While the impact of single extracellular matrix (ECM) proteins and mechanical stiffness on cell function have been thoroughly probed individually, little work has been put into to understanding their interactions in the context of cell function. This is particularly important as the ECM is a complex mixture of proteins that change throughout normal development both in composition and stiffness. Recent work by others has demonstrated that cells respond differently to both static substrate stiffness and mechanical stretch when plated on substrates of different compositions. In addition, the effects of growth factor treatment can also be modulated by substrate composition and stiffness. In this talk I will cover our own recent work investigating the effects of alterations in stiffness and composition of the substrate on cardiac differentiation of stem cells and cardiomyocyte proliferation. The system we use is a polyacrylamide gel system with binding sites generated from solubilized decellularized cardiac ECM. This setup effectively allows us to decouple stiffness and composition to investigate their individual roles and any synergistic/ antagonistic effects. Preliminary data indicate that ECM composition and stiffness interact in a complex manner to effect cardiac differentiation of mesenchymal stem cells. Moreover, fetal cardiac ECM composition enhances neonatal cardiomyocyte proliferation over adult cardiac ECM or normal tissue culture plastic.