequency signals generated by LIV are sensed, transduced, and eventually lead to nuclear YAP expression and ECM production is critical. This proposal aims to address a fundamental gap in bone mechanobiology by mechanistically establishing a mechanosensory function of the cell nucleus to respond to dynamic accelerations produced by LIV. Using our novel team approach, we will test whether LIV generates relative motions of the nucleus within a cell to strengthen nucleo-cytoskeletal scaffolding and to increase force-induced YAP signaling in the cell nuclei to elicit ECM production. We will address this through three sub-hypotheses and specific aims. The aims of this study are to determine in live cells if 1) vibration frequency and acceleration modulate the LIV- induced nuclear motions and resulting F-actin remodeling, 2) LIV-induced perinuclear F-actin remodeling will increase cytoskeletal tension on the nucleus, 3) the magnitude of cytoskeletal tension on the nucleus determines the magnitude of YAP nuclear entry and ECM production. Completion of these aims will provide knowledge on (1) how to enhance the efficacy of LIV-based regenerative modalities in clinic, and (2) foundational structure-function relationships in MSCs. Results will ultimately enable engineering LIV-based approaches that target nucleo-cytoskeletal connectivity with application in many areas including, but not limited to, tissue regeneration, tissue engineering, and aging. 1.3 Benefit of team science effort: This proposed supplement cannot be accomplished by any single investigator and requires an orchestrated effort by three investigators working in different fields: cell mechanobiology, machine learning, and computational biomechanics. Co-Project Lead (CPL) Uzer will work on establishing experimental methods to measure nuclear motion, high-fidelity cell and biological outcomes, ECM production, and nuclear YAP levels. CPL Satici will focus on developing machine learning algorithms to reconstruct 3D nuclear and cytoskeletal geometries in response to LIV in live cells. CPL Fitzpatrick will develop finite element (FE) models to quantify cytoskeletal forces on the nucleus under LIV treatment. Upon successful completion of this work, our team will establish, for the first time, a novel pipeline for data-driven, cell-specific FE models for understanding force-function relationships in cells. This novel method will lead us to new grant submissions to study the role of cell-specific forces in maintaining healthy cell function and ECM composition as well as informing new translational studies that use LIV to attenuate disease progression both in vitro and in vivo.