Abstract
The lineage specification of mesenchymal stem cells (MSCs) is dependent on matrix stiffness, with osteogenesis promoted on or within stiff microenvironments. Unlike native extracellular matrix, conventional hydrogels have relatively static and uniform mechanical properties, thereby limiting the study of how spatiotemporally controlled mechanical cues impact MSC behavior. Here, we spatiotemporally control MSC differentiation in 3D hydrogels by actively modulating matrix stiffness using acoustic droplet vaporization (ADV). An acoustically responsive scaffold (ARS) was generated by co-encapsulating MSCs and perfluorohexane-based emulsion (13 μm) within type I collagen. ADV was used to generate bubbles within the ARS. Bubble diameter and the width of the compacted matrix region increased over time following ADV. Atomic force microscopy measurements demonstrated that the hydrogel region proximal to the bubble was significantly stiffer than the distal regions. Significantly greater levels of Runx2 and osteocalcin expression were observed in MSCs proximal to bubbles compared to distal on days 7 and 14. Higher alkaline phosphatase activity also validated the above findings. The significant upregulation of these osteogenic biomarkers suggests ADV-induced, mechanical changes in ARSs were sufficient to enhance osteogenic differentiation of MSCs. This approach is a significant step towards controlling the 3D differentiation of MSCs in a spatially localized, non-invasive, and on-demand manner.
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