Substrate Stiffness Controls the Cell Cycle of Human Mesenchymal Stem Cells Via Cellular Traction

The microenvironment of human mesenchymal stem cells (hMSCs) regulates their self-renewal and differentiation properties. Previously, it was shown that hMSCs remain quiescent on soft (0.25 kPa) polyacrylamide (PA) gels, but re-enter the cell cycle on a stiff (7.5 kPa) gel. However, how cells behave on intermediate stiffness and what intracellular factors transmit mechanical changes to cell interior, thereby regulating cell cycle, remained unknown. In this work, we have demonstrated that PA gels of stiffness between 1 kPa and 5 kPa act as a mechanical switch from quiescence to proliferation in the cell cycle of hMSCs. Further studies with ROCK inhibitor Y-27632 have revealed that contractile proteins, but not cell spread area, account for the sensitivity of hMSCs towards substrate stiffness, and hence correlates with their changes in cell cycle. These observations therefore suggest that substrate stiffness regulates hMSC proliferation through contractile forces, as generated by cellular contractile proteins in a unique pattern that is distinct from the two other cell types studied here, C2C12 and 3T3.