Paramagnetic Beads and Magnetically Mediated Strain Enhance Cardiomyogenesis in Mouse Embryoid Bodies

Mechanical forces play an important role in proper embryologic development, and similarly such forces can directly impact pluripotency and differentiation of mouse embryonic stem cells (mESC) in vitro. In addition, manipulation of the embryoid body (EB) microenvironment, such as by incorporation of microspheres or microparticles, can similarly influence fate determination. In this study, we developed a mechanical stimulation regimen using permanent neodymium magnets to magnetically attract cells within an EB. Arginine-Glycine-Aspartic Acid (RGD)-conjugated paramagnetic beads were incorporated into the interior of the EBs during aggregation, allowing us to exert force on individual cells using short-term magnetization. EBs were stimulated for one hour at different magnetic field strengths, subsequently exerting a range of force intensity on the cells at different stages of early EB development. Our results demonstrated that following exposure to a 0.2 Tesla magnetic field, ESCs respond to magnetically mediated strain by activating Protein Kinase A (PKA) and increasing phosphorylated extracellular signal-regulated kinase 1/2 (pERK1/2) expression. The timing of stimulation can also be tailored to guide ESC differentiation: the combination of bone morphogenetic protein 4 (BMP4) supplementation with one hour of magnetic attraction on Day 3 enhances cardiomyogenesis by increasing contractile activity and the percentage of sarcomeric α-actin-expressing cells compared to control samples with BMP4 alone. Interestingly, we also observed that the beads alone had some impact on differentiation by increasingly slightly, albeit not significantly, the percentage of cardiomyocytes. Together these results suggest that magnetically mediated strain can be used to enhance the percentage of mouse ESC-derived cardiomyocytes over current differentiation protocols.