Temporally-Stiffening Hydrogel Regulates Cardiac Differentiation via Mechanosensitive Signaling

Stiffness of the extracellular matrix (ECM) surrounding cells plays an integral role in affecting how a cell spreads, migrates, and differentiates, in the case of stem cells. For mature cardiomyocytes, stiffness regulates myofibril striation, beating rate, and fiber alignment, but does not induce de-differentiation [1,2]. Despite improved myocyte function on materials which mimic the ∼10 kPa heart stiffness, the heart does not begin as a contractile ∼10 kPa material, but instead undergoes ∼10-fold myocardial stiffening during development [3]. Thiolated hyaluronic acid (HA) hydrogels have been used to mimic these stiffening dynamics by varying hydrogel functionality and component parameters. Recently, we have shown that pre-cardiac mesodermal cells plated on top of these stiffening HA hydrogels improves cardiomyocyte maturation compared to static, compliant polyacrylamide (PA) hydrogels [3]. While active mechanosensing causes maturation, the specific mechanisms responsible for responding to time-dependent stiffness remain unknown. Here we examined protein kinase signaling and mechanics in response to dynamic vs. static stiffness during the commitment process from embryonic stem cells (ESCs) through cardiomyocytes to better understand how developmentally-appropriate temporal changes in stiffness regulate cell commitment.