Relative Impact of Form-induced Stress vs. Uniaxial Alignment on Multipotent Stem Cell Myogenesis

Tissue engineering strategies based on multipotent stem cells (MSCs) hold significant promise for the repair or replacement of damaged smooth muscle tissue. To design scaffolds which specifically induce MSC smooth muscle lineage progression requires a deeper understanding of the relative influence of various microenvironmental signals on myogenesis. For instance, MSC myogenic differentiation has been shown to be promoted by increases in active RhoA and FAK, both of which can be induced via increased cell-substrate stress. Separate studies have demonstrated MSC myogenesis to be enhanced by uniaxial cell alignment. The goal of the present study was to compare the impact of increased peak cell-substrate stresses vs. increased uniaxial cell alignment on MSC myogenic differentiation. To this end, MSC fate decisions were compared within two distinct multicellular "forms". A "stripe" multicellular pattern was designed to induce uniaxial cell alignment. In contrast, a second multicellular pattern was designed with "loops" or curves, which altered cell directionality while simultaneously generating regional peak stresses significantly above that intrinsic to the "stripe" form. As anticipated, the higher peak stress levels of the "loop" pattern were associated with increased fractions of active RhoA and active FAK. In contrast, two markers of early smooth muscle lineage progression, myocardin and SM-a-actin, were significantly elevated in the "stripe" pattern relative to the "loop" pattern. These results indicate that scaffolds which promote uniaxial MSC alignment may be more inductive of myogenic differentiation than those associated with increased peak, cell-substrate stress but in which cell directionality varies.