Bioengineering the bone microenvironment for improved craniofacial regeneration leveraging biomimetic mechanotransduction

The overarching goal of this proposal is to expand fundamental knowledge of bone repair mechanisms and improve regenerative strategies for craniofacial bone repair by investigating microenvironmental factors influencing osteogenic differentiation and bone regeneration. This work will employ a variety of biomimetic hydrogels, designed to recapitulate the extracellular matrix during distinct stages of bone formation: (1) soft collagen represents the initial matrix where undifferentiated stem cells reside, (2) high-density collagen reflects mesenchymal condensation, (3) partially mineralized high-density collagen simulates onset of calcification, and (4) a fully mineralized state indicative of mineralized woven bone. My preliminary data shows an early onset of osteogenic differentiation in human mesenchymal stem/stromal cell (hMSC) cultured on biomineralized hydrogels. However, the mechanisms to explain this accelerated regenerative response has not been thoroughly investigated. I hypothesize that increased in matrix density and mineralization will enhance osteogenic differentiation and bone regeneration. This hypothesis will be tested via Aim 1 by looking at adhesion-driven mechanotransductive mechanisms in vitro and Aim 2 by evaluating the in vivo bone healing response. Specific Aim 1 will examine the effect of the hydrogel model on osteogenic differentiation, focusing on the molecular clutch of mechanotransduction – a process where mechanical stimuli from the microenvironment is translated into biochemical activity, prompting cellular responses. This aim will evaluate the in vitro hMSC behavior in response to these hydrogels, looking at focal adhesion, mechanotransductive, and differentiation markers. Specific Aim 2 extends this analysis to an in vivo setting, testing the regenerative capacity of injectable microgels in rat calvarial (skull) defects. By providing a comparative analysis of bone healing with the aid of microgels of varying density and mineralization, this aim focuses on understanding the systemic host response to bioengineered grafts to be able to identify microenvironmental conditions that enhance bone regeneration. This application outlines a comprehensive training plan at a well-established institution, designed to develop my expertise in bone tissue engineering and stem cell biology, essential for becoming a craniofacial surgeon- scientist. My fellowship training will focus on gaining expertise in these areas, while developing professional skills in communication, leadership, and teaching, which are essential for my career goals of improving clinical outcomes through surgical innovation. Successful completion of this proposal will provide insights into designing scaffolds that support microenvironments conducive to accelerate differentiation, leading to improved bone regeneration outcomes. By elucidating how biomimetic environments influence differentiation and bone healing, my work aims to contribute to the development of next-generation bioengineered bone grafts, facilitating effective bone repair strategies and improving patient outcomes in craniofacial reconstruction.